Practical problems in regulatory toxicology I ：Formulations
Although highly regulated, all non-clinical toxicology studies have the potential to experience practical prob- lems. Anticipation of potential hurdles and...Practical problems in regulatory toxicology I ：Formulations
Although highly regulated, all non-clinical toxicology studies have the potential to experience practical prob- lems. Anticipation of potential hurdles and planning for success are key actions for all investigators to ensure a study is ro- bust and suitable for submission and regulatory acceptance. This presentation will examine an area of study conduct which can be problematic but which if approached correctly will ensure good study conduct. Many test materials under develop- ment today are insoluble. Achieving a suitable formulation for administration to laboratory animalsis difficult and therefore having strategies for determining suitable vehicles and dosing regimen essential. To achieve the optimal bioavailability of the chemical you need to strike a balance between the dose concentration, dose volume and the dosing regimen. The presentation will consider the interactions of factors such as the physical nature of the final formulation, physicalproperties of vehicles and potential side effects which should be considered together when optimising the final dosing formulation. It will demonstrate how a methodical approach can assist the scientist in establishing a formulation that will achieve the best bioavailability possible with minimal effects to the test animal. The presentation will also illustrate how the Study Director can demonstrate strong involvement and active study management as required by the GLP authorities.展开更多
目的通过对重组结核杆菌融合蛋白(EC)[该制品名称是国家药典委员会确定的药品中文通用名称,"EC"为重组融合蛋白"结核分枝杆菌早期分泌性抗原靶6(ESAT-6)和培养滤液蛋白10(CFP-10)"](简称"EC")的免疫特性...目的通过对重组结核杆菌融合蛋白(EC)[该制品名称是国家药典委员会确定的药品中文通用名称,"EC"为重组融合蛋白"结核分枝杆菌早期分泌性抗原靶6(ESAT-6)和培养滤液蛋白10(CFP-10)"](简称"EC")的免疫特性和临床前安全性研究,探讨其临床应用前景。方法(1)免疫特性研究:取6只雌性豚鼠,使用结核分枝杆菌H37Ra活菌菌液致敏,5周后,皮内注射2.5μg/ml EC原液0.2ml;取4只雌性豚鼠,使用卡介菌活菌菌液致敏,5周后,皮内注射20μg/ml EC原液0.2ml;观察注射部位平均硬结或红晕反应直径[(纵径+横径)/2],≥5mm判为阳性,<5mm判为阴性。(2)急性毒性试验:取80只ICR[(美国)Institute of Cancer Research]小鼠,分为单次肌内和皮内注射组,每组40只,雌雄各半;每组再分为4组,每组10只,雌雄各半。高剂量组(注射EC53.61μg/0.1ml)、低剂量组(注射EC 0.2μg/0.1ml)、溶剂对照组(注射EC稀释液0.1ml)、空白对照组(不给予任何受试物),观察小鼠的外观、运动功能、体质量、各脏器和药物注射部位皮肤等是否有异常。(3)豚鼠全身主动过敏试验:取豚鼠24只,分为4组,每组6只,雌雄各半,各组豚鼠分别隔日腹腔注射高剂量(5μg/kg)EC、低剂量(0.5μg/kg)EC、牛血清白蛋白(60mg)和0.9%NaCl(质量分数为0.9%的NaCl溶液)注射液(2ml),连续3次。致敏结束,末次致敏后第12天静脉快速注射2倍致敏剂量对以上相应的各组致敏豚鼠进行激发。致敏期间,每天观察豚鼠的症状,并于初次和最后一次致敏及激发当日测定每只豚鼠的体质量。