Developing New Pharmacological Tools to Modulate Transposable Elements' Activity in Colorectal Cancer

Abstract

LINE-1 retrotransposons, also known as "jumping genes", are repetitive sequences capable of copying, pasting, and reinserting themselves into the genome. These events were documented at high frequency in various types of cancers, including colorectal cancer (CRC). Furthermore, the expression of proteins encoded by these elements, such as L1ORF1p, has been linked to cancer aggressiveness, stemness, and lower patient survival rates. Colorectal cancer stem cells (CCSC), constitute a small subset of cells endowed with pluripotency and self-renewal abilities. They play roles in tumorigenesis, cancer aggressiveness, drug resistance, cancer recurrence, and metastasis. Conventional chemotherapeutics primarily target bulk tumor cells and tend to spare cancer stem cell populations. Consequently, targeting CCSC is expected to significantly increase complete remission and survival rates in CRC patients. Here, I have characterized specific aspects of a novel repurposed drug that effectively targets CCSC, reactivates the expression of transposable elements and, consequently, triggers an innate immune response. Further, I tested an in silico drug screening approach to identify compounds with high predicted affinity for the RNA binding domains of L1ORF1p, a key protein for the LINE1 retrotransposition event to occur, from a virtual drug library. Two lead compounds, both FDA-approved drugs, were identified and evaluated for their capacity to block L1ORF1p nuclear translocation, a needed step to complete the LINE1 lifecycle, as well as their capacity to decrease LINE-1 retrotransposition levels. In addition, I established a protocol for the isolation, culture, propagation, and cryopreservation of patient-derived normal colonic organoids. This protocol is crucial to the establishment of a colonic organoid biobank, representing a powerful resource to assess cancer-selective toxicity of putative CCSC-targeting compounds. Together, this thesis emphasizes the importance of transposable elements in CRC and contributes to the establishment of a gold standard ex vivo disease-modeling system for the discovery of new therapeutic agents.