Pharmacokinetics and pharmacodynamics of PTC518, an oral huntingtin lowering splicing modifier: A first-in-human study.

AIMS: PTC518 is an orally administered, centrally and peripherally distributed huntingtin (HTT) pre-mRNA splicing modifier being developed for the treatment of Huntington's disease (HD) for which there is a high unmet medical need as there are currently no approved disease-modifying treatments. This first-in-human study investigated the safety, tolerability, pharmacokinetics (PK), and pharmacodynamics (PD) of PTC518 in healthy volunteers. METHODS: This phase 1, single-centre, randomized study in 77 healthy male and female volunteers evaluated the safety and tolerability and PK of PTC518 following single ascending doses and multiple ascending doses, PD as assessed by HTT mRNA and HTT protein levels after single and multiple doses, and food effects. RESULTS: PTC518 demonstrated a favourable safety profile. The majority of treatment-emergent adverse events were mild and transient. PTC518 Tmax was reached at 6-7 h and the terminal T1/2 was 54.0-75.3 h following a single oral dose. Exposure increased with dose though less than dose proportionally. The PTC518 concentrations in cerebrospinal fluid were approximately 2.6-fold higher than the unbound free-drug concentrations in plasma. A significant dose-dependent reduction of up to approximately 60% in HTT mRNA and a significant dose-dependent, time-dependent and sustained reduction in HTT protein levels of up to 35% were observed after PTC518 treatment. CONCLUSIONS: PTC518 was well tolerated, and proof of mechanism of this novel splicing modifier was demonstrated by the dose-dependent decrease in systemic HTT mRNA and HTT protein levels. Results from this first-in-human study support further studies in patients with HD and demonstrate the potential for PTC518 as a breakthrough treatment for HD.

Absorption, metabolism and excretion of 14C-emvododstat following repeat daily oral dose administration in human volunteers using a combination of microtracer radioactivity and high radioactivity doses.

Emvododstat is a potent inhibitor of dihydroorotate dehydrogenase and is now in clinical development for the treatment of COVID-19 and acute myeloid leukemia. Since the metabolism and pharmacokinetics of emvododstat in humans is time­dependent, a repeat dose study design using a combination of microtracer radioactivity and high radioactivity doses was employed to evaluate the metabolism and excretion of emvododstat near steady state. Seven healthy male subjects each received 16 mg/0.3 µCi 14C-emvododstat daily oral doses for 6 days followed by a 16 mg/100 µCi high radioactivity oral dose on Day 7. Following the last 16 mg/0.3 µCi 14C­emvododstat dose on Day 6, total radioactivity in plasma peaked at 6 h post-dose. Following a high radioactivity oral dose (16 mg/100 µCi) of 14C-emvododstat on Day 7, both whole blood and plasma radioactivity peaked at 6 h, rapidly declined from 6 h to 36 h post-dose, and decreased slowly thereafter with measurable radioactivity at 240 h post-dose. The mean cumulative recovery of the administered dose was 6.0% in urine and 19.9% in feces by 240 h post-dose, and the mean extrapolated recovery to infinity was 37.3% in urine and 56.6% in feces. Similar metabolite profiles were observed after repeat daily microtracer radioactivity oral dosing on Day 6 and after a high radioactivity oral dose on Day 7. Emvododstat was the most abundant circulating component, M443 and O-desmethyl emvododstat glucuronide were the major circulating metabolites; M474 was the most abundant metabolite in urine, while O­desmethyl emvododstat was the most abundant metabolite in feces. Significance Statement This study provides a complete set of the absorption, metabolism and excretion data of emvododstat, a potent inhibitor of dihydroorotate dehydrogenase, at close to steady state in healthy human subjects. Resolution of challenges due to slow metabolism and elimination of a lipophilic compound highlighted in this study can be achieved by repeat daily microtracer radioactivity oral dosing followed by a high radioactivity oral dosing at therapeutically relevant doses.

A clinical pharmacokinetic drug-drug interaction study between dextromethorphan and emvododstat, a potent anti-SARS-CoV-2 dihydroorotate dehydrogenase inhibitor.

