Phenotypic studies on dopamine receptor subtype and associated signal transduction mutants: insights and challenges from 10 years at the psychopharmacology–molecular biology interface

Background Mutants with targeted gene deletion (‘knockout’) or insertion (transgenic) of D₁, D₂, D₃, D₄ and D₅ dopamine (DA) receptor subtypes are complemented by an increasing variety of double knockout and transgenic-‘knockout’ models, together with knockout of critical components of DA receptor signalling cascades such as Gα_olf[Gγ₇], adenylyl cyclase type 5, PKA [RIIβ] and DARPP-32. However, it is increasingly recognised that these molecular techniques have a number of inherent limitations. Furthermore, there are poorly understood methodological factors that contribute to inconsistent phenotypic findings between laboratories. Objective This review seeks to document the impact of DA receptor subtype and related transduction mutants on our understanding of the behavioural roles of these entities, primarily at the level of unconditioned psychomotor behaviour. Methods It includes ethologically based and orofacial movement studies in our own laboratories, since these are the only studies to systematically compare each of the D₁, D₂, D₃, D₄ and D₅ receptor and DARPP-32 signal transduction ‘knockouts’. Discussion There is a particular emphasis on identifying methodological factors that might influence phenotypic effects and account for inconsistencies. The findings are offered empirically to (1) specify the extent of phenotypic diversity among individual DA receptor subtypes and transduction components and (2) indicate relationships between D₁, D₂, D₃, D₄ and D₅ receptor subtype proteins, associated Gα_i/Gα_s/Gα_olf[Gγ₇]–adenylyl cyclase type 5–PKA [RIIβ]–DARPP-32 signalling cascades and behaviour. The findings are also offered heuristically as a base for such phenotypic comparisons at additional levels of behaviour so that a yet more complete phenotypic profile might emerge.     

A multidisciplinary approach combining molecular biology,bio-analytical chemistry,and immunochemistry to probe the role of bioactivation of naphthalene in cytotoxicity

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Hepatitis C treatment at a Swedish needle exchange program,a successful model of care – the ACTIONNE study

BackgroundTo engage people who inject drugs (PWID) in HCV care, innovative models of care are urgently needed. A needle exchange program (NEP) could serve as an ideal platform for comprehensive HCV management including post treatment follow up.Methods50 actively injecting patients at the Malmö Needle exchange program (MNEP) were consecutively enrolled between April 2018 and May 2019. All patients received a fixed-dose combination of once-daily glecaprevir/pibrentasvir for 8 or 12 weeks. Patients were monitored weekly during treatment and data on adherence and side effects was recorded. The primary endpoint was SVR12. Adherence to treatment was the secondary endpoint.Results47/50 (94%) patients completed treatment. 45/50 were HCV negative at 12 weeks post treatment giving an SVR12 rate per ITT of 90% and an SVR12 rate per protocol of 96%. One patient showed reinfection 12 weeks post treatment and one patient was lost to follow up and did not produce an SVR12 result. The mean adherence per week, according to pill count, was 98%.ConclusionOur study shows that the NEP can be a useful tool for engaging actively injecting PWID in HCV management and that SVR rates, comparable to those in non-PWID settings, can be achieved.     

Are rats the appropriate experimental model to understand age-related renal drug metabolism and toxicity?

For many years, toxicological investigations have shown that the sensitivity of kidney to xenobiotics evolves depending on the stage of life. The increasing requirement for information on the potential nephrotoxic effect of drugs during human embryonic development, childhood, adulthood and senescence has potentiated toxicological studies in vivo. Rodents, specifically rats, are the primary animal models used in toxicology testing. Despite the popularity of this approach, there are a number of doubts about the appropriateness of rats for the examination of changes in toxicological responses during different stages of life. This perspective tackles the issue of evaluating whether rats fail to adequately mimic the human kidney response to xenobiotic agents through a critical analysis of the literature. We conclude that rats constitute a good model for toxicological investigations during embryonic development, youth and adulthood. However, senescent rats frequently undergo spontaneous kidney degeneration caused by chronic progressive nephropathy, making them a poor model for the study of kidney responses to xenobiotics.     

