Shi, Qian

Abstract Introduction: Peripheral blood leukocyte (PBL) telomere length is associated with colon cancer (CC) risk, but the relationship of PBL telomere length to survival in CC patients receiving chemotherapy is unknown. Methods: We measured relative telomere length (RTL) by qPCR of PBL DNA from 2,748 cases (53% male, 85% white, median age 58 yrs) in the North American Phase III Intergroup Trial N0147 that compared disease-free survival (DFS) among completely resected Stage III CC treated with mFOLFOX6 +/- cetuximab. Tumor markers were assessed, and 12% were BRAF mutant; 35% Kras mutant, and 12% deficient mismatch repair (dMMR). Linear and non-linear prognostic associations of PBL RTL to the primary endpoint of DFS were assessed by Cox models. Optimal categories of RTL were developed using cumulative martingale residual plots, designated from shortest to longest RTL (Table 1). Results: Never smokers had longer PBL RTL (p = 0.0006) than past or current smokers. Those with diabetes mellitus (DM) had longer PBL RTL than those without DM (p = 0.0165). PBL RTL was not associated with any tumor markers (MMR status; p = 0.7174; BRAF mutant vs wild-type (wt); p =0.4039; Kras mutant vs wt; p= 0.1149). PBL RTL was significantly associated with DFS in a non-linear form (p = 0.0389). Cases in the longer RTL category had more favorable DFS than those with shorter or longest RTL, (Table), which remained after adjusting for age, sex , T/N stage, grade, tumor markers, and treatment (p = 0.0230). We observed no significant interaction (p = 0.1490) between RTL categories and treatment. Conclusion: We observed a non-linear association between RTL and DFS in stage III CC, suggesting that an optimal range telomere length can influence tumor behavior in CC patients. Future studies of telomere maintenance mechanisms may help to explain the association of RTL with improved DFS. RTL is associated with smoking and DM in CC cases but further studies are needed to determine if either tobacco use or DM affect survival from CC by modulating PBL telomere length. Citation Format: Nivedita Basu, Ruth A. Johnson, Qian Shi, Garth D. Nelson, Julie M. Cunningham, Richard M. Goldberg, Frank A. Sinicrope, Daniel J. Sargent, Steven R. Alberts, Gloria M. Petersen, Stephen N. Thibodeau, Halcyon G. Skinner, Kristin Litzelman, Songwon Seo, Russell J. Vanderboom, Ronald E. Gangnon, Corinne D. Engelman, David N. Rider, Lisa A. Boardman. Telomeres and survival in stage III colon cancers: Findings from NCCTG N0147 (Alliance) study. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5102. doi:10.1158/1538-7445.AM2014-5102

PurposeProgression-free survival (PFS) has previously been established as a surrogate for overall survival (OS) for first-line metastatic colorectal cancer (mCRC). Because mCRC treatment has advanced in the last decade with extended OS, this surrogacy requires re-examination.MethodsIndividual patient data from 16,762 patients were available from 22 first-line mCRC studies conducted from 1997 to 2006; 12 of those studies tested antiangiogenic and/or anti–epidermal growth factor receptor agents. The relationship between PFS (first event of progression or death) and OS was evaluated by using R2statistics (the closer the value is to 1, the stronger the correlation) from weighted least squares regression of trial-specific hazard ratios estimated by using Cox and Copula models.ResultsForty-four percent of patients received a regimen that included biologic agents. Median first-line PFS was 8.3 months, and median OS was 18.2 months. The correlation between PFS and OS was modest (R2, 0.45 to 0.69). Analyses limited to trials that tested treatments with biologic agents, nonstrategy trials, or superiority trials did not improve surrogacy.ConclusionIn modern mCRC trials, in which survival after the first progression exceeds time to first progression, a positive but modest correlation was observed between OS and PFS at both the patient and trial levels. This finding demonstrates the substantial variability in OS introduced by the number of lines of therapy and types of effective subsequent treatments and the associated challenge to the use of OS as an end point to assess the benefit attributable to a single line of therapy. PFS remains an appropriate primary end point for first-line mCRC trials to detect the direct treatment effect of new agents.

