| 61. |
- Ljunggren, Stefan, 1988-
(författare)
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Lipoproteomics : Environmental and Genetic Factors Affecting High-Density Lipoprotein (HDL)
- 2016
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Lipoprotein particles act as lipid transporters in the blood stream, and measuring cholesterol content in specific subclasses of lipoprotein particles has long been, and still is, a frequently used tool to estimate the risk of cardiovascular disease (CVD). High-density lipoprotein (HDL) is a subclass of lipoproteins often regarded as providing protection against CVD via several functions including reverse cholesterol transport and anti-inflammatory capacities. However, the precise relationship between HDL cholesterol levels and health outcome is still unclear. Lately, new approaches to study HDL composition and function have therefore become more important.HDL function is to a large extent dependent on its proteome, containing more than 100 proteins. Investigating the proteome in individuals with altered gene expression for HDL-associated proteins or with known exposure to environmental contaminants may reveal new insights into how HDL metabolism is affected by various factors. This is of interest in order to better understand the role of HDL in CVD.Papers I and II focus on two different mutations in a structural HDL protein, apolipoprotein A-I (L202P and K131del), and one mutation in the scavenger receptor class B-1 (P297S), which is involved in selective lipid uptake of cholesterol mainly into hepatocytes and adrenal cells. The HDL proteome was analyzed using two-dimensional gel electrophoresis and mass spectrometry. The L202P mutation was identified in HDL of the heterozygote carriers together with a significant decrease of apolipoprotein E and increased zinc-alpha-2-glycoprotein. By contrast, the second apolipoprotein AI mutation (K131del) was associated with significantly elevated alpha-1-antitrypsin and transthyretin levels. Protein analyses of the scavenger receptor class B1 P297S heterozygotes showed a significant increase in HDL apoL-1 along with increased free apoE. The carriers showed no difference in antioxidative capability but a significant increase in apoA-I methionine oxidation.Papers III and IV focus on persistent organic pollutants that may influence HDL composition and function. These compounds accumulate in humans, and exposure has been linked to an increased risk of CVD. To provide a better understanding of the HDL system in relation to pollutants, a population living in a contaminated area was studied. Persistent organic pollutants in isolated HDL were quantified using high-resolution gas chromatography mass spectrometry and significantly increased levels were found in individuals with CVD as compared to healthy controls. Furthermore, there was a significant negative association between the pollutants and paraoxonase-1 anti-oxidant activity. Studying the proteome with nano-liquid chromatography tandem mass spectrometry led to the identification of 118 proteins in HDL, of which ten were significantly associated with the persistent organic pollutants.In summary, the present studies demonstrate protein pattern alterations in HDL associated with inherited genetic variants or pollutant exposure. The studies also provide a set of methods that are useful tools to further comprehend the complexity of lipoprotein metabolism and function. The results are important in order to improve our understanding of HDL in CVD and to explain an increased risk of CVD associated with exposure to organic pollutants.
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| 62. |
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| 63. |
- Magnusson, Marie, et al.
(författare)
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A placebo-controlled study of retinal blood flow changes by pentoxifylline and metabolites in humans
- 2006
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Ingår i: British Journal of Clinical Pharmacology. - : Wiley. - 0306-5251 .- 1365-2125. ; 61:2, s. 138-147
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Tidskriftsartikel (refereegranskat)abstract
- AIM: To investigate the possible effects of pentoxifylline metabolites on retinal blood flow in humans. METHODS: A randomized, placebo-controlled, four-period cross-over study that was observer blinded and partly blinded for the eight participants. On one occasion a placebo was given as an intravenous (i.v.) infusion over 100 min. On the other three occasions pentoxifylline was administered as i.v. infusions over 100 min at a rate of 3 mg min(-1). Before two of the pentoxifylline infusions the subjects were pretreated with either ciprofloxacin or rifampicin. Retinal blood flow was measured by scanning laser doppler flowmetry (SLDF) in a selected area of the central temporal retina before, during and until 5 h after the end of infusion. Blood samples for concentration analyses of pentoxifyllin, R-M1, S-M1, M4 and M5 were taken serially and areas under the curves (AUCs) were calculated. Linear mixed models were used for the statistical analyses. RESULTS: Mean AUCs (ng h ml(-1)) were significantly increased for pentoxifylline (1964 vs. 1453) and S-M1 (5804 vs. 4227), but not R-M1 when pentoxifylline was co-administered with ciprofloxacin. The mean AUC for M5 was significantly reduced when subjects were pretreated with rifampicin (2041 vs. 3080). Pentoxifylline with and without pretreatment with rifampicin significantly increased retinal blood flow assessed as mean flow, pulsation (i.e. 1-systole/diastole), and diastolic flow (but not during systole), compared with placebo. The increases over placebo were more pronounced on diastolic flow, 9.7% (95% confidence interval 4.2, 15.5) than on mean flow, 4.6% (1.1, 8.3) after pentoxifylline administration. With pentoxifylline after rifampicin pretreatment the corresponding differences were 11.7% (5.8, 17.9) and 5.1% (1.4, 7.8) over placebo, respectively. After co-administration of pentoxifylline and ciprofloxacin we saw only a nonsignificant trend towards increased flow during diastole, but a significant decrease in pulsation. When AUCs for pentoxifylline and its metabolites were used as regressor variables to retinal mean flow we found that pentoxifylline, R-M1 and M5 had coefficients with a positive sign indicating that they enhanced the retinal blood flow. In contrast, S-M1 and M4 had coefficients with negative sign and thus appeared to decrease the blood flow in subjects treated with pentoxifylline. CONCLUSION: The R-M1 and M5 metabolites of pentoxifylline contributed significantly to the effects of pentoxifylline on retinal blood flow.
