Stewart Campbell

A Small Molecule Inhibitor of Enterocytic Microsomal Triglyceride Transfer Protein; SLx-4090, Biochemical, Pharmacodynamic, Pharmacokinetic and Safety Profile

First-generation microsomal triglyceride transfer protein (MTP) inhibitors were designed to inhibit hepatic MTP and provide a novel treatment of dyslipidemia. Effective at lowering low-density lipoprotein-cholesterol (LDL-C), these inhibitors also elevate liver enzymes and induce hepatic steatosis in animals and humans. MTP is highly expressed in the enterocytes, lining the lumen of the jejunum, and is critical in the production of chylomicrons assembled from lipid/cholesterol and their transfer into systemic circulation. 6-(4′-Trifluoromethyl-6-methoxy-biphenyl-2-ylcarboxamido)-1,2,3,4-tetrahydroisoquinoline-2-carboxylic acid phenyl ester (SLx-4090) (IC50 value ∼8 nM) was designed to inhibit only MTP localized to enterocytes. In Caco-2 cells SLx-4090 inhibited apolipoprotein B (IC50 value ∼9.6 nM) but not apolipoprotein A1 secretion. Administered orally to rats SLx-4090 reduced postprandial lipids by >50% with an ED50 value ∼7 mg/kg. SLx-4090 was not detected in the systemic or portal vein serum of the animals (lower limit of quantitation ∼5 ng/ml) after single or multiple oral doses in fasted rodents. When coadministered with tyloxapol, SLx-4090 did not inhibit the secretion of hepatic triglycerides (TG), consistent with the absence of systemic exposure. Chronic treatment with SLx-4090 in mice maintained on a high-fat diet decreased LDL-C and TG and resulted in weight loss without the elevation of liver enzymes or an increase in hepatic fat. The compound did not result in toxicity when administered to rats for 90 days at a dose of 1000 mg/kg per day. These data support the concept that the inhibition of enterocytic MTP could serve as a useful strategy in the treatment of metabolic disorders.

ROCK-Isoform-Specific Polarization of Macrophages Associated with Age-Related Macular Degeneration

Age is a major risk factor in age-related macular degeneration (AMD), but the underlying cause is unknown. We find increased Rho-associated kinase (ROCK) signaling and M2 characteristics in eyes of aged mice, revealing immune changes in aging. ROCK isoforms determine macrophage polarization into M1 and M2 subtypes. M2-like macrophages accumulated in AMD, but not in normal eyes, suggesting that these macrophages may be linked to macular degeneration. M2 macrophages injected into the mouse eye exacerbated choroidal neovascular lesions, while M1 macrophages ameliorated them, supporting a causal role for macrophage subtypes in AMD. Selective ROCK2 inhibition with a small molecule decreased M2-like macrophages and choroidal neovascularization. ROCK2 inhibition upregulated M1 markers without affecting macrophage recruitment, underlining the plasticity of these macrophages. These results reveal age-induced innate immune imbalance as underlying AMD pathogenesis. Targeting macrophage plasticity opens up new possibilities for more effective AMD treatment.

Soft lithography and surface chemistry: enabling tools for new bioassays

The process of drug discovery can be accelerated by increasing the information content of bioassays and by employing assay platforms that are amenable to high throughput screening techniques. In this paper, we demonstrate how the combination of soft lithography with controlled surface chemistry achieves these goals in a wide spectrum of bioassays. A number of soft lithographic methods can be used to generate micro-structures for the purposes of increasing assay density, diversity of test conditions and improving assay detection qualities. In addition, soft lithography, combined with specific surface chemistry modification procedures and protein engineering, may be used to control the localized molecular and biological properties of substrates, thereby enabling the development of new types of bioassays. The developed methodologies are simple, easily implemented, and lend themselves well to automation. Experimental results and prototypes are presented to illustrate the capabilities of these new techniques. For example, soft lithography and surface chemistry are employed for chemically patterning substrates, stenciling biological entities onto substrates and confining solutions. As a result, information-rich, highdensity bioassays can be obtained where biological targets, surface properties and medium solutions are carefully determined and controlled.