Inhalační podání suchých práškových částic je využívanou aplikační cestou pro dosažení lokálního i systémového působení léčiv. U plicních onemocnění je žádoucí depozice léčiv v místě účinku. Pro efektivní léčbu jsou tak zásadní parametry inhalovaných částic, a to především jejich velikost, tvar nebo aerosolizační parametry. Vhodných parametrů je možné dosáhnout volbou metody přípravy nebo pomocných látek (nosičů, porogenů nebo aerosolizačních činidel). Cílem experimentu byla příprava jedenácti šarží práškových směsí sprejovým sušením, které se lišily použitým nosičem, množstvím leucinu či porogenu. Cílem bylo optimalizovat složení pro navázání léčiva s ohledem na požadavky pro plicní podání. Připravené částice byly zhodnoceny z hlediska morfologie, tokových vlastností, pórovitosti, geometrického i aerodynamického průměru. Bylo zjištěno, že se zvyšující se koncentrací leucinu se sypná hustota částic snižovala a zároveň rostla hodnota FPF. Stejně tak docházelo i ke snižování MMAD. Jako vhodná se jevila šarže obsahující 15 % leucinu. Při stanovení optimální koncentrace porogenu (hydrogenuhličitan amonný) u manitolových částic dosahovala nejlepších výsledků šarže s jeho 1% zastoupením, vzhledem k vyhovující velikosti částic oproti ostatním šaržím (MMAD 5,92 ± 1,32 μm), vhodné pórovitosti a obecně přijatelné morfologii částic. Za účelem formulace částic s navázaným léčivem by tedy bylo vhodné snížit aerodynamický průměr částic např. úpravou procesních parametrů sprejového sušení.
Inhalation administration of dry powder particles is a common application route to achieve local and systemic drug effects. For pulmonary diseases, the deposition of drugs at the site of action is desirable. Thus, the parameters of the inhaled particles, especially their size, shape, or aerosolization, are essential for effective treatment. Suitable parameters can be achieved by choice of preparation method or excipients (carriers, porogens, or aerosolizing agents). This experiment aimed to prepare 11 batches of powder mixtures by spray drying, which differed in the carrier used and the amount of leucine or porogen. The aim was to optimize the formulation for drug binding concerning the requirements for pulmonary administration. The prepared particles were evaluated in terms of morphology, flow properties, porosity, and geometric and aerodynamic diameter. It was found that with increasing concentration of leucine, the bulk density of the particles decreased while the FPF value increased. Similarly, there was a decrease in MMAD. The batch containing 15% leucine was the most suitable. In determining the optimum porogen concentration for mannitol particles, the batch with its 1% gave the best results due to its adequate particle size compared to the other batches (MMAD 5.92 ± 1.32 μm), suitable porosity, and particle morphology. Thus, to formulate drug-loaded particles, it would be advisable to reduce the aerodynamic diameter of the particles, e.g., by spray drying process parameters.
Here, we describe the synthesis and biological characterization of 32 novel phenylalanine and leucine dipeptides modified on both the N and C termini by salicylic acid and aromatic or alicyclic amines, respectively. All compounds displayed antiproliferative activity in the tested cancer cell lines and eight of the compounds exhibited single digit micromolar GI50 values. Treated cells rapidly detached from surface of tissue culture dishes and we found that focal adhesion kinase (FAK), p130CAS and paxillin, which are important regulators of cell adhesion, were dephosphorylated at Y397, Y410 and Y118, respectively. The most potent compound reduced proliferation in the HCT-116 cell line in a dose-dependent manner, as shown by a decrease in 5-bromo-2'-deoxyuridine incorporation into DNA. Furthermore, this compound increased the levels of several apoptotic markers, including activated caspases, and increased site-specific poly-(ADP-ribose)polymerase (PARP) cleavage.
- MeSH
- dipeptidy chemická syntéza chemie farmakologie MeSH
- fenylalanin chemická syntéza chemie farmakologie MeSH
- HCT116 buňky MeSH
- leucin chemická syntéza chemie farmakologie MeSH
- lidé MeSH
- molekulární struktura MeSH
- nádorové buňky kultivované MeSH
- proliferace buněk účinky léků MeSH
- protinádorové látky chemická syntéza chemie farmakologie MeSH
- screeningové testy protinádorových léčiv MeSH
- viabilita buněk účinky léků MeSH
- vztah mezi dávkou a účinkem léčiva MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Inhibition of protein degradation is one of strategies for suppression of uncontrolled proliferation of cancer cells. Proteolytic degradation in cells is mainly ensured by proteasome and its inhibition by bortezomib showed benefit in clinical use for the treatment of multiple myeloma. We report here the library of antiproteasomal O-benzyl salicylamides built from leucine and phenylalanine. Prepared compounds displayed antiproliferative activity on K562, CEM and U266 cancer cell lines, ranging from high micromolar to submicromolar GI50 values. The most potent compounds (series 4 and 6) were further assayed for their inhibition of chymotrypsin-like protease activity of the 26S proteasome in U266 cells. The majority of compounds inhibited the proteasome in mid-nanomolar concentrations (IC50 ranging from 57 to 197 nM) and it correlated with cellular potency. In a cell based assay involving green fluorescence protein (GFP) fused to a short degron that is rapidly degraded by a proteasome the compounds induced accumulation of GFP, visualised and quantified by live-cell imaging. Levels of polyubiquitinated proteins in U266 cells treated by compound 4m were also analyzed by immunoblotting, revealing a typical high molecular mass smear of ubiquitin conjugates. Finally, apoptotic cell death in treated U266 cells was detected biochemically by measuring the activity of caspases 3 and 7 in lysates and by immunoblotting of caspase 7, its substrate poly(ADP-ribose)polymerase, and Mcl-1, which all together showed changes typical for apoptosis. All these observations were in agreement with expected cellular mechanism of action and confirmed proteasome targeting by prepared O-benzyl salicylamides.
