title: Dual inhibition of Bcl-2 and Bcl-xL strikingly enhances PI3K inhibition-induced apoptosis in human myeloid leukemia cells through a GSK3- and Bim-dependent mechanism.
abstract: Effects of concomitant inhibition of the PI3K/AKT/mTOR pathway and Bcl-2/Bcl-xL (BCL2L1) were examined in human myeloid leukemia cells. Tetracycline-inducible Bcl-2 and Bcl-xL dual knockdown sharply increased PI3K/AKT/mTOR inhibitor lethality. Conversely, inducible knockdown or dominant-negative AKT increased, whereas constitutively active AKT reduced lethality of the Bcl-2/Bcl-xL inhibitor ABT-737. Furthermore, PI3K/mTOR inhibitors (e.g., BEZ235 and PI-103) synergistically increased ABT-737-mediated cell death in multiple leukemia cell lines and reduced colony formation in leukemic, but not normal, CD34+ cells. Notably, increased lethality was observed in four of six primary acute myelogenous leukemia (AML) specimens. Responding, but not nonresponding, samples exhibited basal AKT phosphorylation. PI3K/mTOR inhibitors markedly downregulated Mcl-1 but increased Bim binding to Bcl-2/Bcl-xL; the latter effect was abrogated by ABT-737. Combined treatment also markedly diminished Bax/Bak binding to Mcl-1, Bcl-2, or Bcl-xL. Bax, Bak, or Bim (BCL2L11) knockdown or Mcl-1 overexpression significantly diminished regimen-induced apoptosis. Interestingly, pharmacologic inhibition or short hairpin RNA knockdown of GSK3α/β significantly attenuated Mcl-1 downregulation and decreased apoptosis. In a systemic AML xenograft model, dual tetracycline-inducible knockdown of Bcl-2/Bcl-xL sharply increased BEZ235 antileukemic effects. In a subcutaneous xenograft model, BEZ235 and ABT-737 coadministration significantly diminished tumor growth, downregulated Mcl-1, activated caspases, and prolonged survival. Together, these findings suggest that antileukemic synergism between PI3K/AKT/mTOR inhibitors and BH3 mimetics involves multiple mechanisms, including Mcl-1 downregulation, release of Bim from Bcl-2/Bcl-xL as well as Bak and Bax from Mcl-1/Bcl-2/Bcl-xL, and GSK3α/β, culminating in Bax/Bak activation and apoptosis. They also argue that combining PI3K/AKT/mTOR inhibitors with BH3 mimetics warrants attention in AML, particularly in the setting of basal AKT activation and/or addiction.
authors.affiliation: Department of Medicine, Virginia Commonwealth University, The Virginia Institute for Molecular Medicine and Massey Cancer Center, Richmond, VA 23298, USA. mrahmani@vcu.edu
authors.forename: Mohamed
authors.initials: M
authors.lastname: Rahmani
authors.forename: Mandy Mayo
authors.initials: MM
authors.lastname: Aust
authors.forename: Elisa
authors.initials: E
authors.lastname: Attkisson
authors.forename: David C
authors.initials: DC
authors.lastname: Williams
authors.forename: Andrea
authors.initials: A
authors.lastname: Ferreira-Gonzalez
authors.forename: Steven
authors.initials: S
authors.lastname: Grant
mesh_terms: D000818:Animals; D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins / Q000235:genetics / Q000378:metabolism*; D000072224:Bcl-2-Like Protein 11; D001713:Biphenyl Compounds / Q000494:pharmacology; D045744:Cell Line, Tumor; D005260:Female; D055785:Gene Knockdown Techniques; D038362:Glycogen Synthase Kinase 3 / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D015151:Immunoblotting; D007951:Leukemia, Myeloid / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D009596:Nitrophenols / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism*; D010879:Piperazines / Q000494:pharmacology; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000235:genetics / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism*; D011804:Quinolines / Q000494:pharmacology; D013449:Sulfonamides / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism; D014407:Tumor Cells, Cultured; D020298:U937 Cells; D023041:Xenograft Model Antitumor Assays; D050998:bcl-2 Homologous Antagonist-Killer Protein / Q000235:genetics / Q000378:metabolism; D051020:bcl-X Protein / Q000235:genetics / Q000378:metabolism*
chemical_list: D051017:Apoptosis Regulatory Proteins; D000072224:Bcl-2-Like Protein 11; D001713:Biphenyl Compounds; D038362:Glycogen Synthase Kinase 3; D007093:Imidazoles; D008565:Membrane Proteins; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D009596:Nitrophenols; D019869:Phosphatidylinositol 3-Kinases; D010879:Piperazines; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D019253:Proto-Oncogene Proteins c-bcl-2; D011804:Quinolines; D013449:Sulfonamides; D058570:TOR Serine-Threonine Kinases; D050998:bcl-2 Homologous Antagonist-Killer Protein; D051020:bcl-X Protein; C501332:ABT-737; C494772:BAK1 protein, human; C494811:BCL2L1 protein, human; C000606775:BCL2L11 protein, human; C000606776:Bcl2l11 protein, mouse; C579094:Mcl1 protein, mouse; C546842:MTOR protein, human; C531198:dactolisib