title: Pazopanib reveals a role for tumor cell B-Raf in the prevention of HER2+ breast cancer brain metastasis.
abstract: OBJECTIVE
  Brain metastases of breast cancer contribute significantly to patient morbidity and mortality. We have tested pazopanib, a recently approved antiangiogenic drug that targets VEGFR1, VEGFR2, VEGFR3, PDGFRβ, PDGFRα, and c-kit, for prevention of experimental brain metastases and mechanism of action.
  
  
  METHODS
  In vitro assays included B-Raf enzymatic assays, Western blots, and angiogenesis assays. For in vivo assays, HER2 transfectants of the brain seeking sublines of MDA-MB-231 cells (231-BR-HER2) and MCF7 cells (MCF7-HER2-BR3, derived herein) were injected into the left cardiac ventricle of mice and treated with vehicle or pazopanib beginning on day 3 postinjection. Brain metastases were counted histologically, imaged, and immunostained.
  
  
  RESULTS
  Treatment with 100 mg/kg of pazopanib resulted in a 73% decline in large 231-BR-HER2 metastases (P < 0.0001) and a 39% decline in micrometastases (P = 0.004). In vitro, pazopanib was directly antiproliferative to 231-BR-HER2 breast cancer cells and inhibited MEK and ERK activation in vitro despite B-Raf and Ras mutations. Enzymatic assays demonstrated that pazopanib directly inhibited the wild type and exon 11 oncogenic mutant, but not the V600E mutant forms of B-Raf. Activation of the B-Raf targets pERK1/2 and pMEK1/2 was decreased in pazopanib-treated brain metastases whereas blood vessel density was unaltered. In the MCF7-HER2-BR3 experimental brain metastasis model, pazopanib reduced overall brain metastasis volume upon magnetic resonance imaging (MRI) by 55% (P = 0.067), without affecting brain metastasis vascular density.
  
  
  CONCLUSIONS
  The data identify a new activity for pazopanib directly on tumor cells as a pan-Raf inhibitor and suggest its potential for prevention of brain metastatic colonization of HER2(+) breast cancer.
authors.affiliation: Women's Cancers Section, Laboratory of Molecular Pharmacology, Radiation Oncology Branch, Center for Cancer Research, National Cancer Institute, Bethesda, Maryland 20892, USA. grilbrun@mail.nih.gov
authors.forename: Brunilde
authors.initials: B
authors.lastname: Gril
authors.forename: Diane
authors.initials: D
authors.lastname: Palmieri
authors.forename: Yong
authors.initials: Y
authors.lastname: Qian
authors.forename: DeeDee
authors.initials: D
authors.lastname: Smart
authors.forename: Lilia
authors.initials: L
authors.lastname: Ileva
authors.forename: David J
authors.initials: DJ
authors.lastname: Liewehr
authors.forename: Seth M
authors.initials: SM
authors.lastname: Steinberg
authors.forename: Patricia S
authors.initials: PS
authors.lastname: Steeg
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology* / Q000627:therapeutic use; D001932:Brain Neoplasms / Q000235:genetics / Q000517:prevention & control* / Q000556:secondary*; D001943:Breast Neoplasms / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism* / Q000473:pathology*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D004195:Disease Models, Animal; D042783:Endothelial Cells / Q000187:drug effects; D048049:Extracellular Signal-Regulated MAP Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D005260:Female; D006801:Humans; D007191:Indazoles; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D009154:Mutation; D048493:Proto-Oncogene Proteins B-raf / Q000037:antagonists & inhibitors* / Q000378:metabolism; D011743:Pyrimidines / Q000494:pharmacology* / Q000627:therapeutic use; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000235:genetics*; D015398:Signal Transduction / Q000187:drug effects; D013449:Sulfonamides / Q000494:pharmacology* / Q000627:therapeutic use; D023041:Xenograft Model Antitumor Assays
chemical_list: D000970:Antineoplastic Agents; D048049:Extracellular Signal-Regulated MAP Kinases; D007191:Indazoles; D048493:Proto-Oncogene Proteins B-raf; D011743:Pyrimidines; D018719:Erb-b2 Receptor Tyrosine Kinases; D013449:Sulfonamides; C516667:pazopanib; C508053:ERBB2 protein, human