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The use of biomarkers for treatment decisions
in oncology1Timothy A Yap, MD PhD and
2Debra Josephs, MD PhD1The Institute of Cancer Research and Royal Marsden Hospital, London, UK;
2Kings College London, UK
Definition of a biomarker
“Characteristic objectively measured and evaluated as indicator of normal biological processes, pathogenic processes or pharmacologic responses to a therapeutic intervention.”
(US National Institutes of Health Biomarkers Definition Working Group)
Key component of translational research
Biomarkers Definitions Working Group (2001) Clin Pharmacol Ther 69:89-85
Reprinted by permission from Macmillan Publishers Ltd: Yap T A et al. Nat. Rev. Cancer 2010;10: 514–523, copyright (2010)
Need right biomarker for the right drug in the right patient at the right time
Past
Current & future
Biomarker-driven drug treatment
Changing focus of anticancer drug development
Translational research in new drug development
Transfers preclinical research into clinical practice:
Study of the biology of the disease Evaluation of the biological effects of the drugs in animals Study of the biological effects of those drugs in humans
Biomarkers and drug development
Used during different steps of drug development in order to:
Define the target deregulation correlate to tumour development (proof of target)
Verify the interaction between the drug and the target or pathway (proof of principle)
Verify that alteration of a specific target is crucial for a specific tumour in humans (proof of concept)
Verify that the interaction between drug and target is responsible for biological effects (proof of activity)
Define a range of doses (biologically active dose range) Provide rationale for combination therapies (crosstalk and
feedback loops)
Park JW et al. Clin Cancer Res 2004;10:3885-3896
Reprinted by permission from Macmillan Publishers Ltd: Yap TA et al. Nat. Rev. Cancer 2010;10: 514–523, copyright (2010)
Rational framework for assessing the risk of failure of the development of a new agent at any particular stage
Provides basis for making key decisions, such as determining the optimal dose range and schedule of a new compound and whether to continue or terminate a drug development programme
Pharmacological audit trail
Biomarkers classification
Three main categories (US-NCI):
A. Prognostic biomarkers
B. Predictive biomarkers
C. Pharmacodynamic biomarkers
Kellof GJ, et al. Clin Cancer Res 2004;10:3881-3884Sawyers CL, et al. Nature 2008;452:548-552
A. Prognostic biomarkers
Correlate with clinical outcome and allow prediction of the natural course of cancer
Guide the therapeutic decision
They include: 1. Biological progression markers 2. Risk biomarkers (or screening biomarkers)
A. Prognostic biomarkers
1. Biological progression markers
Measures of tumour activity (CEA, alphaFP, PSA, CA-125, hCG)
A. Prognostic biomarkers
2. Risk biomarkers (or screening biomarkers)
Describe risk of cancer occurrence or cancer progression because they are implicated in neoplastic progression and include:
1. Carcinogen exposure2. Genetic predisposition (e.g., BRCA1/2 mutations)3. Over expression of genes (e.g., PTEN, BCR-ABL, HER-2/neu,
RAS, AKT)4. Pharmacogenomic parameters (genetic polymorphisms, DNA
methylation, aneuploidy)5. Environmental factors and lifestyle (e.g., HPV or HBV infection,
tobacco use)6. Multifactorial risk models (e.g., Gail model, Oncotype DX, MammaPrint)
B. Predictive biomarkers
Correlate with probability of clinical response to treatment:
Amplification of HER-2 gene in breast cancer for treatment with anti-HER-2
ER/PgR expression in breast cancer for hormone therapy Presence of Philadelphia chromosome (BCR-ABL fusion gene) in
