Citations

Abstract Purpose: Prosurvival Bcl-2 proteins inhibit the mitochondrial and death receptor-mediated apoptotic pathways. Obatoclax is a small-molecule antagonist of the BH3-binding groove of Bcl-2 proteins that may enhance tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) sensitivity and efficacy. Experimental Design: Human pancreatic cancer cell lines were incubated with obatoclax and/or TRAIL and cell viability, Annexin V labeling, caspase cleavage, and cytochrome c release were measured. In drug-treated cell lines, protein-protein interactions were studied by immunoprecipitation. Bax/Bak activation was analyzed using conformation-specific antibodies. Lentiviral short hairpin RNA was used to knockdown Bim, Bid, and apoptosis-inducing factor (AIF) expression. Results: Obatoclax reduced the viability of PANC-1 and BxPC-3 cell lines and synergistically enhanced TRAIL-mediated cytotoxicity. Obatoclax enhanced TRAIL-mediated apoptosis, as shown by Annexin V labeling, which was accompanied by caspase activation (caspase-8, -9, and -3) and cleavage of Bid. Obatoclax potentiated TRAIL-mediated Bax/Bak activation and the release of mitochondrial cytochrome c, Smac, and AIF. Mechanisms underlying the apoptotic effect of obatoclax include displacement of Bak from its sequestration by Bcl-xL or Mcl-1 and release of Bim from Bcl-2 or Mcl-1. Bid knockdown by short hairpin RNA attenuated caspase cleavage and cytotoxicity of obatoclax plus TRAIL. Bim knockdown failed to inhibit the cytotoxic effect of obatoclax alone or combined with TRAIL yet attenuated TRAIL-mediated cytotoxicity. AIF knockdown attenuated cytotoxicity of the drug combination. Conclusions: Obatoclax potentiates TRAIL-mediated apoptosis by unsequestering Bak and Bim from Bcl-2/Bcl-xL or Mcl-1 proteins. This drug combination enhances Bid-mediated cross-talk between the mitochondrial and death receptor-mediated apoptotic pathways and may represent a novel therapeutic strategy against pancreatic cancer.

The pathobiology of medulloblastoma is reviewed in light of emerging data regarding its immunocytochemical and cytobiologic, as well as molecular biologic, characteristics. The nature of the lesion, particularly its nosologic relation to primitive neuroectodermal tumor, is discussed, as is its place in the World Health Organization classification of tumors of the central nervous system. ( J Child Neurol 1992;7:240-252).

Medulloblastoma, the most common embryonal tumor of the central nervous system, affects both children and adults. It poses a significant therapeutic challenge in that age-dependent differences exist, not only in their pathobiology, but in the efficacy of chemotherapy and radiotherapy. This is particularly the case in very young children, whose still developing nervous system exhibits a low tolerance to radiotherapy. We review the epidemiology, clinical presentation, radiologic features, and current therapeutic concepts relative to this unique neoplasm. Efforts are made to highlight clinical controversies. ( J Child Neurol 1992;7:142-155).

Abstract Purpose: The intrinsic drug resistance of colorectal cancers is related in part to overexpression of prosurvival Bcl-2 family proteins. We determined the effects of ABT-737, a small-molecule inhibitor of Bcl-2/Bcl-xL but not Mcl-1, on apoptosis induction alone and in combination with CPT-11 and explored mechanisms underlying their cooperativity. Experimental Design: Human colorectal carcinoma cell lines (HCT116 wild-type and Bax-/-, HT-29, and RKO) were incubated with ABT-737 alone and combined with CPT-11 or bortezomib, and cell viability, caspase cleavage, and Annexin V labeling were measured. In drug-treated cell lines, protein-protein interactions were analyzed by immunoprecipitation. Lentiviral short hairpin RNA was used to knockdown Noxa expression. Results: ABT-737 induced apoptosis in a dose-dependent manner and its coadministration with the topoisomerase I inhibitor, CPT-11, resulted in a synergistic cytotoxic effect. Apoptosis induction by the drug combination was associated with enhanced caspase-8, caspase-9, and caspase-3 activation and poly(ADP-ribose) polymerase cleavage that were completely abrogated in Bax knockout cells. ABT-737 unsequestered the BH3-only protein Bim from its complex with Bcl-xL or Bcl-2 and disrupted the interaction of Bcl-xL with Bak. CPT-11 treatment up-regulated Noxa expression, as did bortezomib, and enhanced Noxa/Mcl-1 complexes. CPT-11 also disrupted the Mcl-1/Bak interaction. Knockdown of Noxa using short hairpin RNA lentiviral constructs was shown to significantly attenuate the cytotoxic effect of CPT-11 or bortezomib combined with ABT-737 and inhibited caspase-3 cleavage. Conclusions: Induction of Noxa by CPT-11 or bortezomib can sensitize colorectal cancer cells expressing Mcl-1 to ABT-737. Up-regulation of Noxa may therefore represent an important strategy to enhance the therapeutic efficacy of ABT-737 against colorectal cancer and other solid tumors.

