The application of discriminant analysis and Machine Learning methods as tools to identify and classify compounds with potential as transdermal enhancers.

Discriminant analysis (DA) has previously been shown to allow the proposal of simple guidelines for the classification of 73 chemical enhancers of percutaneous absorption. Pugh et al. employed DA to classify such enhancers into simple categories, based on the physicochemical properties of the enhancer molecules (Pugh et al., 2005). While this approach provided a reasonable accuracy of classification it was unable to provide a consistently reliable estimate of enhancement ratio (ER, defined as the amount of hydrocortisone transferred after 24h, relative to control). Machine Learning methods, including Gaussian process (GP) regression, have recently been employed in the prediction of percutaneous absorption of exogenous chemicals (Moss et al., 2009; Lam et al., 2010; Sun et al., 2011). They have shown that they provide more accurate predictions of these phenomena. In this study several Machine Learning methods, including the K-nearest-neighbour (KNN) regression, single layer networks, radial basis function networks and the SVM classifier were applied to an enhancer dataset reported previously. The SMOTE sampling method was used to oversample chemical compounds with ER>10 in each training set in order to improve estimation of GP and KNN. Results show that models using five physicochemical descriptors exhibit better performance than those with three features. The best classification result was obtained by using the SVM method without dealing with imbalanced data. Following over-sampling, GP gives the best result. It correctly assigned 8 of the 12 "good" (ER>10) enhancers and 56 of the 59 "poor" enhancers (ER<10). Overall success rates were similar. However, the pharmaceutical advantages of the Machine Learning methods are that they can provide more accurate classification of enhancer type with fewer false-positive results and that, unlike discriminant analysis, they are able to make predictions of enhancer ability.

Formulation and characterization of a captopril ethyl ester drug-in-adhesive-type patch for percutaneous absorption.

BACKGROUND: The ethyl ester of captopril has been shown to exhibit enhanced permeation across human skin compared to the parent drug. A drug-in-adhesive patch formulation of a captopril ethyl ester was therefore developed for optimum drug release. METHOD: A wide range of transdermal patches were prepared using two commercially available bioadhesive polymers. Investigational screening was conducted on the patches using microscopy, texture profile analysis, and infrared spectroscopy. Drug release profiles of suitable patches were obtained using both polydimethylsiloxane (Silastic) and porcine skin in vitro. RESULTS: Diffusion results across Silastic showed a gradual plateau in flux with increased drug loading that may be attributable to intramolecular interactions while flux across porcine skin was seen to increase with increasing patch thickness and attained a therapeutic level. CONCLUSIONS: This study demonstrated that adhesion and drug loading are significant factors in optimizing a topical patch formulation for the delivery of a captopril prodrug.

The application of Gaussian processes in the prediction of percutaneous absorption.

OBJECTIVES: The aim was to assess mathematically the nature of a skin permeability dataset and to determine the utility of Gaussian processes in developing a predictive model for skin permeability, comparing it with existing methods for deriving predictive models. METHODS: Principal component analysis was carried out in order to determine the nature of the dataset. MatLab software was used to assess the performance of Gaussian process, single linear networks (SLN) and quantitative structure-permeability relationships (QSPRs) using a range of statistical measures. KEY FINDINGS: Principal component analysis showed that the dataset is inherently non-linear. The Gaussian process model yielded a predictive model that provides a significantly more accurate estimate of skin absorption than previous models, particularly QSPRs (which were consistently worse than Gaussian process or SLN models), and does so across a wider range of molecular properties. Gaussian process models appear particularly capable of providing excellent predictions where previous studies have shown QSPRs to fail, such as where penetrants have high log P and high molecular weight. CONCLUSIONS: A non-linear approach was more appropriate than QSPRs or SLNs for the analysis of the dataset employed herein, as the prediction and confidence values in the prediction given by the Gaussian process are better than with other methods examined. Gaussian process provides a novel way of analysing skin absorption data that is substantially more accurate, statistically robust and reflective of our empirical understanding of skin absorption than the QSPR methods so far applied to skin absorption.

