Evaluation of the Drug-Drug Interaction Potential of Ensitrelvir Fumaric Acid with Cytochrome P450 3A Substrates in Healthy Japanese Adults.

BACKGROUND: Management of drug-drug interactions (DDIs) for ensitrelvir, a novel 3-chymotrypsin-like protease inhibitor of SARS-CoV-2 infection is crucial. A previous clinical DDI study of ensitrelvir with midazolam, a clinical index cytochrome P450 (CYP) 3A substrate, demonstrated that ensitrelvir given for 5 days orally with a loading/maintenance dose of 750/250 mg acted as a strong CYP3A inhibitor. OBJECTIVES: The objectives of this study were to investigate the effect of ensitrelvir on the pharmacokinetics of CYP3A substrates, dexamethasone, prednisolone and midazolam, and to assess the pharmacokinetics, safety, and tolerability of ensitrelvir following multiple-dose administration of ensitrelvir. METHODS: This was a Phase 1, multicenter, single-arm, open-label study in healthy Japanese adult participants. The effects of multiple doses of ensitrelvir in the fasted state on the pharmacokinetics of dexamethasone, prednisolone, and midazolam were investigated. Ensitrelvir was administered from Day 1 through Day 5, with a loading/maintenance dose of 750/250 mg for the dexamethasone and prednisolone cohorts whereas 375/125 mg for the midazolam cohort. Either dexamethasone, prednisolone, or midazolam was administered alone (Day - 2) or in combination with ensitrelvir (Day 5) in each of the cohorts. Additionally, dexamethasone or prednisolone was administered on Days 9 and 14. The pharmacokinetic parameters of ensitrelvir, dexamethasone, prednisolone, and midazolam were calculated based on their plasma concentration data with non-compartmental analysis. In safety assessments, the nature, frequency, and severity of treatment-emergent adverse events were evaluated and recorded. RESULTS: The area under the concentration-time curve (AUC) ratio of dexamethasone on Day 5 was 3.47-fold compared with the corresponding values for dexamethasone alone on Day - 2 and the effect diminished over time after the last dose of ensitrelvir. No clinically meaningful effect was observed for prednisolone. The AUC ratio of midazolam was 6.77-fold with ensitrelvir 375/125 mg suggesting ensitrelvir at 375/125 mg strongly inhibits CYP3A similar to that at 750/250 mg. No new safety signals with ensitrelvir were reported during the study. CONCLUSION: The inhibitory effect for CYP3A was confirmed after the last dose of ensitrelvir, and the effect diminished over time. In addition, ensitrelvir at 375/125 mg showed CYP3A inhibitory potential similar to that at 750/250 mg. These findings can be used as a clinical recommendation for prescribing ensitrelvir with regard to concomitant medications. CLINICAL TRIAL REGISTRATION: Japan Registry of Clinical Trials identifier: jRCT2031210202.

Nirmatrelvir/ritonavir treatment in SARS-CoV-2 positive kidney transplant recipients - a case series with four patients.

BACKGROUND: Despite vaccination coronavirus disease 2019 (COVID-19)-associated mortality caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) remains high in kidney transplant recipients. Nirmatrelvir is a protease inhibitor with activity against SARS-CoV-2. Nirmatrelvir reduces the risk for mortality and hospitalization, which is approved for treating adults at risk for severe disease. Nirmatrelvir is metabolized by the cytochrome P-450 (CYP) 3A4 isozyme CYP3A4 and is therefore co-administered with the irreversible CYP3A4 inhibitor ritonavir, which results in a drug interaction with tacrolimus. A limited number of patients with nirmatrelvir/ritonavir and tacrolimus therapy after kidney transplantation have been reported to date. It has been reported that tacrolimus was paused during the five-day nirmatrelvir/ritonavir therapy and subtherapeutic tacrolimus levels were observed after finishing nirmatrelvir/ritonavir in two patients. Therefore, optimization of tacrolimus dosing is urgently needed in transplant recipients receiving nirmatrelvir/ritonavir treatment. CASE PRESENTATION: Here, we present our first-hand experience with four patients receiving tacrolimus therapy following kidney transplantation and nirmatrelvir/ritonavir therapy due to COVID-19. Tacrolimus was paused during nirmatrelvir/ritonavir therapy in all patients, which resulted in stable therapeutic tacrolimus levels. Tacrolimus was continued directly after finishing nirmatrelvir/ritonavir to avoid subtherapeutic levels in the first patient treated. This patient received his usual tacrolimus maintenance dose, which resulted in toxic levels. Based on this observation, tacrolimus therapy was continued 24 h after finishing nirmatrelvir/ritonavir treatment at a reduced dose in the subsequent patients. In these patients, therapeutic to supratherapeutic tacrolimus levels were observed despite the therapeutic break and dose reduction. DISCUSSION AND CONCLUSIONS: Based on altered CYP3A4 metabolism, tacrolimus levels have to be closely monitored after treatment with nirmatrelvir/ritonavir. Our study suggests that tacrolimus treatment should be paused during nirmatrelvir/ritonavir medication and be continued 24 h after completing nirmatrelvir/ritonavir therapy at a reduced dose and under close monitoring. Based on the limited number of patients in this study, results must be interpreted with caution.

