Combining Experimental and Computational Methods to Produce Conjugates of Anticholinesterase and Antioxidant Pharmacophores with Linker Chemistries Affecting Biological Activities Related to Treatment of Alzheimer's Disease.

Effective therapeutics for Alzheimer's disease (AD) are in great demand worldwide. In our previous work, we responded to this need by synthesizing novel drug candidates consisting of 4-amino-2,3-polymethylenequinolines conjugated with butylated hydroxytoluene via fixed-length alkylimine or alkylamine linkers (spacers) and studying their bioactivities pertaining to AD treatment. Here, we report significant extensions of these studies, including the use of variable-length spacers and more detailed biological characterizations. Conjugates were potent inhibitors of acetylcholinesterase (AChE, the most active was 17d IC50 15.1 ± 0.2 nM) and butyrylcholinesterase (BChE, the most active was 18d: IC50 5.96 ± 0.58 nM), with weak inhibition of off-target carboxylesterase. Conjugates with alkylamine spacers were more effective cholinesterase inhibitors than alkylimine analogs. Optimal inhibition for AChE was exhibited by cyclohexaquinoline and for BChE by cycloheptaquinoline. Increasing spacer length elevated the potency against both cholinesterases. Structure-activity relationships agreed with docking results. Mixed-type reversible AChE inhibition, dual docking to catalytic and peripheral anionic sites, and propidium iodide displacement suggested the potential of hybrids to block AChE-induced ß-amyloid (Aß) aggregation. Hybrids also exhibited the inhibition of Aß self-aggregation in the thioflavin test; those with a hexaquinoline ring and C8 spacer were the most active. Conjugates demonstrated high antioxidant activity in ABTS and FRAP assays as well as the inhibition of luminol chemiluminescence and lipid peroxidation in mouse brain homogenates. Quantum-chemical calculations explained antioxidant results. Computed ADMET profiles indicated favorable blood-brain barrier permeability, suggesting the CNS activity potential. Thus, the conjugates could be considered promising multifunctional agents for the potential treatment of AD.

The effect of substituent position and solvent on thermal Z‒E isomerization of dihydroquinolylazotetrazole dyes: kinetic, thermodynamic, and spectral approaches.

Kinetic and thermodynamic parameters have been investigated for the thermal Z‒E isomerization of dihydroquinolylazotetrazole dyes with alkyl substituents (Me, t-Bu, and Adm) at positions 1 (dyes 2) and 2 (dyes 3) of the tetrazole moiety in two solvents of different polarity, acetonitrile (MeCN) and toluene. The experimental results show crucial dependence of these parameters on a substituent position in the tetrazole moiety and on a solvent. For dyes 2, Eact and ΔH‡ are lower in MeCN than in toluene that results in a high increase in the lifetimes of the Z isomers: from milliseconds in MeCN to minutes in toluene. For dyes 3, the difference in Eact and ΔH‡ in the two solvents is opposite: Eact and ΔH‡ are by more than 20 kJ mol-1 higher in MeCN, nevertheless, the rate constants for 3 in toluene are comparable with those in MeCN at the ambient temperature and the difference in the behavior is determined by the value of negative entropy of activation. Quantum-chemical calculations of the thermal Z‒E isomerization show the possibility of the process to occur via crossing from the S0 to the thermally induced T1 state. The contribution of this path is highest for 3 in toluene. The analysis of the absorption spectra demonstrates that for the E isomers, the n‒π* and π‒π* transitions are within the long-wavelength absorption band and their positions relative each other are opposite in the solvents: the n‒π* transition is blue-shifted relative to the π‒π* transition in MeCN and is red-shifted in toluene.

Derivatives of 9-phosphorylated acridine as butyrylcholinesterase inhibitors with antioxidant activity and the ability to inhibit ß-amyloid self-aggregation: potential therapeutic agents for Alzheimer's disease.

We investigated the inhibitory activities of novel 9-phosphoryl-9,10-dihydroacridines and 9-phosphorylacridines against acetylcholinesterase (AChE), butyrylcholinesterase (BChE), and carboxylesterase (CES). We also studied the abilities of the new compounds to interfere with the self-aggregation of ß-amyloid (Aß42) in the thioflavin test as well as their antioxidant activities in the ABTS and FRAP assays. We used molecular docking, molecular dynamics simulations, and quantum-chemical calculations to explain experimental results. All new compounds weakly inhibited AChE and off-target CES. Dihydroacridines with aryl substituents in the phosphoryl moiety inhibited BChE; the most active were the dibenzyloxy derivative 1d and its diphenethyl bioisostere 1e (IC50 = 2.90 ± 0.23 µM and 3.22 ± 0.25 µM, respectively). Only one acridine, 2d, an analog of dihydroacridine, 1d, was an effective BChE inhibitor (IC50 = 6.90 ± 0.55 µM), consistent with docking results. Dihydroacridines inhibited Aß42 self-aggregation; 1d and 1e were the most active (58.9% ± 4.7% and 46.9% ± 4.2%, respectively). All dihydroacridines 1 demonstrated high ABTS•+-scavenging and iron-reducing activities comparable to Trolox, but acridines 2 were almost inactive. Observed features were well explained by quantum-chemical calculations. ADMET parameters calculated for all compounds predicted favorable intestinal absorption, good blood-brain barrier permeability, and low cardiac toxicity. Overall, the best results were obtained for two dihydroacridine derivatives 1d and 1e with dibenzyloxy and diphenethyl substituents in the phosphoryl moiety. These compounds displayed high inhibition of BChE activity and Aß42 self-aggregation, high antioxidant activity, and favorable predicted ADMET profiles. Therefore, we consider 1d and 1e as lead compounds for further in-depth studies as potential anti-AD preparations.

