The objective of this study was to assess the relative reinforcing efficacy of the test article, DMX-101, in Sprague-Dawley rats conditioned to self-administer hydrocodone, a Schedule II controlled substance, during daily access periods.

Three-day substitution test sessions were instituted in rats trained to self-administer hydrocodone. Complete substitution was defined as drug-maintained lever-press responding for 3 days at levels similar to that which was maintained by the maintenance dose of hydrocodone.

Each animal was previously instrumented with a chronic indwelling jugular vein catheter that was connected to a computer-controlled drug delivery system via a stainless-steel tethering system. The tethering system was connected to the exteriorized catheters and protected by nylon vests worn by the animals. An initial pool of 32 male rats was conditioned to self-administer 0.18 mg/kg/injection hydrocodone in a standard operant single-lever training procedure under a fixed-ratio 10 (FR10) schedule of reinforcement until self-administration was established. Replacement rats were added to study as needed to retain an adequate pool of animals. From the pool of trained rats, a total of 27 animals each were tested with the saline control and the 0.18 mg/kg/injection hydrocodone maintenance dose. Following the establishment of hydrocodone as a reinforcer, 6 rats per each treatment were then tested with the positive control article (0.01, 0.018, 0.032, and 0.1 mg/kg/injection hydrocodone), and/or DMX-101 (0, 0.1, 0.18, and 0.28 mg/kg/injection) during the study via intravenous bolus injections through surgically-implanted venous catheters. Operant testing sessions with saline or hydrocodone were considered “test sessions” and exposures to the test article or its vehicle were considered “substitution sessions”. All treatments were administered at a dose volume of approximately 37 μL to 55 μL per injection during 1-hour access periods. Operant testing and substitution sessions were conducted once per day for 3 consecutive sessions at each treatment. There were no DMX-101-related mortalities, clinical or veterinary observations, or body weight effects observed.

Hydrocodone initiated and maintained self-administration across a wide range of doses, Consistent with its robust profile of reinforcing effects, and generated the well-characterized inverted U-shaped dose-effect curve, confirming rats were adequately trained and serving as an internal validation before further assessment.

Following a washout period of at least 12 days following the last DMX-101 dose administered, 6 designated animals were subjected to non-contingent administration and plasma samples were analyzed for DMX-101. DMX-101 was detected in rat plasma samples from all DMX-101-dosed animals and at all intervals (inclusive of predose). Buprenorphine (BPN) was detected in 3 rat plasma samples after DMX-101 dose administration. Plasma levels of BPN (Animal No. 1023) were 1.74 ng/mL at 30 minutes and 1.8 ng/mL at 60 minutes; these values reflect 0.004% and 0.003% of total plasma drug levels, respectively. Plasma level of BPN (Animal No. 1038) was 2.72 ng/mL at 60 minutes; this value reflects 0.003% of total plasma drug level. Per DIMERx, buprenorphine is a metabolic product of the synthetic intermediate, vinyl-oxy buprenorphine.

Plasma concentration analyses indicate when animals were allowed to self-administer unlimited amounts of 0.28 mg/kg/injection DMX-101, average plasma concentrations reached up to 25.5-fold Cmax therapeutic (1402 ng/mL) by session end.

At the group level, DMX-101 did not maintain self-administration in rats conditioned to self-administer 0.18 mg/kg/injection hydrocodone under a fixed-ratio 10 schedule in daily 1-hour unlimited access test sessions, although acute reinforcing effects were observed in a single animal. Response patterns observed within and between sessions for DMX-101 indicate that DMX-101 did not serve as a robust reinforcer in this assay at doses selected to approximate 2.8-, 5.4-, and 8-fold Cmax therapeutic. 

Animals were conditioned to self-administer hydrocodone at 0.18 mg/kg/injection. Once self-administration was established with hydrocodone, various doses of the hydrocodone and its vehicle (saline), as well as the test article and its vehicle, were administered during 1-hour access periods over the course of 3 consecutive days. Substitution tests were interspersed with hydrocodone maintenance training sessions; at least 3 hydrocodone self-administration sessions with ≤ 20% variability were imposed between treatments to ensure that self-administration of the hydrocodone maintenance dose remained stable during the testing period. At least 6 animals were tested at each treatment. A test condition constituted 3 consecutive sessions of testing at any given treatment.

The first 2 tests in the series of tests conducted on this study were with 0.18 mg/kg/injection of hydrocodone and saline to clearly identify that animals were self-administering the hydrocodone maintenance dose prior to initiating any other test sessions. The positive control article dose-response curve was generated prior to test article dosing.

A total of 27 animals from the pool demonstrated adequate stimulus control and were tested with 0.18 mg/kg/injection hydrocodone and saline.

Training/maintenance dose (MD) level of hydrocodone used throughout the study.

The following parameters and endpoints were evaluated in this study: mortality, clinical signs, body weights, and behavioral measures (total number of injections during the session, total drug intake during the session, and rate of responding throughout test session). Bioanalytical assessment was conducted to determine plasma concentrations of the test article from 6 animals at 0.28 mg/kg/injection DMX-101, at designated time points on a separate phase of the study following completion of the self-administration testing phase.

Experimental Design:

Descriptive statistical analysis included the following: Means, standard deviations, least squares means, least squares mean standard errors, ratio, percentages, numbers, and/or incidences were reported as appropriate by dataset.

Inferential statistical analysis: All statistical tests were conducted at the 5% significance level. All pairwise comparisons were conducted using two-sided tests and were reported at the 1% and 5% levels, unless otherwise noted. Equivalent tests were reported depending on the terminology defined in the software used in the statistical analysis. The pairwise comparisons of interest are listed below: 

For each endpoint, the model tested for the effects of treatment, time and the interaction between them.