Analytical Separation of Closantel Enantiomers by HPLC

Closantel is an antiparasitic drug marketed in a racemic form with one chiral center. It is meaningful to develop a method for separating and analyzing the closantel enantiomers. In this work, two enantiomeric separation methods of closantel were explored by normal-phase high-performance liquid chromatography. The influences of the chiral stationary phase (CSP) structure, the mobile phase composition, the nature and proportion of different mobile phase modifiers (alcohols and acids), and the column temperature on the enantiomeric separation of closantel were investigated in detail. The two enantiomers were successfully separated on the novel CSP of isopropyl derivatives of cyclofructan 6 and n-hexane-isopropanol-trifluoroacetic acid (97:3:0.1, v/v/v) as a mobile phase with a resolution (Rs) of about 2.48. The enantiomers were also well separated on the CSP of tris-carbamates of amylose with a higher Rs (about 3.79) when a mixture of n-hexane-isopropanol-trifluoroacetic acid (55:45:0.1, v/v/v) was used as mobile phase. Thus, the proposed separation methods can facilitate molecular pharmacological and biological research on closantel and its enantiomers.

A Systematic Review of Intravenous β-Hydroxybutyrate Use in Humans – A Promising Future Therapy?

Therapeutic ketosis is traditionally induced with dietary modification. However, owing to the time delay involved, this is not a practical approach for treatment of acute conditions such as traumatic brain injury. Intravenous administration of ketones would obviate this problem by rapidly inducing ketosis. This has been confirmed in a number of small animal and human studies. Currently no such commercially available product exists. The aim of this systematic review is to review the safety and efficacy of intravenous beta-hydroxybutyrate. The Web of Science, PubMed and EMBASE databases were searched, and a systematic review undertaken. Thirty-five studies were included. The total beta-hydroxybutyrate dose ranged from 30 to 101 g administered over multiple doses as a short infusion, with most studies using the racemic form. Such dosing achieves a beta-hydroxybutyrate concentration >1 mmol/L within 15 min. Infusions were well tolerated with few adverse events. Blood glucose concentrations occasionally were reduced but remained within the normal reference range for all study participants. Few studies have examined the effect of intravenous beta-hydroxybutyrate in disease states. In patients with heart failure, intravenous beta-hydroxybutyrate increased cardiac output by up to 40%. No studies were conducted in patients with neurological disease. Intravenous beta-hydroxybutyrate has been shown to increase cerebral blood flow and reduce cerebral glucose oxidation. Moreover, beta-hydroxybutyrate reduces protein catabolism and attenuates the production of counter-regulatory hormones during induced hypoglycemia. An intravenous beta-hydroxybutyrate formulation is well tolerated and may provide an alternative treatment option worthy of further research in disease states.

The Target Determination and the Mechanism of Action of Chiral-Antimalarial Drugs: A Docking Approach

Due to an undecided target and the prescription of chiral-aminoquinolines (chloroquine, primaquine and quinacrine) in the racemic form, the mechanism of action as well as the reason of causing side effects become unclear. Based on computationally evaluated literature data, the things determined theoretically were (i) the target of aminoquinolines during antimalarial activity, (ii) the mechanism of action of chiral-aminoquinolines and (iii) biologically active enantiomers of aminoquinolines. For the presented study, the enantiomeric binding affinities of aminoquinolines with all the targets claimed by other scientists were calculated, and then used in interpretation with the help of many investigations done/observed by others. The results were very interesting based on which, a new and acceptable mechanism of action of chiral-aminoquinolines during malaria curing step, is given for the first time. The current docking study not only resolves the questionable point about a definite target of aminoquinolines but also makes the mechanism of action understandable.

Differential Effects of Halofuginone Enantiomers on Muscle Fibrosis and Histopathology in Duchenne Muscular Dystrophy

