What the lab can and cannot do: clinical interpretation of drug testing results.

Urine drug testing is one of the objective tools available to assess adherence. To monitor adherence, quantitative urinary results can assist in differentiating "new" drug use from "previous" (historical) drug use. "Spikes" in urinary concentration can assist in identifying patterns of drug use. Coupled chromatographic-mass spectrometric methods are capable of identifying very small amounts of analyte and can make clinical interpretation rather challenging, specifically for drugs that have a longer half-life. Polypharmacy is common in treatment and rehabilitation programs because of co-morbidities. Medications prescribed for comorbidities can cause drug-drug interaction and phenoconversion of genotypic extensive metabolizers into phenotypic poor metabolizers of the treatment drug. This can have significant impact on both pharmacokinetic (PK) and pharmacodynamic properties of the treatment drug. Therapeutic drug monitoring (TDM) coupled with PKs can assist in interpreting the effects of phenoconversion. TDM-PKs reflects the cumulative effects of pathophysiological changes in the patient as well as drug-drug interactions and should be considered for treatment medications/drugs used to manage pain and treat substance abuse. Since only a few enzyme immunoassays for TDM are available, this is a unique opportunity for clinical laboratory scientists to develop TDM-PK protocols that can have a significant impact on patient care and personalized medicine. Interpretation of drug screening results should be done with caution while considering pharmacological properties and the presence or absence of the parent drug and its metabolites. The objective of this manuscript is to review and address the variables that influence interpretation of different drugs analyzed from a rehabilitation and treatment programs perspective.

FASD: folic acid and formic acid - an unholy alliance in the alcohol abusing mother.

Alcohol consumption during pregnancy remains a significant cause of preventable birth defects and developmental disabilities; however, the mechanism of toxicity remains unclear. Methanol is present as a congener in many alcoholic beverages and is formed endogenously. Because ethanol is preferentially metabolized over methanol, it has been found in the sera and cerebro-spinal fluid of alcoholics. Toxicity resulting from methanol has been attributed to formic acid. Formic acid is present in significantly higher quantities in the biofluids of alcoholics. These higher levels can be cytotoxic and cause neuronal cell death. However, the adverse effects can be mitigated by adequate levels of hepatic folic acid, because formic acid elimination depends on folic acid. During pregnancy, folate concentrations are at least 2-fold higher in cord blood then in maternal blood, owing to increased folate requirements. The reverse has been demonstrated in pregnancies with alcohol abuse, suggesting downregulation of folate transporters and low fetal folate levels. Moreover, formic acid can cross the placenta and its adverse effects can be mitigated by folic acid. Thus, the combination of low fetal folate levels and presence of formic acid form a potent cytotoxic combination that may play a significant role in the etiology of fetal alcohol spectrum disorder.

False-positive meconium test results for fatty acid ethyl esters secondary to delayed sample collection.

BACKGROUND: Meconium analysis for fatty acid ethyl esters (FAEEs) is a validated method for identifying heavy prenatal ethanol (EtOH) exposure. This study investigated whether delayed sample collection can result in false-positive test results for FAEEs because of collection of samples potentially contaminated with postnatally produced stool. METHODS: Serial excretions were prospectively collected from neonates born to nondrinking mothers to capture the transition from meconium to postnatal stool. These were analyzed for FAEEs using headspace-solid phase microextraction and gas chromatography-mass spectrometry. Experiments involving incubation of samples with glucose or EtOH were performed to explore a potential mechanism of FAEE elevation. RESULTS: A total of 136 samples were collected from 30 neonates during their first few days of life (median of 4 samples/baby over a mean period of 68.5 hours postpartum). Although the first-collected meconium sample tested negative for FAEEs in all babies, later samples tested above the 2 nmol/g positive cutoff in 19 of 30 babies. Median time to appearance of FAEE-positive samples was 59.2 hours postpartum. In vitro experiments demonstrated that FAEE levels can be further increased in late samples (likely containing postnatal stool) after incubation with glucose, and that FAEEs are readily formed in meconium in the presence of EtOH. CONCLUSIONS: Collection of samples excreted later in the postpartum period can lead to false-positive test results for FAEEs, which could be because of contamination with dietary components of postnatally produced stool and EtOH-producing microorganisms. Clinically, it is critical to collect the earliest possible excretion for determination of FAEEs to ensure that the FAEE content is representative of in utero EtOH exposure.

