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Figure 1.  Flowchart of PETACC-8 Trial Molecular Study Evaluating the Prognostic Impact of Mismatch Repair, KRAS, and BRAF Status
Flowchart of PETACC-8 Trial Molecular Study Evaluating the Prognostic Impact of Mismatch Repair, KRAS, and BRAF Status
Figure 2.  Kaplan-Meier Curves for Disease-Free Survival According to Mismatch Repair, KRAS, and BRAF Status
Kaplan-Meier Curves for Disease-Free Survival According to Mismatch Repair, KRAS, and BRAF Status

HR indicates hazard ratio.

Figure 3.  Effect of KRAS and BRAF Status on Disease-Free Survival (DFS) in Patients With Microsatellite-Stable and Microsatellite-Unstable Tumors
Effect of KRAS and BRAF Status on Disease-Free Survival (DFS) in Patients With Microsatellite-Stable and Microsatellite-Unstable Tumors

HR indicates hazard ratio.

Table 1.  Clinical and Pathological Characteristics According to Mismatch Repair (MMR), KRAS, and BRAF Status in the Present Study Population
Clinical and Pathological Characteristics According to Mismatch Repair (MMR), KRAS, and BRAF Status in the Present Study Population
Table 2.  Univariate Cox Proportional Hazards Regression Models for Disease-Free Survival (DFS) and Overall Survival (OS)
Univariate Cox Proportional Hazards Regression Models for Disease-Free Survival (DFS) and Overall Survival (OS)
Table 3.  Multivariate Cox Proportional Hazards Regression Models for Disease-Free Survival (DFS) and Overall Survival (OS)
Multivariate Cox Proportional Hazards Regression Models for Disease-Free Survival (DFS) and Overall Survival (OS)
1.
Jemal  A, Bray  F, Center  MM, Ferlay  J, Ward  E, Forman  D.  Global cancer statistics.  CA Cancer J Clin. 2011;61(2):69-90.PubMedGoogle ScholarCrossref
2.
Ionov  Y, Peinado  MA, Malkhosyan  S, Shibata  D, Perucho  M.  Ubiquitous somatic mutations in simple repeated sequences reveal a new mechanism for colonic carcinogenesis.  Nature. 1993;363(6429):558-561.PubMedGoogle ScholarCrossref
3.
Peltomäki  P, Lothe  RA, Aaltonen  LA,  et al.  Microsatellite instability is associated with tumors that characterize the hereditary non-polyposis colorectal carcinoma syndrome.  Cancer Res. 1993;53(24):5853-5855.PubMedGoogle Scholar
4.
Herman  JG, Umar  A, Polyak  K,  et al.  Incidence and functional consequences of hMLH1 promoter hypermethylation in colorectal carcinoma.  Proc Natl Acad Sci U S A. 1998;95(12):6870-6875.PubMedGoogle ScholarCrossref
5.
Weisenberger  DJ, Siegmund  KD, Campan  M,  et al.  CpG island methylator phenotype underlies sporadic microsatellite instability and is tightly associated with BRAF mutation in colorectal cancer.  Nat Genet. 2006;38(7):787-793.PubMedGoogle ScholarCrossref
6.
Domingo  E, Niessen  RC, Oliveira  C,  et al.  BRAF-V600E is not involved in the colorectal tumorigenesis of HNPCC in patients with functional MLH1 and MSH2 genes.  Oncogene. 2005;24(24):3995-3998.PubMedGoogle ScholarCrossref
7.
Roth  AD, Tejpar  S, Delorenzi  M,  et al.  Prognostic role of KRAS and BRAF in stage II and III resected colon cancer: results of the translational study on the PETACC-3, EORTC 40993, SAKK 60-00 trial.  J Clin Oncol. 2010;28(3):466-474.PubMedGoogle ScholarCrossref
8.
Van Cutsem  E, Köhne  CH, Láng  I,  et al.  Cetuximab plus irinotecan, fluorouracil, and leucovorin as first-line treatment for metastatic colorectal cancer: updated analysis of overall survival according to tumor KRAS and BRAF mutation status.  J Clin Oncol. 2011;29(15):2011-2019.PubMedGoogle ScholarCrossref
9.
Karapetis  CS, Khambata-Ford  S, Jonker  DJ,  et al.  K-ras mutations and benefit from cetuximab in advanced colorectal cancer.  N Engl J Med. 2008;359(17):1757-1765.PubMedGoogle ScholarCrossref
10.
Van Cutsem  E, Labianca  R, Bodoky  G,  et al.  Randomized phase III trial comparing biweekly infusional fluorouracil/leucovorin alone or with irinotecan in the adjuvant treatment of stage III colon cancer: PETACC-3.  J Clin Oncol. 2009;27(19):3117-3125.PubMedGoogle ScholarCrossref
11.
De Roock  W, Claes  B, Bernasconi  D,  et al.  Effects of KRAS, BRAF, NRAS, and PIK3CA mutations on the efficacy of cetuximab plus chemotherapy in chemotherapy-refractory metastatic colorectal cancer: a retrospective consortium analysis.  Lancet Oncol. 2010;11(8):753-762.PubMedGoogle ScholarCrossref
12.
Yokota  T, Ura  T, Shibata  N,  et al.  BRAF mutation is a powerful prognostic factor in advanced and recurrent colorectal cancer.  Br J Cancer. 2011;104(5):856-862.PubMedGoogle ScholarCrossref
13.
Andreyev  HJ, Norman  AR, Cunningham  D, Oates  JR, Clarke  PA.  Kirsten ras mutations in patients with colorectal cancer: the multicenter “RASCAL” study.  J Natl Cancer Inst. 1998;90(9):675-684.PubMedGoogle ScholarCrossref
14.
Gavin  PG, Colangelo  LH, Fumagalli  D,  et al.  Mutation profiling and microsatellite instability in stage II and III colon cancer: an assessment of their prognostic and oxaliplatin predictive value.  Clin Cancer Res. 2012;18(23):6531-6541.PubMedGoogle ScholarCrossref
15.
Yoon  HH, Tougeron  D, Shi  Q,  et al; Alliance for Clinical Trials in Oncology.  KRAS codon 12 and 13 mutations in relation to disease-free survival in BRAF-wild-type stage III colon cancers from an adjuvant chemotherapy trial (N0147 Alliance).  Clin Cancer Res. 2014;20(11):3033-3043.PubMedGoogle ScholarCrossref
16.
Blons  H, Emile  JF, Le Malicot  K,  et al.  Prognostic value of KRAS mutations in stage III colon cancer: post hoc analysis of the PETACC-8 phase III trial dataset.  Ann Oncol. 2014;25(12):2378-2385.PubMedGoogle ScholarCrossref
17.
Popovici  V, Budinska  E, Tejpar  S,  et al.  Identification of a poor-prognosis BRAF-mutant-like population of patients with colon cancer.  J Clin Oncol. 2012;30(12):1288-1295.PubMedGoogle ScholarCrossref
18.
Ogino  S, Shima  K, Meyerhardt  JA,  et al.  Predictive and prognostic roles of BRAF mutation in stage III colon cancer: results from intergroup trial CALGB 89803.  Clin Cancer Res. 2012;18(3):890-900.PubMedGoogle ScholarCrossref
19.
Sinicrope  FA, Mahoney  MR, Smyrk  TC,  et al.  Prognostic impact of deficient DNA mismatch repair in patients with stage III colon cancer from a randomized trial of FOLFOX-based adjuvant chemotherapy.  J Clin Oncol. 2013;31(29):3664-3672.PubMedGoogle ScholarCrossref
20.
Ribic  CM, Sargent  DJ, Moore  MJ,  et al.  Tumor microsatellite-instability status as a predictor of benefit from fluorouracil-based adjuvant chemotherapy for colon cancer.  N Engl J Med. 2003;349(3):247-257.PubMedGoogle ScholarCrossref
21.
Sargent  DJ, Marsoni  S, Monges  G,  et al.  Defective mismatch repair as a predictive marker for lack of efficacy of fluorouracil-based adjuvant therapy in colon cancer.  J Clin Oncol. 2010;28(20):3219-3226.PubMedGoogle ScholarCrossref
22.
