Höchster, Howard S.

224 Background: Metabolic pathways are reprogrammed during CRC development, leading to increased glycolysis, glutaminolysis, and fatty acid synthesis. Understanding the impact of metabolic reprogramming on treatment outcomes is paramount. Hence, we investigated whether the tumor gene expression of 4 selected major metabolic genes ( PKM, SLC2A1 , SLC16A1 , and CAV1 ) could affect treatment response in pts enrolled in the CALGB/SWOG 80405 trial. Methods: 433 mCRC pts treated with either bevacizumab (bev, n = 226) or cetuximab (cet, n = 207) in combination with first-line chemotherapy were analyzed. RNA was isolated from FFPE tumor samples and sequenced on the HiSeq 2500 (Illumina). Overall survival (OS) and progression-free survival (PFS) were compared between groups of pts categorized by tertiles of gene expression (high [H], medium [M] and low [L]). Logrank P -values provide a non-parametric unadjusted assessment of differences. Likelihood ratio tests (LRT) were computed from multivariate Cox proportional hazards models, adjusting for age, sex, ECOG, tumor side, number of metastatic sites, KRAS , CMS, and treatment. Results: PKM gene expression was associated with cet treatment outcomes with PKM- low tumors showing significantly longer PFS and OS (median PFS: 14.3 vs 9.8 vs 8.1 months, L vs M vs H, P < 0.0001, LRT P = 0.011; median OS: 45.9 vs 31.2 vs 20.9 months, P < 0.0001, LRT P = 0.12). Low SLC2A1 was also associated with longer OS in cet-treated pts (32.4 vs 35.8 vs 25.2 months, P = 0.02, LRT P < 0.0001). Similar results were observed in bev-treated pts where low SLC2A1 expressing tumors had longer OS (37.4 vs 26.1 vs 29 months, P = 0.037, LRT P = 0.069) and a non-significant trend for longer PFS (13.1 vs 11 vs 9.5 months, P = 0.076). CAV1 low gene expression was associated with longer OS in cet-treated pts (37.4 vs 34 vs 21.5 months, P = 0.0048, LRT P = 0.013), while no significant associations were found in bev-treated (interaction LRT P = 0.0009). No significant results were observed for SLC16A1 . Conclusions: The intratumoral expression of metabolic genes was prognostic and predictive in mCRC pts treated with first-line therapy. GLUT1 (encoded by SLC2A1 ) and PKM2 (encoded by PKM ) promote aerobic glycolysis of cancer cells, known as Warburg effect, supporting rapid cell proliferation and survival. CAV1 has been shown to be involved in mitochondrial bioenergetics and fatty acid metabolism and to play either a tumor suppressor or oncogenic role depending on the cancer type and stage. Lower expression of SLC2A1 and PKM was associated with improved survival and increased benefit from cet treatment in our cohort. Low CAV1 expression was also associated with increase survival suggesting a cancer-promoting role in mCRC. Our results suggest that targeting cancer cell metabolism through novel inhibitors of the aforementioned pathways may be a promising therapeutic strategy.

