- Repair of UV-Induced DNA Damage Independent of Nucleotide Excision Repair Is Masked by MUTYH. [Journal Article]
- MCMol Cell 2017 Nov 16; 68(4):797-807.e7
- DNA lesions caused by UV damage are thought to be repaired solely by the nucleotide excision repair (NER) pathway in human cells. Patients carrying mutations within genes functioning in this pathway ...
DNA lesions caused by UV damage are thought to be repaired solely by the nucleotide excision repair (NER) pathway in human cells. Patients carrying mutations within genes functioning in this pathway display a range of pathologies, including an increased susceptibility to cancer, premature aging, and neurological defects. There are currently no curative therapies available. Here we performed a high-throughput chemical screen for agents that could alleviate the cellular sensitivity of NER-deficient cells to UV-induced DNA damage. This led to the identification of the clinically approved anti-diabetic drug acetohexamide, which promoted clearance of UV-induced DNA damage without the accumulation of chromosomal aberrations, hence promoting cellular survival. Acetohexamide exerted this protective function by antagonizing expression of the DNA glycosylase, MUTYH. Together, our data reveal the existence of an NER-independent mechanism to remove UV-induced DNA damage and prevent cell death.
- Study on the interactions of sulfonylurea antidiabetic drugs with normal and glycated human serum albumin by capillary electrophoresis-frontal analysis. [Journal Article]
- JSJ Sep Sci 2016; 39(18):3631-7
- Diabetes is one of the most widespread diseases characterized by a deficiency in the production of insulin or its ineffectiveness. As a result, the increased concentrations of glucose in the blood le...
Diabetes is one of the most widespread diseases characterized by a deficiency in the production of insulin or its ineffectiveness. As a result, the increased concentrations of glucose in the blood lead not only to damage to many of the body's systems but also cause the nonenzymatic glycation of plasma proteins affecting their drug binding. Since the binding ability influences its pharmacokinetics and pharmacodynamics, this is a very important issue in the development of new drugs and personalized medicine. In this study, capillary electrophoresis-frontal analysis was used to evaluate the affinities between human serum albumin or its glycated form and the first generation of sulfonylurea antidiabetics, since their inadequate concentration may induce hypoglycaemia or on the contrary hyperglycaemia. The binding constants decrease in the sequence acetohexamide > tolbutamide > chlorpropamide > carbutamide both for normal and glycated human serum albumins, with glycated giving lower values. These results provide a more quantitative picture of how these drugs bind with normal and modified human serum albumin and indicate capillary electrophoresis-frontal analysis to be another tool for examining the changes arising from modifications of albumin, or any other protein, with all its benefits like short analysis time, small sample requirement, and automation.
- Analysis of free drug fractions in human serum by ultrafast affinity extraction and two-dimensional affinity chromatography. [Journal Article]
- ABAnal Bioanal Chem 2016; 408(1):131-40
- Ultrafast affinity extraction and a two-dimensional high performance affinity chromatographic system were used to measure the free fractions for various drugs in serum and at typical therapeutic conc...
Ultrafast affinity extraction and a two-dimensional high performance affinity chromatographic system were used to measure the free fractions for various drugs in serum and at typical therapeutic concentrations. Pooled samples of normal serum or serum from diabetic patients were utilized in this work. Several drug models (i.e., quinidine, diazepam, gliclazide, tolbutamide, and acetohexamide) were examined that represented a relatively wide range of therapeutic concentrations and affinities for human serum albumin (HSA). The two-dimensional system consisted of an HSA microcolumn for the extraction of a free drug fraction, followed by a larger HSA analytical column for the further separation and measurement of this fraction. Factors that were optimized in this method included the flow rates, column sizes, and column switching times that were employed. The final extraction times used for isolating the free drug fractions were 333-665 ms or less. The dissociation rate constants for several of the drugs with soluble HSA were measured during system optimization, giving results that agreed with reference values. In the final system, free drug fractions in the range of 0.7-9.5% were measured and gave good agreement with values that were determined by ultrafiltration. Association equilibrium constants or global affinities were also estimated by this approach for the drugs with soluble HSA. The results for the two-dimensional system were obtained in 5-10 min or less and required only 1-5 μL of serum per injection. The same approach could be adapted for work with other drugs and proteins in clinical samples or for biomedical research.
- Molecular and biochemical characterisation of human short-chain dehydrogenase/reductase member 3 (DHRS3). [Journal Article]
- CBChem Biol Interact 2015 Jun 5; 234:178-87
- Dehydrogenase/reductase (SDR family) member 3 (DHRS3), also known as retinal short-chain dehydrogenase/reductase (retSDR1) is a member of SDR16C family. This family is thought to be NADP(H) dependent...