Taieb  J, Tabernero  J, Mini  E,  et al; PETACC-8 Study Investigators.  Oxaliplatin, fluorouracil, and leucovorin with or without cetuximab in patients with resected stage III colon cancer (PETACC-8): an open-label, randomised phase 3 trial.  Lancet Oncol. 2014;15(8):862-873.PubMedGoogle ScholarCrossref
23.
Zaanan  A, Cuilliere-Dartigues  P, Guilloux  A,  et al.  Impact of p53 expression and microsatellite instability on stage III colon cancer disease-free survival in patients treated by 5-fluorouracil and leucovorin with or without oxaliplatin.  Ann Oncol. 2010;21(4):772-780.PubMedGoogle ScholarCrossref
24.
Umar  A, Boland  CR, Terdiman  JP,  et al.  Revised Bethesda Guidelines for hereditary nonpolyposis colorectal cancer (Lynch syndrome) and microsatellite instability.  J Natl Cancer Inst. 2004;96(4):261-268.PubMedGoogle ScholarCrossref
25.
Blons  H, Rouleau  E, Charrier  N,  et al; MOKAECM Collaborative Group.  Performance and cost efficiency of KRAS mutation testing for metastatic colorectal cancer in routine diagnosis: the MOKAECM study, a nationwide experience.  PLoS One. 2013;8(7):e68945.PubMedGoogle ScholarCrossref
26.
Samowitz  WS, Curtin  K, Ma  KN,  et al.  Microsatellite instability in sporadic colon cancer is associated with an improved prognosis at the population level.  Cancer Epidemiol Biomarkers Prev. 2001;10(9):917-923.PubMedGoogle Scholar
27.
Sorbye  H, Dragomir  A, Sundström  M,  et al.  High BRAF mutation frequency and marked survival differences in subgroups according to KRAS/BRAF mutation status and tumor tissue availability in a prospective population-based metastatic colorectal cancer cohort.  PLoS One. 2015;10(6):e0131046.PubMedGoogle ScholarCrossref
28.
Janakiraman  M, Vakiani  E, Zeng  Z,  et al.  Genomic and biological characterization of exon 4 KRAS mutations in human cancer.  Cancer Res. 2010;70(14):5901-5911.PubMedGoogle ScholarCrossref
29.
Popat  S, Hubner  R, Houlston  RS.  Systematic review of microsatellite instability and colorectal cancer prognosis.  J Clin Oncol. 2005;23(3):609-618.PubMedGoogle ScholarCrossref
30.
Lanza  G, Gafà  R, Santini  A, Maestri  I, Guerzoni  L, Cavazzini  L.  Immunohistochemical test for MLH1 and MSH2 expression predicts clinical outcome in stage II and III colorectal cancer patients.  J Clin Oncol. 2006;24(15):2359-2367.PubMedGoogle ScholarCrossref
31.
Halling  KC, French  AJ, McDonnell  SK,  et al.  Microsatellite instability and 8p allelic imbalance in stage B2 and C colorectal cancers.  J Natl Cancer Inst. 1999;91(15):1295-1303.PubMedGoogle ScholarCrossref
32.
Zaanan  A, Fléjou  JF, Emile  JF,  et al.  Defective mismatch repair status as a prognostic biomarker of disease-free survival in stage III colon cancer patients treated with adjuvant FOLFOX chemotherapy.  Clin Cancer Res. 2011;17(23):7470-7478.PubMedGoogle ScholarCrossref
33.
Barnetson  RA, Tenesa  A, Farrington  SM,  et al.  Identification and survival of carriers of mutations in DNA mismatch-repair genes in colon cancer.  N Engl J Med. 2006;354(26):2751-2763.PubMedGoogle ScholarCrossref
34.
Kim  GP, Colangelo  LH, Wieand  HS,  et al; National Cancer Institute.  Prognostic and predictive roles of high-degree microsatellite instability in colon cancer: a National Cancer Institute-National Surgical Adjuvant Breast and Bowel Project collaborative study.  J Clin Oncol. 2007;25(7):767-772.PubMedGoogle ScholarCrossref
35.
Salahshor  S, Kressner  U, Fischer  H,  et al.  Microsatellite instability in sporadic colorectal cancer is not an independent prognostic factor.  Br J Cancer. 1999;81(2):190-193.PubMedGoogle ScholarCrossref
36.
Roth  AD, Delorenzi  M, Tejpar  S,  et al.  Integrated analysis of molecular and clinical prognostic factors in stage II/III colon cancer.  J Natl Cancer Inst. 2012;104(21):1635-1646.PubMedGoogle ScholarCrossref
37.
Fink  D, Zheng  H, Nebel  S,  et al.  In vitro and in vivo resistance to cisplatin in cells that have lost DNA mismatch repair.  Cancer Res. 1997;57(10):1841-1845.PubMedGoogle Scholar
38.
Carethers  JM, Chauhan  DP, Fink  D,  et al.  Mismatch repair proficiency and in vitro response to 5-fluorouracil.  Gastroenterology. 1999;117(1):123-131.PubMedGoogle ScholarCrossref
39.
Shen  L, Catalano  PJ, Benson  AB  III, O’Dwyer  P, Hamilton  SR, Issa  JP.  Association between DNA methylation and shortened survival in patients with advanced colorectal cancer treated with 5-fluorouracil based chemotherapy.  Clin Cancer Res. 2007;13(20):6093-6098.PubMedGoogle ScholarCrossref
40.
Van Rijnsoever  M, Elsaleh  H, Joseph  D, McCaul  K, Iacopetta  B.  CpG island methylator phenotype is an independent predictor of survival benefit from 5-fluorouracil in stage III colorectal cancer.  Clin Cancer Res. 2003;9(8):2898-2903.PubMedGoogle Scholar
41.
Jover  R, Nguyen  TP, Pérez-Carbonell  L,  et al.  5-Fluorouracil adjuvant chemotherapy does not increase survival in patients with CpG island methylator phenotype colorectal cancer.  Gastroenterology. 2011;140(4):1174-1181.PubMedGoogle ScholarCrossref
42.
Han  SW, Lee  HJ, Bae  JM,  et al.  Methylation and microsatellite status and recurrence following adjuvant FOLFOX in colorectal cancer.  Int J Cancer. 2013;132(9):2209-2216.PubMedGoogle ScholarCrossref
43.
Andreyev  HJ, Norman  AR, Cunningham  D,  et al.  Kirsten ras mutations in patients with colorectal cancer: the “RASCAL II” study.  Br J Cancer. 2001;85(5):692-696.PubMedGoogle ScholarCrossref
44.
Westra  JL, Schaapveld  M, Hollema  H,  et al.  Determination of TP53 mutation is more relevant than microsatellite instability status for the prediction of disease-free survival in adjuvant-treated stage III colon cancer patients.  J Clin Oncol. 2005;23(24):5635-5643.PubMedGoogle ScholarCrossref
45.
Ogino  S, Meyerhardt  JA, Irahara  N,  et al; Cancer and Leukemia Group B; North Central Cancer Treatment Group; Canadian Cancer Society Research Institute; Southwest Oncology Group.  KRAS mutation in stage III colon cancer and clinical outcome following intergroup trial CALGB 89803.  Clin Cancer Res. 2009;15(23):7322-7329.PubMedGoogle ScholarCrossref
46.
Bokemeyer  C, Bondarenko  I, Makhson  A,  et al.  Fluorouracil, leucovorin, and oxaliplatin with and without cetuximab in the first-line treatment of metastatic colorectal cancer.  J Clin Oncol. 2009;27(5):663-671.PubMedGoogle ScholarCrossref
47.
Douillard  JY, Oliner  KS, Siena  S,  et al.  Panitumumab-FOLFOX4 treatment and RAS mutations in colorectal cancer.  N Engl J Med. 2013;369(11):1023-1034.PubMedGoogle ScholarCrossref
48.
Van Cutsem  E, Köhne  CH, Hitre  E,  et al.  Cetuximab and chemotherapy as initial treatment for metastatic colorectal cancer.  N Engl J Med. 2009;360(14):1408-1417.PubMedGoogle ScholarCrossref
49.
Van Cutsem  E, Lenz  HJ, Köhne  CH,  et al.  Fluorouracil, leucovorin, and irinotecan plus cetuximab treatment and RAS mutations in colorectal cancer.  J Clin Oncol. 2015;33(7):692-700.PubMedGoogle ScholarCrossref
Original Investigation
May 2016

