A multiple-dose toxicity study of tanezumab in cynomolgus monkeys

Nerve growth factor (NGF) is an important mediator of pain and hyperalgesia and has become a target of novel analgesic therapeutics. Tanezumab is a humanized IgG(2) antibody that binds NGF with high affinity and specificity. In a study to assess the toxicity and pharmacokinetic properties of tanezumab in adult, male and female, cynomolgus monkeys following weekly intravenous administration of 1, 10, or 30 mg/kg for up to 26 weeks (followed by an 8-week recovery period), tanezumab was well tolerated with no macroscopic or microscopic effects on those brain, spinal cord, nerve, or ganglia sections evaluated. One fifth of tanezumab-treated monkeys developed an antibody response to tanezumab that prevented maintenance of tanezumab exposure between dosing. In the antibody-negative animals, accumulation of tanezumab was observed; steady state was achieved approximately 8 weeks after the first dose of study drug, and exposure to tanezumab was approximately dose proportional with no observed difference between male and female animals. One monkey died during the study; this monkey had findings suggestive of hypersensitivity reaction. The favorable toxicity and pharmacokinetic profile of tanezumab seen in this study supports its further evaluation for the treatment of pain in clinical practice.     

食蟹猴静脉注射抗HER2人源化抗体急性毒性研究

目的 评价抗HER2 人源化抗体的急性毒性。方法 将食蟹猴随机分为4 组,包括溶媒对照和抗HER2 人源化抗体75、150 和250 mg/kg 组,单次iv 溶媒对照组或供试品,进行各项毒理学指标检测。结果 给药后各组动物临床症状、体质量、摄食量、体温、心电图、血压和血液学检测均未见明显异常;血清生化结果显示,150 mg/kg 组与250 mg/kg 组动物给药后血清IgG 水平出现一过性增加;各组动物均未见大体病理学改变。结论 食蟹猴单次iv 抗HER2 单抗,总体上动物具有良好的耐受性,最大耐受剂量可达250 mg/kg,这些结果为进一步临床前评价抗HER2 人源化单克隆抗体的安全性奠定了基础。     

Developmental toxicity of dibutyltin dichloride given on three consecutive days during organogenesis in cynomolgus monkeys

We previously reported that the administration of dibutyltin dichloride (DBTCl) by nasogastric intubation during the entire period of organogenesis, days 20–50 of pregnancy, was embryolethal, but not teratogenic, in cynomolgus monkeys. The present study was conducted to further evaluate the developmental toxicity of DBTCl given to pregnant monkeys on 3 consecutive days during organogenesis. Cynomolgus monkeys were given DBTCl at 7.5?mg/kg body weight/day by nasogastric intubation on days 19–21, 21–23, 24–26, 26–28, 29–31, 31–33, or 34–36 of pregnancy, and the pregnancy outcome was determined on day 100 of pregnancy. Embryonic/fetal loss was observed in 1 female given DBTCl on days 19–21, 2 females given DBTCl on days 24–26, and 1 female given DBTCl on days 34–36. There were no effects of DBTCl on developmental parameters in surviving fetuses, including fetal body weight, crown-rump length, tail length, or placental weight. No external, internal, or skeletal malformations were detected in fetuses in any group. DBTCl did not affect the incidence of fetuses with skeletal variation or skeletal ossification of fetuses. These data confirm our previous findings that DBTCl was embryolethal, but not teratogenic, in cynomolgus monkeys.     

Investigation of maternal and fetal exposure to an IgG2 monoclonal antibody following biweekly intravaginal administration to cynomolgus monkeys throughout pregnancy

To assess the potential for male-mediated drug transfer to their female partner and/or developing conceptus, vaginal uptake of a monoclonal antibody (mAb) biotherapeutic was assessed in cynomolgus monkeys. A human IgG2 mAb (IgG2X; bound human and cynomolgus monkey neonatal Fc-receptor, FcRn, with similar high affinity) was administered intravaginally (IvG; 100 mg/dose) to 5 pregnant cynomolgus monkeys biweekly from gestation day (gd) 21 to gd133. In all maternal samples collected before gd119, IgG2X plasma concentrations were below the limit of quantification (BLQ; <25 ng/mL). After dosing on gd119 and 133, maternal IgG2X plasma concentrations remained BLQ in 3/5 monkeys and were very low in 2/5 (up to 116 ng/mL; ∼0.01% of the IvG dose). IgG2X was BLQ in all fetal plasma samples. These data indicate that male-mediated mAb drug transfer via seminal fluid does not present a health risk to the female partner and is not bioavailable to the developing conceptus.     

