Gornalusse Germán G, Hirata Roli K, Funk Sarah E, Riolobos Laura, Lopes Vanda S, Manske Gabriel, Prunkard Donna, Colunga Aric G, Hanafi Laïla-Aïcha, Clegg Dennis O, Turtle Cameron, Russell David W
Department of Medicine, University of Washington, Seattle, Washington, USA.
Center for Stem Cell Biology and Engineering, Department of Molecular, Cellular, and Developmental Biology, University of California, Santa Barbara, USA.
Nat Biotechnol. 2017 Aug;35(8):765-772. doi: 10.1038/nbt.3860. Epub 2017 May 15.
Polymorphisms in the human leukocyte antigen (HLA) class I genes can cause the rejection of pluripotent stem cell (PSC)-derived products in allogeneic recipients. Disruption of the Beta-2 Microglobulin (B2M) gene eliminates surface expression of all class I molecules, but leaves the cells vulnerable to lysis by natural killer (NK) cells. Here we show that this 'missing-self' response can be prevented by forced expression of minimally polymorphic HLA-E molecules. We use adeno-associated virus (AAV)-mediated gene editing to knock in HLA-E genes at the B2M locus in human PSCs in a manner that confers inducible, regulated, surface expression of HLA-E single-chain dimers (fused to B2M) or trimers (fused to B2M and a peptide antigen), without surface expression of HLA-A, B or C. These HLA-engineered PSCs and their differentiated derivatives are not recognized as allogeneic by CD8 T cells, do not bind anti-HLA antibodies and are resistant to NK-mediated lysis. Our approach provides a potential source of universal donor cells for applications where the differentiated derivatives lack HLA class II expression.
人类白细胞抗原（HLA）I类基因中的多态性可导致异基因受体对多能干细胞（PSC）衍生产品产生排斥反应。β2微球蛋白（B2M）基因的破坏会消除所有I类分子的表面表达，但会使细胞易被自然杀伤（NK）细胞裂解。在此，我们表明，通过强制表达最低限度多态性的HLA-E分子，可以防止这种“缺失自我”反应。我们使用腺相关病毒（AAV）介导的基因编辑，以赋予HLA-E单链二聚体（与B2M融合）或三聚体（与B2M和肽抗原融合）可诱导、受调控的表面表达的方式，在人类PSC的B2M位点敲入HLA-E基因，而不产生HLA-A、B或C的表面表达。这些经过HLA工程改造的PSC及其分化衍生物不会被CD8 T细胞识别为异基因，不结合抗HLA抗体，并且对NK介导的裂解具有抗性。我们的方法为分化衍生物缺乏HLA II类表达的应用提供了通用供体细胞的潜在来源。
