GLP-2 receptors in human disease: high expression in gastrointestinal stromal tumors and Crohn's disease

Peptide hormones of the glucagon-like peptide (GLP) family play an increasing clinical role, as reported for GLP-1 in diabetes therapy and insulinoma diagnostics. GLP-2, despite its known trophic and anti-inflammatory intestinal actions translated into preliminary clinical studies using the GLP-2 analogue teduglutide for treatment of short bowel syndrome and Crohn's disease, remains poorly characterized in terms of expression of its receptor in tissues of interest. Therefore, the GLP-2 receptor expression was assessed in 237 tumor and 148 non-neoplastic tissue samples with in vitro receptor autoradiography. A GLP-2 receptor expression was present in 68% of gastrointestinal stromal tumors (GIST). Furthermore, GLP-2 receptors were identified in the intestinal myenteric plexus, with significant up-regulation in active Crohn's disease. The GLP-2 receptors in GIST may be used for clinical applications like in vivo targeting with radiolabelled GLP-2 analogues for imaging and therapy. Moreover, the over-expressed GLP-2 receptor in the myenteric plexus may represent the morphological correlate of the clinical target of teduglutide in Crohn's disease.

Modulation of L-α-lysophosphatidylinositol/GPR55 mitogen-activated protein kinase (MAPK) signaling by cannabinoids

GPR55 is activated by l-α-lysophosphatidylinositol (LPI) but also by certain cannabinoids. In this study, we investigated the GPR55 pharmacology of various cannabinoids, including analogues of the CB1 receptor antagonist Rimonabant®, CB2 receptor agonists, and Cannabis sativa constituents. To test ERK1/2 phosphorylation, a primary downstream signaling pathway that conveys LPI-induced activation of GPR55, a high throughput system, was established using the AlphaScreen® SureFire® assay. Here, we show that CB1 receptor antagonists can act both as agonists alone and as inhibitors of LPI signaling under the same assay conditions. This study clarifies the controversy surrounding the GPR55-mediated actions of SR141716A; some reports indicate the compound to be an agonist and some report antagonism. In contrast, we report that the CB2 ligand GW405833 behaves as a partial agonist of GPR55 alone and enhances LPI signaling. GPR55 has been implicated in pain transmission, and thus our results suggest that this receptor may be responsible for some of the antinociceptive actions of certain CB2 receptor ligands. The phytocannabinoids Δ9-tetrahydrocannabivarin, cannabidivarin, and cannabigerovarin are also potent inhibitors of LPI. These Cannabis sativa constituents may represent novel therapeutics targeting GPR55.

[Bone metastasis in prostate cancer]

Bone metastasis and skeletal complications have a devastating impact on the quality of life and are a major cause of morbidity in prostate cancer patients. In addition to established bone-targeted therapies, new drugs such as endothelin A receptor antagonists, MET and VEGFR-2 antagonists or radiopharmaceuticals are in the focus of development. The standard care in prostate cancer patients with bone metastases to prevent skeletal-related events (SRE) are bisphosphonates. Denosumab, a human monoclonal antibody against RANKL, appeared to be superior to zoledronic acid for prevention of SRE and has been shown to prolong bone metastases-free survival. In contrast to zoledronic acid, denosumab clearance is not dependent on kidney function and can be administered subcutaneously. Similar rates of toxicity were observed for both substances; however, long-term data for denosumab are limited.

Arterial hypertension and proteinuria in pediatric chronic kidney disease

A variety of chronic kidney diseases tend to progress towards end-stage kidney disease. Progression is largely due to factors unrelated to the initial disease, including arterial hypertension and proteinuria. Intensive treatment of these two factors is potentially able to slow the progression of kidney disease. Blockers of the renin-angiotensin-aldosterone system, either converting enzyme inhibitors or angiotensin II receptor antagonists, reduce both blood pressure and proteinuria and appear superior to a conventional antihypertensive treatment regimen in preventing progression to end-stage kidney disease. The most recent recommendations state that in children with chronic kidney disease without proteinuria the blood pressure goal is the corresponding 75th centile for body length, age and gender; whereas the 50th centile should be aimed in children with chronic kidney disease and pathologically increased proteinuria.