Experimental oral toxicity of domoic acid in cynomolgus monkeys (Macaca fascicularis) and rats. Preliminary investigations

A recent outbreak of marine food poisoning in humans was attributed to the consumption of blue mussels (Mytilus edulis L.) contaminated with domoic acid (DA) that was produced by the diatom Nitzschia pungens. The clinical and morphological effects of single oral doses of extracts of mussels contaminated with DA or of DA isolated from toxic mussels were investigated in small groups (one to six) of cynomolgus monkeys (Macaca fascicularis; 0.5-10 mg DA/kg body weight) and of Sprague-Dawley rats (60 to 80 mg DA/kg body weight). Control animals were either given saline or were not treated. To test whether monosodium glutamate, present in the food consumed by some affected humans, and dimethylsulphoxide, suspected of being present in the plankton, enhanced the response, monosodium glutamate (at 0.25% of mussel extract bolus) or dimethylsulphoxide (at 1 g per bolus) were co-administered to two (one each) of the DA-treated monkeys. DA-treated monkeys developed transient excitation characterized by vomiting. DA-treated rats showed withdrawal followed by hyperexcitation and death (in one case). Mild to moderate central nervous system lesions consistent with neuroexcitation were present in both monkeys and rats. The addition of monosodium glutamate and dimethylsulphoxide had no significant effect on the appearance and severity of central nervous system clinical signs and lesions. The wide variations in the response of test animals to orally administered DA were attributed to the protective effect of vomiting, and to suspected incomplete or slow gastro-intestinal absorption of the toxic agent. The results reinforce the view that DA is an emetic and that under appropriate conditions may also inflict excitotoxic central nervous system damage.     

Cetuximab: an IgG1 monoclonal antibody for the treatment of epidermal growth factor receptor-expressing tumours

The epidermal growth factor receptor (EGFR) plays an essential role in normal cell growth and differentiation and in the survival of healthy and cancerous cells. EGFR expression is a common feature of non-haematological malignancies and is associated with poor clinical prognosis. Cetuximab is an IgG₁ monoclonal antibody that blocks EGFR activation. It has efficacy alone, and in combination with irinotecan, in the treatment of metastatic colorectal cancer that has progressed on irinotecan-containing therapy. It has been approved for use in combination with irinotecan in both Switzerland and the US and as monotherapy in the US. Cetuximab also has efficacy in cancers of the head and neck and non-small cell lung cancer. Cetuximab is well-tolerated and does not exacerbate the side effects of co-administered cytotoxic chemotherapy.     

Importance of cynomolgus monkeys in development of monoclonal antibody drugs

Animal species used in the preclinical studies for development of monoclonal antibody (mAb) drugs are surveyed in this review. Relevant animal species for preclinical studies of mAb candidates are those express desired epitope of mAb candidates. Cynomolgus monkeys cross-react with mAb drugs much higher than other animal species commonly used in preclinical studies such as absorption, distribution, metabolism and excretion (ADME), efficacy, and toxicity studies, for development of new drugs. Moreover, plasma exposure of the mAb drugs in humans is predicted well from the exposure in the monkeys, and the placental transfer of immunoglobulin G (IgG, all the mAb drugs contain IgG) from mother to fetus is similar between humans and the monkeys from a viewpoint of time course and plasma level of IgG transferred. These observed findings indicate that the monkeys are the most suitable animal species used in the ADME and toxicity studies for development of new mAb drugs.     

单克隆抗体SZ-95亲和色谱纯化人血小板第4因子

目的用单克隆抗体亲和色谱从人血小板破碎液中纯化血小板第 4因子 (PF4 )。方法将单克隆抗体SZ 95 IgG与溴化氰活化的Sepharose 4B凝胶连接成亲和色谱柱SZ 95 IgG Sepharose 4B ,人血小板破碎液经此亲和色谱柱上样后 ,经洗脱获得PF4 ,采用 15 %SDS 聚丙烯酰胺凝胶电泳鉴定其纯度 ,用点印迹鉴定其免疫活性。结果SZ 95 Sepharose 4B亲和色谱柱的偶联率为 72 % ,每 1ml(约 1× 10 9个血小板 )血小板破碎液中可以纯化到PF4 18μg ,其相对分子量约为 12kD ,经点印迹显示与单抗SZ 95反应显带。结论用SZ 95 Sepharose 4B亲和色谱柱纯化的PF4产品得率高、纯度高、活性好     

