Genomic selection strategies for clonally propagated crops.

KEY MESSAGE: For genomic selection in clonally propagated crops with diploid (-like) meiotic behavior to be effective, crossing parents should be selected based on genomic predicted cross-performance unless dominance is negligible. For genomic selection (GS) in clonal breeding programs to be effective, parents should be selected based on genomic predicted cross-performance unless dominance is negligible. Genomic prediction of cross-performance enables efficient exploitation of the additive and dominance value simultaneously. Here, we compared different GS strategies for clonally propagated crops with diploid (-like) meiotic behavior, using strawberry as an example. We used stochastic simulation to evaluate six combinations of three breeding programs and two parent selection methods. The three breeding programs included (1) a breeding program that introduced GS in the first clonal stage, and (2) two variations of a two-part breeding program with one and three crossing cycles per year, respectively. The two parent selection methods were (1) parent selection based on genomic estimated breeding values (GEBVs) and (2) parent selection based on genomic predicted cross-performance (GPCP). Selection of parents based on GPCP produced faster genetic gain than selection of parents based on GEBVs because it reduced inbreeding when the dominance degree increased. The two-part breeding programs with one and three crossing cycles per year using GPCP always produced the most genetic gain unless dominance was negligible. We conclude that (1) in clonal breeding programs with GS, parents should be selected based on GPCP, and (2) a two-part breeding program with parent selection based on GPCP to rapidly drive population improvement has great potential to improve breeding clonally propagated crops.

American Joint Committee on Cancer acceptance criteria for inclusion of risk models for individualized prognosis in the practice of precision medicine.

The American Joint Committee on Cancer (AJCC) has increasingly recognized the need for more personalized probabilistic predictions than those delivered by ordinal staging systems, particularly through the use of accurate risk models or calculators. However, judging the quality and acceptability of a risk model is complex. The AJCC Precision Medicine Core conducted a 2-day meeting to discuss characteristics necessary for a quality risk model in cancer patients. More specifically, the committee established inclusion and exclusion criteria necessary for a risk model to potentially be endorsed by the AJCC. This committee reviewed and discussed relevant literature before creating a checklist unique to this need of AJCC risk model endorsement. The committee identified 13 inclusion and 3 exclusion criteria for AJCC risk model endorsement in cancer. The emphasis centered on performance metrics, implementation clarity, and clinical relevance. The facilitation of personalized probabilistic predictions for cancer patients holds tremendous promise, and these criteria will hopefully greatly accelerate this process. Moreover, these criteria might be useful for a general audience when trying to judge the potential applicability of a published risk model in any clinical domain. CA Cancer J Clin 2016;66:370-374. © 2016 American Cancer Society.

Duration of Adjuvant Chemotherapy for Stage III Colon Cancer.

BACKGROUND: Since 2004, a regimen of 6 months of treatment with oxaliplatin plus a fluoropyrimidine has been standard adjuvant therapy in patients with stage III colon cancer. However, since oxaliplatin is associated with cumulative neurotoxicity, a shorter duration of therapy could spare toxic effects and health expenditures. METHODS: We performed a prospective, preplanned, pooled analysis of six randomized, phase 3 trials that were conducted concurrently to evaluate the noninferiority of adjuvant therapy with either FOLFOX (fluorouracil, leucovorin, and oxaliplatin) or CAPOX (capecitabine and oxaliplatin) administered for 3 months, as compared with 6 months. The primary end point was the rate of disease-free survival at 3 years. Noninferiority of 3 months versus 6 months of therapy could be claimed if the upper limit of the two-sided 95% confidence interval of the hazard ratio did not exceed 1.12. RESULTS: After 3263 events of disease recurrence or death had been reported in 12,834 patients, the noninferiority of 3 months of treatment versus 6 months was not confirmed in the overall study population (hazard ratio, 1.07; 95% confidence interval [CI], 1.00 to 1.15). Noninferiority of the shorter regimen was seen for CAPOX (hazard ratio, 0.95; 95% CI, 0.85 to 1.06) but not for FOLFOX (hazard ratio, 1.16; 95% CI, 1.06 to 1.26). In an exploratory analysis of the combined regimens, among the patients with T1, T2, or T3 and N1 cancers, 3 months of therapy was noninferior to 6 months, with a 3-year rate of disease-free survival of 83.1% and 83.3%, respectively (hazard ratio, 1.01; 95% CI, 0.90 to 1.12). Among patients with cancers that were classified as T4, N2, or both, the disease-free survival rate for a 6-month duration of therapy was superior to that for a 3-month duration (64.4% vs. 62.7%) for the combined treatments (hazard ratio, 1.12; 95% CI, 1.03 to 1.23; P=0.01 for superiority). CONCLUSIONS: Among patients with stage III colon cancer receiving adjuvant therapy with FOLFOX or CAPOX, noninferiority of 3 months of therapy, as compared with 6 months, was not confirmed in the overall population. However, in patients treated with CAPOX, 3 months of therapy was as effective as 6 months, particularly in the lower-risk subgroup. (Funded by the National Cancer Institute and others.).

