Preclinical Evaluation of T-Cell Prolymphocytic Leukemia Demonstrates Heterogeneous BCL2-Family Dependency That May be Effectively Targeted with Small Molecule Inhibitors

Abstract

T-cell prolymphocytic leukemia (T-PLL) is a rare, aggressive, mature T-cell leukemia with very few treatment options and adverse prognosis. There are currently no approved therapies for patients with relapsed T-PLL, where the median OS is less than 6 months. There are limited reports suggesting clinical activity of the BCL2 inhibitor venetoclax in patients with T-PLL. We aimed to characterize the dependencies of T-PLL on members of the BCL2-family of antiapoptotic proteins using BH3 profiling, a functional assay to assess the propensity of a cell to undergo apoptosis (priming) and the relative dependence of a cell on different antiapoptotic proteins. We then sought to experimentally exploit these dependencies with targeted therapies alone, and in combination, designed to inhibit the antiapoptotic proteins towards a therapeutic goal. Cell lines of mature T cell lymphomas/leukemias (TCL/L) including SUPT11 (T-cell leukemia with rearranged TCL1) (n=10) and bone marrow and peripheral blood samples from untreated patients with T-PLL (n=7) were analyzed by BH3 profiling. We also used a relapsed/refractory T-PLL patient sample that were able to engraft and expand in a PDX model. Finally, scRNA/ATAC seq has also been performed in a set of naïve-treated T-PLL patients. In contrast to TCL/L that demonstrated heterogeneity on BCL2 family member dependency, T-PLL samples were primed and consistently exhibited dependency on BCL2/BCL-xL and MCL1. Integration of scRNA/ATAC seq confirmed high expression of BCL2 and MCL1 in T-PLL cells from four patients supporting the premise that BCL2 and MCL1 could potentially be targeted for therapeutic benefit. Therefore, SUPT11 cells (used as a model of T-PLL) were exposed to venetoclax (25 and 50nM; BCL-2inh), fadraciclib (25 and 50nM; CDK2/9inh - indirect inhibitor of MCL-1 as it decreases mRNAs with high decay rates including MCL1 and MYC) or a combination of venetoclax and fadraciclib. We demonstrate that exposure to fadraciclib caused a rapid decline in the levels of phosphorylation on Serine 2 of RNA polymerase II C-terminal domain, as well as decline in the levels of MCL1 within 4 hours and completely depleted these proteins by 8-12 h of exposure. We also show that there was a synergistic increase both in mitochondrial dysfunction as measured by cytochrome C release by 18h and in cell death in cells exposed to the combination at 24h. These results suggest that combining venetoclax and fadraciclib might represent a potential therapeutic approach for T-PLL. In conclusion, preclinical evaluation of a T-PLL cell line and primary patient samples demonstrate BCL2-family dependency that can be effectively targeted with small molecule inhibitors of BCL2 and CDK2/9. In vivo studies in PDX models of T-PLL are underway to form the basis for clinical trials to study these combinations.