Department of Biological Sciences
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Item Oxidative Stress in Retinoblastoma: Correlations with Clinicopathologic Features and Tumor Invasiveness(Taylor & Francis, 2009) Deepa, P.R.Retinoblastoma (RB) is the most common primary intraocular malignancy of infancy and childhood, where tumor invasion into the choroid, optic nerve, and/or orbit are risk factors for metastasis. Here we have correlated oxidative stress with the clinicopathologic characteristics of retinoblastoma. Methods: Tumor samples were processed for histopathologic examination. Malondialdehyde, a biomarker of oxidative stress, was immunohistochemically analyzed in 34 archival retinoblastoma tumor specimens, which included 17 tumors that did not have any invasion of the choroid, optic nerve, and/or orbit, and another 17 tumors that had some form of invasion of the choroid, optic nerve, and/or orbit. Lipid peroxidation levels were biochemically measured in another cohort of retinoblastoma tissue samples (n = 16), and correlated clinicopathologically.Item Clinico-pathological correlations of fatty acid synthase expression in retinoblastoma: An Indian cohort study(Elsiever, 2011) Deepa, P.R.Retinoblastoma (RB), the most common intra-ocular malignancy in children under 5 years of age, has an estimated incidence of about 2000 a year in India, where most cases are in advanced stage at the time of diagnosis. Newer therapeutic approaches would reduce the morbidity of chemotherapy in children with RB. Fatty Acid Synthase (FASN), a lipogenic multi-enzyme complex, is minimally expressed in normal human tissues and over expressed in many cancers, making it an attractive target for cancer therapy. We analyzed RB tissues for FASN protein expression by immunohistochemistry, western blot, and ELISA, and FASN mRNA expression by RT-PCR. FASN expression was correlated with the clinico-pathological characteristics of the tumors. FASN immunostaining was positive in all the 44 RB tissues analyzed (100%). However, FASN expression was heterogeneous within the tumor samples. Tumors with invasion of choroid, optic nerve, orbit and/or retinal pigment epithelium showed significantly higher FASN immunoreactivity than the tumors without invasion (P < 0.05), supported by western analysis (P < 0.05). FASN expression was significantly high in poorly differentiated retinoblastomas (P < 0.05). FASN protein and FASN mRNA estimated by ELISA and RT-PCR respectively showed multi-fold expression over the non-neoplastic muller glial cells that varied quantitatively between tumor tissues. FASN mRNA over-expression was substantially lower than the corresponding FASN protein expression values. The present study reports (i) markedly high expression of FASN protein in poorly differentiated and in invasive retinoblastomas, and (ii) multi-fold over-expression of FASN mRNA and protein in RB tissues, although at varying levels, indicating FASN to be a potential therapeutic target in retinoblastoma management.Item Microarray Analysis and Biochemical Correlations of Oxidative Stress Responsive Genes in Retinoblastoma(Taylor & Francis, 2012) Deepa, P.R.Oxidative stress, which refers to the biological damage caused by free radicals produced in excess of innate antioxidant defenses, is indicated in the ocular cancer retinoblastoma (RB). Here we have analysed the differential expression of oxidative stress responsive genes in oxidant-induced RB cells, and in RB tumor tissues.Item Biochemical changes accompanying apoptotic cell death in retinoblastoma cancer cells treated with lipogenic enzyme inhibitors(Elsiever, 2013) Deepa, P.R.Retinoblastoma (RB) is a malignant intra-ocular neoplasm that affects children (usually below the age of 5 years). In addition to conventional chemotherapy, novel therapeutic strategies that target metabolic pathways such as glycolysis and lipid metabolism are emerging. Fatty acid synthase (FASN), a lipogenic multi-enzyme complex, is over-expressed in retinoblastoma cancer. The present study evaluated the biochemical basis of FASN inhibition induced apoptosis in cultured