Publications by authors named "Elena Shirokova"

21 Publications

  • Page 1 of 1

Probiotics in hepatology: An update.

World J Hepatol 2021 Sep;13(9):1154-1166

Department of Internal Medicine, Gastroenterology and Hepatology, Sechenov University, Moscow 119435, Russia.

The gut-liver axis plays an important role in the pathogenesis of various liver diseases. Probiotics are living bacteria that may be used to correct disorders of this axis. Notable progress has been made in the study of probiotic drugs for the treatment of various liver diseases in the last decade. It has been proven that probiotics are useful for hepatic encephalopathy, but their effects on other symptoms and syndromes of cirrhosis are poorly studied. Their effectiveness in the treatment of metabolic associated fatty liver disease has been shown both in experimental models and in clinical trials, but their effect on the prognosis of this disease has not been described. The beneficial effects of probiotics in alcoholic liver disease have been shown in many experimental studies, but there are very few clinical trials to support these findings. The effects of probiotics on the course of other liver diseases are either poorly studied (such as primary sclerosing cholangitis, chronic hepatitis B and C, and autoimmune hepatitis) or not studied at all (such as primary biliary cholangitis, hepatitis A and E, Wilson's disease, hemochromatosis, storage diseases, and vascular liver diseases). Thus, despite the progress in the study of probiotics in hepatology over the past decade, there are many unexplored and unclear questions surrounding this topic.
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http://dx.doi.org/10.4254/wjh.v13.i9.1154DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8473492PMC
September 2021

Gut dysbiosis is associated with poorer long-term prognosis in cirrhosis.

World J Hepatol 2021 May;13(5):557-570

Department of Internal Medicine, Gastroenterology and Hepatology, Sechenov University, Moscow 119435, Russia.

Background: Gut dysbiosis is common in cirrhosis.

Aim: To study the influence of gut dysbiosis on prognosis in cirrhosis.

Methods: The case-control study included 48 in-patients with cirrhosis and 21 healthy controls. Stool microbiome was assessed using 16S ribosomal ribonucleic acid gene sequencing. We used modified dysbiosis ratio (MDR): [ (%) + (%)]/[ (%) + (%)]. Patients with MDR more the median made up the group with severe dysbiosis, others did the group with non-severe dysbiosis. The follow-up period was 4 years.

Results: The mortality rate of patients with severe dysbiosis was significantly higher than that of patients with non-severe dysbiosis (54.2% 12.5%; = 0.001). The presence of severe dysbiosis was independent risk factors for death [hazard ratio = 8.6 × (1.9-38.0); = 0.005]. The abundance of ( = 0.002), ( = 0.002), and ( = 0.025) was increased and the abundance of ( = 0.025) and ( = 0.045) was decreased in the deceased patients compared with the survivors. The deceased patients had a higher MDR value than the survivors [0.131 × (0.069-0.234) 0.034 × (0.009-0.096); = 0.004]. If we applied an MDR value of 0.14 as the cutoff point, then it predicted patient death within the next year with a sensitivity of 71.4% and a specificity of 82.9% [area under the curve = 0.767 × (0.559-0.974)]. MDR was higher in patients with cirrhosis than in health controls [0.064 × (0.017-0.131) 0.005 × (0.002-0.007); < 0.001], and in patients with decompensated cirrhosis than in patients with compensated cirrhosis [0.106 × (0.023-0.211) 0.033 × (0.012-0.074); = 0.031]. MDR correlated negatively with prothrombin ( = -0.295; = 0.042), cholinesterase ( = -0.466; = 0.014) and serum albumin ( = -0.449; = 0.001) level and positively with Child-Turcotte-Pugh scale value ( = 0.360; = 0.012).

Conclusion: Gut dysbiosis is associated with a poorer long-term prognosis in cirrhosis.
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http://dx.doi.org/10.4254/wjh.v13.i5.557DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8173342PMC
May 2021

Immune disorders and rheumatologic manifestations of viral hepatitis.

World J Gastroenterol 2021 May;27(18):2073-2089

Department of Internal Medicine, Gastroenterology and Hepatology, Sechenov University, Moscow 119435, Russia.

