P3‐030: NIEMANN‐PICK DISEASE TYPE C2 PROTEIN (NPC2; HE1) IS A CSF BIOMARKER OF MACROAUTOPHAGY

Macroautophagy (MA) is an inducible process for the cellular clearance of proteins, macromolecules and organelles. Perturbations in neuronal MA have been implicated in several neurodegenerative disorders, and the induction of this mechanism to clear toxic protein aggregates is a potential therapeutic approach to protein misfolding disorders. Since MA is an entirely intracellular process, monitoring disease or drug-induced alterations in MA in the CNS poses a dilemma. Therefore, we identified a secreted marker of this process computationally and confirmed our findings in vitro and in vivo. We constructed a protein-protein interaction network of autophagy-associated genes/proteins and used a network building algorithm to discover neighboring interactions with proteins that were known or predicted to be secreted by two different methods (PSORT and Sec-HMMER). Using this approach, we identified Niemann-Pick disease type C2 protein (NPC2: HE1) as a candidate secreted biomarker of MA. To verify NPC2, we induced MA in human H4 neuroglioma cells by treatment with rapamycin (RAP), amitryptaline (AMI: a brain-penetrant inducer of autophagy) or nutrient (serum) starvation, and monitored LC3 II maturation. 3-methyladenine (3MA) treatment was used to inhibit nutrient deprivation-induced MA. In order to verify that NPC2 levels are modulated in vivo by autophagy inducers, we treated rats with AMI and monitored levels of NPC2 in the CSF. Induction of MA by RAP, AMI or nutrient deprivation as monitored by LC3 II maturation resulted in increases in NPC2 levels in conditioned media (CM). Nutrient-deprivation induced MA was inhibited by 3MA, and caused corresponding reductions in NPC2 levels in CM. CSF levels of NPC2 increased in response to AMI treatment in rats. Our data suggest that NPC2 is a secreted CSF biomarker of MA.