A recombinant fungal compound induces anti-proliferative and pro-apoptotic effects on colon cancer cells.

Lili Nimri,Orly Spivak,Yitzhak Hadar,Tomer M Salame,Oded Yarden,Dominik Schälling,Irena Peri,Lutz Graeve,Dana Tal,Betty Schwartz

doi:10.18632/oncotarget.15859

Lili Nimri, Orly Spivak + Show 8 more

Open Access

https://doi.org/10.18632/oncotarget.15859

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Abstract

Finding intracellular pathways and molecules that can prevent the proliferation of colon cancer cells can provide significant bases for developing treatments for this disease. Ostreolysin (Oly) is a protein found in the mushroom Pleurotus ostreatus, and we have produced a recombinant version of this protein (rOly).We measured the viability of several colon cancer cells treated with rOly. Xenografts and syngeneic colon cancer cells were injected into in vivo mouse models, which were then treated with this recombinant protein.rOly treatment induced a significant reduction in viability of human and mouse colon cancer cells. In contrast, there was no reduction in the viability of normal epithelial cells from the small intestine. In the search for cellular targets of rOly, we showed that it enhances the anti-proliferative activity of drugs targeting cellular tubulin. This was accompanied by a reduction in the weight and volume of tumours in mice injected with rOly as compared to their respective control mice in two in vivo models.Our results advance the functional understanding of rOly as a potential anti-cancer treatment associated with pro-apoptotic activities preferentially targeting colon cancer cells.

Highlights

Up to 5 μg RNA was used for cDNA synthesis using SuperScriptTM III First-Strand Synthesis System for RT-PCR (Invitrogen) according to the manufacturers protocol
Ostreolysin (Oly) cDNA was amplified by PCR using the primers (F: 3’-ATGGCATACGCACAATGG-5’; R: 3’-TTAGT TCCCCTTCTTCAAGGTGT-5’), purified from a 1% (w/v) agarose gel and transformed into Escherichia coli JM109 cells (Promega) using the pGEM-T Vector System II (Promega) according to the manufacturer’s protocol
Its purity was determined by SDS-PAGE in the presence of a reducing agent and by analytical gel filtration on a Superdex 75 column developed in the presence of 25 mM Tris-HCl + 300 mM NaCl, pH 8

Summary

Introduction

Ostreolysin (Oly) cDNA was amplified by PCR using the primers (F: 3’-ATGGCATACGCACAATGG-5’; R: 3’-TTAGT TCCCCTTCTTCAAGGTGT-5’), purified from a 1% (w/v) agarose gel and transformed into Escherichia coli JM109 cells (Promega) using the pGEM-T Vector System II (Promega) according to the manufacturer’s protocol. The molecular mass of the protein was 15,404 Da and the specific absorbance at 280 nm (calculated by DNAman program) was 2.62.

Results

Conclusion