Submitted by Marco Antonio Oliveira da Silva (maosilva@ipen.br) on 2016-12-21T18:09:44Z No. of bitstreams: 0 / Made available in DSpace on 2016-12-21T18:09:44Z (GMT). No. of bitstreams: 0 / A Austrália é um país cuja fauna é um repositório de potenciais novos biofármacos, pois se encontram no continente os animais mais mortais do planeta, dentre eles, as serpentes. A serpente Pseudechis australis (Mulga snake) é a maior serpente venenosa da Austrália e tem ampla distribuição geográfica. Os venenos de serpentes são complexas misturas com proteínas e peptídeos que apresentam uma variedade de atividades biológicas. Devido à riqueza de seus componentes, várias moléculas encontradas no veneno vêm sendo utilizadas com fins terapêuticos, como agentes anticoagulantes ou analgésicos. Apesar dessas informações, existem poucos dados disponíveis sobre os componentes específicos deste veneno. O presente trabalho tem como objetivo isolar e caracterizar as proteases desse veneno, ainda não descritas, um primeiro passo para compreender o papel destas enzimas no processo de envenenamento, assim como seus inibidores endógenos. Estes desempenham uma função protetora da glândula de veneno, inibindo a ação das enzimas in loco, prevenindo assim a degradação do tecido glandular por estas toxinas. O interesse nestes inibidores está relacionado ao seu potencial uso na terapia de diversas doenças como distúrbios da coagulação, hipertensão e câncer. / Dissertação (Mestrado em Tecnologia Nuclear) / IPEN/D / Instituto de Pesquisas Energéticas e Nucleares - IPEN-CNEN/SP

Submitted by Daniely Januário (daniely.januario@gmail.com) on 2018-03-02T12:31:14Z No. of bitstreams: 1 joseresendedecastrojunior.pdf: 3078753 bytes, checksum: f22309c68ef54befb8ce6682fdfa7d3e (MD5) / Approved for entry into archive by Adriana Oliveira (adriana.oliveira@ufjf.edu.br) on 2018-03-14T18:29:39Z (GMT) No. of bitstreams: 1 joseresendedecastrojunior.pdf: 3078753 bytes, checksum: f22309c68ef54befb8ce6682fdfa7d3e (MD5) / Made available in DSpace on 2018-03-14T18:29:39Z (GMT). No. of bitstreams: 1 joseresendedecastrojunior.pdf: 3078753 bytes, checksum: f22309c68ef54befb8ce6682fdfa7d3e (MD5) Previous issue date: 2016-09-30 / PROQUALI (UFJF) / A fibrilação atrial é a arritmia cardíaca sustentada mais comum, acometendo principalmente idosos e aumenta o risco de acidente vascular cerebral. É atualmente um problema de saúde pública pela sua elevada morbimortalidade e custos crescentes. A varfarina é o anticoagulante que se mostrou efetivo na prevenção deste evento, contudo apresenta limitações que contribuem para a sua baixa aderência e subutilização. O surgimento de novos anticoagulantes, como a rivaroxabana, evidenciou que esta medicação pode ser tão efetiva quanto à varfarina, mas, com menores taxas de complicações hemorrágicas graves, principalmente cerebrais, não havendo necessidade de exames de monitorização, porém com um custo maior. O objetivo deste estudo foi avaliar a custo-utilidade da rivaroxabana comparada à varfarina nesta arritmia, ou seja, comparar o custo da intervenção com sua efetividade medida como ganho em anos de vida ajustada pela qualidade. Inicialmente foi realizada uma revisão sistemática de estudos econômicos sugerindo que a rivaroxabana pode ser uma opção custo-efetiva, principalmente em países desenvolvidos. Houve apenas um estudo realizado em país em desenvolvimento que mostrou resultado divergente. Posteriormente, foi desenvolvido um modelo econômico de Markov, na perspectiva do Sistema Único de Saúde, que permitiu simular a evolução de uma coorte de idosos com fibrilação atrial, em ciclos trimestrais durante toda a vida. Os resultados evidenciaram que os custos incrementais da rivaroxabana foram superiores à varfarina (R$ 7.135,48), com um discreto aumento de utilidade, resultando numa razão de custo-utilidade incremental de R$ 206.816,45/anos de vida ajustada pela qualidade. Este valor encontra-se acima do limiar proposto pela Organização Mundial da Saúde, podendo não ser uma opção custo-efetiva. Apesar das limitações, este trabalho conseguiu reunir as evidências disponíveis e mostrou a necessidade de se ajustar os protocolos clínicos e diretrizes para uma prática clínica que possa conciliar os princípios de integralidade do cuidado à sustentabilidade do sistema de saúde. / Atrial fibrillation is the most common sustained cardiac arrhythmia, affecting mainly the elderly and increases the risk of stroke. It is currently a public health problem because of its high morbidity and mortality and increasing costs. Warfarin is an anticoagulant that was effective in preventing this type of stroke, but has limitations that contribute to its low adhesion and underutilization. The emergence of new anticoagulants such as rivaroxaban, suggest that this drug may be as effective as warfarin, but with lower rates of major bleeding complications, mainly brain. Also, with this drug, there is no need for monitoring tests, but is more expensive. The aim of this study was to evaluate the cost-utility of rivaroxabana compared to warfarin in this arrhythmia, or compare the cost of a health intervention with effectiveness measured as a gain in years of life adjusted for quality. Initially, a systematic review of economic studies was performed that suggest that rivaroxaban can be a cost-effective option compared to warfarin, especially in developed countries. There was only one study in a developing country which showed divergent results. Later, it was developed an economic model of Markov was developed with the Unified Health System perspective, which allowed simulation of the evolution of a cohort of elderly patients with Atrial fibrillation in quarterly cycles throughout life. The results showed that the incremental costs of rivaroxaban was superior to warfarin (R$ 7.135,48), with a slight increase of utility, resulting in incremental cost-utility ratio of R$ 206.816,45/years of life adjusted for quality. This value is above the threshold proposed by the Wolrd Health Organization, may not be a cost-effective option. Despite the limitations, this study brought together the available evidence and showed the need to adjust the clinical protocols and guidelines for clinical practice that can reconcile the principles of comprehensive care to the sustainability of the health system.