Infection with hepatotropic viruses is not limited to the liver and can lead to the development of various immunological disorders (the formation of cryoglobulins, rheumatoid factor, antinuclear antibodies, autoantibodies specific for autoimmune hepatitis and primary biliary cholangitis, and others), which can manifest as glomerulonephritis, arthritis, uveitis, vasculitis (cryoglobulinemic vasculitis, polyarteritis nodosa, Henoch-Schonlein purpura, isolated cutaneous necrotizing vasculitis), and other rheumatologic disorders, and be a trigger for the subsequent development of autoimmune hepatitis and primary biliary cholangitis. A further study of the association between autoimmune liver diseases and hepatotropic virus infection would be useful to assess the results of treatment of these associated diseases with antiviral drugs. The relationship of these immune disorders and their manifestations with hepatotropic viruses is best studied for chronic hepatitis B and C. Only isolated cases of these associations are described for hepatitis A. These links are least studied, and are often controversial for hepatitis E, possibly due to their relatively rare diagnoses. Patients with uveitis, glomerulonephritis, arthritis, vasculitis, autoimmune liver diseases should be tested for biomarkers of viral hepatitis, and if present, these patients should be treated with antiviral drugs.
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http://dx.doi.org/10.3748/wjg.v27.i18.2073DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8117740PMC
May 2021

Correlation of Objective Endpoints and Subjective Patient-Reported Outcomes in NAFLD Treatment with Essential Phospholipids: Real-World Data Based on Pooled Analysis of Observational Studies.

Drugs Real World Outcomes 2021 Sep 15;8(3):369-382. Epub 2021 May 15.

Department of Medical Affairs, Sanofi, 125009 Business Center "Summit", Tverskaya 22, Moscow, Russia.

Background: While no "gold-standard" pharmacotherapy for nonalcoholic fatty liver disease (NAFLD) is yet established, essential phospholipids (EPLs) are reported to decrease steatosis and improve laboratory parameters.

Objective: This analysis evaluated adherence and satisfaction with EPL treatment as patient-reported outcomes and their relationship with changes in laboratory and ultrasound parameters among Russian patients with NAFLD.

Methods: Data were pooled from three observational Russian studies-MANPOWER (2015-2016), LIDER 1 (2012-2013), and LIDER 2 (2013)-in which EPLs were used for at least 12 weeks in the treatment of liver diseases and which measured both subjective and objective endpoints. Only patients who had NAFLD were included in this analysis. The main endpoints were to determine treatment adherence and satisfaction with 12 weeks of EPL therapy, relationship between adherence/satisfaction and changes in the laboratory and ultrasound parameters. A secondary subgroup analysis was performed to identify patients with NAFLD who responded better (or worse) to 24 weeks of adjunctive EPL treatment.

Results: Overall, 3384 patients were included. A total of 82.2% of patients were adherent to 12 weeks of EPL treatment; high/very high satisfaction was reported by 15.3%/65.9% of clinicians and 15.9%/64.4% of patients. There was positive correlation between patients' adherence and satisfaction and significant improvement in laboratory (transaminases, lipid profile; p < 0.001) and ultrasound (steatosis, p < 0.001) parameters, and improvement in symptoms (p < 0.001) after 24 weeks of EPL. Male patients, patients with unhealthy lifestyles, and those with more comorbidities showed a better response in laboratory and ultrasound parameters.

Conclusions: Patients with NAFLD treated with adjunctive EPL therapy in real-world clinical practice in Russia showed good treatment adherence and treatment satisfaction. Improvements in laboratory and ultrasound parameters, as well as dynamics of patient symptoms, were positively correlated with adherence and satisfaction.
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http://dx.doi.org/10.1007/s40801-021-00250-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8123926PMC
September 2021

Effectiveness of phosphatidylcholine as adjunctive therapy in improving liver function tests in patients with non-alcoholic fatty liver disease and metabolic comorbidities: real-life observational study from Russia.

BMJ Open Gastroenterol 2020 26;7(1):e000368. Epub 2020 Mar 26.

Medical Affairs, Sanofi-Aventis SA, Moscow, Russian Federation.

Objective: Non-alcoholic fatty liver disease (NAFLD) is the most common cause of abnormal results of liver function tests. Earlier research showed that polyenylphosphatidylcholine (PPC) has hepatoprotective effects and thus can be used for the treatment of NAFLD and the prevention of its progression. Accordingly, the aim of this observational study was to evaluate if PPC administered as adjunctive therapy in routine clinical practice can effectively improve liver function tests of NAFLD in Russian patients with associated metabolic comorbidities.

