title: Cotargeting histone deacetylases and oncogenic BRAF synergistically kills human melanoma cells by necrosis independently of RIPK1 and RIPK3.
abstract: Past studies have shown that histone deacetylase (HDAC) and mutant BRAF (v-Raf murine sarcoma viral oncogene homolog B1) inhibitors synergistically kill melanoma cells with activating mutations in BRAF. However, the mechanism(s) involved remains less understood. Here, we report that combinations of HDAC and BRAF inhibitors kill BRAF(V600E) melanoma cells by induction of necrosis. Cotreatment with the HDAC inhibitor suberoylanilide hydroxamic acid (SAHA) or panobinostat (LBH589) and the BRAF inhibitor PLX4720 activated the caspase cascade, but caspases appeared dispensable for killing, in that inhibition of caspases did not invariably block induction of cell death. The majority of dying cells acquired propidium iodide positivity instantly when they became positive for Annexin V, suggesting induction of necrosis. This was supported by caspase-independent release of high-mobility group protein B1, and further consolidated by rupture of the plasma membrane and loss of nuclear and cytoplasmic contents, as manifested by transmission electron microscopic analysis. Of note, neither the necrosis inhibitor necrostatin-1 nor the small interference RNA (siRNA) knockdown of receptor-interacting protein kinase 3 (RIPK3) inhibited cell death, suggesting that RIPK1 and RIPK3 do not contribute to induction of necrosis by combinations of HDAC and BRAF inhibitors in BRAF(V600E) melanoma cells. Significantly, SAHA and the clinically available BRAF inhibitor vemurafenib cooperatively inhibited BRAF(V600E) melanoma xenograft growth in a mouse model even when caspase-3 was inhibited. Taken together, these results indicate that cotreatment with HDAC and BRAF inhibitors can bypass canonical cell death pathways to kill melanoma cells, which may be of therapeutic advantage in the treatment of melanoma.
authors.affiliation: School of Medicine and Public Health, University of Newcastle, Callaghan, NSW, Australia.
authors.forename: F
authors.initials: F
authors.lastname: Lai
authors.forename: S T
authors.initials: ST
authors.lastname: Guo
authors.forename: L
authors.initials: L
authors.lastname: Jin
authors.forename: C C
authors.initials: CC
authors.lastname: Jiang
authors.forename: C Y
authors.initials: CY
authors.lastname: Wang
authors.forename: A
authors.initials: A
authors.lastname: Croft
authors.forename: M N
authors.initials: MN
authors.lastname: Chi
authors.forename: H-Y
authors.initials: HY
authors.lastname: Tseng
authors.forename: M
authors.initials: M
authors.lastname: Farrelly
authors.forename: B
authors.initials: B
authors.lastname: Atmadibrata
authors.forename: J
authors.initials: J
authors.lastname: Norman
authors.forename: T
authors.initials: T
authors.lastname: Liu
authors.forename: P
authors.initials: P
authors.lastname: Hersey
authors.forename: X D
authors.initials: XD
authors.lastname: Zhang
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D004357:Drug Synergism; D055785:Gene Knockdown Techniques; D056572:Histone Deacetylase Inhibitors / Q000494:pharmacology*; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology; D008297:Male; D008545:Melanoma / Q000188:drug therapy*; D051379:Mice; D008819:Mice, Nude; D020125:Mutation, Missense; D009336:Necrosis; D000077767:Panobinostat; D048493:Proto-Oncogene Proteins B-raf / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics; D053422:Receptor-Interacting Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D013449:Sulfonamides / Q000494:pharmacology; D000077484:Vemurafenib; D000077337:Vorinostat; D023041:Xenograft Model Antitumor Assays
chemical_list: D000970:Antineoplastic Agents; D056572:Histone Deacetylase Inhibitors; D006877:Hydroxamic Acids; D007211:Indoles; C528407:PLX 4720; D034741:RNA, Small Interfering; D013449:Sulfonamides; D000077484:Vemurafenib; D000077337:Vorinostat; D000077767:Panobinostat; C482119:BRAF protein, human; D048493:Proto-Oncogene Proteins B-raf; C506249:RIPK1 protein, human; C119705:RIPK3 protein, human; D053422:Receptor-Interacting Protein Serine-Threonine Kinases