Efficacy and safety evaluation of black ginseng (Panax ginseng C.A. Mey.) extract (CJ EnerG): broad spectrum cytotoxic activity in human cancer cell lines and 28-day repeated oral toxicity study in Sprague-Dawley rats.

BACKGROUND: Ginseng (Panax ginseng C.A. Mey.) has been used as a valuable ingredient in traditional medicine for thousands of years mostly in Asian countries due to its therapeutic effects in various diseases. Among the processed ginseng products, black ginseng is produced by a repeated steaming and drying process of ginseng roots and has been known for its superior efficacy based on high accumulation of minor ginsenosides as recently discovered. Despite its popularity and increasing use, the toxicity information on black ginseng still remained largely lacking, raising safety concerns. This study was therefore carried out to determine the repeated oral toxicity of black ginseng extract (BGE; CJ EnerG) with evaluation of cytotoxic activity as validation of its pharmacological activity for toxicity testing. METHODS: Prior to the toxicity test, we examined the cytotoxicity of BGE in six cancer cell lines derived from distinct human tissues in comparison with red ginseng extract (RGE), ginsenosides Rg5 and 20(S)-Rg3, and then assessed 28-day repeated oral toxicity in Sprague-Dawley (SD) rats using daily administration of up to 2000 mg/kg BGE. RESULTS: BGE showed higher cytotoxicity than RGE in all the cell lines used in this study. Interestingly, the efficacy of BGE closely resembled the cytotoxic pattern of Rg5, suggesting Rg5 as the main effector in the cytotoxic activity of BGE. During the toxicity study, BGE-treated groups showed no noticeable abnormality in clinical signs, body weight gain, food and water consumption and urinalysis. Furthermore, hematological, serum biochemical and histopathological analyses did not find any BGE-related toxicity. CONCLUSION: Our findings demonstrated that BGE has broad-spectrum in vitro cytotoxic activity, and that NOAEL of BGE in SD rats is > 2000 mg/kg, providing the essential safety information for human consumption.

Double lock of a potent human therapeutic monoclonal antibody against SARS-CoV-2.

Receptor recognition and subsequent membrane fusion are essential for the establishment of successful infection by SARS-CoV-2. Halting these steps can cure COVID-19. Here we have identified and characterized a potent human monoclonal antibody, HB27, that blocks SARS-CoV-2 attachment to its cellular receptor at sub-nM concentrations. Remarkably, HB27 can also prevent SARS-CoV-2 membrane fusion. Consequently, a single dose of HB27 conferred effective protection against SARS-CoV-2 in two established mouse models. Rhesus macaques showed no obvious adverse events when administrated with 10 times the effective dose of HB27. Cryo-EM studies on complex of SARS-CoV-2 trimeric S with HB27 Fab reveal that three Fab fragments work synergistically to occlude SARS-CoV-2 from binding to the ACE2 receptor. Binding of the antibody also restrains any further conformational changes of the receptor binding domain, possibly interfering with progression from the prefusion to the postfusion stage. These results suggest that HB27 is a promising candidate for immuno-therapies against COVID-19.

S(+)-(2E)-N-(2-Hydroxypropyl)-3-Phenylprop-2-Enamide (KM-568): A Novel Cinnamamide Derivative with Anticonvulsant Activity in Animal Models of Seizures and Epilepsy.

