GENE EXPRESSION PROFILING AFTER ANGIOGENESIS INHIBITOR TREATMENT

The angiogenic process can be summarized as cell activation by a lack of oxygen releases angiogenic molecules that attract inflammatory and endothelial cells and promote their proliferation. Several protein fragments produced by the digestion of the blood-vessel walls intensify the proliferative activity of endothelial cells. Acetyl salicylic acid is often used as an analgesic drug to relieve minor aches and pains, a drug with antitumour activity and an anti-inflammatory medication. In our experiments we propose using the angiogenesis model and photodynamic therapy (PDT) for observing changes in angiogenesis after treatment, and also for increasing the effect of PDT by addition of angiogenesis inhibitors

THE EFFECT OF ACETYLSALICYLIC ACID ON ANGIOGENESIS IN VITRO

Angiogenesis, the formation of new blood vessels, is an essential aspect of, among others, embryonic development, wound healing and the female reproductive cycle. It is also necessary for the expansion of tumour masses beyond a minute volume. Acetylsalicylic acid (ASA) is a non-steroidal anti-inflammatory drug with additional antitumour activity. We tested ASA for its ability to inhibit angiogenesis in a simplified angiogenesis model, hASC+HUVEC co cultured in vitro, using immunocytochemical staining with fluorescence-marked antibodies and observation of tubulelike structures and their branching under a fluorescence microscope. We confirmed that ASA is an efficient and useful angiogenesis inhibitor and deserves further attention. We intend using the designed angiogenesis model and the methods described for observing changes in angiogenesis after anti tumour photodynamic therapy (PDT), and also for enhancing PDT efficiency by addition of angiogenesis inhibitors

Proline-based carbamates as cholinesterase inhibitors

Series of twenty-five benzyl (2S)-2-(arylcarbamoyl)pyrrolidine-1-carboxylates was prepared and completely characterized. All the compounds were tested for their in vitro ability to inhibit acetylcholinesterase (AChE) and butyrylcholinesterase (BChE), and the selectivity of compounds to individual cholinesterases was determined. Screening of the cytotoxicity of all the compounds was performed using a human monocytic leukaemia THP-1 cell line, and the compounds demonstrated insignificant toxicity. All the compounds showed rather moderate inhibitory effect against AChE; benzyl (2S)-2-[(2-chlorophenyl)carbamoyl]pyrrolidine-1-carboxylate (IC50 = 46.35 M) was the most potent agent. On the other hand, benzyl (2S)-2-[(4-bromophenyl)-] and benzyl (2S)-2-[(2-bromophenyl)carbamoyl]pyrrolidine-1-carboxylates expressed anti-BChE activity (IC50 = 28.21 and 27.38 M, respectively) comparable with that of rivastigmine. The ortho-brominated compound as well as benzyl (2S)-2-[(2-hydroxyphenyl)carbamoyl]pyrrolidine-1-carboxylate demonstrated greater selectivity to BChE. The in silico characterization of the structure–inhibitory potency for the set of proline-based carbamates considering electronic, steric and lipophilic properties was provided using comparative molecular surface analysis (CoMSA) and principal component analysis (PCA). Moreover, the systematic space inspection with splitting data into the training/test subset was performed to monitor the statistical estimators performance in the effort to map the probability-guided pharmacophore pattern. The comprehensive screening of the AChE/BChE profile revealed potentially relevant structural and physicochemical features that might be essential for mapping of the carbamates inhibition efficiency indicating qualitative variations exerted on the reaction site by the substituent in the 30-/40-position of the phenyl ring. In addition, the investigation was completed by a molecular docking study of recombinant human AChE

FOTOTOXICKÝ VLIV PORFYRINOVÝCH SENSITIZERŮ A VIDITELNÉHO ZÁŘENÍ NA GRAM-POZITIVNÍ METHICILIN-REZISTENTNÍ KMEN S. AUREUS

The use of antimicrobial photodynamic therapy (aPDT) as a therapeutic modality for the treatment of localized microbial infections represents an developing new field. The emergence of strains resistant to antibiotics has provided the necessary impulse for new drug or technology discoveries to combat these resistant compounds. Although the aPDT is still in infancy, its need is still growing. Like PDT, main components of antimicrobial photodynamic therapy are appropriate light, dye called photosensitizer and created reactive oxygen species. In this article photosensitizers TMPyP and ZnTPPS4 are investigated for antimicrobial photodynamic therapy. We tested these porphyrins on bacterial methicilin – resistant strain MRSA alone and bound in complex created with hp-β-cyclodextrin. The light emitting diodes (414 nm) were used at the doses 0 and 150 J/cm2. Tested concentrations were from 0.78 to 100 μM. This experimental work predicated that TMPyP is very successful compound in aPDT. In contrary to ZnTPPS4 which was efficient for eradication of tested gram-positive bacteria only in higher concentrations

SAR-mediated Similarity Assessment of the Property Profile for New, Silicon-Based AChE/BChE Inhibitors

