Paracetamol analogues conjugated by FAAH induce TRPV1-mediated antinociception without causing acute liver toxicity.

Paracetamol, one of the most widely used pain-relieving drugs, is deacetylated to 4-aminophenol (4-AP) that undergoes fatty acid amide hydrolase (FAAH)-dependent biotransformation into N-arachidonoylphenolamine (AM404), which mediates TRPV1-dependent antinociception in the brain of rodents. However, paracetamol is also converted to the liver-toxic metabolite N-acetyl-p-benzoquinone imine already at therapeutic doses, urging for safer paracetamol analogues. Primary amine analogues with chemical structures similar to paracetamol were evaluated for their propensity to undergo FAAH-dependent N-arachidonoyl conjugation into TRPV1 activators both in vitro and in vivo in rodents. The antinociceptive and antipyretic activity of paracetamol and primary amine analogues was examined with regard to FAAH and TRPV1 as well as if these analogues produced acute liver toxicity. 5-Amino-2-methoxyphenol (2) and 5-aminoindazole (3) displayed efficient target protein interactions with a dose-dependent antinociceptive effect in the mice formalin test, which in the second phase was dependent on FAAH and TRPV1. No hepatotoxicity of the FAAH substrates transformed into TRPV1 activators was observed. While paracetamol attenuates pyrexia via inhibition of brain cyclooxygenase, its antinociceptive FAAH substrate 4-AP was not antipyretic, suggesting separate mechanisms for the antipyretic and antinociceptive effect of paracetamol. Furthermore, compound 3 reduced fever without a brain cyclooxygenase inhibitory action. The data support our view that analgesics and antipyretics without liver toxicity can be derived from paracetamol. Thus, research into the molecular actions of paracetamol could pave the way for the discovery of analgesics and antipyretics with a better benefit-to-risk ratio.

N,N'-Bis(2-mercaptoethyl)isophthalamide Binds Electrophilic Paracetamol Metabolites and Prevents Paracetamol-Induced Liver Toxicity.

Paracetamol overdosing may cause liver injury including fulminant liver failure due to generation of the toxic metabolites, N-acetyl-p-benzoquinone imine (NAPQI) and p-benzoquinone (p-BQ). Herein, the chelating agent, N,N'-Bis(2-mercaptoethyl)isophthalamide (NBMI), was examined for its potential ability to entrap NAPQI and p-BQ and to prevent paracetamol-induced liver injury. Both NBMI and the conventional paracetamol antidote N-acetylcysteine (NAC) were investigated with regard to their abilities to scavenge the NAPQI and p-BQ in a Transient Receptor Potential Ankyrin 1-dependent screening assay. Stoichiometric evaluations indicated that NBMI was able to entrap these metabolites more efficiently than NAC. Furthermore, oral administration of either NBMI (680 mg/kg) or NAC (680 mg/kg) prevented the development of the characteristic liver necrosis and elevation of serum alanine aminotransferase in a mouse model for paracetamol-induced liver injury. In summary, these results show that NBMI is able to entrap the toxic metabolites NAPQI and p-BQ and to prevent paracetamol-induced liver injury in mice.

Selepressin, a novel selective vasopressin V1A agonist, is an effective substitute for norepinephrine in a phase IIa randomized, placebo-controlled trial in septic shock patients.

BACKGROUND: Vasopressin is widely used for vasopressor support in septic shock patients, but experimental evidence suggests that selective V1A agonists are superior. The initial pharmacodynamic effects, pharmacokinetics, and safety of selepressin, a novel V1A-selective vasopressin analogue, was examined in a phase IIa trial in septic shock patients. METHODS: This was a randomized, double-blind, placebo-controlled multicenter trial in 53 patients in early septic shock (aged ≥18 years, fluid resuscitation, requiring vasopressor support) who received selepressin 1.25 ng/kg/minute (n = 10), 2.5 ng/kg/minute (n = 19), 3.75 ng/kg/minute (n = 2), or placebo (n = 21) until shock resolution or a maximum of 7 days. If mean arterial pressure (MAP) ≥65 mmHg was not maintained, open-label norepinephrine was added. Co-primary endpoints were maintenance of MAP >60 mmHg without norepinephrine, norepinephrine dose, and proportion of patients maintaining MAP >60 mmHg with or without norepinephrine over 7 days. Secondary endpoints included cumulative fluid balance, organ dysfunction, pharmacokinetics, and safety. RESULTS: A higher proportion of the patients receiving 2.5 ng/kg/minute selepressin maintained MAP >60 mmHg without norepinephrine (about 50% and 70% at 12 and 24 h, respectively) vs. 1.25 ng/kg/minute selepressin and placebo (p < 0.01). The 7-day cumulative doses of norepinephrine were 761, 659, and 249 µg/kg (placebo 1.25 ng/kg/minute and 2.5 ng/kg/minute, respectively; 2.5 ng/kg/minute vs. placebo; p < 0.01). Norepinephrine infusion was weaned more rapidly in selepressin 2.5 ng/kg/minute vs. placebo (0.04 vs. 0.18 µg/kg/minute at 24 h, p < 0.001), successfully maintaining target MAP and reducing norepinephrine dose vs. placebo (first 24 h, p < 0.001). Cumulative net fluid balance was lower from day 5 onward in the selepressin 2.5 ng/kg/minute group vs. placebo (p < 0.05). The selepressin 2.5 ng/kg/minute group had a greater proportion of days alive and free of ventilation vs. placebo (p < 0.02). Selepressin (2.5 ng/kg/minute) was well tolerated, with a similar frequency of treatment-emergent adverse events for selepressin 2.5 ng/kg/minute and placebo. Two patients were infused at 3.75 ng/kg/minute, one of whom had the study drug infusion discontinued for possible safety reasons, with subsequent discontinuation of this dose group. CONCLUSIONS: In septic shock patients, selepressin 2.5 ng/kg/minute was able to rapidly replace norepinephrine while maintaining adequate MAP, and it may improve fluid balance and shorten the time of mechanical ventilation. TRIAL REGISTRATION: ClinicalTrials.gov, NCT01000649 . Registered on September 30, 2009.