Methadone: a review of drug-drug and pathophysiological interactions.

Numerous established and potential drug interactions with methadone are clinically important in people treated with methadone either for addiction or for chronic pain. Methadone users often have comorbidities and are prescribed drugs that may interact with methadone. Methadone is extensively metabolized by cytochrome P450 (CYP) 3A4 and to a lesser extent by CYP 1A2, 2D6, 2D8, 2C9/2C8, 2C19, and 2B6. Eighty-six percent of methadone is protein bound, predominately to α1-acid glycoprotein (AGP). Polymorphisms in or interactions with CYPs that metabolize methadone, changes in protein binding, and other pathophysiological conditions affect the pharmacokinetic properties of methadone. It is critical for health care providers who treat patients on methadone to have adequate information on the interactions of methadone with other drugs of abuse and other medications. We set out to describe drug-drug interactions as well as physiological and pathophysiological factors that may impact the pharmacokinetics of methadone. Using MEDLINE, we conducted a systematic search for papers and related abstracts published between 1966 and June 2010. Keywords that included methadone, drug-drug interactions, CYP P450 and AGP identified a total of 7709 papers. Other databases, including the Cochrane Database of Systematic Reviews and Scopus, were also searched; an additional 929 papers were found. Final selection of 286 publications was based on the relevance of each paper to the topic. Over 50 such interactions were found. Interactions of methadone with other drugs can lead to increased or decreased methadone drug levels in patients and result in potential overdose or withdrawal, respectively. The former can contribute to methadone's fatality. Prescribers of methadone and pharmacists should enquire about any new medications (including natural products and over-the-counter medications) periodically, and especially when an otherwise stable patient suddenly experiences drug craving, withdrawal or intoxication.

Prevalence of Fetal Alcohol Exposure by Analysis of Meconium Fatty Acid Ethyl Esters; A National Canadian Study.

Our study aimed to estimate the prevalence of heavy fetal alcohol exposure through the analysis of meconium FAEEs as an objective biomarker of fetal exposure. We conducted a study on meconium samples collected nationwide through the Maternal-Infant Research on Environmental Chemicals (MIREC) Study Group. FAEE in meconium was quantified by an established headspace solid-phase microextraction coupled with gas chromatography-mass spectrometry (SPME GC-MS). Out of 1315 samples collected in 10 Canadian obstetric units coast to coast between 2008-2011, the estimated prevalence of positive meconium FAEE ranged between 1.16% and 2.40%, translating into at least 1800 new cases of FASD in Canada each year. Positive maternal self- reports of heavy alcohol use were tenfold lower (0.24%). Use of meconium FAEE revealed tenfold more cases of heavy exposure to maternal drinking than did maternal reports. The use of objective measures of maternal alcohol exposure is critical in accurately estimating risks and in monitoring effective prevention of FASD.

Folate fortification and supplementation--are we there yet?

BACKGROUND: Folic acid fortification of flour has significantly decreased the incidence of neural tube defects (NTDs). We aimed at examining whether Ontario women of child-bearing age exhibit protective levels of RBC folate. METHODS: We reviewed laboratory databases on RBC folic acid from pre and post fortification years. The data included age, gender, RBC folate, hemoglobin, mean cell volume and pregnancy test. We examined a sub-set of females at ages 14-45 years who were non-anemic and normocytic. Complete protection against NTD was defined as RBC folate concentration above 900 nmol/L. RESULTS: In 2006, 40% of the women of child-bearing age and 36% of pregnant women, exhibited RBC folate levels below 900 nmol/L, rendering them sub-optimally protected against NTD. CONCLUSION: A considerable proportion of pregnant women is still at risk of having a baby with NTD. This should be remedied by increasing the mandatory concentrations of folic acid required in flour, complemented by public education and increasing the folic acid in prenatal supplements.

Association of In Utero Exposure to Polybrominated Diphenyl Ethers With the Risk of Hypospadias.

