Inhibition of ApoE4 Endocytosis with LDLR-LA Peptides

The Apolipoprotein E4 (ApoE4) genotype is the most significant genetic risk factor for late-onset Alzheimers disease (AD). A key driver of ApoE4 cellular toxicity is the endo-lysosomal burden resulting from the excessive receptor-mediated uptake of ApoE4 lipoparticles. The high-affinity interaction between lipidated ApoE4 and the Low-Density Lipoprotein Receptor (LDLR) saturates the cellular degradation machinery, correlating with lysosomal alkalinization, lipid accumulation, and cell death. To target this critical interaction interface, which consists of 7 tandem ligand-binding type-A (LA) modules in the human LDLR, we present the design and evaluation of recombinant LDLR minireceptors comprising combinations of these LA modules to competitively antagonize ApoE4 endocytosis. We observe a distinct isoform-dependent uptake dynamic across multiple central nervous system (CNS) cell models, with ApoE4 showing significantly greater total intracellular accumulation than ApoE2. Furthermore, engineered LA peptides selectively bind ApoE4 over human serum LDL and differentially inhibit its uptake, revealing a distinct structural efficacy hierarchy of LA3456 [~] LA345 > LA456 [~] LA45 >> LA34. We establish the resilience of the LA45 minireceptor under physiological serum conditions and identify LA345 as the most stable truncated construct in vitro. Notably, molecular tagging orientation is critical for therapeutic engineering; C-terminal tagging completely preserves the inhibitory function of the minireceptors, whereas N-terminal tagging drastically reduces it. These findings provide a framework for scalable, deliverable inhibition of the ApoE4-LDLR interaction as a potential therapeutic target to mitigate endo-lysosomal accumulation in AD.