Metabolism of captopril carboxyl ester derivatives for percutaneous absorption.

OBJECTIVES: To determine the metabolism of captopril n-carboxyl derivatives and how this may impact on their use as transdermal prodrugs. The pharmacological activity of the ester derivatives was also characterised in order to compare the angiotensin converting enzyme inhibitory potency of the derivatives compared with the parent drug, captopril. METHODS: The metabolism rates of the ester derivatives were determined in vitro (using porcine liver esterase and porcine ear skin) and in silico (using molecular modelling to investigate the potential to predict metabolism). KEY FINDINGS: Relatively slow pseudo first-order metabolism of the prodrugs was observed, with the ethyl ester displaying the highest rate of metabolism. A strong relationship was established between in-vitro methods, while in-silico methods support the use of in-vitro methods and highlight the potential of in-silico techniques to predict metabolism. All the prodrugs behaved as angiotensin converting enzyme inhibitors, with the methyl ester displaying optimum inhibition. CONCLUSIONS: In-vitro porcine liver esterase metabolism rates inform in-vitro skin rates well, and in-silico interaction energies relate well to both. Thus, in-silico methods may be developed that include interaction energies to predict metabolism rates.

Transdermal delivery enhancement of haloperidol from gel formulations by 1,8-cineole.

The feasibility of using 10% 1,8-cineole as an enhancer for transdermal delivery of haloperidol has been examined. In-vitro transdermal delivery across full-thickness human, rabbit and hairless mouse skins was measured from three polymer gel systems, hypromellose (hydroxypropylmethylcellulose), Carbomer (Carbopol) 940 and macrogol (polyethylene glycol) using Franz cells. Values for the permeability coefficient kp, calculated as the product (Kh)x(D/h2) where these two factors were obtained from curve fitting of the non-steady-state equation over 24 h, were similar from the three formulations. The value of kp from hypromellose was significantly enhanced by cineole by factors of 6.2 (4.6-8.1), 5.6 (5.0-6.2) and 3.0 (2.6-3.4) for human, rabbit and mouse, respectively (mean and 95% confidence intervals). Enhancement ratios for K: 13.3 (8.3-20), 3.1 (2.5-3.9) and 2.0 (1.5-2.6), were higher than those for D: 0.47 (0.41-0.55), 1.8 (1.6-2.1) and 1.5 (1.3-1.8). This suggested that the barrier function of the skin lipids was marginally affected and the main effect was to increase the thermodynamic activity of the drug in the barrier. The enhancement achieved in human skin suggested that delivery could be safely enhanced by terpenoids.

Sustained expression of hepatocyte nuclear factor-6 leads to loss of pancreatic beta-cells by apoptosis.

Hepatocyte nuclear factor-6 (HNF-6) is the ONECUT-homeodomain transcription factor that is enriched in liver and also present in pancreas and central nervous system. It is expressed in the pancreatic bud at E10.5. In adult pancreas, its expression is restricted to the exocrine pancreas and duct cells. Since duct cells are thought to be precursors of endocrine cells and HNF-6 is involved in the regulation of the expression of HNF-4alpha and -1beta, genes that cause maturity onset diabetes of the young (MODY), we hypothesized that the sustained expression of HNF-6 would affect beta-cell function. We generated transgenic mice over-expressing human HNF-6 using the mouse insulin I promoter (MIP). We obtained one female founder in which the transgene had been incorporated into two sites; the chromosome (Ch) 14 and the X chromosome. The integration site of the latter was within centromeric heterochromatin and the transgene was inactivated. Studies on mice in which the transgene was integrated into Ch14 showed beta-cell specific defects functionally and pathologically. The insulin secretory response to glucose and arginine in the in situ-perfused pancreas was also significantly impaired in these mice. Immunohistochemical analysis revealed that the islets were smaller and had an abnormal architecture with an inverted ratio of alpha- and beta-cells resulting from beta-cell loss to 30% by 6-wk of age. The decreased number of beta-cells was quantified first time by fluorescent activated cell sorting using entire pancreata from the transgenic mice crossed with MIP-green fluorescent protein (GFP) mice. This severe loss of beta-cells involved programmed cell death.