Agonist specific L-type Ca(2+)-current stimulation in ventricular myocytes by a novel steroid-like compound

In cardiac muscle the amplitude of Ca(2+) transients can be increased by enhancing Ca(2+) influx. Among the processes leading to increased Ca(2+) influx, agonists of the L-type Ca(2+)-channel can play an important role. Known pharmacological Ca(2+)-channel agonists act on different binding sites on the channel protein, which may lead not only to enhanced peak currents, but also to distinct changes in other biophysical characteristics of the current. In this study, membrane currents were recorded with the patch-clamp technique in the whole-cell configuration in guinea pig isolated ventricular myocytes in combination with confocal fluorescence Ca(2+) imaging techniques and a variety of pharmacological tools. Testing a new positive inotropic steroid-like compound, we found that it increased the L-type Ca(2+)-current by 2.5-fold by shifting the voltage-dependence of activation by 20.2 mV towards negative potentials. The dose-response relationship revealed two vastly different affinities (EC(50(high-affinity))=4.5+/-1.7 nM, EC(50(low-affinity))=8.0+/-1.1 microM) exhibiting differential pharmacological interactions with three classes of Ca(2+)-current antagonists, suggesting more than one binding site on the channel protein. Therefore, we identified and characterized a novel positive inotropic compound (F90927) as a member of a new class of Ca(2+)-channel agonists exhibiting unique features, which set it apart from other presently known L-type Ca(2+)-channel agonists.

Grover's disease induced by cetuximab

A 71-year-old man exhibited an acute acneiform rash affecting the face and the upper trunk about 2 weeks after starting cetuximab, an epidermal growth factor (EGF) receptor antagonist treatment for metastatic colon cancer. The skin eruption faded after stopping cetuximab and applying topical corticosteroids. The reexposure to cetuximab 3 weeks later provoked a more extended relapse of the skin rash, which then clinically and histologically corresponded to transient acantholytic dermatosis . While the acneiform cutaneous side effects of the EGF receptor antagonists are interpreted as a result of the direct interference with pilosebaceous follicle homeostasis, in this case an acrosyringium-related pathogenesis might be postulated. Applying topical corticosteroids and emollients, the cetuximab therapy could be pursued.

Snake venom C-type lectins interacting with platelet receptors. Structure-function relationships and effects on haemostasis

Snake venoms contain components that affect the prey either by neurotoxic or haemorrhagic effects. The latter category affect haemostasis either by inhibiting or activating platelets or coagulation factors. They fall into several types based upon structure and mode of action. A major class is the snake C-type lectins or C-type lectin-like family which shows a typical folding like that in classic C-type lectins such as the selectins and mannose-binding proteins. Those in snake venoms are mostly based on a heterodimeric structure with two subunits alpha and beta, which are often oligomerized to form larger molecules. Simple heterodimeric members of this family have been shown to inhibit platelet functions by binding to GPIb but others activate platelets via the same receptor. Some that act via GPIb do so by inducing von Willebrand factor to bind to it. Another series of snake C-type lectins activate platelets by binding to GPVI while yet another series uses the integrin alpha(2)beta(1) to affect platelet function. The structure of more and more of these C-type lectins have now been, and are being, determined, often together with their ligands, casting light on binding sites and mechanisms. In addition, it is relatively easy to model the structure of the C-type lectins if the primary structure is known. These studies have shown that these proteins are quite a complex group, often with more than one platelet receptor as ligand and although superficially some appear to act as inhibitors, in fact most function by inducing thrombocytopenia by various routes. The relationship between structure and function in this group of venom proteins will be discussed.