Disease-free Survival and Local Recurrence for Laparoscopic Resection Compared With Open Resection of Stage II to III Rectal Cancer: Follow-up Results of the ACOSOG Z6051 Randomized Controlled Trial.

OBJECTIVE: To determine the disease-free survival (DFS) and recurrence after the treatment of patients with rectal cancer with open (OPEN) or laparoscopic (LAP) resection. BACKGROUND: This randomized clinical trial (ACOSOG [Alliance] Z6051), performed between 2008 and 2013, compared LAP and OPEN resection of stage II/III rectal cancer, within 12 cm of the anal verge (T1-3, N0-2, M0) in patients who received neoadjuvant chemoradiotherapy. The rectum and mesorectum were resected using open instruments for rectal dissection (included hybrid hand-assisted laparoscopic) or with laparoscopic instruments under pneumoperitoneum. The 2-year DFS and recurrence were secondary endpoints of Z6051. METHODS: The DFS and recurrence were not powered, and are being assessed for superiority. Recurrence was determined at 3, 6, 9, 12, and every 6 months thereafter, using carcinoembryonic antigen, physical examination, computed tomography, and colonoscopy. In all, 486 patients were randomized to LAP (243) or OPEN (243), with 462 eligible for analysis (LAP = 240 and OPEN = 222). Median follow-up is 47.9 months. RESULTS: The 2-year DFS was LAP 79.5% (95% confidence interval [CI] 74.4-84.9) and OPEN 83.2% (95% CI 78.3-88.3). Local and regional recurrence was 4.6% LAP and 4.5% OPEN. Distant recurrence was 14.6% LAP and 16.7% OPEN.Disease-free survival was impacted by unsuccessful resection (hazard ratio [HR] 1.87, 95% CI 1.21-2.91): composite of incomplete specimen (HR 1.65, 95% CI 0.85-3.18); positive circumferential resection margins (HR 2.31, 95% CI 1.40-3.79); positive distal margin (HR 2.53, 95% CI 1.30-3.77). CONCLUSION: Laparoscopic assisted resection of rectal cancer was not found to be significantly different to OPEN resection of rectal cancer based on the outcomes of DFS and recurrence.

An adaptive multi-stage phase I dose-finding design incorporating continuous efficacy and toxicity data from multiple treatment cycles.

Phase I designs traditionally use the dose-limiting toxicity (DLT), a binary endpoint from the first treatment cycle, to identify the maximum-tolerated dose (MTD) assuming a monotonically increasing relationship between dose and efficacy. In this article, we establish a general framework for a multi-stage adaptive design where we jointly model a continuous efficacy outcome and continuous/quasi-continuous toxicity endpoints from multiple treatment cycles. The normalized Total Toxicity Profile (nTTP) is used as an illustration for quasi-continuous toxicity endpoints, and we replace DLT with nTTP to take into account multiple grades and types of toxicities. In addition, the proposed design accommodates non-monotone dose-efficacy relationships, and longitudinal toxicity data in effort to capture the adverse events from multiple cycles. Stage 1 of our design uses toxicity data to perform dose-escalation and identify a set of initially allowable (safe) doses; stage 2 of our design incorporates an efficacy outcome to update the set of allowable doses for each new cohort and randomizes the new cohort of patients to the allowable doses with emphasis towards those with higher predicted efficacy. Stage 3 uses all data from all treated patients at the end of the trial to make final recommendations. Simulations showed that the design had a high probability of making the correct dose selection and good overdose control across various dose-efficacy and dose-toxicity scenarios. In addition, the proposed design allows for early termination when all doses are too toxic. To our best knowledge, the proposed dual-endpoint dose-finding design is the first such study to incorporate multiple cycles of toxicities and a continuous efficacy outcome.

Identification of powdery mildew resistance QTL in strawberry (Fragaria × ananassa).

Key Message Powdery mildew resistance in two strawberry mapping populations is controlled by both stable and transient novel QTL of moderate effect. Some low transferability of QTL across wider germplasm was observed. The obligate biotrophic fungus Podosphaera aphanis is the causative agent of powdery mildew on cultivated strawberry (Fragaria × ananassa). Genotypes from two bi-parental mapping populations 'Emily' × 'Fenella' and 'Redgauntlet' × 'Hapil' were phenotyped for powdery mildew disease severity in a series of field trials. Here, we report multiple QTL associated with resistance to powdery mildew, identified in ten phenotyping events conducted across different years and locations. Six QTL show a level of stable resistance across multiple phenotyping events; however, many other QTL were represented in a single phenotyping event and therefore must be considered transient. Subsequent screening of identified QTL across a validation set determined whether identified QTL remained closely linked to the associated resistance gene in the wider germplasm. Furthermore, a preliminary association analysis identified a novel conserved locus for further investigation. Our data suggest that resistance is highly complex and that multiple, primarily additive, sources of quantitative resistance to powdery mildew exist across strawberry germplasm. Utilisation of the reported markers in marker-assisted breeding or genomic selection would lead to improved powdery mildew-resistant strawberry cultivars, particularly where the studied parents, progeny and close pedigree material are included in breeding germplasm.

