Adhering to the guidelines: rates of BRCA mutation using NCCN genetic testing criteria

Anna Beck MD1, Haimiao Yuan MS1, Junlin Liao PhD1, Pamela Imperiale-Hagerman BS2, Krysten Shipley MS, CGC1, Lillian Erdahl MD1, Sonia Sugg MD1, Ronald Weigel MD,PhD1, Ingrid M Lizarraga MBBS1

1University of Iowa Hospitals and Clinics, 2University of Iowa Carver College of Medicine

Presenter Information: Anna Beck, MD; anna-beck@uiowa.edu; 920-562-0420

Background

BRCA risk assessment is recommended by the National Comprehensive Cancer Network (NCCN) in patients with breast cancer who meet at least one of 7 criteria. Limited data are available regarding the likelihood of detecting a mutation when these criteria are followed.

Methods

At a single institution, 199 patients with breast cancer who underwent BRCA testing were identified. Patients underwent genetic counseling and testing based on NCCN guidelines. A retrospective chart review examined patient and tumor characteristics, NCCN criteria applied and BRCA testing results.

Results

Twelve (6.0%) of the 199 patients had a deleterious BRCA mutation. The most common individual reason for testing was high risk family history. Patients with BRCA mutation were more likely to have ER or PR negative cancer (75%, p=0.003). The criteria which yielded the highest rates of BRCA mutation were: family history of BRCA mutations (50%, p=0.019), age ≤45 at diagnosis (9.7%, p=0.034) and meeting ≥3 NCCN criteria as reasons for testing (13.3%, p =0.03). Having a known family history of BRCA mutation and age ≤45 remained associated with increased rate of BRCA mutation on multivariate analysis (OR 14.3, CI 1.2-166.3; OR 11.6, CI 1.2-108.6). Meeting only one criterion for testing yielded a mutation rate of 2.3%.

Conclusion

Using NCCN guidelines for genetic high-risk assessment, a deleterious BRCA mutation was detected in 6% of breast cancer patients. Criteria associated with higher rates of BRCA mutations were identified and may be higher yield when deciding to delay surgical planning to wait for BRCA testing results.

Poster Abstracts-HCCC Scientific Retreat 2018 *Oral Presentation (PDF page matches poster list numbers)

BACKGROUND: Homologous recombination (HR) is an essential pathway that repairs deleterious forms

of DNA damage such as interstrand DNA crosslinks, stalled replication forks and double strand breaks.

While deficiency in HR is risk factor in cancer development, cancers may also become dependent on

excessive HR, making this process an attractive target for cancer therapy. Following resection of the 5’

end in HR, the 3’ overhang is immediately coated by the ssDNA-binding protein, replication protein A

(RPA). The high affinity of this interaction kinetically blocks the formation of the Rad51 nucleoprotein

filament – the active species in the HR. Rapid displacement of RPA requires the action of a

recombination mediator, BRCA2 in humans or Rad52 in Saccharomyces cerevisiae. The underlying

mechanism by which a recombination mediator promotes the formation of the Rad51 nucleoprotein

filament has remained elusive.

METHODS: RPA contains four DNA binding domains (DBDs), which have been proposed to undergo

dynamic interactions with ssDNA within the RPA-ssDNA complex. We have used RPA with individually

labeled DBDs and single molecule total internal reflection microscopy (smTIRFM) to elucidate domain

level conformational dynamics of RPA on ssDNA.

RESULTS: We observe various levels of fluorescence enhancement that are domain-dependent;

suggesting microscopic dissociation/association events in the bound complex. Both DBD-A and D of RPA

showed 4 states of fluorescence in smTIRFM experiments, though the behavior of the A and D domains

are distinct. Addition of Rad52 resulted in a loss of the highest state of RPA DBD-D fluorescent

enhancement, though had no effect of DBD-A.

CONCLUSIONS: The effect of Rad52 on RPA DBD-D dynamics, but not DBD-A, suggests the Rad52 acts to

limit the dynamics of RPA DBD-D specifically. The limitation of RPA DBD-D dynamics by Rad52 may allow

for nucleation and filament formation of Rad51, thus promoting HR.

Abstract title: Single molecule observation of the Rad52 recombination mediator mechanism

Authors (in order submitted by poster presenter):Colleen Caldwell / Department of Biochemistry, University of Iowa Carver College of MedicineNilisha Pokhrel / Department of Biological Science, Marquette UniversityJoseph Tibbs / Department of Physics, University of Northern IowaAli Tabei / Department of Physics, University of Northern IowaMarc S Wold / Department of Biochemistry, University of Iowa Carver College of MedicineEdwin Antony / Department of Biological Science, Marquette UniversityMaria Spies / Department of Biochemistry, University of Iowa Carver College of Medicine

Despitetheincorporationoftheepidermalgrowthfactorreceptor(EGFR)inhibitorcetuximabintotheclinicalmanagementofheadandnecksquamouscellcarcinoma(HNSCC),limitedtonolong-termchangesinoverallsurvivalareobservedinHNSCCpatientseventhoughEGFRisexpressedathighlevelsinthesetumors.Therefore,theidentificationofnoveltherapeuticapproachestoenhancetheclinicalefficacyofcetuximabcouldleadtoimprovedlong-termsurvivalforHNSCCpatients.Ourpreviousworksuggeststhatcetuximabactivatestheinterleukin-1(IL-1)pathwayviatumorreleaseofIL-1alpha(IL-1α),althoughtheimplicationsofactivatingthispathwayareunclear.TheIL-1pathwayplaysacentralroleinimmuneresponseanddisplaysbothpro-tumorandanti-tumoractivities.IL-1maypromotetumorgrowthbyupregulatingthesecretionofpro-inflammatorymediatorsinvolvedinangiogenesisandmetastasis.Ontheotherhand,IL-1signalingmaypromoteantitumorimmunityspecificallyvianaturalkiller(NK)-cellmediatedantibody-dependentcell-mediatedcytotoxicity(ADCC),whichisalsoanimportantmechanismofactionofcetuximab.ThegoalofourworkistodeterminehowmodulationoftheIL-1pathwayaffectsHNSCCtumorresponsetocetuximab.WefoundthatblockadeofIL-1signalingusinganIL-1-receptorantagonist(IL-1RA,anakinra),neutralizingIL-1α/IL-1βantibodies,andgeneticknockdownoftheIL-1Rallsuppressedtheanti-tumorefficacyofcetuximab,whileIL-1αoverexpressionandtreatmentwithrecombinantIL-1αenhancedHNSCCtumorresponsetocetuximabinimmunodeficientandimmunocompetentHNSCCmousemodels.Mechanistically,theseresultsappeartobeduetomodulationofADCC,aswefoundthatIL-1blockadesignificantlyreducedcetuximab-mediatedADCCinvitro.Additionally,wefoundthatHNSCCpatientswithhighbaselinecirculatinglevelsofIL-1ligands(IL-1α,IL-1β)weresignificantlymorelikelytorespondfavorablytocetuximabmonotherapycomparedtopatientswithlowornobaselinecirculatingIL-1ligands.Altogether,theseresultssuggestthatIL-1signalingisnecessaryforHNSCCtumorresponsetocetuximab.Therefore,IL-1αwarrantsfurtherstudyasanoveltherapeutictoenhanceresponsetocetuximabandasanimmunologicbiomarkerforcetuximabresponse.

Abstract Title: Interleukin-1 signaling is required for HNSCC tumor response to cetuximab

Authors (in order submitted by poster presenter):Madelyn Espinosa-Cotton, Department of Radiation Oncology, University of Iowa Rachel Dahl, Department of Pathology, University of IowaIsaac Jensen, Department of Microbiology and Immunology, University of Iowa Ayana J. McLaren, Lincoln UniversityKenley Miller, San Jacinto CollegeSamuel N. Rodman, Department of Radiation Oncology, University of IowaElana Fertig, Johns Hopkins UniversitySandra Schmitz, Universite catholique de Louvain Andrean L. Simons, Department of Pathology, University of Iowa

Background. Assignment of breast cancer therapy is defined by immunohistochemical analyses that evaluate steroid hormone and growth factor receptors in tumor biopsies. Patients whose tumors are devoid of these receptors have fewer treatment options, and poor survival. This outcome is more commonly measured among Black versus Caucasian patients, and in patients with intact ovarian function. G-protein coupled estrogen receptor, GPER, triggers the release of nascent epidermal growth factor-like ligands from the surface of breast cancer cells holds significance for breast cancer biology and treatment in that it challenges the binary rubric that categorically defines breast cancer as either estrogen or growth factor dependent. Experimental Design. GPER was measured by immunohistochemical analysis using a semi-quantitative scoring method in 497 breast tumors harvested at first diagnosis obtained from either the NCI Cooperative Breast Cancer Tissue Resource or an epidemiological population study, the Nashville Breast Health Study (NBHS), including tumor specimens collected from 389 Caucasian and 108 Black patients. GPER was correlated with disease progression variables and evaluated by race, menopausal status and tumor immunophenotype.

Results. GPER is expressed in more than two-thirds of invasive breast carcinomas with no difference observed between Caucasian and Black women (p=0.069). Co-distribution of ER and GPER was measured in approximately half of all tumors (48%). Additionally, GPER is commonly expressed in ER-negative tumors (107/171= 62%) and in triple-negative breast cancer (TNBC) (42/56= 75%) and its expression did not differ by race (p=0.42). Moreover, while ER-positive tumors are less common in premenopausal (74/153= 48%) versus postmenopausal women (244/336= 73%), GPER expression is not influenced by menopausal status. GPER is positively associated with tumor size in patients from either the NCI or NBHS study, and with frank metastasis and tumor stage in the NCI breast cancer collection. However, no significant association is seen with nodal invasion in the NCI breast cancer collection. In contrast, in patients from the epidemiological study, GPER is directly linked to nodal invasion but neither metastasis nor tumor stage. Survival data in NBHS patients found that of the 17 women who died of breast cancer, 16 expressed GPER in their primary tumors, while nearly half of these tumors were ER-positive (8/17). An equal number of non-survivors had tumors which were ER-negative and thus, were judged by current standards to be ineligible for hormonal therapy.

Abstract title: Association of G-protein-coupled estrogen receptor (GPER) in breast cancer in Caucasian and Black women by tumor immunophenotype and menopause status

Authors (in order submitted by poster presenter):Edward Filardo, Dept of Surgery and Holden Comprehensive Cancer Center, University of Iowa, Iowa City, IASandra Deming-Halverson, Department of Medicine, Vanderbilt-Ingram Cancer Center, Nashville, TNCarl Graeber, Department of Medicine, Brown University Medical School, Providence, RIJason Machan, Department of Medicine and Orthopedics, Brown University Medical School, Providence, RIEdmond Sabo, Department of Pathology, Technion Institute, Haifa, IsraelWei Zheng, Department of Medicine, Vanderbilt-Ingram Cancer Center, Nashville, TN

Authors: Micaela G. Fosdick, Alex Wolff, Ronal Peralta, Michael Y. Zhang, Jon C.D. Houtman, PhD

Abstract Title: Suppression of human T cell activation by glycerol monolaurate derivatives

Background: Glycerol Monolaurate is composed of a 12 carbon chain fatty acid with an ester linkage to a glycerol head group. Interestingly, GML inhibits the growth of bacteria, fungi, and enveloped viruses. It is used commercially in several products as a preservative and an anti-microbial agent; however the effect of GML on mammalian cells is not yet fully established. Previous work in this lab has shown GML inhibits T cell signaling by disrupting cell membrane lipid rafts and localization of Arp 2/3 complex subunits. Methods: GML derivatives were tested to determine what components of GML are responsible for its T cell inhibiting activity. These compounds differ from GML in carbon chain length, head group, linker, and position of the laurate group. Activation was measured via LAT clustering, cytokine production, and calcium signaling. Results: Derivatives with a carbon chain less than 12 carbons did not inhibit T cell activation. In addition, the linker and head group polarity were also found to contribute to GML’s ability to inhibit cytokine production, however changing the position of the laurate group did not affect GML’s activity. Conclusion: None of the GML analogs inhibited T cell activity better than GML. However, some compounds that did not disrupt cell membrane organization still inhibited cytokine release. This suggests GML’s effects are more complex than those seen by the lipid raft disruption alone. Overall, these data contribute to the understanding of a novel class of immune modulating lipids and suggest novel therapeutic applications of GML including targeting immune cancers and altering immune cell function during cancer treatment.

THE ROLE OF PROSTAGLANDINS IN COLLECTIVE, INVASIVE CELL MIGRATION Emily Fox1, Tina Tootle1

1Anatomy and Cell Biology Dept., University of Iowa

Collective cell migration – the coordinated movement of tightly or loosely associated cells – is important for both normal development and tumor invasion. While prostaglandins (PGs), short-range lipid signaling molecules, regulate cell migration, and are known to be up regulated in many cancerous tissues, their mechanisms of action are poorly understood in both single and multicellular migration contexts. To address this knowledge gap we use the collective, invasive, epithelial migration that occurs during Drosophila oogenesis. The Drosophila ovary contains chains of developing follicles composed of 15 germline derived nurse cells and 1 oocyte surrounded by a layer of somatic epithelial cells. During Stage 9 of oogenesis, a cluster of 6-8 of these somatic cells delaminate from the outer epithelium and migrate invasively between the nurse cells to the oocyte border; this migration is termed border cell migration. To study the roles of PGs in border cell migration, we utilize genetic mutations in pxt, the Drosophila cyclooxygenase-like enzyme, which is responsible for all PG synthesis. Using quantitative analyses, I find that loss of Pxt causes aberrant border cell migration. Loss of Pxt results in both a significant delay in border cell migration and an increase in cluster length compared to wild-type controls. We hypothesize that both the delay and alteration in cluster morphology are due to changes in among the border cells and/or between the border cells and the surrounding nurse cells. While E-Cadherin appears to be unaffected by the loss of Pxt, integrin levels on the interface between the border cells and the nurse cells is reduced. As integrin-based adhesion is essential for correctly timed border cell migration and cluster cohesion, our data supports the model that PGs regulate integrins to control border cell migration and cluster morphology. Future work will further investigate this model as well as the role of PGs in the border cell cluster vs the nurse cells. Our studies on PG signaling during border cell migration provide insights into the conserved mechanisms by which PGs regulate collective, invasive cell migrations. Indeed, high levels of PGs and integrins are independently associated with cancer migration and metastasis.

Assessing the molecular heterogeneity of cutaneous T cell lymphoma using single cell RNA-seq

Background

Cutaneous T cell lymphomas (CTCL) are a group of hematopoietic malignancies that have increased in incidence over the last 40 years. Sezary syndrome (SS) is a form of CTCL characterized by the presence of malignant cells in the blood and substantial skin involvement. In the setting of worsening disease, many treatments can initially decrease the tumor burden, but ultimately few therapies are able to prevent the outgrowth of a resistant population. The lack of effective therapies is due in part to an incomplete understanding of disease pathogenesis.

Methods

In order to assess for the presence of subpopulations with disparate gene expression profiles, circulating malignant cells (CD3+CD4+CD5hi) and normal CD4 T cells (CD3+CD4+CD5int) cells were flow sorted from a blood sample taken from a patient with SS. Whole genome gene expression libraries and barcode-linked targeted T cell receptor libraries were generated from approximately 3,531 malignant cells and 4,486 normal CD4 T cells. Paired end 150 bp sequencing was performed on the libraries.

Results

Flow sorted malignant and normal cells largely segregated from each other and into separate populations by whole genome gene expression profiles. Malignant cells could be identified in the population selected as normal by flow sorting and vice versa, indicating that gene expression profiling by single cell RNA-seq may be a better discriminator of malignant and normal cells. Subpopulations were identified within the malignant population that differed in their expression of cell proliferation and differentiation factors, such as FOS and JUN, as well as antigen presentation molecules.

Conclusions

SS is made up of populations of cells that may have a spectrum of molecular profiles. Further work is needed to investigate how the degree of heterogeneity may affect the disease course and response to specific treatments.

Authors (as submitted by poster presenter):Ali Jabbari, Department of Dermatology, University of IowaVincent Liu, Department of Dermatology, University of IowaNick Borcherding, Department of Pathology, University of IowaBrian Link, Department of Internal Medicine, University of IowaWeizhou Zhang, Department of Pathology, University of Iowa

CD177 suppresses breast-cancer relapse by regulating the canonical Wnt/b-catenin pathway PaigeKluz,RyanKolb,QingXie,NicholasBorcherding,LinnaWang,YinanZhang,JamesP.DeAndrade,PhilipM.Spanheimer,WeiLi,ChristopherStipp,KatherineN.Gibson-Corley,KimberlyCole,ChenZhao,AndrewBellizzi,AndyW.Tao,SoniaSugg,RonaldJ.Weigel,WeizhouZhang

Background:TheavailabilityoflargeAffymetrixandRNA-sequencingdatasetsmakesitpossibletoidentifygenesinvolvedincancer.Aimingtoidentifyimmunemoleculeswithanovelfunctionincancerpathogenesis,wefoundtheclusterofdifferentiation177(CD177),aknownneutrophilantigen,tobepositivelycorrelatedwithrelapse-free(RFS),metastasis-free(MFS)oroverallsurvival(OS)inseveralsolidcancersincludingthosefrombreast,prostate,cervix,andlung.Focusingonbreastcancer,wefoundthatCD177isexpressedinnormalbreastepithelialcellsandissignificantlyreducedininvasivecancer.WefoundthatCD177suppressesbreastcancerpathogenesisusinginvivomodels.Mechanistically,CD177formsacomplexwithE-CadherinandBeta-Cateninatadherensjunction.ThisphysicalinteractionbetweenCD177,E-CadherinandBeta-CateninpreventsBeta-CateninactivationviathecanonicalWNTsignaling.WethusidentifiedCD177asanovelcancersuppressivemoleculeanduncoveredaregulatorymechanismforcanonicalWNT/Beta-Catenin-mediatedoncogenicsignalingtransduction.

KeyFindings:WeidentifiedanovelproteincomplexinvolvingCD177andproteinsfromadherensjunctions that can suppress cancer formation via inhibiting the WNT/Beta-Catenin signalingpathway.WediscoveredanovelregulatoryarmforadherensjunctionsandforWNT/Beta-Cateninactivation, two key cellular biological processes that share Beta-Catenin and are relevant to theoncogenesisofmultiplecancertypes.CD177couldpotentiallybeusedasapredictionmarkerforrelapseandmetastasisofseveralsolidcancers.

Targeting a Novel RABL6A-RB1 Pathway Suppresses MPNST Pathogenesis

Jordan Kohlmeyer1, Courtney Kaemmer3, Allison Moreno Samayoa2, Chandra Maharjan3, Vickie Knepper-Adrian4, Dave Gordon5, Rebecca Dodd4, Ben Darbro5, Munir Tanas6 and Dawn E. Quelle1,3,6

Molecular Medicine Graduate Program1, Post Baccalaureate Research Education Program2, and Departments of Pharmacology3, Internal Medicine4, Pediatrics5 and Pathology6 at the University of Iowa, Carver College of Medicine and Holden Comprehensive Cancer Center, Iowa City, IA

Malignant peripheral nerve sheath tumors (MPNSTs) are aggressive, deadly sarcomas that arise from the myelinating nerve sheath. These tumors can arise sporadically (50%) or in association with the cancer predisposition syndrome, neurofibromatosis type I (NF1). The only known cure for MPNSTs is complete surgical resection with wide margins; however, many tumors are non-resectable or cannot be fully resected due to their location and/or large size. In most MPNSTs, the retinoblastoma (RB1) tumor suppressor pathway is inactivated through hyper-activation of CDK4/6 kinases that disable RB1, often through loss of cell cycle inhibitors such as p16 and p27.

RABL6A is an oncogenic GTPase and newly recognized inhibitor of the RB1 pathway whose role in MPNST biology is not known. We examined RABL6A protein levels in human MPNST lines and found it is upregulated compared to non-transformed Schwann cells (NHSC), supporting our hypothesis that RABL6A drives MPNST pathogenesis through inactivation of RB1, and this pathway represents an important, new therapeutic target. Tissue microarray analyses show marked upregulation of RABL6A coincident with p27 loss in human MPNSTs compared to patient-matched PNFs. In vitro assays demonstrate RABL6A is essential for MPNST proliferation and survival. Loss of RABL6A caused MPNST cell death and G1 phase arrest that coincided with p27 upregulation and RB1 hypo-phosphorylation. These data suggested MPNSTs will be inhibited by drugs targeting the RB1 pathway. Indeed, the selective CDK4/6 inhibitor, palbociclib (PD0332991), kills MPNST cells in an RABL6A-dependent manner. Our findings establish a critical role for RABL6A in MPNST pathogenesis and identify RABL6A-RB1 signaling as a novel target for MPNST therapy.

Title: Obesity-associated inflammation promotes angiogenesis via angiopoietin-like 4

Authors: Ryan Kolb1,2, Paige Kluz1,3, Zhen Wei Tan4, Nicholas Borcherding5,6, Nicholas Bormann7,

Louis Balcziak8, Pengcheng Zhu4, Brandon SJ. Davies9, Francois Gourranc10, Katherine Gibson-Corley1,

Aloysius Klingelhutz10, Daniel J. Kelpsch11, Tina L. Tootle11, Khaled Elmasry12,13 Ahmed S. Ibrahim14,15,

Mohamed Al-Shabrawey14, Nguan Soon Tan4,16,17,18, Fayyaz S. Sutterwala19, Weizhou Zhang1,3,5,6,20, *.

Obesity is associated with an increased risk of developing ER-positive breast cancer in postmenopausal

woman and also predicts poor clinical outcomes regardless of menopausal status. We recently found that

obesity leads to an increase in tumor-infiltrating macrophages with activated NLRC4 inflammasome and

increased interleukin (IL)-1β production. IL-1β, in turn, leads to increased angiogenesis and cancer

progression. Using next generation RNA sequencing, we identified an NLRC4/IL-1β-dependent

upregulation of angiopoietin-like 4 (ANGPTL4), a known angiogenic factor in cancer, in tumors from

obese mice. ANGPTL4-deficiency, either by genetic knockout or antibody neutralization, led to a

significant reduction in obesity-induced angiogenesis and tumor growth. At a mechanistic level, IL-1β

induces ANGPTL4 expression mainly from primary adipocytes in a manner dependent on NF-κB- and

MAP kinase-activation, which is in synergy with hypoxia and macrophage-produced 15(S)-

hydroxyeicosatetraenoic acid (15(S)-HETE). This report shows that adipocyte-derived ANGPTL4 drives

disease progression under obese conditions and is a potential therapeutic target for treating obese breast

cancer patients. Author Affiliations (as submitted by poster presenter):Ryan Kolb, University of Iowa, Department of PathologyPaige Kluz, University of IowaZhen Wei Tan, School of Biological Sciences Nanyang Technological UniversityNicholas Borcherding, University of IowaNicholas Bormann, University of IowaLouis Balcziak, University of IowaBrandon SJ Davies, University of IowaFrancois Gourranc, University of IowaKatherine Gibson-Corley, University of IowaAloysius K Lingelhutz, University of IowaDaniel J Kelpsch, University of IowaTina L Tootle, University of IowaKhaled Elmasry, University of IowaNguan Soon Tan, School of Biological Sciences Nanyang Technological UniversityFayyaz S Sutterwala, Cedars-Sinai Medical CenterWeizhou Zhang, University of Iowa

PHF8 promotes resistance to anti-HER2 therapies in breast cancer cells

through IL-6

Authors: Qi Liu1, Nicholas Borcherding2, Peng Shao3, Weizhou Zhang4 and Hank Heng Qi5

1 Ph.D. Department of Anatomy and Cell Biology, Carver College of Medicine, University of Iowa, Iowa City, IA, 52242; Email: qi-liu-2@uiowa.edu; Tel: 217-384-8461 2 M.D., Ph.D. Department of Pathology, Carver College of Medicine, University of Iowa, Iowa City, IA, 52242; 3 Ph.D. Department of Microbiology, Carver College of Medicine, University of Iowa, Iowa City, IA, 52242; 4 Ph.D. Department of Pathology, Carver College of Medicine, University of Iowa, Iowa City, IA, 52242, 5 M.D., Ph.D. Department of Anatomy and Cell Biology, Carver College of Medicine, University of Iowa, Iowa City, IA, 52242.

Background

HER2 plays critical roles in tumorigenesis and is associated with poor prognosis of breast cancers. Although some tumors respond well to anti-HER2 therapies, e.g. trastuzumab and lapatinib, many are resistant de novo or following therapy. Epigenetic mechanisms influencing HER2-driven cancers and drug resistance are largely unknown.

Methods and Results

We recently reported that histone demethylase PHF8 is overexpressed in is essential for growth of HER2-positive (HER2+) breast cancer cells. To further characterize the underling mechanisms, we performed RNA sequencing in four double stable MCF10A cell lines which overexpress HER2 combined with inducible PHF8 knockdown. Importantly, PHF8 knockdown attenuated the upregulation of cell cycle genes, epithelial to mesenchymal transition (EMT) markers and cytokine genes including IL-6 and IL-8 induced by HER2. Furthermore, we confirmed that secreted IL-6 is decreased by PHF8 knockdown. ChIP-qPCR proved that PHF8 is enriched on the promoters of IL-6, and its occupancy significantly increased by HER2 overexpression. Moreover, in primary tumor cells from MMTV- ErbB2/MMTV-Cre/Phf8flox/flox mouse model, we found Il-6 expression is significantly decreased by Phf8 knockout. Thus, these data indicates PHF8 regulates IL-6 in HER2 signaling.

