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S Narula

Publications and source records attributed to S Narula.

At least 19 recordsLinked to original sources
PubMed2005

Protein phosphorylation and signal transduction modulation: chemistry perspectives for small-molecule drug discovery.

Protein phosphorylation has been exploited by Nature in profound ways to control various aspects of cell proliferation, differentiation, metabolism, survival, motility and gene transcription. Cellular signal transduction pathways involve protein kinases, protein phosphatases, and phosphoprotein-interacting domain (e.g., SH2, PTB, WW, FHA, 14-3-3) containing cellular proteins to provide multidimensional, dynamic and reversible regulation of many biological activities. Knowledge of cellular signal transduction pathways has led to the identification of promising therapeutic targets amongst these superfamilies of enzymes and adapter proteins which have been linked to various cancers as well as inflammatory, immune, metabolic and bone diseases. This review focuses on protein kinase, protein phosphatase and phosphoprotein-interacting cellular protein therapeutic targets with an emphasis on small-molecule drug discovery from a chemistry perspective. Noteworthy studies related to molecular genetics, signal transduction pathways, structural biology, and drug design for several of these therapeutic targets are highlighted. Some exemplary proof-of-concept lead compounds, clinical candidates and/or breakthrough medicines are further detailed to illustrate achievements as well as challenges in the generation, optimization and development of small-molecule inhibitors of protein kinases, protein phosphatases or phosphoprotein-interacting domain containing cellular proteins.

Drug Design↗
PubMed2003

Deciduous dentition and enamel defects.

Two hundred eighty children including wellnourished, malnourished and infants with intrauterine growth retardation (IUGR) were examined for dental eruption and enamel hypoplasia. In malnourished and IUGR children eruption of teeth was delayed. The prevalence of enamel hypoplsia in wellnourished children was 20% being significantly higher in females as compared to males in age group 1-2 years. Enamel hypoplasia was seen in 36.6% malnourished subjects. Breast-feeding was protective against enamel hypoplasia.

Dental Enamel Hypoplasia↗
PubMed2001

Ubiquitous transgenic expression of the IL-23 subunit p19 induces multiorgan inflammation, runting, infertility, and premature death.

p19, a molecule structurally related to IL-6, G-CSF, and the p35 subunit of IL-12, is a subunit of the recently discovered cytokine IL-23. Here we show that expression of p19 in multiple tissues of transgenic mice induced a striking phenotype characterized by runting, systemic inflammation, infertility, and death before 3 mo of age. Founder animals had infiltrates of lymphocytes and macrophages in skin, lung, liver, pancreas, and the digestive tract and were anemic. The serum concentrations of the proinflammatory cytokines TNF-alpha and IL-1 were elevated, and the number of circulating neutrophils was increased. In addition, ubiquitous expression of p19 resulted in constitutive expression of acute phase proteins in the liver. Surprisingly, liver-specific expression of p19 failed to reproduce any of these abnormalities, suggesting specific requirements for production of biologically active p19. Bone marrow transfer experiments showed that expression of p19 by hemopoietic cells alone recapitulated the phenotype induced by its widespread expression, pointing to hemopoietic cells as the source of biologically active p19. These findings indicate that p19 shares biological properties with IL-6, IL-12, and G-CSF and that cell-specific expression is required for its biological activity.

Acute-Phase Proteins↗
PubMed2001

Altered T-cell receptor + CD28-mediated signaling and blocked cell cycle progression in interleukin 10 and transforming growth factor-beta-treated alloreactive T cells that do not induce graft-versus-host disease.

The induction of anergy in T cells, although widely accepted as critical for the maintenance of tolerance, is still poorly understood at the molecular level. Recent evidence demonstrates that in addition to blockade of costimulation using monoclonal antibodies (mAbs) directed against cell surface determinants, treatment of mixed lymphocyte reaction (MLR) cultures with interleukin 10 (IL-10) and transforming growth factor-beta (TGF-beta) results in induction of tolerance, rendering alloreactive murine CD4(+) T cells incapable of inducing graft-versus-host disease (GVHD) after in vivo transfer to histoincompatible recipients. The present study, using these cells prior to adoptive transfer, determined that IL-10 + TGF-beta-tolerant CD4(+) T cells exhibit an altered pattern of T-cell receptor (TCR) + CD28-mediated signaling and are incapable of progressing out of the G(1) phase of the cell cycle during stimulation with HLA class II disparate antigen-presenting cells. TGFbeta + IL-10-tolerant cells were incapable of phosphorylating TCR-zeta, or activating ZAP-70, Ras, and MAPK, similarly to T-cell tolerized by blockade of B7/CD28 and CD40/CD40L pathways. Moreover, these cells were incapable of clonal expansion due to defective synthesis of cyclin D3 and cyclin A, and defective activation of cyclin-dependent kinase (cdk)4, cdk6, and cdk2. These cells also exhibited defective down-regulation of p27(kip1) cdk inhibitor and lack of cyclin D2-cdk4 activation, Rb hyperphosphorylation, and progression to the S phase of the cell cycle. These data link anergy-specific proximal biochemical alterations and the downstream nuclear pathways that control T-cell expansion and provide a biochemical profile of IL-10 + TGF-beta-tolerant alloreactive T cells that do not induce GVHD when transferred into MHC class II disparate recipients in vivo.

