Cellular delivery of antibodies: effective targeted subcellular imaging and new therapeutic tool

Marzia Massignani,Nicholas Warren,Steven Armes,Andrew Lewis,Nisa Patikarnmonthon,Irene Canton,Giuseppe Battaglia

doi:10.1038/npre.2010.4427.1

Marzia Massignani, Nicholas Warren + Show 5 more

Open Access

https://doi.org/10.1038/npre.2010.4427.1

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Abstract

AbstractIt is already more than a century since the pioneering work of the Nobel Laureate Ehrlich gave birth to the side chain theory1, which helped to define antibodies and their ability to target specific biological sites. However, the use of antibodies is still restricted to the extracellular space due to the lack of a suitable delivery vehicle for the efficient transport of antibodies into live cells without inducing toxicity. In this work, we report the efficient encapsulation and delivery of antibodies into live cells with no significant loss of cell viability or any deleterious affect on the cell metabolic activity. This delivery system is based on poly(2-(methacryloyloxy)ethyl phosphorylcholine)-block-(2-(diisopropylamino)ethyl methacrylate), (PMPC-PDPA), a pH sensitive diblock copolymer that self-assembles to form nanometer-sized vesicles, also known as polymersomes, at physiological pH. These polymersomes can successfully deliver relatively high antibody payloads within live cells. Once inside the cells, we demonstrate that these antibodies can target their epitope by immune-labelling of cytoskeleton, Golgi, and transcription factor proteins in live cells. We also demonstrate that this effective antibody delivery mechanism can be used to control specific subcellular events, as well as modulate cell activity and pro-inflammatory process.

Highlights

E used to control specific subcellular events, as well as modulate cell activity and pro-inflammatory process
Cytosolic antibody delivery In order to assess effective cytosolic delivery of antibody within live cells, we encapsulated AlexaFluor546-labelled IgG antibodies within poly(2(methacryloyloxy)ethyl phosphorylcholine) (PMPC)-poly(2(diisopropylamino)ethyl methacrylate) (PDPA) polymersomes, followed by incubation with primary live human dermal fibroblasts (HDF) for various time periods
Live cells exhibit strong fluorescent signals from the cytosol (Figure S1a-b). These results were compared with a traditional immunolabelling method (Figure S1cd) where permeabilised fixed cells were treated with the same concentration of antibody (AlexaFluor 546 goat Anti-mouse IgG)

Summary

Introduction

E used to control specific subcellular events, as well as modulate cell activity and pro-inflammatory process. Recent strategies for the intracellular delivery of functional proteins include the use of liposomes to encapsulate both proteins and genes for intracellular delivery[7], antibody delivery via protein transduction domains (PTD)[8], membrane receptors and transport signals[911], viral vectors[12], microinjection[13] or electroporation[14] Each of these methods lack convenience and effectiveness and are often compromised by cellular toxicity[15]. We present a non-cytotoxic delivery system based on a pH-sensitive diblock copolymer that self-assembles at physiological pH to form nanometer-sized polymersomes These polymersomes successfully deliver functional antibodies into the cytoplasm. The presence of PMPC-PDPA polymersomes within cells consistently has a negligible effect on cell viability and stress levels over a broad range of cell types, including both primary cells and cell lines.[18, 21, 22]

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