IL-6 is increased in HER2-overexpressing cancer stem cells, contributing to trastuzumab resistance. We found that the expression of both IL-6 and PHF8 are upregulated in lapatinib-resistant SKBR3 and BT474 cells (-R cells). PHF8 knockdown in BT474-R cells decreased IL-6 mRNA level. Moreover, we also revealed that PHF8 regulates IL-6 in HER2+ HCC1954 cells, which possess higher endogenous IL-6 than BT474 and SKBR3 and are resistant to trastuzumab de novo. Consequentially, stem cell markers NANOG and OCT4 were decreased by PHF8 knockdown in these contexts. Moreover, overexpressing wtPHF8 significantly antagonized the growth inhibition and apoptosis in SKBR3 cells with treatment of anti-HER2 therapies.

Conclusions

In sum, upregulated PHF8 in HER2+ breast cancers may antagonize the efficiency of anti-HER2 therapies by increasing IL-6 expression. This study gains mechanistic insights into the application of PHF8 inhibitors in the resistance of anti-HER2 therapies.

The TAZ-CAMTA1 and YAP-TFE3 fusion proteins transform cells by binding to members of the histone acetyltransferase Ada2a-

containing complex (ATAC)

Nicole Merritt1, Dushyandi Rajendran2, Zhen-Yuan Lin2, Xiaomeng Zhang3, Katrina Mitchell3, Colleen Fullenkamp1, Anne-Claude Gingras2, Kieran Harvey3, Munir Tanas1

1Department of Pathology, University of Iowa, Iowa City, IA, 2Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, ON, 3Peter MacCallum Cancer Centre, Melbourne, VIC.

Background: TAZ (WWTR1) and YAP are oncogenic transcriptional coactivators negatively regulated by the Hippo pathway. Constitutive activation of TAZ and YAP has been observed in many different types of cancer including most sarcomas. Epithelioid hemangioendothelioma is a vascular sarcoma characterized by WWTR1-CAMTA1 (90%) and YAP1-TFE3 gene fusions (10%). The fusion proteins contain the N-terminus of TAZ and YAP fused in frame to the C-termini of CAMTA1 and TFE3, respectively.

Methods: NIH 3T3 and SW872 cells lines were transduced with the gene fusions. In vitro assays were used to assess hallmarks of cancer. Protein-protein interactions were identified using BioID mass spectrometry and a shRNA screen was conducted to determine proteins crucial to driving anchorage-independent growth.

Results: We have shown that TAZ-CAMTA1 (TC) and YAP-TFE3 (YT) promote anchorage-independent growth as well as tumorigenesis and metastasis in vivo. BioID mass spectrometry identified 41 novel interactions mediated by the fusion proteins, 26 of which are epigenetic modifying enzymes. Of these 26 proteins, YEATS2 was identified by shRNA screen as critical to TC driven anchorage independent growth. Mass spectrometry data showed that three subunits (YEATS2, ZZZ3, and KAT14) of the histone acetyltransferase complex, Ada2a-containing complex (ATAC), interacted with both the TC and YT fusion proteins. Kaplan-Meier analysis of the TCGA Illumina HiSeq RNA-Seq data set for sarcomas demonstrates that increased expression of YEATS2 or ZZZ3 predict worse overall survival (p=0.0049 and p=0.0037 respectively) across all sarcomas, independent of histological type.

Conclusions: The data suggest that oncogenic properties of TC and YT can be explained in part by their interaction with key epigenetic modifiers that potentiate their oncogenic transcriptional programs. Future studies will elucidate the mechanisms by which the ATAC complex augments the TAZ and YAP oncogenic transcriptional programs.

Radiologic response of colorectal ovarian metastasis to neoadjuvant chemotherapy

Hannah L. Phillips, Braden S. Jensen, Mary Belding-Schmitt , Carlos H.F. Chan, Department of Surgery,

University of Iowa Hospitals and Clinics, Iowa City, IA

Background:

Ovarian masses are often assumed to be ovarian cancers and commonly treated with upfront surgery,

even in the presence of peritoneal metastasis. However, some cases are ovarian metastases of

colorectal origin and upfront incomplete cytoreductive surgery may have negative impact on the

oncological outcome. Neoadjuvant chemotherapy followed by complete cytoreductive surgery and

hyperthermic intraperitoneal chemotherapy may be a better approach. The aim of our study is to

determine if ovarian metastasis initially treated with chemotherapy exhibit radiologic response.

Methods:

A retrospective review of patients treated at the University of Iowa for stage IV colorectal cancer with

ovarian metastasis between the years 2006-2016 was conducted using the cancer registry data from the

Holden Comprehensive Cancer Center. Patients receiving neoadjuvant chemotherapy (FOLFOX +/-

Avastin) with CT scans prior and within 3 months of starting treatment were identified. Imaging was

reviewed using the revised RECIST guidelines for gauging response to therapy of target lesions.

Results:

Thirty-two patients with ovarian metastasis were identified with only 9 patients (28%) receiving

neoadjuvant chemotherapy. Twenty-five patients underwent surgical resection without neoadjuvant

chemotherapy, 13 (57%) with the incorrect presumptive diagnosis of ovarian cancer. Of the nine

patients who underwent neoadjuvant chemotherapy, 5/9 (56%) demonstrated reduction in size of the

ovarian metastasis or stable lesions. Four of the nine (44%) patients studied had interval increase in size

of ovarian lesions.

Conclusions:

Given over 50% of patients studied had a radiologic response to neoadjuvant chemotherapy and 57% of

patients undergoing resection were incorrectly diagnosed, we argue patients with ovarian metastasis

should be properly identified and undergo neoadjuvant chemotherapy in an attempt to decrease tumor

burden prior to definitive cytoreductive surgery.

RationallyimprovingTcell-mediatedcancerimmunotherapyusingSleepingBeautymutagenesistoidentifynoveldrugtargetsLauraM.Rogers1,AdamJ.Dupuy1,2,andGeorgeJ.Weiner1,31CancerCenterandDepartmentsof2AnatomyandCellBiologyand3InternalMedicine,UniversityofIowa,IowaCity

Tcellscaneliminatetumorcellsthroughoutthebody,asevidencedbytheclinicalsuccessofimmunotherapiesdesignedtoenhanceTcellfunction,includingimmunecheckpointblockadeandchimericantigenreceptorTcell(CAR-T)therapies.Expandingsuccesstoabroadernumberofpatientsisatoppriorityinthefield.TcellinfiltrationintotumorsappearstobeanimportantprerequisiteforthesuccessofbothimmunecheckpointblockadeandCAR-Ttherapies.Thus,increasingTcellinfiltrationintotumorscouldhavebeneficialtherapeuticimpact.

WehavedesignedaforwardgeneticscreentoidentifyTcellgenesthatcontributetointratumoralTcellaccumulationusingSleepingBeautymutagenesisinTcells.Oursystematicscreenapproachallowsustoevaluatebothloss-andgain-of-functionmutationsacrosstheentiregenomeinimmunocompetentmelanomaandlymphomamodelsthatpreservethecomplexityofthetumormicroenvironment.Weidentified312genes,20ofwhichweredetectedinmorethanonemouse,representingstronggenecandidatesforvalidation.

Importantly,thevastmajorityofgenesweidentifiedwerenotpreviouslyknowntobeinvolvedintumor-associatedTcellbiology,andlikelywouldnothavebeenconsideredasdrugtargetstoenhanceimmunotherapy.ThesegeneshavethepotentialtomodifyimportantTcellfunctionsincludingtraffickingtothetumor,clonalexpansion,andsustainedviabilityonceinsidethetumor.WedemonstratethatoneofthesegenecandidatesisfunctionallyassociatedwithTcellcytokineproductioninvitro.

AdditionalscreensusingtheEL4lymphomamodel,andalsoincludinganti-PD-1therapyhavebeenperformed.Preliminarydatafromtheseindicatethatfewgenesareconservedacrosstumormodelsandonlyonegenewasconservedacrossallscreencohorts.WearecurrentlyinvestigatingtheroleofthisgeneinintratumoralTcellaccumulationandimmunotherapyenhancementusinganinvivoapproach.

IdentificationandcharacterizationoftheRhofamilyasadriverofdrugresistanceinBRAFV600

mutantmelanoma.

JacobSchillo,CharlotteFeddersen,HayleyVaughn,AndrewVoigt,EliotZhu,LexyWadsworth,ChristopherStipp,andAdamDupuy

Theserine/threonineproteinkinaseBRAFisfrequentlymutatedincutaneousmelanoma.Approximately60%patientsharboraBRAFmutation(~40%V600E).Thesemutationsareselectivelytargetedbythekinaseinhibitorvemurafenib.Althoughinitialresponseisencouraging,themajorityofpatientsdevelopdrugresistancewithinashorttimeframe.Whiledrugresistancecanbedelayedbycombiningvemurafenibwithcobimetinib,aMEKinhibitor,mechanismsofresistancetothedrugcombinationortovemurafenibalonearenotwellcharacterized.Therefore,weperformedagain-of-functionscreenutilizingtheSleepingBeautytransposonsystemtoidentifynoveldriversofvemurafenibandcombinationtherapyresistanceinBRAFV600mutantmelanoma.Ourscreenidentifiedfourcandidatedriversofdrugresistancethathavebeenvalidatedinmultiplemelanomacelllinesbyassessingcellgrowthandviabilityinvaryingconcentrationsofvemurafenib.

OurscreenidentifiedtwomembersoftheDBLfamilyofguaninenucleotideexchangefactors(GEFs),VAV1andatruncatedformofMCF2.FunctionaltestsofVAV1andMCF2identifiedtheRhofamilymembersRac1andCDC42ashavingincreasedsignalingwithinthehumanA375BRAFV600Emutantmelanomacelllinefollowingtreatmentwithvemurafenib.Inaddition,wedeterminedtheabilityofeachcandidateGEFtoimpactMAPKandPI3K/AKTsignaling,knownmediatorsofvemurafenibresistance.Finally,weevaluatedapanelofBRAFV600mutantmelanomacelllines,includingaseriesofnovelcelllinesderivedfrompatient-derivedxenografts,todeterminethebroaderroleofthecandidateDBLfamilymembersindrivingvemurafenibresistance.

TheRhofamilyisactivateddownstreamofavarietyofextracellularsignalingpathwayswhichareknowntodriveincreasedvemurafenibresistance.Therefore,understandingtheroleoftheDBLfamilyinmediatingdrugresistancemayprovidenoveltargetsforfuturecombinatorialtherapiestoimprovepatientsurvival.

*All authors affiliated with University of Iowa

Transcription factor TFAP2A and its inhibitor KCTD15 regulate the cellular phenotype in melanocytes and melanoma

Hannah Seberg PhD1, Gregory Bonde1, Mikaela Mallin1, Robert Cornell PhD1

1Department of Anatomy and Cell Biology, University of Iowa

Background: Melanoma tumor cells can alternate between a differentiated/proliferative phenotype and an invasive, drug-resistant phenotype. Microphthalmia-associated transcription factor (MITF) may contribute to this switch, as high levels of MITF activity promote proliferation and differentiation, while lower levels promote invasiveness. During embryonic development, the transcription factor TFAP2A collaborates with MITF to directly drive expression of melanocyte differentiation genes. The Cancer Genome Atlas shows that TFAP2A expression is frequently decreased in advanced melanomas, suggesting it may suppress the invasive phenotype. Interestingly, TFAP2A promotes expression of KCTD15, a potent inhibitor of TFAP2A activity, resulting in a negative feedback loop. We hypothesized that overexpressing tfap2a in the melanocyte lineage of melanoma-prone zebrafish would delay tumor formation, while overexpressing kctd15a would accelerate it.

Methods: We cloned tfap2a, kctd15a, or control mCherry cDNAs into the cargo position of a dual cassette vector in which mitfa promoters separately drive expression of mitfa and of a cargo gene. Upon injection into mitfa-/- zebrafish embryos, which otherwise lack melanocytes, this construct rescues melanocytes that all express the cargo gene. We injected these constructs into melanoma-prone mitfa-/- mutants, raised injected animals, and monitored the appearance of tumors.

Results: As expected, compared to mitfa-promoter driven expression of mCherry in melanoma-prone zebrafish, expression of tfap2a led to significantly longer tumor-free survival. Unexpectedly, in preliminary findings, expression of kctd15a also significantly prolonged tumor-free survival compared to controls. Also unexpectedly, kctd15a-overexpressors had significantly more melanocytes as adults than did controls or tfap2a-overexpressors.

Conclusions: These results indicate that the observed loss of TFAP2A in advanced melanoma contributes to its pathology. The unexpected consequences of overexpressing a potent inhibitor of all TFAP2 paralogs suggests an opposing function of other TFAP2 paralogs or other KCTD15-sensitive transcription factors.

A novel RABL6A-PP2A-AKT pathway drives pancreatic neuroendocrine tumor growth

Umesalma Shaikamjad1, Courtney A. Kaemmer1, Blake Letney1, Jordan L. Kohlmeyer2, Jacqueline A. Reilly 1, Angela M. Schab1, Ryan M. Sheehy1,3, Jussara Hagen1, Nitija Tiwari1, Fenghuang Zhan4, Thomas M. O’Dorisio4, James R. Howe5, Andrew M. Bellizzi6, Abbey Perl7, Stefan Strack1, Goutham Narla7, Benjamin W. Darbro8, Frederick W. Quelle1,4, and Dawn E.Quelle1,2,3,6,#

The Department of Pharmacology1, Molecular Medicine Graduate Program2, Free Radical & Radiation Biology Training Program3 and the Departments of Internal Medicine4, Surgery5, Pathology6, and Pediatrics8 in the College of Medicine, Holden Comprehensive Cancer Center, University of Iowa, Iowa City, Iowa. Case Western Reserve University7, Cleveland, Ohio.

Hyperactivated AKT/mTOR signaling is a hallmark of pancreatic neuroendocrine tumors

(PNETs). Drugs targeting the pathway are used clinically but tumor resistance invariably

develops. A better understanding of factors controlling AKT/mTOR signaling and PNET

pathogenesis is needed to improve current therapies. We discovered that RABL6A, a new

oncogenic driver of PNET proliferation, is required for AKT activity. Silencing RABL6A caused

PNET cell cycle arrest that coincided with selective loss of AKT-S473 (not T308)

phosphorylation and AKT/mTOR inactivation. Restoration of AKT phosphorylation rescued the

G1 phase block triggered by RABL6A silencing. Mechanistically, loss of AKT-S473

phosphorylation in RABL6A depleted cells resulted from increased protein phosphatase 2A

(PP2A) activity. Inhibition of PP2A restored phosphorylation of AKT-S473 in RABL6A depleted

cells whereas PP2A reactivation in control cells using a specific small molecule activator of

PP2A (SMAP) abolished that phosphorylation. Importantly, SMAP treatment effectively killed

PNET cells in a RABL6A-dependent manner and suppressed PNET growth in vivo. This work

identifies RABL6A as a new inhibitor of the PP2A tumor suppressor whose expression is

required for AKT signaling in PNET cells. Our findings offer novel targets, PP2A and RABL6A,

for PNET therapy.

Abl family kinases suppress the malignant phenotype of metastatic castrate-resistant prostate cancer via inhibition of AKT signaling pathways

Introduction and Objective: Our recent studies have revealed a suppressor function for Abl kinases in pre-clinical models of metastatic castrate-resistant prostate cancer (mCRPC), but the molecular mechanism was unknown. The aim of this study was to identify the signaling pathways that account for the aggressive behavior of Abl kinase-deficient mCRPC. Materials and Methods: We used a combination of genetic and pharmacological approaches to manipulate Abl kinase activity in a pre-clinical model of mCRPC and determined the impact on malignant behavior, including tumor cell motility and growth in 3D matrix. A reverse phase protein microarray functional proteomics approach was used to identify signaling pathways altered in Abl-deficient cells. Results: Abl kinase-deficient mCRPC cells displayed dramatically increased cell motility and growth in soft 3D extracellular matrix. Stable depletion of both Abl kinase isoforms by RNAi produced the strongest effect, indicating that Abl1 and Abl2 cooperate to limit malignant behavior. Functional proteomic analysis of over 300 signaling proteins revealed that Abl kinase-deficient mCRPC growing out in 3D matrix showed increased activation of AKT kinase and downstream signaling pathways compared to control cells. Key results were independently confirmed by immunoblotting. MK-2206, an AKT inhibitor under clinical development, abolished the increased 3D growth of Abl deficient cells. Conclusion: Abl kinases can have paradoxical tumor suppressor functions in mCRPC linked to their ability to limit survival signaling through the AKT pathway. Abl kinase expression status should be considered in any mCRPC clinical trial involving the use of multi-tyrosine kinase inhibitors that cross inhibit the Abl kinase family.

Authors (as submitted by poster presenter):Christopher Stipp, PhD/BiologyMelissa Marchal/BiologyAfshin Varzavand/BiologyJames Brown, MD/UrologyYousef Zakharia, MD/Internal MedicineMichael Henry, PhD/Molecular Physiology & Biophysics

Contact: Presenter: Sean Tompkins, B.S.; sean-tompkins@uiowa.edu; 630-697-1763

Authors: Ryan D Sheldon, PhD; Adam J Rauckhorst, PhD; Katherine N. Gibson-Corley, DVM, PhD; Adam J Dupuy, PhD; Douglas J Spitz, PhD; Eric B Taylor, PhD

Title: Loss of mitochondrial pyruvate carrier activity short circuits hepatocellular tumorigenesis

Background: HCCs exhibit a profound difference in metabolism compared to normal hepatocytes, the loss of glucneogenesis. The Mitochondrial Pyruvate Carrier (MPC) gates gluconeogenesis by importing pyruvate into the mitochondrial matrix. Yet, data from the cancer genome atlas indicate that across numerous cancers, MPC transcript expression is greatest in HCC. We hypothesized that MPC function is repurposed in HCC to support tumor metabolism.

Methods: WT and MPC liver-specific knockout mice (LivKO) were injected with a priming dose of N-nitrosodiethylamine at age 15 days and then twice weekly with low dose carbon tetrachloride from age 8 to 23 weeks. We analyzed mouse livers and cell culture correlates to understand how MPC disruption decreases tumorigenesis.

Results: Upon euthanasia at age 24 weeks tumor number was markedly lower in MPC LivKO vs WT mice. We compared tumors and adjacent normal tissue by RNAseq and observed systematic differences in transcripts for glutathione metabolism between WT and MPC LivKO tumors. Enzymatic tests for glutathione content revealed that WT but not MPC LivKO tumors significantly increased total glutathione content relative to adjacent normal tissue. Furthermore, relative reduced vs oxidized glutathione content was greater in WT vs MPC LivKO tumors. In cultured mouse hepatoma Hepa1-6 cells, simultaneously impairing glutathione synthesis and disrupting MPC activity synthetically decreased proliferation. Synthetic lethality was rescued by N-acetylcysteine supplementation. In primary mouse hepatocytes, MPC disruption impaired utilization of glutamine to regenerate glutathione content during acute chemical depletion. We propose that HCCs utilize mitochondrial pyruvate import to sustain essential TCA-cycle activity, thereby sparing glutamine for glutathione synthesis. By this model, MPC disruption results in adaptive mitochondrial glutamine utilization, decreased glutathione synthetic capacity, and increased vulnerability to stress, thereby impairing HCC initiation and progression. Thus, disrupting MPC activity breaks a metabolic circuit that may be common in HCCs and amenable to therapeutic targeting.

Oncogenic SnoRNA ACA11 activated by t(4;14), Cooperates with Rb1 deficiency to drive B Cell Proliferation

Chakrapani Tripathi1,3, Vanessa A. de Oliveira1,3, Hongwei Xu1,3, Melissa L. Bates2,3, , Michael H. Tomasson1,3

1Department of Internal Medicine, Hematology Oncology Division, 2Department of Health and Human Physiology and the 3Holden Comprehensive Cancer Center, University of Iowa, Iowa City IA

The candidate oncogene WHSC1 (MMSET) at 4p16.3 encodes a histone methyltransferase over-expressed as a result of the t(4;14) translocation in MM. Mutations that cooperate with t(4;14) in myeloma initiation are unknown, but chromosome 13 deletions (Del(13)) are highly correlated with the t(4;14) in MM. Del(13) is found in 85-94% of patients with t(4;14) and is associated with poor prognosis. Our lab revealed that non-coding snoRNA ACA11 is activated by the t(4;14) translocation and induces cell proliferation by unleashing reactive oxygen species (ROS). On the basis of that finding we hypothesized that additional mutations are certainly required in addition to ACA11 for MM development. To test the hypothesis that ACA11 overexpression cooperates with Rb1 deletion promotes B cell proliferation via excessive ROS production and apoptosis inhibition. We transfected the Rb1 KO (germinal center B-cell) and WT splenic naïve B cells (CD43 negative cells) with ecotropic retrovirus containing ACA11 and MMSET II gene sequence i.e MSCV-ACA11 ires-Td-tomato, MSCV-WHSC1-ires-GFP and empty vector controls (GFP).We observed the B cell proliferation by cell count via trypan blue exclusion for 5 days and apoptosis inhibition via down regulation of apoptotic protein caspases expression. In order to develop targeted therapies in MM, we need to find out the pathways that are activated by recurrent driver mutation like t(4;14). Our data show that ACA11 overexpression cooperates with Rb1 deletion and promotes B cell proliferation via ROS upregulation and apoptosis inhibition. We would like to further explore the mechanism behind ACA11 and/or MMSET and RB1 gene association in MM disease pathogenesis.

Dissectingtumor-infiltratingregulatoryT-cellheterogeneityusingsinglecell

RNA-sequencingandroleofCD177Authors:AjaykumarVishwakarma,KawtherK.Ahmed,NicholasBorcherding,RyanKolb,PaigeKluz,GauravPandey,MichaelJorgensen,KimberlyCole,SoniaSugg,AndrewBellizzi,MunirTanas,DeqinMa,KatherineN.Gibson-Corley,JuliaKlesney-Tait,YuwenZhu,WeizhouZhang

UnderstandingphenotypicandfunctionalheterogeneityofregulatoryT-cell(Tregs)inthetumor

microenvironmentatasinglecelllevelisessentialtoidentifynovelmarkersofimmune

suppressionandimmunologicalmechanismsplayingroleincancerprogression.Analysisofbulk

RNA-sequencingdatasetsfromseveraltypesofcanceridentifiedCD177asatopdifferentially

regulatedgeneintumor-infiltratingregulatoryT-cellsincomparisontopatient-derivedperipheral

bloodTregs.Tofurthercharacterizetumor-infiltrating,weperformedsingle-cellRNA-sequencing

(ScRNA-seq)ofTregsandconventionalCD4Tcells(Tconv)isolatedfromfreshsurgicalbreast

cancerspecimens.InaccordancewiththebulkRNA-seq,weconfirmedexpressionofintra-tumoral

CD177inasubsetoftumor-infiltratingTregsbutnotinTconv.ComparingCD177+TregstoCD177-

Tregs,weidentifiedseveralimmunesuppressivemarkersandchemokinereceptorsthatwere

significantlyupregulated.WeverifiedCD177surfaceproteinexpressionintumor-infiltrating

lymphocytes(TILs)indifferentcancertypesbyflowcytometryandimmunohistochemistry.CD177

wasexpressedonasubsetofTregsanditsexpressionwasassociatedwithincreasedexpressionof

immunecheckpointinhibitorsPD1,CTLA4andactivationmarkerCD27andCCR8.Functional

characterizationofCD177positiveversusnegativeTregsobtainedfrombothCD177deficientmice

andhumantumorsamplesrevealedCD177+Tregsweremoreimmune-suppressivethantheir

negativecounterparts.TheeffectsofhostCD177ontumorgrowthwasexaminedinasyngeneic

orthotopicallyimplantedbreastcancermodelandlossofCD177ledtoreducedbreastcancer

tumorgrowthonlywhenalimitednumberofcellswereimplanted(500)butnotathighernumbers

(>1000).WehavepreviouslyidentifiedabreastcancercellintrinsicfunctionofCD177inregulating

WNT/b-cateninsignaling(Unpublished).Similarly,weobservedthattheoverexpressionofCD177

inaJurkatTcelllinereducesWNT3amediatedb-cateninactivitysuggestingitsroleinmodulating

Tregfunction.OverallourdatasuggeststhatCD177mayprovideamarkerforamoresuppressive

tumor-associatedTregpopulation.

