UCLA researchers in the Department of Pediatrics Allergy and Immunology have developed a gene therapy method to express the full-length DOCK8 protein in cells, aimed at treating DOCK8 deficiency. These method allows for safe and effective DOCK8 expression in hematopoietic stem cells without the need for an HLA-matched donor, enabling patients to undergo transplantation at a younger age and improving overall survival rates.
BACKGROUND: The DOCK8 protein plays an essential role in regulating immune cell functions and maintaining a healthy immune system. Mutations in the DOCK8 gene can result in DOCK8 deficiency, leading to severe infections, autoimmune complications, and an increased risk of cancer. Patients begin to exhibit symptoms within the first few months to early years of life and are severely impacted by the disease. Therefore, early diagnosis and prompt treatment are critical for managing DOCK8 deficiency to improve patient outcomes. While allogeneic hematopoietic stem cell transplantation (HSCT) is a curative option for DOCK8 deficiency, it often leads to various serious complications including chronic graft versus host disease. Recently, ex vivo gene modification of autologous hematopoietic stem and progenitor cells (HSPCs) has emerged as a promising treatment with fewer side effects, eliminating the need for a human leukocyte antigen (HLA)-matched donor.
INNOVATION: UCLA researchers led by Dr. Caroline Kuo at the Medical Center have developed a method to express the full-length DOCK8 protein in cells as a treatment for DOCK8 deficiency using lentiviral vector transduction of human hematopoietic stem cells (HSC). To reduce the risks of insertional mutagenesis from gamma-retroviral vectors, they used modified HIV-1 based lentiviral vectors, offering a safer alternative for modifying HSCs. Testing the lentiviral vectors combined with specific cell treatments showed DOCK8 expression in both Jurkat cell lines and peripheral blood stem cells (PBSCs) from healthy donors. However, DOCK8 expression levels were low in due to the large size of the DOCK8 gene. It is known that lentiviral vectors carrying a large gene cassette are often hindered by low titers and poor transduction rates. To solve this issue, researchers created two lentiviral vectors, each of them expressing one half of the DOCK8 protein with adjacent intein sequences. The intein sequences allowed for ligation of the two DOCK8 protein fragments into a fully functional DOCK8 protein as demonstrated by restoration of DOCK8 expression in primary human T cells and K562 cells.
POTENTIAL APPLICATIONS:
- Expressing a full-length DOCK8 protein in a cell
- Treating DOCK8 deficiency
ADVANTAGES:
- Do not require an HLA-matched donor and allow patients to undergo transplantation at a younger age, thus improving their overall survival
- Allow for safe and effective DOCK8 expression
DEVELOPMENT-TO-DATE:
DOCK8 lentiviral vectors were tested in Jurkat cell lines and PBSCs from healthy donors. The optimized inteins were initially evaluated for producing green fluorescent protein in human K562 cells, and were subsequently tested for restoring DOCK8 protein expression in Jurkat T cell lines and K562 cells.
Keywords: DOCK8 deficiency, immunodeficiency, gene therapy, gene editing, autologous, hematopoietic stem cell, immune cell, bone marrow transplantation, lentiviral vector, intein splicing, protein splicing, HSCT, HSC, HSPC