Abstract Purpose: Recurrence risk assessment to make treatment decisions for early-stage colon cancer patients is a major unmet medical need. The aim of this retrospective multicenter study was to evaluate the clinical utility of guanylyl cyclase C (GCC) mRNA levels in lymph nodes on colon cancer recurrence. Methods: The proportion of lymph nodes with GCC-positive mRNA (LNR) was evaluated in 463 untreated T3N0 patients, blinded to clinical outcomes. One site's (n = 97) tissue grossing method precluded appropriate lymph node assessment resulting in post hoc exclusion. Cox regression models tested the relationship between GCC and the primary endpoint of time to recurrence. Assay methods, primary analyses, and cut points were all prespecified. Results: Final dataset contained 366 patients, 38 (10%) of whom had recurrence. Presence of four or more GCC-positive lymph nodes was significantly associated with risk of recurrence [hazard ratio (HR) = 2.46, 95% confidence interval (CI), 1.07–5.69, P = 0.035], whereas binary GCC LNR risk class (HR = 1.87, 95% CI, 0.99–3.54, P = 0.054) and mismatch repair (MMR) status (HR = 0.77, 95% CI, 0.36–1.62, P = 0.49) were not. In a secondary analysis using a 3-level GCC LNR risk group classification of high (LNR > 0.20), intermediate (0.10 < LNR ≤ 0.20), and low (LNR ≤ 0.10), high-risk patients had a 2.5 times higher recurrence risk compared with low-risk patients (HR = 2.53, 95% CI, 1.24–5.17, P = 0.011). Conclusions: GCC status is a promising prognostic factor independent of traditional histopathology risk factors in a contemporary population of patients with stage IIa colon cancer not treated with adjuvant therapy, but GCC determination must be performed with methodology adapted to the tissue procurement and fixation technique. Clin Cancer Res; 20(16); 4361–9. ©2014 AACR.

AbstractPurpose: A let-7 microRNA-complementary site (LCS6) polymorphism in the 3′ untranslated region of the KRAS gene has been shown to disrupt let-7 binding and upregulate KRAS expression. We evaluated the LCS6 genotype and its association with KRAS mutation status, clinicopathologic features, and disease-free survival (DFS) in patients with stage III colon cancer who enrolled in a phase III clinical trial (NCCTG N0147).Experimental Design: The LCS6 genotype was assayed by real-time PCR in DNA extracted from whole blood (n = 2,834) and compared with paired tumor tissue (n = 977). χ2 and two-sample t tests were used to compare baseline factors and KRAS mutation status between patients defined by LCS6 variant status. Log-rank tests and multivariate Cox models assessed associations between LCS6 status and DFS, respectively.Results: We identified 432 (15.2%) blood samples and 143 (14.6%) tumor samples heterozygous or homozygous for the LCS6 G-allele, and 2,402 of 2,834 (84.8%) blood samples and 834 of 977 (85.4%) tumor samples homozygous for the LCS6 T-allele. Genotype results were highly concordant (99.8%) in cases with paired blood and tumor tissue (n = 977). G-allele carriers were significantly more frequent in Caucasians versus other races (χ2 test, P < 0.0001). The LCS6 genotype was not associated with KRAS mutation status, clinicopathologic features (all P > 0.2), or DFS (log-rank P = 0.49; HR, 0.929; 95% confidence interval, 0.76–1.14), even after combining LCS6 genotype with KRAS mutation status.Conclusions: In the largest association study investigating the LCS6 polymorphism in colon cancers, the germline LCS6 genotype was not associated with KRAS mutation status or with clinical outcome in patients with stage III tumors. Clin Cancer Res; 20(12); 3319–27. ©2014 AACR.

Abstract Purpose: We examined the prognostic impact of specific KRAS mutations in patients with stage III colon adenocarcinoma receiving adjuvant FOLFOX alone or combined with cetuximab in a phase III trial (N0147). Analysis was restricted to BRAF–wild-type tumors, because BRAF mutation was associated with poor prognosis, and BRAF and KRAS mutations are mutually exclusive. Experimental Design: The seven most common KRAS mutations in codon 12 and codon 13 were examined in 2,478 BRAF–wild-type tumors. Because KRAS mutations in codon 12 (n = 779) or 13 (n = 220) were not predictive of adjuvant cetuximab benefit, study arms were pooled for analysis. Disease-free survival (DFS) was evaluated by HRs using Cox models. Results: KRAS mutations in codon 12 (multivariate HR, 1.52; 95% confidence interval, CI, 1.28–1.80; P < 0.0001) or codon 13 (multivariate HR, 1.36; 95% CI, 1.04–1.77; P = 0.0248) were significantly associated with shorter DFS compared with patients with wild-type KRAS/BRAF tumors, independent of covariates. KRAS codon 12 mutations were independently associated with proficient mismatch repair (P < 0.0001), proximal tumor site (P < 0.0001), low grade, age, and sex, whereas codon 13 mutations were associated with proximal site (P < 0.0001). Conclusion: KRAS mutations in either codon 12 or 13 are associated with inferior survival in patients with resected stage III colon cancer. These data highlight the importance of accurate molecular characterization and the significant role of KRAS mutations in both codons in the progression of this malignancy in the adjuvant setting. Clin Cancer Res; 20(11); 3033–43. ©2014 AACR.