Developmental toxicity assessment of the new turf herbicide,methiozolin ([5-(2,6-difluorobenzyl)oxymethyl-5-methyl-3,3(3-methylthiophen-2-yl)-1,2-isoxazoline]), in rabbits

Methiozolin is a new herbicide to control annual bluegrass (Poa annua L.) and large crabgrass (Digitaria sanguinalis (L.) Scop.) in various turfgrasses. The potential of methiozolin to induce maternal and developmental toxicity was investigated in the pregnant New Zealand White Rabbits. Methiozolin was, at dose levels of 0, 125, 250 and 500 mg/kg/day, administered by oral gavage to artificially inseminated rabbits (25 females per group) from days 6 to 28 of gestation. All does were subjected to Cesarean section on day 29 of gestation. At 500 mg/kg/day, treatment-related toxicities including abortion (10/22), decreased mean body weight, weight gain, net body weight change, reduced food consumption and decreased fetal weight were observed. At 125 and 250 mg/kg/day, no signs of maternal and developmental toxicity were observed. There were no treatment-related external, visceral and skeletal abnormalities of fetuses at all doses tested. In the current experimental conditions, the no observed adverse effect levels (NOAELs) of methiozolin are considered to be 250 mg/kg/day for does and prenatal development.     

Characterization of CYP1A in hepatocytes of cynomolgus monkeys (Macaca fascicularis) and induction by different substituted polychlorinated biphenyls (PCBs)

Cynomolgus monkeys (Macaca fascicularis) have been used previously as a model to study effects on cytochrome P450 (CYP) regulation. Until now it has not been elucidated which CYP1A proteins are present in this primate species. The aim of this study was to characterize CYP1A in untreated hepatocytes of cynomolgus monkey using two specific CYP1A inhibitors (α -naphthoflavone and furafylline). The effect of different substituted polychlorinated biphenyls (PCBs) on CYP1A regulation was also studied in these hepatocytes. Small quantities of CYP1A2 have been identified in untreated hepatocytes. Northern blots showed the presence of a CYP1A mRNA in untreated hepatocytes, when hybridizations where performed with human CYP1A2 cDNA. Inhibitions with furafylline and α -naphthoflavone also suggested the presence of CYP1A2 properties. After induction with different PCBs, (probably) CYP1A1 mRNA and enzyme activity were induced in cynomolgus monkey hepatocytes. As expected, 2,3′,4,4′,5-PeCB (PCB no. 118), a mono-ortho substituted congener, was a potent CYP1A inducer but 2,2′,3,4,4′,5′,5′-HpCB (PCB no. 180), a di-ortho and 2,2′,3,4′,5,5′,6-HpCB (PCB no. 187), a tri-ortho substituted PCB, could induce CYP1A mRNA and enzyme activity in cynomolgus monkey hepatocytes as well. Received: 20 April 1998 / Accepted: 1 July 1998     

神经生长因子对抑郁症大鼠模型认知行为的影响研究

目的：探讨神经生长因子（NGF）对实验性抑郁症大鼠模型的认知行为的影响及其机制。方法：32只SD大鼠随机分为对照组、模型组、NGF低剂量组及NGF高剂量组,采用孤养及长期不可预见性刺激法建立抑郁症大鼠模型,NGF低剂量组及NGF高剂量组分别给予不同剂量NGF干预,采用T迷宫实验观察4组大鼠认知行为变化及海马区神经元数目变化。结果：模型组大鼠T迷宫实验错误次数增多,海马区神经元数目降低,NGF干预组T迷宫实验错误次数减少,海马神经元数目较模型组增多,并呈现剂量相关性。结论：NGF能够改善抑郁症大鼠模型认知行为障碍,其可能通过保护海马区神经元发挥作用。     

