Low borderline plasma levels of antithrombin,protein C and protein S are risk factors for venous thromboembolism

Summary. Background: Inherited deficiencies of antithrombin (AT), protein C (PC) and protein S (PS) are risk factors for venous thromboembolism (VTE). They are usually defined by laboratory cut‐offs (in our setting 81, 70 and 63 IU dL^?1, respectively), which give only a rough idea of the VTE risk associated with plasma levels of these proteins. Objectives: We investigated whether the risk of VTE associated with the plasma deficiencies of AT, PC or PS has a dose–response effect, and whether low borderline levels of these proteins are associated with an increased risk of VTE, both in the whole study population and separately in carriers of either factor V Leiden or G20210A prothrombin gene mutation. Patients/Methods: A case–control study of 1401 patients with a first objectively‐documented VTE and 1847 healthy controls has been carried out. Results: A dose–response effect on the VTE risk was observed for all the three anticoagulant proteins. Compared with individuals with AT, PC or PS levels > 100 IU/dL, the adjusted odds ratio (95%CI) of VTE was 2.00 (1.44–2.78) for AT levels between 76 and 85 IUdL^?1, 2.21 (1.54–3.18) and 1.84 (1.31–2.59) for PC and PS levels between 61 and 75 IUdL^?1. The risk of unprovoked VTE in factor V Leiden or prothrombin G20210A carriers appears 2 to 3‐fold increased when levels of AT or PS are low borderline. Conclusions: Low borderline plasma levels of AT, PC and PS are associated with a 2‐fold increased risk of VTE and should be considered in the assessment of the individual VTE risk.     

Use of antithrombin III in critical patients

Objective To evaluate the effect of the AT III concentrates upon the clinical evolution and hemostatic parameters.Design Prospective, open, randomized trial.Patients and participants Septic and multiple trauma patients admitted to our Intensive Care Unit.Setting Levels of AT III below 70% were used as criteria to choose 36 patients, 20 of whom received treatment with AT III and 16 did not.Interventions AT III concentrates were administered at an initial dose of 60 U/kg followed by 10 U/kg every six hours.Results The administration of AT III neither contributes to alterations in haemostasis, nor the clinical evolution (evaluated according to Apache II score).Conclusions The results suggest that the administration of AT III concentrates to critical patients with acquired low levels, but without manifest DIC, may not be justified; although further studies on a larger population are required to establish definite conclussions.     

Antithrombin III in patients with severe sepsis: a pharmacokinetic study

Objectives: To evaluate the safety, pharmacokinetics, and the practicability of two different antithrombin III (AT III) high-dose regimens in patients with severe sepsis.¶Design: Prospective, open, randomized, 2 parallel groups, multinational clinical trial.¶Setting: Eleven academic medical center intensive care units (ICU) in Austria, Belgium, Denmark, Germany, Norway and Sweden.¶Patients: Thirty-three patients with severe sepsis who received standard supportive care and antimicrobial therapy, in addition to the administration of AT III.¶Interventions: Patients received an intravenous loading dose of 6,000 IU AT III followed by either intermittent bolus infusions of 1,000 IU AT III every 4 h or a continuous infusion of 250 IU AT III/h for 4 days, resulting in a total dose for both dosage regimens of 30,000 IU AT III.¶Measurements: All patients were evaluated for safety and all but one for pharmacokinetics.¶Results and conclusions: The administration of AT III was safe and well tolerated. The overall 28-day all-cause mortality was 30 % (43 % intermittent bolus infusions; 21 % continuous infusion). The mean probability of dying according to the SAPS II was 48 %. The difference in mortality between both groups was within the range of chance. AT III plasma levels were elevated from low baseline levels to above 120 % soon after onset of AT III therapy and remained at these levels for the treatment phase of 4 days. Functional and immunologic levels of AT III corresponded very well. With an overall median volume of distribution of 4.5 l (range: 2.4–6.5 l), AT III only moderately extended beyond plasma. The overall median elimination half-life was 18.6 h (range: 5.1–37.4). Overall, median response was 1.75 % per IU/kg (range: 1.14–2.8).¶The variability of elimination parameters was quite noteworthy (CV = 41–59 %), whereas distribution-related parameters showed a moderate variability (CV = 24 %). In spite of this variability, both high-dose IV regimens reliably provided AT III levels above 120 % for all but one patient. An increased mortality was observed for patients with a distribution volume exceeding 4.5 l (or a response < 1.7 % per IU/kg). AT III distribution volumes above 4.5 l might indicate a capillary leak phenomenon. The continuous infusion regimen was slightly preferred by the investigators with regard to practicability.     

