Quantitative assessment of the in-vitro binding kinetics of antisickling aromatic aldehydes with hemoglobin A: A universal HPLC-UV/Vis method to quantitate Schiff-base adduct formation.

Aromatic aldehydes act as allosteric effectors of hemoglobin (AEH), forming Schiff-base adducts with the protein to increase its oxygen (O2) affinity; a desirable property in sickle cell disease (SCD) treatment, as the high-O2 affinity hemoglobin (Hb) does not polymerize and subsequently prevents erythrocytes sickling. This study reports the development, validation, and application of a weak cation-exchange HPLC assay - quantifying the appearance of Hb-AEH adduct - as a "universal" method, allowing for the prioritization of AEH candidates through an understanding of their Hb binding affinity and kinetics. Concentration- and time-dependent Hb binding profiles of ten AEHs were determined with HPLC, followed by the appropriate non-linear modeling to characterize their steady-state binding affinity (KDss), and binding kinetics second-order association (kon) and first-order dissociation (koff) rate constants. Vanillin-derived AEHs exhibited enhanced binding affinity to Hb, primarily due to their faster kon. Across AEH, kon and koff values are strongly correlated (r = 0.993, n = 7), suggesting that modifications of the AEH scaffold enhanced their interactions with Hb as intended, but inadvertently increased their Hb-AEH adduct dissociation. To our knowledge, the present study is the first to provide valuable insight into Hb binding kinetics of antisickling aromatic aldehydes, and the assay will be a useful platform in screening/prioritizing drug candidates for SCD treatment.

Identifying a patient-centered outcome measure for a comparative effectiveness treatment trial in myasthenia gravis.

INTRODUCTION/AIMS: Data regarding the comparative effectiveness of myasthenia gravis (MG) treatments is not available. We used patient input to identify a patient-centered outcome measure (PCOM) for PROMISE-MG, a comparative effectiveness trial of MG treatments. METHODS: First, a questionnaire survey was administered to 58 people with MG at the patient meeting of the Myasthenia Gravis Foundation of America (MGFA), evaluating the impact of MG-related symptoms and MG treatments on patients' lives. Second, an online focus group of 13 patients with MG was conducted. Third, a potential outcome measure was selected. Fourth, the selected PCOM was evaluated by patients to assess how completely and accurately it captured their experiences with MG. RESULTS: The patient survey showed that limb weakness had the most impact on patients' lives. Weight gain, mood swings, insomnia, and diarrhea were the most bothersome treatment side effects. Avoiding hospitalization was very important. Focus group participants reported fatigue as one of the most bothersome symptoms and differentiated it from myasthenic weakness. They defined an ideal treatment as having minimal or no side effects and an 80% improvement in symptoms. DISCUSSION: Based on patient input, the 15-item Myasthenia Gravis Quality of Life-Revised (MG-QOL15R) scale, a validated patient-reported outcome measure (PRO), was selected as the primary PCOM for PROMISE-MG. Avoiding hospitalization and having minimal to no treatment adverse effects were selected as additional outcome measures. The patient-centeredness of a PRO depends on the context of a study: PROs should be evaluated for appropriateness as a PCOM for every study.

VZHE-039, a novel antisickling agent that prevents erythrocyte sickling under both hypoxic and anoxic conditions.

Sickle cell disease (SCD) results from a hemoglobin (Hb) mutation ßGlu6 → ßVal6 that changes normal Hb (HbA) into sickle Hb (HbS). Under hypoxia, HbS polymerizes into rigid fibers, causing red blood cells (RBCs) to sickle; leading to numerous adverse pathological effects. The RBC sickling is made worse by the low oxygen (O2) affinity of HbS, due to elevated intra-RBC concentrations of the natural Hb effector, 2,3-diphosphoglycerate. This has prompted the development of Hb modifiers, such as aromatic aldehydes, with the intent of increasing Hb affinity for O2 with subsequent prevention of RBC sickling. One such molecule, Voxelotor was recently approved by U.S. FDA to treat SCD. Here we report results of a novel aromatic aldehyde, VZHE-039, that mimics both the O2-dependent and O2-independent antisickling properties of fetal hemoglobin. The latter mechanism of action-as elucidated through crystallographic and biological studies-is likely due to disruption of key intermolecular contacts necessary for stable HbS polymer formation. This dual antisickling mechanism, in addition to VZHE-039 metabolic stability, has translated into significantly enhanced and sustained pharmacologic activities. Finally, VZHE-039 showed no significant inhibition of several CYPs, demonstrated efficient RBC partitioning and high membrane permeability, and is not an efflux transporter (P-gp) substrate.

