Hydrophobic Surfactant-DNA Complex (Surf-DNA) Enables DNA-Encoded-Library-Compatible Decarboxylative Arylation under Anhydrous Conditions.

DNA-encoded libraries (DELs) are a key technology for identifying small-molecule hits in both the pharmaceutical industry and academia, but their chemical diversity is largely limited to water-compatible reactions to aid in the solubility and integrity of encoding DNA tags. To broaden the DEL chemical space, we present a workflow utilizing DNA-cationic surfactant complexation that enables dissolution and reactions on-DNA in anhydrous organic solvents. We demonstrate its utility by developing DEL-compatible photoredox decarboxylative C(sp2)-C(sp3) coupling under water-free conditions. The workflow is optimized for the 96-well format necessary for large-scale DEL productions, and it enables screening and optimization of DEL-compatible reactions in organic solvents.

Photoredox-Mediated Deoxygenative Alkylation of DNA-Tagged Alkenes with Activated Alcohols.

DNA-encoded library (DEL) screens have become a key technology to find small molecule binders to biological targets for drug discovery applications. The development of new DNA-compatible chemistries to expand the accessible DEL chemical space is imperative to enhance screen success across broad target classes and modalities. Additionally, reactions that use commonly available building blocks as well as those that enable the fsp3 of library members to be increased would have high impact for accessing diverse drug-like structures. Herein, we report a DNA-compatible Giese-type addition of nonstabilized C-centered radicals generated by the deoxygenation of preactivated alcohols into on-DNA olefins. Although alcohols have been historically underused as a building block class within DEL synthesis, their activation to a xanthate enables Csp3-Csp3 coupling to furnish sp3-rich products. This reaction is compatible with multiple classes of functional groups, does not damage the DNA tag, and is suitable for use in DEL productions.

Promising antimalarials targeting apicoplast DNA polymerase from Plasmodium falciparum.

Malaria is caused by the parasite Plasmodium falciparum, which contains an essential non-photosynthetic plastid called the apicoplast. A single DNA polymerase, apPOL, is targeted to the apicoplast, where it replicates and repairs the genome. apPOL has no direct orthologs in mammals and is considered a promising drug target for the treatment and/or prevention of malaria. We previously reported screening the Malaria Box to identify MMV666123 as an inhibitor of apPOL. Herein we extend our studies and report structure-activity relationships for MMV666123 and identify key structural motifs necessary for inhibition. Although attempts to crystallize apPOL with the inhibitor were not fruitful, kinetic analysis and crystal structure determinations of WT and mutant apo-enzymes, facilitated model building and provided insights into the putative inhibitor binding site. Our results validate apPOL as an antimalarial target and provide an avenue for the design of high potency, specific inhibitors of apPOL and other A-family DNA polymerases.

Palladium-Mediated Carbonylative Suzuki Coupling for DNA-Encoded Library Synthesis.

Developing new DNA-compatible reactions is key to expanding the accessible chemical space of DNA-encoded library (DEL) technology. Here we disclose the first report of a DNA-compatible carbonylative Suzuki coupling of DNA-conjugated (hetero)aryl iodides with (hetero)aryl boronic acids to access di(hetero)aryl ketones, a valuable structural motif present within several approved or clinically advanced small molecules. The reported DNA-compatible, Pd(OAc)2-mediated system is mild, uses a robust protocol, has a wide substrate scope for both coupling partners, is suitable for large-scale DEL productions, and provides a source of previously unexplored chemical matter for DEL screens.

Palladium-Mediated C-N Coupling of DNA-Conjugated (Hetero)aryl Halides with Aliphatic and (Hetero)aromatic Amines.

DNA-encoded chemical library (DEL) screens are a powerful hit generation tool in drug discovery, but the diversity of DEL chemical matter is dependent on developing robust reaction conditions that may be used on hundreds to millions of substrate combinations and that are compatible with the platform. Here, we disclose the first report of a general, aqueous, DNA-compatible C-N coupling condition that can now couple aliphatic amines, in addition to (hetero)aromatic amines, with a variety of (hetero)aryl iodides, bromides, and chlorides. The reported BippyPhos-Pd(OAc)2 catalyst system has a wide substrate scope for both coupling partners, is operationally feasible for large scale DEL productions, uses common DEL building block solution stocks, and enables an expansion of DEL-accessible, drug-like chemical space.

