EGFR-mutant lung adenocarcinoma (LUAD) is typically associated with an immunosuppressive tumor immune microenvironment (TIME) and poor responses to PD-1 blockade. However, the contribution of drug-tolerant persister cells (DTPs) to immunotherapy resistance remains unclear. We hypothesized that DTPs-derived chemokine (C-C motif) ligand 20 (CCL20) promotes immune evasion and impairs PD-1-based immunotherapy in EGFR-mutant. To test this, humanized NSG mice engrafted with EGFR-mutant H1975 cells and human peripheral blood mononuclear cells (n = 6 per group) were administered with anti-PD-1 (200 µg) and/or anti-CCL20 (20 µg) every other day for a total of four doses. The combination reduced mean tumor volume by 65% compared with control treatment (p < 0.01). Single-cell RNA sequencing showed that cotreatment selectively suppressed CCL20 expression in DTP clusters and activated interferon-α and -γ signaling (ISG15, CMPK2). Multiplex immunofluorescence revealed combination treatment increased infiltration of M1-like macrophages, plasmacytoid dendritic cells, and memory B cells, alongside spatial segregation of CD4+ regulatory T cells (p < 0.05). These findings identify DTPs-derived CCL20 as a mediator of immunosuppressive TIME and support combined CCL20/PD-1 blockade as a potential therapeutic strategy for EGFR-mutant lung adenocarcinoma.
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