ix Objectives: To quantify polypharmacy and assess the socio-economic predictors of medication use and expenditure in diabetics. Methods: This study analyzed adult diabetes patients using a nationally representative sample in Medical Expenditure Panel Survey in 2006. Top ten most highly utilized drug classes were identified. Descriptive statistics were used to portray the patients’ medication utilization and spending. Generalized linear models were conducted to assess the socio-economic variants in drug use and spending. Results: On average, a diabetes patient had 45 prescriptions in 2006, for total annual spending of $3,161. A diabetes patient used drugs from 3.43 classes within top ten drug classes. Races and insurance coverage are associated with drug use and spending, holding other factors constant. Conclusion: Diabetes patients use multiple classes of drugs. Insurance coverage and races are related with drug spending and utilization. More research is needed to evaluate the potential risks of drug-drug interactions due to polypharmacy.

This research explored quantitative relationships (QSPKR) between different molecular descriptors and pertinent, systemic PK properties for 14 calcium channel blockers (CCB). Physicochemical properties (PC) such as molecular weight (MW), molar volume (MV), calculated logP (clogP), pKa, calculated logD7.4 (clogD), % ionized at pH 6.3 and pH 7.4, hydrogen bond donors (HBD), hydrogen bond acceptors (HBA), and number of rotatable bonds (nRot) were chosen as possible predictor variables for systemic PK properties for CCB, obtained from pertinent literature, assessing the PK of CCB after intravenous administration to healthy humans. All PC properties and molecular descriptors were computed using ACD-solubility/DB 12.01. Total body clearance (CLtot), steady-state volume of distribution (Vdss), total area under the plasma concentration-time profile (AUCoo), terminal half-life (t1/2), and fraction of drug excreted unchanged in urine (fe), if available, were obtained or derived from original references, exclusively from IV studies that administered CCB to healthy human volunteers. Several articles focused on drug interactions with grapefruit juice or the impact of renal/hepatic dysfunction, and in such cases, data from the healthy control group were used. Each study was evaluated for study design, PK sampling schedule, bioanalytical and PK analysis methods before inclusion into the final database. The assumption of linear systemic PK was verified by assessing AUCoo versus (IV) dose. Plasma protein binding information was collected from in-vitro experiments to obtain the fraction unbound in plasma (fu). Unbound volume of distribution at a steady state (Vdssu), unbound total (CLtotu), renal (CLrenu), and non-renal clearance (CLnonrenu) were estimated and compared with the relevant physiological references for Vdssu (plasma volume, blood volume, extracellular and intracellular spaces, total body water and body weight) and for the unbound clearances (liver blood flow, renal plasma flow, and glomerular filtration rate, GFR). Final PK property values were obtained by averaging across available studies. The distribution of both PC and PK properties were evaluated, and correlation matrices amongst PC properties were constructed to assess for collinearity. If two PC descriptors were found to be collinear, i.e. r, ≥ 0.8, only one of them was used in the final univariate analysis. Finally, univariate linear regression of all PK variables versus each molecular descriptor was performed; any relationship with p<0.05 and r2≥0.30 was considered to be statistically significant. The PC properties of the final 14 CCB were reasonably normally distributed with few exceptions. Overall, CCBs are small (MW range of 316-496 Da), basic and lipophilic (logD7.4 range of 1.5-5.1) molecules. On the other hand, for the PK properties, the distributions were found to be skewed with high standard deviations. Thus, all PK variables (except fu) were log-transformed. Although CCB are mostly highly plasma protein bound (fu range of 0.2-20%), they are characterized by extensive extravascular tissue distribution (Vdss range of 0.6-20.4 l/kg) and high, mainly metabolic, clearance (CLtot range of 3.7-131.7 ml/min/kg). Clevidipine is the only CCB undergoing extensive, extra-hepatic ester hydrolysis, responsible for the highest CLtot value. Urinary excretion for CCB is negligible. Amlodipine is a PK outlier due to its high Vdss (20.4 l/kg) and low CLtot (6.9 ml/min/kg, due to low hepatic extraction) with fu of 2%. Therefore, the final QSPKR analysis was performed including, as well as excluding amlodipine. Excluding amlodipine, the relationship between fu and logD7.4 was negative and significant (r2 of 0.4, n=12). The relationships between CLtotu, CLnonrenu and CLrenu and logD7.4 were found to be positive and significant (r2 between 0.6-0.7, n=3-12); none of the other PC variables affected any of the clearance terms. Although the relationship between Vdssu and logD7.4 was not significant (r2 of 0.25, n=12), it showed the expected positive slope. In fact, after removing bepridil (the remaining outlier in Vdssu), the relationship with logD7.4 became statistically significant (r2=0.46, n=11). The QSPKR obtained in this study for CCB, with logD7.4 being the main PC determinant for systemic PK properties, were similar to those previously reported for opioids, β-adrenergic receptor ligands and benzodiazepines. However, slope estimates for the relationships of CLnonrenu and CLtotu as a function of logD7.4 for CCB were higher compared to these previously studied compounds, which showed higher sensitivity, most likely as a result of their higher lipophilicity. Overall, lipophilicity measured as logD7.4 was found to be a statistically significant and plausible PC determinant for the biologically relevant systemic PK properties for CCB and other classes of drugs.