Conjugates of Tacrine and Salicylic Acid Derivatives as New Promising Multitarget Agents for Alzheimer's Disease.

A series of previously synthesized conjugates of tacrine and salicylamide was extended by varying the structure of the salicylamide fragment and using salicylic aldehyde to synthesize salicylimine derivatives. The hybrids exhibited broad-spectrum biological activity. All new conjugates were potent inhibitors of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) with selectivity toward BChE. The structure of the salicylamide moiety exerted little effect on anticholinesterase activity, but AChE inhibition increased with spacer elongation. The most active conjugates were salicylimine derivatives: IC50 values of the lead compound 10c were 0.0826 µM (AChE) and 0.0156 µM (BChE), with weak inhibition of the off-target carboxylesterase. The hybrids were mixed-type reversible inhibitors of both cholinesterases and displayed dual binding to the catalytic and peripheral anionic sites of AChE in molecular docking, which, along with experimental results on propidium iodide displacement, suggested their potential to block AChE-induced ß-amyloid aggregation. All conjugates inhibited Aß42 self-aggregation in the thioflavin test, and inhibition increased with spacer elongation. Salicylimine 10c and salicylamide 5c with (CH2)8 spacers were the lead compounds for inhibiting Aß42 self-aggregation, which was corroborated by molecular docking to Aß42. ABTS•+-scavenging activity was highest for salicylamides 5a-c, intermediate for salicylimines 10a-c, low for F-containing salicylamides 7, and non-existent for methoxybenzoylamides 6 and difluoromethoxybenzoylamides 8. In the FRAP antioxidant (AO) assay, the test compounds displayed little or no activity. Quantum chemical analysis and molecular dynamics (MD) simulations with QM/MM potentials explained the AO structure-activity relationships. All conjugates were effective chelators of Cu2+, Fe2+, and Zn2+, with molar compound/metal (Cu2+) ratios of 2:1 (5b) and ~1:1 (10b). Conjugates exerted comparable or lower cytotoxicity than tacrine on mouse hepatocytes and had favorable predicted intestinal absorption and blood-brain barrier permeability. The overall results indicate that the synthesized conjugates are promising new multifunctional agents for the potential treatment of AD.

Caspase-3, Caspase-8 and XIAP Gene Expression in the Placenta: Exploring the Causes of Spontaneous Preterm Labour.

A better understanding of the pathogenesis of preterm birth (PTB) will allow us to lower the PTB rate, reducing perinatal morbidity and mortality. This article presents the hypothesis that premature placenta apoptosis could be a potential cause of PTB. We evaluated gene expression involved in apoptosis: caspase-3, caspase-8, and XIAP (X-linked inhibitor of apoptosis) in the placenta during pregnancy (n = 41), at the onset of preterm labour (n = 42), after preterm (n = 44) and term (n = 32) labour. We used RNA extraction, reverse transcription, and PCR. During pregnancy the gene expression of caspase-3 and caspase-8 is low, but XIAP is higher than the caspases. At the onset of preterm labour, we observed a significantly increased expression of both caspase-8 (10.7-fold, p < 0.01) and caspase-3 (2.5-fold, p < 0.01) and XIAP (3-fold; p < 0.05) compared with expression during pregnancy. Our study showed that during pregnancy, the expression of caspase genes in the placenta is low and probably controlled by high XIAP expression. At the onset of preterm labour, the expression of caspase genes increases sharply. This may initiate the onset of preterm labour.

Matrix metalloproteinases MMP-2 and MMP-9 as markers for the prediction of preeclampsia in the first trimester.