PURPOSE: A therapeutic agent that targets both viral replication and the hyper-reactive immune response would offer a highly desirable treatment for severe acute respiratory syndrome corona virus 2 (SARS-CoV-2, coronavirus disease 2019, COVID-19) management. Emvododstat (PTC299; 4-chlorophenyl 6-chloro-1-[4-methoxyphenyl]-1,3, 4,9-tetrahydro-2H-pyrido[3,4-b]indole-2-carboxylate) was found to be a potent inhibitor of immunomodulatory and inflammation-related processes by inhibition of dihydroorotate dehydrogenase to reduce the severity of SARS-CoV-2 infections This drug interaction study was performed to determine if emvododstat was an inhibitor of CYP2D6. METHODS: Potential drug-drug interactions between emvododstat and a CYP2D6 probe substrate (dextromethorphan) were investigated by measuring plasma dextromethorphan and metabolite (dextrorphan) concentrations before and after emvododstat administration. On day 1, 18 healthy subjects received an oral dose of 30 mg dextromethorphan followed by a 4-day washout period. On day 5, subjects received an oral dose of 250 mg emvododstat with food. Two hours later, 30 mg dextromethorphan was administered. RESULTS: When given with emvododstat, plasma dextromethorphan concentrations increased substantially, while metabolite levels (dextrorphan) remained essentially the same. Maximum plasma dextromethorphan concentration (Cmax) increased from 2006 to 5847 pg/mL. Dextromethorphan exposure (AUC) increased from 18,829 to 157,400 h·pg/mL for AUC0-last and from 21,585 to 362,107 h·pg/mL for AUC0-inf following administration of emvododstat. When dextromethorphan parameters were compared before and after emvododstat, least squares mean ratios (90% confidence interval) were found to be 2.9 (2.2, 3.8), 8.4 (6.1, 11.5), and 14.9 (10.0, 22.1) for Cmax, AUC0-last, and AUC0-inf, respectively. CONCLUSION: Emvododstat appears to be a strong CYP2D6 inhibitor. No drug-related treatment emergent adverse effects (TEAEs) were considered to be severe or serious. TRIAL REGISTRATION: EudraCT 2021-004626-29, 11 May 2021.

Absorption, distribution, metabolism and excretion of 14C-vatiquinone in rats, dogs, and human subjects.

Vatiquinone is a potent inhibitor of 15-lipoxygenase and is in clinical development for the treatment of mitochondrial diseases and other disorders characterised by high levels of oxidative stress and dysregulation of energy metabolism.In rats, 14C-vatiquinone-derived radioactivity was quickly and widely distributed throughout the body and cleared from most tissues by 24 h post-dose following a single oral dose of 14C-vatiquinone.Following oral administration, 94% of dose was recovered within seven days in rats, approximately 61% of dose was recovered within seven days in dogs and approximately 93% of dose was recovered within nine days in human subjects (IND 119220). Faecal excretion was the major route (>56% dose) in all species; urinary excretion was minimal in rats and dogs (<3% dose) but was higher in humans (∼ 22% dose).Following oral administration, vatiquinone was the dominant circulating component in rats and dogs but was minor in human subjects. There were no plasma metabolites that were more than 10% of total drug related exposures in all species.Following oral administration, vatiquinone was not detectable in urine but was the most prominent component in faeces in rats, dogs, and humans.

Absorption, distribution, metabolism and excretion of 14C-Emvododstat following a single oral dose in rats and dogs.

Emvododstat is a potent inhibitor of dihydroorotate dehydrogenase and is now in clinical development for the treatment of acute myeloid leukaemia and COVID-19.Following an oral dose administration in Long-Evans rats, 14C-emvododstat-derived radioactivity was widely distributed throughout the body, with the highest distribution in the endocrine, fatty, and secretory tissues and the lowest in central nervous system.Following a single oral dose of 14C-emvododstat in rats, 54.7% of the dose was recovered in faeces while less than 0.4% of dose was recovered in urine 7 days post-dose. Emvododstat was the dominant radioactive component in plasma and faeces.Following a single oral dose of 14C-emvododstat in dogs, 75.2% of the dose was recovered in faeces while 0.5% of dose was recovered in urine 8 days post-dose. Emvododstat was the dominant radioactive component in faeces, while emvododstat and its two metabolites (O-desmethyl emvododstat and emvododstat amide bond hydrolysis product) were the major circulating radioactivity in dog plasma.

In vitro metabolism, pharmacokinetics and drug interaction potentials of emvododstat, a DHODH inhibitor.

