The Isoleucine at Position 118 in Transmembrane 2 Is Responsible for the Selectivity of Xamoterol, Nebivolol, and ICI89406 for the Human ß1-Adrenoceptor.

Known off-target interactions frequently cause predictable drug side-effects (e.g., ß1-antagonists used for heart disease, risk ß2-mediated bronchospasm). Computer-aided drug design would improve if the structural basis of existing drug selectivity was understood. A mutagenesis approach determined the ligand-amino acid interactions required for ß1-selective affinity of xamoterol and nebivolol, followed by computer-based modeling to provide possible structural explanations. 3H-CGP12177 whole cell binding was conducted in Chinese hamster ovary cells stably expressing human ß1, ß2, and chimeric ß1/ß2-adrenoceptors (ARs). Single point mutations were investigated in transiently transfected cells. Modeling studies involved docking ligands into three-dimensional receptor structures and performing molecular dynamics simulations, comparing interaction frequencies between apo and holo structures of ß1 and ß2-ARs. From these observations, an ICI89406 derivative was investigated that gave further insights into selectivity. Stable cell line studies determined that transmembrane 2 was crucial for the ß1-selective affinity of xamoterol and nebivolol. Single point mutations determined that the ß1-AR isoleucine (I118) rather than the ß2 histidine (H93) explained selectivity. Studies of other ß1-ligands found I118 was important for ICI89406 selective affinity but not that for betaxolol, bisoprolol, or esmolol. Modeling studies suggested that the interaction energies and solvation of ß1-I118 and ß2-H93 are factors determining selectivity of xamoterol and ICI89406. ICI89406 without its phenyl group loses its high ß1-AR affinity, resulting in the same affinity as for the ß2-AR. The human ß1-AR residue I118 is crucial for the ß1-selective affinity of xamoterol, nebivolol, and ICI89406 but not all ß1-selective compounds. SIGNIFICANCE STATEMENT: Some ligands have selective binding affinity for the human ß1 versus the ß2-adrenoceptor; however, the molecular/structural reason for this is not known. The transmembrane 2 residue isoleucine I118 is responsible for the selective ß1-binding of xamoterol, nebivolol, and ICI89406 but does not explain the selective ß1-binding of betaxolol, bisoprolol, or esmolol. Understanding the structural basis of selectivity is important to improve computer-aided ligand design, and targeting I118 in ß1-adrenoceptors is likely to increase ß1-selectivity of drugs.

Molecular determinants of pro-arrhythmia proclivity of d- and l-sotalol via a multi-scale modeling pipeline.

Drug isomers may differ in their proarrhythmia risk. An interesting example is the drug sotalol, an antiarrhythmic drug comprising d- and l- enantiomers that both block the hERG cardiac potassium channel and confer differing degrees of proarrhythmic risk. We developed a multi-scale in silico pipeline focusing on hERG channel - drug interactions and used it to probe and predict the mechanisms of pro-arrhythmia risks of the two enantiomers of sotalol. Molecular dynamics (MD) simulations predicted comparable hERG channel binding affinities for d- and l-sotalol, which were validated with electrophysiology experiments. MD derived thermodynamic and kinetic parameters were used to build multi-scale functional computational models of cardiac electrophysiology at the cell and tissue scales. Functional models were used to predict inactivated state binding affinities to recapitulate electrocardiogram (ECG) QT interval prolongation observed in clinical data. Our study demonstrates how modeling and simulation can be applied to predict drug effects from the atom to the rhythm for dl-sotalol and also increased proarrhythmia proclivity of d- vs. l-sotalol when accounting for stereospecific beta-adrenergic receptor blocking.

Unique Positive Cooperativity Between the ß-Arrestin-Biased ß-Blocker Carvedilol and a Small Molecule Positive Allosteric Modulator of the ß2-Adrenergic Receptor.

