Population pharmacokinetics of primaquine and its metabolites in African males.

BACKGROUND: Primaquine (PQ) is the prototype 8-aminoquinoline drug, a class which targets gametocytes and hypnozoites. The World Health Organization (WHO) recommends adding a single low dose of primaquine to the standard artemisinin-based combination therapy (ACT) in order to block malaria transmission in regions with low malaria transmission. However, the haemolytic toxicity is a major adverse outcome of primaquine in glucose-6-phosphate dehydrogenase (G6PD)-deficient subjects. This study aimed to characterize the pharmacokinetic properties of primaquine and its major metabolites in G6PD-deficient subjects. METHODS: A single low-dose of primaquine (0.4-0.5 mg/kg) was administered in twenty-eight African males. Venous and capillary plasma were sampled up to 24 h after the drug administration. Haemoglobin levels were observed up to 28 days after drug administration. Only PQ, carboxy-primaquine (CPQ), and primaquine carbamoyl-glucuronide (PQCG) were present in plasma samples and measured using liquid chromatography mass spectrometry. Drug and metabolites' pharmacokinetic properties were investigated using nonlinear mixed-effects modelling. RESULTS: Population pharmacokinetic properties of PQ, CPQ, and PQCG can be described by one-compartment disposition kinetics with a transit-absorption model. Body weight was implemented as an allometric function on the clearance and volume parameters for all compounds. None of the covariates significantly affected the pharmacokinetic parameters. No significant correlations were detected between the exposures of the measured compounds and the change in haemoglobin or methaemoglobin levels. There was no significant haemoglobin drop in the G6PD-deficient patients after administration of a single low dose of PQ. CONCLUSIONS: A single low-dose of PQ was haematologically safe in this population of G6PD-normal and G6PD-deficient African males without malaria. Trial registration NCT02535767.

Comparative pharmacokinetics and tissue distribution of primaquine enantiomers in mice.

BACKGROUND: Primaquine (PQ) has been used for the radical cure of relapsing Plasmodium vivax malaria for more than 60 years. PQ is also recommended for prophylaxis and prevention of transmission of Plasmodium falciparum. However, clinical utility of PQ has been limited due to toxicity in individuals with genetic deficiencies in glucose 6-phosphate dehydrogenase (G6PD). PQ is currently approved for clinical use as a racemic mixture. Recent studies in animals as well as humans have established differential pharmacological and toxicological properties of the two enantiomers of PQ. This has been attributed to differential metabolism and pharmacokinetics of individual PQ enantiomers. The aim of the current study is to evaluate the comparative pharmacokinetics (PK), tissue distribution and metabolic profiles of the individual enantiomers in mice. METHODS: Two groups of 21 male Albino ND4 Swiss mice were dosed orally with 45 mg/kg of S-(+)-PQ and R-(-)PQ respectively. Each of the enantiomers was comprised of a 50:50 mixture of 12C- and 13C- stable isotope labelled species (at 6 carbons on the benzene ring of the quinoline core). Three mice were euthanized from each group at different time points (at 0, 0.5, 1, 2, 4, 8, 24 h) and blood was collected by terminal cardiac bleed. Liver, spleen, lungs, kidneys and brain were removed, extracted and analysed using UPLC/MS. The metabolites were profiled by tandem mass (MS/MS) fragmentation profile and fragments with 12C-13C twin peaks. Non-compartmental analysis was performed using the Phoenix WinNonLin PK software module. RESULTS: The plasma AUC0-last (µg h/mL) (1.6 vs. 0.6), T1/2 (h) (1.9 vs. 0.45), and Tmax (h) (1 vs. 0.5) were greater for SPQ as compared to RPQ. Generally, the concentration of SPQ was higher in all tissues. At Tmax, (0.5-1 h in all tissues), the level of SPQ was 3 times that of RPQ in the liver. Measured Cmax of SPQ and RPQ in the liver were about 100 and 40 times the Cmax values in plasma, respectively. Similar observations were recorded in other tissues where the concentration of SPQ was higher compared to RPQ (2× in the spleen, 6× in the kidneys, and 49× in the lungs) than in the plasma. CPQ, the major metabolite, was preferentially generated from RPQ, with higher levels in all tissues (> 10× in the liver, and 3.5× in the plasma) than from SPQ. The PQ-o-quinone was preferentially formed from the SPQ (> 4× compared to RPQ), with higher concentrations in the liver. CONCLUSION: These studies show that in mice, PQ enantiomers are differentially biodistributed and metabolized, which may contribute to differential pharmacologic and toxicity profiles of PQ enantiomers. The findings on higher levels of PQ-o-quinone in liver and RBCs compared to plasma and preferential generation of this metabolite from SPQ are consistent with the higher anti-malarial efficacy of SPQ observed in the mouse causal prophylaxis test, and higher haemolytic toxicity in the humanized mouse model of G6PD deficiency. Potential relevance of these findings to clinical use of racemic PQ and other 8-aminoquinolines vis-à-vis need for further clinical evaluation of individual enantiomers are discussed.