Adaptor Proteins, Signal Transducing↗
PubMed2001

Type 1 T regulatory cells.

Suppression by T regulatory (Tr) cells is essential for induction of tolerance. Many types of Tr cells have been described in a number of systems, and their biology has been the subject of intensive investigation. Although many aspects of the mechanisms by which these cells exert their effects remain to be elucidated, it is well established that Tr cells suppress immune responses via cell-to-cell interactions and/or the production of interleukin (IL)-10 and transforming growth factor (TGF)-beta. Type-1 T regulatory (Tr1) cells are defined by their ability to produce high levels of IL-10 and TGF-beta. Tr1 cells specific for a variety of antigens arise in vivo, but may also differentiate from naive CD4+ T cells in the presence of IL-10 in vitro. Tr1 cells have a low proliferative capacity, which can be overcome by IL-15. Tr1 cells suppress naive and memory T helper type 1 or 2 responses via production of IL-10 and TGF-beta. Further characterisation of Tr1 cells at the molecular level will define their mechanisms of action and clarify their relationship with other subsets of Tr cells. The use of Tr1 cells to identify novel targets for the development of new therapeutic agents, and as a cellular therapy to modulate peripheral tolerance, can be foreseen.

Animals↗
PubMed2001

Expression and isolation of antimicrobial small molecules from soil DNA libraries.

Natural products have been a critically important source of clinically relevant small molecule therapeutics. However, the discovery rate of novel structural classes of antimicrobial molecules has declined. Recently, increasing evidence has shown that the number of species cultivated from soil represents less than 1% of the total population, opening up the exciting possibility that these uncultured species may provide a large untapped pool from which novel natural products can be discovered. We have constructed and expressed in E. coli a BAC (bacterial artificial chromosome) library containing genomic fragments of DNA (5-120kb) isolated directly from soil organisms (S-DNA). Screening of the library resulted in the identification of several antimicrobial activities expressed by different recombinant clones. One clone (mg1.1) has been partially characterized and found to express several small molecules related to and including indirubin. These results show that genes involved in natural product synthesis can be cloned directly from S-DNA and expressed in a heterologous host, supporting the idea that this technology has the potential to provide novel natural products from the wealth of environmental microbial diversity and is a potentially important new tool for drug discovery.

Amino Acid Sequence↗
PubMed2000

Structure-based design of an osteoclast-selective, nonpeptide src homology 2 inhibitor with in vivo antiresorptive activity.

Targeted disruption of the pp60(src) (Src) gene has implicated this tyrosine kinase in osteoclast-mediated bone resorption and as a therapeutic target for the treatment of osteoporosis and other bone-related diseases. Herein we describe the discovery of a nonpeptide inhibitor (AP22408) of Src that demonstrates in vivo antiresorptive activity. Based on a cocrystal structure of the noncatalytic Src homology 2 (SH2) domain of Src complexed with citrate [in the phosphotyrosine (pTyr) binding pocket], we designed 3',4'-diphosphonophenylalanine (Dpp) as a pTyr mimic. In addition to its design to bind Src SH2, the Dpp moiety exhibits bone-targeting properties that confer osteoclast selectivity, hence minimizing possible undesired effects on other cells that have Src-dependent activities. The chemical structure AP22408 also illustrates a bicyclic template to replace the post-pTyr sequence of cognate Src SH2 phosphopeptides such as Ac-pTyr-Glu-Glu-Ile (1). An x-ray structure of AP22408 complexed with Lck (S164C) SH2 confirmed molecular interactions of both the Dpp and bicyclic template of AP22408 as predicted from molecular modeling. Relative to the cognate phosphopeptide, AP22408 exhibits significantly increased Src SH2 binding affinity (IC(50) = 0.30 microM for AP22408 and 5.5 microM for 1). Furthermore, AP22408 inhibits rabbit osteoclast-mediated resorption of dentine in a cellular assay, exhibits bone-targeting properties based on a hydroxyapatite adsorption assay, and demonstrates in vivo antiresorptive activity in a parathyroid hormone-induced rat model.

