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Research Articles: Targets

Discovery and characterization of inhibitors of human palmitoyl acyltransferases

¹ Apogee Biotechnology Corporation; ² Department of Pharmacology, Penn State College of Medicine, Hershey, Pennsylvania; ³ Department of Pharmaceutical Sciences, Medical University of South Carolina, Charleston, South Carolina; and ⁴ Department of Biochemistry and Molecular Biology, Penn State University, University Park, Pennsylvania

Requests for reprints: Charles D. Smith, Department of Pharmaceutical Sciences, Medical University of South Carolina, 280 Calhoun Street, Box 250140, Charleston, SC 29425. Phone: 843-792-3420; Fax: 843-792-3420. E-mail: smithchd{at}musc.edu

The covalent attachment of palmitate to specific proteins by the action of palmitoyl acyltransferases (PAT) plays critical roles in the biological activities of several oncoproteins. Two PAT activities are expressed by human cells: type 1 PATs that modify the farnesyl-dependent palmitoylation motif found in H- and N-Ras, and type 2 PATs that modify the myristoyl-dependent palmitoylation motif found in the Src family of tyrosine kinases. We have previously shown that the type 1 PAT HIP14 causes cellular transformation. In the current study, we show that mRNA encoding HIP14 is up-regulated in a number of types of human tumors. To assess the potential of HIP14 and other PATs as targets for new anticancer drugs, we developed three cell-based assays suitable for high-throughput screening to identify inhibitors of these enzymes. Using these screens, five chemotypes, with activity toward either type 1 or type 2 PAT activity, were identified. The activity of the hits were confirmed using assays that quantify the in vitro inhibition of PAT activity, as well as a cell-based assay that determines the abilities of the compounds to prevent the localization of palmitoylated green fluorescent proteins to the plasma membrane. Representative compounds from each chemotype showed broad antiproliferative activity toward a panel of human tumor cell lines and inhibited the growth of tumors in vivo. Together, these data show that PATs, and HIP14 in particular, are interesting new targets for anticancer compounds, and that small molecules with such activity can be identified by high-throughput screening. [Mol Cancer Ther 2006;5(7):1647–59]

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

⁵ J.D. Diller, unpublished data.

⁶ Z. Xia and C.D. Smith. Synthesis of fluorescent peptide substrates for cellular palmitoyl acyltransferase, In preparation.

⁷ http://chembridge.com/chembridge/compound.html

⁸ C.E. Ducker, et al. HIP14 directly affects Ras-mediated transformation, submitted for publication.

⁹ Y. Zhuang, et al. Synthesis and evaluation of substituted cyclohexyl-octahydro-pyrrolo[1,2-a]pyrazines that inhibit palmitoyl acyltransferases, in preparation.

¹⁰ J.M. Draper, et al. Cellular analyses of protein palmitoyl acyltransferases, submitted for publication.

Received 2/28/06; revised 5/ 2/06; accepted 5/12/06.

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