Design: A total of 2843 adult patients with newly diagnosed NAFLD, who had a least one of four comorbidities, namely, overweight/obesity, hypertension, type 2 diabetes mellitus, and hypercholesterolaemia, and who were prescribed 1.8 g/day of PPC as an adjunctive treatment to standard care, were enrolled during 2015-2016. Laboratory data were collected at baseline and 12 and 24 weeks of the study, and included liver function tests (aspartate aminotransferase (AST), alanine aminotransferase (ALT), gamma-glutamyl transferase (GGT)), fasting plasma glucose, and lipid profile.

Results: Overall, 2263 patients (79.6%) had at least two metabolic comorbidities associated with NAFLD, and overweight/obesity was the most common comorbidity reported in 2298 (80.8%) patients. At 24 weeks, there was a significant decrease in liver enzyme levels (all p<0.001 compared with baseline). Across the four comorbidity subgroups, there was a mean drop of ALT levels ranging from 19.7 to 22.0 U/L, AST from 16.9 to 18.4 U/L, and GGT from 17.2 to 18.7 U/L. Similar findings were reported in subgroups with either one, two, three, or four comorbidities, with a significant decrease in liver enzyme levels ranging from 18.4 to 22.4 U/L for ALT, 14.8 to 18.7 U/L for AST, and 15.5 to 19.5 U/L for GGT.

Conclusions: Adjuvant treatment with PPC resulted in consistent improvements in liver enzymes in patients with newly diagnosed NAFLD and associated metabolic comorbidities.

Trial Registration Number: NCT00063622.
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http://dx.doi.org/10.1136/bmjgast-2019-000368DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7170405PMC
November 2021

Effectiveness of phosphatidylcholine in alleviating steatosis in patients with non-alcoholic fatty liver disease and cardiometabolic comorbidities (MANPOWER study).

BMJ Open Gastroenterol 2020 13;7(1):e000341. Epub 2020 Jan 13.

Medical Affairs, Sanofi-Aventis SA, Moscow, Russian Federation.

Objective: The concept of using naturally occurring compounds such as polyenylphosphatidylcholine (PPC) as an adjunctive therapy to treat non-alcoholic fatty liver disease (NAFLD) and alleviate or reverse hepatic steatosis appears a very attractive option for liver protection. We aim to evaluate if PPC adjunctive therapy can effectively improve the ultrasonographic features of NAFLD in routine clinical practice in Russian patients with cardiometabolic comorbidities.

Design: This 24-week, observational, prospective study was carried out in 174 medical sites across 6 federal districts of Russia. A total of 2843 adult patients with newly diagnosed NAFLD, who had a least one of four comorbidities, namely overweight/obesity, hypertension, type 2 diabetes mellitus and hypercholesterolaemia, and who received PPC as an adjunctive treatment to standard care, were enrolled. The assessment of liver ultrasonography was qualitative.

Results: Overall, 2263 (79.6%) patients had at least two metabolic comorbidities associated with NAFLD, and overweight/obesity was the most common comorbidity reported in 2298 (80.8%) patients. Almost all study participants (2837/2843; 99.8%) were prescribed 1.8 g of PPC administered three times daily. At baseline, the most frequently identified abnormalities on ultrasound were liver hyperechogenicity (84.0% of patients) and heterogeneous liver structure (62.9%). At 24 weeks, a significant (p<0.05) improvement in liver echogenicity and in liver structure was observed in 1932/2827 (68.3%) patients (95% CI 66.6% to 70.1%) and in 1207/2827 (42.7%) patients (95% CI 40.9% to 44.5%), respectively. The analysis of ultrasonographic signs by number of comorbidities revealed similar findings-liver echogenicity improved in 67.2%-69.3% and liver structure in 35.6%-45.3% of patients depending on the number of comorbidities.

Conclusion: This study showed that PPC adjunctive therapy may be useful in improving the ultrasonographic features of NAFLD in patients with associated cardiometabolic comorbidities. It also supports evidence regarding the role of PPC in the complex management of NAFLD.
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http://dx.doi.org/10.1136/bmjgast-2019-000341DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7011021PMC
November 2021

Real-world comorbidities and treatment patterns among patients with non-alcoholic fatty liver disease receiving phosphatidylcholine as adjunctive therapy in Russia.

