Surveying the Proteome-Wide Landscape of Mitoxantrone and Examining Drug Sensitivity in BRCA1-Deficient Ovarian Cancer Using Quantitative Proteomics

<b>Background</b>: Mitoxantrone (MX) is regularly used to treat several cancers. Despite its long history in the clinic, recent studies continue to unveil novel protein targets. These targets may contribute to the cytotoxic effects of the drug, as well as potential non-canonical antitumor activity. A better understanding of MX's cellular targets is required to fully comprehend the molecular consequences of treatment and to interpret MX sensitivity in homologous recombination (HR)-deficient cancer. <b>Methods</b>: Here, we evaluated MX activity in HR-deficient UWB1.289 (BRCA1-) ovarian cancer cells and surveyed the binding profile of MX using TMT-labeled quantitative proteomics and chemoproteomics. <b>Results</b>: Mass spectrometry (MS) analysis of cellular extracts from MX-treated BRCA1-UWB1.289 cells revealed unique downregulation of pathways instrumental in maintaining genomic stability, including single-strand annealing. Moreover, the BRCA1- cells exhibited a significant upregulation of proteins involved in ribosome biogenesis and RNA processing. Additional MS analyses following affinity-purification using a biotinylated-mitoxantrone probe corroborated these findings, which showed considerable targeting of proteins involved in genome maintenance and RNA processing. <b>Conclusions</b>: Our results suggest that an interplay of both canonical and non-canonical MX-antitumor activity overwhelms the BRCA1- UWB1.289 cells. Furthermore, this study characterizes the target landscape of MX, providing insights into off-target effects and MX action in HR-deficient cancer.