重组人胰高血糖素类肽-1(7-36)的药动学与组织分布

目的研究重组人胰高血糖素类肽-1(7-36)[rhGLP-1(7-36)]在动物体内的药动学与组织分布。方法猕猴给予不同剂量(3.5、14和56μg.kg-1)rhGLP-1(7-36),酶联免疫吸附分析法(ELISA)测定猕猴血浆中rhGLP-1(7-36)浓度,[125I]标记示踪法结合分子排阻色谱法(SEC)观察rh-GLP-1(7-36)在Wistar大鼠各组织中的分布。结果猕猴单次sc不同剂量rhGLP-1(7-36)后,血药浓度及AUC随剂量呈线性增加,各组达峰时间(Tmax)均为15 min,末端相半衰期(T12β)35～45 min,全身清除率(CL)1.0～1.3 L.min-1.kg-1,组间均无差别。sc给药绝对生物利用度22.2%。以3.5μg.kg-1剂量连续sc 7 d体内无蓄积。大鼠sc[125I]rh-GLP-1(7-36)后10 min血浆原形药物浓度达峰,降解物迅速出现。肾脏与胰腺此时总放射性浓度最高。结论在研究剂量范围内rhGLP-1(7-36)呈线性动力学。吸收、降解迅速是其动力学的特点。药物主要通过肾脏排泄,胰腺可能为药物的靶器官。     

重组人胰高血糖素样多肽-1(7-36)在健康人体的药代动力学

目的研究重组人胰高血糖素类多肽-1(7-36)[rhGLP-1(7-36)](促胰岛素生成药)在健康志愿者的药代动力学。方法选择12,9名健康志愿者,分别单次(0.1,0.15,0.2mg)和多次(0.2mg)皮下注射rhGLP-1(7-36),连续5日,用荧光酶联免疫分析法测定其血药浓度,用DAS软件计算其药代动力学参数。结果单次:药-时曲线符合一房室模型,Cmax、AUC0-t随剂量增加而增加,tmax约为19~21min,t1/2为10~14min。多次:首次与末次给药后的Cmax分别为(726.76±94.07),(737.15±72.12)ng·L-1;tmax为18min左右;t1/2为15~17min。结论在0.1~0.2mg内,过程呈一级线性动力学特征,体内无蓄积,耐受性较好。     

rhGLP-1(7-36)对糖尿病肾病大鼠肾脏的保护机制研究

目的 探讨rhGLP-1(7-36)对糖尿病肾病(diabetic kidney disease,DKD)大鼠肾脏的保护机制.方法 DKD模型大鼠随机分成模型组、rhGLP-1(7-36)低剂量组(20μg·kg-1)、rhGLP-1(7-36)中剂量组(40 μg·kg-1)、rhGLP-1(7-36)高剂量组(80...     

健康志愿者单次皮下注射重组人胰高血糖素类多肽-1(7-36)[rhGLP-1(7-36)]耐受性研究

目的：在中国健康成年志愿者中评价单次皮下注射重组人胰高血糖素类多肽-1（7-36）[rhGLP-1（7-36）]的安全性、耐受性。方法：根据GCP设计试验方案，并获得伦理委员会批准。受试者须自愿签署知情同意书。选择42名18～50岁健康成人，将受试者随机分至0．10～0．45mg7个剂量组，每组6名，男女各半，分别接受单次皮下注射rhGLP-1（7-36），进行临床和实验室检查，考察药物耐受性。结果：单次皮下注射rhGLP-1（7-36）耐受性试验中，各组受试者各项指标测定值均在正常范围内，条件均衡，具较好可比性。因剂量至0．30mg时，不良事件（恶心、呕吐）在该组发生率超过50％，故于该剂量组试验完成后终止了下一剂量组试验，仅有4个剂量组共24名健康受试者完成了本试验。24例受试者完成的4个剂量组耐受性试验中，给药后实验室检查未见有临床意义的改变。试验中出现10例（共15例次）可能与药物有关的不良反应，如头晕、恶心、呕吐等，但均可耐受，且为一过性反应，于给药后1h内自行消失。其中，不良反应主要发生在0.25、0．30mg组（共7例12例次），而低剂量组（0．10、0．20mg）仅有3例发生轻度不良反应。15例次不良反应中，头晕、恶心有10例次，呕吐有5例次；整个试验过程未见严重不良事件。结论：24名中国健康成年受试者分别单次皮下注射rhGLP-1（7-36），最大剂量至0．20mg，比较安全、耐受性较好，为最大耐受剂量。而单次给药剂量至0．25mg或0．30mg则不良反应发生率较高，最大耐受剂量为0．20mg。建议单次用药剂量不宜超过0．20mg。在Ⅱ期临床试验中需严密观察恶心、呕吐这些与药物可能有关的不良反应及其发生机制。     

