Targeted Nucleases—Human Gene Therapy 2.0?

Targeted nucleases, which include zinc finger nucleases and CRIPSR/Cas9, are poised to revolutionize gene therapy by potentially offering safer and more precise methods to engineer human genes. Although several different nuclease platforms are available, they all act in the same way, by first introducing a DNA break at a specific targeted sequence. Subsequent repair of the break by host cell pathways can then be exploited to achieve three possible outcomes—gene disruption, gene editing (mutation correction), or the regulated addition of new genetic material at a defined genetic locus or “safe harbor.” The gene editing and addition outcomes also require the introduction into a cell of a homologous “donor sequence,” to serve as a repair template to direct such changes.

My group is using targeted nucleases in hematopoietic stem cells (HSC), with the specific goal of developing anti-HIV therapies. Our initial efforts focused on disrupting the CCR5 gene, which codes for an entry co-receptor molecule used by most strains of HIV. We demonstrated using humanized mice—immune-deficient mice transplanted with human HSC—that CCR5 disruption could suppress HIV replication without any adverse impact on human hematopoiesis. A clinical trial based on knocking-out this non-essential human gene is now enrolling HIV/AIDS patients.

More recently, we have also developed methods that allow us to edit genes in HSC at high frequency. At some loci, we can reach levels that are comparable to those achieved with more standard gene therapy tools such as lentiviral vectors, but without the concern that randomly integrating vectors could drive insertional mutagenesis events. This capability, combined with a growing clinical experience with zinc finger nucleases, means that targeted nucleases are poised to be applied to other diseases of the blood and immune system, and beyond.