TPS311 Background: Approximately 45% of dMMR/MSI-H metastatic colorectal cancer (mCRC) in the immunotherapy arm progressed at 12 mos (KEYNOTE 177). We hypothesize that dMMR/MSI-H mCRC patients (pts) may be more effectively treated with the combination of PD-1/PD-L1 (PD-1) pathway blockade and mFOLFOX6/bevacizumab (bev) rather than with anti-PD-L1 therapy (atezo) alone. Preclinical work demonstrated synergistic effects between anti-PD-1/anti-VEGF as well as between oxaliplatin/anti-PD-1 in murine CRC models, and phase II data showed activity of anti-PD-1/anti-VEGF in chemotherapy refractory colon cancer. Within the AtezoTRIBE 8-pt dMMR CRC subgroup treated with FOLFOXIRI+bev+atezo, median PFS was not reached, with the first progression event at ~16 mos. Additionally, in other solid tumor malignancies, anti-PD-1 plus anti-VEGFr (i.e., HCC and RCC) as well as anti-PD-1 plus chemotherapy (i.e., gastroesophageal and lung cancers) combinations are standard first-line treatments. Methods: This two-arm prospective phase III open-label trial randomizes (1:1) mCRC dMMR/MSI-H to atezo monotherapy v mFOLFOX6/bev+atezo combination. Key inclusion criteria have been simplified on recent amendments to better mirror clinical practice for pts receiving mFOLFOX6/bev+atezo: One cycle of FOLFOX or CAPOX, with or without bev (or biosimilar) prior to enrollment allowed, dMMR tumor determined by local CLIA-certified IHC assay (MLH1/MSH2/MSH6/PMS2) or MSI-H by local CLIA-certified PCR or NGS panel; pts with total bilirubin ≤4.0 x ULN; duration of therapy for up to two years for both arms; imaging frequency on post-treatment follow-up has been reduced; as has measurable disease per RECIST. Primary endpoint is PFS. Assuming the atezo monotherapy control arm has a 48% PFS at 24 mos as assessed by site investigator, we have 80% power to detect a hazard ratio of 0.6 (equivalent to 64.4% PFS at 24 mos) with alpha 0.025 one-sided. Stratification factors include BRAFV600E status, metastatic site, and prior adjuvant CRC therapy. Secondary endpoints include overall survival, objective response rate, safety profile, disease control rate, and duration of response. Archived tumor tissue and blood samples will be collected for correlative studies. Harmonization of translational analyses is planned between GI004 (COMMIT) and A021502 (ATOMIC). Sample size has been modified with the accrual goal of 120 pts randomized between the two immunotherapy arms needed for study completion. Enrollment actively continues at U.S. sites. Current accrual (as of 9-20-2024): 100/120. Clinical trial information: NCT02997228 .

142 Background: Fruquintinib (F) is a highly selective, oral tyrosine kinase inhibitor of all 3 vascular endothelial growth factor receptors (VEGFRs -1, -2, -3) that is approved by the US FDA and in the EU, UK, and Japan for previously treated metastatic colorectal cancer (mCRC), regardless of biomarker status. FRESCO-2 (NCT04322539) met its primary endpoint demonstrating significantly improved overall survival (OS) with F + best supportive care (BSC) vs placebo (P) + BSC. This subgroup analysis assessed landmark survival of all FRESCO-2 patients (pts), and baseline (BL) characteristics and safety data of pts treated with F who had OS ≥10 months (mos) compared with the intent-to-treat (ITT) population (pop). Methods: Pts were randomized 2:1 to receive F 5 mg or matching P, by mouth, once daily, 3 weeks on, 1 week off, + BSC. Pts had received prior chemotherapy, anti-VEGF therapy and, if RAS wild type, anti-EGFR therapy; and had prior exposure to trifluridine/tipiracil (TAS-102) and/or regorafenib. The hazard ratio (HR) between the two treatment arms was calculated from a stratified Cox proportional hazard model with treatment group as the only covariate