Chirality Matters: Fine-Tuning of Novel Monoamine Reuptake Inhibitors Selectivity through Manipulation of Stereochemistry.

The high structural similarity, especially in transmembrane regions, of dopamine, norepinephrine, and serotonin transporters, as well as the lack of all crystal structures of human isoforms, make the specific targeting of individual transporters rather challenging. Ligand design itself is also rather limited, as many chemists, fully aware of the synthetic and analytical challenges, tend to modify lead compounds in a way that reduces the number of chiral centers and hence limits the potential chemical space of synthetic ligands. We have previously shown that increasing molecular complexity by introducing additional chiral centers ultimately leads to more selective and potent dopamine reuptake inhibitors. Herein, we significantly extend our structure-activity relationship of dopamine transporter-selective ligands and further demonstrate how stereoisomers of defined absolute configuration may fine-tune and direct the activity towards distinct targets. From the pool of active compounds, using the examples of stereoisomers 7h and 8h, we further showcase how in vitro activity significantly differs in in vivo drug efficacy experiments, calling for proper validation of individual stereoisomers in animal studies. Furthermore, by generating a large library of compounds with defined absolute configurations, we lay the groundwork for computational chemists to further optimize and rationally design specific monoamine transporter reuptake inhibitors.

Novel Selective and Low-Toxic Inhibitor of LmCPB2.8ΔCTE (CPB) One Important Cysteine Protease for Leishmania Virulence.

Leishmaniasis is a highly prevalent, yet neglected disease caused by protozoan parasites of the genus Leishmania. In the search for newer, safer, and more effective antileishmanial compounds, we herein present a study of the mode of action in addition to a detailed structural and biological characterization of LQOF-G6 [N-benzoyl-N'-benzyl-N″-(4-tertbutylphenyl)guanidine]. X-ray crystallography and extensive NMR experiments revealed that LQOF-G6 nearly exclusively adopts the Z conformation stabilized by an intramolecular hydrogen bond. The investigated guanidine showed selective inhibitory activity on Leishmania major cysteine protease LmCPB2.8ΔCTE (CPB) with ~73% inhibition and an IC50-CPB of 6.0 µM. This compound did not show any activity against the mammalian homologues cathepsin L and B. LQOF-G6 has been found to be nontoxic toward both organs and several cell lines, and no signs of hepatotoxicity or nephrotoxicity were observed from the analysis of biochemical clinical plasma markers in the treated mice. Docking simulations and experimental NMR measurements showed a clear contribution of the conformational parameters to the strength of the binding in the active site of the enzyme, and thus fit the differences in the inhibition values of LQOF-G6 compared to the other guanidines. Furthermore, the resulting data render LQOF-G6 suitable for further development as an antileishmanial drug.

Application of a pharmacokinetic model in characterizing sources of polychlorinated biphenyl exposure and determining threshold daily intakes for adverse health effects in infants and toddlers.

Characterization of PCB exposure sources for vulnerable population groups is essential to minimize the health effects of PCB exposure. At the same time, it is important to consolidate the knowledge on threshold intakes of PCBs for infants and toddlers to prevent health effects. We estimated total PCB concentrations from birth to 2 years of age in children from Slovak and Czech populations, which continue to have high PCB concentrations in breast milk. Using a pharmacokinetic (PK) model, we characterized dominant PCB exposure sources and estimated new threshold estimated daily intakes (TEDI) (above which adverse effects cannot be excluded) for postnatal PCB exposure in infants and toddlers. In the PK model, concentrations of seven indicator PCBs in breast milk and cord blood samples from 291 mother-child pairs from the Slovak birth cohort, and 396 breast milk samples from Czech mothers we used, together with their physiological characteristics and PCB concentrations from other exposure sources (food, dust, air). The estimated total PCB concentrations in children's blood at different ages were compared with threshold PCB concentrations of 500, 700 and 1000 ng·glipid-1 in serum proposed by the French Agency for Food, Environmental and Occupational Health & Safety (ANSES) and the German Environment Agency (UBA), above which possible adverse health effects may be expected. We estimated that up to 20.6% of Slovak children and up to 45.7% of Czech children at two years of age exceeded the threshold value of 700 ng·glipid-1 in blood. Mean TEDIs leading to values of 500 ng·glipid-1 in blood for children up to two years ranged between 110 and 220 ng·kg-1·bw·day-1, varying according to breastfeeding duration. Breast milk and prenatal exposure contributed to 71%-85% of PCBs exposure at two years of age. In contrast, the contributions of PCBs from dust and indoor air were negligible.