ELISA试剂盒方法测定重组人胰高血糖素类多肽-1(7-36)浓度

目的：定量检测血浆样本中重组人胰高血糖素类多肽-1（7-36）[rhGLP-1（7-36）]的含量。方法：用LINCO公司生产的试剂盒对rhGLP-1（7-36）的标准品和血浆样本进行定量检测。结果：测定的特异性、灵敏度、测量范围、重现性和回收率满足生物制品药代动力学研究的要求。结论：用LINCO公司生产的试剂盒定量血浆样品中的rhGLP-1（7-36），该检测方法符合生物制品检测方法，可以作为其药代动力学研究血药浓度的检测方法。     

重组人干扰素-β1b对恒河猴免疫原性的影响

目的:研究重组人干扰素-β1b(rhIFN-β1b)对恒河猴的免疫原性及其对疗效的潜在影响.方法:实验分溶剂对照组及rhIFN-β1b低、中、高剂量组,分别皮下注射1.25%人血清白蛋白0.36mL·kg-1及rhIFN-β1b 4.0×106,1.2×107及3.6×107IU·kg-1,连续90d.血清结合抗体测定采用ELISA法,中和抗体测定采用细胞病变抑制法.结果:给药3周后,3个给药组动物均出现rhIFN-β1b结合抗体,4周后抗体滴度达最大值,不同剂量组间在结合抗体滴度方面未见明显的剂量-反应关系.同时,给药3周后血清中出现中和抗体,以后抗体滴度逐步升高.停药30d后,中和抗体滴度逐渐降低.结论:恒河猴重复给予rhIFN-β1b后,体内产生结合抗体和中和抗体,但给药组动物未观察到明显的病理学和生化指标异常,提示该生物制品重复使用是安全的.     

重组EPO-Fc 融合蛋白对恒河猴的长期毒性和免疫原性研究*

目的:观察重组EPO-Fc对恒河猴的长期毒性和免疫原性。方法:健康恒河猴随机分为EPO-Fc高、中、低剂量组(80,25,8μg.kg-1)和对照组,每组6只,雌雄各半。皮下注射给药,每周1次,连续24周。末次给药后24 h处死2/3,剩下动物停药后继续观察8周。观察症状和检测指标包括:一般情况、血液学、血生化、血电解质、心电图、粪尿、骨髓、病理组织学及血清抗体等各项检测。结果:连续给药后,高、中剂量组的红细胞计数(RBC)、血细胞比容(HCT)、血红蛋白(HGB)等升高,这与其药理作用有关。给药第4周高剂量组开始出现抗EPO-Fc抗体,第16周各剂量组大多数动物产生抗体;恢复期间,抗体滴度逐渐降低。其余各项观察指标均未见异常。结论:长期给予EPO-Fc对恒河猴无明显毒性,其安全剂量为80μg.kg-1,但具有免疫原性。     

人参三醇-3,6-二琥珀酸酯钠I期临床耐受性研究

目的:评价抗肿瘤新药人参三醇-3,6-二琥珀酸酯钠的人体安全性,观察毒副反应并确定人体安全耐受剂量。方法:采用改良的Fibonacci方法进行剂量递增。选择30例健康受试者和9例肿瘤患者。21例健康受试者进行7个剂量组单次给药试验(20,40,70,100,140,190,240 mg);9例健康受试者进行3个剂量组(70,100,140 mg)连续给药30 d的试验,9例肿瘤患者进行100 mg连续给药30 d试验。结果:21例健康受试者单次给药试验未观察到不良反应;9例健康受试者和9例肿瘤患者连续给药30 d试验主要的不良反应为乏力、体内热感、咽痛和大便干燥等。结论:人参三醇-3,6-二琥珀酸酯钠毒性反应轻,患者耐受性好。推荐Ⅱ期临床使用剂量及方法为70~100 mg.d-1,静脉滴注,连用30 d。     

多糖-蛋白结合菌苗免疫婴儿对b型流感杆菌的血清调理活性

b型流感杆菌(Hib)多糖菌苗对18～24月龄以下婴儿的免疫原性很差.本文作者用精制Hib多糖(PRP)菌苗和Hib多糖-白喉类毒素(PRP-D)结合菌苗免疫9～13月龄婴儿,进行了血清调理活性比较试验.两种菌苗分别接种14名婴儿,接种两针,间隔1个月.采集免疫前及每次注射后1个月的血清样品,测定PRP抗体总浓度及检查     

