Abstracts

Rationale: TARPγ8, a transmembrane AMPA receptor regulatory protein (TARP), is enriched in regions associated with seizures (cortex, amygdala, hippocampus) with little to no expression in hindbrain. Therefore, TARPγ8 provides a mechanism to target epilepsy-associated brain regions with an improved tolerability profile. Here, two TARPγ8 negative allosteric modulators (NAMs), RAP-219 and RTX-1738, were assessed in preclinical seizure models.

Methods: Subjects were adult male CF-1 mice (Charles River, Portage, MI).

RAP-219 (single dose, po, vs vehicle): pentylenetetrazol (PTZ) and corneal kindling (CK) models. PTZ was infused intravenously 2h post-dose (vehicle or RAP-219: 0.01, 0.1, 1 mg/kg [n=10]); time from start of infusion to first twitch and onset of sustained clonus were recorded. Mice were fully kindled (Racine scale 5) for the CK model one day pre-treatment (vehicle or RAP-219: 0.01, 0.02, 0.03, 0.1, 0.3, 1, 3 mg/kg, [n=8]). Two hours post-dose and immediately prior to corneal stimulation challenge, animals were tested on rotarod to assess motoric impairment, with failure defined as 3 falls.

RTX-1738 (0.003, 0.01, 0.03, 0.1, 0.3 mg/kg, po, vs vehicle): acute (single dose) or chronic (7 days, QD) in the CK model. Locomotor activity was assessed with the open-field test (15 min) 24h after the final dose in the chronic study. Total distance traveled and horizontal/vertical activity count were recorded.

Receptor occupancy (RO) was determined via ex vivo autoradiography 4h post-dose as a function of plasma concentration in rat. Group means were compared using ANOVA, followed by Dunnett’s post hoc analysis. Data were fitted to Hill functions using nonlinear least-squares analysis to estimate ED₅₀ and plasma EC₅₀.

Results: RAP-219: 0.1 and 1 mg/kg doses in the PTZ model were associated with significantly higher threshold vs vehicle (P< 0.01). ED₅₀ values for twitch and clonus were 0.02 mg/kg, with EC₅₀ values of 2.9 and 2.4 ng/mL, respectively (Fig 1). In CK, mice were protected from seizures (ED₅₀, 0.02 mg/kg; EC₅₀, 2.3 ng/mL) by RAP-219 treatment, which also showed minimal, non-dose-dependent failures on the rotarod (therapeutic index >150 [Fig 2]).

RTX-1738: In CK, mice were protected from seizures (acute: ED₅₀, 0.05 mg/kg; EC₅₀, 4.7 ng/mL; chronic: ED₅₀, 0.01 mg/kg; EC₅₀, 3.6 ng/mL). No differences in locomotor activity were observed between vehicle and any RTX-1738 dose.

RO analysis of RAP-219 and RTX-1738 yielded EC₅₀ values of 3.3 ng/mL and 3.2 ng/mL, respectively. In the CK model, RAP-219 and RTX-1738 showed full protection at RO of 70% and 90%, corresponding to plasma concentrations of 7 ng/mL and 10 ng/mL, respectively.

Conclusions: Selective inhibition of AMPA receptors containing TARPγ8 provided potent, dose-dependent antiseizure effects that were maintained after 7-day dosing period in preclinical models of focal and generalized seizures. Exposures that produced antiseizure effects did not impair motor function, yielding a high therapeutic index. TARPγ8 NAMs may provide a new precision medicine option for patients with epilepsy with improved tolerability.

Funding: Funded by Rapport