Keywords:RegulatoryTcells,Single-cellRNA-seq,CD177,Cancer

TRAF3 regulates Pim2- and Myc-mediated B cell survival

Amy Whillock1,2,3, Nurbek Mambetsariev1,2,3, Wai Lin1,2, Laura Stunz1,5, Gail Bishop1,2,4,5,6

University of Iowa Carver College of Medicine 1Department of Microbiology, 2Interdisciplinary Graduate Program in Immunology, 3Medical Scientist Training Program, 4Department of Internal

Medicine, 5Holden Comprehensive Cancer Center, 6VA Medical Center

Background Tumor necrosis factor receptor associated factor 3 (TRAF3) is an adaptor protein that plays an important role in B cell homeostasis. TRAF3 deletions or inactivating mutations are common in human B cell malignancies. In human and mouse B cells, TRAF3 negatively regulates signaling of several receptors important for B cell homeostasis (including BAFFR, CD40, and IL-6R). B cell specific TRAF3-deficient (B-Traf3-/-) mice have enlarged spleens and lymph nodes and an increased frequency of B cell tumors as they age. B cells from B-Traf3-/- mice have markedly increased survival and autoantibody production. The mechanisms by which TRAF3 regulates B cell survival are not yet well defined. Through a microarray, we identified the pro-survival serine/threonine kinase Pim2 as being significantly upregulated in TRAF3-deficient B cells. We hypothesized that TRAF3 regulates B cell survival through suppression of Pim2 and its targets.

Results Confirming what we saw in the microarray, TRAF3-deficient mouse B cells and human malignant B cell lines with low expression of TRAF3 had increased Pim2 mRNA and protein which was independent of the non-canonical NF-κB pathway. Additionally, TRAF3-deficient mouse B cells had increased phosphorylation of Pim2 targets (BAD, p70S6K, and 4E-BP1). TRAF3-deficient B cells showed increased survival relative to TRAF3-sufficient B cells after treatment with the Pim inhibitors SGI-1776 and TP-3654. Pim2 can promote survival by phosphorylating and stabilizing the oncogenic transcription factor c-Myc. Loss of TRAF3 led to transcription-independent c-Myc protein elevation that was dependent on Pim2. K48 polyubiquitination of c-Myc was decreased in the absence of TRAF3, consistent with increased c-myc protein stability. TRAF3 deficiency correlated with B cell resistance to apoptosis mediated by the c-Myc transcription inhibitor JQ1.

Conclusions Our results show that TRAF3 promotes B cell survival by regulating expression of and interaction between Pim2 and c-Myc. Our results identify a novel pathway that promotes survival in the absence of TRAF3 and have important roles in determining responsiveness to cytotoxic agents. This information can be used for therapeutic strategies targeting TRAF3 deficient B cell malignancies.

Epithelial-to-mesenchymal transition conveys resistance to restrictive growth

conditions through increased autophagic flux

Tumor progression requires cancer cells to adapt to varying microenvironments both in the

primary tumor and in metastatic end organs. Metastasis is enhanced by epithelial-to-

mesenchymal transition (EMT), which facilitates invasiveness and confers resistance to local

stresses, such as hypoxia and nutrient limitation, although EMT is not necessarily required for

these processes. Among these stressors, growth-factor (GF) deprivation may induce or select for

cells that bear the hallmarks of EMT (EMT-like cells). The mechanisms by which EMT-like

cells survive GF deprivation (restrictive growth conditions), which may occur when metastatic

cancer cells are removed from GFs resident in the primary tumor microenvironment, remain

largely uncertain. Herein we show that EMT confers upon cells the ability to maintain

proliferative potential, following extended serum deprivation. We found that EMT-like prostate

cancer cells (TEM 4-18) demonstrate elevated survival and reduced senescence compared to

their more epithelial counterparts (PC-3E) in both short- and long-term exposure to serum

withdrawal. In addition, resistance to anchorage-dependent growth-induced apoptosis (Anoikis)

due to elevated autophagy is also evident in EMT-like cells. Forced expression of snail1

conferred survival advantages to PC-3 cells. Conversely, CRISPR-mediated knockout of Zeb1

reversed EMT status in TEM4-18 cells and survival advantage in restrictive growth conditions.

We found that EMT-like cells exhibit elevated autophagic flux. Genetic interference with

autophagy in EMT-like cells reversed their survival advantage and increased senescence in

restrictive growth conditions as well as anoikis resistance. Our results suggest that the metastatic

capabilities of EMT-like cells depend not only on their established invasive characteristics, but

also on increased autophagy tied to the EMT-like state, and its role in surviving restrictive

growth conditions.

Authors (as submitted by poster presenter):Lei Zhao; Jones T. Nauseef; Marisa R. Buchakjian; J. Matthew Barnes; Michael D. HenryAll authors: Department of Molecular Physiology and Biophysics, University of Iowa Carver College of Medicine

TRAF3-mediated regulation of the T cell receptor complex

Tina Arkee1, 2

, Alicia Wallis2, and Gail A. Bishop

1, 2, 3, 4

1University of Iowa Medical Scientist Training Program,

2Interdisciplinary Graduate Program in

Immunology, 3Department of Microbiology,

4 VA Medical Center, Iowa City IA

TRAF3 is an adaptor protein with diverse cell type-specific functions that plays an important role

in T cell biology. T cell conditional TRAF3-/-

mice, despite normal numbers of T cells, have poor

responses to immunization and infection. Further investigation revealed that early events in T

cell receptor (TCR) signaling are defective in TRAF3-deficient T cells. Examining the basis of

this defect, we recently reported that TRAF3 normally inhibits localization of the negative

regulators Csk and PTPN22 to the TCR membrane complex. We now wish to understand how

TRAF3 is recruited to this complex. TRAF3 associates with the TCR-CD28 complex upon

stimulation; engagement of both the TCR and CD28 is required for TRAF3 recruitment. We

hypothesize that TRAF3 associates with the TCR complex either indirectly, by binding to other

CD28-associated adaptor proteins, or directly, by binding to motifs in the cytoplasmic tail of

CD28. Results of preliminary experiments suggest that the TRAF3 TRAF-C domain associates

with Linker of Activated T cells (LAT), but not with the LAT-binding adaptor protein Gads. We

are currently investigating the structural requirements for the association of TRAF3 with CD28,

the adaptor protein Grb2, and other TCR signaling proteins, to determine how TRAF3 is

recruited to the TCR complex. Based upon our previous finding that TRAF3 enhances TCR

signaling by inhibiting the localization of negative regulatory proteins associated with the TCR,

as well as our preliminary data indicating that TRAF3 interacts with members of the LAT

complex, we are also investigating the interactions between TRAF3 and negative regulatory

proteins associated with the LAT complex. TRAF3 has been suggested as important for the

growth of certain types of human T cell lymphoma, and T cells play an important role in

elimination of tumor cells, so the information to be gained from this project has implications for

pathogenesis and treatment of these malignancies.

Matrix metalloproteinases and immunosuppressive biomarker profiles of 7 HNSCC cell lines

Amber M. Bates1, Maria Paula Gomez Hernandez1, Emily A. Lanzel2, Fang Qian1,3, and Kim A. Brogden1

1Iowa Institute for Oral Health Research, College of Dentistry, The University of Iowa, Iowa City, IA 52242, USA, amber-bates@uiowa.edu; paula-gomez@uiowa.edu; kim-brogden@uiowa.edu 2Department of Oral Pathology, Radiology and Medicine, College of Dentistry, The University of Iowa, Iowa City, IA 52242, USA, emily-lanzel@uiowa.edu 3Division of Biostatistics and Research Design, College of Dentistry, The University of Iowa, Iowa City, IA 52242, USA, fang-qian@uiowa.edu

Support: NIH/NIDCR T90 DE023520, R01 DE014390 Keywords: matrix metalloproteinases, cytokine profiling, HNSCC

Objectives: Biomarkers like programmed death ligand-1 (PD-L1) have become a focal point for immunotherapeutic checkpoint inhibition in head and neck squamous cell carcinoma (HNSCC). However, it’s only part of the total immunosuppressive biomarker profile of HNSCC cells. Matrix metalloproteinases (MMPs) are enzymes that break down the basement membrane allowing cancer cells to metastasize. MMPs also play an important role in the tumor microenvironment. MMPs can activate certain cytokines, growth factors, and chemokines post-translationally. The objective of this study was to determine MMP and biomarker profiles of seven different HNSCC cell lines. Methods: Cell lines were grown in minimal media at 1*106 viable cells/mL and incubated at 37°C. After 24hrs supernatants were collected, and adhering cells were removed with cell lysis buffer. Multiplex immunoassays were used to determine MMP1, MMP7, MMP9, IL-6, VEGFA, IL-1α, TNFα, GM-CSF, IL-1RA, and IL-8 concentrations in supernatants. ELISAs were used to determine PD-L1, CD47, FASL, and IDO concentrations in cell lysates. A one-way ANOVA was fit to examine log-transformed concentrations of biomarkers between seven HNSCC cell lines, and pairwise group comparisons were conducted using post-hoc Tukey’s Honest Significant Differences test (alpha=0.05). Results: Significant differences (p<0.05) in both MMP and biomarker concentrations were found between the seven HNSCC cell lines. For example, MMP9 was highest in SCC25 and UM-SCC99, MMP7 was highest in SCC25 and UM-SCC19, and MMP1 was highest in SCC25. This suggests different patients’ HNSCC cells can express varying amounts of certain biomarkers and MMPs. These differences could be due to metastatic stage of the cancer, primary tumor site, type of tissue the tumor originated from, or genomic differences between patients. Conclusion: MMP and biomarker expression profiles should be considered when choosing cell lines for future studies. The results support the reason for personalized medicine and the need to further investigate how it can be used to treat HNSCC.

Uncovering the developmental functions of nuclear actin Daniel J. Kelpsch, Camille M. Jamie, and Tina L. Tootle

Several decades of nuclear actin research implicate it in the regulation of transcription, chromatin remodeling, nuclear organization, replication, cell cycle, and the DNA damage response. Recent studies have implicated misregulation of nuclear actin in escape from cellular senescence – implicating nuclear actin levels in cancer progression. However, what functions nuclear actin play in vivo and how the structure of nuclear actin impacts its functions remain largely unexplored. We have identified three reagents to examine nuclear actin during Drosophila oogenesis, i.e. follicle development. Follicles are composed of roughly 1000 somatic follicle cells and 16 germline cells, including 15 nurse or support cells and a single oocyte. Follicles progress through a series of 14 morphological stages, from the germanium to Stage 14 (S14). We find that monomeric actin (G-actin) is present in all cells and fairly uniform levels throughout oogenesis. Conversely, one actin antibody (C4) recognizes nuclear actin during early oogenesis (germarium-S9), including germline and somatic stem cells, mitotic follicle cells, and in transcriptionally silent nurse cells. These data suggest C4 nuclear actin promotes stemness, regulates the cell cycle, and inhibits transcription. This nuclear actin appears to be largely monomeric and does not localize to the chromatin. As C4 nuclear actin decreases with follicle development, another actin antibody (AC15) begins to label nuclear actin. Specifically, the nurse cells exhibit AC15 nuclear actin starting at S8 and this increases with development; the follicle cells exhibit AC15 nuclear actin weakly at S9 and highly starting at S10. AC15 nuclear actin appears to be polymeric and localizes to the DNA; this and other data suggest it promotes transcription in both the nurse cells and the follicle cells. To further explore the roles of nuclear actin, we manipulate nuclear actin levels by blocking its nuclear import (Importin 9) and export (Exportin 6). Knockdown of Importin 9, results in female sterility and defects within the germarium, supporting a role for nuclear actin in stemness. Additionally, reduced Importin 9 levels cause chromatin defects. Loss or knockdown of Exportin 6 results reduced female fertility, distinct chromatin defects, heterochromatin issues, and alterations in the nuclear envelope. These data indicate nuclear actin regulates nuclear organization, and we predict such a function impacts transcription. Together these studies reveal nuclear actin plays critical roles in Drosophila follicle development. Considering the evolutionary conservation of actin structure and function, these studies will also provide insight into the roles of actin in the nucleus in the development of other tissues and organisms, and during tissue homeostasis and cancer.

*All authors: Anatomy and Cell Biology, University of Iowa

Investigating the roles of Fascin in collective cell migration using Drosophila border cell migration

Maureen C. Lamb1, Kelsey Anilker1, and Tina L. Tootle, Ph.D.1 1Anatomy and Cell Biology, University of Iowa Carver College of Medicine, Iowa City, IA 52242

Fascin, an actin binding protein, is a regulator of many developmental processes and contributes to cancer aggressiveness. Functioning to bundle actin filaments, Fascin promotes cell motility, invasion, and adhesion through its canonical role of forming filopodia and invadopodia. Fascin controls cell migration during development such as, growth cone extension and dendrite formation. In addition, Fascin is highly upregulated in certain types of cancer, and elevated expression is associated with increased invasiveness, aggressiveness and mortality of these cancers. While Fascin’s role in regulating cell migration has largely been attributed to its function as an actin bundler, Fascin has other functions including, interaction with mechanotransduction machinery, and nuclear localization, that may contribute to cell migration. While Fascin has been studied in the context of single cell migration and 2D migration, the role of Fascin in 3D collective cell migration has yet to be investigated. To study the role of Fascin in invasive, collective cellular migration in vivo we use Drosophila border cells as a model. Border cell migration occurs during Stage 9 of oogenesis in which a specified group of follicular epithelial cells cluster together and migrate posteriorly in between the nurse cells to the nurse cell - oocyte border. This process is crucial for oocyte development since aberrant or delayed border cell migration leads to female sterility. Fascin is highly expressed in the border cells and fascin-null flies are sterile. These and other findings led us to hypothesize that Fascin plays a critical role in promoting border cell migration during oogenesis. Contrary to prior reports, we find that follicles from young fascin-null flies display a significant delay in border cell migration. Cell-specific knockdown studies suggest that Fascin is required within the somatic and germline cells to mediate migration. Furthermore, rescue of Fascin expression in the germline cells, but not somatic cells, fails to rescue border cell migration. Together these findings implicate a cell-autonomous role for Fascin in collective cell migration in vivo during Drosophila oogenesis. Furthermore, these findings provide a system to investigate the actin bundling-independent functions of Fascin in invasive cellular migration. Overall, this research will lead to a more complete understanding of the function of Fascin in developmental cell migrations and cancer metastasis.

The impact of intra-operative bacterial contamination on peritoneal recurrence of pancreatic cancer

Ann M. Miller1, Aaron T. Scott1, Ann M. Tomanek-Chalkley1, Carlos H.F. Chan1 1Department of Surgery, University of Iowa, Iowa City, IA

Background: Pre-operative biliary stenting is commonly done for pancreatic head adenocarcinoma and spillage of bile contaminated with intestinal flora is therefore unavoidable during pancreaticoduodenectomy. Emerging evidence suggests that post-operative infection links to cancer recurrence. Thus, we hypothesize that intra-operative bacterial contamination facilitates peritoneal recurrence after curative surgery. In this study, we aim to investigate the effects of bacterial lipopolysaccharides (LPS) in peritoneal metastasis of pancreatic cancer.

Methods: LPS (1 mg/kg) or PBS (vehicle control) was injected into the peritoneal cavity of C57BL/6 mice along with mouse pancreatic cancer cells (Panc02). Tumor burden was determined by both number and weight of peritoneal tumors at four weeks post-implantation. Immune cells infiltrating into the peritoneum were examined via flow cytometry following peritoneal lavage two weeks post-implantation. The effect of LPS on cell proliferation in vitro was determined by live-cell fluorescent stain after incubating Panc02 cells with various doses of LPS (1-100 ug/ml) for 24 hours. The impact of LPS on anoikis was determined by colony-forming assay after maintaining Panc02 cells in suspension for 24 hours in the presence of various doses of LPS (1-100 ug/ml). The effect of LPS on apoptosis was determined by Annexin V/PI staining after 24 hours of LPS treatment.

Results: Mice co-treated with LPS had significant higher peritoneal tumor burden (mean tumor number/mouse: 6 vs. 2, P = 0.0192; mean total tumor weight/mouse: 0.54 vs. 0.17 g, P = 0.0296). Mice co-treated with LPS also displayed a significant decrease in frequency of macrophages in the peritoneum (mean macrophage frequency: 7.64 vs 1.46, P =0.0299). While there was no significant difference in apoptosis and anoikis of Panc02 cells after LPS treatment, cell proliferation decreased with increasing LPS doses (1ug/ml: 82%, 10ug/ml: 68%, 100 ug/ml: 44%, P=0.0206).

Conclusions: LPS exposure increases tumor burden in our peritoneal tumor model. But the lack of proliferative and anti-apoptotic effect of LPS on cancer cells in vitro suggests that LPS promotes cancer progression in vivo by modifying the host peritoneal tumor microenvironment.

Title: Loss of the NIAM tumor suppressor cooperates with Myc activation in B-cell malignancies

Authors: Chandra K. Maharjan, Angela M. Schab, Ryan M. Sheehy, Amy L. Whillock, Michael Pisano,

Chandini Reddi, Jacqueline E. Reilly, Jackson Nteeba, Maureen C. Lamb, Timothy Ginader, Brian J. Smith,

Nicholas Borcherding, Xuefang Jing, Van Tompkins, Fenghuang Zhan, Gail Bishop, David K. Meyerholz,

Carol J. Holman, Siegfried Janz and Dawn E. Quelle

Background and Rationale: Diffuse large B-cell lymphoma (DLBCL) is the most common and an

aggressive subtype of B-cell malignancies. A better understanding of molecular mechanisms underlying

its development will identify novel chemotherapeutic targets and treatment strategies. Myc oncogene

activation is a primary initiating event in B-cell lymphoma development. Subsequent alterations in the

ARF-Mdm2-p53 tumor suppressor pathway, including inactivation of the ARF and p53 tumor

suppressors or overexpression of the Mdm2 oncoprotein, cooperate with Myc activation to drive

mature B-cell tumor progression and connote worse outcome. NIAM (Nuclear Interactor of ARF and

Mdm2) is a novel regulator of ARF-Mdm2-p53 signaling whose loss causes B-cell lymphoma at an

incidence of ~20-30% in older animals. NIAM-deficient mice were crossed with B-cell specific Myc

transgenic mice (which develop a spectrum of B-cell lymphomas) to test if NIAM loss promotes Myc-

driven B-cell lymphomagenesis.

Key Findings: Dual mutant mice (Myc transgenic; NIAM-deficient) showed reduced survival, increased

tumor incidence, and accelerated spontaneous lymphoma development compared to Myc transgenic or

NIAM-deficient animals. NIAM deficiency on a Myc transgenic background shifted tumor type from

follicular lymphoma (FL) to the more aggressive DLBCL. Molecular studies revealed that NIAM is a new

binding partner of c-Myc whose loss increases endogenous c-Myc expression. Analyses of endogenous

c-Myc mRNA levels indicated that its regulation by NIAM is post-transcriptional. Functionally, NIAM loss

enhances the proliferation of splenic B-cells without altering their apoptosis. In a pristane model of Myc-

dependent plasmacytomagenesis, NIAM deficiency promoted a more diverse histopathological

phenotype including leukemic involvement of spleen, kidney and other organs. This work uncovers

NIAM as a new negative regulator of Myc that suppresses Myc-driven B-lineage malignancies.

The influence of the tumor microenvironment on CRISPR/Cas9-generated mouse models of sarcoma

Victoria Stephens,1,3 Amanda Scherer,2,3 Vickie Knepper-Adrian,2,3 Rebecca Dodd,2,3

1PREP@Iowa, University of Iowa, Iowa City, Iowa 52242

2Department of Internal Medicine, University of Iowa, Iowa City, Iowa 52242

3Holden Comprehensive Cancer Center, University of Iowa, Iowa City, Iowa 52242

Soft-tissue sarcomas (STSs) are rare mesenchymal tumors that arise from muscle, fat and connective

tissue. The rarity and heterogeneity of patient samples has complicated clinical investigations of

sarcoma biology. Therefore, we aim to find an efficient in vivo animal model using patient-relevant

mutations that allow for preclinical investigations to study the genetics present in STSs, specifically

malignant peripheral nerve sheath tumors (MPNSTs). Recently, we were the first group to generate a

CRISPR/Cas9-mediated mouse model of MPNSTs that allows us to develop these tumors in wild-type

mice. To induce tumor formation, we injected an adenovirus that contains Cas9 and guide RNAs for NF1

and p53 into the sciatic nerve of mice of different background strains including 129XV/Jae, 129X,

C57/BL6, and Balb/C. Initial tumor growth data showed that Balb/C mice have accelerated tumor

development. To investigate mechanisms contributing to the accelerated tumor initiation in Balb/C

mice, we used real-time PCR and immunohistochemistry (IHC) approaches to characterize the tumor

microenvironment (TME) of the various mouse strains. Here, we report the cellular and genetic

differences between the TME of each mouse strain used in these studies. This study is particularly

important because it shows how the influences of the TME between mouse strains can impact tumor

biology.

Exploring the relationships between nuclear actin, the nucleolus, and prostaglandins Dylane Wineland and Tina Tootle Department of Anatomy and Cell Biology, University of Iowa, Carver College of Medicine Actin is not simply a cytoskeletal component, but localizes to and functions within the nucleus. Cell culture studies have implicated nuclear actin in regulating transcription, DNA damage repair, and chromatin organization. Recent work has uncovered that high nuclear actin levels promote the escape from senescence and contribute to cancer development. The mechanisms by which nuclear actin mediates this remain unknown. Here we take advantage of the robust genetic system of Drosophila and the well-characterized process of oogenesis to uncover the functions of nuclear actin. During oogenesis, follicles made up of ~1000 somatic follicle cells and 16 germline cells, consisting of 15 nurse cells and 1 oocyte, progress through a series of 14 morphological stages, from the germanium to Stage 14 (S14). Previous research in the Tootle lab has shown that nuclear actin is dynamically regulated during Drosophila oogenesis. Specifically, germline expression of GFP-Actin induces nuclear actin rods in the nurse cells during Stages 5-9. Similarly, using an antibody to actin (C4), a subset of the nurse cells exhibit structured or blobby nuclear actin beginning around S5 and decreasing until S9. Using both GFP-Actin rod formation and anti-Actin C4 staining we identified prostaglandins (PGs) as a regulator of nuclear actin during oogenesis. PGs are lipid signals made downstream of cyclooxygenase (COX) enzymes and regulate many processes in the body, including reproduction and cancer. In Drosophila, Pxt is the COX-like enzyme and regulates follicle maturation and the actin cytoskeleton. We find that loss of Pxt leads to an increase in the number of nuclear actin rods being formed when GFP-Actin is expressed. It also results in increased levels of endogenous structured nuclear actin (C4) in the nurse cells. These data lead us to propose that prostaglandins negatively regulate nuclear actin. We next sought to uncover the functions of nuclear actin. Given the unusual appearance of the structured nuclear actin, we reasoned that identifying where in the nucleus nuclear actin localizes to would provide functional insight. We find that the structured nuclear actin resides within the nucleolus. Previous studies in the lab have shown that loss of Exportin 6, a nuclear export protein, also leads to increased levels of structured C4 nuclear actin in the nurse cells similar to what we see with loss of Pxt. Interestingly, both show an altered nucleolar morphology through Fibrillarin antibody staining. This finding led us to ask if increased nuclear actin levels were leading to a disrupted nucleolar phenotype. Prior studies have shown that PGs regulate nucleolar structure during oogenesis, leading us to speculate that PGs regulate nuclear actin to control nucleolar structure and function. Future studies will further explore the interplay between nuclear actin, the nucleolus, and PGs. These findings are particularly intriguing in relation to cancer, as high PG levels, increased nuclear actin, and nucleolar changes are all biomarkers of aggressive cancers.

Nuclear GRB2 as a transcriptional regulator and functional modulator of T cells Aline Sandouk, BS1,2, Mahmood Y. Bilal, PhD3, Micaela Fosdick, BS4, Liyang Zhang, PhD5, Zhen Xu, PhD6, Jon C. D. Houtman, PhD1,3 1Interdisciplinary Graduate Program in Immunology, 2Medical Scientist Training Program, and 3Department of Microbiology, 4Interdisciplinary Graduate Program in Molecular Medicine, 5Department of Biochemistry, 6Protein Crystallography Facility, University of Iowa Carver

Background: Growth factor receptor-bound protein 2 (GRB2) is a cytoplasmic adaptor protein present in all human cells. Aberrant function is associated with multiple cancers and chronic diseases. GRB2 deficiency in T cells inhibits clustering of key T cell receptor (TCR) signaling components following activation, leading to downstream inhibition of calcium influx and cytokine production critical for effector activity. However, emerging evidence shows GRB2 localization to the nucleus of T cells, yet no studies have investigated the nuclear function of GRB2. Methods: We have developed recombinant microRNAs that suppress the expression of GRB2 in Hut78 T cells and primary human T cells to examine the effect of GRB2 deficiency on T cell effector function. We have used subcellular fractionation to determine changes in GRB2 levels in the cytoplasm and the nucleus in response to activation. We have also used EMSA to investigate the interaction of GRB2 with DNA oligonucleotides designed on the basis of SELEX-derived consensus and antisensus sequences. Results: GRB2-deficient T cells exhibit greatly reduced gene expression and release of IFN-γ compared to controls after stimulation with anti-CD3, anti-CD28 antibodies. Bypassing TCR signaling downstream of GRB2 with PMA and ionomycin does not rescue the GRB2-deficient phenotype. A strong association of GRB2 with the IFN-γ promoter region was observed, as well as with the transcriptional proteins Rbp1 and CBP. Nuclear translocation of GRB2 increased upon TCR stimulation of primary human T cells, which was enriched for the dimeric versus monomeric forms of GRB2. Conclusions: Together, these findings suggest that upon TCR stimulation, GRB2 interacts with nuclear transport proteins to translocate into the nucleus, where it is recruited to key genomic regions, binds transcriptional machinery, and regulates genes important for T cell effector function.