von Willebrand factor/factor VIII concentrate (Haemate® P) dosing based on pharmacokinetics: a prospective multicenter trial in elective surgery

BACKGROUND: While plasma-derived concentrates containing large amounts of von Willebrand factor (VWF) are effective in treating von Willebrand disease (VWD), optimal dosing remains to be fully characterized. OBJECTIVES: To determine the feasibility of dosing Haemate P VWF/factor VIII (FVIII) concentrate based on pharmacokinetics (PK) in the management of surgical subjects with VWD. METHODS: VWD subjects scheduled for elective surgery were enrolled in a prospective multicenter open-label cohort study. A pre-operative loading dose of VWF/FVIII concentrate based upon prior individual subject PK analysis was administered followed by postoperative therapeutic/maintenance infusions. RESULTS: Twenty-eight subjects with types 1, 2A or 3 VWD and one with type 2 M were enrolled. Median in vivo recovery of VWF ristocetin cofactor (VWF:RCo) was 1.9 IU dL(-1) (IU kg(-1))(-1) with an interquartile range (IQR) of 1.6-2.5 IU dL(-1) (IU kg(-1))(-1). Median response, half-life and clearance were 74.0% (IQR, 55.5-100%), 15.6 h (IQR, 9.0-28.4 h) and 3.26 mL kg(-1) h(-1) (IQR, 2.29-5.21 mL kg(-1) h(-1)), respectively. A PK-guided median VWF:RCo loading dose of 62.4 IU kg(-1) (IQR, 50.1-87.0 IU kg(-1)) was administered. Postoperative mean trough VWF:RCo levels of 62-73 IU dL(-1) were sufficient to prevent bleeding. Investigators rated hemostasis excellent or good in 96.3% of subjects on the day of surgery and 100% on the next day and on day 14. A subject with multiple risk factors developed pulmonary embolism, which resolved without sequelae. CONCLUSIONS: Haemate P provided effective and safe hemostasis in VWD subjects undergoing elective surgery. Selection of Haemate P loading dose on the basis of VWF PK proved feasible.     

Corifollitropin alfa for female infertility

INTRODUCTION: Follicle-stimulating hormone (FSH) is essential for the development of ovarian follicles. Urinary-derived and recombinant FSH (rFSH) preparations are widely used in infertility treatment but have to be administered daily to achieve steady-state serum levels. Corifollitropin alfa is a hybrid molecule with a prolonged half-life. AREAS COVERED: The development and clinical testing of corifollitropin alfa, including the pharmacodynamics and kinetics, efficacy and drug safety. Searches were performed using the Medline database. EXPERT OPINION: Corifollitropin alfa is composed of the FSH α-subunit and a hybrid of the FSH β-subunit and the C-terminal peptide (CTP) of the human chorionic gonadotropin (hCG) β-subunit. The rationale of developing such a molecule was to reduce patient burden, by reducing the number of injections required to sustain multifollicular growth. Two strengths of corifollitropin are available (for patients ≤ 60 kg and > 60 kg). Compared with a daily dose of 200 IU of rFSH, 150 mcg of corifollitropin is equivalent in safety and pregnancy outcomes in women > 60 and < 90 kg using an antagonist protocol. Another RCT in women ≤ 60 kg also confirmed safety and efficacy of follicular stimulation (100 mcg of corifollitropin versus 150 IU of rFSH), but it was not powered to demonstrate equivalence in terms of pregnancy rates.     

Pharmacokinetics and pharmacodynamics of epoetin delta in two studies in healthy volunteers and two studies in patients with chronic kidney disease