Among ß-blockers that are clinically prescribed for heart failure, carvedilol is a first-choice agent with unique pharmacological properties. Carvedilol is distinct from other ß-blockers in its ability to elicit ß-arrestin-biased agonism, which has been suggested to underlie its cardioprotective effects. Augmenting the pharmacologic properties of carvedilol thus holds the promise of developing more efficacious and/or biased ß-blockers. We recently identified compound-6 (cmpd-6), the first small molecule positive allosteric modulator of the ß2-adrenergic receptor (ß2AR). Cmpd-6 is positively cooperative with orthosteric agonists at the ß2AR and enhances agonist-mediated transducer (G-protein and ß-arrestin) signaling in an unbiased manner. Here, we report that cmpd-6, quite unexpectedly, displays strong positive cooperativity only with carvedilol among a panel of structurally diverse ß-blockers. Cmpd-6 enhances the binding affinity of carvedilol for the ß2AR and augments its ability to competitively antagonize agonist-induced cAMP generation. Cmpd-6 potentiates ß-arrestin1- but not Gs-protein-mediated high-affinity binding of carvedilol at the ß2AR and ß-arrestin-mediated cellular functions in response to carvedilol including extracellular signal-regulated kinase phosphorylation, receptor endocytosis, and trafficking into lysosomes. Importantly, an analog of cmpd-6 that selectively retains positive cooperativity with carvedilol acts as a negative modulator of agonist-stimulated ß2AR signaling. These unprecedented cooperative properties of carvedilol and cmpd-6 have implications for fundamental understanding of G-protein-coupled receptor (GPCR) allosteric modulation, as well as for the development of more effective biased beta blockers and other GPCR therapeutics. SIGNIFICANCE STATEMENT: This study reports on the small molecule-mediated allosteric modulation of the ß-arrestin-biased ß-blocker, carvedilol. The small molecule, compound-6 (cmpd-6), displays an exclusive positive cooperativity with carvedilol among other ß-blockers and enhances the binding affinity of carvedilol for the ß2-adrenergic receptor. Cooperative effects of cmpd-6 augment the ß-blockade property of carvedilol while potentiating its ß-arrestin-mediated signaling functions. These findings have potential implications in advancing G-protein-coupled receptor allostery, developing biased therapeutics and remedying cardiovascular ailments.

ß-Blocker Use and Risk of Mortality in Heart Failure Patients Initiating Maintenance Dialysis.

RATIONAL & OBJECTIVE: Beta-blockers are recommended for patients with heart failure (HF) but their benefit in the dialysis population is uncertain. Beta-blockers are heterogeneous, including with respect to their removal by hemodialysis. We sought to evaluate whether ß-blocker use and their dialyzability characteristics were associated with early mortality among patients with chronic kidney disease with HF who transitioned to dialysis. STUDY DESIGN: Retrospective cohort study. SETTING & PARTICIPANTS: Adults patients with chronic kidney disease (aged≥18 years) and HF who initiated either hemodialysis or peritoneal dialysis during January 1, 2007, to June 30, 2016, within an integrated health system were included. EXPOSURES: Patients were considered treated with ß-blockers if they had a quantity of drug dispensed covering the dialysis transition date. OUTCOMES: All-cause mortality within 6 months and 1 year or hospitalization within 6 months after transition to maintenance dialysis. ANALYTICAL APPROACH: Inverse probability of treatment weights using propensity scores was used to balance covariates between treatment groups. Cox proportional hazard analysis and logistic regression were used to investigate the association between ß-blocker use and study outcomes. RESULTS: 3,503 patients were included in the study. There were 2,115 (60.4%) patients using ß-blockers at transition. Compared with nonusers, the HR for all-cause mortality within 6 months was 0.79 (95% CI, 0.65-0.94) among users of any ß-blocker and 0.68 (95% CI, 0.53-0.88) among users of metoprolol at transition. There were no observed differences in all-cause or cardiovascular-related hospitalization. LIMITATIONS: The observational nature of our study could not fully account for residual confounding. CONCLUSIONS: Beta-blockers were associated with a lower rate of mortality among incident hemodialysis patients with HF. Similar associations were not observed for hospitalizations within the first 6 months following transition to dialysis.

Beta-adrenergic receptor antagonism is proinflammatory and exacerbates neuroinflammation in a mouse model of Alzheimer's Disease.