Importance: Polybrominated diphenyl ethers (PBDEs) are added to many consumer products as flame retardants, and their endocrine-disrupting properties are a growing health concern in pregnancy. Objective: To investigate whether in utero PBDE exposure as measured in maternal hair is associated with increased risk for hypospadias. Design, Setting, and Participants: In this case-control study, the setting was the urology clinic of a tertiary pediatric hospital between January 3, 2011, and April 1, 2013. Participants were children diagnosed as having hypospadias and their mothers and a control group of children without hypospadias and their mothers. Dates of data analysis were September 3, 2017, to December 28, 2017. Exposures: Gestational exposure to 8 PBDEs as measured in the 3-cm segment closest to the skull of maternal hair by gas chromatography-mass spectroscopy as a proxy for in utero exposure. The mothers resided in the same household for the duration of their pregnancy. Main Outcomes and Measures: Difference in total maternal hair PBDE levels between the hypospadias and control groups. Results: Total PBDE levels were significantly higher among mothers of infants with hypospadias (n = 152) (total PBDE level, 51.4 pg/mg; interquartile range, 35.8-78.5 pg/mg) than among controls (n = 64) (total PBDE level, 35.8 pg/mg; interquartile range, 18.1-69.9 pg/mg) (P = .02). Of the 152 women with sufficient hair samples for analysis in the case group, 89 completed a questionnaire and were included in a multivariable analysis, and of the 64 women with sufficient hair samples for analysis in the control group, 54 completed a questionnaire and were included in a multivariable analysis. Adjusting for potential confounders, hypospadias was associated with a relative 48.2% (95% CI, 23.2%-65.4%) higher maternal level of total PBDE levels in the multivariable analysis. Conclusions and Relevance: In this analysis, mothers of children with hypospadias were exposed during pregnancy to significantly higher levels of PBDEs. The results of this study suggest that level of exposure to PBDEs during gestation may have a role in the etiology of hypospadias.

Formic acid, a novel metabolite of chronic ethanol abuse, causes neurotoxicity, which is prevented by folic acid.

BACKGROUND: Methanol is endogenously formed in the brain and is present as a congener in most alcoholic beverages. Because ethanol is preferentially metabolized over methanol (MeOH) by alcohol dehydrogenase, it is not surprising that MeOH accumulates in the alcohol-abusing population. This suggests that the alcohol-drinking population will have higher levels of MeOH's neurotoxic metabolite, formic acid (FA). FA elimination is mediated by folic acid. Neurotoxicity is a common result of chronic alcoholism. This study shows for the first time that FA, found in chronic alcoholics, is neurotoxic and this toxicity can be mitigated by folic acid administration. OBJECTIVE: To determine if FA levels are higher in the alcohol-drinking population and to assess its neurotoxicity in organotypic hippocampal rat brain slice cultures. METHODS: Serum and CSF FA was measured in samples from both ethanol abusing and control patients, who presented to a hospital emergency department. FA's neurotoxicity and its reversibility by folic acid were assessed using organotypic rat brain hippocampal slice cultures using clinically relevant concentrations. RESULTS: Serum FA levels in the alcoholics (mean +/- SE: 0.416 +/- 0.093 mmol/l, n = 23) were significantly higher than in controls (mean +/- SE: 0.154 +/- 0.009 mmol/l, n = 82) (p < 0.0002). FA was not detected in the controls' CSF (n = 20), whereas it was >0.15 mmol/l in CSF of 3 of the 4 alcoholic cases. Low doses of FA from 1 to 5 mmol/l added for 24, 48 or 72 hours to the rat brain slice cultures caused neuronal death as measured by propidium iodide staining. When folic acid (1 micromol/l) was added with the FA, neuronal death was prevented. CONCLUSIONS: Formic acid may be a significant factor in the neurotoxicity of ethanol abuse. This neurotoxicity can be mitigated by folic acid administration at a clinically relevant dose.

Toxicology: then and now.

Toxicology is "the science of poisons"; more specifically the chemical and physical properties of poisons, their physiological or behavioral effects on living organisms, qualitative, and quantitative methods for their analysis and the development of procedures for the treatment of poisoning. Although the history of poisons dates to the earliest times, the study and the science of toxicology can be traced to Paracelsus (1493-1541) and Orfila (1757-1853). Modern toxicology is characterized by sophisticated scientific investigation and evaluation of toxic exposures. The 20th century is marked by an advanced level of understanding of toxicology. DNA and various biochemicals that maintain cellular functions were discovered. Our level of knowledge of toxic effects on organs and cells is now being revealed at the molecular level. This paper will review the historical progress of clinical and forensic toxicology by exploring analytical techniques in drug analysis, differing biological matrices, clinical toxicology, therapeutic drug management, workplace drug testing, and pharmacodynamic monitoring and pharmacogenetics.

