Mode of Action of SQSTM1 Mutations in Paget's Disease Bone

SQSTM1 突变在佩吉特病骨中的作用方式

• 批准号: 7139867
• 负责人: MARC F HANSEN
• 金额: $ 16.28万
• 依托单位: UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-07-15 至 2008-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7139867
• 关键词: bone; emotions; gene mutation; osteosarcoma

DESCRIPTION (provided by applicant): Paget's Disease of bone is a condition in which rapid bone formation occurs, altering the strength and shape of the bone. Predisposition to familial Paget's Disease has been linked to a number of loci, including the Sequestosome 1 (SQSTM1) locus where germline mutations have been identified in 40% of the familial cases. Germline SQSTM1 mutations have also been found in some apparent sporadic cases of Paget's Disease, suggesting that these may be de novo familial cases. In all but one case, the constitutional mutations have been found to be heterozygous. This suggests that the mutation may be acting in a dominant manner. However, it is also possible that the mutation is acting as a dominant negative or haploinsufficiency mutation, or as a recessive mutation that has undergone loss of heterozygosity in the affected bone. We were curious as to the nature of the mutation in the affected bone. We chose to examine the SQSTM1 locus in the affected bones of sporadic patients to see: 1) whether somatic mutations in SQSTM1 occurred in these bones and 2) whether the mutation was heterozygous or homozygous in the affected bone. When we sequenced the SQSTM1 gene in both the affected bone and matched peripheral blood, we found a P392L mutation in 4 of 5 samples of affected bone and none of the matched blood samples. What was more surprising was that in the affected bone, the relative frequency of the P392L mutation in the samples ranged from 10% to 28%, suggesting that the bone was mosaic for the mutation. The relative frequency of the mutation in each individual sample was consistent upon repeated resampling and resequencing. Next, since osteosarcoma is a complication of Paget's Disease, we were curious to see whether SQSTM1 mutations were present in pagetic osteosarcomas. When we screened the SQSTM1 gene in these tumors, we found that 4 of 5 osteosarcomas had homozygous P392L mutations. Again, no mutations were detected in the matched normal bone. Together, this evidence causes a paradigm shift in our model of Paget's Disease. It suggests that while mutations in SQSTM1 are predisposing in familial Paget's Disease, they are not the initiating event in sporadic Paget's Disease. Secondly, this suggested that the tumors arose from osteoblastic cells that contained the SQSTM1 mutation and that osteoblasts play a role in the etiology of Paget's Disease. Our hypothesis is that mutations in SQSTM1 occur somatically in a subset of osteoblast-like cells in pagetic bone and that these somatic mutations become homozygous during pagetic osteosarcoma tumorigenesis. To test this hypothesis, we propose the following specific aims: 1) To test whether somatic SQSTM1 mutations are heterozygous and clonal in the pagetic bone. 2) To test whether loss of heterozygosity at the SQSTM1 locus occurs in the pagetic bone or during pagetic tumorigenesis.

描述（由申请人提供）：Paget的骨骼疾病是发生快速骨形成的疾病，从而改变了骨骼的强度和形状。家族性paget病的易感性与许多基因座有关，包括隔离的1（SQSTM1）基因座，在40％的家族病例中已经鉴定出种系突变。在某些明显的偶然病例的paget病例中，还发现了种系SQSTM1突变，这表明这些可能是从头家族病例。除一种情况外，所有宪法突变被发现是杂合的。这表明突变可能以主导方式起作用。但是，该突变也可能充当主导性阴性或单倍宽度突变，或者是导致受影响骨骼中杂合性丧失的隐性突变。我们对受影响骨骼突变的性质感到好奇。我们选择检查偶发患者受影响的骨骼中的SQSTM1基因座，以查看：1）SQSTM1中的体细胞突变是否发生在这些骨骼中，以及2）突变是在受影响骨骼中的杂合性还是纯合子。当我们在受影响的骨骼和匹配的外周血中对SQSTM1基因进行了测序时，我们在5个受影响的骨骼样本中的4个中发现了P392L突变，没有匹配的血液样本。更令人惊讶的是，在受影响的骨骼中，样品中p392l突变的相对频率范围为10％至28％，这表明该突变的骨骼是镶嵌的。在重复重新采样和重新取代时，每个样品中突变的相对频率是一致的。接下来，由于骨肉瘤是Paget疾病的并发症，因此我们很好奇地看到SQSTM1突变是否存在于Pagetic骨肉瘤中。当我们在这些肿瘤中筛选SQSTM1基因时，我们发现5个骨肉瘤中有4个具有纯合P392L突变。同样，在匹配的正常骨中未检测到突变。这些证据共同导致我们的paget病模型发生了范式转变。这表明，尽管SQSTM1中的突变在家族性Paget的疾病中易于体现，但它们并不是零星Paget病的起始事件。其次，这表明肿瘤来自含有SQSTM1突变的成骨细胞，并且成骨细胞在Paget病的病因中起作用。我们的假设是，SQSTM1中的突变在pagetic骨中成骨细胞样细胞的子集中发生，并且这些体细胞突变在Pagotic骨肉瘤肿瘤发生过程中变得纯合。为了检验这一假设，我们提出以下特定目的：1）测试体细胞SQSTM1突变是否是杂合骨中的杂合子和克隆的。 2）测试SQSTM1基因座的杂合性丧失是在雌蕊骨中还是在雌蕊肿瘤发生期间发生。