Efficient transduction of LEDGF/p75 mutant cells by complementary gain-of-function HIV-1 integrase mutant viruses

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 (IN)–binding domain from the lentiviral integration host cofactor lens epithelium–derived growth factor (LEDGF)/p75 are a promising retargeting strategy. We previously proposed the utility of IN mutant lentiviral vectors that are selectively activated by complementary LEDGF/p75 variants, and our initial modifications in human immunodeficiency virus type 1 IN 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 IN, which greatly improves the efficiency of this system. Both K42E and initial reverse-charge mutations in IN negatively affected 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 IN 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 IN mutant viral and LEDGF/p75 host proteins.