Triplex-mediated DNA repair as a novel therapy for Hurler Syndrome patients

描述（由申请人提供）：这是一项测试使用三链体分子用作治疗剂以纠正Hurler综合征（HS）的可行性的建议。溶酶体贮积病（LSD），如HS，代表了具有破坏性临床特征的单基因疾病的多样化集合。LSD由编码溶酶体酶和其他对溶酶体功能至关重要的蛋白质的基因突变引起。HS是由戊-L-艾杜糖醛酸酶基因（IDUA）突变引起的。对于许多LSD，治疗包括酶替代疗法和支持性护理。然而，酶替代疗法的费用从90，000美元到565，000美元不等。或者，异基因造血干细胞移植（HSCT）代表了一种非常有效的治疗方法。然而，同种异体HSCT需要HLA匹配的供体，与显著的发病率和死亡率相关，并且昂贵，每个患者约20万美元。 使用CD 34+细胞的自体HSCT的基因修饰正在成为同种异体HSCT的可行治疗替代方案。自体HSCT是首选，因为它比同种异体HSCT便宜得多，也更安全。 在CD 34+细胞中靶向基因组修饰的一种新兴方法是使用三链体分子，其形式为三链体形成寡核苷酸或肽核酸。这些分子以序列特异性方式结合双链体DNA，并在与供体DNA分子结合时刺激重组。基因修饰通过细胞自身DNA修复机制的募集发生，而不需要病毒载体。病毒载体已被证明在此类疗法中存在问题。 我们的具体目标是测试使用三链体分子纠正IDUA基因中最突出的基因突变之一W 402 X的可行性。为了使这项技术作为商业疗法可行，我们必须证明我们可以在CD 34+细胞中实现至少3%的基因靶向校正成功率。接下来，我们必须证明我们可以从这些干细胞中产生足够数量的单核细胞，以用作大规模HSCT的桥接治疗的一部分。在第二阶段，我们将进行体内研究，以证明体内疗效，并致力于开发和IND使能研究，以将该技术推向临床研究。 公共卫生相关性：Hurler综合征（HS）是一种毁灭性的遗传性疾病，导致多器官衰竭，中枢神经系统进行性恶化，是由单个基因突变引起的，该基因参与了我们细胞内大糖分子的加工。L2 Diagnostics LLC正在开发一种治疗性基因靶向方案，以纠正导致HS的最突出突变之一。我们的方案将修复HS患者分离的血液干细胞中的这种突变，然后将纠正的细胞重新引入患者体内，可能对疾病的症状产生重大影响。

DESCRIPTION (provided by applicant): This is a proposal to test the feasibility of using triplex molecules for use as a therapeutic to correct Hurler Syndrome (HS). Lysosomal storage diseases (LSDs), such as HS, represent a diverse collection of monogenic disorders that have devastating clinical features. LSDs are caused by mutations in genes that encode lysosomal enzymes and other proteins that are critical for lysosomal function. HS is caused by mutations in the ¿ -L-iduronidase gene (IDUA). For many LSDs, treatment consists of enzyme replacement therapy and supportive care. However, enzyme replacement therapy costs from $90,000 to $565,000 per patient. Alternatively, allogeneic hematopoietic stem cell transplantation (HSCT) represents a highly effective treatment. However, allogeneic HSCT requires an HLA-matched donor, is associated with significant morbidity and mortality and is expensive, about $200,000 per patient. Genetic modification of autologous HSCT, using CD34+ cells, is emerging as a viable therapeutic alternative to allogeneic HSCT. Autologous HSCT is preferred because it is far less expensive and safer than allogeneic HSCT. One emerging approach to targeted genome modification in CD34+ cells is the use of triplex molecules, either in the form of triplex-forming oligonucleotides or peptide nucleic acids. These molecules bind to duplex DNA in a sequence-specific manner and stimulate recombination when combined with donor DNA molecules. Gene modification occurs via recruitment of the cells own DNA repair machinery, without the need for viral vectors. Viral vectors have proven problematic in such therapies. Our Specific Aim is to test the feasibility of using triplex molecules to correct one of the most prominent gene mutations in the IDUA gene, W402X. For this technology to be viable as a commercial therapy, we must demonstrate that we can achieve a success rate of at least 3% for correction of our gene target in CD34+ cells. Next, we must demonstrate that we can generate a sufficient number of monocytes from these stem cells to be utilized as part of a bridging therapy to wholesale HSCT. In Phase II, we will carry out in vivo studies to demonstrate in vivo efficacy and work on developmental and IND enabling studies to move this technology toward clinical studies. PUBLIC HEALTH RELEVANCE: Hurler Syndrome (HS), a devastating genetic disease that results in multiple organ failure with progressive deterioration in the central nervous system, is caused by mutations in a single gene that is involved in the processing of large sugar molecules within compartments in our cells. L2 Diagnostics LLC is proposing to develop a therapeutic gene targeting protocol to correct one of the most prominent mutations responsible for HS. Our protocol would repair this mutation in isolated blood stem cells from HS patients and then reintroduce the corrected cells back into the patient with the potential of having a significant impact on the symptoms of the disease.