Prognostic Effect of BRAF and KRAS Mutations in Patients With Stage III Colon Cancer Treated With Leucovorin, Fluorouracil, and Oxaliplatin With or Without Cetuximab: A Post Hoc Analysis of the PETACC-8 Trial

Author Affiliations
  • 1Paris Descartes University, Department of Digestive Oncology, European Georges Pompidou Hospital, Assistance Publique-Hôpitaux de Paris, France
  • 2Centre de Recherché UMR-S 1147, Médecine Personnalisée, Pharmacogénomique, Optimisation Thérapeutique, Institut National de la Santé et de la Recherche Médicale, Paris, France
  • 3Department of Statistics, Fédération Francophone de Cancérologie Digestive, Dijon, France
  • 4Department of Pathology, Ambroise Paré Hospital, Assistance Publique–Hôpitaux de Paris, Boulogne-Billancourt, France
  • 5Versailles Saint-Quentin-en-Yvelines University, Boulogne-Billancourt, France
  • 6Paris Descartes University, Department of Biology, European Georges Pompidou Hospital, Assistance Publique–Hôpitaux de Paris, France
  • 7Department of Oncology, Sainte Catherine Institute, Avignon, France
  • 8Department of Oncology, Cancérologie de l’Ouest Institute, Nantes, France
  • 9Medical Oncology Department, Vall d’Hebron University Hospital and Institute of Oncology, Barcelona, Spain
  • 10Department of Experimental and Clinical Medicine, University of Florence, Florence, Italy
  • 11First Medical Department, University Hospital Carl Gustav Carus, Dresden, Germany
  • 12Department of Gastroenterology, Erasme Hospital University, Brussels, Belgium
  • 13Department of Hepato-Gastroenterology, Dijon University Hospital, Dijon, France
  • 14Centre de Recherche UMR 866, Lipides, Nutrition, Cancer, Institut National de la Santé et de la Recherche Médicale, Dijon, France
JAMA Oncol. 2016;2(5):643-653. doi:10.1001/jamaoncol.2015.5225
Abstract

Importance  The prognostic value of BRAF and KRAS mutations in patients who have undergone resection for colon cancer and have been treated with combination leucovorin, fluorouracil, and oxaliplatin (FOLFOX)-based adjuvant chemotherapy is controversial, possibly owing to a lack of stratification on mismatch repair status.

Objective  To examine the prognostic effect of BRAF and KRAS mutations in patients with stage III colon cancer treated with adjuvant FOLFOX with or without cetuximab.

Design, Setting, and Participants  This study included patients with available tumor blocks of resected stage III colon adenocarcinoma who participated between December 2005 and November 2009 in the PETACC-8 phase III randomized trial. Mismatch repair, BRAF V600E, and KRAS exon 2 mutational status were determined on prospectively collected tumor blocks from 2559 patients enrolled in the PETACC-8 trial. The data were analyzed in April 2015.

Intervention  Patients were randomly assigned to receive 6 months of FOLFOX4 or FOLFOX4 plus cetuximab after surgical resection for stage III colon cancer.

Main Outcomes and Measures  Associations between these biomarkers and disease-free survival (DFS) and overall survival (OS) were analyzed with Cox proportional hazards models. Multivariate models were adjusted for covariates (age, sex, tumor grade, T/N stage, tumor location, Eastern Cooperative Oncology Group performance status).

Results  Among the 2559 patients enrolled in the PETACC-8 trial (42.9% female; median [range] age, 60.0 [19.0-75.0] years), microsatellite instability (MSI) phenotype, KRAS, and BRAF V600E mutations were detected in, respectively, 9.9% (177 of 1791), 33.1% (588 of 1776), and 9.0% (148 of 1643) of cases. In multivariate analysis, MSI (hazard ratio [HR] for DFS: 1.10 [95% CI, 0.73-1.64], P = .67; HR for OS: 1.02 [95% CI, 0.61-1.69], P = .94) and BRAF V600E mutation (HR for DFS: 1.22 [95% CI, 0.81-1.85], P = .34; HR for OS: 1.13 [95% CI, 0.64-2.00], P = .66) were not prognostic, whereas KRAS mutation was significantly associated with shorter DFS (HR, 1.55 [95% CI, 1.23-1.95]; P < .001) and OS (HR, 1.56 [95% CI, 1.12-2.15]; P = .008). The subgroup analysis showed in patients with microsatellite-stable tumors that both KRAS (HR for DFS: 1.64 [95% CI, 1.29-2.08], P < .001; HR for OS: 1.71 [95% CI, 1.21-2.41], P = .002) and BRAF V600E mutation (HR for DFS: 1.74 [95% CI, 1.14-2.69], P = .01; HR for OS: 1.84 [95% CI, 1.01-3.36], P = .046) were independently associated with worse clinical outcomes. In patients with MSI tumors, KRAS status was not prognostic, whereas BRAF V600E mutation was associated with significantly longer DFS (HR, 0.23 [95% CI, 0.06-0.92]; P = .04) but not OS (HR, 0.19 [95% CI, 0.03-1.24]; P = .08).

Conclusions and Relevance  BRAF V600E and KRAS mutations were significantly associated with shorter DFS and OS in patients with microsatellite-stable tumors but not in patients with MSI tumors. Future trials in the adjuvant setting will have to take into account mismatch repair, BRAF, and KRAS status for stratification.

Trial Registration  EudraCT 2005-003463-23

Introduction

Colorectal cancer (CRC) is the third most common cancer worldwide, causing more than 600 000 deaths each year.1 Disease stage remains the strongest prognostic variable and is the key determinant of treatment. The majority of newly diagnosed cases of CRC are in patients with nonmetastatic disease that can potentially be cured by surgery, either alone or combined with adjuvant chemotherapy. However, there is considerable stage-independent prognostic variability, likely due to tumor characteristics. Colorectal cancer is a biologically heterogeneous disease that develops via 2 well-described pathways of colorectal carcinogenesis, including chromosomal instability and, less commonly, microsatellite instability (MSI), which occurs in approximately 15% of cases. Microsatellite instability is a consequence of deficient DNA mismatch repair (MMR) that results in the accumulation of insertion and/or deletion mutations within microsatellite DNA regions.2 Deficient MMR can result from inheritance of a germline mutation in an MMR gene (MLH1, MSH2, MSH6, PMS2), causing Lynch syndrome,3 or more commonly, from epigenetic inactivation of MLH1,4 which is associated with hypermethylation of the promoter regions of cancer-specific genes, a situation known as the CpG island methylator phenotype (CIMP).5 Sporadic MSI CRC tumors are enriched with the BRAF-activating somatic V600E mutation (BRAF V600E), which is absent from MSI tumors associated with Lynch syndrome.6 The BRAF V600E mutation has an overall frequency of approximately 10% in all CRCs and is mutually exclusive of KRAS mutations, which are detected in 40% to 45% of cases.7,8