Emvododstat was identified as a potent inhibitor of dihydroorotate dehydrogenase and is now in clinical development for the treatment of acute myeloid leukaemia and COVID-19. The objective of this paper is to evaluate the metabolism, pharmacokinetics, and drug interaction potentials of emvododstat.Emvododstat showed high binding to plasma protein with minimal distribution into blood cells in mouse, rat, dog, monkey, and human whole blood.O-Demethylation followed by glucuronidation appeared to be the major metabolic pathway in rat, dog, monkey, and human hepatocytes. CYP2C8, 2C19, 2D6, and 3A4 were involved in O-desmethyl emvododstat metabolite formation. Both emvododstat and O-desmethyl emvododstat inhibited CYP2D6 activity and induced CYP expression to different extents in vitro.Emvododstat and O-desmethyl emvododstat inhibited BCRP transporter activity but did not inhibit bile salt transporters and other efflux or uptake transporters. Neither emvododstat nor O-desmethyl emvododstat was a substrate for common efflux or uptake transporters investigated.Emvododstat is bioavailable in mice, rats, dogs, and monkeys following a single oral dose. The absorption was generally slow with the mean plasma Tmax ranging from 2 to 5 h; plasma exposure of O-desmethyl emvododstat was lower in rodents, but relatively higher in dogs and monkeys.

Metabolism and Disposition of Ataluren after Oral Administration to Mice, Rats, Dogs, and Humans.

Ataluren is a unique small molecule developed for the treatment of diseases caused by nonsense mutations, which result in premature termination of ribosomal translation and lack of full-length protein production. This study investigated the in vivo metabolism and disposition of ataluren in mice, rats, dogs, and humans. After single oral administration of [14C]ataluren, the overall recovery of radioactivity was ≥93.7%, with approximately 39%, 17%-21%, 12%, and 55% in the urine and 54%, 70%-72%, 80%, and 47% in the feces from intact mice, rats, dogs, and humans, respectively. In bile duct-cannulated (BDC) rats, approximately 10%, 7%, and 82% of the dose was recovered in the urine, feces, and bile, respectively, suggesting that biliary secretion was a major route for the elimination of ataluren in the rats. Ataluren was extensively metabolized after oral administration, and the metabolic profiles of ataluren were quantitatively similar across all species. Unchanged ataluren was the dominant radioactive component in plasma. Ataluren acyl glucuronide was the most prominent metabolite in plasma of all species and the dominant metabolite in BDC rat bile and human urine, whereas the oxadiazole cleavage products were the major or prominent metabolites in the feces of all species. Overall, the results indicate that phase I metabolism is negligible and that the pathway largely involves glucuronidation. No other circulatory conjugation metabolite was detected across investigated species. SIGNIFICANCE STATEMENT: Ataluren is a novel carboxylic acid-containing small molecule drug for treating nonsense mutation Duchenne muscular dystrophy. In vivo metabolism and disposition after a single dose of the drug were investigated in mice, rats, dogs, and humans. Phase I metabolism of ataluren was negligible, and the pathway largely involves glucuronidation. No other circulatory conjugation metabolite was detected across investigated species.

Clinical Assessment of Breast Cancer Resistance Protein (BCRP)-Mediated Drug-Drug Interactions of Sepiapterin with Curcumin and Rosuvastatin in Healthy Volunteers.

BACKGROUND AND OBJECTIVE: Sepiapterin, also known as PTC923 and CNSA-001, is a synthetic form of endogenous sepiapterin being developed as a novel oral treatment for phenylketonuria. Sepiapterin is a natural precursor of tetrahydrobiopterin (BH4) and, when orally administered, is converted to BH4 via the pterin salvage pathway. In vitro studies have demonstrated that both sepiapterin and BH4 are both substrates and inhibitors of the breast cancer resistance protein (BCRP) transporter. This phase I study investigated BCRP-mediated drug-drug interactions of sepiapterin as a victim and as a perpetrator. METHODS: An open-label, fixed-sequence, four-period, crossover, single-dose study was conducted in adult male and female healthy volunteers (18-55 years of age). In a given treatment period, subjects received a single oral dose of sepiapterin (20 mg/kg), sepiapterin (20 mg/kg) plus curcumin (2 g), rosuvastatin (10 mg), or rosuvastatin (10 mg) plus sepiapterin (60 mg/kg). The pharmacokinetics of sepiapterin, its metabolite BH4, and rosuvastatin were studied, and geometric mean ratios of exposures in the presence and absence of the BCRP inhibitor curcumin or sepiapterin were estimated. The presence of the BCRP c.421C>A polymorphism was evaluated in all subjects. RESULTS: A total of 29 subjects were enrolled and included in the safety analysis. Among them, 26 subjects were included in the pharmacokinetic and drug-drug interaction analyses. Following oral administration 20 mg/kg sepiapterin, sepiapterin was rapidly and extensively converted to BH4, and BH4 maximum observed concentration (415.0 ng/mL) was observed 4.95 h (time to maximum observed concentration) post-dose. Sepiapterin maximum observed concentration and area under the concentration-time curve from time 0 to time of the last quantifiable measurement or the last sample collection time (AUClast) were <1% of BH4 values. Coadministration of the BCRP inhibitor curcumin (2 g) increased BH4 maximum observed concentration, AUClast, and area under the concentration-time curve from time 0 extrapolated to infinity by 24%, 21%, and 20%, respectively. When sepiapterin was coadministered with the BCRP substrate rosuvastatin, there was no effect on the pharmacokinetics of rosuvastatin. BCRP c.421C/A carriers (n = 4) had higher plasma exposures of BH4 (1.39 × for AUClast) and rosuvastatin (1.61 × for AUClast) than c.421C/C carriers (n = 22). Greater increases in BH4 exposures (1.33 vs 1.18 for AUClast) were observed in c.421C/A carriers compared with c.421C/C carriers when sepiapterin was coadministered with curcumin. All treatments were well tolerated during the study. CONCLUSIONS: Oral coadministration of the BCRP inhibitor curcumin slightly increased the plasma exposure of sepiapterin and its metabolite BH4 in healthy volunteers. This modest increase was deemed not clinically meaningful. Sepiapterin did not alter the pharmacokinetics of the BCRP substrate rosuvastatin.

Effect of Itraconazole, a CYP3A4 Inhibitor, and Rifampin, a CYP3A4 Inducer, on the Pharmacokinetics of Vatiquinone.

Vatiquinone is a small molecule inhibitor of 15-lipoxygenase in development for patients with Friedreich's ataxia. The objective of this analysis was to determine the effect of a cytochrome P450 isoform 3A4 (CYP3A4) inhibitor and inducer on vatiquinone pharmacokinetics (PKs). The coadministration of 400 mg of vatiquinone with 200 mg of itraconazole (a CYP3A4 inhibitor) resulted in increased maximum observed concentration (Cmax) of vatiquinone and systemic exposure (AUC0-inf) by approximately 3.5- and 2.9-fold, respectively. The coadministration of 400 mg of vatiquinone with 600 mg of rifampin (a CYP3A4 inducer) resulted in decreased vatiquinone Cmax and AUC0-inf by approximately 0.64- and 0.54-fold, respectively. The terminal half-life of vatiquinone was not affected by itraconazole or rifampin. These clinical study results confirm the in vitro reaction phenotyping data that shows that CYP3A4 plays an important role in vatiquinone metabolism. The result of this analysis together with phase 3 efficacy and safety data, population PK analysis, and the exposure-response relationship will determine if the extent of vatiquinone changes in the presence of CYP3A4 inhibitors and inducers are considered clinically relevant.

Determination of Vatiquinone Drug-Drug Interactions, as CYP450 Perpetrator and Victim, Using Physiologically Based Pharmacokinetic (PBPK) Modeling and Simulation.

Vatiquinone, a 15-lipoxygenase inhibitor, is in development for patients with Friedreich's ataxia. Physiologically based pharmacokinetic (PBPK) modeling addressed drug-drug interaction gaps without additional studies. A PBPK model (Simcyp Simulator version 21, full model) was developed using parameters obtained from in vitro studies, in silico estimation and optimization, and two clinical studies. A venous blood dosing model best characterized vatiquinone lymphatic absorption. Apparent oral clearance (CL/F) was used to optimize intrinsic clearance (CLint). Intestinal availability (Fg) was estimated using the hybrid flow term (Qgut), unbound fraction in the enterocytes (fugut), and gut intrinsic metabolic clearance (CLuG,int). Renal clearance (CLR) was set to zero. Assuming an Fa of 1, CYP3A4 contribution (fmCYP3A4) was further optimized. The PBPK model was verified with two clinical studies and demonstrated that it adequately characterized vatiquinone PK. As a perpetrator, the model predicted no risk for vatiquinone to significantly alter the drug exposures of CYP3A4 and CYP1A2 substrates as evident bynegligible reduction in both midazolam and caffeine area under the curve (AUC)inf and Cmax. As a victim, the model predicted that vatiquinone exposures are weakly influenced by moderate CYP3A4 inhibitors and inducers. With fluconazole coadministration, vatiquinone AUCinf and Cmax increased by nearly 50% and 25%, respectively. With efavirenz coadministration, vatiquinone AUCinf and Cmax decreased by approximately 20% and 10%, respectively. Results suggested that vatiquinone does not significantly impact CYP3A4 and CYP1A2 substrates and that moderate CYP3A4 inhibitors and inducers weakly impact vatiquinone AUC.

