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

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

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47 Scopus citations


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 Ca2+-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 (Δψm) and weak caspase-9 activation, whereas thapsigargin had no effect. (R)-4-(3-Dimethylamino-1- phenylsulfanylmethylpropylamino)-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 Δψm 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.

Original languageEnglish (US)
Pages (from-to)667-678
Number of pages12
JournalMolecular Pharmacology
Issue number3
StatePublished - Sep 2009


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