P15: Relationship of Repeat RNA Expression in Cancer Metastasis

Introduction: Single cell RNA-sequencing of circulating tumor cells in genetically engineered mouse models has revealed the abundant expression of repetitive element RNAs implicating their role in the metastatic cascade. Many of these repeat RNAs have recently been found to behave like viruses and undergo reverse transcription to replicate in the genome. Moreover, these viral-like repeat RNAs have been shown to alter the innate immune response in cancers. This study aimed to understand the relationship of repeat RNA expression with metastasis, the immune microenvironment, and the potential effect of nucleoside reverse transcriptase inhibitors (NRTIs), commonly used in the treatment of viral infections, on metastatic propensity in mouse cancer models.


Methods: Mouse-derived cancer cell lines RIL175 (hepatocellular carcinoma), NB508 (pancreatic ductal adenocarcinoma), and CT26 (colon cancer) were grown in the presence of emtricitabine, lamivudine, tenofovir, and islatravir, at varying concentrations. Cytotoxicity was assessed using PrestoBlue cell viability reagent. BALB/cJ mice and CT26 cells were used to generate syngeneic mouse models of colon cancer metastases. Lung metastases were generated via tail vein injection of cells and liver metastases via injection of cells directly into the liver. Initial NRTI in vivo testing has been performed with tail vein injection of CT26 cells grown in the presence of tenofovir or vehicle control. The mice were then treated with twice weekly intraperitoneal (IP) injection of drug or PBS, and tumor seeding was monitored using in vivo bioluminescence.


Results: In cells treated with tenofovir and islatravir, cytotoxicity was seen at 200 uM and 50 uM, respectively. Emtricitabine and lamivudine had no cytotoxic effects. Lung metastases were successfully generated in 19 of 22 mice and liver metastases in 9 of 11 mice. Initial in vivo experiments with tenofovir pretreatment of CT26 followed by tail vein inoculation and IP injection of tenofovir resulted in a trend of reduced lung metastases ((3.02 ± 4.36) x 10⁶ vs (8.71 ± 8.5) x 10⁵) average radiance; p = 0.61). Lung and livers were removed for formalin fixed paraffin embedding for H&E histology, immune cell IHC (CD8, CD163, CD27), and mouse repeat RNA-ISH for digital image quantification analysis.


Conclusions: We have successfully generated syngeneic liver and lung metastatic colon cancer models for comparative analysis of repeat RNA expression with associated histological features and immune infiltrate. This study indicates a potential novel oncologic application of antiretroviral medications that have been used safely in humans for viral diseases.