Dose Range-Finding Report on the Immunotoxicity of Methadone in Female B6C3F1 Mice (CAS No. 1095-90-5)
The following abstract presents results of a study conducted by a contract laboratory for the National Toxicology Program. The findings have not been peer reviewed and were not evaluated in accordance with the levels of evidence criteria established by NTP in March 2009. The findings and conclusions for this study should not be construed to represent the views of the NTP or the U.S. Government.
Since methadone has a good oral bioavailability, it has been used in maintenance programs for the past 29 years in an attempt to limit the use of the intravenous route of administration by abusers of opiates, such as heroin. Previously, the effects of morphine on a number of immune parameters were explored. The present study sought to determine whether methadone had similar effects on the immune status of female B6C3F1 mice.
Methadone was nominated to the NTP for toxicological evaluation and was selected for immunotoxicity studies by the chemical manager. Thus, the purpose of these range-finding studies was to select the doses and time of administration of methadone for the protocol to evaluate the potential effects on the immune system.
The studies were conducted in female B6C3F1 mice. The animals were administered methadone injections on day 1 through day 5 every 6 hours at doses of 5, 10, and 20 mg/kg to evaulate body weight, organ weight and the T-dependent antibody response. Methadone was administered by subcutaneous injection as a solution in sterile distilled water. Additional studies were performed to determine the methadone serum levels. In these range-finding studies, serum methadone and corticosterone levels peaked one hour following a single subcutaneous injection of 20 mg/kg methadone HCl (Figure 3). After a single injection with 20 mg/kg methadone, the pharmacokinetics revealed a serum half-life of nearly equal to 2 hours (Figure 2). Following five injections over a 24 hour period (every 6 hours), methadone levels were elevated as would be expected; however, corticosterone levels were not increased (Figure 4). This suggested that the ability of methadone to elevate corticosterone became uncoupled following repeated dosing, indicative of a tolerance mechanism. Moreover, dosing every 6 hours for five days induced an increase in the catabolism of methadone itself (Figure 5). Therefore, all protocol assays began one hour after subcutaneous administration of methadone HCl at doses of 10, 20, and 30 mg/kg, a time at which both methadone and corticosterone serum levels were elevated.
For the range-finding study, body weight, body weight change and organ weights were measured for toxicolological parameters. Results showed a 72% decrease in body weight change after five days at the low dose of 5 mg/kg and a 70% decrease at the high dose of 20 mg/kg. Within the organ weight data, the spleen was significantly decreased in weight at the 10 and 20 mg/kg doses by 12% and 24%, respectively. The spleen percent body weight was also decreased by 11% at the 10 mg/kg dose and 22% at the 20 mg/kg dose. The percent body weight of the liver at the 20 mg/kg dose was increased by 7%. The thymus weight was decreased at the 10 and 20 mg/kg doses by 43% and 55%, respectively. The percent body weight of the thymus was also decreased 42% at the 10 mg/kg dose and 53% at the 20 mg/kg dose. The percent body weight of the lungs was increased 17% at the low dose of 5 mg/kg and 15% at the high dose of 20 mg/kg.
The immunologic parameter evaluated in the range-finding study was the T-dependent antibody response. Mice exposed to methadone HCl on day 1 up to day 5 produced no effect on the IgM antibody-forming cell per 106 spleen cells. Along with the spleen weight decrease, the spleen cell number was decreased at doses of 5, 10 and 20 mg/kg by 14%, 21% and 36%, respectively. The immunologic assay was performed one hour after the last exposure to methadone. In addition, an in vitro T-dependent antibody response was performed and the data showed no effect.
In conclusion, all assays for the protocol began one hour after one subcutaneous administration of methadone HCl at doses of 10, 20, and 30 mg/kg, a time at which both methadone and corticosterone serum levels were elevated.