Design, synthesis and characterization of captopril prodrugs for enhanced percutaneous absorption.

Most drugs are designed primarily for oral administration, but the activity and stability profiles desirable for this route often make them unsuitable for transdermal delivery. We were therefore interested in designing analogues of captopril, a model drug with poor percutaneous penetration, for which the sustained steady-state blood plasma level associated with transdermal delivery (and which is unattainable orally) would be particularly beneficial. Quantitative structure-permeability relationships (QSPRs) predicted that ester and thiol prodrug derivatives of captopril would have lower maximal transdermal flux (J(m)) than the parent drug, since the increases in permeability coefficient (k(p)) of prodrugs would be outweighed by the reductions in aqueous solubility. Therefore, the aim of this study was to synthesize a series of prodrugs of captopril and to determine if a QSPR model could be used to design therapeutically viable prodrugs. Molecules with the highest predicted k(p) values were synthesized and characterized, and J(m) measured in Franz diffusion cells from saturated aqueous donor across porcine skin (fresh and frozen). In-vitro metabolism was also measured. Captopril and the prodrugs crossed the skin relatively freely, with J(m) being highest for ethyl to butyl esters. Substantial first-order metabolism of the prodrugs was observed, suggesting that their enhanced percutaneous absorption was complemented by their metabolic performance. The results suggested that QSPR models provided excellent enhancements in drug delivery. This was not seen at higher lipophilicities, suggesting that issues of solubility need to be considered in conjunction with any such use of a QSPR model.

Gastroretentive dosage forms: overview and special case of Helicobacter pylori.

The challenge to develop efficient gastroretentive dosage forms began about 20 years ago, following the discovery of Helicobacter pylori by Warren and Marshall. In order to understand the real difficulty of increasing the gastric residence time of a dosage form, we have first summarized the important physiologic parameters, which act upon the gastric residence time. Afterwards, we have reviewed the different drug delivery systems designed until now, i.e. high-density, intragastric floating, expandable, superporous hydrogel, mucoadhesive and magnetic systems. Finally, we have focused on gastroretentive dosage forms especially designed against H. pylori, including specific targeting systems against this bacterium.

Discriminant analysis as a tool to identify compounds with potential as transdermal enhancers.

Structure-activity relationships were sought for 73 enhancers of hydrocortisone permeation from propylene glycol across hairless mouse skin. Enhancers had chain lengths (CC) from 0 to 16 carbon atoms, 1 to 8 H-bonding atoms (HB), molecular weight 60 to 450, log P (calculated) -1.7 to 9.7 and log S (calculated) -7.8 to 0.7. These predictive properties were chosen because of their ready availability. Enhancement ratio (ER) was defined as hydrocortisone transferred after 24 h relative to control. Values for the ER ranged from 0.2 to 25.3. Multiple regression analysis failed to predict activity; ER values for the 'good' enhancers (ER > 10) were underestimated. Simple guidelines suggested that high ER was associated with CC > 12 and HB 2-5. This was refined by multivariate analysis to identify significant predictors. Discriminant analysis using CC, HB, and molecular weight correctly assigned 11 of the 12 'good' enhancers (92%). The incorrectly assigned compound was a known, idiosyncratic Br compound. Seventeen of the 61 'poor' enhancers (28%) were incorrectly assigned but four could be considered marginal (ER > 8). The success of this simple approach in identifying potent enhancers suggested its potential in predicting novel enhancer activity.

Topical delivery of retinyl ascorbate co-drug. 2. Comparative skin tissue and keratin binding studies.