Abstract Purpose: Proapoptotic BH3-only proteins (Bim, Bad, Bid, Puma, and Noxa) initiate apoptosis by binding to regulatory sites on antiapoptotic Bcl-2 proteins, directly neutralizing their cytoprotective function. Expression of these proteins in colon cancer patients may account for differences in recurrence and survival rates. Experimental Design: Archival tumor-node-metastasis stage II and III primary colon carcinomas from patients treated in 5-fluorouracil–based adjuvant therapy trials were studied. Immunohistochemical analysis of Bim, Puma, and Noxa proteins was done using tissue microarrays (n = 431). Immunoscores were determined and correlated with clinicopathologic variables and disease-free survival (DFS) and overall survival (OS) rates. Results: Elevated expression of proapoptotic Bim (hazard ratio, 0.65; 95% confidence interval, 0.44-0.97; P = 0.033) and Puma (hazard ratio, 0.59; 95% confidence interval, 0.37-0.93; P = 0.022), but not Noxa, proteins in the tumor cytoplasm was significantly associated with more favorable OS in a univariate analysis, and elevated Bim expression was also associated with better DFS (P = 0.023). Patient age, tumor stage, and histologic grade were also prognostic. Multivariate Cox analysis showed that Bim (DFS, P = 0.030; OS, P = 0.045) and Puma (OS, P = 0.037) expression were independent predictors of OS after adjustment for histologic grade, tumor stage, age, and treatment. Furthermore, the combined variable of Bim and Puma was highly discriminant for both DFS (P = 0.0034) and OS (P = 0.0011). Conclusions: The proapoptotic BH3-only proteins Bim and Puma can provide prognostic information for stage II and III colon cancer patients receiving 5-fluorouracil–based adjuvant chemotherapy. Furthermore, our results support BH3-only proteins as molecular targets of novel anticancer drugs.

Abstract Purpose: Aberrant expression of T-cell coregulatory molecules has been investigated as a mechanism by which certain cancers may evade host immune surveillance. We evaluated expression of the T-cell coregulators B7-H1, B7-H3, and PD-1 in urothelial cell carcinoma (UCC) of the bladder. Experimental Design: Immunohistochemistry for B7-H1, B7-H3, and PD-1 was done on paraffin-embedded sections from 318 consecutive patients with UCC who underwent radical cystectomy. Expression was correlated with clinicopathologic outcomes and postoperative survival. Results: B7-H3 was widely expressed in UCC, as 222 of 314 (70.7%) tumors showed positive staining. Expression of B7-H3 in UCC was significantly increased compared with adjacent, nontumor urothelium, as a median of 70% of tumor cells expressed B7-H3, compared with 20% of cells in nontumor specimens (P < 0.001). The increase in B7-H3 expression was independent of tumor stage (P = 0.13). Expression of B7-H1 by UCC tumors (P < 0.001) and PD-1 by tumor-infiltrating lymphocytes (P = 0.012) were significantly associated with increased pathologic stage. Patients who had received intravesical bacillus Calmette-Guerin before cystectomy tended to show increased expression of B7-H3 (P = 0.023) and PD-1 (P = 0.071) but were less likely to express B7-H1 (P = 0.027). Moreover, for the subset of patients with organ-confined disease (n = 167), B7-H1 expression independently predicted all-cause mortality after cystectomy (hazard ratio, 3.18; 95% confidence interval, 1.74-5.79; P < 0.001). Conclusions: B7-H3 is highly expressed in UCC across tumor stages, whereas B7-H1 and PD-1 expression are associated with advanced disease. B7-H1 expression predicts mortality after cystectomy for patients with organ-confined tumors. These molecules may represent novel diagnostic or prognostic markers, as well as therapeutic targets, for patients with UCC.