抗乙肝病毒新药ZBH201001的一般毒性研究

发布时间：2018-01-25 19:30

本文关键词： 核苷类抗乙肝病毒药物 ZBH201001 临床前安全性评价 Beagle犬单次给药毒性研究重复给药毒性研究　出处：《广西医科大学》2015年硕士论文　论文类型：学位论文

【摘要】：目的：ZBH201001是军事医学科学院毒物药物研究所研究人员在阿米福韦化学结构的基础上设计合成的一种核苷类抗乙肝病毒药物。本研究依据药物临床前安全性评价相关指导原则,以Beagle犬为研究对象,分别进行单次给药毒性研究和重复给药毒性研究,观察动物出现的毒性反应,了解ZBH201001的急性毒性作用,以及重复给药毒性作用的靶器官及其损害的可逆性,确定未观察到损害作用的剂量(NOAEL),为其临床试验的用药剂量和用药安全剂量范围提供参考,同时为该新药临床申请和新药注册申请提供数据支持。方法：(1)Beagle犬经口急性毒性试验：采用近似致死剂量法,单次经口灌胃给药,给药剂量分别为118、264、594、890、1334、2000 mg/kg体重。毒性检测指标包括一般临床观察、体重、摄食量、体温、心电图、血液学、血液生化、凝血指标以及尿液化学,给药观察3周后,对所有动物进行组织病理学检查。(2) Beagle犬经口重复给药1个月毒性试验：实验设6.0、12.0、24.0mg/kg体重组及玉米油溶剂对照组,每天经口给药1次,给药期30天,给药结束后恢复4周。连续给药9天后,因各剂量组动物均出现了明显的毒性反应,故将剂量调低为3.0、6.0与9.0 mg/kg体重,继续给药至实验结束。实验期间每日进行一般临床观察与体重、摄食量测定,给药中期、给药结束与恢复期分别进行体温、心电图、血液学、血液生化、凝血指标与尿液化学检测,并于给药结束与恢复期结束进行组织病理学检查。结果：(1)Beagle犬经口急性毒性试验：Beagle犬单次经口灌胃给药,594 mg/kg体重及以上剂量组动物给药后笼旁观察显示,给药后和/或药后次日可见动物呕吐,偶见稀便。随后观察到动物摄食状况普遍较差、体重持续降低；约自第3周起,动物摄食状况逐渐好转,体重逐渐增加。890 mg/kg体重剂量组1只雌性动物于给药后第12天意外死亡,重复给予该剂量雄性动物存活。118 mg/kg及264 mg/kg组给药后观察到动物摄食减少,体重轻度降低。较高剂量组动物给药后个别时间点个别血液学指标、尿液化学指标出现异常变化,体温、心电图等参数未见有毒理学意义的改变。组织病理学检查：890 mg/kg组意外死亡动物尸检除体型消瘦外未见明显异常,镜下检查见肺严重水肿、出血、肺泡上皮脱落,胃肠系统出现病变；其余6只存活动物剖杀,体表和内脏检查均未见异常。1334 mg/kg体重组动物肝脏灶性肝细胞空泡变性、散在坏死,结肠出血、上皮细胞坏死、炎性细胞浸润；2000mg/kg体重组动物胃黏膜坏死、水肿；其他动物各脏器未见明显异常。(2) Beagle犬经口重复给药1个月毒性试验：结果显示,以24.0、12.0、6.0 mg/kg体重剂量给药9天后,雌、雄动物均表现出胃肠毒性反应,包括稀便、动物摄食状况差、体重降低等体征,并呈现较明显的量效与时效关系。因此将剂量将低为9.0、6.0与3.0 mg/kg体重继续给药,我们观察到高剂量9.0 mg/kg体重组动物摄食量、体重持续降低,并陆续有稀便和/或血便等现象,并呈现消瘦、活动减少等临床体征；3.0、6.0mg/kg组动物体重虽仍低于同期对照组,但基本保持在稳定的水平,未进一步降低。无论剂量调整前还是调整后,高、中、低剂量组动物的摄食量变化表现出与体重变化基本相一致的变化趋势。各期高、中、低剂量组心电图、体温及眼科检查均未见有毒理学意义的异常变化。给药中期与给药结束后血液学检查提示高剂量组动物APTT明显降低,并且这种变化在恢复期恢复正常,提示受试药物可能会影响动物的内源性凝血系统。各期尿液分析高、中、低剂量组未见异常改变。组织病理学检查,中剂量和/或高剂量组对消化管与胃肠系统具有明显的毒性作用,中剂量和/或高剂量组动物胸腺、睾丸、前列腺、胰腺萎缩,骨髓红系造血抑制、脂肪化,肝细胞肿胀、变性。上述各组织/脏器病变在恢复期均减轻和/或恢复,说明其毒性具有一定的可恢复性。给药结束高、中、低剂量组均有部分动物出现肺水肿、肺泡上皮脱落、炎性细胞浸润等病变,恢复期结束各组动物未见上述病变；考虑到口服给予供试品混悬液时动物偶有呛咳反应,并且肺部病变的严重程度及发生率无量效关系,因此认为所观察到的肺部病变与给药操作有关。结论：在本试验条件下,(1)Beagle犬经口急性毒性试验MTD2000mg/kg体重,最小毒性反应剂量为118 mg/kg体重,胃肠道可能是ZBH201001对Beagle犬急性毒性作用的靶器官/靶组织。(2)Beagle犬经口重复给药1个月毒性试验,中剂量和/或高剂量可引起食管、肝脏、胸腺、睾丸、前列腺、胰腺、骨髓、肾脏、胃肠系统一定的毒性反应,并可能影响内源性凝血系统,并且这些毒性反应停药后可恢复；因此,ZBH201001对Beagle犬重复给药的NOAEL为3.0 mg/kg体重(人临床拟用剂量以10 mg/日计,约相当于临床拟用剂量的18倍)。
[Abstract]:Objective: ZBH201001 is the Military Medical Science Academy of the PLA Drug Research Institute researchers design a synthesis of nucleoside anti HBV drugs based on the chemical structure of Amie. On the basis of tenofovir drug preclinical safety evaluation guidelines in Beagle dogs as the research object, respectively for single dose toxicity study and repeated toxicity of drugs the toxic reaction of the animal, the acute toxicity of ZBH201001, reversibility and repeated drug toxicity and target organ damage, determine the observed damage dose (NOAEL), to provide reference for the clinical trials of the dosage and safety dose range, and provide data support for the clinical application and new drug application. Methods: (1) Beagle canine acute toxicity test: the approximate lethal dose, single