Retinyl ascorbate (RA-AsA), an ester co-drug of vitamins A (RA) and C (AsA), is proposed as a topical antioxidant/cell division regulator for reducing UV-induced generation of free radicals and disrupted dermal cell growth. The efficacy of dermatological agents is influenced by their retention within the skin, which is increased by the interaction with skin components. Keratin is the major protein (approximately 95%) in the skin, and this paper reports the binding of RA-AsA, RA, AsA, retinol, ascorbic acid palmitate and retinol palmitate to three tissues-human callus, pig ear skin and bovine horn keratin. Tissue samples were incubated with solutions of compounds and the uptake measured as the ratio of bound/free compound at equilibrium. Binding to keratin was assessed using delipidised tissue, and was much higher for the polar compounds, suggesting dipolar/H-bonding interaction. Binding strength was ranked as human > porcine > bovine, but there was no distinction for highly lipophilic compounds. The binding characteristic of native tissues was complicated by lipid content of the tissues. There seemed to be a dual effect. The binding of very lipophilic materials increased with lipid content, implying that a substantial amount is dissolved in the lipid matrix. For highly polar AsA, lipid content decreased the binding, suggesting that the lipid reduced the strong polar interactions with skin protein/keratin.

Receptor regulatory properties evident in the molecular similarity of serotonin receptor ligands and purine nucleotides.

Previous computational studies have explored the relative molecular similarity inherent in the ligands of neurotransmitter-regulated cell receptors and purine nucleotides. This study presents the results of an investigation of the major serotonin (5-HT) receptor classes, using molecular superimposition and fitting data. Ligands for 5HT(1B/C/D) and 5HT(4/7) receptors identified pharmacophores in the adenine ring of ATP. 5-HT(2) and 5-HT(3) receptor ligands identified pharmacophores in the guanosine nucleotide and cyclic nucleotide, respectively. The described molecular similarity is consistent with the cyclic nucleotide responses observed during signal transduction events initiated by 5-HT, and the reported similarity between ligands of the 5-HT(1B) and 5-HT(1D), 5-HT(1A) and 5-HT(7), and 5-HT(4) and 5-HT(3) receptors. The results are discussed in terms of current pharmacophoric models and signal transduction events involving interaction between G-protein receptors and catalytic sites.

Simple rules defining the potential of compounds for transdermal delivery or toxicity.

PURPOSE: Simple rules based on readily accessible physicochemical properties enable identification of solutes that penetrate skin very slowly or rapidly. METHODS: Literature in vitro maximal flux values (Jmax) across human skin were collected for 87 penetrants. Penetrants were assigned as "good" (Jmax > 10(-5.52) mole x cm(-2) x h(-1)), "bad" (Jmax < 10(-8.84) mole x cm(-2) x h(-1)) or "intermediate" based on mean +/- 1SD. The feasibility of using readily available physicochemical properties, such as molecular weight (MW), melting point (MP, degrees K), octanol-water partition coefficient (K), water solubility (S, molarity), number of atoms available for H-bonding (HB), in assigning solutes was examined. RESULTS: Good penetrants had MW < or = 152, log S > -2.3, HB < or = 5, log K < 2.6, MP < or = 432. Bad penetrants had MW > 213, log S < -1.6, HB > or = 4, log K > 1.2, MP > or = 223. Discriminant analysis using MW, HB, log K correctly assigned 70% of compounds. Individual success rates were good (88%), intermediate (58%), bad (93%). Aqueous Jmax data for 148 test solutes were used for validation. Discriminant analysis assigned 76% of compounds, with individual rates of good (76%), intermediate (67%), and bad (97%). No good penetrants were misclassified as bad or vice versa. CONCLUSIONS: These rules enable rapid screening of potential drug delivery candidates and environmental exposure risks.

Effect of vehicle pretreatment on the flux, retention, and diffusion of topically applied penetrants in vitro.