Presenter contact information: Aline Sandouk Bachelor of Science aline-sandouk@uiowa.edu (319) 335-7597

Title: Toll-like receptor 8 agonist (VTX-2337) enhances HNSCC response to cetuximab

Author: Yinwen Cheng1,2, Rachel Dahl2, Douglas Laux3, Madelyn Espinosa-Cotton2,4, Andrean Simons1,2,4

1Interdisciplinary Graduate Program in Human Toxicology, 2Department of Pathology, 3Department of Internal Medicine - Hematology, Oncology and Blood and Marrow Transplantation, 4Free Radical and Radiation Biology Program, Department of Radiation Oncology, University of Iowa, Iowa City, IA

Background: Epidermal growth factor receptor (EGFR)-targeted therapy involving the monoclonal EGFR antibody cetuximab is routinely used for the treatment of recurrent/metastatic head and neck squamous cell carcinoma (HNSCC). Although cetuximab-based therapy has been proven beneficial, only a subset of patients respond (response rate of 15-20%) despite robust tumor expression of EGFR. In addition to cetuximab’s inhibitory effects on EGFR, its IgG1 backbone triggers additional antitumor activity in the form of natural killer (NK) cell-mediated antibody-dependent cell-mediated cytotoxicity (ADCC). NK cell-mediated ADCC is believed to be the major mechanism of cetuximab’s clinical activity, which is supported by the fact that EGFR TKIs and IgG2 antibodies that are unable to trigger NK cell-mediated ADCC, provide little to no clinical benefit for HNSCCs. Recently, toll-like receptor (TLR) agonists have garnered much interest due to their ability to invoke NK cell-mediated anti-tumor responses. In particular, TLR8 is widely spread on myeloid dendritic cells, monocytes and monocyte-derived dendritic cells and anti-tumor effects have been observed with a small molecule TLR8 agonist VTX-2337. Based on these data, we propose that VTX-2337 would enhance HNSCC response to cetuximab by promoting NK cell activation.

Method/Results: Over a 3-week treatment period in a TUBO-EGFR/BALB/c mouse model, we found that VTX-2337 did not affect tumor growth as a single agent or in combination with cetuximab. However, the survival rate of mice treated with the combination of VTX-2337 and cetuximab (median survival = 65 d) was dramatically longer than either agent alone (median survival ~ 25 d) suggesting that this combination may result in long term changes in survival and not immediate changes in response. In mechanistic studies we observed that VTX-2337 resulted in the secretion of the proinflammatory cytokines IL-1β, TNF-α and IFN-γ (measured by ELISA) from human peripheral blood mononuclear cells (PBMCs). VTX-2337 also significantly promoted NK cell activation (CD54+, CD16-) compared to controls when in combination with cetuximab.

Conclusion: Our results suggest that the anti-tumor efficacy of cetuximab can be enhanced with a TLR8 agonist and provides a rationale for further investigation of cetuximab in combination with other immunotherapeutic agents that enhance NK cell activity.

Diagnostic accuracy of 68Ga-DOTATOC positron emission tomography(PET) in neuro-endocrine tumors. Silvia N. Ghobrial, MD; Yusuf Menda, MD; Sarah L Bell, MS; Gideon K Zamba,PhDand M. Sue O’Dorisio MD,PhD.

Background: Neuroendocrine tumors (NETs) are solid tumors withmalignant potential that arise from neuroendocrine cells throughout thebody. NETs characteristically express somatostatin receptors (SSTR)which is conventionally utilized as a target for diagnostic and therapeuticinterventions. Octreoscan single photon emission tomography (SPECT)and high resolution, contrast enhanced CT scan constitute the current goldstandard of imaging for the detection and the staging of neuroendocrinetumors and other somatostatin receptor positive cancers. This novelimaging modality has the relative advantage of cutting down imaging timeto two hours as opposed to two days with SPECT. The aim of this study isto investigate the diagnostic accuracy of 68Ga-DOTATOC PET in NETscompared to the currently standard of care.Method: In this prospective comparative study, subjects with confirmedNETs seen at the University of Iowa Hospitals and Clinic between June2012 and June 2014 were screened and consented. A total of 67 subjectsunderwent 68Ga-DOTATOC PET in addition to their standard of careimaging studies comprising Octereoscan and high-resolution contrast-enhanced CT. Out of the 67 subjects, 39 subjects underwent all threeimaging studies within the predetermined 4 months period and wereincluded in the final analysis. Two reviewers independently interpreted theanonymized imaging studies and recorded the presence or absence ofdisease in each body segment across the three imaging modalities.Disease burden as determined by each imaging modality was directlycompared to assess diagnostic accuracy.Results: A significant difference in disease burden was seen on directcomparison between imaging modalities. On average, 68Ga-DOTATOCPET identified lesions at 1.2 more sites compared to Octreoscan and CT(P<0.01). Whilst Specificity and positive predictive values were similar at100% for both techniques. 68Ga-DOTATOC PET showed superiorsensitivity (97% vs 88%) and negative predictive value (86% vs 60%).

Conclusion: 68 Ga-DOTATOC PET is a novel imaging technique withsuperior lesion detection rate and accuracy compared to the currentimaging techniques.Corresponding author:Silvia N. Gobrial, MDPostdoctoral ScholarDepartment of Pediatric Hematology and Oncology University of IowaHospitals and ClinicsSilvia-ghobrial@uiowa.edu

*all authors: University of Iowa Hospitals and Clinics

Thrombotic potential in pediatric patients with acute lymphoblastic leukemia at baseline and during induction therapy

Rahul Kumar, PhD1, Roxane M Mitten, B.A., 2, Anjali Sharathkumar2, ,M.D., and Sanjana Dayal, PhD1 1Departments of Internal Medicine and 2Stead Family Department of Pediatrics, University of Iowa, Carver College of Medicine, Iowa City, IA

Background: Thromboembolism is one of the comorbidities in children with acute lymphoblastic leukemia (ALL), affecting up to 30% of patients; yet the mechanisms of thrombogenesis remain elusive. We examined if the cell free DNA (cfDNA) and histones are associated with thrombogenesis in ALL. This concept was translated from recent observations suggesting that components of neutrophil extracellular traps (NETs), like cfDNA and histones contribute to thrombosis. During tumor lysis in ALL it’s possible that cfDNA and histones are released from leukemic cells leading to thrombogenesis. We hypothesized that circulating levels of cfDNA and nucleosomes increases during the induction therapy in pediatric ALL and are associated with increased thrombogenic potential. Methods: In a longitudinal study, we recruited consecutive ALL patients and the blood samples were collected at the time of diagnosis and weekly during the induction therapy for up to 4 weeks. Blood samples were also collected from age matched healthy subjects, and patients with sepsis and deep vein thrombosis (DVT) as a positive control. Plasma levels of cfDNA were measured using Qubit DNA kit and nucleosomes were measured using Cell Death Detection ELISA Kit. Calibrated Automated Thrombogram was used to assess thrombin generation. Results: We recruited a total of 31 children: ALL: 18; healthy children: 5; Sepsis: 3; DVT: 5. We first established that the levels of cfDNA and endogenous thrombotic potential (ETP) were significantly higher in patients with sepsis and DVT as compared with healthy controls. Interestingly, in ALL samples, significant increase in cfDNA and ETP were observed at the baseline as well as during induction. Analysis using one way ANOVA showed overall significant increase in cfDNA and ETP compared to healthy control. Plasma nucleosomes also showed a nonsignificant increasing trend at baseline and during induction compared to healthy controls. However, the levels of cfDNA or nucleosomes did not correlate with ETP. Conclusions: Our findings suggest that a higher thrombogenic potential exists in pediatric ALL at the diagnosis as well as during induction phase, which is independent of plasma cfDNA and nucleosomes. Future efforts will focus on understanding the association between common ALL mutations and thrombogenesis.

Thyroid Hormone Receptor Interacting Protein 13 AAA-ATPase Regulates Cellular Deubiquitination

Can Li* MS; Ivana Frech DSc, PhD; Reinaldo Franqui-Machin PhD; Jiliang Xia PhD; Guido Tricot MD, PhD; Fenghuang Zhan MD, PhD.

Division of Hematology, Oncology, and Blood Marrow Transplantation, Department of Internal Medicine, University of Iowa, Iowa City, IA, USA.

*Presenter: Can Li, MS, can-li@uiowa.edu, Phone # 319-384-1339

BackgroundDeubiquitination like ubiquitination is a highly regulated process and implicated in almost everycellular process. Multiple Deubiquitinating enzymes (DUBs) have been identified but theirfunctional regulation is poorly defined. Mechanistic understanding of DUBs activity includingUbiquitin carboxyl-terminal hydrolase 7 (USP7) is cryptic. It appears that energy probablyprovided by a scaffolding protein is required for USP7 to correctly associate with its substrates.

Methods The Thyroid Hormone Receptor Interacting Protein 13 AAA-ATPase (TRIP13) was highly expressed in multiple myeloma cells conferring resistance to Bortezomib treated cells. Thus, we hypothesize that TRIP13 could be involved in cellular protein degradation. To address this, we performed a drug screening to overcome the Bortezomib resistance. Using western blot analysis and in vitro assay we discovered that TRIP13 regulates cellular de-ubiquitination and by co-immunoprecipitation we showed that TRIP13 binds USP7 and provides energy to allow USP7 to activate its targets. To further confirm the role of TRIP13 in vivo we generated lymphocytes-specific transgenic mice. Cellular fractionation and western blots were used to analyze USP7 substrates such us p53 and PTEN.

Results Here, we found that TRIP13 when overexpressed in multiple cancer and other disorders regulates ubiquitination negatively by increasing cellular deubiquitination. Overexpression of TRIP13 in mice and culture cells resulted in excess cellular deubiquitination by modulating the association of the USP7 with its substrates (PTEN, p53, NEK2). Deletion of TRIP13 resulted in USP7 reduced-substrate interaction linked to diseases ranging from cancer to other disorders. TRIP13 lymphocytes-specific showed lower cellular ubiquitination in thymus and spleen with consequent changes in USP7 substrates localization and protein levels such us PTEN and p53.

Conclusion Our studies presented here demonstrate that TRIP13 plays a unique and novel role in regulating cellular ubiquitination and specifically on USP7 activation. TRIP13 proves to be an important target of therapeutic drug action in multiple cancers.

Integration of selective single-cell capture at a wireless electrodearray with on-chip electrical lysis and high-throughput analysis

Min Li, Robbyn K. Anand*

Department of Chemistry, Iowa State University, Ames, IA 50010, USA.

Email: rkanand@iastate.edu

Integration of versatile selective isolation techniques with subsequent single-cell analysiscontinues to be an important challenge. Separation based on dielectrophoresis features antibody-independent separation and also avoids leukocyte contamination. However, many of the currentapproaches suffer from low throughput and are not scalable. These limitations stem from designconstraints such as the requirement that all electrodes must be connected via wire leads to thepower source. Further, in DEP devices that employ insulating posts to shape the electric field,integration of these structures intended for cell capture with other features, such as chambers foron-chip analysis, is non-trivial. Here we report a DEP strategy that addresses these concerns.First, DEP is employed to selectively capture and isolate CTCs in micropockets, while blood cellsare excluded and flow past. The capture methodology used here eliminates sequential screeningof all cell populations one-by-one, the case in droplet separation, thus greatly increasing sortingefficiency. Moreover, high-fidelity single-cell capture could be readily achieved when the pocketdimensions matched to those of the cells. Second, the leak channel design reported here opensa flow circuit that enables valve-free transfer of individually isolated cells into reaction chambers,while a ‘split electrode’ design allows recapture within the chambers and electrical lysis forsubsequent assay evaluation. Third, the use of arrays of wireless electrodes removes therequirement of ohmic contact to individual array elements, thus enabling device to be scalablealong both x- and y- directions. This wireless electrode array not only provides a high-throughputmodule in cell capture, but also in cell imaging and analysis. A key difference that distinguishesthis from previous approaches is that DEP-based sorting, electrical lysis and analysis of singlecells are integrated while retaining high-throughput and valve-free control. Most importantly, thefunctions are accomplished with minimal peripheral equipment – a power supply and amicroscope – thus increasing the relevance of this platform to point-of-care application.

Preclinical evaluation of CXCR4 as novel radio-theranostic target for high grade neuroendocrine and neuroblastoma tumors

Presenter: Dijie Liu, DVM, PhD, dijie-liu@uiowa.edu, 319-356-6506

Co-authors: Mengshi Li, PhD candidate; Andrew M. Bellizzi, MD; Dongyoul Lee, PhD candidate, Yusuf

Menda, MD; Kimia Nourmahnad, MS; Michael K. Schultz, PhD; M Sue. O’Dorisio, MD, PhD

Background: Somatostatin receptor subtype 2 (SSTR2) targeting is a mainstay of diagnosis and therapy

for low grade neuroendocrine tumors (NETs), and SSTR2 directed peptide-receptor radionuclide therapy is

effective in patients with G1 and G2 NETs. However, the approach is relatively ineffective for

neuroendocrine carcinomas (NECs) and G3 NETs because SSTR2 expression is low in these tumor types.

Advanced neuroblastoma (NB), as the most common extracranial solid tumor of childhood, remains a

clinical challenge with poor survival. Thus, there is a critical need for new theranostic targets that are

expressed in NECs and NB. Chemokine receptor 4 (CXCR4) has been proved to stimulate cell

proliferation, angiogenesis, migration, and metastasis of cancer as a poor prognostic factor. In this study

we evaluated the expression of CXCR4 in high grade NETs and NB cancer cell lines in vitro, in patient

tissue microarray and evaluated the microPET/CT imaging profile of 68Ga-Pentixafor (a CXCR4

antagonist), in mice bearing high-grade NETs and NB xenografts.

Methods: CXCR4 mRNA and protein expression was quantified by qGPCR array assay and flow

cytometry in high-grade tumor cell lines. Tumor xenografts from mice were collected for

immunohistochemistry staining evaluation. Pentixafor was labeled with 68GaCl3 for microPET/CT imaging

in tumor-bearing mice, followed by euthanasia and assessment of biodistribution.

Results: IMR32 neuroblastoma cells expressed high levels of SSTR2 and CXCR4 while H727 lung

neuroendocrine cells expressed a moderate level of CXCR4 and minimal SSTR2, as evidenced by both in

vitro and in vivo tests. MicroPET/CT imaging of 68Ga-Pentixafor demonstrated high retention in IMR32 and

H727 tumor xenografts (5.8% and 1.1% ID/g, respectively) 1 hr post-injection with primarily renal clearance

and partial distribution to digestive tract. 68Ga-Pentixafor tumor uptake was specifically blocked by co-

injection of excess mass of unlabeled antagonist, Plerixafor.

Conclusion: CXCR4 is expressed at variable levels in G3 NETs and NB cell lines. 68Ga-Pentixafor is a

potential radiopharmaceutical for microPET/CT imaging of CXCR4 expression in mice bearing high-grade

NETs and NB. 68Ga/177Lu-, 68Ga/90Y-Pentixafor are promising radio-theranostic pairs for targeting CXCR4

in patients with G3 NETs or NECs and NB.

DNA damaging agents, cisplatin and carboplatin, have been widely used as a primary treatment

for many types of cancer including endometrial cancer and triple-negative breast cancers (TNBC). Endometrial cancer is the most frequent gynecologic malignancy. TNBC is about 10-20% of breast cancers, which are negative for hormone receptors of estrogen, progesterone and HER2 expression. Drug resistance is one of the major causes of therapeutic failure for patients with metastatic or recurrent disease, thus highlighting the need to identify novel factors

driving resistance to DNA damaging agents. Acquired drug resistance prevents cancer therapies from achieving stable and complete responses. Emerging evidence implicates a key role for non-mutational drug resistance mechanisms underlying the survival of residual cancer ‘persister’ cells. Metadherin (MTDH, also known as AEG-1 and LYRIC) has been

consistently associated with resistance to multiple chemotherapeutic agents and metastasis. Disruption of MTDH can increase sensitivity to chemotherapy. MTDH is positively connected with c-myc via positive feedback loop. There are no known small chemical inhibitors to target c-

myc or MTDH. GPX4 inhibitor was found to induce the reduction of MTDH and c-myc during ferroptosis in endometrial cancer and TNBC cancer cell lines. These findings suggest that targeting of GPX4 may represent a therapeutic strategy to prevent acquired drug resistance.

Abstract Title: Ferroptosis induction by GPX4 inhibitor in endometrial cancer and triple negative breast cancer

Authors (as submitted by poster presenter):Xiangbing Meng, Department of Obstetrics and GynecologyJianling Bi,Department of Obstetrics and GynecologyShujie Yang, Department of Obstetrics and GynecologyBrett Wagner,Free Radical Radiation Biology, and Division of the Department of Radiation OncologyMary Li,Department of Obstetrics and GynecologyZihan Wang, Department of Obstetrics and GynecologyGarry R. Buettner,Free Radical Radiation Biology, and Division of the Department of Radiation OncologyDouglas Spitz, Free Radical Radiation Biology, and Division of the Department of Radiation OncologyMeng Wu, High Throughput Screening Facility at University of Iowa (UIHTS), Department of Biochemistry, Carver College of Medicine, Division of Medicinal and Natural Products Chemistry, Department of Pharmaceutical Sciences and Experimental Therapeutics, College of PharmacyKimberly K Leslie,Department of Obstetrics and Gynecology

Intratumoral Toll-like Receptor 9 (TLR9) agonist, CMP-001, in combination with pembrolizumab can reverse resistance to PD-1 inhibition in a phase 1b trial in subjects with advanced melanoma

Mohammed Milhem1, Sue Blackwell1, George Weiner1, Yousef Zakharia1, Melanie Frees1, Jill Corlette1, Michele Freesmeier1, Jaime

Bonner1, Rene Gonzales2, Theresa Medina2, John M. Kirkwood 3, Elizabeth Buchbinder4, Inderjit Mehmi5, Jiaxin Niu6, Montaser

Shaheen7, Ryan Weight8, Kim Margolin9, Jason Luke10, Aaron Morris11, David Mauro11, Arthur M. Krieg11 and Antoni Ribas12

1University of Iowa, 2

University of Colorado Denver, 3

University of Pittsburgh Medical Center, 4

Dana Farber Cancer Institute,5West Virginia University, 6Banner MD Anderson Cancer Center, 7University of Arizona, 8Thomas Jefferson University, 9City of Hope, 10University

of Chicago, 11Checkmate Pharmaceuticals, and 12University of California Los Angeles

Background : CMP-001 comprises a CpG-A oligodeoxynucleotide packaged within a virus-like particle. It is designed to activate tumor-associated

plasmacytoid dendritic cells via TLR9 inducing an interferon-rich tumor microenvironment and anti-tumor CD8+ T cell responses.

Materials and Methods : CMP-001-001 is an ongoing Phase 1b trial evaluating intratumoral (IT) CMP-001 in combination with pembrolizumab

(administered per label) in subjects with advanced melanoma resistant (either did not respond or progressed) on prior anti-PD-1 monotherapy or

in combination. During dose escalation, subjects were enrolled to cohorts of ≥ 3 subjects at CMP-001 doses of 1, 3, 5, 7.5, and 10 mg in two

dosing schedules (weekly for 7w, followed by q3w; or weekly for 2w, followed by q 3w). CMP-001 was administered IT into an accessible lesion(s),

and response assessed in all target lesions (injected and non-injected) by RECIST v1.1. Study therapy was continued until progression, toxicity,

investigator decision or withdrawal of consent. Baseline and on-therapy serum was collected for cytokine analysis. Immunohistochemical and

RNA-Seq analysis was performed on available pre- and post-treatment tumor biopsies.

Results: As of December 31, 2017, 68 subjects have been treated (44 in Escalation and 24 in Expansion). Safety data from 63 subjects

demonstrated a manageable acute toxicity profile consisting predominately of fever, N/V, headache, hypotension and rigors. Grade 3/4 related

AEs reported in ≥1 subject; hypotension (n=7), anemia (n=2), chills (n=2), hypertension (n=2) and fever (n=2). The Objective Response Rates

(ORR) across all dose cohorts on weekly (n=40) and q3week schedules (n=13) were 22.5% (9/40; 95 % CI 11-39%) and 7.7%% (1/13; 95% CI 0-36%)

respectively. For subjects dosed weekly at 3 and 5 mg, the ORR was 33.3% (6/18 95% CI 13-59%). Of the 10 responders, 1 progressed (w36), 2

withdrew consent (w13, w25), 7 remain on study with 2 subjects maintaining their response though w72. Regression of non-injected tumors

occurred in cutaneous, nodal, hepatic, and splenic metastases. CMP-001 induced TLR9 activation with a median 5.9 fold increase in serum

CXCL10 (range of 0.9 – 276.3; mean fold increase of 21.8 with SD=48.8; n=39). Immunohistochemical and RNA-Seq analysis of tumor biopsies

revealed increases in tumor-infiltrating CD8 T cells (>5 fold), PD-L1 expression (>3 fold increase in H score), and transcriptional signature of

inflammation in 2/4 subjects with analyzable pre-and post-treatment samples.

Conclusions: CMP-001 in combination with pembrolizumab resulted in objective, durable tumor responses with tolerable toxicities in subjects

with advanced melanoma resistant to prior anti-PD-1 therapy. CMP-001 dosing at 5 mg/weekly has been selected for further evaluation in the

ongoing dose expansion phase of this study.

RhoA-myosinII axis protects circulating tumor cells from fluid shear stress-induced damage.

Devon Moose, Benjamin Krog, Lei Zhao, Gretchen Burke, Lillian Rhodes, and Michael Henry

Circulating tumor cells (CTCs) are exposed to hemodynamic forces, which have long been thought to be mechanically destructive to CTCs. However, recent studies show that cancer cell lines from diverse histologies exhibit resistance to brief (millisecond) pulses of high-level (750-6400 dyn/cm2)fluid shear stress (FSS), whereas non-transformed epithelial cells are sensitive to this mechanical insult (PMID: 23226552, 26447202). Moreover, exposure of cancer cells to FSS results in cortical stiffening (PMID: 25908902). Herein, we elucidate the mechanism of FSS resistance in cancer cells, and extend these findings to experimental models of CTCs. We show that although some cancer cell lines exhibit elevated levels of membrane repair relative to non-transformed counterparts, intrinsic resistance to plasma membrane damage is a more consistent feature distinguishing cancer cells. FSS resistance is detectable in cancer cells acutely isolated from primary mouse TP53/PTEN mutant prostate tumors, not just a feature of cultured cancer cell lines. Our findings indicate that cancer cells respond to FSS by activation of RhoA-myosinII contractility which protects them from nanometer-scale damage to the plasma membrane. Moreover, we present evidence that the RhoA myosinII axis protects CTCs from mechanical damage in animal models. Treatment of PC-3 cancer cells with a non-toxic dose of the myosin II inhibitor blebbistatin (20µM; 3h) reduced the number of intact cells arrested in the lung microvasculature immediately following tail vein injection. Additionally, treatment of mice bearing orthotopically implanted, metastatic PC-3 derived prostate tumors with blebbistatin (2.5mg/kg; 3h) acutely reduced steady-state CTC levels by approximately 10-fold. Taken together, our data indicate that viable CTCs actively resist destruction by hemodynamic forces and are likely to be more mechanically robust than is commonly thought.

Author affiliation (as submitted by poster presenter):Devon Moose / Molecular Physiology & Biophysics, Cancer BiologyBenjamin Krog / Molecular Physiology & Biophysics, SynderBioLei Zhao / Molecular Physiology & BiophysicsGretchen Burke / Molecular Physiology & BiophysicsLillian Rhodes / Molecular Physiology & BiophysicsMichael Henry / Molecular Physiology & Biophysics, Cancer Biology, SynderBio

Nuclease-activated probe (NucAP) technology for cancer detection

Shambhavi Shubham1, Sven Kruspe1, David D. Dickey1, Kevin Urak1,2, Giselle N. Blanco1, Brian J Smith3,4, Melissa

Curry4, Michael E Wright5, James O. McNamara II1,2, Carlos H. Chan6 Paloma H. Giangrande1,2,7.

