Discovery of potent, novel, non-toxic anti-malarial compounds via quantum modelling, virtual screening and in vitro experimental validation

<p>Abstract</p> <p>Background</p> <p>Developing resistance towards existing anti-malarial therapies emphasize the urgent need for new therapeutic options. Additionally, many malaria drugs in use today have high toxicity and low therapeutic indices. Gradient Biomodeling, LLC has developed a quantum-model search technology that uses quantum similarity and does not depend explicitly on chemical structure, as molecules are rigorously described in fundamental quantum attributes related to individual pharmacological properties. Therapeutic activity, as well as toxicity and other essential properties can be analysed and optimized simultaneously, independently of one another. Such methodology is suitable for a search of novel, non-toxic, active anti-malarial compounds.</p> <p>Methods</p> <p>A set of innovative algorithms is used for the fast calculation and interpretation of electron-density attributes of molecular structures at the quantum level for rapid discovery of prospective pharmaceuticals. Potency and efficacy, as well as additional physicochemical, metabolic, pharmacokinetic, safety, permeability and other properties were characterized by the procedure. Once quantum models are developed and experimentally validated, the methodology provides a straightforward implementation for lead discovery, compound optimizzation and <it>de novo </it>molecular design.</p> <p>Results</p> <p>Starting with a diverse training set of 26 well-known anti-malarial agents combined with 1730 moderately active and inactive molecules, novel compounds that have strong anti-malarial activity, low cytotoxicity and structural dissimilarity from the training set were discovered and experimentally validated. Twelve compounds were identified <it>in silico </it>and tested <it>in vitro</it>; eight of them showed anti-malarial activity (IC50 ≤ 10 μM), with six being very effective (IC50 ≤ 1 μM), and four exhibiting low nanomolar potency. The most active compounds were also tested for mammalian cytotoxicity and found to be non-toxic, with a therapeutic index of more than 6,900 for the most active compound.</p> <p>Conclusions</p> <p>Gradient's metric modelling approach and electron-density molecular representations can be powerful tools in the discovery and design of novel anti-malarial compounds. Since the quantum models are agnostic of the particular biological target, the technology can account for different mechanisms of action and be used for <it>de novo </it>design of small molecules with activity against not only the asexual phase of the malaria parasite, but also against the liver stage of the parasite development, which may lead to true causal prophylaxis.</p

Performance of Sulfide-Driven Fuel Cell Aerated by Venturi Tube Ejector

Hydrogen sulfide is frequently met in natural waters, like mineral springs, but mostly it is found in marine water with low renewal rate. The Black Sea has extremely high hydrogen sulfide content. It can be utilized in different ways, but the most promising one is direct conversion into electricity. This result can be attained by a sulfide-driven fuel cell (SDFC), converting sulfide to sulfate thus releasing electric energy up to 24 GJ/t. One of the most important problems is the mass transfer limitation on oxygen transfer in the cathode space of the fuel cell. This problem can be solved using a gas diffusion electrode or highly efficient saturation by oxygen in an ejector of the Venturi tube type. This work presents experimental data in laboratory-scale SDFC for sulfide conversion into sulfate, sulfite and polysulfide releasing different amounts of electric energy. Two types of aeration are tested: direct air blow and Venturi-tube ejector. Besides pure graphite, two catalysts, i.e., cobalt spinel and zirconia-doped graphite were tested as anodes. Experiments were carried out at initial sulfide concentrations from 50 to 300 mg/L. Sulfate, sulfite and thiosulfate ions were detected in the outlet solutions from the fuel cell. The electrochemical results show good agreement with the chemical analyses. Most of the results show attained high efficiencies of the fuel cell, i.e., up to 80%. The practical applications of this method can be extended for other purposes, like treatment of polluted water together with utilization as energy.</jats:p

Discovery of Novel Liver-Stage Antimalarials through Quantum Similarity.

