Abstract：
The study was conducted to assess the feasibility of integrating state-of-the-art sequencing techniques and flow cytometry into diagnostic workup of pediatric lymphoma. RNA sequencing (RNAseq), whole exome sequencing, and flow cytometry were implemented into routine diagnostic workup of pediatric biopsies with lymphoma in the differential diagnosis. Within 1 year, biopsies from 110 children (122 specimens) were analyzed because of suspected malignant lymphoma. The experience with a standardized workflow combining histology and immunohistochemistry, flow cytometry, and next-generation sequencing technologies is reported. Flow cytometry was performed with fresh tissue in 83% (102/122) of specimens and allowed rapid diagnosis of T-cell and B-cell non-Hodgkin lymphomas. RNAseq was performed in all non-Hodgkin lymphoma biopsies and 42% (19/45) of Hodgkin lymphoma samples. RNAseq detected all but one of the translocations found by fluorescence in situ hybridization and PCR. RNAseq and whole exome sequencing identified additional genetic abnormalities not detected by conventional approaches. Finally, 3 cases are highlighted to exemplify how synergy between different diagnostic techniques and specialists can be achieved. This study demonstrates the feasibility and discusses the added value of integrating modern sequencing techniques and flow cytometry into a workflow for routine diagnostic workup of lymphoma. The inclusion of RNA and DNA sequencing not only supports diagnostics but also will lay the ground for the development of novel research-based treatment strategies for pediatric lymphoma patients.
摘要：
进行该研究是为了评估将最先进的测序技术和流式细胞术整合到小儿淋巴瘤的诊断检查中的可行性。RNA测序（RNAseq），在鉴别诊断中,将全外显子组测序(WES)和流式细胞术应用于患有淋巴瘤的儿科活检的常规诊断检查中.一年之内,我们分析了110例儿童(122份样本)的活检,因为怀疑是恶性淋巴瘤.结合组织学和免疫组织化学的标准化工作流程的经验，流式细胞术,报告了下一代测序技术。对83％（102/122）的标本中的新鲜组织进行流式细胞术，并可以快速诊断T细胞和B细胞非霍奇金淋巴瘤（NHL）。在所有NHL活检和42%(19/45)的霍奇金淋巴瘤(HL)样品中进行RNAseq。RNAseq检测到通过FISH和PCR发现的除一个以外的所有易位。RNAseq和WES鉴定了常规方法未检测到的其他遗传异常。最后,突出显示了三个案例，以举例说明如何实现不同诊断技术和专家之间的协同作用。这项研究证明了可行性，并讨论了将现代测序技术和流式细胞术整合到淋巴瘤常规诊断工作流程中的附加值。RNA和DNA测序的纳入不仅支持诊断，而且还将为儿童淋巴瘤患者开发新的基于研究的治疗策略奠定基础。
