Implementation of the CYP Index for the Design of Selective Tryptophan-2,3-dioxygenase Inhibitors.

A class of imidazoisoindole (III) heme-binding indoleamine-2,3-dioxygenase (IDO1) inhibitors were optimized via structure-based drug design into a series of tryptophan-2,3-dioxygenase (TDO)-selective inhibitors. Kynurenine pathway modulation was demonstrated in vivo, which enabled evaluation of TDO as a potential cancer immunotherapy target. As means of mitigating the risk of drug-drug interactions arising from cytochrome P450 inhibition, a novel property-based drug design parameter, herein referred to as the CYP Index, was implemented for the design of inhibitors with appreciable selectivity for TDO over CYP3A4. We anticipate the CYP Index will be a valuable design parameter for optimizing CYP inhibition of any small molecule inhibitor containing a Lewis basic motif capable of binding heme.

Structure-based design of small-molecule inhibitors of EBNA1 DNA binding blocks Epstein-Barr virus latent infection and tumor growth.

Epstein-Barr virus (EBV) is a DNA tumor virus responsible for 1 to 2% of human cancers including subtypes of Burkitt's lymphoma, Hodgkin's lymphoma, gastric carcinoma, and nasopharyngeal carcinoma (NPC). Persistent latent infection drives EBV-associated tumorigenesis. Epstein-Barr nuclear antigen 1 (EBNA1) is the only viral protein consistently expressed in all EBV-associated tumors and is therefore an attractive target for therapeutic intervention. It is a multifunctional DNA binding protein critical for viral replication, genome maintenance, viral gene expression, and host cell survival. Using a fragment-based approach and x-ray crystallography, we identify a 2,3-disubstituted benzoic acid series that selectively inhibits the DNA binding activity of EBNA1. We characterize these inhibitors biochemically and in cell-based assays, including chromatin immunoprecipitation and DNA replication assays. In addition, we demonstrate the potency of EBNA1 inhibitors to suppress tumor growth in several EBV-dependent xenograft models, including patient-derived xenografts for NPC. These inhibitors selectively block EBV gene transcription and alter the cellular transforming growth factor-ß (TGF-ß) signaling pathway in NPC tumor xenografts. These EBNA1-specific inhibitors show favorable pharmacological properties and have the potential to be further developed for the treatment of EBV-associated malignancies.

Stereochemistry of mephedrone neuropharmacology: enantiomer-specific behavioural and neurochemical effects in rats.

BACKGROUND AND PURPOSE: Synthetic cathinones, commonly referred to as 'bath salts', are a group of amphetamine-like drugs gaining popularity worldwide. 4-Methylmethcathinone (mephedrone, MEPH) is the most commonly abused synthetic cathinone in the UK, and exerts its effects by acting as a substrate-type releaser at monoamine transporters. Similar to other cathinone-related compounds, MEPH has a chiral centre and exists stably as two enantiomers: R-mephedrone (R-MEPH) and S-mephedrone (S-MEPH). EXPERIMENTAL APPROACH: Here, we provide the first investigation into the neurochemical and behavioural effects of R-MEPH and S-MEPH. We analysed both enantiomers in rat brain synaptosome neurotransmitter release assays and also investigated their effects on locomotor activity (e.g. ambulatory activity and repetitive movements), behavioural sensitization and reward. KEY RESULTS: Both enantiomers displayed similar potency as substrates (i.e. releasers) at dopamine transporters, but R-MEPH was much less potent than S-MEPH as a substrate at 5-HT transporters. Locomotor activity was evaluated in acute and repeated administration paradigms, with R-MEPH producing greater repetitive movements than S-MEPH across multiple doses. After repeated drug exposure, only R-MEPH produced sensitization of repetitive movements. R-MEPH produced a conditioned place preference whereas S-MEPH did not. Lastly, R-MEPH and S-MEPH produced biphasic profiles in an assay of intracranial self-stimulation (ICSS), but R-MEPH produced greater ICSS facilitation than S-MEPH. CONCLUSIONS AND IMPLICATIONS: Our data are the first to demonstrate stereospecific effects of MEPH enantiomers and suggest that the predominant dopaminergic actions of R-MEPH (i.e. the lack of serotonergic actions) render this stereoisomer more stimulant-like when compared with S-MEPH. This hypothesis warrants further study.

