Sinicrope, Frank A.

PURPOSE The microsatellite instability (MSI) or deficient mismatch repair (dMMR) phenotype is usually regarded as a single biologic entity, given the absence of comparative analyses regarding prognosis and response to chemotherapy between sporadic and familial dMMR cancers. PATIENTS AND METHODS Patients with stage III colon cancers were randomly assigned to FOLFOX (leucovorin, fluorouracil, and oxaliplatin) with or without cetuximab in 2 large adjuvant phase III trials (N = 5,577). Among patients with MSI and KRAS exon 2 wild-type (WT) tumors, the prognostic and predictive impacts of sporadic versus familial dMMR cancers and BRAF V600E mutational status were determined. Multivariable Cox proportional hazards models were used to assess disease-free survival (DFS) by treatment arm, adjusting for age, sex, tumor grade, Eastern Cooperative Oncology Group performance status, pT/pN stage, and primary tumor location. RESULTS Among patients with MSI status with complete data for dMMR mechanism analysis (n = 354), 255 (72%) had sporadic ( BRAF mutation and/or MLH1 methylation) and 99 (28%) had familial tumors ( BRAF WT and unmethylated MLH1 or loss of MSH2/MSH6/PMS2 protein expression). A large proportion of dMMR sporadic tumors were mutated for BRAF (n = 200). In patients treated with FOLFOX, DFS did not differ statistically by dMMR mechanism, whereas in patients treated with FOLFOX plus cetuximab, those with sporadic tumors had worse DFS than those with familial cancers (multivariable hazard ratio, 2.69; 95% CI, 1.02 to 7.08; P = .04). Considering the predictive utility, the interaction between treatment and dMMR mechanism was significant ( P = .03). Furthermore, a nonsignificant trend toward a deleterious effect of adding cetuximab to FOLFOX was observed in patients with BRAF-mutant but not BRAF WT tumors. CONCLUSION The addition of cetuximab to adjuvant FOLFOX was associated with shorter DFS in patients with sporadic dMMR colon cancer. Additional studies are needed to validate these results in metastatic disease.

AbstractBackgroundThe addition of bevacizumab to chemotherapy improved outcomes for patients with metastatic colon cancer. E5204 was designed to test whether the addition of bevacizumab to mFOLFOX6, following neoadjuvant chemoradiation and definitive surgery, could improve overall survival (OS) in patients with stage II/III adenocarcinoma of the rectum.Subjects, Materials, and MethodsPatients with stage II/III rectal cancer who had completed neoadjuvant 5-fluorouracil-based chemoradiation and had undergone complete resection were enrolled. Patients were randomized to mFOLFOX6 (Arm A) or mFOLFOX6 with bevacizumab (Arm B) administered every 2 weeks for 12 cycles.ResultsE5204 registered only 355 patients (17% of planned accrual goal) as it was terminated prematurely owing to poor accrual. At a median follow-up of 72 months, there was no difference in 5-year overall survival (88.3% vs. 83.7%) or 5-year disease-free survival (71.2% vs. 76.5%) between the two arms. The rate of treatment-related grade ≥ 3 adverse events (AEs) was 68.8% on Arm A and 70.7% on Arm B. Arm B had a higher proportion of patients who discontinued therapy early as a result of AEs and patient withdrawal than did Arm A (32.4% vs. 21.5%, p = .029).The most common grade 3–4 treatment-related AEs were neutropenia, leukopenia, neuropathy, diarrhea (without prior colostomy), and fatigue.ConclusionAt 17% of its planned accrual, E5204 did not meet its primary endpoint. The addition of bevacizumab to FOLFOX6 in the adjuvant setting did not significantly improve OS in patients with stage II/III rectal cancer.Implications for PracticeAt 17% of its planned accrual, E5204 was terminated early owing to poor accrual. At a median follow-up of 72 months, there was no significant difference in 5-year overall survival (88.3% vs. 83.7%) or in 5-year disease-free survival (71.2% vs. 76.5%) between the two arms. Despite significant advances in the treatment of rectal cancer, especially in improving local control rates, the risk of distant metastases and the need to further improve quality of life remain a challenge. Strategies combining novel agents with chemoradiation to improve both distant and local control are needed.