BACKGROUND: Epoetin delta, unlike recombinant erythropoietins, is produced in a human cell line and therefore has a human-type glycosylation profile. OBJECTIVES: The pharmacokinetics of epoetin delta were examined in 2 studies in healthy volunteers and 2 studies in patients with chronic kidney disease. METHODS: In study 1, 21 healthy men were randomized to receive epoetin delta 15, 40, or 100 IU/kg IV tiw or placebo for 4 weeks. In study 2, an open-label, cross-over study, 32 healthy volunteers were randomized to receive single doses of epoetin delta 75 IU/kg IV or SC. In study 3, 40 patients receiving hemodialysis were withdrawn from epoetin alfa and randomized to receive epoetin delta or epoetin alfa 50 or 100 IU/kg tiw for 4 weeks. Study 4 was a single-dose study comparing epoetin delta 150 and 300 IU/kg IV or SC in 28 hemodialysis patients. RESULTS: In study 1, after repeated dosing (day 24) in healthy men, mean C(max) values ranged from 219.9 to 1793.0 enzyme-linked immunosorbent assay units (EU)/L; AUC from 827 to 9318 h x EU/L; C1 from 0.014 to 0.024 L/h per kg; Vd from 0.067 to 0.076 L/kg; and t(1/2) from 2.23 to 3.35 hours. There was evidence of a dose-dependent effect of epoetin delta on hemoglobin levels and hematocrit, with doses of 40 and 100 IU/kg associated with significant increases compared with 15 IU/kg (P < 0.001 for dose trend). The only adverse event occurring in > or = 10% of healthy individuals in study 1 was headache (1 [20.0%] in the epoetin delta 15 IU-kg group, 3 [60.0%] in the epoetin delta 100-IU/kg group, 2 [33.3%] in the placebo group). In study 2 in healthy volunteers, mean values for epoetin delta 75 IU/kg IV were 1771 EU/L for C(max), 10,632 h x EU/L for AUC, 0.010 L/h per kg for Cl, 0.074 L/kg for Vd, and 5.12 hours for t(1/2); the corresponding values for epoetin delta 75 IU/kg SC were 113 EU/L, 3231 h x EU/L, 0.035 L/h per kg, 0.760 L/kg, and 14.90 hours. The serum epoetin delta concentration peaked after 10.9 hours with subcutaneous administration. The most common adverse event in study 2 was back pain (10 [31.3%] individuals). In study 3 in patients receiving hemodialysis, mean values for C(max) and AUC with a single dose of epoetin delta 50 IU/kg were 1103 EU/L and 10,896 h x EU/L, respectively, and with the corresponding dose of epoetin alfa were 1354 EU/L and 9957 h x EU/L. Values for the 100-IU/kg doses were approximately double those for the 50-IU/kg doses. Values for Cl, Vd, and t(1/2) were numerically similar for epoetin delta and epoietin alfa across doses. Epoetin delta 100 IU/kg was associated with a numerically greater rate of increase in hemoglobin compared with the 50-IU/kg dose (mean, 0.025 vs -0.004, respectively); the results were similar for epoetin alfa (0.029 vs -0.001). The difference between epoetin alfa and epoetin delta was not statistically significant. The most common adverse events were related to edema (peripheral edema: 60%/50% for epoetin delta 50/100 IU/kg and 60%/60% for epoetin alfa 50/100 IU/kg; facial edema: 30%/30% and 50%/70%, respectively; generalized edema: 50%/30% and 40%/40%). In study 4 in patients receiving hemodialysis, mean C(max) values with epoetin delta 150 and 300 IU/kg IV were 3257 and 4770 EU/L, respectively; the corresponding mean values were 36,208 and 77,736 h x EU/L for AUC, 0.007 and 0.005 L/h per kg for Cl; 0.097 L/kg for Vd in both groups; and 9.9 and 13.2 hours for t(1/2). With epoetin delta 150 and 300 IU/kg SC, the respective values were 162.2 and 467.7 EU/L, 9547 and 27,888 h x EU/L, 0.026 and 0.020 L/h per kg, 1.28 and 0.78 L/kg, and 33.1 and 27.8 hours. The only adverse event occurring in > or = 10% of subjects was headache (2 [40.0%] in the epoetin delta 150-IU/kg IV group, 3 [50.0%] in the epoetin delta 300-IU/kg SC group). No neutralizing anti-erythropoietin antibodies were detected in any individual. The bioavailability of subcutaneous epoetin delta is approximately 30%, and concentrations peak later and decline more slowly than with intravenous injection. Pharmacokinetic parameters in hemodialysis patients were similar to those in healthy individuals, although AUC and t(1/2) were numerically higher (by 49% and 34%, respectively). CONCLUSIONS: These studies in healthy volunteers and patients with chronic kidney disease indicate that the pharmacokinetics of epoetin delta are dose dependent but nonlinear, leading to dose-dependent increases in hemoglobin levels. The pharmacodynamic response to epoetin delta appeared to be as expected for an epoetin.     