leukaemia for anti-BCR-ABL Mutations of EGFR in lung tumours for EGFR inhibitors Extended RAS mutation wildtype (exons 2, 3 and 4 of KRAS and
NRAS) in colorectal cancer for EGFR monoclonal antibodies BRAF V600E mutations in melanoma for BRAF inhibitors EML4/ALK translocations in lung cancers for ALK inhibitors
C. Pharmacodynamic biomarkers
Correlated to biological and clinical effects of the drug on the tumour Measured in tumour biopsies or normal tissue including platelet-rich
plasma, hair follicles, circulating tumour cells, saliva, or urine
They could be: Lower expression or activity of a molecular target
(e.g., EGFR expression) Decrease in phosphorylation signal of target (e.g. phosphoSer473 AKT) Induction of metabolism (e.g., interference with cytochrome P-450) Interference with cell growth processes
(e.g., Ki-67, BCL-2, cytokeratins) Vascularization and metabolism (imaging biomarkers)
Sarker D et al. Biomarkers Med 2007;1:399-417Yap TA et al. Nat. Rev. Cancer 2010; 10: 514–523 (2010)
“Proximal” and “Distal” biomarkers
Proximal: reflect the immediate effect of the drug on its target (e.g., decrease in a protein substrate of phosphorylation downstream the target kinase such as the phosphorylation of the AKT substrate PRAS40 to reflect the effect of an AKT inhibitor on AKT)
Distal: reflect the effect of the drug downstream the molecular target (e.g., Ki-67 expression for reduction in proliferation)
Applications of biomarkers in drug development
1. Target discovery and validation
2. Preclinical studies
3. First-in-human phase I clinical studies
4. Phase II/III studies
1. Applications of biomarkers in drug development: Target discovery and validation
Identify targets for therapy(e.g., HER-2, VEGF, VEGFR, BCR-ABL, BRAF…):
HER-2 amplification in breast cancer for anti-HER-2 therapy
2. Applications of biomarkers in drug development preclinical studies
Selection of animal models and lead compounds to test
Definition of mechanisms of action in vitro and in vivo
Prediction of the effects of drug combinations
3. Applications of biomarkers in drug development: First-in-human Phase I clinical studies
Evaluate safety and tolerability of a novel compound and select optimal dose for subsequent efficacy trials by establishing the biologically effective dose range and maximum tolerated dose (MTD)
Pharmacokinetic studies (what the body does to the drug) should be undertaken
Pharmacodynamic studies (what the drug does to the body) should be conducted in both tumour and normal tissue, such as platelet-rich plasma and hair follicles
Preliminary signals of antitumour activity may be correlated with predictive biomarkers of response in individual patients
3. Application of biomarkers in Phase I clinical studies: Case studies
PARP inhibitors
AKT inhibitors
MEK and BRAF inhibitors
Immune checkpoint inhibitors
PARP-1 is a key enzyme involved in the repair of single-strand DNA breaks
PAR chains are degraded via PARG
RepairedDNA
PARPDNA damage
Binds directly to SSBs
Repair enzymes
PAR
Nicotinamide+pADPr
NAD+
Once bound to damaged DNA, PARP modifies itself producing large branched chains of PAR
Illustration courtesy of AstraZeneca
3. Application of biomarkers in Phase I clinical studies: PARP inhibitors
Pharmacokinetic (PK) and pharmacodynamic (PD) data
Redrawn from: Fong PC et al. N Engl J Med 2009; 361:123-134
0 100
50
10152025
Cm
ax(μ
g/m
l)
200 300 400 500 600Olaparib dose (mg)
0 100
40
0
6080
100120
AUC
10(μ
gxhr
/ml)
200 300 400 500 600Olaparib dose (mg)
20
0 20
40
0
6080
100
PAR
P in
hibi
tion
(% o
f bas
elin
e)
40 60 100Olaparib steady-state AUC
(μgxhr/ml)
20
80
8 31 35 36 42Patient no.
