Safety and accuracy of 68Ga-DOTATOC PET/CT in children and young adults with solid tumors.

68Ga-DOTA-tyr3-Octreotide (68Ga-DOTATOC) PET/CT has been shown to have high accuracy in adults with neuroendocrine tumors, however has not been studied in pediatric patients. This study evaluated the safety and accuracy of 68Ga-DOTATOC PET/CT in children and young adults with solid tumors that express somatostatin receptor type 2. A series of three prospective, IRB approved, clinical trials evaluating safety and efficacy of 68Ga-DOTATOC PET/CT were conducted for subjects aged 6 months to 90 years. This study reports the results for the 26 children and young adults, aged 16 months to 29 years who participated in these trials. The administered activity of 68Ga-DOTATOC was 1.59 MBq/kg with an upper limit of 111 MBq for subjects < 18 years and 148 MBq for young adults. Safety was assessed with laboratory studies and patient/parent report of symptoms before and after the scan. Scans were interpreted in consensus by two board-certified nuclear medicine physicians. Each scan was categorized on a patient basis as true positive, true negative, false negative or false positive against a reference standard that included a combination of histopathology, other imaging modalities and clinical follow-up. Nine Grade I adverse events (AEs) occurred among 26 subjects, none of which were attributable to 68Ga-DOTATOC. Sensitivity of 68Ga-DOTATOC PET/CT was 88% (14 true positive, 2 false negative) and specificity was 100% (10 true negative, 0 false positive). 68Ga-DOTATOC PET/CT is safe and accurate in children and young adults with solid tumors expressing somatostatin receptor type 2.

Differentiation of small bowel and pancreatic neuroendocrine tumors by gene-expression profiling.

BACKGROUND: Between 10% and 20% of patients with neuroendocrine tumors (NETs) present with metastases of unknown primary site. Because knowledge of the primary site has important implications for treatment, we set out to define gene-expression profiles to differentiate between small-bowel NETs (SBNETs) and pancreatic NETs (PNETs). METHODS: RNA was extracted from tumor and normal tissues in 11 patients with SBNETs and 15 patients with PNETs, and qPCR was performed for 367 GPCR genes. Differentially expressed genes were identified using the RT2 Profiler. Whole genome expression analysis was performed on 11 SBNETs, 5 PNETS, and corresponding normal tissues. Statistical significance was evaluated by the Student t test and ANOVA. RESULTS: Whole-genome analysis revealed 173 significantly differentially expressed genes in SBNETs and normal tissues and in 52 in PNETs. GPCR arrays identified 28 genes in SBNETs and 18 in PNETs, with significant expression differences from normal tissues. In all SBNETs, 2 genes were significantly upregulated by more than fivefold: OXTR and GPR113. No PNETs shared this profile, whereas 73% had a greater than fivefold downregulation of ADORA1 and SCTR. These genes also allowed for determination of the primary site in 8 of 10 liver metastases. CONCLUSION: Differential expression patterns using as few as 2 to 4 GPCR genes successfully discriminated primary sites in small bowel and pancreatic NETs.

Characterization of intestinal and pancreatic dysfunction in VPAC1-null mutant mouse.

OBJECTIVES: These studies examined the effect of homozygous deletion of vasoactive intestinal peptide receptor type 1 (VPAC1) on development and function of intestines and pancreas. METHODS: Genetically engineered VPAC1-null mutant mice were monitored for growth, development, and glucose homeostasis. Expression of VPAC1 was examined during embryonic development using VPAC1 promoter-driven ß-galactosidase transgenic mice. RESULTS: Homozygous deletion of VPAC1 resulted in fetal, neonatal, and postweaning death owing to failure to thrive, intestinal obstruction, and hypoglycemia. Histological findings demonstrated disorganized hyperproliferation of intestinal epithelial cells with mucus deposition and bowel wall thickening. The pancreas demonstrated small dysmorphic islets of Langerhans containing α, ß, and δ cells. Expression of a VPAC1 promoter-driven transgene was observed in E12.5 and E14.5 intestinal epithelial and pancreatic endocrine cells. Vasoactive intestinal peptide receptor type 1-null mutant animals had lower baseline blood glucose levels compared to both heterozygous and wild-type littermates. Vasoactive intestinal peptide receptor type 1-deficient mice responded to oral glucose challenge with normal rise in blood glucose followed by rapid hypoglycemia and failure to restore baseline glucose levels. Insulin challenge resulted in profound hypoglycemia and inadequate glucose homeostasis in VPAC1-null mutant animals. CONCLUSIONS: These observations support a role for VPAC1 during embryonic and neonatal development of intestines and endocrine pancreas.

