A Micro RNA Polymorphism (MiRSNP) in 3’UTR of K-ras gene was associated with clinical outcome in mCRC patients

treated with either single agent cetuximab (IMCL-0144) or in combination with irinotecan (EPIC)

Thomas Winder1, Wu Zhang1, Anthony El-Khoueiry1, Dongyun Yang1, Alexandra Pohl1, Georg Lurje1, Eric Rowinsky2,

Shirin Khambata-Ford3, Christiane Langer3, Melissa Awad3 and Heinz-Josef Lenz1

1.USC/Norris Comprehensive Cancer Center, Los Angeles, CA 2. Imclone Systems, New York, NY 3. Bristol Myers Squibb Pharmaceutical Res Ins., New York, NY

Introduction

Patients and method

Results

Conclusion

Recent studies have found K-ras mutation status predict resistance to EGFR inhibitors in mCRC(1,2). An in vitro study demonstrated let-7 family of microRNAs can regulate RAS expression by binding to the 3’-UTR of RAS gene(3). Several studies had shown a MicroRNA single nucleotide polymorphism (MiRSNP) in a let-7 microRNA complementary site (lcs6) in the K-ras 3’-UTR region was associated with increased cancer risk in NSCLC and reduced overall survival in oral cancers(4,5). We tested the hypothesis whether this MiRSNP will be associated with response and progression free survival in 130 mCRC patients treated with single agent cetuximab(IMCL-0144) and in 186 mCRC patients treated with cetuximab in combination irinotecan (EPIC) independent of K-ras mutation status.

Our data suggest for the first time that the functional germline polymorphism in K-ras lcs6 may be a potential predictive marker in mCRC patients treated with cetuximab-based chemotherapy independent of K-ras mutation status. This finding warranted further confirmative clinical trials.

K-ras lcs6 SNP was tested in 130 mCRC patients enrolled in IMC-0144 phase II clinical trial (single agent cetuximab) and in 186 mCRC patients enrolled in a second line phase III trial of cetuximab plus irinotecan versus irinotecan alone (EPIC). Patients enrolled in the correlative studies had similar clinical characteristics with the patients enroll in the IMCL-0144 trial but no tissue available. For EPIC trial, the analyzed population is not representative of the treated population. A small percentage of the overall population was analyzed, which is reflected in the efficacy data for the ITT versus analyzed population. In contrast to the results from the overall population, the PFS in the analyzed population is not significantly different between the arms (RR and PFS favor the Cetuximab arm in overall population) and the OS too trends the opposite way. Genomic DNA was extracted from dissected formalin fixed paraffin embedded tumor tissue. K-ras mutation status and the polymorphism were analyzed using direct sequencing and PCR-RFLP technique. Briefly, forward 5’-TTAGGAGAGACGGGGTTTCA-3’ and reverse primer 5’-AAATGAGTTCTGCAAAACAGG-3’ were used for PCR amplification, PCR products were digested by restriction enzyme TfiI (New England Biolab), and alleles were separated on 4% NuSieve ethidium bromide stained agarose gel.

Cetuximab + Irinotecan (n=84)

Irinotecan alone (n=102)

Cetuximab alone (n=130)

Frequency % Frequency % Frequency % Median age, yr (range) 59 (34-85) 61 (25-90) 60 (29-85) Sex Female 44 52 39 38 66 51 Male 40 48 63 62 64 49 Race Asian 5 6 3 3 3 2 Black 13 15 15 15 3 2 Caucasian 61 73 80 78 121 93 Other 5 6 4 4 3 2 ECOG performance status 0 45 54 42 41 52 41 1 36 43 56 55 76 59 2 3 4 4 4 No. of disease sites 1 26 31 29 29 ≥ 2 58 69 72 71 Missing 1 Kras mutation status Wild type 57 68 64 63 88 68 Mutant 27 32 38 37 42 32

Baseline characteristics in patients enrolled in either EPIC or IMCL-0144 correlative study

PR

PRSD

SD

PD

PD

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

K-ras lcs6 TT genotype (n=55) K-ras lcs6 TG+GG genotype (n=12)

P = 0.0189(Fisher's exact)

EPIC IMCL-0144

0.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1.0

0 6 12

Months Since Starting Irinotecan + Cetuximab (Arm A)

Estim

ate

d

Pro

babili

ty

of

PF

S

Log-rank test P = 0.037

K-ras lcs6 T/T (n=21)

K-ras lcs6 TG+GG(n=6)

Progression-free survival by K-ras lcs6 MiRSNP in mutant K-ras of EPIC Arm A(cetuximab+irinotecan)

Tumor response by K-ras lcs6 MiRSNP in wild type K-ras mCRC treat with single agent cetuximab(IMCL-0144)

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