A stereochemical journey around spirocyclic glutamic acid analogs.

A practical divergent synthetic approach is reported for the library of regio- and stereoisomers of glutamic acid analogs built on the spiro[3.3]heptane scaffold. Formation of the spirocyclic scaffold was achieved starting from a common precursor - an O-silylated 2-(hydroxymethyl)cyclobutanone derivative. Its olefination required using the titanium-based Tebbe protocol since the standard Wittig reaction did not work with this particular substrate. The construction of the second cyclobutane ring of the spirocyclic system was achieved through either subsequent dichloroketene addition or Meinwald oxirane rearrangement as the key synthetic steps, depending on the substitution patterns in the target compounds (1,6- or 1,5-, respectively). Further modified Strecker reaction of the resulting racemic spirocyclic ketones with the Ellman's sulfinamide as a chiral auxiliary had low to moderate diastereoselectivity; nevertheless, all stereoisomers were isolated in pure form via chromatographic separation, and their absolute configuration was confirmed by X-ray crystallography. Members of the library were tested for the inhibitory activity against H. pylori glutamate racemase.

Small molecule SWELL1 complex induction improves glycemic control and nonalcoholic fatty liver disease in murine Type 2 diabetes.

Type 2 diabetes is associated with insulin resistance, impaired pancreatic ß-cell insulin secretion, and nonalcoholic fatty liver disease. Tissue-specific SWELL1 ablation impairs insulin signaling in adipose, skeletal muscle, and endothelium, and impairs ß-cell insulin secretion and glycemic control. Here, we show that ICl,SWELL and SWELL1 protein are reduced in adipose and ß-cells in murine and human diabetes. Combining cryo-electron microscopy, molecular docking, medicinal chemistry, and functional studies, we define a structure activity relationship to rationally-design active derivatives of a SWELL1 channel inhibitor (DCPIB/SN-401), that bind the SWELL1 hexameric complex, restore SWELL1 protein, plasma membrane trafficking, signaling, glycemic control and islet insulin secretion via SWELL1-dependent mechanisms. In vivo, SN-401 restores glycemic control, reduces hepatic steatosis/injury, improves insulin-sensitivity and insulin secretion in murine diabetes. These findings demonstrate that SWELL1 channel modulators improve SWELL1-dependent systemic metabolism in Type 2 diabetes, representing a first-in-class therapeutic approach for diabetes and nonalcoholic fatty liver disease.

Suppression of human T cell activation by derivatives of glycerol monolaurate.

Glycerol monolaurate (GML), a naturally occurring monoglyceride, is widely used commercially for its antimicrobial properties. Interestingly, several studies have shown that GML not only has antimicrobial properties but is also an anti-inflammatory agent. GML inhibits peripheral blood mononuclear cell proliferation and inhibits T cell receptor (TCR)-induced signaling events. In this study, we perform an extensive structure activity relationship analysis to investigate the structural components of GML necessary for its suppression of human T cell activation. Human T cells were treated with analogs of GML, differing in acyl chain length, head group, linkage of acyl chain, and number of laurate groups. Treated cells were then tested for changes in membrane dynamics, LAT clustering, calcium signaling, and cytokine production. We found that an acyl chain with 12-14 carbons, a polar head group, an ester linkage, and a single laurate group at any position are all necessary for GML to inhibit protein clustering, calcium signaling, and cytokine production. Removing the glycerol head group or replacing the ester linkage with a nitrogen prevented derivative-mediated inhibition of protein cluster formation and calcium signaling, while still inhibiting TCR-induced cytokine production. These findings expand our current understanding of the mechanisms of action of GML and the of GML needed to function as a novel immunosuppressant.

One-Pot Reductive Alkylation of 2,4-Dihydroxy Quinolines and Pyridines.

A one-pot, Hantzsch ester-mediated Knoevenagel condensation-reduction reaction has been developed for alkylation of a wide range of substituted 2,4-quinoline diols and 2,4-pyridine diols with aldehydes. The process is operationally simple to perform, scalable, and provides highly useful C-3 alkylated quinoline and pyridine diols in yields of 58-92%. The alkylation products can be converted to 2,4-dihaloquinoline and pyridine substrates for further functionalization.