BH3 domains other than Bim and Bid can directly activate Bax/Bak.

Bcl-2 family proteins regulate a critical step in apoptosis referred to as mitochondrial outer membrane permeabilization (MOMP). Members of a subgroup of the Bcl-2 family, known as the BH3-only proteins, activate pro-apoptotic effectors (Bax and Bak) to initiate MOMP. They do so by neutralizing pro-survival Bcl-2 proteins and/or directly activating Bax/Bak. Bim and Bid are reported to be direct activators; however, here we show that BH3 peptides other than Bim and Bid exhibited various degrees of direct activation of the effector Bax or Bak, including Bmf and Noxa BH3s. In the absence of potent direct activators, such as Bim and Bid, we unmasked novel direct activator BH3 ligands capable of inducing effector-mediated cytochrome c release and liposome permeabilization, even when both Bcl-xL- and Mcl-1-type anti-apoptotic proteins were inhibited. The ability of these weaker direct activator BH3 peptides to cause MOMP correlated with that of the corresponding full-length proteins to induce apoptosis in the absence of Bim and Bid. We propose that, in certain contexts, direct activation by BH3-only proteins other than Bim and Bid may significantly contribute to MOMP and apoptosis.

Mitochondrial outer membrane proteins assist Bid in Bax-mediated lipidic pore formation.

Mitochondrial outer membrane permeabilization (MOMP) is a critical step in apoptosis and is regulated by Bcl-2 family proteins. In vitro systems using cardiolipin-containing liposomes have demonstrated the key features of MOMP induced by Bax and cleaved Bid; however, the nature of the "pores" and how they are formed remain obscure. We found that mitochondrial outer membranes contained very little cardiolipin, far less than that required for liposome permeabilization, despite their responsiveness to Bcl-2 family proteins. Strikingly, the incorporation of isolated mitochondrial outer membrane (MOM) proteins into liposomes lacking cardiolipin conferred responsiveness to cleaved Bid and Bax. Cardiolipin dependence was observed only when permeabilization was induced with cleaved Bid but not with Bid or Bim BH3 peptide or oligomerized Bax. Therefore, we conclude that MOM proteins specifically assist cleaved Bid in Bax-mediated permeabilization. Cryoelectron microscopy of cardiolipin-liposomes revealed that cleaved Bid and Bax produced large round holes with diameters of 25-100 nm, suggestive of lipidic pores. In sum, we propose that activated Bax induces lipidic pore formation and that MOM proteins assist cleaved Bid in this process in the absence of cardiolipin.

The antiproliferative agent MLN944 preferentially inhibits transcription.

MLN944 is a novel compound currently being codeveloped by Millennium Pharmaceuticals and Xenova Ltd. as a cancer therapeutic and is in a phase I clinical trial for solid tumors. Although MLN944 was originally proposed to function as a topoisomerase I and II inhibitor, more recent data has shown that it is a DNA-intercalating agent that does not inhibit the catalytic activity of topoisomerase I or II. We show here that MLN944 inhibits incorporation of radiolabeled precursors into RNA preferentially over incorporation into DNA and protein in HCT116 and H460 cells. To determine if MLN944 inhibits transcription, a human RNA polymerase II in vitro transcription system was used. MLN944 inhibited initiation when added before or after the formation of preinitiation complexes and inhibited elongation at higher concentrations. The preferential inhibition of initiation differentiates MLN944 from actinomycin D, which more strongly inhibits elongation. Transcription of all RNA polymerases was inhibited in nuclei isolated from HeLa cells treated with low concentrations of MLN944. Our data are consistent with transcription as the target of the potent cytotoxic effects of MLN944.