Celecoxib alleviates tamoxifen-instigated angiogenic effects by ROS-dependent VEGF/VEGFR2 autocrine signaling

Abstract

BackgroundTamoxifen (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.MethodsThe 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.ResultsHere, 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.ConclusionsTaken 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.

Synergistic Effects of Tamoxifen and Tranilast on VEGF and MMP-9 Regulation in Cultured Human Breast Cancer Cells

“…Tamoxifen is a competitive antagonist of estrogen, but seems to have antiangiogenic effects independent of, and in addition to, estrogen receptor content or its anti-estrogen effects (Kousidoua et al, 2008). It have been reported that tamoxifen treatment of breast cancer, induces an antiangiogenic response by an efficient decrease of VEGF both in vitro and in vivo (Garvin and Dabrosin, 2003;Garvin et al, 2005;Nilsson and Dabrosin, 2006;Kumar et al, 2013;Rajput et al, 2013).…”

Molecular inhibition of prostaglandin E2 with GW627368X: Therapeutic potential and preclinical safety assessment in mouse sarcoma model

“…IHC for specific proteins were carried out on tumor tissue sections using Biogenex IHC detection system according to manufacturer's guidelines. 24,25 Prostaglandin E2 and VEGF quantification by ELISA Snap frozen tissue samples preserved in ¡80…”

“…From the stored sample, level of VEGF in serum of treated and untreated groups were quantified using VEGF quantikine ELISA kit, R&D systems according to kit guidelines. 25 Toxicological study Gross behavioral examination All animals were inspected twice a day during the course of the study and monitored carefully during weighing/feeding period. Animals were observed for reaction to treatment such as changes in skin, fur, eyes, and mucous membranes.…”

“…The suspension was centrifuged at 400 g for 5mins and supernatant discarded. Following another wash with PBS the spleenocytes were seeded in RPMI medium in 96 well (3,000cells/well) and 6 well (5 £ 10 5 cell/well) plates for MTT assay 25 and crystal violet staining respectively.…”

Cancer Recurrence and Regional Anesthesia: The Theories, the Data, and the Future in Outcomes

“…Therefore, postulated mechanisms proposed for the antitumor activity of nonsteroidal anti-inflammatory drugs (NSAIDs) include reduction in prostaglandin E2, resulting in activation of apoptosis in tumor cells [49], restricted angiogenesis via down-regulation of VEGF expression [50], and reduced inhibition of NK cell activity [5,51].…”