Background: Inflammation is a precursor for many cancers. NF-kB plays an important role in inflammation, is associated with cancerous growth and has been identified as a potential target for development of anticancer therapies. Lung cancer is a classic example of inflammation-induced carcinogenesis where it prevents apoptosis and stimulates production of pro-inflammatory cytokines. CLEFMA or 4-[3,5-bis(2-chlorobenzylidene-4-oxo-piperidine-1-yl)-4-oxo-2-butenoic acid] is a new molecule that potently inhibits proliferation of various cancer cells without affecting normal cells. In this study, we evaluated the antiproliferative activity of CLEFMA in a mouse xenograft model of human lung cancer.
Methods: Cell viability was determined in human lung cancer cells H441 and A549 after treatment with CLEFMA. Immunobloting studies were performed in cell lysates and tissue homogenates for anti-apoptotic and pro-apoptotic markers. These were further confirmed by immunohistochemical studies. The ability of CLEFMA in reducing the metabolic turnover in tumor tissue was examined by positron emission tomography using F-18-FDG as a biomarker of glucose metabolism.
Results: We found that CLEFMA-induced suppression of lung adenocarcinoma H441 and A549 cell proliferation was associated with cleavage of caspases 3/9 and PARP, suggesting apoptotic cell death. In vivo, CLEFMA demonstrated a dose-dependent suppression of tumor growth associated with reduction of tumor proliferation marker Ki-67. Non-invasive PET imaging with F-18-flurodeoxyglucose also showed abrogation of glucose uptake in tumor tissue. In the tumor tissue homogenates, we found inhibition of anti-apoptotic markers cIAP1, Bcl-XL, BCL2 and survivin, and upregulation of pro-apoptotic BID cleavage, BAX and PARP. Further, CLEFMA inhibited NF-kB at mRNA level and reduced phospho-p65 translocation into the nucleus. It also reduced the expression of COX-2 in tumor tissue, and significantly reduced serum pro-inflammatory cytokines TNF-α and IL-6.
Conclusions: The findings suggest that CLEFMA inhibits growth of lung cancer xenografts by activating caspase-dependent apoptotic pathway, and the tumor suppression is associated with anti-inflammatory and anti-metastatic effects, perhaps mediated by NF-kB.
Citation Format: Vivek R. Yadav, Kaustuv Sahoo, Vibhudutta Awasthi. CLEFMA induces apoptosis and downregulates inflammatory markers in lung cancer model. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4866. doi:10.1158/1538-7445.AM2013-4866