(4)皮内刺激试验:取6只新西兰兔,单次皮内注射EC 10μg(0.2ml)/点,每侧5个点,观察皮内注射后的刺激反应情况。结果(1)免疫特性研究结果:EC原液对卡介菌活菌菌液致敏的4只豚鼠的皮肤试验,阳性为0只;EC原液对结核分枝杆菌H37Ra活菌菌液致敏的6只豚鼠的皮肤试验,阳性为6只。(2)急性毒性试验结果:所有小鼠未出现明显的急性中毒反应和显示急性中毒靶器官。小鼠皮内注射EC前、注射EC后1d、3d、5d、7d、8d、10d、12d、14d高剂量组、低剂量组小鼠平均体质量分别为(20.6±1.3)g^(24.9±2.1)g、(20.5±1.6)g^(26.0±3.1)g;注射后不同观察时间各组平均体质量与空白对照组[(21.0±1.1)g^(25.3±2.3)g]比较差异均无统计学意义(t值分别为0.571~0.392,0.695~0.615;P值分别为0.575~0.700,0.496~0.546);小鼠肌内注射EC前、注射EC后1d、3d、5d、7d、8d、10d、12d、14d高剂量组、低剂量组小鼠平均体质量分别为(21.0±1.5)g^(26.2±1.9)g、(20.5±2.1)g^(25.8±3.8)g;注射后不同观察时间各组平均体质量与空白对照组[(21.2±1.7)g^(25.8±3.1)g]比较差异均无统计学意义(t值分别为0.360~0.318,0.900~0.006;P值分别为0.723~0.754,0.380~0.995)。(3)豚鼠全身主动过敏试验结果:豚鼠无过敏反应。高剂量组、低剂量组豚鼠首次致敏、末次致敏、激发的平均体质量分别为(327.5±24.3)g、(347.2±32.7)g、(402.2±34.9)g;(331.3±26.7)g、(346.2±32.0)g、(411.3±38.9)g,与相应观察时间注射0.9%NaCl的阴性对照组[(329.5±27.4)g、(348.3±27.0)g、(399.4±25.4)g]比较,差异均无统计学意义(t值分别为0.328、0.181、0.284,0.474、0.366、0.875;P值分别为0.757、0.864、0.788,0.656、0.730、0.422)。(4)兔皮内刺激试验结果:单次皮内注射10μg(0.2ml)/点EC,无明显刺激反应。结论EC能够鉴别结核感染与卡介苗免疫,临床前动物试验安全性好,有望应用于人群结核感染的诊断与鉴别诊断。展开更多
文摘Practical problems in regulatory toxicology I ：Formulations
Although highly regulated, all non-clinical toxicology studies have the potential to experience practical prob- lems. Anticipation of potential hurdles and planning for success are key actions for all investigators to ensure a study is ro- bust and suitable for submission and regulatory acceptance. This presentation will examine an area of study conduct which can be problematic but which if approached correctly will ensure good study conduct. Many test materials under develop- ment today are insoluble. Achieving a suitable formulation for administration to laboratory animalsis difficult and therefore having strategies for determining suitable vehicles and dosing regimen essential. To achieve the optimal bioavailability of the chemical you need to strike a balance between the dose concentration, dose volume and the dosing regimen. The presentation will consider the interactions of factors such as the physical nature of the final formulation, physicalproperties of vehicles and potential side effects which should be considered together when optimising the final dosing formulation. It will demonstrate how a methodical approach can assist the scientist in establishing a formulation that will achieve the best bioavailability possible with minimal effects to the test animal. The presentation will also illustrate how the Study Director can demonstrate strong involvement and active study management as required by the GLP authorities.
