Upfront Xpert MTB/RIF for diagnosis of pediatric TB-Does it work? Experience from India.

Background: Diagnosis of TB in pediatric population poses several challenges. A novel initiative was implemented in several major cities of India aimed at providing upfront access to free-of-cost Xpert MTB/RIF to presumptive pediatric TB cases. This paper aims to describe the experience of implementing this large initiative and assess feasibility of the intervention in high TB burden settings.

[³H]Chiba-1001(methyl-SSR180711) has low in vitro binding affinity and poor in vivo selectivity to nicotinic alpha-7 receptor in rodent brain.

Neuronal nicotinic acetylcholine receptor (nAChR) agonists active at the alpha-7 (α-7) receptor subtype are potential therapeutics for cognitive deficits in schizophrenia, Alzheimer's disease, and other mental disorders. SSR180711, an α-7 selective partial agonist, has been shown to improve preclinical cognition. A novel positron emission tomography (PET) radioligand, ¹¹C-Chiba1001, is a close analog of SSR180711.

Preclinical profile of a novel metabotropic glutamate receptor 5 positive allosteric modulator.

Recent reports have indicated that patients with schizophrenia have a profound hypo-functionality of glutamatergic signaling pathways. Positive allosteric modulation of mGlu(5) receptor has been postulated to augment NMDA function and thereby alleviate the glutamatergic hypo-function observed in schizophrenic patients. Here we report the in vitro and in vivo characterization of CPPZ (1-(4-(2-chloro-4-fluorophenyl)piperazin-1-yl)-2-(pyridin-4-ylmethoxy)ethanone), a structurally novel positive allosteric modulator selective for mGlu(5) receptor.

4-aryl piperazine and piperidine amides as novel mGluR5 positive allosteric modulators.

Positive allosteric modulation of metabotropic glutamate receptor 5 (mGluR5) is regarded as a potential novel treatment for schizophrenic patients. Herein we report the synthesis and SAR of 4-aryl piperazine and piperidine amides as potent mGluR5 positive allosteric modulators (PAMs). Several analogs have excellent activity and desired drug-like properties.

Absence of direct effects on the dopamine D2 receptor by mGluR2/3-selective receptor agonists LY 354,740 and LY 379,268.

We previously reported the absence of high-affinity binding of the group II metabotropic glutamate receptor agonists LY 354,740 and LY 379,268 to the D2L dopamine receptor. A rebuttal to our findings has since been reported (see Introduction section); this study represents our response. Analysis by LCMS of LY 354,740 and LY 379,268 used in this study revealed the correct molecular mass for these compounds.

Acute pharmacological modulation of mGluR8 reduces measures of anxiety.

Metabotropic glutamate receptors (mGluRs), which are coupled to second messenger pathways via G proteins, modulate glutamatergic and GABAergic neurotransmission. Because of their role in modulating neurotransmission, mGluRs are attractive therapeutic targets for anxiety disorders. Previously we showed that mGluR8(-/-) male mice showed higher measures of anxiety in the open field and elevated plus maze than age-matched wild-type mice.

Synthesis and Biological activity of kappa opioid receptor agonists. Part 2: preparation of 3-aryl-2-pyridone analogues generated by solution- and solid-phase parallel synthesis methods.

New analogues of the previously described 3-aryl pyridone KOR agonists have been synthesised by parallel synthetic methods, both in solution- and with solid-phase chemistry, making use of the well known and versatile Mitsunobu, Suzuki and Buchwald reactions. Opioid receptor binding data for the compounds produced is reported.

Tissue distribution and differential expression of melanocortin 1 receptor, a malignant melanoma marker.

The melanocortin 1 receptor is a G-protein-coupled receptor, described to be expressed on melanomas and melanocytes. Subsequent RT-PCR studies demonstrated the presence of melanocortin 1 receptor mRNA in other tissues such as pituitary gland and testis. Previously, we have demonstrated that three HLA-A2 binding nonamer peptides derived from melanocortin 1 receptor can elicit peptide-specific CTL which can recognize target cells transfected with the melanocortin 1 receptor gene and MHC class I matched melanoma lines.

3-Aryl pyridone derivatives. Potent and selective kappa opioid receptor agonists.

A new series of 3-aryl pyridone based kappa opioid receptor agonists was designed and synthesised, based on an understanding of the classical kappa opioid receptor pharmacophore. The most potent of the new compounds were comparable to U-69,593 in receptor affinity, selectivity and functional agonist effect at the cloned human kappa opioid receptor.

Cysteine residues are involved in structure and function of melanocortin 1 receptor: Substitution of a cysteine residue in transmembrane segment two converts an agonist to antagonist.

Reduction of disulfide bonds in human melanocortin 1 receptor (hMC1R) with increasing concentrations of DTT (dithiothreitol) resulted in a decrease in the binding of [125I]-ACTH (adrenocorticotropic hormone, L-isomer) in an uniphasic manner and a decrease in [125I]-NDP-MSH ([Nle(4),D-Phe(7)]-alpha-melanocyte stimulating hormone; D-isomer) binding in a biphasic manner. Pretreatment of hMC1R with 10 mM DTT resulted in a 36-fold loss of affinity for alpha-MSH (L-isomer) without affecting the affinity of NDP-MSH (D-isomer). To characterize the role of individual cysteine residues, we employed site-directed mutagenesis to substitute cysteine by glycine at all fourteen positions in hMC1R and analysed wild-type and mutant receptors for ligand binding and cAMP signalling.

Normal feeding behavior, body weight and leptin response require the neuropeptide Y Y2 receptor.

Neuropeptide Y (NPY), a 36-amino-acid peptide widely expressed in the brain is involved in many physiological responses, including hypothalamic control of food intake and cardiovascular homeostasis. NPY mediates its effects through binding to the Y1, Y2 and Y5 G-protein-coupled receptors. Little is known of the role of the Y2 receptor in mediating the different NPY effects.

