Bio

Professional Education


  • Doctor of Philosophy, Tufts University (2008)
  • Master's, University of Montreal, Chemistry (2003)

Stanford Advisors


Publications

Journal Articles


  • CYP3A4-Catalyzed Simvastatin Metabolism as a Non-Invasive Marker of Small Intestinal Health in Celiac Disease. American journal of gastroenterology Morón, B., Verma, A. K., Das, P., Taavela, J., Dafik, L., DiRaimondo, T. R., Albertelli, M. A., Kraemer, T., Mäki, M., Khosla, C., Rogler, G., Makharia, G. K. 2013; 108 (8): 1344-1351

    Abstract

    Histological examination of duodenal biopsies is the gold standard for assessing intestinal damage in celiac disease (CD). A noninvasive marker of disease status is necessary, because obtaining duodenal biopsies is invasive and not suitable for routine monitoring of CD patients. As the small intestine is a major site of cytochrome P450 3A4 (CYP3A4) activity and also the location of the celiac lesion, we investigated whether patients with active CD display abnormal pharmacokinetics of an orally administered CYP3A4 substrate, simvastatin (SV), which could potentially be used for noninvasive assessment of their small intestinal health.Preclinical experiments were performed in CYP3A4-humanized mice to examine the feasibility of the test. Subsequently, a clinical trial was undertaken with 11 healthy volunteers, 18 newly diagnosed patients with CD, and 25 celiac patients who had followed a gluten-free diet (GFD) for more than 1 year. The maximum concentration (Cmax) of orally administered SV plus its major non-CYP3A4-derived metabolite SV acid (SV equivalent (SVeq)) was measured, and compared with clinical, histological, and serological parameters.In CYP3A4-humanized mice, a marked decrease in SV metabolism was observed in response to enteropathy. In the clinical setting, untreated celiac patients displayed a significantly higher SVeq Cmax (46±24 nM) compared with treated patients (21±16 nM, P<0.001) or healthy subjects (19±11 nM, P<0.005). SVeq Cmax correctly predicted the diagnosis in 16/18 untreated celiac patients, and also the recovery status of all follow-up patients that exhibited normal or near-normal biopsies (Marsh 0-2). All patients with abnormal SVeq Cmax showed a reduction in the value after 1 year of following a GFD.SVeq Cmax is a promising noninvasive marker for assessment of small intestinal health. Further studies are warranted to establish its clinical utility for assessing gut status of patients with CD.

    View details for DOI 10.1038/ajg.2013.151

    View details for PubMedID 23732466

  • Anti-Oligomannose Antibodies as Potential Serum Biomarkers of Aggressive Prostate Cancer DRUG DEVELOPMENT RESEARCH Wang, D., Dafik, L., Nolley, R., Huang, W., Wolfinger, R. D., Wang, L., Peehl, D. M. 2013; 74 (2): 65-80

    View details for DOI 10.1002/ddr.21063

    View details for Web of Science ID 000316463700002

  • G alpha(h)/transglutaminase-2 activity is required for maximal activation of adenylylcyclase 8 in human and rat glioma cells CELLULAR SIGNALLING Obara, Y., Yanagihata, Y., Abe, T., Dafik, L., Ishii, K., Nakahata, N. 2013; 25 (3): 589-597