Secondary metabolites derived from plants, especially those used by traditional healers, are at the forefront of new drug development in combating diseases such as cancer and diabetes. Garlic is employed in indigenous medicine all over the world for the treatment of a variety of diseases. Dietary garlic has been recognized for its beneficial health effects. In particular, garlic consumption has been correlated with (i) reduction of risk factors for cardiovascular diseases and cancer, (ii) stimulation of immune function, (iii) enhanced detoxification of foreign compounds, (iv) hepatoprotection, (v) antimicrobial effects, (vi) antioxidant effects, and most importantly (vii) its hypoglycemic and anticoagulant properties. Due to these beneficial properties, garlic and its closely related genera which includes Tulbaghia violacea, may be useful as coadjuvant therapy in the treatment of type 2 diabetes and some of its physiological complications. The aim of this study was to determine if T. violacea has antidiabetic and anticoagulant properties. This was performed in vitro using both aqueous and organic extracts of the roots, leaves and bulbs. An organic extract was able to improve glucose-stimulated insulin secretion (GSIS) in INS-1 pancreatic β-cells and glucose uptake in Chang liver cells. The BO extract had no effect on the glucose uptake in 3T3-L1 an adipose cell line and reduced glucose utilisation in C2C12, a skeletal muscle cell line. Some of the properties displayed by T. violacea in this study are consistent with those found in similar studies with garlic extracts. It was observed that the BO extract increased the membrane potential and Glut-2 expression in INS-1 cells cultured at hyperglycemic levels, however, at normoglycemic levels a reduction was observed. The oxygen consumption increased at both glycemic levels due to treatment with the BO extract. Platelets were exposed to the extracts to determine their effects upon platelet aggregation, adhesion and protein secretion. Since the BO extract displayed the highest potential at inhibiting platelet aggregation and adhesion. A rat model was used in ex vivo studies to determine if the extract exhibited the same effect in a physiological model. It was noted that the BO extract exhibited a higher degree of inhibition on platelet aggregation and adhesion than the positive control, aspirin. The BO extract reduced clotting times in the prothrombin time (PT) test, but prolonged the clotting time in the actived partial thromboplastin time (APTT) assay in the ex vivo model; however, it had no affect on these clotting assays in the in vitro model using human blood. The BO extract increased the D-dimer and Fibrinogen-C levels in the in vitro model, but had no effect on the D-dimer concentrations and lowered the Fibrinogen-C levels in the ex vivo model. The active compounds in the extract remain to be elucidated.

Rotas sintéticas foram empregadas no grupo funcional ligado ao carbono 4 da trans-hidroxi-L-prolina para a obtenção de quatro análogos de prolina com amino e guanido grupos nesta posição e isomeria espacial cis e trans. Adicionalmente, o aminoácido guanidino fenilalanina foi comparado com os análogos de prolina em todos os ensaios realizados. Entre os análogos de prolina sintetizados, o peptídeo contendo o aminoácido não natural com o grupo funcional guanido e isomeria trans apresentou a melhor atividade inibitória contra trombina. Entretanto, o peptídeo sintetizado com o aminoácido guanidino fenilalanina ainda demonstra ter uma melhor atividade inibitória em comparação com os análogos de prolina. / Synthetic routes were employed in the functional group attached to the carbon 4 of trans-4-hydroxy-L-proline to obtain four proline analogues with amino and guanido groups in this position and cis and trans spatial isomerism. Additionally, the amino acid guanidino phenylalanine was compared with the analogues of proline in all tests. Among the proline analogues synthesized, the peptide containing the unnatural amino acid functional group with guanido and trans isomerism showed the best inhibitory activity against thrombin. However, the peptide synthesized with the amino acid guanidino phenylalanine exhibits an even better inhibitory activity in comparison to proline analogues.

Background Fewer than one-third of nursing home residents with atrial fibrillation were treated with the only available oral anticoagulant, warfarin, historically. Management of atrial fibrillation has transformed in recent years with the approval of 4 direct-acting oral anticoagulants (DOACs) since 2010. Methods Using the national Minimum Data Set 3.0 linked to Medicare Part A and D claims, we first described contemporary (2011-2016) warfarin and DOAC utilization in the nursing home population (Aim 1). In Aim 2, we linked residents to nursing home and county level data to study associations between resident, facility, county, and state characteristics and anticoagulant treatment. Using a new-user active comparator design, we then compared the incidence of safety (i.e., bleeding), effectiveness (i.e., ischemic stroke), and mortality outcomes between residents initiating DOACs versus warfarin (Aim 3). Results The proportion of residents with atrial fibrillation receiving treatment increased from 42.3% in 2011 to 47.8% as of December 31, 2016, at which time 48.2% of treated residents received DOACs. Demographic and clinical characteristics of residents using DOACs and warfarin were similar in 2016. Half of the 8,734 DOAC users received standard dosages and most were treated with apixaban (54.4%) or rivaroxaban (35.8%) in 2016. Compared with warfarin, bleeding rates were lower and ischemic stroke rates were higher for apixaban users. Ischemic stroke and bleeding rates for dabigatran and rivaroxaban were comparable to warfarin. Mortality rates were lower versus warfarin for each DOAC. Conclusions In nursing homes, DOACs are being used commonly and with equal or greater benefit than warfarin.