Adrenergic systems regulate both cognitive function and immune function. The primary source of adrenergic signaling in the brain is norepinephrine (NE) neurons of the locus coeruleus (LC), which are vulnerable to age-related degeneration and are one of the earliest sites of pathology and degeneration in neurodegenerative disorders such as Alzheimer's Disease (AD). Loss of adrenergic tone may potentiate neuroinflammation both in aging and neurodegenerative conditions. Importantly, beta-blockers (beta-adrenergic antagonists) are a common treatment for hypertension, co-morbid with aging, and may further exacerbate neuroinflammation associated with loss of adrenergic tone in the central nervous system (CNS). The present studies were designed to both examine proinflammatory consequences of beta-blocker administration in an acute lipopolysaccharide (LPS) model as well as to examine chronic effects of beta-blocker administration on neuroinflammation and behavior in an amyloid-beta protein precursor (APP) mouse model of AD. We provide evidence for robust potentiation of peripheral inflammation with 4 different beta-blockers in an acute model of LPS. However, beta-blockers did not potentiate CNS inflammation in this model. Notably, in this same model, the genetic knockdown of either beta1- or beta2-adrenergic receptors in microglia did potentiate CNS inflammation. Furthermore, in an APP mouse model of amyloid pathology, chronic beta-blocker administration did potentiate CNS inflammation. The beta-blocker, metoprolol, also induced markers of phagocytosis and impaired cognitive behavior in both wild-type and APP mice. Given the induction of markers of phagocytosis in vivo, we examined phagocytosis of synaptosomes in an in vitro primary microglia culture and showed that beta-blockers enhanced whereas beta-adrenergic agonists inhibited phagocytosis of synaptosomes. In conclusion, beta-blockers potentiated inflammation peripherally in a systemic model of inflammation and centrally in an amyloidosis model of neuroinflammation. Additionally, beta-blockers impaired learning and memory and modulated synaptic phagocytosis with implications for synaptic degeneration. These findings warrant further consideration of the proinflammatory consequences of chronic beta-blocker administration, which are not restricted to the periphery in patients with neurodegenerative disorders.

Presence and function of ß-adrenergic receptors in primary equine bronchial epithelia cells.

The ß-adrenergic receptor (ß-AR) plays an important role in regulating a variety of cell and organ functions in different animal species and is an important target in asthma pathogenesis and therapy. The ß-AR expression and function in equine bronchial epithelial cells (EBEC) were not known but innervation and significant decrease in receptor level were reported in the equine bronchial tissues from asthmatic horses. 125I-iodocyanopindolol (ICYP) binding studies were undertaken in primary freshly isolated and cultured EBEC to identify the presence of the ß-ARs. The receptor distribution was assessed using subtype-selective ß-AR antagonists (ICI 118 551 (ß2) and CGP 20712A (ß1). The ß-AR function was confirmed by measuring the agonist-induced intracellular cAMP accumulation in freshly isolated and cultured EBEC. In both freshly isolated and cultured EBEC, the specific ICYP binding was saturable and of high affinity. The maximal receptor density (Bmax) was 9763 ± 140 binding sites/cell (mean ± SEM, n = 7) and 10575 ± 194 binding sites/cell (mean ± SEM, n = 5) in freshly isolated and cultured EBEC, respectively. The receptor affinity to the ligand (KD) was also not different between the two cell conditions. ICI 118.551 displaced ICYP with 25 000-fold higher affinity than CGP 20712A. Moreover, in both fresh isolated and cultured EBEC, cAMP-accumulation was stimulated with a rank-order of potency of isoproterenol > adrenaline > noradrenaline. These results highlight the ß2-AR to be a key subtype in both freshly isolated and cultured primary EBEC.

Stereochemical facets of clinical ß-blockers: An overview.

The modern ß-adrenergic agonists (ß-blockers) possess one or more than one chiral center in their structure. Two enantiomers exhibit distinct pharmacodynamic and pharmacokinetic behaviors. Current progress in drug designing has resulted in the ability to understand the role of chirality in modern therapeutics. Furthermore, with a greater understanding of the molecular structure of precise drug targets, development of new drugs is directed towards the pure enantiomers instead of its racemates. The present review deals with a discussion on the stereochemical facets of chiral clinical ß-blockers. This review provides details of stereo-selectivity in the pharmacological behavior of some of ß-blockers and their metabolites. An effort has been made on highlighting the distinction between the therapeutic behavior of the racemic mixtures and pure enantiomers.

Allosteric "beta-blocker" isolated from a DNA-encoded small molecule library.

