Study on the Mechanisms of Antitumor Activity of Two Type I Ribosome Inactivating Proteins

Ribosome inactivating proteins (RIPs), belonging to the family of defense proteins, manifest enzymatic activity that brings about the cleavage of a specific adenine base from ribosomal RNA. RIPs, which have been widely purified from plants, fungi and bacteria, are found to exhibit antiproliferative/antitumor, immunomodulatory, antimicrobial, and anti-insect activities. This study intends to elucidate the mechanisms of antitumor activity of two type I RIPs through in vitro and in vivo models.;In the present investigation, marmorin, a type I RIP from fruiting bodies of the edible mushroom Hypsizigus marmoreus, inhibited the survival of breast cancer in vitro and in vivo. It evinced more potent cytotoxicity toward estrogen receptor (ER)-positive MCF7 breast cancer cells than ER-negative MDA-MB-231 cells. Further study disclosed that marmorin undermined the expression level of estrogen receptor a (ERalpha) and significantly inhibited the proliferation of MCF7 cells induced by 17beta-estradiol. Knockdown of ERalpha in MCF7 cells significantly attenuated the inhibitory effect of marmorin on proliferation, suggesting that the ERalpha-mediated pathway was implicated in the suppressive action of marmorin on ER-positive breast cancer cells. Moreover, marmorin induced time-dependent apoptosis in both MCF7 and MDA-MB-231 cells. It brought about G2/M-phase arrest, mitochondrial membrane potential depolarization and caspase-9 activation in MCF7 cells, and to a lesser extent in MDA-MB-231 cells. Marmorin triggered the death receptor apoptotic pathway (e.g. caspase-8 activation) and endoplasmic reticulum stress (ERS, as evidenced by phosphorylation of PERK and IRE1alpha, cleavage of caspase-12, and up-regulation of CHOP expression) in both MCF7 and MDA-MB-231 cells. In summary, marmorin exhibited inhibitory effect on breast cancer partially via diminution of ERalpha and apoptotic pathways mediated by mitochondrial, death receptor and ERS. The results advocate that marmorin is a potential candidate for breast cancer therapy.;The other type I RIP alpha-momorcharin (alpha-MMC), which was isolated from seeds of the bitter gourd Momordica charantia, exerted its inhibitory effect on cell viability and clonogenic formation of nasopharyngeal carcinoma (NPC) CNE2 and HONE1 cells in vitro. To further ascertain its molecular mechanism, it was found that alpha-MMC targeted endoplasmic reticulum and down-regulated unfolded protein response (UPR) in CNE2 (e.g. down-regulation of expression level of PERK, IRE1alpha and CHOP) and HONE1 (e.g. down-regulation of expression level of PERK and CHOP) cells. Moreover, alpha-MMC induced dose- and time-dependent apoptosis in both CNE2 and HONE1 cells. Further study disclosed that alpha-MMC initiated mitochondrial- and death-receptor mediated apoptotic signaling cascades in CNE2 cells (as evidenced by activation of caspase-9, caspase-8 and caspase-3, and mitochondrial membrane potential depolarization). However, alpha-MMC brought about ablation in HONE1 cells. alpha-MMC caused G0/G1 phase cell cycle arrest in CNE2 cells and S phase arrest in HONE1 cells. Besides, alpha-MMC retained its cytotoxicity on hypoxic CNE2 and HONE1 cells involving down-regulation of HIF1alpha and VEGF expression. These observations indicate the potential utility of alpha-MMC for NPC therapy.;Taken together, type I RIPs marmorin and alpha-MMC inhibited the growth of breast cancer and nasopharyngeal carcinoma, respectively through triggering of the apoptotic pathway. The results advocate that type RIPs are potential candidates for cancer therapy.