Non-alcoholic fatty liver disease (NAFLD) – pathogenesis,classification, and effect on drug metabolizing enzymes and transporters

Non-alcoholic fatty liver disease (NAFLD) is a spectrum of liver disorders. It is defined by the presence of steatosis in more than 5% of hepatocytes with little or no alcohol consumption. Insulin resistance, the metabolic syndrome or type 2 diabetes and genetic variants of PNPLA3 or TM6SF2 seem to play a role in the pathogenesis of NAFLD. The pathological progression of NAFLD follows tentatively a “three-hit” process namely steatosis, lipotoxicity and inflammation. The presence of steatosis, oxidative stress and inflammatory mediators like TNF-α and IL-6 has been implicated in the alterations of nuclear factors such as CAR, PXR, PPAR-α in NAFLD. These factors may result in altered expression and activity of drug metabolizing enzymes (DMEs) or transporters.

Existing evidence suggests that the effect of NAFLD on CYP3A4, CYP2E1 and MRP3 is more consistent across rodent and human studies. CYP3A4 activity is down-regulated in NASH whereas the activity of CYP2E1 and the efflux transporter MRP3 is up-regulated. However, it is not clear how the majority of CYPs, UGTs, SULTs and transporters are influenced by NAFLD either in vivo or in vitro. The alterations associated with NAFLD could be a potential source of drug variability in patients and could have serious implications for the safety and efficacy of xenobiotics. In this review, we summarize the effects of NAFLD on the regulation, expression and activity of major DMEs and transporters. We also discuss the potential mechanisms underlying these alterations.     

The lay user perspective on the quality of pharmaceuticals, drug therapy and pharmacy services – results of focus group discussions

Background: This article presents the results of a study on quality of pharmacy services and perceived risk of pharmaceuticals. The results presented here are part of a multi-study evaluation of major changes in drug distribution in Iceland. Objectives: This sub-study addressed the question: what is the lay user perspective on pharmaceuticals and pharmacy services, including their perception of risk? Methods: To answer this question, seven focus group discussions were conducted with pharmacy customers in different locations in Iceland following new drug distribution legislation in 1996.Results: The lay perspective emphasizes a definite split between lay and expert views on the value and quality of pharmaceuticals, drug therapy and pharmacy services, as well as in their assessment of risk. Participants voiced spontaneous criticism of the roles of both physicians and pharmacists in drug therapy; and expressed concern about the quality and safety of pharmaceuticals. Some scope for shared values was noted between the legislative goals and the lay user perspective, despite the fact that the public was in no way involved in the drafting of the new legislation. Conclusion: The results of this study raise questions about the nature and extent of the perceived gap between the medical and pharmacy professions on the one side and the lay public and health policy decision-makers on the other side in their views on the quality and safety of pharmaceuticals and pharmacy services.     

Compliance with the legal age limits for alcohol,tobacco and gambling – A comparative study on test purchasing in retail outlets

Aim: To assess whether retail outlets comply with the minimum legal age of 18 for the purchase of alcohol and tobacco and for gambling on slot machines in Finland. Methods: A test-purchase research in 117 retail outlets selling alcohol, tobacco and supplying slot machines in two towns. Five-hundred fifty-seven purchase attempts were made for alcohol (n?=?173), tobacco (n?=?177) and gambling on slot machines (n?=?170) in private outlets and 37 attempts in government alcohol retail monopoly outlets. The differences in denial rates and factors of potential significance for denials were tested using logistic regression. Findings: The denial rate for gambling on slot machines was 4%. The odds that an alcohol or a tobacco purchase was denied were 25- and 20-fold, respectively, compared to gambling. The odds of denial of an alcohol purchase were 12-fold in alcohol monopoly stores compared to private outlets. Conclusion: The low compliance with the age limit for gambling is a challenge in a gambling policy system where slot machines are decentralized in private retail outlets and are widely available. The high denial rate in government alcohol retail monopoly outlets indicates that monopoly outlets are more capable of enforcing legal age limits effectively compared to private outlets.     