<p>Venous thromboembolism (VTE), which encompasses deep vein thrombosis (DVT) and pulmonary embolism (PE), is the most preventable cause of death in hospitalized patients. Due to its high mortality, morbidity, and cost, health care providers are obligated to not only effectively diagnose and treat VTE, but also to prevent it if possible. This has been reinforced by a number of national and international quality initiatives to prevent hospital-acquired VTE. Despite the existence of well-accepted clinical practice guidelines on VTE prophylaxis, 1 in 3 hospitalized medical patients receives an inappropriate VTE prophylaxis strategy. Both underuse of prophylaxis in patients with VTE risk, and overuse of prophylaxis in patients without VTE risk are problems. The use of inappropriate VTE prophylaxis strategies is likely due to the complexity and heterogeneity of hospitalized medical patients, and the difficulty of applying “one size fits all” practice guidelines to this group. Institution-wide knowledge translation strategies are required to close the gap between evidence and practice, and promote evidence-based VTE prophylaxis strategies in hospitalized medical patients. The objective of this thesis is to design a cluster randomized controlled trial to determine if a standardized electronic order set, with an embedded computerized decision support system and audit and feedback component, affects the use of appropriate VTE prophylaxis in hospitalized medical patients. The unit of randomization in this study is the hospital, which serves as the cluster. The unit of observation in this study is the individual patient. The primary outcome of this study is the proportion of in-hospital days during which appropriate VTE prophylaxis is administered, in intervention versus control hospitals. Secondary outcomes are the rates of hospital-acquired VTE, major bleeding and mortality, in intervention versus control hospitals. Design, analytic and ethical challenges unique to cluster randomized trials will also be discussed. Strategies to overcome them in this trial will be presented.</p> / Master of Science (MSc)

Background Treatment of ischemic stroke with recombinant tissue plasminogen activator for intravenous thrombolysis (IVT) must be delivered within a narrow time window after symptom onset. This effective hyperacute treatment can be administered after ruling out active anticoagulation with direct oral anticoagulants (DOACs). Whenever this is impractical, e.g., due to aphasia, plasmatic DOAC levels are measured with a consequent delay in the IVT decision-making process ranging from 30 to 60 minutes of time. This study will test the hypothesis that hyperacute point-of-care assessment of clotting time in the patient's whole blood has sufficient diagnostic accuracy to determine immediately whether stroke patients are pretreated with DOAC. Methods and Design This will be a prospective single-center diagnostic accuracy study in 1,850 consecutive acute ischemic stroke patients at a tertiary stroke center in Saxony, Germany. Presence of active anticoagulation with DOAC will be determined by point-of-care quantification of clotting time via whole blood viscoelastic testing (ClotPro) using Russell venom viper and ecarin assay compared with high-performance liquid chromatography-tandem mass spectrometry as the reference standard. Discussion Viscoelastic point-of-care assessment of clotting time in whole blood might improve swift delivery of time-sensitive hyperacute treatment with IVT in stroke patients.

BACKGROUND:Anticoagulation with vitamin K antagonists such as warfarin has historically been used for the long term management of patients with thromboembolic disease. However, these agents have a slow onset of action which requires bridging therapy with heparin and its analogues, which are available only in parenteral route. To overcome these limitations, new oral anticoagulants such as factor Xa inhibitors and direct thrombin inhibitors have been developed. The aim of this article is to systematically review the phase 3 clinical trials of new oral anticoagulants in common medical conditions.METHODS:We searched PubMed (Medline) from January 2007 to February 2013 using "Oral anticoagulants", "New oral anticoagulants", "Randomized controlled trial", "Novel anticoagulants", "Apixaban", "Rivaroxaban", "Edoxaban", "Dabigatran etexilate", "Dabigatran" and a combination of the above terms. The available evidence from the phase 3 RCTs was summarized on the basis of individual drug and the medical conditions categorized into "atrial fibrillation", "acute coronary syndrome", "orthopedic surgery", "venous thromboembolism" and "medically ill patients".RESULTS:Apixaban, rivaroxaban and dabigatran have been found to be either non-inferior or superior to enoxaparin in prophylaxis of venous thromboembolism in knee and hip replacement with similar bleeding risk, superior to warfarin for stroke prevention in atrial fibrillation with significant reduction in the risk of major bleeding, non-inferior to aspirin for reducing cardiovascular death and stroke in acute coronary syndrome with significant increase in the risk of major bleed. Rivaroxaban and dabigatran are also superior to the conventional agents in the management of symptomatic venous thromboembolism. However, compared to enoxaparin, apixaban and rivaroxaban use lead to significantly increased bleeding risk in medically ill patients. Additional studies evaluating the specific reversal agents of these new drugs for the management of life-threatening bleeding or other adverse effects are necessary.CONCLUSION:Considering their pharmacological properties, their efficacy and bleeding complications, the new oral agents offer a net favourable clinical profile in orthopedic surgery, atrial fibrillation, acute coronary syndrome and increase the risk of bleeding in critically ill patients. Further studies are necessary to determine the long term safety and to identify the specific reversal agents of these new drugs.