BACKGROUNDHuman epidermal growth factor receptor 2 (HER2) is a therapeutic target in patients with esophageal adenocarcinoma (EAC), with gene amplification used as a selection criterion for treatment, although to the authors' knowledge the concordance between amplification and HER2 protein expression remains undefined in EAC. Furthermore, the association between HER2 and its interacting partner, human epidermal growth factor receptor 3 (HER3), is unknown yet appears to be of potential therapeutic relevance.METHODSPatients with untreated EACs (N = 673) were analyzed for HER2 amplification and polysomy 17 by fluorescence in situ hybridization in parallel with immunohistochemistry (IHC) (IHC scores of 0‐1+, 2+, and 3+). Amplification was defined as HER2/CEP17 ≥ 2. HER3 expression by IHC was analyzed in randomly selected cases (n = 224). IHC and fluorescence in situ hybridization results were compared using least squares linear regression.RESULTSOverall, 17% of the EACs (116 of 673 EACs) were HER2‐amplified with an amplification frequency that was highest among IHC3+ cases (89%) and declined among IHC2+ cases (13%) and IHC0 to IHC1+ cases (4%). Among HER2‐amplified cases, the level of amplification increased linearly with HER2 membranous expression (HER2/CEP17 ratio: 7.9 in IHC3+ and 5.5 in IHC2+ vs 2.8 in IHC0 to IHC1+ [P < .0001]), with 14% of amplified tumors demonstrating absent/faint expression (IHC0 to IHC1+). Polysomy 17 was not found to be associated with HER2 expression. Cytoplasmic HER3 expression was detected in 87% of tumors (195 of 224 tumors) and was found to be significantly associated with better differentiation (P < .0001). Stepwise increases in HER3 expression were associated with higher HER2 expression levels (P = .0019).CONCLUSIONSLevels of HER2 protein expression and amplification were found to be linearly associated and highly concordant. Among amplified tumors with absent/faint expression, the level of amplification was low. Frequent expression of HER3 suggests its relevance as a therapeutic target, and its significant association with HER2 supports ongoing efforts to inhibit HER2/HER3 in patients with EAC. Cancer 2014;120:415–424. © 2013 American Cancer Society.

Purpose With improved patient care, better diagnosis, and more treatment options after tumor recurrence, outcomes after fluorouracil (FU) -based treatment are expected to have improved over time in early-stage colon cancer. Data from 18,449 patients enrolled onto 21 phase III trials conducted from 1978 to 2002 were evaluated for potential differences in time to recurrence (TTR), time from recurrence to death (TRD), and overall survival (OS) with regard to FU-based adjuvant regimens. Methods Trials were predefined as old versus newer era using initial accrual before or after 1995. Outcomes were compared between patients enrolled onto old- or newer-era trials, stratified by stage. Results Within the first 3 years, recurrence rates were lower in newer- versus old-era trials for patients with stage II disease, with no differences among those with stage III disease. Both TRD and OS were significantly longer in newer-era trials overall and within each stage. The lymph node (LN) ratio (ie, number of positive nodes divided by total nodes harvested) in those with stage III disease declined over time. TTR improved slightly, with larger number of LNs examined in both stages. Conclusion Improved TRD in newer trials supports the premise that more aggressive intervention (oxaliplatin- and irinotecan-based chemotherapy and/or surgery for recurrent disease) improves OS for patients previously treated in the adjuvant setting. Lower recurrence rates with identical treatments in those with stage II disease enrolled onto newer-era trials reflect stage migration over time, calling into question historical data related to the benefit of FU-based adjuvant therapy in such patients.

