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

描述（由申请人提供）：抗肿瘤药物顺铂是治疗各种上皮和脑肿瘤所必需的。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。顺铂的细胞分布仅通过病理学观察、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进行生物化学检测间接检测。这项研究的长期目标是预防耳毒性和永久性耳聋。我们已经开发了一种荧光标记的顺铂分子，它具有生物活性，并且在全身给药后可以在内耳，肾脏和其他器官的单个细胞内检测到。此外，我们可以使用这种结合程序，以确定细胞质顺铂结合蛋白，可能揭示新的细胞内机制的细胞毒性或保护。工作假设是：顺铂通过内淋巴转运到耳蜗毛细胞，并与细胞溶质蛋白结合。本项目的具体目标是：目的1：在三种啮齿动物模型中表征顺铂的耳蜗分布，目的2：确定体外调节顺铂细胞摄取的条件，目的3：验证顺铂从体内内淋巴进入耳蜗毛细胞，目的4：确定顺铂结合蛋白及其在耳蜗中的分布。通过确定哪些细胞吸收顺铂，顺铂进入细胞的机制，并确定其蛋白结合伙伴，我们可以开始开发新的策略，以防止顺铂诱导的耳毒性。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，这对于获得语言和教育技能的儿科患者尤其重要。了解顺铂如何穿过血迷路屏障进入耳蜗的机制对于预防顺铂诱导的耳毒性至关重要。拟议的研究将能够开发新的策略，以防止耳蜗吸收顺铂及其衍生物和随后的耳毒性后遗症，特别是终身耳聋，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.