Cholesterol-Modified Oligonucleotides as Internal Reaction Controls during DNA-Encoded Chemical Library Synthesis.

We report two cholesterol-modified oligonucleotides for use as internal controls for on-DNA reactions during the pooled stages of a DNA-encoded chemical library (DECL) synthesis. As these cholesterol-tagged oligonucleotides are chromatographically separable from normal DECL intermediates, they can be directly monitored by mass spectrometry to track reaction progression within a complex pool of DNA. We observed similar product conversions for reactions on substrates linked to a standard DECL DNA headpiece, to the cholesterol-modified oligonucleotides, and to the cholesterol-modified oligonucleotides while in the presence of pooled DECL synthetic intermediates-validating their use as a representative control. We also highlight an example from a DECL production in which the use of the cholesterol-modified oligonucleotides provided quality control information that guided synthetic decisions. We conclude that the use of cholesterol-modified oligonucleotides as a regular control will significantly improve the quality of DECL productions.

A Novel Triphenylphosphonium Carrier to Target Mitochondria without Uncoupling Oxidative Phosphorylation.

Mitochondrial dysfunction is an underlying pathology in numerous diseases. Delivery of diagnostic and therapeutic cargo directly into mitochondria is a powerful approach to study and treat these diseases. The triphenylphosphonium (TPP+) moiety is the most widely used mitochondriotropic carrier. However, studies have shown that TPP+ is not inert; TPP+ conjugates uncouple mitochondrial oxidative phosphorylation. To date, all efforts toward addressing this problem have focused on modifying lipophilicity of TPP+-linker-cargo conjugates to alter mitochondrial uptake, albeit with limited success. We show that structural modifications to the TPP+ phenyl rings that decrease electron density on the phosphorus atom can abrogate uncoupling activity as compared to the parent TPP+ moiety and prevent dissipation of mitochondrial membrane potential. These alterations of the TPP+ structure do not negatively affect the delivery of cargo to mitochondria. Results here identify the 4-CF3-phenyl TPP+ moiety as an inert mitochondria-targeting carrier to safely target pharmacophores and probes to mitochondria.

Decrypting a Cryptic Allosteric Pocket in H. pylori Glutamate Racemase.

One of our greatest challenges in drug design is targeting cryptic allosteric pockets in enzyme targets. Drug leads that do bind to these cryptic pockets are often discovered during HTS campaigns, and the mechanisms of action are rarely understood. Nevertheless, it is often the case that the allosteric pocket provides the best option for drug development against a given target. In the current studies we present a successful way forward in rationally exploiting the cryptic allosteric pocket of H. pylori glutamate racemase, an essential enzyme in this pathogen's life cycle. A wide range of computational and experimental methods are employed in a workflow leading to the discovery of a series of natural product allosteric inhibitors which occupy the allosteric pocket of this essential racemase. The confluence of these studies reveals a fascinating source of the allosteric inhibition, which centers on the abolition of essential monomer-monomer coupled motion networks.

Decrypting a cryptic allosteric pocket in H. pylori glutamate racemase.

One of our greatest challenges in drug design is targeting cryptic allosteric pockets in enzyme targets. Drug leads that do bind to these cryptic pockets are often discovered during HTS campaigns, and the mechanisms of action are rarely understood. Nevertheless, it is often the case that the allosteric pocket provides the best option for drug development against a given target. In the current studies we present a successful way forward in rationally exploiting the cryptic allosteric pocket of H. pylori glutamate racemase, an essential enzyme in this pathogen's life cycle. A wide range of computational and experimental methods are employed in a workflow leading to the discovery of a series of natural product allosteric inhibitors which occupy the allosteric pocket of this essential racemase. The confluence of these studies reveals a fascinating source of the allosteric inhibition, which centers on the abolition of essential monomer-monomer coupled motion networks.

Effect of mitoquinone (Mito-Q) on neuropathic endpoints in an obese and type 2 diabetic rat model.

