Elucidating BBIT20’s disruption of the BRCA1/BARD1 interaction: A new therapeutic avenue in pancreatic cancer

Introduction: Pancreatic cancer (PC) is highly resistant to treatment, remaining one of the deadliest malignancies worldwide [1]. Deficiency in the homologous recombination (HR) DNA damage repair pathway renders tumours more sensitive to therapeutic approaches [2,3]. However, because only a minority of PCs exhibit deficiencies in this pathway, effective treatment alternatives are essential [4]. BBIT20 was recently developed as a first-in-class molecule that inhibits the BRCA1/BARD1 interaction, a crucial complex in HR. In vitro and in vivo data revealed its potent antitumor activity, both as a monotherapy and combined with chemotherapy, in PC [5]. In silico analyses indicated that the BRCA1 RING domain is the preferential target of BBIT20. Nonetheless, the exact interaction site and mode of action of BBIT20 still require in vitro validation.

Methods: A recombinant BRCA1 RING domain was produced and purified from Escherichia coli BL21 (DE3). The interaction between BBIT20 and BRCA1 was assessed by fluorescent quenching and far-UV circular dichroism.

Results: Fluorescent quenching confirmed that BBIT20 induced a dose-dependent quenching effect in the BRCA1 RING domain (K_sv = 3,97*10⁴ M^-1). Circular dichroism further revealed dose‑dependent alterations in the protein’s secondary structure, indicating an interaction with BBIT20.

Conclusions: This study indicated that BBIT20 directly interacts with BRCA1 RING domain, inducing structural alterations that hinder formation of the BRCA1/BARD1 heterodimer and ultimately disrupt the HR pathway. Nuclear magnetic resonance (NMR) will be performed to gain deeper insight into the molecular dynamics of this interaction, which will support the rational design of improved BBIT20 derivatives.

Acknowledgments: This work received support from the PT national funds (FCT/MECI, Fundação para a Ciência e Tecnologia and Ministério da Educação, Ciência e Inovação) through the project 2024.13556.PEX (https://doi.org/10.54499/2024.13556.PEX) and the PhD studentship 2024.03484.BDANA (Inês Mota).