Although KRAS mutations are predictive of resistance to epidermal growth factor receptor inhibitors in metastatic CRC,9-11 and although BRAF V600E is now recognized as a marker of poor prognosis in this setting,8,12 the prognostic effect of these mutations in nonmetastatic CRC is controversial. KRAS mutations have been linked to disease recurrence and poorer overall survival in some studies but not in others, and there is some evidence that its role depends on the tumor site.7,13-16 Consistent data indicate that BRAF V600E mutation is associated with poor outcomes after relapse,14,17 but its direct effect on recurrence for patients in the adjuvant setting requires clarification.7,14,18,19 Most studies have shown an association of MSI phenotype with a better survival in earlier tumor stage, whereas the effect in stage III tumors is more controversial, and data in patients treated by the current combination leucovorin, fluorouracil, and oxaliplatin (FOLFOX) standard are scarce.19-21

Prognostic evaluation of these biomarkers is hampered by the fact that published data often mix prospective and retrospective studies, colon and rectal cancer, stage I to III tumors, and patients who did or did not receive a variety of adjuvant regimens; also, tumors are often not uniformly analyzed for all these biological molecular markers together (MSI, KRAS, BRAF). In fact, the frequency of KRAS and BRAF V600E mutations differs according to MMR status, and this may have impaired our interpretation of the effect of these mutations on clinical outcomes. We therefore examined disease-free survival (DFS) and overall survival (OS) according to MMR, KRAS, and BRAF status, determined on stage III colon cancer specimens collected prospectively from patients who received adjuvant FOLFOX alone or combined with cetuximab in the PETACC-8 randomized clinical trial. We also examined the prognostic value of KRAS and BRAF V600E mutations according to MMR status.

Box Section Ref ID

Key Points

  • Question What is the prognostic value of BRAF and KRAS mutations in patients who have undergone resection for colon cancer and have been treated with combination leucovorin, fluorouracil, and oxaliplatin (FOLFOX)-based adjuvant chemotherapy?

  • Findings This post hoc analysis of patients with stage III colon cancer who participated in the PETACC-8 phase III randomized clinical trial found that in patients with microsatellite-stable tumors, both KRAS and BRAF V600E mutations were independently linked to shorter disease-free and overall survival. In patients with microsatellite-unstable tumors, KRAS status was not prognostic, whereas BRAF V600E mutation was associated with significantly longer disease-free but not overall survival.

  • Meaning Microsatellite, KRAS, and BRAF V600E status assessment should be mandatory to stratify adequately in future adjuvant trials and must be discussed in our daily practice.

Methods
Study Population

This study included all patients with biological informed consent signed and available tumor blocks of resected stage III (any T, N1 or N2, M0) colon adenocarcinoma who have participated in the PETACC-8 phase III randomized trial.22 Patients in this trial were randomized to receive 6 months of FOLFOX4: 85 mg/m2 oxaliplatin (2-hour infusion) on day 1, and leucovorin 200 mg/m2 followed by fluorouracil (bolus) 400 mg/m2 and a 22-hour continuous infusion of fluorouracil 600 mg/m2 for 2 consecutive days, or FOLFOX4 plus cetuximab. The PETACC-8 study included 2559 patients between December 2005 and November 2009 and was amended in June 2008 to restrict random assignment to patients whose tumors expressed wild-type KRAS. Among the 2096 patients randomized before amendment, 1881 (90%) were retrospectively screened for KRAS mutations. The PETACC-8 trial, which received ethical approval for the study protocol and the translational data research integration, demonstrates that the addition of cetuximab to FOLFOX4 did not improve DFS compared with FOLFOX4 alone in patients with KRAS exon 2 wild-type resected stage III colon cancer.

MMR Status Determination

Mismatch repair tumor status was determined by immunohistochemical analysis (IHC), or by MSI testing when IHC was indeterminate. Microsatellite instability phenotype tumors were defined as exhibiting the loss of expression of 1 or more MMR proteins by IHC or exhibiting high-level tumor DNA MSI on MSI testing. Microsatellite-stable (MSS) phenotype tumors were defined by normal MMR protein expression in IHC, or MSS or low-level MSI status on MSI testing.

Immunohistochemical Analysis

Mismatch repair protein (MLH1, MSH2, MSH6, PMS2) expression was analyzed by IHC on tissue microarrays. Primary monoclonal antibodies against MLH1 (clone G168-728, diluted 1:100; BD Pharmingen), MSH2 (clone FE11, diluted 1:100; Oncogene Research Products), MSH6 (clone 44, diluted 1:100; BD Pharmingen), and PMS2 (clone A16-4, diluted 1:100; BD Pharmingen) were applied. Mismatch repair protein loss was defined as the absence of nuclear staining in neoplastic cells but positive nuclear staining in lymphocytes and normal adjacent colonic epithelium.23

MSI Testing

DNA was extracted from formalin-fixed paraffin-embedded (FFPE) tumor tissues for MSI analysis using 5 monomorphic mononucleotide markers (BAT-25, BAT-26, NR-21, NR-24, NR-27).24 Specimens with at least 3 unstable markers were scored as highly unstable, and specimens with fewer than 3 unstable markers were scored as stable.

BRAF and KRAS Gene Mutations

BRAF and KRAS mutation status was determined on genomic DNA extracted from FFPE tissues, using the QIAamp DNA Mini Kit (Qiagen). Molecular analysis was centralized and carried out retrospectively for patients included before trial amendment and prospectively for all other patients. Testing for 7 KRAS exon 2 mutations and the BRAF V600E hotspot exon 15 mutation (c.1799T>A/p.V600E) was based on real-time polymerase chain reaction using TaqMan probes (Applied Biosystems). Each assay was validated to detect 10% of mutated alleles.25

Statistical Analysis

Biomarker status was analyzed by investigators blinded to patient outcomes, and then transmitted for survival analyses to the data center. Disease-free survival was defined as the time between randomization and local or metastatic recurrence, diagnosis of a second colon or rectal cancer, or death, whichever occurred first. Overall survival was measured from randomization until death from any cause. For comparisons of baseline characteristics, categorical outcomes were analyzed with χ2 tests, and continuous outcomes, with standard parametric or nonparametric tests. Continuous variables are presented as the mean (SD) and median with interquartile range.

Disease-free and overall survival curves were estimated with the Kaplan-Meier method. Differences between groups of patients were analyzed with unstratified log-rank tests. Patients in the 2 treatment arms were combined because no difference was found between the 2 arms for efficacy analyses. An unstratified Cox regression model was used to estimate hazard ratios (HRs), 95% confidence intervals, and P values for candidate prognostic factors. Multivariate analyses were adjusted for stratification factors (nodal category, T stage, and obstruction or perforation status), sex, age, Eastern Cooperative Oncology Group performance status, tumor grade, primary tumor location, vascular invasion or lymphatic infiltration (VELI), and MMR, KRAS, and BRAF status.

Analyses were carried out with a 2-sided significance level of 5%. Results were unadjusted for multiple comparisons. All statistical analyses were performed with the SAS statistical software package, version 9.4.

Results
Patients

Among the 2559 patients included in the PETACC-8 phase III trial, 2043 signed the biological informed consent form, including 1791 patients with available FFPE specimen and no technical failure for the MMR status analysis. One hundred forty-eight (8.3%) and 15 (0.8%) of these patients were respectively excluded from BRAF V600E and KRAS exon 2 mutation analysis because of a technical issue (insufficient material or test failure) (Figure 1).

Characteristics of the study population are presented in Table 1. No noteworthy difference was observed between the present study population and the entire PETACC-8 trial population in terms of sex (male, 57.2% vs 57.1%), age (≤70 years, 89.8% vs 89.4%), and other clinical features, as well as pathological characteristics (eTable 1 in the Supplement). In the present study population, MSI phenotype and KRAS and BRAF V600E mutations were detected in 9.9% (177 of 1791), 33.1% (588 of 1776), and 9.0% (148 of 1643) of cases, respectively (Figure 1).