A hierarchical Bayesian design for randomized Phase II clinical trials with multiple groups.

Enhanced knowledge of the biological and genetic basis of cancer is re-defining the target population for new treatments. In oncology, potential targets for a new therapeutic agent often include various solid and hematologic malignancies that share common signaling pathways. New agents are often tested in multiple tumor types across which information can be borrowed. We propose a hierarchical Bayesian design (HBD) to simultaneously test a novel agent in multiple groups for randomized Phase II clinical trials with binary endpoints. Compared to parallel design for individual tumor groups, the HBD has greatly reduced sample size. Therefore, this improves efficiency and decreases the financial cost of conducting randomized Phase II clinical trials. An R package hbdct has been developed to implement the HBD and streamline the sample size calibration.

Estimation of tumour regression and growth rates during treatment in patients with advanced prostate cancer: a retrospective analysis.

BACKGROUND: We applied mathematical models to clinical trial data available at Project Data Sphere LLC (Cary, NC, USA), a non-profit universal access data-sharing warehouse. Our aim was to assess the rates of cancer growth and regression using the comparator groups of eight randomised clinical trials that enrolled patients with metastatic castration-resistant prostate cancer. METHODS: In this retrospective analysis, we used data from eight randomised clinical trials with metastatic castration-resistant prostate cancer to estimate the growth (g) and regression (d) rates of disease burden over time. Rates were obtained by applying mathematical models to prostate-specific antigen levels as the representation of tumour quantity. Rates were compared between study interventions (prednisone, mitoxantrone, and docetaxel) and off-treatment data when on-study treatment had been discontinued to understand disease behaviour during treatment and after discontinuation. Growth (g) was examined for association with a traditional endpoint (overall survival) and for its potential use as an endpoint to reduce sample size in clinical trials. FINDINGS: Estimates for g, d, or both were obtained in 2353 (88%) of 2678 patients with data available for analysis; g differentiated docetaxel (a US Food and Drug Administration-approved therapy) from prednisone and mitoxantrone and was predictive of overall survival in a landmark analysis at 8 months. A simulated sample size analysis, in which g was used as the endpoint, compared docetaxel data with mitoxantrone data and showed that small sample sizes were sufficient to achieve 80% power (16, 47, and 25 patients, respectively, in the three docetaxel comparator groups). Similar results were found when the mitoxantrone data were compared with the prednisone data (41, 39, and 41 patients in the three mitoxantrone comparator groups). Finally, after discontinuation of docetaxel therapy, median tumour growth (g) increased by nearly five times. INTERPRETATION: The application of mathematical models to existing clinical data allowed estimation of rates of growth and regression that provided new insights in metastatic castration-resistant prostate cancer. The availability of clinical data through initiatives such as Project Data Sphere, when combined with innovative modelling techniques, could greatly enhance our understanding of how cancer responds to treatment, and accelerate the productivity of clinical development programmes. FUNDING: None.

Validation of Progression-Free Survival as a Surrogate Endpoint for Overall Survival in Malignant Mesothelioma: Analysis of Cancer and Leukemia Group B and North Central Cancer Treatment Group (Alliance) Trials.

PURPOSE: The aim of this study was to investigate whether progression-free survival (PFS) can be considered a surrogate endpoint for overall survival (OS) in malignant mesothelioma. MATERIALS AND METHODS: Individual data were collected from 15 Cancer and Leukemia Group B (615 patients) and 2 North Central Cancer Treatment Group (101 patients) phase II trials. The effects of 5 risk factors for OS and PFS, including age, histology, performance status (PS), white blood cell count, and European Organisation for Research and Treatment of Cancer (EORTC) risk score, were used in the analysis. Individual-level surrogacy was assessed by Kendall's tau through a Clayton bivariate Copula survival (CBCS) model. Summary-level surrogacy was evaluated via the association between logarithms of the hazard ratio (log HR)-log HROS and log HRPFS-measured in R2 from a weighted least-square (WLS) regression model and the CBCS model. RESULTS: The median PFS for all patients was 3.0 months (95% confidence interval [CI], 2.8-3.5 months) and the median OS was 7.2 months (95% CI, 6.5-8.0 months). Moderate correlations between PFS and OS were observed across all risk factors at the individual level, with Kendall's tau ranging from 0.46 to 0.47. The summary-level surrogacy varied among risk factors. The Copula R2 ranged from 0.51 for PS to 0.78 for histology. The WLS R2 ranged from 0.26 for EORTC and PS to 0.67 for age. CONCLUSIONS: The analyses demonstrated low to moderate individual-level surrogacy between PFS and OS. At the summary level, the surrogacy between PFS and OS varied significantly across different risk factors. With a short postprogression survival and a moderate correlation between PFS and OS, there is no evidence that PFS is a valid surrogate endpoint for OS in malignant mesothelioma. The Oncologist 2017;22:189-198Implications for Practice: For better disease management and for more efficient clinical trial designs, it is important to know if progression-free survival (PFS) is a good surrogate endpoint for overall survival in malignant mesothelioma. With a relatively large database of 17 phase II trials and 716 patients from Cancer and Leukemia Group B and North Central Cancer Treatment Group, we conducted statistical analyses and found that there is no evidence to suggest that PFS is a valid surrogate endpoint for OS for malignant mesothelioma. Future research work is needed to find alternative surrogate endpoints for OS.

