Background Colorectal cancers (CRCs) expressing programmed death ligand 1 (PD-L1) have poor prognosis. In the multicohort KEYNOTE-028 trial, the anti–PD-1 antibody pembrolizumab was evaluated in 20 PD-L1–positive advanced solid tumors. Herein, we report results for the advanced CRC cohort. Methods Patients with advanced, treatment-resistant PD-L1–positive carcinoma of the colon or rectum were enrolled, regardless of microsatellite instability (MSI) status. Pembrolizumab 10 mg/kg was administered every 2 weeks for up to 2 years or until disease progression/unacceptable toxicity. Response was assessed every 8 weeks for the first 6 months and every 12 weeks thereafter. Primary end points were safety and overall response rate by investigator review per Response Evaluation Criteria in Solid Tumors version 1.1. Data cutoff was June 20, 2016. Results Of 137 patients with CRC and samples evaluable for PD-L1 expression, 33 (24%) had PD-L1–positive tumors, of which 23 were enrolled. Median follow-up was 5.3 months, and 8 patients (35%) reported treatment-related adverse events (AEs), most commonly fatigue (n = 3, 13%), stomatitis (n = 2, 9%), and asthenia (n = 2, 9%). One patient (4%) experienced grade 4 treatment-related increased blood bilirubin. No grade 3 AEs, discontinuations, or deaths were attributed to treatment. Most patients (n = 15, 65%) experienced progressive disease. One partial response occurred in a patient (4%) with MSI-high CRC. Conclusion Pembrolizumab demonstrated a favorable safety profile in advanced PD-L1–positive CRC. Antitumor activity was observed in a single patient with MSI-high CRC, warranting further evaluation in this patient population.