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| 64. |
- Marco, M., et al.
(författare)
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Pterocarpans and isoflavones from the root bark of Millettia micans and of Millettia dura
- 2017
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Ingår i: Phytochemistry Letters. - : Elsevier BV. - 1874-3900. ; 21, s. 216-220
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Tidskriftsartikel (refereegranskat)abstract
- From the CH2Cl2/CH3OH (1:1) extract of the root bark of Millettia micans, a new pterocarpan, (6aR, 11aR)-3-hydroxy- 7,8,9-trimethoxypterocarpan (1), named micanspterocarpan, was isolated. Similar investigation of the CH2Cl2/CH3OH (1:1) extract of the root bark of Millettia dura gave a further new pterocarpan, (6aR, 11aR)-8,9-methylenedioxy-3-prenyloxypterocarpan (2), named 3-O-prenylmaackiain, along with six known isoflavones (38) and a chalcone (9). All purified compounds were identified by NMR and MS, whereas the absolute configurations of the new pterocarpans were established by chriptical data analyses including quantum chemical ECD calculation. Among the isolated constituents, calopogonium isoflavone B (3) and isoerythrin A-4'-(3-methylbut-2-enyl) ether (4) showed marginal activities against the 3D7 and the Dd2 strains of Plasmodium falciparum (70-90% inhibition at 40 mu M). Maximaisoflavone B (5) and 7,2'-dimethoxy-4', 5'-methylenedioxyisoflavone (7) were weakly cytotoxic (IC50 153.5 and 174.1 mu M, respectively) against the MDA-MB-231 human breast cancer cell line. None of the tested compounds showed in-vitro translation inhibitory activity or toxicity against the HEK-293 human embryonic kidney cell line at 40 mu M.
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| 65. |
- Matricon, Pierre, et al.
(författare)
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Structure-based virtual screening discovers potent and selective adenosine A1 receptor antagonists
- 2023
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Ingår i: European Journal of Medicinal Chemistry. - : Elsevier BV. - 0223-5234 .- 1768-3254. ; 257
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Tidskriftsartikel (refereegranskat)abstract
- Development of subtype-selective leads is essential in drug discovery campaigns targeting G protein-coupled receptors (GPCRs). Herein, a structure-based virtual screening approach to rationally design subtype-selective ligands was applied to the A1 and A2A adenosine receptors (A1R and A2AR). Crystal structures of these closely related subtypes revealed a non-conserved subpocket in the binding sites that could be exploited to identify A1R selective ligands. A library of 4.6 million compounds was screened computationally against both receptors using molecular docking and 20 A1R selective ligands were predicted. Of these, seven antagonized the A1R with micromolar activities and several compounds displayed slight selectivity for this subtype. Twenty-seven analogs of two discovered scaffolds were designed, resulting in antagonists with nanomolar potency and up to 76-fold A1R-selectivity. Our results show the potential of structure-based virtual screening to guide discovery and optimization of subtype-selective ligands, which could facilitate the development of safer drugs.
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| 66. |
- Mitra, Sanhita, et al.
(författare)
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Subcellular distribution of p53 by the p53-responsive lncRNA NBAT1 determines chemotherapeutic response in neuroblastoma.
