Review of embryo-fetal developmental toxicity studies performed for recent FDA-approved pharmaceuticals

Details of embryo-fetal development (EFD) studies were compiled from published FDA approval documents for 43 small molecule drugs (2014–2015) and 37 monoclonal antibodies (mAbs, 2002–2015). Anti-cancer agents were analyzed separately. Rats and rabbits were the species used for EFD studies on 93% of small molecule drugs. Overall, the rat and rabbit were equally sensitive to maternal and fetal toxicity (including teratogenicity). Dosages equivalent to more than 50-times the human exposure (or 10-times for mAbs) were frequently used, but were unnecessary for 90% of drugs. EFD studies were not required for several recently approved mAbs owing to pre-existing scientific knowledge. The cynomolgus monkey was used for developmental toxicity testing of 75% of mAbs, frequently using an ePPND study design. Studies in pregnant rodents using homologous murine antibodies supplemented or replaced monkey studies under some circumstances. Most anti-cancer small molecules and mAbs were tested for developmental toxicity in at least one species.     

A reconsideration of the relevance of recent animal studies for development of treatment procedures for alcoholics

Amit and Sutherland's conclusions concerning the use of conditioned taste aversions for alcoholism treatment are critically evaluated. Their conclusion that painful electric shock is contraindicated as a basis for alcohol taste aversions is consistent with the animal and human literature which depicts nausea as a more biologically appropriate US for taste aversion formation. However, Amit and Sutherland also conclude that alcoholics will not develop illness-induced alcohol aversions because animal studies show that aversion acquisition is disrupted by preconditioning familiarity with the conditioned stimulus (CS) — flavor — or unconditioned stimulus (US) — illness. This conclusion is untenable because Amit and Sutherland only considered animal conditioning methods that differed markedly from aversion therapy practices. Other animal studies modeled after aversion therapy procedures clearly show CS and US preexposure effects to be transitory phenomena. Moreover, experimental and clinical data show humans to be quite susceptible to taste aversion formation, and that many alcoholics do form strong alcohol aversions under appropriate conditioning parameters. Additional implications of the animal literature for effective aversion therapy are explored, and the paper concludes with a discussion of covert sensitization, a promising verbal aversion therapy which has resulted in the development of strong alcohol aversions in many volunteer subjects at the Augusta Veterans Administration Medical Center.     

Review of recent toxicology studies on p-dichlorobenzene

Results from recent long-term inhalation, mutagenicity, embryotoxicity and metabolism studies on p-dichlorobenzene (p-DCB) are reviewed. Groups of male and female rats and female mice were exposed for 5 hr/day on 5 days/wk to p-DCB at concentrations of 0, 75 or 500 ppm for a total period of c. 76 wk (rats) or 57 wk (female mice) followed by 36 wk (rats) or 19 wk (female mice) without p-DCB exposure. No overt signs of toxicity were apparent at any exposure level nor were there treatment-related effects on the biochemical determinations, urine analyses or haematological parameters. Slightly elevated urinary coproporphyrin excretion and increased liver and kidney weights were regarded as treatment-related effects in the 500-ppm exposure group of the rats. The non-tumour and tumour pathology did not indicate any treatment-related effect in any group of either species. An embryotoxicity and teratology study on rats exposed to 0, 75, 200 or 500 ppm p-DCB vapour in air during the period of organogenesis did not demonstrate any signs of embryo- or foetotoxicity or teratogenicity at any exposure level. In a series of mutagenicity tests including the Salmonella typhimurium, dominant lethal and cytogenetic assays, p-DCB did not produce a mutagenic response. Studies using oral or inhalation routes of exposure demonstrated rapid metabolic transformation of p-DCB and excretion of the products, even after long-term exposure.     

Cholinergic hypothesis of human memory: Review of basic and clinical studies

Recent neurochemical data from autopsy and biopsy studies of Alzheimer patients lends strong support for a selective impairment of cholinergic functioning. A number of pharmacological trials are described testing the effects of cholinergic agonists and antagonists on cognitive functions of both young normal volunteers as well as geriatric, demented patients. Paradigms included prelearning and postlearning pharmacological manipulations as well as testing the effect of a combination of a cholinergic precursor (lecithin) plus an anticholinesterase (tetrahydroaminoacridine). The results indicated great variability of responses among subjects, with mild-to-moderate enhancement of memory recall in specific subgroups of normal controls and demented patients. Further research should be aimed at developing reliable in vivo cholinergic markers and validation of these measures as response predictors of specific cholinergic treatments in patients with dementia.     

