骨巨细胞瘤中H3F3A G34W蛋白表达阴性病例的基因突变类型分析

Types of Gene Mutation in H3F3A G34W-Negative Cases in Giant Cell Tumor of Bone

目的:骨巨细胞瘤(giant cell tumor of bone,GCTB)的病理特征是病灶中出现单核细胞、基质细胞和破骨细胞样多核巨细胞,高达95%的GCTB携带有H3F3A G34W突变。本文评估了GCTB中H3F3A G34W蛋白表达阴性病例出现的其他罕见H3F3A分子突变类型。方法:收集2017年1月至2022年8月中山大学肿瘤防治中心收治的15例免疫组织化学染色评估H3F3A G34W蛋白表达为阴性的GCTB病例,并对阴性病例采用Sanger测序验证H3F3A的突变类型。结果:对15例H3F3A G34W蛋白表达阴性的GCTB患者病例采用DNA Sanger测序法检测H3F3A突变情况,测序结果显示10例存在互斥突变,分别为:7例G34L(Gly 34 Leu,7/15,46.7%),2例G34V(Gly 34Val,2/15,13.3%),1例G34R(Gly 34 Arg,1/15,6.7%),其余5例均为野生型(Gly 34,5/15,33.3%)。所有阴性病例通过Sanger测序验证后发现并无G34W突变。H3F3A突变蛋白表达阳性及分子突变皆可支持GCTB的诊断。结论:H3F3A突变是骨巨细胞瘤高度敏感和特异的标志物。免疫组织化学是一种快速、灵敏和可靠的方法,用于识别骨巨细胞瘤中H3F3A蛋白的突变,但仅可检测H3F3A G34W突变亚型,对其他罕见突变类型需Sanger测序甚至高通量测序进一步验证。

Objective : Giant cell tumor of bone(GCTB) is characterized by mononuclear cells, stromal cells and osteoclast-like multi-nucleated giant cells;up to 95% of GCTB are with H3F3A G34W mutation. This article was designed to evaluate other rare molecular types of H3F3A mutation in protein-negative cases in GCTB. Methods: A total of 15 GCTB cases with negative H3F3A G34W protein expression assessed by immunohistochemical staining in Cancer Center of Sun Yat-sen University from January 2017 to August 2022 were collected, and Sanger sequencing was used to verify the type of H3F3A mutation in H3F3A G34W-negative cases. Results: The H3F3A mutation was detected by Sanger sequencing in 15 GCTB patients with negative H3F3A G34W protein expression. The sequencing results showed that there were mutually exclusive mutations in 10 cases, namely, 7 cases of G34L(Gly 34 Leu, 7/15, 46.7%), 2 cases of G34V(Gly 34 Val, 2/15, 13.3%), and 1 case of G34R(34 Arg, 1/15, 6.7%), and the remaining 5 cases were wild type(Gly 34, 5/15, 33.3%). All H3F3A G34W-negative cases were verified by Sanger sequencing, and no G34W mutation was found. Detection of mutant H3F3A protein and molecular mutation can support the diagnosis of GCTB. Conclusion: H3F3A mutation is a highly sensitive and specific marker for giant cell tumor of bone. Immunohistochemistry is a rapid, sensitive and reliable method for identifying H3F3A G34W mutation in H3F3A protein in giant cell tumors of bone. Further validation of other rare mutation types should be performed by Sanger sequencing,or even high throughput sequencing.