A project of clinical application in a gene therapy for chronic granulomatous disease with gp91-phox deficiency

由于A. Fischer对X-SCID患者的基因治疗方案中已经报道了3-淋巴细胞增生性疾病，因此临床基因治疗的安全性需要更多的基础和临床证据。本文首次分析了200多例慢性肉芽肿病（CGD）患者，并分析了gp91-phox缺乏症剪合突变患者ifn - γ的遗传机制（Blood 98: 436-441, 2001）。在我们与Sugimoto博士合作的为期3年的项目中，我们构建了一个双电子逆转录病毒载体Ha-MDR-IRES-gp91，用于MDR1和gp91的共表达，gp91是x -联性慢性肉芽肿病（X-CGD）的基因。Epstein-Barr病毒转化X-CGD患者的B细胞和X-CGD患者的人cd34阳性细胞，通过Ha-MDR-IRES-gp91转导共同表达p糖蛋白和gp91，并获得超氧化物生成活性。用ha - mdr - ires -gp91转导的骨髓细胞移植小鼠，外周血单核细胞中P-gl . More蛋白和gp91共表达。经紫杉醇多次给药后，p -糖蛋白阳性细胞与gp91阳性细胞的比例维持1年以上。这些结果表明，人类多药耐药基因（MDR1）与治疗基因的共表达为转基因细胞提供了可选择的生长优势（J gene Med vol. 4, 2003）。在与Sumimoto博士的合作中，我们阐明了第四种细胞质因子p40 （phox）通过PB1-PC与p67 （phox）的相互作用，通过调节p67 （phox）和p47 （phox）的膜募集来参与吞噬细胞氧化酶的激活。[J] .生物医学工程学报；2002;21(23):6312-20。此外，我们克隆了与p47phox和p67phox同源的新蛋白cdna，分别命名为p41nox和p51nox。基于组合研究，我们发现新的同源物p41nox和p51nox可能一起或与经典同源物结合起作用，从而激活两种Nox家族氧化酶（J Biol Chem 2003 in press）。少

Clinical gene therapy was required more fundamental and clinical evidences for safety since 3-lymopoproliferative disease had been reported in A. Fischer's gene therapy protocol for X-SCID patients. More than 200 patients with chronic granulomatous disease (CGD) were first analyzed and a genetic mechanisms of IFN-gamma responsibility was analyzed in the patients with the splice mutation of gp91-phox deficiency (Blood 98 : 436-441, 2001).In our 3-year's project, collaborating with Dr. Sugimoto, we constructed a bicistronic retrovirus vector, Ha-MDR-IRES-gp91, for the co-expression of MDR1 and gp91, a gene responsible for X-linked chronic granulomatous disease (X-CGD). Epstein-Barr virus-transformed B cells from X-CGD patients and human CD34-positive cells from an X-CGD patient transduced with Ha-MDR-IRES-gp91 co-expressed P-glycoprotein and gp91, and acquired superoxide-generating activity. Mice transplanted with Ha-MDR-IRES-gp91-transduced bone marrow cells showed co-expression of P-gl … More ycoprotein and gp91 in peripheral blood mononuclear cells. After repeatedly administration of paclitaxel, the ratio of P-glycoprotein- and gp91-positive cells were maintained for over 1 year. These results suggest that co-expression of a human multidrug resistance gene (MDR1) with a therapeutic gene affords selectable growth advantage to genetically modified cells (J Gene Med vol. 4, 2003).In a collaboration with Dr. Sumimoto, we clarified the fourth cytosolic factor p40 (phox) participates in activation of the phagocyte oxidase by regulating membrane recruitment of p67 (phox) and p47 (phox) via the PB1-PC interaction with p67 (phox). (EMBO J 2002 Dec 2 ; 21(23) : 6312-20). In addition, we cloned human cDNAs for novel proteins homologous to p47phox and p67phox, designated p41nox and p51nox, respectively. Based on the combination studies, we found that the novel homologues p41nox and p51nox likely function together or in combination with a classical one, thereby activating the two Nox family oxidases (J Biol Chem 2003 in press). Less