The ß2-adrenergic receptor (ß2AR) has been a model system for understanding regulatory mechanisms of G-protein-coupled receptor (GPCR) actions and plays a significant role in cardiovascular and pulmonary diseases. Because all known ß-adrenergic receptor drugs target the orthosteric binding site of the receptor, we set out to isolate allosteric ligands for this receptor by panning DNA-encoded small-molecule libraries comprising 190 million distinct compounds against purified human ß2AR. Here, we report the discovery of a small-molecule negative allosteric modulator (antagonist), compound 15 [([4-((2S)-3-(((S)-3-(3-bromophenyl)-1-(methylamino)-1-oxopropan-2-yl)amino)-2-(2-cyclohexyl-2-phenylacetamido)-3-oxopropyl)benzamide], exhibiting a unique chemotype and low micromolar affinity for the ß2AR. Binding of 15 to the receptor cooperatively enhances orthosteric inverse agonist binding while negatively modulating binding of orthosteric agonists. Studies with a specific antibody that binds to an intracellular region of the ß2AR suggest that 15 binds in proximity to the G-protein binding site on the cytosolic surface of the ß2AR. In cell-signaling studies, 15 inhibits cAMP production through the ß2AR, but not that mediated by other Gs-coupled receptors. Compound 15 also similarly inhibits ß-arrestin recruitment to the activated ß2AR. This study presents an allosteric small-molecule ligand for the ß2AR and introduces a broadly applicable method for screening DNA-encoded small-molecule libraries against purified GPCR targets. Importantly, such an approach could facilitate the discovery of GPCR drugs with tailored allosteric effects.

Effects of atorvastatin on talinolol absorption: A potential drug-drug interaction.

In this study, we aimed to determine the drug-drug interaction potential between atorvastatin (ATOR), and talinolol (TAL). Concentration-dependent effects of ATOR on the intestinal permeability of TAL were investigated by an in situ intestinal perfusion method. Dose-dependent effects of ATOR on TAL exposure were evaluated by measuring plasma concentrations after oral administration in rats. ATOR slightly changed the intestinal secretion of TAL in jejunum but not in colon. Plasma AUC levels of TAL were elevated by co-administration of ATOR at low and high doses whereas medium doses of ATOR resulted in a decrease in TAL bioavailability. However, these changes were not statistically significant. In our study, the pharmacokinetics of TAL were not affected by the concurrent use of ATOR in rats. In conclusion, it should be considered that complex interplay between the efflux and uptake transporters in the tissues and inhibition of these transporters by modulating agents may overshadow individual effects of each other.

Intractable Ocular Diseases and Treatment Progress.

In recent years, with the aging of the population and the frequent use of electronic devices, many eye diseases have shown a linear upward trend, such as dry eye disease, glaucoma, cataract, age-related macular degeneration, and diabetic retinopathy. These diseases are often chronic and difficult to cure. Based on the structure and barrier of the human eye, this review describes the pathogenesis and treatments of several intractable eye diseases and summarizes the advanced ocular drug delivery systems to provide new treatment ideas for these diseases. Finally, we also look forward to the prospect of RNAi therapy in the treatment of eye diseases.

Comprehensive MS-based screening and identification of pharmaceutical transformation products formed during enzymatic conversion.

In this study, transformation products (TPs) of diclofenac, mefenamic acid, and sotalol derived from peroxidase- and laccase-catalyzed transformations were studied with different mass spectrometry (MS)-based workflows. A straightforward pre-screening of enzymatic degradation rate was performed using a robotic nano-ESI source coupled to single quadrupole MS. Accurate mass data and information on molecular hydrophobicity were obtained from a serial coupling of reversed phase liquid chromatography (RPLC) with hydrophilic interaction liquid chromatography (HILIC) to a time-of-flight-mass spectrometer (ToF-MS). These parameters were combined with fragmentation information from product ion scan operated in enhanced mode (EPI) with precursor selection in Q3 and data from multiple reaction monitoring (MRM) modes using a hybrid triple quadrupole-linear ion trap-mass spectrometer (QqQ/LIT-MS). "Suspect" MRM modes did not provide a significant sensitivity improvement compared to EPI experiments. The complementarity of the data from different MS-based workflows allowed for an increase of identification confidence. Overall, this study demonstrated that dimerization, hydroxylation, and dehydration reactions were the predominant mechanisms found for diclofenac and mefenamic acid during enzyme-catalyzed transformation, whereas a degradation product was observed for the peroxidase-catalyzed conversion of sotalol. Results can contribute to understand enzymatic mechanisms and provide a basis for assessing risks and benefits of enzyme-based remediation. Graphical abstract ᅟ.

