Molecular analysis demonstrates high prevalence of chloroquine resistance but no evidence of artemisinin resistance in Plasmodium falciparum in the Chittagong Hill Tracts of Bangladesh

Background Artemisinin resistance is present in the Greater Mekong region and poses a significant threat for current anti-malarial treatment guidelines in Bangladesh. The aim of this molecular study was to assess the current status of drug resistance in the Chittagong Hill Tracts of Bangladesh near the Myanmar border. Methods Samples were obtained from patients enrolled into a Clinical Trial (NCT02389374) conducted in Alikadam, Bandarban between August 2014 and January 2015. Plasmodium falciparum infections were confirmed by PCR and all P. falciparum positive isolates genotyped for the pfcrt K76T and pfmdr1 N86Y markers. The propeller region of the kelch 13 (k13) gene was sequenced from isolates from patients with delayed parasite clearance. Results In total, 130 P. falciparum isolates were available for analysis. The pfcrt mutation K76T, associated with chloroquine resistance was found in 81.5% (106/130) of cases and the pfmdr1 mutation N86Y in 13.9% (18/130) cases. No single nucleotide polymorphisms were observed in the k13 propeller region. Conclusion This study provides molecular evidence for the ongoing presence of chloroquine resistant P. falciparum in Bangladesh, but no evidence of mutations in the k13 propeller domain associated with artemisinin resistance. Monitoring for artemisinin susceptibility in Bangladesh is needed to ensure early detection and containment emerging anti-malarial resistance.</p

An Estimation of Hybrid Quantum Mechanical Molecular Mechanical Polarization Energies for Small Molecules Using Polarizable Force-Field Approaches

In this work, we report two polarizable molecular mechanics (polMM) force field models for estimating the polarization energy in hybrid quantum mechanical molecular mechanical (QM/MM) calculations. These two models, named the potential of atomic charges (PAC) and potential of atomic dipoles (PAD), are formulated from the ab initio quantum mechanical (QM) response kernels for the prediction of the QM density response to an external molecular mechanical (MM) environment (as described by external point charges). The PAC model is similar to fluctuating charge (FQ) models because the energy depends on external electrostatic potential values at QM atomic sites; the PAD energy depends on external electrostatic field values at QM atomic sites, resembling induced dipole (ID) models. To demonstrate their uses, we apply the PAC and PAD models to 12 small molecules, which are solvated by TIP3P water. The PAC model reproduces the QM/MM polarization energy with a R2 value of 0.71 for aniline (in 10,000 TIP3P water configurations) and 0.87 or higher for other 11 solute molecules, while the PAD model has a much better performance with R2 values of 0.98 or higher. The PAC model reproduces reference QM/MM hydration free energies for 12 solute molecules with a RMSD of 0.59 kcal/mol. The PAD model is even more accurate, with a much smaller RMSD of 0.12 kcal/mol, with respect to the reference. This suggests that polarization effects, including both local charge distortion and intramolecular charge transfer, can be well captured by induced dipole type models with proper parametrization

Kinetic Modeling of API Oxidation: 1. The AIBN/H2O/CH3OH Radical "Soup"

While different flavors of API stress testing systems have been used in experimental investigations for decades, the detailed kinetics of such systems as well as the chemical composition of prominent reactive species, specifically reactive oxygen species, are unknown.As a first step toward understanding and modeling API oxidation in stress testing, we investigated a typical radical "soup" solution an API is subject to during stress testing.Here we applied ab-initio electronic structure calculations to automatically generate and refine a detailed chemical kinetics model, taking a fresh look at API oxidation.We generated a detailed kinetic model for a representative azobisisobutyronitrile (AIBN)/H2O/CH3OH stress-testing system with varied co-solvent ratio (50%/50% -- 99.5%/0.5% vol. water/methanol) and for representative pH values (4--10) at 40oC stirred and open to the atmosphere.At acidic conditions hydroxymethyl alkoxyl is the dominant alkoxyl radical, and at basic conditions, for most studied initial methanol concentrations, cyanoisopropyl alkoxyl becomes the dominant alkoxyl radical, albeit at an overall lower concentration.At acidic conditions the levels of cyanoisopropyl peroxyl, hydroxymethyl peroxyl, and hydroperoxyl radicals are relatively high and comparable, while at both neutral and basic pH conditions, superoxide becomes the prominent radical in the system.The present work reveals the prominent species in a common model API stress testing system at various co-solvent and pH conditions, sets the stage for an in-depth quantitative API kinetic study, and demonstrates usage of novel software tools for automated chemical kinetic model generation and ab-initio refinement