Abstract Difluoromethylornithine (DFMO), an inhibitor of polyamine synthesis, was shown to act synergistically with a NSAID for chemoprevention of colorectal neoplasia. We determined the efficacy and safety of DFMO plus aspirin for prevention of colorectal adenomas and regression of rectal aberrant crypt foci (ACF) in patients with prior advanced adenomas or cancer. A double-blinded, placebo-controlled trial was performed in 104 subjects (age 46–83) randomized (1:1) to receive daily DFMO (500 mg orally) plus aspirin (325 mg) or matched placebos for one year. All polyps were removed at baseline. Adenoma number (primary endpoint) and rectal ACF (index cluster and total) were evaluated at a one year colonoscopy. ACF were identified by chromoendoscopy. Toxicity was monitored, including audiometry. Eighty-seven subjects were evaluable for adenomas or ACF modulation (n = 62). At one year of treatment, adenomas were detected in 16 (38.1%) subjects in the DFMO plus aspirin arm (n = 42) versus 18 (40.9%) in the placebo arm (n = 44; P = 0.790); advanced adenomas were similar (n = 3/arm). DFMO plus aspirin was associated with a statistically significant reduction in the median number of rectal ACF compared with placebo (P = 0.036). Total rectal ACF burden was also reduced in the treatment versus the placebo arm relative to baseline (74% vs. 45%, P = 0.020). No increase in adverse events, including ototoxicity, was observed in the treatment versus placebo arms. While adenoma recurrence was not significantly reduced by one year of DFMO plus aspirin, the drug combination significantly reduced rectal ACF number consistent with a chemopreventive effect.

Abstract Resistance to chemotherapy remains a major obstacle to improving outcomes of cancer treatment. Recent data suggest intrinsic functions of immune checkpoint proteins, including PD-L1, that may be distinct from their immunoregulatory roles. We found that colorectal cancer (CRC) cells with mutant BRAF(V600E) or KRASalleles show increased PD-L1 transcripts and expression of cell surface PD-L1 protein compared to those with wild-type copies. Furthermore, ectopic expression of BRAFV600E or mutant KRAS induced PD-L1 whose upregulation was attenuated by inhibition of RAS-RAF-MEK-ERK signaling. Combined knockdown of MEK/ERK effectors c-JUN and YAP markedly reduced PD-L1. Knockout of PD-L1 in RKO cells resulted in a reduction in chemotherapy-induced DNA double strand breaks (pH2AX) and caspase-3 cleavage compared to parental cells. Results were confirmed in PD-L1 knockout MC38 murine CRC cells where re-expression of PD-L1, but not its deletion mutants, was shown to induced pH2AX and apoptosis. In cells with knockout of PD-L1, reductions in p-AKT and the BH3-only proteins BIM and BIK were observed that could be restored by re-expression of PD-L1. Treatment of cells with an anti-PD-L1 antibody reduced p-AKT, BIM and BIK, and also attenuated chemotherapy-induced apoptosis. Re-expression of BIM in PD-L1 knockout cells restored apoptosis. Murine tumor xenografts generated from cells with knockout of PD-L1 displayed resistance to oxaliplatin-induced tumor regression compared to tumors derived from parental cells. Using TCGA datasets, human CRCs with high vs low PD-L1 mRNA levels were significantly associated with better survival. In summary, we identified a non-immune function of tumor cell-intrinsic PD-L1 that upregulates BH3-only proteins BIM and BIK to promote apoptosis, thereby suggesting its potential as a predictive biomarker. Citation Format: Daofu Feng, Krishnendu Pal, Debabrata Mukhopadhyay, Xin Liu, Haidong Dong, Shengbing Huang, Frank A. Sinicrope. Tumor intrinsic PD-L1 is mediated by BRAFV600E and can regulate drug-induced apoptosis in human colon cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 705.