<p>UVOD: Komplikacije terminalne bubrežne isuficijencije (TBI) kada se jačina glomerularne filtracije (JGF) smanji ispod 10 mL/min moguće je lečiti jedino hroničnom dijalizom ili transplantacijom bubrega tj. nadoknadom potpuno ili delimično izgubljene bubrežne funkcije. Uz blagovremenu edukaciju bolesnika o progresivnom toku hronične bubrežne bolesti, mogućnostima dijaliznog tretmana i transplantacije bubrega, treba na vreme obezbediti stalni funkcionalni vaskularni pristup za hemodijalizu (HD) hirur&scaron;kom intervencijom kreiranja arteriovenske fistule (AVF), po mogućnosti najmanje 6 meseci pre anticipiranog započinjanja HD, jer je za njenu maturaciju potrebno 4 do 6 nedelja. Primarna AVF je op&scaron;tepreporučeni najbolji stalni vaskularni pristup za bolesnike kod kojih se planira hemodijaliza. Najče&scaron;ći razlog za disfunkciju vaskularnog pristupa za hemodijalizu su u 80% slučajeva trombozne komplikacije, koje se u 90% slučajeva javljaju na venskom segmentu AVF i posledica su progresivne venske neointimalne hiperplazije. Pored histolo&scaron;kih karakteristika zida venskog krvnog suda i hemodinamskih uslova, u etiopatogenezi ovog &raquo;adaptivnog odgovora&laquo; vrlo značajnu ulogu igraju endotel i ostale komponente hemostaznog sistema (trombocitna, koagulaciona i fibrinolizna), imunolo&scaron;ki i citolo&scaron;ki činioci i genetski faktori. Prevencija nastanka rane tromboze vaskularnog pristupa za hemodijalizu kod bolesnika sa TBI je moguća primenom antitromboznih lekova, tj. antitrombocitne ili antikoagulantne terapije. CILJ: Proceniti efikasnost primenjenih antitromboznih lekova (tiklopidina i nadroparin-kalcijuma) u prevenciji nastanka rane tromboze/afunkcionalnosti AVF za hemodijalizu za vreme njene maturacije unutar 6 nedelja od kreiranja u bolesnika sa TBI. Ispitati nivo biomarkera hemostaznog sistema i markere trombofilije u bolesnika sa TBI pre kreiranja AVF u cilju dopune uzroka nastanka rane tromboze/afunkcionalnosti arteriovenskih fistula za hemodijalizu. Ispitati učestalost trombofilije i njen uticaj na funkcionalnost AVF i uporediti efikasnost primenjenih preventivnih režima između bolesnika sa i bez trombofilije. MATERIJAL I METODE: U ispitivanje su uključene osobe oba pola sa prethodno postavljenom dijagnozom TBI kod kojih nisu postojale kontraindikacije za planirno hirur&scaron;ko kreiranje prvog stalnog vaskularnog pristupa za hemodijalizu u vidu autologne arteriovenske fistule (AAVF). Nakon hirur&scaron;kog kreiranja radiocefalične arteriovenske fisule u distalnoj trećini podlaktice nedominantne ruke (89/121), intermedijalne (4/121) ili proksimalne (28/121) AAVF u studiju je uključen 121 ispitanik, koji su u cilju procene uticaja različitih antitromboznih lekova na sprečavanje nastanka rane tromboze fistula za hemodijalizu kod bolesnika sa TBI ispitanici su podeljeni u 3 grupe: Grupa I, kontrolna; 40 ispitanika koji nakon kreiranja AVF nisu dobijali antitromboznu terapiju, Grupa II; 42 ispitanika kod kojih je dan nakon kreiranja AVF započeta primena antitrombocitnog leka iz grupe tienopiridina, Ticlodix&reg; (ticlopidin) tbl a 250 mg, 2 x &frac12; tbl dnevno tokom 6 nedelja i Grupa III; 39 ispitanika kod kojih je dan nakon kreiranja AVF započeta subkutana primena antikoagulantnog leka iz grupe niskomolekularnih heparina, Fraxiparine&reg; (nadroparin-kalcijum) 2850 anti Xa i.j. (0.3 ml) dnevno tokom 6 nedelja. Jednokratno određivanje laboratorijskih parametara pokazatelja bubrežne funkcije, metabolizma glukoze i hroničnog zapaljenja, funkcionalnosti hemostaznog sistema, trombofilnih markera i genskog polimorfizma vr&scaron;eno je unutar dve nedelje pre hirur&scaron;kog kreiranja AAVF. Kriterijum za utvrđivanje ishoda uticaja antitrombozne terpije predstavlja maturacija AVF koja je definisana kao uspe&scaron;na ako je započeto sprovođenje efikasne hemodijalize najranije 6 nedelja nakon njenog hirur&scaron;kog kreiranja po proceni nadležnog nefrologa. Dijagnoza prisustva tromboze AVF postavljena je od strane nadležnog vaskularnog hirurga/nefrologa fizikalnim pregledom tokom njene maturacije, koji je podrazumevao inspekciju, palpatorno utvrđivanje odsustva karakterističnog trila i auskultatornih karakteristika protočnosti AVF ili ultarsonografskim pregledom od strane radiologa. REZULTATI: Između ispitivanih grupa u odnosu na broj tromboziranih/ afunkcionalnih AVF tokom njene maturacije (12/40 vs. 4/42 vs. 5/39; P=0.033), ustanovljena je značajna statistička razlika kao i poređenjem broja tromboziranih/afunkcionalnih AVF tokom sazrevanja u kontrolnoj grupi u odnosu na grupu ispitanika (objedinjene Grupe II i Grupa III) koja je primala antitromboznu profilaksu (12/40 vs. 9/81; P=0.009). Daljom analizom ispitivanih grupa, utvrđena je statistički značajna razlika u broju tromboziranih/afunkcionih AV fistula između kontrolne Grupe I i Grupe II (P=0.019). Testiranjem razlike u broju tromboziranih/ afunkcionalnih AVF između ispitanika kontrolne Grupe I i Grupe III nije dobijena statistički značajna razlika, kao ni između Grupe II i Grupe III. Zastupljenost broja tromboziranih/afunkcionalnih distalnih AVF za vreme njihove maturacije (12/33 vs 2/31 vs. 3/24; P=0.008) se između ispitivanih grupa značajno statistički razlikovala kao i zastupljenost tromboziranih/afunkcionalnih distalnih AVF tokom sazrevanja u kontrolnoj grupi u odnosu na grupu ispitanika koja je primala antitromboznu profilaksu (12/34 vs. 5/55; P=0.002). Testiranjem statističke razlike u broju tromboziranih/afunkcionalnih distalnih AVF između ispitanika kontrolne Grupe