Formation primaquine-5,6-orthoquinone, the putative active and toxic metabolite of primaquine via direct oxidation in human erythrocytes.

BACKGROUND: The activity and haemolytic toxicity associated with primaquine has been linked to its reactive metabolites. The reactive metabolites are thought to be primarily formed through the action of cytochrome P450-mediated pathways. Human erythrocytes generally are not considered a significant contributor to drug biotransformation. As erythrocytes are the target of primaquine toxicity, the ability of erythrocytes to mediate the formation of reactive oxidative primaquine metabolites in the absence of hepatic enzymes, was evaluated. METHODS: Primaquine and its enantiomers were incubated separately with human red blood cells and haemoglobin. Post-incubation analysis was performed with UPLC-MS/MS to identify products of biotransformation. RESULTS: The major metabolite detected was identified as primaquine-5,6-orthoquinone, reflecting the pathway yielding putative active and haematotoxic metabolites of primaquine, which was formed by oxidative demethylation of 5-hydroxyprimaquine. Incubation of primaquine with haemoglobin in a cell-free system yielded similar results. It appears that the observed biotransformation is due to non-enzymatic processes, perhaps due to reactive oxygen species (ROS) present in erythrocytes or in the haemoglobin incubates. CONCLUSION: This study presents new evidence that primaquine-5,6-orthoquinone, the metabolite of primaquine reflecting the oxidative biotransformation pathway, is generated in erythrocytes, probably by non-enzymatic means, and may not require transport from the liver or other tissues.

Modulation of Cytochrome P450, P-glycoprotein and Pregnane X Receptor by Selected Antimalarial Herbs-Implication for Herb-Drug Interaction.

Seven medicinal plants popularly used for treating malaria in West Africa were selected to assess herb-drug interaction potential through a series of in vitro methods. Fluorescent cytochrome P450 (CYP) assays were conducted using the recombinant CYP enzymes for CYP1A2, CYP2A6, CYP2B6, CYP2C9, CYP2C19, CYP2D6 and CYP3A4 to assess the effect of the methanolic extracts on the metabolic activity of CYPs. Secondly, the inhibitory effect of the extracts was evaluated on P-glycoproteins (P-gp) using calcein-AM, a fluorescent substrate, in MDCK-II and hMDR1-MDCK-II cells. The inhibition of P-gp activity was determined as a reflection of increase in calcein-AM uptake. Additionally, the enzyme induction potential of the extracts was assessed through the modulation of PXR activity in HepG2 cells transiently transfected with pSG5-PXR and PCR5 plasmid DNA. Significant inhibition of CYP activity (IC50 < 10 µg/mL) was observed with the following herbs: A. muricata [CYP2C9, 3A4 and CYP2D6]; M. indica [CYP2C9]; M. charantia [CYP2C9 and CYP2C19]; P. amarus [CYP2C19, CYP2C9 and CYP3A4]; T. diversifolia [CYP2C19 and CYP3A4]. Extracts of four herbs (P. amarus, M. charantia, T. diversifolia and A. muricata) exhibited significant inhibition of P-gp with IC50 values (µg/mL) of 17 ± 1, 16 ± 0.4, 26 ± 1, and 24 ± 1, respectively. In addition, four herbs (A. mexicana, M. charantia, P. amarus and T. diversifolia) showed a >two-fold increase in induction in PXR activity. These findings suggest that these herbs may be capable of eliciting herb-drug interactions if consumed in high quantities with concomitant use of conventional therapies.

Pathway-specific inhibition of primaquine metabolism by chloroquine/quinine.

