Vladimir Dobričić

Prediction of blood–brain barrier permeation of α-adrenergic and imidazoline receptor ligands using PAMPA technique and quantitative-structure permeability relationship analysis

Imidazoline receptor ligands are a numerous family of biologically active compounds known to produce central hypotensive effect by interaction with both α2-adrenoreceptors (α2-AR) and imidazoline receptors (IRs). Recent hypotheses connect those ligands with several neurological disorders. Therefore some IRs ligands are examined as novel centrally acting antihypertensives and drug candidates for treatment of various neurological diseases. Effective Blood-Brain Barrier (BBB) permeability (P(e)) of 18 IRs/α-ARs ligands and 22 Central Nervous System (CNS) drugs was experimentally determined using Parallel Artificial Membrane Permeability Assay (PAMPA) and studied by the Quantitative-Structure-Permeability Relationship (QSPR) methodology. The dominant molecules/cations species of compounds have been calculated at pH = 7.4. The analyzed ligands were optimized using Density Functional Theory (B3LYP/6-31G(d,p)) included in ChemBio3D Ultra 13.0 program and molecule descriptors for optimized compounds were calculated using ChemBio3D Ultra 13.0, Dragon 6.0 and ADMET predictor 6.5 software. Effective permeability of compounds was used as dependent variable (Y), while calculated molecular parametres were used as independent variables (X) in the QSPR study. SIMCA P+ 12.0 was used for Partial Least Square (PLS) analysis, while the stepwise Multiple Linear Regression (MLR) and Artificial Neural Networks (ANN) modeling were performed using STASTICA Neural Networks 4.0. Predictive potential of the formed models was confirmed by Leave-One-Out Cross- and external-validation and the most reliable models were selected. The descriptors that are important for model building are identified as well as their influence on BBB permeability. Results of the QSPR studies could be used as time and cost efficient screening tools for evaluation of BBB permeation of novel α-adrenergic/imidazoline receptor ligands, as promising drug candidates for treatment of hypertension or neurological diseases.

Inorganically modified diatomite as a potential prolonged-release drug carrier

Inorganic modification of diatomite was performed with the precipitation product of partially neutralized aluminum sulfate solution at three different mass ratios. The starting and the modified diatomites were characterized by SEM-EDS, FTIR, thermal analysis and zeta potential measurements and evaluated for drug loading capacity in adsorption batch experiments using diclofenac sodium (DS) as a model drug. In vitro drug release studies were performed in phosphate buffer pH6.8 from comprimates containing: the drug adsorbed onto the selected modified diatomite sample (DAMD), physical mixture of the drug with the selected modified diatomite sample (PMDMD) and physical mixture of the drug with the starting diatomite (PMDD). In vivo acute toxicity testing of the modified diatomite samples was performed on mice. High adsorbent loading of the selected modified diatomite sample (~250mg/g in 2h) enabled the preparation of comprimates containing adsorbed DS in the amount near to its therapeutic dose. Drug release studies demonstrated prolonged release of DS over a period of 8h from both DAMD comprimates (18% after 8h) and PMDMD comprimates (45% after 8h). The release kinetics for DAMD and PMDMD comprimates fitted well with Korsmeyer-Peppas and Bhaskar models, indicating that the release mechanism was a combination of non-Fickian diffusion and ion exchange process.

17β-carboxamide steroids – in vitro prediction of human skin permeability and retention using PAMPA technique

In this paper, twenty-two 17β-carboxamide steroids were synthesized from five corticosteroids (hydrocortisone, prednisolone, methylprednisolone, dexamethasone and betamethasone) in two steps. The first step was periodic acid oxydation of these corticosteroids to corresponding cortienic acids and the second step was amidation of thus obtained cortienic acids with esterified l-amino acids. These compounds are potential soft corticosteroids with local anti-inflammatory activity in the skin. Parallel artificial membrane permeability assay (PAMPA) was applied in order to predict permeability and retention of these compounds in human skin. Comparison of permeability and retention parameters between 17β-carboxamide steroids and corresponding corticosteroids was performed. Compounds with significantly higher retention were identified and the derivative that does not have significantly higher permeability was underlined. Molecular structures of all compounds were optimized by use of Gaussian semiempirical/PM3 method. Geometrical, thermodynamic, physicochemical and electronical molecular parameters of the optimized structures were calculated and quantitative structure-property relationship (QSPR) analysis was performed in order to explain permeability and retention of these compounds. ANN-, PLS- and MLR-QSPR models were created. Quality of these models was evaluated by commonly used statistical parameters and the most reliable models were selected. Analyzing descriptors in the selected models, main molecular properties that influence permeability and retention in the PAMPA artificial membrane were identified. Based on these data, further structural modifications could be applied in order to increase retention without significant increase of permeability, which can positively affect potential local anti-inflammatory activity of these compounds. Selected QSPR models could be used as in silico tool for predicting human skin permeability and retention of novel 17β-carboxamide steroids without performing PAMPA experiments.

