Nalfurafine

Formulation

The table below describes the formulation of nalfurafine hydrochloride and the controls morphine sulfate and gabapentin for the experiments shown on this page.

Experiment(s)	Compound	Dose(s)	Correction factor	Dose volume mL/kg	Route	Vehicle	Suspension / Solution
Pharmacokinetics	Nalfurafine	100, 1000	1.08	1	SC	PBS	Solution
Pharmacokinetics	Nalfurafine	µg/kg	1.08	1	SC	PBS	Solution
Irwin (1-30 µg/kg),	Nalfurafine	1, 3, 10, 30	1.08	1	SC	PBS	Solution
Rotarod (1-30 µg/kg)	Nalfurafine	µg/kg	1.08	1	SC	PBS	Solution
Irwin (100-3000 µg/kg),	Nalfurafine	100, 300, 1000, 3000	1.08	3	SC	PBS	Solution
Rotarod (100-3000 µg/kg)	Nalfurafine	µg/kg	1.08	3	SC	PBS	Solution
Plantar incision,	Nalfurafine	0.3, 1, 3, 10	1.08	1	SC	PBS	Solution
L5/L6 SNL	Nalfurafine	µg/kg	1.08	1	SC	PBS	Solution
Plantar incision	Morphine	6 mg/kg	1.33	1	SC	Saline	Solution
L5/L6 SNL	Gabapentin	60 mg/kg	1.00	1	PO	Saline	Solution

L5/L6 SNL = L5/L6 spinal nerve ligation; PO = per os; SC = subcutaneous; PBS = phosphate buffered saline (pH 7.2)

Results

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Method: Pharmacokinetics

Nalfurafine concentrations were measured in serially collected plasma samples and terminally collected brain samples (shown on two graphs) from males and females treated with 100 or 1000 µg/kg nalfurafine (SC) in a vehicle of PBS. There were 4 rats/group for plasma and 2-3 rats/group for brain.

Method: Pharmacokinetics

A table shows pharmacokinetics parameters calculated for nalfurafine using non-compartmental analysis in male and female rats treated with nalfurafine (100 or 1000 µg/kg, SC, vehicle of PBS). The means for the 4 groups (males and females treated with 100 µg/kg nalfurafine and males and females treated with 1000 µg/kg nalfurafine, respectively) are as follows. The highest concentration (Cmax) was 0.029, 0.023, 0.29, and 0.32 µM; the time at the maximum concentration (Tmax) was 0.5, 0.44, 0.31, and 0.44 hours; the half-life (T1/2) was 1.34, 1.42, 1.17, and 1.17 hours; area under the curve for time 0 to 24 hours was 2.7, 2.7, 30.3, and 39.6 minutes*µM; area under the curve for time 0 to infinity was 2.8, 2.9, 31, and 40.9 minutes*µM; the brain-to-plasma ratio at 2 hours was 0.74, 0.54, 1.02, and 0.96; the brain-to-plasma ratio at 24 hours could not be calculated because all samples were below the lower limit of quantification.

Method: Modified Irwin functional observational battery

Two reviewers observed and scored 39 behaviors in a modified Irwin test. For each animal, the scores for behavioral observations from each reviewer are summed then averaged for male and female rats (shown on two graphs). Responses are shown at the following time points: baseline (before treatment) and at 1, 2, 4, and 6 hours after treatment with vehicle (0.5% methylcellulose, delivered PO) or carbamazepine (10, 30, 100, or 300 mg/kg, delivered PO). There were 4 rats per group. Kruskal-Wallis tests were performed at each timepoint for each sex, followed by Dunn’s tests when a significant effect was observed. Dunn’s tests found an increase in the number of observations relative to vehicle in males at 2 hours and 6 hours post-treatment with 300 mg/kg carbamazepine (p<0.05 for both comparisons). Dunn’s tests found an increase in the number of observations relative to vehicle in females at 4 hours post-treatment with 300 mg/kg carbamazepine (p<0.01) and at 6 hours post-treatment with 300 mg/kg carbamazepine (p<0.05). Data were analyzed using GraphPad Prism version 10.0.2.

