Exploration of Piperidinols as Potential Antitubercular Agents

Novel drugs to treat tuberculosis are required and the identification of potential targets is important. Piperidinols have been identified as potential antimycobacterial agents (MIC < 5 μg/mL), which also inhibit mycobacterial arylamine N-acetyltransferase (NAT), an enzyme essential for mycobacterial survival inside macrophages. The NAT inhibition involves a prodrug-like mechanism in which activation leads to the formation of bioactive phenyl vinyl ketone (PVK). The PVK fragment selectively forms an adduct with the cysteine residue in the active site. Time dependent inhibition of the NAT enzyme from Mycobacterium marinum (M. marinum) demonstrates a covalent binding mechanism for all inhibitory piperidinol analogues. The structure activity relationship highlights the importance of halide substitution on the piperidinol benzene ring. The structures of the NAT enzymes from M. marinum and M. tuberculosis, although 74% identical, have different residues in their active site clefts and allow the effects of amino acid substitutions to be assessed in understanding inhibitory potency. In addition, we have used the piperidinol 3-dimensional shape and electrostatic properties to identify two additional distinct chemical scaffolds as inhibitors of NAT. While one of the scaffolds has anti-tubercular activity, both inhibit NAT but through a non-covalent mechanism

“MedChemVR”: A Virtual Reality Game to Enhance Medicinal Chemistry Education

Medicinal chemistry (MC) is an indispensable component of the pharmacy curriculum. The pharmacists’ unique knowledge of a medicine’s chemistry enhances their understanding of the pharmacological activity, manufacturing, storage, use, supply, and handling of drugs. However, chemistry is a challenging subject for both teaching and learning. These challenges are typically caused by the inability of students to construct a mental image of the three-dimensional (3D) structure of a drug molecule from its two-dimensional presentations. This study explores a prototype virtual reality (VR) gamification option, as an educational tool developed to aid the learning process and to improve the delivery of the MC subject to students. The developed system is evaluated by a cohort of 41 students. The analysis of the results was encouraging and provided invaluable feedback for the future development of the proposed system

“MedChemVR”: A Virtual Reality Game to Enhance Medicinal Chemistry Education

Medicinal chemistry (MC) is an indispensable component of the pharmacy curriculum. The pharmacists’ unique knowledge of a medicine’s chemistry enhances their understanding of the pharmacological activity, manufacturing, storage, use, supply, and handling of drugs. However, chemistry is a challenging subject for both teaching and learning. These challenges are typically caused by the inability of students to construct a mental image of the three-dimensional (3D) structure of a drug molecule from its two-dimensional presentations. This study explores a prototype virtual reality (VR) gamification option, as an educational tool developed to aid the learning process and to improve the delivery of the MC subject to students. The developed system is evaluated by a cohort of 41 students. The analysis of the results was encouraging and provided invaluable feedback for the future development of the proposed system

Identifying the Characteristics of Virtual Reality Gamification for Complex Educational Topics

Multidisciplinary topics in education pose a major challenge for traditional learning and teaching methods. Such topics can deter students from selecting particular courses or hinder their study progress. This study focused on the subject of medicinal chemistry, which is a discipline combining medicine and chemistry. This combination of applied and basic science creates a complex field of education that is challenging to both teach and learn. Chemical and pharmacological principles are typically presented in 2D molecular structures and, recently, 3D molecular models have been utilized to improve the visualization of chemical compounds and their chemical interactions. Contemporary studies have presented Virtual Reality (VR) as an alternative method for improving the learning and teaching of multidisciplinary specialties such as this. However, current educational efforts employing VR offer limited interactivity and a traditional teaching method previously presented in 2D. This reduces students’ interest and concentration in the taught subjects. This paper presents the development rationale of a novel VR educational application based on the evaluation of the user requirements by 405 pharmacy undergraduate students. The results informed the development and preliminary evaluation of a proposed VR serious game application, which was deployed in a real-life class environment and evaluated in contrast to traditional teaching methods by 15 students. The derived results confirmed the advantages of VR technology as a learning and teaching tool, in addition to the end-users’ willingness to adopt VR systems as a learning aid

Identifying the Characteristics of Virtual Reality Gamification for Complex Educational Topics

Multidisciplinary topics in education pose a major challenge for traditional learning and teaching methods. Such topics can deter students from selecting particular courses or hinder their study progress. This study focused on the subject of medicinal chemistry, which is a discipline combining medicine and chemistry. This combination of applied and basic science creates a complex field of education that is challenging to both teach and learn. Chemical and pharmacological principles are typically presented in 2D molecular structures and, recently, 3D molecular models have been utilized to improve the visualization of chemical compounds and their chemical interactions. Contemporary studies have presented Virtual Reality (VR) as an alternative method for improving the learning and teaching of multidisciplinary specialties such as this. However, current educational efforts employing VR offer limited interactivity and a traditional teaching method previously presented in 2D. This reduces students’ interest and concentration in the taught subjects. This paper presents the development rationale of a novel VR educational application based on the evaluation of the user requirements by 405 pharmacy undergraduate students. The results informed the development and preliminary evaluation of a proposed VR serious game application, which was deployed in a real-life class environment and evaluated in contrast to traditional teaching methods by 15 students. The derived results confirmed the advantages of VR technology as a learning and teaching tool, in addition to the end-users’ willingness to adopt VR systems as a learning aid