BACKGROUND: There has been some evidence to suggest that the addition of chloroquine (CQ) or quinine (QN) to 8-aminoquinoline (8-AQ) treatment regimens may increase the therapeutic efficacy of the 8-AQ and simultaneously mitigate against its haemolytic toxicity. However, both CQ and QN are considered effective, although perhaps moderate inhibitors of CYP2D6, an enzyme now regarded as necessary for primaquine (PQ) pharmacologic activity. An understanding of the influence of CQ and QN on the metabolism of PQ may shed light on the potential mechanisms of the beneficial interaction. METHODS: Differential metabolism of PQ enantiomers by recombinant human CYP2D6, monoamine oxidase A (MAO), and cryopreserved human hepatocytes in the presence/absence of CQ and QN. RESULTS: Both CQ and QN significantly inhibited the activity of CYP2D6. PQ depletion by MAO and human hepatocytes was not affected significantly by the presence of CQ and QN. CYP2D6-mediated hydroxylation was largely suppressed by both CQ and QN. The formation of the primary deaminated metabolites, including carboxyprimaquine (CPQ) and cyclized side chain derivative from the aldehyde (m/z 241), was not sensitive to the presence of CQ and QN. However, the appearance of the glucuronides of CPQ and PQ alcohol were significantly suppressed. CQ and QN also inhibited the appearance of the m/z 257 metabolite with a similar pattern, suggesting that it may be derived from the CPQ conjugate. The apparent quinone-imine of CPQ (m/z 289) was only partially suppressed by both QN and CQ, but with a differential pattern of inhibition for the two drugs. The m/z 274 (quinone-imine of a ring-hydroxylated PQ metabolite) and m/z 422 (an apparent glucose conjugate of PQ) metabolites in hepatocytes were strongly suppressed by both QN and CQ, perhaps a reflection of the 2D6 inhibition by these drugs. The formation of the carbamoyl glucuronide of PQ (m/z 480) was not affected by CQ/QN. CONCLUSION: The metabolite-specific interactions in the current studies seem at variance with earlier reports of the dependence of PQ on CYP2D6 metabolism, and enhanced PQ anti-malarial activity/reduced toxicity in the presence of CQ/QN. These results suggest a complex picture in which CQ/QN may shift metabolite pathway balances towards a profile that retains efficacy, while reducing the formation or availability of toxic metabolites to erythrocytes. Alternatively, these drugs may alter transport or distribution of PQ metabolites in a fashion that reduces toxicity while maintaining efficacy against the parasite.

Differential kinetic profiles and metabolism of primaquine enantiomers by human hepatocytes.

BACKGROUND: The clinical utility of primaquine (PQ), used as a racemic mixture of two enantiomers, is limited due to metabolism-linked hemolytic toxicity in individuals with genetic deficiency in glucose-6-phosphate dehydrogenase. The current study investigated differential metabolism of PQ enantiomers in light of the suggestions that toxicity and efficacy might be largely enantioselective. METHODS: Stable isotope (13)C-labelled primaquine and its two enantiomers (+)-PQ, (-)-PQ were separately incubated with cryopreserved human hepatocytes. Time-tracked substrate depletion and metabolite production were monitored via UHPLC-MS/MS. RESULTS: The initial half-life of 217 and 65 min; elimination rate constants (λ) of 0.19 and 0.64 h(-1); intrinsic clearance (Clint) of 2.55 and 8.49 (µL/min)/million cells, which when up-scaled yielded Clint of 6.49 and 21.6 (mL/min)/kg body mass was obtained respectively for (+)- and (-)-PQ. The extrapolation of in vitro intrinsic clearance to in vivo human hepatic blood clearance, performed using the well-stirred liver model, showed that the rate of hepatic clearance of (+)-PQ was only 45 % that of (-)-PQ. Two major primary routes of metabolism were observed-oxidative deamination of the terminal amine and hydroxylations on the quinoline moiety of PQ. The major deaminated metabolite, carboxyprimaquine (CPQ) was preferentially generated from the (-)-PQ. Other deaminated metabolites including PQ terminal alcohol (m/z 261), a cyclized side chain derivative from the aldehyde (m/z 241), cyclized carboxylic acid derivative (m/z 257), a quinone-imine product of hydroxylated CPQ (m/z 289), CPQ glucuronide (m/z 451) and the glucuronide of PQ alcohol (m/z 437) were all preferentially generated from the (-)-PQ. The major quinoline oxidation product (m/z 274) was preferentially generated from (+)-PQ. In addition to the products of the two metabolic pathways, two other major metabolites were observed: a prominent glycosylated conjugate of PQ on the terminal amine (m/z 422), peaking by 30 min and preferentially generated by (+)-PQ; and the carbamoyl glucuronide of PQ (m/z 480) exclusively generated from (+)-PQ. CONCLUSION: Metabolism of PQ showed enantioselectivity. These findings may provide important information in establishing clinical differences in PQ enantiomers.

Antimalarial and cytotoxic properties of Chukrasia tabularis A. Juss and Turraea vogelii Hook F. Ex. Benth.

Malaria, caused by plasmodium parasite, is at the moment the highest cause of morbidity and mortality in the tropics. Recently, there is increasing efforts to develop more potent antimalarials from plant sources that will have little or no adverse effects. This study is aimed at investigating the in vivo mice antimalarial and in vitro antiplasmodial effects of two Meliaceae plants commonly used in Nigerian ethnomedicine as part of recipe for treating malaria infection: Chukrasia tabularis and Turraea vogelii. Hot water decoction and methanol extract of both plants were evaluated for their antimalarial activity in vivo using the mice model assay and in vitro using the parasite lactate dehydrogenase (pLDH) assay. The extracts were also assessed for toxicity with brine shrimp lethality assay and in mammalian cell lines using the neural red assay. The in vivo mice model antimalarial study showed that the methanol extract of the stem bark of C. tabularis exhibited the highest % chemosuppression (83.65 ± 0.66) at the highest dosage administered (800 mg/kg) when compared with chloroquine diphosphate, the standard reference drug which had a % suppression of 90.36 ± 0.04 (p < 0.05). The in vitro antiplasmodial study indicated that the aqueous extract of the stem bark of C. tabularis displayed good activity against Plasmodium falciparum chloroquine-sensitive (D6) strain (IC50 of 10.739 µg/mL) and chloroquine-resistant (W2) strain. Chloroquine and artemisinin had <0.163 and <0.0264, respectively.