Broadening the scope of binding free energy calculations using a Separated Topologies approach

Binding free energy calculations predict the potency of compounds to protein binding sites in a physically rigorous manner and see broad application in prioritizing the synthesis of novel drug candidates. Relative binding free energy calculations (RBFE) have emerged as an industry standard approach to achieve highly accurate rank-order predictions of the potency of related compounds; however, this approach requires that the ligands share a common scaffold and a common binding mode, restricting the methods\u27 domain of applicability. This is a critical limitation, since complex modifications to the ligands, especially core hopping, are very common in drug design. Absolute Binding Free Energy calculations (ABFE) are an alternate method, which can be used for ligands that are not congeneric. However, ABFE suffer from a known problem of long convergence times, due to the need to sample additional degrees of freedom within each system, such as sampling rearrangements necessary to open and close the binding site. Here, we report on an alternative method for RBFE, called Separated Topologies (SepTop), which overcomes the issues in both of the aforementioned methods, by enabling large scaffold changes between ligands with a convergence time comparable to traditional RBFE. Instead of only mutating atoms that vary between two ligands, this approach performs two absolute free energy calculations at the same time in opposite directions, one for each ligand. Defining the two ligands independently allows the comparison of binding of diverse ligands without the artificial constraints of identical poses or a suitable atom-atom mapping. This approach also avoids the need to sample the unbound state of the protein, making it more efficient than absolute binding free energy calculations. Here, we introduce an implementation of SepTop. We developed a general and efficient protocol for running SepTop, and we demonstrated the method on four diverse, pharmaceutically relevant systems. Here we report the performance of the method, as well as our practical insights into strengths, weaknesses, and challenges of applying this method in an industrial drug design setting. We find that the accuracy of the approach is sufficiently high to rank order ligands with an accuracy comparable to traditional RBFE calculations, while maintaining the additional flexibility of SepTop

Infrared spectroscopy of small protonated water clusters, H ⁺(H<inf>2</inf>O)<inf>n</inf> (n = 2-5): Isomers, argon tagging, and deuteration

Infrared photodissociation spectroscopy is reported for mass-selected H+(H2O)n complexes and their deuterated analogues with and without argon "tagging." H+(H 2O)nArm and D+(D2O) nArm complexes are studied in the O-H (O-D) stretching region for clusters in the small size range (n = 2-5). Upon infrared excitation, these clusters fragment by the loss of either argon atoms or one or more intact water molecules. Their excitation spectra show distinct bands in the region of the symmetric and asymmetric stretches of water and in the hydrogen bonding region. Experimental studies are complemented by computational work that explores the isomeric structures, their energetics and vibrational spectra. The addition of an argon atom is essential to obtain photodissociation for the n = 2-3 complexes, and specific inclusion of the argon in calculations is necessary to reproduce the measured spectra. For n = 3-5, spectra are obtained both with and without argon. The added argon atom allows selection of a subset of colder clusters and it increases the photodissociation yield. Although most of these clusters have more than one possible isomeric structure, the spectra measured correspond to a single isomer that is computed to be the most stable. Deuteration in these small cluster sizes leads to expected lowering of frequencies, but the spectra indicate the presence of the same single most-stable isomer for each cluster size. © 2010 American Chemical Society