Dehydrogenase/reductase (SDR family) member 3 (DHRS3), also known as retinal short-chain dehydrogenase/reductase (retSDR1) is a member of SDR16C family. This family is thought to be NADP(H) dependent and to have multiple substrates; however, to date, only all-trans-retinal has been identified as a DHRS3 substrate. The reductive reaction catalysed by DHRS3 seems to be physiological, and recent studies proved the importance of DHRS3 for maintaining suitable retinoic acid levels during embryonic development in vivo. Although it seems that DHRS3 is an important protein, knowledge of the protein and its properties is quite limited, with the majority of information being more than 15 years old. This study aimed to generate a more comprehensive characterisation of the DHRS3 protein. Recombinant enzyme was prepared and demonstrated to be a microsomal, integral-membrane protein with the C-terminus oriented towards the cytosol, consistent with its preference of NADPH as a cofactor. It was determined that DHRS3 also participates in the metabolism of other endogenous compounds, such as androstenedione, estrone, and DL-glyceraldehyde, and in the biotransformation of xenobiotics (e.g., NNK and acetohexamide) in addition to all-trans-retinal. Purified and reconstituted enzyme was prepared for the first time and will be used for further studies. Expression of DHRS3 was shown at the level of both mRNA and protein in the human liver, testis and small intestine. This new information could open other areas of DHRS3 protein research.
- Computer aided screening of secreted frizzled-related protein 4 (SFRP4): a potential control for diabetes mellitus. [Journal Article]
- MMolecules 2014; 19(7):10129-36
- Diabetes mellitus is a life threatening disease and scientists are doing their best to find a cost effective and permanent treatment of this malady. The recent trend is to control the disease by targ...
Diabetes mellitus is a life threatening disease and scientists are doing their best to find a cost effective and permanent treatment of this malady. The recent trend is to control the disease by target base inhibiting of enzymes or proteins. Secreted frizzled-related protein 4 (SFRP4) is found to cause five times more risk of diabetes when expressed above average levels. This study was therefore designed to analyze the SFRP4 and to find its potential inhibitors. SFRP4 was analyzed by bio-informatics tools of sequence tool and structure tool. A total of three potential inhibitors of SFRP4 were found, namely cyclothiazide, clopamide and perindopril. These inhibitors showed significant interactions with SFRP4 as compared to other inhibitors as well as control (acetohexamide). The findings suggest the possible treatment of diabetes mellitus type 2 by inhibiting the SFRP4 using the inhibitors cyclothiazide, clopamide and perindopril.
- Reductive metabolism of nabumetone by human liver microsomal and cytosolic fractions: exploratory prediction using inhibitors and substrates as marker probes. [Journal Article]
- EJEur J Drug Metab Pharmacokinet 2015; 40(2):127-35
- The metabolic reduction of nabumetone was examined by inhibition and correlation studies using human liver microsomes and cytosol. This reduction was observed in both fractions, with the V(max) value...
The metabolic reduction of nabumetone was examined by inhibition and correlation studies using human liver microsomes and cytosol. This reduction was observed in both fractions, with the V(max) values for reduction activity being approximately fourfold higher, and the V(max)/K(m) values approximately three-fold higher, in the microsomes than in the cytosol. The reduction of nabumetone was inhibited by 18β-glycyrrhetinic acid, an 11β-hydroxysteroid dehydrogenase (11β-HSD) inhibitor, in the microsomal fraction. The reduction activity was also inhibited by quercetin and menadione [carbonyl reductase (CBR) inhibitors], and by phenolphthalein and medroxyprogesterone acetate [potent inhibitors of aldo-keto reductase (AKR) 1C1, 1C2 and 1C4] in the cytosol. A good correlation (r² = 0.93) was observed between the reduction of nabumetone and of cortisone, as a marker of 11β-HSD activity, in the microsomal fractions. There was also an excellent relationship between reduction of nabumetone and of the AKR1C substrates, acetohexamide, and ethacrynic acid (r 2 = 0.92 and 0.93, respectively), in the cytosol fractions. However, a poor correlation was observed between the formation of 4-(6-methoxy-2-naphthyl)-butan-2-ol (MNBO) from nabumetone and CBR activity (with 4-benzoyl pyridine reduction as a CBR substrate) in the cytosol fractions (r² = 0.24). These findings indicate that nabumetone may be metabolized by 11β-HSD in human liver microsomes, and primarily by AKR1C4 in human liver cytosol, although multiple enzymes in the AKR1C subfamily may be involved. It cannot be completely denied that CBR is involved to some extent in the formation of MNBO from nabumetone in the cytosol fraction.
- Effects of Fatty Acids and Glycation on Drug Interactions with Human Serum Albumin. [Journal Article]
- CMCurr Metabolomics 2013 Sep 01; 1(3):239-250
- The presence of elevated glucose concentrations in diabetes is a metabolic change that leads to an increase in the amount of non-enzymatic glycation that occurs for serum proteins. One protein that i...
