Inhibition of Diacylglycerol Acyltransferase 2 Versus Diacylglycerol Acyltransferase 1: Potential Therapeutic Implications of Pharmacology.

PURPOSE: Hepatic steatosis due to altered lipid metabolism and accumulation of hepatic triglycerides is a hallmark of nonalcoholic fatty liver disease (NAFLD). Diacylglycerol acyltransferase (DGAT) enzymes, DGAT1 and DGAT2, catalyze the terminal reaction in triglyceride synthesis, making them attractive targets for pharmacologic intervention. There is a common misconception that these enzymes are related; however, despite their similar names, DGAT1 and DGAT2 differ significantly on multiple levels. As we look ahead to future clinical studies of DGAT2 inhibitors in patients with NAFLD and nonalcoholic steatohepatitis (NASH), we review key differences and include evidence to highlight and support DGAT2 inhibitor (DGAT2i) pharmacology. METHODS: Three Phase I, randomized, double-blind, placebo-controlled trials assessed the safety, tolerability, and pharmacokinetic properties of the DGAT2i ervogastat (PF-06865571) in healthy adult participants (Single Dose Study to Assess the Safety, Tolerability and Pharmacokinetics of PF-06865571 [study C2541001] and Study to Assess the Safety, Tolerability, and Pharmacokinetics of Multiple Doses of PF-06865571 in Healthy, Including Overweight and Obese, Adult Subjects [study C2541002]) or participants with NAFLD (2-Week Study in People With Nonalcoholic Fatty Liver Disease [study C2541005]). Data from 2 Phase I, randomized, double-blind, placebo-controlled trials of the DGAT1i PF-04620110 in healthy participants (A Single Dose Study of PF-04620110 in Overweight and Obese, Otherwise Healthy Volunteers [study B0961001] and A Multiple Dose Study of PF-04620110 in Overweight and Obese, Otherwise Healthy Volunteers [study B0961002]) were included for comparison. Safety outcomes were the primary end point in all studies, except in study C2541005, in which safety was the secondary end point, with relative change from baseline in whole liver fat at day 15 assessed as the primary end point. Safety data were analyzed across studies by total daily dose of ervogastat (5, 15, 50, 100, 150, 500, 600, 1000, and 1500 mg) or PF-04620110 (0.3, 1, 3, 5, 7, 10, 14, and 21 mg), with placebo data pooled separately across ervogastat and PF-04620110 studies. FINDINGS: Published data indicate that DGAT1 and DGAT2 differ in multiple dimensions, including gene family, subcellular localization, substrate preference, and specificity, with unrelated pharmacologic inhibition properties and differing safety profiles. Although initial nonclinical studies suggested a potentially attractive therapeutic profile with DGAT1 inhibition, genetic and pharmacologic data suggest otherwise, with common gastrointestinal adverse events, including nausea, vomiting, and diarrhea, limiting further clinical development. Conversely, DGAT2 inhibition, although initially not pursued as aggressively as a potential target for pharmacologic intervention, has consistent efficacy in nonclinical studies, with reduced triglyceride synthesis accompanied by reduced expression of genes essential for de novo lipogenesis. In addition, early clinical data indicate antisteatotic effects with DGAT2i ervogastat, in participants with NAFLD, accompanied by a well-tolerated safety profile. IMPLICATIONS: Although pharmacologic DGAT1is are limited by an adverse safety profile, data support use of DGAT2i as an effective and well-tolerated therapeutic strategy for patients with NAFLD, NASH, and NASH with liver fibrosis. CLINICALTRIALS: gov identifiers: NCT03092232, NCT03230383, NCT03513588, NCT00799006, and NCT00959426.

Pharmacokinetics, mass balance, metabolism, and excretion of the liver-targeted acetyl-CoA carboxylase inhibitor PF-05221304 (clesacostat) in humans.