Piperidinols that show anti-tubercular activity as inhibitors of arylamine N-acetyltransferase: an essential enzyme for mycobacterial survival inside macrophages

Latent M. tuberculosis infection presents one of the major obstacles in the global eradication of tuberculosis (TB). Cholesterol plays a critical role in the persistence of M. tuberculosis within the macrophage during latent infection. Catabolism of cholesterol contributes to the pool of propionyl-CoA, a precursor that is incorporated into cell-wall lipids. Arylamine N-acetyltransferase (NAT) is encoded within a gene cluster that is involved in the cholesterol sterol-ring degradation and is essential for intracellular survival. The ability of the NAT from M. tuberculosis (TBNAT) to utilise propionyl-CoA links it to the cholesterol-catabolism pathway. Deleting the nat gene or inhibiting the NAT enzyme prevents intracellular survival and results in depletion of cell-wall lipids. TBNAT has been investigated as a potential target for TB therapies. From a previous high-throughput screen, 3-benzoyl-4-phenyl-1-methylpiperidinol was identified as a selective inhibitor of prokaryotic NAT that exhibited antimycobacterial activity. The compound resulted in time-dependent irreversible inhibition of the NAT activity when tested against NAT from M. marinum (MMNAT). To further evaluate the antimycobacterial activity and the NAT inhibition of this compound, four piperidinol analogues were tested. All five compounds exert potent antimycobacterial activity against M. tuberculosis with MIC values of 2.3-16.9 µM. Treatment of the MMNAT enzyme with this set of inhibitors resulted in an irreversible time-dependent inhibition of NAT activity. Here we investigate the mechanism of NAT inhibition by studying protein-ligand interactions using mass spectrometry in combination with enzyme analysis and structure determination. We propose a covalent mechanism of NAT inhibition that involves the formation of a reactive intermediate and selective cysteine residue modification. These piperidinols present a unique class of antimycobacterial compounds that have a novel mode of action different from known anti-tubercular drugs

Arylamine N-Acetyltransferases from mycobacteria: investigations of a potential yarget for anti-tubercular therapy

Reactivation of latent infection is the major cause of tuberculosis (TB). Cholesterol is a critical carbon source during latent infection. Catabolism of cholesterol contributes to the pool of propionyl-CoA, a precursor that is incorporated into cell-wall lipids. Arylamine N-acetyltransferase (NAT) is encoded within a gene cluster that is involved in the sterol-ring degradation and is essential for intracellular survival. NAT from M. tuberculosis (TBNAT) can utilise propionyl-CoA and therefore was proposed as a target for TB-drug development. Deleting the nat gene or inhibiting the NAT enzyme prevents intracellular survival and results in depletion of cell-wall lipids. NAT inhibitors, including the piperidinol class, were identified by high-throughput screening. The insolubility of recombinant TBNAT has been a major limitation in pursuing it as a drug target. Subcloning tbnat into a pVLT31 vector resulted in a yield of 6-16 mg/litre-bacterial-culture of pure-soluble recombinant TBNAT. The increased yield allowed for extensive screening for crystallisation conditions. However, since a structure was not obtained, the model NAT from M. marinum (MMNAT) was employed to further understand NAT as a target. Screening against a panel of Acyl-CoA cofactors showed that MMNAT can also utilise propionyl-CoA. The MMNAT structure in complex with the high affinity substrate hydralazine was determined (2.1 Å) and the architecture of the arylamine pocket was delineated. A novel mechanism for the acetylation reaction of hydralazine has emerged. It is proposed that the acetyl group is transferred from acetyl-CoA to the heterocyclic aromatic nitrogen of hydralazine, which explains the immediate cyclisation of the acetylated metabolite into an N-methyltriazolophthalazine. By employing mass spectroscopy, enzyme assays, computational docking and structural studies, a covalent mechanism of inhibition by the piperidinol class was established, and the inhibitor-binding pocket was identified. Inhibitors with new scaffolds were identified using the in silico 3D-shape screening and thermal shift assay.</p

Mycobacterial arylamine N-acetyltrasferases and tuberculosis

An N-acetyltransferase (NAT) enzyme has been identified in many mycobacterial species. It has been suggested that the enzyme plays a role in cholesterol degradation and maintaining the homeostasis of acetyl-Coenzyme A, a central metabolite in lipid metabolism. The nat genes from Mycobacterium tuberculosis and Mycobacterium bovis bacillus Calmette–Guerin (BCG) are identical and are encoded in highly similar operons in both organisms (the nat operon). The other genes of the operon are involved in catabolism of the sterol rings of cholesterol. The gene cluster is essential for the survival of M. bovis BCG within macrophages, where cholesterol is likely to be the fuel for intracellular survival. Strains missing these genes are unable to grow on minimal medium with cholesterol, although they retain the ability to grow on rich media