Anti-leprosy drug Clofazimine binds to human Raf1 kinase inhibitory protein and enhances ERK phosphorylation

Abstract

Human Raf1 kinase inhibitory protein (hRKIP) is an important modulator of the Ras/Raf1/MEK/ERK signaling pathway. Here, we demonstrated that anti-leprosy drug Clofazimine can bind to hRKIP with a significantly stronger affinity than the endogenous substrate phosphatidylethanolamine (PE) by using Biolayer interference technology. Moreover, we identified that residues P74, S75, K80, P111, P112, V177, and P178 play crucial roles in the binding of hRKIP to Clofazimine by using a combination of Nuclear Magnetic Resonance spectroscopy and molecular docking approach. These residues are located at the conserved ligand-binding pocket of hRKIP. Furthermore, we found that 3.2 μM Clofazimine could significantly increase the ERK phosphorylation level by about 37%. Our results indicate that Clofazimine can enhance Ras/Raf1/MEK/ERK signaling transduction pathway via binding to hRKIP. This work provides valuable hints for exploiting Clofazimine as a potential lead compound to efficiently treat the diseases related to RKIP or the Ras/Raf/MEK/ERK pathway.

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Proapoptotic effects of novel thiazole derivative on human glioma cells

The aim of the present study was to investigate the antiproliferative and proapoptotic actions of N-(5-benzyl-1,3-thiazol-2-yl)-3,5-dimethyl-1-benzofuran-2-carboxamide derivative (compound 5) in glioma cells in comparison with the actions of temozolomide (TMZ) and doxorubicin (Dox), used as positive controls. The antiproliferative activity of the compound 5, TMZ, and Dox on human glioblastoma U251 and human glioblastoma multiform T98G cells was measured using the MTT test. Western blot analysis, fluorescent microscopy, agarose gel retardation assay, flow cytometric analysis, and the DNA comet assay under alkaline conditions were carried out to study the effect of compound 5 on U251 cells. This compound showed ~20 times higher cytotoxicity toward U251 and T98G cells compared with the effects of TMZ and approximately two times higher activity than that of the Dox. Compound 5 induced apoptosis in U251 cells by PARP1 and caspase 3 cleavage mechanisms, also inducing an increase in the level of Bax and Bim proapoptotic proteins and a decrease in the level of phosho-ERK1/2 kinase. The cytotoxicity of compound 5 was associated with an increase in the production of the hydrogen peroxide and the formation of DNA single-strand breaks. This compound 5 did not intercalate into a DNA molecule. Thus, the novel thiazole derivative (compound 5) proved to be a potential antiglioma drug that showed much higher cytotoxic action on human glioma cells compared with the effects of TMZ and Dox. Its cytotoxicity is associated with apoptosis induction, production of the reactive oxygen species, and formation of DNA single-strand breaks without significant DNA intercalation. Correspondence to Andriy Babsky, PhD, Biology Faculty, Ivan Franko National University of Lviv, Hrushevskoho Street, 4, 79005 Lviv, Ukraine Tel: +380 322 394 790; fax: +380 322 616 048; e-mail: andriy.babsky@lnu.edu.ua Received December 5, 2017 Accepted July 23, 2018 Copyright © 2018 Wolters Kluwer Health, Inc. All rights reserved.

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Long non-coding RNA H19 contributes to hypoxia-induced CPC injury by suppressing Sirt1 through miR-200a-3p

Abstract

Cardiomyocytes death is the chief obstacle that prevents the heart function recovery in myocardial infarction (MI)-induced heart failure (HF). Cardiac progenitor cells (CPCs)-based myocardial regeneration has provided a promising method for heart function recovery after MI. However, CPCs can easily lose their proliferation ability due to oxygen deficiency in infarcted myocardium. Revealing the underlying molecular mechanism for CPC proliferation is critical for effective MI therapy. In the present study, we set up a CoCl₂-induced hypoxia model in CPCs. We found that the expression of long non-coding RNA H19 was significantly down-regulated in CPCs after hypoxia stimuli. In addition, H19 suppression attenuated the proliferation and migration of CPCs under hypoxia stress. Furthermore, we discovered that H19 regulated the proliferation and migration of CPCs through mediating the expression of Sirt1 which is a target of miR-200a-3p under hypoxia. In conclusion, our findings demonstrate a novel regulatory mechanism for the proliferation and migration of CPCs under hypoxia condition, which provides useful information for the development of new therapeutic targets for MI therapy.

https://ift.tt/2nVWKIu

Tafa-2 plays an essential role in neuronal survival and neurobiological function in mice

Abstract

Tafa is a family of small secreted proteins with conserved cysteine residues and restricted expression in the brain. It is composed of five highly homologous genes referred to as Tafa-1 to -5. Among them, Tafa-2 is identified as one of the potential genes responsible for intellectual deficiency in a patient with mild mental retardation. To investigate the biological function of Tafa-2 in vivo, Tafa-2 knockout mice were generated. The mutant mice grew and developed normally but exhibited impairments in spatial learning and memory in Morris water maze test and impairments in short- and long-term memory in novel object recognition test, accompanied with increased level of anxiety-like behaviors in open-field test and elevated plus maze test, and decreased level of depression-like behaviors in forced-swim test and tail-suspension test. Further examinations revealed that Tafa-2 deficiency causes severe neuronal reduction and increased apoptosis in the brain of Tafa-2^−/− mice via downregulation of PI3K/Akt and MAPK/Erk pathways. Conformably, the expression levels of CREB target genes including BDNF, c-fos and NF1, and CBP were found to be reduced in the brain of Tafa-2^−/− mice. Taken together, our data indicate that Tafa-2 may function as a neurotrophic factor essential for neuronal survival and neurobiological functions.

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MitoCPR: a novel protective mechanism in response to mitochondrial protein import stress

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Trail armed oncolytic poxvirus suppresses lung cancer cell by inducing apoptosis

Abstract

Lung cancer has a high morbidity rate worldwide and is often resistant to therapy. Oncolytic virus therapy is a developing trend for cancer treatment. Thus, we constructed an oncolytic poxvirus carrying human trail gene that expresses a membrane-binding tumor necrosis factor and associated apoptosis-inducing ligand (TRAIL, Onco^pox-trail). We hypothesized that the expression of trail would increase the efficacy of the oncolytic poxvirus. The effect of the TRAIL protein depends on the death receptors on the surface of different cancer cells. The expression of death receptors in lung cancer cell lines was analyzed by western blot analysis. In vitro, the oncolytic poxvirus carrying the trail gene displayed a better cytotoxicity at the cell level in the lung cancer cell line than that carrying the Onco^pox-empty. TRAIL protein mainly induced apoptosis and inhibited necrosis. In vivo, two transplanted tumor models of human A549 lung cancer cells and mouse Lewis lung cancer cells were used to verify the anti-cancer effect of the oncolytic poxvirus carrying the trail gene. TUNEL staining results of the tumor histological sections also verified the anti-cancer effect. Similarly, through systemic administration of Onco^pox-trail, the oncolytic poxvirus also exhibited anti-cancer effect.

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Altered expressions of memory genes in food-entrained circadian rhythm

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