Altered regulation of TERMINAL FLOWER 1 causes the unique vernalisation response in an arctic woodland strawberry accession.

Vernalisation requirement is an agriculturally important trait that postpones the development of cold-sensitive floral organs until the spring. The family Rosaceae includes many agriculturally important fruit and berry crops that suffer from crop losses caused by frost injury to overwintering flower buds. Recently, a vernalisation-requiring accession of the Rosaceae model woodland strawberry (Fragaria vesca) has been identified in northern Norway. Understanding the molecular basis of the vernalisation requirement in this accession would advance the development of strawberry cultivars better adapted to temperate climate. We use gene silencing, gene expression analysis, genetic mapping and population genomics to study the genetic basis of the vernalisation requirement in woodland strawberry. Our results indicate that the woodland strawberry vernalisation requirement is endemic to northern Norwegian population, and mapping data suggest the orthologue of TERMINAL FLOWER1 (FvTFL1) as the causal floral repressor. We demonstrate that exceptionally low temperatures are needed to downregulate FvTFL1 and to make these plants competent to induce flowering at low postvernalisation temperatures in the spring. We show that altered regulation of FvTFL1 in the northern Norwegian woodland strawberry accession postpones flower induction until the spring, allowing plants to avoid winter injuries of flower buds that commonly occur in temperate regions.

A Bayesian dose-finding design incorporating toxicity data from multiple treatment cycles.

Phase I oncology trials are designed to identify a safe dose with an acceptable toxicity profile. The dose is typically determined based on the probability of severe toxicity observed during the first treatment cycle, although patients continue to receive treatment for multiple cycles. In addition, the toxicity data from multiple types and grades are typically summarized into a single binary outcome of dose-limiting toxicity. A novel endpoint, the total toxicity profile, was previously developed to account for the multiple toxicity types and grades. In this work, we propose to account for longitudinal repeated measures of total toxicity profile over multiple treatment cycles, accounting for cumulative toxicity during dosing-finding. A linear mixed model was utilized in the Bayesian framework, with addition of Bayesian risk functions for decision-making in dose assignment. The performance of this design is evaluated using simulation studies and compared with the previously proposed quasi-likelihood continual reassessment method (QLCRM) design. Twelve clinical scenarios incorporating four different locations of maximum tolerated dose and three different time trends (decreasing, increasing, and no effect) were investigated. The proposed repeated measures design was comparable with the QLCRM when only cycle 1 data were utilized in dose-finding; however, it demonstrated an improvement over the QLCRM when data from multiple cycles were used across all scenarios. Copyright © 2016 John Wiley & Sons, Ltd.

Repeated measures dose-finding design with time-trend detection in the presence of correlated toxicity data.

BACKGROUND: Phase I trials are designed to determine the safety, tolerability, and recommended phase 2 dose of therapeutic agents for subsequent testing. The dose-finding paradigm has thus traditionally focused on identifying the maximum tolerable dose of an agent or combination therapy under the assumption that there is a non-decreasing relationship between dose-toxicity and dose-efficacy. The dose is typically determined based on the probability of severe toxicity observed during the first treatment cycle. A novel endpoint, the total toxicity profile, was previously developed to account for the multiple toxicity types and grades experienced in the first cycle. More recently, this was extended to a repeated measures design based on the total toxicity profile to account for longitudinal toxicities over multiple treatment cycles in the absence of within-patient correlation. METHODS: In this work, we propose to extend the design in the presence of within-patient correlation. Furthermore, we provide a framework to detect a toxicity time trend (toxicity increasing, decreasing, or stable) over multiple treatment cycles. We utilize a linear mixed model in the Bayesian framework, with the addition of Bayesian risk functions for decision-making in dose assignment. RESULTS: The performance of this design was evaluated using simulation studies and real data from a phase I trial. We demonstrated that using available toxicity data from all cycles of treatment improves the accuracy of maximum tolerated dose identification and allows for the detection of a time trend. The performance is consistent regardless of the strength of the within-patient correlation. In addition, the use of a quasi-continuous total toxicity profile score significantly increased the power to detect time trends compared to when binary data only were used. CONCLUSION: The increased interest in molecularly targeted agents and immunotherapies in oncology necessitates innovative phase I study designs. Our proposed framework provides a tool to tackle some of the challenges presented by these novel agents, specifically through the ability to understand patterns of toxicity over time, which is important in the cases of cumulative or late toxicities.