Progressive loss of muscle and muscle function is associated with significant fibrosis in Duchenne muscular dystrophy (DMD) patients. Halofuginone, an analog of febrifugine, prevents fibrosis in various animal models, including those of muscular dystrophies. Effects of (+)/(−)-halofuginone enantiomers on motor coordination and diaphragm histopathology in mdx mice, the mouse model for DMD, were examined. Four-week-old male mice were treated with racemic halofuginone, or its separate enantiomers, for 10 weeks. Controls were treated with saline. Racemic halofuginone-treated mice demonstrated better motor coordination and balance than controls. However, (+)-halofuginone surpassed the racemic form’s effect. No effect was observed for (−)-halofuginone, which behaved like the control. A significant reduction in collagen content and degenerative areas, and an increase in utrophin levels were observed in diaphragms of mice treated with racemic halofuginone. Again, (+)-halofuginone was more effective than the racemic form, whereas (−)-halofuginone had no effect. Both racemic and (+)-halofuginone increased diaphragm myofiber diameters, with no effect for (−)-halofuginone. No effects were observed for any of the compounds tested in an in-vitro cell viability assay. These results, demonstrating a differential effect of the halofuginone enantiomers and superiority of (+)-halofuginone, are of great importance for future use of (+)-halofuginone as a DMD antifibrotic therapy.

The folding propensity of α/sulfono-γ-AA peptidic foldamers with both left- and right-handedness

The discovery and application of new types of helical peptidic foldamers have been an attractive endeavor to enable the development of new materials, catalysts and biological molecules. To maximize their application potential through structure-based design, it is imperative to control their helical handedness based on their molecular scaffold. Herein we first demonstrate the generalizability of the solid-state right-handed helical propensity of the 413-helix of L-α/L-sulfono-γ-AA peptides that as short as 11-mer, using the high-resolution X-ray single crystallography. The atomic level folding conformation of the foldamers was also elucidated by 2D NMR and circular dichroism under various conditions. Subsequently, we show that the helical handedness of this class of foldamer is controlled by the chirality of their chiral side chains, as demonstrated by the left-handed 413-helix comprising 1:1 D-α/D-sulfono-γ-AA peptide. In addition, a heterochiral coiled-coil-like structure was also revealed for the first time, unambiguously supporting the impact of chirality on their helical handedness. Our findings enable the structure-based design of unique folding biopolymers and materials with the exclusive handedness or the racemic form of the foldamers in the future.

A P(V)-Platform for Oligonucleotide Synthesis

<div><div><div><p>The early promise of gene-based therapies is currently being realized at an accelerated pace with over 155 active clinical trials for antisense compounds and multiple FDA-approved oligonucleotide therapeutics. Fundamental advances in this area are vital and present an unprecedented opportunity to both address disease states that have been resistant to other common modalities and improve the significant sustainability challenges associated with production of these complex molecules on a commercial scale. The advent of phosphoramidite coupling chemistry and solid-phase synthesis 40 years ago democratized oligonucleotide synthesis to the scientific community, paving the way for many of these stunning developments. The reliability and generality of this approach for the preparation of native phosphate-diesters is attributed to the high reactivity of phosphorus when in the P(III)-oxidation state versus the desired P(V), as it enables rapid P-heteroatom bond formation. However, the growing demand for more diverse phosphorus-based linkages has challenged the limits of this technology. For example, the phosphorothioate (PS) linkage, which stabilizes oligonucleotides towards nuclease cleavage, is universally employed in current oligonucleotide therapeutics but is generally incorporated in racemic form. Stereodefined PS oligonucleotides may have desirable biological and physical properties but are accessed with difficulty using phosphoramidite chemistry. Here we report a flexible and efficient [P(V)]-based platform that can install a wide variety of phosphate linkages at will into oligonucleotides. This approach uses readily accessible reagents and can efficiently install not only stereodefined or racemic thiophosphates, but can install any combination of (S, R or rac)-PS with native phosphodiester (PO2) and phosphorodithioate (PS2) linkages into DNA and other modified nucleotides. Importantly this platform easily accesses this diversity under a standardized coupling protocol with sustainably prepared, stable, P(V) reagents.</p></div></div></div>

