Caughey, Bennett Adam

PURPOSE This study aimed to investigate the clinicomolecular profiles and the efficacy of human epidermal growth factor receptor 2 (HER2)-targeted therapy in HER2 -amplified biliary tract cancer (BTC). METHODS This study was an international collaboration that used combined data from the prospective SCRUM-Japan GOZILA and MONSTAR-SCREEN in Japan and retrospective reviews in the United States; patients with advanced BTC who had received systemic therapy were included. The clinicomolecular profiles were evaluated in an exploratory cohort, whereas the efficacy of HER2-targeted therapy was assessed in a biomarker-selected cohort. RESULTS Of the 439 patients in the exploratory cohort, 43 (10%) had HER2 amplification. The frequencies of coalterations were higher in patients with HER2 amplification versus patients without HER2 amplification including HER2 mutations (26% v 5%, P < .001), TP53 mutations (84% v 61%, P = .003), and BRAF amplification (9% v 2%, P = .030). There were no KRAS mutations identified in patients with HER2- amplified BTC. No significant difference in overall survival (OS) was observed between patients with and without HER2 amplification (median, 17.7 v 16.9 months; hazard ratio [HR], 0.95 [95% CI, 0.65 to 1.40]). Of the 60 patients with HER2 -amplified BTC in the biomarker-selected cohort (43 from Japan and 17 from the United States), the OS was significantly longer in 29 patients who received HER2-targeted therapy than in those who did not receive HER2-targeted therapy (median, 24.3 v 12.1 months; HR, 0.39 [95% CI, 0.23 to 0.82]). Multivariate analysis identified HER2-targeted therapy as an independent prognostic factor for OS (HR, 0.29 [95% CI, 0.14 to 0.58]; P < .001). CONCLUSION HER2 amplification was found in 10% of advanced BTC and was not identified as an independent prognostic factor for OS. Patients with HER2 -amplified BTC derive significant benefit from HER2-targeted therapy.

544 Background: Biliary tract cancer (BTC) is an aggressive malignancy with poor prognosis and limited treatment options. The prevalence of HER2 amplification in BTC has been reported to be 5-20%, and clinical trials suggested the clinical benefit of HER2-targeted treatment in this disease subpopulation. In a global collaboration, we investigated the comprehensive clinico-molecular characterization as well as the efficacy of anti-HER2 therapy in HER2-amplified BTC. Methods: Patients with advanced BTC who received systemic therapy were included from the GOZILA and MONSTAR studies in Japan and from the COLOMATE trial in the US. Genomic alterations were detected by tissue next-generation sequencing (NGS), including FoundationOne CDx and TempuslxT, and plasma circulating tumor DNA NGS including Guardant360 and FoundationOne Liquid CDx. The clinico-molecular characteristics were evaluated in an exploratory cohort comprised of patients from Japan, whereas the efficacy of anti-HER2 therapy was assessed in a biomarker selected cohort with patients from both Japan and the US. Results: Of 439 patients included in the exploratory cohort, 43 (10%) had HER2 amplification. BTC with primary tumor location in the gallbladder accounted for 58% of primary sites of patients with HER2 amplification. TP53 mutation (84% vs 61%, p=0.003) and BRAF amplification (9% vs 2%, p=0.030) were significantly more frequent in patients with HER2 amplification compared to those without, and no KRAS mutation co-occurred with HER2 amplification. There was no significant difference in overall survival (OS) between patients with and without HER2 amplification (17.7 months [M] vs 16.9 M, hazard ratio [HR] 0.95, 95% confidence interval [CI] 0.65-1.40, p=0.799). Of 58 patients with HER2-amplified BTC included in the biomarker selected cohort (43 Japan, 15 US), 27 (47%) received anti-HER2 therapy. The median OS of patients treated with anti-HER2 therapy was significantly longer than that of patients who did not receive anti-HER2 therapy (24.3 M vs. 12.1 M, HR 0.43, 95% CI 0.23-0.82, p=0.011). Multivariate analysis also identified anti-HER2 therapy as an independent prognostic factor for OS (HR 0.42, 95% CI 0.20-0.85, p=0.015). Conclusions: HER2 amplifications were found in 10% of advanced BTC and did not represent an independent predictive factor for OS. Of clinical significance, patients with HER2-amplified BTC derive a significant benefit from anti-HER2 therapy.

