Brain Tumor Update

William A. Freije, M.D.

How to summarize 140,140,533 individual

measurements in twenty five slides

William A. Freije, M.D.

Math

22,283 probesets in 133A chip22,645 probesets in 133B chip44,928 probesets total

20 pairs for each probeset

151 samples

140,140,533 data points total

Tumor types examined to dateGlial tumors

AstrocytomaPilocytic/infantile 2/2Grade II 2Grade III 9

OligodendrogliomaGrade II 8Grade III 11

Mixed oligo/astrocytoma (Grade III) 9Glioblastoma multiforme 63Gliosarcoma 3Ependymoma 3

Poorly differentiated embryonal tumorsMedulloblastoma 5

Menigothelial tumorsMeningioma 17

Nerve sheath tumorsSchwannoma 3

Metastatic tumors 4 TOTAL 141

Summary of completed arrays

• 143 samples from 129 patients– 141 tumor samples– 2 normal brain specimens from GBM pts

• 8 normal brain controls– From autopsy patients

• 151 samples completed on 133A and 133B Affymetrix oligonucleotide arrays

Patient samples- replicates, multiple sampling, progression, and autopsy

• Replicates: same tumor, same biopsy– Mixed oligo/astro (2,1) proceeding to GBM (1)– Oligo III (2) proceeding to GBM (1)

• Multiple sampling with progression analysis– GBM tumor (2) 2nd resection of GBM (2)

• Progression– Oligo III (1) 2 pts proceeding to GBM (1) 2 pts– GBM (1) 2 pts proceeding to 2nd cranio GBM (1) 2 pts

• Autopsy– GBM (1) 2 pts with normal contralateral brain (1) 2 pts

Objectives

• Global microarray classifier to discriminate the major types of primary brain tumors: medulloblastoma, glioma, and meningothelial.– Complement analysis with normal tissue data base.

• Dissect gene expression profiles of glial tumors:– Replace histologic classification scheme with expressed gene

profile classification– Prognosis- identify high and low risk disease– Identify tumors with susceptibility to chemotherapeutic agents – Tumor subclass identification– Define molecular changes associated with disease progression

Objectives

• Explore the different mathematical models and software used to summarize microarray data.– Dchip– Genespring– Statistical analysis

• t test • Multi-dimensional scaling• Random forest prediction• Fisher linear discriminant analysis• Log rank and Wilcoxon nonparametric tests• Kaplan-Meier survival plots

Global microarray classifier

Global microarray classifier

• Gene expression profiles of the three major brain tumor types- medulloblastoma, meningioma, and glial- were examined with:– Multidimensional scaling– Random forest prediction- all gene analysis– Fisher linear discriminant analysis- univariate