Without quantum theory any understanding of molecular interactions is incomplete. In principal, chemistry, and even biology, can be fully derived from non-relativistic quantum mechanics. In practice, conventional quantum chemical calculations are computationally too intensive and time consuming to be useful for drug discovery on more than a limited basis. A previously described, original, quantum-based computational process for drug discovery and design bridges this gap between theory and practice, and allows the application of quantum methods to large-scale in silico identification of active compounds. Here, we show the results of this quantum-similarity approach applied to the discovery of novel liver-stage antimalarials. Testing of only five of the model-predicted compounds in vitro and in vivo hepatic stage drug inhibition assays with P. berghei identified four novel chemical structures representing three separate quantum classes of liver-stage antimalarials. All four compounds inhibited liver-stage Plasmodium as a single oral dose in the quantitative PCR mouse liver-stage sporozoites-challenge model. One of the newly identified compounds, cethromycin [ABT-773], a macrolide-quinoline hybrid, is a drug with an extensive (over 5,000 people) safety profile warranting its exploitation as a new weapon for the current effort of malaria eradication. The results of our molecular modeling exceed current state-of-the-art computational methods. Drug discovery through quantum similarity is data-driven, agnostic to any particular target or disease process that can evaluate multiple phenotypic, target-specific, or co-crystal structural data. This allows the incorporation of additional pharmacological requirements, as well as rapid exploration of novel chemical spaces for therapeutic applications

<i>in vitro</i> inhibition of liver stage malaria.

<p><b>A.</b> Three of the new compounds indicated 30%, 96% and 55% inhibition while cethromycin alone at 20 μM had 54% inhibition. The individual components of cethromycin-quinoline and erythromycin, were inactive. Error is average of duplicate wells performed in two independent biologic experiments with different preparations of sporozoites. <b>B</b>. Microscopic image taken at 20x magnification of near 95% inhibition by T5531873. 50,000 Hepa1-6 cells were seeded in each well 24 hrs prior to infection with ~60,000 <i>P</i>. <i>berghei</i> sporozoites. The 2E6 anti-HSP70 antibody was used for immunofluorescent numeration of infected cells.</p

Matching quantum scores of input and output molecules from monesin and telithromycin.

<p>Matching quantum scores of input and output molecules from monesin and telithromycin.</p

Liver-stage Quantum Components.

<p>Quantum similarity of <b>A.</b> GNF-Pf-1498 and the quinoline-macrolide hybrid <b>B.</b> cethromycin that is related to CHEMBL440116 as well as chemical structures of <b>C.</b> GNF-Pf-1498 and <b>D.</b> cethromycin.</p

Matching quantum scores of input and output molecules from cyclosporine.

<p>Matching quantum scores of input and output molecules from cyclosporine.</p

Compound Structures from Quantum modeling.

<p>Chemical structures of CHEMBL440116 and four other identified molecules which were acquired and tested.</p

<i>in vivo</i> inhibition.

<p>10,000 sporozoites were inoculated by tailvein injection and mice were sacrificed 40 hours later, livers were harvested, placed in RNAzol and parasite levels determined by realtime PCR from cDNA from reverse transcription. Relative fluorescent units were compared to control to determine percent inhibition. Cethromycin CET was administered only once while the other drugs were given twice 24 hours apart from each dose. Two drugs related to CET, quinolone (QN) and erythromycin (ERY), had only marginal effect on parasite growth. CET’s effectiveness increased with dosage, reaching 60% reduction at 50 mg/kg. CET was also able to eliminate parasite infection when combined with low dose of PQ. All three novel compounds (T0507-9950, T5531873, T0510-7064) demonstrated significant inhibitory effect on parasite proliferation. Error is standard error of mean of three mice with real time PCR performed in duplicate for transcript levels in each mouse. B. Actual log values of relative fluorescent units from controls, cethromycin 12 mg/kg, primaquine 15 mg/kg and in combination are depicted. The combination has a 4 fold drop in relative fluorescent units compared to primaquine.</p

Summary of quantum scores with molecule identifications and reference molecules for scoring.

<p>Summary of quantum scores with molecule identifications and reference molecules for scoring.</p