A set of 25 novel, silicon-based carbamate derivatives as potential acetyl- and butyrylcholinesterase (AChE/BChE) inhibitors was synthesized and characterized by their in vitro inhibition profiles and the selectivity indexes (SIs). The prepared compounds were also tested for their inhibition potential on photosynthetic electron transport (PET) in spinach (Spinacia oleracea L.) chloroplasts. In fact, some of the newly prepared molecules revealed comparable or even better inhibitory activities compared to the marketed drugs (rivastigmine or galanthamine) and commercially applied pesticide Diuron®, respectively. Generally, most compounds exhibited better inhibition potency towards AChE; however, a wider activity span was observed for BChE. Notably, benzyl N-[(1S)-2-[(tert-butyldimethylsilyl)oxy]-1-[(2-hydroxyphenyl)carbamoyl]ethyl]-carbamate and benzyl N-[(1S)-2-[(tert-butyldimethylsilyl)oxy]-1-[(3-hydroxyphenyl) carbamoyl]ethyl]-carbamate were characterized by fairly high selective indexes. Specifically, the first compound was prescribed with the lowest IC50 value that corresponds quite well with galanthamine inhibition activity, while the inhibitory profiles of the second molecule and benzyl-N-[(1S)-2-[(tert-butyldimethylsilyl)oxy]-1-[(4-hydroxyphenyl)carbamoyl]ethyl]-carbamate are in line with rivastigmine activity. Moreover, a structure-activity relationship (SAR)-driven similarity evaluation of the physicochemical properties for the carbamates examined appeared to have foreseen the activity cliffs using a similarity-activity landscape index for BChE inhibitory response values. The ‘indirect’ ligand-based and ‘direct’ protein-mediated in silico approaches were applied to specify electronic/steric/lipophilic factors that are potentially valid for quantitative (Q)SAR modeling of the carbamate analogues. The stochastic model validation was used to generate an ‘average’ 3D-QSAR pharmacophore pattern. Finally, the target-oriented molecular docking was employed to (re)arrange the spatial distribution of the ligand property space for BChE and photosystemII (PSII).Soubor 25 nových karbamátových derivátů na bázi křemíku jako potenciálních inhibitorů acetyl- a butyrylcholinesterázy (AChE/BChE) byl syntetizován a charakterizován jejich inhibičními profily in vitro a indexem selektivity (SIs). Připravené sloučeniny byly také testovány na jejich inhibiční potenciál ve fotosyntetickém elektronovém transportu (PET) ve špenátových (Spinacia oleracea L.) chloroplastech. Ve skutečnosti některé nově připravené molekuly odhalily srovnatelné nebo dokonce lepší inhibiční aktivity ve srovnání s obchodovanými léčivy (rivastigmin nebo galanthamin) a komerčně aplikovaným pesticidem Diuron®. Obecně většina sloučenin vykazovala lepší inhibiční účinnost vůči AChE; bylo však pozorováno širší rozpětí aktivity u BChE. Zejména benzyl N-[(1S)-2-[(terc-butyldimethylsilyl) oxy]-1-[(2-hydroxyfenyl)karbamoyl]ethyl]-karbamát a benzyl N-[(1S)-2-[(terc-butyldimethylsilyl)oxy]-1-[(3-hydroxyfenyl)karbamoyl]ethyl]-karbamát byl charakterizován poměrně vysokými indexy selektivity. Konkrétně první sloučenina měla nejnižší hodnotou IC50, která docela dobře odpovídá inhibiční aktivitě galantaminu, zatímco inhibiční profily druhé molekuly a benzyl-N-[(1S)-2-[(terc-butyldimethylsilyl)oxy]-1-[(4-hydroxyfenyl)karbamoyl]ethyl]karbamátu jsou v souladu s rivastigminovou aktivitou. Navíc vyhodnocení podobnosti fyzikálně-chemických vlastností zkoumaných karbamátů na základě vztahu struktura-aktivita (SAR) předpovídalo jistou aktivitu pomocí indexu podobnost-aktivita pro hodnoty inhibiční odpovědi BChE. „Nepřímé“ ligandy a „přímé“ proteiny zprostředkované v silikonových přístupech byly použity ke stanovení elektronických/sterických/lipofilních faktorů, které jsou potenciálně platné pro kvantitativní (Q)SAR modelování karbamátových analogů. Validace stochastického modelu byla použita k vytvoření „průměrného“ 3D-QSAR farmakoforu. Nakonec bylo použito cílené molekulární dokování aby se změnila prostorová distribuce vlastností ligandu pro BChE a fotosystém II (PSII)

Proline-Based Carbamates as Cholinesterase Inhibitors

Series of twenty-five benzyl (2S)-2-(arylcarbamoyl)pyrrolidine-1-carboxylates was prepared and completely characterized. All the compounds were tested for their in vitro ability to inhibit acetylcholinesterase (AChE) and butyrylcholinesterase (BChE), and the selectivity of compounds to individual cholinesterases was determined. Screening of the cytotoxicity of all the compounds was performed using a human monocytic leukaemia THP-1 cell line, and the compounds demonstrated insignificant toxicity. All the compounds showed rather moderate inhibitory effect against AChE; benzyl (2S)-2-[(2-chlorophenyl)carbamoyl]pyrrolidine-1-carboxylate (IC50 = 46.35 μM) was the most potent agent. On the other hand, benzyl (2S)-2-[(4-bromophenyl)-] and benzyl (2S)-2-[(2-bromophenyl)carbamoyl]pyrrolidine-1-carboxylates expressed anti-BChE activity (IC50 = 28.21 and 27.38 μM, respectively) comparable with that of rivastigmine. The ortho-brominated compound as well as benzyl (2S)-2-[(2-hydroxyphenyl)carbamoyl]pyrrolidine-1-carboxylate demonstrated greater selectivity to BChE. The in silico characterization of the structure–inhibitory potency for the set of proline-based carbamates considering electronic, steric and lipophilic properties was provided using comparative molecular surface analysis (CoMSA) and principal component analysis (PCA). Moreover, the systematic space inspection with splitting data into the training/test subset was performed to monitor the statistical estimators performance in the effort to map the probability-guided pharmacophore pattern. The comprehensive screening of the AChE/BChE profile revealed potentially relevant structural and physicochemical features that might be essential for mapping of the carbamates inhibition efficiency indicating qualitative variations exerted on the reaction site by the substituent in the 3′-/4′-position of the phenyl ring. In addition, the investigation was completed by a molecular docking study of recombinant human AChE