Among the 1791 patients tested for MMR status, all tumors were tested by IHC assay, except for 105 cases that were also tested by MSI assay because of indeterminate IHC results. Among the 177 tumors with MSI phenotype, 130 (73.4%) had lost MLH1 protein expression, 23 (13.0%) MSH2 expression, 6 (3.4%) MSH6 expression, and 12 (6.8%) PMS2 expression. The remaining 6 MSI tumors were considered indeterminate by means of IHC assay but positive by means of MSI testing. As expected, MSI compared with MSS phenotype was significantly associated with proximal tumors, high grade, and female sex (Table 1).19-21,26 Mutated vs nonmutated KRAS tumor was significantly associated with proximal site and VELI-positive status, whereas mutated vs nonmutated BRAF tumor was significantly associated with female sex, proximal site, higher N and T stage, higher grade, and VELI-positive status (Table 1). KRAS and BRAF V600E mutations were mutually exclusive except in 4 patients (Table 1). Even if the co-occurrence of KRAS and BRAF V600E mutations is rare in CRC, this event has already been described with a frequency of approximately 0.2%, which is in line with our results.27,28 The prevalence of KRAS mutations was higher in patients with MSS tumors (34.9%) than in patients with MSI tumors (17.0%) (P < .001).7,19 In contrast, BRAF V600E mutation was significantly more frequent in patients with MSI tumors (32.3%) than in those with MSS tumors (6.4%) (P < .001)7,19 (Table 1).

Prognostic Factors in the Overall Population

With an overall median follow-up of 3.52 years (95% CI, 3.46-3.64 years), higher T and N stage were independently associated with shorter DFS, whereas proximal site, higher N stage, and higher tumor grade were independently associated with shorter OS (Tables 2 and 3). In the biomarker analysis, no interaction was found between treatment (with or without cetuximab) and MMR status in terms of DFS or OS, but an interaction was found between both BRAF V600E and KRAS mutation and MMR status in terms of DFS and OS. Furthermore, no interaction was found between treatment (FOLFOX vs FOLFOX plus cetuximab) and KRAS status in terms of DFS (P = .82) and OS (P = .73); and the treatment administered significantly influenced neither DFS (HR, 0.88 [95% CI, 0.66-1.16]; P = .36) nor OS (HR, 0.98 [95% CI, 0.66-1.45]; P = .91) in patients with KRAS-mutated tumors.

The 3-year DFS rates were 77.9% and 73.9% among patients with MSI and MSS tumors, respectively (Figure 2A). In multivariate analysis, MSI phenotype was not significantly associated with DFS (HR, 1.10 [95% CI, 0.73-1.64]; P = .67) or OS (HR, 1.02 [95% CI, 0.61-1.69]; P = .94) (Table 3).

The 3-year DFS rates were 69.4% vs 77.1% among patients with mutated vs wild-type KRAS tumors (Figure 2B), and 73.8% vs 74.7% among patients with mutated vs wild-type BRAF tumors (Figure 2C). In multivariate analysis, KRAS mutation was significantly associated with shorter DFS (HR, 1.55 [95% CI, 1.23-1.95]; P < .001) and shorter OS (HR, 1.56 [95% CI, 1.12-2.15]; P = .008), whereas BRAF V600E mutation influenced neither outcome (Tables 2 and 3).

Prognostic Effect of KRAS and BRAF V600E Mutations in Patients With MSS Tumors

The 3-year DFS rates were 68.7% and 77.1%, respectively, among MSS patients with mutated and wild-type KRAS tumors (Figure 3A). In multivariate analysis, MSS patients with mutated vs wild-type KRAS tumors had significantly shorter DFS (HR, 1.64 [95% CI, 1.29-2.08]; P < .001) and shorter OS (HR, 1.71 [95% CI, 1.21-2.41]; P = .002) (Table 3). A similar negative effect was observed for BRAF V600E mutation. The 3-year DFS rates were 67.0% and 74.7%, respectively, among MSS patients with mutated and wild-type BRAF tumors (Figure 3B). In multivariate analysis, BRAF V600E mutation in patients with MSS tumors remained significantly associated with shorter DFS (HR, 1.74 [95% CI, 1.14-2.69]; P = .01) and shorter OS (HR, 1.84 [95% CI, 1.01-3.36]; P = .046) (Table 3).

Prognostic Effect of KRAS and BRAF V600E Mutations in Patients With MSI Tumors

The 3-year DFS rates were 82.8% and 77.5%, respectively, among MSI patients with mutated and wild-type KRAS tumors (Figure 3C). In multivariate analysis, KRAS status in patients with MSI tumors was not significantly associated with DFS (HR, 0.94 [95% CI, 0.32-2.74]; P = .91) or OS (HR, 0.90 [95% CI, 0.23-3.45]; P = .88) (Table 3). As observed in patients with MSS tumors, BRAF V600E mutation was again associated with clinical outcome in patients with MSI tumors, but the prognostic effect was in the opposite direction. Indeed, the 3-year DFS rates were 85.2% and 74.3%, respectively, among MSI patients with mutated and wild-type BRAF tumors (Figure 3D). In multivariate analysis, MSI tumors harboring BRAF V600E mutation were associated with longer DFS (HR, 0.23 [95% CI, 0.06-0.92]; P = .04) but not longer OS (HR, 0.19 [95% CI, 0.03-1.24]; P = .08) (Table 3).

Discussion

The aim of this study was to determine the prognostic value of MMR, KRAS, and BRAF status determined on prospectively collected stage III colon cancer specimens from patients receiving FOLFOX with or without cetuximab in a randomized trial of adjuvant therapy. We found that MMR status was not predictive of either DFS or OS. Most previous studies have shown a favorable prognostic effect of the MSI phenotype,20,21,26,29-32 but others showed no significant difference in clinical outcome between patients with MSI and MSS tumors.33-35 This discrepancy might be explained by a lack of adjustment for BRAF and KRAS status, tumor stage, or the adjuvant treatment regimen. Indeed, although our cohort was composed only of patients with stage III colon cancer, studies showing longer survival among patients with MSI vs MSS tumors generally combined stage II and III tumors, and the favorable prognostic effect seemed to be stronger in stage II disease.21,36 In the NCCTG N0147 study, which had a design similar to that of the PETACC-08 trial, analysis of the 2580 patients with stage III colon cancer participating in this trial showed that MMR status had no prognostic value.19 Furthermore, in vitro studies have shown that oxaliplatin,37 contrary to fluorouracil,38 is active on CRC cell lines independently of MMR status, suggesting that the clinical effect of MMR status might be attenuated in patients receiving FOLFOX-based adjuvant chemotherapy.

We found that BRAF V600E mutation influenced neither DFS nor OS in the overall study population, whereas a negative effect on DFS and OS was observed in the MSS subgroup. Recently, data from the NCCTG N0147 trial showed that BRAF V600E mutation was significantly associated with shorter DFS in multivariate analysis (HR, 1.37 [95% CI, 1.08-1.70]; P = .009).19 However, the adverse effect of BRAF V600E mutation was limited to patients with MSS tumors after stratification on MMR status.19 It seems important to adjust the BRAF prognostic value to the MMR status to identify more precisely the patients with poor clinical outcomes in stage III colon cancer. Three retrospective analyses of randomized adjuvant trials suggested that BRAF V600E mutation was independently associated with shorter OS but not with disease-free or recurrence-free survival7,14,18 (eTable 2 in the Supplement). However, when MMR status was taken into account in these studies, the worse prognostic value of BRAF V600E mutation was attenuated in patients with MSI tumors. Indeed, MSI/BRAF wild-type patients had the best prognosis, whereas the MSS/BRAF V600E mutation subgroup had the worst prognosis. Patients with MSS/BRAF wild-type or MSI/BRAF V600E–mutated tumors had intermediate survival.14,18