Nirmatrelvir/ritonavir Use With Tacrolimus in Lung Transplant Recipients: A Single-center Case Series.

BACKGROUND: Limited data and guidelines exist for using nirmatrelvir/ritonavir in solid organ transplant recipients stabilized on tacrolimus for the treatment of mild-to-moderate coronavirus disease. Concern exists regarding the impact of utilizing a 5-d course of nirmatrelvir/ritonavir with calcineurin inhibitors because of significant drug-drug interactions between ritonavir, a potent cytochrome P450 3A inhibitor, and other cytochrome P450 3A substrates, such as tacrolimus. METHODS: We report the successful use of nirmatrelvir/ritonavir in 12 outpatient lung transplant recipients with confirmed severe acute respiratory syndrome coronavirus 2 infection stabilized on tacrolimus immunosuppression. All patients stopped tacrolimus and started nirmatrelvir/ritonavir 10 to 14 h after the last dose of tacrolimus. Tacrolimus was withheld and then reinitiated at a modified dose 48 h following the completion of nirmatrelvir/ritonavir therapy. Tacrolimus trough levels were checked during nirmatrelvir/ritonavir therapy and tacrolimus reinitiation. RESULTS: Ten (10/12) patients were able to resume their original tacrolimus dose within 4 d of completing nirmatrelvir/ritonavir therapy and maintain therapeutic levels of tacrolimus. No patients experienced tacrolimus toxicity or acute rejection during the 30-d postcompletion of nirmatrelvir/ritonavir therapy. CONCLUSIONS: In this cohort of lung transplant recipients on tacrolimus, we demonstrated that nirmatrelvir/ritonavir can be safely used with close monitoring of tacrolimus levels and appropriate dose adjustments of tacrolimus. Further confirmatory studies are needed to determine the appropriate use of therapeutic drug monitoring and tacrolimus dose following completion of nirmatrelvir/ritonavir in the solid organ transplant population.

Prevalence of Potential Drug-drug Interactions With Ritonavir-containing COVID-19 Therapy in the United States: An Analysis of the National Health and Nutrition Examination Survey.

PURPOSE: The evolving epidemiology and treatment landscape of COVID-19 necessitates research into potential drug-drug interactions (pDDIs) from the use of new treatments for COVID-19, particularly those that contain ritonavir, a potent inhibitor of the cytochrome P350 3A4 (CYP3A4) metabolic pathway. In this study, we assessed the prevalence of pDDIs between medications for chronic conditions metabolized through the CYP3A4 metabolic pathway and ritonavir-containing COVID-19 medications in the US general population. METHODS: This study combined National Health and Nutrition Examination Survey (NHANES) waves 2015 to 2016 and 2017 to March 2020 to observe pDDI prevalence between ritonavir-containing therapy and coadministered medications among US adults 18 years or older. CYP3A4-mediated medications were identified from affirmative medication questionnaire response and associated prescription examination by surveyors. CYP3A4-mediated medications with associated pDDIs with ritonavir and assessed pDDI severity (minor, major, moderate, and severe) were obtained from the University of Liverpool's COVID-19 online drug interaction checker, Lexicomp, and US Food and Drug Administration fact sheets. pDDI prevalence and severity were evaluated by demographic characteristics and COVID-19 risk factors. FINDINGS: A total of 15,685 adult participants were identified during the 2015 to 2020 NHANES waves. Survey participants used a mean (SD) of 2.7 (1.8) drugs with likelihood of a pDDI. The weighted prevalence of major to contraindicated pDDIs among the US population was 29.3%. Prevalence rates among those 60 years and older, with serious heart conditions, with moderate chronic kidney disease (CKD), with severe CKD, with diabetes, and with HIV were 60.2%, 80.7%, 73.9%, 69.5%, 63.4%, and 68.5%, respectively. Results remained largely unchanged after removal of statins from the list of drugs associated with ritonavir-based pDDIs. IMPLICATIONS: Approximately one-third of the US population would be at risk for a major or contraindicated pDDI should they receive a ritonavir-containing regimen, and this risk increases significantly among individuals 60 years or older and with comorbidities such as serious heart conditions, CKD, diabetes, and HIV. The state of polypharmacy in the US population and the quickly changing COVID-19 landscape indicate significant risk of pDDIs among those requiring treatment with ritonavir-containing COVID-19 medications. Practitioners should take polypharmacy, age, and comorbidity profile into account when prescribing COVID-19 therapies. Alternative treatment regimens should be considered, especially for those of older age and those with risk factors for progression to severe COVID-19.

Nirmatrelvir combined with ritonavir for preventing and treating COVID-19.