Prognosis of patients with peritoneal metastatic colorectal cancer given systemic therapy: an analysis of individual patient data from prospective randomised trials from the Analysis and Research in Cancers of the Digestive System (ARCAD) database.

BACKGROUND: Patients with peritoneal metastatic colorectal cancer have reduced overall survival compared with patients with metastatic colorectal cancer without peritoneal involvement. Here we further investigated the effect of the number and location of metastases in patients receiving first-line systemic chemotherapy. METHODS: We analysed individual patient data for previously untreated patients enrolled in 14 phase 3 randomised trials done between 1997 and 2008. Trials were included if protocols explicitly pre-specified and solicited for patients with peritoneal involvement in the trial data collection process or had done a formal peritoneum-focused review of individual pre-treatment scans. We used stratified multivariable Cox models to assess the prognostic associations of peritoneal metastatic colorectal cancer with overall survival and progression-free survival, adjusting for other key clinical-pathological factors (age, sex, Eastern Cooperative Oncology Group (ECOG) performance score, primary tumour location [colon vs rectum], previous treatment, and baseline BMI). The primary endpoint was difference in overall survival between populations with and without peritoneal metastases. FINDINGS: Individual patient data were available for 10 553 patients. 9178 (87%) of 10 553 patients had non-peritoneal metastatic colorectal cancer (4385 with one site of metastasis, 4793 with two or more sites of metastasis), 194 (2%) patients had isolated peritoneal metastatic colorectal cancer, and 1181 (11%) had peritoneal metastatic colorectal cancer and other organ involvement. These groups were similar in age, ethnic origin, and use of targeted treatment. Patients with peritoneal metastatic colorectal cancer were more likely than those with non-peritoneal metastatic colorectal cancer to be women (565 [41%] of 1371 vs 3312 [36%] of 9169 patients; p=0·0003), have colon primary tumours (1116 [84%] of 1334 patients vs 5603 [66%]; p<0·0001), and have performance status of 2 (136 [10%] vs 521 [6%]; p<0·0001). We recorded a higher proportion of patients with mutated BRAF in patients with peritoneal-only (eight [18%] of 44 patients with available data) and peritoneal metastatic colorectal cancer with other sites of metastasis (34 [12%] of 289), compared with patients with non-peritoneal metastatic colorectal cancer (194 [9%] of 2230; p=0·028 comparing the three groups). Overall survival (adjusted HR 0·75, 95% CI 0·63-0·91; p=0·003) was better in patients with isolated non-peritoneal sites than in those with isolated peritoneal metastatic colorectal cancer. Overall survival of patients with two of more non-peritoneal sites of metastasis (adjusted HR 1·04, 95% CI 0·86-1·25, p=0.69) and those with peritoneal metastatic colorectal cancer plus one other site of metastasis (adjusted HR 1·10, 95% CI 0·89-1·37, p=0·37) was similar to those with isolated peritoneal metastases. Compared with patients with isolated peritoneal metastases, those with peritoneal metastases and two or more additional sites of metastasis had the shortest survival (adjusted HR 1·40; CI 1·14-1·71; p=0·0011). INTERPRETATION: Patients with peritoneal metastatic colorectal cancer have significantly shorter overall survival than those with other isolated sites of metastases. In patients with several sites of metastasis, poor survival is a function of both increased number of metastatic sites and peritoneal involvement. The pattern of metastasis and in particular, peritoneal involvement, results in prognostic heterogeneity of metastatic colorectal cancer. FUNDING: None.

Alcohol consumption and colon cancer prognosis among participants in north central cancer treatment group phase III trial N0147.

Alcohol consumption is associated with a modest increased risk of colon cancer, but its relationship with colon cancer survival has not been elucidated. Using data from a phase III randomized adjuvant trial, we assessed the association of alcohol consumption with colon cancer outcomes. Patients completed a risk factor questionnaire before randomization to FOLFOX or FOLFOX + cetuximab (N = 1984). Information was collected on lifestyle factors, including smoking, physical activity and consumption of different types of alcohol. Cox models assessed the association between alcohol consumption and outcomes of disease-free survival (DFS), time-to-recurrence (TTR) and overall survival (OS), adjusting for age, sex, study arm, body mass, smoking, physical activity and performance status. No statistically significant difference in outcomes between ever and never drinkers were noted [hazard ratio (HR)DFS = 0.86, HRTTR = 0.87, HROS = 0.86, p-values = 0.11-0.17]. However, when considering alcohol type, ever consumers of red wine (n = 628) had significantly better outcomes than never consumers (HRDFS = 0.80, HRTTR = 0.81, HROS = 0.78, p-values = 0.01-0.02). Favorable outcomes were confirmed in patients who consumed 1-30 glasses/month of red wine (n = 601, HR = 0.80-0.83, p-values = 0.03-0.049); there was a suggestion of more favorable outcomes in patients who consumed >30 glasses/month of red wine (n = 27, HR = 0.33-0.38, p-values = 0.05-0.06). Beer and liquor consumption were not associated with outcomes. Although alcohol consumption was not associated with colon cancer outcomes overall, mild to moderate red wine consumption was suggestively associated with longer OS, DFS and TTR in stage III colon cancer patients.