BMJ Open Gastroenterol 2019 18;6(1):e000307. Epub 2019 Aug 18.

Medical Affairs, Sanofi-Aventis SA, Moscow, Russia.

Objective: Previous research conducted in Russia showed that the number of patients with non-alcoholic fatty liver disease (NAFLD) and associated metabolic comorbidities is large. We conducted an observational study to describe the management of NAFLD in patients with metabolic syndrome in Russia.

Design: A total of 2843 adult patients from 174 medical sites across 6 federal districts of Russia with newly diagnosed NAFLD, who had at least one of four comorbidities, namely overweight/obesity, hypertension, type 2 diabetes mellitus, and hypercholesterolaemia, and who received phosphatidylcholine (PPC) as an adjunctive treatment to standard care, were enrolled during 2015-2016.

Results: Overall, 2263 patients (79.6%) had at least two metabolic comorbidities associated with NAFLD; overweight/obesity was the most common comorbidity reported in 2298 patients (80.8%). Simple steatosis was the most frequently identified clinical form of NAFLD, diagnosed in 2128 patients (74.9%). Among hypertensive patients, ACE inhibitors, statins, and sartans were most commonly prescribed. Biguanides were administered in more than half of diabetic patients. In patients with overweight/obesity and hypercholesterolaemia, statins were the most frequently prescribed medications. Almost all patients (2837/2843; 99.8%) were treated with 1.8 g of PPC three times per day. PPC therapy was associated with a 90.5% 6-month compliance rate, high treatment satisfaction, and a favourable safety profile. However, almost 15% of diabetic patients and 40% of overweight/obese patients received no further treatment.

Conclusions: In Russia, patients with newly diagnosed NAFLD represent a population heavily burdened by comorbidities, mainly overweight/obesity and hypercholesterolaemia. A significant part of these patients did not receive a comprehensive pharmacotherapy, highlighting the existing unmet need in the current management of NAFLD patients with metabolic syndrome in Russia.
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http://dx.doi.org/10.1136/bmjgast-2019-000307DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6711446PMC
August 2019

Platelet count to spleen diameter ratio non-invasively identifies severe fibrosis and cirrhosis in patients with autoimmune hepatitis.

J Gastroenterol Hepatol 2016 Dec;31(12):1956-1962

Department of Hepatology, V.H. Vasilenko Clinic of Internal Diseases, Gastroenterology and Hepatology, First Moscow State Medical University n.a. I.M. Sechenov, Moscow, Russian Federation.

Background And Aim: Non-invasive markers are essential to assess the progression of chronic liver diseases to fibrosis/ cirrhosis and the effectiveness of therapeutic strategies. The aim of this study was to evaluate the ability of non-invasive markers to identify significant fibrosis, severe fibrosis, and cirrhosis in patients with autoimmune hepatitis (AIH).

Methods: Seventy-six patients with AIH were enrolled in the study and analyzed for the following parameters of liver fibrosis: Fibrosis 4 score (FIB-4), aspartate aminotransferase (AST) to alanine aminotransferase (ALT) ratio (AAR), AST to platelet count ratio (APRI), and platelet count to spleen diameter (PC/SD) ratio. All patients underwent liver biopsy. The diagnostic accuracy of tests was evaluated by the area under the receiver operating characteristic curve (AUROC).

Results: Among the 76 AIH patients, 55 (72.3%) had significant fibrosis (≥ F2), 37 (48.7%) had severe fibrosis (≥ F3), and 29 (38.2%) had cirrhosis (F4). PC/SD ratio (AUROC = 0.840) was superior to AAR (AUROC = 0.756), FIB-4 (AUROC = 0.702), and APRI (AUROC = 0.626) in discriminating between mild and significant fibrosis (≥ F2). The AUROCs of PC/SD ratio, FIB-4, AAR, and APRI were 0.884, 0.742, 0.731, and 0.707, respectively, for severe fibrosis (≥ F3); 0.968, 0.795, 0.744, and 0.723, respectively, for cirrhosis (F4). PC/SD ratio correctly identified 85.1% of patients with severe fibrosis, and 89.6% of patients with cirrhosis.

