title: Molecular characterization of PS-341 (bortezomib) resistance: implications for overcoming resistance using lysophosphatidic acid acyltransferase (LPAAT)-beta inhibitors.
abstract: PS-341 (bortezomib, Velcadetrade mark) is a promising novel agent for treatment of advanced multiple myeloma (MM); however, 65% of patients with relapsed refractory disease in a phase II study do not respond to PS-341. We have previously shown that lysophosphatidic acid acyltransferase (LPAAT)-beta inhibitor CT-32615 triggers caspase-dependent apoptosis, and can overcome resistance to conventional therapeutics (i.e., dexamethasone, doxorubicin, melphalan) in MM cells. In this study, we therefore determined whether CT-32615 could also overcome resistance to PS-341. We first characterized molecular mechanisms of resistance to PS-341 in DHL-4 cells. DHL-4 cells express low levels of caspase-3 and caspase-8; furthermore, no cleavage in caspase-8, caspase-9, caspase-3, poly ADP-ribose polymerase (PARP), or DNA fragmentation factor 45 was triggered by PS-341 treatment. We have previously shown that PS-341 treatment triggers phosphorylation of c-Jun NH(2)-terminal kinase (JNK), which subsequently induces caspase-dependent apoptosis; conversely, JNK inhibition blocks PS-341-induced apoptosis. We here show that phosphorylation of SEK-1, JNK, and c-Jun are not induced by PS-341 treatment, suggesting that PS-341 does not trigger a stress response in DHL-4 cells. Importantly, CT-32615 inhibits growth of DHL-4 cells in a time- and dose-dependent fashion: a transient G2/M cell cycle arrest induced by CT-32615 is mediated via downregulation of cdc25c and cdc2. CT-32615 triggered swelling and lysis of DHL-4 cells, without caspase/PARP cleavage or TUNEL-positivity, suggesting a necrotic response. Our studies therefore demonstrate that LPAAT-beta inhibitor CT-32615 triggers necrosis, even in PS-341-resistant DHL-4 cells, providing the framework for its evaluation to overcome clinical PS-341 resistance and improve patient outcome.
authors.affiliation: Jerome Lipper Multiple Myeloma Center, Department of Medical Oncology, Dana-Farber Cancer Institute and Harvard Medical School, 44 Binney Street, Boston, MA 02115, USA.
authors.forename: Teru
authors.initials: T
authors.lastname: Hideshima
authors.forename: Dharminder
authors.initials: D
authors.lastname: Chauhan
authors.forename: Kenji
authors.initials: K
authors.lastname: Ishitsuka
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Yasui
authors.forename: Noopur
authors.initials: N
authors.lastname: Raje
authors.forename: Shaji
authors.initials: S
authors.lastname: Kumar
authors.forename: Klaus
authors.initials: K
authors.lastname: Podar
authors.forename: Constantine
authors.initials: C
authors.lastname: Mitsiades
authors.forename: Hiromasa
authors.initials: H
authors.lastname: Hideshima
authors.forename: Lynn
authors.initials: L
authors.lastname: Bonham
authors.forename: Nikhil C
authors.initials: NC
authors.lastname: Munshi
authors.forename: Paul G
authors.initials: PG
authors.lastname: Richardson
authors.forename: Jack W
authors.initials: JW
authors.lastname: Singer
authors.forename: Kenneth C
authors.initials: KC
authors.lastname: Anderson
mesh_terms: D000217:Acyltransferases / Q000037:antagonists & inhibitors*; D000255:Adenosine Triphosphate / Q000378:metabolism; D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins; D001897:Boronic Acids / Q000494:pharmacology*; D000069286:Bortezomib; D016203:CDC2 Protein Kinase / Q000378:metabolism; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000378:metabolism; D002455:Cell Division; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation; D019008:Drug Resistance, Neoplasm; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005434:Flow Cytometry; D016195:G2 Phase; D006801:Humans; D015151:Immunoblotting; D020287:In Situ Nick-End Labeling; D009101:Multiple Myeloma / Q000188:drug therapy / Q000378:metabolism; D009336:Necrosis; D010766:Phosphorylation; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D011480:Protease Inhibitors / Q000494:pharmacology*; D061988:Proteasome Inhibitors; D011506:Proteins / Q000378:metabolism; D011719:Pyrazines / Q000494:pharmacology*; D013997:Time Factors; D016896:Treatment Outcome; D020687:cdc25 Phosphatases / Q000378:metabolism
chemical_list: D051017:Apoptosis Regulatory Proteins; D001897:Boronic Acids; D018797:Cell Cycle Proteins; D004791:Enzyme Inhibitors; D011480:Protease Inhibitors; D061988:Proteasome Inhibitors; D011506:Proteins; D011719:Pyrazines; C109664:caspase-activated DNase inhibitor; D000069286:Bortezomib; D000255:Adenosine Triphosphate; D000217:Acyltransferases; C018764:2-acylglycerophosphate acyltransferase; D011065:Poly(ADP-ribose) Polymerases; D016203:CDC2 Protein Kinase; C517009:CDC25C protein, human; D020687:cdc25 Phosphatases; C505407:CASP3 protein, human; C505530:CASP8 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases