Molecular Analysis and Gene Therapy in Inherited dysmyelinating disorder

我们研究了酶替代疗法（ERT）和骨髓移植（BMT）对15例日本戈谢病患者（ERT：12例，BMT：3例）的疗效。其表型分为可能的1型（12例）和3b型（3例）。实验室检查结果（血红蛋白、血小板、血管紧张素转换酶和酸性磷酸酶）和严重程度评分指数（SSI）在大多数情况下通过治疗得到改善。然而，治疗后，体格生长，特别是身高仍然严重迟缓（前-2.7SD，后-2.2SD）。骨髓移植对生长发育迟缓的改善优于激素替代疗法，基因型和脾切除对治疗效果无影响。低剂量方案（60 U/kg/剂量< 6个月）导致3例患者在治疗期间发生骨受累。这些数据表明，在治疗儿童戈谢病1型患者时应注意体格生长，而酶的初始剂量是获得生长的最重要因素。 关于我们 我们分析了3例日本患者阿萨基因的核苷酸序列，（病例1、12和13）。在晚期婴儿型病例1中，发现了两个新的突变：366 a *g跃迁（指定为366 g）在第一内含子的3'剪接位点的-2位，和外显子5中的1542 T *C（指定为1542 C），导致亮氨酸298被丝氨酸取代。通过RT-PCR扩增的患者cDNA片段的分析显示，366 g等位基因的转录本被异常剪接。在瞬时表达研究中，携带298 Ser突变cDNA的转染子没有显示阿萨活性的增加，这证实了该突变是晚期婴儿MLD的一个原因。在幼年型病例12中，发现了一个假缺陷（PD）等位基因，该等位基因消除了一个N-糖基化位点（350 Asn *Ser）。这是日本人中首例阿萨基因PD等位基因。然而，在病例12中未检测到致病突变。成人型MLD患者13例为父系445 A和母系2330 T突变等位基因复合杂合子。2330 T，影响剪接受体位点的选择，被认为是负责在病人的轻度表型。对阿萨基因在MLD患者中的进一步分析，将有助于对MLD的表型-基因型相关性的思考。少

We investigated the effects of enzyme replacement therapy (ERT) and bone marrow transplantation (BMT) for 15 Japanese patients with Gaucher disease (ERT : 12 cases, BMT : 3 cases). Their phenotypes were classified into possible type 1 (12 cases) and type 3b (3 cases). Laboratory findings (values of hemoglobin, platelet, angiotensin converting enzyme and acid phosphatase) and severity score index (SSI) were improved by treatment in most of cases. However, physical growth, particulary height, was still severely retarded after treatment (pre--2.7SD,post--2.2SD). BMT made physical growth retardation more improved than ERT.Genotype and splenectomy did not influence the responce of treatment. Low dose protocol (60U/kg/dose < 6 months) resulted in bone involvement during treatment in three patients. These data suggest that one should pay attention to physical growth in treatment for pediatric Gaucher disease type 1 patients and the initial dosage of enzyme is the most important factor to obta … More in sufficient clinical effects in ERT.We have analyzed on the nucleotide sequence of ASA genes in three Japanese patient (case 1,12, and 13) with MLD.In case 1 with late infantile form, two novel mutations were found : a 366a*g transition (designated 366g) in the position -2 of 3' splice site of first intron, and 1542T*C in exon 5 (designated 1542C) causing leucine 298 to be substituted by serine. The analysis of the patient's cDNA fragments amplified by RT-PCR revealed that transcripts of the 366g allele were spliced aberrantly. In a transient expression study, transfectant with the mutant cDNA carrying 298Ser did not show an increase of ASA activity, which confirms the mutation is a cause of late infantile MLD.In case 12 with juvenile form, a pseudodeficiency (PD) allele, which abolishes an N-glycosylation site (350Asn*Ser), was found. This is the first case with PD allele of ASA gene in Japanese origin. However, no disease causing mutation was detected in the case 12. Case 13 with adult form MLD was a compound heterozygote of mutant alleles : 445A of paternal origin and 2330T of maternal origin. The 2330T,affecting splice acceptor site selection, was suggested to be responsible for the mild phenotype in the patient. The further analysis of ASA gene in MLD patients should provide insight into the consideration on the phenotype-genotype correlation in the disorder. Less