Heparin is a highly sulfated glycosaminoglycan with potent anticoagulant, antimetastatic, and anti-inflammatory effects. Polymeric and polyanionic nature of heparin makes dosing and side effects a nightmare for healthcare professionals. Our laboratory has proposed appropriately designed, small, highly sulfated aromatic molecules as potential mimetics of heparin. These easier-to-synthesize small molecules have been shown to possess interesting pharmacological and improved toxicological profiles. However, the detection and characterization of these highly sulfated molecules is challenging. A robust RP-IP UPLC-MS method was developed to successfully retain, resolve and quantify sulfated non-saccharide GAG mimetics without the requirement of pre- or post- column derivatization. Comparative analysis reveals intricate dependence of resolution and ionization on the structure of ion-pairing agents. This is the first report showing systematic use of MS cone voltage to fingerprint sulfated GAG mimetics, perhaps eliminating the need for tandem MS techniques.

Background. Allometric scaling (AS) is widely used in predicting human clearance (CL) based on animal data. Substantial prediction errors have been commonly observed and various modifications to AS have not provided a broad reliable improvement. In this study, an extensive data set was assembled including animal and human systemic CL and physiochemical properties. The allometric exponents were calculated based on multiple species AS and single-species AS methods. The correlations between the allometic exponents and physiochemical properties were evaluated in an attempt to find covariates that may explain the inter-compound variability in the allometric exponents. Lastly, the statistical approaches in analyzing the allometric function were evaluated with the collected data. Methods. 1- A nonlinear mixed effect modeling (MEM) approach was performed to investigate the central tendency and distribution of AS exponents as well as to identify whether there are any correlations between the allometric exponent, and coefficient, with the physicochemical and drug metabolism and pharmacokinetics (DMPK) properties of the compounds. 2- Single-species AS was performed to estimate the single-species AS exponent distributions and their corresponding central tendencies. The correlation between the estimated single-species AS exponents and the physicochemical and DMPK properties of the compounds were also examined. 3- The methodologies of log-log transformation followed by linear regression (LL-LR) and direct nonlinear regression methods (NLS) with different weighting schemes on the AS power function were investigated. The central tendency and distribution of the allometric exponents were evaluated and compared across methods. Furthermore, the human CL prediction performance was evaluated among methods. Results. The estimated central tendency and distribution of AS exponents from the nonlinear MEM as well as the single-species AS approaches were consistent with literature reports. There were no significant correlations identified between the estimated AS exponents and the physicochemical or DMPK properties. The methods of LL-LR and the NLS with 1/w2 weighting (variance weighted by CL2 during the variance minimization process) results in the most similar allometric exponent with central tendency around 0.668 and provided the best human CL prediction among methods investigated. Conclusion. The knowledge gained in this work by extensive modeling and simulations contributed to a better understanding of the variability in AS exponents and better practice in performing AS in human CL prediction