Polyphenol oxidase and enzymatic browning in apricot (Prunus armeniaca L.): Effect on phenolic composition and deduction of main substrates.

In this study, we investigate the effect of enzymatic browning on the phenolic composition of apricot in vivo and in vitro. The in vitro browning was caused by the recombinant latent apricot polyphenol oxidase (L-PaPPO). Successful heterologous expression of PaPPO in Escherichia coli yielded substantial amounts of enzyme containing both copper ions in the catalytic active site. The expressed L-PaPPO was characterized with regard to its molecular mass (56531.3 Da), pH optimum (7.0), activation by SDS, and enzyme kinetics. LC-MS/MS was used to compare the phenolic profiles of brown and non-brown apricots. The browning reactions did significantly decrease total phenolics and antioxidant capacity (measured with DPPH and CUPRAC assays). Catechin, epicatechin, and B-type procyanidins were the individual phenolics most affected by browning, followed by chlorogenic and neochlorogenic acid. These phenolics are most likely the main endogenous substrates of L-PaPPO, as they were oxidized much faster than the other identified phenolics.

Pre- and postnatal polychlorinated biphenyl exposure and cognitive and behavioral development at age 45 Months in a cohort of Slovak children.

Evidence of associations of pre- and postnatal exposure to polychlorinated biphenyls (PCBs) with cognitive development beyond early childhood is inconsistent. A previous report from this cohort observed adverse associations between early life PCB exposures and infant Bayley scores at age 16 months. The present study examines pre- and postnatal PCB exposures in relation to both behavior and cognitive development at age 45 months. Participants were 472 mother-child pairs residing in an area of eastern Slovakia characterized by environmental contamination with PCBs, which resulted in elevated blood serum concentrations. PCB-153 and PCB-118 concentrations were measured in maternal and in infant 6-, 16-, and 45-month serum samples. At age 45 months, children were administered five subtests of the Wechsler Preschool and Primary Scale of Intelligence (WPPSI-III), and mothers completed the Child Behavior Checklist (CBCL). Negative binomial and multiple linear regressions were used to estimate PCB-CBCL and PCB-WPPSI-III subtest score associations, respectively. Pre- and postnatal levels of PCB-153 and PCB-118 were not associated with cognitive performance on the WPPSI-III in this cohort. There was some suggestion that higher postnatal PCB concentrations were associated with more sleep problems and feelings of depression and anxiousness.

Synthesis and dopamine receptor binding of dihydrexidine and SKF 38393 catecholamine-based analogues.

Dopamine is an important neurotransmitter that regulates numerous essential functions, including cognition and voluntary movement. As such, it serves as an important scaffold for synthesis of novel analogues as part of drug development effort to obtain drugs for treatment of neurodegenerative diseases, such as Parkinson's disease. To that end, similarity search of the ZINC database based on two known dopamine-1 receptor (D1R) agonists, dihydrexidine (DHX) and SKF 38393, respectively, was used to predict novel chemical entities with potential binding to D1R. Three compounds that showed the highest similarity index were selected for synthesis and bioactivity profiling. All main synthesis products as well as the isolated intermediates, were properly characterized. The physico-chemical analyses were performed using HRESIMS, GC/MS, LC/MS with UV-Vis detection, and FTIR, 1H NMR and 13C NMR spectroscopy. Binding to D1 and D2 receptors and inhibition of dopamine reuptake via dopamine transporter were measured for the synthesized analogues of DHX and SKF 38393.