1 Internal Medicine, University of Iowa, Iowa City, IA, USA 2 Molecular & Cellular Biology Program, Iowa City, IA, USA 3 Department of Biostatistics, University of Iowa, Iowa City, IA, USA 4 Holden Comprehensive Cancer Center, University of Iowa, Iowa City, IA, USA 5 Department of Physiology, University of Iowa, Iowa City, IA, USA 6 Department of Surgery, University of Iowa, Iowa City, IA, USA 7 Radiation Oncology, University of Iowa, Iowa City, IA, USA

Nucleases have been proposed as powerful biomarkers for cancer detection. Advantages of nucleasesfor cancer detection include: 1) their elevated expression in most cancer cells (including those cancercells in blood implicated in metastasis that have undergone an epithelial-to-mesenchymal transition –EMT), and 2) their enzymatic activity, which can be exploited for signal-amplification in detectionmethods. We recently developed a novel technology to enable the rapid and robust detection of cancer-derived nucleases in patient blood (Fig. 1). This technology, Nuclease Activated Probe (NucAP), isbased on chemically-modified, quenched fluorescent probes that are activated by cancer nucleases butnot nucleases present in human serum (e.g. RNase A). Proof-of-concept studies to date exploring thisapproach have focused on circulating tumor cell (CTC)-derived intracellular nucleases from advanced(stage IV) breast cancer patients.1 These studies have identified 3 probes (dsDNA, ssDNA and 2′F-RNA)which exhibited robust performance in distinguishing patients with stage IV breast cancer from healthycontrols. Given the difficulty in capturing CTCs from blood, more recently, we have begun to evaluate theNucAP technology with cancer-derived nucleases secreted in serum/plasma. To date, we have observed

elevated cancer-derived nuclease activity in both breast andpancreatic cancer cell lines grown in culture and patient serum.Elevated nuclease activity in patient serum was consistent with thepresence of DNAse-like nucleases. In contrast, DNAse-likenuclease activity was reduced in the plasma samples of prostatecancer patients compared to healthy donors and inverselycorrelated with disease stage. Proteomic analysis of the prostatecancer plasma samples has revealed the identity of severalDNAse-like nucleases whose levels decrease with diseaseprogression. Current studies are focused on confirming the identityof the specific nucleases and determining the mechanism forreduced nuclease activity in prostate cancer. We are alsoinvestigating potential serum vs. plasma collection effects on assayturnout. In summary, the NucAP approach is simple (which can

facilitate point-of-care), inexpensive, and can be applied to both CTC-derived intracellular nucleases aswell as cancer-derived nucleases secreted in serum without the CTC capture step. The implications ofthese findings could be transformative for several reasons. 1) they could lead to the development ofeffective assays for the early detection of various cancers including breast, pancreatic and prostatecancer and for monitoring treatment response that is both straightforward and cost effective. 2) theyprovide proof-of-principle for a new point-of-care platform that would enable clinicians to capitalize on thevast potential of nucleases as biomarkers for cancer diagnostics.

1. Kruspe, S. et al. Rapid and Sensitive Detection of Breast Cancer Cells in Patient Blood with Nuclease-Activated

Probe Technology. Mol Ther Nucleic Acids 8, 542-557 (2017).

Novel assay using ligand-receptor complex-binding RNA aptamers to measure occupancy of IL2 receptor a (IL2Ra) by IL2

Suresh Veeramani, Sue E. Blackwell, William H. Thiel, Paloma H. Giangrande and George J. Weiner

Holden Comprehensive Cancer Center and Department of Internal Medicine, University of Iowa, Iowa City, IA 52241

Background: RNA aptamers are oligonucleotides that bind to antigens in a manner analogous to antibodies. Using novel IL2- IL2Ra complex-binding RNA aptamers, we report a novel assay that allows for quantitation of receptor occupancy by its ligand.

Methods: A whole cell Systematic Evolution of Ligands by EXponential enrichment (SELEX) strategy was used to select T regulatory (Treg) cell-specific RNA aptamers. Aptamers against common T cell antigens were precleared using normal donor CD4+IL2Raneg T cells, which was followed by selecting for CD4+IL2Ra+ Treg-binding aptamers. The process was repeated for eight rounds with each round using T cells from different donors. High-throughput sequencing and bioinformatics analysis followed to select top candidate aptamers. Binding of the top candidates to IL2Ra and IL2-bound IL2Ra was determined. A TaqMan RT-qPCR-based binding assay was designed to measure IL2Ra receptor occupancy using aptamers showing differential binding to unoccupied and IL2-occupied IL2Ra. Aptamers that bound preferentially to unoccupied receptor (e.g. Tr-8) or to IL2-occupied receptor (e.g. Tr-7) were paired in equimolar quantity and added to IL2Ra-coated Dynabeads incubated with various concentrations of IL2. Aptamer binding was quantified in RT-qPCR by using aptamer-specific fluorescent probes.

Results: Five of the top 12 Treg-binding aptamers recognized IL2Ra. Among these, some aptamers bound preferentially to unoccupied IL2Ra (Tr-6 and Tr-8) and others to IL2-IL2Ra complex (Tr-1 and Tr-7). A standard curve capable of measuring IL2Ra occupancy by IL2 was constructed by comparing the percentage of IL2-occupied IL2Ra against the ratio of bound Tr-7:Tr-8 (R2=0.946; p=0.001).

Conclusions: We identified novel aptamer pairs showing differential binding preferences towards unoccupied IL2Ra or the IL2-IL2Ra complex. Utilizing this differential binding property, we developed a novel assay that measures receptor occupancy by IL2. The ability to measure receptor occupancy by such aptamers is highly novel and significant that could be valuable to study IL2-IL2Ra complex as a diagnostic as well as therapeutic target in anti-tumor immunity. Importantly, modification of this assay to measure other receptor-ligand complexes can have significant impact in the cancer immunotherapy field.

Title : A novel tool for monitoring endogenous progesterone receptor expression by mCherry using CRISPR-Cas9 genome editing technique

Contact information for presenter: Full name: Shujie Yang, PhD Email: Shujie-yang@uiowa.edu Tel: 353-3071

Authors: Yiyang Li , Wei Zhou, Abby Fronk, Xiangbing Meng 1, 2, Kuo-kuang Wen, Meng Wu, Adam Dupuy, Kimberly Leslie 1, 2, Shujie Yang 1, 2, *

Abstract Background: Endometrial cancer is the most common gynecologic cancer with incidences and deaths increasing over the past 30 years, likely due to the ineffectiveness of standard treatments. Progesterone therapy has over 60 years of history in treating endometrial cancer with relatively good outcomes for primary endometrial cancer, but less promising for metastatic and recurrent disease. Literature supports high PR expression in correlation with better clinical outcomes.

Methods: We have designed a PR reporter gene with mCherry and inserted it into the PR gene by homologous recombination after CRISPR-Cas9 generated the DNA double strand break at the end of exon8, to monitor the endogenous PR expression. Two endometrial cancer cell lines, Ishikawa and Hec50, were used to test the effect of various drugs on PR and illustrated through the red fluorescence of mCherry.

Results: We have validated a PR expression increase in parallel with mCherry expression after treating PR reporter gene transfected Ishikawa cells with histone deacetylase inhibitor, Romidepsin. The PR reporter gene transfected Ishikawa cells have been used to screen the FDA-approved 1018 library in search of more drugs effective in increasing PR expression. We hope a new FDA drug can restore functional PR expression and further sensitize endometrial cancer to progesterone therapy. This reporter gene was also been used to identify novel PR repressors by using GeCKO library. We demonstrated that potential novel PR repressors are APH1A, SOX9, GLUD2 and BCAS4.

Conclusion: This technique can also be used to generate endogenous reporter gene for many candidate target genes.

CD138+/CD24+ As Tumor-Initiating Cell Biomarkers in Multiple Myeloma

Fenghuang Zhan MD, PhD1*; Minjie Gao, PhD1; Hua Bai, BS1; Yogesh Jethava, MD1; Jiliang Xia, PhD1; Reinaldo Franqui-Machin, PhD1; Kalyan Nadiminti, MD1; Michael Tomasson, MD1; Siegfried Janz, MD2; Ivana Frech, DSc, PhD1; Guido Tricot, MD, PhD1. 1Division of Hematology, Oncology, and Blood and Marrow Transplantation, Department of Internal Medicine, University of Iowa, Iowa City, Iowa, USA. 2 Department of Pathology, University of Iowa, Iowa City, Iowa, USA.

*Presenter: Fenghuang Zhan MD, PhD, fenghuang-zhan@uiowa.edu, Phone # 319-384-0066

Background Treatment failure in cancers, including multiple myeloma (MM), are mostly likely due to the persistence of a minor population of tumor initiating cells (TICs), which are non-cycling or low-cycling and very drug-resistant tumor cells. Methods We particularly interested in genes encoding cell surface proteins and found that CD24 was the one of top upregulated gene in the light chain-restricted/Side Population MM cells compared to CD138+ MM cells. We, thus, hypothesize that CD24 was a TIC marker. To address this, we examined self-renewal potential by clonogenicity and drug resistance of CD138+ CD24+ sorted cells. To confirm the presence of MM CD24+ subpopulation in primary MM patients, we analyzed 60 patients’ samples by flow cytometry using the panel of plasma cell markers (CD138, CD38, k+ or λ+ light chain, and CD56), and B cell markers (CD45, CD19), as well as the CD24 antibody. We also analyzed CD24 expression in 84 newly diagnosed MM patients using immunohistochemistry (IHC) with CD138 and CD24 antibodies. To identify CD24 signaling pathways for maintaining TIC features, the transcriptional factor activation assay and gene expression profiles were performed on CD24+ and CD24– MM cells Results We have identified that the cell surface antigen CD24 is a TIC biomarker in multiple myeloma (MM). CD24+ MM cells show increased clonogenecity, drug resistance, and tumor formation evidenced that as few as 10 CD24+ MM cells developed tumors in vivo. CD24+ MM cells are enriched after chemotherapy in complete remission (CR) MM patients with minimal residual disease (MRD) and therefore linked to drug resistance. The induced pluripotent or embryonic stem cell (iPS/ES) genes, such as NANOG, OCT4, KLF4, and SOX2, are significantly upregulated in CD24+ MM cells. STAT3 binds with CD24 and mediates CD24-induced stemness. Immunotargeting of CD24+ MM cells prevents tumor progression in vivo. Conclusion Our studies presented here demonstrate that CD24 maintains the feature of self-renewal and drug resistance in MM, providing a biomarker for myeloma genesis and targeted therapy.

The combination of all-trans retinoic acid-loaded nanoparticles and an adenoviral tumor vaccine has a benefit on survival in a tumor model system

Clinical benefit of cancer immunotherapies is mostly noted as prolonged survival. Better

results will most likely be obtained after identification of the best antigens, adjuvants, delivery

vehicles, administration routes, and appropriate co-treatments to alleviate tumor

immunosuppression. Preclinical and clinical studies have shown that all-trans retinoic acid

(ATRA) drives the differentiation of myeloid-derived suppressor cells (MDSC), which are key

drivers of resistance, into mature functional cells, therefore reducing tumor immunosuppression.

However, the exact mechanism of action that regulates the fate of immune cells after ATRA

treatment remains unknown. Nonetheless, several other important observations revealed a

valuable immunoregulatory property of ATRA as a drug for treatment of cancer, where ATRA

may be able to boost the development of antitumor immunity. However, the use of this drug is

limited due to the occurrence side effects, unfavorable physico-chemical characteristics, and

poor pharmacokinetic profile. Here we prepared retinoic acid-loaded poly(lactic-co-glycolic acid)

nanoparticles (NP) and also investigated if combining ATRA-NP with a therapeutic vaccination

using an adenovirus encoding a model tumor-associated antigen, ovalbumin (Ad5-OVA), can

enhance the circulating levels of OVA-specific CD8+ T lymphocytes and improve survival in

tumor-challenged mice. In vitro, the combination of ATRA-NP and Ad5-OVA up-regulated the

expression of CD86 in bone marrow-derived dendritic cells. For the in vivo studies, mice were

challenged with E.G7-OVA cells and treated s.c. with Ad5OVA (108 PFU) peritumorally and/or

ATRA formulations (15 mg/kg) once the tumors were palpable (3 or 4 days after tumor

challenge). ATRA-NP or ATRA in solution was administered during 7 consecutive days. It was

observed an increase, albeit not statistically significant, of OVA-specific CD8+ T lymphocytes for

the animals treated with the combination of ATRA-NP and Ad5-OVA compared to Ad5-OVA

treatment only. Untreated animals and mice treated with ATRA only had tumors that

progressively grew and most of these mice were sacrificed between day 18 and 25 post-tumor

challenge. The survival of animals treated with ATRA-NP + Ad5OVA was significantly higher

than the survival of animals treated with Ad5-OVA alone. These results suggest that the

beneficial effects of Ad5-OVA and ATRA-NP combinatory treatment may be due to

improvement of the tumor-specific immune response and decreased tumor immunosuppression.

Authors (as submitted by poster presenter):Cristina Maria de Barros / University of Iowa College of PharmacySean Geary / University of Iowa College of PharmacyAliasger Salem / University of Iowa College of Pharmacy

Title: Cancer immunotherapy based on in situ immunization with a TLR9-agonist-

containing virus-like particle in mice

Authors & Affiliations:

Caitlin D. Lemke-Miltner1, Sue E. Blackwell1, Anna E. Krug1, Sarah L. Mott1, Brian J.

Smith1,2, Aaron J. Morris3, Arthur M. Krieg3, and George J. Weiner1

1) Holden Comprehensive Cancer Center and Department of Internal Medicine,University of Iowa

2) Department of Biostatistics, University of Iowa3) Checkmate Pharmaceuticals

ABSTRACT

The current studies were designed to evaluate, in preclinical studies, the immunologic

and therapeutic effects of intratumoral (IT) delivery of a novel virus-like particle (VLP) as

a cancer immunotherapy. CMP-001 is a VLP composed of the Qβ bacteriophage

capsid protein encapsulating an immunostimulatory CpG-A oligodeoxynucleotide TLR9

agonist. The impact in vitro of CMP-001 on human and murine cells, and the in vivo

impact of IT CMP-001 on syngeneic tumors in immunocompetent mice, was evaluated.

Additional studies evaluated the role of anti-Qβ antibody, and the addition of anti-PD1

antibody, on both the therapeutic effect of CMP-001, and the role T cells play in that

response. In vitro, the immunostimulatory effects of CMP-001 were dependent on anti-

Qβ. In vivo, IT CMP-001 treatment of murine A20 lymphoma enhanced survival, and

reduced tumor growth of both injected and contralateral noninjected tumors. IT CMP-

001 augmented T cell infiltration in injected A20 tumors. Depletion of T cells eliminated

the CMP-001-induced anti-tumor effects. Systemic anti-PD-1 enhanced the anti-tumor

response of both injected and noninjected contralateral tumors. CMP-001 had similar

effects in a unilateral B16 melanoma model. We conclude IT CMP-001 induces a

robust anti-tumor T cell response in an anti-Qβ dependent manner and results in

systemic anti-tumor effects that are enhanced by anti-PD-1 in two murine models. Early

phase clinical evaluation of the combination of CMP-001 and anti-PD1 is ongoing based

in part on these results.

Cellular vaccines employ APCs, typically dendritic cells (DCs), to deliver tumor antigens to patients that result in anti-tumor T cell responses. However, DCs are difficult to isolate and differentiate in vitro. B cell vaccines (Bvac) have advantages in addressing these challenges. Easily isolated in large numbers from blood, we demonstrated that in vitro-activated B cells are effective APCs, inducing strong antigen-specific CD8 T cell responses.

We applied the Bvac system to the B16 mouse melanoma model and demonstrated that Bvac presenting melanoma tumor cell lysates provide an effective antigen stimulus, resulting in statistically significant survival compared to tumor challenged naive mice and Bvac pulsed with three known melanoma expressing peptides. This finding demonstrates that known tumor antigenic peptides are not required, enhancing Bvac clinical applicability and versatility.

Recent In vitro assays have demonstrated that Bvac upregulate extracellular receptors typical of extravasation and homing to secondary lymphoid organs. In migration experiments Bvac migrated to the lymph node chemokines CCR19 and CCR21; concurrently CD4 and CD8 T cells migrated into Bvac conditioned media. Finally, T cell activation assays of co-cultured Bvacs with naive CD4 or CD8 results in rapidly induced T cell effector and activated populations. These findings indicate that B cell cancer vaccines are worth further exploration as a therapeutic option.

Abstract title: B lymphocyte antigen-presenting cell (APC) vaccines are effective in a mouse melanoma model

Authors (as submitted by poster presenter):Kyp L. Oxley, Depts. of Microbiology & Internal Medicine, University of Iowa and VA Medical Center, Iowa City, IABrett M. Hanson, Depts. of Microbiology & Internal Medicine, University of IowaAshley N. Zani , Depts. of Microbiology & Internal Medicine, University of IowaGail A. Bishop, Depts. of Microbiology & Internal Medicine, University of Iowa and VA Medical Center, Iowa City, IA

Validation Studies of a Functional Genomic shRNA Screen Investigating Vemurafenib Resistance in BRAF V600E-Mutated Melanoma

Marion Vanneste, Tyler Foley, Lei Zhao, Charlotte Feddersen, Afshin Varzavand, Eliot Zhu, Rob Piper, Christopher Stipp, Adam Dupuy, Michael Henry

BACKGROUND: BRAF Mutations, which constitutively activate the MAPK pathway, are present in 50% of cutaneous melanomas. Vemurafenib, a small molecule inhibitor of the BRAF V600E mutation, elicited a 63% reduction in the risk of death and 74% reduction in risk of disease progression in a phase III clinical trial of patients with metastatic melanoma. However, acquired resistance to single-agent BRAF inhibitor therapy is common and the underlying mechanisms have yet to be fully elucidated. Through a combined effort of labs at the University of Iowa, a functional genomic shRNA screen was conducted to identify loss-of-function mutations causing vemurafenib resistance in BRAF V600E-mutated A375 melanoma cells.

METHODS: A375 cells were transduced with lentiviral ShRNA constructs targeting the 9 selected genes. Dose response experiments were performed to evaluate short-term response of cell lines to Vemurafenib (72h). Emergence of acquired drug resistance was evaluated in a long term growth assay were cumulative population doubling was calculated over a 70 days period.

RESULTS: Expression of NF1, CUL3, SUV420H1 and TAOK1 was successfully reduced and further validated. Development of the ShRNA cell lines for the 5 additional genes (TF, FJX1, FADS2, ZYG11B, ERBB3) is still in process. Only CUL3 KD significantly decreased sensitivity of cells to Vemurafenib in our dose response experiment while both NF1 and CUL3 KD conferred resistance to the drug before control cells in the long-term growth assay. However, SUV420H1 and TAOK1 KD failed to confer resistance in both assays.

CONCLUSION: Only CUL3 and NF1 KD conferred resistance to Vemurafenib in A375 melanoma cells. Although NF1 and CUL3 had been previously implicated in similar shRNA or CRISPR-Cas9 screens, their identification in our screen validated our approach. Validation of additional hits is still in process, hoping to identify novel mutational markers in the development of resistance to Vemurafenib.

Authors (as submitted by poster presenter):Marion Vanneste / Molecular Physiology and BiophysicsTyler Foley / University of Iowa, Carver College of MedicineLei Zhao / Molecular Physiology and BiophysicsCharlotte Feddersen / MSTP, Anatomy and Cell BiologyAfshin Varzavand / BiologyEliot Zhu / MSTP, Cancer BiologyRob Piper / Molecular Physiology and BiophysicsChristopher Stipp / Biology, Molecular Physiology and BiophysicsAdam Dupuy / Anatomy and Cell Biology, Cancer BiologyMichael Henry / Molecular Physiology and Biophysics, SynderBio

Randomized, placebo-controlled, double-blind P2b trial of GC4419 to reduce severe radiation-related oral mucositis (SOM) in oral cavity/oropharynx pts

Carryn M. Anderson, Christopher M. Lee, Deborah Saunders, Amarinthia Curtis, Neal Dunlap, Chaitali Nangia, Arielle S. Lee, Jon Holmlund, Jeffrey Mark Brill, Stephen T. Sonis, John M. Buatti; University of Iowa Hospitals and Clinics, Iowa City, IA; Cancer Care Northwest, Spokane, WA; Northeast Cancer Center/Health Sciences North, Sudbury, ON; Spartanburg Medical Center, Spartanburg, SC; University of Louisville/James Graham Brown Cancer Center, Louisville, KY; UC/Irvine Medical Center, Orange, CA; HOPE Cancer Center of East Texas, Tyler, TX; Galera Therapeutics, Inc, Malvern, PA; BioModels, Boston, MA

Purpose/Objectives: Intensity-modulated radiotherapy (IMRT) plus cisplatin is established treatment for

locally advanced OC/OP cancer, but appx. 70% of patients develop SOM, defined as WHO Grade 3 or 4,

which limits patients' ability to eat solids (Gr 3) or liquids (Gr 4, requiring artificial nutrition). An RT-

induced burst of superoxide initiates oral mucositis (OM) development.

GC4419, a superoxide dismutase mimetic, interrupts this process by potently converting superoxide to

H2O2. It showed promising reductions of SOM in a published open-label Phase 1b/2a trial (IJROBP 1 Feb

2018).

Materials/Methods: 223 pts with OC or OP cancer receiving 70 Gy IMRT (>50 Gy to > 2 oral sites) plus

cisplatin (qwk or q3wk), were randomized 1:1:1 to PBO, 30 or 90 mg of GC4419, by 60-minute IV

infusion, M-F before each RT fraction. OM by the WHO scale was assessed by trained evaluators biw

during RT & qwk for up to 8 wks post RT. Primary endpoint was duration of SOM. Secondary endpoints

included incidence & time to onset of SOM. Analyses (each active dose v PBO, ITT) proceeded by a

sequential, conditional approach; 2-sided α=0.05.

Results: 90 mg GC4419 reduced SOM across endpoints, including a statistically significant reduction in

the primary endpoint of duration. Efficacy results with 30 mg were intermediate and did not reach

significance. Baseline patient & tumor characteristics, & treatment delivery, were well-balanced. Safety

was comparable across arms with no significant GC4419-specific toxicity or increased toxicity of

IMRT/cisplatin.

PBO 30mg 90mg 90mg vs. PBO

N 74 73 76 Relative δ p=

Duration SOM, median days 19 8 1.5 92% 0.024

Incidence SOM thru 60 Gy 58% 40% 37% 36% 0.010*

Incidence SOM thru last RT 65% 60% 43% 34% 0.009*

Incidence Grade 4 OM 30% 21% 16% 47% 0.045*

Onset SOM, median days 39 47 61 56% 0.080*

*nominal p value, pre-specified secondary endpoint

Conclusions: GC4419 provides a clinically meaningful reduction of SOM in terms of duration, incidence

and severity (Grade 4), with a safety profile comparable to placebo.

Cancer cells have increased steady state levels of reactive oxygen species (ROS; O2•-

and H2O2) compared to normal cells. It has been proposed that using redox active agents that further increase ROS levels will result is selective cancer cell death. This metabolic frailty can be targeted using drugs deemed safe for human use, ascorbate (ASC) and disulfiram (DSF), via a mechanism of H2O2 production. CuATSM is a drug being used in clinical trials to treat ALS disease. Imaging studies have shown that CuATSM preferentially concentrates in hypoxic tissues, releasing its Cu after entering cells. Copper (Cu) can participate in oxidation reactions that result in highly toxic hydroxyl radicals. Tumors often have areas of hypoxic tissue that exhibit resistance to ionizing radiation (IR). Our hypothesis is that CuATSM can be used to sensitize hypoxic regions of tumors to IR, ASC and/or DSF. Method: H2O2 flux after addition of ASC or DSF was measured in small cell lung cancer (SCLC) cells DMS53 using the 3-aminotriazole method. The Cu uptake of these cells was measured in normoxia and hypoxia after treatment with CuATSM. DMS53 and DMS273 (SCLC lines) were treated with ASC, CuATSM, and/or DSF+CuSO4 at varying oxygen tensions with and without IR and clonogenic assays were performed. Female nude mice bearing DMS273 xenografts were treated with CuATSM 30 mg/Kg/day oral gavage, ASC 4 g/Kg/day i.p., 90Y-DOTATOC 20 MBq tail vein on day one, or 90Y-DOTATOC + 3 doses of ASC. Average tumor volumes and tumor Cu concentration was measured. Results: Pharmacologically relevant dosing of DSF and ASC resulted in significantly higher fluxes of H2O2 compared to control. Both DSF and ASC enhanced IR clonogenic cell death in SCLC lines. Treatment with CuATSM resulted in increased intracellular Cu and enhanced cell death from ASC and IR in hypoxic conditions. Oral CuATSM resulted in elevated Cu levels in tumor xenografts and the combination of ASC + 90Y-DOTATOC resulted in slower tumor growth however the mice lost significant weight. Conclusion: These observations support the hypothesis that the differences in steady-state level of ROS in small cell lung cancer cells can be exploited to develop effective therapies using ASC and DSF. Furthermore CuATSM can enhance responses in hypoxic tumor tissues to both DSF and ASC. Significant elevation of intratumoral Cu in can be reached after oral administration of CuATSM demonstrating its potential as an anticancer agent. Most importantly, the addition of ASC to targeted neuroendocrine therapy may represent a translatable strategy for neuroendocrine tumors. (Supported by Neuroendocrine Spore Developmental grant P50 CA174521, T32 CA078586, P30 CA086862 and Carver Research Program of Excellence in Redox Biology and Medicine.)

