The CaMKIIα hub ligand Ph-HTBA promotes neuroprotection after focal ischemic stroke by a distinct molecular interaction

Ca2+/calmodulin-dependent protein kinase II alpha (CaMKII alpha) is a potential target for acute neuroprotection due to its key role in physiological and pathological glutamate signaling. The hub domain organizes the CaMKII holoenzyme into large oligomers, and additional functional effects on holoenzyme activation have lately emerged. We recently reported that compounds related to the proposed neuromodulator gamma-hydroxybutyrate (GHB) selectively bind to the CaMKII alpha hub domain and increase hub thermal stabilization, which is believed to have functional consequences and to mediate neuroprotection. However, the detailed molecular mechanism is unknown. In this study, we functionally characterize the novel and brain permeable GHB analog (E)-2-(5-hy-droxy-2-phenyl-5,7,8,9-tetrahydro-6H-benzo[7]annulen-6-ylidene)acetic acid (Ph-HTBA). Administration of a single dose of Ph-HTBA at a clinically relevant time point (3-6 h after photothrombotic stroke) promotes neu-roprotection with a superior effect at low doses compared to the smaller GHB analog 3-hydroxycyclopent-1-ene-carboxylic acid (HOCPCA). In contrast to HOCPCA, Ph-HTBA reduces Ca2+-stimulated CaMKII alpha Thr286 autophosphorylation in primary cortical neurons and substrate phosphorylation of recombinant CaMKII alpha, potentially contributing to its neuroprotective effect. Supported by previous in silico docking studies, we suggest that Ph-HTBA makes distinct molecular interactions with the hub cavity, which may contribute to its differential functional profile and superior neuroprotective effect compared to HOCPCA. Together, this highlights Ph-HTBA as a promising tool to study hub functionality, but also as a good candidate for clinical development.

Automated summary

Ca2+/calmodulin-dependent protein kinase II alpha (CaMKII alpha) is a potential target for acute neuroprotection. A novel GHB analog, Ph-HTBA, was found to promote neuroprotection after stroke, potentially through reducing CaMKII alpha phosphorylation. Ph-HTBA may have distinct molecular interactions with the hub cavity, contributing to its superior neuroprotective effect compared to other GHB analogs.