title: RasGRPs are targets of the anti-cancer agent ingenol-3-angelate.
abstract: Ingenol-3-angelate (I3A) is a non-tumor promoting phorbol ester-like compound identified in the sap of Euphoria peplus. Similar to tumor promoting phorbol esters, I3A is a diacylglycerol (DAG) analogue that binds with high affinity to the C1 domains of PKCs, recruits PKCs to cellular membranes and promotes enzyme activation. Numerous anti-cancer activities have been attributed to I3A and ascribed to I3A's effects on PKCs. We show here that I3A also binds to and activates members of the RasGRP family of Ras activators leading to robust elevation of Ras-GTP and engagement of the Raf-Mek-Erk kinase cascade. In response to I3A, recombinant proteins consisting of GFP fused separately to full-length RasGRP1 and RasGRP3 were rapidly recruited to cell membranes, consistent with direct binding of the compound to RasGRP's C1 domain. In the case of RasGRP3, IA3 treatment led to positive regulatory phosphorylation on T133 and activation of the candidate regulatory kinase PKCδ. I3A treatment of select B non-Hodgkin's lymphoma cell lines resulted in quantitative and qualitative changes in Bcl-2 family member proteins and induction of apoptosis, as previously demonstrated with the DAG analogue bryostatin 1 and its synthetic analogue pico. Our results offer further insights into the anticancer properties of I3A, support the idea that RasGRPs represent potential cancer therapeutic targets along with PKC, and expand the known range of ligands for RasGRP regulation.
authors.affiliation: Department of Biochemistry, University of Alberta, Edmonton, Alberta, Canada.
authors.forename: Xiaohua
authors.initials: X
authors.lastname: Song
authors.forename: Ana
authors.initials: A
authors.lastname: Lopez-Campistrous
authors.forename: Lucy
authors.initials: L
authors.lastname: Sun
authors.forename: Nancy A
authors.initials: NA
authors.lastname: Dower
authors.forename: Noemi
authors.initials: N
authors.lastname: Kedei
authors.forename: Jing
authors.initials: J
authors.lastname: Yang
authors.forename: Jessica S
authors.initials: JS
authors.lastname: Kelsey
authors.forename: Nancy E
authors.initials: NE
authors.lastname: Lewin
authors.forename: Tim E
authors.initials: TE
authors.lastname: Esch
authors.forename: Peter M
authors.initials: PM
authors.lastname: Blumberg
authors.forename: James C
authors.initials: JC
authors.lastname: Stone
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D002460:Cell Line; D004224:Diterpenes / Q000494:pharmacology*; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D018613:Microscopy, Confocal; D011494:Protein Kinases / Q000378:metabolism; D051381:Rats; D018631:ras Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism
chemical_list: C486592:3-ingenyl angelate; D000970:Antineoplastic Agents; D004224:Diterpenes; D011494:Protein Kinases; D018631:ras Proteins