BACKGROUND: Oral nirmatrelvir/ritonavir (Paxlovid®) aims to avoid severe COVID-19 in asymptomatic people or those with mild symptoms, thereby decreasing hospitalization and death. Due to its novelty, there are currently few published study results. It remains to be evaluated for which indications and patient populations the drug is suitable.  OBJECTIVES: To assess the efficacy and safety of nirmatrelvir/ritonavir (Paxlovid®) plus standard of care compared to standard of care with or without placebo, or any other intervention for treating COVID-19 and for preventing SARS-CoV-2 infection. To explore equity aspects in subgroup analyses. To keep up to date with the evolving evidence base using a living systematic review (LSR) approach and make new relevant studies available to readers in-between publication of review updates. SEARCH METHODS: We searched the Cochrane COVID-19 Study Register, Scopus, and WHO COVID-19 Global literature on coronavirus disease database, identifying completed and ongoing studies without language restrictions and incorporating studies up to 11 July 2022.  This is a LSR. We conduct monthly update searches that are being made publicly available on the open science framework (OSF) platform. SELECTION CRITERIA: Studies were eligible if they were randomized controlled trials (RCTs) comparing nirmatrelvir/ritonavir plus standard of care with standard of care with or without placebo, or any other intervention for treatment of people with confirmed COVID-19 diagnosis, irrespective of disease severity or treatment setting, and for prevention of SARS-CoV-2 infection. We screened all studies for research integrity. Studies were ineligible if they had been retracted, or if they were not prospectively registered including appropriate ethics approval. DATA COLLECTION AND ANALYSIS: We followed standard Cochrane methodology and used the Cochrane risk of bias 2 tool. We rated the certainty of evidence using the GRADE approach for the following outcomes: 1. to treat outpatients with mild COVID-19; 2. to treat inpatients with moderate-to-severe COVID-19: mortality, clinical worsening or improvement, quality of life, (serious) adverse events, and viral clearance; 3. to prevent SARS-CoV-2 infection in post-exposure prophylaxis (PEP); and 4. pre-exposure prophylaxis (PrEP) scenarios: SARS-CoV-2 infection, development of COVID-19 symptoms, mortality, admission to hospital, quality of life, and (serious) adverse events. We explored inequity by subgroup analysis for elderly people, socially-disadvantaged people with comorbidities, populations from LICs and LMICs, and people from different ethnic and racial backgrounds. MAIN RESULTS: As of 11 July 2022, we included one RCT with 2246 participants in outpatient settings with mild symptomatic COVID-19 comparing nirmatrelvir/ritonavir plus standard of care with standard of care plus placebo. Trial participants were unvaccinated, without previous confirmed SARS-CoV-2 infection, had a symptom onset of no more than five days before randomization, and were at high risk for progression to severe disease. Prohibited prior or concomitant therapies included medications highly dependent on CYP3A4 for clearance and CYP3A4 inducers.  We identified eight ongoing studies. Nirmatrelvir/ritonavir for treating COVID-19 in outpatient settings with asymptomatic or mild disease For the specific population of unvaccinated, high-risk patients nirmatrelvir/ritonavir plus standard of care compared to standard of care plus placebo may reduce all-cause mortality at 28 days (risk ratio (RR) 0.04, 95% confidence interval (CI) 0.00 to 0.68; 1 study, 2224 participants; estimated absolute effect: 11 deaths per 1000 people receiving placebo compared to 0 deaths per 1000 people receiving nirmatrelvir/ritonavir; low-certainty evidence, and admission to hospital or death within 28 days (RR 0.13, 95% CI 0.07 to 0.27; 1 study, 2224 participants; estimated absolute effect: 61 admissions or deaths per 1000 people receiving placebo compared to eight admissions or deaths per 1000 people receiving nirmatrelvir/ritonavir; low-certainty evidence). Nirmatrelvir/ritonavir plus standard of care may reduce serious adverse events during the study period compared to standard of care plus placebo (RR 0.24, 95% CI 0.15 to 0.41; 1 study, 2224 participants; low-certainty evidence). Nirmatrelvir/ritonavir plus standard of care probably has little or no effect on treatment-emergent adverse events (RR 0.95, 95% CI 0.82 to 1.10; 1 study, 2224 participants; moderate-certainty evidence), and probably increases treatment-related adverse events such as dysgeusia and diarrhoea during the study period compared to standard of care plus placebo (RR 2.06, 95% CI 1.44 to 2.95; 1 study, 2224 participants; moderate-certainty evidence). Nirmatrelvir/ritonavir plus standard of care probably decreases discontinuation of study drug due to adverse events compared to standard of care plus placebo (RR 0.49, 95% CI 0.30 to 0.80; 1 study, 2224 participants; moderate-certainty evidence). No study results were identified for improvement of clinical status, quality of life, and viral clearance.  Subgroup analyses for equity Most study participants were younger than 65 years (87.1% of the : modified intention to treat (mITT1) population with 2085 participants), of white ethnicity (71.5%), and were from UMICs or HICs (92.1% of study centres). Data on comorbidities were insufficient.  The outcome 'admission to hospital or death' was investigated for equity: age (< 65 years versus ≥ 65 years) and ethnicity (Asian versus Black versus White versus others). There was no difference between subgroups of age. The effects favoured treatment with nirmatrelvir/ritonavir for the White ethnic group. Estimated effects in the other ethnic groups included the line of no effect (RR = 1). No subgroups were reported for comorbidity status and World Bank country classification by income level. No subgroups were reported for other outcomes. Nirmatrelvir/ritonavir for treating COVID-19 in inpatient settings with moderate to severe disease No studies available. Nirmatrelvir/ritonavir for preventing SARS-CoV-2 infection (PrEP and PEP) No studies available. AUTHORS' CONCLUSIONS: There is low-certainty evidence that nirmatrelvir/ritonavir reduces the risk of all-cause mortality and hospital admission or death based on one trial investigating unvaccinated COVID-19 participants without previous infection that were at high risk and with symptom onset of no more than five days. There is low- to moderate-certainty evidence that nirmatrelvir/ritonavir is safe in people without prior or concomitant therapies including medications highly dependent on CYP3A4. Regarding equity aspects, except for ethnicity, no differences in effect size and direction were identified. No evidence is available on nirmatrelvir/ritonavir to treat hospitalized people with COVID-19 and to prevent a SARS-CoV-2 infection. We will continually update our search and make search results available on OSF.