in the model. Results: In the ITT pop, OS was improved with F vs P (HR 0.66); OS rates (95% confidence intervals [CIs]) at 6 and 9 mos were 60.4% (55.9–64.9) vs 41.5% (35.0–48.0) and 41.1% (36.4–45.8) vs 28.2% (22.1–34.3), respectively. Progression-free survival (PFS) rates (95% CIs) at 6 and 9 mos were 23.8% (19.7–28.0) vs 1.1% (0–2.6) and 11.3% (8.1–14.6) vs 0.5% (0–1.6), respectively. Of 461 pts who received F, 113 (24.5%) had OS ≥10 mos (range 10.0–18.9) with a median duration of F treatment of 6.3 mos (range 0.7–19.1). Among pts receiving F, a higher proportion (≥10% difference) of pts with OS ≥10 mos had no liver metastases (mets) and an Eastern Cooperative Oncology Group performance status (ECOG PS) of 0 at BL compared with the ITT pop (Table). In pts with OS ≥10 mos, 61.9% experienced a grade ≥3 treatment-emergent adverse event compared with 62.7% in the ITT pop. Conclusions: Longer-term OS and PFS rates at 6 and 9 mos were higher with F vs P in the FRESCO-2 ITT pop. The data show a higher proportion of pts with OS ≥10 mos had an absence of liver mets and ECOG PS 0 at BL than the ITT pop. The safety profile of F in pts with OS ≥10 mos was consistent with the ITT pop. Clinical trial information: NCT04322539 . BL characteristics of pts treated with F with OS ≥10 mos vs ITT pop. BL characteristic Pts with OS ≥10 mos (n=113) ITT pop(n=461) ECOG PS (0 / 1), % 54.0 / 46.0 42.5 / 57.5 Median time since 1 st CRC diagnosis, mos 52.0 47.2 Primary tumor location at 1 st diagnosis (colon / rectum / both), % 51.3 / 35.4 / 13.3 60.5 / 31.0 / 8.5 Primary site at 1 st diagnosis (colon left / colon right), % 42.5 / 15.9 41.6 / 21.0 Liver mets, % 58.4 73.5 Median duration of mCRC, mos 42.7 37.9 Prior lines of therapy for mCRC (≤3 / >3), % 26.5 / 73.5 27.1 / 72.9 Prior TAS-102, % 46.9 52.1 Prior regorafenib, % 12.4 8.7

PURPOSE SWOG S1815 was a randomized, open label phase III trial, evaluating gemcitabine, nab-paclitaxel, and cisplatin (GAP) versus gemcitabine and cisplatin (GC) in patients with newly diagnosed advanced biliary tract cancers (BTCs). METHODS Patients with newly diagnosed locally advanced unresectable or metastatic BTC, including intrahepatic cholangiocarcinoma (ICC) and extrahepatic cholangiocarcinoma (ECC) and gallbladder carcinoma (GBC), were randomly assigned 2:1 to either GAP (gemcitabine 800 mg/m 2 , cisplatin 25 mg/m 2 , and nab-paclitaxel 100 mg/m 2 intravenously once per day on days 1 and 8 of a 21-day cycle) or GC (gemcitabine 1,000 mg/m 2 and cisplatin 25 mg/m 2 intravenously once per day on days 1 and 8 of a 21-day cycle). RESULTS Among 452 randomly assigned participants, 441 were eligible and analyzable, 67% with ICC, 16% with GBC, and 17% with ECC. There was no significant difference in overall survival (OS) between GAP versus GC. Median OS with GAP was 14.0 months (95% CI, 12.4 to 16.1) and 13.6 months with GC (95% CI, 9.7 to 16.6); hazard ratio (HR), 0.91 (95% CI, 0.72 to 1.14); P = .41. Median progression-free survival (PFS) was similar between groups with median PFS for GAP being 7.5 months (95% CI, 6.4 to 8.5) versus 6.3 months for GC (95% CI, 4.4 to 8.2); HR, 0.89 (95% CI, 0.71 to 1.12); P = .32. In exploratory subset analyses, the OS and PFS benefits of GAP versus GC treatment were greater in locally advanced disease compared with metastatic disease, although not statistically significant (interaction P = .14 for OS and P = .17 for PFS). Moreover, GAP versus GC showed greater improvement in PFS among participants with GBC than those with ICC or ECC (interaction P = .01), but not OS (interaction P = .28). CONCLUSION The addition of a taxane in the GAP regimen to the standard gemcitabine-cisplatin regimen did not improve OS in newly diagnosed BTC. More toxicity was encountered with GAP versus GC.

