Efficient Transduction of LEDGF/p75 Mutant Cells by Gain-of-Function HIV-1 Integrase Mutant Viruses.

Hao Wang (1, 2) , Ming-Chieh Shun (1, 2) , Xiang Li (1, 2) , Francesca Di Nunzio (1, 2) , Stephen Hare (3) , Peter Cherepanov (4, 3) , Alan Engelman (1, 2)

1 Dana-Farber Cancer Institute [Boston]
2 HMS - Harvard Medical School [Boston]
3 Imperial College London
4 Cancer Research UK

Controlling the specificity of retroviral DNA integration could improve the safety of gene therapy vectors, and fusions of heterologous chromatin binding modules to the integrase-binding domain from the lentiviral integration host cofactor LEDGF/p75 are a promising retargeting strategy. We previously proposed the utility of integrase mutant lentiviral vectors that are selectively activated by complementary LEDGF/p75 variants, and our initial modifications in HIV-1 integrase and LEDGF/p75 supported about 13% of wild-type vector transduction activity. Here we describe the selection and characterization of the K42E gain-of-function mutation in HIV-1 integrase, which greatly improves the efficiency of this system. Both K42E and initial reverse-charge mutations in integrase negatively impacted reverse transcription and integration, yet when combined together boosted viral transduction efficiency to ~75% of the wild-type vector in a manner dependent on a complementary LEDGF/p75 variant. Although the K42E mutation conferred functional gains to integrase mutant viral reverse transcription and integration, only the integration boost depended on the engineered LEDGF/p75 mutant. We conclude that the specificity of lentiviral retargeting strategies based on heterologous LEDGF/p75 fusion proteins will benefit from our optimized system that utilizes the unique complementation properties of reverse-charge integrase mutant viral and LEDGF/p75 host proteins.

Domaines: Sciences du Vivant [q-bio]

Hao Wang, Ming-Chieh Shun, Xiang Li, Francesca Di Nunzio, Stephen Hare, et al.. Efficient Transduction of LEDGF/p75 Mutant Cells by Gain-of-Function HIV-1 Integrase Mutant Viruses.. Molecular Therapy - Methods and Clinical Development, 2014, 1 (2), ⟨10.1038/mtm.2013.2⟩. ⟨pasteur-01536152⟩