Abstract Title: Enhancing small cell lung cancer therapy responses via disruption of copper ion & peroxide balance

Authors (as submitted by poster presenter):Melissa A Fath, University of Iowa, Radiation OncologySebastian J. Sciegienka/ University of Iowa Free Radical and Radiation Biology Samuel Rodman/ University of Iowa Free Radical and Radiation BiologyAnn Tomanek-Chalkley/ University of Iowa Free Radical and Radiation BiologyDongyoul Lee/ University of Iowa Department of Radiology Collin Heer/ University of Iowa Free Radical and Radiation BiologyDijie Li/u Pediatric OncologyMoustafa Gabr/ University of Iowa Department of ChemistryKelly Falls/ University of Iowa Free Radical and Radiation BiologySue O'Dorisio/Pediatric OncologyF. Christopher Pigge/ University of Iowa Department of ChemistryDouglas R. Spitz/ University of Iowa Free Radical and Radiation Biology

Background: One mechanism by which to selectively enhance the sensitivity of cancer cells toward

standard-of-care treatments is targeting cancer cell mitochondrial oxidative metabolism. It has been

proposed that cancer cells increase the production of reducing equivalents in the form of NADPH in

order to combat increased levels of O2•- and H2O2, relative to normal cells. Phosphorylation of NAD by

NAD kinase is the sole source of NADP(H). The two primary sources of cellular NAD are the de novo and

salvage pathways. Cancer cells rely more heavily (relative to normal cells) on the salvage pathway driven

by nicotinamide phosphoribosyltransferase (NAMPT) as a source of NAD. We hypothesize targeting of

NAMPT will selectively decrease NAD(H) and NADP(H) levels in cancer cells, resulting in selective

metabolic oxidative stress, cell death, and sensitization to agents that induce oxidative stress.

Methods: Clonogenic survival of Cal27 head and neck cancer cells, H1299 and DMS273 lung cancer cells, and normal human fibroblasts (NHFs) was determined following exposure to 1-100 nM FK866 (NAMPT inhibitor) for 72 h with or without 10-20 pmol cell-1 ascorbate for 1 h or 1 mM nicotinamide mononucleotide (NMN) for 72 hours. Doxycycline (1 µg/mL) treatment was used to induce catalase expression in H1299T cells containing a Tet-inducible promoter upstream of the human catalase gene. Cellular NAD(H) and NADP(H) as well as GSH content were determined by MTT or DTNB-reduction-based spectrophotometric assays, respectfully.

Results and Conclusions: Treatment with NAMPT inhibitor FK866 (1-50 nM) results in clonogenic cancer

cell death while having no effect on NHFs. FK866 treatment induces metabolic oxidative stress in cancer

cells as evidenced by decreases in cellular NAD(H) and NAPD(H) levels accompanied by a decrease in

total glutathione (GSH) and increase in glutathione disulfide (GSSG). Cancer cell death induced by FK866

treatment is completely abrogated by NMN supplementation. In addition, FK866 treatment

preferentially sensitizes cancer cells to AscH- in a manner that is greater-than-additive and dependent

on H2O2. Based on these findings, we propose that specific targeting of NAMPT has the potential to

selectively deplete NAD(H) and NADP(H) and induce metabolic oxidative stress in cancer cells, while

having little to no effect on normal cells. These results could provide a novel biochemical rationale for

developing new combined modality approaches for cancer therapy. (supported by T32 CA078586, P30

CA086862, R01 CA182804)

Abstract Title: Induction of metabolic oxidative stress and sensitization to standard-of-care therapies via targeting of NAD+ salvage pathway in cancer cells

Authors (as submitted by poster presenter):Collin D. Heer / Free Radical and Radiation Biology ProgramSebastian J. Sciegienka / Free Radical and Radiation Biology ProgramDavid B. Riffe / Free Radical and Radiation Biology ProgramKranti A. Mapuskar / Free Radical and Radiation Biology ProgramDouglas R. Spitz / Free Radical and Radiation Biology Program

A G1-lipid checkpoint regulates cell cycle dependent radiation therapy response ofhuman head and neck cancer

Ehab H. Sarsour, Amanda L. Kalen, Wafa AM Asha, Jyungmean Son, Yusuf Menda,John Buatti and Prabhat C. Goswami

Division of Radiation and Free Radical Biology; Department of Radiation Oncology, TheUniversity of Iowa, Iowa City, IOWA, USA

F-18 fluorothymidine (FLT) positron emission tomography is a non-invasive imagingmethod that is used to measure proliferation of tumors. Results from a Phase I trial(NCT00721799; University of Iowa) showed that head and neck squamous cell cancer(HNSCC) subjects with a higher pre-therapy tumor FLT uptake (high proliferative index;HPI) had a better outcome (16 subjects) compared to subjects with a lower pre-therapytumor FLT uptake (low proliferative index, LPI; 12 subjects), suggesting that the pre-therapy proliferative status of HNSCC is a critical determinant of therapy outcome.Results from HNSCC (Cal27, FaDu, and SCC25) cell lines showed that HPI (>60%S+G2+M) cultures are radiosensitive and LPI (>70% G0+G1) cultures from the same cellline are relatively radioresistant. Radiation sensitivity of HPI cultures was associated withhigher metabolic activities and lower antioxidant enzyme levels resulting in a higheroxidation status (ROS: LOO●, O2●‒, and H2O2) compared to LPI cultures that exhibited alower oxidation status. Results from RNAseq analysis identified G0-G1 switch gene 2(G0S2) as a potential regulator for the differential radiation sensitivity of HPI and LPIcultures. G0S2 regulates progression (higher expression in G0+G1 compared to S+G2+M)and lipid metabolism (negative regulator of ATGL, a cytosolic neutral lipase) during thecell cycle. siRNA knockdown of G0S2 recruited more cells to the proliferative cycleresulting in radiosensitization. In contrast, inhibition of ATGL activity using atglistatinsuppressed radiation induced toxicity of Cal27 cells. Interestingly, HNSCC subjects witha higher compared to a lower pre-therapy tumor G0S2 expression correlated with asignificantly shorter survival (n = 270; p = 0.009; NCBI). These results support thehypothesis that G0S2 coordinates a G1-lipid checkpoint: G0+G1 cells prior to thischeckpoint are radioresistant, whereas S+G2+M cells that are beyond this checkpoint areradiosensitive.

Radiomic Features from Perinodular Parenchyma Enhance Lung Cancer Computer-aided

Diagnosis Tools

Johanna Uthoff (1,2), Matthew J. Stephens(3), John D. Newell Jr. (1,2), Eric A. Hoffman (1,2),

Jared Larson (2), Nicholas Koehn (2), Frank A. De Stefano (2), Chrissy M. Lusk (4), Angela S.

Wenzlaff (4), Donovan Watza (4), Christine Neslund-Dudas (7), Laurie L. Carr (5), David A.

Lynch (6), Ann G. Schwartz (4), Jesscia C. Sieren (1,2)

1. Department of Biomedical Engineering, University of Iowa, Iowa City, Iowa.

2. Department of Radiology, University of Iowa, Iowa City, Iowa.

3. Department of Radiology, University of Cincinnati, Cincinnati, Ohio

4. Karmanos Cancer Institute, Wayne State University, Detroit, MI

5. Department of Medicine, National Jewish Health, Denver Colorado

6. Department of Radiology, National Jewish Health, Denver, Colorado

7. Department of Public Health Sciences, Henry Ford Health System, Detroit, MI

Rationale: With growing use of low-dose computed tomography for lung cancer screening, the

detection frequency of lung nodules has increased dramatically. Non-invasive distinction of

malignant from benign nodules will result in notable reduction in procedure-related morbidity

and healthcare costs.

Objectives: To examine the benefit of including perinodular parenchymal features in computer-

aided diagnosis (CADx) tools for pulmonary nodule assessment.

Methods, Measurements, and Main Results: Lung nodule cases with pathology confirmed

diagnosis (74 malignant, 289 benign) were used to extract quantitative imaging characteristics

from computed tomography scans of the nodule and perinodular parenchyma tissue. A CADx

development pipeline was employed using k-medoids clustering and information theory to

determine efficient predictor sets for different amounts of parenchyma inclusion and build an

artificial neural network classifier The resulting CADx tool was validated using an independent

cohort (50 malignant, 50 benign). The inclusion of parenchymal imaging features produced

statistically better CADx tools over exclusively nodular features (p <0.001). The best performing

CADx (AUC-ROC =1.0) included features derived from nodule diameter-based surrounding

parenchyma tissue quartile-bands. We demonstrate similar high performance values on the

independent validation cohort (AUC-ROC =0.965). A comparison using the independent

validation cohort with the Fleischner pulmonary nodule follow-up guidelines demonstrated a

theoretical reduction in recommended follow-up imaging and procedures.

Conclusions: Radiomic features extracted from the parenchyma surrounding lung nodules

contain valid signals with spatial relevance for the task of lung cancer risk classification.

Through optimization of feature extraction regions from the parenchyma, CADx validation

performance of 100% sensitivity and 96% specificity was achieved.

To determine if mitochondrial pyruvate transport could represent a therapeutic target for sensitizing cancer cells to oxidative stress, lung and breast cancer cells were treated with 5 µM UK5099 to inhibit the mitochondrial pyruvate carrier (MPC). Treatment with UK5099 selectively sensitized lung and breast cancer cells to clonogenic cell killing when combined with depletion of glutathione using 1 mM buthionine sulfoximine (BSO; a glutathione synthesis inhibitor) for 48 h and 72 h, relative to normal lung and breast epithelial cells. Furthermore, cancer cell killing mediated by UK5099 combined with BSO was inhibited by the thiol antioxidant, N-acetylcysteine (NAC; 20 mM), independent of GSH levels indicating a mechanism of toxicity involving reduced thiols and metabolic oxidative stress. In addition, treatment with UK5099 alone for 48 h also decreased levels of total glutathione in cancer cells that could be reversed by NAC. Using oxidation sensitive fluorescent dyes (CDCFH2 and MitoSOX), treatment of lung and breast cancer cells for 24 h and 48 h with UK5099 induced increases in steady state levels of pro-oxidants (presumably hydroperoxides and mitochondrial superoxide) which were further increased with BSO. Finally, treatment of breast cancer cells with UK5099 for 24 and 48 hours significantly sensitized breast cancer cells to clonogenic cell killing mediated by paclitaxel. These data support the hypothesis that inhibition of the MPC selectively causes an impairment of antioxidant capability in cancer cells that is enhanced by depletion of glutathione. Furthermore, these results also support the hypothesis that inhibition of the MPC represents a significant target for sensitizing human breast cancer cells to chemotherapy agents thought to induce oxidative stress. (Supported by R01CA182804.)

Abstract Title: Inhibition of mitochondrial pyruvate transport selectively sensitizes cancer cells to metabolic oxidative stre

Authors (as submitted by poster presenter): Shane R Solst / Radiation OncologySamuel N Rodman / Radiation OncologyMelissa A Fath / Radiation OncologyEric B Taylor / BiochemistryDouglas R Spitz / Radiation Oncology

Radiomic biomarkers of breast cancer in screening mammography aided by peri-lesional signal

Breast cancer is the most common cancer diagnosis for women in the United States. Annual breast cancer

screening with mammography is recommended for average-risk women starting at the age of 40. False-

positive lesions in screening can lead to unnecessary follow-up procedures and stresses on the patient.

Computer-aided diagnosis (CADx) tools can offer an independent and objective risk assessment to be

used in conjunction with radiologist assessment to further decrease the false-positive rate. Quantitative

imaging features of intensity, texture, and shape were extracted from breast lesions and surrounding tissue

in 287 mammograms (150 malignant, 137 benign). A feature set reduction method to remove highly intra-

correlated features was devised using k-medoids clustering and k-fold cross validation. A novel feature

selection method using information theory was introduced which builds a feature set for classification by

determining a group of class-informative features with low set co-information. An artificial neural

network was built from the selected feature set using 10-hidden layer nodes and the tanh activation

function. The resulting CADx tool achieved a training accuracy of 96.2%, sensitivity of 97.6%,

specificity of 95.2%, and area-under-the-curve of 0.971 along with 97.1% sensitivity and 94.9%

specificity a blinded validation set.

Authors (as submitted by poster presenter):Johanna Uthoff / Radiology and Biomedical EngineeringMaheen Rajput M.D. / RadiologyJessica C. Sieren Ph.D. / Radiology and Biomedical Engineering

Optimization of selenium in cell culture media to maximize selenoenzyme activity

Jeffrey M. Stolwijk1, Kelly Falls1, Brett A. Wagner1, Michael McCormick1, Yousef Zakharia1, Douglas Spitz1 and Garry R. Buettner1 1University of Iowa, Iowa City, IA

Cell culture is a widely-used approach to examine basic biochemical mechanisms, including the biochemistry and biology of selenium (Se). Se is an essential trace element needed for optimal health. It has long been known that most cell culture is deficient in Se, which can limit expression of Se-proteins. Thus, supplementation of cell culture media with Se should always be a consideration. However, this supplementation must be within a relatively narrow range to overcome deficiency and yet not induce toxicity. The window to avoid potential toxicity can be widened by using seleno-L-methionine (SLM) instead of inorganic forms of Se, such as selenite or selenate. However, the optimal level of supplementation has not been fully characterized.

Many of the 25 known Se-proteins have central roles in the redox biochemistry of cells and tissues, e.g. the glutathione peroxidases (GPx) and thioredoxin reductases (TrxR). In a wide variety of cell-types we have seen dose-dependent increases in both GPx protein expression and enzyme activity when media are supplemented with SLM. However, the different members of the GPx family have different dose-response relationships. We have observed that when over-expressing Se-enzymes, using adenoviral techniques, it is especially important to supplement media with Se to maximize Se-enzyme activity. There is evidence for a hierarchy for Se-utilization among the selenoproteins. This hierarchy appears to extend to GPx1 and GPx 4; first results examining this issue will be presented.

In a clinical trial at The University of Iowa, subjects received ~30x the RDA (55 µg d-1) of Se as SLM daily for more than 7 mo. The level of total Se in blood increased about 50-fold. However, the activity of GPx1 in the blood remained constant. This points to the importance of supplementing cell culture media with Se to maximize the activity of GPx1. This simple maneuver will increase the rigor and reproducibility of experiments addressing basic redox biology.

Qualitative Analysis of Rectal Cancer Patients’ Decisions on Where to Receive Surgery in a Rural State

AUTHORS: Mary Charlton1,2, Ariana Shahnazi3, Irena Gribovskaja-Rupp4, Chi Lin5, Elizabeth A. Chrischilles1, Charles F. Lynch1,2, Lisa Hunter2, and Marcia M. Ward6

1Department of Epidemiology, University of Iowa College of Public Health, Iowa City, IA 2Iowa Cancer Registry, University of Iowa College of Public Health, Iowa City, IA 3Department of Communications, University of Iowa College of Liberal Arts and Sciences, Iowa City, IA 4Department of Surgery, University of Iowa Carver College of Medicine, Iowa City, IA 5Department of Radiation Oncology, University of Nebraska Medical Center, Omaha, NE 6Department of Health Management and Policy, University of Iowa College of Public Health, Iowa City, IA

Presenter email address: mary-charlton@uiowa.edu

BACKGROUND: Current literature suggests surgeons and hospitals that perform large volumes of rectal cancer care achieve superior outcomes, but only about half of rectal cancer resections are performed by high-volume surgeons in comprehensive hospitals. Little is known about considerations of patients with rectal cancer when deciding where to receive surgery, particularly among those residing in rural areas.

METHODS: A purposive sample of stage II/III rectal adenocarcinoma survivors diagnosed 2013-2015 was identified through the Iowa Cancer Registry and interviewed by telephone about factors influencing decisions on where to receive rectal cancer surgery. Interviews were recorded and transcribed, and a thematic analysis was conducted using a multiple coding approach among research team members.

RESULTS: Thematic saturation was reached after interviewing 15 survivors; their mean age was 63; 60% were male, 53% resided in non-metropolitan areas and 60% received surgery at low-volume centers. Almost all patients considered surgeon volume and experience to be important determinants of outcomes, but few actually assessed it. Most characterized surgeon experience based on subjective assessments including interpersonal skills, ability to explain the procedure, and perceived confidence of the surgeon. Recommendation from a trusted source, usually a physician, appeared to be a main driver of where patients received surgery. Patients who chose low-volume centers noted comfort and familiarity as important decision factors. Several patients reported not trusting online sources for information about treatment.

CONCLUSIONS: Most rectal cancer patients in our sample relied on physician referrals to decide where to receive surgery. Further research is needed to determine rectal cancer patients’ preferences for obtaining information about surgeon/hospital volume and experience since our findings suggest they are neither discussing these factors with their surgeon nor researching them on their own. Once preferences are determined, targeted interventions facilitating more informed decision-making by patients can be developed.

Gut Check: The Gastrointestinal Microbiome of Solid Tumor Patients Compared to Healthy Controls, A Feasibility Study of a Stool Collection Protocol

Catherine Cherwin, PhD, RN

University of Iowa, College of Nursing Catherine-cherwin@uiowa.edu

319-335-7654

Back ground: An imbalance in the florae of the gut, the GI microbiome, has been associated with

diseases like inflammatory bowel disease and colorectal cancer and may lead to high GI symptom

burden (such as constipation, diarrhea, gas, and bloating). Chemotherapy can inhibit replacement of GI

epithelial cells, compromising the protective GI barrier. Few microbiome studies have explored

microbiome changes in people with cancer. More research is needed to describe potential

chemotherapy-related GI microbiome changes, including in people with solid tumors. People with

cancer receiving cytotoxic chemotherapy, including platinum-based chemotherapy, can experience a

high symptom burden and stool collection procedures may be overly complex and difficult for them to

complete. Further, social stigma associated with collecting stool samples my impact study recruitment

rates.

Purpose: The purpose of this study was to determine feasibility of a stool collection protocol for GI

microbiome analysis in people with breast or lung cancer receiving platinum-based chemotherapy.

Methods: N=10 patients with breast or lung cancer receiving ≥ cycle 3 of platinum-based chemotherapy

were recruited to provide stool samples following chemotherapy administration. Participants also

completed a feasibility questionnaire. Feasibility of study protocol was evaluated through rate of

recruitment, attrition, and responses given on the feasibility questionnaire.

Results: In all, N=10 provided signed consent and n=9 provided stool samples and questionnaire data

(90%). A majority of the sample reported that the collection was acceptable (88.9%) and with few

complications. Problems reported by participants included: difficulty timing bowel movements, needing

to urinate during the sample collection, and difficulty using the stool collection kit.

Conclusions: Results of this study indicate that stool sample collection is feasible in people with cancer

receiving chemotherapy with regard to ease and acceptability of sample collection but not with regard

to rate of participant recruitment. To improve protocol feasibility, eligibility criteria will be expanded to

include other solid tumor diagnoses receiving cytotoxic chemotherapy, and to other cytotoxic

chemotherapies with high symptom burden.

Rural-urban differences in treatment at high-volume hospitals among rectal cancer

patients

AUTHORS: Catherine Chioreso1, Mary Charlton1,2, Irena Gribovskaja-Rupp3, Chi Lin4, Marcia

M. Ward5, Charles F. Lynch1,2, Elizabeth A. Chrischilles1

1Department of Epidemiology, University of Iowa College of Public Health, Iowa City, IA 2Iowa Cancer Registry, University of Iowa College of Public Health, Iowa City, IA 3Department of Surgery, University of Iowa Carver College of Medicine, Iowa City, IA 4Department of Radiation Oncology, University of Nebraska Medical Center, Omaha, NE 5Department of Health Management and Policy, University of Iowa College of Public Health,

Iowa City, IA

Presenter contact information:

Mary Charlton, PhD

Email: mary-charlton@uiowa.edu

Phone: 319-384-1564

Background: Hospitals performing high volumes of rectal cancer resections achieve superior

rates of sphincter preservation and survival, but many rectal cancer resections are still performed

in low-volume centers, particularly among rural populations. Our objective was to determine

factors associated with receiving surgery at high-volume hospitals (HVHs).

Methods: Data were extracted from the SEER-Medicare database for beneficiaries (age 66+)

with stage II/III rectal adenocarcinoma diagnosed 2007-2011 who received rectal cancer-directed

surgery. Hospital volume was classified by the number of rectal cancer resections in Medicare

claims from 2007-2011. Rural-Urban Commuting Area codes were used to classify patients as

urban or rural. Travel times were calculated from the patient’s zip code to: 1) hospital where they

received surgery, and 2) nearest HVH. Logistic regression was used to determine factors

associated with travel time and receiving surgery at HVHs.

Results: Of 2,053 patients, 21% were rural and 79% were urban. Rural patients compared to

urban patients were less likely to have surgery at a HVH (43% vs. 61%; p<.0001). In order to

reach the nearest HVH, median travel time for rural and urban patients was 78 and 16 minutes,

respectively. Among rural patients, odds of receiving surgery at a HVH were significantly lower

for every 10-minute increase in travel time (OR=0.992; 0.988-0.997), and nearly significantly

higher for patients in areas with higher percentages of college-educated people (OR=1.024;

0.998-1.051). Those who received surgery in HVHs more often had guideline-recommended

staging (TRUS/MRI) and neoadjuvant chemoradiation.

Conclusions: Compared to urban patients, rural patients were less likely to travel to HVHs,

which had substantially greater travel times. Given receipt of surgery at HVHs is associated with

guideline-recommended rectal cancer management, further research is needed to understand

barriers to receiving care at HVHs.

Introduction: An elevated lifetime risk of breast cancer of ≥20% should prompt evaluation for high risk

screening or risk reduction. Atypical (ductal or lobular) hyperplasia (AH) and lobular carcinoma in situ

(LCIS) are associated with a 4-8 fold increase in breast cancer risk. It is unclear how often these lesions

found on workup of abnormal imaging influence patients’ breast cancer risk management.

Methods: The institutional pathology database was queried for AH or LCIS without cancer found on core

needle biopsy (CNB) or excisional biopsy (ExB) from 2013-2016, and chart review was performed.

Univariate and multivariate analysis examined factors predicting recommendations.

Findings: Of 56 patients identified, 13 had LCIS and 42 had AH. Most (98.2%) were referred to breast

clinic. Recommended interventions included: high risk screening with MRI (37.5%), genetic testing

(16.1%), risk reducing medication (RRM) (39.3%) and prophylactic mastectomy (5.3%). Lifetime risk of

breast cancer was calculated using a risk model for 26/56 (46.4%). On multivariate analysis having a risk

model calculated was independently associated with receiving recommendation of high risk screening

with MRI (p=0.00). Recommendation of RRM were associated with a higher level of lifetime risk as

calculated by risk model (average risk estate 28.1% vs 29.6%, p<0.04). Genetic testing identified a

deleterious high risk mutation in 1/6 patients tested. Three (5.4%) patients developed subsequent

breast cancer over a mean follow up period of 2.2 years.

Conclusions: Identification of atypia on CNB or ExB resulted in changes in breast cancer risk

management recommendations for 57.1% (32/56) of patients. Establishment of a systematic algorithm

that includes routine use of breast cancer risk models to evaluate these patients should identify more

individuals who would benefit from high risk screening and risk reducing interventions.

Abstract Title: Management of breast cancer risk in patients with atypia: variability in a single academic institution

Authors (as submitted by poster presenter):Sophia L Fu, MD, MS, FACS/UIHCAnna Beck, MD/UIHCLeonel Vasquez, MD/UIHCAmani Bashir, MD/UIHCLillian Erdahl, MD, FACS/UIHCSonia L Sugg, MD, FACS/UIHCRonald J Weigel, MD, FACS/UIHCIngrid Lizarraga, MBBS, MD, FACS/UIHC

Use Patterns of a Symptom Management Website for Patients in Rural Areas

Background: Patients with advanced cancers may experience multiple distressing symptoms

associated with their cancer or it’s treatments. Patients living in rural areas have decreased access

to cancer symptom management specialists such as supportive oncology and palliative care

services. Web-based interventions have the potential to close this access gap. Oncology

Associated Symptoms and Individualized Strategies (OASIS) is a web-based intervention for

patients living in rural areas designed to provide symptom management education and tools to

improve symptom control. One strategy OASIS employs is the use of a symptom diary to help

patients better understand potential triggers and mitigating factors for their symptoms. However,

it is not known how patients in rural areas will engage with this type of web-based intervention.

Purpose: The purpose of this feasibility study is to evaluate website use patterns of patients

enrolled in OASIS. Specifically, what pages they visit and how often as well as what symptoms

and strategies they choose to track in the diary.

Methods: A sample of n=36 patients will be enrolled. Eligibility includes over 18 years old,

diagnosis of advanced cancer, and living in a rural zip code. Data will be extracted from the

OASIS website user logs. A summary of the page visits and symptom diary entries were

generated. Frequencies and means were calculated. Differences in use patterns based on age, sex,

and internet fluency were evaluated.

Results: N=13 patients completed at least one week of the intervention. Mean age was 68 years,

majority white, and majority men. 135 entries were made in the daily tracker. Average number of

tracking 7.5x/week for all active participants. Participants engage with the site for an average of 4

weeks. Numbness & tingling, fatigue, memory problems, depression, constipation, and lack of

appetite were the most commonly tracked symptoms. The mostly commonly tracked strategies

were sleep hygiene and nutrition. A wide range of sub-strategies were used by participants

Conclusions: The results from this study will be used to refine the OASIS intervention to meet

the needs of these patients. Future research is needed to evaluate the efficacy of the OASIS

intervention on symptom severity and distress.

