In-silico and In-vitro Evaluation of Novel Carboxamide Analogue on the Metastasis of Triple Negative Breast Cancer Cells Utilizing Novel PCPTC-loaded PEGylated-PLGA Nanocarriers.

This study aimed to determine the effects of novel N-{3-[(pyridin-4-yl)carbamoyl] phenyl} thiophene-2-carboxamide or PCPTC chemical moiety loaded Poly(lactic-co-glycolic acid)-Poly (Ethylene glycol) or (PLGA-PEGylated) NP as an anti-metastatic Ran GTPase therapeutic agent on MDA-MB231 triple-negative human breast cancer cells. Molecular docking and MD simulation was done to determine the binding potential of novel carboxamide PCPTC with Ran GTPase. PLGA and PLGA-PEG based NP encapsulating PCPTC were fabricated using the Modified Double Emulsion Solvent Evaporation Technique and characterized for size, zeta potential, polydispersity and morphology. In vitro evaluation of loaded nanoparticles such as cellular localization study, cell proliferation, cell migration, cell invasion and Ran Pull Down assay were carried out on MDA-MB231 breast cancer cells. Ran downregulation was determined by pull down assay. PCPTC with Ran GTPase exhibited strong structural stability based on in silico analysis. The average sizes of PCPTC loaded NP ranged between 166.3 nm to 257.5 nm and were all negatively charged. Scanning electron microscopy data showed that loaded NP were smooth and spherical. Fluorescence microscopy data confirmed the intracellular localization of loaded nanoparticles inside the MDA-MB231 cells. Cell proliferation assay (MTT assay) confirmed the cytotoxic effect of the loaded-NP when compared to blank nanoparticles. PCPTC-loaded NP inhibited metastasis and invasion of MDA-MB231 cells. This anti-metastatic and the anti-invasive effect was due to the Ran GTPase cycle blockage, which was confirmed by performing Ran Pull down assay. we propose that PCPTC is a promising compound to inhibit Ran GTPase and may act as a potential therapeutic agent against breast cancer. PCPTC-loaded NP successfully stopped the metastasis of MDA-MB231 cells by disrupting the Ran cycle.