Novel Triple-Negative Breast Cancer vulnerability governed by exoribonuclease-mediated RNA decay
Introduction
Breast cancer is the most common cancer and the 2nd leading cause of cancer death in females. Triple-negative breast cancer (TNBC) is a highly aggressive and lethal subtype of breast cancer with limited treatment options. A significant portion of TNBCs harbor hyperactivation of transcription factor MYC, which drives gene dysregulation and promote the elevated growth and proliferation of TNBCs. On the other hand, it has also been shown that transcriptional amplification in cancers, including those driven by MYC hyperactivation, places specific burdens on molecular machines involved in RNA metabolism. As one such example, we have shown that inhibiting molecules involved in RNA splicing can lead to significant cell stress and death of MYC+ TNBCs. However, while groundbreaking research has focused on the dysregulation of RNA synthesis and processing in cancer, far less is known about how oncogenes such as MYC deregulate RNA decay pathways to drive aberrant gene regulation. Exoribonuclease-mediated RNA decay is a critical component of RNA metabolism. Although defects in exoribonucleases present in various disease settings have been linked to the cellular accumulation of misprocessed RNA transcripts, such relevance in cancer is far from understood. We hypothesize that exoribonucleases play critical roles in the removal of aberrant RNA transcripts and prevent the induction of dsRNA-mediated cell stresses, thereby conferring survival benefits to TNBCs.
Methods
Using datasets generated from prior genome-wide forward genetic screens and online databases, we analyzed the dependence of TNBC on exoribonucleases expression. Leveraging cutting-edge chemical-genetic (PROTAC) tools, we engineered human TNBC SUM159 cell lines to express inducible degradable forms of select exoribonuclease proteins, including DIS3, PNPT1, and EXOSC10. We used these engineered human TNBC cells to evaluate the accumulation of RNA, changes in cell fitness, and cellular alterations induced by the perturbation of exoribonuclease expressions.
Results
Through prior genome-wide forward genetic screen, we identified that silencing of several exoribonucleases resulted in a significant reduction in cell fitness in MYC hyperactivated cells as compared to those without MYC hyperactivation. Subsequent analysis of Cancer Dependency Map (DepMap) database further revealed that the function of many exoribonucleases is critical across models of TNBCs. Competition assay results demonstrated that engineered SUM159 cells exhibit reduced fitness upon degradation of exoribonucleases. qRT-PCR analysis demonstrated intracellular accumulation of various RNA transcripts downstream of exoribonuclease perturbation. Ongoing RNA-sequencing has been performed to evaluate global changes to RNA transcriptome upon perturbation of exoribonucleases, and ongoing characterization suggests degradation of various exoribonucleases leads to induction of cell cycle arrest and activation of immune signaling.
Conclusion
Through this study, we hope to shed light on how exoribonuclease-mediated RNA decay helps sustain TNBC growth and survival. Further, the study will serve as a foundation for the development of novel therapeutic approaches targeting exoribonucleases for the potential treatment of TNBCs. Additionally, the study will support the evaluation of exoribonucleases and RNA decay pathways as potential vulnerabilities of other cancers harboring dysregulated RNA metabolism.