Enantioselective pharmacokinetics of primaquine in healthy human volunteers.

Primaquine (PQ), a racemic drug, is the only treatment available for radical cure of relapsing Plasmodium vivax malaria and blocking transmission of P. falciparum malaria. Recent studies have shown differential pharmacologic and toxicologic profiles of individual PQ enantiomers in rodent, dog, and primate animal models. This study was conducted in six healthy adult human volunteers to determine the plasma pharmacokinetic profile of enantiomers of PQ and carboxyprimaquine (cPQ), the major plasma metabolite. The individuals were orally administered PQ diphosphate, equivalent to 45-mg base, 30 minutes after a normal breakfast. Blood samples were collected at different time intervals, and plasma samples were analyzed for enantiomers of PQ and cPQ. Plasma PQ concentrations were low and variable for both parent enantiomers and peaked around 2-4 hours. Peak (-)-(R)-PQ concentrations ranged from 121 ng/ml to 221 ng/ml, and peak (+)-(S)-PQ concentrations ranged from 168 ng/ml to 299 ng/ml. The cPQ concentrations were much higher and were surprisingly consistent from subject to subject. Essentially all the cPQ detected in plasma was (-)-cPQ. The peak concentrations of (-)-cPQ were observed at 8 hours (range: 1104-1756 ng/ml); however, very high concentrations were sustained through 24 hours. (+)-cPQ was two orders of magnitude lower than (-)-cPQ, and in a few subjects it was detected but only under the limit of quantification. In vitro studies with primary human hepatocytes also suggested more rapid metabolism of (-)-PQ compared with (+)-PQ. The results suggest more rapid metabolism of (-)-PQ to (-) cPQ compared with (+)-PQ. Alternatively, (+)-PQ or (+)-cPQ could be rapidly converted to another metabolite(s) or distributed to tissues. This is the first clinical report on enantioselective pharmacokinetic profiles of PQ and cPQ and supports further clinical evaluation of individual PQ enantiomers.

Enantioselective metabolism of primaquine by human CYP2D6.

BACKGROUND: Primaquine, currently the only approved drug for the treatment and radical cure of Plasmodium vivax malaria, is still used as a racemic mixture. Clinical use of primaquine has been limited due to haemolytic toxicity in individuals with genetic deficiency in glucose-6-phosphate dehydrogenase. Earlier studies have linked its therapeutic effects to CYP2D6-generated metabolites. The aim of the current study was to investigate the differential generation of the CYP2D6 metabolites by racemic primaquine and its individual enantiomers. METHODS: Stable isotope 13C-labelled primaquine and its two enantiomers were incubated with recombinant cytochrome-P450 supersomes containing CYP2D6 under optimized conditions. Metabolite identification and time-point quantitative analysis were performed using LC-MS/MS. UHPLC retention time, twin peaks with a mass difference of 6, MS-MS fragmentation pattern, and relative peak area with respect to parent compound were used for phenotyping and quantitative analysis of metabolites. RESULTS: The rate of metabolism of (+)-(S)-primaquine was significantly higher (50% depletion of 20 µM in 120 min) compared to (-)-(R)-primaquine (30% depletion) when incubated with CYP2D6. The estimated Vmax (µmol/min/mg) were 0.75, 0.98 and 0.42, with Km (µM) of 24.2, 33.1 and 21.6 for (±)-primaquine, (+)-primaquine and (-)-primaquine, respectively. Three stable mono-hydroxylated metabolites, namely, 2-, 3- and 4-hydroxyprimaquine (2-OH-PQ, 3-OH-PQ, and 4-OH-PQ), were identified and quantified. 2-OH-PQ was preferentially formed from (+)-primaquine in a ratio of 4:1 compared to (-)-primaquine. The racemic (±)-primaquine showed a pattern similar to the (-)-primaquine; 2-OH-PQ accounted for about 15-17% of total CYP2D6-mediated conversion of (+)-primaquine. In contrast, 4-OH-PQ was preferentially formed with (-)-primaquine (5:1), accounting for 22% of the total (-)-primaquine conversion. 3-OH-PQ was generated from both enantiomers and racemate. 5-hydroxyprimaquine was unstable. Its orthoquinone degradation product (twice as abundant in (+)-primaquine compared to (-)-primaquine) was identified and accounted for 18-20% of the CYP2D6-mediated conversion of (+)-primaquine. Other minor metabolites included dihydroxyprimaquine species, two quinone-imine products of dihydroxylated primaquine, and a primaquine terminal alcohol with variable generation from the individual enantiomers. CONCLUSION: The metabolism of primaquine by human CYP2D6 and the generation of its metabolites display enantio-selectivity regarding formation of hydroxylated product profiles. This may partly explain differential pharmacologic and toxicologic properties of primaquine enantiomers.