Lopinavir (LPV) is a potent protease inhibitor specific for HIV-1. However, LPV has poor placental penetration due to substrate activity for efflux transporter by P-glycoprotein (P-gp). Since fatty acid transporters are highly expressed in the placenta during pregnancy, we designed fatty acid ester prodrug of lopinavir as substrates of fatty acid transporter in order to improve their uptake into placenta. Seven dicarboxylic acid esters of lopinavir have been made in our lab. The structures were characterized by 1H-NMR, 13C-NMR, LC-MS/MS, HRMS, IR and melting points. After making the prodrugs, an LC-MS/MS method with high specificity and sensitivity, as well as simultaneous quantitative analyses of lopinavir and SLPV, GLPV and DLPV in the BeWo cells methanol extraction was established and validated. The uptake of prodrugs (SLPV, GLPV and DLPV) in the BeWo cells was then determined. GLPV has the highest uptake followed by SLPV and then DLPV. The results suggest that the carbon length of the promoiety may have a positive relationship with the uptake. Ideal prodrugs should be stable before they reach placenta and can be hydrolyzed in the placenta and/or in fetal plasma. We did a series of stability and hydrolysis studies in human tissue fractions. The results showed that GLPV and SLPV were very stable in HIC, HLC and human adult plasma. DLPV was stable in HIC, HLC, but can be hydrolyzed in human adult plasma. GLPV and SLPV cannot be hydrolyzed in either human placenta or fetal plasma, while DLPV can be hydrolyzed in both human placenta and fetal plasma. Anti-HIV activities study of prodrugs was also conducted. The results showed that the EC50 of three prodrugs (GLPV, SLPV and DLPV) are 0.86 μM, 0.84 μM and 0.05 μM, which are much lower than 50 μM (The active drug criteria for this assay). It suggests that prodrugs have apparently anti-HIV activity. DLPV has comparable apparent anti-HIV activity to LPV (<0.02 μM). After incubation with CEM-SS cells for 6 days, almost half of DLPV was hydrolyzed into LPV. Therefore, the high anti-HIV potent of DLPV may be due to the anti-HIV activity of generated LPV.

Background: Despite the numerous pharmaceutical applications of alcohols andglycols, the interspecies differences with respect to their pharmacokinetics (PK) arepoorly understood. The aim of this research was to use in-vivo and in-vitro approaches to compare and model the PK characteristics across various species.Methods: Appropriate published in-vivo studies (in different species) foralcohols and glycols were carefully selected. PK analysis was performed using (a) noncompartmental analysis and (b) compartmental modeling to estimate relevant dose-independent PK parameters. Next, six alcohols (methanol, ethanol, 1 -propanol, 1 -butanol, 1-hexanol, and 1-octanol), two glycols (ethylene glycol and propylene glycol)and one secondary alcohol (2-propanol) were examined as in-vitro substrates for equine ADH using a UV spectrophotometric assay to evaluate the effect of molecular structure. Furthermore, in-vitro metabolism of ethanol and propylene glycol was also characterized in hepatic cytosolic fractions from rat, rabbit, dog and human and in-vitro in-vivo correlation for the hepatic disposition parameters was assessed. Finally, allometric scaling relationships for ethanol and propylene glycol PK parameters (in-vivo and in-vitro) were developed and validated.Results: Alcohols and glycols exhibited nonlinear PK due to saturable hepaticmetabolism in all species. The reported in-vivo data were well described by oneltwo compartment PK models with parallel saturable metabolism and first-order renalexcretion. In-vitro equine ADH experiments revealed differences in affinity andturnover between the substrates: Enzyme affinity (1/Km) and in-vitro intrinsic clearance (CLintin-vitro) correlated positively with logP values; glycols showed lower CLintin-vitro values than straight-chain alcohols. In-vitro hepatic cytosol studies yielded acceptable in-vivo predictions for the metabolic clearance (CLmet) of ethanol and propylene glycol in the rat, dog and human, but not the rabbit. Vdss, Vmax, CLintin-vitro, CLmet scaled allometrically across species with similar powers for both ethanol and propylene glycol, and good agreement between in-vivo and in-vitro scaling was noted. The allometric scaling models gave excellent predictions when externally validated against in-vivo concentration-time data. Conclusions: The present research demonstrates .the successful application of amodeling-based approach to elucidate interspecies relationships for alcohols andglycols, compounds which exhibit nonlinear PK and mainly low hepatic extractionbehavior. The in-vitro experimental systems have been used successfully forcharacterizing alcohol/glycol metabolism and predicting in-vivo disposition.

Problem: There are a significant proportion of patients taking acetylcholinesterase inhibitors (ChEi) for cognitive dysfunction also taking medications with anticholinergic (ACh) properties that may counteract their effects. As the number of ACh medications, burden, increases so does the likelihood of an adverse outcome.Background: ACh medications are frequently used in the elderly population (Carnahan 2004) even those with dementia or AD (Roe et al., 2002; Giron et al., 2001; Altavela 2003; Gill et al., 2005; Kogut et al., 2005). Methods: Hospitalized patients > 65 years of age with dementia (AD, other dementias, or with inferred dementia based on ChEi or NMDA antagonist medication use) were studied using UHC clinical database. This document was created in Microsoft Word 2000. Results: Dementia patients on ChEi therapy were more likely to receive an ACh (chi-square 70.1, df=l, pConclusion: A person's age and mental health status along with their current drug regimen, such as ChEi therapy, need to be closely and carefully considered before deciding to use unnecessary ACh drugs in this population which can have detrimental effects.