下肢深静脉血栓形成患者抗凝蛋白缺陷的临床研究

目的：研究中国人群下肢深静脉血栓形成（IDVT）患者抗凝蛋白缺陷的发生率，探讨中国人群LDVT的主要发病机制。方法：应用ACLPutLlm型全自动血凝仪检测1幔）例LDVT患者（73例初发，27例复发）和100例健康人的抗凝血酶（AT）、蛋白S（PS）、蛋白C（PC）活性及活化蛋白C抵抗性（APCR）。结果：LDVT组与正常对照组相比、LDVT复发组与初发组相比，AT、PS、PC活性明显降低，APCR阳性率明显升高，均有极显著差异（P〈0．01）；本组100例LDVT患者中共有25例患者存在有抗凝蛋白缺陷，以PS缺陷的总发生率最高，为13％（13例），其次是PC缺陷，为8％（8例）；AT缺陷占5％（5例），APCR缺陷的总发生率最小，为4％（4例）。结论：先天性或获得性抗凝蛋白缺陷是中国人LDVT发病和复发的重要机制之一，因此有必要对LDVT患者进行抗凝蛋白水平的筛选。     

Immune tolerance induction with high-dose FVIII and pulsed intravenous immunoglobulin

The development of neutralizing allo-antibodies against factor VIII (FVIII) or FVIII inhibitors is a severe complication in the treatment of haemophilia A. About 25% of the children with severe haemophilia A develop FVIII inhibitors. Here we report on a boy with severe haemophilia A and intron 22 inversion of the FVIII gene who was diagnosed at ten months of age. After 16 exposure days to FVIII (81 days after initial exposure) he developed a FVIII inhibitor (maximum: 9.76 BU/ml). Therapy: We started immune tolerance induction (ITI) according to the Bonn protocol with high dose plasma derived FVIII concentrate (100 IU per kg body weight) twice daily. For additional inhibitor elimination treatment the patient received intravenous immunoglobulin (ivIg) at a dose of 1-2 g/kg body weight every 4 to 6 weeks. After start of treatment a rapid decline of the inhibitor level was observed, nevertheless low FVIII inhibitor levels persisted (<5 BU/ml). Furthermore, the FVIII half-life was still accelerated. However, after every course of ivIg the inhibitor level declined and FVIII half-life was prolonged. Currently, the FVIII half-life is approaching normal values after more than seven months of ITI duration. Conclusion: Additional application of immunoglobulin is beneficial for immune tolerance induction.     

血栓性疾病患者抗凝蛋白检测的临床意义

目的 探讨血栓性疾病患者蛋白C(PC)、蛋白S(PS)和抗凝血酶(AT)活性水平检测在排除常见获得性血栓危险因素中的临床意义.方法 检测85例血栓性疾病患者与50名正常健康对照者血浆PC、PS、AT活性水平,并作比较分析.结果 85例血栓性疾病患者中位年龄42(17～69)岁.其中≤45岁者60例(70.6%).动脉血栓组和静脉血栓组患者PC、PS、AT活性均明显低于正常对照组(P<0.01);复发组PC、PS、AT平均活性低于初发组(P<0.01);年龄≤45岁患者组PC、PS、AT平均活性低于45岁以上组(P<0.01).共有26例(30.6%)患者存在抗凝蛋白活性降低;PS活性降低的发生率最高(10.6%),其次为PC活性降低(8.2%),AT活性降低和联合活性降低(各占5.9%).结论 无明确常见获得性血栓危险因素的血栓患者发病年龄较轻,且普遍存在抗凝蛋白水平低下;抗凝蛋白活性降低不仅与血栓性疾病的发生有关,而且与血栓复发密切相关.     

Protein C concentrate to restore physiological values in adult septic patients

OBJECTIVE: To describe the efficacy and safety of protein C (PC) concentrate to restore physiological values in adult septic patients having clinical contraindications to activated PC. DESIGN: Case series (pilot study). SETTING: Three adult ICUs of a University Hospital. PATIENTS AND PARTICIPANTS: Twenty adult patients affected by severe sepsis or septic shock with plasma values of PC < 50%. INTERVENTIONS: Patients were treated with PC concentrate (Ceprotin ((R))--Baxter) with a starting bolus followed by a continuous infusion for 72 h [3 IU/(kg h)]. MEASUREMENTS AND RESULTS: PC activity, WBC, platelets, D: -Dimer, fibrinogen, PT, aPTT, AT III, lactate, Sepsis-related Organ Failure Assessment (SOFA), Disseminated Intravascular Coagulation (DIC) score, adverse events, and mortality were measured. Baseline plasma PC activity was 34.5 +/- 9.1%. PC concentrate normalized the PC activity in all patients within 48 h, and then remained stable for the following days. At baseline, several patients showed abnormal PT, aPTT, platelets values, and lactate levels. During the study period, there was a significant increase of platelets, fibrinogen, PT, AT III, and a significant decrease of D: -Dimer, aPTT, DIC score, and lactate. No adverse reactions (hemorrhage or thrombosis) were observed. Mortality at 28 days was 35%. CONCLUSIONS: Our pilot study shows that the administration of PC concentrate to patients having contraindications to the treatment with activated PC was safe and possibly useful to control the coagulopathy triggered and sustained by sepsis. A randomized, double blind study in patients with severe sepsis and contraindications to activated PC administration would be advisable to state the safety and the possible role of this product in the treatment of severe sepsis.     

