CCL20 Released by Drug–Tolerant Persisters Impairs Immunotherapy in EGFR– Mutant Lung Adenocarcinoma

ABSTRACT

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.