The disposition of the hepatoselective ACC inhibitor PF-05221304 (Clesacostat) was studied after a single 50-mg oral dose of [14C]-PF-05221304 to healthy human subjects.Mass balance was achieved with 89.9% of the administered dose recovered in urine and faeces, over the 11-day study period. The total administered radioactivity excreted in faeces and urine was 81.7 and 8.2%, respectively. Unchanged PF-05221304 accounted for 35.6% of the radioactive dose in faeces, suggesting ∼64% of the administered dose was absorbed.PF-05221304 was principally metabolised via oxidative and reductive pathways involving: (a) N-dealkylation, (b) isopropyl group monohydroxylation to yield enantiomeric metabolites (M2a and M2b), (c) hydroxylation on the 3-azaspiro[5.5]undecan-8-one moiety to metabolites M5 and 519c, and (d) carbonyl group reduction to enantiomeric alcohol metabolites M3, and M4. Secondary metabolites (521a, 521b, and 533), derived from a combination of oxidation and reduction of the primary metabolites accounted for ∼14.8% of the dose. In plasma, unchanged PF-05221304 represented 96.1% circulating radioactivity. Metabolites M1, M2b, and M2a represented 1.94, 1.76, and 0.18% of circulating radioactivity, respectively.Overall, these data suggest that PF-05221304 is well absorbed in humans and eliminated largely via phase I metabolism.

Efficacy and safety of an orally administered DGAT2 inhibitor alone or coadministered with a liver-targeted ACC inhibitor in adults with non-alcoholic steatohepatitis (NASH): rationale and design of the phase II, dose-ranging, dose-finding, randomised, placebo-controlled MIRNA (Metabolic Interventions to Resolve NASH with fibrosis) study.

INTRODUCTION: Small molecule inhibitors of the terminal step in intrahepatic triglyceride synthesis (diacylglycerol acyltransferase 2 inhibitor (DGAT2i, PF-06865571, ervogastat)) and upstream blockade of de novo lipogenesis via acetyl-coenzyme A carboxylase inhibitor (ACCi, PF-05221304, clesacostat) showed promise in reducing hepatic steatosis in early clinical trials. This study assesses efficacy and safety of these metabolic interventions to resolve non-alcoholic steatohepatitis (NASH) with fibrosis. METHODS AND ANALYSIS: This phase II, randomised, dose-ranging, dose-finding study evaluates DGAT2i 25-300 mg two times per day (BID) or 150-300 mg once a day, DGAT2i 150-300 mg BID+ACCi 5-10 mg BID coadministration or matching placebo in a planned 450 adults with biopsy-confirmed NASH and liver fibrosis stages 2-3 from approximately 220 sites in 11 countries across North America, Europe and Asia. A triage approach including double-confirmation via non-invasive markers is included prior to screening/baseline liver biopsy. On confirmation of histological diagnosis, participants enter a ≥6-week run-in period, then a 48-week double-blind, double-dummy dosing period. The primary endpoint is the proportion of participants achieving histological NASH resolution without worsening fibrosis, ≥1 stage improvement in fibrosis without worsening NASH, or both, assessed by central pathologists. Other endpoints include assessment of hepatic steatosis (imaging substudy), overall safety and tolerability, and evaluation of blood-based biomarkers and quantitative ultrasound parameters over time. ETHICS AND DISSEMINATION: Metabolic Interventions to Resolve NASH with fibrosis (MIRNA) is conducted in accordance with the Declaration of Helsinki and Council for International Organisations of Medical Sciences (CIOMS) International Ethical Guidelines, International Council on Harmonisation Good Clinical Practice guidelines, applicable laws and regulations, including privacy laws. Local independent review board/ethics committees (IRB/ECs) review/approve the protocol, any amendments, informed consent and other forms. Participants provide written informed consent. Details of all IRB/ECs, as well as results, will be published in a peer-reviewed journal and publicly disclosed through ClinicalTrials.gov, EudraCT, and/or www.pfizer.com and other public registries as per applicable local laws/regulations. TRIAL REGISTRATION NUMBER: NCT04321031.

ACC inhibitor alone or co-administered with a DGAT2 inhibitor in patients with non-alcoholic fatty liver disease: two parallel, placebo-controlled, randomized phase 2a trials.

