Mechanism and Uses of Transmembrane Helix Insertion by Soluble Peptides

 描述(由申请人提供)：基础研究将导致医学上的新方法，这是NIH的一个中心思想，我们相信我们已经找到了一个。由于美国国立卫生研究院早期的资助，我们发现了一系列多肽：(1)靶向体内的酸性组织，包括肿瘤；(2)可以将极性分子输送到细胞内，在细胞质中释放它们。对多肽的研究从结构、能量和动力学方面对多肽插入膜的基础科学有了新的见解，随着我们对原理的更多了解，我们将能够为特定的应用设计更好的多肽版本。这些多肽，我们称之为“PHLIPs”(即pH(低)插入多肽)，主要以非结构单体的形式溶解在水溶液中，以非结构单体的形式结合到双层或膜的表面，并折叠成螺旋，在酸性环境中插入跨膜。我们已经建立了多肽插入的基本能量学和动力学。我们已经证明了PHLIP的标记版本在小鼠模型中靶标和成像多种肿瘤，并且成像显示非常小的肿瘤(1毫米)并准确地识别肿瘤边界。我们还证实了小的货物分子可以在PHLIP的插入端被运送到癌细胞。水溶性治疗分子可以通过在细胞内不稳定但在细胞外稳定的键以货物的形式连接到PHLIP的插入端。当PHLIP在低pH值下折叠并通过膜运送货物时，键断裂，货物在胞浆中释放，在那里它可能有治疗效果。当注射到小鼠体内时，最近已经证明一种形式的PHLIP递送治疗在小鼠模型中成功地靶向并有效地治疗了淋巴瘤。我们建议的研究旨在完善PHLIP设计，以用于毒素和肽核酸的靶向递送。我们还计划了一种更先进的肿瘤荧光标记方法和一种新的测量细胞表面pH的方法，这是PHLIPs酸性靶向的关键参数。如果成功，在了解更多关于多肽-膜相互作用的生物物理学和细胞表面酸性的生理学的同时，我们将把这项技术推向临床应用。

 DESCRIPTION (provided by applicant): It is a central idea of the NIH that basic research will lead to new approaches in medicine, and we believe that we have found one. As a result of earlier funding by the NIH, we have discovered a family of peptides that (1) target acidic tissues in vivo, including tumors, and (2) can deliver polar molecules into cells, releasing them in the cytoplasm. Studies of the peptides have led to new insights about the basic science of peptide insertion into membranes in terms of structure, energetics and kinetics, and as we learn more about the principles, we will be able to design better versions of peptides for specific applications. The peptides, which we call "pHLIPs" (for pH (Low) Insertion Peptides) are soluble as mostly unstructured monomers in aqueous solution, bind as unstructured monomers to the surfaces of bilayers or membranes, and fold to make helices that insert across membranes when the environment is acidic. We have established the basic energetics and kinetics of peptide insertion. We have shown that a labeled version of pHLIP targets and images many kinds of tumors in mouse models, and that the imaging reveals very small tumors (1mm) and accurately identifies tumor borders. We have also established that small cargo molecules can be delivered into cancer cells on the inserting end of pHLIP. A water-soluble therapeutic molecule can be attached as cargo to the inserting end of pHLIP by a bond that is unstable in side a cell, but stable outside the cell. When pHLIP folds at low pH and delivers the cargo across the membrane, the bond breaks and the cargo is released in the cytosol, where it may have a therapeutic effect. When injected into a mouse, a form of pHLIP-delivered treatment has been recently demonstrated to successfully target and effectively treat lymphoma in a mouse model. Our proposed studies are aimed at refining pHLIP designs for specific uses in targeted delivery of toxins and Peptide Nucleic Acids. We also plan a more advanced method for fluorescence labeling of tumors and a new method for measuring the pH at cell surfaces, a key parameter for the acidic targeting of pHLIPs. If successful, while learning more about the biophysics of peptide-membrane interactions and the physiology of cell surface acidity, we will have advanced the technology toward clinical applications.