Desmethyl macrolides: synthesis and evaluation of 4-desmethyl telithromycin.

Novel sources of antibiotics are needed to address the serious threat of bacterial resistance. Accordingly, we have launched a structure-based drug design program featuring a desmethylation strategy wherein methyl groups have been replaced with hydrogens. Herein we report the total synthesis, molecular modeling, and biological evaluation of 4-desmethyl telithromycin (6), a novel desmethyl analogue of the third-generation ketolide antibiotic telithromycin (2) and our final analogue in this series. While 4-desmethyl telithromycin (6) was found to be equipotent with telithromycin (2) against wild-type bacteria, it was 4-fold less potent against the A2058G mutant. These findings reveal that strategically replacing the C4-methyl group with hydrogen (i.e., desmethylation) did not address this mechanism of resistance. Throughout the desmethyl series, the sequential addition of methyls to the 14-membered macrolactone resulted in improved bioactivity. Molecular modeling methods indicate that changes in conformational flexibility dominate the increased biological activity; moreover, they reveal 6 adopts a different conformation once bound to the A2058G ribosome, thus impacting noncovalent interactions reflected in a lower MIC value. Finally, fluorescence polarization experiments of 6 with E. coli ribosomes confirmed 6 is indeed binding the ribosome.

Intramolecular Rearrangement of α-Amino Acid Amide Derivatives of 2-Aminobenzothiazoles.

We have found that α-amino acid amide derivatives of 2-aminobenzothiazoles undergo a time-dependent, thermal rearrangement in which the amine group attacks the 2-position carbon of the thiazole ring to form a 5,5-spiro ring system. This is followed by sulfur leaving and air oxidation to the corresponding symmetrical disulfide. The isolated yields of such products are quite high (>70%) if there is conformational bias to further promote the intramolecular reaction such as for the 2-aminobenzothiazole amides derived from proline or 4-aminopiperidine-4-carboxylic acid. This rearrangement has not been described previously for α-amino acid amide derivatives of 2-aminobenzothiazoles. However, a related reaction involving 2-semicarbazido benzothiazoles has been recently reported.

Facile Synthesis of Mutagen X (MX): 3-Chloro-4-(dichloromethyl)-5-hydroxy-5H-furan-2-one.

3-Chloro-4-(dichloromethyl)-5-hydroxy-5H-furan-2-one (Mutagen X, MX) was synthesized in six steps from commercially-available and inexpensive starting materials (27% overall yield). This synthesis enables the preparation of MX analogs and does not require the use of chlorine gas, as do previously reported methods.

Characterization of a series of 4-aminoquinolines that stimulate caspase-7 mediated cleavage of TDP-43 and inhibit its function.

Dysfunction of the heterogeneous ribonucleoprotein TAR DNA binding protein 43 (TDP-43) is associated with neurodegeneration in diseases such as amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). Here we examine the effects of a series of 4-aminoquinolines with affinity for TDP-43 upon caspase-7-induced cleavage of TDP-43 and TDP-43 cellular function. These compounds were mixed inhibitors of biotinylated TG6 binding to TDP-43, binding to both free and occupied TDP-43. Incubation of TDP-43 and caspase-7 in the presence of these compounds stimulated caspase-7 mediated cleavage of TDP-43. This effect was antagonized by the oligonucleotide TG12, prevented by denaturing TDP-43, and exhibited a similar relation of structure to function as for the displacement of bt-TG6 binding to TDP-43. In addition, the compounds did not affect caspase-7 enzyme activity. In human neuroglioma H4 cells, these compounds lowered levels of TDP-43 and increased TDP-43 C-terminal fragments via a caspase-dependent mechanism. Subsequent experiments demonstrated that this was due to induction of caspases 3 and 7 leading to increased PARP cleavage in H4 cells with similar rank order of the potency among the compounds tests for displacement of bt-TG6 binding. Exposure to these compounds also reduced HDAC-6, ATG-7, and increased LC3B, consistent with the effects of TDP-43 siRNA described by other investigators. These data suggest that such compounds may be useful biochemical probes to further understand both the normal and pathological functions of TDP-43, and its cleavage and metabolism promoted by caspases.