The inhibitory and inducing effects of ritonavir on hepatic and intestinal CYP3A and other drug-handling proteins.

Ritonavir, originally developed as HIV protease inhibitor, is widely used as a booster in several HIV pharmacotherapy regimens and more recently in Covid-19 treatment (e.g., Paxlovid). Its boosting capacity is due to the highly potent irreversible inhibition of the cytochrome P450 (CYP) 3 A enzyme, thereby enhancing the plasma exposure to coadministered drugs metabolized by CYP3A. Typically used booster doses of ritonavir are 100-200 mg once or twice daily. This review aims to address several aspects of this booster drug, including the possibility to use lower ritonavir doses, 20 mg for instance, resulting in partial CYP3A inactivation in patients. If complete CYP3A inhibition is not needed, lower ritonavir doses could be used, thereby reducing unwanted side effects. In this context, there are contradictory reports on the actual recovery time of CYP3A activity after ritonavir discontinuation, but probably this will take at least one day. In addition to ritonavir's CYP3A inhibitory effect, it can also induce and/or inhibit other CYP enzymes and drug transporters, albeit to a lesser extent. Although ritonavir thus exhibits gene induction capacities, with respect to CYP3A activity the inhibition capacity clearly predominates. Another potent CYP3A inhibitor, the ritonavir analog cobicistat, has been reported to lack the ability to induce enzyme and transporter genes. This might result in a more favorable drug-drug interaction profile compared to ritonavir, although the actual benefit appears to be limited. Indeed, ritonavir is still the clinically most used pharmacokinetic enhancer, indicating that its side effects are well manageable, even in chronic administration regimens.

Safety, Tolerability, and Pharmacokinetics of the Novel Antiviral Agent Ensitrelvir Fumaric Acid, a SARS-CoV-2 3CL Protease Inhibitor, in Healthy Adults.

Ensitrelvir is a novel selective inhibitor of the 3C-like protease of SARS-CoV-2, which is essential for viral replication. This phase 1 study of ensitrelvir assessed its safety, tolerability, and pharmacokinetics of single (part 1, n = 50) and multiple (part 2, n = 33) ascending oral doses. Effect of food on the pharmacokinetics of ensitrelvir, differences in pharmacokinetics of ensitrelvir between Japanese and white participants, and effect of ensitrelvir on the pharmacokinetics of midazolam (a cytochrome P450 3A [CYP3A] substrate) were also assessed. In part 1, Japanese participants were randomized to placebo or ensitrelvir at doses of 20, 70, 250, 500, 1,000, or 2,000 mg. In part 2, Japanese and white participants were randomized to placebo or once-daily ensitrelvir at loading/maintenance dose 375/125 mg or 750/250 mg for 5 days. Most treatment-related adverse events observed were mild in severity and were resolved without treatment. Plasma exposures showed almost dose proportionality, and geometric mean half-life of ensitrelvir following the single dose was 42.2 to 48.1 h. Food intake reduced Cmax and delayed Tmax of ensitrelvir but did not impact the area under the curve (AUC), suggesting suitability for administration without food restriction. Compared with Japanese participants, plasma exposures were slightly lower for white participants. Ensitrelvir affected the pharmacokinetics of CYP3A substrates because of increase in AUC of midazolam coadministered with ensitrelvir 750/250 mg on day 6. In conclusion, ensitrelvir was well-tolerated and demonstrated favorable pharmacokinetics, including a long half-life, supporting once-daily oral dosing. These results validate further assessments of ensitrelvir in participants with SARS-CoV-2 infection.

The Mechanism-Based Inactivation of CYP3A4 by Ritonavir: What Mechanism?

