title: Celecoxib alleviates tamoxifen-instigated angiogenic effects by ROS-dependent VEGF/VEGFR2 autocrine signaling.
abstract: BACKGROUND
  Tamoxifen (TAM) is widely used in the chemotherapy of breast cancer and as a preventive agent against recurrence after surgery. However, extended TAM administration for breast cancer induces increased VEGF levels in patients, promoting new blood vessel formation and thereby limiting its efficacy. Celecoxib (CXB), a selective COX-2 inhibitor, suppresses VEGF gene expression by targeting the VEGF promoter responsible for its inhibitory effect. For this study, we had selected CXB as non-steroidal anti-inflammatory drug in combination with TAM for suppressing VEGF expression and simultaneously reducing doses of both the drugs.
  
  
  METHODS
  The effects of CXB combined with TAM were examined in two human breast cancer cell lines in culture, MCF7 and MDA-MB-231. Assays of proliferation, apoptosis, angiogenesis, metastasis, cell cycle distribution, and receptor signaling were performed.
  
  
  RESULTS
  Here, we elucidated how the combination of TAM and CXB at nontoxic doses exerts anti-angiogenic effects by specifically targeting VEGF/VEGFR2 autocrine signaling through ROS generation. At the molecular level, TAM-CXB suppresses VHL-mediated HIF-1α activation, responsible for expression of COX-2, MMP-2 and VEGF. Besides low VEGF levels, TAM-CXB also suppresses VEGFR2 expression, confirmed through quantifying secreted VEGF levels, luciferase and RT-PCR studies. Interestingly, we observed that TAM-CXB was effective in blocking VEGFR2 promoter induced expression and further 2 fold decrease in VEGF levels was observed in combination than TAM alone in both cell lines. Secondly, TAM-CXB regulated VEGFR2 inhibits Src expression, responsible for tumor progression and metastasis. FACS and in vivo enzymatic studies showed significant increase in the reactive oxygen species upon TAM-CXB treatment.
  
  
  CONCLUSIONS
  Taken together, our experimental results indicate that this additive combination shows promising outcome in anti-metastatic and apoptotic studies. In a line, our preclinical studies evidenced that this additive combination of TAM and CXB is a potential drug candidate for treatment of breast tumors expressing high levels of VEGF and VEGFR2. This ingenious combination might be a better tailored clinical regimen than TAM alone for breast cancer treatment.
authors.affiliation: School of Medical Science and Technology, Indian Institute of Technology Kharagpur, Kharagpur-721302, West Bengal PIN-721302, India.
authors.forename: B N Prashanth
authors.initials: BN
authors.lastname: Kumar
authors.forename: Shashi
authors.initials: S
authors.lastname: Rajput
authors.forename: Kaushik Kumar
authors.initials: KK
authors.lastname: Dey
authors.forename: Aditya
authors.initials: A
authors.lastname: Parekh
authors.forename: Subhasis
authors.initials: S
authors.lastname: Das
authors.forename: Abhijit
authors.initials: A
authors.lastname: Mazumdar
authors.forename: Mahitosh
authors.initials: M
authors.lastname: Mandal
mesh_terms: D000818:Animals; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000494:pharmacology*; D019898:Autocrine Communication / Q000187:drug effects*; D015153:Blotting, Western; D001943:Breast Neoplasms / Q000378:metabolism*; D000068579:Celecoxib; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D005260:Female; D006801:Humans; D051379:Mice; D009389:Neovascularization, Pathologic / Q000139:chemically induced; D011720:Pyrazoles / Q000008:administration & dosage; D017382:Reactive Oxygen Species / Q000378:metabolism; D040262:Receptors, Vascular Endothelial Growth Factor / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects; D013449:Sulfonamides / Q000008:administration & dosage; D013629:Tamoxifen / Q000008:administration & dosage / Q000009:adverse effects; D014162:Transfection; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism*; D023041:Xenograft Model Antitumor Assays
chemical_list: D011720:Pyrazoles; D017382:Reactive Oxygen Species; D013449:Sulfonamides; D042461:Vascular Endothelial Growth Factor A; D013629:Tamoxifen; D040262:Receptors, Vascular Endothelial Growth Factor; D000068579:Celecoxib