PBDE serum concentration and preschool maturity of children from Slovakia.

Polybrominated diphenylethers (PBDEs) are persistent organic pollutants (POPs), they are considered endocrine disruptors and can bioaccumulate in nature, and in living tissue. Human exposure to and the presence of PBDEs in human samples is of concern due to their potential health risks. Young children are one of the most vulnerable populations to PBDE's potential health effects. Ninety-one serum samples of 6-year-old children, residing in a contaminated location, due to former production of polychlorinated biphenyls (PCBs), were analysed to examine children's exposure to PBDEs in Slovakia. Median serum concentrations found for individual PBDE congeners BDE-28+33, -47, -99, -100, -153, -154 and -183 were 0.015, 0.184, 0.079, 0.046, 0.176, 0.014, and 0.097 ng g-1 lipid weight, respectively. Children's preschool maturity was measured using the Wechsler Preschool and Primary Scale of Intelligence (WPPSI-III) test. In multivariate analyses BDE-153 serum concentrations were significantly inversely associated with WPPSI-III composite score (p = 0.011, ß = -23.6), while adjusting for PCB-153 and sex. Significant negative associations were observed for BDE-153 serum concentrations (p = 0.002, ß = -29.8) and WPPSI-III composite score, after controlling for PCB-118 and sex. Negative associations were also observed for BDE-47, BDE-100 and BDE-153, with different individual WPPSI subtest scores, after adjustment with PCB-153 and/or PCB-118 and sex. Serum concentrations of PCB-153 and PCB-118 were not statistically significantly associated with WPPSI-III composite score and individual subtest scores. These findings demonstrate adverse effects of PBDE serum exposure on preschool maturity of children, and PBDEs potentially negative impact on child neuropsychological development.

A Birth Cohort Study of Maternal and Infant Serum PCB-153 and DDE Concentrations and Responses to Infant Tuberculosis Vaccination.

BACKGROUND: Reasons for the highly variable and often poor protection conferred by the Mycobacterium bovis bacille Calmette-Guérin (BCG) vaccine are multifaceted and poorly understood. OBJECTIVES: We aimed to determine whether early-life exposure to PCBs (polychlorinated biphenyls) and DDE [1,1-dichloro-2,2-bis(p-chlorophenyl)ethylene] reduces 6-month infant BCG vaccine response. METHODS: Data came from families participating in a prospective birth cohort in eastern Slovakia. At birth, maternal and cord blood were collected for chemical analyses, and infants were immunized with BCG. Blood was collected from infants for chemical analyses and to determine 6-month BCG-specific immunoglobulin (Ig) G and IgA levels. Multivariable linear regression models were fit to examine chemical-BCG associations among approximately 500 mother-infant pairs, with adjustment for confounders. RESULTS: The median 6-month infant concentration of the prevalent congener PCB-153 was 113 ng/g lipid [interquartile range (IQR): 37-248], and 388 ng/g lipid (IQR: 115-847) for DDE. Higher 6-month infant concentrations of PCB-153 and DDE were strongly associated with lower 6-month BCG-specific antibody levels. For instance, BCG-specific IgG levels were 37% lower for infants with PCB-153 concentrations at the 75th percentile compared to the 25th percentile (95% CI: -42, -32; p < 0.001). Results were similar in magnitude and precision for DDE. There was also evidence of PCB-DDE additivity, where exposure to both compounds reduced anti-BCG levels more than exposure to either compound alone. CONCLUSIONS: The associations observed in this study indicate that environmental exposures may be overlooked contributors to poorer responses to BCG vaccine. The overall association between these exposures and tuberculosis incidence is unknown. CITATION: Jusko TA, De Roos AJ, Lee SY, Thevenet-Morrison K, Schwartz SM, Verner MA, Palkovicova Murinova L, Drobná B, Kocan A, Fabisiková A, Conka K, Trnovec T, Hertz-Picciotto I, Lawrence BP. 2016. A birth cohort study of maternal and infant serum PCB-153 and DDE concentrations and responses to infant tuberculosis vaccination. Environ Health Perspect 124:813-821; http://dx.doi.org/10.1289/ehp.1510101.

