Abstracts

Rationale: Fenfluramine (FFA) treatment provides clinically significant reductions in convulsive seizure frequency and improved executive function in patients with Dravet syndrome (DS). FFA also reduces sudden unexpected death in epilepsy (SUDEP) in mouse models of DS. Epidemiological evidence suggests SUDEP rates are reduced in patients treated with FFA relative to standard of care. Recent studies have begun to elucidate FFA’s mechanism of action at serotonin and Sigma-1 receptors (S-1R). Studies in an scn1a mutant zebrafish model of DS demonstrate that FFA reduces epileptiform activity, reduces hypo-locomotion, and preserves dendritic arborization through activity at both S-1R and 5HT receptor subtypes, most notably 5HT2A, 5HT2C, and 5HT1D. In this study, we investigated FFA effects upon survival, myelination, and neuroinflammation in a mouse model of DS.

Methods: Starting on postnatal day [PND] 7, scn1a+/- mice (DS mice) were treated subcutaneously once daily with FFA (15 mg/kg; n=17), diazepam (10 mg/kg; n=24), or no treatment (n=115) until PND 35-37. Survival was evaluated by Kaplan-Meier/Wilcoxon test. FFPE sagittal brain sections were immunostained with D-MBP antibody for degenerated myelin and CD11b antibody for inflammatory microglia. CD11b+ microglia phagocytose damaged myelin and thus are a secondary marker for myelin damage. For D-MBP, whole cortex and hippocampus were scored semi-quantitatively on a (0-5) scale, and areas of staining in the parietal cortex and CA3 region were quantitated. For CD11b, areas of corpus callosum and hippocampus were scored semiquantitatively on a (0-5) scale. Statistical significance for histopathological markers was assessed by t test.

Results: By PND 38, 55% and 62% of no treatment and diazepam-treated DS mice died; all deaths that were observed appeared to be due to SUDEP. In animals receiving FFA treatment, the mortality rate was reduced to 24% (P< 0.01; Figure 1). Degenerated myelin was enriched in DS mouse cortex and hippocampus compared to wild-type controls. FFA treatment, but not diazepam treatment, significantly reduced degenerated myelin, seen by semiquantitative scoring (P< 0.05) and quantitation (P< 0.001) of D-MBP staining in these DS tissues. Activated CD11b+ microglia were enriched in DS mouse corpus collosum and hippocampus relative to wild-type controls. FFA treatment, but not diazepam treatment, significantly reduced CD11b+ activated microglia in these DS tissues (P=0.05).

Conclusions: This is the first report of increased survival and reductions in myelin degeneration and neuroinflammation in DS mice treated with FFA compared with vehicle or diazepam. Improved survival and reduced myelin damage and neuroinflammation corroborate findings in zebrafish models of DS, demonstrating the claim that FFA exhibits disease-modifying effects in a second animal model of DS. These observations in mice support the clinical benefits observed with fenfluramine treatment in seizure control, executive function, and all-cause mortality/SUDEP.

Funding: Please list any funding that was received in support of this abstract.: Zogenix, Inc.