title: Inhibition of MCL-1 enhances lapatinib toxicity and overcomes lapatinib resistance via BAK-dependent autophagy.
abstract: Prior studies demonstrated that resistance to the ERBB1/2 inhibitor Lapatinib in HCT116 cells was mediated by increased MCL-1 expression. We examined whether inhibition of BCL-2 family function could restore Lapatinib toxicity in Lapatinib adapted tumor cells and enhance Lapatinib toxicity in naive cells. The BCL-2 family antagonist Obatoclax (GX15-070), that inhibits BCL-2/BCL-X(L)/MCL-1 function, enhanced Lapatinib toxicity in parental HCT116 and Lapatinib adapted HCT116 cells. In breast cancer lines, regardless of elevated ERBB1/2 expression, GX15-070 enhanced Lapatinib toxicity within 3-12 h. The promotion of Lapatinib toxicity neither correlated with cleavage of caspase 3 nor was blocked by inhibition caspases; and was not associated with changes in the activities of ERK1/2, JNK1/2 or p38 MAPK but with reduced AKT, mTOR and S6K1 phosphorylation. The promotion of Lapatinib toxicity by GX15-070 correlated with increased cytosolic levels of apoptosis inducing factor (AIF) and expression of ATG8 (LC3), and the formation of large vesicles that intensely stained for a transfected LC3-GFP construct. Knock down of the autophagy regulatory proteins ATG5 or Beclin1 suppressed the induction of LC3-GFP vesicularization and significantly reduced cell killing, whereas knock down of MCL-1 and BCL-X(L) enhanced the induction of LC3-GFP vesicularization and significantly enhanced cell killing. Knockdown of Beclin1 and AIF abolished cell killing. Collectively, our data demonstrate that Obatoclax mediated inhibition of MCL-1 rapidly enhances Lapatinib toxicity in tumor cells via a toxic form of autophagy and via AIF release from the mitochondrion.
authors.affiliation: Department of Biochemistry and Molecular Biology, Virginia Commonwealth University, Richmond, VA, USA.
authors.forename: Aditi Pandya
authors.initials: AP
authors.lastname: Martin
authors.forename: Clint
authors.initials: C
authors.lastname: Mitchell
authors.forename: Mohamed
authors.initials: M
authors.lastname: Rahmani
authors.forename: Kenneth P
authors.initials: KP
authors.lastname: Nephew
authors.forename: Steven
authors.initials: S
authors.lastname: Grant
authors.forename: Paul
authors.initials: P
authors.lastname: Dent
mesh_terms: D000971:Antineoplastic Combined Chemotherapy Protocols / Q000494:pharmacology*; D001343:Autophagy / Q000187:drug effects / Q000502:physiology; D016923:Cell Death / Q000187:drug effects; D019008:Drug Resistance, Neoplasm; D004357:Drug Synergism; D004789:Enzyme Activation / Q000187:drug effects; D045325:HCT116 Cells; D006801:Humans; D007211:Indoles; D000077341:Lapatinib; D009154:Mutation; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D011758:Pyrroles / Q000008:administration & dosage / Q000494:pharmacology*; D011799:Quinazolines / Q000008:administration & dosage / Q000494:pharmacology*; D050998:bcl-2 Homologous Antagonist-Killer Protein / Q000378:metabolism*
chemical_list: C494772:BAK1 protein, human; D007211:Indoles; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D019253:Proto-Oncogene Proteins c-bcl-2; D011758:Pyrroles; D011799:Quinazolines; D050998:bcl-2 Homologous Antagonist-Killer Protein; D000077341:Lapatinib; C520962:obatoclax