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Design, intracellular expression, and activity of a human anti-human immunodeficiency virus type 1 gp120 single-chain
W A Marasco1, W A Haseltine, S Y Chen
1Department of Medicine, Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA 02115.
Insights
Researchers engineered a single-chain antibody for intracellular expression to combat HIV-1. This novel antibody targets the virus within cells, significantly reducing HIV-1 infectivity and offering a new therapeutic strategy.
Area of Science:
- Immunology
- Virology
- Molecular Biology
Background:
- Human immunodeficiency virus type 1 (HIV-1) infection remains a global health challenge.
- Intracellular targeting of viral proteins presents a promising therapeutic avenue.
- Previous strategies often face challenges with delivery and stability.
Purpose of the Study:
- To design and evaluate a single-chain antibody for intracellular expression against HIV-1.
- To assess the antibody's stability, localization, and inhibitory effects on HIV-1 replication.
- To demonstrate the feasibility of intracellular antibody-based antiviral strategies.
Main Methods:
- A single-chain antibody was constructed using variable regions from a human monoclonal antibody targeting the HIV-1 envelope protein's CD4 binding site.
- The antibody was engineered for intracellular expression in eukaryotic cells, including a signal sequence for endoplasmic reticulum retention.
- Functional assays were performed to assess antibody expression, binding to HIV-1 envelope protein, inhibition of viral processing, syncytia formation, and infectivity of produced viral particles.
Main Results:
- The engineered single-chain antibody was stably expressed and retained within the endoplasmic reticulum of host cells without causing toxicity.
- The intracellular antibody effectively bound to the HIV-1 envelope protein, inhibiting its precursor processing and preventing syncytia formation.
- Cells expressing the single-chain antibody produced HIV-1 particles with substantially reduced infectivity.
Conclusions:
- Intracellular expression of single-chain antibodies targeting viral proteins is a feasible strategy for controlling infectious diseases.
- This approach offers a novel mechanism for inhibiting viral replication at the intracellular level.
- Engineered intracellular antibodies hold potential as research tools and therapeutic agents for HIV-1 and other viral infections.
Abstract:
A single-chain antibody, derived from a human monoclonal antibody that recognizes the CD4 binding region of the human immunodeficiency virus type 1 (HIV-1) envelope protein, has been designed for intracellular expression in eukaryotic cells. The single-chain antibody is composed of an immunoglobulin heavy-chain leader sequence and heavy- and light-chain variable regions that are joined by an interchain linker. The antibody is stably expressed and retained in the endoplasmic reticulum and is not toxic to the cells. The antibody binds to the envelope protein within the cell and inhibits processing of the envelope precursor and syncytia formation. The infectivity of the HIV-1 particles produced by cells that express the single-chain antibody is substantially reduced. These studies illustrate the feasibility of designing antibodies that bind and inactivate molecules intracellularly. Antibodies that act on target molecules within cells should provide a useful tool for research as well as for control of infectious and other diseases.