The effect of beta-adrenergic blockade on inflammatory and cardiovascular responses to acute mental stress.

Acute mental stress elicits increases in plasma cytokine concentrations in humans, but the underlying mechanisms remain poorly understood. We assessed the impact of beta-adrenergic blockade on plasma interleukin 6 (IL-6) and IL-1 receptor antagonist (IL-1Ra) responses in a parallel group, double-blind randomised placebo-controlled trial involving 64 healthy young adult volunteers. Participants were administered 80 mg slow-release propranolol or placebo daily for 7 days before the stress testing session in which responses to 3 behavioural challenges (public speaking, mirror tracing, mental arithmetic) were evaluated. Propranolol administration was associated with reduced baseline levels of heart rate and IL-1Ra, and systolic blood pressure (BP) in men. Tasks stimulated increased plasma IL-6 concentrations sampled 45 min and 75 min after challenge, but these responses were blocked by propranolol in men (p < 0.001). Propranolol did not influence IL-6 responses in women, or IL-1Ra in either sex. Blood pressure and heart rate increased markedly during the tasks, but there was no differential stress reactivity in propranolol and placebo conditions. The results of the study support a role of sympathetic nervous system activation in stimulating acute IL-6 responses to stress, but only in men. The reasons for the differences between men and women remain to be resolved.

Effect of dosing regimen and microneedle pretreatment on in vitro skin retention of topically applied beta-blockers.

Topical beta-blocker formulations are commonly used to treat infantile hemangiomas (IHs); however, the skin concentrations and drug permeation through the skin have not been quantified. Microneedles (MNs) may increase local skin concentrations, which could further enhance lesion clearance and improve dosing regimens. The objective of this study was to quantify skin concentrations and drug permeation of two beta-blockers, propranolol and timolol, in vitro after application to intact skin and skin pretreated with solid MNs of two lengths. Propranolol skin concentrations and drug permeation were significantly higher than timolol skin concentrations for all study conditions, which is likely due to the lipophilic nature of propranolol compared to the hydrophilicity of timolol. Propranolol skin concentrations were significantly influenced by dosing regimen, as skin concentrations increased with increasing drug application. Pretreatment of the skin with solid 250 µm and 500 µm length MNs increased local skin concentrations of timolol; propranolol skin concentrations did not significantly increase after MN pretreatment. Propranolol and timolol permeation through the skin increased after MN pretreatment with both MN lengths for both compounds. Taken together, solid MN pretreatment prior to application of topical timolol may be beneficial for deep or mixed IHs upon further optimization of the MN treatment paradigm.

Dual Activity of Hydroxypropyl-ß-Cyclodextrin and Water-Soluble Carriers on the Solubility of Carvedilol.

Carvedilol (CAR) is a non-selective α and ß blocker categorized as class II drug with low water solubility. Several recent studies have investigated ways to overcome this problem. The aim of the present study was to combine two of these methods: the inclusion complex using hydroxypropyl-ß-cyclodextrin (HPßCD) with solid dispersion using two carriers: Poloxamer 188 (PLX) and Polyvinylpyrrolidone K-30 (PVP) to enhance the solubility, bioavailability, and the stability of CAR. Kneading method was used to prepare CAR-HPßCD inclusion complex (KD). The action of different carriers separately and in combination on Carvedilol solubility was investigated in three series. CAR-carrier and KD-carrier solid dispersions were prepared by solvent evaporation method. In vitro dissolution test was conducted in three different media: double-distilled water (DDW), simulative gastric fluid (SGF), and PBS pH 6.8 (PBS). The interactions between CAR, HPßCD, and different carriers were explored by Fourier transform infrared spectroscopy (FTIR), powder X-ray diffractometry (XRD), and differential scanning colorimetry (DSC). The results showed higher solubility of CAR in KD-PVP solid dispersions up to 70, 25, and 22 fold compared to pure CAR in DDW, SGF, and PBS, respectively. DSC and XRD analyses indicated an improved degree of transformation of CAR in KD-PVP solid dispersion from crystalline to amorphous state. This study provides a new successful combination of two polymers with the dual action of HPßCD and PLX/PVP on water solubility and bioavailability of CAR.

Ocular Metabolism of Levobunolol: Historic and Emerging Metabolic Pathways.