BACKGROUNDA point mutation (V600E) in the BRAF oncogene is a prognostic biomarker and may predict for nonresponse to anti‐EGFR antibody therapy in patients with colorectal carcinoma. BRAFV600E mutations are frequently detected in tumors with microsatellite instability and indicate a sporadic origin. We used a mutation‐specific antibody to examine mutant BRAFV600E protein expression and its concordance with BRAFV600E mutation data.METHODSPrimary stage III colon carcinomas were analyzed for BRAFV600E mutations in exon 15, and 50 BRAFV600E mutation carriers and 25 wild‐type tumors were selected for analysis of BRAF proteins by immunohistochemistry (IHC). IHC was performed in archival tissue specimens using a pan‐BRAF antibody and a mutation‐specific antibody against BRAFV600E proteins. Staining was scored by 2 pathologists who were blinded to clinical and mutation data.RESULTSUsing a pan‐BRAF antibody, total BRAF protein expression was observed in the tumor cell cytoplasm in 74 of 75 colon carcinomas. A mutation‐specific antibody identified diffuse cytoplasmic staining of mutant BRAFV600E proteins in 49 of 74 cancers. Analysis using a polymerase chain reaction‐based assay revealed that all 49 of these cancers carried BRAFV600E mutations. In contrast, BRAFV600E staining was absent in all 25 tumors that carried wild‐type copies of BRAF.CONCLUSIONSA BRAF mutation‐specific (V600E) antibody detected tumors with BRAFV600E mutations and exhibited complete concordance with a DNA‐based method. These results support the use of IHC as a simplified strategy to screen colorectal cancers for BRAFV600E mutations in clinical practice. Cancer 2013;119:2765–2770. © 2013 American Cancer Society.

AbstractA well‐designed pilot study can advance science by providing essential preliminary data to support or motivate further research, refine study logistics, and demonstrate proof of concept. Often, the outcomes of such studies can be quantified by a success/failure dichotomy. For example, a novel compound may show activation of a neural pathway, or it may not. When an intervention's efficacy is quantified using a dichotomous outcome, probability mass functions can be enumerated to determine the probability that the observed result from a pilot study supports further evaluation of the intervention since there is only a finite, and often small, number of sample configurations possible. The purpose of this research was to determine the probability of an “efficacy signal” for pilot studies using one‐ and two‐sample pilot study designs. Efficacy signal was defined as the probability of observing a more favorable response proportion relative to a historical control (one‐sample setting) or to a concurrent control (two‐sample setting). An enumeration study (exact simulation) was conducted to calculate the efficacy signal probability. One‐sample study designs yielded higher probability of determining an efficacy signal than the two‐sample setting; however, sampling variation must be accounted for. A 68% score confidence interval is recommended for this purpose.

Purpose By using data from North Central Cancer Treatment Group Phase III Trial N0147, a randomized adjuvant trial of patients with stage III colon cancer, we assessed the relationship between smoking and cancer outcomes, disease-free survival (DFS), and time to recurrence (TTR), accounting for heterogeneity by patient and tumor characteristics. Patients and Methods Before random assignment to infusional fluorouracil, leucovorin, and oxaliplatin (FOLFOX) or FOLFOX plus cetuximab, 1,968 participants completed a questionnaire on smoking history and other risk factors. Cox models assessed the association between smoking history and the primary trial outcome of DFS (ie, time to recurrence or death), as well as TTR, adjusting for other clinical and patient factors. The median follow-up was 3.5 years among patients who did not experience events. Results Compared with never-smokers, ever smokers experienced significantly shorter DFS (3-year DFS proportion: 70% v 74%; hazard ratio [HR], 1.21; 95% CI, 1.02 to 1.42). This association persisted after multivariate adjustment (HR, 1.23; 95% CI, 1.02 to 1.49). There was significant interaction in this association by BRAF mutation status (P = .03): smoking was associated with shorter DFS in patients with BRAF wild-type (HR, 1.36; 95% CI, 1.11 to 1.66) but not BRAF mutated (HR, 0.80; 95% CI, 0.50 to 1.29) colon cancer. Smoking was more strongly associated with poorer DFS in those with KRAS mutated versus KRAS wild-type colon cancer (HR, 1.50 [95% CI, 1.12 to 2.00] v HR, 1.09 [95% CI, 0.85 to 1.39]), although interaction by KRAS mutation status was not statistically significant (P = .07). Associations were comparable in analyses of TTR. Conclusion Overall, smoking was significantly associated with shorter DFS and TTR in patients with colon cancer. These adverse relationships were most evident in patients with BRAF wild-type or KRAS mutated colon cancer.