Here we found that BRAF V600E mutation was significantly associated with longer DFS but not OS in patients with MSI tumors. This suggests that the prognostic effect of BRAF V600E mutation in MSI patients treated with FOLFOX with or without cetuximab adjuvant chemotherapy may be indirectly related to the CIMP phenotype. Indeed, there is considerable overlap among tumors characterized as MSI, mutated BRAF V600E, and CIMP.5 Tumors with BRAF V600E mutation and MSI phenotype occur almost exclusively as a consequence of CIMP, associated with methylation of the MLH1 promoter. The prognostic value of the CIMP phenotype in patients receiving fluorouracil-based adjuvant chemotherapy is controversial.39-41 To our knowledge, only 1 retrospective study has evaluated the prognostic effect of both MMR and CIMP status in patients with stage III colon cancer receiving FOLFOX-based adjuvant chemotherapy, showing that MSI/CIMP-positive tumor status was associated with poorer DFS than MSI/CIMP-negative tumor status.42 However, in the MSI/CIMP-positive subgroup analysis, patients with MLH1 methylation had a longer DFS than those with methylation at other loci.42

We found that KRAS mutations were linked to survival defined by a shorter DFS and OS in the overall study population. Stratification on MMR status showed that this effect was restricted to the MSS subgroup whereas no effect of KRAS status was seen in the MSI subgroup. Large population-based cohorts and retrospective analyses of randomized adjuvant trials have reported conflicting results concerning the prognostic value of KRAS exon 2 mutations7,13,43-45 (eTable 2 in the Supplement). Retrospective analyses of 3 randomized adjuvant trials (CKVO 90-11, CALGB 89803, and PETACC-3) failed to demonstrate any association between KRAS codon 12 and 13 mutations and recurrence-free survival or OS in patients with stage II and III colon cancer.7,44,45 In contrast, multivariate analysis of data from patients with stage III colon cancer included in the NCCTG N0147 study showed that KRAS mutations in either codon 12 or 13 were independently associated with poorer DFS.15 More recently, data on patients enrolled in the PETACC-8 trial showed that both KRAS codon 12 and 13 mutations were related to prognosis in patients with distal tumors only.16 We report here that KRAS mutations were associated with a poor outcome in patients with MSS tumors, which represent approximately 90% of all stage III colon cancer. This is in accordance with our previous report16 because MSS tumors are more frequently distal, whereas MSI tumors are mainly located in the proximal site. A lack of stratification on MMR tumor status may, at least in part, explain discrepancies concerning the prognostic value of KRAS mutation among previously published studies.

In randomized studies including patients with KRAS-mutated metastatic CRC, oxaliplatin-based chemotherapy in combination with epidermal growth factor receptor inhibitors (cetuximab and panitumumab) is associated with worse survival than oxaliplatin-based chemotherapy alone,46,47 which is not the case for irinotecan-based chemotherapy.48 In our study, cetuximab in combination with oxaliplatin-based chemotherapy did not worsen the clinical outcomes of patients with stage III colon cancer with KRAS-mutated tumors. Further analysis based on the complete assessment of KRAS/NRAS exons 2, 3, and 4, as well as survival after recurrence and treatments received after recurrence, is needed to better elucidate the real predictive effect of KRAS mutations in the adjuvant setting.

Strengths of our study include biomarker analysis in a prospective collection of tumor blocks from patients with stage III colon cancer treated in a randomized trial with the current standard FOLFOX-based adjuvant chemotherapy. The present study population was representative of the entire PETACC-8 population because no statistically significant difference was observed in terms of clinical and pathological characteristics (see eTable 1 in the Supplement). This large study allowed us to analyze the prognostic effect of BRAF V600E and KRAS mutations according to MMR status. Study limitations include the lack of patients treated with fluorouracil alone, making it difficult to analyze the predictive value of MMR status for the response to oxaliplatin-based adjuvant chemotherapy. The percentages of patients with mutated KRAS tumors were lower than expected in the MSI and MSS groups,7 following amendment of the PETACC-8 trial eligibility criteria to restrict random assignment to patients with KRAS wild-type tumors. We also recognize the inherent limitations related to the lack of assessment of activating hotspot mutations in KRAS/NRAS exons 2, 3, and 4 on clinical outcomes of patients with colon cancer treated with FOLFOX with or without cetuximab in the adjuvant setting. Indeed, recent data from randomized studies demonstrated that patients with metastatic CRC who have RAS wild-type tumors derived a significant survival benefit from the addition of epidermal growth factor receptor inhibitors (panitumumab or cetuximab) to chemotherapy, whereas patients with RAS tumor mutations did not.47,49 Finally, our hypothesis suggesting that CIMP status may play a role in the good prognostic effect of BRAF V600E mutation in patients with MSI tumors should be interpreted with caution because of the small number of patients and the lack of CIMP profiling of the tumors. These results need to be confirmed on pooled data from large phase III adjuvant trials using FOLFOX.

Conclusions

This large analysis of patients with stage III colon cancer receiving FOLFOX-based adjuvant chemotherapy shows that MMR status should be taken into account in future prognostic studies involving KRAS and BRAF V600E mutations. BRAF V600E and KRAS exon 2 mutations are independently linked to shorter DFS and OS in patients with MSS tumors. In contrast, KRAS mutations have no prognostic value in patients with MSI tumors, whereas the BRAF V600E mutation could be associated with longer survival.

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Article Information

Accepted for Publication: October 20, 2015.

Corresponding Author: Julien Taieb, MD, PhD, Department of Gastroenterology and Digestive Oncology, European Georges Pompidou Hospital, and Paris Descartes University, 20 rue Leblanc, 75015 Paris, France (julien.taieb@egp.aphp.fr).

Published Online: January 14, 2016. doi:10.1001/jamaoncol.2015.5225.

Author Contributions: Dr Taieb and Ms Le Malicot had full access to all of the data in the study and take responsibility for the integrity of the data and the accuracy of the data analysis. Drs Taieb and Zaanan and Ms Le Malicot contributed equally to this work as first authors. Drs Emile and Laurent-Puig contributed equally to this work as senior authors.

Study concept and design: Taieb, Zaanan, Tabernero, Mini, Folprecht, Laurent-Puig.

Acquisition, analysis, or interpretation of data: All authors.

Drafting of the manuscript: Taieb, Zaanan, Le Malicot, Julié, Blons, Tabernero, Lepage, Laurent-Puig.

Critical revision of the manuscript for important intellectual content: Taieb, Zaanan, Le Malicot, Julié, Mineur, Bennouna, Tabernero, Mini, Folprecht, Van Laethem, Emile, Laurent-Puig.

Statistical analysis: Le Malicot, Tabernero, Laurent-Puig.

Obtained funding: Taieb, Laurent-Puig.

Administrative, technical, or material support: Taieb, Julié, Blons, Mineur, Bennouna, Tabernero, Folprecht, Van Laethem, Lepage, Emile, Laurent-Puig.

Study supervision: Taieb, Zaanan, Laurent-Puig.

Conflict of Interest Disclosures: Dr Taieb has participated in consulting and/or advisory boards for Merck, Sanofi, Roche Genentech, Pfizer, and Amgen. Dr Zaanan has participated in consulting and/or advisory boards for Roche, Merck Serono, Amgen, Sanofi, and Lilly. Dr Julié has received honoraria from Roche and Merck Sereno. Dr Bennouna has participated in consulting and/or advisory boards for Roche, Boehringer Ingelheim, Novartis, and Pierre Fabre and has received honoraria from Roche, Boehringer Ingelheim, Novartis, and Pierre Fabre. Dr Tabernero has participated in consulting and/or advisory boards for Amgen, ImClone Systems, Lilly, Millennium, Novartis, Roche/Genentech, Sanofi, Celgene, Chugai Pharma, Taiho Pharmaceutical, Boehringer Ingelheim and Merck Serono. Dr Folprecht has participated in consulting and/or advisory boards for Roche, Merck KGaA, Lilly, and Bristol and has received honoraria from Merck KGaA, Lilly, and Bayer and research funding from Merck KGaA. Dr Laurent-Puig has participated in consulting and/or advisory boards for and received honoraria from Sanofi, Merck Serono, Amgen, Roche, Genomic Health, Myriad Genetics, and Pfizer. No other disclosures are reported.