神经生长因子(NGF)对骨折愈合影响的初步临床观察

目的探讨神经生长因子(NGF)局部注射促进骨折愈合的临床作用。方法对48例四肢新鲜骨折病人进行骨折端经皮局部注射NGF促进骨折愈合的临床治疗观察,其中股骨骨折11例、肱骨骨折7例、胫腓骨骨折16例、尺桡骨骨折14例。据骨折类型、治疗方法相同或相似的原则设立相应的对照组,对骨痂生长情况、骨折临床愈合的时间进行对比研究分析。结果2组病例均获随访,时间为3~9个月,2组骨痂出现的时间、骨痂量的多少2周内无明显不同,3周后骨痂量治疗组多于对照组,治疗组各类型骨折临床愈合时间均较对照组有不同程度的缩短,疗效优于对照组,经统计学处理(P<0.01),2组对比有极显著差异。结论临床应用神经生长因子(NGF)经皮局部注射在骨折中后期有促进骨折愈合修复的作用。     

鼠抗人多聚体蛋白聚糖单克隆抗体对骨髓瘤细胞株生长的抑制作用

目的研究多聚体蛋白聚糖(syndecan-1,CD138)单克隆抗体(mAb)4B3对天然表达syndecan-1的人多发性骨髓瘤(MM)细胞XC-1和XG-2的生物学效应。方法采用间接免疫荧光标记和流式细胞术分析,通过4B3mAb观察XC-1和XC-2细胞膜上syndecan-1荧光标记阳性率;按常规培养方式对XC-1、XG-2进行细胞培养和台盼蓝染色记数,观察不同浓度的4B3mAb对XC-1和XG-2细胞体外生长作用。结果4B3mAb能识别XG-1和XG-2细胞表达的syndecan-1分子,表达率与商品化的鼠抗人syndecan-1 mAb(BB4)相似;同时,4133mAb与XG-1和XG-2细胞膜上syndecan-1荧光标记阳性率基本一致,反应均呈强阳性;加入不同浓度的4B3mAb,XC-1细胞数量2d后开始下降,而XG-2细胞3d后数量出现减少。结论4B3mAb抗体对XC-1和XG-2均具有明显的生长抑制作用,但XG-2细胞的生长抑制作用早于XC-1,可为MM的生物治疗提供新途径。     

Developmental toxicity and neurotoxicity of two matrine-type alkaloids,matrine and sophocarpine,in zebrafish (Danio rerio) embryos/larvae

Matrine and sophocarpine are two major matrine-type alkaloids included in the traditional Chinese medicine (TCM) Kushen (the root of Sophora flavescens Ait.). They have been widely used clinically in China, however with few reports concerning their potential toxicities. This study investigated the developmental toxicity and neurotoxicity of matrine and sophocarpine on zebrafish embryos/larvae from 0 to 96/120 h post fertilization (hpf). Both drugs displayed teratogenic and lethal effects with the EC₅₀ and LC₅₀ values at 145 and 240 mg/L for matrine and 87.1 and 166 mg/L for sophocarpine, respectively. Exposure of matrine and sophocarpine significantly altered spontaneous movement and inhibited swimming performance at concentrations below those causing lethality and malformations, indicating a neurotoxic potential of both drugs. The results are in agreement with most mammalian studies and clinical observations.     

Nonclinical safety of mavrilimumab,an anti-GMCSF receptor alpha monoclonal antibody,in cynomolgus monkeys: Relevance for human safety

Mavrilimumab (CAM-3001) is an investigational human IgG4 monoclonal antibody (MAb) targeting GM-CSF receptor alpha which is currently being developed for the treatment of RA. GM-CSF plays a central role in the pathogenesis of rheumatoid arthritis (RA) through the activation, differentiation, and survival of macrophages and neutrophils. To support clinical development, the nonclinical safety of mavrilimumab was evaluated in several studies with cynomolgus monkeys as the pharmacologically relevant species. Comprehensive toxicity parameters were assessed in each study, and treatment duration ranged from 4 to 26 weeks. Mavrilimumab has an acceptable safety profile in monkeys with no changes in any parameters other than microscopic findings in lung. In several studies, minimal accumulation of foamy alveolar macrophages was observed. This finding was only seen in studies of at least 11 weeks duration, was reversible following a dose-free recovery period and was considered non-adverse. At higher dose levels (≥ 30 mg/kg/week), in a 26-week repeat-IV dose study, the presence of lung foreign material, cholesterol clefts, and granulomatous inflammation was also observed in a few animals and was considered adverse. The dose- and time-related accumulation of foamy macrophages in lung following exposure to mavrilimumab observed in several NHP studies was expected based upon the known role of GM-CSFRα signaling in the function of alveolar macrophages. Overall, a clean no-observed-adverse-effect-level (NOAEL) without any effects in lung was established and provided adequate clinical safety margins. In clinical studies in RA patients, mavrilimumab has demonstrated good clinical activity with adequate safety to support further clinical development. A Phase 2b study of mavrilimumab in subjects with RA is in progress.     