不同吸附体系及因素水平对PCC中4类抗凝蛋白的影响研究

目的 探讨不同吸附体系及其柠檬酸钠、氯化钠、pH、电导4因素对凝血酶原复合物(PCC)中蛋白C、蛋白S、蛋白Z、抗凝血酶收率及比活的影响.方法 选择柠檬酸钠、氯化钠、pH3因素3水平设计正交试验,配制9种PCC平衡吸附体系;凝胶平衡后,吸附血浆,洗涤、洗脱,制备PCC浓缩物;测定浓缩物中蛋白C和S、抗凝血酶活性、蛋白Z含量、总体积及蛋白浓度,获得四类抗凝蛋白收率及比活,比较不同吸附体系收率、比活总趋势;利用正交试验直观分析法分析柠檬酸钠、氯化钠、pH对蛋白C、S和Z收率及比活的影响;测定吸附体系电导率,分析电导率与4类抗凝蛋白收率及比活的关系.结果 不同吸附体系4类抗凝蛋白收率及比活有较大差异,抗凝血酶二者均极低.柠檬酸钠对蛋白C收率及比活、蛋白S收率影响较大;pH对蛋白S比活、蛋白Z收率及比活影响极差分别为:0.120、6.847、0.716,高于其他因素;氯化钠对其影响均最小.柠檬酸钠浓度越低,蛋白C收率及比活越高,蛋白S收率则越低,0.015 mol/L水平蛋白Z收率为49.11％,高于其他水平,而蛋白S和Z比活最低;pH越低,蛋白Z收率则越高,pH7.0水平蛋白C和S收率及比活、蛋白Z比活最高;氯化钠浓度0.075 mol/L水平,蛋白C收率及比活、蛋白S和Z比活分别为:29.37％、0.533 IU/mg、0.401 IU/mg、2.362 μg/mg,均高于其他水平,而蛋白S、Z收率最低.不同电导率条件,蛋白C收率及蛋白Z比活变化较大,其中蛋白Z与S收率、蛋白Z与S比活、蛋白C收率与比活变化趋势基本一致;(10.00±0.20) ms/cm条件,蛋白C、S和Z比活较高.结论 不同吸附体系抗凝血酶收率及比活均极低,蛋白C、S和Z收率及比活间无规律性.氯化钠对蛋白C、S和Z收率及比活影响较小,柠檬酸钠、氯化钠、pH不同水平对蛋白C、S和Z收率影响效应无共性,0.01 mol/L柠檬酸钠,0.075 mol/L氯化钠、pH7.0水平蛋白C、S和Z比活最佳.电导率与四类抗凝蛋白收率及比活间无线性关系.     

Vitamin E supplementation in hyperlipidaemic patients: effect of increasing doses on in vitro and in vivo low-density lipoprotein oxidation

BACKGROUND: Vitamin E supplementation is associated with a reduced risk of developing atherosclerotic events; probably because it inhibits low-density lipoprotein (LDL) oxidation, an initial step in atherosclerosis. Metal ion-dependent LDL oxidation is a commonly used method to estimate oxidizability of LDL, but the effect of antioxidant supplementation on the levels of autoantibodies to oxidised LDL (ox-LDL), an in vivo indicator of LDL oxidation, is unknown. DESIGN: This double-blind, placebo-controlled study investigated the susceptibility of LDL to copper induced oxidation and malondialdehyde (MDA) derivatized-LDL (MDA-LDL) in hyperlipidaemic patients on supplements of vitamin E. The vitamin E group (n = 20) took vitamin E 100 IU daily and the dose was doubled at six-weekly intervals to 1600 IU daily. The control group (n = 17) received placebo in the same fashion. Blood samples were obtained at baseline and each subsequent visit to measure vitamin E status and oxidation of LDL. RESULTS: A significant increase in both alpha-tocopherol levels and the lengths of lag phase was seen in the vitamin E group after first week of supplementation (100 IU day-1). This continued to rise in a dose-dependent fashion with a doubling of the lag phase on 1600 IU daily. However, the titre of antibodies to MDA-LDL was not altered. CONCLUSIONS: The results suggest that although regarded as an in vivo marker of LDL oxidation, antibodies to MDA-LDL may not be a suitable measure to evaluate the effect of short-term antioxidant supplementation. The failure of autoantibody titres to fall despite reduced oxidizability of LDL may possibly be attributable to a long half-life of the antibody or, once initiated, a continuous immunological response to ox-LDL contained in atherosclerotic lesions of the arterial wall.     