oral gavage Drug dosage was 11826459489013342000 mg/kg weight. Toxicity indexes included clinical observation, average weight, food intake, body temperature, electrocardiogram, hematology, blood biochemistry, blood coagulation index and urine chemistry, 3 weeks after the administration of observation, pathological examination of all animal. (2) Beagle dogs by mouth repeated dose toxicity test: 1 months experiment set 6.0,12.0,24.0mg/kg and body weight of corn oil solvent control group, daily oral administration of 1 times, delivery period of 30 days after the end of dosing recovery for 4 weeks. After 9 days of medication, for each dose group animal showed obvious toxicity, it will be the dose was 3.0,6.0 and 9 mg/kg by weight, continued administration until the end of the experiment. During the experiment, daily general clinical observation and determination of body weight, food intake, drug administration and the end of the mid recovery period were temperature, ECG, hematology, blood biochemistry, coagulation Blood and urine chemical detection, and recovery period to the end of dosing and end for histopathological examination. Results: (1) Beagle canine acute toxicity test: Beagle canine single oral gavage, and more than 594 mg/kg body weight dose group after administration of animal cage observation showed that the drug after the drug and / or the day after the visible animal vomiting, occasionally stools. Then observed animal feeding conditions are generally poor, continue to reduce weight; since about third weeks, animal feeding conditions gradually improved, the body weight increased.890 mg/kg body weight dose group of 1 female animal in twelfth days after administration of accidental death, repeated administration the dose of the male animal survival.118 mg/kg and 264 mg/kg were observed after the administration of animal food intake, body weight decreased slightly. The higher dose group after administration of animal individual time points of individual blood index, urine chemical indicators of abnormal ECG, body temperature. There is no toxicological significance map and other parameters change. Histopathological examination: 890 mg/kg group in addition to the accidental death of animal body weight and autopsy showed no abnormalities, endoscopic examination showed severe pulmonary edema, hemorrhage, alveolar epithelial shedding, gastrointestinal system disease; the remaining 6 were killed only activities, superficial and visceral examination showed no abnormal.1334 mg/kg recombinant animal liver focal liver cell degeneration, scattered necrosis, bleeding in the colon, epithelial cell necrosis, inflammatory cell infiltration; 2000mg/kg recombinant animal gastric mucosa necrosis, edema; other animal organs had no obvious abnormalities. (2) Beagle dogs by mouth repeated administration of 1 month toxicity test: the results showed that after 9 days of treatment, the 24.0,12.0,6.0 dose of mg/kg body weight of female and male animal showed gastrointestinal