Conclusions: PC/SD ratio proved to be a simple non-invasive tool to correctly identify AIH patients with severe fibrosis and cirrhosis, thereby reducing the need for a liver biopsy in these patients.
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http://dx.doi.org/10.1111/jgh.13407DOI Listing
December 2016

Anti-HIV therapy with AZT prodrugs: AZT phosphonate derivatives, current state and prospects.

Expert Opin Drug Metab Toxicol 2010 Jun;6(6):701-14

Engelhardt Institute of Molecular Biology, Vavilov 32, Moscow, Russian Federation.

Importance Of The Field: AIDS, a disease caused by human immunodeficiency virus, was called 'plague of the twentieth century'. 3'-Azido-3'-deoxythymidine (AZT), the first compound approved for the treatment of HIV, is still a mandatory component of treatment schemes. However, its toxicity stimulated a search for new agents.

Areas Covered In This Review: This review presents the history and current state of the design of AZT prodrugs based on its phosphonate derivatives.

What The Reader Will Gain: Although every effort was made to include as many AZT structures bearing phosphonate residues and demonstrate the variety they offer, we also concentrated on the studies performed in our laboratory. Special attention was also paid to AZT 5'-H-phosphonate (phosphazide, Nikavir) approved in the Russian Federation as a drug for the prevention and treatment of HIV infection.

Take Home Message: The prodrug strategy applied to AZT phosphonate derivatives enriched chemistry, biology and medicine not only with new knowledge, methods and structures, but also with a new anti-HIV drug Nikavir. Currently, study of another phosphonate, AZT 5'-aminocarbonylphosphonate, is underway. Slow release of AZT following oral administration and penetration into cells, decreased toxicity and the lack of cumulative properties make the compounds of this group promising as extended-release forms of AZT.
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http://dx.doi.org/10.1517/17425251003713501DOI Listing
June 2010

5'-aminocarbonyl phosphonates as new zidovudine depot forms: antiviral properties, intracellular transformations, and pharmacokinetic parameters.

Drug Metab Dispos 2009 Mar 23;37(3):494-501. Epub 2008 Dec 23.

Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow, Russia.

The main disadvantages of 3'-azido-3'-deoxythymidine (zidovudine, AZT), the most common anti-HIV drug, are toxicity and a short half-life in the organism. The introduction of an H-phosphonate group into the AZT 5' position resulted in significant improvement of its therapeutic properties and allowed a new anti-HIV drug, Nikavir (AZT H-phosphonate). In this work, we described a new group of AZT derivatives, namely, AZT 5'-aminocarbonylphosphonates. The synthesized compounds displayed antiviral properties in cell cultures infected with HIV-1 and the capacity to release the active nucleoside in animals (rabbits and dogs) in a dose-dependent manner. The compounds were less toxic in MT-4 and HL-60 cell cultures and experimental animals compared with AZT. Major metabolites found in MT-4 cells after their incubation with AZT 5'-aminocarbonylphosphonate 1 were AZT and AZT 5'-phosphate (25 and 55%, respectively). Among the tested compounds, phosphonate 1 was the most effective AZT donor, and its longest t(1/2) and T(max) values in the line phosphonate 1--AZT H-phosphonate--AZT imply that compound 1 is an extended depot form of AZT. Although bioavailability of AZT after oral administration of phosphonate 1 was lower than those of AZT H-phosphonate and AZT (8 against 14 and 49%), we expect that this reduction would not cause essential decrease of antiviral activity but noticeably decrease toxicity as a result of gradual accumulation of AZT in blood and the absence of sharp difference between C(max) and C(min). Such a combination of properties makes the compounds of this group promising for further studies as extended-release forms of AZT.
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http://dx.doi.org/10.1124/dmd.108.022269DOI Listing
March 2009

The human vomeronasal type-1 receptor family--detection of volatiles and cAMP signaling in HeLa/Olf cells.

FASEB J 2008 May 20;22(5):1416-25. Epub 2007 Dec 20.

Department of Molecular Genetics, German Institute of Human Nutrition Potsdam-Rehbruecke, Arthur-Scheunert-Allee 114-116, 14558 Nuthetal, Germany.