- 2021
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Ingår i: Cancer research. - 1538-7445. ; 81:6, s. 1457-1471
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Tidskriftsartikel (refereegranskat)abstract
- Neuroblastoma has a low mutation rate for the p53 gene. Alternative ways of p53 inactivation have been proposed in neuroblastoma, such as abnormal cytoplasmic accumulation of wild-type p53. However, mechanisms leading to p53 inactivation via cytoplasmic accumulation are not well investigated. Here we show that the neuroblastoma risk-associated locus 6p22.3-derived tumor suppressor NBAT1 is a p53-responsive lncRNA that regulates p53 subcellular levels. Low expression of NBAT1 provided resistance to genotoxic drugs by promoting p53 accumulation in cytoplasm and loss from mitochondrial and nuclear compartments. Depletion of NBAT1 altered CRM1 function and contributed to the loss of p53-dependent nuclear gene expression during genotoxic drug treatment. CRM1 inhibition rescued p53-dependent nuclear functions and sensitized NBAT1-depleted cells to genotoxic drugs. Combined inhibition of CRM1 and MDM2 was even more effective in sensitizing aggressive neuroblastoma cells with p53 cytoplasmic accumulation. Thus, our mechanistic studies uncover an NBAT1-dependent CRM1/MDM2-based potential combination therapy for high-risk neuroblastoma patients.
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| 67. |
- Rehling, Daniel, et al.
(författare)
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Crystal structures of NUDT15 variants enabled by a potent inhibitor reveal the structural basis for thiopurine sensitivity
- 2021
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Ingår i: Journal of Biological Chemistry. - : Elsevier BV. - 0021-9258 .- 1083-351X. ; 296
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Tidskriftsartikel (refereegranskat)abstract
- The enzyme NUDT15 efficiently hydrolyzes the active metabolites of thiopurine drugs, which are routinely used for treating cancer and inflammatory diseases. Loss-of-function variants in NUDT15 are strongly associated with thiopurine intolerance, such as leukopenia, and preemptive NUDT15 genotyping has been clinically implemented to personalize thiopurine dosing. However, understanding the molecular consequences of these variants has been difficult, as no structural information was available for NUDT15 proteins encoded by clinically actionable pharmacogenetic variants because of their inherent instability. Recently, the small molecule NUDT15 inhibitor TH1760 has been shown to sensitize cells to thiopurines, through enhanced accumulation of 6-thio-guanine in DNA. Building upon this, we herein report the development of the potent and specific NUDT15 inhibitor, TH7755. TH7755 demonstrates a greatly improved cellular target engagement and 6-thioguanine potentiation compared with TH1760, while showing no cytotoxicity on its own. This potent inhibitor also stabilized NUDT15, enabling analysis by X-ray crystallography. We have determined high-resolution structures of the clinically relevant NUDT15 variants Arg139Cys, Arg139His, Val18Ile, and V18_V19insGlyVal. These structures provide clear insights into the structural basis for the thiopurine intolerance phenotype observed in patients carrying these pharmacogenetic variants. These findings will aid in predicting the effects of new NUDT15 sequence variations yet to be discovered in the clinic.
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| 68. |
- Rhodin, Malin M., et al.
(författare)
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Human renal function maturation : a quantitative description using weight and postmenstrual age
- 2009
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Ingår i: Pediatric nephrology (Berlin, West). - : Springer Science and Business Media LLC. - 0931-041X .- 1432-198X. ; 24:1, s. 67-76
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Tidskriftsartikel (refereegranskat)abstract
- This study pools published data to describe the increase in glomerular filtration rate (GFR) from very premature neonates to young adults. The data comprises measured GFR (using polyfructose, Cr-51-EDTA, mannitol or iohexol) from eight studies (n=923) and involved very premature neonates (22 weeks postmenstrual age) to adulthood (31 years). A nonlinear mixed effects approach (NONMEM) was used to examine the influences of size and maturation on renal function. Size was the primary covariate, and GFR was standardized for a body weight of 70 kg using an allometric power model. Postmenstrual age (PMA) was a better descriptor of maturational changes than postnatal age (PNA). A sigmoid hyperbolic model described the nonlinear relationship between GFR maturation and PMA. Assuming an allometric coefficient of 3/4, the fully mature (adult) GFR is predicted to be 121.2 mL/min per 70 kg [95% confidence interval (CI) 117-125]. Half of the adult value is reached at 47.7 post-menstrual weeks (95% CI 45.1-50.5), with a Hill coefficient of 3.40 (95% CI 3.03-3.80). At 1-year postnatal age, the GFR is predicted to be 90% of the adult GFR. Glomerular filtration rate can be predicted with a consistent relationship from early prematurity to adulthood. We propose that this offers a clinically useful definition of renal function in children and young adults that is independent of the predictable changes associated with age and size.
