The Use of Intransally‐delivered Elastin‐like Polypeptide for Drug Delivery to the Central Nervous System
Authors: McGowan Jeremy W.D. , Vig Parminder J.S. , Bidwell Gene L. 
Published: November 2023

Drug delivery to the central nervous system (CNS) ishindered by the blood brain barrier (BBB). Intranasal administration (IN) hasbeen demonstrated to bypass the BBB, leading to increased drug concentrations in the CNS. Cell penetrating peptides (CPP) have also been used to increase uptake in different tissues including the CNS. Determining the effectiveness of IN with or without the addition of CPPs will inform future delivery strategies. Elastin like polypeptide (ELP) is a thermally responsive polypeptide used as a carrier and stabilizer, making conjugated biomolecules more resistant to proteolysis and extending their half‐lives in vivo. In our lab, we have used CPP‐fused ELPs conjugated to the therapeutic peptides with promising results in disease models. Currently, we are assessing the use of IN to bypass the BBB and facilitate delivery of ELP‐fused therapeutics to the CNS. We chose two well‐studied CPPs to conjugate to the ELP carrier: TAT and SynB1. Fluorescently‐labeled CPP‐ELP conjugates were delivered via IN and intravenous (IV) routes. Polypeptide levels were determined using direct fluorescence measurements in fluid samples and quantitative tissue imaging. Total brain ELP levels were 12.1 ± 0.8 μg/g of tissue after IN delivery and 17.9 ± 1.0 μg/g of tissue after IV delivery. SynB1‐ELP levels were lower than ELP levels after both IN and IV delivery (2.2 ± 0.7 μg/g and 7.1 ± 0.4 μg/g, respectively). Both the higher brain levels achieved by the IV route and the decrease in brain levels seen after SynB1 conjugation were statistically significant (two‐way ANOVA, p < 0.05, post‐hoc Tukey multiple comparison). TAT‐ELP was not detectable in the brain via either route. Though IV administration resulted in higher overall brain ELP levels, the whole‐organ fluorescence assay did not account for intravascular versus extravascular protein levels. To determine if the administered protein was sequestered on the vascular side of the BBB, the experiment was repeated, but mice were perfused with heparinized saline to remove all blood from the CNS. For IN administered ELP, 79.3% of the fluorescence signal remained in the brain after perfusion, contrasted with only2.9% remaining fluorescence after IV administration. These data indicate that the vast majority of the protein is extravascular after IN delivery versus sequestered in the vasculature after IV delivery. Polypeptide distribution in the brain was determined by scanning thin sections of tissue and fluorescence microscopy. These data confirm the data from the whole‐organ fluorescence demonstrating primarily extravascular ELP following IN administration and intravascular ELP following IV administration. In summary, these results indicate that both SynB1 and TAT, when conjugated directly to the ELP carrier, significantly decrease CNS levels when compared to ELP alone following either IV or IN. Additionally, IN administration of ELP resulted inmost protein localized across the BBB, while IV administered protein was localized in the vascular compartment. We conclude from these data that the IN route is more effective for CNS delivery of ELP and may therefore hold promise for translational applications of ELP‐based drug delivery for CNS disorders.Support or Funding InformationJWDM is supported by the Robert Currier Pre‐doctoral Fellowship in Translational Neuroscience.