Pharmacology education in the medical curriculum: Challenges and opportunities for improvement.

The knowledge and application of pharmacology is essential for safe prescribing and administration of drugs. In this narrative review, the challenges to pharmacology education in the medical curricula were broadly identified to include issues around content and pedagogies. The increasing number of approved drugs and drug targets, expanding field of pharmacology and the often-changing treatment guidelines and board-defined competencies can make pharmacology education in the medical curriculum daunting. There has been a consensus around the deployment of innovative medical curricula with emphasis on vertical and horizontal integration. This strategy, effective as it has been, presents new challenges to pharmacology education. As a discipline often perceived by students to be hard-to-learn, the future of pharmacology education must include heavy reliance on active learning strategies. The continuing utilization of problem-based, team-based and case-based learning can be complemented with personalized learning which aims to identify the learning gaps in individual students. Technology-inspired student engagement can foster pharmacology learning and retention. Early exposure to pharmacology from premedical preparation through an enduring across-the-level integration can be an effective way to enhance pharmacology learning in the medical curricula.

Metabolic characterization of the new benzimidazole synthetic opioids - nitazenes.

The recent re-emergence and the increasing popularity of nitazenes, a group of new synthetic opioids (NSO) that belong to the benzimidazole chemical class, has raised public health concerns. As a class of potential opioid analgesic agents whose development was discontinued in the 1960s due to their high potential for abuse, very little is known about their metabolism and physiologic disposition. In the current study, three nitazenes-butonitazene, isotonitazene and protonitaze were incubated in human liver microsomes (HLM), human S9 (HS9) fractions and recombinant cytochrome P450 enzymes. All three nitazenes were rapidly metabolized in both HLM and HS9 with over 95% depletion within 60 min. In HLM, butonitazene, isotonitazene and protonitazene had in vitro intrinsic clearance (CLint) (µL/min/mg protein) values of 309, 221 and 216 respectively compared to 150 of verapamil, the positive control. In HS9, CLint values were 217, 139, and 150 for butonitazene, isotonitazene and protonitazene respectively compared to only 35 for testosterone, the control probe substrate. Putative metabolite identified from this study include products of hydroxylation, desethylation, dealkylation, desethylation followed by dealkylation, and desethylation followed by hydroxylation. The metabolic phenotyping showed CYP2D6, CYP2B6 and CYP2C8 and the major hepatic enzymes responsible for the metabolism of nitazenes. Within 30 min of incubation, CYP2D6 depleted butonitazene (99%), isotonitazene (72%) and butonitazene (100%) significantly. The rapid metabolism of nitazenes may be an important factor in accurate and timely detections and quantitation of the unchanged drugs in human matrices following intoxication or in forensic analysis. The involvement of multiple polymorphic CYPs in their metabolism may play important roles in the susceptibility to intoxication and/or addiction, depending on the activity of the metabolites.

The potential of Sutherlandia frutescens for herb-drug interaction.

In Africa, Sutherlandia frutescens is a popular medicinal herb widely consumed by people living with human immunodeficiency virus/AIDS. Concomitant use with antiretroviral drugs has generated concerns of herb-drug interaction (HDI). This study investigated the inhibitory effects of the crude extracts of S. frutescens on the major cytochrome P450 isozymes with the use of pooled human liver microsomes. Its effect on the metabolic clearance of midazolam using cryopreserved hepatocytes was also monitored. The potential of S. frutescens to inhibit human ATP-binding cassette transporters (P-gp and BCRP) and the human organic anion transporting polypeptide (OATP1B1 and OATP1B3) activity was assessed using cell lines overexpressing the transporter proteins. S. frutescens showed inhibitory potency for CYP1A2 (IC(50) = 41.0 µg/ml), CYP2A6 (IC(50) = 160 µg/ml), CYP2B6 (IC(50) = 20.0 µg/ml), CYP2C8 (IC(50) = 22.4 µg/ml), CYP2C9 (IC(50) = 23.0 µg/ml), CYP2C19 (IC(50) = 35.9 µg/ml), and CYP3A4/5 (IC(50) = 17.5 µg/ml [with midazolam1'-hydroxylation]; IC(50) = 28.3 µg/ml [with testosterone 6ß-hydroxylation]). Time-dependent (irreversible) inhibition by S. frutescens was observed for CYP3A4/5 (K(I) = 296 µg/ml, k(inact) = 0.063 min(-1)) under the conditions of this study. S. frutescens also delays the production of midazolam metabolites in the hepatocytes, decreasing its clearance by 40%. Furthermore, S. frutescens inhibited P-gp (IC(50) = 324.8 µg/ml), OATP1B1 (IC(50) = 10.4 µg/ml), and OATP1B3 (IC(50) = 6.6 µg/ml). The result indicates the potential for HDI between S. frutescens and the substrates of the affected enzymes, if sufficient in vivo concentration of the extract is attained.

