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Pharmacoinformatics Based Elucidation and Designing of Potential Inhibitors Against Plasmodium Falciparum to Target Importin Α/Β Mediated Nuclear Importation

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Pharmacoinformatics Based Elucidation and Designing of Potential Inhibitors Against Plasmodium Falciparum to Target Importin Α/Β Mediated Nuclear Importation bioRxiv preprint doi: https://doi.org/10.1101/2020.12.29.424688; this version posted December 30, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. Pharmacoinformatics based elucidation and designing of potential inhibitors against Plasmodium falciparum to target importin α/β mediated nuclear importation Arafat Rahman Oany1,2*, Tahmina Pervin3, Mohammad Ali Moni 4, 5, 6* 1 Department of Biotechnology and Genetic Engineering, Faculty of Life Science, Mawlana Bhashani Science and Technology University, Tangail, Bangladesh. 2 Aristopharma Limited, Bangladesh 3 Biotechnology and Genetic Engineering Discipline, Life Science School, Khulna University, Khulna, Bangladesh. 4Department of Computer Science & Engineering, Green University Bangladesh 5Department of Computer Science & Engineering, Pabna University of Science & Technology, Bangladesh 6WHO Collaborating Centre on eHealth, UNSW Digital Health, School of Public Health and Community Medicine, Faculty of Medicine, UNSW Sydney, Australia * Corresponding Author Mohammad Ali Moni, PhD (Cantab) Professor, Green University of Bangladesh Professor, Pabna University of Science & Technology Senior Fellow, WHO Collaborating Centre on eHealth, School of Public Health and Community Medicine UNSW Sydney, Australia E mail: [email protected] [email protected] & Arafat Rahman Oany (http://orcid.org/0000-0003-2228-457X) Department of Biotechnology and Genetic Engineering, Faculty of Life Science, Mawlana Bhashani Science and Technology University, Tangail-1902, Bangladesh Email: [email protected] Page 1 of 17 bioRxiv preprint doi: https://doi.org/10.1101/2020.12.29.424688; this version posted December 30, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. Abstract: Plasmodium falciparum, the prime causative agent of malaria, is responsible for 4, 05,000 deaths per year and fatality rates are higher among the children aged below 5 years. The emerging distribution of the multi-drug resistant P. falciparum becomes a worldwide concern, so the identification of unique targets and novel inhibitors is a prime need now. In the present study, we have employed pharmacoinformatics approaches to analyze 265 lead-like compounds from PubChem databases for virtual screening. Thereafter, 15 lead-like compounds were docked within the active side pocket of importin alpha. Comparative ligand properties and absorption, distribution, metabolism, excretion, and toxicity (ADMET) profile were also assessed. Finally, a novel inhibitor was designed and assessed computationally for its efficacy. From the comparative analysis we have found that our screened compounds possess better results than the existing lead ivermectin; having the highest binding energy of -15.6 kcal/mol, whereas ivermectin has -12.4kcal/mol. The novel lead compound possessed more fascinating output without deviating any of the rules of Lipinski. It also possessed higher bioavailability and the drug-likeness score of 0.55 and 0.71, respectively compared to ivermectin. Furthermore, the binding study was confirmed by molecular dynamics simulation over 25 ns by evaluating the stability of the complex. Finally, all the screened compounds and the novel compound showed promising ADMET properties likewise. To end, we hope that our proposed screened compounds, as well as the novel compound, might give some advances to treat malaria efficiently in vitro and in vivo. Page 2 of 17 bioRxiv preprint doi: https://doi.org/10.1101/2020.12.29.424688; this version posted December 30, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. Keywords: ADMET, Molecular docking, Importin α/β, Drug-resistance, Drug-designing 1.0 Introduction: Malaria, a disease caused by the infection through protozoan parasites of the genus Plasmodium, disseminates to humans by female anopheles mosquitoes and is responsible for about 228 million medical cases in recent years. It is also accountable for nearly half a million deaths, and the severity rates are higher among the children (Carter & Mendis 2002), (Snow et al. 2005). Among the five species of malaria, P. falciparum is the deadliest and responsible for 99.7% of malaria cases in the African region. That is considered as a hot-spot of malaria infection, having 213 million or 93% cases in that region while the total worldwide cases were 228 millions.