Chloride extrusion enhancers as novel therapeutics for neurological diseases

Abstract

The K+-Cl− cotransporter KCC2 is responsible for maintaining low Cl− concentration in neurons of the central nervous system (CNS), essential for postsynaptic inhibition through GABAA and glycine receptors. While no CNS disorders have been associated with KCC2 mutations, loss of activity of this transporter has emerged as a key mechanism underlying several neurological and psychiatric disorders including epilepsy, motor spasticity, stress, anxiety, schizophrenia, morphine-induced hyperalgesia and chronic pain1–9. Recent reports indicate that enhancing KCC2 activity may be the favoured therapeutic strategy to restore inhibition and normal function in pathological condition involving impaired Cl− transport10–12. We designed an assay for high-throughput screening which led to the identification of KCC2 activators that reduce [Cl−]i. Optimization of a first-in-class arylmethylidine family of compounds resulted in a KCC2-selective analog (CLP257) that lowers [Cl−]i. CLP257 restored impaired Cl− transport in neurons with diminished KCC2 activity. The compound rescued KCC2 plasma membrane expression, renormalised stimulus-evoked responses in spinal nociceptive pathways sensitized after nerve injury and alleviated hypersensitivity in a rat model of neuropathic pain. Oral efficacy for analgesia equivalent to that of Pregabalin but without motor impairment was achievable with a CLP257 prodrug. These results validate KCC2 as a druggable target for CNS diseases.

KCC2 function modulates in vitro ictogenesis

“…Suppression of KCC2 during enhanced excitation is thought to alter the balance of excitation and inhibition that in turn may lead to anomalous activity such as seizures (Huberfeld et al, 2007;Rivera et al, 2002Rivera et al, , 2004. Surprisingly, we have found that enhancing KCC2 activity with CLP257 (Gagnon et al, 2013) increased the duration of ictal events without affecting their interval of occurrence. These results suggest that downregulation of KCC2 might not be the cause of seizures in epilepsy but rather an adaptive mechanism to prevent the increase in extracellular [K + ] due to the excessive activation of GABA A receptors and KCC2 activity.…”

“…In addition, expression and function of KCC2 are tightly regulated by neuronal activity in a way that enhanced excitation results in suppression of KCC2 activity (Huberfeld et al, 2007;Rivera et al, 2002Rivera et al, , 2004. Indeed, restoring KCC2 activity by application of CLP257 increased the duration of isolated GABAergic events (Gagnon et al, 2013).…”

A Novel Regulatory Locus of Phosphorylation in the C Terminus of the Potassium Chloride Cotransporter KCC2 That Interferes with N-Ethylmaleimide or Staurosporine-mediated Activation*♦

“…This might be important for finetuning the operational mode of KCC2 and to integrate different signaling pathways. These findings also imply that drugs aimed to modify KCC2 phosphorylation for therapeutic benefit (36,73) have to be investigated under different conditions as their effect might depend on the conformational state of KCC2 and therefore be highly context-specific. Functional cross-talk between different regions has also been suggested for NKCC1, where phosphorylation of the N terminus causes movement of the C terminus, which in turn entails altered interactions between different transmembrane domains (74).…”

Regulation of neuronal chloride homeostasis by neuromodulators

“…2013). Although the precise mechanism of the drug action on KCC2 is currently unknown, CLP257 was able to act as a chloride extrusion enhancer in a neuropathic pain model, and restore Cl − transport in adult spinal cord slices with impaired KCC2 function.…”

“…While the compound itself exhibited modest binding to adenosine receptors and other classes of GPCRs (Gagnon et al . 2013), future studies must systematically examine whether G q ‐GPCR agonists could also work as novel Cl − extrusion enhancers under physiological and/or pathological states.…”