Alterations in lipid metabolism might contribute to the pathogenesis of non-alcoholic fatty liver disease (NAFLD). However, no pharmacological agents are currently approved in the United States or the European Union for the treatment of NAFLD. Two parallel phase 2a studies investigated the effects of liver-directed ACC1/2 inhibition in adults with NAFLD. The first study ( NCT03248882 ) examined the effects of monotherapy with a novel ACC1/2 inhibitor, PF-05221304 (2, 10, 25 and 50 mg once daily (QD)), versus placebo at 16 weeks of treatment; the second study ( NCT03776175 ) investigated the effects of PF-05221304 (15 mg twice daily (BID)) co-administered with a DGAT2 inhibitor, PF-06865571 (300 mg BID), versus placebo after 6 weeks of treatment. The primary endpoint in both studies was percent change from baseline in liver fat assessed by magnetic resonance imaging-proton density fat fraction. Dose-dependent reductions in liver fat reached 50-65% with PF-05221304 monotherapy doses ≥10 mg QD; least squares mean (LSM) 80% confidence interval (CI) was -7.2 (-13.9, 0.0), -17.1 (-22.7, -11.1), -49.9 (-53.3, -46.2), -55.9 (-59.0, -52.4) and -64.8 (-67.5, -62.0) with 16 weeks placebo and PF-05221304 2, 10, 25 and 50 mg QD, respectively. The overall incidence of adverse events (AEs) did not increase with increasing PF-05221304 dose, except for a dose-dependent elevation in serum triglycerides (a known consequence of hepatic acetyl-coenzyme A carboxylase (ACC) inhibition) in 23/305 (8%) patients, leading to withdrawal in 13/305 (4%), and a dose-dependent elevation in other serum lipids. Co-administration of PF-05221304 and PF-06865571 lowered liver fat compared to placebo (placebo-adjusted LSM (90% CI) -44.6% (-54.8, -32.2)). Placebo-adjusted LSM (90% CI) reduction in liver fat was -44.5% (-55.0, -31.7) and -35.4% (-47.4, -20.7) after 6 weeks with PF-05221304 or PF-06865571 alone. AEs were reported for 10/28 (36%) patients after co-administered PF-05221304 and PF-06865571, with no discontinuations due to AEs, and the ACC inhibitor-mediated effect on serum triglycerides was mitigated, suggesting that PF-05221304 and PF-06865571 co-administration has the potential to address some of the limitations of ACC inhibition alone.

Overview of the Clinical Pharmacology of Ertugliflozin, a Novel Sodium-Glucose Cotransporter 2 (SGLT2) Inhibitor.

Ertugliflozin, a selective inhibitor of sodium-glucose cotransporter 2 (SGLT2), is approved in the US, EU, and other regions for the treatment of adults with type 2 diabetes mellitus (T2DM). This review summarizes the ertugliflozin pharmacokinetic (PK) and pharmacodynamic data obtained during phase I clinical development, which supported the registration and labeling of this drug. The PK of ertugliflozin was similar in healthy subjects and patients with T2DM. Oral absorption was rapid, with time to peak plasma concentrations (Tmax) occurring at 1 h (fasted) and 2 h (fed) postdose. The terminal phase half-life ranged from 11 to 18 h and steady-state concentrations were achieved by 6 days after initiating once-daily dosing. Ertugliflozin exposure increased in a dose-proportional manner over the tested dose range of 0.5-300 mg. Ertugliflozin is categorized as a Biopharmaceutical Classification System Class I drug with an absolute bioavailability of ~ 100% under fasted conditions. Administration of the ertugliflozin 15 mg commercial tablet with food resulted in no meaningful effect on ertugliflozin area under the plasma concentration-time curve (AUC), but decreased peak concentrations (Cmax) by 29%. The effect on Cmax is not clinically relevant and ertugliflozin can be administered without regard to food. Mild, moderate, and severe renal impairment were associated with a ≤ 70% increase in ertugliflozin exposure relative to subjects with normal renal function, and no dose adjustment in renal impairment patients is needed based on PK results. Consistent with the mechanism of action of SGLT2 inhibitors, 24-h urinary glucose excretion decreased with worsening renal function. In subjects with moderate hepatic impairment, a decrease in AUC (13%) relative to subjects with normal hepatic function was observed and not considered clinically relevant. Concomitant administration of metformin, sitagliptin, glimepiride, or simvastatin with ertugliflozin did not have clinically meaningful effects on the PK of ertugliflozin or the coadministered medications. Coadministration of rifampin decreased ertugliflozin AUC and Cmax by 39% and 15%, respectively, and is not expected to affect efficacy in a clinically meaningful manner. This comprehensive evaluation supports administration to patients with T2DM without regard to prandial status and with no dose adjustments for coadministration with commonly prescribed drugs, or in patients with renal impairment or mild-to-moderate hepatic impairment based on ertugliflozin PK.