Desmethyl Macrolides: Synthesis and Evaluation of 4,10-Didesmethyl Telithromycin.

Novel sources of antibiotics are required to keep pace with the inevitable onset of bacterial resistance. Continuing with our macrolide desmethylation strategy as a source of new antibiotics, we report the total synthesis, molecular modeling and biological evaluation of 4,10-didesmethyl telithromycin (4), a novel desmethyl analogue of the 3rd-generation drug telithromycin (2). Telithromycin is an FDA-approved ketolide antibiotic derived from erythromycin (1). We found 4,10-didesmethyl telithromycin (4) to be four times more active than previously prepared 4,8,10-tridesmethyl congener (3) in MIC assays. While less potent than telithromycin (2), the inclusion of the C-8 methyl group has improved biological activity suggesting it plays an important role in antibiotic function.

Total synthesis of (-)-4,8,10-tridesmethyl telithromycin.

Novel sources of antibiotics are required to address the serious problem of antibiotic resistance. Telithromycin (2) is a third-generation macrolide antibiotic prepared from erythromycin (1) and used clinically since 2004. Herein we report the details of our efforts that ultimately led to the total synthesis of (-)-4,8,10-tridesmethyl telithromycin (3) wherein methyl groups have been replaced with hydrogens. The synthesis of desmethyl macrolides has emerged as a novel strategy for preparing bioactive antibiotics.

Desmethyl Macrolide Analogues to Address Antibiotic Resistance: Total Synthesis and Biological Evaluation of 4,8,10-Tridesmethyl Telithromycin.

There is an urgent need to discover new drugs to address the pressing problem of antibiotic-resistance. Macrolide antibiotics such as erythromycin (1) are safe, broad-spectrum antibiotics used in the clinic since 1954. Herein we report the synthesis and evaluation of 4,8,10-tridesmethyl telithromycin (3), a novel desmethyl analogue of the 3rd-generation drug telithromycin (2), which is a semisynthetic derivative of 1. Analogue 3 was found to possess antibiotic activity and was superior to telithromycin (2) when tested against resistant strains of S. aureus possessing an A→T mutation at position 2058 (E. coli numbering).

Synthesis of polysubstituted pyrroles from sulfinimines (N-sulfinyl imines).

Polysubstituted 2-carboxylate and 2-phosphonate pyrroles are prepared by aromatization of the corresponding 3-oxo 2-carboxylate and 2-phosphonate NH-pyrrolidines using air. Reaction of electrophiles with 3-oxo pyrrolidine dianions readily introduces substituents, regioselectively at C-4 in these pyrrolidines.

Concise syntheses of D-desosamine, 2-thiopyrimidinyl desosamine donors, and methyl desosaminide analogues from D-glucose.

A concise synthesis of d-desosamine has been accomplished in five steps and in 15% overall yield from methyl alpha-d-glucopyranoside. Desosamine was then transformed into two known 2-thiopyrimidinyl donors (Woodward and Tatsuta donors), each in two steps. Finally, analogues of methyl desosaminide at the C-3 position were prepared (3-pyrrolidino, 3-piperidino, 3-morpholino) from a common 2,3-anhydrosugar intermediate.