Amplification of Extrachromosomal MYC Paralogs Shapes Immunosuppressive Tumor Microenvironment in Small Cell Lung Cancer

PURPOSE: Immunotherapy has demonstrated promise in small cell lung cancer (SCLC), but certain patients encounter limited benefits, highlighting the need for immunosuppressive biomarkers. Extrachromosomal circular DNA (ecDNA) promotes amplification of MYC paralogs (MYC, MYCN, and MYCL), driving cross-resistance in SCLC. In this study, we aim to investigate whether ecDNA-mediated MYC paralog amplification (ecMYC+) represents immunosuppressive features in SCLC. EXPERIMENTAL DESIGN: Bulk RNA sequencing data were retrieved from public database and paraffin-embedded samples. The overexpression and amplification of MYC paralogs were identified using IHC and FISH. Imaging mass cytometry and multiplex IHC were used to characterize spatial distribution of the tumor immune microenvironment. The copy number of MYC paralogs was investigated using qRT-PCR. RNA sequencing and flow cytometry were performed in SCLC cell lines. RESULTS: The mean copy number of ecDNAs and the frequency of ecMYC+ cell lines were higher in SCLC than those in the other lineages (SCLC 22/47 vs. others 15/282). In ecMYC+ SCLC, multiple immune-related pathways were downregulated whereas nucleotide metabolism processes were upregulated. Inhibition of nucleotide metabolism induced ecDNA elimination, along with activated antigen presenting pathways. Highly dispersed MYC paralog amplifications were detected in resected treatment-naïve SCLC samples. Through the resolution of 103,341 cells from 24 pathologic regions, we observed higher expression of Ki67, VEGFA, FAP, and FOXP3 and reduced T-cell infiltration in ecMYC+ samples. Moreover, ecMYC+ samples exhibited elevated cellular neighborhoods dominated by Ki67+ tumors, with reduced spatial interaction with immune cells. CONCLUSIONS: Extrachromosomal amplification of MYC paralogs shapes the suppressive tumor immune microenvironment, identifying potential subgroups of immunotherapy-resistant patients.