Ritonavir is the most potent cytochrome P450 (CYP) 3A4 inhibitor in clinical use and is often applied as a booster for drugs with low oral bioavailability due to CYP3A4-mediated biotransformation, as in the treatment of HIV (e.g., lopinavir/ritonavir) and more recently COVID-19 (Paxlovid or nirmatrelvir/ritonavir). Despite its clinical importance, the exact mechanism of ritonavir-mediated CYP3A4 inactivation is still not fully understood. Nonetheless, ritonavir is clearly a potent mechanism-based inactivator, which irreversibly blocks CYP3A4. Here, we discuss four fundamentally different mechanisms proposed for this irreversible inactivation/inhibition, namely the (I) formation of a metabolic-intermediate complex (MIC), tightly coordinating to the heme group; (II) strong ligation of unmodified ritonavir to the heme iron; (III) heme destruction; and (IV) covalent attachment of a reactive ritonavir intermediate to the CYP3A4 apoprotein. Ritonavir further appears to inactivate CYP3A4 and CYP3A5 with similar potency, which is important since ritonavir is applied in patients of all ethnicities. Although it is currently not possible to conclude what the primary mechanism of action in vivo is, it is unlikely that any of the proposed mechanisms are fundamentally wrong. We, therefore, propose that ritonavir markedly inactivates CYP3A through a mixed set of mechanisms. This functional redundancy may well contribute to its overall inhibitory efficacy.

In Silico Prediction of the Metabolic Resistance of Vitamin D Analogs against CYP3A4 Metabolizing Enzyme.

The microsomal cytochrome P450 3A4 (CYP3A4) and mitochondrial cytochrome P450 24A1 (CYP24A1) hydroxylating enzymes both metabolize vitamin D and its analogs. The three-dimensional (3D) structure of the full-length native human CYP3A4 has been solved, but the respective structure of the main vitamin D hydroxylating CYP24A1 enzyme is unknown. The structures of recombinant CYP24A1 enzymes have been solved; however, from studies of the vitamin D receptor, the use of a truncated protein for docking studies of ligands led to incorrect results. As the structure of the native CYP3A4 protein is known, we performed rigid docking supported by molecular dynamic simulation using CYP3A4 to predict the metabolic conversion of analogs of 1,25-dihydroxyvitamin D2 (1,25D2). This is highly important to the design of novel vitamin D-based drug candidates of reasonable metabolic stability as CYP3A4 metabolizes ca. 50% of the drug substances. The use of the 3D structure data of human CYP3A4 has allowed us to explain the substantial differences in the metabolic conversion of the side-chain geometric analogs of 1,25D2. The calculated free enthalpy of the binding of an analog of 1,25D2 to CYP3A4 agreed with the experimentally observed conversion of the analog by CYP24A1. The metabolic conversion of an analog of 1,25D2 to the main vitamin D hydroxylating enzyme CYP24A1, of unknown 3D structure, can be explained by the binding strength of the analog to the known 3D structure of the CYP3A4 enzyme.

Oral Drugs Against COVID-19.

BACKGROUND: Five-day oral therapies against early COVID-19 infection have recently been conditionally approved in Europe. In the drug combination nirmatrelvir + ritonavir (nirmatrelvir/r), the active agent, nirmatrelvir, is made bioavailable in clinically adequate amounts by the additional administration of a potent inhibitor of its first-pass metabolism by way of cytochrome P450 [CYP] 3A in the gut and liver. In view of the central role of CYP3A in the clearance of many different kinds of drugs, and the fact that many patients with COVID-19 are taking multiple drugs to treat other conditions, it is important to assess the potential for drug interactions when nirmatrelvir/r is given, and to minimize the risks associated with such interactions. METHODS: We defined the interaction profile of ritonavir on the basis of information derived from two databases (Medline, GoogleScholar), three standard electronic texts on drug interactions, and manufacturer-supplied drug information. We compiled a list of drugs and their potentially relevant interactions, developed a risk min - imization algorithm, and applied it to the substances in question. We also compiled a list of commonly prescribed drugs for which there is no risk of interaction with nirmatrelvir/r. RESULTS: Out of 190 drugs and drug combinations, 57 do not need any special measures when given in combination with brief, low-dose ritonavir treatment, while 15 require dose modification or a therapeutic alternative, 8 can be temporarily discontinued, 9 contraindicate ritonavir use, and 102 should preferably be combined with a different treatment. CONCLUSION: We have proposed measures that are simple to carry out for the main types of drug that can interact with ritonavir. These measures can be implemented under quarantine conditions before starting a 5-day treatment with nirmatrelvir/r.

Interactions of the protease inhibitor, ritonavir, with common anesthesia drugs.