Method: Modified Irwin functional observational battery

A heat map depicts the severity of the behaviors observed by two reviewers in a modified Irwin test at baseline and at 1, 2, 4, and 6 hours following the administration of vehicle (0.5% methylcellulose, delivered PO) or carbamazepine (10, 30, 100, or 300 mg/kg, delivered PO). There were 8 rats per group (4 male and 4 female), except for the 300 mg/kg carbamazepine group at 6 hours, which had 7 rats (4 male and 3 female). Severity score was calculated as the sum of scores given by the two reviewers for all animals at that dose and timepoint divided by the maximum possible score, transformed into a percentage. In summary, carbamazepine administration led to decreased locomotor activity, flaccid abdominal tone, and decreased visual placement, at the 300 mg/kg dose.

Method: Modified Irwin functional observational battery

Two graphs show the body temperature of male or female rats during the modified Irwin test. Responses are shown at the following timepoints: baseline (before treatment) and at 1 and 6 hours after treatment with vehicle (0.5% methylcellulose, delivered PO) or carbamazepine (10, 30, 100, or 300 mg/kg, delivered PO). There were 4 rats per group, except the 300 mg/kg carbamazepine group of females had 3 rats at 6 hour post-treatment. A significant effect of treatment was found in a two-way repeated measures ANOVA for female rats. Data were analyzed using GraphPad Prism version 10.0.2.

Method: Modified Irwin functional observational battery

Method: Rat Rotarod

Two graphs show the latency for male or female rats to fall off a rotarod apparatus. Responses are shown at the following time points: baseline (before treatment) and at 30, 60, and 90 minutes after treatment with vehicle (phosphate buffered saline, delivered SC) or nalfurafine (1, 3, 10, or 30 μg/kg, delivered SC). There were 10 rats per group. Males but not females had a significant interaction of time x treatment in a two-way repeated measures ANOVA. In males, the latency to fall in the 30 μg/kg nalfurafine group was approximately 30 seconds at each post-treatment time point, but this was not significantly different from the 40 second latency seen in the vehicle group. Data were analyzed using GraphPad Prism version 10.0.2.

Method: Rat Rotarod

Model: Plantar Incision Model in Rat (Brennan Model)
Endpoint: Rat Hind Paw Mechanical Allodynia Behavior (von Frey Filaments) Method

Two graphs show ipsilateral hind paw mechanical allodynia behavior assessed with von Frey filaments in male or female rats that have undergone plantar incision surgery. Responses are shown at the following time points: baseline (before surgery), prior to treatment at 1 day after surgery, and at 1, 2, 4, and 6 hours after treatment at 1 day after surgery. The treatments are vehicle (PBS, delivered SC), nalfurafine (0.3, 1, 3, or 10 μg/kg, delivered SC), or the positive control morphine (6 mg/kg, delivered SC). There were 10 rats per group. A significant interaction of time x treatment was found in a two-way repeated measures ANOVA for each sex. Dunnett’s tests found significantly higher withdrawal thresholds relative to vehicle in males at 1 hour post-treatment with morphine (p<0.0001) and at 2 hours post-treatment with 0.3, 1, and 10 μg/kg nalfurafine (p<0.05 for all comparisons) and morphine (p<0.01). Dunnett’s tests found significantly higher withdrawal thresholds relative to vehicle in females at 1 hour post-treatment with morphine (p<0.0001); at 2 hours post-treatment with 3 μg/kg nalfurafine (p<0.05), 10 μg/kg nalfurafine (p<0.01), and morphine (p<0.001); and at 4 hours post-treatment with 3 μg/kg nalfurafine (p<0.05), 10 μg/kg nalfurafine (p<0.05), and morphine (p<0.01).