Variation in human platelet glycoprotein VI content modulates glycoprotein VI-specific prothrombinase activity

- Glycoprotein VI (GPVI) is a platelet-specific receptor for collagen that figures prominently in signal transduction. An addition to binding to type I and III collagens, GPVI is also bound specifically by collagen-related peptide and convulxin (CVX), a snake venom protein. We developed a quantitative assay of platelet GPVI in which biotin-conjugated CVX binds selectively to GPVI in separated total platelet proteins by a ligand blot procedure. Using this approach, we have documented a 5-fold range in platelet GPVI content among 23 normal healthy subjects. In addition, we have determined that CVX-induced or collagen-related peptide-induced prothrombinase activity is directly proportional to the platelet content of GPVI. A statistically significant correlation was observed at 2 CVX concentrations: 14.7 ng/mL (R(2)=0.854 and P<0.001, n=11) and 22 ng/mL (R(2)=0.776 and P<0.001, n=12). In previous studies, we established a similar range of expression of the integrin collagen receptor alpha(2)beta(1) on platelets of normal subjects. Among 15 donors, there is a direct correlation between platelet alpha(2)beta(1) density and GPVI content (R(2)=0.475 and P=0.004). In view of the well-documented association of GPVI with platelet procoagulant activity, this study suggests that the variation in GPVI content is a potential risk factor that may predispose individuals to hemorrhagic or thromboembolic disorders.

Effects of antihypertensive drugs on blood pressure and proteinuria in childhood

BACKGROUND: Historically, there have been few drug trials for antihypertensive treatment in childhood and recommendations have been extrapolated from data obtained in adulthood. During the last decade an increased awareness of the risks of childhood hypertension stimulated clinical trials of antihypertensive agents in children. OBJECTIVE: The aim of this article is to systematically review the studies published between 1995 and 2006 that deal with the effect of antihypertensive drugs on childhood hypertension or proteinuria. METHODS: Medline, Current Contents, personal files and reference lists were used as data sources. RESULTS: Fifty-two out of 79 initially found reports were excluded. Consequently 27 articles were retained for the final analysis. The blood pressure reduction was similar with angiotensin-converting enzyme inhibitors (10.7/8.1 mmHg), angiotensin II receptor antagonists (10.5/6.9 mmHg) and calcium-channel blockers (9.3/7.2 mmHg). In addition angiotensin-converting enzyme inhibitors (by 49%) and angiotensin II receptor antagonists (by 59%) significantly reduced pathological proteinuria. CONCLUSIONS: The blood pressure reduction of angiotensin-converting enzyme inhibitors, angiotensin II receptor antagonists and calcium-channel blockers is almost identical. In children with pathological proteinuria angiotensin-converting enzyme inhibitors or angiotensin II antagonists are superior to calcium-channel blockers.

[Perspectives in the treatment of subarachnoid-hemorrhage-induced cerebral vasospasm]

Cerebral vasospasm is still the most important cause of death and disability after rupture of intracranial aneurysms. The therapeutic strategies in the treatment of subarachnoid hemorrhage induced vasospasm vasospasm include four groups: 1) prevention of vasospasm; 2) reversion of vasospasm; 3) improvement of cerebral perfusion; and 4) neuroprotection and rescue therapies. Recent experimental studies allowed the design of phase II clinical studies which demonstrated positive results with medications and compounds such as statins (simvastatin and pravastatin) and endothelin-1 receptor antagonists (clasozentan). Moreover, experimental and clinical evidences showed the advantages of early cerebrospinal fluid drainage, intrathecal administration of NO-donors, effects of Ca2+ protein kinase inhibitor (Fasudil) and catecholamines on the cerebral vessels. This review article summarizes the stage of investigation of these medications and therapeutic strategies which will be relevant in the treatment of cerebral vasospasm.