The potential of Hypoxis hemerocallidea for herb-drug interaction.

CONTEXT: Aqueous decoction of Hypoxis hemerocallidea Fisch. & C.A. Mey. (Hypoxidaceae) (Hypoxis) is widely consumed in Southern Africa by people living with HIV/AIDS, some of whom are on ARV and other medications. OBJECTIVE: The aim of this study was to investigate the potential of the crude aqueous extracts of Hypoxis to inhibit major forms of CYP450 and transport proteins. MATERIALS AND METHODS: Corms of Hypoxis were water-extracted and incubated (in graded concentrations: 1-100 µg/mL) with human liver microsomes (20 min) to monitor the effects on phenacetin O-deethylation, coumarin 7-hydroxylation, bupropion hydroxylation, paclitaxel 6α-hydroxylation, diclofenac 4'-hydroxylation, S-mephenytoin 4'-hydroxylation, bufuralol 1'-hydroxylation, chlorzoxazone 6-hydroxylation, midazolam 1'-hydroxylation and testosterone 6ß-hydroxylation as markers for the metabolic activities of CYP1A2, 2A6, 2B6, 2C8, 2C9, 2C19, 2D6, 2E1 and 3A4/5, respectively. The generation of metabolites were monitored and quantified with the aid of LC-MS/MS. The potential of the extracts to inhibit human ATP-binding cassette transporter activity was assessed using recombinant MDCKII and LLC-PK1 cells over-expressing human breast cancer resistant protein and human P-glycoprotein , respectively (with Ko143 and cyclosporin A as positive controls). Similar assessment was performed with human organic anion transporting polypeptide (OATP1B1 and OATP1B3) using recombinant HEK293 cells over-expressing OATP1B1 and OATP1B3, respectively (with rifamycin and 10 µM atorvastatin as positive controls). RESULTS: Extracts of Hypoxis inhibited the production of the metabolites of the substrates of the following enzymes (as compared to controls) with the indicated IC50 values (µg/mL): CYP1A2 (120.6), CYP2A6 (210.8), CYP2B6 (98.5), CYP2C8 (195.2), CYP2C9 (156) and CYP3A4/5 (185.4). The inhibition of the uptake activity of OATP1B1 and OATP1B3 were also observed with IC50 values of 93.4 and 244.8 µg/mL, respectively. DISCUSSION: Extract concentrations higher than the estimated IC50 values are achievable in the gastrointestinal tract when traditional doses of Hypoxis are considered. This may have profound effects on presystemic metabolism of the drug substrates. If absorbed, systemic inhibition of metabolic enzymes/transporters by Hypoxis may be expected. CONCLUSION: The result suggests that there is the potential for HDI between Hypoxis and the substrates of the affected enzymes/transporters, if sufficient in vivo concentration of Hypoxis extracts is attained.

Herb-drug Interactions in Neuropsychiatric Pharmacotherapy - A Review of Clinically Relevant Findings.

The management of neuropsychiatric disorders relies heavily on pharmacotherapy. The use of herbal products as complimentary medicine, often concomitantly, is common among patients taking prescription neuropsychiatric drugs. Herb-drug interaction, a clinical consequence of this practice, may jeopardize the success of pharmacotherapy in neuropsychiatry. Besides the wellknown ability of phytochemicals to inhibit and/or induce drug-metabolizing enzymes and transport proteins, several phytoconstituents are capable of exerting pharmacological effects on the central nervous system. This study reviewed the relevant literature and identified 13 commonly used herbal products - celery, echinacea, ginkgo, ginseng, hydroxycut, kava, kratom, moringa, piperine, rhodiola, St. John's wort, terminalia/commiphora ayurvedic mixture and valerian - which have shown clinically relevant interactions with prescription drugs used in the management of neuropsychiatric disorders. The consequent pharmacokinetic and pharmacodynamic interactions with orthodox medications often result in deleterious clinical consequences. This underscores the importance of caution in herb-drug co-medication.

Rare Heterogeneous Adverse Events Associated with mRNA-Based COVID-19 Vaccines: A Systematic Review.