Selection of subjects for human milk surveillance and research studies

Subject selection for studies investigating environmental chemicals in human milk requires thoughtful consideration of multiple factors. Studies often need to produce generalizable and representative data that can be compared to other similar studies. Goals of the study determine the selection of subjects. Exposure is only one factor that influences the levels of environmental chemicals in human milk. Many maternal and infant characteristics should be considered in subject selection. Collection procedures of human milk samples also affect subject selection, as subject burden may be enough to reduce compliance with the collection protocols. Decisions must be made about pooling of samples both within subjects and within populations. Guidelines for subject selection are provided by the World Health Organization for human milk monitoring, but distinct differences in the lactational practices, geography, and ethnic and racial diversity of the U.S. population require somewhat different approaches.     

Pharmacokinetic and clinical studies on amphetamine dependent subjects

Summary Eighteen subjects with amphetamine psychosis were studied with respect to fluid balance, intensity and duration of psychotic symptoms, urinary and plasma amphetamine levels and the relative amounts of unchanged drug and metabolites in urine. On admission to hospital about half of the psychotic patients were dehydrated, the water lack being up to 6.7% of total body weight. The dehydrated subjects had lower renal clearances of amphetamine because of lower rates of urine production. As noted previously there was a strongly positive correlation between urinary pH and the half life (T _1/2) of plasma amphetamine, with an increase inT _1/2 of about 7 h for every unit increase in urinary pH. Patients with alkaline urine had intense psychoses lasting for about 4 1/2 days after the last dose of amphetamine. In patiens with acid urine, the psychotic symptoms were milder, and of about 2 days duration. No correlation was found between the degree of psychosis in different subjects and the plasma levels of the drug. — The ratio between the amounts of labelled metabolites and unchanged drug excreted in urine rose for each day after administration of³H-amphetamine, implying a slower excretion rate for the metabolites than for the parent drug. The relative proportion of metabolites was higher in patients with an alkaline urine, being more than 90% after the first day. — When amphetamine (200 mg i.v.) was given to nonpsychotic, dependent subjects, the peak plasma levels (mean 423 ng/ml) exceeded the highest levels observed during the first day in psychotic patients. However, no psychotic symptoms were observed in these subjects. The volumes of distribution calculated from the monoexponential elimination curves were higher than those previously reported in nondependent subjects. — With an alkaline urine a group of nonpsychotic amphetamine-dependent subjects had significantly longer plasmaT _1/2 (p<0.05) than a group of drug-naive control subjects. The results suggest that increased tissue binding may be a component in tolerance to amphetamine in dependent humans.     

Estimation of human maximum tolerable intake for methylmercury based on two recent studies in monkeys

Results of long-term toxicity studies of methylmercury (MeHg) in monkeys have been reported. The aim of this study was to estimate the threshold body burden, blood level and threshold daily intake (TDI) of MeHg for monkey and human. The concepts of this study stood on that body burden of MeHg would follow the accumulation theory, and that the more intake of MeHg, the earlier the neurotoxicity appeared, vice versa. The threshold blood level (TBL) of monkey was estimated to be 0.71 as Hg mg/L and the body burden was estimated to be 4.83 as Hg mg/kg. The TDI was estimated to be 0.025 as Hg mg/kg day. In human, the TBL was estimated with compensation by elimination constants of human and monkey. The blood threshold limit and TDI of human were estimated to be 0.33 as Hg mg/L and 0.0046 as Hg mg/kg day, respectively. The estimated body burden was 0.46 as Hg mg/kg.     

Synthesis of isotopically labeled daclatasvir for use in human clinical studies

Daclatasvir is a novel hepatitis C virus NS5A inhibitor developed by Bristol‐Myers Squibb and marketed as Daklinza®. The need to support the development of daclatasvir required the synthesis of carbon‐14 labeled material for use in human absorption, distribution, metabolism, and excretion studies. A total of 7.53 mCi of [¹⁴C]‐daclatasvir was synthesized in eight steps from commercially available [¹⁴C]‐copper cyanide. The radiochemical purity was 99.6%, and specific activity was 3.86 μCi/mg. To support a human absolute bioavailability study, 5.56 g of [¹³C₂, ¹⁵N₄]‐daclatasvir was synthesized in four steps.     

Analytical methods of biological monitoring for exposure to pesticides: recent update

Extensive use of synthetic pesticides for agricultural and nonagricultural purposes began in the past 50 years. As a result of their wide and extensive application, exposure to hazardous pesticides is a concern to the general population and occupationally exposed persons. Robust methods are therefore needed for measuring markers of pesticide exposure. This article presents a review of the most recently published analytical methodologies and instrumentations developed for and applied to biological monitoring of exposure to pesticides of various classes. Most of the methods reviewed here are based on chromatography combined with mass spectrometry detection. This work clearly demonstrates that although gas chromatography still appears to be the most widely employed technique for pesticide analysis in various biological samples, recently a trend has been observed toward the use of liquid chromatography coupled with tandem mass spectrometry.     