Healthcare professionals’ knowledge,attitude and practice towards adverse drug reaction (ADR) reporting at the health center level in Ethiopia

International Journal of Clinical Pharmacy - Background Adverse drug reactions (ADRs) are major health problems which are of global concern. Spontaneous reporting of adverse drug reactions...     

Reasons for supply side driven drug shortages – A mixed-methods study on first-level,higher-level,and root causes from the perspective of marketing authorization holders

BackgroundDrug shortages impact multiple stakeholders and are detrimental to patient safety. Additionally, drug shortages are an extensive financial burden. In Germany, drug shortages, according to data from the federal ministry for drug and medical products (BfArM), have been increasing by 18% between 2018 and 2021. Studies show that shortages are most frequently supply side driven and that often reasons remain unknown.ObjectiveThe aim is to develop a holistic understanding of supply side causes for drug shortages in Germany from marketing authorization holders’ perspectives and to derive implications for shortage mitigation.MethodsA mixed-methods research design, with a grounded theory approach based on a structured literature review, BfArM data analysis, and semi-structured interviews, was used.ResultsInput factor supply issues, manufacturing issues, logistics issues, product recalls, and product discontinuations were identified as first-level causes. Furthermore, a theory on their connection to higher-level causes related to business decision-making, as well as root causes linked to regulations, company values, internal processes, market dynamics, external shocks, and macroeconomic factors, was developed.ConclusionActions to mitigate drug shortages in Germany (e.g., improving business processes, diversifying tender criteria) were derived. These may thus increase patient safety and decrease the financial burden on the healthcare system.     

In vitro profiling of the metabolism and drug–drug interaction of tofogliflozin,a potent and highly specific sodium-glucose co-transporter 2 inhibitor,using human liver microsomes,human hepatocytes,and recombinant human CYP

Abstract

1. The metabolism and drug–drug interaction (DDI) risk of tofogliflozin, a potent and highly specific sodium-glucose co-transporter 2 inhibitor, were evaluated by in vitro studies using human liver microsomes, human hepatocytes, and recombinant human CYPs.

2. The main metabolite of tofogliflozin was the carboxylated derivative (M1) in human hepatocytes, which was the same as in vivo. The metabolic pathway of tofogliflozin to M1 was considered to be as follows: first, tofogliflozin was catalyzed to the primary hydroxylated derivative (M4) by CYP2C18, CYP4A11 and CYP4F3B, then M4 was oxidized to M1.

3. Tofogliflozin had no induction potential on CYP1A2 and CYP3A4. Neither tofogliflozin nor M1 had inhibition potential on CYPs, with the exception of a weak CYP2C19 inhibition by M1.

4. Not only are multiple metabolic enzymes involved in the tofogliflozin metabolism, but the drug is also excreted into urine after oral administration, indicating that tofogliflozin is eliminated through multiple pathways. Thus, the exposure of tofogliflozin would not be significantly altered by DDI caused by any co-administered drugs. Also, tofogliflozin seems not to cause significant DDI of co-administered drugs because tofogliflozin has no CYP induction or inhibition potency, and the main metabolite M1 has no clinically relevant CYP inhibition potency.     

