Avaliação toxicológica da exposição à Cannabis e cocaína na gravidez em cordão umbilical humano: validação de método analítico e prospecção de biomarcadores proteicos de toxicidade / Toxicological assessment of Cannabis and cocaine exposure during pregnancy in human umbilical cord tissue: analytical method validation and prospection of protein biomarkers related to fetotoxicity

O abuso de drogas atinge aproximadamente 35 milhões de pessoas em todo planeta, sendo um problema alarmante em decorrência de graves danos à saúde, como a dependência química e intoxicações fatais. No Brasil, o número de usuários tem crescido principalmente para o consumo de produtos da Cannabis e cocaína, drogas amplamente consumidas, inclusive entre mulheres em período gestacional, trazendo à tona um novo grupo de risco. A exposição gestacional a drogas de abuso está diretamente relacionada a malformações fetais e complicações de saúde para mãe e bebê nos períodos pré- e pós-natal. Tradicionalmente, a avaliação toxicológica da exposição é realizada pela detecção da droga parental e de seus produtos de biotransformação em matrizes materno-fetais por meio de métodos bioanalíticos. Entretanto, estes ensaios não fornecem informações acerca dos impactos fisiológicos ocasionados pela exposição, deixando uma lacuna no que tange às informações sobre os mecanismos e moléculas subjacentes envolvidos em processos de toxicidade. Desse modo, o desenvolvimento de análises toxicológicas mais robustas utilizando tecnologia de ponta, que possam comprovar o uso drogas e também elucidar aspectos de toxicidade é de suma importância, pois auxiliam na compreensão do impacto biológico relativo à exposição humana a xenobióticos. Neste trabalho foram desenvolvidos ensaios bioanalíticos, utilizando o tecido do cordão umbilical para a avaliação da exposição in utero à canabinoides. Foi desenvolvido e validado método QuECheRS adaptado como preparo de amostra, no qual etapas simultâneas de extração e hidrólise alcalina de canabinoides são alcançadas, utilizando cromatografia em fase gasosa acoplada a espectrômetro de massas para detecção de delta-9-tetraidrocanabinol (THC), canabinol (CBN), 11-hidroxi-delta-9-tetraidrocanabinol (11-OHTHC) e 11-nor-9-carboxi-tetrahidrocanabinol (THC-COOH). Também foram desenvolvidas metodologias utilizando LC-MS/MS e Trapped Ion Mobility Mass Spectrometry para análise de proteoma de cordão umbilical humano em diferentes regiões, no intuito de identificar biomarcadores proteicos relativos à fetotoxicidade do uso de drogas na gravidez. Até o presente momento, QuECheRS é utilizado pela primeira vez como abordagem bioanalítica para avaliação de drogas ilícitas em matrizes teciduais materno-fetais e mostrou-se satisfatório para detecção de produtos da Cannabis. Nos ensaios proteômicos, foram identificados potenciais biomarcadores de fetotoxicidade, como as moléculas ACTA 2, Collagen alpha-1 (XVIII), SMC1A, KNL1, KMT2A, em tecidos expostos à Cannabis e/ou cocaína. Tais macromoléculas estão correlacionadas a malformações embriogênicas e complicações de saúde na vida intra-uterina. As metodologias desenvolvidas neste trabalho podem ser úteis para uma melhor avaliação da toxicidade do uso de drogas na gravidez, fornecendo novas pistas sobre a exposição e/ou efeitos tóxicos significativos considerados na avaliação de risco