Toxicity, biological activity, and pharmacokinetics of TXU (anti-CD7)-pokeweed antiviral protein in chimpanzees and adult patients infected with human immunodeficiency virus

The purpose of the present study was to evaluate the toxicity and pharmacokinetics of TXU (anti-CD7)-pokeweed antiviral protein (PAP) in human immunodeficiency virus (HIV)-infected chimpanzees and adult patients. At a total dose of 100 microg/kg, TXU-PAP did not cause severe (grade >/= 3) toxicity in any of the four HIV type 1 (HIV-1)-infected or two healthy chimpanzees. The only side effects were a transient elevation of the liver enzyme alanine aminotransferase between days 2 and 14 without a concomitant rise in total bilirubin levels and a decrease in the serum albumin levels between days 1 and 5 without any concomitant weight gain or peripheral edema. TXU-PAP showed favorable pharmacokinetics in chimpanzees with a plasma elimination half-life of 5.1 to 12.0 h and a systemic clearance of 5.8 to 15.1 ml/h/kg. At 2 months after initiation of the TXU-PAP infusions, the HIV-1 burden was reduced to below-detection levels in three of the four chimpanzees, and in the remaining chimpanzee, the HIV burden was <500 RNA copies/ml at 2 weeks but returned to the pretreatment levels by 2 months. TXU-PAP was well tolerated by HIV-1-infected adult patients who received a single 5 microg/kg i.v. infusion of TXU-PAP. TXU-PAP showed very favorable pharmacokinetics in these patients with a relatively long plasma elimination half-life of 12.4 +/- 1.4 h, a mean residence time of 17.9 +/- 2.0 h, and a slow systemic clearance of 2.7 +/- 0.7 ml/h/kg. Concentrations of TXU-PAP required for effective inhibition of HIV-1 replication in preclinical models were achieved in HIV-1-infected patients at the 5 microg/kg dose level without any adverse reactions, and the mean value for AUC was 3059 +/- 721 ng. h/ml. The 1-h postinfusion plasma samples from TXU-PAP-treated patients showed potent anti-HIV activity in vitro and inhibited the replication of HIV in normal peripheral blood mononuclear cells (PBMCs) even at a 1:100 dilution. Although treatment with TXU-PAP at the 5 microg/kg dose level does not provide sustained therapeutic levels, it was capable of reducing the viral burden in six of six patients evaluated. To our knowledge, this is the first report of a clinical pharmacokinetics study of a PAP immunoconjugate in HIV-infected patients. The favorable long plasma elimination half-life of TXU-PAP in combination with its low toxicity provides the basis for further investigation of TXU-PAP as a potential anti-HIV agent.     

两个遗传性蛋白C缺陷症家系临床表型和基因型变化的研究

目的对两个遗传性蛋白C（PC）缺陷症家系进行临床表型和基因突变检测。方法血浆蛋白C活性（PC：A）和抗原（PC：Ag）分别用发色底物法和ELISA法测定，蛋白S活性（PS：A）和抗凝血酶活性（AT：A）用发色底物法测定。用PCR法对先证者PC基因的9个外显子及其侧翼、内含子序列进行扩增，PCR产物纯化后直接测序，检测其基因突变。仅对先证者家系成员基因突变部位的外显子及其侧翼序列进行PCR扩增和测序。突变位点经限制性内切酶酶切分析或直接测序证实。结果先证者1（Ⅱ7）的PC：A和PC：Ag分别为1．2％和0。基因测序显示，先证者1在PC基因外显子5存在C3135G杂合错义突变，致C（TGC）64W（TGG），同时在外显子7存在T6128G杂合错义突变，致F（TTC）139V（GTC）。家系成员中，先证者的父亲（Ⅰ4）和女儿（Ⅲ3）存在T6128G杂合突变，先证者的舅舅（Ⅱ）存在C3135G杂合突变，先证者丈夫（Ⅱ8）存在外显子76161-6163或6164～6166AAG（K150或K151）杂合缺失，而其女儿（Ⅲ3）亦有此突变。先证者2（Ⅲ1）的PC：A和PC：Ag分别为50．3％、1．9mg／L，基因测序显示其存在K150或K151杂合缺失，该突变遗传自其父亲。2个家系中所有成员在PC基因启动子区中存在-1654C／T、-1641A／G、-1476A／T多态性，先证者2为CC／GG／TT纯合型。限制性内切酶PSp5Ⅱ酶切分析显示T6168G不是多态性。所有成员的PS：A和AT：A均在正常范围。结论复合杂合性PC基因突变（C64W和F139V）是导致先证者1遗传性Ⅰ型PC缺陷症的原因，杂合性Lys150或151缺失突变和PC基因启动子区CC／GG／TI纯合多态性是致先证者2遗传性Ⅰ型PC缺陷症的原因。C64W为国际首次报道，F139V、K150或151缺失突变为国内首次报道。     