MDS plot with 127 samples(normal not included) 12021 genes

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ASTRO.2165

ASTRO.II.1420

ASTRO.II.578

ASTRO.III.1425

ASTRO.III.1704

ASTRO.III.1723

ASTRO.III.587

ASTRO.III.659

ASTRO.III.671

ASTRO.III.672 ASTRO.III.747

DIASTRO.873

G30

G31

G32

G33

G34

G35

G36

G37

G38

G39

G40G41

G42

G43

G44

G45G46

G47G48G49

G50

G51

G52G53G54

G55

G56

G57

G58G59

G60

G61

G62

G63 G64

GBM.1038

GBM.1354

GBM.1414

GBM.1423

GBM.1469

GBM.1478

GBM.1495

GBM.1516

GBM.1521

GBM.1656

GBM.1667

GBM.1675

GBM.1681

GBM.1745

GBM.1798

GBM.1876

GBM.2166

GBM.585

GBM.597GBM.604

GBM.636

GBM.660

GBM.697

GBM.706

GBM.824

GBM.839

GBM.931

GBM.976

GBM.995

GLIOS.631

GLIOS.818

GLIOS.988

MEDUL.1497

MEDUL.1833

MEDUL.745

MEDUL.855 MEDUL.982

MENIN.HIGH.609

MENIN.HIGH.623

MENIN.HIGH.624

MENIN.HIGH.679MENIN.HIGH.776

MENIN.HIGH.795

MENIN.HIGH.822MENIN.HIGH.984

MENIN.LOW.1024

MENIN.LOW.1048MENIN.LOW.1391

MENIN.LOW.600MENIN.LOW.606

MENIN.LOW.607MENIN.LOW.612

MENIN.LOW.682MENIN.LOW.758

MIXED.III.1721MIXED.III.615

MIXED.III.664

MIXED.III.713

MIXED.III.799

MIXED.III.886MIXED.III.912

OLIGO.II.619OLIGO.II.627OLIGO.II.705 OLIGO.II.725

OLIGO.II.766

OLIGO.II.909

OLIGO.II.934

OLIGO.II.936

OLIGO.III.1433

OLIGO.III.1804

OLIGO.III.613

OLIGO.III.650

OLIGO.III.695OLIGO.III.742

OLIGO.III.744

OLIGO.III.975

PILO.1036

PILO.633

PXASTRO.620

Top 30 genes separating medulloblastoma, meningioma, and

gliomas: Random Forest Basic Statistics for Glimos Basic Statistics for Medullo Basic Statistics for Meningomaprobe set gene N MIN MAX MEAN STD N MIN MAX MEAN STD N MIN MAX MEAN STD218730_s_atosteoglycin (osteoinductive factor, mimecan)105 -9.37058 919.505 47.55679 107.9226 5 39.0109 231.761 134.144 77.63038 17 1375.27 4002.25 2574.354 608.1179214628_at nescient helix loop helix 1105 -47.2857 987.054 44.21422 108.4981 5 325.683 3104.73 1867.303 1010.653 17 -29.7336 11.0561 -2.12149 10.34009213033_s_atConsensus includes gb:AI186739 /FEA=EST /DB_XREF=gi:3737377 /DB_XREF=est:qe79c01.x1 /CLONE=IMAGE:1745184 /UG=Hs.326416 Homo sapiens mRNA; cDNA DKFZp564H1916 (from clone DKFZp564H1916)105 44.034 338.369 158.2436 65.31504 5 430.84 1033 764.0866 258.5299 17 70.707 191.878 121.1015 30.3049205908_s_atosteomodulin 105 0.295823 545.358 32.18404 54.58468 5 4.3044 79.9094 32.05136 28.60105 17 528.462 1705.21 1284.898 347.42201131_s_atcadherin 1, type 1, E-cadherin (epithelial)105 -25.3469 267.363 36.28974 51.80603 5 -17.3901 -0.199821 -10.00296 8.435334 17 836.099 3352.95 1631.388 686.7499211354_s_atleptin receptor 105 7.35465 178.686 43.21471 24.93849 5 41.388 135.453 73.77012 36.78924 17 46.3518 3430.86 1561.565 1027.232209333_at unc-51-like kinase 1 (C. elegans)105 82.9791 421.414 229.8967 73.94169 5 332.812 889.844 561.0934 207.8361 17 114.158 368.692 206.9113 65.03922215304_at Consensus includes gb:U79293.1 /DEF=Human clone 23948 mRNA sequence. /FEA=mRNA /DB_XREF=gi:1710274 /UG=Hs.159264 Human clone 23948 mRNA sequence105 67.1945 172.208 108.4256 21.3167 5 84.0341 148.279 113.7452 23.98376 17 162.874 531.842 342.5836 113.1816210613_s_atsynaptogyrin 1 105 46.5661 257.959 119.1586 38.00912 5 75.8846 152.798 104.1902 32.02954 17 260.392 865.304 496.6782 181.8693201951_at activated