INTRODUCTION: Preeclampsia is a life-threatening condition for the mother and foetus. Globally, it is dia-gnosed in 10 mil. women every year, which accounts for 3% to 8% of all pregnancies. Currently there is no proven effective treatment for preeclampsia. The aforesaid text actualises the issue of predicting this complication. To determine the prognostic significance of matrix metalloproteinases-2 and -9 levels as early markers of preeclampsia, the present prospective study was conducted. MATERIALS AND METHODS: The levels of matrix metalloproteinases-2 and -9 were assessed in 72 patients. Thirty-four of them subsequently developed preeclampsia during pregnancy (20 patients with moderate preeclampsia, 14 patients with severe preeclampsia), and constituted the basic group; 38 patients made up the control group. RESULTS: In pregnant women with the subsequent development of preeclampsia, the level of matrix metalloproteinase-2 at 11-13 weeks of gestation was 155 ± 73.4 ng/mL and significantly exceeded its level in pregnant women without hypertensive disorders - 75.0 ± 32.8 ng/mL. The study conducted demonstrates a significantly lower concentration of matrix metalloproteinase-9 in pregnant women with preeclampsia compared to the control - 749 ± 296 ng/mL and 1,667 ± 552 ng/mL (P < 0.001). The performed research figures that in the first trimester, the cut-off value of matrix metalloproteinase-2 for predicting the development of preeclampsia is 102 ng/mL (sensitivity 88.24% and specificity 82.76%). For matrix metalloproteinase-9, a level of 980 ng/mL in the first trimester predicts the development of preeclampsia with a sensitivity of 85.29% and a specificity of 84.48%. CONCLUSION: The study established the cut-off values of matrix metalloproteinases-2 and -9 for predicting the development of preeclampsia in the first trimester.

Matrix Metalloproteinases MMP-2 and MMP-9 Occupy a New Role in Severe Preeclampsia.

INTRODUCTION: Preeclampsia (PE) is a life-threatening condition for the mother, the fetus, and the newborn. Matrix metalloproteinases (MMP) participate in the two primary stages of PE: remodeling of blood vessels at the stage of placental formation and the development of hypertension due to damage to the basement membrane of blood vessels. The object of the present study was to reveal the role of MMP-2 and MMP-9 in the development of severe preeclampsia. MATERIALS AND METHODS: We conducted a retrospective study that included 92 pregnant women at a gestational age of 26-38 weeks, of which the principal group consisted of 61 patients with severe PE. We divided the principal group into two subgroups: the first subgroup was designated the severe early-onset preeclampsia (EO-PE) group and consisted of 30 pregnant women. The second group was designated the severe late-onset preeclampsia (LO-PE) group, comprising 31 patients. We determined the plasma concentrations of MMPs 2 and 9 in the groups with an ELISA. RESULTS: In the group of PE patients with both EO-PE and LO-PE, the level of MMP-2 was significantly higher compared to the women undergoing normal pregnancy; and we observed no significant differences when we compared the levels of MMP-2 in the subgroups with EO-PE and LO-PE. Analysis of the concentrations of MMP-9 in EO-PE and LO-PE subgroups revealed attenuated levels of MMP-9 in both groups relative to the control group. We also noted a diminished level of MMP-9 in the EO-PE group compared to the LO-PE group. CONCLUSIONS: The significantly increased levels of MMP-2 in women-both in the EO-PE and LO severe PE subgroups-explain the participation of this enzyme in endothelial dysfunction in the second stage of severe PE. A diminution in MMP-9 in the EO-PE group confirmed the participation of MMP-9 in the process of spiral artery transformation.

Maternal Cardiac Function after Normal Delivery, Preeclampsia, and Eclampsia: A Prospective Study.

INTRODUCTION: The aim of this study is to assess maternal cardiac function in the postpartum period, after 2 and 6 months in the parturient with preeclampsia and eclampsia. MATERIALS AND METHODS: Prospective study: 90 postpartum women after preeclampsia and eclampsia and 55 patients after an uncomplicated pregnancy. The parameters of maternal hemodynamics were recorded on days 1, 3, 5, 9, and 14 of postpartum period, after 2 and 6 months. The cardiac parameters were assessed. RESULTS: PE is accompanied by increased peripheral vascular resistance. The indicator of vascular resistance, SVR, is elevated for both mild and severe PE. With mild PE, a significant increase in SVR is observed up to 5 days of postpartum period, with severe PE/E up to 9 days. We found that in case of severe PE, SVR remains elevated to 6 months after delivery. The parameters of the contractile function of the heart (ESV, EDV, SV, SI, CO, СI, MVCF) were significantly decreased: with mild PE up to 5-9 days, with severe up to 9-14 days of puerperia. ESV, SV, SI, CO, and CI remain low with severe PE up to 6 months. The revealed decreasing of contractile function of the heart is a sign of asymptomatic heart failure. CONCLUSIONS: The hemodynamics of the puerperas after PE and E is characterized by impaired contractility of the myocardium and an increase in the indices of peripheral resistance. The degree of deviation in the parameters of cardiac hemodynamics and vascular resistance depended on the severity of hypertensive complications of pregnancy.

Surface tension model for surfactant solutions at the critical micelle concentration.

A model for the limiting surface tension of surfactant solutions (surface tension at and above the critical micelle concentration, cmc) was developed. This model takes advantage of the equilibrium between the surfactant molecules on the liquid/vacuum surface and in micelles in the bulk at the cmc. An approximate analytical equation for the surface tension at the cmc was obtained. The derived equation contains two parameters, which characterize the intermolecular interactions in the micelles, and the third parameter, which is the surface area per surfactant molecule at the interface. These parameters were calculated using a new atomistic modeling approach. The performed calculations of the limiting surface tension for four simple surfactants show good agreement with experimental data (~30% accuracy). The developed model provides the guidance for design of surfactants with low surface tension values.