PURPOSE: The flux of a topically applied drug depends on the activity in the skin and the interaction between the vehicle and skin. Permeation of vehicle into the skin can alter the activity of drug and the properties of the skin barrier. The aim of this in vitro study was to separate and quantify these effects. METHODS: The flux of four radiolabeled permeants (water, phenol, diflunisal, and diazepam) with log Koct/water values from 1.4 to 4.3 was measured over 4 h through heat-separated human epidermis pretreated for 30 min with vehicles having Hildebrand solubility parameters from 7.9 to 23.4 (cal/cm3)1/2. RESULTS: Enhancement was greatest after pretreatment with the more lipophilic vehicles. A synergistic enhancement was observed using binary mixtures. The flux of diazepam was not enhanced to the same extent as the other permeants, possibly because its partitioning into the epidermis is close to optimal (log Koct 2.96). CONCLUSION: An analysis of the permeant remaining in the epidermis revealed that the enhancement can be the result of either increased partitioning of permeant into the epidermis or an increasing diffusivity of permeants through the epidermis.

Physico-chemical determinants of dermal drug delivery: effects of the number and substitution pattern of polar groups.

The aim of this study was to investigate the effect of number and substitution pattern of -OH groups of a set of phenols on the in vitro permeation of heat-separated human epidermis. The diffusion was calculated from Log(D/x)=logk(p)-0.59logK(oct)+0.024 (D, diffusion coefficient; x, pathlength; k(p), permeability coefficient (cm/h); and K(oct), octanol-water partition coefficient). The main factors reducing D were the dipolar and hydrogen bonding capabilities of the permeants quantified as their Hansen partial solubility parameters delta(p) and delta(h). These parameters are significantly reduced by the degree of symmetry of the molecule, so that phloroglucinol (1,3,5-benzenetriol), with three -OH groups, diffuses more rapidly that phenol. When symmetry is absent, as in 1,2,4-benzenetriol, the number of -OH groups results in very slow diffusion. D/x (cm/h) was related to the combined solubility parameter delta(a) defined as radical(delta(p)(2)+delta(h)(2)) by: (D/x)=0.0024-0.000065delta(a) (n=7, R(2)=0.70, P=0.012).

Receptor regulatory properties evident in the molecular similarity of dopamine receptor ligands and purine nucleotides.

Computational studies have revealed similarities in the relative configurations of purine nucleotides and ligands for histamine, acetylcholine and adrenergic receptors. In common with other G-protein-regulated receptors, dopamine receptors are associated with specific changes in nucleotide levels during signal transduction processes. The purpose of this study was to investigate molecular similarity in dopamine receptor ligands and purine nucleotides. Molecular superimposition and fitting data for D1-like receptor ligands identified a pharmacophore in the adenine and ribose rings of ATP. D2-like agonists and antagonists related to a pharmacophore in the guanine and ribose rings of GTP. The results are consistent with the hypothesis that the dopamine receptor family may have evolved from receptors for the ATP and GTP nucleotides.

The effect of the nature of H-bonding groups on diffusion through PDMS membranes saturated with octanol and toluene.

The permeation of a series of structurally related compounds across silicone membranes (PDMS) was studied. The PDMS was saturated either with toluene, to mimic a functionally inert barrier, or octanol, to mimic the polar/hydrogen bonding environment of the stratum corneum lipid barrier. Phenol, salicylic acid, benzoic acid, anisole, phenylethanol and benzyl alcohol were chosen in an attempt to relate permeation to their different H-bonding capabilities. The flux was lower through the octanol system suggesting retardation by polar/H-bonding interactions. Separation of the permeability coefficient into its thermodynamic (partition coefficient) and kinetic (diffusion coefficient) terms suggests that the effect of altering polarity within the membrane has a greater impact on the diffusion of permeant rather than its chemical potential within the membrane.

Regulation of pancreatic beta-cell growth and survival by the serine/threonine protein kinase Akt1/PKBalpha.