Background: Since the successful development, approval, and administration of vaccines against SARS-CoV-2, the causative agent of COVID-19, there have been reports in the published literature, passive surveillance systems, and other pharmacovigilance platforms of a broad spectrum of adverse events following COVID-19 vaccination. A comprehensive review of the more serious adverse events associated with the Pfizer-BioNTech and Moderna mRNA vaccines is warranted, given the massive number of vaccine doses administered worldwide and the novel mechanism of action of these mRNA vaccines in the healthcare industry. Methods: A systematic review of the literature was conducted to identify relevant studies that have reported mRNA COVID-19 vaccine-related adverse events. Results: Serious and severe adverse events following mRNA COVID-19 vaccinations are rare. While a definitive causal relationship was not established in most cases, important adverse events associated with post-vaccination included rare and non-fatal myocarditis and pericarditis in younger vaccine recipients, thrombocytopenia, neurological effects such as seizures and orofacial events, skin reactions, and allergic hypersensitivities. Conclusions: As a relatively new set of vaccines already administered to billions of people, COVID-19 mRNA-based vaccines are generally safe and efficacious. Further studies on long-term adverse events and other unpredictable reactions in close proximity to mRNA vaccination are required.

Comparative single dose pharmacokinetics and metabolism of racemic primaquine and its enantiomers in human volunteers.

Primaquine (PQ) is a racemic drug used in treatment of malaria for six decades. Recent studies suggest that the two enantiomers of PQ are differentially metabolized in animals, and this results in different pharmacological and toxicological profiles. The current study characterizes the pharmacokinetic (PK) properties, metabolism and tolerability of the individual enantiomers of PQ in healthy human volunteers with normal glucose-6-phosphate dehydrogenase (G6PD) activity. Two cohorts (at two dose levels), each with 18 subjects, participated in three study arms in a crossover fashion: a single dose of the (-)-R enantiomer (RPQ), a single dose of the (+)-S enantiomer (SPQ), and a single dose of racemic PQ (RSPQ). PQ and its key metabolites carboxyprimaquine (cPQ) and PQ-N-carbamoyl glucuronide (PQ-N-CG) were analyzed. Clear differences were observed in PK and metabolism of the two enantiomers. Relative PQ exposure was higher with SPQ as compared to RPQ. PQ maximum plasma concentration (Cmax) and area under the plasma concentration-time curve were higher for SPQ, while the apparent volume of distribution and total body clearance were higher for RPQ. Metabolism of the two enantiomers showed dramatic differences: plasma PQ-N-CG was derived solely from SPQ, while RPQ was much more efficiently converted to cPQ than was SPQ. Cmax of cPQ and PQ-N-CG were 10 and 2 times higher, respectively, than the parent drugs. The study demonstrates that the PK properties of PQ enantiomers show clear differences, and metabolism is highly enantioselective. Such differences in metabolism suggest potentially distinct toxicity profiles in multi-dose regimens, especially in G6PD-deficient subjects.

Sexually Transmitted Neisseria gonorrhoeae Infections-Update on Drug Treatment and Vaccine Development.

Background: Sexually transmitted gonorrhea, caused by the Gram-negative diplococcus Neisseria gonorrhoeae, continues to be a serious global health challenge despite efforts to eradicate it. Multidrug resistance among clinical N. gonorrhoeae isolates has limited treatment options, and attempts to develop vaccines have not been successful. Methods: A search of published literature was conducted, and information extracted to provide an update on the status of therapeutics and vaccine development for gonorrheal infection. Results: Recommended pharmacological treatment for gonorrhea has changed multiple times due to increasing acquisition of resistance to existing antibiotics by N. gonorrhoeae. Only broad-spectrum cephalosporin-based combination therapies are currently recommended for treatment of uncomplicated urogenital and anorectal gonococcal infections. With the reported emergence of ceftriaxone resistance, successful strategies addressing the global burden of gonorrhea must include vaccination. Century-old efforts at developing an effective vaccine against gonorrhea, leading to only four clinical trials, have not yielded any successful vaccine. Conclusions: While it is important to continue to explore new drugs for the treatment of gonorrhea, the historical trend of resistance acquisition suggests that any long-term strategy should include vaccine development. Advanced technologies in proteomics and in silico approaches to vaccine target identification may provide templates for future success.

Clinical Management of COVID-19: A Review of Pharmacological Treatment Options.

