Comparison of Droplet Digital PCR and Metagenomic Next-Generation Sequencing Methods for the Detection of Human Herpesvirus 6B Infection Using Cell-Free DNA from Patients Receiving CAR-T and Hematopoietic Stem Cell Transplantation.
Comparison of Droplet Digital PCR and Metagenomic Next-Generation Sequencing Methods for the Detection of Human Herpesvirus 6B Infection Using Cell-Free DNA from Patients Receiving CAR-T and Hematopoietic Stem Cell Transplantation.
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使用来自接受 CAR-T 和造血干细胞移植的患者的游离 DNA 检测人疱疹病毒 6B 感染的液滴数字 PCR 和宏基因组下一代测序方法的比较
DOI：
10.2147/idr.s379439
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发表时间：
2022
影响因子：
3.9
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中科院分区：
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本研究的目的是检查和比较液滴数字PCR（ddPCR）和宏基因组下一代测序（mNGS）在人类疱疹病毒6 B（HHV-6 B）检测中的差异。在接受嵌合抗原受体修饰的T细胞（CAR-T）治疗和造血干细胞移植（HSCT）的患者中，长期监测HHV-6 B病毒载量可用于识别免疫效应细胞相关神经毒性综合征（ICANS）并指导药物治疗。回顾性分析了27例同时具有mNGS和ddPCR检测结果的疑似HHV-6 B感染患者，其中19例接受了CAR T细胞治疗，8例接受了HSCT。设计HHV-6 B探针和引物，并评价ddPCR检测的性能。随后，使用血液和尿液进行ddPCR。收集临床信息和mNGS研究数据。ddPCR检测结果与mNGS检测结果呈显著相关（P < 0.001，R2 = 0.672）。在27份时间配对样本中，ddPCR显示20份样本HHV-6 B检测阳性，而单独mNGS显示12份样本HHV-6 B检测阳性。ddPCR在8个样本中检测到额外的HHV-6 B感染，如果仅使用mNGS，则会遗漏这些感染。此外，首次HHV-6 B感染事件在CAR T细胞输注后14天（中位数）检测到（范围为8至19天）。通过ddPCR对HHV-6 B进行纵向监测，以评估抗病毒治疗的有效性。数据显示，随着抗病毒治疗，HHV-6 B病毒载量逐渐下降。结果表明，ddPCR提高了HHV-6 B的阳性检出率，是mNGS方法的有效辅助手段。此外，在接受CAR T细胞治疗和HSCT的患者中纵向检测和定量HHV-6 B病毒载量可以作为药物治疗的指导。
The aim of this study was to examine and compare the differences between droplet digital PCR (ddPCR) and metagenomic next-generation sequencing (mNGS) in the detection of human herpesvirus 6B (HHV-6B). Long-term monitoring of HHV-6B viral load in patients receiving chimeric antigen receptor-modified T-cell (CAR-T) therapy and hematopoietic stem cell transplantation (HSCT) can be used to identify immune effector cell-associated neurotoxicity syndrome (ICANS) and guide drug therapy. Twenty-seven patients with suspected HHV-6B infection who had both mNGS and ddPCR test results were analyzed retrospectively, including 19 patients who received CAR T-cell therapy and 8 who received HSCT. The HHV-6B probe and primers were designed, and the performance of the ddPCR assay was evaluated. Subsequently, ddPCR was performed utilizing blood and urine. Data on clinical information and mNGS investigations were collected. The ddPCR test results correlated significantly with the mNGS test results (P < 0.001, R2 = 0.672). Of the 27 time-paired samples, ddPCR showed positive HHV-6B detection in 20 samples, while mNGS alone showed positive HHV-6B detection in 12 samples. ddPCR detected additional HHV-6B infections in 8 samples that would have been missed if only mNGS were used. In addition, the first HHV-6B infection event was detected at a median of 14 days after CAR T-cell infusion (range, 8 to 19 days). Longitudinal monitoring of HHV-6B by ddPCR was performed to assess the effectiveness of antiviral therapy. The data showed that with antiviral treatment HHV-6B viral load gradually decreased. Our results indicated that ddPCR improved the HHV-6B positive detection ratio and was an effective adjunct to mNGS methods. Furthermore, the longitudinal detection and quantification of HHV-6B viral load in patients undergoing CAR T-cell therapy and HSCT may serve as a guide for drug treatment.