toxicity, including stools, animal feeding condition, weight loss and other symptoms, and showed obvious dose effect With the aging relationship. So the dose will continue to give medicine for low 9.0,6.0 and 3 mg/kg weight, we observed high dose 9 mg/kg recombinant animal food intake, body weight decreased, and there were rare and / or bloody stool phenomenon, and showed clinical signs such as weight loss, reduced activity; 3.0,6.0mg/kg group was lower than the weight of an animal compared with the control group, but remained at a stable level, not further reduced. No dose adjustment before or after adjustment, high, low dose group showed a trend of animal food intake and body weight change change accord. The high, low dose group, ECG, abnormal changes in toxicological significance body temperature and ophthalmic examination showed no medication. The middle and afteradministration hematology examination showed high dose group animal APTT decreased significantly, and the change in the recovery period returned to normal, suggesting that subjects may affect The endogenous coagulation system. Analysis of the animal urine during the high and low dose group showed no abnormal changes. Histopathological examination, middle dose and / or high dose group has obvious toxic effect on the digestive tract and gastrointestinal system, middle dose and / or high dose group of animal thymus, testis, prostate, pancreatic atrophy. Bone marrow hematopoietic suppression, fat, liver cell swelling, degeneration. The tissue / organ diseases during the recovery period were reduced and / or recovery, the toxicity can be recovered. The end of administration, and pulmonary edema occurred in low dose group were part of an animal, alveolar epithelial shedding, inflammatory cell infiltration lesions were not found in the end of the recovery period of animal disease; considering the oral administration of the sample suspension when the animal occasionally cough, and the severity of lung disease and there was no dose-response relationship, so that the observed lung Department of administration and operation of the lesions. Conclusion: under the experimental conditions, (1) Beagle dogs were acute toxicity test of MTD2000mg/kg weight, minimum dose toxicity was 118 mg/kg body weight, the gastrointestinal tract may be the target organ / ZBH201001 on acute toxicity of Beagle dog target tissue. (2) Beagle canine oral repeat for 1 months drug toxicity test, dose and / or high doses can cause the esophagus, liver, thymus, testis, prostate, pancreas, bone marrow, kidney toxicity, gastrointestinal system, and may affect the endogenous coagulation system, and the toxic reaction after drug withdrawal can be recovered; therefore, ZBH201001 repeat in Beagle dogs administration of NOAEL for 3 mg/kg weight (clinical dose of 10 mg/ to date, equivalent to about clinical dose of 18 times).

【学位授予单位】：广西医科大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R965

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