MUTUAL REGULATION OF P53, MDM2 AND NFKB:

（申请人的摘要）尽管在本发明中有实质性的进展， 治疗儿童急性淋巴细胞白血病（ALL），约25-35% 的患儿会复发或诱导治疗失败。失败 对治疗有反应可能是由于对细胞凋亡的抵抗 由化疗药物诱导。在申请人的儿童ALL研究中 在复发时，他发现表达突变p53基因的白血病细胞， 过表达p53抑制性MDM 2癌基因表达组成性高， 活化的核因子κ B（NF κ B）水平，其与 抗DNA损伤后的凋亡。相反，表达 野生型（WT）-p53通常显示没有或非常低水平的活化的 NFkB。由于wt-p53功能在DNA损伤后的细胞中被诱导，并且由于 在表达mut-p53等位基因或过表达mut-p53等位基因的细胞中， MDM 2，申请人假设， wt-p53可能抑制NF κ B活性。因此，mut-p53的表达 或高水平的MDM 2会增加激活的NFkB的水平， 抑制wt-p53的NF κ B抑制功能。在正常细胞中，p53 在DNA损伤时，NF κ B和NF κ B相互作用，导致细胞凋亡 在细胞中持续不可修复的DNA损伤;这种相互调节的丧失， p53突变或MDM 2过表达将导致对 促骨质疏松剂。为了验证他的假设，他将研究 突变型p53或MDM 2在ALL细胞中上调NF κ B活性。 重要的是，他将研究直接抑制 MDM 2和NFkB激活。这些研究可能具有直接的临床应用， 提示抑制白血病中NF κ B介导的治疗抗性的方法， 其他恶性肿瘤。本研究的具体目的是：（1）评估 突变型p53和过表达的MDM 2激活NF κ B的能力， 转录调节NF κ B/p65基因表达，包括鉴定 p65/RelA中可能的p53或MDM 2结合和反应元件 2）研究mut-p53和MDM 2蛋白在启动子下表达的能力， 通过增强NFkB抑制剂IkBa的周转来激活NFkB，和3）为了 评估阻断MDM 2过表达和增加IkB水平的作用 利用抗MDM 2反义和蛋白酶体对细胞凋亡的敏感性 抑制剂，分别。

DESCRIPTION: (Applicant's Abstract) Despite substantial advances in the treatment of childhood acute lymphoblastic leukemia (ALL), approximately 25-35% of children with this disease will relapse or fail induction therapy. Failure to respond to treatment may result from development of resistance to apoptosis induced by chemotherapeutic agents. In the applicant's studies of pediatric ALL in relapse, he has found that leukemic cells expressing a mutant p53 gene or overexpressing the p53-inhibitory MDM2 oncogene express constitutively high levels of activated Nuclear Factor kappa B (NFkB), which has been linked to resistance to apoptosis following DNA damage. Conversely, ALL cells expressing wild type (wt)-p53 typically show either no or very low levels of activated NFkB. Since wt-p53 function is induced in cells following DNA damage, and since this function is lost in cells expressing a mut-p53 allele or overexpressing MDM2, the applicant hypothesizes that an important apoptosis-inducing effect of wt-p53 may be inhibition of NFkB activity. Accordingly, expression of mut-p53 or high levels of MDM2 would increase the level of activated NFkB by suppressing the NFkB-inhibitory function of wt-p53. Thus, in normal cells, p53 and NFkB would mutually interact in response to DNA damage to permit apoptosis in cells sustaining irreparable DNA damage; loss of this mutual regulation by either p53 mutation or MDM2 overexpression would result in resistance to apoptosis-inducing agents. To test his hypothesis, he will study the potential upregulation of NFkB activity by either mutant p53 or MDM2 in ALL cells. Importantly, he will examine the cellular consequences of directly inhibiting MDM2 and NFkB activation. These studies may have direct clinical application in suggesting ways to inhibit NFkB-mediated therapy resistance in leukemia and other malignancies. The specific aims of this study are 1) To evaluate the ability of mutant p53 and overexpressed MDM2 to activate NFkB by transcriptionally regulating NFkB/p65 gene expression, including identification of possible p53- or MDM2-binding and response elements in the p65/RelA promoter; 2) To investigate the ability of mut-p53 and MDM2 proteins to activate NFkB by enhancing turnover of the NFkB inhibitor IkBa and 3) To evaluate the effect of blocking MDM2 overexpression and augmenting IkB levels on sensitivity to apoptosis utilizing anti-MDM2 antisense and proteasome inhibitors, respectively.