Hing L. Sham

Polo-like Kinase 2 (PLK2) Phosphorylates α-Synuclein at Serine 129 in Central Nervous System

Several neurological diseases, including Parkinson disease and dementia with Lewy bodies, are characterized by the accumulation of α-synuclein phosphorylated at Ser-129 (p-Ser-129). The kinase or kinases responsible for this phosphorylation have been the subject of intense investigation. Here we submit evidence that polo-like kinase 2 (PLK2, also known as serum-inducible kinase or SNK) is a principle contributor to α-synuclein phosphorylation at Ser-129 in neurons. PLK2 directly phosphorylates α-synuclein at Ser-129 in an in vitro biochemical assay. Inhibitors of PLK kinases inhibited α-synuclein phosphorylation both in primary cortical cell cultures and in mouse brain in vivo. Finally, specific knockdown of PLK2 expression by transduction with short hairpin RNA constructs or by knock-out of the plk2 gene reduced p-Ser-129 levels. These results indicate that PLK2 plays a critical role in α-synuclein phosphorylation in central nervous system.

Validation of diacyl glycerolacyltransferase I as a novel target for the treatment of obesity and dyslipidemia using a potent and selective small molecule inhibitor.

A highly potent and selective DGAT-1 inhibitor was identified and used in rodent models of obesity and postprandial chylomicron excursion to validate DGAT-1 inhibition as a novel approach for the treatment of metabolic diseases. Specifically, compound 4a conferred weight loss and a reduction in liver triglycerides when dosed chronically in DIO mice and depleted serum triglycerides following a lipid challenge in a dose-dependent manner, thus, reproducing major phenotypical characteristics of DGAT-1(-/-) mice.

Selective Chk1 inhibitors differentially sensitize p53-deficient cancer cells to cancer therapeutics

The majority of cancer therapeutics induces DNA damage to kill cells. Normal proliferating cells undergo cell cycle arrest in response to DNA damage, thus allowing DNA repair to protect the genome. DNA damage induced cell cycle arrest depends on an evolutionarily conserved signal transduction network in which the Chk1 kinase plays a critical role. In mammalian cells, the p53 and RB pathways further augment the cell cycle arrest response to prevent catastrophic cell death. Given the fact that most tumor cells suffer defects in the p53 and RB pathways, it is likely that tumor cells would depend more on the Chk1 kinase to maintain cell cycle arrest than would normal cells. Therefore Chk1 inhibition could be used to specifically sensitize tumor cells to DNA‐damaging agents. We have previously shown that siRNA‐mediated Chk1 knockdown abrogates DNA damage‐induced checkpoints and potentiates the cytotoxicity of several DNA‐damaging agents in p53‐deficient cell lines. In this study, we have developed 2 potent and selective Chk1 inhibitors, A‐690002 and A‐641397, and shown that these compounds abrogate cell cycle checkpoints and potentiate the cytotoxicity of topoisomerase inhibitors and γ‐radiation in p53‐deficient but not in p53‐proficient cells of different tissue origins. These results indicate that it is feasible to achieve a therapeutic window with 1 or more Chk1 inhibitors in potentiation of cancer therapy based on the status of the p53 pathway in a wide spectrum of tumor types.

Synthesis and Biological Evaluation of 2-Indolyloxazolines as a New Class of Tubulin Polymerization Inhibitors. Discovery of A-289099 as an Orally Active Antitumor Agent

A series of indole containing oxazolines has been discovered as a result of structural modifications of the lead compound A-105972. The compounds exert their anticancer activity through inhibition of tubulin polymerization by binding at the colchicine site. A-289099 was identified as an orally active antimitotic agent active against various cancer cell lines including those that express the MDR phenotype. The anticancer activity, pharmacokinetics, and an efficient and enantioselective synthesis of A-289099 are described.

X-ray Crystallographic Structure of ABT-378 (Lopinavir) Bound to HIV-1 Protease

The crystal structure of ABT-378 (lopinavir), bound to the active site of HIV-1 protease is described. A comparison with crystal structures of ritonavir, A-78791, and BILA-2450 shows some analogous features with previous reported compounds. A cyclic urea unit in the P(2) position of ABT-378 is novel and makes two bidentate hydrogen bonds with Asp 29 of HIV-1 protease. In addition, a previously unreported shift in the Gly 48 carbonyl position is observed. A discussion of the structural features responsible for its high potency against wild-type HIV protease is given along with an analysis of the effect of active site mutations on potency in in vitro assays.