Drug abuse affects approximately 35 million people worldwide and can be considered a significant burden on society due to severe health problems, e.g. drug addiction and fatal poisonings. In Brazil, the number of users has been growing related to Cannabis and cocaine products, drugs widely used, including among women in gestational period, bringing up a new risk group. Gestational exposure to drugs of abuse is directly related to fetal malformations and health complications for mother and babies in the pre- and postnatal periods. Traditionally, toxicological assessment of exposure is performed by detecting the parent drug and its biotransformation products in maternal-fetal matrices using bioanalytical methods. However, these assays do not provide information about the physiological impacts caused by exposure, leaving a lack of information about the pathways and molecules involved in toxicity processes. Thus, the development of robust toxicological analyzes using cutting-edge technologies in order to prove drug use and also elucidate aspects of toxicity is very important, as they help in understanding the biological impact of human exposure to xenobiotics. Herein, bioanalytical methods using umbilical cord tissue to assess in utero exposure to cannabinoids were developed. A QuECheRS method was developed fully validated as a sample preparation technique for simultaneous extraction and alkaline hydrolysis of cannabinoids, using gas chromatography coupled to mass spectrometry to detect the analytes delta-9-tetrahydrocannabinol (THC), cannabinol (CBN), 11-hydroxydelta-9-tetrahydrocannabinol (11-OH-THC) and 11-nor-9-carboxy-tetrahydrocannabinol (THC-COOH). LC-MS/MS based proteomics and Trapped Ion Mobility Mass Spectrometry were also developed in order to identify protein biomarkers related to fetotoxicity of drug use in pregnancy. Our works represents the first use of QuECheRS for evaluation of illicit drugs in maternal-fetal tissue and was suitable for detection of Cannabis products. In the proteomic assays, potential biomarkers of fetotoxicity were identified in the exposed tissues, such as ACTA 2, Collagen alpha-1 (XVIII), SMC1A, KNL1, KMT2A. These proteins are related to embryogenic malformations and health complications in intrauterine life. The methodologies developed in this project may be useful for a better assessment of the toxicity of drug use in pregnancy, providing new clues about exposure and/or significant toxic effects that should be considered in the risk assessment

Warning about drinking during pregnancy: lessons from the French experience.

BACKGROUND: In France, since 2007, there is a compulsory warning recommending abstinence during pregnancy on every container of alcohol. Awareness of this warning, which consists of a small pictogram, is unknown. The aim of this study was to assess awareness of the warning and risk perceptions about prenatal drinking in pregnant and postpartum women. METHODS: A cross-sectional survey was carried out by telephone five years after the introduction of the warning label. A total of 3603 pregnant or postpartum French women participated. A quota sampling method was used to ensure the sample reflected the population. Multivariate analyses examined the characteristics associated with knowledge of risks and with awareness of the warning label. RESULTS: The warning label had been noticed by 66.1% of women and 77.3% of drinkers. Of those who had noticed the warning, 98.6% thought that it suggested abstinence. Overall, 40.8% of the women thought that spirits were more harmful than wine or beer, and 8.9% thought that drinking beer was recommended for lactation. CONCLUSION: Awareness of the warning is high but knowledge about the risks associated with wine and beer is poor. PRACTICE IMPLICATIONS: Future information campaigns should educate women about standard drinks and their pure alcohol equivalent. They should emphasize the risks associated with drinking during breastfeeding.

Exposure to environmentally persistent free radicals during gestation lowers energy expenditure and impairs skeletal muscle mitochondrial function in adult mice.

We have investigated the effects of in utero exposure to environmentally persistent free radicals (EPFRs) on growth, metabolism, energy utilization, and skeletal muscle mitochondria in a mouse model of diet-induced obesity. Pregnant mice were treated with laboratory-generated, combustion-derived particular matter (MCP230). The adult offspring were placed on a high-fat diet for 12 wk, after which we observed a 9.8% increase in their body weight. The increase in body size observed in the MCP230-exposed mice was not associated with increases in food intake but was associated with a reduction in physical activity and lower energy expenditure. The reduced energy expenditure in mice indirectly exposed to MCP230 was associated with reductions in skeletal muscle mitochondrial DNA copy number, lower mRNA levels of electron transport genes, and reduced citrate synthase activity. Upregulation of key genes involved in ameliorating oxidative stress was also observed in the muscle of MCP230-exposed mice. These findings suggest that gestational exposure to MCP230 leads to a reduction in energy expenditure at least in part through alterations to mitochondrial metabolism in the skeletal muscle.

Patterns of opioid utilization in pregnancy in a large cohort of commercial insurance beneficiaries in the United States.

