Low molecular weight compounds from mushrooms as potential Bcl-2 inhibitors: docking and virtual screening studies

Mestrado com dupla diplomação com o Institut Superieur de Biotechnologie de MonastirMushrooms have the ability to promote apoptosis in tumor cell lines, but the mechanism of action is not quite well understood. Inhibition of the interaction between Bcl-2 and pro-apoptotic proteins could be an important step that leads to apoptosis. Therefore, the discovery of compounds with the ability to inhibit Bcl-2 is an ongoing research topic in drug discovery. In this study, we started by analyzing Bcl-2 experimental structures that are currently available in Protein Data Bank database. After analysis of the more relevant Bcl-2 structures, 4 were finally selected. An analysis of the best docking methodology was then performed using a cross-docking and re-docking approach while testing 2 docking softwares: AutoDock 4 and AutoDock Vina. Autodock4 provided the best docking results and was selected to perform a virtual screening study applied to a dataset of 40 Low Molecular Weight (LMW) compounds present in mushrooms, using the selected Bcl-2 structures as target. Results suggest that steroid are the more promising family, among the analyzed compounds, and may have the ability to interact with Bcl-2 and this way promoting tumor apoptosis. The steroids that presented lowest estimated binding energy (ΔG) were: Ganodermanondiol, Cerevisterol, Ganoderic Acid X and Lucidenic Lactone; with estimated ΔG values between -8,45 and -8,23 Kcal/mol. A detailed analysis of the docked conformation of these 4 top ranked LMW compounds was also performed and illustrates a plausible interaction between the 4 top raked steroids and Bcl-2, thus substantiating the accuracy of the predicted docked poses. Therefore, tumoral apoptosis promoted by mushroom might be related to Bcl-2 inhibition mediated by steroid family of compounds.Os cogumelos apresentam a capacidade de promover a apoptose em linhas células tumorais, No entanto o seu mecanismo de ação não é completamente conhecido. A inibição da interação entre Bcl-2 e proteínas pro-apoptóticas pode ser um passo importante na iniciação do processo de apoptose tumoral. Por essa razão, a descoberta de compostos que inibam a proteína Bcl-2 é uma área importante na descoberta de novos fármacos antitumorais. Neste estudo, começou-se por analisar as estruturas experimentais de Bcl-2 atualmente presentes na base de estruturas Protein Data Bank. Após análise das estruturas de Bcl-2 mais relevantes, 4 foram escolhidas. Um estudo de “cross-docking” e “re-docking” foi então realizado para escolher a metodologia de “docking” mais adequada. Testaram-se 2 softwares, o AutoDock 4 e o AutoDock Vina, e verificou-se que o AutoDock 4 apresentava melhores resultados, tendo sido o selecionado para realizar os ensaios de “screening” virtual dos 40 compostos de baixo peso molecular presentes em cogumelos, utilizando as 4 estruturas selecionadas. Os resultados obtidos sugerem que os esteroides são a família de compostos mais prometedores de entre as famílias de compostos estudadas. Os esteroides que apresentaram valores de energia de ligação (ΔG) mais baixos foram: Ganodermanondiol, Cerevisterol, Ácido Ganoderico X and Lactona Lucidénica, com valores de ΔG estimado entre -8,45 e -8,23 Kcal/mol. Uma análise detalhada da conformação de ligação foi também realizada dos 4 melhores compostos de baixo peso molecular melhor classificados. Esta análise demonstra um modo de interação plausível entre os compostos e a estrutura da Bcl-2, consubstanciando a eficácia dos resultados obtidos por “docking”. Conclui-se que o processo inibição de apoptose tumoral observada em cogumelos pode estar relacionado com a inibição da Bcl-2 por esteroides presentes nos cogumelos

Investigating mushroom LMW compounds as potential Bcl-2 inhibitors: docking studies using AutoDock4

The B cell CLL/lymphoma-2 (Bcl-2) family is functionally classified as either anti-apoptotic or pro-apoptotic, and the regulation of its interactions dictates survival or commitment to apoptosis. Bcl-2 family is also implicated in a wide range of diseases. In some types of cancers, including lymphomas and epithelial cancers, protein overexpression of anti-apoptotic Bcl-2 family, such as the Bcl-2 protein is indicative of cancer in an advanced stage, with a poor prognosis and resistant to chemotherapy [1]. Several reports indicate that mushrooms have the ability to promote apoptosis in tumour cell lines, but the mechanism of action is not fully understood. Inhibition of the interaction between Bcl-2 (anti-apoptotic protein) and proapoptotic proteins could be an important step in the mechanism of mushroom induced apoptosis. Therefore, the discovery of compounds with the capacity to inhibit Bcl-2 is an ongoing research topic on cancer therapy. In this work, docking studies were performed using a dataset of 40 low molecular weight (LMW) compounds present in mushrooms. The docking software AutoDock 4 was used and docking studies were performed using 5 selected Bcl-2 crystal structures as targets. Compounds with the lowest predicted binding energy (predΔG) are expected to be the more potent inhibitors. Among the tested compounds, steroids presented the lowest predΔG with several exhibiting values below -9 kcal/mol. The results are corroborated by several reports that state that steroids induce apoptosis in several tumor cells. It is thus feasible that they might act by preventing Bcl-2 from forming complexes with the respective proapoptotic protein interaction partners, namely Bak, Bax, and Bim. Moreover, previous studies on our research group demonstrated that 48 h treatment of MCF-7 cells (breast carcinoma) with Suillus collinitus methanolic extract caused a decrease in Bcl-2, highlighting the antitumor potential of this mushroom species [2]. In conclusion, the process of apoptosis promoted by mushroom extracts may be related to the inhibition of Bcl-2 by the steroid derivatives herein studied. However, further studies are needed to confirm this hypothesis