Deciphering insights of novel recombinant tmTNFα in cell growth inhibition.

Abstract

Soluble TNFα, a member of TNF superfamily attributes dual roles in apoptosis and cell proliferation whereas its precursor transmembrane TNFα (tmTNFα) has potential for tumor reduction without initiating proliferation. In this perspective, we recombinantly expressed functional tmTNFα and explored its potential in cell growth inhibition. While structural characterizations of purified tmTNFα revealed integrity of the protein, cell viability assays demonstrated significant antiproliferative effect on HepG2 (IC50: 36nM) and HeLa (IC50: 23nM) cells. Mechanistic insights into mode of cell death unveiled G1 arrest in HepG2 and G2/M arrest in HeLa cells accompanied with disruption of mitochondrial membrane potential and activation of executioner caspases. Subsequent, flow cytometry based assays resulted confirmatory evidence of apoptosis after treatment with the recombinant protein. Additionally, effect of the recombinant protein on 3D tumor spheroids was explored, which rendered reduction in tumor size due to cell death as evident from confocal microscopy studies. Effectiveness of the tmTNFα in 2D monolayer as well as in complex 3D spheroids demonstrate the therapeutic significance of the protein, featuring recombinant tmTNFα as an attractive option for cancer therapeutics in days to come.