Abstract C030: LINE-1 expression imbalance is linked to immune modulation and pluripotency heterogeneity

Abstract Immune-repressive tumor microenvironments and cancer clonal heterogeneity remain major barriers to effective therapy. Cancer cells also display features of partial reprogramming, making the choice of an appropriate “normal” reference challenging. Although induced pluripotent stem cells (iPSCs) provide a comparative system, they too exhibit notable clonal heterogeneity. Such heterogeneity in both cancer and iPSCs often maps to Polycomb-covered domains enriched for transposable elements (TEs). LINE-1 (L1) retrotransposons make up a major fraction of these elements and have been associated with both innate and adaptive immune responses. Their activity is tightly regulated in embryonic stem cells (ESCs) via KRAB-containing zinc finger proteins (KZFPs) targeting L1s for epigenetic silencing, but increasing evidence indicates that these elements are variably activated in cancer. Here, we compare L1 activation across ESCs, iPSCs, and cancers to determine how L1 heterogeneity contributes to immune modulation. We analyzed stranded RNA-seq datasets to characterize sense and antisense L1 expression across fibroblasts, ESCs, iPSCs, and cancer. Cancers and ESCs showed elevated antisense L1 activation relative to their controls, whereas iPSCs expressed higher overall L1 activation compared to fetal fibroblasts but maintained a sense-biased pattern compared to ESCs. Interferon-stimulated gene (ISG) scoring revealed that samples with the highest sense L1 activation had the highest ISG score, whereas antisense-high samples showed reduced ISG score. Moreover, the number of KZFP genes showing negative correlations with activated L1 increased specifically for antisense L1 compared with sense L1, suggesting an antisense-linked regulatory mechanism associated with immune repression. A distance-weighted L1 activation score for each gene identified gene-level effects of L1 activation, with high-scoring genes in ESC enriched for ligase-related functions. Based on this scoring, the oncogene ZNF93 exhibited L1 influence in ESCs but not in iPSCs. Notably, the iPSC sample with the highest ISG score also exhibited the most L1-gene chimeric events. Finally, long-read datasets validated L1-driven transcripts such as LINC01801, with L1-derived isoforms detected in both ESCs and iPSCs. Together, these findings show that dedifferentiation states impose distinct L1 regulatory programs with immune implications. Both ESCs and cancers display antisense-biased L1 activation linked to reduced ISG signaling and expanded negative KZFP correlations, consistent with an immune-repressed environment. Heterogeneous L1 activation may represent a shared mechanism driving immune modulation and transcriptional variability in pluripotent and cancer contexts. Further studies are warranted to define the direct interactions between L1 transcripts, KZFP networks, and immune-related pathways. Citation Format: Qianhui Wan, Amy Leung, Abner Portillo, Mahek Vinod. Bhandari, Joo Song, Dustin Schones. LINE-1 expression imbalance is linked to immune modulation and pluripotency heterogeneity [abstract]. In: Proceedings of the AACR Immuno-Oncology Conference (AACR IO): Discovery and Innovation in Cancer Immunology: Revolutionizing Treatment through Immunotherapy; 2026 Feb 18-21; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Immunol Res 2026;14(2 Suppl):Abstract nr C030.