UC San Diego

Traumatic brain injury (TBI) is a major public health concern that can result in long-term neurological impairments. Calpain is a calcium-dependent cysteine protease that is activated within minutes after TBI and sustained calpain activation is known to contribute to neurodegeneration and blood-brain barrier dysregulation. Based on its role in disease progression, calpain inhibition has been identified as a promising therapeutic target. Efforts to develop therapeutics for calpain inhibition would benefit from the ability to measure calpain activity with spatial precision within the injured tissue. In this work, I have engineered nanomaterials that can be used to measure and/or inhibit calpain activity after TBI. First, I used a hyaluronic acid-targeted activity-based nanosensor for TBI (HA-targeted TBI-ABN) to measure calpain activity in the cortex, hippocampus, and hindbrain after injury in both female and male mice. In a mouse model of TBI, I show that the HA-targeted TBI-ABN detects high calpain activity in the cortex and hippocampus ipsilateral to injury in both female and male mice. Second, I designed an activity-based nanotheranostic for TBI (TBI-ABNT) that can both sense and inhibit calpain activity. To inhibit calpain activity, I incorporated calpastatin peptide (CAST), an endogenous inhibitor of calpain. In a mouse model of TBI, I show that the TBI-ABNT construct is able to sense and inhibit calpain activity in the injured cortex and hippocampus. In an analysis of cellular calpain activity, I observe ABNT-mediated inhibition of calpain activity in neurons, endothelial cells, and microglia of the cortex. In a comparison of neuronal calpain activity by brain structure, I observe greater ABNT-mediated inhibition of calpain activity in cortical neurons compared to hippocampal neurons. Furthermore, I found that apoptosis was dependent on both calpain inhibition and by brain structure. Lastly, I designed a calpasatin peptide-based nanomaterial (CAST-PEG) to improve the pharmacokinetics of therapeutic calpastatin peptide. I show that a high valency of CAST per polymeric scaffold improves inhibitory activity, blood half-life, and biodistribution compared to free CAST. In a mouse model of TBI, I see a dose-dependent improvement in the accumulation of CAST-PEG in the injured brain compared to off-target organs. Finally, I show that neither CAST nor CAST-PEG delivery reduces calpain-specific α-spectrin breakdown products at 24 hours after injury. I present three platforms that can be used to understand the regional- and cell-specific activity of calpain and its inhibition to inform therapeutic interventions for TBI.