Identification and Characterization of TONSL as an Immortalizing Oncogene

Abstract

The global issue of exponentially increasing breast cancer cases necessitates investigating the early genomic aberrations leading to tumorigenesis. To address this, we employed our unique biobank of healthy breast tissue to generate an isogeneic cell line model system, comprising primary breast epithelial cells and their immortalized counterparts. By comparing the genetic alterations between these cell types, we discovered that TONSL upregulation is one of the early events during breast tumorigenesis. TONSL is a Tonsoku-like DNA repair protein located on chromosome 8q24.3. Our findings reveal that TONSL is an immortalizing oncogene, capable of transforming primary breast epithelial cells in conjunction with defined oncogenes, resulting in Estrogen Receptor-positive breast adenocarcinomas. Furthermore, we observed that TONSL-amplified breast cancer cells are dependent on TONSL for tumor growth, and these TONSLHigh cells and tumors exhibit an upregulated homologous recombination DNA repair pathway, which may contribute to chemotherapy resistance. It is noteworthy that higher levels of TONSL protein in primary breast cancers, particularly in ER+ breast cancers, are associated with poor outcomes. Approximately 20% of breast cancers display recurring genomic amplification involving chromosome 8q24.3, with TONSL amplification potentially being the initial hit leading to tumorigenesis. In an attempt to target these TONSL/chr. 8q24.3 amplified breast tumors, we observed that breast cancer cells with this amplification are sensitive to CBL0137 – a TONSL-FACT complex inhibitor, both in vitro and in vivo. TONSL interacts with multiple proteins and functions in multiple cellular processes. To study the TONSL specific interactome, we performed immunoprecipitation with a TONSL antibody using protein lysates from TONSL-immortalized primary breast epithelial cells, followed by mass spectrometry analysis of the immunoprecipitates. Our results identified several proteins selectively enriched with the TONSL antibody, with the most significant being ETS variant transcription factor 6 (ETV6). ETV6 is known to play a role as a transcriptional repressor during embryonic development and hematopoiesis. Further studies on TONSL/chromosome 8q24.3 amplification will contribute to our understanding of breast tumor initiation, progression, and metastasis processes, as well as facilitate the development of novel therapeutic agents targeting the TONSL interactome.