Abstract A038: LINE-1 remodel pancreatic ductal adenocarcinoma metastatic plasticity

Abstract

Retrotransposons comprise most of the human dark genome and are frequently upregulated during cancer progression. Long interspersed nuclear element-1 (LINE-1), the only autonomously active human retrotransposon, is aberrantly reactivated in pancreatic ductal adenocarcinoma (PDAC), yet how LINE-1 regulates metastatic plasticity remains unknown. LINE-1 encodes the RNA-binding protein ORF1p and the endonuclease-reverse transcriptase ORF2p, whose activities contribute to genome instability and innate immune signaling. We hypothesized that LINE-1 promotes PDAC metastatic plasticity by remodeling tumor-intrinsic programs and the metastatic microenvironment. In this study, we established doxycycline-inducible LINE-1 knockdown (PDAC8) and overexpression (PDAC6 and PDAC9) models derived from patient-derived xenografts established at Massachusetts General Hospital. We demonstrated that knockdown of LINE-1 ORF1 reduces tumor clonogenicity, proliferation, as well as three-dimensional (3D) tumor sphere size and formation ability. Next, transcriptomic analysis revealed that LINE-1 depletion downregulated other repetitive elements such as HERV-K and AluYg6 that are dsRNA contributors. LINE-1 knockdown also attenuated IRF7, a master regulator of type I interferon (IFN-I), together with downstream interferon-stimulated genes (ISGs), including OAS1, MX1, and IFIT1, indicating suppression of antiviral innate immune signaling mediated by dsRNA pattern-recognition receptors (PRRs). Furthermore, reduced LINE-1 resulted in diminished STING expression. Our findings suggest that LINE-1-derived repetitive RNAs and reverse-transcribed cDNA sustain antiviral-like innate immune activation through PRR- and cGAS/STING-associated pathways. Reduced LINE-1 expression also downregulated SERPINE1, accompanied by reduced tumor cell migration. Using experimental lung and liver metastasis models, we evaluated the role of LINE-1 during early metastatic niche colonization through Dox-induced knockdown at tumor inoculation and assessed its therapeutic potential by inducing knockdown after micrometastases were established. We showed that LINE-1 ORF1 targeted shRNA knockdown reduced tumor burden in both lung and liver metastasis. Collectively, these findings identify LINE-1 as a regulator of repeatome remodeling, innate immune signaling, and metastatic plasticity in PDAC, supporting LINE-1 ORF1 as a therapeutic target. We are currently developing and optimizing antisense oligonucleotides targeting LINE-1 ORF1 to translate these findings into therapies to suppress PDAC metastasis.