Exploration of Structure-Activity Relationship of Aromatic Aldehydes Bearing Pyridinylmethoxy-Methyl Esters as Novel Antisickling Agents.

Aromatic aldehydes elicit their antisickling effects primarily by increasing the affinity of hemoglobin (Hb) for oxygen (O2). However, challenges related to weak potency and poor pharmacokinetic properties have hampered their development to treat sickle cell disease (SCD). Herein, we report our efforts to enhance the pharmacological profile of our previously reported compounds. These compounds showed enhanced effects on Hb modification, Hb-O2 affinity, and sickling inhibition, with sustained pharmacological effects in vitro. Importantly, some compounds exhibited unusually high antisickling activity despite moderate effects on the Hb-O2 affinity, which we attribute to an O2-independent antisickling activity, in addition to the O2-dependent activity. Structural studies are consistent with our hypothesis, which revealed the compounds interacting strongly with the polymer-stabilizing αF-helix could potentially weaken the polymer. In vivo studies with wild-type mice demonstrated significant pharmacologic effects. Our structure-based efforts have identified promising leads to be developed as novel therapeutic agents for SCD.

Pharmacokinetic profile analyses for inhaled drugs in humans using the lung delivery and disposition model.

The kinetic clarification of lung disposition for inhaled drugs in humans via pharmacokinetic (PK) modeling aids in their development and regulation for systemic and local delivery, but remains challenging due to its multiplex nature. This study exercised our lung delivery and disposition kinetic model to derive the kinetic descriptors for the lung disposition of four drugs [calcitonin, tobramycin, ciprofloxacin and fluticasone propionate (FP)] inhaled via different inhalers from the published PK profile data. With the drug dose delivered to the lung (DTL) estimated from the corresponding γ-scintigraphy or in vivo predictive cascade impactor data, the model-based curve-fitting and statistical moment analyses derived the rate constants of lung absorption (ka ) and non-absorptive disposition (knad ). The ka values differed substantially between the drugs (0.05-1.00 h-1 ), but conformed to the lung partition-based membrane diffusion except for FP, and were inhaler/delivery/deposition-independent. The knad values also varied widely (0.03-2.32 h-1 ), yet appeared to be explained by the presence or absence of non-absorptive disposition in the lung via mucociliary clearance, local tissue degradation, binding/sequestration and/or phagocytosis, and to be sensitive to differences in lung deposition. For FP, its ka value of 0.2 h-1 was unusually low, suggesting solubility/dissolution-limited slow lung absorption, but was comparable between two inhaler products. Thus, the difference in the PK profile was attributed to differences in the DTL and the knad value, the latter likely originating from different aerosol sizes and regional deposition in the lung. Overall, this empirical, rather simpler model-based analysis provided a quantitative kinetic understanding of lung absorption and non-absorptive disposition for four inhaled drugs from PK profiles in humans.

Discovery of Second-Generation NLRP3 Inflammasome Inhibitors: Design, Synthesis, and Biological Characterization.

NLRP3 inflammasomes have recently emerged as an attractive drug target for neurodegenerative disorders. In our continuing studies, a new chemical scaffold was designed as selective inhibitors of NLRP3 inflammasomes. Initial characterization of the lead HL16 demonstrated improved, however, nonselective inhibition on the NLRP3 inflammasome. Structure-activity relationship studies of HL16 identified a new lead, 17 (YQ128), with an IC50 of 0.30 ± 0.01 µM. Further studies from in vitro and in vivo models confirmed its selective inhibition on the NLRP3 inflammasome and its brain penetration. Furthermore, pharmacokinetic studies in rats at 20 mg/kg indicated extensive systemic clearance and tissue distribution, leading to a half-life of 6.6 h. However, the oral bioavailability is estimated to be only 10%, which may reflect limited GI permeability and possibly high first-pass effects. Collectively, these findings strongly encourage development of more potent analogues with improved pharmacokinetic properties from this new chemical scaffold.

