Strategy for the consistent preparation of sufficient non-viral large chromosomal vectors for biopharmaceutical applications

现代药物和疗法对于特定疾病或患者群体变得越来越复杂和具体。一种非常特殊的治疗方法是基因治疗。基因治疗是将基因用作药物。这些基因可以通过使用转基因病毒携带基因或通过使用非病毒方法传递给患者，该方法使用从细菌中分离的称为质粒的环状DNA分子。这两种技术仍处于起步阶段，但已经在疫苗接种、癌症治疗和遗传疾病矫正方面取得了巨大的医学进展。然而，病毒有几个缺点，包括安全考虑和限制携带大量的遗传信息。另一方面，被称为BAC（细菌人工染色体）的大质粒DNA分子可以被修饰以掺入广泛的重要控制区域，这些区域允许基因在正确的组织中和正确的时间表达。这种较大的尺寸构成了科学和工业挑战，因为尺寸限制了单个细菌中可以产生的量，此外，由于BAC是脆弱的，尺寸还需要特殊的加工考虑。目前的挑战是能够制备BAC分子，其数量足以治疗能够从这些药物中受益的人数。我们需要研究并开发在生物制造业中大规模制造DNA分子的方法。DNA分子的质量也是至关重要的。它们需要以最适合于递送给人类（和动物）并且不含污染物质的形式存在。我们提出的研究将使工业能够制造可以大规模制造的DNA分子。拟议的研究成果将允许其他参与基因治疗的人制备用于治疗癌症的大DNA分子，这项研究还将使相关学科的其他研究人员受益于制造和操纵大型DNA结构的能力。我们还将探索以正确的三维形式制造DNA的方法，以便有效地被细胞吸收，DNA是一种有效的药物

Modern medicines and therapies are becoming increasingly complex and specific for a particular disease or group of patients. One very specific type of therapy is Gene Therapy. Gene therapy is the use of genes as medicines. These genes can be delivered to patients either by the use of genetically modified viruses to carry the genes or by using non-viral methods which employ circular DNA molecules called plasmids isolated from bacteria. Both techniques are still in their infancy but are already promising huge medical advances in vaccination, cancer therapy and the correction of genetic disorders. However, viruses suffer from several drawbacks including safety considerations and limited to carry large genetic information. On the other hand large plasmid DNA molecules called BACs ( Bacterial Artificial Chromosomes) can be modified to incorporate a wide range of important control regions which allow expression of the gene in the correct tissue and at the correct time. This larger size poses a scientific and industrial challenge because the size limits the amount that can be made in a single bacterium, in addition, the size also calls for special processing considerations because BACs are fragile. The challenge is to be able to prepare the BAC molecules in the amounts needed to treat the numbers of people who could benefit from these medicines. We need to investigate then develop methods of making the DNA molecules at large scales in the biomanufacturing industry. The quality of the DNA molecules is also paramount. They need to be in the form that is most appropriate for delivering to humans (and animals) and free of contaminating material. The research we propose will enable industry to make DNA molecules that can be made at the large scales. The proposed research outcomes will allow others involved in gene therapy to prepare large DNA molecules for the treatment of cancers, for vaccines and to correct genetic disorders The science proposed will also allow other researchers in related disciplines to benefit from being able to make and manipulate large DNA constructs .We will also explore ways of making the DNA in the correct three dimensional form needed for efficient uptake into cells so that the DNA is an effective medicine.