Czynniki rokownicze śmiertelności po operacji pękniętych tętniaków tętnicy szyjnej wewnętrznej

Background and purpose An analysis of predictors of mortality in patients with ruptured intracranial aneurysm is an important aspect in the assessment of outcome. The aim of the study was to analyse factors determining mortality risk after the surgical treatment of ruptured internal carotid artery (ICA) aneurysm. Material and methods This study comprised 242 patients operated on between 1997 and 2006 in the Neurosurgery Department of the Medical University Hospital in Gdansk, Poland. Multivariate logistic regression, ROC curves (for model assessment as a mortality classifier) and population attributable risk for contribution of individual factor mortality explanation were used to assess factors related to in-hospital mortality. Results 14.9% of patients died postoperatively. In univariate analysis, increased risk of death was related to the Glasgow Coma Scale score, WFNS score, Hunt-Hess and Fisher grade, preoperative neurological deficit, delayed cerebral ischaemia (DCI), trapping and bypass operative method. Multivariate analysis revealed two independent predictors of in-hospital mortality: DCI and Hunt-Hess grade. 91% of mortality risk was attributed to grade 4 or 5 in Hunt-Hess scale and DCI. The dominant predictor of survival was the Hunt-Hess scale. Increase by one grade in the Hunt-Hess scale resulted in two-fold increase of in-hospital mortality risk. Conclusions Postoperative mortality after ICA aneurysm rupture is determined by clinical status at admission and the occurrence of DCI.Wstęp i cel pracy Analiza czynnikowa śmiertelności wśród chorych z pękniętym tętniakiem wewnątrzczaszkowym stanowi ważny aspekt w ocenie wyników leczenia. Celem pracy była analiza czynników mających wpływ na zwiększone ryzyko zgonu po operacji krwawiącego tętniaka tętnicy szyjnej wewnętrznej. Materiał i metody W badaniu wzięło udział 242 chorych operowanych w latach 1997—2006 w Klinice Neurochirurgii Gdańskiego Uniwersytetu Medycznego. Czynniki rokownicze śmiertelności w tej grupie analizowano za pomocą wieloczynnikowej regresji logistycznej, porównania krzywych ROC (celem oceny poprawności całego modelu jako klasyfikatora śmiertelności) i wskaźników ryzyka przypisanego populacji (PAR) dla wskazania udziału poszczególnych czynników w wyjaśnieniu śmiertelności. Analizowano jedynie przypadki zgonów występujące w trakcie hospitalizacji. Wyniki Śmiertelność pooperacyjna wyniosła 14,9%. W analizie jednoczynnikowej ryzyko zgonu zależało od: stopnia w skali śpiączki Glasgow, punktacji w skali WFNS, Hunta–Hessa, Fishera, przedoperacyjnego deficytu neurologicznego, rozpoznania opóźnionego niedokrwienia mózgu (delayed cerebral ischaemia — DCI), metody operacyjnej polegającej na zamknięciu naczynia lub wytworzeniu obejścia naczyniowego. Spośród nich analiza wieloczynnikowa wyłoniła dwie niezależne zmienne, które miały istotny wpływ na śmiertelność: DCI oraz stan kliniczny pacjentów przy przyjęciu mierzony w skali Hunta-Hessa. Stopień 4. i 5. w tej skali oraz rozpoznanie DCI odpowiadały za 91% ryzyka zgonu po operacji tętniaka tętnicy szyjnej wewnętrznej. Dominujące znaczenie w przewidywaniu przeżycia miała skala Hunta-Hessa. Ocena stanu klinicznego chorego o jeden stopień wyżej w tej skali zwiększała ponaddwukrotnie ryzyko zgonu pooperacyjnego. Wnioski Śmiertelność pooperacyjna po pęknięciu tętniaka tętnicy szyjnej wewnętrznej uwarunkowana jest stanem klinicznym przy przyjęciu i wystąpieniem DCI

Mapping the substrate-binding subsite specificity of a Porphyromonas gingivalis Tpr peptidase

Calcium-dependent peptidases of the calpain family are widespread in eukaryotes but uncommon in prokaryotes. A few bacterial calpain homologs have been discovered but none of them have been characterized in detail. Here we present an in-depth substrate specificity analysis of the bacterial calpain-like peptidase Tpr from Porphyromonas gingivalis. Using the positional scanning hybrid combinatorial substrate library method, we found that the specificity of Tpr peptidase differs substantially from the papain family of cysteine proteases, showing a strong preference for proline residues at positions P2 and P3. Such a degree of specificity indicates that this P. gingivalis cell-surface peptidase has a more sophisticated role than indiscriminate protein degradation to generate peptide nutrients, and may fulfil virulence-related functions such as immune evasion

Calpeptin is a potent cathepsin inhibitor and drug candidate for SARS-CoV-2 infections