Use of generally recognized as safe or dietary compounds to inhibit buprenorphine metabolism: potential to improve buprenorphine oral bioavailability.

The present study evaluated the potential of five generally recognized as safe (GRAS) or dietary compounds (α-mangostin, chrysin, ginger extract, pterostilbene and silybin) to inhibit oxidative (CYP) and conjugative (UGT) metabolism using pooled human intestinal and liver microsomes. Buprenorphine was chosen as the model substrate as it is extensively metabolized by CYPs to norbuprenorphine and by UGTs to buprenorphine glucuronide. Chrysin, ginger extract, α-mangostin, pterostilbene and silybin were tested for their inhibition of the formation of norbuprenorphine or buprenorphine glucuronide in both intestinal and liver microsomes. Pterostilbene was the most potent inhibitor of norbuprenorphine formation in both intestinal and liver microsomes, with IC50 values of 1.3 and 0.8 µM, respectively, while α-mangostin and silybin most potently inhibited buprenorphine glucuronide formation. The equipotent combination of pterostilbene and ginger extract additively inhibited both pathways in intestinal microsomes. Since pterostilbene and ginger extract showed potent CYP and/or UGT inhibition of buprenorphine metabolism, their equipotent combination was tested to assess the presence of synergistic inhibition. However, because the combination showed additive inhibition, it was not used while performing IVIVE analysis. Based on quantitative in vitro-in vivo extrapolation, pterostilbene (21 mg oral dose) appeared to be most effective in improving the mean predicted Foral and AUC∞ PO of buprenorphine from 3 ± 2% and 340 ± 330 ng*min/ml to 75 ± 8% and 36,000 ± 25,000 ng*min/ml, respectively. At a 10-fold lower dose of pterostilbene, the predicted buprenorphine Foral approximated sublingual bioavailability (~35%) and showed a 2-4 fold reduction in the variability around the predicted AUC∞ PO of buprenorphine. These results demonstrate the feasibility of using various GRAS/dietary compounds to inhibit substantially the metabolism by CYP and UGT enzymes to achieve higher and less variable oral bioavailability. This inhibitor strategy may be useful for drugs suffering from low and variable oral bioavailability due to extensive presystemic oxidative and/or conjugative metabolism.

Rational modification of vanillin derivatives to stereospecifically destabilize sickle hemoglobin polymer formation.

Increasing the affinity of hemoglobin for oxygen represents a feasible and promising therapeutic approach for sickle cell disease by mitigating the primary pathophysiological event, i.e. the hypoxia-induced polymerization of sickle hemoglobin (Hb S) and the concomitant erythrocyte sickling. Investigations on a novel synthetic antisickling agent, SAJ-310, with improved and sustained antisickling activity have previously been reported. To further enhance the biological effects of SAJ-310, a structure-based approach was employed to modify this compound to specifically inhibit Hb S polymer formation through interactions which perturb the Hb S polymer-stabilizing αF-helix, in addition to primarily increasing the oxygen affinity of hemoglobin. Three compounds, TD-7, TD-8 and TD-9, were synthesized and studied for their interactions with hemoglobin at the atomic level, as well as their functional and antisickling activities in vitro. X-ray crystallographic studies with liganded hemoglobin in complex with TD-7 showed the predicted mode of binding, although the interaction with the αF-helix was not as strong as expected. These findings provide important insights and guidance towards the development of molecules that would be expected to bind and make stronger interactions with the αF-helix, resulting in more efficacious novel therapeutics for sickle cell disease.

Rational design of pyridyl derivatives of vanillin for the treatment of sickle cell disease.

Hypoxia-induced polymerization of sickle hemoglobin (Hb S) is the principal phenomenon that underlays the pathophysiology and morbidity associated with sickle cell disease (SCD). Opportunely, as an allosteric protein, hemoglobin (Hb) serves as a convenient and potentially critical druggable target. Consequently, molecules that prevent Hb S polymerization (Hb modifiers), and the associated erythrocyte sickling have been investigated-and retain significant interest-as a viable therapeutic strategy for SCD. This group of molecules, including aromatic aldehydes, form high oxygen affinity Schiff-base adducts with Hb S, which are resistant to polymerization. Here, we report the design and synthesis of novel potent antisickling agents (SAJ-009, SAJ-310 and SAJ-270) based on the pharmacophore of vanillin and INN-312, a previously reported pyridyl derivative of vanillin. These novel derivatives exhibited superior in vitro binding and pharmacokinetic properties compared to vanillin, which translated into significantly enhanced allosteric and antisickling properties. Crystal structure studies of liganded Hb in the R2 quaternary state in complex with SAJ-310 provided important insights into the allosteric and antisickling properties of this group of compounds. While these derivatives generally show similar in vitro biological potency, significant structure-dependent differences in their biochemical profiles would help predict the most promising candidates for successful in vivo pre-clinical translational studies and inform further structural modifications to improve on their pharmacologic properties.