Metabolism and Disposition of the HIV-1 Protease Inhibitor Lopinavir (ABT-378) Given in Combination with Ritonavir in Rats, Dogs, and Humans

AbstractPurpose. The objective of this study was to examine the metabolism and disposition of the HIV protease inhibitor lopinavir in humans and animal models. Methods. The plasma protein binding of [14C]lopinavir was examined in vitro via equilibrium dialysis technique. The tissue distribution of radioactivity was examined in rats dosed with [14C]lopinavir in combination with ritonavir. The metabolism and disposition of [14C]lopinavir was examined in rats, dogs, and humans given alone (in rats only) or in combination with ritonavir. Results. The plasma protein binding of lopinavir was high in all species (97.4-99.7% in human plasma), with a concentration-dependent decrease in binding. Radioactivity was extensively distributed into tissues, except brain, in rats. On oral dosing to rats, ritonavir was found to increase the exposure of lopinavir-derived radioactivity 13-fold. Radioactivity was primarily cleared via the hepato-biliary route in all species (>82% of radioactive dose excreted via fecal route), with urinary route of elimination being significant only in humans (10.4% of radioactive dose). Oxidative metabolites were the predominant components of excreted radioactivity. The predominant site of metabolism was found to be the carbon-4 of the cyclic urea moiety, with subsequent secondary metabolism occurring on the diphenyl core moiety. In all the three species examined, the primary component of plasma radioactivity was unchanged lopinavir (>88%) with small amounts of oxidative metabolites. Conclusions. Lopinavir was subject to extensive metabolism in vivo. Co-administered ritonavir markedly enhanced the pharmacokinetics of lopinavir-derived radioactivity in rats, probably due to inhibition of presystemic and systemic metabolism, leading to an increased exposure to this potent HIV protease inhibitor.

Discovery of piperidine-aryl urea-based stearoyl-CoA desaturase 1 inhibitors

A series of structurally novel stearoyl-CoA desaturase1 (SCD1) inhibitors has been identified via molecular scaffold manipulation. Preliminary structure-activity relationship (SAR) studies led to the discovery of potent, and orally bioavailable piperidine-aryl urea-based SCD1 inhibitors. 4-(2-Chlorophenoxy)-N-[3-(methyl carbamoyl)phenyl]piperidine-1-carboxamide 4c exhibited robust in vivo activity with dose-dependent desaturation index lowering effects.

Novel sulfonate analogues of combretastatin A-4: Potent antimitotic agents

Sulfonate analogues of combretastatin A-4 have been prepared. These compounds compete with colchicine and combretastatin A-4 for the colchicine binding site on tubulin and are potent inhibitors of tubulin polymerization and cell proliferation. Importantly, these compounds also inhibit the proliferation of P-glycoprotein positive (+) cancer cells, which are resistant to many other antitumor agents.

Design of an orally efficacious hydroxyethylamine (HEA) BACE-1 inhibitor in a preclinical animal model.

In this Letter, we describe our efforts to design HEA BACE-1 inhibitors that are highly permeable coupled with negligible levels of permeability-glycoprotein activity. These efforts culminate in producing 16 which lowers Αβ by 28% and 32% in the cortex and CSF, respectively, in the preclinical wild type Hartley guinea pig animal model when dosed orally at 30mpk BID for 2.5days.

Design of orally bioavailable, symmetry-based inhibitors of HIV protease.

A series of novel inhibitors of HIV-1 protease with excellent oral bioavailability is described. Differential acylation of the two amino groups of symmetry-based diamine core groups 2-5 led to unsymmetrically substituted inhibitors 17-43, many of which inhibited HIV protease at subnanomolar concentrations. Anti-HIV activity in vitro was observed at 0.1-1 microM. A systematic evaluation of the pharmacokinetic behavior of these inhibitors in rats identified the influence of aqueous solubility, molecular size and hydrogen-bonding functionality. Compound 30 (A-80987) was selected for further evaluation based on a favorable Cmax/ ED50 ratio (> 20) and half-life (> 2 h).