Anti-PARantibody
Anti-PARP1antibody
Anti-actinantibody
100
30
0
506070
Nuc
lei w
ith ≥
10 s
mal
l or
≥3
larg
e fo
ci
200 600Twice-daily olaparib dose
(mg)
20
400
40
10
Before first doseHr 6 (Day 1)Day 8Day 14Day 21, 2nd cycle
100
30
0
506070
Nuc
lei w
ith ≥
10 s
mal
l or
≥3
larg
e fo
ci
200 600Twice-daily olaparib dose
(mg)
20
400
40
10
Before first doseMaximum 1st cycle
Yap et al, ASCO 2007
γH2AX
DAPI
PD analysis: Hair follicles for γ-H2AX foci
Reprinted with permission. © 2007 American Society of Clinical Oncology. All rights reserved. Yap T et al. J Clin Oncol; 25(18S), 2007: abst 3529
3. Application of biomarkers in Phase I clinical studies: PARP inhibitors
RECIST efficacy in BRCA1/2 mutated tumours 1st 60 patients 19 BRCA carriers (15 ovarian, 3 breast, 1 prostate) 8 PR ovarian; 1 CR breast; 1 PR prostate; 2 SD
Redrawn from: Fong PC et al. N Engl J Med 2009; 361:123-134
Progressive disease Stable disease Partial response Complete response
Ovarian cancer Partial response Complete response
0
20
60
80
Trea
tmen
t dur
atio
n (w
k)
40
3. Application of biomarkers in Phase I clinical studies: AKT inhibitors
Reprinted by permission from Macmillan Publishers Ltd: Yap TA et al. Nat. Rev. Cancer 2009;9: 167-181, copyright (2009)
3. Application of biomarkers in Phase I clinical studies: AKT inhibitors
Yap TA et al., J Clin Oncol 29, 2011:4688-4695. Reprinted with permission. © 2011 American Society of Clinical Oncology. All rights reserved.
Pair tumour biopsy PD biomarker studies
Hair follicle PD biomarker studies
3. Application of biomarkers in Phase I clinical studies: AKT inhibitors
Yap TA et al., J Clin Oncol 29, 2011:4688-4695. Reprinted with permission. © 2011 American Society of Clinical Oncology. All rights reserved.
Biomarker-driven case study of responding patient on MK-2206
3. Application of biomarkers in Phase I clinical studies: AKT inhibitors
Reprinted from Yap TA et al., Clin.Cancer Res, 20: 5672-5685, copyright 2014, with permission from AACR
Platelet-rich plasma PD biomarker studies
3. Application of biomarkers in Phase I clinical studies: MEK and BRAF inhibitors
Reprinted from Clin. Cancer Res, copyright 2010, 16(13): 3329-3334, Pratilas C et al., Targeting the Mitogen-Activated Protein Kinase Pathway: Physiological Feedback and Drug response, with permission from AACR
3. Application of biomarkers in Phase I clinical studies: MEK inhibitor
Inhibition of ERK phosphorylation has been proposed as a PD proof-of-mechanism biomarker of MEK inhibitor activity
A phase I study of a MEK inhibitor (AZD6244) was designed in two parts
The first, to determine the maximum tolerated dose, was also used to document a dose-dependent inhibition of ERK phosphorylation in a surrogate tissue (PBMC)
The second part demonstrated consistent inhibition of ERK phosphorylation and reduction in the Ki-67 labelling index, a marker of cell proliferation, when comparing pre- and post-treatment tumour biopsies at all doses
Therefore a lower dose level (50% of the MTD) could be recommended as the tolerable phase II dose
Adjei AA et al., J Clin Oncol. 2008; 26(13): 2139-2146
3. Application of biomarkers in Phase I clinical studies: MEK inhibitor
Inhibition of ERK phosphorylation
Adjei AA et al., J Clin Oncol. 2008; 26(13): 2139-2146 Reprinted with permission.© 2008 American Society of Clinical Oncology. All rights reserved
In peripheral-blood mononuclear cell samples
Pre-treatment Post-treatment
In paired tumour samples Tumor cell nuclei H-score for pERK
ERK phosphorylation
Before study (%inhibition) Day 7-21 (% inhibition)Before dose
(n=50)*After 1 hour
(n=50)Before dose
(n=25)After 1 hour
(n=22)Mean 0 65.4 51.0 69.2
Median 0 66.0 59.0 76.0
Range 0 5-100 3-90 15-97
3. Application of biomarkers in Phase I clinical studies: MEK inhibitor
Reduction in Ki-67 labelling indexPre-treatment Post-treatment
Proportion of tumour cell nuclei staining for Ki-67
Change in target lesion size