LC-MS/MS methods for direct measurement of sepiapterin and tetrahydrobiopterin in human plasma and clinical applications.

Aim: Tetrahydrobiopterin (BH4), a natural cofactor of aromatic amino acid hydroxylases, and sepiapterin, a natural precursor of BH4, are endogenously present in human plasma. This is the first report on methods for direct quantification of sepiapterin and BH4 in human plasma by LC-MS/MS for pharmacokinetic assessment. Materials & methods: The analytes in plasma were harvested from blood that were treated with 10% ascorbic acid (AA) to a final concentration of 1% AA. Results & conclusion: The quantification methods were validated for calibration ranges of 0.75-500 ng/ml and 0.5-500 ng/ml for sepiapterin and BH4, respectively. Quantification of analytes was challenging due to their susceptibility to redox reactions. The validated methods were utilized successfully to support clinical development of sepiapterin.

Lack of Concentration-QTc Relationship and Cardiac Risk With Vatiquinone Therapeutic and Supratherapeutic Doses.

Vatiquinone, a 15-lipoxygenase inhibitor, is in development for patients with Friedreich ataxia. The study determined the effect of vatiquinone on electrocardiogram parameters. This was a 2-part, single-center, randomized, double-blinded, and placebo-controlled study. Part 1 used an adaptive approach to identify a supratherapeutic dose, while Part 2 evaluated the effect of vatiquinone on Fridericia corrected QT interval (QTcF). A safe and tolerated supratherapeutic dose of 1400 mg was identified. Concentration-QTcF analysis confirmed there was no statistically significant relationship between vatiquinone concentration and QTcF. QTcF effect (ie, ΔΔQTcF) exceeding 10 milliseconds was excluded for concentrations up to approximately 11,500 ng/mL. By-time-point analysis confirmed that least-squares mean ΔΔQTcF was below 10 milliseconds. Largest least-squares mean ΔΔQTcF of 1.5 milliseconds was observed at 2 hours after dosing. Vatiquinone did not have a clinically relevant effect on heart rate nor on cardiac conduction (PR interval and QRS interval). No new safety signals were found, as safety data are consistent with the known safety profile of vatiquinone. These findings altogether demonstrated that there is a minimal cardiac risk for vatiquinone concentrations up to the supratherapeutic dose level.

Relative Oral Bioavailability and Food Effects of Two Sepiapterin Formulations in Healthy Participants.

Sepiapterin is an orally administered drug in development for the treatment of phenylketonuria, an inborn error of metabolism characterized by the deficiency of the phenylalanine-metabolizing enzyme phenylalanine hydroxylase. This study characterized the pharmacokinetics, safety, and tolerability of 2 clinical sepiapterin formulations (Phase 1/2, Phase 3) and the effects of food on the pharmacokinetics of the Phase 3 formulation in healthy participants. In Part A, 18 participants were randomized to one of 2 treatment sequences, each with 4 dosing periods comprising a single dose (20 or 60 mg/kg) of the Phase 1/2 or the Phase 3 formulation with a low-fat diet. In Part B, 14 participants were randomized to one of 2 sequences, each comprising 4 dosing periods of a single dose (20 or 60 mg/kg) of the Phase 3 formulation under fed (high-fat) or fasted conditions. Following oral administration, sepiapterin was quickly absorbed and rapidly and extensively converted to tetrahydrobiopterin (BH4). BH4 was the major circulating active moiety. Under low-fat conditions, the Phase 3 formulation was bioequivalent to the Phase 1/2 formulation at 20 mg/kg, while slightly lower BH4 exposure (approximately 0.81×) for the Phase 3 formulation was observed at 60 mg/kg. BH4 exposure increased to approximately 1.7× under the low-fat condition and approximately 2.8× under the high-fat condition at a dose of either 20 or 60 mg/kg for the Phase 3 formulation, compared with the fasted condition. Both sepiapterin formulations were well tolerated, with no serious or severe adverse events reported. All treatment-emergent adverse events were mild or moderate in severity.

