Caspase-dependent Mcl-1 cleavage and effect of Mcl-1 phosphorylation in ABT-737-induced apoptosis in human acute lymphoblastic leukemia cell lines

Exp Biol Med (Maywood). 2014 Oct;239(10):1390-402. doi: 10.1177/1535370214538745. Epub 2014 Jun 20.

Abstract

ABT-737 is a BH3-mimetic that has a wide spectrum of single-agent activity against acute lymphoblastic leukemia (ALL) cell lines and xenografts. Previously, we reported that in response to ABT-737, ABT-737-resistant ALL cell lines showed an apparent increase in Mcl-1 (an anti-apoptotic Bcl-2 family protein that is not effectively inhibited by ABT-737) while ABT-737-sensitive ALL cell lines showed decreased Mcl-1 levels. Here we explored the mechanism of Mcl-1 cleavage by ABT-737 and the effect of adjacent phosphorylation sites on Mcl-1 cleavage and apoptosis induced by ABT-737 in a human B-lineage ALL cell line. Caspase cleavage sites in Mcl-1 and the effect of mutation in Mcl-1 phosphorylation sites were determined by transducing Mcl-1 variants tagged with the V5 epitope into human ALL cells. Cytotoxicity was by fluorescence-based DIMSCAN, and changes in protein by immunoblotting. ABT-737 induced a caspase-dependent cleavage of Mcl-1. Of the two Mcl-1 caspase cleavage sites (D127 and D157), D157 was the site of ABT-737-induced cleavage in ALL cells. Cells with exogenously expressed Mcl-1 Δ157 fragment showed greater caspase-3 and caspase-9 activation when they were treated with ABT-737 compared with cells expressing wild-type or D157A mutant Mcl-1. Cells with mutated phosphorylation sites on Mcl-1 (S159A and T163A) were less susceptible to Mcl-1 cleavage and apoptosis induced by ABT-737. Our data showed that Mcl-1 is post-translationally regulated in response to ABT-737 treatment, primarily via a caspase-dependent cleavage that generates a pro-apoptotic Mcl-1 fragment.

Keywords: ABT-737; Mcl-1; acute lymphoblastic leukemia; caspase-dependent cleavage.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Apoptosis*
  • B-Lymphocytes / drug effects*
  • B-Lymphocytes / physiology*
  • Biphenyl Compounds / toxicity*
  • Caspases / metabolism*
  • Cell Line, Tumor
  • Cytological Techniques
  • Humans
  • Hydrolysis
  • Immunoblotting
  • Myeloid Cell Leukemia Sequence 1 Protein / metabolism*
  • Nitrophenols / toxicity*
  • Phosphorylation
  • Piperazines / toxicity
  • Sulfonamides / toxicity*

Substances

  • ABT-737
  • Biphenyl Compounds
  • MCL1 protein, human
  • Myeloid Cell Leukemia Sequence 1 Protein
  • Nitrophenols
  • Piperazines
  • Sulfonamides
  • Caspases