Design, Synthesis, and Biological Evaluation of Ester and Ether Derivatives of Antisickling Agent 5-HMF for the Treatment of Sickle Cell Disease.

Candidate drugs to counter intracellular polymerization of deoxygenated sickle hemoglobin (Hb S) continue to represent a promising approach to mitigating the primary cause of the pathophysiology associated with sickle cell disease (SCD). One such compound is the naturally occurring antisickling agent, 5-hydroxymethyl-2-furfural (5-HMF), which has been studied in the clinic for the treatment of SCD. As part of our efforts to develop novel efficacious drugs with improved pharmacologic properties, we structurally modified 5-HMF into 12 ether and ester derivatives. The choice of 5-HMF as a pharmacophore was influenced by a combination of its demonstrated attractive hemoglobin modifying and antisickling properties, well-known safety profiles, and its reported nontoxic major metabolites. The derivatives were investigated for their time- and/or dose-dependent effects on important antisickling parameters, such as modification of hemoglobin, corresponding changes in oxygen affinity, and inhibition of red blood cell sickling. The novel test compounds bound and modified Hb and concomitantly increased the protein affinity for oxygen. Five of the derivatives exhibited 1.5- to 4.0-fold higher antisickling effects than 5-HMF. The binding mode of the compounds with Hb was confirmed by X-ray crystallography and, in part, helps explain their observed biochemical properties. Our findings, in addition to the potential therapeutic application, provide valuable insights and potential guidance for further modifications of these (and similar) compounds to enhance their pharmacologic properties.

Phase 1 study evaluating the safety and pharmacokinetics of pralatrexate in relapsed/refractory advanced solid tumors and lymphoma patients with mild, moderate, and severe renal impairment.

PURPOSE: Pralatrexate is a folate analogue indicated for the treatment of relapsed or refractory peripheral T-cell lymphoma. It has not been formally tested in patients with renal impairment. This study evaluated the pharmacokinetic (PK) profile of pralatrexate in patients with renal impairment and with relapsed/refractory advanced solid tumors and lymphoma. METHODS: This was an open-label, nonrandomized, phase 1 study. Eligible patients received pralatrexate administered as an IV push over 3-5 min once weekly for 6 weeks in 7-week cycles until progressive disease or intolerable toxicity. Four cohorts of 6 patients were planned for a total of 24 patients. Patients with normal renal function (Cohort A), mild (Cohort B), and moderate renal impairment (Cohort C) received 30 mg/m2 pralatrexate once weekly for 6 weeks in 7-week cycles, and patients with severe renal impairment (Cohort D) were to be administered 20 mg/m2 once weekly for 6 weeks. Plasma and urine samples were collected at pre-specified time points to determine the PK profile of pralatrexate in each treatment cohort. Patients were followed for safety and tolerability. RESULTS: A total of 29 patients were enrolled and 27 patients (14 male) received at least 1 dose of pralatrexate. Because of a qualifying toxicity in Cohort C, the starting dose for Cohort D was reduced to 15 mg/m2. Chronic renal impairment led to a decrease in renal clearance of the pralatrexate diastereomers, PDX-10a and PDX-10b, but systemic exposure to these diastereomers was not dramatically affected by renal impairment. Pralatrexate exposure in Cohort D (15 mg/m2) was similar to the exposure in other cohorts (30 mg/m2). No apparent difference in toxicity between the four treatment cohorts was observed, except for an increase in cytopenias in patients with severe renal impairment. CONCLUSION: Pralatrexate exposure, at a dose of 30 mg/m2, in patients with mild or moderate renal impairment was similar to the exposure in patients with normal renal function. For patients with severe renal impairment only, a pralatrexate dose of 15 mg/m2 is recommended.

