Almost one fourth of patients with apparently sporadic pheochromocytoma may be carriers of mutations; routine analysis for mutations of RET, VHL, SDHD, and SDHB is indicated to identify pheochromocytoma-associated syndromes that would otherwise be missed.

No abstract

The gamma isoform of the peroxisome proliferator-activated receptor, PPAR gamma, regulates adipocyte differentiation and has recently been shown to be expressed in neoplasia of the colon and other tissues. We have found four somatic PPAR gamma mutations among 55 sporadic colon cancers: one nonsense, one frameshift, and two missense mutations. Each greatly impaired the function of the protein. c.472delA results in deletion of the entire ligand binding domain. Q286P and K319X retain a total or partial ligand binding domain but lose the ability to activate transcription through a failure to bind to ligands. R288H showed a normal response to synthetic ligands but greatly decreased transcription and binding when exposed to natural ligands. These data indicate that colon cancer in humans is associated with loss-of-function mutations in PPAR gamma.

Activation of PPAR␥ by synthetic ligands, such as thiazolidinediones, stimulates adipogenesis and improves insulin sensitivity. Although thiazolidinediones represent a major therapy for type 2 diabetes, conflicting studies showing that these agents can increase or decrease colonic tumors in mice have raised concerns about the role of PPAR␥ in colon cancer. To analyze critically the role of this receptor, we have used mice heterozygous for Ppar␥ with both chemical and genetic models of this malignancy. Heterozygous loss of PPAR␥ causes an increase in ␤-catenin levels and a greater incidence of colon cancer when animals are treated with azoxymethane. However, mice with preexisting damage to Apc, a regulator of ␤-catenin, develop tumors in a manner insensitive to the status of PPAR␥. These data show that PPAR␥ can suppress ␤-catenin levels and colon carcinogenesis but only before damage to the APC͞␤-catenin pathway. This finding suggests a potentially important use for PPAR␥ ligands as chemopreventative agents in colon cancer.

Few surveillance systems have been specifically designed for collecting and analyzing data for the early detection of a bioterrorist event. Because current evaluations of surveillance systems for detecting bioterrorism and emerging infections are insufficient to characterize the timeliness or sensitivity and specificity, clinical and public health decision making based on these systems may be compromised.

Purpose To develop diagnostic criteria for nonparaneoplastic autoimmune retinopathy (AIR) through expert panel consensus and to examine treatment patterns among clinical experts. Design Modified Delphi process. Methods A survey of uveitis specialists in the American Uveitis Society (AUS), a face-to-face meeting (AIR Workshop) held at the National Eye Institute (NEI), and two iterations of expert panel surveys were utilized in a modified Delphi process. The expert panel consisted of 17 experts including uveitis specialists and researchers with expertise in antiretinal antibody detection. Supermajority consensus was used and defined as 75% of experts in agreement. Results There was unanimous agreement among experts regarding the categorization of autoimmune retinopathies as nonparaneoplastic and paraneoplastic, including cancer-associated retinopathy (CAR) and melanoma-associated retinopathy (MAR). Diagnostic criteria and tests essential to the diagnosis of nonparaneoplastic AIR and multiple supportive criteria reached consensus. For treatment, experts agreed that corticosteroids and conventional immunosuppressives should be used (prescribed) as 1st or 2nd line treatments, though a consensus agreed that biologics and intravenous immunoglobulin were considered appropriate in the treatment of nonparaneoplastic AIR patients regardless of the stage of disease. Experts agreed that more evidence is needed to treat nonparaneoplastic AIR patients with long-term immunomodulatory therapy and that there is enough equipoise to justify randomized, placebo-controlled trials to determine if nonparaneoplastic AIR patients should be treated with long-term immunomodulatory therapy. Regarding antiretinal antibody detection, consensus agreed that a standardized assay system is needed to detect serum antiretinal antibodies. Consensus agreed that an ideal assay should have a two-tier design and that western blot (WB) and immunohistochemistry (IHC) should be the methods used to identify antiretinal antibodies. Conclusions Consensus was achieved using a modified Delphi process to develop diagnostic criteria for nonparaneoplastic AIR. There is enough equipoise to justify randomized, placebo-controlled trials to determine whether patients with nonparaneoplastic AIR should be treated with long-term immunomodulatory therapy. Efforts to develop a standardized two-tier assay system for the detection of antiretinal antibodies have been initiated as a result of this study.