加替沙星滴耳液Ⅰ期临床耐受性试验

目的评价健康人体对加替沙星滴耳液的耐受性。方法采用随机、双盲、安慰剂对照的单中心临床研究。52名健康受试者,男女各半,分两部分试验,均点耳后耳浴10 min。单次给药耐受性试验纳入36例,随机分为5组(6、6、8、8、8例,每组包含2例安慰剂),给药剂量分别为0.45、0.9、1.35、1.8、2.25 mg(即3、6、9、12、15滴)。连续给药耐受性试验纳入16例,随机分为2组,每组8例(每组包含2例安慰剂),给药剂量分别为1.8(12滴)、2.25 mg(15滴),3次·d-1,疗程为7 d。观察受试者用药前后生命体征、临床症状及实验室指标的变化,记录不良事件。结果 51例受试者完成试验,1例脱落。单次给药和连续给药耐受性试验中,受试者用药前后生命体征、全身症状、实验室检查均未出现有临床意义的改变。连续给药试验中有2例出现耳部不适感,考虑与试验药物肯定有关。11例实验室检查结果异常(血糖、血脂升高,血常规、尿常规、肝功能、凝血功能异常),单次给药7例,连续给药4例。所有不良事件均为轻度,考虑与试验药物可能有关,未经干预,均自行缓解。结论健康受试者耳点加替沙星滴耳液,单次用药2.25 mg·次-1以内,连续用药1.8 mg·次-1以内,每日3次,疗程7 d,可安全耐受。     

重组人胰高血糖素样肽-1对实验性糖尿病大鼠血糖的影响

目的观察重组人胰高血糖素样肽 1[rhGLP 1(7 36 )NH2 和rhGLP 1(7 37) ]对实验性糖尿病大鼠模型血糖和血浆胰岛素水平的影响。方法正常SD大鼠按 3.2 4g/kg腹腔注射 5 0 %葡萄糖 ,诱发暂时性高血糖动物模型。腹腔注射不同剂量 (2 0、4 0、80 μg/kg)rhGLP 1(7 36 )NH2 和rhGLP 1(7 37) ,间隔取血测定血糖与血浆胰岛素水平。正常SD大鼠按 6 0mg/kg腹腔注射链脲佐菌素 ,建立实验性 1型糖尿病大鼠模型。静脉输注rhGLP 1(7 36 )NH2 [4.0ng/(kg·min) ]12 0min并观察血糖变化。结果腹腔注射 4 0、80 μg/kgrhGLP 1(7 36 )NH2 和rhGLP 1(7 37)处理组大鼠的血糖、血浆胰岛素水平与对照组之间的差别具有非常显著性意义 (P <0 .0 0 1) ,且呈现一定的量效关系。二者之间促胰岛素分泌和降血糖活性差异无显著性。静脉输注rhGLP 1(7 36 )NH2 6 0min后 ,血糖水平 (17.0 4± 1.31)mmol/L(P <0 .0 5 ) ,12 0min后血糖水平 (11.98± 1.0 5 )mmol/L(P <0 .0 0 1)。结论rhGLP 1能显著改善实验性糖尿病大鼠的血糖水平 ,这与其促胰岛素分泌活性有关。     

重组人胰高血糖素类多肽-1(7-36)对化学致糖尿病模型动物的降血糖作用

目的　本文研究了重组人胰高血糖素类多肽 1(7 36 )[rhGLP 1(7 36 ) ]对化学所致糖尿病模型动物的降血糖和促胰岛素分泌作用。方法　对四氧嘧啶型糖尿病小鼠及链脲霉素型糖尿病大鼠皮下注射不同剂量的rhGLP 1,分别于给药 4天后采血 ,收取血清 ,测定血糖及胰岛素值。结果　四氧嘧啶型糖尿病小鼠皮下注射 4 0、80 μg·kg-1rhGLP 1后 ,血糖值显著下降 (P <0 0 1) ,而 2 0 μg·kg-1剂量组则无显著性改变 ;链脲霉素型糖尿病大鼠皮下注射 2 8、5 6 μg·kg-1rhGLP 1后 ,血糖值显著下降 (P <0 0 5、P <0 0 1) ,而 14 μg·kg-1剂量组则无显著性改变。结论　rhGLP 1对实验动物部分 β细胞破坏的胰岛仍有促分泌胰岛素及降糖作用     