Y79 RB cells. FASN inhibitors (cerulenin, triclosan and orlistat) significantly inhibited FASN enzyme activity (P < 0.05) in Y79 RB cells. This was accompanied by a decrease in palmitate synthesis (end-product depletion), and increased malonyl CoA levels (substrate accumulation). Differential lipid profile was biochemically estimated in neoplastic (Y79 RB) and non-neoplastic (3T3) cells subjected to FASN inhibition. The relative proportion of phosphatidyl choline to neutral lipids (triglyceride + total cholesterol) in Y79 RB cancer cells was found to be higher than the non-neoplastic cells, indicative of altered lipid distribution and utilization in tumor cells. FASN inhibitor treated Y79 RB and fibroblast cells showed decrease in the cellular lipids (triglyceride, cholesterol and phosphatidyl choline) levels. Apoptotic DNA damage induced by FASN inhibitors was accompanied by enhanced lipid peroxidation.Item Integrated Analysis of Dysregulated miRNA-Gene Expression in HMGA2-Silenced Retinoblastoma Cells(Sage, 2014) Deepa, P.R.Retinoblastoma (RB) is a primary childhood eye cancer. HMGA2 shows promise as a molecule for targeted therapy. The involvement of miRNAs in genome-level molecular dys-regulation in HMGA2-silenced RB cells is poorly understood. Through miRNA expression microarray profiling, and an integrated array analysis of the HMGA2-silenced RB cells, the dysregulated miRNAs and the miRNA-target relationships were modelled. Loop network analysis revealed a regulatory association between the transcription factor (SOX5) and the deregulated miRNAs imiR-29a, miR-9*, miR-9-3). Silencing of HMGA2 deregulated the vital oncomirs (miR-7, miR-331, miR-26a, miR-221, miR-17∼92 and miR-106b∼25) in RB cells. From this list, the role of the miR-106b∼25 cluster was examined further for its expression in primary RB tumor tissues (n = 20). The regulatory targets of miR-106b∼25 cluster namely p21 (cyclin-dependent kinase inhibitor) and BIM (pro-apoptotic gene) were elevated, and apoptotic cell death was observed, in RB tumor cells treated with the specific antagomirs of the miR-106b∼25 cluster. Thus, suppression of miR-106b∼25 cluster controls RB tumor growth. Taken together, HMGA2 mediated anti-tumor effect present in RB is, in part, mediated through the miR-106b∼25 clusterItem Computational and in vitro Investigation of miRNA-Gene Regulations in Retinoblastoma Pathogenesis: miRNA Mimics Strategy(Sage, 2015) Deepa, P.R.Retinoblastoma (RB), a primary pediatric intraocular tumor, arises from primitive retinal layers. Several novel molecular strategies are being developed for the clinical management of RB. miRNAs are known to regulate cancer-relevant biological processes. Here, the role of selected miRNAs, namely, miR-532-5p and miR-486-3p, has been analyzed for potential therapeutic targeting in RB.Item Targeting HSP90/Survivin using a cell permeable structure based peptido-mimetic shepherdin in retinoblastoma(Elsiever, 2016) Deepa, P.R.Retinoblastoma (RB) is a childhood retinal malignancy. Effective therapeutic strategies are still being investigated in RB disease management. Here, the anti-cancer effect of shepherdin, a peptido-mimetic inhibiting heat shock protein (HSP90)-Survivin interaction has been analyzed.Item Targeting HMGA2 in Retinoblastoma Cells in vitro Using the Aptamer Strategy(Karger, 2016) Deepa, P.R.High-mobility group A2 (HMGA2) protein regulates retinoblastoma (RB) cancer cell proliferation. Here, a stable phosphorothioate-modified HMGA2 aptamer was used to block HMGA2 protein function in RB cells. HMGA2-aptamer internalisation in RB cells (Y79, Weri Rb1) and non-neoplastic human retinal cells (MIO-M1) were optimised. Aptamer induced dose-dependent cytotoxicity in RB cancer cells (0.25-1.5 µM). Increased expression of TGFβ, SMAD4, CDH1, BAX, CASP 3, PARP mRNA and decreased SNAI1, Bcl2 mRNA levels in aptamer-treated RB cells suggests the activation of TGFβ-SMAD4-mediated apoptotic pathway. Synergistic effect with etoposide was observed in aptamer treated RB cells (p value ≤0.05). No significant toxicity was observed in non-neoplastic retinal cells.