Specific PAF antagonist WEB‐2086 induces terminal differentiation of murine and human leukemia cells

Abstract

A pharmacological approach to neoplasia by differentiation therapy relies on the availability of cytodifferentiating agents whose antitumor efficacy is usually assayed first on malignant cells in vitro. Using murine erythroleukemia cells (MELCs) as the model, we found that WEB-2086, a triazolobenzodiazepine-derived PAF antagonist originally developed as an anti-inflammatory drug, induces a dose-dependent inhibition of MELC growth and hemoglobin accumulation as a result of a true commitment to differentiation. MELCs treated for 5 days with 1 mM WEB-2086 show greater than or equal to 85% benzidine-positive cells, increased expression of alpha- and beta-globin genes, and down-regulation of c-Myb. This differentiation pattern, which does not involve histone H4 acetylation and is abrogated by the action of phorbol 12-myristate 13-acetate, recalls the pattern induced by hexamethylene bisacetamide (HMBA). In addition to MELCs, human erythroleukemia K562 and HEL and myeloid HL60 cells are massively committed to maturation by WEB-2086 and, with some differences, by its analog, WEB-2170. This suggests that WEB-2086, structurally distant from other known inducers, might be a member of a new class of cytodifferentiation agents active on a broad range of transformed cells in vitro and useful, prospectively, for anticancer therapy due to their high tolerability in vivo.

WEB-2086 and WEB-2170 trigger apoptosis in both ATRA-sensitive and -resistant promyelocytic leukemia cells and greatly enhance ATRA differentiation potential

“…This confirmed our previous observations on functional similarities between HMBA and WEBs exerting no HDACi activity 14 …”

“…This hypothesis is supported by the facts that (a) WEBs -originally synthesized from benzodiazepineshave maintained the ability to bind PBR; 27 (b) specific PBRligands were reported to induce differentiation and/or apoptosis in a variety of malignant cells including human leukemia cell lines, 28 as well as to sensitize acute myeloid leukemia cells to cytotoxic agents and elicit apoptosis by altering mitochondrial membrane potential. 29 Overall, our findings showed that WEBs can be regarded as very attractive and multifaced molecules which, beyond exerting anti-inflammatory, 30 antineoangiogenetic 31 and antiproliferative and differentiative activities, 14 also have the ability of inducing in vitro apoptosis of both ATRA-sensitive andresistant APL established cell lines as well as primary blasts from patients with t(15;17) APL. Moreover, highly effective synergism Cultures were maintained for 3 days without (control) and with either 1 mM WEB-2086, 0.5 mM WEB-2170 or 1 mM ATRA.…”

“…14 Here, we report that WEBs, as single agents, exerted a marked apoptotic effect in vitro on both ATRA-sensitive andresistant APL cell lines and on blasts from patients with t(15;17) APL. Moreover, it was found that suboptimal amounts of WEBs, which could not trigger apoptosis themselves, were, nevertheless, very effective in enhancing up to 40-fold ATRA differentiation potential in NB4 cells to bring benefits to pharmacological treatment of APL.…”

Growth inhibition and differentiation of human breast cancer cells by the PAFR antagonist WEB-2086

“…For ERa, 5 0 -CAAGCCCGCTCATGATCA-3 0 and 5 0 -CACCATGCCCTCTACACA-3 0 (388 bp); for peripheral benzodiazepine receptor (PBR), 5 0 -CACGCTCTACTCAG-CCATGG-3 0 and 5 0 -GCAGTAGTTGAGTGTGGTCGC-3 0 (298 bp); for PAFR, 5 0 -A CCAACACAGTGCCCGACAGTGCT-3 0 and 5 0 -GGGTGACCTGATG TGCATCA-TTAAT-3 0 (363 bp); for b 2 -microglobulin, 5 0 -CTCGCG CTACTCTCT-CTTTCT-3 0 and 5 0 -ACATGGAGACAGCACTCAA AG-3 0 (514 bp). Reverse transcription-polymerase chain reaction (RT -PCR) products were analysed as described previously (Cellai et al, 2002).…”

Mechanistic insight into WEB-2170-induced apoptosis in human acute myelogenous leukemia cells: The crucial role of PTEN

“…Indeed, epigenetic modulators are functional to set up an environment permissive for cell differentiation and/or apoptosis, but may not always be sufficient themselves to trigger these processes without the cooperation of other proactive signals. It is of great interest, therefore, to identify and characterize new agents capable of inducing cytostasis, differentiation, and apoptosis in AML cells, and supporting the action of epigenetic modulators and conventional chemotherapy so as to decrease effective therapeutic doses and untoward consequences to the host.We previously showed that WEB-2170 da ligand of platelet-activating factor receptor (PAF-r) originally synthesized from an anxiolytic triazolobenzodiazepine [11,12]d could also promote murine erythroleukemia cell maturation [13] and trigger apoptosis in ATRA-sensitive and -resistant APL cell lines, and in blasts from patients with APL [14]. The present study aimed to understand mechanisms of WEB-2170-induced cytostasis and apoptosis in the APL cell line NB4, as well as to assess drug efficacy in other AML cell lines and, eventually, ex vivo in blasts from patients with different (M0 through M5) AML subtypes.…”