EACR25-1853

Unraveling CIP2A’s impact on DNA damage response in basal- like triple-negative breast cancer

P. Doan¹, O. Kauko¹, S. Nagelli², J. Westermarck^1,3

¹Turku Bioscience Centre, University of Turku and Åbo Akademi University, Turku, Finland
²Turku Bioscience Centre, University of Turku and Åbo Akademi University, Finland
³Institute of Biomedicine, University of Turku, Turku, Finland

Introduction:

Basal-like triple-negative breast cancer (BL-TNBC) is an aggressive malignancy characterized by a high mutational burden, frequently driven by BRCA mutations or defects in homologous recombination (HR) pathways. CIP2A, a known inhibitor of protein phosphatase 2A (PP2A), is a driver protein for BL-TNBC initiation and progression. Emerging phosphoproteomic data suggest that CIP2A impairs PP2A-mediated dephosphorylation of several DNA damage response (DDR) proteins. However, the functional consequences of these phosphorylation events in BL-TNBC remain unclear. This study investigates the role of CIP2A in DDR regulation and its impact on BL-TNBC cell survival and therapeutic response.

Material and method:

A functional dependency analysis using DepMap data was conducted to assess CIP2A’s association with key DDR proteins. Site-specific CRISPR/Cas9 mutagenesis screening was employed to examine the impact of CIP2A-regulated phosphosites in BL-TNBC cells. Additionally, CRISPR/Cas9 mutagenesis under ATR inhibitor treatment was performed to evaluate phosphosite-specific contributions to therapy response. Functional assays, including cytotoxicity and colony formation assays, were carried out in TNBC cells expressing a PP2A-binding defective CIP2A mutant in combination with DDR-targeting agents to further elucidate the relationship between CIP2A and DDR.

Result and discussion:

CIP2A was found to exhibit functional co-dependency with multiple DDR proteins. Phosphoproteomic analysis confirmed CIP2A-mediated inhibition of PP2A-dependent dephosphorylation within the TopBP1-associated DDR complex. CRISPR/Cas9 mutagenesis revealed differential cell fitness effects upon phosphosite-specific modifications. Furthermore, TNBC cells expressing the PP2A-binding defective CIP2A mutant displayed altered sensitivity to DDR inhibitors, underscoring the functional significance of CIP2A-mediated regulation in therapy response.

Conclusion:

Our findings demonstrate that CIP2A plays a crucial role in DDR phosphoregulation, influencing BL-TNBC cell survival and therapeutic vulnerability. By leveraging site-specific CRISPR/Cas9 mutagenesis, we aim to identify additional DDR targets modulated by CIP2A. These insights may pave the way for novel therapeutic strategies targeting CIP2A-dependent DDR vulnerabilities in BL-TNBC.