HPLC assay for albendazole and metabolites in human plasma for clinical pharmacokinetic studies

A sensitive and selective HPLC chromatography method using UV detection (295 nm) was developed for the determination of albendazole, albendazole sulfoxide (ABZSO), and albendazole sulfone (ABZSO2) in human plasma. Albendazole, ABZSO, ABZSO2, and the internal standard, oxibendazole, were extracted from human plasma by loading onto a conditioned C(18) SPE cartridge, rinsing with 15% methanol, and eluting with 90% methanol. Samples were evaporated under a stream of nitrogen, reconstituted with mobile phase, 1.25% triethylamine in water-methanol-acetonitrile (72:15:13, v/v/v) (pH* 3.1), and injected onto a Waters muBondapak Phenyl 3.9 x 300 mm HPLC column. Mobile phase flow rate was 1.0 ml/min. The retention times of albendazole, ABZSO, ABZSO2, and the internal standard were approximately 24.4, 7.9, 13.4, and 11.3 min, respectively. Total run time was 30 min. The assay was linear for concentration ranges in human plasma of 20-600 ng/ml for albendazole, 20-1000 ng/ml for ABZSO, and 20-300 ng/ml for ABZSO2. The analysis of quality control samples demonstrated excellent precision. Coefficients of variation for albendazole (20, 400, 600 ng/ml) were 6.7, 8.1 and 7.0%; ABZSO (20, 400, 800 ng/ml) were 6.0, 8.5 and 5.9%; ABZSO2 (20, 150, 300 ng/ml) were 3.1, 3.9 and 2.3%, respectively. The method appears to be robust and has been applied to a pharmacokinetic study of albendazole in healthy volunteers.     

Neuropsychiatric evaluation in subjects chronically exposed to organophosphate pesticides.

Long-term exposure to low levels of organophosphate pesticides (OP) may produce neuropsychiatric symptoms. We performed clinical, neuropsychiatric, and laboratory evaluations of 37 workers involved in family agriculture of tobacco from southern Brazil who had been exposed to OP for 3 months, and in 25 of these workers, after 3 months without exposure to OP. Plasma acetylcholinesterase activity levels of all subjects were within the normal range (3.2 to 9.0 U/l) and were not different between on- and off-exposure periods (4.7 +/- 0.9 and 4.5 +/- 1.1 U/l, respectively). Clinically significant extrapyramidal symptoms were present in 12 of 25 subjects, which is unexpected in such a population. There was a significant reduction of extrapyramidal symptoms after 3 months without exposure to OP, but 10 subjects still had significant parkinsonism. Mini-mental and word span scores were within the expected range for this population and were not influenced by exposure to OP. Eighteen of the 37 subjects (48%) had current psychiatric diagnoses in the first interview (13 with generalized anxiety disorder and 8 with major depression). Among the 25 subjects who completed both evaluations, the total number of current psychiatric diagnoses, after 3 months without using OP, dropped from 24 to 13 and the number of affected individuals with any psychiatric diagnosis dropped from 11 to 7. In conclusion, this study reinforces the need for parameters other than acetylcholinesterase activity to monitor for chronic consequences of chronic low-dose OP exposure, and it suggests that subjects have not only transient motor and psychiatric consequences while exposed, but may also develop enduring extrapyramidal symptoms.     

An efficient HPLC method for the quantitative determination of atazanavir in human plasma suitable for bioequivalence and pharmacokinetic studies in healthy human subjects

An efficient HPLC method for the determination of atazanavir in human plasma has been developed and validated. A relatively simple mobile phase consisting of acetonitrile–ammonium formate buffer (pH 3; 10 mM) (45:55, v/v) was pumped at a low flow rate of 0.3 ml/min through a reverse phase Phenomenex^® Luna C18 (2) (5 μm, 150 mm × 2.0 mm i.d.) column maintained at 30 °C. Diazepam was used as an internal standard and the eluent was monitored at 210 nm. The major advantage of this method over previously reported procedures is that the narrow-bore HPLC column used resulted in relatively short retention times for the internal standard (6.8 min) and atazanavir (8.3 min) with excellent peak resolution and associated reduction in solvent usage. Sample preparation involved liquid–liquid extraction using 400 μl plasma treated with sodium carbonate (2 M) and extracted with a mixed organic solvent consisting of ethyl acetate–n-hexane (50:50, v/v). The organic layer was removed and evaporated to dryness under nitrogen. Samples were reconstituted in mobile phase (100 μl) and 20 μl was injected onto the column. The procedures were validated according to international standards with good reproducibility and linear response with correlation coefficients (r) consistently ≥0.999. The intra- and inter-day accuracies were 97.1 ± 5.04 and 98.0 ± 11.3 respectively at the LLOQ and between 101 ± 4.48% and 104 ± 2.09% for the QC samples. The intra- and inter-day precision were ≤11.6% RSD at the LLOQ and ≤6.78% RSD across the entire QC concentration range. Mean recovery based on high, medium and low quality control standards ranged between 94.4 ± 1.07% and 100 ± 2.22%. Plasma samples were evaluated under short-term (ambient temperature for 6 h) and long-term (−10 ± 2 °C for 2 months) storage conditions and were found to be stable. The method described is efficient and has the necessary accuracy and precision for the rapid quantitative determination of atazanavir in human plasma and is thus highly suitable for use in pharmacokinetic/bioavailability/bioequivalence studies in healthy human subjects.     