The protease inhibitor, ritonavir, is a strong inhibitor of CYP 3A. The drug is used for management of the human immunovirus and is currently part of an oral antiviral drug combination (nirmatrelvir-ritonavir) for the early treatment of SARS-2 COVID-19-positive patients aged 12 years and over who have recognized comorbidities. The CYP 3A enzyme system is responsible for clearance of numerous drugs used in anesthesia (e.g., alfentanil, fentanyl, methadone, rocuronium, bupivacaine, midazolam, ketamine). Ritonavir will have an impact on drug clearances that are dependent on ritonavir concentration, anesthesia drug intrinsic hepatic clearance, metabolic pathways, concentration-response relationship, and route of administration. Drugs with a steep concentration-response relationship (ketamine, midazolam, rocuronium) are mostly affected because small changes in concentration have major changes in effect response. An increase in midazolam concentration is observed after oral administration because CYP 3A in the gastrointestinal wall is inhibited, causing a large increase in relative bioavailability. Fentanyl infusion may be associated with a modest increase in plasma concentration and effect, but the large between subject variability of pharmacokinetic and pharmacodynamic concentration changes suggests it will have little impact on an individual patient, especially when used with adverse effect monitoring. It has been proposed that drugs that have no or only a small metabolic pathway involving the CYP 3A enzyme be used during anesthesia, for example, propofol, atracurium, remifentanil, and the volatile agents. That anesthesia approach denies children of drugs with considerable value. It is better that the inhibitory changes in clearance of these drugs are understood so that rational drug choices can be made to tailor drug use to the individual patient. Altered drug dose, anticipation of duration of effect, timing of administration, use of reversal agents and perioperative monitoring would better behoove children undergoing anesthesia.

Impact of Inflammation on Midazolam Metabolism in Severe COVID-19 Patients.

Midazolam is a benzodiazepine frequently used for sedation in patients hospitalized in the intensive care unit (ICU) for coronavirus disease 2019 (COVID-19). This drug is primarily metabolized by cytochrome P450 3A (CYP3A) isoenzymes. Several studies have suggested that inflammation, frequently observed in these patients, could modulate CYP3A activity. The objective of this work was to study the impact of inflammation on midazolam pharmacokinetics in patients with COVID-19. Forty-eight patients hospitalized in the ICU for COVID-19 and treated with midazolam administered by continuous infusion were included in this study. Midazolam and α-hydroxymidazolam concentrations were measured and patient data, including the use of CYP3A inhibitors, were collected. Total and unbound concentrations of midazolam and α-hydroxymidazolam were measured in plasma using a validated liquid-chromatography coupled with mass spectrometry method. Inflammatory condition was evaluated by C-reactive protein (CRP) level measurement. Both drug concentrations and CRP measurements were performed on 354 plasma samples. CRP elevation was significantly associated with the α-hydroxymidazolam/midazolam plasma ratio decrease, whether for the unbound fraction or for the total fraction. Conversely, inflammation was not associated with protein binding modifications. Logically, α-hydroxymidazolam/midazolam plasma ratio was significantly reduced when patients were treated with CYP3A inhibitors. In this study, we showed that inflammation probably reduces the metabolism of midazolam by CYP3A. These results suggest that molecules with narrow therapeutic margins and metabolized by CYP3A should be administrated with care in case of massive inflammatory situations.

Interplay of Vitamin D and CYP3A4 Polymorphisms in Endocrine Disorders and Cancer.

Vitamin D has received considerable optimistic attention as a potentially important factor in many pathological states over the past few decades. However, the proportion of the active form of vitamin D metabolites responsible for biological activity is highly questionable in disease states due to flexible alterations in the enzymes responsible for their metabolism. For instance, CYP3A4 plays a crucial role in the biotransformation of vitamin D and other drug substances. Food-drug and/or drug-drug interactions, the disease state, genetic polymorphism, age, sex, diet, and environmental factors all influence CYP3A4 activity. Genetic polymorphisms in CYP450-encoding genes have received considerable attention in the past few decades due to their extensive impact on the pharmacokinetic and dynamic properties of drugs and endogenous substances. In this review, we focused on CYP3A4 polymorphisms and their interplay with vitamin D metabolism and summarized the role of vitamin D in calcium homeostasis, bone diseases, diabetes, cancer, other diseases, and drug substances. We also reviewed clinical observations pertaining to CYP3A4 polymorphisms among the aforementioned disease conditions. In addition, we highlighted the future perspectives of studying the pharmacogenetics of CYP3A4, which may have potential clinical significance for developing novel diagnostic genetic markers that will ascertain disease risk and progression.

In vitro characterization of the furin inhibitor MI-1851: Albumin binding, interaction with cytochrome P450 enzymes and cytotoxicity.

The substrate-analog furin inhibitor MI-1851 can suppress the cleavage of SARS-CoV-2 spike protein and consequently produces significant antiviral effect on infected human airway epithelial cells. In this study, the interaction of inhibitor MI-1851 was examined with human serum albumin using fluorescence spectroscopy and ultrafiltration techniques. Furthermore, the impacts of MI-1851 on human microsomal hepatic cytochrome P450 (CYP) 1A2, 2C9, 2C19, 2D6 and 3A4 activities were assessed based on fluorometric assays. The inhibitory action was also examined on human recombinant CYP3A4 enzyme and on hepatocytes. In addition, microsomal stability (60 min) and cytotoxicity were tested as well. MI-1851 showed no relevant interaction with human serum albumin and was significantly depleted by human microsomes. Furthermore, it did not inhibit CYP1A2, 2C9, 2C19 and 2D6 enzymes. In human hepatocytes, CYP3A4 was significantly suppressed by MI-1851 and weak inhibition was noticed in regard to human microsomes and human recombinant CYP3A4. Finally, MI-1851 did not impair the viability and the oxidative status of primary human hepatocytes (up to 100 µM concentration). Based on these observations, furin inhibitor MI-1851 appears to be potential drug candidates in the treatment of COVID-19, due to the involvement of furin in S protein priming and thus activation of the pandemic SARS-CoV-2.