Since the outbreak and subsequent declaration of COVID-19 as a global pandemic in March 2020, concerted efforts have been applied by the scientific community to curtail the spread of the disease and find a cure. While vaccines constitute a vital part of the public health strategy to reduce the burden of COVID-19, the management of this disease will continue to rely heavily on pharmacotherapy. This study aims to provide an updated review of pharmacological agents that have been developed and/or repurposed for the treatment of COVID-19. To this end, a comprehensive literature search was conducted using the PubMed, Google Scholar, and LitCovid databases. Relevant clinical studies on drugs used in the management of COVID-19 were identified and evaluated in terms of evidence of efficacy and safety. To date, the FDA has approved three therapies for the treatment of COVID-19 Emergency Use Authorization: convalescent plasma, remdesivir, and casirivimab/imdevimab (REGN-COV2). Drugs such as lopinavir/ritonavir, umifenovir, favipiravir, anakinra, chloroquine, hydroxychloroquine, tocilizumab, interferons, tissue plasminogen activator, intravenous immunoglobulins, and nafamosat have been used off-label with mixed therapeutic results. Adjunctive administration of corticosteroids is also very common. The clinical experience with these approved and repurposed drugs is limited, and data on efficacy for the new indication are not strong. Overall, the response of the global scientific community to the COVID-19 pandemic has been impressive, as evident from the volume of scientific literature elucidating the molecular biology and pathophysiology of SARS-CoV-2 and the approval of three new drugs for clinical management. Reviewed studies have shown mixed data on efficacy and safety of the currently utilized drugs. The lack of standard treatment for COVID-19 has made it difficult to interpret results from most of the published studies due to the risk of attribution error. The long-term effects of drugs can only be assessed after several years of clinical experience; therefore, the efficacy and safety of current COVID-19 therapeutics should continue to be rigorously monitored as part of post-marketing studies.

Quantitative determination of primaquine-5,6-ortho-quinone and carboxyprimaquine-5,6-ortho-quinone in human erythrocytes by UHPLC-MS/MS.

The antimalarial drug primaquine (PQ) causes methemoglobinemia and hemolysis in individuals with a genetic deficiency of glucose 6-phosphate dehydrogenase. Reactive oxygen species (ROS) generated by redox cycling of the metabolite primaquine-5,6-orthoquinone (POQ) in erythrocytes has been attributed to be responsible for the toxicity of PQ. Carboxyprimaquine (CPQ), the major human plasma metabolite of PQ, can also form the analogous carboxyprimaquine-5,6-orthoquinone (CPOQ) metabolite, which can also generate ROS in erythrocytes by redox cycling, thus contributing to the hematotoxicity of this drug. In order to study these pathways and characterize such effects in vivo, methods are needed for characterization and quantification of POQ and CPOQ in human erythrocytes. The purpose of this work was to develop a validated method for the quantitative determination of CPOQ and POQ metabolites in human erythrocytes, suitable for clinical studies of PQ metabolism. Several liquid-liquid extraction methods using different organic solvents had been investigated. The solvent mixture of water-methanol-acetonitrile (9:9:5, v/v) was shown to yield the best results for the two analytes. Chromatographic analysis of POQ and CPOQ in human erythrocytes was achieved on a high strength silica (HSS) column and gradient elution (water and acetonitrile, both containing 0.1% formic acid) by ultra-high-performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS). Quantitative estimation of POQ and CPOQ was executed by monitoring ion pairs of m/z 260.23 > 175.03 and m/z 275.19 > 175.04, respectively. The method, which was validated for precision, accuracy, selectivity, and linearity, was successfully applied for the quantitative determination of POQ and CPOQ, the key metabolites of PQ in human erythrocytes in PQ clinical study.

Pitolisant and Other Histamine-3 Receptor Antagonists-An Update on Therapeutic Potentials and Clinical Prospects.

Background: Besides its well-known role as a peripheral chemical mediator of immune, vascular, and cellular responses, histamine plays major roles in the central nervous system, particularly in the mediation of arousal and cognition-enhancement. These central effects are mediated by the histamine-3 auto receptors, the modulation of which is thought to be beneficial for the treatment of disorders that impair cognition or manifest with excessive daytime sleepiness. Methods: A database search of PubMed, Google Scholar, and clinicaltrials.gov was performed in June 2020. Full-text articles were screened and reviewed to provide an update on pitolisant and other histamine-3 receptor antagonists. Results: A new class of drugs-histamine-3 receptor antagonists-has emerged with the approval of pitolisant for the treatment of narcolepsy with or without cataplexy. At the recommended dose, pitolisant is well tolerated and effective. It has also been evaluated for potential therapeutic benefit in Parkinson disease, epilepsy, attention deficit hyperactivity disorder, Alzheimer's disease, and dementia. Limited studies have shown pitolisant to lack abuse potential which will be a major advantage over existing drug options for narcolepsy. Several histamine-3 receptor antagonists are currently in development for a variety of clinical indications. Conclusions: Although limited clinical studies have been conducted on this new class of drugs, the reviewed literature showed promising results for future additions to the clinical indications of pitolisant, and the expansion of the list of approved drugs in this class for a variety of indications.