BACKGROUND: There are few data regarding the utilization of opioids during pregnancy. The objective of this study was to define the prevalence and patterns of opioid use in a large cohort of pregnant women who were commercial insurance beneficiaries. METHODS: Data for the study were derived from a deidentified research database of women from across the United States who had both medical and prescription benefits. By using diagnostic codes, the authors defined a cohort of 534,500 women with completed pregnancies who were enrolled in a commercial insurance plan from 6 months before pregnancy through delivery. RESULTS: Overall, 76,742 women (14.4%) were dispensed an opioid at some point during pregnancy. There were 30,566 women (5.7%) dispensed an opioid during the first trimester, 30,434 women (5.7%) during the second trimester, and 34,906 women (6.5%) during the third trimester. Of these, 11,747 women (2.2%) were dispensed opioids three or more times during pregnancy. The most commonly dispensed opioids during pregnancy were hydrocodone (6.8%), codeine (6.1%), and oxycodone (2.0%). The prevalence of exposure at anytime during pregnancy decreased slightly during the study period from 14.9% for pregnancies that delivered in 2005 to 12.9% in 2011. The prevalence of exposure varied significantly by region and was lowest in the Northeast and highest in the South. CONCLUSIONS: This study demonstrates that opioids are very common exposures during pregnancy. Given the small and inconsistent body of literature on their safety in pregnancy, these findings suggest a need for research in this area.

[Safety assessment of nanomaterials in reproductive developmental field].

A diverse array of nanomaterials (NMs) such as amorphous nanosilica and carbon nanotubes have become widespread in use due to the development of nanotechnology. NMs are already being applied in universal fields because they have unique physicochemical properties. On the other hand, the increasing use of NMs has raised public concern about their potential risks to human health. In particular, recent reports indicated that carbon nanotubes induced mesothelioma-like lesions in mice, in a way similar to those induced by crocidolite asbestos. However, current knowledge of the potential risk of nanomaterials is considered insufficient. Because NMs have the potential to improve the quality of human life, it is essential to ensure the safety of NMs and provide information for designing NMs with safety. Especially, few studies have examined the effect of NMs on maintenance of pregnancy. Similar to the cases of thalidomide, a lot of evidence shows that fetuses are affected more than adults by a variety of environmental toxins because of physiological immaturity. Therefore it is essential to examine the effect of NMs on fetuses and pregnancies. Here we introduce the potential risk of amorphous nanosilica, most widely used NMs in food and the cosmetics field, to induce fetotoxicity and useful information for developing NMs with safety.

The use during pregnancy of prescription, over-the-counter, and alternative medications among Hispanic women.

BACKGROUND: Despite lack of scientific evidence about the safety of complementary and alternative medicines, the reported use of such remedies during pregnancies has increased. This study was undertaken to investigate the use of herbs, vitamins, and over-the-counter and prescription medications among pregnant Hispanic women and reasons for use, and to assess physician-patient level of communication about women's use. METHODS: A total of 485 Hispanic women were surveyed by means of a self-administered questionnaire immediately postpartum in a public hospital in Houston, Texas. The primary outcome was use of alternative therapies during the prenatal period. RESULTS: During their pregnancies, 19 percent of the participants took herbs and 47 percent took vitamin supplements, other than prenatal vitamins. The most common reason for using herbs and vitamins was to improve the woman's general health and energy level (59%); a few women (12%) had used them for specific pregnancy-related problems. Overall, 77 percent took prenatal vitamins and 21 percent supplemented with folic acid. The rates of use of over-the-counter and prescription medications were 23 and 29 percent, respectively. The use of prescription medication was two-and-a-half times higher among women with history of medical problems (adjusted OR = 2.59, 95% CI = 1.59-4.25, p = 0.0001). No other factor studied was independently associated with supplement or medication use. One in five women (20%) believed that herbs and vitamins were safer to use than prescription medication or were better at treating medical problems than prescription medicine, and one-third had not disclosed information about supplement use to their physicians. CONCLUSIONS: Use of herbal remedies does not appear to be a replacement for conventional medicine among most pregnant Hispanic women. Patient education about the risks of alternative therapies may lead to a reduction in intake of alternative medicines and greater disclosure to medical practitioners among this ethnic group.

Safety assessment of coenzyme Q10 (CoQ10).