The human genome harbors 5 remnant genes coding for vomeronasal type-1 receptors, compared with 187 of such receptors in mice. In rodents, vomeronasal type-1 receptors are typically expressed in the vomeronasal organ. They are believed to be highly selective and sensitive pheromone detectors, as may be inferred from one receptor, V1rb2, responding to picomolar concentrations of the mouse pheromone 2-heptanone. Expression patterns, ligands, and signal transduction of human vomeronasal type-1 receptors have, however, remained largely obscure. Our aim was to deorphan and functionally characterize these 5 putative human pheromone receptors. Here, we report functional expression for all 5 receptors in HeLa/Olf cells. The recombinant, N-terminally tagged receptors expressed at the plasma membrane of HeLa/Olf cells and responded differentially to 19 of 140 odorants in a combinatorial way. C9-C10 aliphatic alcohols or aldehydes emerged as the best agonists at submicromolar concentrations above human odorant thresholds. Surprisingly, and in contrast to mouse V1rb2, all human vomeronasal type-1 receptors activated cAMP signaling via G protein alphaolf, when expressed in HeLa/Olf cells. While a biological function of human vomeronasal type-1 receptors is still elusive, our data show that their major functional characteristics are similar to those of odorant receptors.
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http://dx.doi.org/10.1096/fj.07-9233comDOI Listing
May 2008

5'-carbamoylphosphonyl-[6-3H]-AZT as a tool for studying metabolic transformations of the nonradioactive counterpart, an inhibitor of HIV replication.

Nucleosides Nucleotides Nucleic Acids 2007 ;26(8-9):897-900

Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow, Russia.

An effective synthesis of 5'-carbamoylphosphonyl-[6-3H]-AZT was developed from [6-3H]-AZT. For the synthesized compound, chemical and enzymatic stability were determined and its penetration across HL-60 cell membranes was studied.
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http://dx.doi.org/10.1080/15257770701505899DOI Listing
March 2008

Structural determinants of odorant recognition by the human olfactory receptors OR1A1 and OR1A2.

J Struct Biol 2007 Sep 25;159(3):400-12. Epub 2007 May 25.

German Institute of Human Nutrition, Potsdam-Rehbruecke, Department of Molecular Genetics, Arthur-Scheunert-Allee 114-116, 14558 Nuthetal, Germany.

An interaction of odorants with olfactory receptors is thought to be the initial step in odorant detection. However, ligands have been reported for only 6 out of 380 human olfactory receptors, with their structural determinants of odorant recognition just beginning to emerge. Guided by the notion that amino acid positions that interact with specific odorants would be conserved in orthologs, but variable in paralogs, and based on the prediction of a set of 22 of such amino acid positions, we have combined site-directed mutagenesis, rhodopsin-based homology modelling, and functional expression in HeLa/Olf cells of receptors OR1A1 and OR1A2. We found that (i) their odorant profiles are centred around citronellic terpenoid structures, (ii) two evolutionary conserved amino acid residues in transmembrane domain 3 are necessary for the responsiveness of OR1A1 and the mouse ortholog Olfr43 to (S)-(-)-citronellol, (iii) changes at these two positions are sufficient to account for the differential (S)-(-)-citronellol responsiveness of the paralogs OR1A1 and OR1A2, and (iv) the interaction sites for (S)-(-)-citronellal and (S)-(-)-citronellol differ in both human receptors. Our results show that the orientation of odorants within a homology modelling-derived binding pocket of olfactory receptor orthologs is defined by evolutionary conserved amino acid positions.
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http://dx.doi.org/10.1016/j.jsb.2007.04.013DOI Listing
September 2007

Synthesis of novel alkyl triphosphates and their substrate properties toward terminal deoxynucleotidyltransferase.

Nucleosides Nucleotides Nucleic Acids 2007 ;26(4):323-34

Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow, Russian Federation.

Novel triphosphate derivatives bearing bulky or small groups at alpha-position attached to the triphosphate residue through linkers of different structures and lengths were synthesized and studied as substrates toward terminal deoxynucleotidyltransferase. The substrate efficacy depends on the structure of substituents, linker length, and nature of metal activator. The replacement of hydrophobic groups by small substituents decreased the substrate efficacy by about 20 times in respect to hydrophobic residues. The dependence on metal activator is the following: Co(2+) > Mn(2+) >> Mg(2+). The model of interaction of alkyl triphosphates with linkers of different lengths bearing TdT active site is presented.
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http://dx.doi.org/10.1080/15257770701296713DOI Listing
July 2007

AZT 5'-Cholinephosphate as an anti-HIV agent: the study of biochemical properties and metabolic transformations using its 32P-labelled counterpart.

