Capturing Altered Brain Proteomic Signaling

Background: The prevalence of cannabis use is highest amongst adolescents and young adults, who are also more vulnerable to the development of Cannabis Use Disorder (CUD) and its neuropsychiatric and cognitive consequences. This is supported by preclinical data demonstrating neuronal/synaptic pathology and immunomodulatory changes with repeated cannabinoid exposure especially in adolescents. The underlying neuropathology in humans, however, remains unclear, in part due to difficulty in obtaining brain tissues from patients with CUD. Exosomes are vesicles, that are secreted from various cell types in the brain including neurons, and which can be isolated from blood. Importantly, exosomes cross the blood brain barrier with their contents intact, making them particularly attractive vehicles for potential signatures of neuropathology in the periphery. Proteomic analyses of exosome protein cargo may help identify patterns of altered protein signaling in the brain associated with recurrent cannabis exposure. Advances in Mass Spectrometry (MS) approaches present an unprecedented opportunity for profiling proteomes in body-fluids or within organelles of interest (e.g. exosomes). MS approaches have been used to evaluate the exosomal proteome of disease states but this promising approach has not been examined in CUD. We propose to examine the proteomic changes related to recurrent cannabis exposure in plasma neuron derived exosomes (NDE) using a label-free proteomics approach to identify broad proteomic signatures of altered signaling in CUD.

Specific Aim 1: To collect plasma samples from 20 adolescents and young adults with CUD and those without any exposure to cannabis (n = 10, 5 females in each group), from the ongoing studies. Extraction of NDE from the total pool of plasma exosomes (TPE) will be done under an MTA with NeuroDex (https://www.neurodex.co/) a company with proprietary technology, ExoSORT for capturing NDE.

Specific Aim 2: To examine the broad differential proteomic profile of TPE and NDE in CUD compared to matched HCs in a label-free, discovery proteomics approach using MS. To examine the enrichment of the differentially expressed proteins to cellular components, biological processes, and molecular functions with in silico bioinformatics approaches with a focus on brain specific proteins. To specifically examine the enrichment of proteins of relevance to synaptic function using SynGo database. To validate selected proteins with western blot.

Exploratory Aim: To examine the impact of acute exposure to cannabinoids (delta-9-tetrahydrocannabinol and cannabidiol) on plasma composition and size distributions of TPEs and NDEs.

Future directions: The data collected in this pilot project will be used to support larger grant applications to NIH for examining proteomic signatures in CUD. The results of this analysis will provide valuable preliminary confirmation regarding the feasibility of deriving CNS relevant signals from the periphery in developmental neuropsychiatric disorders. This would also allow us to explore the biophysical properties of TPE and NDE in further detail in future studies. Replication of the results of this study in larger studies will have numerous translational implications for research and clinical care of persons with substance use disorders.