Safety, Tolerability, Pharmacokinetics, and Pharmacodynamics of a Liver-Targeting Acetyl-CoA Carboxylase Inhibitor (PF-05221304): A Three-Part Randomized Phase 1 Study.

PF-05221304 is a liver-targeted inhibitor of acetyl-CoA carboxylase, an enzyme that catalyzes the first committed step in de novo lipogenesis (DNL). This first-in-human study investigated safety/tolerability and pharmacokinetics of single and multiple ascending oral PF-05221304 doses, and fructose-stimulated DNL inhibition with repeated oral doses. Healthy subjects (n = 96) received single (1-240 mg) or repeated (2-200 mg daily) doses for 14 days or single 100-mg doses with and without food. PF-05221304 was well tolerated at all doses. Repeated PF-05221304 doses inhibited hepatic DNL in a dose-dependent manner, with near-complete inhibition seen at higher doses. With doses yielding ≥90% DNL inhibition, asymptomatic increases in fasting/postprandial serum triglyceride levels (≥40 mg/day) and declines in platelet count (≥60 mg/day) occurred; these were not observed at ≤80% DNL inhibition. Steady-state pharmacokinetics generally increased dose-proportionally, with a half-life of 14-18 hours and a minimal food effect on plasma exposure. The observed safety and tolerability, pharmacokinetics, and pharmacodynamics support the continued evaluation of PF-05221304 for the treatment of nonalcoholic steatohepatitis.

Targeting diacylglycerol acyltransferase 2 for the treatment of nonalcoholic steatohepatitis.

Nonalcoholic steatohepatitis (NASH) is characterized by the accumulation of hepatocyte triglycerides, the synthesis of which is catalyzed by diacylglycerol acyltransferases (DGATs). Here, we investigate DGAT2 as a potential therapeutic target using an orally administered, selective DGAT2 inhibitor, PF-06427878. Treatment with PF-06427878 resulted in the reduction of hepatic and circulating plasma triglyceride concentrations and decreased lipogenic gene expression in rats maintained on a Western-type diet. In a mouse model of NASH, histological improvements in steatosis, ballooning, and fibrosis were evident in the livers of animals receiving PF-06427878 compared with mice treated with vehicle alone. We extended these nonclinical studies to two phase 1 studies in humans [NCT02855177 (n = 24) and NCT02391623 (n = 39; n = 38 completed)] and observed that PF-06427878 was well tolerated and influenced markers of liver function (alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, and total bilirubin) in healthy adults, with statistically significant reductions from baseline at day 14 in participants treated with PF-06427878 1500 milligrams per day (P < 0.05). Moreover, magnetic resonance imaging using proton density fat fraction showed that PF-06427878 1500 milligrams per day reduced hepatic steatosis in healthy adult participants. Our findings highlight DGAT2 inhibition by a small, potent, selective compound as a potential therapeutic approach for the treatment of NASH.

Glycemic Effect and Safety of a Systemic, Partial Glucokinase Activator, PF-04937319, in Patients With Type 2 Diabetes Mellitus Inadequately Controlled on Metformin-A Randomized, Crossover, Active-Controlled Study.

Glucokinase enhances glucose conversion to glucose-6-phosphate, causing glucose-stimulated insulin secretion from pancreatic ß cells and increased hepatic glucose uptake. PF-04937319 is a partial glucokinase activator designed to maintain efficacy with reduced hypoglycemia risk. In this randomized, double-blind, double-dummy, 3-period crossover phase 1b study, patients aged 18-70 years with type 2 diabetes mellitus and on metformin received once-daily PF-04937319 (300 mg), split-dose PF-04937319 (150+100 mg; breakfast+lunch), or sitagliptin (100 mg once daily). The primary end point was day 14 weighted mean daily glucose (WMDG) change from period-specific baseline. Secondary end points included change from baseline in fasting plasma glucose, premeal C-peptide and insulin, and safety, including hypoglycemia frequency. Mean decrease from baseline in observed WMDG (mg/dL) was greater for PF-04937319 (split-dose, -31.24; once daily, -31.33) versus sitagliptin (-19.24). Using the integrated glucose red-cell HbA1c model, the observed WMDG effect with both PF-04937319 dosing regimens was projected to yield a clinically superior effect on mean glycated hemoglobin (HbA1c ; split-dose, -0.88%; once daily, -0.94%) compared with sitagliptin (-0.63%). There was no difference in premeal C-peptide or insulin levels, and although the effect on WMDG with both PF-04937319 regimens was similar, the split-dose regimen appeared to offer some advantage in safety and tolerability.

