Arredondo-Vega Francisco X, Santisteban Ines, Richard Eva, Bali Pawan, Koleilat Majed, Loubser Michael, Al-Ghonaium Abdulaziz, Al-Helali Mariam, Hershfield Michael S
Department of Medicine, Duke University Medical Center, Durham, NC 27710, USA.
Blood. 2002 Feb 1;99(3):1005-13. doi: 10.1182/blood.v99.3.1005.
Four patients from 3 Saudi Arabian families had delayed onset of immune deficiency due to homozygosity for a novel intronic mutation, g.31701T>A, in the last splice acceptor site of the adenosine deaminase (ADA) gene. Aberrant splicing mutated the last 4 ADA amino acids and added a 43-residue "tail" that rendered the protein unstable. Mutant complementary DNA (cDNA) expressed in Escherichia coli yielded 1% of the ADA activity obtained with wild-type cDNA. The oldest patient, 16 years old at diagnosis, had greater residual immune function and less elevated erythrocyte deoxyadenosine nucleotides than his 4-year-old affected sister. His T cells and Epstein-Barr virus (EBV) B cell line had 75% of normal ADA activity and ADA protein of normal size. DNA from these cells and his whole blood possessed 2 mutant ADA alleles. Both carried g.31701T>A, but one had acquired a deletion of the 11 adjacent base pair, g.31702-12, which suppressed aberrant splicing and excised an unusual purine-rich tract from the wild-type intron 11/exon 12 junction. During ADA replacement therapy, ADA activity in T cells and abundance of the "second-site" revertant allele decreased markedly. This finding raises an important issue relevant to stem cell gene therapy.
来自3个沙特阿拉伯家庭的4名患者因腺苷脱氨酶(ADA)基因最后一个剪接受体位点的一个新的内含子突变g.31701T>A纯合而出现免疫缺陷发病延迟。异常剪接使ADA的最后4个氨基酸发生突变,并添加了一个43个残基的“尾巴”,导致蛋白质不稳定。在大肠杆菌中表达的突变互补DNA(cDNA)产生的ADA活性仅为野生型cDNA的1%。最年长的患者在诊断时为16岁,其残余免疫功能较强,红细胞脱氧腺苷核苷酸升高程度低于他4岁的患病妹妹。他的T细胞和爱泼斯坦-巴尔病毒(EBV)B细胞系具有正常ADA活性的75%,且ADA蛋白大小正常。来自这些细胞和他全血的DNA携带2个突变的ADA等位基因。两者都携带g.31701T>A,但其中一个发生了11个相邻碱基对g.31702-12的缺失,这抑制了异常剪接,并从野生型内含子11/外显子12连接处切除了一段不寻常的富含嘌呤的片段。在ADA替代治疗期间,T细胞中的ADA活性和“第二位点”回复等位基因的丰度显著下降。这一发现提出了一个与干细胞基因治疗相关的重要问题。