In Vitro Tests for Aerosol Deposition. IV: Simulating Variations in Human Breath Profiles for Realistic DPI Testing.

BACKGROUND: The amount of drug aerosol from an inhaler that can pass through an in vitro model of the mouth and throat (MT) during a realistic breath or inhalation flow rate vs. time profile (IP) is designated the total lung dose in vitro, or TLDin vitro. This article describes a clinical study that enabled us to recommend a general method of selecting IPs for use with powder inhalers of known airflow resistance (R) provided subjects followed written instructions either alone or in combination with formal training. METHODS: In a drug-free clinical trial, inhaler-naïve, nonsmoking healthy adult human volunteers were screened for normal pulmonary function. IPs were collected from each volunteer inhaling through different air flow resistances after different levels of training. IPs were analyzed to determine the distribution of inhalation variables across the population and their dependence on training and airflow resistance. RESULTS: Equations for IP simulation are presented that describe the data including confidence limits at each resistance and training condition. Realistic IPs at upper (90%), median (50%), and lower (10%) confidence limits were functions of R and training. Peak inspiratory flow rates (PIFR) were inversely proportional to R so that if R was assigned, values for PIFR could be calculated. The time of PIFR, TPIFR, and the total inhaled volume (V) were unrelated to R, but dependent on training. Once R was assigned for a powder inhaler to be tested, a range of simulated IPs could be generated for the different training scenarios. Values for flow rate acceleration and depth of inspiration could also be varied within the population limits of TPIFR and V. CONCLUSIONS: The use of simulated IPs, in concert with realistic in vitro testing, should improve the DPI design process and the confidence with which clinical testing may be initiated for a chosen device.

Design, Synthesis, and Investigation of Novel Nitric Oxide (NO)-Releasing Prodrugs as Drug Candidates for the Treatment of Ischemic Disorders: Insights into NO-Releasing Prodrug Biotransformation and Hemoglobin-NO Biochemistry.

We have developed novel nitric oxide (NO)-releasing prodrugs of efaproxiral (RSR13) for their potential therapeutic applications in a variety of diseases with underlying ischemia. RSR13 is an allosteric effector of hemoglobin (Hb) that decreases the protein's affinity for oxygen, thereby increasing tissue oxygenation. NO, because of its vasodilatory property, in the form of ester prodrugs has been found to be useful in managing several cardiovascular diseases by increasing blood flow and oxygenation in ischemic tissues. We synthesized three NO-donor ester derivatives of RSR13 (DD-1, DD-2, and DD-3) by attaching the NO-releasing moieties nitrooxyethyl, nitrooxypropyl, and 1-(pyrrolidin-1-yl)diazen-1-ium-1,2-diolate, respectively, to the carboxylate of RSR13. In vitro studies demonstrated that the compounds released NO in a time-dependent manner upon being incubated with l-cysteine (1.8-9.3%) or human serum (2.3-52.5%) and also reduced the affinity of Hb for oxygen in whole blood (ΔP50 of 4.9-21.7 mmHg vs ΔP50 of 25.4-32.1 mmHg for RSR13). Crystallographic studies showed RSR13, the hydrolysis product of the reaction between DD-1 and deoxygenated Hb, bound to the central water cavity of Hb. Also, the hydrolysis product, NO, was observed exclusively bound to the two α hemes, the first such HbNO structure to be reported, capturing the previously proposed physiological bis-ligated nitrosylHb species. Finally, nitrate was observed bound to ßHis97. Ultraperformance liquid chromatography-mass spectrometry analysis of the compounds incubated with matrices used for the various studies demonstrated the presence of the predicted reaction products. Our findings, beyond the potential therapeutic application, provide valuable insights into the biotransformation of NO-releasing prodrugs and their mechanism of action and into hemoglobin-NO biochemistry at the molecular level.

Effect of uptake transporters OAT3 and OATP1B1 and efflux transporter MRP2 on the pharmacokinetics of eluxadoline.

