Abstracts

Rationale: Many brain insults including traumatic brain injury, stroke and status epilepticus (SE) cause the later development of epilepsy. However, there are currently no treatments that prevent epileptogenesis following brain injuries. Recent data shows that inhibiting the JAK/STAT pathway with the compound WP1066 at the time of brain insult (SE) attenuates the severity of the ensuing epilepsy; specifically, rats given WP1066 show a significant reduction in the frequency of spontaneous seizures compared to vehicle-injected animals up to 4 weeks following SE. However, unfavorable pharmacokinetic (PK) properties of WP1066 limit its potential for preclinical development as WP1066 is rapidly cleared from blood and brain and only transiently reduces activation of the JAK/STAT pathway. Therefore, the purpose of this study is to determine stability and PK characteristics of a novel JAK/STAT inhibitor to determine its potential for clinical development as a disease modifying therapy for epilepsy. Methods: We have recently initiated PK studies on a novel JAK/STAT inhibitor, Ruxolitinib, aimed at increasing the efficacy of JAK/STAT inhibition in brain following peripheral administration. Ruxolitinib was also tested for its ability to inhibit STAT3 phosphorylation (pSTAT3) via administration 30 min after onset of SE. Animals were sacrificed at various time points following SE and expression of pSTAT3 was measured via Western blot analysis. Video-EEG monitoring was performed to assess the chronic effects of drug injections on the frequency, duration, and severity of spontaneous seizures. Results: Compared to WP1066, Ruxolitinib shows increased stability, improved blood-brain barrier penetration (IC₅₀ = 8.58 nM), and greater inhibition of pSTAT3 in both cultured hippocampal neurons and in brain following pilocarpine-induced SE in vivo. In our preliminary studies, Ruxolitinib showed a 20-fold increase in cortex concentrations compared to WP1066 in normal rats 1 hour after injection (n = 4). In addition, Western blot analysis of rat hippocampi 1 hour after SE showed almost complete inhibition (p < 0.001, n = 4) and ~45% reduction of pSTAT3 levels at 3 hours (p < 0.05, n = 4) compared to vehicle-injected controls. Ruxolitinib also appears to reduce the frequency of spontaneous seizures compared to vehicle-injected animals. Conclusions: These findings demonstrate that Ruxolitinib has improved PK properties and pSTAT3 inhibition compared to previously investigated JAK/STAT inhibitors, and suggest that Ruxolitinib is a promising candidate for further pre-clinical development as a disease-modifying agent with the potential to reduce development and/or severity of epilepsy following brain insults.