leucocyte cell adhesion molecule105 76.1528 1854.63 851.7832 402.9412 5 -14.0933 69.4731 24.80122 37.15646 17 803.548 2397.63 1741.075 477.9259201779_s_atring finger protein 13105 650.239 2596.45 1283.324 382.4834 5 288.457 683.202 447.2462 184.8749 17 669.096 1612.62 1251.202 227.229215894_at prostaglandin D2 receptor (DP)105 -11.5355 149.623 10.46443 16.16079 5 0.098112 28.2224 17.84952 11.79279 17 164.109 2138.8 867.4683 546.0132205817_at sine oculis homeobox homolog 1 (Drosophila)105 -11.5102 288.029 46.83396 59.61075 5 -3.13016 8.2915 3.875953 4.888203 17 338.796 1867.88 794.6298 421.2563218445_at core histone macroH2A2.2105 28.59 649.193 247.5455 160.4923 5 541.18 1719.58 1016.707 455.6534 17 131.34 774.38 347.0988 158.517212753_at ring finger protein 3105 382.061 881.465 605.1936 106.0061 5 723.43 1030.33 940.576 124.7212 17 441.633 654.843 549.1284 68.42664218019_s_athypothetical protein FLJ 21324105 413.144 1846.68 979.5283 341.2896 5 141.728 484.967 282.6928 137.4829 17 643.823 2808.3 1449.522 588.4715210220_at frizzled homolog 2 (Drosophila)105 -60.139 251.71 33.05856 56.01755 5 100.461 254.455 180.3838 67.45427 17 302.127 1073.77 650.7259 244.8725203140_at B-cell CLL/lymphoma 6 (zinc finger protein 51)105 506.601 3326.01 1392.414 543.1683 5 69.5947 1271.02 429.0395 478.9433 17 583.225 2263.6 1082.263 418.0388217767_at complement component 3105 132.734 6229.87 2230.454 1447.201 5 7.08325 157.612 57.71707 59.87464 17 281.284 3261.5 1331.695 783.892206502_s_atinsulinoma-associated 1105 -16.3835 1839.12 278.1431 348.4116 5 1918.54 3830.36 2970.23 760.8547 17 -14.2498 6.39007 -6.585445 4.951107220464_at hypothetical protein FLJ 12122105 31.6528 306.665 200.9388 46.49041 5 164.053 354.348 294.0548 75.569 17 100.774 204.925 158.7797 32.45294209170_s_atglycoprotein M6B 105 467.012 7060.63 4307.835 1029.579 5 346.769 966.088 709.8858 251.6957 17 -48.4947 447.49 36.63054 110.1389204334_at Kruppel-like factor 7 (ubiquitous)105 152.804 556.368 286.0404 78.03113 5 529.31 1130.07 723.2574 245.5566 17 154.924 401.675 243.1059 67.33611203453_at sodium channel, nonvoltage-gated 1 alpha105 -29.2737 236.908 -2.439193 25.22252 5 -33.0986 -9.38667 -26.06129 9.550012 17 41.113 1774.18 344.4032 439.8941204284_at protein phosphatase 1, regulatory (inhibitor) subunit 3C105 125.231 2996.61 506.8364 370.4084 5 36.3986 60.0269 48.63366 10.05597 17 82.8482 3605.78 963.3741 849.123201161_s_atcold shock domain protein A105 -13.3354 1003.34 313.6028 231.9014 5 -36.8945 224.99 53.52776 104.3266 17 1098.48 3888.19 2268.86 707.769203723_at inositol 1,4,5-trisphosphate 3-kinase B105 78.5692 3389.8 1061.958 560.2612 5 42.027 131.529 91.4661 39.70138 17 467.452 1633.87 805.1482 318.71201160_s_atcold shock domain protein A105 217.168 2057.08 819.1187 432.0559 5 103.204 635.205 315.257 195.9336 17 2101.58 3852.77 2782.311 439.4509212812_at Consensus includes gb:AI700633 /FEA=EST /DB_XREF=gi:4988533 /DB_XREF=est:we38g03.x1 /CLONE=IMAGE:2343412 /UG=Hs.288232 Homo sapiens cDNA: FLJ 22642 fis, clone HSI06970105 405.402 2165.57 1155.531 409.5289 5 169.065 401.065 263.3256 85.53858 17 373.612 2244.18 806.5341 516.4613221472_at adenosine deaminase105 327.501 1514.87 690.4505 202.7465 5 211.109 380.162 289.5644 67.21175 17 496.196 1092.41 681.4449 165.4245

