Alzheimer’s disease (AD) is the most common age-related neurodegenerative disease. Biologics with potential to affect underlying pathophysiology have failed to display significant therapeutic benefits in the clinic. The inability of biologics to penetrate the blood-brain barrier (BBB) represents their biggest hurdle to clinical efficacy. A promising strategy to address this challenge is to fuse IgGs to a second affinity ligand that engages a cerebrovascular endothelial target and induces transport across the BBB. While nearly all prior efforts have focused on the transferrin receptor (TfR-1) as the prototypical endothelial target despite inherent delivery and safety challenges, we have developed a bispecific antibody shuttle that engages CD98hc, the heavy chain of the large neutral amino acid transporter (LAT-1). Notably, our preliminary data show preserved affinity and specificity of off-the-shelf IgGs incorporated into our first-generation CD98hc bispecific shuttle, superior brain retention of IgGs shuttled via CD98hc as compared to TfR-1, and preserved endothelial levels of CD98hc and brain uptake after saturating CD98hc shuttle doses.The overall objective of this proposal is to further the development of the CD98hc BBB shuttle by: i) defining the optimal characteristics for maximal IgG brain parenchymal delivery; ii) evaluating age- and disease-dependent effects on CD98hc-mediated transport; and iii) generating proof-of-concept efficacy data in an AD mouse model using agonist (anti-TrkB) and antagonist (anti- phospho-tau) antibodies alone and in combination. Our central hypothesis is that IgG transport via the CD98hc shuttles will increase parenchymal delivery and achieve brain concentrations and target engagement comparable to viral delivery, generating robust neuroprotection in a tau mouse model, including potential additive or synergistic neuroprotective effects by targeting two different neuroprotective pathways. Therefore, we propose in Aim 1 to optimize brain parenchymal delivery of a second-generation CD98hc antibody shuttle. We will test the impact of the valency, affinity, and epitope of CD98hc interaction on brain uptake and parenchymal delivery. The optimal dose and construct identified using adult mice will be tested in aged wild-type and PS19 (tau P301S) mice to evaluate the integrity of the transport pathway. Next, in Aim 2, we will assess the efficacy of individual neuroprotective antibody/CD98hc shuttles in PS19 mice. Validated antibodies against TrkB (C20, agonist) and phosphorylated tau (PHF1) will be individually delivered via our best CD98hc shuttle. Efficacy outcomes will be levels of phosphorylated and insoluble tau, neuronal and synaptic loss, and cognitive deficits. Efficacy will be tested in three modes of treatment, beginning at early, intermediate, and late stages of progression of AD pathology and cognitive decline. Finally, in Aim 3, the efficacy of combinations of antibody shuttles targeting multiple neuroprotective pathways in PS19 mice will be evaluated. These studies are expected to provide new fundamental knowledge related to the CD98hc transport pathway and preclinical data for bispecific AD therapeutics, including combination therapies that target mechanistically unique neuroprotective pathways.

The goal of this research is to improve the delivery of neuroprotective antibodies to the brain by generating bifunctional antibodies that target both a protein at the blood brain barrier and a target within the brain. This research will involve optimizing bifunctional antibodies to achieve high and sustained antibody levels in the brain, and then using them to prevent Alzheimer's disease-like pathology in animal models. The bifunctional antibodies and insights gained in this work are expected to accelerate the development of therapeutics for treating Alzheimer's disease and other neurodegenerative disorders.