Funding/Support: The study was sponsored by the Fédération Francophone de Cancérologie Digestive (FFCD) with funds from a grant from the Ligue Nationale Contre le Cancer. Merck KGaA and Sanofi supported the study, Merck provided the study cetuximab and financial support for study management, and Sanofi provided financial support for the provision of oxaliplatin to Belgian sites when necessary.

Role of the Funder/Sponsor: The FFCD was responsible for the design and conduct of the study; collection, management, analysis, and interpretation of the data; preparation, review, and approval of the manuscript; and decision to submit the manuscript for publication. Merck KGaA and Sanofi-Aventis reviewed the manuscript but had no role in the design and conduct of the study; collection, management, analysis, and interpretation of the data; preparation of the manuscript; and decision to submit the manuscript for publication.

Additional Information: The PETACC-8 study investigators are listed in the eAppendix in the Supplement.

Additional Contributions: We thank all participating patients and their families, and the study groups and investigators from the participating countries; we also thank the team from the FFCD data center.

References
1.
Jemal  A, Bray  F, Center  MM, Ferlay  J, Ward  E, Forman  D.  Global cancer statistics.  CA Cancer J Clin. 2011;61(2):69-90.PubMedGoogle ScholarCrossref
2.
Ionov  Y, Peinado  MA, Malkhosyan  S, Shibata  D, Perucho  M.  Ubiquitous somatic mutations in simple repeated sequences reveal a new mechanism for colonic carcinogenesis.  Nature. 1993;363(6429):558-561.PubMedGoogle ScholarCrossref
3.
Peltomäki  P, Lothe  RA, Aaltonen  LA,  et al.  Microsatellite instability is associated with tumors that characterize the hereditary non-polyposis colorectal carcinoma syndrome.  Cancer Res. 1993;53(24):5853-5855.PubMedGoogle Scholar
4.
Herman  JG, Umar  A, Polyak  K,  et al.  Incidence and functional consequences of hMLH1 promoter hypermethylation in colorectal carcinoma.  Proc Natl Acad Sci U S A. 1998;95(12):6870-6875.PubMedGoogle ScholarCrossref
5.
Weisenberger  DJ, Siegmund  KD, Campan  M,  et al.  CpG island methylator phenotype underlies sporadic microsatellite instability and is tightly associated with BRAF mutation in colorectal cancer.  Nat Genet. 2006;38(7):787-793.PubMedGoogle ScholarCrossref
6.
Domingo  E, Niessen  RC, Oliveira  C,  et al.  BRAF-V600E is not involved in the colorectal tumorigenesis of HNPCC in patients with functional MLH1 and MSH2 genes.  Oncogene. 2005;24(24):3995-3998.PubMedGoogle ScholarCrossref
7.
Roth  AD, Tejpar  S, Delorenzi  M,  et al.  Prognostic role of KRAS and BRAF in stage II and III resected colon cancer: results of the translational study on the PETACC-3, EORTC 40993, SAKK 60-00 trial.  J Clin Oncol. 2010;28(3):466-474.PubMedGoogle ScholarCrossref
8.
Van Cutsem  E, Köhne  CH, Láng  I,  et al.  Cetuximab plus irinotecan, fluorouracil, and leucovorin as first-line treatment for metastatic colorectal cancer: updated analysis of overall survival according to tumor KRAS and BRAF mutation status.  J Clin Oncol. 2011;29(15):2011-2019.PubMedGoogle ScholarCrossref
9.
Karapetis  CS, Khambata-Ford  S, Jonker  DJ,  et al.  K-ras mutations and benefit from cetuximab in advanced colorectal cancer.  N Engl J Med. 2008;359(17):1757-1765.PubMedGoogle ScholarCrossref
10.
Van Cutsem  E, Labianca  R, Bodoky  G,  et al.  Randomized phase III trial comparing biweekly infusional fluorouracil/leucovorin alone or with irinotecan in the adjuvant treatment of stage III colon cancer: PETACC-3.  J Clin Oncol. 2009;27(19):3117-3125.PubMedGoogle ScholarCrossref
11.
De Roock  W, Claes  B, Bernasconi  D,  et al.  Effects of KRAS, BRAF, NRAS, and PIK3CA mutations on the efficacy of cetuximab plus chemotherapy in chemotherapy-refractory metastatic colorectal cancer: a retrospective consortium analysis.  Lancet Oncol. 2010;11(8):753-762.PubMedGoogle ScholarCrossref
12.
Yokota  T, Ura  T, Shibata  N,  et al.  BRAF mutation is a powerful prognostic factor in advanced and recurrent colorectal cancer.  Br J Cancer. 2011;104(5):856-862.PubMedGoogle ScholarCrossref
13.
Andreyev  HJ, Norman  AR, Cunningham  D, Oates  JR, Clarke  PA.  Kirsten ras mutations in patients with colorectal cancer: the multicenter “RASCAL” study.  J Natl Cancer Inst. 1998;90(9):675-684.PubMedGoogle ScholarCrossref
14.
Gavin  PG, Colangelo  LH, Fumagalli  D,  et al.  Mutation profiling and microsatellite instability in stage II and III colon cancer: an assessment of their prognostic and oxaliplatin predictive value.  Clin Cancer Res. 2012;18(23):6531-6541.PubMedGoogle ScholarCrossref
15.
Yoon  HH, Tougeron  D, Shi  Q,  et al; Alliance for Clinical Trials in Oncology.  KRAS codon 12 and 13 mutations in relation to disease-free survival in BRAF-wild-type stage III colon cancers from an adjuvant chemotherapy trial (N0147 Alliance).  Clin Cancer Res. 2014;20(11):3033-3043.PubMedGoogle ScholarCrossref
16.
Blons  H, Emile  JF, Le Malicot  K,  et al.  Prognostic value of KRAS mutations in stage III colon cancer: post hoc analysis of the PETACC-8 phase III trial dataset.  Ann Oncol. 2014;25(12):2378-2385.PubMedGoogle ScholarCrossref
17.
Popovici  V, Budinska  E, Tejpar  S,  et al.  Identification of a poor-prognosis BRAF-mutant-like population of patients with colon cancer.  J Clin Oncol. 2012;30(12):1288-1295.PubMedGoogle ScholarCrossref
18.
Ogino  S, Shima  K, Meyerhardt  JA,  et al.  Predictive and prognostic roles of BRAF mutation in stage III colon cancer: results from intergroup trial CALGB 89803.  Clin Cancer Res. 2012;18(3):890-900.PubMedGoogle ScholarCrossref
19.
Sinicrope  FA, Mahoney  MR, Smyrk  TC,  et al.  Prognostic impact of deficient DNA mismatch repair in patients with stage III colon cancer from a randomized trial of FOLFOX-based adjuvant chemotherapy.  J Clin Oncol. 2013;31(29):3664-3672.PubMedGoogle ScholarCrossref
20.
Ribic  CM, Sargent  DJ, Moore  MJ,  et al.  Tumor microsatellite-instability status as a predictor of benefit from fluorouracil-based adjuvant chemotherapy for colon cancer.  N Engl J Med. 2003;349(3):247-257.PubMedGoogle ScholarCrossref
21.
Sargent  DJ, Marsoni  S, Monges  G,  et al.  Defective mismatch repair as a predictive marker for lack of efficacy of fluorouracil-based adjuvant therapy in colon cancer.  J Clin Oncol. 2010;28(20):3219-3226.PubMedGoogle ScholarCrossref
22.
Taieb  J, Tabernero  J, Mini  E,  et al; PETACC-8 Study Investigators.  Oxaliplatin, fluorouracil, and leucovorin with or without cetuximab in patients with resected stage III colon cancer (PETACC-8): an open-label, randomised phase 3 trial.  Lancet Oncol. 2014;15(8):862-873.PubMedGoogle ScholarCrossref