e15169 Background: In metastatic colorectal cancer with deficient DNA mismatch repair (dMMR), anti-PD-1 antibody monotherapy produced high tumor response rates and extended progression-free survival compared to lack of benefit for cancers with proficient MMR. In an ongoing phase III randomized trial, we will determine if the addition of the anti-PD-L1 antibody, atezolizumab (Genentech), to adjuvant FOLFOX can improve patient disease-free survival (DFS) vs FOLFOX alone in patients with stage III colon cancers with dMMR. By blocking the PD-1/PD-L1 interaction, atezolizumab may activate T cells, thereby, restoring their ability to detect and attack tumor cells. Limited data suggest that FOLFOX may increase intratumoral cytotoxic CD8+ T cells that could serve as ‘immune priming'. Methods: Patients with curatively resected stage III colon carcinomas with evidence of dMMR are randomized to modified FOLFOX6 for 6 months (12 cycles) alone (control arm) or combined with atezolizumab (840 mg IV q2 wk) with continuation of the antibody as monotherapy for an additional 6 months (total duration of 12 months) [experimental arm]. Patients will be stratified by T, N stage and tumor sidedness. Local testing for MMR proteins is allowed. Atezolizumab must begin by/with cycle 2. One cycle of FOLFOX is allowed pre-registration. The targeted accrual goal of 700 patients and 165 disease-free survival (DFS) events will provide 90% power to detect an effect size expressed as hazard ratio of 0.6 for the primary endpoint of DFS at two-sided alpha of 0.05. Interim analyses are planned at 50% and 75% of events. Secondary endpoints include overall survival, treatment tolerability, and quality of life. Results: This study is being conducted by the Alliance for Clinical Trials in Oncology, was approved by NCI CTEP and activated in 09/2017. The study is actively accruing and, as of 02/11/2019, 152 patients are enrolled. We are actively exploring an international collaboration. Conclusions: This is a current clinical trial in progress. Clinical trial information: NCT02912559.

3583 Background: The Microsatellite instability (MSI) or deficient mismatch repair (dMMR) phenotype is usually taken as a single biological entity whereas no data are available concerning prognosis and response to chemotherapy between sporadic and familial dMMR cases. Methods: Resected KRAS exon 2 wild-type (WT) tumor stage III colon cancers (N = 4596) from patients (pts) randomly assigned to FOLFOX +/- cetuximab in two adjuvant large phase III trials were prospectively analyzed for MSI status and dMMR mechanism (sporadic vs familial). Stratified Cox models were used to assess prognostic and predictive values of dMMR mechanism by treatment arms, adjusting for age, gender, tumor grade, ECOG PS, pT/pN stage and primary tumor location. Results: Among dMMR patients with complete data for dMMR mechanism analysis (N = 354), there were 255 (72%) sporadic ( BRAF mutated or WT with MLH1 methylation) and 99 (28%) familial (loss of MSH2 or MSH6, or loss MLH1 with BRAF WT and unmethylated MLH1) cases. A large proportion of dMMR sporadic cases were mutated for BRAF (n = 200; 80%). In pts treated with FOLFOX, the disease-free survival (DFS) was not statistically different by dMMR mechanism, while for pts treated with FOLFOX + cetuximab, the sporadic cases did worse than familial cases (DFS; adjusted (adj) HR, 2.69; 95% CI, 1.02-7.08; P= 0.04). Considering the predictive value, a deleterious effect of adding cetuximab to FOLFOX was observed in sporadic (DFS; adjHR, 1.68; 95% CI, 1.01-2.79; P= 0.04) but not in familial dMMR pts (interaction P value regarding treatment effect = 0.03). Furthermore, a non-significant trend to a deleterious effect of adding cetuximab to FOLFOX was observed in BRAF mutant (DFS; adjHR, 1.66; 95% CI, 0.95-2.92; P= 0.07) but not in BRAF WT pts. Conclusions: The addition of cetuximab to FOLFOX was associated with reduced DFS in patients with sporadic dMMR cases. Further studies including the methylator phenotype (CIMP) analysis are needed to validate these results. Clinical trial information: NCT00265811 and NCT00079274.