Nucleosides Nucleotides Nucleic Acids 2007 ;26(1):23-36

Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow, Russian Federation.

Biochemical and metabolic transformations of 3'-azido-3'-deoxythymidine 5'-choline phosphate (1) were studied using its 32P-labelled counterpart for the evaluation of possible reasons for its enhanced anti-HIV activity. An effective synthesis of 32P-labelled 1 with a specific activity >1,000 Ci/mmol was developed by esterification of 32P-phosphoric acid with choline in the presence of BrCN followed by the coupling of the resulting choline phosphate with 3'-azido-3'-deoxythymidine (AZT). Chemical and enzymatic stabilities of 1 as well as the dynamics of penetration through HL-60 cell membranes were studied at the concentrations comparable to its antiviral concentrations. The products of intracellular transformations of the studied nucleotide were identified.
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http://dx.doi.org/10.1080/15257770601052257DOI Listing
April 2007

Podocin and MEC-2 bind cholesterol to regulate the activity of associated ion channels.

Proc Natl Acad Sci U S A 2006 Nov 1;103(46):17079-86. Epub 2006 Nov 1.

Renal Division, University Hospital Freiburg, 79106 Freiburg, Germany.

The prohibitin (PHB)-domain proteins are membrane proteins that regulate a variety of biological activities, including mechanosensation, osmotic homeostasis, and cell signaling, although the mechanism of this regulation is unknown. We have studied two members of this large protein family, MEC-2, which is needed for touch sensitivity in Caenorhabditis elegans, and Podocin, a protein involved in the function of the filtration barrier in the mammalian kidney, and find that both proteins bind cholesterol. This binding requires the PHB domain (including palmitoylation sites within it) and part of the N-terminally adjacent hydrophobic domain that attaches the proteins to the inner leaflet of the plasma membrane. By binding to MEC-2 and Podocin, cholesterol associates with ion-channel complexes to which these proteins bind: DEG/ENaC channels for MEC-2 and TRPC channels for Podocin. Both the MEC-2-dependent activation of mechanosensation and the Podocin-dependent activation of TRPC channels require cholesterol. Thus, MEC-2, Podocin, and probably many other PHB-domain proteins by binding to themselves, cholesterol, and target proteins regulate the formation and function of large protein-cholesterol supercomplexes in the plasma membrane.
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http://dx.doi.org/10.1073/pnas.0607465103DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1859892PMC
November 2006

Identification of specific ligands for orphan olfactory receptors. G protein-dependent agonism and antagonism of odorants.

J Biol Chem 2005 Mar 14;280(12):11807-15. Epub 2004 Dec 14.

Department of Molecular Genetics, German Institute of Human Nutrition Potsdam-Rehbruecke, Arthur-Scheunert-Allee 114-116, 14558 Nuthetal, Germany.

Olfactory receptors are the largest group of orphan G protein-coupled receptors with an infinitely small number of agonists identified out of thousands of odorants. The de-orphaning of olfactory receptor (OR) is complicated by its combinatorial odorant coding and thus requires large scale odorant and receptor screening and establishing receptor-specific odorant profiles. Here, we report on the stable reconstitution of OR-specific signaling in HeLa/Olf cells via G protein alphaolf and adenylyl cyclase type-III to the Ca2+ influx-mediating olfactory cyclic nucleotide-gated CNGA2 channel. We demonstrate the central role of Galphaolf in odorant-specific signaling out of OR. The employment of the non-typical G protein alpha15 dramatically altered the odorant specificities of 3 of 7 receptors that had been characterized previously by different groups. We further show for two OR that an odorant may be an agonist or antagonist, depending on the G protein used. HeLa/Olf cells proved suitable for high-throughput screening in fluorescence-imaging plate reader experiments, resulting in the de-orphaning of two new OR for the odorant (-)citronellal from an expression library of 93 receptors. To demonstrate the G protein dependence of its odorant response pattern, we screened the most sensitive (-)citronellal receptor Olfr43 versus 94 odorants simultaneously in the presence of Galpha15 or Galphaolf. We finally established an EC50-ranking odorant profile for Olfr43 in HeLa/Olf cells. In summary, we conclude that, in heterologous systems, odorants may function as agonists or antagonists, depending on the G protein used. HeLa/Olf cells provide an olfactory model system for functional expression and de-orphaning of OR.
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http://dx.doi.org/10.1074/jbc.M411508200DOI Listing
March 2005

Intracellular metabolism and pharmacokinetics of 5'-hydrogenphosphonate of 3'-azido-2',3'-dideoxythymidine, a prodrug of 3'-azido-2',3'-dideoxythymidine.