Quantitative Systems Toxicology Modeling Informed Safe Dose Selection of Emvododstat in Acute Myeloid Leukemia Patients.

Clinical investigation of emvododstat for the treatment of solid tumors was halted after two patients who were heavily treated with other anticancer therapies experienced drug-induced liver failure. However, preclinical investigations supported that emvododstat at lower doses might be effective in treating acute myeloid leukemia (AML) and against severe acute respiratory syndrome-coronavirus 2 as a dihydroorotate dehydrogenase inhibitor. Therefore, a quantitative systems toxicology model, DILIsym, was used to predict liver safety of the proposed dosing of emvododstat in AML clinical trials. In vitro mechanistic toxicity data of emvododstat and its desmethyl metabolite were integrated with in vivo exposure within DILIsym to predict hepatotoxicity responses in a simulated human population. DILIsym simulations predicted alanine aminotransferase elevations observed in prior emvododstat clinical trials in patients with solid tumors, but not in the prospective AML clinical trial with the proposed dosing regimens. Exposure predictions based on physiologically-based pharmacokinetic modeling suggested that reduced doses of emvododstat would produce clinical exposures that would be efficacious to treat AML. In the AML clinical trial, only eight patients experienced aminotransferase elevations, all of which were mild (grade 1), all resolving within a short period of time, and no patient showed symptoms of hepatotoxicity, confirming the prospective prediction of liver safety. Overall, retrospective DILIsym simulations adequately predicted the liver safety liabilities of emvododstat in solid tumor trials and prospective simulations predicted the liver safety of reduced doses in an AML clinical trial. The modeling was critical to enabling regulatory approval to proceed with the AML clinical trial wherein the predicted liver safety was confirmed.

Phase Ib Study of Unesbulin (PTC596) Plus Dacarbazine for the Treatment of Locally Recurrent, Unresectable or Metastatic, Relapsed or Refractory Leiomyosarcoma.

PURPOSE: This multicenter, single-arm, open-label, phase Ib study was designed to determine the recommended phase II dose (RP2D) and to evaluate the safety and preliminary efficacy of unesbulin plus dacarbazine (DTIC) in patients with advanced leiomyosarcoma (LMS). PATIENTS AND METHODS: Adult subjects with locally advanced, unresectable or metastatic, relapsed or refractory LMS were treated with escalating doses of unesbulin orally twice per week in combination with DTIC 1,000 mg/m2 intravenously (IV) once every 21 days. The time-to-event continual reassessment method was used to determine the RP2D on the basis of dose-limiting toxicities (DLTs) assessed during the first two 21-day treatment cycles. All explored doses of unesbulin (200 mg up to 400 mg) were in combination with DTIC. An expansion cohort was enrolled to evaluate the safety and efficacy of unesbulin at the RP2D. RESULTS: Unesbulin 300 mg administered orally twice per week in combination with DTIC 1,000 mg/m2 IV once every 21 days was identified as the RP2D. On the basis of data from 27 subjects who were deemed DLT-evaluable, toxicity was higher in the unesbulin 400 mg group, with three of four subjects (75%) experiencing DLTs versus one of four subjects (25%) in the 200 mg group and three of 19 subjects (15.8%) in the 300 mg group. The most commonly reported DLTs and treatment-related grade 3 and 4 adverse events were thrombocytopenia and neutropenia. At the RP2D, seven subjects who were efficacy evaluable achieved partial response for an objective response rate of 24.1%. CONCLUSION: Unesbulin 300 mg twice per week plus DTIC 1,000 mg/m2 once every 21 days was identified as the RP2D, demonstrating a favorable benefit-risk profile in a heavily pretreated population of adults with advanced LMS.

Validation and application of volumetric absorptive microsampling (VAMS) dried blood method for phenylalanine measurement in patients with phenylketonuria.

