Small molecule XIAP inhibitors sensitize childhood acute leukemia cells for CD95‐induced apoptosis

Abstract

Escape of apoptosis may contribute to treatment failure in childhood acute lymphoblastic leukemia (ALL) calling for new approaches to overcome apoptosis resistance. Here, we provide for the first time evidence that small molecule inhibitors that target the anti-apoptotic protein X-linked inhibitor of apoptosis (XIAP) sensitize ALL cells for CD95-induced apoptosis. XIAP inhibitors at subtoxic concentrations, but not a structurally related control compound, act synergistically with agonistic anti-CD95 antibodies or MegaFasL, a hexameric form of CD95 ligand, to induce apoptosis in ALL cells. Further, XIAP inhibitors co-operate with MegaFasL to reduce clonogenic survival of ALL cells demonstrating their effect also on long-term survival. In contrast, XIAP inhibitors show little effect on MegaFasL-mediated apoptosis in normal peripheral blood lymphocytes (PBLs), pointing to some tumor selectivity. Molecular studies reveal that XIAP inhibitors enhance CD95-induced activation of caspases, loss of mitochondrial membrane potential and cytochrome c release in a caspase-dependent manner. Importantly, XIAP inhibitors sensitize primary leukemic blasts from children with ALL for MegaFasL-induced apoptosis. Thus, small molecule XIAP inhibitors present a promising novel approach to enhance CD95-induced apoptosis in childhood acute leukemia.Acute lymphocytic leukemia (ALL) presents the most frequent malignancy of childhood. 1 Although the overall prognosis for children with ALL is good with a cure rate of 80%, the outcome of patients who relapse remains poor. 1,2 Thus, novel approaches are required to overcome resistance mechanisms in a subgroup of children with ALL. 3 Resistance to apoptosis (programmed cell death) is one of the hallmarks of human cancers including leukemia. 4-7 Apoptosis can be induced via two major pathways, i.e., the death receptor (extrinsic) pathway or the mitochondrial (extrinsic) pathway. 8,9 Ligation of death receptors of the tumor necrosis factor (TNF) receptor superfamily such as CD95 (APO-1/ Fas) by their cognate ligands or agonistic antibodies results in caspase-8 activation at the death inducing signaling complex (DISC) which induces direct cleavage of downstream effector caspases such as caspase-3. 10 In the mitochondrial pathway the release of apoptogenic factors such as cytochrome c, apoptosis inducing factor (AIF), second mitochondria-derived activator of caspase (Smac)/direct IAP binding protein with low pI (DIABLO) or Omi/high temperature requirement protein A2 (HtrA2) from mitochondria into the cytosol results in caspase-3 activation via formation of the cytochrome c/Apaf-1/caspase-9-containing apoptosome complex. 11 The failure of antileukemic therapy has been largely attributed to evasion of apoptosis, because many anticancer treatments including chemo-, radio-or immuno-therapy primarily act by triggering apoptotic cell death in cancer cells. 9 In principle, resistance to apoptosis may be caused by the lack or reduced function of pro-apoptotic molecules or alternatively, by the dominance o...

RIP1 is required for IAP inhibitor-mediated sensitization of childhood acute leukemia cells to chemotherapy-induced apoptosis

“…As the IAP inhibitors antagonize XIAP besides cIAP1 and cIAP2, 18 the neutralization of the XIAPimposed inhibition of effector caspases by IAP inhibitors might contribute to promote IAP inhibitor-and AraC-induced apoptosis, as we recently reported for death receptor-induced apoptosis. 17,28 Also, our findings indicate that DNA damage might promote RIP1/FADD/caspase-8 formation independently of autocrine/paracrine TNFa signaling.…”

Smac mimetic and glucocorticoids synergize to induce apoptosis in childhood ALL by promoting ripoptosome assembly

“…Previously, we reported that small-molecule inhibitors of IAP proteins such as Smac mimetics act in concert with other cytotoxic stimuli, for example death receptor ligands such as TRAIL and CD95 ligand, chemotherapeutic drugs, or irradiation, in ALL or other cancer entities. 24,25,38,[40][41][42][43] Second, we identify a novel underlying molecular mechanism of this synergistic interaction by showing that BV6 and dexamethasone act in concert to trigger proteasomal degradation of IAP proteins, which in turn promotes the assembly of the ripoptosome, a cytosolic cell death complex ( Figure 6). Several lines of evidence support this conclusion, as follows:…”

“…22 We previously reported that small-molecule IAP inhibitors enhance death receptoror chemotherapy-induced apoptosis in childhood leukemia. [23][24][25] However, the question of whether or not targeting IAP proteins represents a suitable strategy to bypass GC resistance in ALL has not yet been answered. Therefore, in the present study we investigated the question of whether or not neutralization of XIAP, cIAP1, and cIAP2 by the small-molecule Smac mimetic BV6 19 can enhance the antileukemic activity of GC against childhood ALL.…”

Molecular Pathways: Targeting Death Receptors and Smac Mimetics

“…For example, in ALL subtoxic concentrations of IAP inhibitors were described to synergistically induce apoptosis and to reduce clonogenic survival in combination with agonistic anti-CD95 antibodies or MegaFas ligand, a hexameric form of CD95 ligand, whereas they failed to sensitize normal peripheral blood lymphocytes for CD95-mediated apoptosis (47). The clinical relevance of this combination strategy was underscored by experiments performed in primary leukemic blasts freshly isolated from children with ALL, which similarly showed a synergistic induction of apoptosis of IAP inhibitors and MegaFas ligand (47). Also, the Smac mimetic BV6 was shown to sensitize Jurkat T-ALL cells that express key components of the death receptor pathway such as FADD or caspase-8 to CD95-mediated apoptosis, whereas FADD-or caspase-8-deficient cells remained resistant to this combination (48).…”