Effect of Degarelix, a Gonadotropin-Releasing Hormone Receptor Antagonist for the Treatment of Prostate Cancer, on Cardiac Repolarisation in a Randomised, Placebo and Active Comparator Controlled Thorough QT/QTc Trial in Healthy Men.

BACKGROUND AND OBJECTIVES: Degarelix is a gonadotropin-releasing hormone antagonist registered for the treatment of advanced hormone-dependent prostate cancer. Treatment causing androgen deprivation is associated with QT prolongation and this study investigated whether degarelix at supratherapeutic concentrations has an intrinsic effect per se on cardiac repolarisation and the QT interval. METHODS: This was a single-centre, randomised, crossover study comparing the effect of degarelix, placebo, and the positive control moxifloxacin on the QT interval. Degarelix and placebo treatments were double-blind, whereas moxifloxacin treatment was open-label. Eighty healthy men, aged 18-45 years, received single intravenous doses of degarelix 2.8 mg, and placebo, as well as a single oral dose of moxifloxacin 400 mg. Electrocardiograms were collected up to 24 h after the start of administration, with the QT interval assessed and plasma concentrations of degarelix concomitantly analysed. RESULTS: Time-matched, one-sided 95% upper confidence boundaries for baseline-corrected average changes from placebo for the QT interval, corrected using the Fridericia method (ΔΔQTcF), did not exceed 10 ms at any timepoint, with maximum degarelix concentrations reaching approximately threefold the concentrations seen in the treatment of prostate cancer. Furthermore, concentration-exposure analysis indicated absence of any QT prolongation effects of degarelix. No significant effect on any other cardiac parameter was observed. The lower bound of the 98.3% confidence interval for moxifloxacin ΔΔQTcF exceeded 5 ms, thus verifying assay sensitivity. CONCLUSION: The results showed that the study was validated to detect a significant effect on the QT interval, and that degarelix by itself does not have any effect on the QT interval and cardiac repolarisation at supratherapeutic concentrations.

Characterisation of Population Pharmacokinetics and Endogenous Follicle-Stimulating Hormone (FSH) Levels After Multiple Dosing of a Recombinant Human FSH (FE 999049) in Healthy Women.

OBJECTIVE: The aim of this study was to characterise the population pharmacokinetics of FE 999049, a novel recombinant human follicle-stimulating hormone (FSH), after multiple dosing in healthy women, taking into account endogenous FSH levels and the reproductive hormone dynamics. METHODS: Longitudinal measurements of FSH, luteinising hormone, progesterone, estradiol, and inhibin B levels were collected after repeated subcutaneous dosing with 225 IU of FE 999049 in 24 gonadotropin downregulated healthy women. The FSH data were described using nonlinear mixed-effects modelling. RESULTS: The measured FSH levels were modelled as a sum of endogenous FSH and FE 999049. The FE 999049 population pharmacokinetics were best described using a one-compartment model with first-order absorption and elimination, and a transit model for delayed absorption. The apparent clearance and volume of distribution increased with body weight in accordance with an allometrically scaled power exponent of 0.75 and 1, respectively. Endogenous FSH levels were lower in individuals with higher progesterone levels at baseline and were further suppressed over time with increasing inhibin B levels. CONCLUSIONS: This characterisation of FE 999049 population pharmacokinetics after repeated dosing is in line with previous findings after single-dose administration. The results provide a basis for study design and data evaluation in the future development of recombinant FSH products, and show it can be of importance to account for endogenous FSH levels and its variation over time for accurate estimation of exogenously administered FSH pharmacokinetic parameters. Thus, correcting FSH concentrations by the observed endogenous FSH baseline value at all time points may be incorrect.

Population Pharmacokinetic Modelling of FE 999049, a Recombinant Human Follicle-Stimulating Hormone, in Healthy Women After Single Ascending Doses.

