肿瘤原位液中肿瘤来源DNA实时监测脑胶质母细胞瘤术后替莫唑胺诱导超突变的临床应用价值

Clinical application value of tumor-derived DNA in tumor in situ fluid for real-time monitoring temozolamine-induced hypermutations after glioblastoma surgery

目的:探讨脑肿瘤原位液(tumor in situ fluid,TISF)来源的肿瘤DNA(tumor-derived DNA)测序实时监测脑胶质母细胞瘤(GBM)术后替莫唑胺(temozolomide,TMZ)诱导超突变的临床应用价值。方法:收集2019年3月至2021年6月于我院神经外科接受诊治的21例GBM患者的肿瘤组织标本及术后TMZ辅助治疗期间的TISF标本和CSF标本,共50个样本(4个CSF、19个组织标本、27个TISF),从TISF和CSF中提取无细胞DNA(cell-free DNA,cfDNA)进行肿瘤来源DNA靶向测序,表征脑胶质母细胞瘤术后基因组景观,并与肿瘤组织标本测序结果对比,鉴定出在术后TMZ辅助化疗期间基因组发生超突变的GBM患者。结果:21例GBM患者平均年龄(53.2±13.5)岁,18例为IDH野生型,3例为IDH突变型,初始MGMT甲基化均阳性。共检测到67个不同基因的785个突变,每个个体平均突变数为15.7,最常见的改变基因为TP53(35%)、NF1(33%)、TSC2(29%)、PTEN(27%)、PTCH1(23%),最常见的突变类型是错义突变(65%),其次是多重突变(13%)和移码突变(12%)。与初始肿瘤组织标本测序结果对比,共有2例患者经TISF肿瘤来源DNA测序检测出在用药期间发生基因组超突变,并证实为TMZ诱导产生的超突变。结论:通过TISF肿瘤来源DNA在体动态监测,可实时在体表征脑胶质母细胞瘤术后肿瘤基因景观及监测替莫唑胺诱导的超突变,通过基因组分析,早期发现肿瘤耐药,提示临床调整后续治疗方案。

Objective:To explore the clinical value of real-time monitoring of temozolamide-induced supermutations after glioblastoma surgery by tumor DNA sequencing from brain tumor in situ.Methods:The tumor tissue specimens of 21 GBM patients who were diagnosed and treated in our hospital from March 2019 to June 2021,as well as TISF specimens and CSF specimens during postoperative TMZ adjuvant treatment,were collected,a total of 50 specimens(4 CSF,19 tissue specimens,27 TISF),extract cell-free DNA(cfDNA)from TISF and CSF for tumor-derived DNA targeted sequencing,characterize the postoperative genomic landscape of glioblastoma,and correlate it with the tumor.Comparison of the sequencing results of tissue samples identified GBM patients with hypermutation in their genome during postoperative TMZ adjuvant chemotherapy.Results:The average age of 21 GBM patients was(53.2±13.5)years,18 were IDH wild-type,3 were IDH mutant,and the initial MGMT methylation was positive.A total of 785 mutations in 67 different genes were detected.The average number of mutations per individual was 15.7.The most common altered genes were TP53(35%),NF1(33%),TSC2(29%),PTEN(27%)and PTCH1(23%),the most common type of mutation is missense mutation(65%),followed by multiple mutations(13%)and frameshift mutations(12%).Compared with the sequencing results of initial tumor tissue specimens,a total of 2 patients were detected by TISF tumor-derived DNA sequencing to have genomic hypermutations during medication,and it was confirmed that they were hypermutations induced by TMZ.Conclusion:Through the dynamic monitoring of TISF tumor-derived DNA in vitro,the tumor gene landscape after glioblastoma surgery can be characterized in real time and the supermutations induced by temozolidomide can be monitored.Through genomic analysis,tumor drug resistance can be found early,indicating clinical adjustment of subsequent treatment plans.