PP2A modulation overcomes multidrug resistance in chronic lymphocytic leukemia via mPTP-dependent apoptosis
Kallesh D. Jayappa, Brian Tran, Vicki L. Gordon, Christopher Morris, Shekhar Saha, Caroline C. Farrington, Caitlin M. O’Connor, Kaitlin P. Zawacki, Krista M. Isaac, Mark Kester, Timothy P. Bender, Michael E. Williams, Craig A. Portell, Michael J. Weber, Goutham Narla
Kallesh D. Jayappa, Brian Tran, Vicki L. Gordon, Christopher Morris, Shekhar Saha, Caroline C. Farrington, Caitlin M. O’Connor, Kaitlin P. Zawacki, Krista M. Isaac, Mark Kester, Timothy P. Bender, Michael E. Williams, Craig A. Portell, Michael J. Weber, Goutham Narla
View: Text | PDF
Research Article Cell biology Oncology

PP2A modulation overcomes multidrug resistance in chronic lymphocytic leukemia via mPTP-dependent apoptosis

Abstract

Targeted therapies such as venetoclax (VEN) (Bcl-2 inhibitor) have revolutionized the treatment of chronic lymphocytic leukemia (CLL). We previously reported that persister CLL cells in treated patients overexpress multiple antiapoptotic proteins and display resistance to proapoptotic agents. Here, we demonstrated that multidrug-resistant CLL cells in vivo exhibited apoptosis restriction at a pre-mitochondrial level due to insufficient activation of the Bax and Bak (Bax/Bak) proteins. Co-immunoprecipitation analyses with selective BH domain antagonists revealed that the pleiotropic proapoptotic protein (Bim) was prevented from activating Bax/Bak by “switching” interactions to other upregulated antiapoptotic proteins (Mcl-1, Bcl-xL, Bcl-2). Hence, treatments that bypass Bax/Bak restriction are required to deplete these resistant cells in patients. Protein phosphatase 2A (PP2A) contributes to oncogenesis and treatment resistance. We observed that small-molecule activator of PP2A (SMAP) induced cytotoxicity in multiple cancer cell lines and CLL samples, including multidrug-resistant leukemia and lymphoma cells. The SMAP (DT-061) activated apoptosis in multidrug-resistant CLL cells through induction of mitochondrial permeability transition pores, independent of Bax/Bak. DT-061 inhibited the growth of wild-type and Bax/Bak double-knockout, multidrug-resistant CLL cells in a xenograft mouse model. Collectively, we discovered multidrug-resistant CLL cells in patients and validated a pharmacologically tractable pathway to deplete this reservoir.

Authors

Kallesh D. Jayappa, Brian Tran, Vicki L. Gordon, Christopher Morris, Shekhar Saha, Caroline C. Farrington, Caitlin M. O’Connor, Kaitlin P. Zawacki, Krista M. Isaac, Mark Kester, Timothy P. Bender, Michael E. Williams, Craig A. Portell, Michael J. Weber, Goutham Narla

×

Figure 5

PP2A modulation by DT-061 activates apoptosis in CLL cells by releasing mPTPs.

Options: View larger image (or click on image) Download as PowerPoint
PP2A modulation by DT-061 activates apoptosis in CLL cells by releasing ...
(A and B) PBMCs from patients with CLL pretreated with increasing concentrations of the CypD inhibitor NIM811 or CspA for 1 hour were incubated with DT-061 (16 μM) or VEN (25 nM) for 12 hours. Apoptosis induction was determined by analyzing cleaved caspase 9, cleaved PARP, and viability dye staining in CLL cells using flow cytometry (cleaved PARP and viability dye data are included in Supplemental Figure 7). Data are presented as box plots showing the percentage of CLL (CD5+CD19+) cells positive for cleaved caspase-9. (C) Samples from patients with CLL pretreated with the CypD inhibitor NIM811 (10 μM) or CspA (10 μM) were incubated with DT-061 (12, 16, and 20 μM) for 6 hours, and mPTP opening in CLL (CD5+CD19+) cells was assessed using flow cytometry as described in Methods. Stacked histograms show calcein AM staining in CLL cells subjected to various treatments (left panel). Box plots show the percentage CLL cells positive for Calcein AM staining in multiple patient samples treated with DT-061 with or without NIM811 or CspA pretreatment (right panel). Statistical significance was determined by ANOVA with Šidák’s post hoc test for multiple comparisons. *P < 0.05 and **P < 0.01. Data are presented as the mean ± SD.