Modified local diatomite as potential functional drug carrier--A model study for diclofenac sodium.

Diatomite makes a promising candidate for a drug carrier because of its high porosity, large surface area, modifiable surface chemistry and biocompatibility. Herein, refined diatomite from Kolubara coal basin, which complied with the pharmacopoeial requirements for heavy metals content and microbiological quality, was used as a starting material. Inorganic modification of the starting material was performed through a simple, one-step procedure. Significant increase in adsorbent loading with diclofenac sodium (DS) was achieved after the modification process (∼373mg/g) which enabled the preparation of comprimates containing therapeutic dose of the adsorbed drug. Adsorption of DS onto modified diatomite resulted in the alteration of the drugs XRD pattern and FTIR spectrum. In vitro drug release studies in phosphate buffer pH 7.5 demonstrated prolonged DS release over 8h from comprimates containing DS adsorbed on modified diatomite (up to 37% after 8h) and those containing physical mixture of the same composition (up to 45% after 8h). The results of in vivo toxicity testing on mice pointed on potential safety of both unmodified (starting) and modified diatomite. All these findings favor the application of diatomite as a potential functional drug carrier.

Biopartitioning micellar chromatography as a predictive tool for skin and corneal permeability of newly synthesized 17β-carboxamide steroids

In this paper, human skin and corneal permeability of twenty-two newly synthesized 17β-carboxamide steroids was predicted using biopartitioning micellar chromatography (BMC). These compounds are potential soft glucocorticoids with local anti-inflammatory activity when applied to the skin or eye. BMC systems are used to simulate physicochemical properties of human skin (BMC-skin) and cornea (BMC-cornea). Micellar mobile phase, consisted of 0.04 M solution of polyoxyethylene (23) lauryl ether (Brij 35), was prepared at different pH values - 5.50 (BMC-skin) and 7.50 (BMC-cornea). Retention factors (k), obtained by use of BMC, were calculated for all newly synthesized 17β-carboxamide steroids as well as for parent glucocorticoids (hydrocortisone, prednisolone, methylprednisolone, dexamethasone and betamethasone). Good correlation was obtained between BMC-skin retention factors and permeability coefficients calculated by use of the artificial membrane that simulates stratum corneum of the human skin. Quantitative structure-retention relationship (QSRR) study was performed in order to explain retention factors of these compounds in the tested BMC systems. ANN-QSRR(k), PLS-QSRR(k) and MLR-QSRR(k) models, created by use of BMC-skin retention data, were compared and optimal model (PLS-QSRR(k)) was selected. Molecular descriptors of the selected model indicate that lipophilicity and number of short C-C fragments of tested compounds have the strongest influence on the retention in the BMC-skin system and presumably on their in vivo permeability through human skin. The same model can be applied to the BMC-cornea system and the same conclusion can be drawn for corneal permeability. This model could be used as a predictive tool for the synthesis of novel 17β-carboxamide steroids with desirable permeability through human skin or cornea, depending on their potential pharmacological application.

Evaluation of critical formulation parameters in design and differentiation of self-microemulsifying drug delivery systems (SMEDDSs) for oral delivery of aciclovir.

The study investigated the influence of formulation parameters for design of self-microemulsifying drug delivery systems (SMEDDSs) comprising oil (medium chain triglycerides) (10%), surfactant (Labrasol(®), polysorbate 20, or Kolliphor(®) RH40), cosurfactant (Plurol(®) Oleique CC 497) (q.s. ad 100%), and cosolvent (glycerol or macrogol 400) (20% or 30%), and evaluate their potential as carriers for oral delivery of a poorly permeable antivirotic aciclovir (acyclovir). The drug loading capacity of the prepared formulations ranged from 0.18-31.66 mg/ml. Among a total of 60 formulations, three formulations meet the limits for average droplet size (Z-ave) and polydispersity index (PdI) that have been set for SMEDDSs (Z-ave≤100nm, PdI<0.250) upon spontaneous dispersion in 0.1M HCl and phosphate buffer pH 7.2. SMEDDSs with the highest aciclovir loading capacity (24.06 mg/ml and 21.12 mg/ml) provided the in vitro drug release rates of 0.325 mg cm(-2)min(-1) and 0.323 mg cm(-2)min(-1), respectively, and significantly enhanced drug permeability in the parallel artificial membrane permeability assay (PAMPA), in comparison with the pure drug substance. The results revealed that development of SMEDDSs with enhanced drug loading capacity and oral delivery potential, required optimization of hydrophilic ingredients, in terms of size of hydrophilic moiety of the surfactant, surfactant-to-cosurfactant mass ratio (Km), and log P of the cosolvent.