The tumour suppressor gene PTEN encodes a dual-specificity phosphatase that recognizes protein substrates and phosphatidylinositol-3,4,5-triphosphate. PTEN seems to play multiple roles in tumour suppression and the blockade of phosphoinositide-3-kinase signalling is important for its growth suppressive effects, although precise mechanisms are not fully understood. In this study, we show that PTEN plays a unique role in the insulin-signalling pathway in a breast cancer model. Ectopic expression of wild-type PTEN in MCF-7 epithelial breast cancer cells resulted in universal inhibition of Akt phosphorylation in response to stimulation by diverse growth factors and selective inhibition of MEK/extracellular signal-regulated kinase (ERK) phosphorylation stimulated by insulin or insulin-like growth factor 1 (IGF-1). The latter was accompanied by a decrease in the phosphorylation of insulin receptor substrate 1 (IRS-1) and the association of IRS-1 with Grb2/Sos, without affecting the phosphorylation status of the insulin receptor and Shc, nor Shc/Grb2 complex formation. The MEK inhibitor, PD980059, but not the PI3K inhibitor, wortmannin, abolished the effect of PTEN on insulin-stimulated cell growth. Without addition of insulin, wortmannin reduced PTEN-mediated growth suppression, whereas PD980059 had little effect, suggesting that PTEN suppresses insulin-stimulated cell growth by blocking the mitogen-activated protein kinase (MAPK) pathway. Furthermore, PD980059 treatment led to the downregulation of cyclin D1 and the suppression of cell cycle progression. Our data suggest that PTEN blocks MAPK phosphorylation in response to insulin stimulation by inhibiting the phosphorylation of IRS-1 and IRS-1/Grb2/Sos complex formation, which leads to downregulation of cyclin D1, inhibition of cell cycle progression and suppression of cell growth.

A novel pathway for degradation of the cyanobacterial heptapeptide hepatotoxin microcystin LR was identified in a newly isolated Sphingomonas sp. (Bourne et al. 1996 Appl. Environ. Microbiol. 62: 4086-4094). We now report the cloning and molecular characterisation of four genes from this Sphingomonas sp. that exist on a 5.8-kb genomic fragment and encode the three hydrolytic enzymes involved in this pathway together with a putative oligopeptide transporter. The heterologously expressed degradation pathway proteins are enzymatically active. Microcystinase (MlrA), the first enzyme in the degradative pathway, is a 336-residue endopeptidase, which displays only low sequence identity with a hypothetical protein from Methanobacterium thermoautotrophicum. Inhibition of microcystinase by EDTA and 1,10-phenanthroline suggests that it is a metalloenzyme. The most likely residues that could potentially chelate an active-site transition metal ion are in the sequence HXXHXE, which would be unique for a metalloproteinase. Situated immediately downstream of mlrA with the same direction of transcription is a gene mlrD, whose conceptual translation (MlrD, 442 residues) shows significant sequence identity and similar potential transmembrane spanning regions to the PTR2 family of oligopeptide transporters. A gene mlrB is situated downstream of the mlrA and mlrD genes, but transcribed in the opposite direction. The gene encodes the enzyme MlrB (402 residues) which cleaves linear microcystin LR to a tetrapeptide degradation product. This enzyme belongs to the "penicillin-binding enzyme" family of active site serine hydrolases. The final gene in the cluster mlrC, is located upstream of the mlrA gene and is transcribed in the opposite direction. It codes for MlrC (507 residues) which mediates further peptidolytic degradation of the tetrapeptide. This protein shows significant sequence identity to a hypothetical protein from Streptomyces coelicolor. It is suspected to be a metallopeptidase based on inhibition by metal chelators. It is postulated on the basis of comparison with other microorganisms that the genes in this cluster may all be involved in cell wall peptidoglycan cycling and subsequently act fortuitously in hydrolysis of microcystin LR.