Background: Unresectable cholangiocarcinoma (CCA) remains a cancer of dismal prognosis as these tumors tend to be highly chemotherapy-refractory, providing patients (pts) a median survival of only around nine months. We herein provide an interim update to this phase II, single-arm, open-label study, where we investigate the safety and efficacy of olaparib plus pembrolizumab in pts with CCA after first-line gemcitabine-based therapy. We hypothesize that olaparib will increase tumoral response to pembrolizumab by inducing DNA damage and increasing tumor antigen number to produce a durable immune response against CCA - thereby increasing the overall response rate (ORR) of pts from 17.5% (historic chemotherapy control) to 35%. Methods: In total, 36 histologically confirmed advanced cholangiocarcinoma pts will be enrolled onto this trial. Pts receive pembrolizumab, 200 mg, on day 1 of each 21-day cycle, and oral olaparib, 300 mg, twice daily. The primary objective is to assess ORR, while a secondary aim is to evaluate safety. Tumor biopsies are collected at baseline, the third week after treatment initiation, and disease progression; the tissue is tested for ERCC1, PD-1/PD-L1 expression, IDH1/2 mutation status, and immune cell (CD3 and CD8) response. CT or MRI is carried out every 6 weeks for the first 6 months and every 9 weeks for the next 6 months to assess clinical response. Results: At the cutoff date of September 26, 2021, 12 pts had been enrolled and received at least one dose of olaparib plus pembrolizumab. The median age is 62 years (range 47-74); most are female (N = 10) and Caucasian (N = 8). Ten pts were diagnosed with cholangiocarcinoma and 2 with gallbladder carcinoma. According to RECIST v1.1, partial response (PR) was seen in one pt (-50%), stable disease (SD) in 4 pts (one with promising ongoing response at 23% tumor shrinkage), and disease progression in 7 pts. Three pts are actively on trial (1 PR and 2 SD). Complete molecular analysis by FoundationOne CDx revealed mutations in ATM (N = 2), SKT11 (N = 1), ARID1A (N = 2), and IDH1 (N = 2); the full tumor profile will be presented at the ASCO-GI meeting. Primary grade 3 treatment-related adverse events (AEs) included anemia (N = 2) and diarrhea (N = 1. Grade 1/2 AEs included thrombocytopenia (N = 2), anemia (N = 1), rash (N = 1), and diarrhea (N = 1). Conclusions: Interim trial results indicate that combination olaparib and pembrolizumab has acceptable safety and manageable toxicity in pts with advanced cholangiocarcinoma. Clinical trial information: NCT04306367.