3532 Background: Colorectal cancer with dMMR display heterogeneity in the extent of intratumoral T-cell infiltration which may explain their variable responsiveness to PD-1 blockade. We examined the association of intratumoral CD3+ and CD8+ T-cell densities (TCD) with objective response rate (ORR) and response duration in patients with dMMR mCRCs receiving pembrolizumab (PEM). Methods: Record review was performed on 12 patients with dMMR mCRC treated with PEM (200 mg intravenously every 3 weeks) after failure of prior chemotherapy [median no. of regimens was 1 (range 1-4)] between 01/2015 and 12/2017. CD3+ and CD8+ TCDs were analyzed in the primary tumor core (CT) and at the invasive margin (IM) by immunohistochemistry and automated image analysis to determine density score (0 to 100) for each T-cell subtype and compartment (Ventana Medical Systems, Inc.). Patients were categorized as 1) responders [CR (complete response) + PR (partial response)] vs. non responders [SD (stable disease) + PD (progressive disease)] per RECIST version 1.1, and 2) by duration of response (< or > 12 months). Results: Median follow-up post PEM was 19.5 (9-41) months. Responders included 2 CR and 5 PR; non-responders included 4 SD and 1 PD. The ORR and median time to response were 58.3% (7/12) and 12 weeks (range 9-40), respectively. CD3+ and CD8+ TCD scores were higher in responders vs. in non-responders as well as in patients who had disease control for > 12 months; differences were greatest for CD8+ CT (Table). Conclusions: Among patients treated with PEM, data suggest higher intratumoral CD3+ and CD8+ TCDs in responders versus non-responders and in those with a longer duration of disease control. If confirmed, TCDs may potentially predict responsiveness to PD-1 blockade in dMMR mCRCs. [Table: see text]

3520 Background: Duration of adjuvant FOLFOX or CAPOX for stage III CC is being guided by pt stratification into low (T1-3N1) and high (T4 or N2) risk groups based on the IDEA study. We determined the relative contributions of clinical and molecular features for prediction of time-to-recurrence (TTR) and survival after recurrence (SAR) within each risk group. Methods: Stage III CC (N=5,430) from 2 trials of adjuvant FOLFOX ± cetuximab with similar outcome by study arm [NCCTG N0147 (Alliance), PETACC-8] were used. Tumors were analyzed for mismatch repair (dMMR vs pMMR), mutations in KRAS (exon 2) and BRAFV600E. Median pt follow-up was 83.4 months. Relative contributions to predicting outcome were assessed using χ2 (Harrell’s rms) based on multivariable (MV) Cox models. Results: N (50.8%) and T (31.8%) stage were the top two contributors to prediction of TTR which supports risk grouping. High risk (n=2566) vs low risk (n=2774) pts had poorer TTR (HR 2.7, 95% CI, 2.4-3.0) and SAR [HR 1.6 (1.4-1.9)], both p<.0001. TTR: KRAS contributed the most to predicting TTR among high (58.6%) and low (51.1%) risk pts (Table). Contribution of MMR (16%) to predicting TTR was limited to low risk pts. Contribution of BRAFV600E to TTR was nearly 3-fold increased in high vs low risk pts. SAR: BRAFV600E contributed the most to predicting SAR, especially in high vs low risk pts (2-fold increase). Tumor sidedness and performance status (PS) were key contributors to SAR, but not TTR. MV associations: TTR: low risk, KRAS [HR 1.7 (1.4-2.3], MMR [HR 0.55 (.35-.87), gender (M/F) [HR 1.3 (1.0-1.5)], all p<.04]; high risk: BRAF [HR 1.3 (1.1-1.7)], sidedness (R vs L) [HR 1.14 (1.0-1.3)], KRAS [HR 1.4 (1.2-1.6)], all p<.04]. SAR: BRAF, sidedness, PS (all p<.05). Conclusions: KRAS mutation was the strongest predictor of shorter TTR in both risk groups whereas BRAFV600E was the primary driver of SAR, especially in high risk pts. Support: U10CA180821, U10CA180882, U24CA196171; BMS, Pfizer, Sanofi. NCT00079274.[Table: see text]