Background: Intra-amniotic infections (IAIs) are common complications of labor and delivery. If inadequately treated, these infections can lead to significant morbidity and mortality in the mother and the fetus. Intrapartum aminoglycoside (AG) administration is recommended for the management of IAIs. AGs are known to cross the placenta and achieve bactericidal concentrations in fetal serum. However, the highest and most persistent fetal levels are achieved in renal tissue. So, the fetus may be vulnerable to the nephrotoxic effects of AGs. Megalin, a 600 kDaendocytic receptor, is responsible for the uptake of AGs into renal proximal tubular epithelial cells. This receptor is also expressed in human term placenta and it is reasonable to speculate that it is similarly involved in the placental transport of AGs. However, the mechanisms responsible for placental AG uptake and transport have not yet been characterized. Objective: To evaluate the role of megalin in the transport of AGs across human placenta. Specific aims: (1) To assess and compare megalin expression in term and preterm placental villous tissue, and (2) assess the functional activity of megalin in in vitro placental models. Methods: (1) Following IRB approval, placental tissue samples were collected from pregnant women undergoing term or preterm deliveries. Placental villous tissueswere used to quantify megalin expression by western blotting and q-PCR (2) The human choriocarcinoma cell line (BeWo cells) were grown on Transwell plates, and then megalin expression and function were assessed. Results: Megalin protein and mRNA expression were confirmed in samples of human placental villous tissues. Megalin mRNA expression declined steeply with gestational age till week 31 of gestation then it plateaued thereafter. Also, the expression in the early preterm (n=2) was six fold higher than that of both late preterm (n=3) and term placenta (n=10) (p<0.05). The uptake of 3H-gentamicin by the BeWo cells was time-dependent, saturable (Vmax=42.9 ± 4.9 nmol/mg protein/min; Km=2.93±0.68mM) and partially inhibited by megalin inhibitors. Conclusion: Megalin is expressed in human placental villous tissues as well as the BeWo cells. When grown on Transwell® plates, the BeWo cells appear to be the most appropriate model to study the in vitro transport of AGs across the apical membrane. Time, temperature and concentration dependence of gentamicin uptake in the BeWo cells indicate protein-mediated transport. The inhibition data are consistent with megalin-mediated endocytosis of AGs.

The kinetics and mechanism of accumulation for liposomal ciprofloxacin (Lipo-CPFX) into the rat alveolar macrophage NR8383 cells were studied in vitro, in comparison to unformulated ciprofloxacin (CPFX). Upon incubation with CPFX or Lipo-CPFX, cellular drug accumulation was determined from the cell lysates or efflux was from the extracellular media by fluorescence-HPLC. The accumulation for Lipo-CPFX reached the asymptotic values at ≥ 2 hours, which was a result of uptake and efflux. The uptake appeared to be due to liposomes, mediated via cellular energy-independent mechanism like lipid fusion. In contrast, the efflux appeared to be due to ciprofloxacin, partly cellular energy-dependent, and involve probenecid-sensitive multidrug resistance proteins (MRPs). Overall, Lipo-CPFX enabled greater drug accumulation into the NR8383 cells than CPFX. This logically suggests a greater potential to treat respiratory infections especially caused by bacteria resistant to phagocytic killing.

Thrombosis is one of the leading causes of mortality and morbidity that is associated with myocardial infarction, stroke and pulmonary embolism. Anti-thrombotic agents which intend to reduce the occurrence and severity of thrombosis usually target the enzymes of the coagulation cascade. FXIa, a 160 kDa homodimer is gaining popularity of late as a potential target for anti-thrombotic agents due to its relative safety. A number of inhibitors which target the active site of FXIa have been reported but to our knowledge there have been no inhibitors which act via an allosteric mechanism. The aim of this project was to screen for allosteric inhibitors of FXIa from of pool of sulfated small-molecules.These molecules were primarily designed to act as heparin mimetics; heparin being a natural anti-coagulant. These compounds were then analyzed to determine whether inhibition was via an allosteric mechanism.