Adsorption↗
PubMed2000

Investigating protein-ligand interactions with a mutant FKBP possessing a designed specificity pocket.

Using structure-based design and protein mutagenesis we have remodeled the FKBP12 ligand binding site to include a sizable, hydrophobic specificity pocket. This mutant (F36V-FKBP) is capable of binding, with low or subnanomolar affinities, novel synthetic ligands possessing designed substituents that sterically prevent binding to the wild-type protein. Using binding and structural analysis of bumped compounds, we show here that the pocket is highly promiscuous-capable of binding a range of hydrophobic alkyl and aryl moieties with comparable affinity. Ligand affinity therefore appears largely insensitive to the degree of occupancy or quality of packing of the pocket. NMR spectroscopic analysis indicates that similar ligands can adopt radically different binding modes, thus complicating the interpretation of structure-activity relationships.

Acetamides↗
PubMed2000

Endocannabinoid 2-arachidonyl glycerol is a full agonist through human type 2 cannabinoid receptor: antagonism by anandamide.

The endocannabinoids anandamide and 2-arachidonyl glycerol (2-AG) bind to G protein-coupled central and peripheral cannabinoid receptors CB1 and CB2, respectively. Due to the relatively high expression of the CB2 isotype on peripheral immune cells, it has been hypothesized that this receptor mediates the immunosuppressive effects of cannabinoids. Unfortunately, there was a dearth of pharmacological studies with the endocannabinoids and human CB2 (hCB2). These studies compare and contrast the potency and efficacy of anandamide, 2-AG, and the synthetic cannabinoid HU210 at hCB2. Using [(35)S]guanosine-5'-O-(3-thio)triphosphate (GTPgammaS) and radioligand bindings in insect Sf9-hCB2 membranes, we showed that both endocannabinoids bound hCB2 with similar affinity and that the cannabinoids acted as full agonists in stimulating [(35)S]GTPgammaS exchange, although 2-AG was 3-fold more potent than anandamide (EC(50) = 38.9 +/- 3.1 and 121 +/- 29 nM, respectively). In a mammalian expression system (Chinese hamster ovary-hCB2 cells), HU210 and 2-AG maximally inhibited forskolin-stimulated cAMP synthesis (IC(50) = 1.61 +/- 0.42 nM and 1.30 +/- 0.37 microM, respectively) although anandamide was ineffective. In Chinese hamster ovary-hCB2 membranes, HU210 and 2-AG were also full agonists in stimulating [(35)S]GTPgammaS binding (EC(50) = 1.96 +/- 0.35 and 122 +/- 17 nM, respectively), but anandamide was a weak partial agonist (EC(50) = 261 +/- 91 nM; 34 +/- 4% of maximum). Due to its low intrinsic activity, coincubation with anandamide effectively attenuated the functional activity of 2-AG at hCB2. Collectively, the data showed that both endocannabinoids bound hCB2 with similar affinity, but only 2-AG functioned as a full agonist. Moreover, the agonistic activity of 2-AG was attenuated by anandamide.

Animals↗
PubMed1999

Selective epimerization of rapamycin via a retroaldol/aldol mechanism mediated by titanium tetraisopropoxide.

We describe the efficient and selective epimerization of the immunosuppressant rapamycin to 28-epirapamycin under mild conditions. The mechanism of epimerization involves an equilibrium of the four C28/C29 diastereomers through a two-step retroaldol/aldol (macrocycle ring-opening/ring-closing) sequence. This retroaldol/aldol equilibration is not restricted to rapamycin but is also applicable to acyclic beta-hydroxyketones. A potentially useful extension of the method--the use of beta-hydroxyketones as enolate synthons for effecting inter- or intramolecular aldol reactions under neutral conditions--is demonstrated.

Anti-Bacterial Agents↗
PubMed1999

Induction of CD4+ T cell alloantigen-specific hyporesponsiveness by IL-10 and TGF-beta.