3579 Background: Fruquintinib (F) is a highly selective, oral tyrosine kinase inhibitor of all 3 VEGF receptors. Based on FRESCO (NCT02314819) and FRESCO-2 (NCT04322539) data, F was approved by the US FDA for previously treatedmetastatic colorectal cancer (mCRC). Efficacy and safety of F + best supportive care (BSC) vs placebo (P) + BSC were analyzed according to prior treatment (tx) sequence using FRESCO and FRESCO-2 data. Methods: In FRESCO and FRESCO-2, patients (pts) were randomized 2:1 to receive F 5 mg or matching P, PO daily x 21 days every 28 days. In FRESCO, eligible pts had received ≥2 prior chemotherapy regimens and were permitted to have received prior anti-VEGF and/or anti-EGFR therapies, whereas in FRESCO-2, in addition to standard chemotherapies, anti-VEGF, and anti-EGFR therapies if indicated, prior exposure to trifluridine/tipiracil (TAS) and/or regorafenib (rego) was required. FRESCO-2 pts must have also received a BRAF inhibitor and/or immunotherapy, if indicated and available in their country. Results: In FRESCO (F n=278 vs P n=138), no pts had received rego and 16/416 pts (4%) had received TAS. InFRESCO-2, prior therapies were balanced between F vs P arms; pts had received prior TAS or rego in these sequences: prior TAS only (n=240 vs n=121); prior rego only (n=40 vs n=18); prior rego then TAS (n=84 vs n=45); prior TAS then rego (n=95 vs n=44). Improvements in OS and PFS were seen with F vs P regardless of prior tx sequence (Table), with similar magnitudes of benefit for OS (HRs 0.53–0.77) and PFS (HRs 0.25–0.37). F safety profile was consistent regardless of prior tx sequence (Table). Conclusions: F is effective in mCRC with a manageable safety profile regardless of prior TAS or rego tx and sequence. Improvement with F was similar for OS and PFS despite the number and sequence of prior regimens. F provides a new tx option for pts with previously treated mCRC regardless of prior TAS or rego exposure. Clinical trial information: NCT04322539 . [Table: see text]

TPS3632 Background: Approximately 45% of dMMR/MSI-H mCRC in the immunotherapy arm progressed at 12 mos (KEYNOTE 177). We hypothesize that dMMR/MSI-H mCRC patients (pts) may be more effectively treated with the combination of PD-1/PD-L1 (PD-1) pathway blockade and mFOLFOX6/bevacizumab (bev) rather than with anti-PD-L1 therapy (atezo) alone. Preclinical work demonstrated synergistic effects between anti-PD-1/anti-VEGF as well as between oxaliplatin/anti-PD-1 in murine CRC models, and phase II data showed activity of anti-PD-1/anti-VEGF in chemotherapy refractory colon cancer. Upon AtezoTRIBE subgroup analysis of the 8 pts with dMMR mCRC treated with FOLFOXIRI+bev+atezo, median PFS was not reached, with the first pt having progression ~16 mos. Additionally, in other solid tumor malignancies, anti-PD-1 plus anti-VEGFr (i.e., HCC and RCC) as well as anti-PD-1 plus chemotherapy (i.e., gastroesophageal and lung cancers) combinations are standard first-line treatments. Methods: This two-arm prospective phase III open-label trial randomizes (1:1) mCRC dMMR/MSI-H to atezo monotherapy v mFOLFOX6/bev+atezo combination. Key inclusion criteria have been simplified on recent amendments to better mirror clinical practice for pts receiving mFOLFOX6/bev+atezo: One cycle of FOLFOX or CAPOX, with or without bev (or biosimilar) prior to enrollment allowed, dMMR tumor determined by local CLIA-certified IHC assay (MLH1/MSH2/MSH6/PMS2) or MSI-H by local CLIA-certified PCR or NGS panel; pts with total bilirubin ≤4.0 x ULN; duration of therapy for up to two years for both arms; imaging frequency on post-treatment follow-up have been reduced; as has measurable disease per RECIST. Primary endpoint is PFS. Assuming the atezo monotherapy control arm has a 48% PFS at 24 mos as assessed by site investigator, we have 80% power to detect a hazard ratio of 0.6 (equivalent to 64.4% PFS at 24 mos) with alpha 0.025 one-sided. Stratification factors include BRAFV600E status, metastatic site, and prior adjuvant CRC therapy. Secondary endpoints include overall survival, objective response rate, safety profile, disease control rate, and duration of response. Archived tumor tissue and blood samples will be collected for correlative studies. Harmonization of translational analyses is planned between GI004 (COMMIT) and A021502 (ATOMIC). Sample size has been modified with the accrual goal of 120 pts randomized between the two immunotherapy arms needed for study completion. Enrollment actively continues. Current accrual (as of 02-02-2024): 96/120. NCT02997228. Support: U10CA180868, -180822, -180888, UG1CA189867, U24CA196067; Genentech, Inc. Clinical trial information: NCT02997228 .