I i Grupe II utvrđena je statistički značajna razlika (P=0.005), dok između Grupe I i Grupe III (P=0.051), kao ni između Grupe II i Grupe III (P=0.439) nije dobijena statistički značajna razlika. Između podgrupa ispitanika kod kojih je do&scaron;lo do tromboze/afunkcionalnosti AVF 21/121 (17.35%) i podgrupe ispitanika sa funkcionalno maturiranom AVF 90/121 (82.64%), značajna statistička razlika ispitanih hemostaznih parametara je bila prisutna u vrednostima agregabilnosti trombocita uz kolagen kao induktor (59.33&plusmn;33.1 vs. 75.04&plusmn;29.6; P=0.033). Značajna statistička razlika je zabeležena i u zastupljenosti sledećih trombofilnih markera: deficita PC (3/21 vs. 3/100; P=0.030), APC-R (4/21 vs. 5/100; P=0.026), prisustva antifosfolipidnih ACL IgM antitela (1/21 vs. 0/100; P=0.028), heterozigotnog polimorfizma FV G1691A (3/21 vs. 3/100; P=0.03) i homozigotne mutacije gena FII G20210A (1/21 vs. 0/100; P=0.028), između podgrupa bolesnika sa tromboziranom afunkcionalnom i funkcionalnom AVF. Takođe je značajna statistička razlika između podgrupa bolesnika kod kojih je do&scaron;lo tromboze/afunkcionalnosti AVF i podgrupe ispitanika sa funkcionalno maturiranom AVF bila prisutna u odnosu na postojanje ranijih tromboza (23/21 vs 19/100; P=0.000) kao i zastupljenosti izolovanih venskih tromboza (9/21 vs. 2/100; P=0.000). Prediktivni potencijal pojedinačnih parametara za maturaciju AVF ispitan je univarijantnom logističkom regresionom analizom. Prilikom ispitivanja uticaja pojedinačnih parametara na maturaciju fistule, zapazili smo da su ispitanici koji su primali antitromboznu terapiju imali 3 puta veću &scaron;ansu za funkcionalno maturiranu AVF [OR 3.45 (1.3-9.03)] u odnosu na bolesnike bez terapije. Ispitanici koji su imali prethodne tromboze su imali vi&scaron;estruko povi&scaron;en rizik [OR 6.92 (2.51-19.06)] za nastanak tromboze/afunkcionalnost AVF tokom maturacije. Prilikom ispitivanja uticaja pojedinačnih parametara na rizik od pojave tromboze/afunkcionalnosti distalne AVF, zapažamo da primena antitrombozne terapije [OR 5.4 (CI 1.7 - 17.35)] petostruko snižava rizik za nastanak tromboze/ afunkcionalnosti distalne AVF, odnosno da primena antitrombozne terapije petostruko povećava &scaron;ansu za adekvatnu maturaciju distalne AVF. Ispitanici koji su imali aterosklerotske KVB [OR 0.32 (0.1-0.98)] i ranije tromboze [OR 0.14 (0.04-0.44)] su imali za 68% i 86% manju verovatnoću za adekvatnu maturaciju distalne AVF (334). Trombofilija je bila prisutna u 59/121 (48.8%) ispitanika. U odnosu na markere aktivacije koagulacione komponente hemostaznog sistema i inflamatorne pokazatelje, između podgrupa ispitanika sa ili bez trombofilije statistički značajna razlika je bila prisutna u vrednostima koncentracije FVIII (170.35&plusmn;103.97 vs. 235.26&plusmn;124.80; P=0.02) i odnosa trombociti/limfociti (181&plusmn;64.58 vs. 148.11&plusmn;66.15; P=0.026). U odnosu na lokalizaciju AVF, u podgrupi ispitanika sa trombofilijom i tromboziranom/ afunkcionalnom AVF, njih 8/11 su pripadale distalnim AVF, 3/11 proksimalnim AVF, dok je u podgrupi ispitanika bez trombofilije i tromboziranom/afunkcionalnom AVF, njih 9/10 imalo distalnu, a 1/10 proksimalnu AVF. U grupi bolesnika sa trombofilijom nije zabeleženo prisustvo statistički značajne razlike u efikasnosti primenjenih antitromboznih režima merene učestalo&scaron;ću tromboza/afunkcionalnosti AVF u odnosu na bolesnike sa trombofilijom koji nisu primali antitromboznu terapiju (5/19 vs. 2/18 vs. 4/22; P=0.493). U grupi ispitanika bez trombofilije utvrđeno je postojanje statistički značajne razlike u učestalosti tromboza/afunkcionalnosti AVF između grupe sa i bez primene antitromboznih lekova kako u ukupnom broju tromboziranih/afunkcionalnih AVF (7/21 vs. 2/24 vs. 1/17; P=0.030). Iako je zastupljenost tromboza/afunkcionalnosti AVF u bolesnika sa kombinovanom trombofilijom če&scaron;ća u odnosu na ispitanike koji su imali drugu vrstu ili uop&scaron;te nisu imali trombofiliju (6/18 vs. 15/103; P=0.052), ona nije dostigla statistički značajnu vrednost. ZAKLJUČAK: Profilaktička primena antitromboznih lekova (tiklopidina i nadroparin-kalcijuma) smanjuje učestalost pojave rane tromboze i pojavu primarne nefunkcionalnosti AVF za hemodijalizu tokom njene maturacije. Primena antitrombozne terapije petostruko snižava rizik za nastanak tromboze/ afunkcionalnosti distalne AVF tokom njene maturacije. Bolesnici koji su imali prethodne tromboze imaju vi&scaron;estruko povi&scaron;en rizik za nastanak tromboze AVF tokom njene maturacije. Kod bolesnika koji su imali aterosklerotske KVB i ranije tromboze verovatnoća za adekvatnu maturaciju distalne AVF je niža za 68% , odnosno 86%. U na&scaron;em istraživanju nije utvrđeno postojanje superiornosti antikoagulantne u odnosu na antitrombocitnu profilaksu tj. oba primenjena režima su bila podjednako efikasna. U terminalnoj bubrežnoj insuficijenciji prisutan je značajan poremećaj funkcionalnosti hemostaznog sistema koji se očituje u disfunkciji endotela i poremećenoj (sniženoj) funkcionalnosti trombocita, prisustvu prokoagulantnog stanja koje se manifestuje povi&scaron;enom trombinskom aktivno&scaron;ću, povi&scaron;enom koncentracijom činilaca koagulacije i smanjenom fibrinoliznom aktivno&scaron;ću. Če&scaron;ća zastupljenost ukupnih ranijih tromboza (arterijskih i venskih), če&scaron;ća zastupljenost izolovanih venskih tromboza i učestalije prisustvo trombofilije prezentovano deficitom PC, prisustvom rezistencije na APC, prisusustvom antifosfolipidnih antikardiolipinskih antitela IgM, heterozigotnog