Abstract 4500: What distinguishes a PKC activator like phorbol ester from a PKC functional antagonist like bryostatin 1? Bryologues permit mechanistic insight
Authors: Kedei Noemi , Telek Andrea , Czifra Gabriella , Czap Alexandra , Chen Jinqiu , Goldsmith Paul K. , Keck Gary E. , Blumberg Peter M. 
Published: February 2011

Abstract
Bryostatin 1, a promising anticancer agent in clinical trials, is a potent PKC activator in vitro that paradoxically often antagonizes the effects of the typical PKC activator phorbol 12-myristate 13-acetate (PMA) in cellular systems. Understanding the underlying mechanism(s) responsible for this functional PKC antagonism should afford a new generation of drugs for PKC, a validated therapeutic target. We use the synthetic bryostatin look-alike (bryologue) Merle 23 that differs structurally from bryostatin 1 to only a modest degree as a tool to dissect mechanisms responsible for the unique effects of bryostatin 1. We have previously reported that Merle 23 is PMA like, bryostatin-1 like, intermediate or unique in its behavior depending on the system and response examined. In LNCaP prostate cancer cells Merle 23 is bryostatin 1 like in failing to inhibit cell proliferation, to induce apoptosis, or to induce secretion of TNF-alpha, responses induced by PMA. Using siRNAs and PKC inhibitors we show that PKC delta, but not PKC alpha or epsilon, was responsible for induction of apoptosis. Nuclear localized PKC delta was previously shown to be responsible for the apoptotic response, and we found that Merle 23 resembled bryostatin 1 and was unlike PMA in that failed to induce translocation of PKC delta and phospho-PKD1 to the nucleus. In contrast, Merle 23 was PMA-like in inducing nuclear translocation of PKC alpha and PKC epsilon. Bryostatin 1 is unique in down-regulating PKC alpha very efficiently in LNCaP cells and Merle 23 is unique for quick and very efficient down-regulation of PKC delta. If proteosome inhibitors were used to prevent down-regulation of PKC delta, the bryostatin 1-like effects of Merle 23 on proliferation and TNF-alpha secretion were transformed to PMA-like effects: Merle 23 inhibited LNCaP cell proliferation and induced secretion of TNF-alpha when co-applied with lactacystin or MG-132. Similarly, for tyrosine phosphorylation of PKC delta at position 311 and for cFos activation, responses where Merle 23 had effects intermediate between bryostatin 1 and PMA, Merle 23 became more PMA-like when co-applied with proteosome inhibitors. Finally, consistent with stability of PKC signaling elements varying between ligands, we found that phosphorylation of ERK, Mek1 and Mek2, and AKT in response to bryostatin 1, PMA, and Merle 23 was similar at 30 min but returned to the basal level at 150 min for bryostatin 1, but not PMA or Merle 23, as determined using capillary iso-electrofocusing immune-assay. We conclude that differential regulation of PKC isoforms by bryostatin and the bryologues is an important contributor to their differential biological activity relative to PMA.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4500.