Fig. 4. Persistent mutations are retained during cancer evolution under selective pressure of ICB.

a, The presence of loss-prone, multi-copy and only-copy mutations identified in pre-ICB treatment NSCLC tumors was evaluated post-ICB therapy in tumors from eight patients with NSCLC and durable clinical benefit from ICB. Serial tumor samples were acquired with a minimum time difference of 6 months between biopsies and were analyzed by means of whole exome sequencing. A marked difference in the frequency of loss between clonal loss-prone and persistent mutation sets was observed, with an odds ratio of 61.46 (P < 2.2 × 10⁻¹⁶). Across 16 serially biopsied tumors from 8 patients, a total of 363 out of 2,836 clonal mutations that were detected in the baseline tumors were lost in the descendent tumors; of these 358 (98.6%) were clonal loss-prone mutations. In six patients analyzed, we did not identify any clonal persistent mutation that was lost in the descendent tumor. In two patients, we detected four clonal multi-copy mutations that were not detected in the descendent tumors, resulting in an extremely low rate of loss in this mutation category (clonal multi-copy mutations: 4 out of 1,031 lost, 0.4% loss frequency; clonal only-copy mutations: 1 out of 117 lost, 0.9% loss frequency). b, While the loss frequency was higher for subclonal loss-prone compared to persistent mutations, this did not reach statistical significance (odds ratio = 1.24, P = 0.44). Notably, the loss frequency for subclonal multi-copy mutations was 9.3% compared with 14.8% for subclonal loss-prone mutations, potentially indicating differential selection pressures for these alterations.