3144 Background: Longitudinal brain structure changes across the lifespan are associated with neurodegenerative disorders. We previously showed that genetic variants in dopamine and Parkinson’s disease genes are predictive and prognostic in mCRC. Indeed, the brain-gut axis is emerging as a critical player in CRC biology. Hence, we investigated whether the tumor gene expression of 3 metabolic genes involved in brain development and aging could affect treatment response in patients enrolled in the CALGB/SWOG 80405 trial. Methods: 433 mCRC pts treated with either bevacizumab (bev, n = 226) or cetuximab (cet, n = 207) in combination with first-line chemotherapy were included in the analysis. RNA was isolated from FFPE tumor samples and sequenced on the HiSeq 2500 (Illumina). Overall survival (OS) and progression-free survival (PFS) were compared between groups of pts categorized by tertiles of expression of 3 genes: APOE, DACH1, and GPR139. Likelihood ratio tests, hazard ratios and 95% confidence intervals were computed from multivariate Cox proportional hazards models, adjusting for age, sex, ethnicity, ECOG PS, tumor location, number of metastatic sites, KRAS, MSI status, and treatment. Logrank P-values describe differences without adjustment for patient characteristics. Results: Both APOE and DACH1 (but not GPR139) met the 0.05 false discovery rate threshold for association with both PFS ( P = .00085, P = .015) and OS ( P = .021, P = .021, respectively) in the full Cox models. In cet-treated pts, APOE-high (H) showed significantly shorter PFS (median 9.2 vs 11.8 vs 13.0 months (mo), high vs low (as reference) logrank P = .0021) and OS (median 21.9 vs 35.8 vs 34.6 mo; logrank P = .0017) compared to APOE-median and -low, respectively. Opposite results were observed for DACH1, where DACH1-H tumors had longer PFS (median 14.5 vs 10.6 vs 8.07 mo; logrank P = .0011) and OS (median 39.4 vs 30.9 vs 21.5 mo; logrank P = .0021) when treated with cet. In bev-treated pts, similar results were observed with APOE-H showing shorter OS (median 25.8 vs 26.3 vs 37.5 mo; logrank P = .0016) and DACH1-H longer OS (median 36.5 vs 28.3 vs 22.4 mo; logrank P = .0017). No significant gene-treatment interaction was observed for either gene for PFS and OS. Conclusions: The tumor expression of metabolic genes associated with changes in the brain structure was prognostic in mCRC pts treated with first-line therapy. Notably, APOE and DACH1 have been recently reported to play a role in CRC where APOE overexpression has been associated with aggressive biological behavior whereas DACH1 has been shown to suppress CRC cell growth and invasion. Our results support this evidence and suggest that further investigation into the role of APOE and DACH1 in CRC biology and into their potential as targets for drug development is warranted. Trial Identifier: NCT00265850.