Authors (as submitted by poster presenter):Stephanie Gilbertson-White, PhD, APRNSeyedehtanaz Saeidzadeh, MSN, RN / College of NursingChi Yeung, MA / College of Education, Department of Counseling Psychology Todd Papke, PhD / College of Nursing

Presenter:    Sloane  H.  Henry,  MA,  sloane-­‐henry@uiowa.edu,  319-­‐512-­‐9968  Other  Authors:      

       Lindsay  Schultz,  BA,  schultz@canceriowa.edu          Kelly  W.  Sittig,  CCPH,  sittig@canceriowa.edu  

Title:    The  role  of  the  Iowa  Cancer  Consortium  in  reducing  the  burden  of  cancer  in  Iowa  using  the  Iowa  Cancer  Plan  as  a  roadmap  

The  Iowa  Cancer  Consortium  is  a  statewide  nonprofit  coalition  of  health  care  providers,  public  health  professionals,  caregivers,  researchers,  cancer  survivors,  volunteers  and  advocates  working  together  to  reduce  the  burden  of  cancer  in  Iowa.    The  Iowa  Cancer  Consortium  convenes  partners  and  stakeholders  in  the  state  of  Iowa  using  a  variety  of  methods  including  statewide  workgroups,  the  Iowa  Cancer  Summit,  partner  meetings,  networking  opportunities,  webinars  and  educational  materials  and  opportunities.      The  Consortium’s  network  also  extends  into  communities  where  the  need  is  often  the  greatest  by  supporting  work  being  done  by  local  partners  with  community  grants  through  staff  outreach.    Staff  collaborates  with  many  community  agencies  and  individuals  to  bring  quality  cancer  control  work  to  underserved  populations  such  as  rural,  Native  American,  LGBTQ,  African  American  and  Hispanic  populations.  A  major  component  of  the  Consortium’s  collaborative  work  is  the  2018-­‐2022  Iowa  Cancer  Plan.    This  plan  is  used  as  a  roadmap  or  guide  for  cancer  control  in  Iowa.    The  Iowa  Cancer  Plan  identifies  five  priority  areas  to  be  addressed  in  order  to  reduce  the  burden  of  cancer  in  Iowa.    The  five  priority  areas  are  prevention,  screening,  treatment,  quality  of  life  and  health  equity.    These  priorities  help  guide  cancer  control  work  being  done  throughout  the  state  by  our  many  partners  and  stakeholders.    The  goals  and  data  targets  found  in  the  Iowa  Cancer  Plan  can  also  serve  as  a  guide  to  help  address  policy  and  even  research  in  the  area  of  comprehensive  cancer  control.  Participation  in  the  Iowa  Cancer  Consortium  connects  cancer  control  professionals  and  researchers  with  a  network  that  extends  into  communities  across  the  state.  The  diverse  network  of  individuals  and  organizations  working  with  the  Iowa  Cancer  Consortium  will  further  help  reduce  the  burden  of  cancer  for  all  Iowans.

Background: Men with a low-risk prostate cancer (PCa) can opt for active surveillance (AS), a monitoring strategy deferring active treatment (AT) in the absence of disease progression. AS can minimize the harms of overtreatment, but historically most eligible men select AT. We evaluated the associations of decision-making factors (knowledge, decisional processes, and psychological factors), socio-demographic and clinical factors with selecting AS.

Methods: We enrolled 1139 men from Kaiser Permanente Northern California (KPNC) with a low-risk PCa (PSA < 10 ng/mL, Gleason < 7, stage ≤ 2a) diagnosed between 2012-14. We conducted telephone surveys within 30 days of diagnosis, collecting data onsocio-demographics and clinical history, PCa knowledge, psychological factors and decisional processes. We abstracted medical record data on comorbidities and tumor characteristics. We classified men as selecting AS if they remained continually enrolled in KPNC and did not undergo active treatment (surgery, radiotherapy, or hormone therapy) within a year of diagnosis. We used multivariable logistic regression analyses to identify factors associated with selecting AS. We used the c-statistic to assess predictive accuracy.

Results: We evaluated 1117 subjects, median age 62, 81% white, 82% married, 19% < any college education, 34% no comorbidities. During one-year follow-up, 639 (57%) opted for AS. Significant predictors for selecting AS were being aware of low-risk status (OR 1.7; 95% CI, 1.0, 3.0); knowing that AS was a treatment option (3.5; 1.6, 7.8); wanting to avoid treatment complications (1.2, 1.1, 1.4) a urologist recommending only AS (8.8; 5.0, 15.5); spouse/partner preferring AS/WW (4.6, 1.9, 11.2). Conversely, valuing cancer control (2.3; 1.3, 4.1), greater anxiety (1.5; 1.1, 2.1), greater decision confidence (2.2; 1.5, 3.1), and having higher PSA levels (1.2 per unit change; 1.1, 1.3), clinical stage (T2a vs. T1c, 2.2; 1.2, 4.1), and percent positive biopsy cores (>25% vs. <10%, 3.3; 2.2, 5.0) were associated with AT. The c-statistic for the model with socio-demographic and clinical variables was 0.71; the c-statistic was 0.87 for the full model with decision-making factors.

Conclusions: A substantial proportion of subjects selected AT, and decisions were highly associated with tumor characteristics. However, decision-making factors also independently predicted treatment selection. Men selecting AS were more knowledgeable than those selecting AT about PCa prognosis and treatment options, and were more concerned about avoiding treatment harms. Although supported by urologists in selecting AS, these men were less confident in their decision than those selecting AT. Efforts to provide comprehensive early decision support to men with low-risk cancers may facilitate better informed decision making and potentially increase uptake of AS.

Abstract Title: Selecting active surveillance: decision-making factors for men with a low-risk prostate cancer

Authors (as submitted by poster presenter):Richard M. Hoffman/University of IowaTania Lobo/Georgetown UniversityStephen Van Den Eden/Kaiser Permanente Northern CaliforniaGeorge Luta/Georgetown UniversityKimberly Davis/Georgetown UniversityDavid AronsonKaiser Permanente Northern California\Kathryn Taylor/Georgetown University

TITLE: PERCH and ORIEN: Implementation of cancer center wide biorespository aimed to support national collaborative effort in driving research and discovery of personalized treatments for cancer

AUTHORS: Laura Jacobus1,2 MS CCRC, Ashley McCarthy1 MPH, George Weiner MD1,2, Kenneth Nepple1,2 MD

INSTITUTIONS: 1. Holden Comprehensive Cancer Center, the University of Iowa, Iowa City, IA United States2. Carver College of Medicine, The University of Iowa, Iowa City, IA United States

Email laura-jacobus@uiowa.edu

ashley-mccarthy@uiowa.edu

kenneth-nepple@uiowa.edu

george-weiner@uiowa.edu

Phone 319-384-5154 319-467-5839 319-356-2114 319-353-8620

Background: Late in 2017, Holden Comprehensive Cancer Center (HCCC) joined the Oncology Research Information Exchange Network (ORIEN), a national initiative among leading cancer centers promoting collaboration to drive research and discovery of personalized treatments for cancer. ORIEN partners 17 U.S. cancer centers with M2Gen, a leading health informatics company, and pharmaceutical industry support. Patients treated at ORIEN member centers consent to share their clinical, biospecimens, and molecular data contributing to one of the world’s largest clinically annotated tissue repositories. The aim is to leverage big data and genomic technology across institutional boundaries to learn more about cancer and cancer treatment. ORIEN returns molecular data to researchers while also creating molecularly based ‘communities’ of patients with similar tumors and diseases in hopes of increasing opportunity and efficiency in clinical trial enrollment.

Methods: The implementation of ORIEN included the convergence of enrollment methods across previously independent biorepositories within HCCC. Thoughtful consideration of efficiency and the patient experience lead to the development of a single Unified Biorepository Consent which has been named ‘PERCH’ (Patients Enhancing Research Collaborations at Holden). To accommodate increased volume of patients approached for enrollment, a Research Appointment was created taking place prior to a patient’s clinical appointment. During this appointment, research opportunities, primarily related to biorespository contribution, are introduced without disrupting clinical workflow. New workflows were established through the collaboration of many members of HCCC. Participation in the national collaboration has been initially piloted in urologic cancer with a plan to expand to additional tumor sites over time. Importantly, the ORIEN project functions to expand the research of HCCC personnel, but not to replace any current efforts or projects.

Results/Conclusions: As of April 1, 2018 over 800 patients have been approached for enrollment and over 600 have agreed to participate in biorepository related projects the umbrella of PERCH, including ORIEN. In urologic cancer, 60 specimens have been contributed to ORIEN and return of molecular data will be forthcoming.

Head and Neck Cancer Survivorship from the Patient Perspective Head and Neck

Nicholas Kendell, MS; Aaron Seaman, PhD; Alan J Christensen, PhD; Timothy A Thomsen, MD; Nitin A Pagedar, MD, MPH

Background: Survivorship care is increasingly recognized as critical. Despite published guidelines, many aspects of head and neck cancer survivorship care lack both an evidence base and understanding of patient priorities. The UI Otolaryngology clinic instituted a survivorship clinic model in 2011. We sought to assess patients’ satisfaction with and priorities for UI survivorship care and to characterize the care provided by the clinic.

Methods: Patients with history of cancer of the upper aerodigestive tract, salivary gland, or head and neck skin; 18 months or more from last cancer-directed treatment; and seen in a surgeon’s clinic or the survivorship clinic were identified, approached, and given an anonymous survey at an office visit. Descriptive statistical analysis was performed using SAS version 9.4.

Results: There were 144 respondents, of whom 31% were women. Forty percent reported high school diploma or less; 37% reported college degree or greater. Half were 3 years or fewer from cancer treatment, and 28% were more than 5 years out. Most respondents identified as ‘very satisfied’ with their care, and 120, 86%, replied they were very satisfied with their access to the clinic resources. Evaluation for recurrent cancer was the most common priority for follow up, while help with cancer- or treatment-related symptoms and general health maintenance were the next most common priorities. 30% reported having received a survivorship care plan, and 72% reported little or no discussion of their cancer follow up with a primary care physician.

Discussion: Patients reported high levels of satisfaction with survivorship care. Findings indicate that although guidelines place great emphasis on health maintenance, survivors’ focus remains on cancer and recurrence, even years after treatment. Guidelines recommending primary care delivery of survivorship care are unlikely to be successful given low level of primary care involvement in follow up.

Plans for Future Work: This study is ongoing, and we aim to double the size of the cohort. We have used this study to build a cohort with whom we are planning two studies: one focusing on quality of life and the effect of rurality, and a second, qualitative one to examine the patient and caregiver survivorship experience.

Authors (as submitted by poster presenter):Nitin A Pagedar, MD, MPHAaron Seaman, PhDNicholas D Kendell, MSAlan J Christensen, PhDTimothy A Thomsen, MD

Communication Preferences for Breast Cancer Patients

Skillful communication is at the heart of the patient-centered care model. With this in mind, there has been increased interest in improving clinician skills in delivering bad news to patients. The aim of this study was to determine the preferred method of receiving test results in a sample of breast cancer patients, using a questionnaire designed by the investigators.

Patients aged 18 and over with a diagnosis of breast cancer or DCIS seen at the Holden Comprehensive Cancer Center between 6/12/17 -10/13/17 were offered a questionnaire at the time of check-in to their clinic visit. Effects of patient and disease characteristics on most preferred method of communicating test results were evaluated using multinomial logistic regression models.

Out of 582 patients offered a questionnaire, 452 (78%) were returned and 395 (68%) were deemed adequate for analysis. Most patients preferred a call ASAP regardless of the type of test (biopsy or imaging) or whether the result was good or bad news. Patients with Stage IV cancer were more likely to want in-person communication than patients with Stage I-III cancer for all types of results. Those with no college education were less likely than those with at least some college education to want a message via the electronic patient portal for good news on an imaging test (OR 0.21, CI 0.08-0.55) or a biopsy test (OR 0.32, CI 0.12-0.83). For imaging results, as the age of the patient increased, the odds of preferring in-person communication relative to a phone call ASAP increased (OR 1.04, CI 1.01-1.07). Compared to those with children, patients without children were more likely to want in-person results (OR 2.26, CI 1.17-4.35) or a scheduled call (OR 6.19, CI 1.35-28.41) for bad news on an imaging test versus a call ASAP. Most patients had no preference for any particular day of the week or time of day for a phone call.

These results highlight that patients largely prioritize getting results as quickly as possible but that communication preferences may differ depending on factors such as stage of breast cancer, education level, and family type. Thus, each patient should be asked about their preferences for communication at the initial visit and perhaps again each time a test is ordered, as their preferences may vary in response to personal circumstances.

Authors (as submitted by poster presenter):Sneha Phadke/University of IowaBradley McDowell/Holden Comprehensive Cancer CenterNajla Itani/University of IowaNicole Grogan/University of IowaMark Vander Weg/University of IowaTim Ginader/Holden Comprehensive Cancer CenterSarah Bell/Holden Comprehensive Cancer Center

Survival benefit of obesity in stage IV colorectal cancer: Better tolerability of chemotherapy?

Catherine G. Tran, MD1, Braden S. Jensen, BS1, Elise E. Hill, BS1, Austin C. Stark1, Meghan E. Flannery, MD1, Daniel J. Berg, MD2,3, Carlos H.F. Chan, MD, PhD1,3 Departments of Surgery1 and Internal Medicine2, Holden Comprehensive Cancer Center3, University of Iowa Carver College of Medicine, Iowa City, IA, USA

Background: Obesity is a known risk factor for the development of colorectal cancer (CRC) and has long been associated with increased mortality of CRC. However, recent studies have shown that obesity is associated with improved cancer survival, a phenomenon termed “obesity paradox.” The purpose of this study is to revisit the impact of obesity on survival of stage IV CRC. Methods: Retrospective analysis of CRC patients presenting to the Holden Comprehensive Cancer Center between January 2006 and April 2016 was conducted under an approved IRB protocol. Clinicopathological and follow-up data were prospectively gathered in the institution’s cancer registry. Body mass index (BMI) at time of diagnosis and treatment details were obtained from patients’ electronic medical records. Patients were grouped by BMI as normal weight (NW, 18.5-24.9 kg/m2), overweight (OW, 25-30 kg/m2) and obese (OB, >30 kg/m2). Continuous and categorical variables were analyzed using ANOVA and chi-square statistical methods, respectively. Survival probabilities of different study groups were estimated and plotted using the Kaplan-Meier method. Results: Records of 375 patients with stage IV CRC were obtained, including 126 NW, 129 OW, and 120 OB patients. With a median follow-up of 16.1 months, median overall survival (OS) was higher in OB than NW patients (20.7 vs. 15.7 months, P<0.001). No difference in age, gender, race, primary tumor site, KRAS/BRAF mutational status, metastatic sites, and surgical treatment was found between study groups. Duration of chemotherapy was significantly longer for OB than NW patients (12.1 vs. 5.4 months, p=0.006). Conclusions: OB patients have improved survival after diagnosis of stage IV CRC and longer duration of chemotherapy compared to NW patients. While the causal relationship would be difficult to establish, these results suggest OB patients may tolerate chemotherapy longer due to the presence of excessive nutritional reserve, leading to better survival.

Background

Small tumor diagnostic tools including ultrasound-guided fine-needle aspiration (US-guided

FNA) and computed tomography (CT) could be causing rising and racially/ethnically different

thyroid cancer incidence rates due to variable overdiagnosis of indolent tumors. Papillary tumors

and <40mm tumors are most likely to be overdiagnosed as indolent tumors by FNA and CT.

Methods

Age-adjusted incidence rates (AAIRs) for years 2007-2014 were calculated for race/ethnicity

(White, Hispanic, Asian, African American, Native American) by patient/tumor characteristics

for microscopically confirmed malignant thyroid cancer cases in SEER 18 (N=93,607).

Multivariate analysis determined cancer patients’ odds ratios (ORs) of diagnosis with papillary

thyroid carcinoma (vs. other histologies) and tumors <40mm (vs. ≥40mm).

Results

For both males and females, there were statistically significant differences in incidence rates

between race/ethnicity with Whites having the highest AAIRs and African Americans the lowest

AAIRs. Among thyroid cancer patients, tumor size and histology differed significantly by race

and insurance coverage, after controlling for gender, age, stage, and tumor sequence. Non-

Whites with thyroid cancer (vs. Whites) were less associated with small tumors (ORs 0.51-0.79;

all P<0.0001). Medicaid and uninsured patients with thyroid cancer were less associated with

tumors <40mm (OR 0.55-0.71, 95% CI=0.49-0.76) and papillary carcinoma (OR 0.86, 95%

CI=0.80-0.93)

Conclusion

Diagnosis of small tumors is occurring at greater rates in Whites (relative to non-Whites) and

insured (vs. Medicaid and uninsured) patients; consequently, these groups may be vulnerable to

unnecessary tests and treatments, or potentially aided with early detection. Guidelines may be

needed that define post-detection interventions to limit overtreatment of indolent, small-sized,

papillary carcinomas.

Abstract Title: Racial/Ethnic Differences in Thyroid Cancer Incidence in the United States, 2007-2014

Authors (as submitted with abstract information):Kristin Weeks, MSTP Amanda Kahl/ MPH Epidemiology UICharles Lynch/ MD, PhD Epidemiology UIMary Charlton/ PhD Epidemiology UI

Intermittent hypoxia alters the bone marrow microenvironment and promotes the engraftment of malignant plasma cells in Multiple Myeloma. Authors: Mahmoud Ali, Sandeep Kowkuntla, Hongwei Xu, Chakrapani Tripathi, Jisung Yuk, Fenghuang Zhan, Deep Hath, Monica Shokeen, Michael H. Tomasson, and Melissa L. Bates

Multiple myeloma (MM) is a plasma cell cancer, characterized by bone destruction, hypercalcemia, anemia, and renal failure with 30,280 new cases in the United State each year. Obesity is associated

with increased MM risk in humans. Obstructive sleep apnea (OSA) is another condition associated with obesity that shares overlapping risk factors with MM. Both conditions are most commonly prevalent among the elderly above 65 years old, men, and with high body mass index. Chronic intermittent hypoxia (CIH) resulting from sleep apnea has emerged as a potent stimulator of tumorigenesis in solid tumors. In our study, we exposed MM-resistant C57BL/6 mice to a CIH profile that mimics human sleep apnea followed by 5TGM1 MM cell injection. CIH exposed mice developed paralysis indicative of symptomatic MM more frequently compared to unexposed controls (Odds ratio of developing paralysis 17.5 (1.6-192.3) compared to controls, p=0.005). Flow cytometry and monoclonal immunoglobulin ELISA quantification data also showed that CIH-exposed mice have malignant plasma cells primarily in the bone marrow. Preliminary data suggests that CIH promoted the engraftment of malignant plasma cells in bone marrow through stimulating angiogenesis and altering the stromal microenvironment rendering it hospitable to cancer. We have developed a novel method to evaluate angiogenesis by clearing the whole bone and examining the bone marrow blood vessels distribution under the confocal microscope without disturbing its anatomy. Blood vessels density in CIH exposed mice was higher compared to controls (median= 19.3% vs 12.5% respectively, P=0.03). RNASeq results show increases in VCAM expression and markers of b-cell proliferation and differentiation in CIH exposed mice. These experiments demonstrate that brief pre-conditioning with CIH promotes the engraftment of malignant tumor cells as well as tumor angiogenesis. This suggests that sleep apnea may be a targetable risk factor for MM.

Title: Preexisting mental illness in SEER-Medicare ER+ breast cancer patients

Authors: Haskins CB; McDowell BD; Carnahan RM; Fiedorowicz JG; Smith BJ; Wallace RB; Chrischilles EA

Character Count (abstract body): 1809

Background: Despite survival benefits, many patients with breast cancer (BC) are nonadherent to endocrine therapies (ET). Mental illness may impact treatment behaviors. We established a cohort using SEER-Medicare, a nationally representative cancer registry, linked to Medicare claims.

Methods: Included women were part of a study about preexisting mental illness and influence on endocrine therapy use, aged 66+ at their first estrogen receptor positive (ER+) BC diagnosed 2007-2013. All had continuous Medicare parts A and B for 12+ months prior and 18+ months after BC diagnosis, 18+ months of continuous part D after BC diagnosis, and a record of surgery. Mental illness was identified using one inpatient or two outpatient ICD-9 diagnoses occurring 30+ days apart.

Results: Increasing lookback periods resulted in cohorts with 37905, 34955 (7.8% fewer) and 32336 (15% fewer) patients for 12, 24, and 36 month periods, respectively. Estimates of any mental illness prevalence increased with longer lookback periods: 5,066 (13.4%), 7,012 (20.1%), and 7,971 (24.7%). Mood (11.3% at 36 months, 5.0% at 12 months), anxiety (9.5%, 4.2%), and psychoses (5.3%, 3.1%) were the most prevalent disorders, followed by dementias (4.4%, 2.5%), drug use (4.3%, 1.9%), and bipolar disorders (1.3%, 0.8%). Less common were delirium (1.2%, 0.5%), adjustment disorders (0.4%, 0.1%), alcohol disorders (0.3%, 0.1%), and personality disorders (0.3%, 0.1%).

Mood, anxiety, and drug use disorder prevalence increased in more recent diagnosis years. Mood disorder and dementia prevalence increased with age, whereas psychoses, drug use, and bipolar decreased; no age association was observed with anxiety. Any mental illness prevalence was highest in black, white, and Hispanic women (12-month prevalence 13.4%, 16.6%, 14.6%, respectively) and less common among Asian (8.3%) and other race (8.4%) women.

Conclusions: As many as one-fourth of women have evidence of mental illness preceding BC diagnosis. A 36 month lookback identified more diagnoses but resulted in a 15% smaller cohort. However, the remaining sample size is sufficiently robust to test relevant, specific hypotheses.

Presenter: Derek B. Danahy, B.S., derek-danahy@uiowa.edu, 913-291-4377 Co-authors: Isaac J. Jensen, B.S.

Thomas S. Griffith, PhD Vladimir P. Badovinac, PhD

Sepsis reinvigorates CD8 T cell responses in tumor bearing mice and prolongs survival

Sepsis is a systemic infection that initiates an exaggerated immune response characterized by acute production of pro- and anti-inflammatory mediators leading to significant lymphopenia and resulting in host damage or destruction. Sepsis strikes regardless of patient age or health-status, however, malignancy is the greatest risk factor for sepsis onset with cancer patients displaying a 10-fold increase in sepsis incidence. Improvements in rapid detection and treatment help many septic patients survive the acute phase of sepsis, however, it is not clear if/how sepsis influences growth/development of existing tumors and underlying anti-tumor immune responses. Our recent data obtained in tumor-free hosts suggest that sepsis has the capacity to reduce quantity and quality (effector function) of naïve/memory CD8 T cell responses, diminishing CD8 T cell-mediated immunity and contributing to a sepsis-induced long-term state of immunoparalysis. Therefore, sepsis might also diminish efficacy of existing anti-tumor CD8 T cell responses resulting in increased tumor development and worsening the outcome. To test this, mice received SQ administration of B16 melanoma, a tumor model in which CD8 T cells provide partial control, followed by cecal ligation and puncture (CLP) model of sepsis induction at the time when tumor was palpable (>14 days). Surprisingly, sepsis survivors displayed reduced progression of B16 melanoma and prolonged survival compared to non-septic Sham controls. Interestingly, increased in situ proliferation, survival, and effector IFN- production was observed in tumor-infiltrating CD8 T cells obtained from CLP hosts. Those cells also re-expressed their activating/inhibitory receptors (ex PD-1, LAG3) suggesting that sepsis can, potentially by destroying tumor cells along the way, reinvigorate existing but inefficient anti-tumor CD8 T cells. Future experiments will also assess the contribution of pro-inflammatory mediators (e.g. TNF-) produced during the cytokine storm and/or by tumor-specific CD8 T cells to further define sepsis-induced beneficial reductions in tumor size observed. Thus, these data suggest while sepsis leads to high mortality in cancer patients, those that survive could have alterations (perhaps beneficial) in anti-tumor CD8 T cell responses improving outcome.

Clinical parameters predicting response and outcomes to immunotherapy in metastatic cancers. Rohan Garje, Adithya Chennamadhavuni, Sarah Bell, Gerald H. Clamon; University of Iowa Hospitals and Clinics, Holden Comprehensive Cancer Center, Iowa City, IA; University of Iowa Hospitals and Clinics, Iowa City, IA; University of Iowa, Iowa City, IA; University of Iowa Hospital and Clinics, Iowa City, IA

Background: Checkpoint inhibitors are approved as effective therapy in a number of malignancies. The response rates range between 20-40% in various cancers. Predictive as well as prognostic clinical characteristics and bio-markers are needed to identify the population that benefits with immunotherapy. Prior studies suggested that the presence of hepatic metastases is a marker of poor prognosis. In this study of patients treated with checkpoint immunotherapy, we evaluated if it also predicts poor response to immunotherapy. Other clinical factors such as age, BMI, baseline LDH, albumin, lymphocyte count, neutrophil count and neutrophil lymphocyte ratio (NLR) were explored as predictive markers of response to checkpoint immunotherapy.

