Dynamic BH3 Profiling As Pharmacodynamic Biomarker for the Activity of BH3 Mimetics

Abstract

Pharmacodynamic (PD) biomarkers provide information about the pharmacologic effects of a drug on its target, i.e. to assess whether a given agent is engaging its target in the expected manner. There is currently no convenient PD marker for BH3 mimetics. BH3 profiling (BP) is a functional assay developed to measure cells susceptibility to apoptosis and cell's dependence on certain Bcl-2 anti-apoptotic proteins for survival. Dynamic BH3 profiling (DBP) measures the changes of BP signals after drug perturbations. In this study, we studied whether DBP with lymphoid cells can serve as a PD biomarker for the activity of BH3 mimetics. Using DBP and cell lines with defined dependency, we first showed that BH3 mimetics BCL201 (S55746) and S63845 are highly selective for Bcl-2 or Mcl-1, respectively. Next, we treated primary human lymphoid cells with BCL201 and then conducted DBP. We asked if pretreatment with BCL201 altered mitochondrial sensitivity to different BH3 peptides. After treatment with 1 μM BCL201, an amount consistent with achievable levels in vivo, T cell sensitivity to the Mcl-1 selective MS1 peptide, but not Bad peptide or Bfl-1 selective FS1 peptide, increased significantly, from 20% to 51%. These results suggest that BCL201 treatment increases T cell dependency on Mcl-1 and that DBP of T cells with MS1 peptide may serve as a PD marker for BCL201 activity. Similar results were also observed in B cells. Next, we conducted DBP on healthy T cells treated with S63845. T cell mitochondrial sensitivity to MS1 peptide or Bcl-xL selective HRK peptide did not change. However, the treatment increased mitochondrial sensitivity to the Bad and FS1 peptides significantly, boosting the signal from 10.2% to 57.2% and 46.8% respectively, suggesting DBP of T cells with Bad or FS1 peptides can be robust PD markers for S63845 activity. This results also indicate that S63845 treatment augmented T cell dependency on Bcl-2 and Bfl-1 for survival. Currently, we are investigating whether DBP can be used as a PD biomarker for in vivo studies. Our preliminary data suggest that DBP works well across different species (rat, mouse, and human). More importantly, DBP can detect changes in mitochondrial apoptotic signaling of ex vivo rat cells after in vivo treatment with BH3 mimetics. Conclusions: Exposure to BH3 mimetics causes changes in mitochondrial apoptotic signaling of T/B cells which are readily detectable by DBP. DBP with lymphoid cells may provide a robust PD biomarker for clinical trials testing BH3 mimetics, either as monotherapy or in combination with other agents. Disclosures Wang: Novartis Institutes for Biomedical Research: Current Employment. Qiu:Novartis Institutes for Biomedical Research: Current Employment. Halilovic:Novartis Institutes for Biomedical Research: Current Employment, Current equity holder in publicly-traded company.