3523 Background: Evidence suggests neurotransmitter signaling may play a crucial role in tumorigenesis of gastrointestinal (GI) cancers, including migration, proliferation, and apoptosis, as well as angiogenesis and the microenvironment. We developed and applied a strategy to identify upstream regulators of neurotransmitters and their associations with patient outcomes. Methods: Prior literature yielded 85 neurotransmitter signaling genes mapped to RNA-seq gene expression in tumor tissue from 545 patients with mCRC enrolled in the phase III CALGB/SWOG 80405 trial. These samples were also sequenced by FoundationOne CDx for DNA damage. Using Cox proportional hazards regression, we identified a subset of CDx genes associated with overall survival (OS) or progression-free survival (PFS). We extended the Causal Inference Test (CIT) for time-to-event data and identified a subset of the 85 signaling genes consistent with mediation of DNA mutation effects on OS and PFS. Another CIT analysis identified potential upstream neurotransmitter regulators, gene expression mediators of mutation effects on the signaling genes. Top genes were then assessed in whole-transcriptome sequencing (WTS) expression data with Caris MI profile collected from 398 patients with advanced CRC enrolled in the MONSTAR-SCREEN-2 trial. Results: At the 0.05 alpha level, we identified 20 signaling genes with gene expression associated with PFS and/or OS, and 68 genes with evidence of DNA mutations associated with PFS and/or OS. False discovery rate (FDR) < 0.05 was applied to filter the 56,674 expression features on PFS and OS associations, yielding 173 unique genes as possible mediators. The CIT yielded 53 putative mediators at FDR < 0.05, suggesting upstream regulation of neurotransmitter signaling. Five genes were identified as targeting at least 8 signaling genes each, PIGU, ANO9, LY6G6D, and GPR160, and FSCN1, where the first 4 are favorable and only FSCN1 is a poorer prognostic. Following up on these 5 genes in MONSTAR-SCREEN-2 by dichotomizing at the median value of WTS expression, 3 were nominally associated with OS, PIGU ( P = 0.050; HR, 0.59; 95%CI, 0.34-1.00), ANO9( P = 0.029; HR, 0.55; 95%CI, 0.32-0.95), LY6G6D( P = 0.0024; HR, 0.44; 95%CI, 0.25-0.76), and FSCN1 was suggestive ( P = 0.079; HR, 1.62; 95%CI, 0.94-2.77), with direction of effects in agreement with 80405 results for all four. Conclusions: Our novel approach identified previously unknown upstream regulators of neurotransmitter signaling associated with treatment outcomes in mCRC. PIGU, FSCN1, and LY6G6D have been suggested in prior literature as targets for CRC and ANO9 has been suggested as a target for GI cancer. These results bolster the strategy of using mediation analysis with large-scale molecular data to identify new therapeutic targets in CRC. NCT00265850.

PURPOSE The phase III Cancer and Leukemia Group B (CALGB)/SWOG 80405 trial found no difference in overall survival (OS) in patients with metastatic colorectal cancer receiving first-line chemotherapy in combination with either bevacizumab or cetuximab. We investigated the potential prognostic and predictive value of HER2 amplification and gene expression using next-generation sequencing (NGS) and NanoString data. PATIENTS AND METHODS Primary tumor DNA from 559 patients was profiled for HER2 amplification by NGS (FoundationOne CDx). Tumor tissue from 925 patients was tested for NanoString gene expression using an 800-gene panel. OS and progression-free survival (PFS) were the time-to-event end points. RESULTS High HER2 expression (dichotomized at median) was associated with longer PFS (11.6 v 10 months, P = .012) and OS (32 v 25.3 months, P = .033), independent of treatment. An OS benefit for cetuximab versus bevacizumab was observed in the high HER2 expression group ( P = .02), whereas a worse PFS for cetuximab was seen in the low-expression group ( P = .019). When modeled as a continuous variable, increased HER2 expression was associated with longer OS (hazard ratio [HR], 0.83 [95% CI, 0.75 to 0.93]; adjusted P = .0007) and PFS (HR, 0.82 [95% CI, 0.74 to 0.91]; adjusted P = .0002), reaching a plateau effect after the median. In patients with HER2 expression lower than median, treatment with cetuximab was associated with worse PFS (HR, 1.38 [95% CI, 1.12 to 1.71]; adjusted P = .0027) and OS (HR, 1.28 [95% CI, 1.02 to 1.59]; adjusted P = .03) compared with that with bevacizumab. A significant interaction between HER2 expression and the treatment arm was observed for OS ( P intx = .017), PFS ( P intx = .048), and objective response rate ( P intx = .001). CONCLUSION HER2 gene expression was prognostic and predictive in CALGB/SWOG 80405. HER2 tumor expression may inform treatment selection for patients with low HER2 favoring bevacizumab- versus cetuximab-based therapies.