23.
Zaanan  A, Cuilliere-Dartigues  P, Guilloux  A,  et al.  Impact of p53 expression and microsatellite instability on stage III colon cancer disease-free survival in patients treated by 5-fluorouracil and leucovorin with or without oxaliplatin.  Ann Oncol. 2010;21(4):772-780.PubMedGoogle ScholarCrossref
24.
Umar  A, Boland  CR, Terdiman  JP,  et al.  Revised Bethesda Guidelines for hereditary nonpolyposis colorectal cancer (Lynch syndrome) and microsatellite instability.  J Natl Cancer Inst. 2004;96(4):261-268.PubMedGoogle ScholarCrossref
25.
Blons  H, Rouleau  E, Charrier  N,  et al; MOKAECM Collaborative Group.  Performance and cost efficiency of KRAS mutation testing for metastatic colorectal cancer in routine diagnosis: the MOKAECM study, a nationwide experience.  PLoS One. 2013;8(7):e68945.PubMedGoogle ScholarCrossref
26.
Samowitz  WS, Curtin  K, Ma  KN,  et al.  Microsatellite instability in sporadic colon cancer is associated with an improved prognosis at the population level.  Cancer Epidemiol Biomarkers Prev. 2001;10(9):917-923.PubMedGoogle Scholar
27.
Sorbye  H, Dragomir  A, Sundström  M,  et al.  High BRAF mutation frequency and marked survival differences in subgroups according to KRAS/BRAF mutation status and tumor tissue availability in a prospective population-based metastatic colorectal cancer cohort.  PLoS One. 2015;10(6):e0131046.PubMedGoogle ScholarCrossref
28.
Janakiraman  M, Vakiani  E, Zeng  Z,  et al.  Genomic and biological characterization of exon 4 KRAS mutations in human cancer.  Cancer Res. 2010;70(14):5901-5911.PubMedGoogle ScholarCrossref
29.
Popat  S, Hubner  R, Houlston  RS.  Systematic review of microsatellite instability and colorectal cancer prognosis.  J Clin Oncol. 2005;23(3):609-618.PubMedGoogle ScholarCrossref
30.
Lanza  G, Gafà  R, Santini  A, Maestri  I, Guerzoni  L, Cavazzini  L.  Immunohistochemical test for MLH1 and MSH2 expression predicts clinical outcome in stage II and III colorectal cancer patients.  J Clin Oncol. 2006;24(15):2359-2367.PubMedGoogle ScholarCrossref
31.
Halling  KC, French  AJ, McDonnell  SK,  et al.  Microsatellite instability and 8p allelic imbalance in stage B2 and C colorectal cancers.  J Natl Cancer Inst. 1999;91(15):1295-1303.PubMedGoogle ScholarCrossref
32.
Zaanan  A, Fléjou  JF, Emile  JF,  et al.  Defective mismatch repair status as a prognostic biomarker of disease-free survival in stage III colon cancer patients treated with adjuvant FOLFOX chemotherapy.  Clin Cancer Res. 2011;17(23):7470-7478.PubMedGoogle ScholarCrossref
33.
Barnetson  RA, Tenesa  A, Farrington  SM,  et al.  Identification and survival of carriers of mutations in DNA mismatch-repair genes in colon cancer.  N Engl J Med. 2006;354(26):2751-2763.PubMedGoogle ScholarCrossref
34.
Kim  GP, Colangelo  LH, Wieand  HS,  et al; National Cancer Institute.  Prognostic and predictive roles of high-degree microsatellite instability in colon cancer: a National Cancer Institute-National Surgical Adjuvant Breast and Bowel Project collaborative study.  J Clin Oncol. 2007;25(7):767-772.PubMedGoogle ScholarCrossref
35.
Salahshor  S, Kressner  U, Fischer  H,  et al.  Microsatellite instability in sporadic colorectal cancer is not an independent prognostic factor.  Br J Cancer. 1999;81(2):190-193.PubMedGoogle ScholarCrossref
36.
Roth  AD, Delorenzi  M, Tejpar  S,  et al.  Integrated analysis of molecular and clinical prognostic factors in stage II/III colon cancer.  J Natl Cancer Inst. 2012;104(21):1635-1646.PubMedGoogle ScholarCrossref
37.
Fink  D, Zheng  H, Nebel  S,  et al.  In vitro and in vivo resistance to cisplatin in cells that have lost DNA mismatch repair.  Cancer Res. 1997;57(10):1841-1845.PubMedGoogle Scholar
38.
Carethers  JM, Chauhan  DP, Fink  D,  et al.  Mismatch repair proficiency and in vitro response to 5-fluorouracil.  Gastroenterology. 1999;117(1):123-131.PubMedGoogle ScholarCrossref
39.
Shen  L, Catalano  PJ, Benson  AB  III, O’Dwyer  P, Hamilton  SR, Issa  JP.  Association between DNA methylation and shortened survival in patients with advanced colorectal cancer treated with 5-fluorouracil based chemotherapy.  Clin Cancer Res. 2007;13(20):6093-6098.PubMedGoogle ScholarCrossref
40.
Van Rijnsoever  M, Elsaleh  H, Joseph  D, McCaul  K, Iacopetta  B.  CpG island methylator phenotype is an independent predictor of survival benefit from 5-fluorouracil in stage III colorectal cancer.  Clin Cancer Res. 2003;9(8):2898-2903.PubMedGoogle Scholar
41.
Jover  R, Nguyen  TP, Pérez-Carbonell  L,  et al.  5-Fluorouracil adjuvant chemotherapy does not increase survival in patients with CpG island methylator phenotype colorectal cancer.  Gastroenterology. 2011;140(4):1174-1181.PubMedGoogle ScholarCrossref
42.
Han  SW, Lee  HJ, Bae  JM,  et al.  Methylation and microsatellite status and recurrence following adjuvant FOLFOX in colorectal cancer.  Int J Cancer. 2013;132(9):2209-2216.PubMedGoogle ScholarCrossref
43.
Andreyev  HJ, Norman  AR, Cunningham  D,  et al.  Kirsten ras mutations in patients with colorectal cancer: the “RASCAL II” study.  Br J Cancer. 2001;85(5):692-696.PubMedGoogle ScholarCrossref
44.
Westra  JL, Schaapveld  M, Hollema  H,  et al.  Determination of TP53 mutation is more relevant than microsatellite instability status for the prediction of disease-free survival in adjuvant-treated stage III colon cancer patients.  J Clin Oncol. 2005;23(24):5635-5643.PubMedGoogle ScholarCrossref
45.
Ogino  S, Meyerhardt  JA, Irahara  N,  et al; Cancer and Leukemia Group B; North Central Cancer Treatment Group; Canadian Cancer Society Research Institute; Southwest Oncology Group.  KRAS mutation in stage III colon cancer and clinical outcome following intergroup trial CALGB 89803.  Clin Cancer Res. 2009;15(23):7322-7329.PubMedGoogle ScholarCrossref
46.
Bokemeyer  C, Bondarenko  I, Makhson  A,  et al.  Fluorouracil, leucovorin, and oxaliplatin with and without cetuximab in the first-line treatment of metastatic colorectal cancer.  J Clin Oncol. 2009;27(5):663-671.PubMedGoogle ScholarCrossref
47.
Douillard  JY, Oliner  KS, Siena  S,  et al.  Panitumumab-FOLFOX4 treatment and RAS mutations in colorectal cancer.  N Engl J Med. 2013;369(11):1023-1034.PubMedGoogle ScholarCrossref
48.
Van Cutsem  E, Köhne  CH, Hitre  E,  et al.  Cetuximab and chemotherapy as initial treatment for metastatic colorectal cancer.  N Engl J Med. 2009;360(14):1408-1417.PubMedGoogle ScholarCrossref
49.
Van Cutsem  E, Lenz  HJ, Köhne  CH,  et al.  Fluorouracil, leucovorin, and irinotecan plus cetuximab treatment and RAS mutations in colorectal cancer.  J Clin Oncol. 2015;33(7):692-700.PubMedGoogle ScholarCrossref
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