Disposition of Nirmatrelvir, an Orally Bioavailable Inhibitor of SARS-CoV-2 3C-Like Protease, across Animals and Humans.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) 3C-like protease inhibitor PF-07321332 (nirmatrelvir), in combination with ritonavir (Paxlovid), was recently granted emergency use authorization by multiple regulatory agencies for the treatment of coronavirus disease 2019 (COVID-19) in adults and pediatric patients. Disposition studies on nirmatrelvir in animals and in human reagents, which were used to support clinical studies, are described herein. Plasma clearance was moderate in rats (27.2 ml/min per kg) and monkeys (17.1 ml/min per kg), resulting in half-lives of 5.1 and 0.8 hours, respectively. The corresponding oral bioavailability was moderate in rats (34%-50%) and low in monkeys (8.5%), primarily due to oxidative metabolism along the gastrointestinal tract in this species. Nirmatrelvir demonstrated moderate plasma protein binding in rats, monkeys, and humans with mean unbound fractions ranging from 0.310 to 0.478. The metabolism of nirmatrelvir was qualitatively similar in liver microsomes and hepatocytes from rats, monkeys, and humans; prominent metabolites arose via cytochrome P450 (CYP450)-mediated oxidations on the P1 pyrrolidinone ring, P2 6,6-dimethyl-3-azabicyclo[3.1.0]hexane, and the tertiary-butyl group at the P3 position. Reaction phenotyping studies in human liver microsomes revealed that CYP3A4 was primarily responsible (fraction metabolized = 0.99) for the oxidative metabolism of nirmatrelvir. Minor clearance mechanisms involving renal and biliary excretion of unchanged nirmatrelvir were also noted in animals and in sandwich-cultured human hepatocytes. Nirmatrelvir was a reversible and time-dependent inhibitor as well as inducer of CYP3A activity in vitro. First-in-human pharmacokinetic studies have demonstrated a considerable boost in the oral systemic exposure of nirmatrelvir upon coadministration with the CYP3A4 inhibitor ritonavir, consistent with the predominant role of CYP3A4 in nirmatrelvir metabolism. SIGNIFICANCE STATEMENT: The manuscript describes the preclinical disposition, metabolism, and drug-drug interaction potential of PF-07321332 (nirmatrelvir), an orally active peptidomimetic-based inhibitor of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) 3CL protease, which has been granted emergency use authorization by multiple regulatory agencies around the globe for the treatment of coronavirus disease 2019 (COVID-19) in COVID-19-positive adults and pediatric patients who are at high risk for progression to severe COVID-19, including hospitalization or death.

Physiologically-Based Pharmacokinetic-Led Guidance for Patients With Cystic Fibrosis Taking Elexacaftor-Tezacaftor-Ivacaftor With Nirmatrelvir-Ritonavir for the Treatment of COVID-19.

Cystic fibrosis transmembrane conductance regulator (CFTR) modulating therapies, including elexacaftor-tezacaftor-ivacaftor, are primarily eliminated through cytochrome P450 (CYP) 3A-mediated metabolism. This creates a therapeutic challenge to the treatment of coronavirus disease 2019 (COVID-19) with nirmatrelvir-ritonavir in people with cystic fibrosis (CF) due to the potential for significant drug-drug interactions (DDIs). However, the population with CF is more at risk of serious illness following COVID-19 infection and hence it is important to manage the DDI risk and provide treatment options. CYP3A-mediated DDI of elexacaftor-tezacaftor-ivacaftor was evaluated using a physiologically-based pharmacokinetic modeling approach. Modeling was performed incorporating physiological information and drug-dependent parameters of elexacaftor-tezacaftor-ivacaftor to predict the effect of ritonavir (the CYP3A inhibiting component of the combination) on the pharmacokinetics of elexacaftor-tezacaftor-ivacaftor. The elexacaftor-tezacaftor-ivacaftor models were verified using independent clinical pharmacokinetic and DDI data of elexacaftor-tezacaftor-ivacaftor with a range of CYP3A modulators. When ritonavir was administered on Days 1 through 5, the predicted area under the curve (AUC) ratio of ivacaftor (the most sensitive CYP3A substrate) on Day 6 was 9.31, indicating that its metabolism was strongly inhibited. Based on the predicted DDI, the dose of elexacaftor-tezacaftor-ivacaftor should be reduced when coadministered with nirmatrelvir-ritonavir to elexacaftor 200 mg-tezacaftor 100 mg-ivacaftor 150 mg on Days 1 and 5, with delayed resumption of full-dose elexacaftor-tezacaftor-ivacaftor on Day 9, considering the residual inhibitory effect of ritonavir as a mechanism-based inhibitor. The simulation predicts a regimen of elexacaftor-tezacaftor-ivacaftor administered concomitantly with nirmatrelvir-ritonavir in people with CF that will likely decrease the impact of the drug interaction.