Vitamin D status in relation to age,bone mineral density of the spine and femur in obese Saudi females – A hospital-based study

The aim of the present study was to evaluate the association between Bone mineral density in lumber spine and femoral neck with serum total levels of vitamin D, sun exposure and Consumption of vitamin D Supplement in obese Saudi females aged between 30 and 54?years old. Recent attention to the high prevalence of osteoporosis and its association with low vitamin D levels in adults has raised the importance of vitamin D evaluation. A low level of vitamin D is considered to be one of the most important risk factors for osteoporosis. In this study; 120 obese Saudi females with no diagnosed chronic diseases attending the Outpatient clinic at king Khalid University hospital in Riyadh. Saudi Arabia, recruited randomly in period of 12?months. In this study, Serum levels of total Vitamin D were considered to be severe deficient if it was lower than 25?ng/mL, mild to moderate deficient if it was between 25 and 60?ng/mL and optimum level if it was 61–200?ng/mL. The results showed that; sun exposure was significantly affect and Correlate with serum level of Vitamin D in the subjects. In addition, daily consumption of Vitamin D supplement was significantly affect and Correlate with serum level of Vitamin D in the subjects of this study. Moreover, the results showed that; 50% of the age group (40–49?years old) having severe deficiency of Vitamin D. While, 50% of the age group (50–59?years old) having optimal level of Vitamin D. And these results mean that age is not Correlated with vitamin D deficiency in subjects of this study.     

Characterisation of the thermal,spectroscopic and drug dissolution properties of mefenamic acid and polyoxyethylene–polyoxypropylene solid dispersions

The aim of this study was to investigate the solubility of mefenamic acid (MA), a highly cohesive, poorly water-soluble drug in a copolymer of polyoxyethylene–polyoxypropylene (Lutrol F68®), and to understand the effect drug polymer solubility has on in vitro dissolution of MA. Solid dispersions (SD) of MA were prepared by a hot melt method, using Lutrol F68® as a thermoplastic polymeric platform. High-speed differential scanning calorimetry (Hyper-DSC), Raman spectroscopy, powder X-ray diffractometry (PXRD) and hot-stage/fluorescence microscopy were used to assess the solubility of the drug in molten and solid polymer. Drug dissolution studies were subsequently conducted on single-phase solid solutions and biphasic SD using phosphate buffer pH 6.8 as dissolution media. Solubility investigations using Hyper-DSC, Raman spectroscopy and hot-stage microscopy suggested MA was soluble in molten Lutrol F68® up to a concentration of 35% (w/w). Conversely, the solubility in the solid-state matrix was limited to <15% (w/w); determined by Raman spectroscopy, PXRD and fluorescence microscopy. As expected the dissolution properties of MA were significantly influenced by the solubility of the drug in the polymer matrix. At a concentration of 10% (w/w) MA (a single phase solid solution) dissolution of MA in phosphate buffer 6.8 was rapid, whereas at a concentration of 50% (w/w) MA (biphasic SD) dissolution was significantly slower. This study has clearly demonstrated the complexity of drug–polymer binary blends and in particular defining the solubility of a drug within a polymeric platform. Moreover, this investigation has demonstrated the significant effect drug solubility within a polymeric matrix has upon the in vitro dissolution properties of solid polymer/drug binary blends. © 2009 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 98:4545–4556, 2009     

A contrast in safety,pharmacokinetics and pharmacodynamics across age groups after a single 50 mg oral dose of the γ-secretase inhibitor avagacestat

AIM

To evaluate the single dose pharmacokinetics, pharmacodynamics, and preliminary tolerability of the γ-secretase inhibitor BMS-708163 (avagacestat) in young and elderly men and women.

METHODS

All subjects received double-blinded administration of a single 50 mg dose of avagacestat in capsule form or matching placebo. Main evaluations included pharmacokinetics, safety, plasma amyloid-β (Aβ)_1–40 concentratios and exploration of Notch biomarkers.

RESULTS

Avagacestat 50 mg capsule was well tolerated and rapidly absorbed among young and elderly subjects, with a median t_max between 1 and 2 h post dose and an average half-life between 41 and 71 h. In general, subjects aged 75 years or more had higher AUC(0,∞) values than those aged less than 75 years. An exploratory analysis of Aβ_1–40 serum concentrations showed a pattern of decreasing concentrations over the first 4–6 h followed by a rise above baseline that was maintained until the end of the assessment period. Adverse events were generally mild, occurring more frequently in elderly subjects, with no observed difference between subjects receiving avagacestat and placebo. No dose limiting gastrointestinal effects of avagacestat were observed and exploratory biomarkers of Notch inhibition did not change significantly.