Pharmacokinetic studies on Wilfactin®, a von Willebrand factor concentrate with a low factor VIII content treated with three virus-inactivation/removal methods

OBJECTIVE: In order to correct the primary von Willebrand factor (VWF) defect and avoid supra-physiologic plasma levels of factor VIII, a pure VWF concentrate almost devoid of FVIII was developed and used in France since 1989. METHODS: The pharmacokinetic (PK) profile of the most recent version of this concentrate (Wilfactin; LFB, Les Ulis, France), treated with three virus-inactivation/removal methods (solvent/detergent, 35 nm filtration, dry heat treatment), was investigated in 25 patients. Seventeen patients with various types of clinically severe von Willebrand disease (VWD) were included in a crossover, randomized trial carried out in five European centers and comparing Wilfactin with concentrates containing both FVIII and VWF (FVIII/VWF). Eight type 3 VWD patients were included in another trial carried out in six French centers comparing Wilfactin with its previous version (Facteur Willebrand-LFB; LFB) that adopted one virus-inactivation method only. RESULTS: For both the measurements evaluated in this study (VWF antigen, VWF:Ag; and VWF ristocetin co-factor activity, VWF:RCo), Wilfactin had a PK profile similar to that of the FVIII/VWF concentrates and of Facteur Willebrand-LFB. VWF:RCo and VWF:Ag recoveries were 2.1 +/- 0.3 and 1.8 +/- 0.3 per IU kg(-1), respectively, and the half-lives were 12.4 +/- 1.8 and 15.9 +/- 1.5 h. The FVIII synthesis rate was 5.8 +/- 1.0 IU dL(-1) h(-1), with a half-life of 15.8 +/- 2.4 h. CONCLUSION: The PK of VWF and FVIII have not been altered by the three virus-inactivation/removal steps during the manufacturing of Wilfactin.     

两个终止密码子导致的遗传性蛋白C和蛋白S缺陷症

目的:研究1个蛋白C(PC)和1个蛋白S(PS)缺陷症家系的表型诊断和基因特征。方法:PC活性(PC:A)和PS活性(PS:A)用发色底物法测定;PC抗原(PC:Ag)和PS抗原(PS:Ag)用ELISA方法测定。用PCR扩增PC和PS基因各个外显子及其侧翼序列,用直接测序法检测突变点。利用逆转录PCR(RT-PCR)分析PCmRNA水平变化。同时利用蛋白印迹分析血浆中PC含量的变化。结果:先证者1的PC:A为49%,PC:Ag为1.34mg/L,基因检测发现PC基因9号外显子的8831有G→A杂合无义突变,导致Trp372Stop;先证者2的PS:A为29%,PC:Ag为8.3mg/L,14号外显子Gln522(CAG)→Stop(TAG)。结论:G8831A杂合突变引起Trp372Stop,其可导致遗传性PC缺陷症;Gln522Stop可导致遗传性PS缺陷症。     

Kidney-selective prodrugs of 6-mercaptopurine: biochemical basis of the kidney selectivity of S-(6-purinyl)-L-cysteine and metabolism of new analogs in rats

Recently, we have reported that S-(6-purinyl)-L-cysteine (PC) is a kidney-selective prodrug of 6-mercaptopurine. In the present study, the in vivo metabolism of PC and the biochemical basis of its renal selectivity were further investigated. In addition, several PC analogs were synthesized and evaluated as prodrugs of 6-mercaptopurine by determining the concentrations of 6-mercaptopurine and its metabolites, 6-methylmercaptopurine and 6-thiouric acid, in urine after rats were given the analogs. At 30 min after PC treatments, kidney metabolite concentrations were dependent on the PC dose at 40 to 130 mumol/kg and were not increased when a 400 mumol PC/kg dose was given. At the 400 mumol PC/kg dose, metabolite concentrations in the kidneys were higher at 30 min than at 1 or 3 hr, and were nearly 2.5- and 100-fold higher than those in liver and plasma, respectively. Rates of PC in vitro metabolism by liver and kidney cytosolic cysteine conjugate beta-lyases (beta-lyases) were similar, but metabolism by renal mitochondrial beta-lyase occurred at a 3-fold higher rate than the rate obtained with hepatic mitochondrial beta-lyase. When rats were given aminooxyacetic acid (500 mumol/kg) or probenecid (270 mumol/kg) before PC (130 mumol/kg), total kidney metabolite concentrations were reduced by 55 and 36%, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