    Abstract

    Gα(h) (or transglutaminase-2 (TG2)) is an atypical guanine nucleotide binding-protein that associates with G protein-coupled receptors. TG2 also exerts transglutaminase activity that catalyzes posttranslational protein cross-linking with the formation of ε-(γ-glutamyl) lysine or (γ-glutamyl) polyamine bonds. Here, the role of Gα(h)/TG2 in signal transduction in glial cells was examined in detail. In 1321N1 human astrocytoma cells that lack Gα(h)/TG2, overexpression of Gα(h)/TG2 caused an enhancement of cAMP accumulation stimulated with the β-adrenergic receptor agonist, isoproterenol, or the adenylylcyclase activator, forskolin. This cAMP-enhancement was reversed by the TG2 inhibitor, ERW1069. In rat C6 glioma cells that express endogenous Gα(h)/TG2, cAMP accumulation induced by isoproterenol or forskolin was significantly inhibited by overexpression of Gα(h)/TG2-C277V, a dominant-negative mutant that lacks transglutaminase activity, but was not inhibited by the Gα(h)/TG2-S171E mutant that cannot bind GTP/GDP. These results suggest Gα(h)/TG2 potentiates adenylylcyclase activity by its transglutaminase activity and not by its G-protein activity. Gα(h)/TG2 also increased the activities of the cAMP response element and interleukin-6 promoter, accompanied by an of cAMP in both glioma cells. Since adenylylcyclase 8 plays a major role in cAMP production, we focused on post-translational modification of adenylylcyclase 8 by Gα(h)/TG2. Adenylylcyclase 8 is expressed in both 1321N1 and C6 cells; however, Gα(h)/TG2 affected neither adenylylcyclase 8 expression levels, glycosylation, nor dimerization status. In contrast, pentylamine, a substrate of Gα(h)/TG2, was incorporated into adenylylcyclase 8 in a transglutaminase activity-dependent manner. Taking these results together, Gα(h)/TG2 promotes cAMP production accompanied by a modification of adenylylcyclase 8 in glioma cells.

    View details for DOI 10.1016/j.cellsig.2012.11.021

    View details for Web of Science ID 000315932000002

    View details for PubMedID 23200849

  • Activation and Inhibition of Transglutaminase 2 in Mice PLOS ONE Dafik, L., Albertelli, M., Stamnaes, J., Sollid, L. M., Khosla, C. 2012; 7 (2)

    Abstract

    Transglutaminase 2 (TG2) is an allosterically regulated enzyme with transamidating, deamidating and cell signaling activities. It is thought to catalyze sequence-specific deamidation of dietary gluten peptides in the small intestines of celiac disease patients. Because this modification has profound consequences for disease pathogenesis, there is considerable interest in the design of small molecule TG2 inhibitors. Although many classes of TG2 inhibitors have been reported, thus far an animal model for screening them to identify promising celiac drug candidates has remained elusive. Using intraperitoneal administration of the toll-like receptor 3 (TLR3) ligand, polyinosinic-polycytidylic acid (poly(I?C)), we induced rapid TG2 activation in the mouse small intestine. Dose dependence was observed in the activation of TG2 as well as the associated villous atrophy, gross clinical response, and rise in serum concentration of the IL-15/IL-15R complex. TG2 activity was most pronounced in the upper small intestine. No evidence of TG2 activation was observed in the lung mucosa, nor were TLR7/8 ligands able to elicit an analogous response. Introduction of ERW1041E, a small molecule TG2 inhibitor, in this mouse model resulted in TG2 inhibition in the small intestine. TG2 inhibition had no effect on villous atrophy, suggesting that activation of this enzyme is a consequence, rather than a cause, of poly(I?C) induced enteropathy. Consistent with this finding, administration of poly(I?C) to TG2 knockout mice also induced villous atrophy. Our findings pave the way for pharmacological evaluation of small molecule TG2 inhibitors as drug candidates for celiac disease.

    View details for DOI 10.1371/journal.pone.0030642

    View details for Web of Science ID 000301979000009

    View details for PubMedID 22319575

  • Dihydroisoxazole Analogs for Labeling and Visualization of Catalytically Active Transglutaminase 2 CHEMISTRY & BIOLOGY Dafik, L., Khosla, C. 2011; 18 (1): 58-66

    Abstract

    We report the synthesis and preliminary characterization of "clickable" inhibitors of human transglutaminase 2 (TG2). These inhibitors possess the 3-halo-4,5-dihydroisoxazole warhead along with bioorthogonal groups such as azide or alkyne moieties that enable subsequent covalent modification with fluorophores. Their mechanism for inhibition of TG2 is based on halide displacement, resulting in the formation of a stable imino thioether. Inhibition assays against recombinant human TG2 revealed that some of the clickable inhibitors prepared in this study have comparable specificity as benchmark dihydroisoxazole inhibitors reported earlier. At low micromolar concentrations they completely inhibited transiently activated TG2 in a WI-38 fibroblast scratch assay and could subsequently be used to visualize the active enzyme in situ. The potential use of these inhibitors to probe the role of TG2 in celiac sprue as well as other diseases is discussed.