Polychlorinated biphenyls and selected organochlorine pesticides in serum of Slovak population from industrial and non-industrial areas.

The concentrations of indicator polychlorinated biphenyls (PCBs No. 28, 52, 101, 138, 153 and 180) and organochlorine pesticides (HCB, p,p'-DDE and p,p'-DDT) in 121 blood serum specimens collected from non-occupationally exposed adults living in contaminated and comparison areas were determined using high-resolution gas chromatography/electron capture detection (HRGC/ECD). The sum of the serum concentrations of the three most abundant PCB congeners (No. 138, 153 and 180) found in participants (N = 81) living in industrial areas near incinerators, metallurgical and chemical plants (Krompachy, Kosice, Nemecka and Sala) was significantly higher (p < 0.001) than in participants living in villages and towns without known sources causing persistent organic pollutant (POP) contamination (N = 40). Similarly, significant differences were observed for p,p'-DDE (p < 0.0001) and p,p'-DDT (p < 0.002). However, a Mann-Whitney U test between groups showed that the difference for HCB was not statistically significant (p = 0.089). Age was positively correlated with the sum of PCBs (No. 138, 153 and 180), HCB and the sum of p,p'-DDE and p,p'-DDT (p < 0.01 for all).

PCDDs, PCDFs, PCBs and OCPs in sediments from selected areas in the Slovak Republic.

Within our research project, 34 river bottom sediments were collected in 2006-2007 at five areas across Slovakia with industrial sources of persistent organic pollutants (Kosice, Krompachy, Nemecká, Sala, Nováky) and one background area (Starina). Sediments were analyzed for seven 2,3,7,8-substituted polychlorinated dibenzo-p-dioxins (PCDDs) and 10 dibenzofurans (PCDFs), 12 dioxin-like and 6 indicator polychlorinated biphenyls (PCBs), hexachlorobenzene (HCB) and 1,1,1,-trichloro-2,2-bis(p-chlorophenyl)ethane (p,p'-DDT) with 1,1-dichloro-2,2-bis(p-chlorophenyl)ethylene (p,p'-DDE). Analytical procedure based on accelerated solvent extraction followed by a semi-automated clean-up and fractionation was used. Determination of target compounds was performed by HRGC-HRMS analysis. Total WHO toxic equivalent WHO1998-PCDD/F/dl-PCB-TEQ concentrations ranged from 0.26 to 559 pg TEQ g(-1) dry matter (dm), with a median 2.2 pg TEQ g(-1) dm. The sums of six indicator PCBs were in the range 0.56-1014 ng g(-1) dm, with a median 11.8 ng g(-1) dm. The concentrations of organochlorine pesticides HCB and p,p'-DDE/DDT varied from 0.15 to 34.8 ng g(-1) dm, with a median 0.91 ng g(-1) dm and 0.46-34.1 ng g(-1)dm, with a median 6.7 ng g(-1)dm, respectively. The most abundant congeners in all sediment samples among dioxins, furans and PCBs were OCDD, 1,2,3,4,6,7,8-HpCDF, PCB-118 and PCB-153.

Anthropometric, socioeconomic, and maternal health determinants of placental transfer of organochlorine compounds.