The effects of OATP1B1, OAT3, and MRP2 on the pharmacokinetics of eluxadoline, an oral, locally active, opioid receptor agonist/antagonist being developed for treatment of IBS-d were assessed in vivo. Coadministration of a single 200 mg dose of eluxadoline with cyclosporine, and probenecid increased eluxadoline systemic exposure [AUC(0-inf) ] by 4.4- and 1.4-fold, respectively, whereas peak exposure (Cmax ) increased 6.2-fold and 1.3-fold, respectively. Cyclosporine had little effect on renal clearance (CLren ) of eluxadoline whereas probenecid reduced CLren by nearly 50%. These study results suggested that sinusoidal OATP1B1-mediated hepatic uptake of eluxadoline (during first-pass and systemic extraction) plays a major role in its absorption and disposition, whereas OAT3-mediated basolateral uptake in the proximal renal tubules and MRP2-mediated canalicular and renal tubular apical efflux play only minor roles in its overall disposition. All treatments were safe and well tolerated.

Cumulative organic anion transporter-mediated drug-drug interaction potential of multiple components in salvia miltiorrhiza (danshen) preparations.

PURPOSE: To evaluate organic anion transporter-mediated drug-drug interaction (DDI) potential for individual active components of Danshen (Salvia miltiorrhiza) vs. combinations using in vitro and in silico approaches. METHODS: Inhibition profiles for single Danshen components and combinations were generated in stably-expressing human (h)OAT1 and hOAT3 cells. Plasma concentration-time profiles for compounds were estimated from in vivo human data using an i.v. two-compartment model (with first-order elimination). The cumulative DDI index was proposed as an indicator of DDI potential for combination products. This index was used to evaluate the DDI potential for Danshen injectables from 16 different manufacturers and 14 different lots from a single manufacturer. RESULTS: The cumulative DDI index predicted in vivo inhibition potentials, 82% (hOAT1) and 74% (hOAT3), comparable with those observed in vitro, 72 ± 7% (hOAT1) and 81 ± 10% (hOAT3), for Danshen component combinations. Using simulated unbound Cmax values, a wide range in cumulative DDI index between manufacturers, and between lots, was predicted. Many products exhibited a cumulative DDI index > 1 (50% inhibition). CONCLUSIONS: Danshen injectables will likely exhibit strong potential to inhibit hOAT1 and hOAT3 function in vivo. The proposed cumulative DDI index might improve prediction of DDI potential of herbal medicines or pharmaceutical preparations containing multiple components.

Human organic cation transporters 1 (SLC22A1), 2 (SLC22A2), and 3 (SLC22A3) as disposition pathways for fluoroquinolone antimicrobials.

Fluoroquinolones (FQs) are important antimicrobials that exhibit activity against a wide range of bacterial pathogens and excellent tissue permeation. They exist as charged molecules in biological fluids, and thus, their disposition depends heavily on active transport and facilitative diffusion. A recent review of the clinical literature indicated that tubular secretion and reabsorption are major determinants of their half-life in plasma, efficacy, and drug-drug interactions. In particular, reported in vivo interactions between FQs and cationic drugs affecting renal clearance implicated organic cation transporters (OCTs). In this study, 13 FQs, ciprofloxacin, enoxacin, fleroxacin, gatifloxacin, levofloxacin, lomefloxacin, moxifloxacin, norfloxacin, ofloxacin, pefloxacin, prulifloxacin, rufloxacin, and sparfloxacin, were screened for their ability to inhibit transport activity of human OCT1 (hOCT1) (SLC22A1), hOCT2 (SLC22A2), and hOCT3 (SLC22A3). All, with the exception of enoxacin, significantly inhibited hOCT1-mediated uptake under initial test conditions. None of the FQs inhibited hOCT2, and only moxifloxacin inhibited hOCT3 (~30%), even at a 1,000-fold excess. Gatifloxacin, moxifloxacin, prulifloxacin, and sparfloxacin were determined to be competitive inhibitors of hOCT1. Inhibition constants (K(i)) were estimated to be 250 ± 18 µM, 161 ± 19 µM, 136 ± 33 µM, and 94 ± 8 µM, respectively. Moxifloxacin competitively inhibited hOCT3-mediated uptake, with a K(i) value of 1,598 ± 146 µM. Despite expression in enterocytes (luminal), hepatocytes (sinusoidal), and proximal tubule cells (basolateral), hOCT3 does not appear to contribute significantly to FQ disposition. However, hOCT1 in the sinusoidal membrane of hepatocytes, and potentially the basolateral membrane of proximal tubule cells, is likely to play a role in the disposition of these antimicrobial agents.

