Crystal Pharmatech (United States)

Abstract This paper discloses technical details of an early approach we explored for molecular glue induced ternary complex structure prediction using Feynman-Kac (FK) steering in the diffusion modules of AlphaFold3-type models. While we recently reported the successful results of our YDS-GlueFold model in predicting 8 novel molecular glue induced ternary structures validated by experimental data, here we share insights from our earlier FK steering experiments which were conducted prior to developing YDS-GlueFold. Notably, the unmodified AlphaFold3-type baseline failed to predict any of the eight complexes, underscoring the difficulty of sampling ternary conformations. By contrast, our FK-steered diffusion approach—FKSFold—successfully predicted 3 of the 8 cases in these early trials. We detail how we leveraged FK formalism to modify the diffusion process, using interface predicted TM-score (ipTM) as a reward function to guide sampling toward high-quality protein-protein interfaces in ternary complexes. This strategy of using ipTM as a reward function aligns well with the mechanism of action (MOA) of molecular glues, which function by inducing or stabilizing protein-protein interfaces. By implementing stochastic control theory within the diffusion module, our approach attempted to steer the reverse diffusion process toward conformations with superior interface quality in ternary protein-protein-small molecule complexes. In these early trials, FK steering successfully predicted 3 out of the 8 cases that were later all correctly predicted by YDS-GlueFold. This work provides insights into the evolution of our technical approaches toward molecular glue-induced complex prediction, documenting an important step in the development pathway that eventually led to YDS-GlueFold. Code is available athttps://github.com/YDS-Pharmatech/FKSFold.

Abstract Helicobacter pylori infection can cause a variety of gastrointestinal diseases. In severe cases, there is a risk of gastric cancer. Antibiotics are often used for clinical treatment of H. pylori infections. However, because of antibiotic overuse in recent years and the emergence of multidrug-resistant bacteria, there is an urgent need to develop new treatment methods and drugs to achieve complete eradication of H. pylori. Endolysins and holins encoded by bacterial viruses (i.e., phages) represent a promising avenue of investigation. These lyase-based antibacterial drugs act on the bacterial cell wall to destroy the bacteria. Currently, a type of endolysin that has been studied more frequently acts on the amide bond between peptidoglycans, and holin is a transmembrane protein that can punch holes in the cell membrane. However, as a Gram-negative bacterium, H. pylori possesses a layer of impermeable lipopolysaccharides on the cell wall, which prevents endolysin interaction with the cell wall. Therefore, we designed a genetic linkage between an endolysin enzyme and a holin enzyme with a section of polypeptides (e.g., polycations and hydrophobic peptides) that enable penetration of the outer membrane. These complexes were designated “artilysins” and were efficiently expressed in Escherichia coli. In vitro bacteriostasis experiments showed that the purified artilysins had strong bacteriostatic effects on H. pylori. In addition, the surface of H. pylori was perforated and destroyed, as confirmed by electron microscopy, which was proved that artilysins had bacteriolytic effect on H. pylori.

An efficient two-step method has been developed for the separation of β-cypermethrin stereoisomers by supercritical fluid chromatography with polysaccharide chiral stationary phases. With respect to retention, selectivity, and resolution of β-cypermethrin, the effects of chiral stationary phases, cosolvents, mobile phases, and column temperature have been studied in detail. Through a two-step separation, β-cypermethrin was firstly separated by using a cellulose-derived chiral stationary phase to obtain two stereoisomeric pairs, and further resolved on an amylose-based chiral stationary phase to produce four enantiopure stereoisomers. The electronic circular dichroism patterns of the first- and the third-eluted isomers in methanol solution showed the mirror image of each other in the wavelength range 200∼300nm, indicating that they were a pair of enantiomers. Moreover, the second- and the fourth-eluted isomers were also enantiomers. This proposed two-step strategy showed low solvent consumption, fast separation speed, and high-purity, which may provide an effective approach for preparative separation of compounds with multiple chiral centers and difficult-to-separate multicomponent samples.