polimorfizma FV G1691A, homozigotne mutacije FII G201210A i niža vrednost agregabilnosti trombocita uz kolagen kao induktor su markeri koji su u na&scaron;em ispitivanju signifikantno če&scaron;će zastupljeni kod ispitanika sa trombozom/ afunkcijom AVF za hemodijalizu tokom njenog sazrevanja. Trombofilija je prisutna kod 48.8% bolesnika saTBI, ali na&scaron;im ispitivanjem nije utvrđen njen uticaj na nastanak rane tromboze/afunkcionalnosti AVF izuzev u grupi bolesnika sa kombinovanom trombofilijom. Mali broj krvarećih komplikacija u na&scaron;oj studiji ukazuje na bezbednost primenjenog preventivnog režima. Na osnovu dobijenih rezultata može se preporučiti profilaktička primena tiklopidina ili nadroparin-kalcijuma u preventivnim dozama kod bolesnika sa TBI neposredno nakon kreiranja AVF. Primenu profilakse tromboznih komplikacija kod bolesnika sa novokreiranom AVF preporučujemo posebno kod bolesnika koji su imali prethodne tromboze i/ili kliničke manifestacije aterosklerotskih kardiovaskularnih bolesti.</p> / <p>INTRODUCTION: Complications in end stage renal disease (ESRD) when the glomerular filtration rate (GFR) decreases below 10mL/min can only be treated by chronic dialysis or kidney transplant ie. total or partial renal replacement therapy. With prompt education of the patient regarding the progressive course of the chronic kidney disease, possibilities of dialysis treatment and kidney transplantation, the patient should timely be granted permanent functional vascular hemodialysis (HD) access through surgical intervention by creating arteriovenous fistula (AVF), preferably at least 6 months prior to the anticipated start of HD, as period for its maturation is between 4 and 6 weeks. Primary AVF is the generally best recommended permanent vascular access for patients planned for dialysis. The most common reason for dysfunction of the vascular access for hemodialysis are thrombotic complications in 80% of the cases, 90% of which appear in the venous segment of AVF as the consequence of progressive venous neointimal hyperplasia. Beside the histological characteristics of the venous blood vessel wall and hemodynamic conditions, in the etiopathogenesis of this &ldquo;adaptive answer&rdquo;, endothel and other components of the hemostatic system (platelet, coagulation and fibrinolysis), immunological and cytological components as well as genetic factors play a very important role. Prevention of occurrence of early thrombosis of vascular access for hemodialysis in patients with ESRD is possible by treatment with antithrombotic drugs, ie. antiplatelet or anticoagulant therapy. OBJECTIVE: Estimate the efficiency of applied antithrombotic drugs (ticlopidine and nadroparincalcium) in prevention of occurrence of early thrombosis/dysfunction of AVF for hemodialysis during its time of maturation within the 6 week period. Examine the level of biomarkers of the hemostatic system and thrombophilic markers in patients with ESRD before the creation of AVF with the goal of finding additional causes of occurrence of early thrombosis/dysfunction of arteriovenous fistula for hemodialysis. Determine the incidence of thrombophilia and its impact on the functionality of AVF and compare the efficiency of applied preventive regimen between patients with and without thrombophilia. MATERIAL AND METHODS: The study included persons of both sexes with previously established diagnosis of ESRD in which there were no contraindications for the planned surgical creation of the first permanent vascular access for hemodialysis in the form of autologous arteriovenous fistula (AAVF). After the surgical creation of the radiocephalic arteriovenous fistula in the distal third of the forearm of the non-dominant hand (89/121), intermedial (4/121) or proximal (28/121) AAVF, the total number of 121 patients were included in the study and divided into three groups in order to estimate the influence of different antithrombotic drugs in prevention of early thrombosis for hemodialysis in patients with ESRD: Group I, control; 40 subjects which did not receive antithrombotic therapy after the creation of AVF, Group II; 42 subjects which started receiving an antithrombotic drug from the tienopiridine group, Ticlodix&reg; (ticlopidine) 2 x &frac12; of 250mg tbl, daily, during the period of 6 weeks, after the creation of AVF, and Group III; 39 subjects which started subcutaneously receiving a drug from the low-molecular weight herapin group, Fraxiparine&reg; (nadroparine-calcium) 2850 anti Xa i.j. (0.3 ml) daily, during the period of 6 weeks. One-time determination of laboratory parameters and renal function, glucose metabolism and chronic inflammation, hemostatic system functionality, thrombophilic markers and gene polymorphism was performed within two weeks prior to surgical creation of AAVF. The criteria for determining the outcome of the impact of antithrombotic therapy is the maturation of AVF, which is defined as successful if the implementation of effective hemodialysis started at least 6 weeks after its creation, where the effectiveness of hemodialysis is estimated by a competent nephrologist. The diagnosis of the presence of AVF thrombosis was set by a competent vascular surgeon/nephrologist through physical examination during its maturation, which included inspection, palpatory determination of absence of the characteristic thrill and auscultatory characteristics of the flow of AVF, or by ultrasonographic examination by the radiologist. RESULTS: Between the groups in terms of number of