Fluoroquinolone disposition: identification of the contribution of renal secretory and reabsorptive drug transporters.

INTRODUCTION: Fluoroquinolones (FQs) exist as charged molecules in blood and urine making their absorption, distribution, and elimination likely to be influenced by active transport mechanisms. Greater understanding of in vivo FQ clearance mechanisms should help improve the predictability of drug-drug interactions, enhance the clinical safety and efficacy, and aid future novel drug design strategies. AREAS COVERED: The authors present an overview of FQ development and associated drug-drug interactions, followed by systematic quantitative review of the physicochemical and in vivo pharmacokinetic properties for 15 representative FQs using historical clinical literature. These results were correlated with in vitro studies implicating drug transporters in FQ clearance to link clinical and in vitro evidence supporting the contribution of drug transport mechanisms to FQ disposition. Specific transporters likely to handle FQs in human renal proximal tubule cells are also identified. EXPERT OPINION: Renal handling, that is, tubular secretion and reabsorption, appears to be the main determinant of FQ plasma half-life, clinical duration of action, and drug-drug interactions. Due to their zwitterionic nature, FQs are likely to interact with organic anion and cation transporters within the solute carrier (SLC) superfamily, including OAT1, OAT3, OCT2, OCTN1, OCTN2, MATE1, and MATE2. The ATP-binding cassette (ABC) transporters MDR1, MRP2, MRP4, and BCRP also may interact with FQs.

Clinical pharmacokinetics and drug-drug interactions of endothelin receptor antagonists in pulmonary arterial hypertension.

The authors review the basic pharmacology and potential for adverse drug-drug interactions (DDIs) of bosentan and ambrisentan, the 2 endothelin receptor antagonists currently approved for pulmonary arterial hypertension (PAH) treatment. Bosentan, an endothelin (ET) receptor-type ET(A) and ET(B) antagonist, is metabolized to active metabolites by and an inducer of cytochrome P450 (CYP)2C9 and CYP3A. Ambrisentan, a selective ET(A) receptor antagonist, is metabolized primarily by uridine 5'diphosphate glucuronosyltransferases (UGTs) 1A9S, 2B7S, and 1A3S and, to a lesser extent, by CYP3A and CYP2C19. Drug interactions observed with bosentan DDI studies have demonstrated a potential for significant clinical implications during PAH management: bosentan is contraindicated with cyclosporine A and glyburide, and additional monitoring/dose adjustments are required when coadministered with hormonal contraceptives, simvastatin, lopinavir/ritonavir, and rifampicin. As bosentan carries a boxed warning regarding risks of liver injury and showed dose-dependant increases in serum aminotransferase abnormalities, drug interactions that increase bosentan exposure are of particular clinical concern. Ambrisentan DDI studies performed to date have shown only one clinically relevant DDI, an interaction with cyclosporine A that requires ambrisentan dose reduction. As the treatment of PAH moves toward multimodal combination therapy, scrutiny should be placed on ensuring that drug combinations achieve maximal clinical benefit while minimizing side effects.

Crystallographic analysis of human hemoglobin elucidates the structural basis of the potent and dual antisickling activity of pyridyl derivatives of vanillin.

Vanillin has previously been studied clinically as an antisickling agent to treat sickle-cell disease. In vitro investigations with pyridyl derivatives of vanillin, including INN-312 and INN-298, showed as much as a 90-fold increase in antisickling activity compared with vanillin. The compounds preferentially bind to and modify sickle hemoglobin (Hb S) to increase the affinity of Hb for oxygen. INN-312 also led to a considerable increase in the solubility of deoxygenated Hb S under completely deoxygenated conditions. Crystallographic studies of normal human Hb with INN-312 and INN-298 showed that the compounds form Schiff-base adducts with the N-terminus of the α-subunits to constrain the liganded (or relaxed-state) Hb conformation relative to the unliganded (or tense-state) Hb conformation. Interestingly, while INN-298 binds and directs its meta-positioned pyridine-methoxy moiety (relative to the aldehyde moiety) further down the central water cavity of the protein, that of INN-312, which is ortho to the aldehyde, extends towards the surface of the protein. These studies suggest that these compounds may act to prevent sickling of SS cells by increasing the fraction of the soluble high-affinity Hb S and/or by stereospecific inhibition of deoxygenated Hb S polymerization.