Coenzyme Q10 (CoQ10) is a naturally occurring component present in living cells. Its physiological function is to act as an essential cofactor for ATP production, and to perform important antioxidant activities in the body. In most countries, CoQ10 has been widely used as a dietary supplement for more than 20 years. Recently, the use of CoQ10 as a dietary supplement has grown with a corresponding increase in daily dosage. The present review describes the safety profile of CoQ10 on the basis of animal and human data. The published reports concerning safety studies indicate that CoQ10 has low toxicity and does not induce serious adverse effects in humans. The acceptable daily intake (ADI) is 12mg/kg/day, calculated from the no-observed-adverse-effect level (NOAEL) of 1200 mg/kg/day derived from a 52-week chronic toxicity study in rats, i.e., 720 mg/day for a person weighing 60 kg. Risk assessment for CoQ10 based on various clinical trial data indicates that the observed safety level (OSL) for CoQ10 is 1200 mg/day/person. Evidence from pharmacokinetic studies suggest that exogenous CoQ10 does not influence the biosynthesis of endogenous CoQ9/CoQ10 nor does it accumulate into plasma or tissues after cessation of supplementation. Overall, these data from preclinical and clinical studies indicate that CoQ10 is highly safe for use as a dietary supplement. Additionally, analysis of CoQ10 bioavailability or its pharmacokinetics provides the pertinent safety evaluation for CoQ10.

Prescription of hazardous drugs during pregnancy.

BACKGROUND: Prescribing drugs to pregnant women requires the balancing of benefits and risks. Only a small proportion of drugs are known to be harmful to the fetus, but for the vast majority of drugs little evidence of fetal safety exists. AIM: To determine the prescription pattern of potentially and clearly harmful prescription drugs during pregnancy with reference to drug safety categorisation, and to define the drug groups primarily responsible for multiple drug use during pregnancy. STUDY DESIGN: A retrospective, register-based cohort study. METHODS: Linkage of three nationwide registers in Finland. Data collection included prescription drugs purchased during the preconception period and each trimester in the pregnant cohort, and the corresponding time periods in the non-pregnant controls. The pregnancy safety categorisation was determined for each drug (Anatomic Therapeutic Chemical [ATC] code) by using the Swedish classification of approved medicinal products (Farmaceutiska Specialiteter i Sverige [FASS]) and if not available, the corresponding Australian (Australian Drug Evaluation Committee [ADEC]) or US categorisation (FDA). GROUPS STUDIED: Women applying for maternity support (maternal grants) during the year 1999 (n = 43 470) plus non-pregnant control women matched by age and hospital district (n = 43 470). RESULTS: In the pregnant cohort, 20.4% of women purchased at least one drug classified as potentially harmful during pregnancy, and 3.4% purchased at least one drug classified as clearly harmful. A significant decline occurred in the number of pregnant women purchasing potentially and clearly harmful drugs during the first trimester when compared with the preconception period, and the decline continued from the first to the second trimester. In the pregnant cohort, 107 (0.2%) women purchased at least ten different drugs during pregnancy. The drugs most commonly purchased in this group were topical corticosteroids and nasal preparations. CONCLUSION: The use of hazardous prescription drugs declines during pregnancy but prescriptions of known teratogens and the relatively frequent practice of polypharmacy in epilepsy place emphasis on the need for careful pre-pregnancy counselling. However, drug safety classifications give a very crude estimation of risk and should only be used as general guidelines when planning treatment. Risk assessment must always be made on an individual basis, and pregnant women with illnesses requiring treatment must be treated adequately.

Reproductive toxicity assessment of lasofoxifene, a selective estrogen receptor modulator (SERM), in female rats.