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Boxplots of probesets identified by Random Forest Predictionwhich separate medulloblastoma, meningioma, and glioma

Basic Statistics for Glimos Basic Statistics for Medullo Basic Statistics for Meningomaprobe set gene N MIN MAX MEAN STD N MIN MAX MEAN STD N MIN MAX MEAN STD213033_s_atUG= Hs.326416 Homo sapiens mRNA 105 44.034 338.369 158.244 65.315 5 430.84 1033 764.087 258.53 17 70.707 191.878 121.101 30.3049

Probeset 213033_s_at: medulloblastoma specific gene

Normaltissueproject

Brain tumor data:

complementary

Probe Set Gene Name GENE EXPRESSION PROFILE Rsq218730_s_atosteoglycin (osteoinductive factor, mimecan)osteoglycin (osteoinductive factor, mimecan) 0.929119200606_at desmoplakin (DPI, DPII) desmoplakin (DPI, DPII) 0.917703205908_s_atosteomodulin osteomodulin 0.911471213260_at 0 0.893029208613_s_atfilamin B, beta (actin binding protein 278) filamin B, beta (actin binding protein 278) 0.892358205907_s_atosteomodulin osteomodulin 0.891166213075_at 0 0.862349208614_s_atfilamin B, beta (actin binding protein 278) filamin B, beta (actin binding protein 278) 0.857919209211_at Kruppel-like factor 5 (intestinal) Kruppel-like factor 5 (intestinal) 0.857414219525_at hypothetical protein FLJ10847 hypothetical protein FLJ 10847 0.853462204049_s_atKIAA0680 gene product KIAA0680 gene product 0.847858209212_s_atKruppel-like factor 5 (intestinal) Kruppel-like factor 5 (intestinal) 0.839219901_at hypothetical protein FLJ11183 hypothetical protein FLJ 11183 0.833329219561_at COPZ2 for nonclathrin coat protein zeta-COPCOPZ2 for nonclathrin coat protein zeta-COP 0.827962201131_s_atcadherin 1, type 1, E-cadherin (epithelial) cadherin 1, type 1, E-cadherin (epithelial) 0.827628209264_s_attransmembrane 4 superfamily member 7 transmembrane 4 superfamily member 7 0.823607204048_s_atKIAA0680 gene product KIAA0680 gene product 0.817975214106_s_atGDP-mannose 4,6-dehydratase GDP-mannose 4,6-dehydratase 0.815178210220_at frizzled homolog 2 (Drosophila) frizzled homolog 2 (Drosophila) 0.81113201161_s_atcold shock domain protein A cold shock domain protein A 0.807587221814_at tumor endothelial marker 5 precursor tumor endothelial marker 5 precursor 0.806426206307_s_atforkhead box D1 forkhead box D1 0.804335220419_s_atubiquitin specific protease 25 ubiquitin specific protease 25 0.800923213429_at 0 0.799371204526_s_atvascular Rab-GAP/TBC-containing vascular Rab-GAP/TBC-containing 0.798332217889_s_atduodenal cytochrome b duodenal cytochrome b 0.796698201332_s_atsignal transducer and activator of transcription 6, interleukin-4 inducedsignal transducer and activator of transcription 6, interleukin-4 induced0.785545208796_s_atcyclin G1 cyclin G1 0.785375209894_at leptin receptor leptin receptor 0.784983

Top 30 genes separating medulloblastoma, meningioma, and gliomas: Fisher Linear Discriminant Assay

GBM analysis

GBM samples: de novo vs progressive samples

• 63 samples total from 57 patients

• 48 de novo samples from 46 patients

• 15 progressive samples from 11 patients

• 3 de novo samples are autopsy specimens

• 1 progressive sample is from autopsy

• De novo– Treated– Untreated– Total 48

• Progressive– Treated– Untreated– Total 15

GBM samples: de novo vs progressive samples

Clinical parameters

• TTP- time to progression– Time in days from diagnosis or intervention (at UCLA)

to the first evidence of disease progression

• TTS- time to survival– Time in days from diagnosis or intervention (at UCLA)

to the current day if alive or to the day of death

• Survival time– Time in days from initial diagnosis to the current day if

alive or to the day of death

Birth Clinicalidentification

Disease

1st Rx

DEATH

2nd Rx

Progression

TTS 1stTTP 1st

TTS 2ndTTP 2nd

TTP and TTS are relatedto time of treatment

Unknown

Survival

• TTP (Days)– Range 7-1031 – Mean 188– Median 115– SD 226

• TTP Alive Dead– Median 219 79

GBM clinical parameters- TTP

• TTS (Days)– Range 7-1247– Mean 367– Median 236 – SD 328

• TTS Alive Dead– Median 927 187

GBM clinical parameters- TTS

• Survival (Days)– Range 56-8330– Mean 850– Median 541 – SD 1191

• Survival Alive Dead– Median 936 520

GBM clinical parameters- Survival

GBM clinical parameters- outliers

• Long term survivors: all patients with TTS> 723 days. This group of patients all have TTP that are > 612 days (TTP mean + 2 SD). 10 patients in this group

• Short term survivors: all patients with TTP < 60 days. This group of patients all had a TTS < 150 days. 13 patients in this group.