A P(V)-Platform for Oligonucleotide Synthesis

<div><div><div><p>The early promise of gene-based therapies is currently being realized at an accelerated pace with over 155 active clinical trials for antisense compounds and multiple FDA-approved oligonucleotide therapeutics. Fundamental advances in this area are vital and present an unprecedented opportunity to both address disease states that have been resistant to other common modalities and improve the significant sustainability challenges associated with production of these complex molecules on a commercial scale. The advent of phosphoramidite coupling chemistry and solid-phase synthesis 40 years ago democratized oligonucleotide synthesis to the scientific community, paving the way for many of these stunning developments. The reliability and generality of this approach for the preparation of native phosphate-diesters is attributed to the high reactivity of phosphorus when in the P(III)-oxidation state versus the desired P(V), as it enables rapid P-heteroatom bond formation. However, the growing demand for more diverse phosphorus-based linkages has challenged the limits of this technology. For example, the phosphorothioate (PS) linkage, which stabilizes oligonucleotides towards nuclease cleavage, is universally employed in current oligonucleotide therapeutics but is generally incorporated in racemic form. Stereodefined PS oligonucleotides may have desirable biological and physical properties but are accessed with difficulty using phosphoramidite chemistry. Here we report a flexible and efficient [P(V)]-based platform that can install a wide variety of phosphate linkages at will into oligonucleotides. This approach uses readily accessible reagents and can efficiently install not only stereodefined or racemic thiophosphates, but can install any combination of (S, R or rac)-PS with native phosphodiester (PO2) and phosphorodithioate (PS2) linkages into DNA and other modified nucleotides. Importantly this platform easily accesses this diversity under a standardized coupling protocol with sustainably prepared, stable, P(V) reagents.</p></div></div></div>

Breast Cancer Therapeutics Based on Fusarochromanone and EGFR Inhibitors

Background: Fusarochromanone (FC101) is a small molecule with potent anti-cancer activity. It was originally derived from the fungal plant pathogen, Fusarium equiseti, and it has also been synthesized in non-racemic form in our lab. Numerous studies reveal the promising biological activity of FC101, including potent anti-angiogenic and anti-cancer activity. While FC101 is potent as a single drug treatment across many cancer cell lines, current cancer therapies often incorporate a combination of drugs in order to increase efficacy and decrease the development of drug resistance. In this study, we leverage drug combinations and cellular phenotypic screens to address important questions about FC101’s mode of action and its potential synergies as an anti-cancer therapeutic agent in triple negative breast cancer (TNBC).Method: We hypothesized that FC101’s activity against TNBC is similar to the known mTOR inhibitor, everolimus, because FC101 reduces the phosphorylation of two key mTOR substrates, S6K and S6. Since everolimus synergistically enhances the anti-cancer activities of known EGFR inhibitors (erlotinib or lapatinib) in TNBC, we performed analogous studies with FC101. Phenotypic cellular assays helped assess whether FC101 (in both single and combination treatments) acts similarly to everolimus.Results: FC101 outperformed all other single treatments in both cell proliferation and viability assays. Unlike everolimus, however, FC101 brought about a sustained decrease in cell viability in drug washout studies. None of the other drugs were able to maintain comparable effects upon removal of the treatment agents. Although we observed slightly additive effects when the TNBC cells were treated with FC101 and either EGFR inhibitor, those effects were not truly synergistic in the manner displayed with everolimus. Conclusion: Our results rule out direct inhibition of mTOR by FC101 and suggest that FC101 acts through a different mechanism than everolimus. This lays the foundation for the refinement of our hypothesis in order to better understand FC101’s mode of action as a novel anti-cancer agent.

Production of (R)-mandelic acid from styrene, L-phenylalanine, glycerol, or glucose via cascade biotransformations

Abstract(R)-mandelic acid is an industrially important chemical, especially used for producing antibiotics. Its chemical synthesis often uses highly toxic cyanide to produce its racemic form, followed by kinetic resolution with 50% maximum yield. Here we report a green and sustainable biocatalytic method for producing (R)-mandelic acid from easily available styrene, biobased L-phenylalanine, and renewable feedstocks such as glycerol and glucose, respectively. An epoxidation-hydrolysis-double oxidation artificial enzyme cascade was developed to produce (R)-mandelic acid at 1.52 g/L from styrene with > 99% ee. Incorporation of deamination and decarboxylation into the above cascade enables direct conversion of L-phenylalanine to (R)-mandelic acid at 913 mg/L and > 99% ee. Expressing the five-enzyme cascade in an L-phenylalanine-overproducing E. coli NST74 strain led to the direct synthesis of (R)-mandelic acid from glycerol or glucose, affording 228 or 152 mg/L product via fermentation. Moreover, coupling of E. coli cells expressing L-phenylalanine biosynthesis pathway with E. coli cells expressing the artificial enzyme cascade enabled the production of 760 or 455 mg/L (R)-mandelic acid from glycerol or glucose. These simple, safe, and green methods show great potential in producing (R)-mandelic acid from renewable feedstocks.