thrombosed/dysfunctional AVF during its maturation (12/40 vs. 4/42 vs. 5/39, P = 0.033), a significant statistical difference was established, as well as by comparing the number of thrombosed/dysfunctional AVF during maturation in the control group compared to the group of respondents (unified Group II and Group III) which received antithrombotic prophylaxis (12/40 vs. 9/81, P = 0.009). Through further analysis of the examined groups, a statistically significant difference was observed in the number of thrombosed/dysfunctional AV fistula between the control Group I and Group II (P = 0.019). There was no statistically significant difference noticed in the numbers of thrombosed/dysfunctional AVF between the subjects in the control Group I and Group III, as well as between Group II and Group III. Presence of the number of thrombosed/dysfunctional distal AVF during their maturation (12/33 vs 2/31 vs. 3/24, P = 0.008) between the groups statistically significantly varied, as well as the presence of the number of thrombosed/dysfunctional distal AVF during the maturation in the control group as compared to the group of subjects who received antithrombotic prophylaxis (12/34 vs. 5/55; P=0.002). By testing statistical differences in the number of thrombosed/dysfunctional distal AVF between the subjects in the control Group I and Group II a statistically significant difference (P = 0.005) was established, while there was no statistically significant difference between Group I and Group III (P = 0.051), nor between Group II and Group III (P = 0.439). Among the subgroup of patients with thrombosis/dysfunction of AVF 21/121 (17.35%) and the subgroup of subjects with functionally maturated AVF 90/121 (82.64%), a statistically significant difference of the examined hemostasis parameters was present in the values of platelet aggregation with collagen as the inducer (59.33 &plusmn; 75.04 vs. 33.1 &plusmn; 29.6; P = 0.033). A significant statistical difference was recorded in the presence of the following thrombophilic markers: deficit of PC (3/21 vs. 3/100; P = 0.030), APC-R (4/21 vs. 5/100; P = 0.026), the presence of antiphospholipid ACL IgM antibodies ( 1/21 vs. 0/100; P = 0.028), heterozygous FV G1691A polymorphism (3/21 vs. 3/100; P = 0.03) and homozygous gene mutation FII G20210A (1/21 vs. 0/100; P = 0.028), between the subgroups of patients with thrombosed/dysfunctional and functional AVF. There also was a significant statistical difference between the groups of patients which encountered thrombosis/dysfunction of AVF and subgroups of subjects with functional maturated AVF in relation to the existence of previous thrombosis (23/21 vs. 19/100; P = 0.000) and the presence of isolated venous thrombosis (9/21 vs. 2/100; P = 0.000). Predictive potential of individual parameters for AVF maturation was tested by univariate logistic regression analysis. During the examination of the influence of individual parameters on fistula maturation, we observed that subjects who received antithrombotic therapy were 3 times more likely to develop functionally maturated AVF [OR 3.45 (1.3-9.03)] as compared to subjects who did not receive any treatment. Subjects which previously had thrombosis had a multiple times increased risk [OR 6.92 (2:51 to 19:06)] of developing thrombosis/dysfunctional AVF during its maturation. When examining the influence of individual parameters on the risk of thrombosis/dysfunction of the distal AVF, we noted that the implementation of antithrombotic therapy [OR 5.4 (CI 1.7 - 17:35)] reduced risk of thrombosis/dysfunction of the distal AVF by five times, ie. that the implementation of antithrombotic therapy increases the chance for adequate distal AVF maturation by five times. The subjects that had atherosclerotic cardiovascular diseases (CVD) [OR 0.32 (0.1-0.98)] or previous thrombosis [OR 0.14 (0.04-00.44)] had a 68% or 86% less chance for adequate distal AVF maturation (334). Thrombophilia was present in 59/121 (48.8%) patients. In relation to the markers of activation of coagulation components of the hemostatic system and inflammatory markers, among subgroups of subjects with or without thrombophilia a statistically significant difference was present in the FVIII concentration (170.35 &plusmn; 103.97 vs. 235.26 &plusmn; 124.80; P = 0.02) and the platelets/lymphocytes ratio (181 &plusmn; 64.58 vs. 148.11 &plusmn; 66.15; P = 0.026). In relation to the localization of AVF, in the subgroup of subjects with thrombophilia and thrombosed/dysfunctional AVF, 8/11 of them belonged to distal AVF, 3/11 proximal AVF, while in the subgroup of subjects without thrombophilia and thrombosed/dysfunctional AVF, had 9/10 distal and 1/10 proximal AVF. In the group of subjects with thrombophilia there was no record of the presence of statistically significant differences in the efficiency of antithrombotic regimen which was measured by the frequency of thrombosis/dysfunction of AVF as compared to subjects with thrombophilia which did not receive antithrombotic therapy (5/19 vs. 2/18 vs. 4/22, P = 0.493). In the group of subjects without thrombophilia statistically significant differences were found in the frequency of thrombosis/dysfunctions of AVF among groups with and without the use of antithrombotic drugs in the total number of thrombosed/dysfunctional AVF (7/21 vs. 2/24 vs. 1/17, P = 0.030). Although the presence of thrombosis/dysfunction of AVF in patients with combined thrombophilia was more frequent compared to those who had other types of, or did not have thrombophilia (6/18 vs. 15/103; P = 0.052), it did not reach a statistically significant value. CONCLUSION: Prophylactic use of antithrombotic drugs (ticlopidine and nadroparin-calcium) reduces the incidence of early thrombosis and the occurrence of primary AVF dysfunction for hemodialysis during its maturation. Implementation of antithrombotic therapy reduced risk of thrombosis/ dysfunction of the distal AVF during its maturation by five times. Patients who have had previous thrombosis have multiple times greater risk of AVF thrombosis during its maturation. In patients who had atherosclerotic CVD or previous thrombosis, the probability for adequate maturation of distal AVF is lower by 68% or 86%. In our study there was no evidence of superiority of anticoagulant compared to antiplatelet prophylaxis ie. both regimens were equally effective. In ESRD there is significant disarrangement of hemostatic system functionality, which is reflected in endothelial dysfunction and disturbed (reduced) platelet functionality, the presence of procoagulant condition that is manifested by elevated thrombin activity, increased levels of clotting factors and reduced fibrinolytic activity. More frequent presence of total previous thrombosis (arterial and venous), higher frequency of isolated venous thrombosis and frequent presence of thrombophilia presented by the deficit of PC, the presence of resistance to APC, presence of anticardiolipin antiphospholipid antibodies IgM, heterozygous FV G1691A polymorphism, homozygous mutation FII G201210A and lower value of collagen induced platelet aggregation are the markers in our study which are significantly more frequent in patients with thrombosis/dysfunction of AVF for hemodialysis during its maturation. Thrombophilia is present in 48.8% of patients with ESRD, however our study does not determine its impact on early thrombosis/dysfunction of AVF except in the group of patients with combined thrombophilia. A small number of bleeding complications in our study points to the safety of the applied preventive regimen. Based on the obtained results, prophylactic use of ticlopidine or nadroparin-calcium in preventive doses can be recommended for patients with ESRD immediately after AVF creation. Prophylactic treatment of thrombotic complications in patients with newly created AVF is recommended especially in patients who have had previous thrombosis and/or clinical manifestations of atherosclerotic cardiovascular diseases.</p>

Anticoagulants are the basis for treatment and prevention of thrombotic diseases. The currently available medicines are associated with a wide range of adverse reactions that mandates developing new anticoagulants. Several lines of evidence support the superiority of factor Xa (FXa) as a promising target to develop novel anticoagulants. This work focuses on the design of direct and indirect FXa inhibitors using an interdisciplinary approach. As indirect FXa inhibitors, a focused library of tetrasulfated N–arylacyl tetrahydroisoquinoline (THIQ) nonsaccharide allosteric antithrombin activators was designed, synthesized, and biochemically evaluated to establish their structure–activity relationship (SAR). An N–arylacyl THIQ analog having carboxylate at position–3, two sulfate groups at positions–5 and –8 of THIQ moiety, butanoyl linker, and two sulfate groups at positions–2 and –5 of the phenolic monocyclic moiety was identified as the most promising nonsaccharide antithrombin activator with KD of 1322 ± 237 μM and acceleration potential of 80–fold. Its biochemical profile indicates a strong possibility that it activates antithrombin by the pre–equilibrium pathway rather than the induced–fit mechanism utilized by heparin analogs. A similar interdisciplinary approach was exploited to design direct FXa inhibitors that possess high selectivity and are potentially orally bioavailable. Structurally, the designed direct FXa inhibitors are neutral THIQ dicarboxamides. THIQ dicarboxamide is a privileged structure with a semi–rigid character, a structural feature that potentially offers high selectivity for targeting FXa over other coagulation and digestive proteases. It can also be thought of as an amino acid–like structure, which affords accessibility to a large number of compounds using well established peptide chemistry. Mechanistically, the designed inhibitors were expected to bind to FXa in the active site and function as orthosteric inhibitors. These direct FXa active site inhibitors are also likely to inhibit clot–bound enzyme. Nearly 60 THIQ dicarboxamides were synthesized and biochemically evaluated. Through detailed SAR analysis, the most potent analog was designed and found to exhibit an IC50 of 270 nM (Ki = 135 nM), an improvement of more than 207–fold over the first inhibitor synthesized in the study. The most potent inhibitor displayed at least 1887–fold selectivity for FXa over other coagulation enzymes and a selectivity index of at least 279–fold over the digestive serine proteases. This analog doubled plasma clotting times at 17–20 μM, which are comparable to those of agents being currently studied in clinical trials. Overall, allosteric and orthosteric approaches led to the design of indirect and direct small molecule inhibitors of FXa based on the THIQ scaffold. This work introduces two promising molecules, a tetrasulfated N–arylacyl THIQ analog as a heparin mimetic and a neutral THIQ dicarboxamide as a potent, selective, and potentially bioavailable peptidomimetic, for further advanced medicinal chemistry studies.