• All short term survivors are being reviewed for neurodecline secondary to other causes.

GBM living patients

• 13/57 GBM patients are alive

• 4/13 alive GBM patients have a TTP>300 (range 659-1311). All of these patients were identified in the outlier group.

GBM survival analysisstatistical methods employed

• R square of LDA to determine number of groups

• Best split measures

• Wilcoxon test statistics

• Logrank test statistics

• Kaplan-Meier survival plots

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Determine Number of Clusters by Separating the 63 samples with gene expression ( with top 2000 brain tissue genes by LDA)

probe set gene improve cutpoints wilcoxon logrank Mean SD CV204081_at neurogranin (protein kinase C substrate, RC3)16.35477 118.091 5.072859 5.349787 1176.9 1446.504 122.908202341_s_attripartite motif-containing 2 17.72539 257.5935 4.644394 6.444972 295.2165 245.6886 83.22319205967_at H4 histone family, member G 17.34052 589.2245 13.90282 20.00598 814.3877 487.3 59.83636204792_s_atKIAA0590 gene product 17.61183 40.50325 6.24666 6.385589 42.88363 22.33911 52.0924221974_at imprinted in Prader-Willi syndrome 16.50345 408.8255 12.79754 12.31084 355.81 154.2864 43.36203213038_at Consensus includes gb:AL031602 /DEF=Human DNA sequence from clone RP5-1174N9 on chromosome 1p34.1-35.3. Contains the gene for a novel protein with IBR domain, a (pseudo?) gene for a novel protein similar to MT1E (metallothionein 1E (functional)), ESTs, S19.92423 186.636 10.94211 8.488861 155.3839 65.39862 42.08841214943_s_atKIAA0117 protein 15.83049 145.8245 5.38903 5.387896 116.9904 44.42918 37.97677215673_at KIAA1655 protein 15.09382 215.703 7.150397 9.296527 188.744 67.29094 35.65197215173_at Consensus includes gb:N46852 /FEA=EST /DB_XREF=gi:1188018 /DB_XREF=est:yy73d05.s1 /CLONE=IMAGE:279177 /UG=Hs.285625 Homo sapiens mRNA; cDNA DKFZp434A119 (from clone DKFZp434A119)17.80204 81.40775 15.59632 16.52646 66.71456 22.28061 33.39692210479_s_atRAR-related orphan receptor A 16.54388 99.41775 9.233742 11.65061 97.49982 31.57559 32.38528209689_at hypothetical protein MGC13033 16.69765 71.9006 6.222114 5.233312 74.11629 23.77174 32.07357211605_s_atgb:U41742.1 /DEF=Human nucleophosmin-retinoic acid receptor alpha fusion protein NPM-RAR long form mRNA, complete cds. /FEA=mRNA /PROD=nucleophosmin-retinoic acid receptor alphafusion protein NPM-RAR long form /DB_XREF=gi:1314307 /FL=gb:U41742.115.24191 52.1697 8.269468 8.870645 49.07062 15.38787 31.35863207101_at vesicle-associated membrane protein 1 (synaptobrevin 1)15.84204 57.28035 13.1415 14.20455 57.17401 17.12364 29.95004215869_at Consensus includes gb:AK022254.1 /DEF=Homo sapiens cDNA FLJ12192 fis, clone MAMMA1000851. /FEA=mRNA /DB_XREF=gi:10433612 /UG=Hs.288451 Homo sapiens cDNA FLJ12192 fis, clone MAMMA100085118.53496 31.6195 2.239083 5.616346 39.0193 11.44799 29.33929206837_at cartilage paired-class homeoprotein 1 16.42133 56.0869 7.84732 10.2639 55.89611 16.33592 29.22551215816_at immunoglobulin lambda-like polypeptide 215.29086 88.26045 14.23423 15.20453 84.10287 23.13337 27.50603202119_s_atcopine III 17.20225 1008.16 7.887961 6.846188 895.8745 241.0367 26.90518215581_s_atConsensus includes gb:AK022303.1 /DEF=Homo sapiens cDNA FLJ12241 fis, clone MAMMA1001274. /FEA=mRNA /DB_XREF=gi:10433670 /UG=Hs.287503 Homo sapiens cDNA FLJ12241 fis, clone MAMMA100127418.02766 229.978 8.241964 12.61092 204.1595 53.87562 26.38898221136_at growth