The emergence of the threat of radiological terrorism and other radiological incidents has led to the need for development of fast, accurate and noninvasive methods for detection of radiation exposure. The purpose of this study was to extend radiation metabolomic biomarker discovery to humans, as previous studies have focused on mice. Urine was collected from patients undergoing total body irradiation at Memorial Sloan-Kettering Cancer Center prior to hematopoietic stem cell transplantation at 4–6 h postirradiation (a single dose of 1.25 Gy) and 24 h (three fractions of 1.25 Gy each). Global metabolomic profiling was obtained through analysis with ultra performance liquid chromatography coupled to time-of-flight mass spectrometry (TOFMS). Prior to further analyses, each sample was normalized to its respective creatinine level. Statistical analysis was conducted by the nonparametric Kolmogorov-Smirnov test and the Fisher’s exact test and markers were validated against pure standards. Seven markers showed distinct differences between pre- and post-exposure samples. Of those, trimethyl-l-lysine and the carnitine conjugates acetylcarnitine, decanoylcarnitine and octanoylcarnitine play an important role in the transportation of fatty acids across mitochondria for subsequent fatty acid β-oxidation. The remaining metabolites, hypoxanthine, xanthine and uric acid are the final products of the purine catabolism pathway, and high levels of excretion have been associated with increased oxidative stress and radiation induced DNA damage. Further analysis revealed sex differences in the patterns of excretion of the markers, demonstrating that generation of a sex-specific metabolomic signature will be informative and can provide a quick and reliable assessment of individuals in a radiological scenario. This is the first radiation metabolomics study in human urine laying the foundation for the use of metabolomics in biodosimetry and providing confidence in biomarker identification based on the overlap between animal models and humans.

Extensive preclinical data implicate corticotropin-releasing hormone (CRH), acting through its CRH1 receptor, in stress-and dependence-induced alcohol seeking. We evaluated pexacerfont, an orally available, brain penetrant CRH1 antagonist for its ability to suppress stress-induced alcohol craving and brain responses in treatment seeking alcohol-dependent patients in early abstinence. Fiftyfour anxious alcohol-dependent participants were admitted to an inpatient unit at the NIH Clinical Center, completed withdrawal treatment, and were enrolled in a double-blind, randomized, placebo-controlled study with pexacerfont (300 mg/day for 7 days, followed by 100 mg/day for 23 days). After reaching steady state, participants were assessed for alcohol craving in response to stressful or alcoholrelated cues, neuroendocrine responses to these stimuli, and functional magnetic resonance imaging (fMRI) responses to alcohol-related stimuli or stimuli with positive or negative emotional valence. A separate group of 10 patients received open-label pexacerfont following the same dosing regimen and had cerebrospinal fluid sampled to estimate central nervous system exposure. Pexacerfont treatment had no effect on alcohol craving, emotional responses, or anxiety. There was no effect of pexacerfont on neural responses to alcohol-related or affective stimuli. These results were obtained despite drug levels in cerebrospinal fluid (CSF) that predict close to 90% central CRH1 receptor occupancy. CRH1 antagonists have been grouped based on their receptor dissociation kinetics, with pexacerfont falling in a category characterized by fast dissociation. Our results may indicate that antagonists with slow offset are required for therapeutic efficacy. Alternatively, the extensive preclinical data on CRH1 antagonism as a mechanism to suppress alcohol seeking may not translate to humans.

Synthetic cathinones are novel stimulants derived from cathinone, with amphetamines or cocaine-like effects, often labeled "not for human consumption" and considered "legal highs". Emergence of these new designer drugs complicate interpretation of forensic and clinical cases, with introduction of many new analogs designed to circumvent legislation and vary effects and potencies. We developed a method for the simultaneous quantification of 28 synthetic cathinones, including four metabolites, in urine by liquid chromatography coupled to high resolution mass spectrometry (LC-HRMS). These cathinones include cathinone, methcathinone, and synthetic cathinones position-3'-substituted, N-alkyl-substituted, ring-substituted, methylenedioxy-substituted, and pyrrolidinyl-substituted. One mL phosphate buffer pH 6 and 25 μL IStd solution were combined with 0.25 mL urine, and subjected to solid phase cation exchange extraction (SOLA SCX). The chromatographic reverse-phase separation was achieved with a gradient mobile phase of 0.1 % formic acid in water and in acetonitrile in 20 min. We employed a Q Exactive high resolution mass spectrometer, with compounds identified and quantified by target-MSMS experiments. The assay was linear from 0.5-1 to 100 μg/L, with limits of detection of 0.25-1 μg/L. Imprecision (n = 20) was <15.9 % and accuracy (n = 20) 85.2-118.1 %. Extraction efficiency was 78.9-116.7 % (CV 1.4-16.7 %, n = 5), process efficiency 57.7-104.9 %, and matrix effects from -29.5 % to 1.5 % (CV 1.9-13.1 %, n = 10). Most synthetic cathinones were stable at 4 °C for 72 h (n = 27) and after 3 freeze-thaw cycles (n = 26), but many (n = 19) were not stable at room temperature for 24 h (losses up to -67.6 %). The method was applied to authentic urine specimens from synthetic cathinone users. This method provides a comprehensive confirmation method for 28 synthetic cathinones in urine, with good selectivity and specificity.

3,4-Methylenedioxypyrovalerone (MDPV) is a psychoactive component of so-called bath salts products that has caused serious medical consequences in humans. In this chapter, we review the neuropharmacology of MDPV and related analogs, and supplement the discussion with new results from our preclinical experiments. MDPV acts as a potent uptake inhibitor at plasma membrane transporters for dopamine (DAT) and norepinephrine (NET) in nervous tissue. The MDPV formulation in bath salts is a racemic mixture, and the S isomer is much more potent than the R isomer at blocking DAT and producing abuse-related effects. Elevations in brain extracellular dopamine produced by MDPV are likely to underlie its locomotor stimulant and addictive properties. MDPV displays rapid pharmacokinetics when injected into rats (0.5–2.0 mg/kg), with peak plasma concentrations achieved by 10–20 min and declining quickly thereafter. MDPV is metabolized to 3,4-dihydroxypyrovalerone (3,4-catechol-PV) and 4-hydroxy-3-methoxypyrovalerone (4-OH-3-MeO-PV) in vivo, but motor activation produced by the drug is positively correlated with plasma concentrations of parent drug and not its metabolites. 3,4-Catechol-PV is a potent uptake blocker at DAT in vitro but has little activity after administration in vivo. 4-OH-3-MeO-PV is the main MDPV metabolite but is weak at DAT and NET. MDPV analogs, such as α-pyrrolidinovalerophenone (α-PVP), display similar ability to inhibit DAT and increase extracellular dopamine concentrations. Taken together, these findings demonstrate that MDPV and its analogs represent a unique class of transporter inhibitors with a high propensity for abuse and addiction.