In the phase I KEYNOTE-001 study, pembrolizumab demonstrated durable antitumor activity in patients with advanced non-small-cell lung cancer (NSCLC). We sought to characterize the relationship between pembrolizumab dose, exposure, and response to define an effective dose for these patients. Patients received pembrolizumab 2 mg/kg every 3 weeks (Q3W) (n = 55), 10 mg/kg Q3W (n = 238), or 10 mg/kg Q2W (n = 156). Response (RECIST v1.1) was assessed every 9 weeks. The relationship between the estimated pembrolizumab area under the concentration–time curve at steady state over 6 weeks (AUCss–6weeks) and the longitudinal change in tumor size (sum of longest diameters) was analyzed by regression and non-linear mixed effects modeling. This model was simultaneously fit to all tumor size data, then used to simulate response rates, normalizing the trial data across dose for prognostic covariates (tumor PD-L1 expression and EGFR mutation status). The exposure–safety relationship was assessed by logistic regression of pembrolizumab AUCss–6weeks versus occurrence of adverse events (AEs) of interest based on their immune etiology. Overall response rates were 15% 95% confidence interval (CI) 7%–28% at 2 mg/kg Q3W, 25% (18%–33%) at 10 mg/kg Q3W, and 21% (95% CI 14%–30%) at 10 mg/kg Q2W. Regression analyses of percentage change from baseline in tumor size versus AUCss–6weeks indicated a flat relationship (regression slope P > 0.05). Simulations showed the exposure–response relationship to be similarly flat, thus indicating that the lowest evaluated dose of 2 mg/kg Q3W to likely be at or near the efficacy plateau. Exposure–safety analysis showed the AE incidence to be similar among the clinically tested doses. No significant exposure dependency on efficacy or safety was identified for pembrolizumab across doses of 2–10 mg/kg. These results support the use of a 2 mg/kg Q3W dosage in patients with previously treated, advanced NSCLC. NCT01295827.