The aim of this study was to relate placental transfer, quantified by the cord to maternal serum concentration ratio (C/M), of five organochlorine pesticides (OCP) hexachlorobenzene (HCB), ß-hexachlorocyclohexane (ß-HCH), γ-hexachlorocyclohexane (γ-HCH) , p,p'-DDT, p,p'-DDE and 15 polychlorinated biphenyl (PCB) congeners (28, 52, 101, 105, 114, 118, 123(+149), 138(+163), 153, 156(+171), 157, 167, 170, 180, and 189) to anthropometric, socioeconomic, and maternal health characteristics. We included into the study 1,134 births during the period 2002-2004 from two districts in eastern Slovakia with high organochlorine concentrations relative to other areas of the world. Only concentrations >LOD were taken into account. Variables as age, weight and height of mothers, parity, ethnicity, alcohol consumption, illness during pregnancy, smoking during pregnancy, hypertension, respiratory diseases, rheumatoid arthritis and diabetes mellitus, and birth weight were related to C/M. Results of regression analyses showed that C/M was predicted by several factors studied. Positive associations were observed for gestational alcohol consumption, fewer illnesses during pregnancy, maternal age, and maternal weight. Caucasians had a greater C/M compared to Romani for wet weight data of congeners 170 and 180 and in contrast C/M for HCB was greater in Romani. Our results show that drinking mothers compared to abstaining expose their fetuses not only to alcohol but to an increased level of several PCB congeners. A straightforward explanation of associations between C/M shifts and factors studied is very difficult, however, with regard to the high lipophilicity of OCPs and PCBs, changes in their kinetics probably reflect lipid kinetics.

PCDDs, PCDFs, dioxin-like PCBs and indicator PCBs in soil from five selected areas in Slovakia.

There is a lack of information regarding persistent organic pollutants (POPs) in soil from Slovakia. This paper reports the concentrations of 17 polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs), 12 dioxin-like polychlorinated biphenyls (dl-PCBs) and 6 indicator PCBs associated with 32 soil samples collected in 2007 from areas in the vicinity up to several kilometres from four selected industrial PCDD/F sources in Slovakia and one background area. The results show that the total WHO(1998)-PCDD/F/dl-PCB-TEQ concentrations varied from 0.34 to 18.05 pg g(-1) dry weight (dw) and the average total TEQ concentrations in samples collected at the Sala, Kosice, Krompachy and Dubová areas were 3.18, 2.64, 7.80, and 3.19 pg g(-1)dw respectively. The average of the total WHO(1998)-TEQ values in three soil samples representing the background Starina area was 0.66 pg g(-1) dw. The predominant contaminants among 2,3,7,8-substituted PCDD/Fs, dl-PCB, and indicator PCBs was OCDD, PCB 118, and PCB 153 respectively. The major contributors to the TEQ were 2,3,4,7,8-PeCDF, PCB 126, 1,2,3,7,8-PeCDD, PCB 156 and 1,2,3,4,7,8-HxCDF+1,2,3,4,7,9-HxCDF in descending order.

PCDD/PCDF, dl-PCB and PBDE serum levels of Slovak general population.

Blood serum specimens from 81 non-occupationally exposed adults residing in four areas close to municipal and waste incinerators as well as metallurgical industry plant and 44 adult subjects coming from control area of Slovakia were analysed for polychlorinated dibenzo-p-dioxins (PCDDs), polychlorinated dibenzofurans (PCDFs), dioxin-like polychlorinated biphenyls (dl-PCBs) and polybrominated diphenylethers (PBDEs). The concentration of total WHO(98)TEQ PCDD/F/dl-PCBs in whole group of donors from areas where known sources causing dioxin contamination are present was significantly higher than in control group of donors (p<0.001). Correlation between the age of donors and PCDD/F and dl-PCB levels was confirmed (Spearman's r(PCDD/Fs)=0.543, r(dl-PCBs)=0.521, p<0.001). Furthermore, this study presents first results concerning the PBDE congeners in human serum of Slovak general population. The total concentration (congeners 28, 47, 99, 100, 153, 154, 183) in control group was approximately 1.5-times higher in comparison to that of residents coming from areas with supposed environmental pollution. The most abundant congeners in all samples were BDE-47 and BDE-153 with median values of 0.24 ng g(-1) lipid and 0.23ngg(-1) lipid, respectively. The positive association between PBDE values and age of donors was not found.

Maternal and early postnatal polychlorinated biphenyl exposure in relation to total serum immunoglobulin concentrations in 6-month-old infants.