Metabolites in safety testing assessment in early clinical development: a case study with a glucokinase activator.

The present article summarizes Metabolites in Safety Testing (MIST) studies on a glucokinase activator, N,N-dimethyl-5-((2-methyl-6-((5-methylpyrazin-2-yl)carbamoyl)benzofuran-4-yl)oxy)pyrimidine-2-carboxamide (PF-04937319), which is under development for the treatment of type 2 diametes mellitus. Metabolic profiling in rat, dog, and human hepatocytes revealed that PF-04937319 is metabolized via oxidative (major) and hydrolytic pathways (minor). N-Demethylation to metabolite M1 [N-methyl-5-((2-methyl-6-((5-methylpyrazin-2-yl)carbamoyl)benzofuran-4-yl)oxy)pyrimidine-2-carboxamide] was the major metabolic fate of PF-04937319 in human (but not rat or dog) hepatocytes, and was catalyzed by CYP3A and CYP2C isoforms. Qualitative examination of circulating metabolites in humans at the 100- and 300-mg doses from a 14-day multiple dose study revealed unchanged parent drug and M1 as principal components. Because M1 accounted for 65% of the drug-related material at steady state, an authentic standard was synthesized and used for comparison of steady-state exposures in humans and the 3-month safety studies in rats and dogs at the no-observed-adverse-effect level. Although circulating levels of M1 were very low in beagle dogs and female rats, adequate coverage was obtained in terms of total maximal plasma concentration (∼7.7× and 1.8×) and area under the plasma concentration-time curve (AUC; 3.6× and 0.8× AUC) relative to the 100- and 300-mg doses, respectively, in male rats. Examination of primary pharmacology revealed M1 was less potent as a glucokinase activator than the parent drug (compound PF-04937319: EC50 = 0.17 µM; M1: EC50 = 4.69 µM). Furthermore, M1 did not inhibit major human P450 enzymes (IC50 > 30 µM), and was negative in the Salmonella Ames assay, with minimal off-target pharmacology, based on CEREP broad ligand profiling. Insights gained from this analysis should lead to a more efficient and focused development plan for fulfilling MIST requirements with PF-04937319.

Influence of i.v. haloperidol on ventricular repolarization and monophasic action potential duration in anesthetized dogs.

INTRODUCTION: i.v. haloperidol is used commonly for sedation in critically ill patients. However, i.v. haloperidol has been shown to cause the life-threatening ventricular tachyarrhythmia torsades de pointes. Mechanisms by which haloperidol causes torsades de pointes have not been widely investigated in controlled studies. STUDY OBJECTIVES: To determine the effects of i.v. haloperidol on electrophysiologic parameters known to promote torsades de pointes. INTERVENTIONS: Monophasic action potential catheters were guided under fluoroscopy into the right and left ventricles of 14 chloralose-anesthetized dogs (haloperidol, nine dogs; placebo, five dogs). Effective refractory period (ERP), action potential duration at 90% repolarization (APD90), and QTc interval measurements were performed at baseline and after each of four doses of haloperidol (0.15, 0.5, 2.0, and 3.0 mg/kg) or placebo at three different pacing cycle lengths (450, 300, and 250 ms). MEASUREMENTS AND RESULTS: i.v. haloperidol significantly prolonged left and right ventricular ERP by a magnitude of 12 to 20% at all pacing cycle lengths. ERP values in the placebo group did not change significantly from pretreatment values in either ventricle. Haloperidol significantly prolonged left ventricular APD90 at a pacing cycle length of 300 ms. The effects of haloperidol on right ventricular APD90 approached significance at a cycle length of 450 ms. Overall, haloperidol prolonged APD90 by 7 to 11%, with less consistent and more variable effects than those for the ERP. APD90 was not significantly altered in the placebo groups. Haloperidol produced significant prolongation in QTc intervals. The electrophysiologic effects of haloperidol were related to dose, with a plateau reached at the 0.5 mg/kg dose for ERP measurements and at the 2 mg/kg dose for the APD90 and QTc interval measurements. CONCLUSIONS: i.v. haloperidol prolongs ventricular ERP and APD90 in intact canine hearts. These electrophysiologic effects are likely associated with the clinical torsades de pointes-inducing actions of i.v. haloperidol in critically ill patients.