Engineering threshold-based selection systems.

Using model organisms to identify novel therapeutic targets is frequently constrained by pre-existing genetic toolkits. To expedite positive selection for identification of novel downstream effectors, we engineered conditional expression of activated CED-10/Rac to disrupt Caenorhabditis elegans embryonic morphogenesis, titrated to 100% lethality. The strategy of engineering thresholds for positive selection using experimental animals was validated with pharmacological and genetic suppression and is generalizable to diverse molecular processes and experimental systems.

Indinavir Increases Midazolam -Glucuronidation in Humans: Identification of an Alternate CYP3A Inhibitor Using an In Vitro to In Vivo Approach.

Midazolam is a widely used index substrate for assessing effects of xenobiotics on CYP3A activity. A previous study involving human hepatocytes showed the primary route of midazolam metabolism, 1'-hydroxylation, shifted to -glucuronidation in the presence of the CYP3A inhibitor ketoconazole, which may lead to an overprediction of the magnitude of a xenobiotic-midazolam interaction. Because ketoconazole is no longer recommended as a clinical CYP3A inhibitor, indinavir was selected as an alternate CYP3A inhibitor to evaluate the contribution of the -glucuronidation pathway to midazolam metabolism.

Selection of Priority Natural Products for Evaluation as Potential Precipitants of Natural Product-Drug Interactions: A NaPDI Center Recommended Approach.

Pharmacokinetic interactions between natural products (NPs) and conventional medications (prescription and nonprescription) are a longstanding but understudied problem in contemporary pharmacotherapy. Consequently, there are no established methods for selecting and prioritizing commercially available NPs to evaluate as precipitants of NP-drug interactions (NPDIs). As such, NPDI discovery remains largely a retrospective, bedside-to-bench process.

Impact of organic solvents on cytochrome P450 probe reactions: filling the gap with (S)-Warfarin and midazolam hydroxylation.

(S)-Warfarin 7-hydroxylation and midazolam 1'-hydroxylation are among the preferred probe substrate reactions for CYP2C9 and CYP3A4/5, respectively. The impact of solvents on enzyme activity, kinetic parameters, and predicted in vivo hepatic clearance (Cl(H)) associated with each reaction has not been evaluated. The effects of increasing concentrations [0.

Genetic and functional characterization of putative Ras/Raf interaction inhibitors in C. elegans and mammalian cells.

Background: Activation of the mammalian Ras-Raf-MEK-ERK MAPK signaling cascade promotes cellular proliferation, and activating Ras mutations are implicated in cancer onset and maintenance. This pathway, a therapeutic target of interest, is highly conserved and required for vulval development in C. elegans.

Use of Caenorhabditis elegans to evaluate inhibitors of Ras function in vivo.

The human RAS genes constitute the most frequently mutated oncogenes in human cancers, and the critical role of aberrant Ras protein function in oncogenesis is well established. Consequently, considerable effort has been devoted to the development of anti-Ras inhibitors for cancer treatment. An important facet of molecularly targeted cancer drug discovery is the validation of a target-based mechanism of action, as well as the identification of potential off-target effects.