Engineering a Human Organ for Dry Storage

描述(由申请人提供)： 组织工程是一个具有巨大潜力的领域，目前的收入(2000年2.32亿美元，《遗传工程新闻》2001年9月15日)远远低于1000亿美元的估计市场(Barron‘s，5/15/00)。这一差距在很大程度上是由于保质期短和储存成本高的重大障碍。Stratech建议开发克服这些障碍的技术，方法是利用源细胞的遗传操作，加上保存液工程的最新进展，以创造能够在室温下保存活细胞和器官的方法。 目前，组织是通过缓慢冷冻来保存的，在此过程中，它们遭受了许多破坏作用。快速冷冻或玻璃化通过将样品保存在无定形玻璃中避免了这些问题，但玻璃化可能会导致致命的细胞内冰形成。海藻糖是一种葡萄糖二糖，可以增强保存液的玻璃化特性；不幸的是，海藻糖不能穿透细胞膜，因此必须使用新的方法来传递细胞内的海藻糖。此外，直到最近，对医学上有用的样品大小进行玻璃化处理还不现实，但最近的研究表明，精心设计保存液可以极大地提高海藻糖提高玻璃化转变温度的能力。 Stratech正在开发一种用于治疗应用的人类角质形成细胞系，称为NIKS细胞。NKS细胞是这一提议的理想载体，因为它们可以在体外永久培养，并可以用来制造一个有功能的人体器官。此外，它们是无病原体、不会致癌的，它们概括了人类皮肤的正确结构。NKS细胞和从中培养出来的皮肤组织能够进行稳定的基因修饰，从而有可能优化它们在干燥状态下的生存。

DESCRIPTION (provided by applicant): Tissue engineering is a field with tremendous untapped potential, current revenues ($232 million in 2000, Genetic Engineering News 9/15/01) fall far short of the estimated market of $100 billion (Barron's, 5/15/00). This gap is in large part due to the significant barriers of short shelf life and high storage cost. Stratatech proposes to develop technology overcoming these hurdles using genetic manipulations of source cells coupled with recent advances in preservation solution engineering to create methods enabling room temperature storage of living cells and organs. Currently tissues are preserved by slow freezing during which they suffer numerous damaging effects. Rapid freezing or vitrification avoids these issues by preserving samples in an amorphous glass, but vitrification can give rise to lethal intracellular ice formation. Agents such as trehalose, a disaccharide of glucose, enhance the vitrification properties of preservation solutions; unfortunately, trehalose cannot penetrate cell membranes so novel methods must be used to deliver intracellular trehalose. Furthermore, until recently, vitrification of medically useful sample sized was not practical, but recent work has shown that careful engineering of the preservation solution can greatly enhance trehalose's ability to raise glass transition temperatures. Stratatech is developing a line of human keratinocytes for therapeutic applications called NIKS cells. NIKS cells are an ideal vehicle for this proposal in that they can be perpetually cultured in vitro and they can be used to manufacture a functional human organ. In addition they are pathogen-free, non-tumorigenic and they recapitulate the correct architecture of human skin. NIKS cells and skin tissue grown from them are capable of stable genetic modifications making it possible to optimize their survival in the dry state.