BACKGROUND: Lasofoxifene is a nonsteroidal selective estrogen receptor modulator (SERM). With high affinity to the alpha and beta human estrogen receptors and greater potency than other SERMs, lasofoxifene is potentially a superior treatment for postmenopausal osteoporosis. In light of the known effects of estrogen-modulating compounds on female reproductive indices, two studies were conducted to evaluate the effects of lasofoxifene on female rat cyclicity, reproduction, and parturition. METHODS: One study evaluated effects of lasofoxifene on estrous cyclicity, and the second study assessed effects on implantation and parturition. In the cyclicity study, lasofoxifene was administered to female rats at doses of 0.1, 0.3, and 1.0 mg/kg/day for 14 consecutive days. After treatment, there was a 3-week reversibility phase followed by a mating phase. In the implantation study, lasofoxifene was administered to pregnant female rats at doses of 0.01, 0.03, and 0.1 mg/kg/day for 7 consecutive days (gestation day [GD] 0-6). Some animals were euthanized on GD 21, and the remainder of the group was allowed to deliver the F1 generation. Several developmental indices were evaluated in the F1 pups through post-natal day (PND) 21. RESULTS: In the cyclicity study, all lasofoxifene-treated females were anestrous by Study Day 7 (1.0 mg/kg) or 9 (0.3 and 0.1 mg/kg). The reversibility phase resulted in restoration of normal estrous cycles by the end of 1 (0.1 mg/kg) or 2 weeks (0.3 and 1.0 mg/kg). During the mating phase, no adverse effects occurred in pregnancy success or reproductive parameters. In the implantation study, all doses of lasofoxifene increased pre- and post-implantation losses, increased gestation length, and reduced litter size. None of the developmental parameters measured on the F1 generation was adversely affected. CONCLUSION: Lasofoxifene reversibly altered the estrous cycle and inhibited implantation, consistent with what would be expected from a member of the SERM class.

[Drug information and prescription guidance: role of regional centers of pharmacovigilance]. / Information du prescripteur et aide à la prescription: rôle des centres régionaux de pharmacovigilance.

MISSIONS: Drug information is one of the missions of the French Regional Pharmaco-Vigilance Centers together with the evaluation of adverse drug reaction (ADR) reports, expertise, teaching and research. SERVICES: Physicians and other health professionals call their regional center (the address and phone numbers are on the first pages of the Vidal, the French national drug compendium) for any kind of information pertaining to adverse drug reactions or proper use of drugs such as drug use in pregnancy or lactation. RELIABILITY: This information activity is successful because of the competence and impartiality of the regional centers which are based in university hospitals under an agreement signed with the French Medicines Agency.

Dermatologic drugs, pregnancy, and lactation. A conservative guide.

No database for determination of precise risk of drug use during pregnancy and lactation is available. There are, however, educated opinions concerning the advisability of use of a drug during the childbearing years from manufacturers, the Food and Drug Administration, various teratologists, the American Academy of Pediatrics, and the World Health Organization. Not all medications are absolutely contraindicated during pregnancy and lactation. Some drugs have been extensively used without apparent adverse effects in the mother or infant. When it is necessary to select a medication for use during pregnancy or lactation, the medication should have minimal risk. This article summarizes dermatologic drugs whose known risk is low.

A risk assessment of topical tretinoin as a potential human developmental toxin based on animal and comparative human data.

BACKGROUND: Although topically applied all-trans-retinoic acid (tretinoin) undergoes minimal absorption and adds negligibly to normal endogenous levels, its safety in humans is occasionally questioned because oral ingestion of retinoids at therapeutic levels is known to entail teratogenic risks. OBJECTIVE: To assess the actual potential for developmental toxicity from treatment with topical tretinoin. METHODS: Risk assessments were conducted on four known human developmental toxicants (valproic acid, methotrexate, thalidomide, and isotretinoin) and a potential developmental toxicant (acetylsalicylic acid). The margin of safety for each chemical was calculated from the ratio of animal no-observed adverse effect levels to human lowest-observed adverse effect levels or estimated exposure doses. RESULTS: The derived safety margin of more than 100 for topical tretinoin (with 2% absorption) contrasted sharply with the near unity values for valproic acid, methotrexate, thalidomide, and isotretinoin and was larger than that for acetylsalicylic acid. CONCLUSION: These data support other epidemiologic and animal data that topical tretinoin is not a potential human developmental toxicant.

The exclusion of pregnant, pregnable, and once-pregnable people (a.k.a. women) from biomedical research.

The barriers to women's participation as subjects in biomedical research are currently being challenged as a matter of legislative policy, medicine, and law. This Article catalogs the ways in which women have been disadvantaged by their exclusion and recent developments to redress them, and goes on to dissect the underlying rationales for excluding women from clinical trials. The author reveals the 'fundamental misconception' behind exclusionary rationales, and argues that research sponsors in fact have more to fear in the way of potential liability from the exclusion of women, even pregnant women and women of child-bearing capacity, than from their inclusion. Finally, the Article suggests strategies for achieving reform of these exclusionary practices.