Although ocular transport and delivery have been well studied, metabolism in the eye is not well documented, even for clinically available medications such as levobunolol, a potent and nonselective ß-adrenergic receptor antagonist. Recently, we reported an in vitro methodology that could be used to evaluate ocular metabolism across preclinical species and humans. The current investigation provides detailed in vitro ocular and liver metabolism of levobunolol in rat, rabbit, and human S9 fractions, including the formation of equipotent active metabolite, dihydrolevobunolol, with the help of high-resolution mass spectrometry. 11 of the 16 metabolites of levobunolol identified herein, including a direct acetyl conjugate of levobunolol observed in all ocular and liver fractions, have not been reported in the literature. The study documents the identification of six human ocular metabolites that have never been reported. The current investigation presents evidence for ocular and hepatic metabolism of levobunolol via non-cytochrome P450 pathways, which have not been comprehensively investigated to date. Our results indicated that rat liver S9 and human ocular S9 fractions formed the most metabolites. Furthermore, liver was a poor in vitro surrogate for eye, and rat and rabbit were poor surrogates for human in terms of the rate and extent of levobunolol metabolism.

Negative cooperativity across ß1-adrenoceptor homodimers provides insights into the nature of the secondary low-affinity CGP 12177 ß1-adrenoceptor binding conformation.

At the ß1-adrenoceptor, CGP 12177 potently antagonizes agonist responses at the primary high-affinity catecholamine conformation while also exerting agonist effects of its own through a secondary low-affinity conformation. A recent mutagenesis study identified transmembrane region (TM)4 of the ß1-adrenoceptor as key for this low-affinity conformation. Others suggested that TM4 has a role in ß1-adrenoceptor oligomerization. Here, assessment of the dissociation rate of a fluorescent analog of CGP 12177 [bordifluoropyrromethane-tetramethylrhodamine-(±)CGP 12177 (BODIPY-TMR-CGP)] at the human ß1-adrenoceptor expressed in Chinese hamster ovary cells revealed negative cooperative interactions between 2 distinct ß1-adrenoceptor conformations. The dissociation rate of 3 nM BODIPY-TMR-CGP was 0.09 ± 0.01 min(-1) in the absence of competitor ligands, and this was enhanced 2.2- and 2.1-fold in the presence of 1 µM CGP 12177 and 1 µM propranolol, respectively. These effects on the BODIPY-TMR-CGP dissociation rate were markedly enhanced in ß1-adrenoceptor homodimers constrained by bimolecular fluorescence complementation (9.8- and 9.9-fold for 1 µM CGP 12177 and 1 µM propranolol, respectively) and abolished in ß1-adrenoceptors containing TM4 mutations vital for the second conformation pharmacology. This study suggests that negative cooperativity across a ß1-adrenoceptor homodimer may be responsible for generating the low-affinity pharmacology of the secondary ß1-adrenoceptor conformation.

The Nonmetabolized ß-Blocker Nadolol Is a Substrate of OCT1, OCT2, MATE1, MATE2-K, and P-Glycoprotein, but Not of OATP1B1 and OATP1B3.

Nadolol is a nonmetabolized ß-adrenoceptor antagonist and is a substrate of OATP1A2, but not of OATP2B1. However, other drug transporters involved in translocation of nadolol have not been characterized in detail. We therefore investigated nadolol as a potential substrate of the hepatic uptake transporters OATP1B1, OATP1B3, and OCT1 and of the renal transporters OCT2, MATE1, and MATE2-K expressed in HEK cells. Moreover, the importance of P-glycoprotein (P-gp) for nadolol transport was studied using double transfected MDCK-OCT1-P-gp cells. Nadolol was not transported by OATP1B1 and OATP1B3. In contrast, a significantly higher nadolol accumulation (at 1 and 10 µM) was found in OCT1, OCT2, MATE1, and MATE2-K cells compared to control cells (P < 0.01). Km values for OCT2-, MATE1-, and MATE2-K-mediated nadolol uptake were 122, 531, and 372 µM, respectively. Cimetidine (100 µM, P < 0.01) and trimethoprim (100 µM, P < 0.001) significantly inhibited OCT1-, OCT2-, MATE1-, and MATE2-K-mediated nadolol transport. The P-gp inhibitor zosuquidar significantly reduced basal to apical nadolol transport in monolayers of MDCK-OCT1-P-gp cells. In summary, nadolol is a substrate of the cation transporters OCT1, OCT2, MATE1, MATE2-K, and of P-gp. These data will aid future in vivo studies on potential transporter-mediated drug-drug or drug-food interactions with involvement of nadolol.