Adjei AA et al., J Clin Oncol. 2008; 26(13): 2139-2146, Reprinted with permission. © 2008 American Society of Clinical Oncology. All rights reserved
3. Application of biomarkers in Phase I clinical studies: BRAF inhibitor
Somatic mutations of BRAF kinases are present in approximately 60% melanomas. An activated single substitution V600E accounts for 80% of BRAF mutations in malignant melanoma
Phase I study of PLX4032, an orally available inhibitor of V600E mutated BRAF
Patients with melanoma with BRAF V600E mutation were overrepresented because of selective activity of PLX4032 against such tumours in preclinical testing
PLX4032 induced complete or partial tumour regression in 81% of patients who had melanoma with the V600E BRAF mutation proving this as an excellent predictive biomarker
Flaherty KT et al., N Engl J Med 2010; 363:809-19; Bollag G et al., Nature 2010; 467:596–599
3. Application of biomarkers in Phase I clinical studies: BRAF inhibitor
Anti-tumour response in patients with melanoma carrying V600E BRAF mutation
Flaherty KT et al., N Engl J Med 2010;363:809-19, Copyright © 2010 Massachusetts Medical Society. Reprinted with permission from Massachusetts Medical Society Bollag G et al., Nature 2010; 467: 596–599
3. Application of biomarkers in Phase I clinical studies: BRAF inhibitor
A cohort of patients had paired tumour biopsies collected before treatment and following two weeks of treatment, to evaluate pathway inhibition
Phosphorylated-ERK (pERK) levels were determined by immunohistochemistry (IHC) to monitor ERK pathway activity. To monitor proliferation, Ki67 levels were also measured
Levels of pERK and Ki67 were decreased in most biopsies following two weeks of dosing, even in patients with modest drug exposure
However, only patients exposed to plasma levels of PLX4032 higher than 300 μM/h experienced tumour regression. In these patients, pathway analysis typically showed >80% inhibition of cytoplasmic pERK indicating that near-complete inhibition of ERK signaling may be needed for significant tumour response
Flaherty KT et al., N Engl J Med 2010;363:809-19; Bollag G et al., Nature 2010; 467: 596–599
3. Application of biomarkers in Phase I clinical studies: BRAF inhibitor
IHC of pERK, Cyclin D1 and Ki67 in paired tumour biopsies
Flaherty KT et al., N Engl J Med 2010;363:809-19, Copyright © 2010 Massachusetts Medical Society. Reprinted with permission from Massachusetts Medical Society Reprinted by permission from Macmillan Publishers Ltd: Bollag G et al., Nature 2010; 467: 596–599 copyright (2010)
3. Application of biomarkers in Phase I clinical studies: BRAF inhibitor
Correlation of reduction in cytoplasmic pERK with tumour responses
Reprinted by permission from Macmillan Publishers Ltd: Bollag G et al., Nature 2010; 467: 596–599 copyright (2010)
3. Application of biomarkers in Phase I clinical studies: BRAF inhibitor
Oncogenic BRAF has been demonstrated as an important stimulator of metabolic activity
In preclinical studies, PLX4032 rapidly inhibited fluorodeoxyglucose(FDG) uptake specifically in BRAFV600E mutant melanoma cell lines
Therefore, FDG uptake in patients on the PLX4032 phase I trial was assessed using PET imaging before treatment and following two weeks of dosing
All of the assessable patients treated with PLX4032 experienced major reductions in FDG uptake
Bollag G et al., Nature 2010; 467: 596–599
3. Application of biomarkers in Phase I clinical studies: BRAF inhibitor
PET scans for patients taken pre-dose and following 2 weeks of dosing
Reprinted by permission from Macmillan Publishers Ltd: Bollag G et al., Nature 2010; 467: 596–599, copyright (2010)
3. Application of biomarkers in Phase I clinical studies: Immune checkpoint inhibitors
Reprinted by permission from Macmillan Publishers Ltd: Drake, C G et al. Nat Rev Clin Oncol. 2014 Jan; 11(1): 24–37, published on-line 19.3.2013, copyright (2013)
Anti-CTLA-4 antibodies
Anti-PD-1 and anti-PD-L1antibodies
The Immune Synapse
3. Application of biomarkers in Phase I clinical studies: MPDL3280A (anti-PD-L1 antibody)