Several drug screening campaigns identified Calpeptin as a drug candidate against SARS-CoV-2. Initially reported to target the viral main protease (Mpro), its moderate activity in Mpro inhibition assays hints at a second target. Indeed, we show that Calpeptin is an extremely potent cysteine cathepsin inhibitor, a finding additionally supported by X-ray crystallography. Cell infection assays proved Calpeptin’s efficacy against SARS-CoV-2. Treatment of SARS-CoV-2-infected Golden Syrian hamsters with sulfonated Calpeptin at a dose of 1 mg/kg body weight reduces the viral load in the trachea. Despite a higher risk of side effects, an intrinsic advantage in targeting host proteins is their mutational stability in contrast to highly mutable viral targets. Here we show that the inhibition of cathepsins, a protein family of the host organism, by calpeptin is a promising approach for the treatment of SARS-CoV-2 and potentially other viral infections

Unnatural amino acids as achemical tool for the development of protease substrates, inhibitors and activity - baseproblems

Proteolytic enzymes are molecular scissors that are responsible for the amide bond breakdown in peptide and protein substrates. Over the years, the view on proteases has been considerably changed from non-specific digestive enzymes to sophisticated biocatalysts, which by performing limited proteolysis control virtually all biological processes. In order to better understand how proteases work and what are their biologically relevant target substrates, it is indispensable to determine their catalytic preferences. This knowledge can be further utilized to develop selective substrates, inhibitors and activity-based probes (ABPs) enabling the monitoring of proteases activity in various settings, from in vitro analysis on recombinant enzymes or cell lysates to ex vivo and in vivo imaging at the single cell level. Among many chemical-based approaches that have been developed and applied over the years, the Hybrid Combinatorial Substrate Library (HyCoSuL) technology has emerged as one of the most powerful one. HyCoSuL is a combinatorial peptide-based library of fluorogenic substrates, that comprise natural and unnatural amino acids, that can deeply explore the chemical space in proteases active site, providing a structural framework for the development of highly-selective chemical tools. In this review we present the most prominent examples of proteolytic enzymes that have been profiled with HyCoSuL approach yielding selective substrates, potent inhibitors, and very sensitive activity-based probes

Fluorescent Activity-Based Probe for the Selective Detection of Factor VII Activating Protease (FSAP) in Human Plasma

The zymogen form of circulating Factor VII activating protease (FSAP) is activated by histones that are released as a consequence of tissue damage or excessive inflammation. This is likely to have consequences in a number of disease conditions such as stroke, atherosclerosis, liver fibrosis, thrombosis and cancer. To investigate the existence, as well as the concentration of active FSAP (FSAPa) in complex biological systems an active site probe is needed. We used Hybrid Combinatorial Substrate Library (HyCoSuL) to screen for natural and unnatural amino acids that specifically bind to P4-P2 pockets of FSAPa. This information was used to designing a fluorogenic substrate (Ac-Pro-DTyr-Lys-Arg-ACC) as well as an irreversible, fluorogenic activity-based probe Cy5-6-Ahx-Pro-DTyr-Lys-ArgP(OPh)2. In normal human plasma the probe showed very low non-specific reactivity with some plasma proteins but upon activation of pro-FSAP with histones, strong labelling of FSAPa was observed. This labelling could be inhibited by aprotinin and was not found in the plasma of a subject that was homozygous for a polymorphism, which leads to loss of activity, or in plasma that was depleted of FSAP by antibodies. This 2nd generation substrate exhibited 6-fold higher catalytic efficiency than the 1st generation substrate and a much higher selectivity for FSAPa over other plasma proteases. This substrate and probe can be useful to detect and localize FSAPa in normal and pathological tissue and plasma to gain more insight into its functions

SARS-CoV-2 Mpro^{pro} inhibitors and activity-based probes for patient-sample imaging

In December 2019, the first cases of infection with a novel coronavirus, SARS-CoV-2, were diagnosed. Currently, there is no effective antiviral treatment for COVID-19. To address this emerging problem, we focused on the SARS-CoV-2 main protease that constitutes one of the most attractive antiviral drug targets. We have synthesized a combinatorial library of fluorogenic substrates with glutamine in the P1 position. We used it to determine the substrate preferences of the SARS-CoV and SARS-CoV-2 main proteases. On the basis of these findings, we designed and synthesized a potent SARS-CoV-2 inhibitor (Ac-Abu-dTyr-Leu-Gln-VS, half-maximal effective concentration of 3.7 µM) and two activity-based probes, for one of which we determined the crystal structure of its complex with the SARS-CoV-2 M$^{pro}$. We visualized active SARS-CoV-2 M$^{pro}$ in nasopharyngeal epithelial cells of patients suffering from COVID-19 infection. The results of our work provide a structural framework for the design of inhibitors as antiviral agents and/or diagnostic tests