Huduti, E. | bioRxiv Science

bioRxiv2026

Mapping the immune landscape in small cell lung cancer unveils a distinct tumor-reactive CD8+ T cell molecular signature

Small cell lung cancer (SCLC) is a highly aggressive malignancy with limited therapeutic advances. Unlike many other cancers, its immune landscape, particularly immune competence and T cell recognition, remains poorly characterized. Here, we generate a single-cell transcriptome atlas of the SCLC immune microenvironment with paired T cell receptor (TCR) sequencing. By linking T cell states with clonality and a multilayered functional screening, we identify 6 tumor-reactive TCRs that recognize and eradicate autologous SCLC cell lines. We delineate a novel SCLC-reactive CD8+ T cell signature (SCLC_TR), enabling the identification of 47 further SCLC-reactive TCRs. The SCLC_TR signature performs extremely well in pancreatic ductal adenocarcinoma (PDAC), another immune-cold tumor indication, and, most strikingly, patients with elevated SCLC_TR signature scores exhibited significantly improved survival, underlining its prognostic potential. Comparative cell-cell interaction analyses implicate several immunosuppressive mechanisms, with myeloid cells and CD4 regulatory T cells possibly acting as counterbalances to effector T cell activity in SCLC. In summary, our study challenges the prevailing notion of SCLC as an immune-cold tumor type by providing direct evidence of tumor-reactive T cell responses and introduces the SCLC_TR signature as a tool to identify tumor-specific T cells and their microenvironmental restraints and escape mechanisms, ultimately shaping next-generation immunotherapeutic strategies. O_FIG O_LINKSMALLFIG WIDTH=195 HEIGHT=200 SRC="FIGDIR/small/735200v1_ufig1.gif" ALT="Figure 1"> View larger version (69K): org.highwire.dtl.DTLVardef@1c25f2eorg.highwire.dtl.DTLVardef@1f6f519org.highwire.dtl.DTLVardef@553e57org.highwire.dtl.DTLVardef@6fe7e1_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2023

Incorporation of TRDV segments into TCR alpha chains

In human tumor models we tried to identify and clone the TCR of tumor-reactive T cells enriched in mixed lymphocyte tumor-cell cultures (MLTC). In a particular MLTC, we identified a predominant TCR beta chain using the Beta Mark Vbeta Kit, but a corresponding alpha chain could not be amplified via RT-PCR using TRAV-specific forward primers. Therefore, we applied 5RACE to obtain the TCR alpha chain sequences. The 5RACE product revealed an alpha chain that encompassed 89bp of the TRDV1 5UTR, followed by the TRDV1 coding sequence joined in frame to TRAJ24. The ORF reaching from the TRDV1 start codon to the TRAC segment was intact, suggesting a functional TCR. To analyze this MLTC population in greater depth we conducted 10X VDJ sequencing. CellRanger identified the beta chain known from the Beta Mark analysis, but no corresponding alpha chain in the filtered results. The corresponding TRDV-containing TCR alpha chain could, however, be detected in the "all_contig_annotations" files. In a separate project, we performed TCR sequencing of tumor-infiltrating lymphocytes (TILs) in a murine tumor model. Also here, a predominant clonotype contained a TCR alpha chain joining Trdv2-2 in frame to Traj49. Transfection of both TCR cDNAs resulted in cell surface localization of TCR and CD3 as validated by FACS. Tumor recognition of the human, TRDV1-containing TCR could be demonstrated by IFNgamma ELISpot whereas the murine TCR did not recognize a tumor-derived cell line. TRDV-containing alpha chains have been reported in the literature for two HLA I-restricted TCRs against HIV peptides (Ueno et al, Eur J Immunol, 2003). To determine whether such TDRV-containing TCRs are unique events or whether Vdelta segments are commonly incorporated into TCR alpha chains, we queried the NCBI Sequence Read Archive (SRA) for 10X VDJ data and analyzed 21 human and 23 murine datasets. We found that especially TRDV1, Trdv1 and to some extent Trdv2-2 are more commonly incorporated into TCR alpha chains than some TRAV genes, making the TRDV segments a relevant contribution to TCR alpha diversity. For apparently solitary beta chains in 10X VDJ datasets, we suggest to scrutinize the "all_contig_annotations" files as these may contain an accompanying, TRDV-containing alpha chain.

immunology↗