SCID-X1 gene therapy successful with 3D genome architecture

Patients with X-linked severe combined immunodeficiency disorder (SCID-X1), termed “bubble boy disease,” are born with a defective gene that stops the production of immune cells. Gene therapy from St. Jude Children’s Research Hospital restored the immune system in multiple infants with SCID-X1 in 2019 by supplying copies of the corrected gene. Published in Science Advances, St. Jude scientists documented where the gene copies integrate into patient DNA in an effort to monitor patient safety.  This documentation has provided a foundation to understand the biology and safety of lentiviral vectors.

Senior co-corresponding author and St. Jude Department of Computational Biology interim chair Dr Jiyang Yu said: “We now have a robust pipeline to monitor the safety of lentiviral gene therapies.”  He continued: “This really gives hope to patients with genetic diseases that can be cured by lentiviral gene therapy. It looks like we cured bubble boy disease safely in these patients.” The St. Jude team published the entire computational pipeline together with the manuscript.

Unlike earlier retroviral approaches, the St. Jude lentiviral gene therapy for SCID-X1 appears both effective and safe several years after treatment. The researchers detected where the gene was added to patients’ DNA and why the vector integrated in that  place. Scientists had prior knowledge that lentiviral gene therapies integrated into different areas of DNA appeared safer than earlier technologies, but they could not address why.

 3D genome structure

The researchers discovered that copies of the corrected gene were inserted into certain genomic hotspots for the patients in the study and the reason was simple. From a 3D structural perspective, the lentiviral vector first encounters hotspots after entering the cell’s nucleus through the nuclear pore.

“It’s like someone coming into a room and taking the first available seat near the door,” said co-corresponding author Dr Stephen Gottschalk, Chair of St. Jude Department of Bone Marrow Transplantation and Cellular Therapy. “The room is the nucleus. The seats are these DNA elements right near the door of the nuclear pore. That never occurred to me before this study, but it’s a very simple principle in the end.”

It was discovered that the integration site pattern of the gene therapy into patient cells revealed the safety and efficacy of the approach.

First author Dr Koon-Kiu Yan, St. Jude Department of Computational Biology, explained: “We performed a comprehensive single-cell multi-omic analysis of this gene therapy to understand whether a functional copy of the corrected gene was in patient cells, to what extent the gene was expressed, and the chromatin organization at a single-cell level.”  Yan continued: “Before this study, we could measure the general gene expression of a bulk group of cells. But with bone marrow samples from two patients, we saw at a single-cell level which genes were expressed in which cell types.”

Along with their other work, Yan used single-cell analysis to show that the safety and efficacy of treatment are also related to integration into compartments near the nuclear pore. Previous gene therapy efforts integrated into promoters near to or directly into oncogenes, causing cancer. To improve safety, the lentiviral vector used by St. Jude selectively avoids promoters without disrupting oncogenes. The same pattern was observed with a lentiviral gene therapy used to create chimeric antigen receptor (CAR) T cells, suggesting that the phenomenon may be a general mechanism not limited to the SCID-X1 vector.

“The integration pattern data could serve as a map of potentially safe integration sites,” Yu and Gottschalk said. “The single-cell analysis is like deep cartography, a map with a near pixel-perfect resolution. The large number of integration sites could be used as a safety reference for future lentiviral gene therapies.”

One notable case was a patient who needed a second dose of the gene therapy to achieve a cure. They were found to have a different integration pattern than the patients who had responded to the first dose. After receiving the second dose, the patient’s integration pattern resembled the others’ and the therapy was effective.