文摘目的通过对重组结核杆菌融合蛋白(EC)[该制品名称是国家药典委员会确定的药品中文通用名称,"EC"为重组融合蛋白"结核分枝杆菌早期分泌性抗原靶6(ESAT-6)和培养滤液蛋白10(CFP-10)"](简称"EC")的免疫特性和临床前安全性研究,探讨其临床应用前景。方法(1)免疫特性研究:取6只雌性豚鼠,使用结核分枝杆菌H37Ra活菌菌液致敏,5周后,皮内注射2.5μg/ml EC原液0.2ml;取4只雌性豚鼠,使用卡介菌活菌菌液致敏,5周后,皮内注射20μg/ml EC原液0.2ml;观察注射部位平均硬结或红晕反应直径[(纵径+横径)/2],≥5mm判为阳性,<5mm判为阴性。(2)急性毒性试验:取80只ICR[(美国)Institute of Cancer Research]小鼠,分为单次肌内和皮内注射组,每组40只,雌雄各半;每组再分为4组,每组10只,雌雄各半。高剂量组(注射EC53.61μg/0.1ml)、低剂量组(注射EC 0.2μg/0.1ml)、溶剂对照组(注射EC稀释液0.1ml)、空白对照组(不给予任何受试物),观察小鼠的外观、运动功能、体质量、各脏器和药物注射部位皮肤等是否有异常。(3)豚鼠全身主动过敏试验:取豚鼠24只,分为4组,每组6只,雌雄各半,各组豚鼠分别隔日腹腔注射高剂量(5μg/kg)EC、低剂量(0.5μg/kg)EC、牛血清白蛋白(60mg)和0.9%NaCl(质量分数为0.9%的NaCl溶液)注射液(2ml),连续3次。致敏结束,末次致敏后第12天静脉快速注射2倍致敏剂量对以上相应的各组致敏豚鼠进行激发。致敏期间,每天观察豚鼠的症状,并于初次和最后一次致敏及激发当日测定每只豚鼠的体质量。(4)皮内刺激试验:取6只新西兰兔,单次皮内注射EC 10μg(0.2ml)/点,每侧5个点,观察皮内注射后的刺激反应情况。结果(1)免疫特性研究结果:EC原液对卡介菌活菌菌液致敏的4只豚鼠的皮肤试验,阳性为0只;EC原液对结核分枝杆菌H37Ra活菌菌液致敏的6只豚鼠的皮肤试验,阳性为6只。(2)急性毒性试验结果:所有小鼠未出现明显的急性中毒反应和显示急性中毒靶器官。小鼠皮内注射EC前、注射EC后1d、3d、5d、7d、8d、10d、12d、14d高剂量组、低剂量组小鼠平均体质量分别为(20.6±1.3)g^(24.9±2.1)g、(20.5±1.6)g^(26.0±3.1)g;注射后不同观察时间各组平均体质量与空白对照组[(21.0±1.1)g^(25.3±2.3)g]比较差异均无统计学意义(t值分别为0.571~0.392,0.695~0.615;P值分别为0.575~0.700,0.496~0.546);小鼠肌内注射EC前、注射EC后1d、3d、5d、7d、8d、10d、12d、14d高剂量组、低剂量组小鼠平均体质量分别为(21.0±1.5)g^(26.2±1.9)g、(20.5±2.1)g^(25.8±3.8)g;注射后不同观察时间各组平均体质量与空白对照组[(21.2±1.7)g^(25.8±3.1)g]比较差异均无统计学意义(t值分别为0.360~0.318,0.900~0.006;P值分别为0.723~0.754,0.380~0.995)。(3)豚鼠全身主动过敏试验结果:豚鼠无过敏反应。高剂量组、低剂量组豚鼠首次致敏、末次致敏、激发的平均体质量分别为(327.5±24.3)g、(347.2±32.7)g、(402.2±34.9)g;(331.3±26.7)g、(346.2±32.0)g、(411.3±38.9)g,与相应观察时间注射0.9%NaCl的阴性对照组[(329.5±27.4)g、(348.3±27.0)g、(399.4±25.4)g]比较,差异均无统计学意义(t值分别为0.328、0.181、0.284,0.474、0.366、0.875;P值分别为0.757、0.864、0.788,0.656、0.730、0.422)。(4)兔皮内刺激试验结果:单次皮内注射10μg(0.2ml)/点EC,无明显刺激反应。结论EC能够鉴别结核感染与卡介苗免疫,临床前动物试验安全性好,有望应用于人群结核感染的诊断与鉴别诊断。
