Engineering Extracellular Vesicles Enriched with Palmitoylated ACE2 as COVID‐19 Therapy

Abstract

Angiotensin converting enzyme 2 (ACE2) is a key receptor present on cell surfaces that directly interacts with the viral spike (S) protein of the severe acute respiratory syndrome coronavirus‐2 (SARS‐CoV‐2). It is proposed that inhibiting this interaction can be promising in treating COVID‐19. Here, the presence of ACE2 in extracellular vesicles (EVs) is reported and the EV‐ACE2 levels are determined by protein palmitoylation. The Cys141 and Cys498 residues on ACE2 are S‐palmitoylated by zinc finger DHHC‐Type Palmitoyltransferase 3 (ZDHHC3) and de‐palmitoylated by acyl protein thioesterase 1 (LYPLA1), which is critical for the membrane‐targeting of ACE2 and their EV secretion. Importantly, by fusing the S‐palmitoylation‐dependent plasma membrane (PM) targeting sequence with ACE2, EVs enriched with ACE2 on their surface (referred to as PM‐ACE2‐EVs) are engineered. It is shown that PM‐ACE2‐EVs can bind to the SARS‐CoV‐2 S‐RBD with high affinity and block its interaction with cell surface ACE2 in vitro. PM‐ACE2‐EVs show neutralization potency against pseudotyped and authentic SARS‐CoV‐2 in human ACE2 (hACE2) transgenic mice, efficiently block viral load of authentic SARS‐CoV‐2, and thus protect host against SARS‐CoV‐2‐induced lung inflammation. The study provides an efficient engineering protocol for constructing a promising, novel biomaterial for application in prophylactic and therapeutic treatments against COVID‐19.

Elucidating design principles for engineering cell-derived vesicles to inhibit SARS-CoV-2 infection

“…17 Most recently, intravenously administered ACE2-EVs lowered the viral load of authentic SARS-CoV-2 in a mouse model, reduced the levels of pro-inflammatory cytokines in lung tissue, and mitigated lung tissue injury. 22 These studies validated the fundamental concept that decoy EVs could address this disease and raised a number of interesting questions. However, the diversity of experimental systems and designs employed across these studies makes it difficult to synthesize the results of these efforts to evaluate the relationship between specific EV design choices and efficacy.…”

“…This finding contradicts a report in which improving ACE2 loading conferred improvements in potency. 22 Surprisingly, WT-ACE2 and Mut-ACE2 EVs exhibited similar potency despite an estimated 5–10 fold difference in binding affinity of the individual ACE2 variants for Spike, 23 and this finding held across EV subtypes. One possible interpretation for the similar potency across all four EV-ACE2 combinations is that ACE2 loading for these EV samples is high enough that the effect of loading upon inhibitor potency has saturated (which might not have been the case in prior reports 22 ).…”

“… 22 Surprisingly, WT-ACE2 and Mut-ACE2 EVs exhibited similar potency despite an estimated 5–10 fold difference in binding affinity of the individual ACE2 variants for Spike, 23 and this finding held across EV subtypes. One possible interpretation for the similar potency across all four EV-ACE2 combinations is that ACE2 loading for these EV samples is high enough that the effect of loading upon inhibitor potency has saturated (which might not have been the case in prior reports 22 ). Control HS-EVs and UC-EVs, which lacked ACE2, had no effect on transduction efficiency, confirming that inhibition of Spike-lenti is ACE2-dependent at all doses evaluated.…”

“…Moreover, we lack understanding as to how decoy EV performance varies across various emerging viral strains, which is an open question of recognized importance. 22 Resolving these knowledge gaps could help optimize and deploy decoy EV treatments for SARS-CoV-2, novel variants thereof, and perhaps novel viral infections.…”

Recent Progress of Exosome Isolation and Peptide Recognition-Guided Strategies for Exosome Research

“…Nowadays, more and more studies have shown that exosomes play important roles in organisms, such as cell-to-cell communication ( Ruhland et al, 2020 ; Schneider et al, 2021 ), immune response ( Kalluri and LeBleu, 2020 ; Su et al, 2020 ), cell growth and differentiation ( Yin et al, 2021 ; Zarnowski et al, 2021 ), as well as molecular transport ( Zhu Q. et al, 2021 ). Crucially, the concentration and phenotype of exosomes have been proved to reflect the state of their parental cells and associate with the occurrence and development of various diseases, such as cancers ( Jiang et al, 2021 ), infectious diseases ( Ning et al, 2021 ; Xie et al, 2021 ), metabolic, and cardiovascular diseases ( Kalluri and LeBleu, 2020 ). Therefore, developing exosome isolation methods with high efficiency and selectivity will definitely promote the deep understanding of the functions of exosome, as well as the accurate diagnosis and treatment of diseases.…”