title: Vemurafenib resistance reprograms melanoma cells towards glutamine dependence.
abstract: BACKGROUND
  (V600) BRAF mutations drive approximately 50% of metastatic melanoma which can be therapeutically targeted by BRAF inhibitors (BRAFi) and, based on resistance mechanisms, the combination of BRAF and MEK inhibitors (BRAFi + MEKi). Although the combination therapy has been shown to provide superior clinical benefits, acquired resistance is still prevalent and limits the overall survival benefits. Recent work has shown that oncogenic changes can lead to alterations in tumor cell metabolism rendering cells addicted to nutrients, such as the amino acid glutamine. Here, we evaluated whether melanoma cells with acquired resistance display glutamine dependence and whether glutamine metabolism can be a potential molecular target to treat resistant cells.
  
  
  METHODS
  Isogenic BRAFi sensitive parental (V600) BRAF mutant melanoma cell lines and resistant (derived by chronic treatment with vemurafenib) sub-lines were used to assess differences in the glutamine uptake and sensitivity to glutamine deprivation. To evaluate a broader range of resistance mechanisms, isogenic pairs where the sub-lines were resistant to BRAFi + MEKi were also studied. Since resistant cells demonstrated increased sensitivity to glutamine deficiency, we used glutaminase inhibitors BPTES [bis-2-(5 phenylacetamido-1, 2, 4-thiadiazol-2-yl) ethyl sulfide] and L-L-DON (6-Diazo-5-oxo-L-norleucine) to treat MAPK pathway inhibitor (MAPKi) resistant cell populations both in vitro and in vivo.
  
  
  RESULTS
  We demonstrated that MAPKi-acquired resistant cells uptook greater amounts of glutamine and have increased sensitivity to glutamine deprivation than their MAPKi-sensitive counterparts. In addition, it was found that both BPTES and L-DON were more effective at decreasing cell survival of MAPKi-resistant sub-lines than parental cell populations in vitro. We also showed that mutant NRAS was critical for glutamine addiction in mutant NRAS driven resistance. When tested in vivo, we found that xenografts derived from resistant cells were more sensitive to BPTES or L-DON treatment than those derived from parental cells.
  
  
  CONCLUSIONS
  Our study is a proof-of-concept for the potential of targeting glutamine metabolism as an alternative strategy to suppress acquired MAPKi-resistance in melanoma.
authors.affiliation: Department of Cancer Biology, Beckman Research Institute of City of Hope Cancer Center, Duarte, CA, 91010, USA. jedavies@coh.org.
authors.forename: Jenny E
authors.initials: JE
authors.lastname: Hernandez-Davies
authors.affiliation: Department of Cancer Biology, Beckman Research Institute of City of Hope Cancer Center, Duarte, CA, 91010, USA. thatran@coh.org.
authors.forename: Thai Q
authors.initials: TQ
authors.lastname: Tran
authors.affiliation: Department of Cancer Biology, Beckman Research Institute of City of Hope Cancer Center, Duarte, CA, 91010, USA. mreida@coh.org.
authors.forename: Michael A
authors.initials: MA
authors.lastname: Reid
authors.affiliation: Department of Cancer Biology, Beckman Research Institute of City of Hope Cancer Center, Duarte, CA, 91010, USA. krosales@coh.org.
authors.forename: Kimberly R
authors.initials: KR
authors.lastname: Rosales
authors.affiliation: Department of Cancer Biology, Beckman Research Institute of City of Hope Cancer Center, Duarte, CA, 91010, USA. xlowman@coh.org.
authors.forename: Xazmin H
authors.initials: XH
authors.lastname: Lowman
authors.affiliation: Department of Cancer Biology, Beckman Research Institute of City of Hope Cancer Center, Duarte, CA, 91010, USA. Minoan@coh.org.
authors.forename: Min
authors.initials: M
authors.lastname: Pan
authors.affiliation: Division of Dermatology/Department of Medicine, David Geffen School of Medicine, University of California, Los Angeles, CA, 90095-1662, USA. gmoriceau@mednet.ucla.edu.|Jonsson Comprehensive Cancer Center, David Geffen School of Medicine, University of California, Los Angeles, CA, 90095-1662, USA. gmoriceau@mednet.ucla.edu.
authors.forename: Gatien
authors.initials: G
authors.lastname: Moriceau
authors.affiliation: Department of Cancer Biology, Beckman Research Institute of City of Hope Cancer Center, Duarte, CA, 91010, USA. yiyang@coh.org.
authors.forename: Ying
authors.initials: Y
authors.lastname: Yang
authors.affiliation: Animal Tumor Model Program, Division of Comparative Medicine, Beckman Research Institute of City of Hope Cancer Center, Duarte, CA, 91010, USA. jwu@coh.org.
authors.forename: Jun
authors.initials: J
authors.lastname: Wu
authors.affiliation: Division of Dermatology/Department of Medicine, David Geffen School of Medicine, University of California, Los Angeles, CA, 90095-1662, USA. rlo@mednet.ucla.edu.|Jonsson Comprehensive Cancer Center, David Geffen School of Medicine, University of California, Los Angeles, CA, 90095-1662, USA. rlo@mednet.ucla.edu.
authors.forename: Roger S
authors.initials: RS
authors.lastname: Lo
authors.affiliation: Department of Cancer Biology, Beckman Research Institute of City of Hope Cancer Center, Duarte, CA, 91010, USA. mekong@coh.org.
authors.forename: Mei
authors.initials: M
authors.lastname: Kong
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D065150:Cellular Reprogramming / Q000187:drug effects; D019008:Drug Resistance, Neoplasm / Q000187:drug effects*; D004791:Enzyme Inhibitors / Q000494:pharmacology / Q000627:therapeutic use; D020558:GTP Phosphohydrolases / Q000378:metabolism; D055785:Gene Knockdown Techniques; D005972:Glutaminase / Q000037:antagonists & inhibitors / Q000378:metabolism; D005973:Glutamine / Q000378:metabolism*; D006801:Humans; D007211:Indoles / Q000494:pharmacology* / Q000627:therapeutic use*; D008545:Melanoma / Q000188:drug therapy* / Q000378:metabolism* / Q000473:pathology; D008565:Membrane Proteins / Q000378:metabolism; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D013449:Sulfonamides / Q000494:pharmacology* / Q000627:therapeutic use*; D000077484:Vemurafenib
chemical_list: D004791:Enzyme Inhibitors; D007211:Indoles; D008565:Membrane Proteins; D013449:Sulfonamides; D005973:Glutamine; D000077484:Vemurafenib; D005972:Glutaminase; D020558:GTP Phosphohydrolases; C579846:NRAS protein, human