Investigation of Solvolysis Kinetics of New Synthesized Fluocinolone Acetonide C-21 Esters—An In Vitro Model for Prodrug Activation

In this study the solvolysis of newly synthesized fluocinolone acetonide C-21 esters was analysed in comparison with fluocinonide during a 24-hour period of time. The solvolysis was performed in an ethanol-water (90:10 v/v) mixture using the excess of NaHCO3. The solvolytic mixtures of each investigated ester have been assayed by a RP-HPLC method using isocratic elution with methanol-water (75:25 v/v); flow rate 1 mL/min; detection at 238 nm; temperature 25 °C. Solvolytic rate constants were calculated from the obtained data. Geometry optimizations and charges calculations were carried out by Gaussian W03 software. A good correlation (R = 0.9924) was obtained between solvolytic rate constants and the polarity of the C-O2 bond of those esters. The established relation between solvolytic rate constant (K) and lipophilicity (cLogP) with experimental anti-inflammatory activity could be indicative for topical corticosteroid prodrug activation.

Design, Synthesis, and Local Anti‐Inflammatory Activity of 17β‐Carboxamide Derivatives of Glucocorticoids

Molecular docking studies were performed on 18 17β‐carboxamide steroids in order to select compounds with potential local anti‐inflammatory activity. These derivatives are amides of cortienic acids (obtained from hydrocortisone, prednisolone, and methylprednisolone) with methyl or ethyl esters of six amino acids. Interactions with the glucocorticoid receptor (GR), binding energies and ligand efficiency values of these compounds were compared with dexamethasone and cortienic acid obtained from prednisolone (inactive metabolite). On the basis of molecular docking studies, seven compounds were selected and their binding affinities for the GR were predicted by use of the exponential model created in this study. Subsequently, selected compounds were synthesized in good yields by use of modified N,N′‐dicyclohexylcarbodiimide (DCC)/1‐hydroxybenzotriazole (HOBt) coupling procedure. Finally, the local anti‐inflammatory activity of the synthesized compounds was examined by use of the croton oil‐induced ear edema test. In vivo evaluation of systemic side effects as well as in silico prediction of metabolism were performed on the derivative with the best local anti‐inflammatory activity. The combination of molecular docking studies and the exponential model for the GR binding affinity prediction could be used as an in silico tool for the rational design of novel 17β‐carboxamide steroids with potentially better biological profile than dexamethasone.

Design and Synthesis of Novel Deuterated Ligands Functionally Selective for the γ-Aminobutyric Acid Type A Receptor (GABAAR) α6 Subtype with Improved Metabolic Stability and Enhanced Bioavailability

Recent reports indicate that α6β2/3γ2 GABAAR selective ligands may be important for the treatment of trigeminal activation-related pain and neuropsychiatric disorders with sensori-motor gating deficits. Based on 3 functionally α6β2/3γ2 GABAAR selective pyrazoloquinolinones, 42 novel analogs were synthesized, and their in vitro metabolic stability and cytotoxicity as well as their in vivo pharmacokinetics, basic behavioral pharmacology, and effects on locomotion were investigated. Incorporation of deuterium into the methoxy substituents of the ligands increased their duration of action via improved metabolic stability and bioavailability, while their selectivity for the GABAAR α6 subtype was retained. 8b was identified as the lead compound with a substantially improved pharmacokinetic profile. The ligands allosterically modulated diazepam insensitive α6β2/3γ2 GABAARs and were functionally silent at diazepam sensitive α1β2/3γ2 GABAARs, thus no sedation was detected. In addition, these analogs were not cytotoxic, which render them interesting candidates for treatment of CNS disorders mediated by GABAAR α6β2/3γ2 subtypes.

In vitro prediction of gastrointestinal absorption of novel β-hydroxy-β-arylalkanoic acids using PAMPA technique

&NA; Prediction of gastrointestinal absorption of thirteen newly synthesized &bgr;‐hydroxy‐&bgr;‐arylalkanoic acids (HAA) and ibuprofen was performed using PAMPA test. The highest values of PAMPA parameters (%T and Papp) were calculated for 1C, 1B and 2C and these parameters were significantly lower in comparison to ibuprofen. QSPR analysis was performed in order to identify molecular descriptors with the highest influence on %T and − logPapp and to create models which could be used for the design of novel HAA with improved gastrointestinal absorption. Obtained results indicate that introduction of branched side chain, as well as introduction of substituents on one phenyl ring (which disturb symmetry of the molecule) could have positive impact on gastrointestinal absorption. On the basis of these results, six novel HAA were designed and PAMPA parameters %T and − logPapp were predicted by use of selected QSPR models. Designed derivatives should have better gastrointestinal absorption than HAA tested in this study. Graphical abstract Figure. No caption available.