Flupirtine and retigabine as templates for ligand-based drug design of KV7.2/3 activators.

Drug induced liver injury (DILI) and tissue discoloration led to the recent discontinuation of the therapeutic use of the closely related drugs flupirtine and retigabine, respectively. Experience gained with these drugs strongly suggests that heterotetramer, voltage-gated potassium channels 2 and 3 (KV7.2/3) are valid targets for effective treatment of pain and epilepsy. Because the adverse effects are not related to the mechanism of action, it appears promising to investigate chemical modifications of these clinically validated, drug-like leads. In the present retro-metabolic drug design study, a series of 43 compounds were synthesized and characterized with regard to KV7.2/3 opening activity and efficacy. The most active compound 22d displays excellent potency (EC50 = 4 nM) and efficacy (154%) as a KV7.2/3 opener. Limited aqueous solubility hampered toxicity testing at concentrations higher than 63 µM, but this concentration was nontoxic to two hepatocellular cell lines (HEP-G2 and TAMH) in culture. The slightly less active but more soluble compound 25b (EC50 = 11 nM, efficacy 111%) showed an improved toxicity/activity ratio compared to flupirtine by three orders of magnitude and represents an attractive lead structure for the development of safer analgesics and antiepileptics.

Synthesis and potassium KV7 channel opening activity of thioether analogues of the analgesic flupirtine.

Flupirtine, an opener of neuronal voltage gated potassium channels (KV7.2/3), has been used as a therapeutic alternative for pain treatment in patients refractory to NSAIDs and opioids. Because flupirtine is associated with rare but fatal drug-induced liver injury that may result from the formation of toxic metabolites upon metabolic oxidation, we synthesized novel derivatives with the goal of identifying equally active and ultimately safer KV7.2/3 channel openers. Four thioether analogues were designed to lack a nitrogen atom that would be a prerequisite for the formation of toxic para-quinone diimines, and form sulfoxide and sulfone metabolites instead. KV7.2/3 channel opening activity and hepatotoxicity data of twelve novel flupirtine analogues, four thioethers and their respective sulfoxide and sulfone metabolites are reported.

Challenges and Opportunities for Drug Discovery in Developing Countries: The Example of Cutaneous Leishmaniasis.

The current drug discovery paradigm has failed to address the treatment need for diseases of high priority to developing countries. Cutaneous leishmaniasis is a good example of such diseases with virtually no new effective drug developed in the past 70 years. The past two decades had witnessed relatively increased attention toward neglected diseases by stimulating pharmaceutical industries through introductions of Priority Review Vouchers (PRVs) and Product Development Partnerships (PDPs). However, the lion's share of resources allocated for research and development by PRVs and PDPs is directed toward research organizations and pharmaceutical industries in developed countries. This new approach has also not led to the development of drugs for most neglected diseases including cutaneous leishmaniasis. Improving the medical discovery capacity of countries where these diseases are prevalent and enabling exploration of the hitherto untapped natural resources are an effective and sustainable solution.

Sulfide Analogues of Flupirtine and Retigabine with Nanomolar KV 7.2/KV 7.3 Channel Opening Activity.

The potassium channel openers flupirtine and retigabine have proven to be valuable analgesics or antiepileptics. Their recent withdrawal due to occasional hepatotoxicity and tissue discoloration, respectively, leaves a therapeutic niche unfilled. Metabolic oxidation of both drugs gives rise to the formation of electrophilic quinones. These elusive, highly reactive metabolites may induce liver injury in the case of flupirtine and blue tissue discoloration after prolonged intake of retigabine. We examined which structural features can be altered to avoid the detrimental oxidation of the aromatic ring and shift oxidation toward the formation of more benign metabolites. Structure-activity relationship studies were performed to evaluate the KV 7.2/3 channel opening activity of 45 derivatives. Sulfide analogues were identified that are devoid of the risk of quinone formation, but possess potent KV 7.2/3 opening activity. For example, flupirtine analogue 3-(3,5-difluorophenyl)-N-(6-(isobutylthio)-2-(pyrrolidin-1-yl)pyridin-3-yl)propanamide (48) has 100-fold enhanced activity (EC50 =1.4 nm), a vastly improved toxicity/activity ratio, and the same efficacy as retigabine in vitro.

Simultaneous quantification of acidic and basic flupirtine metabolites by supercritical fluid chromatography according to European Medicines Agency validation.

Supercritical fluid chromatography (SFC) holds the potential to become an orthogonal method to HPLC/UHPLC in xenobiotic metabolism studies, due to its outstanding capacity to simultaneously separate highly similar (as HPLC) and physicochemically different analytes (problematic using HPLC). Paucity of guideline-conform validation, however, has been a major obstacle to clinical application of SFC, even in cases where biotransformation yields chemically dissimilar metabolites that require more than one HPLC method for comprehensive analysis. Here, a method based on supercritical fluid chromatography coupled to single quadrupole MS detection was developed to simultaneously quantify the divisive analgesic flupirtine and its acidic and basic metabolites, represented by 4-fluorohippuric acid (4-FHA) and the active metabolite D-13223 respectively, using custom-made synthetic internal standards. Experimental data on the fundamental retention mechanisms under supercritical conditions, indicating the importance of halogen and π-π-bonding for specific retention on polysaccharide-based stationary-phases, is discussed. Compared to previous HPLC methods, the novel method offers higher versatility in terms of the target metabolite range (addressing both acidic and basic metabolites within a singular method), faster analysis (7.5 min), and compliance with green chemistry principles. Validation was performed according to EMA criteria on bioanalytical method validation, demonstrating selectivity, carry-over, calibration curve parameters (LLOQ, range, and linearity), within- and between-run accuracy and precision, dilution integrity, matrix effect and stability. For proof-of-concept, the SFC method was applied to clinical samples of human urine obtained after single intravenous (100 mg), single oral (100 mg), and repeated oral administration (400 mg). Flupirtine, D-13223, and 4-FHA could be quantified, shedding light on the extent of oxidative flupirtine metabolism in humans in the context of the unresolved biotoxification that has led to the withdrawal of specific neuronal KV7 openers.

