Dual role of IL-16 in dysregulated growth of CTCL cells

描述(由申请人提供)：与Sezary综合征相关的T细胞数量的增加可能是由于过度增殖和细胞凋亡减少所致。然而，导致这些影响的机制尚不清楚。我们以前报道过IL-16是一种双功能蛋白，参与调节T细胞的生长。分泌(成熟)部分被归类为一种能力因子，通过与CD4细胞外结合来调节T细胞的生长，促进CD25的表达，并从G0期过渡到G1期。最近在T细胞的核中发现了前体(Pro-IL-16)，它通过调节Skp-2的转录来影响T细胞的增殖。Pro-IL-16作为支架蛋白与辅助因子GABP1和HDAC-3相互作用，抑制Skp2的转录，从而间接调节与T细胞增殖相关的主要周期蛋白激酶抑制剂之一p27kip1的降解。我们的初步数据表明，在CTCL患者的T细胞中，存在以核优先丢失为特征的细胞内PRO-IL-16的重新分布。这种缺失可以归因于PDZ1结构域的突变，与核伴侣蛋白HSC70的结合缺失有关。随着细胞内存储的耗尽，在I-III期，核亲IL-16的丢失似乎与成熟IL-16分泌的增加结合在一起。此外，IL-16的产生总体上呈进行性减少，这可能与启动子的超甲基化有关。因此，我们的假设是，CTCL的发生与IL-16启动子的高甲基化有关，从而导致蛋白质产量的进行性减少。此外，原IL-16的PDZ1突变导致HSC70结合丧失，显著降低核表达或功能，同时增加成熟蛋白的分泌。分泌的IL-16与表面表达的CD4相互作用，进一步诱导肿瘤细胞的细胞周期进展。在这些研究中，我们建议研究甲基化对IL-16启动子和编码区的影响；描述阻止前IL-16核转位的机制，并建立野生型前IL-16表达后Sezary T细胞的细胞效应；最后确定分泌的IL-16对T细胞增殖和T调节细胞潜在扩增的作用。公共卫生相关性：我们已经证明，在细胞周期内，核前IL-16具有调节T细胞进程的功能，而在Sezary T细胞中，前IL-16已经失去了移位到细胞核内的能力，使细胞过度增殖。在这些研究中，我们建议确定参与调节原-IL-16核转位的机制；表达的野生型原-IL-16调节恶性T细胞的过度增殖的能力；以及研究分泌的IL-16在诱导肿瘤抑制基因表达缺失和由此导致的过度增殖中的作用。这些研究的完成不仅将确定Sezary T细胞异常生长的机制，还将阐明潜在治疗的新方法。

DESCRIPTION (provided by applicant): The increase in T cell numbers associated with Sezary Syndrome likely occurs as a result of hyperproliferation as well as decreased apoptosis. The mechanisms involved resulting in these effects however are not clearly understood. We have previously reported that IL-16 is a bi-functional protein involved in regulation of T cell growth. The secreted (mature) portion has been classified as a competence factor that regulates T cell growth through extracellular association with CD4, promoting CD25 expression and transition from the G0 phase to G1. The pro-piece (pro-IL-16) has been recently detected in the nucleus of T cells where it affects proliferation through the regulation of Skp-2 transcription. Pro-IL-16 functions as a scaffold protein which interacts with co- factors GABP1 and HDAC-3 to suppress Skp2 transcription, therefore indirectly regulating degradation of one of the major cyclin kinase inhibitors associated with T cell proliferation, p27kip1. Our preliminary data now indicates that in T cells from CTCL patients there is an initial redistribution of intracellular pro-IL-16 characterized by preferential loss in the nucleus. This loss can be attributable to mutations in the PDZ1 domain, correlating to loss of binding by the nuclear chaperone protein HSC70. The loss of nuclear pro-IL-16 also appears to be combined with an observed increase in secretion of mature IL-16 in stages I-III as intracellular stores become depleted. In addition, there is an overall progressive reduction in the production of IL-16 that may be associated with promoter hypermethylation. It is our hypothesis therefore that the onset of CTCL is associated with hypermethylation of the IL-16 promoter resulting in a progressive reduction in protein production. In addition, mutations in PDZ1 of pro-IL-16 results in loss of HSC70 binding, significantly reducing nuclear expression or function, while increasing secretion of mature protein. Secreted IL-16 interacts with surface expressed CD4 to further induce cell cycle progression in the malignant cells. In these studies we propose to investigate the effects of methylation on the IL-16 promoter as well as within the coding region; to delineate the mechanism by which pro-IL-16 is prevented from nuclear translocation and establish cellular effects of Sezary T cells following expression of wild type pro-IL-16; and finally to determine the role of secreted IL-16 on T cell proliferation and potential expansion of T regulatory cells. PUBLIC HEALTH RELEVANCE: We have shown that nuclear pro-IL-16 functions to regulate T cell progression within the cell cycle and that in Sezary T cells pro-IL-16 has lost the ability to translocate into the nucleus rendering the cells hyperproliferative. In these studies we propose to identity the mechanisms involved in regulating nuclear translocation of pro-IL-16; the ability of expressed wild type pro-IL-16 to regulate hyperproliferation in malignant T cells; and to investigate the role of secreted IL-16 to induce loss of tumor suppressor expression with resultant hyperproliferation. Completion of these studies will not only identify a mechanism for dysregulated growth in Sezary T cells but will elucidate a novel approach to potential therapeutics.