This study sought to determine whether the addition of mitoquinone (Mito-Q) in the diet is an effective treatment for peripheral neuropathy in animal models of diet-induced obesity (pre-diabetes) and type 2 diabetes. Unlike other anti-oxidative stress compounds investigated as a treatment for peripheral neuropathy, Mito-Q specifically targets mitochondria. Although mito-Q has been shown to reduce oxidative stress generated by mitochondria there have been no studies performed of the effect of Mito-Q on peripheral neuropathy induced by diet-induced obesity or type 2 diabetes. Diet-induced obese (12 weeks after high fat diet) or type 2 diabetic rats (12 weeks of high fat diet and 4 weeks after the onset of hyperglycemia) were treated via the diet with Mito-Q (0.93 g/kg diet) for 12 weeks. Afterwards, glucose utilization, vascular reactivity of epineurial arterioles to acetylcholine and peripheral neuropathy related endpoints were examined. The addition of Mito-Q to the diets of obese and diabetic rats improved motor and/or sensory nerve conduction velocity, cornea and intraepidermal nerve fibre density, cornea sensitivity and thermal nociception. Surprisingly, treating obese and diabetic rats with Mito-Q did not improve glucose utilization or vascular reactivity by epineurial arterioles to acetylcholine. These studies imply that mitochondrial dysfunction contributes to peripheral neuropathy in animal models of pre-diabetes and late-stage type 2 diabetes. However, improvement in peripheral neuropathy following treatment with Mito-Q was not associated with improvement in glucose utilization or vascular reactivity of epineurial arterioles to acetylcholine.

Identification of an ethyl 5,6-dihydropyrazolo[1,5-c]quinazoline-1-carboxylate as a catalytic inhibitor of DNA gyrase.

Fluoroquinolones are a class of antibacterial agents used clinically to treat a wide array of bacterial infections and target bacterial type-II topoisomerases (DNA gyrase and topoisomerase IV). Fluoroquinolones, however potent, are susceptible to bacterial resistance with prolonged use, which limits their use in the clinic. Quinazoline-2,4-diones also target bacterial type-II topoisomerases and are not susceptible to bacterial resistance similar to fluoroquinolones, however, their potency pales in comparison to fluoroquinolones. To meet the increasing demand for antibacterial development, nine modified quinazoline-2,4-diones were developed to probe quinazoline-2,4-dione structure modification for possible new binding contacts with the bacterial type-II topoisomerase, DNA gyrase. Evaluation of compounds for inhibition of the supercoiling activity of DNA gyrase revealed a novel ethyl 5,6-dihydropyrazolo[1,5-c]quinazoline-1-carboxylate derivative as a modest inhibitor of DNA gyrase, having an IC50 of 3.5 µM. However, this ethyl 5,6-dihydropyrazolo[1,5-c]quinazoline-1-carboxylate does not trap the catalytic intermediate like fluoroquinolones or typical quinazoline-2,4-diones do. Thus, the ethyl 5,6-dihydropyrazolo[1,5-c]quinazoline-1-carboxylate derivative discovered in this work acts as a catalytic inhibitor of DNA gyrase and therefore represents a new structural type of catalytic inhibitor of DNA gyrase.

Probing structural requirements for human topoisomerase I inhibition by a novel N1-Biphenyl fluoroquinolone.

Fluoroquinolones substituted with N-1 biphenyl and napthyl groups were discovered to act as catalytically inhibitors of human topoisomerases I and II, and to possess anti-proliferative activity in vivo. Structural requirements for these novel quinolones to inhibit catalytic activity of human topoisomerase I have not been explored. In this work novel derivatives of the N-1 biphenyl fluoroquinolone were designed, synthesized and evaluated to understand structural requirements of the C-3 carboxylic acid, C-6 fluorine, C-7 aminomethylpyrrolidine, C-8 methoxy, and the N-1 biphenyl functional groups for hTopoI inhibition. Characterization of each analog for inhibition of hTopoI catalytic inhibition reveals critical insight into structural requirements of these novel quinolones for activity. Additionally, results of DNA binding and modeling studies suggest that N-1 biphenyl fluoroquinolones intercalate between the DNA base pairs with the N-1 biphenyl functional group, rather than the quinolone core, and that this mode of DNA intercalation contributes to inhibition of hTopoI by these novel structures. The results presented here support further development and evaluation of N-1 biphenyl fluoroquinolone analogs as a novel class of anti-cancer agents that act through catalytic inhibition of hTopoI.

