title: Bcl-2 family proteins contribute to apoptotic resistance in lung cancer multicellular spheroids.
abstract: Combinatorial therapies using the proteasome inhibitor, bortezomib, have been found to induce synergistic apoptosis in cancer cells grown as monolayers; however, three-dimensional spheroid culture may be a better model for the multicellular resistance found in solid tumors, such as lung cancer. We tested the combinatorial apoptotic strategy of using bortezomib together with TNF-related apoptosis-inducing ligand (TRAIL), both in monolayers and in spheroids of A549 lung cancer cells. Indeed, bortezomib plus TRAIL induced synergistic apoptosis in A549 cells grown as monolayers, but had little effect on A549 cells grown as three-dimensional multicellular spheroids. The acquired resistance of spheroids was not due to a limitation of diffusion, to survival pathways, such as NF-kappaB or PI3K/Akt/mTOR, or to the up-regulation of FLIP(S) (Fas-associated death domain-like IL-1 beta-converting enzyme inhibitory protein, short). We then investigated a role for the Bcl-2 family of anti- and proapoptotic proteins. When cells formed spheroids, antiapoptotic Bcl-2 increased, whereas antiapoptotic Mcl-1 decreased. ABT-737, a small molecule that inhibits Bcl-2, but not Mcl-1, abolished the multicellular resistance of A549 spheroids to bortezomib plus TRAIL. In another lung cancer cell line, H1299, acquisition of multicellular resistance in spheroids was also accompanied by an increase in Bcl-2 and decrease in Mcl-1. In H1299 spheroids compared with those of A549, however, Mcl-1 remained higher, and Mcl-1 knockdown was more effective than ABT-737 in removing multicellular resistance. Our study suggests that the balance of Bcl-2 family proteins contributes to the acquired multicellular resistance of spheroids, and suggests a possible target for improving the response of lung cancer to bortezomib therapies.
authors.affiliation: Division of Pulmonary and Critical Care Medicine, Chang Gung Memorial Hospital, Chiayi, Taiwan.
authors.forename: Tsung-Ming
authors.initials: TM
authors.lastname: Yang
authors.forename: Dario
authors.initials: D
authors.lastname: Barbone
authors.forename: Dean A
authors.initials: DA
authors.lastname: Fennell
authors.forename: V Courtney
authors.initials: VC
authors.lastname: Broaddus
mesh_terms: D000971:Antineoplastic Combined Chemotherapy Protocols / Q000378:metabolism / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects* / Q000235:genetics; D001713:Biphenyl Compounds / Q000494:pharmacology; D001897:Boronic Acids / Q000494:pharmacology; D000069286:Bortezomib; D053446:CASP8 and FADD-Like Apoptosis Regulating Protein / Q000378:metabolism; D002450:Cell Communication / Q000187:drug effects; D045744:Cell Line, Tumor; D004058:Diffusion; D019008:Drug Resistance, Neoplasm; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D016328:NF-kappa B / Q000378:metabolism; D009596:Nitrophenols / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010879:Piperazines / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D061988:Proteasome Inhibitors; D011494:Protein Kinases / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D011719:Pyrazines / Q000494:pharmacology; D011994:Recombinant Proteins / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects; D018874:Spheroids, Cellular; D013449:Sulfonamides / Q000494:pharmacology; D053221:TNF-Related Apoptosis-Inducing Ligand / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases; D013997:Time Factors
chemical_list: D001713:Biphenyl Compounds; D001897:Boronic Acids; D000069286:Bortezomib; D053446:CASP8 and FADD-Like Apoptosis Regulating Protein; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D016328:NF-kappa B; D009596:Nitrophenols; D019869:Phosphatidylinositol 3-Kinases; D010879:Piperazines; D011480:Protease Inhibitors; D046988:Proteasome Endopeptidase Complex; D061988:Proteasome Inhibitors; D011494:Protein Kinases; D051057:Proto-Oncogene Proteins c-akt; D019253:Proto-Oncogene Proteins c-bcl-2; D011719:Pyrazines; D011994:Recombinant Proteins; D013449:Sulfonamides; D053221:TNF-Related Apoptosis-Inducing Ligand; D058570:TOR Serine-Threonine Kinases; C501332:ABT-737; C546842:MTOR protein, human