Association between DPYD c.1129-5923 C>G/hapB3 and severe toxicity to 5-fluorouracil-based chemotherapy in stage III colon cancer patients: NCCTG N0147 (Alliance).

Severe (grade≥3) adverse events (AEs) to 5-fluorouracil (5-FU)-based chemotherapy regimens can result in treatment delays or cessation, and, in extreme cases, life-threatening complications. Current genetic biomarkers for 5-FU toxicity prediction, however, account for only a small proportion of toxic cases. In the current study, we assessed DPYD variants suggested to correlate with 5-FU toxicity, a deep intronic variant (c.1129-5923 C>G), and four variants within a haplotype (hapB3) in 1953 stage III colon cancer patients who received adjuvant FOLFOX±cetuximab. Logistic regression was used to assess multivariable associations between DPYD variant status and AEs common to 5-FU (5FU-AEs). In our study cohort, 1228 patients (62.9%) reported any grade≥3 AE (overall AE), with 638 patients (32.7%) reporting any grade≥3 5FU-AE. Only 32 of 78 (41.0%) patients carrying DPYD c.1129-5923 C>G and the completely linked hapB3 variants c.1236 C>G and c.959-51 T>C showed at least one grade≥3 5FU-AE, resulting in no statistically significant association (adjusted odds ratio=1.47, 95% confidence interval=0.90-2.43, P=0.1267). No significant associations were identified between c.1129-5923 C>G/hapB3 and overall grade≥3 AE rate. Our results suggest that c.1129-5923 C>G/hapB3 have limited predictive value for severe toxicity to 5-FU-based combination chemotherapy.

Relationship Between Metformin Use and Recurrence and Survival in Patients With Resected Stage III Colon Cancer Receiving Adjuvant Chemotherapy: Results From North Central Cancer Treatment Group N0147 (Alliance).

BACKGROUND: Preclinical and epidemiological data suggest that metformin might have antineoplastic properties against colon cancer (CC). However, the effect of metformin use on patient survival in stage III CC after curative resection is unknown. The survival outcomes were comparable regardless of the duration of metformin use. PATIENTS AND METHODS: Before randomization to FOLFOX (folinic acid, 5-fluorouracil, oxaliplatin) with or without cetuximab, 1,958 patients with stage III CC enrolled in the N0147 study completed a questionnaire with information on diabetes mellitus (DM) and metformin use. Cox models were used to assess the association between metformin use and disease-free survival (DFS), overall survival (OS), and the time to recurrence (TTR), adjusting for clinical and/or pathological factors. RESULTS: Of the 1,958 patients, 1,691 (86%) reported no history of DM, 115 reported DM with metformin use (6%), and 152 reported DM without metformin use (8%). The adjuvant treatment arms were pooled, because metformin use showed homogeneous effects on outcomes across the two arms. Among the patients with DM (n = 267), DFS (adjusted hazard ratio [aHR], 0.90; 95% confidence interval [CI], 0.59-1.35; p = .60), OS (aHR, 0.99; 95% CI, 0.65-1.49; p = .95), and TTR (aHR, 0.87; 95% CI, 0.56-1.35; p = .53) were not different for the metformin users compared with the nonusers after adjusting for tumor and patient factors. The survival outcomes were comparable regardless of the duration of metformin use (<1, 1-5, 6-10, ≥11 years) before randomization (ptrend = .64 for DFS, ptrend = .84 for OS, and ptrend = .87 for TTR). No interaction effects were observed between metformin use and KRAS, BRAF mutation status, tumor site, T/N stage, gender, or age. CONCLUSIONS: Patients with stage III CC undergoing adjuvant chemotherapy who used metformin before the diagnosis of CC experienced DFS, OS, and TTR similar to those for non-DM patients and DM patients without metformin use. IMPLICATIONS FOR PRACTICE: The present study did not find any relationship between metformin use or its duration and disease-free survival, time to recurrence, and overall survival in a large cohort of patients with resected stage III colon cancer receiving adjuvant FOLFOX (folinic acid, fluorouracil, oxaliplatin)-based chemotherapy. This relationship was not modified by KRAS or BRAF mutation or DNA mismatch repair status. Metformin use did not increase or decrease the likelihood of chemotherapy-related grade 3 or higher adverse events.

Molecular markers identify subtypes of stage III colon cancer associated with patient outcomes.

