Positron emission tomography imaging of novel AAV capsids maps rapid brain accumulation

Jai Woong Seo,R Holland Cheng,Michelle L James,Michael Chavez,Eduardo A Silva,Ryan D Leib ,Spencer K Tumbale ,Lisa M Mahakian ,Tatyana Dobreva,Poorva Jain,Viviana Gradinaru,Xiaozhe Ding,Nick Goeden,Sadaf Aghevlian,Elizabeth S Ingham,David J Segal,Kratika Singhal,Shahin Shams,Nicholas C Flytzanis,Bo Wu,Mo Baikoghli,Katherine W Ferrara

doi:10.1038/s41467-020-15818-4

Abstract

Adeno-associated viruses (AAVs) are typically single-stranded deoxyribonucleic acid (ssDNA) encapsulated within 25-nm protein capsids. Recently, tissue-specific AAV capsids (e.g. PHP.eB) have been shown to enhance brain delivery in rodents via the LY6A receptor on brain endothelial cells. Here, we create a non-invasive positron emission tomography (PET) methodology to track viruses. To provide the sensitivity required to track AAVs injected at picomolar levels, a unique multichelator construct labeled with a positron emitter (Cu-64, t1/2 = 12.7 h) is coupled to the viral capsid. We find that brain accumulation of the PHP.eB capsid 1) exceeds that reported in any previous PET study of brain uptake of targeted therapies and 2) is correlated with optical reporter gene transduction of the brain. The PHP.eB capsid brain endothelial receptor affinity is nearly 20-fold greater than that of AAV9. The results suggest that novel PET imaging techniques can be applied to inform and optimize capsid design.

Highlights

Adeno-associated viruses (AAVs) are typically single-stranded deoxyribonucleic acid encapsulated within 25-nm protein capsids
We have developed a bioorthogonal approach for coupling a multichelator and AAV, based on conjugation to AAV surface lysines and cysteines and used this approach to compare the PK of AAV serotype 9 (AAV9)-PHP.eB (AAV9PHP.eB is denoted as PHP.eB hereafter) with AAV9 and AAV9tetracysteine (AAV9-TC)
Based on surface solvent accessibility in the X-ray structure of the AAV9 capsid[30], the estimated number of exposed lysines on AAV9 and PHP.eB ranges from 420 to 480 out of 1185 and 1245 total lysines, respectively (Fig. 1b). This includes 7–8 lysines per viral protein (VP), with one viral particle composed of 60 units of viral protein subunits (VPs)

Summary

Introduction

Adeno-associated viruses (AAVs) are typically single-stranded deoxyribonucleic acid (ssDNA) encapsulated within 25-nm protein capsids. Engineered adenoassociated viruses (AAVs), single-stranded deoxyribonucleic acid (ssDNA) encapsulated within 25-nm protein capsids, have recently shown potential to greatly increase transduction as compared with previous therapeutics[2,3,4]. AAVs can infect dividing and non-dividing cells and result in highly efficient long-term transduction in a broad range of tissues[5,6] This is significant as AAV gene therapy has a solid safety profile, was first approved by the FDA in December 20177 and more than 200 clinical trials have been conducted since 19898. Mer peptide insertion in a surface exposed loop of the capsid, enhanced neuronal transduction throughout the brain compared to the conventionally used AAV serotype 9 (AAV9)[3] This 40 to 90-fold increased efficiency is believed to result from a novel interaction between virus and the brain endothelial cell receptor LY6A10,11. The biodistribution of viral vectors has previously been evaluated by real-time PCR, Southern blotting of the transduced gene, western blotting, immunohistochemistry (IHC), and in vivo imaging of reporter proteins[12]

Methods

Results

Conclusion