The physiological performance of an organ depends on an interplay between changes in cellular function and organ size, determined by cell growth, proliferation and death. Nowhere is this more evident than in the endocrine pancreas, where disturbances in function or mass result in severe disease. Recently, the insulin signal-transduction pathway has been implicated in both the regulation of hormone secretion from beta cells in mammals as well as the determination of cell and organ size in Drosophila melanogaster. A prominent mediator of the actions of insulin and insulin-like growth factor 1 (IGF-1) is the 3'-phosphoinositide-dependent protein kinase Akt, also known as protein kinase B (PKB). Here we report that overexpression of active Akt1 in the mouse beta cell substantially affects compartment size and function. There was a significant increase in both beta-cell size and total islet mass, accompanied by improved glucose tolerance and complete resistance to experimental diabetes.

Probing the effect of vehicles on topical delivery: understanding the basic relationship between solvent and solute penetration using silicone membranes.

PURPOSE: In the present study we examined the relationship between solvent uptake into a model membrane (silicone) with the physical properties of the solvents (e.g., solubility parameter, melting point, molecular weight) and its potential predictability. We then assessed the subsequent topical penetration and retention kinetics of hydrocortisone from various solvents to define whether modifications to either solute diffusivity or partitioning were dominant in increasing permeability through solvent-modified membranes. METHODS: Membrane sorption of solvents was determined from weight differences following immersion in individual solvents, corrected for differences in density. Permeability and retention kinetics of 3H-hydrocortisone, applied as saturated solutions in the various solvents, were determined over 48 h in horizontal Franz-type glass diffusion cells. RESULTS: Solvent sorption into the membrane could be related to differences in solubility parameters, MW and hydrogen bonding (r2=0.76). The actual and predicted volume of solvent sorbed into the membrane was also found to be linearly related to Log hydrocortisone flux, with changes in both diffusivity and partitioning of hydrocortisone observed for the different solvent vehicles. CONCLUSIONS: A simple structure-based predictive model can be applied to the sorption of solvents into silicone membranes. Changes in solute diffusivity and partitioning appeared to contribute to the increased hydrocortisone flux observed with the various solvent vehicles. The application of this predictive model to the more complex skin membrane remains to be determined.

Calpains play a role in insulin secretion and action.

Studies of the genetic basis of type 2 diabetes suggest that variation in the calpain-10 gene affects susceptibility to this common disorder, raising the possibility that calpain-sensitive pathways may play a role in regulating insulin secretion and/or action. Calpains are ubiquitously expressed cysteine proteases that are thought to regulate a variety of normal cellular functions. Here, we report that short-term (4-h) exposure to the cell-permeable calpain inhibitors calpain inhibitor II and E-64-d increases the insulin secretory response to glucose in mouse pancreatic islets. This dose-dependent effect is observed at glucose concentrations above 8 mmol/l. This effect was also seen with other calpain inhibitors with different mechanisms of action but not with cathepsin inhibitors or other protease inhibitors. Enhancement of insulin secretion with short-term exposure to calpain inhibitors is not mediated by increased responses in intracellular Ca2+ or increased glucose metabolism in islets but by accelerated exocytosis of insulin granules. In muscle strips and adipocytes, exposure to both calpain inhibitor II and E-64-d reduced insulin-mediated glucose transport. Incorporation of glucose into glycogen in muscle also was reduced. These results are consistent with a role for calpains in the regulation of insulin secretion and insulin action.

Review of the Naval Health Research Center's development of medical information systems for far-forward echelons of care, 1983 to 1997.

Medical treatment information must be gathered quickly and accurately to ensure continuity of care at far-forward echelons. The manual documentation methods in use during the Vietnam War revealed the need for considerable improvements in medical information documentation, patient tracking, and effective transfer of data throughout each of the first three echelons of care. The U.S. military determined that automation would result in the greatest enhancement of documentation techniques. The Naval Health Research Center (NHRC) has been an integral part of the development of medical information systems for the Navy's far-forward echelons of care. The prototypes designed by NHRC for echelons I and II can successfully raise the standard of treatment while simultaneously reducing the number of individuals needed for administrative duties and increasing the number of medical staff available for patient care. An overview of NHRC's work in medical information systems from 1983 to 1997 is presented.