Gossypol, a BH3 mimetic, induces apoptosis in chronic lymphocytic leukemia cells

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Blood First Edition Paper, prepublished online June 19, 2008; DOI

10.1182/blood-2007-12-126946.

Gossypol, a BH3 mimetic, induces apoptosis in chronic lymphocytic leukemia cells

Kumudha Balakrishnan, G Wierda, J Keating, and Varsha Gandhi*

Experimental Therapeutics, M.D. Cancer Center, Houston, TX

Leukemia Department, M.D. Cancer Center, Houston, TX

* Corresponding author; email: vgandhi@... .

Gossypol, a cottonseed extract derivative, acts as a BH3-mimetic, binding to the

BH3 pocket of anti-apoptotic proteins and displacing pro-death partners to

induce apoptosis. However, knowledge on the molecular underpinnings of its

downstream effects is limited. Since chronic lymphocytic leukemia (CLL) cells

express high levels of anti-apoptotic proteins that act as a survival mechanism

for these replicationally quiescent lymphocytes, we investigated whether

gossypol induces apoptosis in these cells and what mechanism underlies

gossypol-mediated cytotoxicity. Gossypol induced cell death in a concentration-

and time-dependent manner; 24-hour incubation with 30 µM gossypol resulted in

50% cell death (median; range, 10-80%; n = 47) that was not abrogated by

pan-specific caspase inhibitor. Starting at 4 hours, the mitochondrial outer

membrane was significantly permeabilized (median, 77%; range, 54-93%; n = 15).

Mitochondrial outer membrane permeablistaion (MOMP) was concurrent with

increased production of reactive oxygen species (ROS); however, antioxidants did

not abrogate gossypol-induced cell death. Mitochondrial membrane

permeabilization was also associated with loss of intracellular ATP, activation

of BAX, and release of cytochrome c and apoptosis-inducing factor (AIF), which

was translocated to the nucleus. Blocking AIF translocation resulted in a

decreased apoptosis, suggesting that AIF contributes to gossypol-mediated

cytotoxicity in CLL lymphocytes.