临床试验机构研究者标准操作规程

依据SFDA《药品临床试验管理规范》．ICH GCP，WHO GCP，以及我院临床试验的实践，制订临床试验机构研究者的标准操作规程，包括研究者资格与条件，试验前的准备，受试者的招募和筛选，受试者的知情同意，方案的依从性，受试者的医疗．随机化程序和破盲，安全性报告，源文件和源数据．病例报告表，试验用药的管理，试验的终止或暂停，进展报告和总结报告，档案等。     

Pharmacodynamic interaction studies of Ginkgo biloba with cilostazol and clopidogrel in healthy human subjects

AIMS: Ginkgo biloba is available as an over-the-counter drug and reported to cause haemorrhage when coadministered with other antiplatelet agents. We set out to study the interactions of G. biloba with cilostazol and clopidogrel. METHODS: A randomized, open-label, crossover study of 10 healthy male volunteers. The dosage schedules were 120 mg G. biloba, 240 mg G. biloba, 100 mg cilostazol, 200 mg cilostazol, 75 mg clopidogrel, 150 mg clopidogrel, 120 mg G. biloba+ 100 mg cilostazol and 120 mg G. biloba+ 75 mg clopidogrel. Platelet aggregation, platelet count, bleeding time and clotting time were measured 0 and 6 h after drug administration. Platelet aggregation was performed using a dual channel aggregometer, by the turbimetric technique using adenosine diphosphate 5 micromol and 10 micromol, and collagen 1 microg ml(-1). RESULTS: Platelet inhibition with the combination of G. biloba and clopidogrel or cilostazol was not statistically significant compared with individual doses of drugs, with all the three aggregants. There was significant (P < 0.05) potentiation of prolongation of bleeding time with the combination of cilostazol and G. biloba compared with individual doses of both the drugs. There was no significant change in clotting time and platelet count. CONCLUSIONS: Coadministration of G. biloba either with cilostazol or clopidogrel did not enhance antiplatelet activity compared with individual agents. Ginkgo biloba potentiated the bleeding time prolongation effect of cilostazol. There was no significant correlation between prolongation of bleeding time and inhibition of platelet aggregation.     

Gas chromatographic assay of diethylcarbamazine in human plasma for application to clinical pharmacokinetic studies

A sensitive and selective gas chromatography method using flame ionization detection was developed for the determination of diethylcarbamazine (DEC) in human plasma. DEC and the internal standard, 1-diethylcarbamyl-4-ethyl piperazine HCl (E-DEC), were extracted from human plasma after loading onto a conditioned C(18) solid phase extraction cartridge, rinsed with water and eluted with methanol. After evaporation under a stream of nitrogen and reconstitution in methanol, 3 microl were injected onto the GC system. Separation was achieved on a A Heliflex(R) AT-35 capillary column (length 30 m, internal diameter 0.32 mm). Gas flow rates were: hydrogen, 35 ml/min; carrier gas (helium), 1.5 ml/min, make-up gas (helium), 25 ml/min; and air 420 ml/min. The retention times of DEC and internal standard were approximately 5.5 and 7.28 min, respectively. The GC run time was 22 min. The assay was linear in concentration range 100-2000 ng/ml for DEC in human plasma. The analysis of quality control samples for DEC (120, 1000, 2000 ng/ml) demonstrated excellent precision with coefficients of variation of 4.5,1.3, and 1.6%, respectively (n=6). The method was accurate with all intra-day (n=6) and inter-day (n=12) mean concentrations within 4.3% from nominal at all quality control sample concentrations. DEC was found to be stable after 3 freeze-thaw cycles, and with storage at -20 degrees C for 12 weeks. The method is currently being used for pharmacokinetic studies of DEC in healthy volunteers.