BACKGROUND & AIMS: Categorization of colon cancers into distinct subtypes using a combination of pathway-based biomarkers could provide insight into stage-independent variability in outcomes. METHODS: We used a polymerase chain reaction-based assay to detect mutations in BRAF (V600E) and in KRAS in 2720 stage III cancer samples, collected prospectively from patients participating in an adjuvant chemotherapy trial (NCCTG N0147). Tumors deficient or proficient in DNA mismatch repair (MMR) were identified based on detection of MLH1, MSH2, and MSH6 proteins and methylation of the MLH1 promoter. Findings were validated using tumor samples from a separate set of patients with stage III cancer (n = 783). Association with 5-year disease-free survival was evaluated using Cox proportional hazards models. RESULTS: Tumors were categorized into 5 subtypes based on MMR status and detection of BRAF or KRAS mutations which were mutually exclusive. Three subtypes were MMR proficient: those with mutations in BRAF (6.9% of samples), mutations in KRAS (35%), or tumors lacking either BRAF or KRAS mutations (49%). Two subtypes were MMR deficient: the sporadic type (6.8%) with BRAF mutation and/or or hypermethylation of MLH1 and the familial type (2.6%), which lacked BRAF(V600E) or hypermethylation of MLH1. A higher percentage of MMR-proficient tumors with BRAF(V600E) were proximal (76%), high-grade (44%), N2 stage (59%), and detected in women (59%), compared with MMR-proficient tumors without BRAF(V600E) or KRAS mutations (33%, 19%, 41%, and 42%, respectively; all P < .0001). A significantly lower proportion of patients with MMR-proficient tumors with mutant BRAF (hazard ratio = 1.43; 95% confidence interval: 1.11-1.85; Padjusted = .0065) or mutant KRAS (hazard ratio = 1.48; 95% confidence interval: 1.27-1.74; Padjusted < .0001) survived disease-free for 5 years compared with patients whose MMR-proficient tumors lacked mutations in either gene. Disease-free survival rates of patients with MMR-deficient sporadic or familial subtypes was similar to those of patients with MMR-proficient tumors without BRAF or KRAS mutations. The observed differences in survival rates of patients with different tumor subtypes were validated in an independent cohort. CONCLUSIONS: We identified subtypes of stage III colon cancer, based on detection of mutations in BRAF (V600E) or KRAS, and MMR status that show differences in clinical and pathologic features and disease-free survival. Patients with MMR-proficient tumors and BRAF or KRAS mutations had statistically shorter survival times than patients whose tumors lacked these mutations. The tumor subtype found in nearly half of the study cohort (MMR-proficient without BRAF(V600E) or KRAS mutations) had similar outcomes to those of patients with MMR-deficient cancers.

New insights into the evaluation of randomized controlled trials for rare diseases over a long-term research horizon: a simulation study.

Large sample sizes are required in randomized clinical trials designed to meet typical one-sided 2.5% α-level and 80% power. This may not be achievable when the disease is rare. We simulated a series of two-arm superiority trials over a 15-year period. The design parameters examined were the α-level and the number of trials conducted over the 15-year period (thus, trial sample size). Different disease severities and accrual rates were considered. The future treatment effect was characterized by its associated hazard rate; different hypotheses of how treatments improve over time were considered. We defined the total survival benefit as the relative difference of the hazard rates at year 15 versus year 0. The optimal design was defined by maximizing the expected total survival benefit, provided that the risk of selecting at year 15 a treatment inferior to the initial control treatment remains below 1%. Compared with two larger trials with typical one-sided 2.5% α-level, performing a series of small trials with relaxed α-levels leads on average to larger survival benefits over a 15-year research horizon, but also to higher risk of selecting a worse treatment at the end of the research period. Under reasonably optimistic assumptions regarding the future treatment effects, optimal designs outperform traditional ones when the disease is severe (baseline median survival ≤ 1 year) and the accrual is ≥100 patients per year, whereas no major improvement is observed in diseases with better prognosis. Trial designs aiming to maximize survival gain over a long research horizon across a series of trials are worth discussing in the context of rare diseases. Copyright © 2016 John Wiley & Sons, Ltd.

Analysis of circulating DNA and protein biomarkers to predict the clinical activity of regorafenib and assess prognosis in patients with metastatic colorectal cancer: a retrospective, exploratory analysis of the CORRECT trial.

