Dual mechanisms of sHA 14-1 in inducing cell death through endoplasmic reticulum and mitochondria.

Abstract

HA 14-1 is a small-molecule Bcl-2 antagonist that promotes apoptosis in malignant cells, but its mechanism of action is not well defined. We recently reported that HA 14-1 has a half-life of only 15 min in vitro, which led us to develop a stable analog of HA 14-1 (sHA 14-1). The current study characterizes its mode of action. Because of the antiapoptotic function of Bcl-2 family proteins on the endoplasmic reticulum (ER) and mitochondria, the effect of sHA 14-1 on both organelles was evaluated. sHA 14-1 induced ER calcium release in human leukemic cells within 1 min, followed by induction of the ER stress-inducible transcription factor ATF4. Similar kinetics and stronger intensity of ER calcium release were induced by the sarcoendoplasmic reticulum Ca(2+)-ATPase (SERCA) inhibitor thapsigargin, accompanied by similar kinetics and intensity of ATF4 induction. sHA 14-1 directly inhibited SERCA enzymatic activity but had no effect on the inositol triphosphate receptor. Evaluation of the mitochondrial pathway showed that sHA 14-1 triggered a loss of mitochondrial transmembrane potential (Delta psi m) and weak caspase-9 activation, whereas thapsigargin had no effect. (R)-4-(3-Dimethylamino-1-phenylsulfanylmethyl-propylamino)-N-{4-[4-(4'-chloro-biphenyl-2-ylmethyl)-piperazin-1-yl]-benzoyl}-3-nitrobenzenesulfonamide (ABT-737), a well established small-molecule Bcl-2 antagonist, rapidly induced loss of Delta psi m and caspase-9 activation but had no effect on the ER. The pan-caspase inhibitor N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone had some protective effect on sHA 14-1-induced cell death. These collective results suggest a unique dual targeting mechanism of sHA 14-1 on the apoptotic resistance machinery of tumor cells that includes antiapoptotic Bcl-2 family proteins and SERCA proteins.

• Authors: David Hermanson, Sadiya N Addo, Anna A Bajer, Jonathan S Marchant, Sonia Goutam Kumar Das, Balasubramanian Srinivasan, Fawaz Al-Mousa, Francesco Michelangeli, David D Thomas, Tucker W Lebien, Chengguo Xing
• Journal: Molecular pharmacology (Mol Pharmacol) Vol. 76 Issue 3 Pg. 667-78 (Sep 2009) ISSN: 1521-0111 [Electronic] United States
• PMID: 19561125 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't)

Chemical References

• Benzopyrans
• Nitriles
• Proto-Oncogene Proteins c-bcl-2
• ethyl 2-amino-6-(3,5-dimethoxyphenyl)-4-(2-ethoxy-2-oxoethyl)-4H-chromene-3-carboxylate
• ethyl 2-amino-6-bromo-4-(1-cyano-2-ethoxy-2-oxoethyl)-4H-chromene-3-carboxylate
• Sarcoplasmic Reticulum Calcium-Transporting ATPases

Topics

• Apoptosis
• Benzopyrans (chemistry, pharmacology)
• Cell Line, Tumor
• Endoplasmic Reticulum (drug effects, metabolism)
• Humans
• Mitochondria (drug effects, metabolism)
• Nitriles (pharmacology)
• Proto-Oncogene Proteins c-bcl-2 (antagonists & inhibitors, metabolism)
• Sarcoplasmic Reticulum Calcium-Transporting ATPases (antagonists & inhibitors, metabolism)