Pharmacokinetics and metabolism studies on the glucagon-like peptide-1 (GLP-1)-derived metabolite GLP-1(9-36)amide in male Beagle dogs

Abstract

1.?Glucagon-like peptide-1 (GLP-1)(7-36)amide is a 30-amino acid peptide hormone that is secreted from intestinal enteroendocrine L-cells in response to nutrients. GLP-1(7-36)amide possesses potent insulinotropic actions in the augmentation of glucose-dependent insulin secretion. GLP-1(7-36)amide is rapidly metabolized by dipeptidyl peptidase-IV to yield GLP-1(9-36)amide as the principal metabolite. Contrary to the earlier notion that peptide cleavage products of native GLP-1(7-36)amide [including GLP-1(9-36)amide] are pharmacologically inactive, recent studies have demonstrated cardioprotective and insulinomimetic effects with GLP-1(9-36)amide in mice, dogs and humans.

2.?In the present work, in vitro metabolism and pharmacokinetic properties of GLP-1(9-36)amide have been characterized in dogs, since this preclinical species has been used as an animal model to demonstrate the in vivo vasodilatory and cardioprotective effects of GLP-1(9-36)amide. A liquid chromatography tandem mass spectrometry assay was developed for the quantitation of the intact peptide in hepatocyte incubations as opposed to a previously reported enzyme-linked immunosorbent assay. Although GLP-1(9-36)amide was resistant to proteolytic cleavage in dog plasma and bovine serum albumin (t_1/2?>?240?min), the peptide was rapidly metabolized in dog hepatocytes with a t_1/2 of 110?min. Metabolite identification studies in dog hepatocytes revealed a variety of N-terminus cleavage products, most of which, have also been observed in human and mouse hepatocytes. Proteolysis at the C-terminus was not observed in GLP-1(9-36)amide.

3.?Following the administration of a single intravenous bolus dose (20?µg/kg) to male Beagle dogs, GLP-1(9-36)amide exhibited a mean plasma clearance of 15?ml/min/kg and a low steady state distribution volume of 0.05?l/kg, which translated into a short elimination half life of 0.05?h. Following subcutaneous administration of GLP-1(9-36)amide at 50?µg/kg, systemic exposure of GLP-1(9-36)amide as ascertained from maximal plasma concentrations and area under the plasma concentration–time curve from zero to infinity was 44?ng/ml and 32?ng?h/ml, respectively. The subcutaneous bioavailability of GLP-1(9-36)amide in dogs was 57%.

4.?Our findings raise the possibility that the cardioprotective effects of GLP-1(9-36)amide in the conscious dog model of pacing-induced heart failure might be due, at least in part, to the actions of additional downstream metabolites, which are obtained from proteolytic cleavage of the peptide backbone in the parent compound in the liver.     

Procainamide-induced antinuclear antibodies in beagle dogs

Procainamide produces a high incidence of autoimmune reactions in humans. To establish an animal model, we treated three groups of beagle dogs with increasing doses of procainamide HCl (100 to 200 mg/kg) for 11 to 14 months. Sera were examined before and during treatment for antinuclear antibodies (ANA) at a 1:10 dilution by an indirect immunofluorescence assay. The first group of six dogs, which were 1 year old and born in our colony, remained generally negative for ANA. In the second group of four dogs, which were of the same origin but were 3 to 6 years old, two developed ANA after 4 months of treatment. The third group of four dogs were 3 to 6 years old and had been born in the FDA colony to parents originating from another source. These dogs developed ANA after 1 to 2 months and remained generally positive throughout the 11 months of treatment. Dogs of the control groups remained negative except for one that was ANA-positive on several occasions. Three of the ANA-positive treated dogs and the ANA-positive control dog had circulating immune complexes. Serum procainamide concentrations were comparable in dogs of each group; acetylprocainamide was not detected. No alterations were found in serum proteins or in the hemograms, and no clinical signs were seen. The data suggest that age plays a contributory role in the development of this autoimmune response in the dog.     