The C7-aminomethylpyrrolidine group rescues the activity of a thio-fluoroquinolone.

A Mg2+-water bridge between the C-3, C-4 diketo moiety of fluoroquinolones and the conserved amino acid residues in the GyrA/ParC subunit is critical for the binding of a fluoroquinolone to a topoisomerase-DNA covalent complex. The fluoroquinolone UING-5-249 (249) can bind to the GyrB subunit through its C7-aminomethylpyrrolidine group. This interaction is responsible for enhanced activities of 249 against the wild type and quinolone-resistant mutant topoisomerases. To further evaluate the effects of the 249-GyrB interaction on fluoroquinolone activity, we examined the activities of decarboxy- and thio-249 against DNA gyrase and conducted docking studies using the structure of a gyrase-ciprofloxacin-DNA ternary complex. We found that the 249-GyrB interaction rescued the activity of thio-249 but not that of decarboxy-249. A C7-group that binds more strongly to the GyrB subunit may allow for modifications at the C-4 position, leading to a novel compound that is active against the wild type and quinolone-resistant pathogens.

Meta-analysis of Failure and Survival Rate of Implant-supported Single Crowns, Fixed Partial Denture, and Implant Tooth-supported Prostheses.

BACKGROUND: Dental implants have become the most viable option for rehabilitation. Although, many studies report the success of these reconstructions using implants, a cumulative data about the various studies and the failure rate still remain unaddressed. Therefore, the purpose of this systematic review was to analyze these data and to derive the cumulative survival rate of different implant-supported prosthesis. MATERIALS AND METHODS: Manual searches followed by a MEDLINE search were conducted to select prospective and retrospective cohort studies on single crowns (SCs), fixed partial denture (FPD), and tooth implant connected prostheses with a mean follow-up time of minimum of 5 years. Random-effects Poisson's regression models have been used to obtain summary estimates for implant failure and survival rates. RESULTS: Data were extracted from the final selected 63 studies. In a meta-analysis of these studies, the survival rate of SCs supported by implants (95% CI) was 96.363%, for FPDs was 94.525% and implant tooth-supported prostheses was 91.27% after 5 years of function. The cumulative failure rate per 100 FPD years of the SCs, FPDs, and implant tooth-supported prostheses were 0.684, 0.881, and 1.514, respectively. CONCLUSION: The study concludes high survival rates for implant-supported SCs followed by implant-supported FPDs can be expected over an observation period of 5 years. However, tooth implant-supported prostheses can be provided if there are certain limitations prohibiting the completely implant-supported prostheses.

Rehabilitation of an Orbital Defect: A Simplified Technique.

Loss of tissue, whether congenital or traumatic or resulting from malignancy or radical surgery, is accompanied by esthetic and psychologic effects. This loss is more pronounced when the affected part is the eye and all orbital contents, resulting in gross mutilation. Success in maxillofacial prosthetics depends on the full cognizance of the principles that underlie facial harmony, color matching, anchorage and retention, weight bearing and leverage, durability and strength of materials used, tissue compatibility and tolerance. The restoration of orbital defects presents a challenge in maxillofacial prosthetics. Many variations exist in techniques and materials for fabricating orbital prostheses. Careful positioning of the ocular portion of the orbital prosthesis is one of the requirements for a successful esthetic result. A change of this position, which may occur during fabrication or may be due to distortion of the prosthetic housing or loss of retention of the prosthesis, may result in an unsatisfactory appearance. This article presents a simplified technique for fabricating an orbital prosthesis.

Synthesis and biotinylation of oligosaccharide fragments of mannosylated and 5-deoxy-5-methylthio-xylofuranosylated lipoarabinomannan from Mycobacterium tuberculosis.

The attachment of biotin to a molecule provides a powerful tool in biology. Here, we report an efficient synthesis and biotinylation of mannosylated and 5-deoxy-5-methylthio-xylofuranosylated Lipoarabinomannan from Mycobacterium tuberculosis. Preparation of the oligosaccharides involved the sequential addition of thioglycoside donors with arabinofuranosyl-containing acceptors. Methylthio group was introduced near the end of the synthesis.