PD-L1 expression (by tumour cells or tumour-infiltrating immune cells) has emerged as a potential predictive biomarker for PD-1–directed therapy
A urothelial bladder cancer (UBC) expansion cohort of a phase I trial of anti-PD-L1 antibody, MPDL3280A addressed this
Immune cell (but not tumour cell) PD-L1 levels predicted response to MPDL3280A
Results demonstrated that the likelihood of response could be increased by determining the PD-L1 status of tumour-infiltrating immune cells (predictive biomarker)
Powles T et al, Nature, 2014 Nov 27;515(7528):558-62
3. Application of biomarkers in Phase I clinical studies: MPDL3280A (anti-PD-L1)
Immune cell (but not tumour cell) PD-L1 levels predict response to MPDL3280A in UBC
Reprinted by permission from Macmillan Publishers Ltd: Powles T et al., Nature. 2014 Nov 27;515(7528):558-62, copyright (2014)
Positive PD-L1 expression
3. Application of biomarkers in Phase I clinical studies: MPDL3280A (anti-PD-L1)
Immune cell PD-L1 levels predict response to MPDL3280A in UBC
Reprinted by permission from Macmillan Publishers Ltd: Powles T et al., Nature. 2014 Nov 27;515(7528):558-562, copyright (2014)
3. Application of biomarkers in Phase I clinical studies: MPDL3280A (anti-PD-L1 antibody)
Novel biomarkers associated with clinical response to PD-1 directed therapy are still required
Part A of the phase 1 study, in all tumour types, analysed other mechanisms associated with clinical response to the anti-PD-L1 antibody, MPDL3280A
PD-L1 expression, T-helper type 1 gene expression, and CTLA4 expression in pre-treatment samples was associated with clinical response to MPDL3280A
RNA isolated from regressing lesions displayed expression patterns indicative of a generalized activation of CD8 and TH1 T-cell responses
This data suggest that MPDL3280A is most effective in patients in which pre-existing immunity is suppressed by PD-L1, and is re-invigorated on antibody treatment
Herbst RS et al., Nature 2014; 515: 563–567
3. Application of biomarkers in Phase I clinical studies: MPDL3280A (anti-PD-L1 antibody)
PD-L1 expression in pre-treatment samples was associated with clinical response to MPDL3280A
Reprinted by permission from Macmillan Publishers Ltd: Herbst RS et al., Nature 2014; 515: 563–567 copyright (2014)
3. Application of biomarkers in Phase I clinical studies: MPDL3280A (anti-PD-L1 antibody)
Expression of CTLA4 in pre-treatment tumours correlated strongly with response after MPDL3280A
Suggests that CTLA4 may also be a marker of the presence of activated T cells whose functional role as a negative regulator of intra-tumouralT cells appears to be less important than that of PD-L1
Reprinted by permission from Macmillan Publishers Ltd: Herbst RS et al., Nature 2014; 515: 563–567 copyright (2014)
3. Application of biomarkers in Phase I clinical studies: MPDL3280A (anti-PD-L1 antibody)
RNA isolated from regressing lesions, analysed for the presence of transcripts of immunological importance, displayed expression patterns indicative of a generalised activation of CD8 and TH1 T-cell responses
Reprinted by permission from Macmillan Publishers Ltd: Herbst RS et al., Nature 2014; 515: 563–567 copyright (2014)
4. Applications of biomarkers in drug development: Phase II/III clinical studies
Prospective or retrospective collection of tissue samples and performance of molecular analyses to identify mechanisms responsible of biological effects
Could be used as prediction of clinical outcome to identify active drugs for phase III trials
Conclusions
Biomarkers are key components of translational studies, useful in the process of drug development of molecular targeted agents
Biomarkers are useful in the process of dose escalation, definition of schedule and doses for subsequent phase II studies
There is still insufficient knowledge of the dynamic interaction between drug and target to justify a “pharmacodynamically-only” driven strategy.
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