3531 Background: Homologous recombination deficient (HRD) colorectal cancer (CRC) has improved overall survival (OS) when exposed to DNA damaging agents (DDA) oxaliplatin (OX) and irinotecan (IR). However, this OS benefit is only observed in HRD mutated CRC. Low expression of wild type (WT) BRCA1 RNA displays prolonged OS in ovarian cancer; this finding has not been investigated in CRC or outside BRCA. We investigate if low expression of WT HR genes is associated with prolonged OS in response to DDA. Methods: A total of 12,860 CRC patients tumor biopsies were analyzed by next-generation sequencing (592, NextSeq; WES, WTS NovaSeq). Patients with HRD mutations and MSI-H were excluded (N = 935). 11,925 patients were included in the analysis. A total of 11 core ( BARD1, BLM, BRCA1, BRCA2, BRIP1, MRE11, NBN, PALB2, RAD50, RAD51, RAD51B) and 7 related ( BAP1, WRN, DNMT3A, ERCC1, FANCA, FANCF, RECQL4) HR genes were analyzed. Samples were classified by RNA expression percentiles within the studied cohort. Real world OS was extracted from insurance claims and calculated using Kaplan-Meier estimates for molecularly defined cohorts from first of OX or IR to last contact. Results: OS benefit is seen in low RNA expression (bottom 10%) compared to high expression (top 10%) of all core WT HR genes except RAD51B, BARD1 and BRCA2 in response to IR. We observe a progressively longer OS with lower associated RNA expression in response to DDA. OS of nearly 7 years was observed in low expressing BRCA1 and RAD51 following IR exposure and 4.5-year OS benefit when compared to high expression of these genes. OS benefit in response to OX was less robust but significantly prolonged in BRCA1 and BRIP1 (Table). Patients harboring HRD mutations had similar OS to WT HR RNA low (bottom 10%) in response to OX (HR = 0.75; p = 0.1) and IR (HR = 0.82; p = 0.36). Conclusions: Here we report a novel subclass of CRC defined as patients with low RNA expressing WT HR genes that exhibit differential sensitivity to DDA. Significantly longer survival is noted in CRC with low expression BRCA1, RAD51 and BLM, while post-OX survival was significantly prolonged with low expression of BRCA1, BRIP1 and FANCA. Further characterization of sensitive HR genes will better predict DDA sensitivity and impact treatment sequencing. [Table: see text]

