Constitutive IP(3) signaling underlies the sensitivity of B-cell cancers to the Bcl-2/IP(3) receptor disruptor BIRD-2

Anti-apoptotic Bcl-2 proteins are upregulated in different cancers, including diffuse large B-cell lymphoma (DLBCL) and chronic lymphocytic leukemia (CLL), enabling survival by inhibiting pro-apoptotic Bcl-2-family members and inositol 1,4,5-trisphosphate (IP(3)) receptor (IP(3)R)-mediated Ca(2+)-signaling. A peptide tool (Bcl-2/IP(3)R Disruptor-2; BIRD-2) was developed to abrogate the interaction of Bcl-2 with IP(3)Rs by targeting Bcl-2′s BH4 domain. BIRD-2 triggers cell death in primary CLL cells and in DLBCL cell lines. Particularly, DLBCL cells with high levels of IP(3)R2 were sensitive to BIRD-2. Here, we report that BIRD-2-induced cell death in DLBCL cells does not only depend on high IP(3)R2-expression levels, but also on constitutive IP(3) signaling, downstream of the tonically active B-cell receptor. The basal Ca(2+) level in SU-DHL-4 DLBCL cells was significantly elevated due to the constitutive IP(3) production. This constitutive IP(3) signaling fulfilled a pro-survival role, since inhibition of phospholipase C (PLC) using U73122 (2.5 µM) caused cell death in SU-DHL-4 cells. Milder inhibition of IP(3) signaling using a lower U73122 concentration (1 µM) or expression of an IP(3) sponge suppressed both BIRD-2-induced Ca(2+) elevation and apoptosis in SU-DHL-4 cells. Basal PLC/IP(3) signaling also fulfilled a pro-survival role in other DLBCL cell lines, including Karpas 422, RI-1 and SU-DHL-6 cells, whereas PLC inhibition protected these cells against BIRD-2-evoked apoptosis. Finally, U73122 treatment also suppressed BIRD-2-induced cell death in primary CLL, both in unsupported systems and in co-cultures with CD40L-expressing fibroblasts. Thus, constitutive IP(3) signaling in lymphoma and leukemia cells is not only important for cancer cell survival, but also represents a vulnerability, rendering cancer cells dependent on Bcl-2 to limit IP(3)R activity. BIRD-2 seems to switch constitutive IP(3) signaling from pro-survival into pro-death, presenting a plausible therapeutic strategy.