Gluten Exorphins Promote Cell Proliferation through the Activation of Mitogenic and Pro-Survival Pathways.

Celiac disease (CD) is a chronic and systemic autoimmune disorder that affects preferentially the small intestine of individuals with a genetic predisposition. CD is promoted by the ingestion of gluten, a storage protein contained in the endosperm of the seeds of wheat, barley, rye, and related cereals. Once in the gastrointestinal (GI) tract, gluten is enzymatically digested with the consequent release of immunomodulatory and cytotoxic peptides, i.e., 33mer and p31-43. In the late 1970s a new group of biologically active peptides, called gluten exorphins (GEs), was discovered and characterized. In particular, these short peptides showed a morphine-like activity and high affinity for the δ-opioid receptor (DOR). The relevance of GEs in the pathogenesis of CD is still unknown. Recently, it has been proposed that GEs could contribute to asymptomatic CD, which is characterized by the absence of symptoms that are typical of this disorder. In the present work, GEs cellular and molecular effects were in vitro investigated in SUP-T1 and Caco-2 cells, also comparing viability effects with human normal primary lymphocytes. As a result, GEs treatments increased tumor cell proliferation by cell cycle and Cyclins activation as well as by induction of mitogenic and pro-survival pathways. Finally, a computational model of GEs interaction with DOR is provided. Altogether, the results might suggest a possible role of GEs in CD pathogenesis and on its associated cancer comorbidities.

Linker residues regulate the activity and stability of hexokinase 2, a promising anticancer target.

Hexokinase (HK) catalyzes the first step in glucose metabolism, making it an exciting target for the inhibition of tumor initiation and progression due to their elevated glucose metabolism. The upregulation of hexokinase-2 (HK2) in many cancers and its limited expression in normal tissues make it a particularly attractive target for the selective inhibition of cancer growth and the eradication of tumors with limited side effects. The design of such safe and effective anticancer therapeutics requires the development of HK2-specific inhibitors that will not interfere with other HK isozymes. As HK2 is unique among HKs in having a catalytically active N-terminal domain (NTD), we have focused our attention on this region. We previously found that NTD activity is affected by the size of the linker helix-α13 that connects the N- and C-terminal domains of HK2. Three nonactive site residues (D447, S449, and K451) at the beginning of the linker helix-α13 have been found to regulate the NTD activity of HK2. Mutation of these residues led to increased dynamics, as shown via hydrogen deuterium exchange analysis and molecular dynamic simulations. D447A contributed the most to the enhanced dynamics of the NTD, with reduced calorimetric enthalpy of HK2. Similar residues exist in the C-terminal domain (CTD) but are unnecessary for HK1 and HK2 activity. Thus, we postulate these residues serve as a regulatory site for HK2 and may provide new directions for the design of anticancer therapeutics that reduce the rate of glycolysis in cancer through specific inhibition of HK2.

Utilization of Biased G Protein-Coupled ReceptorSignaling towards Development of Safer andPersonalized Therapeutics.

G protein-coupled receptors (GPCRs) are involved in a wide variety of physiologicalprocesses. Therefore, approximately 40% of currently prescribed drugs have targeted this receptorfamily. Discovery of ß-arrestin mediated signaling and also separability of G protein and b-arrestinsignaling pathways have switched the research focus in the GPCR field towards development ofbiased ligands, which provide engagement of the receptor with a certain effector, thus enrichinga specific signaling pathway. In this review, we summarize possible factors that impact signalingprofiles of GPCRs such as oligomerization, drug treatment, disease conditions, genetic background,etc. along with relevant molecules that can be used to modulate signaling properties of GPCRssuch as allosteric or bitopic ligands, ions, aptamers and pepducins. Moreover, we also discuss theimportance of inclusion of pharmacogenomics and molecular dynamics simulations to achieve aholistic understanding of the relation between genetic background and structure and function ofGPCRs and GPCR-related proteins. Consequently, specific downstream signaling pathways can beenriched while those that bring unwanted side effects can be prevented on a patient-specific basis.This will improve studies that centered on development of safer and personalized therapeutics,thus alleviating the burden on economy and public health.

Mechanistic insight into the impact of a bivalent ligand on the structure and dynamics of a GPCR oligomer.

Development of effective bivalent ligands has become the focus of intensive research toward modulation of G protein-coupled receptor (GPCR) oligomers, particularly in the field of GPCR pharmacology. Experimental studies have shown that they increased binding affinity and signaling potency compared to their monovalent counterparts, yet underlying molecular mechanism remains elusive. To address this, we performed accelerated molecular dynamics simulations on bivalent-ligand bound Adenosine 2A receptor (A2AR) dimer in the context of a modeled tetramer, which consists of A2AR and dopamine 2 receptor (D2R) homodimers and their cognate G proteins. Our results demonstrate that bivalent ligand impacted interactions between pharmacophore groups and ligand binding residues, thus modulating allosteric communication network and water channel formed within the receptor. Moreover, it also strengthens contacts between receptor and G protein, by modulating the volume of ligand binding pocket and intracellular domain of the receptor. Importantly, we showed that impact evoked by the bivalent ligand on A2AR dimer was also transmitted to apo D2R, which is part of the neighboring D2R dimer. To the best of our knowledge, this is the first study that provides a mechanistic insight into the impact of a bivalent ligand on dynamics of a GPCR oligomer. Consequently, this will pave the way for development of effective ligands for modulation of GPCR oligomers and hence treatment of crucial diseases such as Parkinson's disease and cancer.