SCD1 inhibition causes cancer cell death by depleting mono-unsaturated fatty acids.

Increased metabolism is a requirement for tumor cell proliferation. To understand the dependence of tumor cells on fatty acid metabolism, we evaluated various nodes of the fatty acid synthesis pathway. Using RNAi we have demonstrated that depletion of fatty-acid synthesis pathway enzymes SCD1, FASN, or ACC1 in HCT116 colon cancer cells results in cytotoxicity that is reversible by addition of exogenous fatty acids. This conditional phenotype is most pronounced when SCD1 is depleted. We used this fatty-acid rescue strategy to characterize several small-molecule inhibitors of fatty acid synthesis, including identification of TOFA as a potent SCD1 inhibitor, representing a previously undescribed activity for this compound. Reference FASN and ACC inhibitors show cytotoxicity that is less pronounced than that of TOFA, and fatty-acid rescue profiles consistent with their proposed enzyme targets. Two reference SCD1 inhibitors show low-nanomolar cytotoxicity that is offset by at least two orders of magnitude by exogenous oleate. One of these inhibitors slows growth of HCT116 xenograft tumors. Our data outline an effective strategy for interrogation of on-mechanism potency and pathway-node-specificity of fatty acid synthesis inhibitors, establish an unambiguous link between fatty acid synthesis and cancer cell survival, and point toward SCD1 as a key target in this pathway.

Implications of promiscuous Pim-1 kinase fragment inhibitor hydrophobic interactions for fragment-based drug design.

We have studied the subtleties of fragment docking and binding using data generated in a Pim-1 kinase inhibitor program. Crystallographic and docking data analyses have been undertaken using inhibitor complexes derived from an in-house surface plasmon resonance (SPR) fragment screen, a virtual needle screen, and a de novo designed fragment inhibitor hybrid. These investigations highlight that fragments that do not fill their binding pocket can exhibit promiscuous hydrophobic interactions due to the lack of steric constraints imposed on them by the boundaries of said pocket. As a result, docking modes that disagree with an observed crystal structure but maintain key crystallographically observed hydrogen bonds still have potential value in ligand design and optimization. This observation runs counter to the lore in fragment-based drug design that all fragment elaboration must be based on the parent crystal structure alone.

Bioinspired total synthesis and human proteasome inhibitory activity of (-)-salinosporamide A, (-)-homosalinosporamide A, and derivatives obtained via organonucleophile promoted bis-cyclizations.

A full account of concise, enantioselective syntheses of the anticancer agent (-)-salinosporamide A and derivatives, including (-)-homosalinosporamide, that was inspired by biosynthetic considerations is described. The brevity of the synthetic strategy stems from a key bis-cyclization of a ß-keto tertiary amide, which retains optical purity enabled by A(1,3)-strain rendering slow epimerization relative to the rate of bis-cyclization. Optimization studies of the key bis-cyclization, enabled through byproduct isolation and characterization, are described that ultimately allowed for a gram scale synthesis of a versatile bicyclic core structure with a high degree of stereoretention. An optimized procedure for zincate generation by the method of Knochel, generally useful for the synthesis of salino A derivatives, led to dramatic improvements in side-chain attachment and a novel diastereomer of salino A. The versatility of the described strategy is demonstrated by the synthesis of designed derivatives including (-)-homosalinosporamide A. Inhibition of the human 20S and 26S proteasome by these derivatives using an enzymatic assay are also reported. The described total synthesis of salino A raises interesting questions regarding how biosynthetic enzymes leading to the salinosporamides proceeding via optically active ß-keto secondary amides, are able to maintain the stereochemical integrity at the labile C2 stereocenter or if a dynamic kinetic resolution is operative.

Zebrafish: an animal model for toxicological studies.

Zebrafish (Danio rerio) has been extensively studied and well described for environmental toxicity studies. Molecular biology and genetics have recently been used to elucidate the underlying mechanisms of toxicity in zebrafish and to predict effects in mammals. The versatile zebrafish is now incorporated in many areas of toxicological programs for assessing human risk and for preclinical drug discovery and screening.