Animal data indicate that developmental tetrachlorodibenzo-p-dioxin exposure alters immune function; however, the potential immunotoxicity of dioxin-like and non-dioxin-like polychlorinated biphenyls (PCBs) in the developing infant is an understudied area. The aim of the current study is to examine the association between maternal and early postnatal PCB concentrations in relation to total infant serum immunoglobulin concentrations determined at 6-months-of-age. We selected 384 mother-infant pairs participating in a birth cohort study in Eastern Slovakia. PCB concentrations of several congeners were determined in maternal and cord serum samples and in infant serum samples collected at 6-months-of-age using gas chromatography with electron capture detection. Total immunoglobulin (Ig) G, A, and M concentrations were determined by nephelometry, and IgE concentrations were determined by enzyme-linked immunoassay. Linear regression models with adjustment for potential confounding factors were used to estimate the associations between maternal, cord, and 6-month infant PCB concentrations and total serum immunoglobulins. The median maternal serum concentration of PCB-153 was 140?ng/g lipid, ?10-fold higher than concentrations in childbearing-age women in the United States during the same period. Maternal, cord, or 6-month infant PCB concentrations were not associated with total serum immunoglobulin levels at 6 months, regardless of the timing of PCB exposure, PCB congener, or specific immunoglobulin. In this population, which has high PCB concentrations relative to most populations in the world today, we did not observe any association between maternal and early postnatal PCB concentrations and total immunoglobulin measures of IgG, IgA, IgM, or IgE.

Half-lives of serum PCB congener concentrations in environmentally exposed early adolescents.

The aim was to determine half-life of six most abundant PCB congeners in the body of early adolescents. In 304 environmentally exposed children, PCB serum concentration was determined at the age of 8 and 12years. Half-life was determined for each child assuming exponential decrease or for the whole cohort using multiple regression. Results obtained by both approaches were in agreement. PCB reuptakes corrupting half-life estimates for each child and each congener were evaluated. If one of the serum PCB concentration values fell below the level of detection (LOD) the pair was excluded and if PCB half-life value exceeded the arbitrary value of 30years. The following median half-lives in years 4.46, 10.59, 9.7, 4.7, 9.1 and 9.8 were obtained for PCB congeners 118, 138(+163), 153, 156(+171), 170 and 180, respectively. The elimination half-life values were not systematically related to PCB serum concentration at any examination age. Between half-life values, percentage of children with significant reuptakes and PCB congener abundance in serum were found significant associations.

A cohort study of developmental polychlorinated biphenyl (PCB) exposure in relation to post-vaccination antibody response at 6-months of age.

BACKGROUND: Extensive experimental data in animals indicate that exposure to polychlorinated biphenyls (PCBs) during pregnancy leads to changes in offspring immune function during the postnatal period. Whether developmental PCB exposure influences immunologic development in humans has received little study. METHODS: The study population was 384 mother-infant pairs recruited from two districts of eastern Slovakia for whom prospectively collected maternal, cord, and 6-month infant blood specimens were available. Several PCB congeners were measured in maternal, cord, and 6-month infant sera by high-resolution gas chromatography with electron capture detection. Concentrations of IgG-specific anti-haemophilus influenzae type b, tetanus toxoid, and diphtheria toxoid were assayed in 6-month infant sera using ELISA methods. Multiple linear regression was used to estimate the relation between maternal, cord, and 6-month infant PCB concentrations and the antibody concentrations evaluated at 6-months of age. RESULTS: Overall, there was little evidence of an association between infant antibody concentrations and PCB measures during the pre- and early postnatal period. In addition, our results did not show specificity in terms of associations limited to a particular developmental period (e.g. pre- vs. postnatal), a particular antibody, or a particular PCB congener. CONCLUSIONS: At the PCB concentrations measured in this cohort, which are high relative to most human populations today, we did not detect an association between maternal or early postnatal PCB exposure and specific antibody responses at 6-months of age.