SFC-MS/MS as an orthogonal technique for improved screening of polar analytes in anti-doping control.

HPLC is considered the method of choice for the separation of various classes of drugs. However, some analytes are still challenging as HPLC shows limited resolution capabilities for highly polar analytes as they interact insufficiently on conventional reversed-phase (RP) columns. Especially in combination with mass spectrometric detection, limitations apply for alterations of stationary phases. Some highly polar sympathomimetic drugs and their metabolites showed almost no retention on different RP columns. Their retention remains poor even on phenylhexyl phases that show different selectivity due to π-π interactions. Supercritical fluid chromatography (SFC) as an orthogonal separation technique to HPLC may help to overcome these issues. Selected polar drugs and metabolites were analyzed utilizing SFC separation. All compounds showed sharp peaks and good retention even for the very polar analytes, such as sulfoconjugates. Retention times and elution orders in SFC are different to both RP and HILIC separations as a result of the orthogonality. Short cycle times could be realized. As temperature and pressure strongly influence the polarity of supercritical fluids, precise regulation of temperature and backpressure is required for the stability of the retention times. As CO2 is the main constituent of the mobile phase in SFC, solvent consumption and solvent waste are considerably reduced. Graphical Abstract SFC-MS/MS vs. LC-MS/MS.

Chemoenzymatic Route for the Synthesis of (S)-Moprolol, a Potential ß-Blocker.

A biocatalytic route for the synthesis of a potential ß-blocker, (S)-moprolol is reported here. Enantiopure synthesis of moprolol is mainly dependent on the chiral intermediate, 3-(2-methoxyphenoxy)-propane-1,2-diol. Various commercial lipases were screened for the enantioselective resolution of (RS)-3-(2-methoxyphenoxy)propane-1,2-diol to produce the desired enantiomer. Among them, Aspergillus niger lipase (ANL) was selected on the basis of both stereo- and regioselectivity. The optimized values of various reaction parameters were determined such as enzyme (15 mg/mL), substrate concentration (10 mM), organic solvent (toluene), reaction temperature (30 °C), and time (18 h).The optimized conditions led to achieving >49% yield with high enantiomeric excess of (S)-3-(2-methoxyphenoxy)propane-1,2-diol. The lipase-mediated catalysis showed regioselective acylation with dual stereoselectivity. Further, the enantiopure intermediate was used for the synthesis of (S)-moprolol, which afforded the desired ß-blocker.

Topical Timolol for Infantile Hemangiomas: Evidence for Efficacy and Degree of Systemic Absorption.

BACKGROUND: Topical use of timolol for infantile hemangiomas has recently emerged with promising results. It is unknown whether topical ß-blockers act locally or if their effect is partly due to systemic absorption. This study investigates whether topically applied timolol is absorbed and reports on the efficacy of this treatment. METHODS: We treated 40 infants with small proliferating hemangiomas with topical timolol gel 0.5% twice daily and assessed urinary excretion and serum levels in a proportion of patients. Clinical response was evaluated on a visual analog scale of standardized photographs after 1, 2, 3, and 5 months. RESULTS: Forty infants with a median age of 18 weeks (range 2-35 wks) were included; 23 (58%) had superficial and 17 (42%) mixed-type hemangiomas. The median size was 3 cm(2) (range 0.1-15 cm(2) ) and nine hemangiomas were ulcerated. The hemangiomas improved significantly during treatment, with a median increase in visual analog scale of 7 points after 5 months (p < 0.001). Urinalysis for timolol was performed in 24 patients and was positive in 20 patients (83%). In three infants, serum levels of timolol were also measured and were all positive (median 0.16 ng/mL [range 0.1-0.18 ng/mL]). No significant side effects were recorded. CONCLUSION: Topical therapy with timolol is effective for infantile hemangiomas, but systemic absorption occurs. Serum levels in our patients were low, suggesting that using timolol for small hemangiomas is safe, but caution is advised when treating ulcerated or large hemangiomas, very young infants, or concomitantly using systemic propranolol.