Targeting aberrant PI3K/Akt activation by PI103 restores sensitivity to TRAIL-induced apoptosis in neuroblastoma.

PURPOSE

Because we recently identified Akt activation as a novel poor prognostic indicator in neuroblastoma, we investigated whether phosphoinositide 3'-kinase (PI3K) inhibition sensitizes neuroblastoma cells for TRAIL-induced apoptosis.

EXPERIMENTAL DESIGN

The effect of pharmacological or genetic inhibition of PI3K or mTOR was analyzed on apoptosis induction, clonogenic survival, and activation of apoptosis signaling pathways in vitro and in a neuroblastoma in vivo model. The functional relevance of individual Bcl-2 family proteins was examined by knockdown or overexpression experiments.

RESULTS

The PI3K inhibitor PI103 cooperates with TRAIL to synergistically induce apoptosis (combination index < 0.1), to suppress clonogenic survival, and to reduce tumor growth in a neuroblastoma in vivo model. Similarly, genetic silencing of PI3K significantly increases TRAIL-mediated apoptosis, whereas genetic or pharmacological blockage of mTOR fails to potentiate TRAIL-induced apoptosis. Combined treatment with PI103 and TRAIL enhances cleavage of Bid and the insertion of tBid into mitochondrial membranes, and reduces phosphorylation of Bim(EL). Additionally, PI103 decreases expression of Mcl-1, XIAP, and cFLIP, thereby promoting Bax/Bak activation, mitochondrial perturbations, and caspase-dependent apoptosis. Knockdown of Bid or Noxa or overexpression of Bcl-2 rescues cells from PI103- and TRAIL-induced apoptosis, whereas Mcl-1 silencing potentiates apoptosis. Bcl-2 overexpression also inhibits cleavage of caspase-3, caspase-8, and Bid pointing to a mitochondria-driven feedback amplification loop.

CONCLUSIONS

PI103 primes neuroblastoma cells for TRAIL-induced apoptosis by shifting the balance toward proapoptotic Bcl-2 family members and increased mitochondrial apoptosis. Thus, PI3K inhibitors represent a novel promising approach to enhance the efficacy of TRAIL-based treatment protocols in neuroblastoma.