Folate status of women in Toronto: Implications of folate fortification and supplementation.

OBJECTIVES: To assess the percentage of women of childbearing age with suboptimal levels of folate for protecting against neural tube defects (<906 nM), and assess folate status among the elderly. METHODS: A total of 1,035 anonymous blood samples from a centralized clinical laboratory, with a catchment area across the Greater Toronto Area, were assessed for red blood cell (RBC) folate concentrations using a chemiluminescent immunoassay. Folate analysis was requested by physicians as part of clinical care. Available data included age, sex, and RBC folate concentration. Descriptive statistics were used to characterize the percent of women who had suboptimal blood folate concentrations, and a multiple regression was used to analyze determinants of folate status. RESULTS: Our data from 2013 show that 7% of women of childbearing age (15-45 years) had RBC folate concentrations below 906 nM, a substantially lower percentage than in our 2006 study (40%). Results from the multiple regression showed that age is a significant positive predictor of higher RBC folate status (p < 0.001). CONCLUSION: Compared to our earlier data, we report a significant decrease in the suboptimal folate status among women of childbearing age. We also show that age is a predictor of higher RBC folate levels. Our data are limited due to a lack of information regarding patient or physician characteristics, and to the nature of our sample, yet our results are consistent with the continued increase in folate status observed among several population-level studies in the US and Canada post-fortification. Further research is needed to determine the reasons for and future implications of this continued increase in the elderly.

Pharmacogenetics of chronic pain management.

OBJECTIVE: The experience of chronic pain is one of the commonest reasons individuals seek medical attention, making the management of chronic pain a major issue in clinical practice. Drug metabolism and responses are affected by many factors, with genetic variations offering only a partial explanation of an individual's response. There is a paucity of evidence for the benefits of pharmacogenetic testing in the context of pain management. DESIGN AND METHODS: We reviewed the literature between 2000 and 2013, and references cited therein, using various keywords related to pain management, pharmacology and pharmacogenetics. RESULTS: Opioids continue to be the mainstay of chronic pain management. Several non-opioid based therapies, such as treatment with cannabinoids, gene therapy and epigenetic-based approaches are now available for these patients. Adjuvant therapies with antidepressants, benzodiazepines or anticonvulsants can also be useful in managing pain. Currently, laboratory monitoring of pain management patients, if performed, is largely through urine drug measurements. CONCLUSIONS: Drug half-life calculations can be used as functional markers of the cumulative effect of pharmacogenetics and drug-drug interactions. Assessment of half-life and therapeutic effects may be more useful than genetic testing in preventing adverse drug reactions to pain medications, while ensuring effective analgesia. Definitive, mass spectrometry-based methods, capable of measuring parent drug and metabolite levels, are the most useful assays for this purpose. Urine drug measurements do not necessarily correlate with serum drug concentrations or therapeutic effects. Therefore, they are limited in their use in monitoring efficacy and toxicity.

Folic acid transport to the human fetus is decreased in pregnancies with chronic alcohol exposure.

BACKGROUND: During pregnancy, the demand for folic acid increases since the fetus requires this nutrient for its rapid growth and cell proliferation. The placenta concentrates folic acid into the fetal circulation; as a result the fetal levels are 2 to 4 times higher than the maternal level. Animal and in vitro studies have suggested that alcohol may impair transport of folic acid across the placenta by decreasing expression of transport proteins. We aim to determine if folate transfer to the fetus is altered in human pregnancies with chronic alcohol consumption. METHODOLOGY/PRINCIPAL FINDINGS: Serum folate was measured in maternal blood and umbilical cord blood at the time of delivery in pregnancies with chronic and heavy alcohol exposure (n = 23) and in non-drinking controls (n = 24). In the alcohol-exposed pairs, the fetal:maternal serum folate ratio was ≤ 1.0 in over half (n = 14), whereas all but one of the controls were >1.0. Mean folate in cord samples was lower in the alcohol-exposed group than in the controls (33.15 ± 19.89 vs 45.91 ± 20.73, p = 0.04). CONCLUSIONS/SIGNIFICANCE: Our results demonstrate that chronic and heavy alcohol use in pregnancy impairs folate transport to the fetus. Altered folate concentrations within the placenta and in the fetus may in part contribute to the deficits observed in the fetal alcohol spectrum disorders.