BACKGROUND: Frequent blood phenylalanine (Phe) measurement is required for phenylketonuria (PKU) patients for diagnosis and disease status monitoring. Though various methods are available for blood Phe measurement, there is a lack of validated quantitative methods for measuring Phe with less than 15% variability. A method to allow at home blood sample collection for the PKU community is in high demand. METHODS: A volumetric absorptive microsampling (VAMS) dried blood collection high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method was developed and fully validated for blood Phe measurement in compliance with regulatory guidances. The method accuracy, precision, stability, selectivity, matrix and hematocrit effects were assessed. A venous plasma collection HPLC-MS/MS method was developed and validated as a reference method. 311 matching VAMS and plasma samples were collected from 24 PKU subjects in a Phase 2 clinical study. Phe measurements using the two methods were compared. RESULTS: Both VAMS and the plasma sample collection methods met the acceptance criteria for Good Laboratory Practice (GLP) bioanalytical analysis. Comparisons showed a high Pearson's correlation of 0.9813. The Passing-Bablok analysis showed that the difference was estimated to be less than 5% and Bland Altman analysis indicated that the difference was proportional with Phe concentration and for the majority of samples (88.85%) the measurement was within ±20% difference. Following 7 days treatment with 60 or 20 mg/kg/day PTC923 (Sepiapterin) or 20 mg/kg/day sapropterin, PKU patients exhibited respectively -206.4, -146.9, and -91.5 µmol/L reductions of blood Phe as measured by the VAMS method. CONCLUSIONS: Concordant results were obtained using VAMS and plasma methods, which demonstrated that VAMS is a reliable method for clinical applications to monitor blood Phe for PKU patients.

A pharmacokinetic drug-drug interaction study between rosuvastatin and emvododstat, a potent anti-SARS-CoV-2 (COVID-19) DHODH (dihydroorotate dehydrogenase) inhibitor.

A therapeutic agent that targets both viral replication and the hyper-reactive immune response would offer a highly desirable treatment for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2; COVID-19) management. Emvododstat (PTC299) was found to be a potent inhibitor of immunomodulatory and inflammation-related processes by the inhibition of dihydroorotate dehydrogenase (DHODH) to reduce SARS-CoV-2 replication. DHODH is the rate-limiting enzyme of the de novo pyrimidine nucleotide biosynthesis pathway. This drug interaction study was performed to determine whether emvododstat was an inhibitor of breast cancer resistance protein (BCRP) transporters in humans. Potential drug-drug interactions (DDIs) between emvododstat and a BCRP transporter substrate (rosuvastatin) were investigated by measuring plasma rosuvastatin concentrations before and after emvododstat administration. There was no apparent difference in rosuvastatin plasma exposure. The geometric means of maximum plasma rosuvastatin concentrations (Cmax ) were 4369 (rosuvastatin) and 5141 pg/mL (rosuvastatin + emvododstat) at 4 h postdose. Geometric mean rosuvastatin area under the concentration-time curve (AUC) from time 0 to the last measurable plasma concentration was 45 616 and 48 975 h·pg/mL when administered alone and after 7 days of b.i.d. emvododstat dosing, respectively. Geometric least squares mean ratios for Cmax and AUC were approximately equal to 1. Overall, administration of multiple doses of 100 mg emvododstat b.i.d. for 7 days in combination with a single dose of rosuvastatin was safe and well tolerated. Emvododstat can be safely administered with other BCRP substrate drugs. Hence, pharmacokinetic DDI mediated via BCRP inhibition is not expected when emvododstat and BCRP substrates are coadministered.

First-in-Human Studies of Pharmacokinetics and Safety of Utreloxastat (PTC857), a Novel 15-Lipooxygenase Inhibitor for the Treatment of Amyotrophic Lateral Sclerosis.

