毛细血管扩张共济失调是一种由 ATM 基因突变引起的单基因疾病。其编码的蛋白激酶 ATM 在双链断裂 (DSB) 的 DNA 修复中发挥着重要作用。该激酶的功能受损会导致多系统疾病，包括免疫缺陷、进行性小脑变性、辐射敏感性、血管扩张、过早衰老和癌症易感性。由于同种异体造血干细胞 (HSC) 移植改善了疾病结果，因此基于自体造血干细胞的基因治疗是另一种有前景的概念。然而，由于 ATM 的 cDNA (9.2 kb) 较大，逆转录病毒颗粒的有效包装和 HSC 的充分转导仍然具有挑战性。我们生成了带有 GFP.F2A.Atm 融合或 GFP 转基因的慢病毒、γ逆转录病毒和泡沫病毒载体，并系统地比较转导效率。载体滴度随着转基因大小的增加而下降，但尽管它们的包装能力有限，我们仍然能够生产慢病毒和γ逆转录病毒颗粒。滴度的降低不能用病毒基因组包装受损来解释，但主要差异发生在转导后。最后，用表达 Atm 的慢病毒载体转导 Atm 缺陷 (ATM-KO) 小鼠成纤维细胞后，我们可以显示磷酸化其下游底物 (pKap1 和 p-p53) 的 ATM 蛋白的表达。© 2023。 ）。

Ataxia telangiectasia is a monogenetic disorder caused by mutations in the ATM gene. Its encoded protein kinase ATM plays a fundamental role in DNA repair of double strand breaks (DSBs). Impaired function of this kinase leads to a multisystemic disorder including immunodeficiency, progressive cerebellar degeneration, radiation sensitivity, dilated blood vessels, premature aging and a predisposition to cancer. Since allogenic hematopoietic stem cell (HSC) transplantation improved disease outcome, gene therapy based on autologous HSCs is an alternative promising concept. However, due to the large cDNA of ATM (9.2 kb), efficient packaging of retroviral particles and sufficient transduction of HSCs remains challenging.We generated lentiviral, gammaretroviral and foamy viral vectors with a GFP.F2A.Atm fusion or a GFP transgene and systematically compared transduction efficiencies. Vector titers dropped with increasing transgene size, but despite their described limited packaging capacity, we were able to produce lentiviral and gammaretroviral particles. The reduction in titers could not be explained by impaired packaging of the viral genomes, but the main differences occurred after transduction. Finally, after transduction of Atm-deficient (ATM-KO) murine fibroblasts with the lentiviral vector expressing Atm, we could show the expression of ATM protein which phosphorylated its downstream substrates (pKap1 and p-p53).© 2023. The Author(s).