(Organisation 2018). The ever-rising distribution of the multi-drug resistant P. falciparum now becomes a global concern and is mediated by the resistance to conventional antimalarials, chloroquine and artemisinin(Ashley et al. 2014; Takala-Harrison et al. 2015; Coppée et al. 2020). As a consequence, there is a pressing necessity to develop novel antimalarial agents that are effective against drug-resistant malarial parasites and might be working on different or unique targets and also structurally distinct from existing antimalarial agents (Peters et al. 1990; Palmer et al. 1993) . The unambiguous contrasts in gene control components standing in the nucleus between P falciparum and the human could prompt new potential drug targets for antimalarial drug development(Wittayacom et al. 2010). Page 3 of 17 bioRxiv preprint doi: https://doi.org/10.1101/2020.12.29.424688; this version posted December 30, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. Nuclear transport also shows a foremost role in disease conditions such as oncogenesis and viral illness (Hogarth et al. 2006; Yoneda-Kato et al. 2008; Ao et al. 2010). The periphery between nuclear and cytoplasmic compartments is infiltrated by NPC that permit swapping of macromolecules between the nucleus and cytoplasm. On the other hand, nuclear import facilitates higher molecular mass proteins through a selective and energy-dependent manner with the aid of NLS and NES. Importin family proteins identify these targeting sequences that expedite the nuclear import of their cargo proteins mediated by the formation of importin α/β heterodimer(Fontes et al. 2000). Precise inhibitors of nuclear import symbolize valuable tools for scrutinizing nuclear vehicle technology in forthcoming research. Leptomycin B, an antifungal antibiotic, has already been tested and is a well-characterized compound for inhibiting nuclear transport. Ratjadones and some small-molecule inhibitors have also been suggested for its efficacy to inhibit nuclear importation(Kudo et al. 1999; Köster et al. 2003). Ivermectin has also been suggested as a promising candidate for inhibiting nuclear import at a precisely lower concentration and thereby killing the pathogens. It is also very selective to importin α/β mediated import and having no effects on other importin family proteins(Chaccour et al. 2013; Panchal et al. 2014). In the present study, we have tried to design a novel inhibitor of this unique target by using pharmacoinformatics, bioinformatics techniques for drug designing, approaches to combat against deadly malaria. Page 4 of 17 bioRxiv preprint doi: https://doi.org/10.1101/2020.12.29.424688; this version posted December 30, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. 2.0 Materials and Methods: The overall proposed molecular mechanism for drug designing is shown in Figure 1. 2.1 Sequence retrieval: The NCBI database was initially explored to find out P. falciparum importin α and β proteins (Wheeler et al. 2007). Protein importin α (gi|29501524|) and β (gi|29501526|) were selected for the study. The sequences were then stored as a FASTA format for further analysis. 2.2 Homology modelling, structural refinement and model validation: Homology models of the targets were done by MODELLER v9 (Sali et al. 1995),and top-scoring template was selected for the model construction. The templates used for the modeling were 4TNM_A and 3W3W_A for importin α and β, respectively. The probability and the identity scores for importin α were 100% and 49% and for β- 100% and 26%, respectively. The structural refinement of the models were done by GalaxyWEB server (Ko et al. 2012). Then the predicted models were employed for the validation through the PROCHECK (Laskowski et al. 1996)tool of the SWISS-MODEL Workspace (Arnold et al. 2006). Additionally, the ProSA-web server (Wiederstein & Sippl 2007) was also utilized for assessing the model quality through Z-score measurement. 2.3 Electrostatic potential analysis and energy minimization: The electrostatic potential of the predicted models was built by APBS (Unni et al. 2011)server through transforming .pdb to .pqr file and the visualization was done by PyMOL (version 1.3) molecular graphics system (DeLano 2002). For further securing the validation of the predicted Page 5 of 17 bioRxiv preprint doi: https://doi.org/10.1101/2020.12.29.424688; this version posted December 30, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. models, the energy minimizations were also done. The GROMOS 96 (Scott et al. 1999) force field was used for the minimization of the structure and validated by the “Ramachandran Plot” statistic by using the SWISS-MODEL Workspace (Arnold et al. 2006).
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