| Abstract
| - Since 1990, the National Cancer Institute has performed extensive in vitro screening ofcompounds for anticancer activity. To date, more than 70 000 compounds have been screenedfor their antiproliferation activities against a panel of 60 human cancer cell lines. We probedthis database to identify novel structural classes with a pattern of biological activity on thesecell lines similar to that of the phorbol esters. The iridals form such a structural class. Usingthe program Autodock, we show that the iridals dock to the same position on the C1b domainof protein kinase C δ as do the phorbol esters, with the primary hydroxyl group of the iridal atthe C3 position forming two hydrogen bonds with the amide group of Thr12 and with thecarbonyl group of Leu 21 and the aldehyde oxygen of the iridal forming a hydrogen bond withthe amide group of Gly23. Biological analysis of two iridals, NSC 631939 and NSC 631941,revealed that they bound to protein kinase C α with Ki values of 75.6 ± 1.3 and 83.6 ± 1.5 nM,respectively. Protein kinase C is now recognized to represent only one of five families of proteinswith C1 domains capable of high-affinity binding of diacylglycerol and the phorbol esters. NSC631939 and NSC 631941 bound to RasGRP3, a phorbol ester receptor that directly linksdiacylglycerol/phorbol ester signaling with Ras activation, with Ki values of 15.5 ± 2.3 and41.7 ± 6.5 nM, respectively. Relative to phorbol 12,13-dibutyrate, they showed 15- and 6-foldselectivity for RasGRP3. Both compounds caused translocation of green fluorescent proteintagged RasGRP3 expressed in HEK293 cells, and both compounds induced phosphorylation ofERK1/2, a downstream indicator of Ras activation, in a RasGRP3-dependent fashion. Weconclude that the iridals represent a promising structural motif for design of ligands for phorbolester receptor family members.
|