4086 Background: Abnormalities in the MAPK pathway are potential therapeutic targets in various cancers. However, the clinical impact of alterations in the MAPK pathway in BTC have not been elucidated, especially outside of canonical mutations in KRAS and BRAF. We investigated the clinical outcomes of advanced BTC with MAPK pathway alterations treated with chemotherapy in Japan and the United States. Methods: Patients with advanced BTC who received gemcitabine plus cisplatin as first-line therapy were included from the GOZILA study in Japan and Duke Molecular Registry of Tumors in the US. Genetic abnormalities were detected by Guardant360, a cell-free DNA assay, in Japan and by blood or tissue-based next-generation sequencing (NGS) at Duke University. Two hundred and seven patients with BTC from Japan were included in an exploratory cohort to evaluate the association of MAPK alterations with overall survival (OS) according to MAPK alteration status. One hundred and ten patients with BTC from both Japan and the US harboring oncogenic alterations in the MAPK pathway were included in a biomarker selected cohort to assess the association of specific MAPK alterations with OS. Multivariate analysis was performed using a Cox regression model based on a univariate p-value < 0.2. Results: MAPK pathway-related oncogenic alterations detected in each cohort are shown in the table below. In the exploratory cohort, median OS was shorter for patients with MAPK alterations vs. no MAPK alteration (15.9 m vs. 24.9 m, log-rank p = 0.001). Based on univariate analysis, the following covariates were selected for multivariate analyses: age, prior resection, and MAPK pathway alteration in the exploratory cohort; country, the timing of NGS, distant metastasis, KRAS amplification, and BRAF class 2 mutation in the biomarker selected cohort. In the exploratory cohort, multivariate analysis identified MAPK pathway alterations as an independent predictor of shorter OS with a HR of 1.92 (95% CI = 1.28-2.87, p = 0.001). In the biomarker selected cohort, multivariate analysis identified BRAF class 2 mutations and KRAS amplification as independent predictors of shorter OS with a HR of 16.2 (95% CI = 3.26-80.8, p = 0.001) and 5.97 (95% CI = 1.74-19.3, p = 0.002) respectively. Conclusions: MAPK pathway alterations, especially BRAF class 2 mutation and KRAS amplification, had a significant negative impact on clinical outcomes in BTC receiving first-line chemotherapy. These results are newly confirmed in BTC. [Table: see text]

3571 Background: In colorectal cancer (CRC), mutations in KRAS, NRAS, and BRAF predict resistance to anti-EGFR therapies. In pancreatic ductal adenocarcinoma (PDAC), ~90% of patients harbor KRAS mutations, while KRAS wild-type tumors often have clinically actionable fusion alterations. Thus, in both cancers, the accurate ascertainment of RAS and BRAF driver status is essential. Sequencing of cell-free DNA (cfDNA) from plasma allows convenient assessment of a tumor’s molecular profile, but sensitivity can be limited by low ctDNA shedding. We sought to establish a ctDNA shedding threshold at which actionable driver mutations can be reliably detected. Methods: Molecular reports and matched clinical data were obtained from the Duke Molecular Registry of Tumors and the SCRUM-Japan GOZILA and GI-SCREEN. CRC or PDAC patients with a pathogenic KRAS, NRAS, or BRAF activating point mutation (“driver”) present on tissue next-generation sequencing (NGS) assays and who also had cfDNA assay available were included. Tissue NGS included Foundation One CDx and Oncomine Comprehensive Assay. Guardant 360 (G360) was the sole plasma cfDNA assay. 131 CRC and 24 PDAC cases with 189 total G360 assays met criteria and were included. Samples were analyzed according to detection of the driver mutation and the maximum mutant allele frequency (MAF) of non-driver mutations on G360. An optimal cut-point for max MAF was explored among the CRC and PDAC patients using a maximally selected Wilcoxon rank statistic method. Results: 76.8% of driver mutations were in KRAS, 22.6% in BRAF, and 1.9% in NRAS with an overlap of 1 BRAF and 1 NRAS mutation with a KRAS mutation. Overall sensitivity of G360 for drivers was 83.0% for CRC and 54.2% for PDAC. No variants were detected on G360 in 9.1% of CRC and 37.5% of PDAC. Sensitivity for driver mutations increased with higher maximum non-driver MAF, with MAF > 1% predicting sensitivity > 98% (Table). Optimal cut-point analysis identified MAF of > 0.34% (p < 0.0001), above which the driver was identified in 97% of patients and below which only 27%. Conclusions: In our study, non-driver MAF > 1% on cfDNA NGS predicts high sensitivity for RAS and BRAF mutations and thus is adequate to guide clinical decisions such as anti-EGFR therapy in CRC, evaluation for fusions in PDAC, and validity in clinical trials. MAF ≤0.34% is a clear threshold to consider an assay inadequate and thus seek alternative testing. Sensitivity rises for MAF between 0.35 and 1% but will require greater patient numbers to establish clinically relevant sensitivity thresholds. These results will be updated with additional data for final presentation. [Table: see text]