Localization of cisplatin and cisplatin-binding proteins in the inner ear

描述(申请人提供)：抗肿瘤药物顺铂是治疗各种上皮性肿瘤和脑肿瘤的必备药物。然而，它具有耳毒性、神经毒性和肾毒性，在接受这些药物治疗的患者中会导致永久性耳聋和急性肾衰竭。顺铂的细胞分布只能通过病理观察、低分辨率正电子发射断层扫描、放射自显影或使用高效液相色谱法进行生化检测来间接检测。这项研究的长期目标是预防耳毒性和永久性耳聋。我们已经开发出一种荧光标记的顺铂分子，它具有生物活性，在全身给药后，可以在内耳、肾脏和其他器官的单个细胞中检测到。此外，我们可以使用这种连接过程来鉴定胞质顺铂结合蛋白，这些蛋白可能揭示细胞内新的细胞毒性或保护机制。顺铂的工作假设是：顺铂通过内淋巴运输到耳蜗毛细胞，并与胞浆蛋白结合。本项目的具体目的是：目的1：研究顺铂在三种啮齿动物模型中的耳蜗区分布，目的2：确定体外调节细胞摄取顺铂的条件，目的3：验证顺铂在体内从内淋巴进入耳蜗毛细胞，目的4：鉴定顺铂结合蛋白及其在耳蜗组织中的分布。通过确定哪些细胞摄取顺铂、顺铂进入细胞的机制以及确定其蛋白结合伙伴，我们可以开始开发新的策略来预防顺铂引起的耳毒性。这将使临床医生能够更有效地使用顺铂和相关药物，同时保留听觉功能，这对获得语言和教育技能的儿科患者尤其重要。了解顺铂如何通过血迷路屏障进入耳蜗腔的机制对于预防顺铂引起的耳毒性至关重要。这项拟议的研究将有助于开发新的策略，以防止耳蜗顺铂及其衍生物的摄取和随后的耳毒性后遗症，特别是终生耳聋、耳鸣和前庭缺陷。

DESCRIPTION (provided by applicant): The anti-neoplastic drug cisplatin is essential for treating a variety of epithelial and brain tumors. However, it is ototoxic, neurotoxic and nephrotoxic, causing permanent deafness and acute kidney failure in patients treated with these drugs. The cellular distribution of cisplatin has been only detected indirectly via pathological observations, by low-resolution positron emission tomography, autoradiography, or biochemically using high-performance liquid chromatography. The long-term goal of this research is to prevent ototoxicity and permanent deafness. We have developed a fluorescently-tagged cisplatin molecule that is bio-active and can be detected within individual cells in the inner ear, kidney and other organs after systemic administration. In addition, we can use this conjugation procedure to identify cytoplasmic cisplatin-binding proteins that may reveal novel intracellular mechanisms of cytotoxicity or protection. The working hypothesis is: Cisplatin trafficks to cochlear hair cells via endolymph, and binds to cytosolic proteins. The specific aims of this project are to: Aim 1: characterize the cochlear distribution of cisplatin in three rodent models, Aim 2: identify the conditions that modulate cellular uptake of cisplatin in vitro, Aim 3: verify that cisplatin enters cochlear hair cells from endolymph in vivo, Aim 4: identify cisplatin-binding proteins and their distribution in the cochlea. By determining which cells take up cisplatin, the mechanisms by which it enters cells and identifying its protein-binding partners, we can begin to develop new strategies to prevent cisplatin-induced ototoxicity. This will allow clinicians to use cisplatin and related drugs more efficaciously while preserving auditory function, especially important in pediatric patients acquiring language and educational skills. Understanding the mechanisms of how cisplatin crosses the blood-labyrinth barrier to enter the cochlea is crucial to prevent cisplatin-induced ototoxicity. The proposed research will enable the development of new strategies to prevent cochlear uptake of cisplatin and its derivates and subsequent ototoxic sequelae, particularly life-long deafness, tinnitus and vestibular deficits.