Epilepsy is one of the most frequent neurological disorders affecting about 1% of the world's human population. Despite availability of multiple treatment options including antiseizure drugs, it is estimated that about 30% of seizures still remain resistant to pharmacotherapy. Searching for new antiseizure and antiepileptic agents constitutes an important issue within modern medicinal chemistry. Cinnamamide derivatives were identified in preclinical as well as clinical studies as important drug candidates for the treatment of epilepsy. The cinnamamide derivative presented here: S(+)-(2E)-N-(2-hydroxypropyl)-3-phenylprop-2-enamide (S(+)-N-(2-hydroxypropyl)cinnamamide, compound KM-568) showed anticonvulsant activity in several models of epilepsy and seizures in mice and rats. It was active in a genetic animal model of epilepsy (Frings audiogenic seizure-susceptible mouse model, ED50 = 13.21 mg/kg, i.p.), acute seizures induced electrically (maximal electroshock test ED50 = 44.46 mg/kg mice i.p., ED50 = 86.6 mg/kg mice p.o., ED50 = 27.58 mg/kg rats i.p., ED50 = 30.81 mg/kg rats p.o., 6-Hz psychomotor seizure model 32 mA ED50 = 71.55 mg/kg mice i.p., 44 mA ED50 = 114.4 mg/kg mice i.p.), chronic seizures induced electrically (corneal kindled mouse model ED50 = 79.17 mg/kg i.p., hippocampal kindled rat model ED50 = 24.21 mg/kg i.p., lamotrigine-resistant amygdala kindled seizure model in rats ED50 = 58.59 mg/kg i.p.), acute seizures induced chemically (subcutaneous metrazol seizure threshold test ED50 = 104.29 mg/kg mice i.p., ED50 = 107.27 mg/kg mice p.o., ED50 = 41.72 mg/kg rats i.p., seizures induced by picrotoxin in mice ED50 = 94.11 mg/kg i.p.) and the pilocarpine-induced status epilepticus model in rats (ED50 = 279.45 mg/kg i.p., ED97 = 498.2 mg/kg i.p.). The chemical structure of the compound including configuration of the chiral center was confirmed by NMR spectroscopy, LC/MS spectroscopy, elemental analysis, and crystallography. Compound KM-568 was identified as a moderately stable derivative in an in vitro mouse liver microsome system. According to the Ames microplate format mutagenicity assay performed, KM-568 was not a base substitution or frameshift mutagen. Cytotoxicity evaluation in two cell lines (HepG2 and H9c2) proved the safety of the compound in concentrations up to 100 µM. Based on the results of anticonvulsant activity and safety profile, S(+)-(2E)-N-(2-hydroxypropyl)-3-phenylprop-2-enamide could be proposed as a new lead compound for further preclinical studies on novel treatment options for epilepsy.

Acute and repeated dose 26-week oral toxicity study of 20(S)-ginsenoside Rg3 in Kunming mice and Sprague-Dawley rats.

BACKGROUND: 20(S)-ginsenoside-Rg3 (C42H72O13), a natural triterpenoid saponin, is extracted from red ginseng. The increasing use of 20(S)-ginsenoside Rg3 has raised product safety concerns. METHODS: In acute toxicity, 20(S)-ginsenoside Rg3 was singly and orally administrated to Kunming mice and Sprague-Dawley (SD) rats at the maximum doses of 1600 mg/kg and 800 mg/kg, respectively. In the 26-week toxicity study, we used repeated oral administration of 20(S)-ginsenoside Rg3 in SD rats over 26 weeks at doses of 0, 20, 60, or 180 mg/kg. Moreover, a 4-week recovery period was scheduled to observe the persistence, delayed occurrence, and reversibility of toxic effects. RESULTS: The result of acute toxicity shows that oral administration of 20(S)-ginsenoside Rg3 to mice and rats did not induce mortality or toxicity up to 1600 and 800 mg/kg, respectively. During a 26-week administration period and a 4-week withdrawal period (recovery period), there were no significant differences in clinical signs, body weight, food consumption, urinalysis parameters, biochemical and hematological values, or histopathological findings. CONCLUSION: The mean oral lethal dose (LD50) of 20(S)-ginsenoside Rg3, in acute toxicity, is above 1600 mg/kg and 800 mg/kg in mice and rats, respectively. In a repeated-dose 26-week oral toxicity study, the no-observed-adverse-effect level for female and male SD rats was 180 mg/kg.

Repeated-dose 26-week oral toxicity study of ginsenoside compound K in Beagle dogs.

ETHNOPHARMACOLOGICAL RELEVANCE: Ginsenoside compound K (CK), a product produced by the intestinal bacteria-mediated breakdown of ginsenoside, exhibits a wide array of pharmacological activities against diverse targets. However, few of preclinical safety evaluation of CK is reported. AIMS OF THE STUDY: The present study therefore sought to assess the toxicity of oral CK in Beagle dogs over a 26-week period. MATERIAL AND METHODS: All dogs received 4, 12, or 36 mg/kg oral CK doses for 26 weeks with regular monitoring, followed by a 4-week recovery period. Animals were monitored through measurements of temperature, weight, food intake, blood chemistry and hematological findings, electrocardiogram (ECG) measurements, urinalysis, gross necropsy and organ weight and tissue histopathology. RESULTS: Animals in the 36 mg/kg group exhibited an apparent reduction in body weight over the study period, in addition to the presence of focal liver necrosis and increased plasma enzyme levels (alanine aminotransferase, ALT; alkaline phosphatase, ALP) consistent with hepatotoxicity, although there was some evidence suggesting this toxicity was reversible. Animals in the 4 and 12 mg/kg groups did not exhibit any apparent toxicity for any measured parameters. CONCLUSION: These results thus indicate that the no observed adverse effect level (NOAEL) in dogs is 12 mg/kg.