    View details for DOI 10.1016/j.chembiol.2010.11.004

    View details for Web of Science ID 000287540300011

    View details for PubMedID 21276939

  • Modulation of Cellular Adhesion by Glycoengineering JOURNAL OF MEDICINAL CHEMISTRY Dafik, L., d'Alarcao, M., Kumar, K. 2010; 53 (10): 4277-4284

    Abstract

    Aberrant glycosylation of lipid and protein molecules on cellular surfaces is responsible for many of the pathophysiological events in tumor progression and metastasis. Sialic acids in particular, are overexpressed on the glycocalyx of malignant tumor cells and sialic acid-mediated cell adhesion is required for metastasis. We report here that replacement of sialic acids on cell surfaces with fluorinated congeners dramatically decreases cell adhesion to E- and P-selectin-coated surfaces. Comparison of adhesion of fluorinated cells with those modified with nonfluorinated analogues suggests that both reduce binding of the modified sialosides to their cognate lectins to a similar extent on a per molecule basis. The overall reduction in cell adhesion results from greater cell surface presentation of the fluorinated congeners. This work suggests an avenue for inhibition of metastasis by administration of small molecules and concomitant noninvasive imaging of tumor cells by (19)F MRI before they are visible by other means.

    View details for DOI 10.1021/jm100374g

    View details for Web of Science ID 000277766900038

    View details for PubMedID 20438083

  • Fluorinated Lipid Constructs Permit Facile Passage of Molecular Cargo into Living Cells JOURNAL OF THE AMERICAN CHEMICAL SOCIETY Dafik, L., Kalsani, V., Leung, A. K., Kumar, K. 2009; 131 (34): 12091-?

    Abstract

    Fluorinated lipids get rapidly internalized into living cells and are also displayed on the cell surface. The uptake of lipids is energy dependent and is likely via the clathrin-mediated endocytic pathway. Fluorinated lipids are 3-5-fold more efficient in acting as molecular transporters of noncovalently bound proteins than their hydrocarbon counterparts. These materials could serve as efficient molecular transporters for molecules that function in the cytoplasm such as short interfering RNAs (siRNAs).

    View details for DOI 10.1021/ja902777d

    View details for Web of Science ID 000269379600035

    View details for PubMedID 19673473

  • Fluorination of mammalian cell surfaces via the sialic acid biosynthetic pathway BIOORGANIC & MEDICINAL CHEMISTRY LETTERS Dafik, L., d'Alarcao, M., Kumar, K. 2008; 18 (22): 5945-5947

    Abstract

    Metabolic oligosaccharide engineering has been employed to introduce fluorine-containing groups onto mammalian cell surfaces. Incubation of HeLa, Jurkat, and HL60 cells in culture with fluorinated sialic acid and mannosamine analogues resulted in cell-surface presentation of fluorinated glycans. Metabolic conversion of fluorinated precursors was detected and quantified by DMB-derivatization and HPLC ESI-MS analysis. Between 7% and 72% of total membrane-associated sialosides were fluorinated, depending on the precursor used and the cell type. Fluorination of mammalian cell surfaces provides a means for introducing a bioorthogonal surface for modulating noncovalent interactions such as those involved in cell adhesion.

    View details for DOI 10.1016/j.bmcl.2008.09.010

    View details for Web of Science ID 000260966800028

    View details for PubMedID 18819798

  • Bioorthogonal noncovalent chemistry: fluorous phases in chemical biology CURRENT OPINION IN CHEMICAL BIOLOGY Yoder, N. C., Yuksel, D., Dafik, L., Kumar, K. 2006; 10 (6): 576-583

    Abstract

    Chemical entities designed to noncovalently interact with predetermined partners have fashioned a new paradigm in chemical biology. Fluorocarbons are extremely promising as supramolecular synthons toward these objectives. Bioorthogonal noncovalent interactions provide a way to modulate self-assembled systems in environments where such control has hitherto not been possible. Fluorocarbons have now found applications in self-assembly as well as proteomics, biomolecule purification and in the creation of microarray platforms. Other self-assembly motifs with similar attributes might be exploited using the same general approach.

    View details for DOI 10.1016/j.cbpa.2006.10.007

    View details for Web of Science ID 000242919700008

    View details for PubMedID 17055332

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