OBJECTIVE: The purpose of this analysis was to develop a population pharmacokinetic model for a novel recombinant human follicle-stimulating hormone (FSH) (FE 999049) expressed from a human cell line of foetal retinal origin (PER.C6(®)) developed for controlled ovarian stimulation prior to assisted reproductive technologies. METHODS: Serum FSH levels were measured following a single subcutaneous FE 999049 injection of 37.5, 75, 150, 225 or 450 IU in 27 pituitary-suppressed healthy female subjects participating in this first-in-human single ascending dose trial. Data was analysed by nonlinear mixed effects population pharmacokinetic modelling in NONMEM 7.2.0. RESULTS: A one-compartment model with first-order absorption and elimination rates was found to best describe the data. A transit model was introduced to describe a delay in the absorption process. The apparent clearance (CL/F) and apparent volume of distribution (V/F) estimates were found to increase with body weight. Body weight was included as an allometrically scaled covariate with a power exponent of 0.75 for CL/F and 1 for V/F. CONCLUSIONS: The single-dose pharmacokinetics of FE 999049 were adequately described by a population pharmacokinetic model. The average drug concentration at steady state is expected to be reduced with increasing body weight.

Different pharmacokinetic and pharmacodynamic properties of recombinant follicle-stimulating hormone (rFSH) derived from a human cell line compared with rFSH from a non-human cell line.

Pharmacokinetic and pharmacodynamic properties of a novel recombinant follicle-stimulating hormone (rFSH) preparation (FE 999049), expressed by a human cell line (PER.C6), was compared with an rFSH preparation (follitropin α) expressed by a Chinese hamster ovary (CHO) cell line in healthy pituitary-suppressed women. Following single intravenous administration of 225 IU (Steelman-Pohley assay), the clearance was lower, 0.31 versus 0.44 L/h, for FE 999049 than for follitropin α. Likewise, the apparent clearance after repeated daily subcutaneous administrations was lower, 0.58 versus 0.99 L/h, and AUC and C(max) higher, 1.7- and 1.6-fold. The absolute bioavailability after a single subcutaneous dose of 450 IU was similar for both preparations, 60-65%. After repeated subcutaneous administration the elimination half-life was approximately 30 and 24 hours for FE 999049 and follitropin α. The ovarian responses by number of follicles and serum concentrations of inhibin B and estradiol, were higher with FE 999049 than with follitropin α, AUC and C(max) for the two latter being >1.6-fold greater with FE 999049 than with follitropin α. These results indicate that administration of equal doses of FE 999049, expressed in a human cell line, and follitropin α, expressed in a CHO cell line, display different pharmacokinetic and pharmacodynamic properties in humans.

Pharmacokinetics and bioavailability of a new testosterone gel formulation in comparison to Testogel® in healthy men.

This randomized, open-label, multiple-dose three-way cross-over study compared the pharmacokinetics of a new testosterone gel formulation in two strengths, testosterone gel 1% and testosterone gel 2% (FE 999303), with Testogel® in 11 testosterone-suppressed healthy men. Subjects received one of six treatment sequences; 50 mg of testosterone was administered once daily for 7 consecutive days, with different treatments separated by washout-periods of 6-9 days. Testosterone gel 1% and testosterone gel 2% displayed greater relative bioavailability (2.6- and 1.6-fold, respectively) than Testogel on Day 1, which persisted, to a smaller extent, on Day 7. Initial absorption was highest and most rapid for testosterone gel 1% and 2%, showing apparent first-order absorption kinetics. Maximum serum concentrations (Cmax ) were 6.25 and 2.97 ng/mL, respectively, occurring ∼5-6 hours post-application on Day 1 versus Cmax of 1.71 ng/mL after ∼24 hours with Testogel, showing apparent zero-order absorption kinetics. Similar differences were observed on Day 7. All treatments appeared to reach approximately the same steady-state level within the first 24 hours. No application-site skin reactions occurred with any preparation. In conclusion, the new testosterone formulation showed higher bioavailability, and the ability to deliver more testosterone in a smaller volume.

Human neuropeptide Y signal peptide gain-of-function polymorphism is associated with increased body mass index: possible mode of function.

Neuropeptide Y (NPY) has been implicated in the control of food intake and energy balance based on many observations in animals. We have studied single nucleotide polymorphisms (SNPs) within the regulatory and coding sequences of the human NPY gene. One variant (1128 T>C), which causes an amino acid change from leucine to proline at codon 7 in the signal peptide of NPY, was associated with increased body mass index (BMI) in two separate Swedish populations of normal and overweight individuals. In vitro transcription and translation studies indicated the unlikelihood that this signal peptide variation affects the site of cleavage and targeting or uptake of NPY into the endoplasmic reticulum (ER). However, the mutant, and to a lesser extent the wild-type, signal peptide by themselves markedly potentiated NPY-induced food intake, as well as hypothalamic NPY receptor signaling. Our findings in humans strongly indicate that the NPY signaling system is implicated in body weight regulation and suggest a new and unexpected functional role of a signal peptide.