Resolution of chiral compounds has played an important role in the pharmaceutical field, involving detailed studies of pharmacokinetics, physiological, toxicological, and metabolic activities of enantiomers. Herein, a reliable method by high-performance liquid chromatography (HPLC) coupled with an optical rotation detector was developed to separate isoborneol enantiomers. A cellulose tris(3, 5-dimethylphenylcarbamate)-coated chiral stationary phase showed the best separation performance for isoborneol enantiomers in the normal phase among four polysaccharide chiral packings. The effects of alcoholic modifiers and column temperature were studied in detail. Resolution of the isoborneol racemate displayed a downward trend along with an increase in the content of ethanol and column temperature, indicating that less ethanol in the mobile phase and lower temperature were favorable to this process. Moreover, two isoborneol enantiomers were obtained via a semipreparative chiral HPLC technique under optimum conditions, and further characterized by analytical HPLC, and experimental and calculated vibrational circular dichroism (VCD) spectroscopy, respectively. The solution VCD spectrum of the first-eluted component was consistent with the Density Functional Theory (DFT) calculated pattern based on the SSS configuration, indicating that this enantiomer should be (1S, 2S, 4S)-(+)-isoborneol. Briefly, these results have provided reliable information to establish a method for analysis, preparative separation, and absolute configuration of chiral compounds without typical chromophoric groups.

Enantiomeric pairs of triticonazole have been successfully separated by supercritical fluid chromatography coupled with a tris(3,5-dimethylphenylcarbamoyl) cellulose-coated chiral stationary phase in this work. The effects of co-solvent, dissolution solvent, flow rate, backpressure, and column temperature have been studied in detail with respect to retention, selectivity, and resolution of triticonazole. As indicated, the co-solvents mostly affected the retention factors and resolution, due to the different molecular structure and polarity. In addition, the dissolution solvents, namely, chloromethanes and alcohols, have been also important for enantioseparation because of the different interaction with stationary phase. Higher flow rate and backpressure led to faster elution of the triticonazole molecules, and the change of column temperature showed slight effect on the resolution of triticonazole racemate. Moreover, a comparative separation experiment between supercritical fluid chromatography and high performance liquid chromatography revealed that chiral supercritical fluid chromatography gave the 3.5 times value of Rs /tR2 than high performance liquid chromatography, which demonstrated that supercritical fluid chromatography had much higher separation efficiency.

Amorphous solid dispersions (ASDs) are being used with increasing frequency for poorly soluble pharmaceutical compounds in development. These systems consist of an amorphous active pharmaceutical ingredient stabilized by a polymer to produce a system with improved physical and solution stability. ASDs are commonly considered as a means of improving the apparent solubility of an active pharmaceutical ingredient. This review will discuss methods of preparation and characterization of ASDs with an emphasis on understanding and predicting stability. Theoretical understanding of supersaturation and predicting in vivo performance will be stressed. Additionally, a summary of preclinical and clinical development efforts will be presented to give the reader an understanding of risks and key pitfalls when developing an ASD.

Computer assisted, or in silico, drug discovery approaches play an important role in the search for small molecule hits and leads. These include structure- and ligand-based methods, as well as data mining and QSAR. They are used to analyze and predict ligand-receptor binding, as well as pharmacokinentic profiles of compounds with therapeutic potential. A diversity of offerings is publically/commercially available for performing these tasks. Each offering comprises select combinations of in silico methods. Efficient in silico drug discovery requires effective use of combinations of these tools. Unfortunately, no single vendor offering integrates all in silico capabilities. Typically, different vendors offer different "flavors" of the same method and specific "flavors" have associated strengths and weaknesses. Furthermore, significant inter-vendor format incompatibilities exist. Consequently, extensive scripting as well as manual intervention is required in order to overcome disparate data formats. In this article, we introduce the architecture and implementation of a highly efficient, and automated in silico drug discovery engine that integrates multi-vendor software. A single graphical user interface enables the user to 'Click & Configure' modeling tools and permits 'Mix & Matching' components from various vendors. It deploys a 'Divide & Conquer' strategy to marshal the resources of a multi-node compute cluster for compute-intensive tasks. This basic framework in performing in silico modeling activities (work-flow automation) envisions the integration of structure-based, ligand-based, and other modes of in silico drug discovery.

Abstract The relationship between point efficiencies and Murphree tray efficiencies was studied for systems where reactions take place in the liquid within a tray column. For nonreactive systems, several well‐known mixing models are available for predicting Murphree tray efficiency from point efficiencies. For reactive systems, the magnitude and nature of the effect of mixing on tray efficiency are different. An analytical representation is developed of the mixed‐pool and plug‐flow mixing models for reactive systems with first‐order kinetics, and there are different regimes, depending on the reaction kinetics. For each of these regimes, the effect of mixing on tray efficiency differs in magnitude and arises from different mechanisms. The resulting tray efficiencies predicted for reactive systems using the methods may differ by an order of magnitude or more from the values predicted when standard methods for nonreactive trays are applied to reactive trays.