Model: Plantar Incision Model in Rat (Brennan Model)
Endpoint: Guarding Behavior

Two graphs show ipsilateral hind paw guarding behavior evaluated in male or female rats that have undergone plantar incision surgery. Responses are shown at the following time points: baseline (before surgery), prior to treatment at 1 day after surgery, and at 1, 2, 4, and 6 hours after treatment at 1 day after surgery. The treatments are vehicle (PBS, delivered SC), nalfurafine (0.3, 1, 3, or 10 ìg/kg, delivered SC), or the positive control morphine (6 mg/kg, delivered SC). There were 10 rats per group. A significant interaction of time x treatment was found in a two-way repeated measures ANOVA for each sex. Dunnett’s tests found significantly lower guarding scores relative to vehicle in males at 1 hour post-treatment with 1, 3, and 10 ìg/kg nalfurafine (p<0.001, 0.01, 0.01, respectively) and morphine (p<0.0001) and at 2 hours post-treatment with 0.3, 1, and 3 ìg/kg nalfurafine (p<0.05, 0.01, 0.05, respectively) and morphine (p<0.0001). Dunnett’s tests found significantly lower guarding scores relative to vehicle in females at 1 hour post-treatment with 0.3, 1, 3, and 10 ìg/kg nalfurafine and morphine (p<0.0001 for all comparisons); at 2 hours post-treatment with 0.3, 1, 3, and 10 ìg/kg nalfurafine (p<0.001, 0.0001, 0.001, 0.0001, respectively) and morphine (p<0.0001); at 4 hours post-treatment with 0.3, 1, 3, and 10 ìg/kg nalfurafine and morphine (p<0.0001 for all comparisons); and at 6 hours post-treatment with 0.3, 1, 3, and 10 ìg/kg nalfurafine (p<0.0001, 0.001, 0.0001, 0.001, respectively) and morphine (p<0.0001).

Model: L5/L6 Spinal Nerve Ligation Model
Endpoint: Rat Hind Paw Mechanical Allodynia Behavior (von Frey Filaments) Method

Model: L5/L6 Spinal Nerve Ligation Model
Endpoint: Rat Hind Paw Cold Allodynia (Acetone Evaporation) Test

Two graphs show ipsilateral hind paw cold allodynia behavior assessed with 5 trials of the acetone evaporation test in male or female rats that have undergone L5/L6 spinal nerve ligation surgery. Responses are shown at the following time points: baseline (before surgery), prior to treatment at 3 weeks after surgery, and at 1 and 3 hours after treatment at 3 weeks after surgery. The treatments are vehicle (PBS, delivered SC), nalfurafine (0.3, 1, 3, or 10 ìg/kg, delivered SC), or the positive control gabapentin (60 mg/kg, delivered PO). There were 10 rats per group. A significant interaction of time x treatment was found in a two-way repeated measures ANOVA for each sex. Dunnett’s tests found significantly lower acetone responses relative to vehicle in males at 1 hour post-treatment with 1, 3, and 10 ìg/kg nalfurafine (p<0.05 for all comparisons) and gabapentin (p<0.01) and at 3 hours post-treatment with 1 ìg/kg nalfurafine (p<0.05) and gabapentin (p<0.05). Dunnett’s tests found significantly lower acetone responses relative to vehicle in females at 1 hour post-treatment with 0.3 ìg/kg nalfurafine (p<0.05), 10 ìg/kg nalfurafine (p<0.01), and gabapentin (p<0.05).

This work was conducted by PsychoGenics Inc. (Paramus, NJ) in collaboration with PSPP, NINDS, NIH under contract # 75N95019D00026. Prescribing information for clinically used controls can be found at labels.fda.gov.

Nalfurafine

Formulation

Results

Pharmacokinetics: Concentration in Plasma and Brain

Pharmacokinetics: Calculated Parameters

Irwin (1-30 µg/kg): Total Observations

Irwin (1-30 µg/kg): Behavioral Severity Scores

Irwin (1-30 µg/kg): Body Temperature

Irwin (100-3000 µg/kg): Total Observations

Irwin (100-3000 µg/kg): Behavioral Severity Scores

Irwin (100-3000 µg/kg): Body Temperature

Rotarod (1-30 µg/kg)

Rotarod (100-3000 µg/kg)

Plantar Incision: von Frey

Plantar Incision: Guarding Behavior

L5/L6 Spinal Nerve Ligation: von Frey

L5/L6 Spinal Nerve Ligation: Acetone