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| 69. |
- Senkowski, Wojciech
(författare)
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High-throughput screening using multicellular tumor spheroids to reveal and exploit tumor-specific vulnerabilities
- 2017
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- High-throughput drug screening (HTS) in live cells is often a vital part of the preclinical anticancer drug discovery process. So far, two-dimensional (2D) monolayer cell cultures have been the most prevalent model in HTS endeavors. However, 2D cell cultures often fail to recapitulate the complex microenvironments of in vivo tumors. Monolayer cultures are highly proliferative and generally do not contain quiescent cells, thought to be one of the main reasons for the anticancer therapy failure in clinic. Thus, there is a need for in vitro cellular models that would increase predictive value of preclinical research results. The utilization of more complex three-dimensional (3D) cell cultures, such as multicellular tumor spheroids (MCTS), which contain both proliferating and quiescent cells, has therefore been proposed. However, difficult handling and high costs still pose significant hurdles for application of MCTS for HTS.In this work, we aimed to develop novel assays to apply MCTS for HTS and drug evaluation. We also set out to identify cellular processes that could be targeted to selectively eradicate quiescent cancer cells. In Paper I, we developed a novel MCTS-based HTS assay and found that nutrient-deprived and hypoxic cancer cells are selectively vulnerable to treatment with inhibitors of mitochondrial oxidative phosphorylation (OXPHOS). We also identified nitazoxanide, an FDA-approved anthelmintic agent, to act as an OXPHOS inhibitor and to potentiate the effects of standard chemotherapy in vivo. Subsequently, in Paper II we applied the high-throughput gene-expression profiling method for MCTS-based drug screening. This led to discovery that quiescent cells up-regulate the mevalonate pathway upon OXPHOS inhibition and that the combination of OXPHOS inhibitors and mevalonate pathway inhibitors (statins) results in synergistic toxicity in this cell population. In Paper III, we developed a novel spheroid-based drug combination-screening platform and identified a set of molecules that synergize with nitazoxanide to eradicate quiescent cancer cells. Finally, in Paper IV, we applied our MCTS-based methods to evaluate the effects of phosphodiesterase (PDE) inhibitors in PDE3A-expressing cell lines.In summary, this work illustrates how MCTS-based HTS yields potential to reveal and exploit previously unrecognized tumor-specific vulnerabilities. It also underscores the importance of cell culture conditions in preclinical drug discovery endeavors.
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| 70. |
- Shwter, Abdrabuh N., et al.
(författare)
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Chemopreventive effect of Phaleria macrocarpa on colorectal cancer aberrant crypt foci in vivo
- 2016
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Ingår i: Journal of Ethnopharmacology. - : Elsevier BV. - 0378-8741 .- 1872-7573. ; 193, s. 195-206
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Tidskriftsartikel (refereegranskat)abstract
- Ethnopharmacological relevance: Natural products are important ingredients for pharmaceutical applications specifically new entities for treating cancer and other diseases. Phaleria macrocarpa is native of Indonesia and considered as a prolific source of bioactive substances useful for chemoprevention. Aim of the study: To investigate the chemopreventive properties of Phaleria macrocarpa on azoxymethane (AOM)-induced aberrant crypt foci (ACF) in rats. Methods: The biological activities of the ethanol extract of P. macrocarpa fruits were evaluated both in vitro and in vivo. First the extract was investigated for its in vitro antioxidant activity by the total phenolic content and ferric reducing antioxidant power assay. Then the chemopreventive effect of P. macrocarpa was performed on AOM-induced aberrant crypt foci as colorectal carcinoma model in rats. Result: the crude ethanolic extract of P. macrocarpa has high antioxidant activity and modulated the oxidative stress as proved by the up-regulation of glutathione-s-transferase and superoxide dismutase. Immunohistochemical staining of the treated sections showed overexpression of PCNA and Bax, reduced crypt sizes and numbers, indicating the characteristic feature of apoptotic cancer cells. PCNA is a landmark of cell damage and turn-over and can be associated with clinical cancer mutation. The most potent doses were 250 mg/kg and 500 mg/kg as compared to 35 mg/kg 5-fluorouracil. Conclusion: In this sense, the potential modulation of the colorectal pathophysiological pathway by P. macrocarpa natural compounds mostly flavonoids offer a great possibility for the discovery of new leads towards the colorectal cancer.
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