Flupirtine Analogues: Explorative Synthesis and Influence of Chemical Structure on KV7.2/KV7.3 Channel Opening Activity.

Neuronal voltage-gated potassium channels KV7.2/KV7.3 are sensitive to small-molecule drugs such as flupirtine, even though physiological response occurs in the absence of ligands. Clinically, prolonged use of flupirtine as a pain medication is associated with rare cases of drug-induced liver injury. Thus, safety concerns prevent a broader use of this non-opioid and non-steroidal analgesic in therapeutic areas with unmet medical needs such as hyperactive bladder or neonatal seizures. With the goal of studying influences of chemical structure on activity and toxicity of flupirtine, we explored modifications of the benzylamino bridge and the substitution pattern in both rings of flupirtine. Among twelve derivatives, four novel thioether derivatives showed the desired activity in cellular assays and may serve as leads for safer KV channel openers.

Self-reported and actual involvement of community pharmacists in patient counseling: a cross-sectional and simulated patient study in Gondar, Ethiopia.

BACKGROUND: Community pharmacists play a crucial role in reducing medication related health problems and improving the patient's overall wellbeing. Evidence suggests that community pharmacist led counseling services result in a better clinical and self-reported outcome, including a higher level of satisfaction and quality of life. OBJECTIVE: This study aims to document self-reported and actual levels of community pharmacists' involvement in the provision of patient counseling and barriers that limit their involvement in such services. METHODS: Simulated patient visits and a cross-sectional survey of community pharmacists were employed in Gondar town, Ethiopia between March 15 and May 15, 2016 to observe actual counseling practices and to assess their reported counseling practices respectively. Four different scenarios were developed for the simulated patient visit. A well designed questionnaire and an assessment form were used for the survey and simulated patient visit. RESULTS: In the cross-sectional survey, 84 pharmacists were approached and 78 agreed to participate (92.8 % response rate). Of the respondents, 96.1% agreed/strongly agreed that patient counseling is important and 69.3% strongly agreed that patient counseling should be a professional duty. The most frequent information provided to patients were dosing schedule of drugs, how to take medication, and drug-food interaction. Majority of community pharmacists either strongly agreed (42.1%) or agreed (51.3%) that patients are comfortable towards their counseling practice. A total of 48 simulated visits were conducted and a medicine was dispensed in all visits. In all four scenarios, dosage schedule (100%), how to take medication (97.6%) and drug-food interaction (69.1%) were the most common type of information provided while what to do when dose is missed (100%), contraindication (95.2%) and the importance of compliance or adherence (92.9%) were the most commonly ignored types of information. CONCLUSIONS: The present study emphasizes the existing gap in self-reported and actual counseling practices by community pharmacist in Gondar town, Ethiopia. Hence, the ministry of health, local health policy makers and other stakeholders should collaborate to design interventions to improve community pharmacists' dispensing and counseling practice.

Self-reported and actual involvement of community pharmacists in patient counseling: a cross-sectional and simulated patient study in Gondar, Ethiopia

Background: Community pharmacists play a crucial role in reducing medication related health problems and improving the patient’s overall wellbeing. Evidence suggests that community pharmacist led counseling services result in a better clinical and self-reported outcome, including a higher level of satisfaction and quality of life. Objective: This study aims to document self-reported and actual levels of community pharmacists’ involvement in the provision of patient counseling and barriers that limit their involvement in such services. Methods: Simulated patient visits and a cross-sectional survey of community pharmacists were employed in Gondar town, Ethiopia between March 15 and May 15, 2016 to observe actual counseling practices and to assess their reported counseling practices respectively. Four different scenarios were developed for the simulated patient visit. A well designed questionnaire and an assessment form were used for the survey and simulated patient visit. Results: In the cross-sectional survey, 84 pharmacists were approached and 78 agreed to participate (92.8 % response rate). Of the respondents, 96.1% agreed/strongly agreed that patient counseling is important and 69.3% strongly agreed that patient counseling should be a professional duty. The most frequent information provided to patients were dosing schedule of drugs, how to take medication, and drug-food interaction. Majority of community pharmacists either strongly agreed (42.1%) or agreed (51.3%) that patients are comfortable towards their counseling practice. A total of 48 simulated visits were conducted and a medicine was dispensed in all visits. In all four scenarios, dosage schedule (100%), how to take medication (97.6%) and drug-food interaction (69.1%) were the most common type of information provided while what to do when dose is missed (100%), contraindication (95.2%) and the importance of compliance or adherence (92.9%) were the most commonly ignored types of information. Conclusions: The present study emphasizes the existing gap in self-reported and actual counseling practices by community pharmacist in Gondar town, Ethiopia. Hence, the ministry of health, local health policy makers and other stakeholders should collaborate to design interventions to improve community pharmacists’ dispensing and counseling practice (AU)

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