Induction and maintenance of Ag-specific tolerance are pivotal for immune homeostasis, prevention of autoimmune disorders, and the goal of transplantation. Recent studies suggest that certain cytokines, notably IL-10 and TGF-beta, may play a role in down-regulating immune functions. To further examine the role of cytokines in Ag-specific hyporesponsiveness, murine CD4+ T cells were exposed ex vivo to alloantigen-bearing stimulators in the presence of exogenous IL-10 and/or TGF-beta. Primary but not secondary alloantigen proliferative responses were inhibited by IL-10 alone. However, the combined addition of IL-10 + TGF-beta markedly induced alloantigen hyporesponsiveness in both primary and secondary MLR cultures. Alloantigen-specific hyporesponsiveness was observed also under conditions in which nominal Ag responses were intact. In adoptive transfer experiments, IL-10 + TGF-beta-treated CD4+ T cells, but not T cells treated with either cytokine alone, were markedly impaired in inducing graft-vs-host disease alloresponses to MHC class II disparate recipients. These data provide the first formal evidence that IL-10 and TGF-beta have at least an additive effect in inducing alloantigen-specific tolerance, and that in vitro cytokines can be exploited to suppress CD4+ T cell-mediated Ag-specific responses in vivo.

Amino Acid Sequence↗
PubMed1999

Endogenous mouse interleukin-10 is up-regulated by exogenously administered recombinant human interleukin-10, but does not contribute to the efficacy of the human protein in mouse models of endotoxemia.

In murine models of experimental endotoxemia, inflammatory cytokines as well as antiinflammatory interleukin-10 (IL-10) appear in the circulation after the injection of lipopolysaccharide (LPS). There is considerable experimental evidence to suggest that the major function of endogenously produced IL-10 is to down-regulate inflammatory cytokine production. Indeed, the protective effects of exogenously administered IL-10 against murine endotoxin lethality have been shown to correlate with its ability to inhibit the LPS-induced production of tumor necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma). While mouse IL-10 (mIL-10) has been used in the majority of studies in murine endotoxemia, we have found the human homolog to be equally effective in suppressing inflammatory cytokine production and in protecting mice from endotoxin lethality. However, we have recently observed that the LPS-induced endogenous IL-10 response is enhanced when mice are treated with recombinant human IL-10 (rhuIL-10). The upregulation of endogenous IL-10 by exogenously administered rhuIL-10 is particularly evident in mice that are primed with Corynebacterium partum (Proprionibacterium acnes). In the present study, we have examined the potential contributions of the increased circulating levels of mouse IL-10 to the inhibitory effects seen with rhuIL-10 on inflammatory cytokine production and endotoxin lethality. We show that pretreatment with a neutralizing anti-mouse IL-10 monoclonal antibody (mAb) has no effect on the ability of rhuIL-10 to suppress an LPS-induced inflammatory cytokine response in these mice. In contrast, the suppressive effects of the human protein on inflammatory cytokine responses are blocked completely by pretreating the animals with an anti-huIL-10 mAb. These data show that despite the up-regulated endogenous IL-10 response, it is the exogenously administered rhuIL-10 that is directly responsible for the suppressed inflammatory cytokine responses that are observed when the human protein is given to endotoxemic mice.

Animals↗
PubMed1999

Variability in lesion location after microelectrode-guided pallidotomy for Parkinson's disease: anatomical, physiological, and technical factors that determine lesion distribution.

OBJECT: To understand the factors that determine the distribution of lesions after microelectrode-guided pallidotomy for Parkinson's disease, the authors quantitatively characterized lesion location in a cohort of patients who were prospectively followed to determine the effects of pallidotomy on clinical outcome. METHODS: Thirty-three patients underwent volumetric magnetic resonance (MR) imaging after surgery to allow quantitative lesion localization in relation to conventional intraventricular landmarks and, alternatively, more anatomically relevant landmarks. The validity of the method was verified in a cohort of postpallidotomy patients who underwent concurrent volumetric and stereotactic MR imaging in an external head frame. Lesions were distributed over a considerable distance in the anteroposterior (8.8 mm) and mediolateral (8.7 mm) dimensions in relation to the anterior commissure and wall of the third ventricle, respectively. Less variation was seen in lesion location in the dorsoventral dimension (4.8 mm) in relation to the intercommissural plane. CONCLUSIONS: Lesion distribution was not random: lesion locations in the anteroposterior and mediolateral dimensions were highly correlated, such that lesions were distributed from anteromedial to posterolateral, parallel to the border of the globus pallidus internus with the obliquely oriented internal capsule. The factors that led to variability in lesion location were variation in third ventricle width and the oblique anteromedial-to-posterolateral course of the internal capsule. This demonstration of variability of lesion location in a cohort of patients who experienced excellent clinical benefits and minimal postoperative complications emphasizes the importance of anatomical variations in determining lesion position and the need for physiological corroboration for correct lesion placement.

