Endothelin receptor B antagonists decrease glioma cell viability independently of their cognate receptor.

Abstract

BACKGROUND:

Endothelin receptor antagonists inhibit the progression of many cancers, but research into their influence on glioma has been limited.

METHODS:

We treated glioma cell lines, LN-229 and SW1088, and melanoma cell lines, A375 and WM35, with two endothelin receptor type B (ETRB)-specific antagonists, A-192621 and BQ788, and quantified viable cells by the capacity of their intracellular esterases to convert non-fluorescent calcein AM into green-fluorescent calcein. We assessed cell proliferation by labeling cells with carboxyfluorescein diacetate succinimidyl ester and quantifying the fluorescence by FACS analysis. We also examined the cell cycle status using BrdU/propidium iodide double staining and FACS analysis. We evaluated changes in gene expression by microarray analysis following treatment with A-192621 in glioma cells. We examined the role of ETRB by reducing its expression level using small interfering RNA (siRNA).

RESULTS:

We report that two ETRB-specific antagonists, A-192621 and BQ788, reduce the number of viable cells in two glioma cell lines in a dose- and time-dependent manner. We describe similar results for two melanoma cell lines. The more potent of the two antagonists, A-192621, decreases the mean number of cell divisions at least in part by inducing a G2/M arrest and apoptosis. Microarray analysis of the effects of A-192621 treatment reveals up-regulation of several DNA damage-inducible genes. These results were confirmed by real-time RT-PCR. Importantly, reducing expression of ETRB with siRNAs does not abrogate the effects of either A-192621 or BQ788 in glioma or melanoma cells. Furthermore, BQ123, an endothelin receptor type A (ETRA)-specific antagonist, has no effect on cell viability in any of these cell lines, indicating that the ETRB-independent effects on cell viability exhibited by A-192621 and BQ788 are not a result of ETRA inhibition.

CONCLUSION:

While ETRB antagonists reduce the viability of glioma cells in vitro, it appears unlikely that this effect is mediated by ETRB inhibition or cross-reaction with ETRA. Instead, we present evidence that A-192621 affects glioma and melanoma viability by activating stress/DNA damage response pathways, which leads to cell cycle arrest and apoptosis. This is the first evidence linking ETRB antagonist treatment to enhanced expression of DNA damage-inducible genes.

• Authors: Jennifer P Montgomery, Paul H Patterson
• Journal: BMC cancer (BMC Cancer) Vol. 8 Pg. 354 (Nov 28 2008) ISSN: 1471-2407 [Electronic] England
• PMID: 19040731 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• A 192621
• Endothelin B Receptor Antagonists
• Isoquinolines
• Pyrroles
• Pyrrolidines
• RNA, Small Interfering
• Receptor, Endothelin A
• Receptor, Endothelin B
• WM5 compound
• Ciprofloxacin

Topics

• Apoptosis (drug effects)
• Cell Cycle (drug effects)
• Cell Line, Tumor
• Cell Proliferation (drug effects)
• Cell Survival (drug effects)
• Ciprofloxacin (analogs & derivatives, pharmacology)
• DNA Damage (drug effects, genetics)
• Endothelin B Receptor Antagonists
• Gene Expression Profiling
• Gene Expression Regulation, Neoplastic
• Glioma (genetics, metabolism, pathology)
• Humans
• Isoquinolines
• Melanoma (genetics, metabolism, pathology)
• Oligonucleotide Array Sequence Analysis
• Pyrroles
• Pyrrolidines (pharmacology)
• RNA, Small Interfering
• Receptor, Endothelin A (metabolism)
• Receptor, Endothelin B (genetics, metabolism)