BACKGROUND: Tumour mutational status is an important determinant of the response of metastatic colorectal cancer to targeted treatments. However, the genotype of the tissue obtained at the time of diagnosis might not accurately represent tumour genotype after multiple lines of treatment. This retrospective exploratory analysis investigated the clinical activity of regorafenib in biomarker subgroups of the CORRECT study population defined by tumour mutational status or plasma protein levels. METHODS: We used BEAMing technology to identify KRAS, PIK3CA, and BRAF mutations in DNA obtained from the plasma of 503 patients with metastatic colorectal cancer who enrolled in the CORRECT trial. We quantified total human genomic DNA isolated from plasma samples for 503 patients using a modified version of human long interspersed nuclear element-1 (LINE-1) quantitive real-time PCR. We also measured the concentration of 15 proteins of interest-angiopoietin 2, interleukin 6, interleukin 8, placental growth factor, soluble TIE-1, soluble VEGFR1, VEGF-A, VEGF-C, VEGF-D, VEGF-A isoform 121, bone morphogenetic protein 7, macrophage colony-stimulating factor, stromal cell-derived factor-1, tissue inhibitor of metalloproteinase 2, and von Willebrand factor-in plasma samples from 611 patients. We did correlative analyses of overall survival and progression-free survival in patient subgroups based on mutational status, circulating DNA concentration, and protein concentrations. The CORRECT trial was registered with ClinicalTrials.gov, number NCT01103323. FINDINGS: Tumour-associated mutations were readily detected with BEAMing of plasma DNA, with KRAS mutations identified in 349 (69%) of 503 patients, PIK3CA mutations in 84 (17%) of 503 patients, and BRAF mutations in 17 (3%) of 502 patients. We did not do correlative analysis based on BRAF genotype because of the low mutational frequency detected for this gene. Some of the most prevalent individual hot-spot mutations we identified included: KRAS (KRAS G12D, 116 [28%] of 413 mutations; G12V, 72 [17%]; and G13D, 67 [16%]) and PIK3CA (PIK3CA E542K, 27 [30%] of 89 mutations; E545K, 37 [42%]; and H1047R, 12 [14%]). 41 (48%) of 86 patients who had received anti-EGFR therapy and whose archival tumour tissue DNA was KRAS wild-type in BEAMing analysis were identified as having KRAS mutations in BEAMing analysis of fresh plasma DNA. Correlative analyses suggest a clinical benefit favouring regorafenib across patient subgroups defined by KRAS and PIK3CA mutational status (progression-free survival with regorafenib vs placebo: hazard ratio [HR] 0·52, 95% CI 0·35-0·76 for KRAS wild-type; HR 0·51, 95% CI 0·40-0·65 for KRAS mutant [KRAS wild type vs mutant, pinteraction=0·74]; HR 0·50, 95% CI 0·40-0·63 for PIK3CA wild-type; HR 0·54, 95% CI 0·32-0·89 for PIK3CA mutant [PIK3CA wild-type vs mutant, pinteraction=0·85]) or circulating DNA concentration (progression-free survival with regorafenib vs placebo: HR 0·53, 95% CI 0·40-0·71, for low circulating DNA concentrations; HR 0·52, 95% CI 0·40-0·70, for high circulating DNA concentrations; low vs high circulating DNA, pinteraction=0·601). With the exception of von Willebrand factor, assessed with the median cutoff method, plasma protein concentrations were also not associated with regorafenib activity in terms of progression-free survival. In univariable analyses, the only plasma protein that was associated with overall survival was TIE-1, high concentrations of which were associated with longer overall survival compared with low TIE-1 concentrations. This association was not significant in multivariable analyses. INTERPRETATION: BEAMing of circulating DNA could be a viable approach for non-invasive analysis of tumour genotype in real time and for the identification of potentially clinically relevant mutations that are not detected in archival tissue. Additionally, the results show that regorafenib seems to be consistently associated with a clinical benefit in a range of patient subgroups based on mutational status and protein biomarker concentrations. FUNDING: Bayer HealthCare Pharmaceuticals.

Development and preliminary evaluation of a 90 K Axiom® SNP array for the allo-octoploid cultivated strawberry Fragaria × ananassa.

BACKGROUND: A high-throughput genotyping platform is needed to enable marker-assisted breeding in the allo-octoploid cultivated strawberry Fragaria × ananassa. Short-read sequences from one diploid and 19 octoploid accessions were aligned to the diploid Fragaria vesca 'Hawaii 4' reference genome to identify single nucleotide polymorphisms (SNPs) and indels for incorporation into a 90 K Affymetrix® Axiom® array. We report the development and preliminary evaluation of this array. RESULTS: About 36 million sequence variants were identified in a 19 member, octoploid germplasm panel. Strategies and filtering pipelines were developed to identify and incorporate markers of several types: di-allelic SNPs (66.6%), multi-allelic SNPs (1.8%), indels (10.1%), and ploidy-reducing "haploSNPs" (11.7%). The remaining SNPs included those discovered in the diploid progenitor F. iinumae (3.9%), and speculative "codon-based" SNPs (5.9%). In genotyping 306 octoploid accessions, SNPs were assigned to six classes with Affymetrix's "SNPolisher" R package. The highest quality classes, PolyHigh Resolution (PHR), No Minor Homozygote (NMH), and Off-Target Variant (OTV) comprised 25%, 38%, and 1% of array markers, respectively. These markers were suitable for genetic studies as demonstrated in the full-sib family 'Holiday' × 'Korona' with the generation of a genetic linkage map consisting of 6,594 PHR SNPs evenly distributed across 28 chromosomes with an average density of approximately one marker per 0.5 cM, thus exceeding our goal of one marker per cM. CONCLUSIONS: The Affymetrix IStraw90 Axiom array is the first high-throughput genotyping platform for cultivated strawberry and is commercially available to the worldwide scientific community. The array's high success rate is likely driven by the presence of naturally occurring variation in ploidy level within the nominally octoploid genome, and by effectiveness of the employed array design and ploidy-reducing strategies. This array enables genetic analyses including generation of high-density linkage maps, identification of quantitative trait loci for economically important traits, and genome-wide association studies, thus providing a basis for marker-assisted breeding in this high value crop.