Adult↗
PubMed1997

Characterization of potential antagonists of human interleukin 5 demonstrates their cross-reactivity with receptors for interleukin 3 and granulocyte-macrophage colony-stimulating factor.

The ligand-binding alpha-chain of the human interleukin 5 (IL-5) receptor was expressed in its soluble form, lacking the transmembrane and cytoplasmic domains, from recombinant baculovirus. The soluble receptor was used in a scintillation proximity assay to identify two chemical compounds that inhibit binding of human IL-5 to the soluble receptor alpha chain with IC50 of 8 microM and 11 microM. These compounds also inhibited the interaction of human IL-5 with its membrane-bound receptor, composed of the ligand-binding alpha chain and signal-transducing beta chain, and prevented signaling through the receptor. Analysis by surface plasmon resonance and matrix-assisted laser-desorption/ionization mass spectrometry showed that the identified compounds bound irreversibly to the receptor at a 1:1 (mol/mol) ratio, suggesting a covalent interaction with the alpha chain of the human IL-5 receptor. Both compounds also inhibited the interaction of the receptors for interleukin 3 (IL-3) and granulocyte-macrophage colony-stimulating factor (GM-CSF), which are involved in hematopoietic differentiation and activation of immune cells, thus eliminating them as potential therapeutic agents. The inhibition of the structurally closely related receptors for IL-5, IL-3 and GM-CSF by both compounds, while binding of interleukin-4 to its receptor was not affected, suggests that a similar reactive site exists in the ligand-binding domains of the receptors for IL-5, IL-3 and GM-CSF.

Animals↗
PubMed1997

Interferon-alpha gene therapy for cancer: retroviral transduction of fibroblasts and particle-mediated transfection of tumor cells are both effective strategies for gene delivery in murine tumor models.

Stable transfection of tumor cells with IFN-alpha genes has been shown to result in abrogation of tumor establishment and induction of antitumor immunity. However, strategies suitable for the clinical application of IFN-alpha gene therapy for cancer have not been reported. In this study, we investigated two gene delivery systems capable of mediating the local paracrine production of high levels of biologically active IFN-alpha in murine tumor models: retroviral transduction of fibroblasts and particle-mediated transfection of tumor cells. In spite of the antiproliferative effects of IFN-alpha, it was possible to obtain stable retroviral producer cell lines and transduce a variety of murine tumor cells including syngeneic fibroblasts to stably secrete 2000-5000 U (40-100 ng) murine IFN-alpha/10(6) cells/24 h. IFN-alpha transduction of tumor cells abrogated tumorigenicity in establishment models and induced antitumor immunity in several murine tumor model systems. Importantly, IFN-alpha gene delivery using retrovirally transduced syngeneic fibroblasts was capable of suppressing the establishment of the poorly immunogenic TS/A mouse mammary adenocarcinoma and induced antitumor immunity. Particle mediated transient transfection of tumor cells using the gene gun led to the production of up to 20,000 U IFN-alpha/10(6) cells during the first 24 h and proved to be equally effective in suppressing establishment of TS/A adenocarcinoma and inducing antitumor immunity. These results suggest that retroviral transduction of autologous fibroblasts can serve as an effective gene delivery method for IFN-alpha gene therapy of cancer. Particle-mediated transfection of freshly isolated tumor cells may represent a clinically attractive alternative approach for nonviral gene delivery. Both strategies circumvent the difficulties in routinely establishing primary tumor cell lines from the vast majority of human cancers.

Animals↗
PubMed1997

Evaluation of plasma serotonin concentration in acute appendicitis.

BACKGROUND: Due to lack of reliable biochemical/radiological markers, the diagnosis of acute appendicitis is based only on clinical features. METHODS: We estimated plasma serotonin levels in 48 patients with acute appendicitis (histologically proven), 27 patients with abdominal pain of other etiologies, and 20 healthy controls. RESULTS: The plasma serotonin levels were (mean +/- SD) 36.6 +/- 12.5 nmol/L, 12.5 +/- 3.6 nmol/L and 10.4 +/- 3.5 nmol/L in the three groups, respectively. The levels in patients with acute appendicitis were significantly higher (p < 0.001) than in the other groups, giving 93.8% sensitivity and 95.7% specificity to the test. CONCLUSION: Plasma serotonin level is a reliable marker of acute appendicitis, especially in the first 48 hours.

Abdominal Pain↗