Methods: In this retrospective analysis of 215 patients conducted at University of Iowa, we included patients with renal cancer (N = 23), cutaneous melanoma (N = 181), ocular melanoma (N = 9) and other (N = 1) who were treated with ipilimumab, nivolumab or pembrolizumab.

Results: Hepatic metastases were significantly more common with a N/L ratio > 4, increased ALT, and with other sites of metastases including bone, lung and other visceral organs. Patients with hepatic metastases had a significantly lower response rate ( p = 0.05) , were more apt to be deceased ( p < .01) but age, BMI, and baseline LDH were not associated with hepatic metastases. Factors significantly associated with improved overall survival included BMI > 30 and having received prior therapy before immunotherapy ( p = .04) Metastases to bone, liver, brain, and other visceral organs were associated with poorer survival. Patient age, gender and type of immunotherapy agent were not associated with survival. Too few patients were treated with concurrent antacids or had antibiotics within 30 days of immunotherapy to know if these factors were significant.

Conclusions: Presence of hepatic metastases is associated with poor response to immunotherapy. Interestingly, BMI > 30 is associated with improved survival in patients who receive immunotherapy.

Baseline parameters total number of patients (N = 215) Age > 70 27.4% BMI > 30 34.6% LDH > 220 22.2% Albumin > 3.4 91.6% Neutrophil/Lymphocyte (N/L) ratio > 4 28.6% Hepatic metastases 22.8%

TREM1 promotes lung metastasis in breast cancer

Gaurav Pandey1*, Ryan Kolb1,2, Kathy Keck3, Baruah Sankar3, David Meyerholz1, Weizhou Zhang1,4 Julia Klesney-Tait3.

* Poster Presenter,Postdoctoral Research Scholar, 1 Department of Pathology, 2 InterdisciplinaryImmunology Postdoctoral training program, 3 Internal Medicine, 4 Department of Radiation Oncology, Carver College of Medicine, University of Iowa, Iowa city, USA.

Triggering receptor expressed on myeloid cells 1 (TREM-1) is a member of the immunoglobulin superfamily mainly expressed on neutrophils, macrophages and monocytes. Increasing evidence suggests a role for TREM-1 not only in acute pathogen-induced reactions but also in chronic and non-infectious inflammatory disorders, including various types of cancer. Here, we demonstrate that TREM-1deficiency protects against lung metastasis in breast cancer, as TREM-1 knockout mice have decreased lung metastasis. This decrease in lung metastasis is associated with an increase in lymphocyte infiltration, in particular CD4 and CD8 T cells, at the site of metastasis. Moreover, depletion of CD4+ cells by administration of an anti-CD4 antibody increased lung metastasis in TREM-1 deficient mice, suggesting that loss of TREM-1 may promote CD4 T cell mediated clearance of lung metastasis. Mechanistically, we found that TREM-1 deficient mice with lung metastases have an increase in the chemokine CCL11 in the lung compared to wild type mice with lung metastasis. Conversely, CCL11 deficient mice have an increase in breast cancer lung metastasis, suggesting that TREM-1 may promote lung metastasis by suppressing the production of CCL11. Ongoing studies are exploring any correlation between TREM-1, CCL11 in regulating lung metastasis and T cell infiltration. In humans, genomic analysis shows a correlation between TREM-1 expression in breast cancer and shorter overall survival, metastasis-free survival regardless of site and lung metastasis-free survival. Collectively, our findings suggest that TREM-1 may promote the lung metastasis by suppressing CD4 T cell infiltration of clearance suppression of metastasis possibly by decreasing CCL11 secretion. Thus, blocking of TREM-1 signaling in cancer patients may reduce the chances of metastasis and increase the overall survival of cancer patients.

Association of breast cancer disease free survival (DFS) with omeprazole use. Josiah An, Adithya Chennamadhavuni, Sarah L Mott, Rohan Garje, Jose Pablo Leone; University of Iowa Hospitals and Clinics, Iowa City, IA; University of Iowa Hospitals and Clinics, Holden Comprehensive Cancer Center, Iowa City, IA; Dana-Farber Cancer Institute, Boston, MA

Background: Recent studies indicated that aryl-hydrocarbon receptor (AHR) modulators - such as omeprazole - decrease breast cancer cell invasion among mice with estrogen receptor negative (ER-) or triple negative (TN) breast cancer. Yet, it is unknown whether omeprazole use decreases breast cancer metastasis and improves survival in humans. The aim of this study was to assess the potential benefit of omeprazole use on DFS in ER- and TN breast cancer patients (pts) at a large academic center. Methods: A retrospective chart review was conducted on pts with breast cancer diagnosed between 2008 and 2016. Pts were classified by omeprazole use ( > 30 days of use). To investigate differences in pt characteristics by receipt of omeprazole, chi-squared and t-tests were used. To examine whether ER or TN status modified the effect of receipt of omeprazole on DFS and overall survival (OS), Cox regression models were used. Results: A total of 1387 pts were included in the study (non-omeprazole n = 968, omeprazole n = 419). There were 285 ER-tumors and 183 TN tumors. After a median follow up of 4 years, 94% of pts had no recurrence of breast cancer and 88% of pts were still alive. Median duration of omeprazole use for omeprazole group was 227 days. Median age at diagnosis was 58 years. Treatments included surgery (98%), radiation (61%), hormone therapy (65%) and chemotherapy (45%). Utilization of hormone therapy was lower in pts receiving omeprazole (61 vs 66%, p = 0.04), but chemotherapy was higher (49 vs 44%, p = 0.05). No significant differences in terms of sex, race, ER, PR, HER-2, stage, radiation, surgery, and age between omeprazole groups were evidenced. Multivariable results for the interaction between omeprazole use and receptor status (ER or TN) on DFS and OS are reported in table.

Conclusions: Omeprazole use in breast cancer did not confer a survival benefit in DFS and OS for tumor subtypes based on ER or TN status.

DFS* OS*

HR 95% CI P-

value

HR 95% CI P-

value

Omeprazole Use Yes vs No At ER = Negative 1.53 0.93 2.51 0.50 1.16 0.68 1.98 0.57

Omeprazole Use Yes vs No At ER = Positive 1.24 0.85 1.80 1.41 0.94 2.10

Omeprazole Use Yes vs No At TN = No 1.13 0.76 1.66 0.34 1.28 0.84 1.94 0.57

Omeprazole Use Yes vs No At TN = Yes 1.58 0.89 2.79 1.03 0.55 1.91

*Adjusted for chemotherapy and hormone therapy

Symptom management barriers in advanced cancer rural patients

Backgrounds: Symptom management can help patients manage their symptoms.

Representational approach (RA) is a patient education framework that has been used for

designing symptom management interventions for cancer patients. The purpose of this study was

to assess the barriers and misconceptions for symptom management in Oncology Associated

Symptoms and Individualized Strategies (OASIS) intervention. OASIS is designed for advanced

cancer patients in rural areas to help them better manage their symptom.

Methods: The intervention was delivered through videoconferencing platform and clinicians

were meeting weekly with patients for eight weeks. RA was used as a framework for guiding

their discussion and helping patients to identify the misconceptions and barriers. The total

number of eight patients were recruited as a part of the feasibility study. Clinician’s meeting with

patients were audiotaped and analyzed for identifying barrier and misconceptions were using a

qualitative approach.

Results: The barriers that patients mentioned were symptom management strategies on the

website was not helpful, patients were not willing to communicate with oncologist regarding

medications and symptoms, patients not willing to take medication, no need to manage symptom

until the distress level gets very high, being a caregiver preventing them to take care of their own

health, and not being open to try new strategies. Symptom misconceptions were related to the

cause of the symptoms, not being aware that they were doing self-management strategies, and

clinician’s recommendation for symptom management contradicts oncologist recommendation.

Conclusion: Patients with advanced cancer have multiple barriers and misconceptions towards

symptom management.

Authors (as submitted by poster presenter):Seyedehtanaz Saeidzadeh, University of Iowa, College of NursingStephanie Gilbertson-White, PhD, APRN-BC

BioinformaticsSharedResource

Abstract:TheHCCCBioinformaticsSharedResource(BSR)supportstheinformaticsneedsoftheclinical,basicandpopulationscientistsoftheHCCC.Thissharedresourcehastheexpertisetocollectandutilizelarge-scaledatasets,includingmolecularassays,forallHCCCmembers.Activeexamplesincludemappingandanalysisfornext-generationDNA/RNAsequencing,machinelearning,featureextraction,tissueprocurementsupportandphenotyping.Servicesinclude:i)acquisition,installation,andmaintenanceofexisting/establishedbioinformaticstools,includingthosedevelopedbyNCI,foranalyzinggenomicandotherlargedatasets;ii)augmentationofexistingtoolswithadditionalsoftwaretoenablecustomanalysesandapplicationsofexistingtools;iii)consultationwithHCCCmembersonthedesignofexperimentsutilizinglarge-scaleandhigh-throughputmolecularassaysandotherexperimentsinvolvinglargedatasets;iv)workingwithothersharedresearchresourcesoftheHCCCtoassureoptimalexchangeandutilizationofdatasetsamongsharedresources,andbetweensharedresourcesandcancercenterinvestigators.

Genomics Shared Resource (GSR) Abstract

Title: New Gene Expression Profiling Technologies Available from the Genomics Shared Resource: Introducing Single-Cell RNA-Seq and Digital PCR

Presenter: Kevin L. Knudtson, PhD

The HCCC Genomics Shared Resource (GSR) has been providing gene expression profiling services to cancer center members for over 15 years. Over this time, the technologies used to perform have evolved to permit deeper and more sensitive detection of levels of gene expression. Currently, the GSR provides expression services using the NGS- RNA-seq and realtime qPCR-based technologies. Recently, the GSR added the ability to perform single-cell expression profiling using the 10X Genomics Chromium System. The 10X system permits the collection of single-cell expression profiles from hundreds to thousands of cells in a sample. Digital PCR is a highly sensitive qPCR technique that permits absolute quantification of nucleic acid targets. The activities and services that support the research investigator focus around 6 major areas of support: 1) DNA Sequencing and Genotyping, 2) Custom Oligonucleotide Synthesis, 3) Genome Sequencing using Illumina HiSeq 4000 and MiSeq, 4) DNA microarray using the Illumina iScan BeadArray System, 5) Real-Time and Digital PCR, and 6) Biomolecular Computing. The clinical laboratory performs the genome sequencing for 3 clinical tests: 1) Clinical Exome, 2) Drug Metabolism, and 3) KidneySeq. More information about the clinical tests and the GSR research support services can be found at: http://www.medicine.uiowa.edu/humangenetics/.

HCCC Core Resource Poster Central Microscopy Research Facility Randy Nessler, BGS, MBA, Director At the University of Iowa's Central Microscopy Research Facilities (CMRF), we provide a wide variety of microscopy techniques for materials and biomedical investigators, an experienced staff, 24/7 access, and support for the beginner and experienced investigator. As one of the leading university microscopy facilities in the nation, our microscopy team is ready to support your research program. Part of our mission is to provide services and instruction to Holden Comprehensive Cancer Center investigators interested in using CMRF technology in their cancer research. Our three most recently added instruments were; a Leica LMD 7000 laser capture microdissection system, a Leica SP8 laser scanning confocal including STED super resolution and a Leica Aperio Ariol Digital Slide Scanning system. To advance its image analysis services, the CMRF has purchased several new modules for its Imaris 4D Visualization and Analysis software suite, including:

• Imaris Measurement Pro: Shape, size and intensity-based quantification. • Imaris Track: Imaging, tracking and motion analysis of live cells and moving objects in 2D and 3D • Imaris Filament Tracer: Automatic detection of neurons (dendritic trees, axons and spines,)

microtubules, and other filament-like structures in 2D, 3D and 4D • Imaris Coloc: Quantify and document co-distribution of multiple stained biological components • Imaris Cell: Quantitatively examine micro relationships that exist within and between cells • Imaris Vantage: Compare and contrast experimental groups by visualizing image data in five

dimensions as uni- or multi-variate scatterplots • The CMRF designed and built a custom, high-end computer workstation to run Imaris, and also

Huygen’s Pro Deconvolution, FIJI, ZEN and LASX. Computer specifications include: a 10-core Intel Extreme CPU, 128GB RAM, an 8GB video card, and multiple PCIe solid state hard drives.

The CMRF is investigating several new imaging technologies, including Light Sheet Microscopy (LSM.) LSM uses planar illumination to deliver confocal-like imaging of large samples, such as embryos, and cleared brain and other tissues. The CMRF is planning to replace its current Transmission Electron Microscope as well as upgrading to the Zeiss LSM880 laser scanning confocal. In 2017, 46 Holden Comprehensive Cancer Center (HCCC) member laboratories (with 113 Research Assistants) from all 4 HCCC programs have utilized the services of the CMRF. These labs utilized a wide array of instrumentation including but not limited to: laser scanning confocal microscopy, light and fluorescent microscopes, paraffin processing, paraffin sectioning, cryostats, bioluminescence imaging, transmission and scanning electron microscopy. CMRF staff assisted researchers by consulting on the design of experiments, conducting research and providing training. We welcome the opportunity to discuss your research projects and goals. Visit our website at: http://cmrf.research.uiowa.edu/

HCCC Biostatistics Core

Authors: Brian J Smith (Core Director)1, Gideon K Zamba1, Patrick J Breheny1, Sarah L Mott2, Timothy Ginader2 1 Department of Biostatistics 2 Holden Comprehensive Cancer Center

Abstract: The Biostatistics Core is a shared resource of the Holden Comprehensive Cancer Center (HCCC) that assists with the development, conduct, analysis, and reporting of cancer-related research projects. The Core consists of biostatistics faculty, staff, and graduate students who support the HCCC research and training missions through the following activities:

• Collaboration with investigators on basic science, genetic, clinical, and population-based studies.• Support of Molecular Epidemiologic Resource (MER) databases and projects.• Development of specific aims, experimental designs, and statistical analysis plans for study

protocol and grant applications.• Determination of sample size and study power.• Web-based data entry and management.• Statistical analysis of study data.• Presentation of statistical methods and results in journal manuscripts and conference

presentations.• Interaction with other shared resources to help ensure integrated access to study design, data

collection, and data analysis.• Conduct of and participation in educational programs for investigators, faculty, fellows, students

and staff.• Support of HCCC protocol review and study monitoring.

Examples of our involvement in research projects are presented to illustrate our biostatistical activities and support provided. We additionally describe our online support request system that facilitates investigator contact with the Core and the matching of investigator needs with Core expertise.

Flow Cytometry Facility

Zuhair Ballas, M.D, Director Justin Fishbaugh, B.S., Technical Director Heath Vignes, B.S., Core-Facility Research Specialist Mike Shey, B.S., Core-Facility Research Professional

Campus Address: 48 EMRB Phone: 335-8103 Website: http://www.healthcare.uiowa.edu/corefacilities/flowcytometry/

The 1,200-square-foot Flow Cytometry Facility is located in the Eckstein Medical Research Building (EMRB). The Facility has one magnetic-based and 10 laser-based instruments whose major purpose is the identification and isolation of various cell populations. The laser-based instruments accomplish this by the use of antibodies to which various colors or dyes have been attached and are directed at molecules known to exist on the cell surface or in the cytoplasm. By using several colors attached to different antibodies, one can identify and purify cells that express any given configuration of various molecules.

In addition to identification and isolation of various cell populations using antibodies, flow cytometry can analyze:

Cell proliferation response to drugs or treatments, such as those used in chemotherapy

Cell physiological properties, such as calcium flux and pH

DNA content and integrity

Transfection markers such as GFP and mCherry

Quantification of cytokines, gene expression, micro RNA

Facility Instrumentation:

Becton Dickinson FACS Aria II high-speed cell sorter (5-laser excitation, 14-color detection) Becton Dickinson FACS Aria Fusion high-speed cell sorter (3-laser excitation, 11-color detection) Becton Dickinson LSR II (three each with 3-4 laser excitation and 8-12 color detection) Becton Dickinson FACS Calibur (2-laser excitation, 4-color detection) Becton Dickinson FACScan (two each with 1-laser excitation and 3-color detection) BioRad BioPlex (Luminex 200, two each) Miltenyi autoMACS

A web-based reservation system allows investigators to schedule instrument time. The Facility is staffed M-F, 8am to 6pm and is available 24/7 following training.

HCCC Core Resource Poster

The Genome Editing Core Facility

Presenter/Author: William Paradee, PhD (william-paradee@uiowa.edu)

Co-Authors: Patricia Yarolem, JoAnn Schwarting, Norma Sinclair

Affiliations: University of Iowa: François M. Abboud Cardiovascular Research Center

The Genome Editing Core Facility is a full-service core facility of the Carver College of Medicine at the University of Iowa. The Core has a long history of providing Iowa researchers access to the latest state-of-the-art technologies for generating transgenic and gene targeted mice to enable production of animal models for human disease. The core works closely with investigators to design and execute experiments based on their specific needs and provides customizable support to enable the successful completion of the investigator’s experimental aims.

The Genome Editing Core provides additional related services such as embryo cryopreservation, sperm cryopreservation, IVF and strain rederivation. Additional services to be offered in the future include colony management and congenic breeding services.

Facility staff provide the expertise, state-of-the-art laboratory space, and pathogen-free animal quarters necessary for the efficient production of transgenic and gene-edited mice.

www.medicine.uiowa.edu/genomeediting

Presenter: Freda Selk, AAS Freda-selk@uiowa.edu 335-8638

Other authors: Dan Olson, MS Daniel-olson@uiowa.edu

Bradley McDowell, PhD Bradley-mcdowell@uiowa.edu

Charles Platz, MD Charles-platz@uiowa.edu

Charles Lynch, MD, PhD Charles-lynch@uiowa.edu

Background: The State Health Registry of Iowa (SHRI) has two types of formalin-fixed, paraffin-embedded tissue repositories: 1) Iowa Residual Tissue Repository (RTR), and 2) Iowa Virtual Tissue Repository (VTR). The SHRI provides data on the cancer experience of Iowans dating back to 1973. Methods: As of April 2018, the Iowa RTR involved 92,128 pathology reports for 63,317 cancer patients representing 66,416 tumors and 545,898 tissue blocks. Most of these tumors were diagnosed between the years 1994 and 2006, and are obtained through the cooperation of private laboratories at the time blocks are to be otherwise discarded. An electronic tracking system assists in efficiently providing de-identified pathology reports and their tissue blocks linked to patient demographic, tumor, first-course treatment, and follow-up characteristics. In April 2018, Iowa RTR pathology, patient, and tumor variables were provided to UI BioShare to support increased utilization of this resource by U of Iowa researchers. When tissue processing is required for a research study, the SHRI collaborates with the Histology component of the Comparative Pathology Laboratory at the University of Iowa. As of April 2018 for the years 1973-2016, there are an estimated 365,000 tumors with tissue available through the Iowa VTR. These tissue blocks and their path reports reside in their originating laboratories. Results and Conclusions: Since 2010 there have been 46 peer-reviewed publications involving Iowa tissue repositories and addressing etiologic, survival, and/or classification issues for NHL, HPV-related sites, and cancers of the pancreas, prostate, breast, colorectum, vulva and ovary. Most of the authors of these publications reside outside the Holden Comprehensive Cancer Center (HCCC), but HCCC members are strongly encouraged to use the resource. HCCC member access is available through the Population Research Core (e-mail: bradley-mcdowell@uiowa.edu).

State Health Registry of Iowa Tissue Repository

HCCC Core Resource Poster

The Viral Vector Core Facility

Presenter/Author: William Paradee, PhD (william-paradee@uiowa.edu)

Co-Authors: Susan Stamnes, Steven Rhines, Jeremy Coffin, Anni Noble, Diane Cryderman, Zhaohui Shi, Seong-Ae Kang, Amanda Moeller, Kaylee Stookesberry, Patrick Sinn, PhD

Affiliations: University of Iowa: Holden Comprehensive Cancer Center; Center for Gene Therapy of Cystic Fibrosis

The Viral Vector Core is a full-service facility located on the University of Iowa Medical School campus. The main objective of the Core is to provide investigators (both intramural and extramural to UI) state-of-the-art gene transfer technologies. The Vector Core utilizes molecular biology techniques to engineer viral vectors for gene transfer in pre-clinical studies and other basic research applications. These studies provide valuable information critical to the understanding of gene function and development of potentially therapeutic vectors. Collaborations between Vector Core staff and investigators allows cross fertilization of ideas and innovations in vector design.

The Core offers several types of vectors for gene delivery including: Adeno-Associated Virus (AAV), Lentivirus, Adenovirus and Vaccinia. In addition to design, cloning and vector production, the Core provides resources critical for understanding the biological aspects of the vector technology.

www.medicine.uiowa.edu/vectorcore

Title: The Molecular Epidemiology Resource Core (MERC)

Authors: Brian Link, MD, Ashley McCarthy, MPH

Category: Translational Research

The Molecular Epidemiology Resource Core (MERC) is a new shared resource offering Holden

Comprehensive Cancer Center (HCCC) investigators support for high quality disease-specific outcomes

research. The MERC offers meticulous collection of longitudinal clinical data and biologic specimens

including serum, plasma and germline DNA, all linkable to tumor samples. The MERC has focused its

efforts on disease-specific groups that have the necessary clinical and research strength to utilize the

resulting data. Current disease-specific groups supported by the MERC include lymphoma, melanoma,

sarcoma, myeloma, and cancers of the breast, pancreas, biliary, and genitourinary system. The MERC is

a rigorous, prospective observational database linked to a biorepository.

All newly diagnosed patients with appropriate histologies are approached about informed consent.

Following enrollment, MERC personnel abstract clinical information including tumor stage, histology, lab

and imaging data, treatment information, events (progression, death) and comorbidities. In general,

clinical information on each subject is updated twice a year for the first three years and then annually.

Psychosocial data including quality-of-life analyses are collected longitudinally. Serum, plasma, buffy

coat and peripheral blood DNA are collected at diagnosis and selected longitudinal time points. Excess

surgical tissue (tumor and normal) from resections and biopsies are also collected when available.

Currently, over 7,500 patients have been enrolled in the MERC and over 28,000 biomaterials have been

collected since the MERCs inception.

The MERC is funded by HCCC, University of Iowa Foundation, and multiple disease-specific federal/non-

federal grants. We also partner with the ORIEN/PERCH initiative.

Recent evidence indicates that metabolic oxidation/reduction (redox) reactions are disrupted in cancer vs. normal cells. These disruptions contribute to cancer progression, cancer cell resistance to therapy, normal tissue injury processes, inflammatory responses, and therapy outcomes. The goal of the Radiation and Free Radical Research Core (RFRRC) is to provide state of the art technologies to HCCC investigators studying the role of oxidative stress and redox biology as they relate to cancer biology and cancer therapy. New knowledge in the redox biology of cancer will aid in the development of novel biochemical approaches for enhancing cancer prevention and improving cancer therapy. This knowledge will also bring greater understanding to the mechanisms of degenerative diseases associated with aging. The main services provided by the RFRRC are focused on providing free radical and radiation biology expertise, reagents, technologies, and analysis for investigators doing basic, pre-clinical, and clinical research. The RFRRC offers three basic services: (1) EPR Services (EPR); (2) Ionizing Radiation Services (IRS); and (3) Antioxidant Enzyme Services (AES) to researchers doing cancer research. Recent high impact studies, supported by the RFRRC, include: the study of pharmacological ascorbate in cancer therapy; use of SOD mimics in cancer therapy; and studies of metabolic reprogramming associated with aging and its impact on cancer therapy. The long-term goal of the RFRRC is to support the expansion of investigator-initiated studies in the HCCC involving the redox biology of cancer, i.e. moving basic science discoveries in redox cancer biology from the bench to the bedside.

RADIATION AND FREE RADICAL RESEARCH CORE

Title: HCCC Population Research Core Author: Bradley D. McDowell, PhD

The Population Research Core (PopRC) is a centralized resource that supports strong

observational study design and data collection/curation methods. The PopRC works with cancer

investigators to (1) implement appropriate design and methodology to answer population-based

questions; (2) provide efficient use of large population-based datasets; (3) support population-

based field research; and (4) provide biospecimens from the Iowa Residual Tissue Repository.

These services support many types of studies including those focused on comparative

effectiveness, multi-site pragmatic trials, molecular epidemiology, and treatment patterns. In

addition, the PopRC supports behavioral science and prospective data collection through

surveys. The primary resources of the PopRC are the expertise and time of its scientific

personnel and its curated and annotated population-based data, including Surveillance,

Epidemiology, and End Result (SEER) data and its linkage to Medicare claims data. Overall, the

PopRC strives to be a collaborative, efficient, and integrated shared resource for encouraging

and supporting the population science goals of the HCCC.

No abstract - Poster Display only

Promoting Success from Idea to PublicationHardin Library for the Health Sciences

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Research project IDEA Ph.D. project

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Be sure you have all relevant information for your project.We support literature searches by showing how to

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2 . Experts: Together more than 75 Years of Experience in Library Services for Research… and Hardin’s HCCC liaison librarians:

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