differentiation factor 2 17.05797 87.1404 15.90917 18.35625 78.07688 20.16395 25.82577220284_at soggy-1 gene 15.91762 108.1905 6.5466 6.660714 121.6557 31.01458 25.49375209622_at serine/threonine kinase 16 15.07834 107.775 2.072865 4.397244 107.8419 27.26403 25.28147216330_s_atPOU domain, class 6, transcription factor 117.10343 83.9075 5.190436 8.524013 84.41992 20.58942 24.38929202745_at ubiquitin specific protease 8 15.8798 284.2635 5.164615 6.35934 282.3306 67.81733 24.02054211949_s_atnucleolar and coiled-body phosphprotein 116.08631 98.22115 4.111311 7.752579 99.60225 23.09385 23.18608203958_s_atKIAA0478 gene product 15.07929 94.5248 5.454099 7.360786 83.17468 18.92072 22.74817204031_s_atpoly(rC) binding protein 2 15.18957 2386.03 1.702373 3.292649 2126.125 473.6835 22.27919216615_s_atConsensus includes gb:AJ005205 /DEF=Homo sapiens 5HT3 gene for serotonin 3 receptor /FEA=mRNA_2 /DB_XREF=gi:7019744 /UG=Hs.2142 5-hydroxytryptamine (serotonin) receptor 3A15.68437 126.743 5.741019 6.291753 136.4631 30.33506 22.22949208474_at claudin 6 16.63842 121.794 23.78793 20.83049 110.7207 24.46725 22.09817217357_at Consensus includes gb:AB028239 /DEF=Homo sapiens pseudogene for necdin /FEA=CDS /DB_XREF=gi:5672690 /UG=Hs.247843 Homo sapiens pseudogene for necdin19.24788 83.49165 19.70104 18.56972 80.05301 17.46124 21.8121207106_s_atleukocyte tyrosine kinase 15.71643 81.42275 17.83355 21.08793 75.69654 16.37434 21.63156213279_at Consensus includes gb:AL050217.1 /DEF=Homo sapiens mRNA; cDNA DKFZp586I0523 (from clone DKFZp586I0523). /FEA=mRNA /DB_XREF=gi:4884458 /UG=Hs.308467 Homo sapiens mRNA; cDNA DKFZp586I0523 (from clone DKFZp586I0523)15.54995 290.359 6.633992 6.701037 273.3557 58.62482 21.44635218972_at hypothetical protein FLJ10890 15.53518 508.754 5.938228 9.052359 497.6981 105.6474 21.2272209411_s_atgolgi associated, gamma adaptin ear containing, ARF binding protein 315.26845 292.641 6.229391 9.161279 278.7253 57.36923 20.58271214341_at adaptor-related protein complex 1, gamma 2 subunit16.53618 107.8035 8.886534 8.689626 116.7656 21.89439 18.75071222236_s_athypothetical protein FLJ20199 16.61413 484.326 11.77899 10.64769 484.0967 89.21855 18.4299215219_at Consensus includes gb:AK025095.1 /DEF=Homo sapiens cDNA: FLJ21442 fis, clone COL04429, highly similar to HSA237839 Homo sapiens mRNA for hypothetical protein. /FEA=mRNA /DB_XREF=gi:10437542 /UG=Hs.306763 Homo sapiens cDNA: FLJ21442 fis, clone COL04429,17.69959 156.658 9.674779 9.667617 149.0093 23.48816 15.76288

Association between gene expression and survival time in GBM samples:top 36 genes

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204792.s.at , p-value= 1.5e-05

Time to death(days), cut= 40.503

Sur

viva

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lowhigh

Kaplan-Meier survival curves for specific probesets

Future

• Turn this information into clinical difference• Develop a model through insight developed from the

microarray screen• Confirm RT-PCR• Develop specific assays for genes thought to be

important in GBM survival (Ab)• Cell line models• Invasion assay• Mouse models• Example: IGF2 highly expressed in poor survivors:

– Global epigenetic imprint altered in GBM poor survival?

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2

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1 3 5 7 9 11 13 15 17 19 21 23 25 27 29 31 33

Series1

Series2

IGF2 expression in GBM long and short survivors

Graph courtesy of E. Castro