Expression and hypoxic regulation of the endothelin system in endocrine cells of human and rat pancreatic islets

CONTEXT: The success of pancreatic islet transplantation depends largely on the capacity of the islet graft to survive the initial phase immediately after transplantation until revascularization is completed. Endothelin-1 (ET-1) is a strong vasoconstrictor which has been involved in solid organ graft failure but is also known to be a potent mitogenic/anti-apoptotic factor which could also potentially enhance the survival of the transplanted islets. OBJECTIVE: Characterization of the endothelin system with regard to a potential endothelin agonist/antagonist treatment. DESIGN: Regulated expression of the endothelin system in human and rat pancreatic islets and beta-cell lines was assessed by means of immunohistochemistry, competition binding studies, western blot, RT-PCR, real-time PCR and transplant studies. RESULTS: ET-1, ETA- and ETB-receptor immunoreactivity was identified in the endocrine cells of human and rat pancreatic islets. The corresponding mRNA was detectable in rat beta-cell lines and isolated rat and human pancreatic islets. Competition binding studies on rat islets revealed binding sites for both receptor types. ET-1 stimulated the phosphorylation of mitogen-activated protein kinase, which was prevented by ETA- and ETB-receptor antagonists. After exposure to hypoxia equal to post-transplant environment oxygen tension, mRNA levels of ET-1 and ETB-receptor of human islets were robustly induced whereas ETA-receptor mRNA did not show significant changes. Immunostaining signals for ET-1 and ETA-receptor of transplanted rat islets were markedly decreased when compared to native pancreatic sections. CONCLUSIONS: In pancreatic islets, ET-1 and its receptors are differentially expressed by hypoxia and after transplantation. Our results provide the biological basis for the study of the potential use of endothelin agonists/antagonists to improve islet transplantation outcome.

Cardiac toxicity with anti-HER-2 therapies: what have we learned so far?

Trastuzumab, a monoclonal antibody that blocks HER-2 receptor, improves the survival of women with HER-2-positive early and advanced breast cancer when given with chemotherapy. Lapatinib, a dual tyrosine kinase inhibitor of EGFR and HER-2, is approved for the treatment of metastatic breast cancer patients after failure of prior anthracycline, taxanes and trastuzumab therapies in combination with capecitabine. Importantly, cardiac toxicity, manifested as symptomatic congestive heart failure or asymptomatic left ventricular ejection fraction decline, has been reported in some of the patients receiving these novel anti-HER-2 therapies, particularly when these drugs are used following anthracyclines, whose cardiotoxic potential has been recognized for decades. This review will focus on the incidence, natural history, underlying mechanisms, management, and areas of uncertainty regarding trastuzumab-and lapatinib-induced cardiotoxicity.

Cannabinoid receptor ligands as potential anticancer agents--high hopes for new therapies?

OBJECTIVES: The endocannabinoid system is an endogenous lipid signalling network comprising arachidonic-acid-derived ligands, cannabinoid (CB) receptors, transporters and endocannabinoid degrading enzymes. The CB(1) receptor is predominantly expressed in neurons but is also co-expressed with the CB(2) receptor in peripheral tissues. In recent years, CB receptor ligands, including Delta(9)-tetrahydrocannabinol, have been proposed as potential anticancer agents. KEY FINDINGS: This review critically discusses the pharmacology of CB receptor activation as a novel therapeutic anticancer strategy in terms of ligand selectivity, tissue specificity and potency. Intriguingly, antitumour effects mediated by cannabinoids are not confined to inhibition of cancer cell proliferation; cannabinoids also reduce angiogenesis, cell migration and metastasis, inhibit carcinogenesis and attenuate inflammatory processes. In the last decade several new selective CB(1) and CB(2) receptor agents have been described, but most studies in the area of cancer research have used non-selective CB ligands. Moreover, many of these ligands exert prominent CB receptor-independent pharmacological effects, such as activation of the G-protein-coupled receptor GPR55, peroxisome proliferator-activated receptor gamma and the transient receptor potential vanilloid channels. SUMMARY: The role of the endocannabinoid system in tumourigenesis is still poorly understood and the molecular mechanisms of cannabinoid anticancer action need to be elucidated. The development of CB(2)-selective anticancer agents could be advantageous in light of the unwanted central effects exerted by CB(1) receptor ligands. Probably the most interesting question is whether cannabinoids could be useful in chemoprevention or in combination with established chemotherapeutic agents.

