Non-insulin dependent diabetes mellitus (type 2 diabetes) affects tens of millions of people worldwide. The search for novel drug candidates is paramount in diabetes research. We have found that thiolane derivatives increase the rate of glucose transport into insulin-sensitive cells such as adipose cells and skeletal muscle cells, but not into insulin-insensitive cells, such as vascular cells. In addition such molecules augment insulin secretion. We synthesized novel potent thiolane and benzoimidazole derivatives with favorable pharmacokinetic and pharmacodynamic parameters and tested these molecules in both, in vitro and in vivo model systems. Several such compounds dose- and time-dependently increased the rate of glucose uptake in L6 myotubes and insulin secretion in INS-1E cells. The compounds exerted their effects through the activation of the LKB1 (Liver Kinase B1)-AMPK pathway. Subcutaneous administration of the lead compound to hyperglycemic Kuo Kondo mice carrying the Ay-yellow obese gene (KKAy) lowered blood glucose levels toward the normoglycemic range.
ALS is a fatal neurodegenerative disease characterized by progressive muscle weakness and reflective loss of upper and lower motor neurons, predominantly in the spinal cord. No effective drugs were developed against this disease so far. We designed and synthesized novel targeted to intracellular organelles chemical chaperones that increase the degradation of disease related misfolded proteins and increase the survival of motor neurons. One of such molecules was active in ALS mice.
Novel therapies based on mimicking of function and structure of transcription factor: NRF-2.
Nuclear erythroid 2-related factor 2 (NRF-2) is a basic region leucine-zipper transcription factor that binds to the antioxidant response element (ARE), thereby regulating the expression of hundreds of genes that are involved in cellular defense against oxidative stress. In addition, NRF-2 regulates the expression of pro- and anti-inflammatory enzymes, and it also acts as an inhibitor of Fas-induced apoptosis. In our preliminary study we have shown that injection of three genes into the striatum (among them NRF-2) led to the reduction of the glutamate levels, provided neuroprotection and finally improved the motor functions and the typical rotation behavior mice with neurodegenerative diseases. We proposed to replace the injection of NRF-2 gene by administration of its mimetics-small organic molecules which will be able to protect neurons from death via the activation of NRF-2 pathway and subsequent enhancement of neurons resistance against oxidative stress, a major pathomechanism in neurodegenerative diseases. Using a multidisciplinary approach (computer-aided drug design, synthetic organic chemistry, in vivo and in vitro models, biochemistry, molecular biology) we developed several small molecules that stimulate the NRF-2 pathway.
Toll-like receptors (TLRs) serve as pattern recognition receptors in the innate immune response to microbial pathogens. TLR4 specifically mediates cellular responses generated after the binding to the receptor bacterial lipopolysaccharide (LPS). TLR4 also responds to endogenous factors produced during different stress stimuli or cell damage. The inhibition of a TLR4-mediated immune response as a potential therapeutic approach might be used for the development of novel cardioprotective drugs. This approach offers promise in the ability to reduce the inflammatory cascade in the heart, and potentially lead to an improved prognosis following MI- induced cardiodamage. Based on a known TLR4 inhibitory peptide, which is built by 9 amino acids and corresponds to the sequences from the TIR domain containing adapter protein TRAM, we have synthesized several peptidomimetics. We investigated the ability of the synthetized peptidomimetics to reduce hypoxic cardiodamage in vitro and in vivo. In both models, our novel peptidomimetics showed very promising biological results.
1α-Antitrypsin (AAT) is a multi-functional protease inhibitor produced by the liver and lungs; it plays an important role in the regulation of inflammation. The role of AAT as a protease inhibitor has already been intensively studied as well as the AAT sequence required for binding to different proteases. However, AAT functions that are not related to its anti-proteases activity have been investigated much less. We hypothesize that AAT regulates anti-inflammatory response by binding to different proteins (that are not proteases). These interactions differ from their anti-protease binding sequences in the AAT surface. By using molecular modelling, 12 peptides were derived from the crystal structure of AAT. These peptides provide potential binding sites to other proteins. Of these, two peptides: ES-11P and ES-12P significantly reduced the inflammatory response of immune cells in two parameters: (1) decreasing the expression of several proteins which are responsible for the differentiation of cell. (2) The reduction of nitrite release. In addition one of the peptides was active in vivo, in mice wound healing model. Now we are developing drugable peptidomimetic molecules that can mimic interactions between AAT and different inflammation-related proteins.
Inhibitor of leukocyte transendothelial migration
Leukocyte transendothelial migration is one of the most important steps in launching an inflammatory immune response and chronic inflammation. Leukocyte migration inhibitors are considered as promising and potentially effective therapeutic agents to treat inflammatory and auto-immune disorders. We developed a novel molecule: methyl 4-((2-(tert-butyl)-6-((2,4,6-trioxotetrahydropyrimidin-5(2H)-ylidene) methyl) phenoxy) methyl) benzoate, (compound 12 or GT-73), that completely blocked leukocyte transendothelial migration, without any toxic effects on immune or endothelial cells (IC50=2.4 µM). In vivo, compound 12 exhibited significant therapeutic effects in inflammatory bowel disease (IBD)/Crohn’s disease, multiple sclerosis, fatty liver disease, and rheumatoid arthritis models. A detailed acute and chronic toxicity profile of the lead compound in vivo did not reveal any toxic effects. Such a type of molecule might, therefore, provide a unique starting point for designing a novel class of leukocyte transmigration blocking agents with broad therapeutic applications in inflammatory and auto-immune pathologies.
SYNTHETIC CHEMISTRY OF BIOGENIC MOLECULES
Apelin receptor (APJ) is a recently identified G protein-coupled receptor activated by the endogenous peptide apelin. The apelin–APJ system has emerged as an important regulator of cardiovascular homeostasis. Recently, a potent benzimidazole-derived apelin peptidomimetic, CMF-019, was patented by Sanofi-Aventis but without a comprehensive description of its synthesis or full spectroscopic characterization of the intermediates. We developed a detailed preparation of CMF-019 through a modified and improved synthetic pathway.
Inhibiting the copper efflux system in Gram negative microbes by peptidomimetics as a novel approach for developing antibiotics
The spread of resistant pathogenic microbes has become one of the most important and urgent problems that medicine faces today. In this proposal, we present a novel approach for the development of new antibiotics, based on basic research related to copper toxicity. In cells, copper ions are double-edged swords. On the one hand, they are essential cofactors, but on the other hand, they are highly toxic. Prokaryotic organisms are more sensitive to copper than are eukaryotic systems, and therefore they employ additional copper transporters that have no homology in the eukaryotic cells. Blocking the copper transport outside of bacteria might be used as a novel approach for the development of effective treatment against bacterial vector diseases.
Especially taking into consideration that the copper transport system in bacteria is very different from humans and such inhibitors should be non-toxic. We designed in silico peptidomimetics that can block CusB flexibility in Acinetobacter baumannii, Pseudomonas aeruginosa, pathogenic ESBL-producing Escherichia Coli and Klebsiella pneumonia. One of them showed promising results as a bacteriostatic agent.