Nizatidine disposition in subjects with normal and impaired renal function

To test the hypothesis that renal insufficiency alters nizatidine disposition, we determined the pharmacokinetics of nizatidine and its major metabolite after a single oral dose in normal volunteers and patients with various degrees of renal dysfunction, after a single intravenous dose in normal volunteers and patients with severe renal failure and during hemodialysis. After intravenous administration the elimination half-life increased from 1.5 +/- 0.2 hours in normal volunteers to 6.9 +/- 3.3 hours in patients with renal failure. The plasma clearance decreased from 0.59 +/- 0.07 L/kg/hr in normal volunteers to 0.14 +/- 0.02 L/kg/hr in patients with renal failure. Nizatidine bioavailability was nearly 100% in normal volunteers but decreased to 75% in patients with renal failure. The volume of distribution was 1.3 +/- 0.1 L/kg in normal volunteers and was not different in patients with renal failure. Nizatidine protein binding was about 30% in normal and uremic plasma. The drug was not substantially removed by hemodialysis. Patients with creatinine clearances less than 50 ml/min/1.73 m2 should receive 150 mg nizatidine once each evening. Patients with creatinine clearances less than 20 ml/min/1.73 m2 should receive 150 mg nizatidine every other night.     

骨髓增殖性肿瘤患者NGAL、VEGF、PC及PS表达水平及意义研究

目的观察骨髓增殖性肿瘤(MPN)患者中性粒细胞明胶酶相关脂质运载蛋白(NGAL)、血管内皮生长因子(VEGF)、血浆蛋白C(PC)及血浆蛋白S(PS)表达水平,并分析其临床意义。方法回顾性选取2019年1月至2021年8月在沧州市中心医院初诊MPN患者40例、MPN治疗后患者40例,以及健康体检者40例为研究对象,分别作为初诊组、治疗后组、对照组。观察3组研究对象、不同临床特征MPN患者的NGAL、VEGF、PC及PS表达水平,分析影响MPN患者的NGAL、VEGF、PC及PS表达水平的因素。结果初诊组、治疗后组的NGAL[(18.34±2.11)、(10.49±1.87)ng/mL]、VEGF[(239.29±10.45)、(151.34±9.13) pg/mL]水平均显著高于对照组[(2.37±0.45) ng/mL、(78.66±8.35) pg/mL],初诊组、治疗后组的PC[(1 794.35±142.49)、(2 918.21±85.35) ng/mL]、PS[(43 424.64±102.41)、(63 467.47±125.48) ng/mL]水平均显著低于对照组[(3...     

Response of Plasma Histaminase Activity to Small Doses of Heparin in Normal Subjects and Patients with Hyperlipoproteinemia

The release of histaminase activity in plasma after small intravenous of heparin was studied in 85 normal subjects and patients. In normal subjects, plasma histaminase activity (basal level, 1.7+/-0.1 U/ml, mean +/-SEM) increased 1.6+/-0.2 U/ml after 10 U of heparin/kg, 8.5+/-2.4 U/ml after 20 U/kg, and 33+/-4.9 U/ml after 75 U/kg. The extent of the increase varied widely among individuals but in a particular individual the response was constant and dose-dependent. Histaminase activity rose to peak levels within 7-15 min and then declined exponentially with a half-life of 40-120 min. This pattern of response was also observed in two patients with the histaminase-producing tumor, medullary carcinoma of the thyroid. A significantly reduced response was observed, however, in 14 patients with type I hyperlipoproteinemia, a disorder in which high plasma triglyceride levels are associated with low postheparin plasma lipolytic activity. After 10 U heparin/kg, plasma histamine activity increased 0.5+/-0.2 U/ml, and after 75 U heparin/kg, 10.9+/-5.6 U/ml. In contrast, in 27 patients with other types of hyperlipoproteinemia in whom postheparin lipolytic activity was normal, the increase (2.4+/-0.6 U/ml) in plasma histaminase activity after 10 U heparin/kg was not significantly different from that of normal subjects. The reduced response of the plasma histaminase activity to heparin in patients with type I hyperlipoproteinemia did not appear to be due to the presence of lipemia or to an inhibitor of the enzyme in plasma. These findings suggest that many patients with type I hyperlipoproteinemia may have deficient release of both lipolytic and histaminase activities into plasma after heparin administration.