Antiviral Res 2004 Aug;63(2):107-13

Engelhardt Institute of Molecular Biology, RAS, 32 Vavilov St., 119991 Moscow, Russian Federation, Russia.

5'-Hydrogenphosphonate of 3'-azido-2',3'-dideoxythymidine (HpAZT), a novel anti-HIV drug approved for the treatment of HIV-infected patients in Russia, displays some clinical advantages over azidothymidine (AZT). Metabolism in the HL-60 cell culture and pharmacokinetics in mice of [6-3H]-HpAZT (in comparison with [6-3H-AZT) were studied to elucidate the metabolic basis of its lower clinical toxicity. Accumulation of [6-3H]-HpAZT-derived products in cells with time, distribution of its radioactive metabolites among blood and different mouse organs and dependence of drug accumulation on the route of administration were investigated. The rate of accumulation of [3H]-HpAZT metabolites in cells was slower than the rate of accumulation of [3H]-AZT metabolites. [3H]-AZTMP was the dominating metabolite at all time points, achieving the level of 15 +/- 3 pmol/10(6) cells after 25 h incubation. After oral or intravenous administrations of [3H]-HpAZT, the (radioactive) metabolites were rapidly distributed among blood, stomach, intestine and liver and were not found in brain, muscles and spleen. [3H]-HpAZT underwent rapid and extensive metabolism, [3H]-AZT being the dominating product at all time points. Administration of 180 nmol of [3H]-HpAZT resulted in an AZT concentration in blood of 1-3 microM after 5 min, which remained practically constant during the next 25 min and did not depend on the route of administration.
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http://dx.doi.org/10.1016/j.antiviral.2004.03.001DOI Listing
August 2004

Uncharged AZT and D4T derivatives of phosphonoformic and phosphonoacetic acids as anti-HIV pronucleosides.

J Med Chem 2004 Jul;47(14):3606-14

Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, 32 Vavilov str., Moscow 119991, Russian Federation.

Two series of new lipophilic phosphonoformate and phosphonoacetate derivatives of AZT and d4T were synthesized and evaluated as anti-HIV agents. The efficacy of some of the synthesized compounds in cell cultures infected with HIV-1 was higher than that of the parent nucleosides and only slightly correlated to their stability in the phosphate buffer and human blood serum. The synthesized phosphonates are most probably prodrug forms of the corresponding nucleosides.
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http://dx.doi.org/10.1021/jm0310176DOI Listing
July 2004

New lipophilic derivatives of AZT and d4T 5'-phosphonates.

Nucleosides Nucleotides Nucleic Acids 2003 May-Aug;22(5-8):981-5

Engelhardt Institute of Molecular Biology Russian Academy of Sciences, Moscow, Russia.

5'-Aminocarbonylphosphonyl and aminocarbonylmethylphosphonyl diesters of AZT and d4T were synthesized as potential anti-HIV agents.
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http://dx.doi.org/10.1081/NCN-120022718DOI Listing
December 2003

Carbocyclic dinucleoside polyphosphonates: interaction with HIV reverse transcriptase and antiviral activity.

J Med Chem 2002 Mar;45(6):1284-91

Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow, Russia.

Carbocyclic alpha, gamma-bis(nucleoside)-5,5'-triphosphonates and alpha, delta-bis(nucleoside)-5,5'-tetraphosphonates (Ap4A and Gp4G) analogues were shown to be a new type of terminating substrate of HIV reverse transcriptase. They effectively inhibited the DNA synthesis catalyzed by this enzyme in model cell-free systems, but their antiviral activity both in Rat1 fibroblast cell culture bearing MLV reverse transcriptase and in HIV-infected MT-4 cells was low. When a liposome delivery system was used, the antiviral efficacy of the compounds under study was increased.
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http://dx.doi.org/10.1021/jm011011lDOI Listing
March 2002
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