神经生长因子治疗颈脊髓过伸性损伤20例疗效分析

目的评价神经生长因子(NGF)对颈脊髓过伸性损伤的疗效。方法40例颈脊髓过伸性损伤患者随机分为治疗组和对照组,分别以NGF及胞二磷胆碱治疗,观察两组疗效。结果肌力恢复:治疗组总有效率为93.8%,对照组为66.7%,两组比较差异有显著意义(P<0.05)。感觉恢复:治疗组总有效率95%,对照组为65%,两组比较差异有显著意义(P<0.05)。结论NGF对脊颈髓过伸性损伤疗效肯定,优于胞二磷胆碱。进一步证实外源性NGF可以提高神经损伤处内源性NGF含量和活性。     

Nerve growth factor (NGF) has novel antidepressant-like properties in rats

Nerve growth factor, a neurotrophin, may have other functions, including a role in depressive disorders. The present study sought to determine whether NGF would (1) have antidepressant-like effects and (2) behave similarly to or differently from other well-recognized antidepressants. Over a broad dose-range, NGF reduced the exaggerated swim test immobility exhibited by the Flinders Sensitive Line (FSL) rats, but at a standard dose of 40 ng/ml, it was not as effective as desipramine (DMI, 5 mg/kg). The low social interaction behavior and locomotor activity of the FSL rats were less affected by NGF than was the immobility. Acute treatment with NGF did not induce c-fos expression in brain regions known to be activated by other acute antidepressants. The fact that chronic treatment with DMI blunted the corticosterone response to fluoxetine was replicated in this study. However, chronic treatment with NGF did not alter this response. Similarly, chronic treatment with fluoxetine blunted 5-HT_1A and 5-HT_2A receptor-mediated responses, whereas chronic treatment with NGF was without effect. Thus, NGF has antidepressant-like effects but does not appear to have biochemical actions typical of other antidepressants.     

神经生长因子治疗新生儿缺氧缺血性脑病40例临床分析

目的：观察神经生长因子（细胞生长肽）肌内注射与胞二磷胆碱静脉滴注对照治疗新生儿缺氧缺血脑病的疗效比较。方法：治疗组在对照组的基础上给予神经生长因子10mg/d，肌肉注射，1次／d,10-20d为一疗程，结果：治疗组有效率87．5％，对照组有效率65．0％，两组比较（P＜0．05）有显著差异性，结论：神经生长因子治疗新生儿缺氧缺血性脑病，疗效较满意，使用方法，不良反应小，是一种有前途的治疗方法。     

Twenty-six-week repeat-dose toxicity study of a recombinant human granulocyte colony-stimulating factor derivative (nartograstim) in cynomolgus monkeys.

An rhG-CSF derivative, nartograstim (NTG), at dose levels of 0 (saline), 0.1, 1, 10, and 100 microg/kg, was administered subcutaneously to groups of 3 male and 3 female cynomolgus monkeys once daily for 26 weeks to investigate its toxicity. In Week 4 or later, an increase in leukocyte counts consisting mainly of neutrophils was noted in all NTG dose groups, and was considered to be attributable to the pharmacological action of NTG. The degree of this increase was reduced with repetition of dosing. Increases in granulocytic cells and granulocytic cells/erythrocytic cells (G/E) ratio in the bone marrow, increase in serum ALP activity, and enlarged spleens with increase of neutrophils in the red pulp were observed at 10 microg/kg and higher. Anemia was noted at 10 microg/kg and higher in Week 4 and was accompanied by an increase in reticulocytes and a decrease in total cholesterol level at 100 microg/kg. Anti-NTG antibody was detected in 1 female at 100 microg/kg, but neutralizing antibodies were not detected at any dose levels in Week 4. In Weeks 13 and 26, these antibodies were detected sporadically at all dose levels. However, there were considerable individual variations in antibody titer, and no definite correlation could be found between the dose levels and the antibody titer. Seven NTG-dosed animals including 3 high dose-group animals showed obvious increases in leukocyte counts until Week 26 but no obvious elevation of anti-NTG or neutralizing antibody. In these animals, changes including anemia became slighter but were still observed in Week 26. Under the conditions in this study, 1 microg/kg was concluded to be the no-observed-adverse-effect level (NOAEL) in cynomolgus monkeys.