Receptor clustering and activation by multivalent interaction through recognition peptides presented on exosomes

Jayasinghe

J of Extracellular Vesicle

et al. 2021

126

119

Natural extracellular vesicles (EVs) are ideal drug carriers due to their remarkable biocompatibility. Their delivery specificity can be achieved by the conjugation of targeting ligands. However, existing methods to engineer target‐specific EVs are tedious or inefficient, having to compromise between harsh chemical treatments and transient interactions. Here, we describe a novel method for the covalent conjugation of EVs with high copy numbers of targeting moieties using protein ligases. Conjugation of EVs with either an epidermal growth factor receptor (EGFR)‐targeting peptide or anti‐EGFR nanobody facilitates their accumulation in EGFR‐positive cancer cells, both in vitro and in vivo. Systemic delivery of paclitaxel by EGFR‐targeting EVs at a low dose significantly increases drug efficacy in a xenografted mouse model of EGFR‐positive lung cancer. The method is also applicable to the conjugation of EVs with peptides and nanobodies targeting other receptors, such as HER2 and SIRP alpha, and the conjugated EVs can deliver RNA in addition to small molecules, supporting the versatile application of EVs in cancer therapies. This simple, yet efficient and versatile method for the stable surface modification of EVs bypasses the need for genetic and chemical modifications, thus facilitating safe and specific delivery of therapeutic payloads to target cells.

Section: Introductionmentioning

confidence: 99%

Covalent conjugation of extracellular vesicles with peptides and nanobodies for targeted therapeutic delivery

Jayasinghe

J of Extracellular Vesicle

et al. 2021

126

119

“…6–9 Besides, as a type of naturally occurred nanocarrier, exosomes exhibit advantages in size, structure, stability and biocompatibility. 10,11 For instance, nanosized exosomes can readily penetrate biological barriers, escape from phagocytosis, and accumulate in tissue microenvironments, such as tumors, via the enhanced permeability and retention effect. In addition, the bilayer membrane structure of exosomes is enhanced by deformable cytoskeleton and “gel-like” cytoplasm-derived content, making exosomes integral during trafficking and compatible for large quantities of soluble cargos.…”

mentioning

confidence: 99%

Molecular Recognition-Based DNA Nanoassemblies on the Surfaces of Nanosized Exosomes

et al. 2017

Exosomes are membrane-enclosed extracellular vesicles derived from cells, carrying biomolecules that include proteins and nucleic acids for intercellular communication. Owning to their advantages of size, structure, stability, and biocompatibility, exosomes have been used widely as natural nanocarriers for intracellular delivery of theranostic agents. Meanwhile, surface modifications needed to endow exosomes with additional functionalities remain challenging by their small size and the complexity of their membrane surfaces. Current methods have used genetic engineering and chemical conjugation, but these strategies require complex manipulations and have only limited applications. Herein, we present an aptamer-based DNA nanoassemblies on exosome surfaces. This in situ assembly method is based on molecular recognition between DNA aptamers and their exosome surface markers, as well as DNA hybridization chain reaction initiated by an aptamer-chimeric trigger. It further demonstrated selective assembly on target cell-derived exosomes, but not exosomes derived from nontarget cells. The present work shows that DNA nanostructures can successfully be assembled on a nanosized organelle. This approach is useful for exosome modification and functionalization, which is expected to have broad biomedical and bioanalytical applications.

“…7–11 Of particular interest from the perspective of nanomedicine is that the multivalent presentation of EGF ligands on NP (NP-EGF) provides an alternative strategy to amplify signal activation and transduction. 12 Multivalency 13 increases the binding affinity of NP and, thus, ligand receptor-binding, 14–18 which can impact uptake rate 19 and mechanism. 20 Importantly, multiple ligand-receptor contacts within the footprint of a particle can generate a strong local receptor activation.…”

mentioning

confidence: 99%

Ligand Density and Nanoparticle Clustering Cooperate in the Multivalent Amplification of Epidermal Growth Factor Receptor Activation

Zhang

Reinhard

2018

ACS Nano

Multivalent presentation of ligands on nanoparticles (NP) is considered a general strategy for enhancing receptor binding and activation through amplification of ligand-receptor interactions within the footprint of the NP. The spatial clustering of ligand-functionalized NP represents an additional, less well understood mechanism for increasing local ligand-receptor interactions, especially for receptors that form higher order assemblies, such as the epidermal growth factor (EGF) receptor (EGFR). To shed light on the interplay between ligand density (i.e. multivalency) and NP clustering in signal amplification, we apply EGF-functionalized 72±1 nm gold nanoparticles (NP-EGF) with known ligand loading (10 – 200 EGF/NP) as quantifiable and experimentally tractable units of EGFR activation and characterize the NP-mediated amplification of EGFR phosphorylation as function of both EGF surface density and NP-EGF clustering for two cancer cell lines (HeLa and MDA-MB-468). The measurements confirm a strong multivalent amplification of EGFR phosphorylation through NP-EGF on the cellular level that results in EGF-loading-dependent maximum EGFR phosphorylation levels. A microscopic analysis of NP-EGF-induced EGFR phosphorylation reveals a heterogeneous spatial distribution of EGFR activation across the cell surface. Clustering of multivalent NP-EGF on sub-diffraction limit length scales is found to result in a local enhancement of EGFR phosphorylation in signaling “hot-spots” from where the signal can spread laterally in an EGF-independent fashion. Increasing EGF loadings of the NP enhance NP-EGF clustering and intensify EGFR phosphorylation. These observations suggest that NP-EGF-clustering and the associated local enhancement of ligand-receptor interactions are intrinsic components of the multivalent amplification of phosphorylation for the heterogeneously distributed EGFR through NP-EGF.