Genomic and transcriptomic profiling of combined small-cell lung cancer through microdissection: unveiling the transformational pathway of mixed subtype

Abstract

Background: Combined small-cell lung carcinoma (cSCLC) represents a rare subtype of SCLC, the mechanisms governing the evolution of cancer genomes and their impact on the tumor immune microenvironment (TIME) within distinct components of cSCLC remain elusive.

Methods: Here, we conducted whole-exome and RNA sequencing on 32 samples from 16 cSCLC cases.

Results: We found striking similarities between two components of cSCLC-LCC/LCNEC (SCLC combined with large-cell carcinoma/neuroendocrine) in terms of tumor mutation burden (TMB), tumor neoantigen burden (TNB), clonality structure, chromosomal instability (CIN), and low levels of immune cell infiltration. In contrast, the two components of cSCLC-ADC/SCC (SCLC combined with adenocarcinoma/squamous-cell carcinoma) exhibited a high level of tumor heterogeneity. Our investigation revealed that cSCLC originated from a monoclonal source, with two potential transformation modes: from SCLC to SCC (mode 1) and from ADC to SCLC (mode 2). Therefore, cSCLC might represent an intermediate state, potentially evolving into another histological tumor morphology through interactions between tumor and TIME surrounding it. Intriguingly, RB1 inactivation emerged as a factor influencing TIME heterogeneity in cSCLC, possibly through neoantigen depletion.

Conclusions: Together, these findings delved into the clonal origin and TIME heterogeneity of different components in cSCLC, shedding new light on the evolutionary processes underlying this enigmatic subtype.

Keywords: Microdissection; Monoclonal origin; RB1; TP53; Transdifferentiation; Tumor immune microenvironment; cSCLC.

Fig. 1

Genomic Landscape of cSCLC. A Top 10 recurrently mutated genes in cSCLC; Comparison of TMB B and TNB C between ADC/SCC and paired SCLC; Comparison of TMB D and TNB E between LCC and paired SCLC; F Comparison of CIN between SCLC and paired ADC/SCC; G Comparison of CIN between SCLC and paired LCC; H Copy number profile of all cSCLCs

Fig. 2

Transcriptome Profile in cSCLC. A Volcano plot of DEGs between SCLC and ADC/SCC groups; B Volcano plot of DEGs between SCLC and LCC groups; C GSEA analysis based on the pre-ranked gene set by log2FC between AS-SCLC and ADC/SCC groups; D GSEA analysis based on the pre-ranked gene set by log2FC between L-SCLC and LCC/LCNEC groups

Fig. 3

TIME Profile in cSCLC. A Heatmap of immune cell infiltration in cSCLC; B Comparison of stromal score, immune score, microenvironment score, and TIS score between SCLC and ADC/SCC; C Comparison of stromal score, immune score, microenvironment score, and TIS score between SCLC and LCC; D Boxplot showing significant differences in immune cell infiltration between two components of cSCLC

Fig. 4

Phylogenetic Tree in cSCLC. A Density plot of mutations, CCF, and phylogenetic tree in L-SCLC; B Density plot of mutations, CCF, and phylogenetic tree in A/S-SCLC. Each point in the density plot on the left panel represents a mutation, with different colors indicating different positions from the phylogenetic tree in the right panel. The right panel displays the evolutionary tree of one patient, with the trunk clone, the trunk subclone, and the two branches indicated by different colors, respectively

Fig. 5

Mechanism of Immune Evasion in cSCLC. A Percentage of clonal expressed antigens among different levels of immune infiltration; B Copy-number loss ratios between LCC and paired SCLC; C Copy-number loss ratios between ADC/SCC and paired SCLC; D Correlation between copy-number loss ratios and wGII; E Immunoediting score between LCC and paired SCLC; F Immunoediting score between ADC/SCC and paired SCLC; G Immunoediting score of clonal mutations or H subclonal mutations between ADC/SCC and paired SCLC; I Immunoediting score between clonal mutations and subclonal mutations in ADC/SCC, or J AD-SCLC; K Overall overview of immune evasion in cSCLC; L Immunoediting score between patients with wild-type RB1 and those with mutant RB1

Fig. 6

RB1 Mechanism in cSCLC. A Association between RB1 VAF and immune score; B wGII between wild-type RB1 and mutant-type RB1 in the whole cohort; C wGII in SCLC-ADC/SCLC subtypes; D Heatmap of DDR gene expression; E Likelihood of occurring neoantigen expressed in tumors with wild-type RB1 and mutant-type RB1; F Likelihood of generating neoantigen in consistently expressed genes in tumors with wild-type RB1 and mutant-type RB1; G Likelihood of neoantigen occurring in all copy number loss regions in tumors with wild-type RB1 and mutant-type RB1; H Likelihood of neoantigen occurring in clonal copy number loss regions in tumors with wild-type RB1 and mutant-type RB1; I Likelihood of neoantigen occurring in subclonal copy number loss regions in tumors with wild-type RB1 and mutant-type RB1; J Enrichment score of cytokine production; K Chronic inflammatory response up; L Acute inflammatory response up pathways between wild-type RB1 and mutant-type RB1; M Influence of RB1 alteration status on TIME