UNRAVELING PKC EPSILON AS POSSIBLE DRUGGABLE TARGET AGAINST BREAST CANCER

Abstract

Breast cancer is the raising concern in women, and incidence rate is increasing globally. Despite immense advancements in technology, its incidence of recurrence and development of chemotherapy resistance is rising. Through its timely detection, the overall rate of its treatment efficacy and patient survival rate can be achieved. A Protein Kinase C (PKC)’s novel class member, Protein Kinase C epsilon (PRKCE), is a serine/threonine kinase whose expression dysregulation promotes cancer's survival, growth, invasion, metastasis, and angiogenesis. The present study explored the pathogenic role of PRKCE and investigated the anti-cancer potential of Zapotin in human breast cancer. PRKCE genetic variants were studied for their potential impact on PRKCE expression as well as structure and function. Its 5’untranslated region variations affect its ability to bind with repressors such as KLF3 and microRNA such as miR-123. Its most pathogenic variants were predicted to be present in the C2-like domain that may also impact its kinase function. The bioinformatics outcomes were further validated through molecular biology experiments. Genotyping analysis on a total of 150 breast cancer patients and 100 healthy individuals indicated its C2-like variant rs1345511001 homozygous C association with breast cancer risk. PRKCE expression, along with, it’s repressors (KLF3 and miR-123) and downstream and upstream genes (VEGF, HIF-1α, SOCS3, AKT, and TPD52) in breast cancer was also studied at the transcriptional level. Expression down-regulation of miR-124 and KLF3 was observed. Elevated expression of PRKCE as well as VEGF, HIF-1α, SOCS3, and TPD52, suggested PRKCE’s contribution in promoting hypoxia. Molecular docking and dynamics simulation further indicated the Zapotin and PRKCE interaction and pharmacokinetic analysis revealed its therapeutic application. Cell viability assay, colony formation, and cell migratory assay along with expression analysis demonstrated the anti-cancer potential of Zapotin by down-regulation of PRKCE’s expression at mRNA and protein level. Zapotin treatment also down-regulated the expression of VEGF, HIF-1ɑ, SOCS3, and Abstract xvii AKT. Furthermore, the influence of Zapotin treatment on protein expression of PRKCE, HIF-1ɑ, and AKT. Zapotin reduced the expression of PRKCE, HIF-1ɑ, and AKT protein. Zapotin’s potential to promote apoptosis was further affirmed by studying PARP cleavage through immune blotting. In Zapotin treated cell, high levels of cleaved PARP were found. The impact of Zapotin’s treatment on the metabolic profile of MCF-7 and MDA-MD-231 cells was also determined through Gas Chromatography-Mass Spectroscopy (GC-MS), and metabolites were mapped on canonical pathways and protein-protein interaction networks using Ingenuity Pathway Analysis (IPA) tool. The analysis depicted that PRKCE is associated with the HIF-1α pathway and may contribute to hypoxia-mediated angiogenesis in breast cancer. PRKCE also modulates breast cancer cells’ metabolic pathways, glycolysis, and Kreb’s cycle and can be targeted through Zapotin treatment. The outcomes of the present study determined PRKCE as a possible prognostic as well as therapeutic target for breast cancer. Its potential contribution in mediating metabolic rewiring and the Warburg effect in breast cancer hinted its contribution in the development of chemoresistance. PRKCE has great potential to be translated as the theragnostic target at clinical level that can facilitate physicians in timely detection and targeting of breast cancer.