Correlation between levels of human papillomavirus (HPV)-16 and 18 antibodies in serum and cervicovaginal secretions in girls and women vaccinated with the HPV-16/18 AS04-adjuvanted vaccine

This pooled analysis of data from four Phase III clinical trials was undertaken to assess the correlation between levels of anti-human papillomavirus (HPV)-16/18 antibodies in serum and cervicovaginal secretions (CVS) in girls and women vaccinated with the HPV-16/18 AS04-adjuvanted vaccine. Serum and CVS samples were collected from a subset of women aged 10-65 years (N=350) at pre-specified time-points from 7 to 36 months post-vaccination. Anti-HPV-16/18 antibody levels in serum and CVS were measured by enzyme-linked immunosorbent assay. Pearson correlation coefficients between serum and CVS antibody levels, standardized for total immunoglobulin G, were calculated at each time-point in women with detectable antibodies in both serum and CVS. All subjects had seroconverted at Month 7 and remained seropositive through Month 36 for both antigens. Geometric mean titers of anti-HPV-16/18 antibodies in serum were substantially higher at all time-points than those in a control group of women who had cleared a natural HPV infection in another trial. In women with detectable antibodies in both serum and CVS, good correlation was seen between HPV-16/18 antibody levels at all time-points (Pearson correlation coefficient: 0.84-0.92 for HPV-16 and 0.90-0.91 for HPV-18). The strong correlation between levels of HPV-16/18 antibodies in serum and CVS up to 36 months post-vaccination in girls and women vaccinated with the HPV-16/18 AS04-adjuvanted vaccine supports transudation of serum antibodies as the mechanism by which antibodies are introduced into CVS. These CVS antibodies may play a role in the protective efficacy of this vaccine.     

间硝苯地平在Beagle犬体内的药代动力学

目的用反相高效液相色谱法研究间硝苯地平(m-nifedipine，m-Nif)在Beagle犬体内的药代动力学特征。方法正交设计优化色谱分离条件，Beagle犬分别iv给予m-Nif 0.288 mg·kg^-1和ig m-Nif 1.152，3.456，10.370 mg·kg^-1。用反相高效液相色谱法分析血浆中原型药物浓度，血浆药物浓度-时间数据用3P97药代动力学软件分析。结果Beagle犬iv m-Nif，其体内过程符合二室模型，T_1/2β为116.8 min；ig给予m-Nif 后在Beagle犬体内的代谢符合一室模型，其中低剂量(1.152 mg·kg^-1)组C_max为20 μg·L^-1， T_1/2(k_e)为147 min；中剂量(3.456 mg·kg^-1)组C_max为36 μg·L^-1，T_1/2(k_e) 为122 min；高剂量(10.37 mg·kg^-1)组C_max为69 μg·L^-1，T_1/2(k_e)为144 min。结论Beagle犬ig和iv m-Nif 后，血浆中药物消除迅速，口服绝对生物利用度较低。     

Chondrotoxicity of ciprofloxacin in immature Beagle dogs: immunohistochemistry, electron microscopy and drug plasma concentrations

The systemic effects of ciprofloxacin in immature Beagles were studied. Dogs of 10-11 weeks were dosed orally for 5 days with 0 (n=3), 30 (n=5) and 200 (n=5) mg ciprofloxacin/kg body wt. Plasma concentrations were measured by high-performance liquid chromatography (HPLC) 1 h after dosing (assuming to be peak concentrations). In view of the high doses used, the plasma concentrations were rather low and declined during the study period. For example, plasma concentrations in the high dose group were 6.6 +/- 0.9 mg/l (day 1), 3.9 +/- 1.4 mg/l (day 3), and 2.6 +/- 1.6 mg/l (day 5). In control dogs and in dogs treated with the low dose of ciprofloxacin no pathological changes were seen by light microscopy. However, cleft formation and erosions were observed in joint cartilage from two of five dogs treated with 200 mg/kg. It is noteworthy that despite the high dose used cartilage lesions were not detectable in all five dogs of this group by light microscopy. Using antibodies against cell membrane receptors (e.g. the alpha(5)beta(1)-integrin) or matrix components (fibronectin, collagen II) the articular cartilage effects were studied in detail by immunohistochemistry. The most sensitive alteration was an increase in fibronectin which was detectable in the vicinity of the lesions in cartilage samples from the group of dogs administered the high dose. No clear-cut changes were seen with the use of antibodies against other matrix components. Electron microscopy revealed typical alterations in chondrocytes from dogs treated with ciprofloxacin: e.g., swollen mitochondria and enlarged rough endoplasmic reticulum. These changes were much more pronounced in dogs from the high dose group than in dogs from the low dose group. Our main conclusion is that after oral administration ciprofloxacin exhibits rather low chondrotoxicity, even in the most sensitive species known to date. This correlates with the findings in humans that ciprofloxacin seems to be less chondrotoxic than pefloxacin or other quinolones.