Utreloxastat (PTC857) is a 15-lipoxygenase inhibitor being developed to treat amyotrophic lateral sclerosis. This first-in-human study investigated the safety and pharmacokinetics of utreloxastat in healthy volunteers (N = 82) in a double-blind, placebo-controlled trial. The effects of a single ascending dose (100-1000 mg), multiple ascending doses (150-500 mg), and food (500 mg) on the pharmacokinetics and safety of utreloxastat were evaluated. Following single doses, the time to maximum plasma concentration (Cmax ) was observed ≈4 hours after dosing and the terminal half-life ranged from 20 to 25.3 hours. The Cmax and area under the concentration-time curve (AUC) increased slightly over dose proportionally. Following multiple doses (once daily/twice daily), the apparent clearance reduced and terminal half-life was ≥33 hours. There was no apparent difference of exposure following morning or evening doses. Varying diets increased the Cmax and AUCs of utreloxastat but did not alter time to Cmax . There were no gender-based differences in exposure. Utreloxastat showed no marked safety signal following single doses up to 1000 mg and multiple doses over 14 days of 500 mg once daily or 250 mg twice daily. The results support further development of utreloxastat for the treatment of patients with amyotrophic lateral sclerosis at a 250-mg twice-daily dose administered with food.

Pharmacokinetics of Dacarbazine and Unesbulin and CYP1A2-Mediated Drug Interactions in Patients With Leiomyosarcoma.

Unesbulin is being investigated in combination with dacarbazine (DTIC) as a potential therapeutic agent in patients with advanced leiomyosarcoma (LMS). This paper reports the pharmacokinetics (PK) of unesbulin, DTIC, and its unreactive surrogate metabolite 5-aminoimidazole-4-carboxamide (AIC) in 29 patients with advanced LMS. Drug interactions between DTIC (and AIC) and unesbulin were evaluated. DTIC (1000 mg/m2 ) was administered to patients with LMS via 1-hour intravenous (IV) infusion on Day 1 of every 21-day (q21d) cycle. Unesbulin dispersible tablets were administered orally twice weekly (BIW), starting on Day 2 of every cycle, except for Cycle 2 (C2), where unesbulin was dosed either on Day 1 together with DTIC or on Day 2, 1 day after DTIC administration. The PK of DTIC, AIC, and unesbulin in Cycle 1 (C1) and C2 were estimated using noncompartmental analysis. DTIC and AIC were measurable immediately after the start of infusion and reached Cmax immediately or shortly after end of infusion at 1.0 and 1.4 hours (Tmax ), respectively. Coadministration of unesbulin orally at 200 mg or above with DTIC inhibited cytochrome P450 (CYP)1A2-mediated DTIC metabolism, resulting in 66.7% reduction of AIC exposures. Such inhibition could be mitigated when unesbulin was dosed the day following DTIC infusion. Repeated unesbulin dosing demonstrated evidence of clinical CYP1A2 induction and increased AIC Cmax by 69.4% and AUCinf by 57.9%. No meaningful difference in unesbulin PK was observed between C2 and C1. The combination therapy of 1000 mg/m2 IV DTIC q21d and 300 mg unesbulin BIW in a staggered regimen is well tolerated in patients with LMS.

Emvododstat, a Potent Dihydroorotate Dehydrogenase Inhibitor, Is Effective in Preclinical Models of Acute Myeloid Leukemia.

Blocking the pyrimidine nucleotide de novo synthesis pathway by inhibiting dihydroorotate dehydrogenase (DHODH) results in the cell cycle arrest and/or differentiation of rapidly proliferating cells including activated lymphocytes, cancer cells, or virally infected cells. Emvododstat (PTC299) is an orally bioavailable small molecule that inhibits DHODH. We evaluated the potential for emvododstat to inhibit the progression of acute myeloid leukemia (AML) using several in vitro and in vivo models of the disease. Broad potent activity was demonstrated against multiple AML cell lines, AML blasts cultured ex vivo from patient blood samples, and AML tumor models including patient-derived xenograft models. Emvododstat induced differentiation, cytotoxicity, or both in primary AML patient blasts cultured ex vivo with 8 of 10 samples showing sensitivity. AML cells with diverse driver mutations were sensitive, suggesting the potential of emvododstat for broad therapeutic application. AML cell lines that are not sensitive to emvododstat are likely to be more reliant on the salvage pathway than on de novo synthesis of pyrimidine nucleotides. Pharmacokinetic experiments in rhesus monkeys demonstrated that emvododstat levels rose rapidly after oral administration, peaking about 2 hours post-dosing. This was associated with an increase in the levels of dihydroorotate (DHO), the substrate for DHODH, within 2 hours of dosing indicating that DHODH inhibition is rapid. DHO levels declined as drug levels declined, consistent with the reversibility of DHODH inhibition by emvododstat. These preclinical findings provide a rationale for clinical evaluation of emvododstat in an ongoing Phase 1 study of patients with relapsed/refractory acute leukemias.