Hyperinflammation Reduces Midazolam Metabolism in Critically Ill Adults with COVID-19.

BACKGROUND AND OBJECTIVE: Many patients treated for COVID-19 related acute respiratory distress syndrome in the intensive care unit are sedated with the benzodiazepine midazolam. Midazolam undergoes extensive metabolism by CYP3A enzymes, which may be inhibited by hyperinflammation. Therefore, an exaggerated proinflammatory response, as often observed in COVID-19, may decrease midazolam clearance. To develop a population pharmacokinetic model for midazolam in adult intensive care unit patients infected with COVID-19 and to assess the effect of inflammation, reflected by IL-6, on the pharmacokinetics of midazolam. METHODS: Midazolam blood samples were collected once a week between March 31 and April 30 2020. Patients were excluded if they concomitantly received CYP3A4 inhibitors, CYP3A4 inducers and/or continuous renal replacement therapy. Midazolam and metabolites were analyzed with an ultra-performance liquid chromatography-tandem mass spectrometry method. A population pharmacokinetic model was developed, using nonlinear mixed effects modelling. IL-6 and CRP, markers of inflammation, were analyzed as covariates. RESULTS: The data were described by a one-compartment model for midazolam and the metabolites 1-OH-midazolam and 1-OH-midazolam-glucuronide. The population mean estimate for midazolam clearance was 6.7 L/h (4.8-8.5 L/h). Midazolam clearance was reduced by increased IL-6 and IL-6 explained more of the variability within our patients than CRP. The midazolam clearance was reduced by 24% (6.7-5.1 L/h) when IL-6 increases from population median 116 to 300 pg/mL. CONCLUSIONS: Inflammation, reflected by high IL-6, reduces midazolam clearance in critically ill patients with COVID-19. This knowledge may help avoid oversedation, but further research is warranted.

Development and Validation of a Novel HPLC Method to Analyse Metabolic Reaction Products Catalysed by the CYP3A2 Isoform: In Vitro Inhibition of CYP3A2 Enzyme Activity by Aspirin (Drugs Often Used Together in COVID-19 Treatment).

Aspirin (also known as acetylsalicylic acid) is a drug intended to treat fever, pain, or inflammation. Treatment of moderate to severe cases of COVID-19 using aspirin along with dexamethasone has gained major attention globally in recent times. Thus, the purpose of this study was to use High-Performance Liquid Chromatography (HPLC) to evaluate the in vitro inhibition of CYP3A2 enzyme activity using aspirin in rat liver microsomes (RLMs). In this study, an efficient and sensitive HPLC method was developed using a reversed phase C18 column (X Bridge 4.6 mm × 150 mm, 3.5 µm) at 243 nm using acetonitrile and water (70:30 v/v). The linearity (r2 > 0.999), precision (<15%), accuracy and recovery (80-120%), limit of detection (5.60 µM and 0.06 µM), limit of quantification (16.98 µM and 0.19 µM), and stability of the newly developed method were validated for dexamethasone and 6ß-hydroxydexamethasone, respectively, following International Conference on Harmonization (ICH) guidelines. This method was applied in vitro to measure CYP3A2 activity. The results showed that aspirin competitively inhibits 6ß-hydroxylation (CYP3A2 activity) with an inhibition constant (Ki) = 95.46 µM and the concentration of the inhibitor causing 50% inhibition of original enzyme activity (IC50) = 190.92 µM. This indicated that there is a minimal risk of toxicity when dexamethasone and aspirin are co-administrated and a very low risk of toxicity and drug interaction with drugs that are a substrate for CYP3A2 in healthcare settings.

Variation in Expression of Cytochrome P450 3A Isoforms and Toxicological Effects: Endo- and Exogenous Substances as Regulatory Factors and Substrates.

The CYP3A subfamily, which includes isoforms CYP3A4, CYP3A5, and CYP3A7 in humans, plays important roles in the metabolism of various endogenous and exogenous substances. Gene and protein expression of CYP3A4, CYP3A5, and CYP3A7 show large inter-individual differences, which are caused by many endogenous and exogenous factors. Inter-individual differences can cause negative outcomes, such as adverse drug events and disease development. Therefore, it is important to understand the variations in CYP3A expression caused by endo- and exogenous factors, as well as the variation in the metabolism and kinetics of endo- and exogenous substrates. In this review, we summarize the factors regulating CYP3A expression, such as bile acids, hormones, microRNA, inflammatory cytokines, drugs, environmental chemicals, and dietary factors. In addition, variations in CYP3A expression under pathological conditions, such as coronavirus disease 2019 and liver diseases, are described as examples of the physiological effects of endogenous factors. We also summarize endogenous and exogenous substrates metabolized by CYP3A isoforms, such as cholesterol, bile acids, hormones, arachidonic acid, vitamin D, and drugs. The relationship between the changes in the kinetics of these substrates and the toxicological effects in our bodies are discussed. The usefulness of these substrates and metabolites as endogenous biomarkers for CYP3A activity is also discussed. Notably, we focused on discrimination between CYP3A4, CYP3A5, and CYP3A7 to understand inter-individual differences in CYP3A expression and function.