- MeSH
- fenylalanin chemie farmakologie MeSH
- inhibitory proteasomu chemická syntéza chemie farmakologie MeSH
- leucin chemie farmakologie MeSH
- lidé MeSH
- molekulární struktura MeSH
- nádorové buněčné linie MeSH
- proliferace buněk účinky léků MeSH
- proteasomový endopeptidasový komplex metabolismus MeSH
- protinádorové látky chemická syntéza chemie farmakologie MeSH
- salicylamidy chemická syntéza chemie farmakologie MeSH
- screeningové testy protinádorových léčiv MeSH
- vztah mezi dávkou a účinkem léčiva MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
The low density lipoprotein receptor (LDLR) is a transmembrane protein that plays a key role in cholesterol metabolism. It contains 860 amino acids including a 21 amino acid long signal sequence, which directs the protein into the endoplasmic reticulum. Mutations in the LDLR gene lead to cholesterol accumulation in the plasma and results in familial hypercholesterolemia (FH). Knowledge of the impact of a mutation on the LDLR protein structure and function is very important for the diagnosis and management of FH. Unfortunately, for a large proportion of mutations this information is still missing. In this study, we focused on the LDLR signal sequence and carried out functional and in silico analyses of two sequence changes, p.(Gly20Arg) and p.(Leu15Pro), localized in this part of the LDLR. Our results revealed that the p.(Gly20Arg) change, previously described as disease causing, has no detrimental effect on protein expression or LDL particle binding. In silico analysis supports this observation, showing that both the wt and p.(Gly20Arg) signal sequences adopt an expected α-helix structure. In contrast, the mutation p.(Leu15Pro) is not associated with functional protein expression and exhibits a structure with disrupted a α-helical arrangement in the signal sequence, which most likely affects protein folding in the endoplasmic reticulum.
- MeSH
- arginin chemie MeSH
- CHO buňky MeSH
- Cricetulus MeSH
- endoplazmatické retikulum metabolismus MeSH
- glycin chemie MeSH
- heterozygot MeSH
- hyperlipoproteinemie typ II krev genetika MeSH
- konfokální mikroskopie MeSH
- křečci praví MeSH
- LDL-receptory genetika metabolismus MeSH
- leucin chemie MeSH
- lidé MeSH
- mutace MeSH
- prolin chemie MeSH
- proteiny - lokalizační signály genetika MeSH
- rodokmen MeSH
- sbalování proteinů MeSH
- sekundární struktura proteinů MeSH
- zvířata MeSH
- Check Tag
- křečci praví MeSH
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
o-Phthaldialdehyde was used in combination with a series of six thiosugars for the pre-column chiral derivatization of selected primary amino acids. The diastereomers were resolved on a conventional reversed-phase column and with fluorescence detection. A surprisingly effective resolution of 1-isoindolyl-(1-thioglycosides) derived from 1-thio-beta-L-fucose was observed.
BACKGROUND: Ghrelin is an endogenous hormone expressed predominantly in the stomach. Ghrelin controls growth hormone secretion and also affects the body's energy balance. We analyzed the association of ghrelin variants with body mass index (BMI), albumin as a marker of malnutrition and plasma lipids as risk factors for atherosclerosis in hemodialyzed patients, in whom malnutrition and accelerated atherosclerosis are common complications. METHODS: Ghrelin variants Arg51>Gln and Leu72> Met were analyzed by PCR-RFLP in 210 hemodialyzed patients, prospectively followed up for 15 months. Changes in body mass index, triglycerides, total cholesterol and albumin over time (after 3, 6, 9, 12 and 15 months of dialysis) were analyzed in subgroups divided according to ghrelin genotypes. RESULTS: Carriers of at least one of the Gln51 and Met72 alleles lost body weight more quickly than Arg51Arg/Leu72Leu homozygotes (p<0.01). Carriers of the Gln51 allele were at higher risk of developing high cholesterol levels (p<0.01). CONCLUSIONS: Common ghrelin variants may have an effect on changes in biochemical and anthropometric parameters in hemodialyzed patients over time and could be used in the future to plan individualized therapy.
- MeSH
- alely MeSH
- arginin chemie MeSH
- cholesterol krev MeSH
- dialýza ledvin MeSH
- dialýza MeSH
- financování organizované MeSH
- genetická variace MeSH
- ghrelin biosyntéza genetika metabolismus MeSH
- glutamin MeSH
- heterozygot MeSH
- index tělesné hmotnosti MeSH
- leucin chemie MeSH
- lidé MeSH
- methionin chemie MeSH
- renální insuficience genetika terapie MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Klíčová slova
- AdaAhx3L3VS,
- MeSH
- aminokyseliny chemie MeSH
- finanční podpora výzkumu jako téma MeSH
- inhibitory proteas farmakologie MeSH
- kosterní svaly chemie metabolismus účinky léků MeSH
- leucin chemie MeSH
- potkani Wistar MeSH
- proteiny metabolismus MeSH
- proteosyntéza MeSH
- tyrosin chemie MeSH
- vinylové sloučeniny farmakologie MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- MeSH
- finanční podpora výzkumu jako téma MeSH
- kosterní svaly metabolismus účinky léků MeSH
- krysa rodu rattus MeSH
- leucin analýza chemie MeSH
- peptidy farmakologie MeSH
- proteiny metabolismus MeSH
- proteosyntéza MeSH
- sepse metabolismus MeSH
- sulfony farmakologie MeSH
- tyrosin analýza MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- zvířata MeSH
