NLX-101 ameliorates responses to hypercapnia in a mouse model of Rett syndrome

A. Abdala Sheikh¹, I. Charles¹, L. Koolen¹, M. Varney², A. Newman-Tancredi². ¹School of Physiology, Pharmacology and Neuroscience, University of Bristol, Bristol, United Kingdom, ²Neurolixis Inc., Dana Point, CA, USA.

Introduction: Rett syndrome is a neurodevelopmental disorder caused by methyl-CPG-binding protein 2 (MECP2) deficiency. It frequently features ataxic and Cheyne-Stokes breathing. Mouse models of Rett showed reduced hypercapnic ventilatory responses and elevated apnoeic threshold (1) . This phenotype likely contributes to ataxic breathing in other conditions, and hypercapnia improved breathing in a patient with Rett (2). Recently, we demonstrated that NLX-101 (aka F15599), a potent, selective and efficacious agonist with bias for 5-HT_1A heteroreceptors (3), improved ataxic breathing in two Rett mouse models. Thus, we investigated the hypothesis that NLX-101 ameliorates impaired hypercapnic ventilatory responses in MECP2 deficient mice.

Methods: We measured minute ventilation (VE) in Mecp2^tm1.1Coyle heterozygous (R168X) and littermate wild-type (WT) female mice (15±0.3 months-old) using unrestrained whole-body plethysmography. Mice were exposed to increasing fractions of inspired CO₂ (FICO₂; 0, 1, 3 and 5%) at 5min intervals. Hypercapnia was performed 20 min after NLX-101 (1.25mg/kg, ip) or vehicle (2.5% DMSO/H₂O, 5mL/kg ip) treatment. Study arm assignment was randomized. Values shown are mean±SEM and were compared using two-factor ANOVA and Tukey’s post hoc (P<0.05).

Results: Under 5% FICO₂, VE was significantly lower in vehicle-treated R168X mice compared to WT (2.52±0.27 vs 3.64±0.16 mL/min/g, P<0.001, n=8). Treatment with NLX-101 significantly increased VE in R168X mice (3.46±0.33 mL/min/g, n=8) compared with vehicle-treated R168X (P<0.01), rescuing it to WT levels (P>0.99). NLX-101 had no effect on VE in WT mice compared to vehicle-treated WT (3.76±0.21 mL/min/g, P>0.99, n=8). NLX-101-treated R168X mice showed a trend to increased VE at lower FICO₂ but these were not significantly different from vehicle-treated R168X. Under 0% FICO₂, NLX-101 significantly reduced apnoea frequency (60±18 vs 295±71 apnoeas/hr, P<0.01, n=8) and duration (507±110 vs 911±150 ms, P<0.01, n=8) compared to vehicle in R168X. Occurrence of apnoeas in vehicle-treated R168X mice was inversely related to FICO₂.

Conclusion: The results demonstrated that NLX-101 rescued hypercapnic ventilatory responses, and reduced apnoea frequency and duration in R168X mice. This positive effect on CO₂ chemosensitivity may be a mechanism by which NLX-101 rescues ataxic breathing in MECP2 deficient mice. The R168X mouse model recapitulates a common mutation occurring in patients with Rett, and has well characterized face validity for translation (4).

References:

1) Toward MA et al. (2013). Exp Physiol 98: 842-9.

2) Smeets EE et al. (2006). Brain Dev 28: 625-32.

3) Newman-Tancredi A (2011). Neuropsychiatry 1: 149-164.

4) Schaevitz LR et al. (2013). Genes Brain Behav 12: 732-40.

Funding: NIH:1R01AT008632-01 and Neurolixis Inc.