Abstract C006: BID upregulation associates with sensitivity to TTK inhibitors in tumor cell lines

Abstract

Abstract Background: Drugs overriding the spindle assembly checkpoint (SAC), such as Aurora kinase B (AURKB) or dual-specificity protein kinase TTK (=Mps1) inhibitors, are in Phase I/II clinical trials. However, their clinical application has been hampered by the lack of biomarkers for patient selection. We have previously demonstrated that BID upregulation, present in 6% of human tumors, predicts sensitivity to AURKB inhibition in cell line and animal models. Here, we investigated the association of BID expression with response to TTK inhibitors (TTKi). Methods: A panel of seven solid tumor cell lines of lung, breast, melanoma, and pancreas origin were used in the study, together with three EGFR-mut clones derived from the PC9 lung cancer cell line. All of them had been previously characterized for BID expression at the mRNA and protein levels. The effects of three TTKi (BAY1217389, CFI-402257 and BOS172722) on proliferation, cell cycle and cell death were analyzed by MTT and flow cytometry, respectively. Silencing and ectopic expression with lentiviral vectors, together with transfection with a 72-gene knock-out library, were used to investigate mechanisms of action. Results: The concordance in response to the three TTKi among the cell lines and clones analyzed was 100%. Three of the seven tumor cell lines and 2/3 EGFR-mut clones were uniformly sensitive, showing IC50s <100 nM  for CFI-402257 and BOS172722 and <10 nM for BAY1217389. BID expression was significantly associated with response to TTKi (p<0.05). All cells sensitive to TTKi had high BID mRNA and protein levels; while the cells with low BID were resistant. Flow cytometry showed G2/M arrest after TTKi in all lines and clones, followed by cell death only in those with high BID. Silencing using shRNA viral particles significantly reduced sensitivity to the three TTKi in the two sensitive cell lines and clones tested. Conversely, ectopic expression of BID in two resistant cell lines increased sensitivity to BAY1217389 and BOS172722. Finally, among a library of 72 genes, only knock-out of BID and the genes coding for the components of the PIDDosome (PIDD, CRADD and CASP2) triggered resistance to TTKi. Conclusions: BID upregulation associates with sensitivity to TTKi through a mechanism involving the PIDDosome. Our results warrant clinical exploration of TTKi in BID overexpressing tumors. Citation Format: Jordi Bertran-Alamillo, Ana Giménez-Capitán, Silvia García-Román, Rafael Rosell, Miguel A Molina-Vila. BID upregulation associates with sensitivity to TTK inhibitors in tumor cell lines [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr C006.