CONCLUSIONS

The favourable safety profile and pharmacokinetic effects of avagacestat in this study support its continued development, especially in the target population of elderly subjects with mild cognitive impairment or Alzheimer''s disease.     

Applicability of the Øie-Tozer model to predict three types of distribution volume (Vd) in humans: Vd in central compartment,Vd at steady state,and Vd at beta phase

This study aimed to investigate the applicability of the Øie-Tozer model to predict human distribution volume (Vd) in the central compartment (V₁), Vd at steady state (Vd_ss), and Vd at beta phase (Vd_β) based on animal Vd. Twenty compounds that have a human V₁/Vd_ss of 0.053–0.66 were selected from the literature. After intravenous administration of the compounds at 0.1 mg/kg to rats, dogs, and monkeys, plasma concentrations were determined, and pharmacokinetic parameters were obtained by one/two-compartmental analyses. The human V₁, Vd_ss, and Vd_β were predicted from animal Vd using the Øie-Tozer model, and the predictability was compared with that using proportionality and simple allometry. The Øie-Tozer model was the most reliable method for the overall prediction of Vd and applicable for accurately predicting human V₁, Vd_ss, and Vd_β (89%, 85%, and 68% of the compounds within a 3-fold error, respectively) when data of monkey for V₁ and data of three animal species for Vd_ss and Vd_β were used. Additionally, the predicted human Vd with the two-compartment model was applicable for predicting pharmacokinetic profiles/parameters in humans after intravenous administration of 18 compounds [except for valproic acid (monophasic elimination profile) and chlorpromazine (deviation: Vd_ss < V₁)]. The prediction was more accurate than that using the predicted Vd_ss with the one-compartment model (e.g., underestimation of maximum plasma concentrations: 2 vs 8 compounds within a 3-fold error, respectively). In summary, the Øie-Tozer model was applicable for predicting human V₁, Vd_ss, and Vd_β, and their predicted Vd with the two-compartment model can lead to accurate pharmacokinetic prediction of compounds that show biphasic elimination.     

Perazine at therapeutic drug concentrations inhibits human cytochrome P450 isoenzyme 1A2 (CYP1A2) and caffeine metabolism – an in vitro study

The aim of the present study was to estimate the inhibitory effect of perazine, a phenothiazine neuroleptic with piperazine structure in a side chain, on human CYP1A2 activity measured as a rate of caffeine 3-N- and 1-N-demethylation. Moreover, the influence of perazine on other caffeine metabolic pathways such as 7-N-demethylation (CYP1A2, CYP2C8/9, CYP3A4) and 8-hydroxylation (CYP3A4, CYP1A2, CYP2C8/9) was also determined. The Dixon analysis showed that in both human liver microsomes and Supersomes CYP1A2 perazine potently and to a similar degree inhibited caffeine 3-N-demethylation (K_i = 3.5 μM) and 1-N-demethylation (K_i = 5 μM). Perazine moderately diminished the rate of caffeine 7-N-demethylation in Supersomes CYP1A2 (K_i = 11.5 μM) and liver microsomes (K_i = 20 μM), and attenuated C-8-hydroxylation (K_i = 15.5μM) in Supersomes CYP1A2. On the other hand, perazine weakly inhibited caffeine C-8-hydroxylation in liver microsomes (K_i = 98 μM). About 80% of basal CYP1A2 activity was reduced by the therapeutic concentrations of perazine (5–10 μM).The obtained results show that perazine at its therapeutic concentrations is a potent inhibitor of human CYP1A2. Hence, taking account of CYP1A2 contribution to the metabolism of endogenous substances (steroids), drugs (xanthine derivatives, phenacetin, propranolol, imipramine, phenothiazine neuroleptics, clozapine) and carcinogenic compounds, the inhibition of CYP1A2 by perazine may be of physiological, pharmacological and toxicological importance.