IL-2 REGULATION OF IL-24 EXPRESSION LEADS TO GROWTH SUPPRESSION OF MELANOMA CELLS

Melanoma is known to be highly resistant to chemotherapy. Treatment with high dose IL-2 has shown significant clinical benefit in a minority of metastatic melanoma patients and has lead to long term survival in a few cases. However, this treatment is associated with excessive multiorgan toxicities, which severely limits its use. We hypothesize that one mechanism of effective IL-2 therapy is through the direct upregulation of IL-24 production in melanoma tumors and subsequent IL-24 mediated tumor growth suppression. Five melanoma cell lines were treated with high dose recombinant hIL-2 at 1000U/ml. Three of the cell lines (A375, WM1341, WM793) showed statistically significant increases in their levels of IL-24 protein when measured by Western blotting, while the remaining two lines (WM35, MeWo) remained negative for IL-24 message and protein. This increase in IL-24 was abolished by either preincubating with an anti-IL-2 antibody or by blocking the IL-2 receptor directly with antibodies against the receptor chains. We also demonstrated by ELISA that these three cell lines secrete IL-24 protein in higher amounts when stimulated with IL-2 than do untreated cells. These cells were found to contain IL-2R beta and gamma message by RT-PCR and also expressed higher levels of IL-24 when treated with IL-15, which shares the IL-2R beta chain. Thus we propose that IL-2 is signaling through IL-2R beta on some melanoma cells to upregulate IL-24 protein expression. To address the biological function of IL-2 in melanoma cells, five cell lines were treated with IL-2 and cell viability determined. Cell growth was found to be significantly decreased by day 4 in the IL-24 positive cell lines while no effect on growth was seen in WM35 or MeWo. Incubating the cells with anti-IL-24 antibody or transfecting with IL-24 siRNA effectively negated the growth suppression seen with IL-2. These data support our hypothesis that in addition to its immunotherapeutic effects, IL-2 also acts directly on some melanoma tumors and that the IL-24 and IL-2R beta status of a tumor may be useful in predicting patient response to high dose IL-2.

Molecular signals in B lymphocyte activation: The role of intracellular calcium ion concentration, protein kinase C activation, and autocrine interleukin-2

Calcium ionophore, ionomycin, and phorbol myristate acetate (PMA) were used to activate rabbit peripheral blood B cells to study the role of increased intracellular calcium ion concentration ( (Ca$\sp2+\rbrack\sb{\rm i}$), protein kinase C (PKC) activation, and autocrine interleukin (IL-2) in inducing cell cycle entry and maintaining activation to DNA synthesis. When stimulated with a combination of ionomycin and PMA the B cells produced a soluble factor that supported the IL-2 dependent cell line, CTLL-2. The identity of the factor was established as IL-2 and its source was proved to be B cells in further experiments. Absorption studies and limiting dilution analysis indicated that IL-2 produced by B cells can act as an autocrine growth factor. Next, the effect of complete and incomplete signalling on B lymphocyte activation leading to cell cycle entry, IL-2 production, functional IL-2 receptor (IL-2R) expression, and DNA synthesis was examined. It was observed that cell cycle entry could be induced by signals provided by each reagent alone, but IL-2 production, IL-2R expression, and progression to DNA synthesis required activation with both reagents. Incomplete activation with ionomycin or PMA alone altered the responsiveness of B cells to further stimulation only in the case of ionomycin, and the unresponsiveness of these cells was apparently due to a lack of functional IL-2R expression on these cells, even though IL-2 production was maintained. The requirement of IL-2 for maintenance of activation to DNA synthesis was then investigated. The hypothesis that IL-2, acts in late G$\sb1$ and is required for DNA synthesis in B cells was supported by comparing IL-2 production and DNA synthesis in peripheral blood cells and purified B cells, kinetic analysis of these events in B cells, effects of anti-IL-2 antibody and PKC inhibitors, and by the response of G$\sb1$ B cells. Additional signals transduced by the interaction of autocrine IL-2 and functional IL-2 receptor on rabbit B cells were found to be necessary to drive these cells to S phase, after initial activation caused by simultaneous increase in (Ca$\sp2+\rbrack\sb{\rm i}$ and PKC activation had induced cell cycle entry, IL-2 production, and functional IL-2 receptor expression. ^