Examination and assessment of target organ toxicity in toxicologic pathology of preclinical safety evaluation of drugs / 药物评价研究

Examination and assessment of target organ toxicity in toxicologic pathology of preclinical safety evaluation of drugs should combine the results of the gross pathology,histopathology and clinical pathology examination data in a well-considered,stepwise approach.In addition,the nomenclature and diagnostic criteria recommended by INHAND should be used to avoid subjective and inappropriate diagnosis.In this paper,we briefly introduced the basic principles for the examination of organ toxicity in toxicology studies,gross pathology,histopathology,diagnostic approach,procedures,and considerations,international harmonization of diagnostic term and criteria,clinical pathology parameters analysis,results of a well-concerted combination of anatomical and clinical pathology data so as to provide some reference for the examination and assessment of target organ toxicity in toxicologic pathology in the field ofpreclinical safety evaluation of drugs in China.

Major factors influencing quality of neuropathology evaluation of drug toxicology / 药物评价研究

Neurotoxicity is one common adverse effect caused by many drugs or compounds.In the early phase of new drug development,it is necessary to screen for neurotoxicants.Neurotoxicity studies in nonhuman primates (NHP) are used to evaluate the neurotoxicity of small-molecule drugs or vaccines that may affect the nervous system across the blood-brain barrier during preclinical safety assessment.Toxicologic pathological evaluation or neuropathological examination is the "gold standard" for the evaluation of drug neurotoxicity in preclinical drug safety studies.In this paper,the majory factors influencing the quality of neuropathology evaluation in toxicology,including the general strategy of neuropathology evaluation,the optimal timing of evaluation,the specific blood-brain barrier in the nervous system,the method of sampling in the histopathology of nerve tissue,and the interference of artificial artifacts in diagnosis of neuropathology,were detailly analyzed in order to provide a reference for setting guidelines of neurotoxicity risk assessment in China and pathologists and toxicologists engaged in nonclinical neurotoxicity studies.

Validation of computerized system in facilities for preclinical safety evaluation of drugs / 药物评价研究

Computerized system has been played an increasingly important role in preclinical safety evaluation of drugs and has been used directly or indirectly for data acquisition,processing,reporting as well as raw data storage.However,the computerized system has not been widely used in facilities for preclinical safety evaluation of drugs or only some functions of modules of computerized system have been used.Based on the current application status of the computerized system in the facilities for preclinical safety evaluation of drugs in China,this paper briefly introduced the following aspects about validation of computerized system,such as GLP regulatory requirements of validation of the computerized system,validation process of the computerized system,maintenance of validation state of the computerized system,safety precautions of performance of the computerized system,as well as electronic records and electronic signatures with the purpose to provide some references for carrying out and speeding up the validation of computerized system and to further improve the efficiency of the computerized system in facilities for preclinical safety evaluation of drugs in China and to be in line with international practice.

Preclinical safety evaluation of recombinant adeno-associated virus 2 vector encoding human tumor necrosis factor receptor-immunoglobulin Fc fusion gene.

Recombinant adeno-associated virus (rAAV) 2 vector gene therapy offers promise for the healing of Rheumatoid arthritis. To support the clinical development of the candidate gene therapeutic product in China, a comprehensive preclinical safety assessment of rAAV2 encoding human TNF receptor-immunoglobulin Fc fusion gene (rAAV2/human TNFR:Fc), were conducted in 3 species of experimental animals. No abnormal findings were observed in mice following single intravenous administration with test article. Compared with the control group, no differences in mean body weight, food consumption in rats and monkeys following the repeated intraarticular administration with rAAV2/human TNFR:Fc. There were also no significant adverse effects due to treatment noted by clinical chemistry, hematology and pathology assessments. After intraarticular administration with rAAV2/human TNFR:Fc, the vector DNA initially distributed to spleen, lymph nodes, and joint synovium. The vector DNA cleared rapidly as it could be detected mainly at the site of injection by 91 d post-administration (182 d for monkey). Taken together, localized delivery of rAAV2/human TNFR:Fc showed no significant toxicity in mice, rats, and monkeys, which support the planned clinical evaluation of this product.