The cloned guinea pig neuropeptide Y receptor Y1 conforms to other mammalian Y1 receptors.

We have cloned the guinea pig neuropeptide Y (NPY) Y1 receptor and found it to be 92-93% identical to other cloned mammalian Y1 receptors. Porcine NPY and peptide YY (PYY) displayed affinities of 43 pM and 48 pM, respectively. NPY2-36 and NPY3-36 had 6- and 46-fold lower affinity, respectively, than intact NPY.

alpha-MSH and its receptors in regulation of tumor necrosis factor-alpha production by human monocyte/macrophages.

The hypothesis that macrophages contain an autocrine circuit based on melanocortin [ACTH and alpha-melanocyte-stimulating hormone (alpha-MSH)] peptides has major implications for neuroimmunomodulation research and inflammation therapy. To test this hypothesis, cells of the THP-1 human monocyte/macrophage line were stimulated with lipopolysaccharide (LPS) in the presence and absence of alpha-MSH. The inflammatory cytokine tumor necrosis factor (TNF)-alpha was inhibited in relation to alpha-MSH concentration.

Amino acid residues in third intracellular loop of melanocortin 1 receptor are involved in G-protein coupling.

To delineate domains essential for G-protein coupling in melanocortin 1 receptor (MC1R), we mutated polar and basic residues to alanine at eleven positions in the putative third intracellular loop and determined consequent changes in the ligand binding and generation of second messenger cAMP. Results demonstrate that ligand binding affinity was not affected by any of the mutations. However, every mutant displayed reduced functional response as compared to the wild type receptor.

Direct effects of corticotrophin on oral keratinocyte cell proliferation.

Corticotropin is produced by keratinocytes and may have an immunoregulatory role in oral mucosa and skin. We have investigated its effects on a human oral keratinocyte cell line and shown that corticotropin, acting via its specific receptor, stimulates a dose-dependent increase in DNA synthesis and induces cell proliferation. When cells were incubated in the presence of increasing concentrations of corticotropin, there were significant increases in intracellular cAMP levels.

Human pigmentation phenotype: a point mutation generates nonfunctional MSH receptor.

alpha-Melanocyte stimulating hormone (alpha-MSH) regulates skin and hair pigmentation by modulating the activity of MSH receptor (MC1R). We have identified Arg151Cys variant of human MC1R in genomic DNA isolated from a person with red hair and light skin of type I. The Arg151Cys variant of MC1R binds to radio-labelled analogue of alpha-MSH with identical affinity as wild type MC1R but can not be stimulated to produce cyclic AMP (cAMP).

Synthetic peptides derived from the melanocyte-stimulating hormone receptor MC1R can stimulate HLA-A2-restricted cytotoxic T lymphocytes that recognize naturally processed peptides on human melanoma cells.

Human melanoma-specific HLA-A2 restricted CTLs have recently been shown to recognize antigens expressed by melanoma lines and normal melanocytes, including Melan-A/Mart-1, gp100, gp75, and tyrosinase. Herein, we define HLA-A2-restricted CTL epitopes from a recently cloned melanocortin 1 receptor (MC1R), which belongs to a new subfamily of the G-protein-coupled receptors expressed on melanomas and melanocytes. Thirty-one MC1R-derived peptides were selected on the basis of HLA-A2-specific motifs and tested for their HLA-A2 binding capacity.

Glutamine235 and arginine272 in human melanocortin 5 receptor determines its low affinity to MSH.

The human melanocortin 5 receptor (hMC5R) in the melanocortin receptor family has been identified as the receptor with low affinity towards alpha-MSH. Here we show that the glutamine at position 235 and arginine at the position 272 in the hMC5R are contributing to the low affinity of this receptor. Glutamine235 and arginine272 in hMC5R were mutated to lysine (Q235K) and cysteine (R272C), respectively, residues which are conserved at these positions in other melanocortin receptor subtypes.

Immunohistochemical detection of the melanocortin 1 receptor in human testis, ovary and placenta using specific monoclonal antibody.

We describe the immunohistochemical detection of the melanocortin 1 receptor (MC1R) protein in human gonadal tissues using a specific monoclonal antibody. The MC1R was found to be present in Leydig's cells in testis, in lutein cells in the corpus luteum and in the nucleus of the trophoblastic cells of the placenta. Though it has been speculated earlier that MC1R is present in gonadal tissues, this is the first report demonstrating the presence of MC1R protein in these cells.

Identification of ligand binding residues in extracellular loops of the melanocortin 1 receptor.

To investigate whether residues in the extracellular domains of melanocortin 1 receptor (MC1R) are required for ligand binding, a number of mutants were constructed where charged residues were converted to alanine. The residues targeted for mutagenesis were Ser6, Glu102, Arg109, Asp184, Glu269, and Thr272. The mutant receptor DNAs were transiently expressed in COS-1 cells and their ability to bind [N1e4,D-Phe7]-alpha-MSH (NDP-MSH) was evaluated.

Distribution of cDNA for melanocortin receptor subtypes in human tissues.

Distribution of cDNA for five individual melanocortin receptor subtypes in 20 different human tissues was determined by PCR using subtype specific primers. PCR products were first visualised by agarose gel electrophoresis and ethidium bromide staining, and specific products were identified for melanocortin 1 receptor (MC1R) in pituitary and testis, for MC2R in adrenal gland, for MC3R in heart, for MC5R in adrenal gland, fat cells, kidney, leukocytes, lung, lymph node, mammary gland, ovary, pituitary, testis and uterus. The MC4R cDNA could not be detected by ethidium bromide staining.