The presence of elevated glucose concentrations in diabetes is a metabolic change that leads to an increase in the amount of non-enzymatic glycation that occurs for serum proteins. One protein that is affected by this process is the main serum protein, human serum albumin (HSA), which is also an important carrier agent for many drugs and fatty acids in the circulatory system. Sulfonylureas drugs, used to treat type 2 diabetes, are known to have significant binding to HSA. This study employed ultrafiltration and high-performance affinity chromatography to examine the effects of HSA glycation on the interactions of several sulfonylurea drugs (i.e., acetohexamide, tolbutamide and gliclazide) with fatty acids, whose concentrations in serum are also affected by diabetes. Similar overall changes in binding were noted for these drugs with normal HSA or glycated HSA and in the presence of the fatty acids. For most of the tested drugs, the addition of physiological levels of the fatty acids to normal HSA and glycated HSA produced weaker binding. At low fatty acid concentrations, many of these systems followed a direct competition model while others involved a mixed-mode interaction. In some cases, there was a change in the interaction mechanism between normal HSA and glycated HSA, as seen with linoleic acid. Systems with only direct competition also gave notable changes in the affinities of fatty acids at their sites of drug competition when comparing normal HSA and glycated HSA. This research demonstrated the importance of considering how changes in the concentrations and types of metabolites (e.g., in this case, glucose and fatty acids) can alter the function of a protein such as HSA and its ability to interact with drugs or other agents.
- Crystallization of a new polymorph of acetohexamide from 2-hydroxybutyl-β-cyclodextrin solution: form VI with a high aqueous solubility. [Journal Article]
- IJInt J Pharm 2013 Sep 10; 453(2):315-21
- A new polymorph of acetohexamide (Form VI) was prepared via the formation of a complex with 2-hydoxybutyl-β-cyclodextrin (HB-β-CD) in aqueous solution. An alkaline solution of acetohexamide and HB-β-...
A new polymorph of acetohexamide (Form VI) was prepared via the formation of a complex with 2-hydoxybutyl-β-cyclodextrin (HB-β-CD) in aqueous solution. An alkaline solution of acetohexamide and HB-β-CD was adjusted to pH 4.0 by titration with hydrochloric acid. The resulting opaque solution was filtered through paper and allowed to stand at 4°C for 24h. The resulting precipitate was isolated on a filter and analyzed for polymorph content by powder X-ray diffractometry and thermal analysis. The diffraction pattern and thermal behavior of the precipitate was different from those of previously reported acetohexamide polymorphs (Forms I, III, IV and V), indicating that a new polymorph of the drug, i.e. Form VI was produced. This new polymorph was fairly stable against conversion to a stable form even at accelerated storage conditions. Crystalline Form VI was highly soluble in water and dissolved more rapidly than the other known polymorphs. This property was reflected in the blood concentrations of the drug after oral administration to rats.
- In vitro inhibition of breast cancer spheroid-induced lymphendothelial defects resembling intravasation into the lymphatic vasculature by acetohexamide, isoxsuprine, nifedipin and proadifen. [Journal Article]
- BJBr J Cancer 2013 Feb 19; 108(3):570-8
- CONCLUSIONS: The targeting of different mechanisms was most likely the reason for synergistic effects of different drug combinations on the inhibition of CCID formation. Furthermore, the treatment with drug combinations allowed also a several-fold reduction in drug concentrations. These results encourage further screening of approved drugs and their in vivo testing.
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- Epac2: a sulfonylurea receptor? [Journal Article]
- BSBiochem Soc Trans 2012; 40(1):6-10
- Sulfonylureas are widely used oral drugs in the treatment of diabetes mellitus. They function by the inhibition of ATP-sensitive K+ channels in pancreatic β-cells, which are thus considered the 'clas...
Sulfonylureas are widely used oral drugs in the treatment of diabetes mellitus. They function by the inhibition of ATP-sensitive K+ channels in pancreatic β-cells, which are thus considered the 'classical' sulfonylurea receptor. Next to the ATP-sensitive K+ channels, additional sulfonylurea-interacting proteins were identified, which might contribute to the physiological effects of this drug family. Most recently, Epac2 (exchange protein directly activated by cAMP 2) was added to the list of sulfonylurea receptors. However, this finding caused controversy in the literature. The critical discussion of the present paper comes to the conclusion that sulfonylureas are not able to activate Epac2 directly and are unlikely to bind to Epac2. Increased blood glucose levels after food intake result in the secretion of insulin from pancreatic β-cells. Glucose levels are detected 'indirectly' by β-cells: owing to increased glycolysis rates, the ratio of cellular ATP/ADP increases and causes the closure of ATP-sensitive K+ channels. In consequence, cells depolarize and voltage-dependent Ca2+ channels open to cause an increase in the cellular Ca2+ concentration. Finally, Ca2+ induces the fusion of insulin-containing granules with the plasma membrane. Sulfonylureas, such as tolbutamide, glibenclamide or acetohexamide, form a class of orally applicable drugs used in the treatment of non-insulin-dependent diabetes mellitus.