MOLECULAR TARGETS OF TRANSLOCATION T(4;14) IN MULTIPLE MYELOMA PATHOGENESIS

描述（由申请人提供）：多发性骨髓瘤（MM）是美国第二常见的血液恶性肿瘤，每年约有14，500名美国人被诊断出，占所有癌症死亡的2%（SEER.cancer.gov）。MM是分泌抗体的浆B细胞的不可治愈的恶性肿瘤，其病因学知之甚少。染色体易位t（4;14）（p16.3;q32.33）与总生存期缩短相关，见于15%的MM患者。4p16.3的候选癌基因WHSC 1（MMSET）编码组蛋白甲基转移酶，由于MM细胞中的t（4;14）而过度表达。令人惊讶的是，当在小鼠中表达时，WHSC 1 cDNA的过表达未能转化细胞系或诱导任何表型。我们鉴定了ACA 11，一种孤儿盒H/ACA小核仁RNA（snoRNA），其编码宿主于WHSC 1的4p16.3处，在携带t（4;14）的细胞系和MM患者样本中上调。ACA 11调节氧化应激，促进MM细胞生长并赋予化疗抗性。ACA 11是一种新的致癌非编码RNA，也在其他癌症类型中表达。 与t（4;14）共同参与骨髓瘤发生的突变尚不清楚，但13号染色体缺失（Del（13））与MM中的t（4;14）高度相关。Del（13）见于85-94%的t（4;14）患者，并与不良预后相关。我们将MM患者样本中的一个最小缺失区域映射到13 q14的视网膜母细胞瘤（RB 1）基因。我们的模型是，ACA 11的表达与RB 1的损失合作，导致预后不良MM。我们建议的特点ACA 11在多发性骨髓瘤中的作用。具体目标1：ACA 11和组蛋白甲基转移酶WHSC 1之间的合作。我们假设，ACA 11与MMSET蛋白表达在骨髓瘤发病机制中的合作，WHSC 1组蛋白甲基转移酶活性是合作所必需的。我们将在细胞系和小鼠中共表达野生型WHSC 1或HMT死亡WHSC 1单独或与ACA 11。具体目标2：目标2：ACA 11如何影响ROS水平和对化疗的敏感性？这些效应需要ACA 11的哪些域和伙伴？我们的数据表明，ACA 11调节活性氧和骨髓瘤化疗的抵抗。我们将确定ACA 11的哪些残基是该功能所需的。我们推测，ACA 11的RNA结合残基将是ACA 11的氧化应激功能，但ACA 11的蛋白质结合域将是至关重要的。具体目标3：Rb 1缺失背景下ACA 11介导的恶性转化。50%的MM患者和90%的t（4;14）患者的13号染色体长臂缺失。我们假设RB 1缺失与t（4;14）在骨髓瘤发病机制中协同作用。我们将测试ACA 11和WHSC 1表达对B细胞发育、氧化应激和Rb 1缺乏背景下转化的影响。最后，我们将[通过对患者样本进行RNA测序来鉴定t（4;14）的靶基因。]对t（4;14）如何促进骨髓瘤发生的进一步了解对癌症生物学具有广泛的意义，并将促进对预后不良MM患者的新治疗方法的开发。

DESCRIPTION (provided by applicant): Multiple Myeloma (MM) is the second most common hematologic malignancy in the United States, diagnosed in approximately 14,500 Americans each year, and is responsible for 2% of all cancer deaths (SEER.cancer.gov). MM is an incurable malignancy of antibody-secreting plasma B-cells whose etiology is poorly understood. Chromosomal translocation t(4;14)(p16.3;q32.33) is associated with shorter overall survival and is found in 15% of MM patients. The candidate oncogene WHSC1 (MMSET) at 4p16.3 encodes a histone methyltransferase over-expressed as a result of the t(4;14) in MM cells. Surprisingly, over-expression of WHSC1 cDNA's failed to transform cell lines or to induce any phenotype when expressed in mice. We identified ACA11, an orphan box H/ACA small nucleolar RNA (snoRNA) encoded hosted within WHSC1 at 4p16.3, to be up-regulated in cell lines and MM patient samples harboring t(4;14). ACA11 modulates oxidative stress, contributes to MM cell growth and confers resistance to chemotherapy. ACA11 is a novel oncogenic non-coding RNA and is also expressed in other cancer types. Mutations that cooperate with t(4;14) in myeloma initiation are unknown, but chromosome 13 deletions (Del(13)) are highly correlated with the t(4;14) in MM. Del(13) is found in 85-94% of patients with t(4;14) and associated with poor prognosis. We mapped a minimally deleted region in MM patient samples to the retinoblastoma (RB1) gene at 13q14. Our model is that ACA11 expression cooperates with RB1 loss to cause poor-prognosis MM. We propose to characterize the role of ACA11 in multiple myeloma. Specific Aim 1: Cooperation between ACA11 and the histone methyltransferase WHSC1. We hypothesize that ACA11 cooperates with MMSET protein expression in myeloma pathogenesis and that WHSC1 histone methyltransferase activity is required for cooperation. We will co-express wild-type WHSC1 or HMT-dead WHSC1 alone or with ACA11 in cell lines and mice. Specific Aim 2: Aim 2: How does ACA11 affect levels of ROS and sensitivity to chemotherapy? What domains and partners of ACA11 are required for these effects? Our data suggest that ACA11 modulates reactive oxygen species and resistance to chemotherapy in myeloma. We will determine which residues of ACA11 are required for this function. We hypothesize that ACA11 RNA binding residues will be dispensable for ACA11's oxidative stress function, but that ACA11's protein-binding domains will be critical. Specific Aim 3: ACA11-mediated malignant transformation in the context of Rb1 deletion. The long arm of chromosome 13 is deleted in 50% of all MM patients, and in 90% of patients with the t(4;14). We hypothesize that RB1 loss cooperates with the t(4;14) in myeloma pathogenesis. We will test the effects of ACA11 and WHSC1 expression on B-cell development, oxidative stress and transformation in the context of Rb1 deficiency. Lastly, we will [identify target genes of t(4;14) by RNA- sequencing patient samples.] A betting understanding of how the t(4;14) contributes to myelomagenesis has broad implications to cancer biology and will facilitate the development of novel therapeutic approaches to patients with poor-prognosis MM.