Abstract LB-290: Innovative targeted methylation sequencing detects tumor signal as low as 0.003% in early-stage cancer plasma
| Autor: | Bin Zheng, Jianmin Wang, Shengrong Lin, Yun Bao, Grace Q. Zhao, Heng Wang |
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| Rok vydání: | 2020 |
| Předmět: | |
| Zdroj: | Cancer Research. 80:LB-290 |
| ISSN: | 1538-7445 0008-5472 |
| DOI: | 10.1158/1538-7445.am2020-lb-290 |
| Popis: | Methylation analysis in cell-free DNA holds great potential for early cancer detection. In the plasma of early stage cancer patient, the tumor content is estimated to be less than 0.1%, often down to 0.01% or lower, therefore demands a highly sensitive assay. Currently there are two major approaches used for cancer screening: the global approach, including WGBS/RRBS or affinity-based enrichment, and large targeted panels containing 10,000 or more of potential methylation markers. Targeted Methylation Sequencing (TMS) provides the most sensitive and specific analysis of methylation markers. However, the sensitivity and specificity of conventional TMS is compromised by low efficiency and low recovery of target enrichment, and further hampered by background noise associated with large panels. The ideal solution would be an in-depth analysis using a focused small cancer-specific methylation biomarker panel, but is not supported by existing technologies. Here we present a new technology designed for TMS analysis in cfDNA: Point-n-Seq, featuring an enrichment of target molecules directly from cfDNA before bisulfite conversion and amplification. Particularly, this technology enables small focused panel that interrogates the methylation status of 1 to ~1000 markers. We designed a CRC panel covering 100 methylation markers in 3 steps: identify ~1000 CRC-specific markers from public databases; eliminate makers with high background signal in baseline cfDNA of healthy population; finalize the list with the most differentiating markers between patient and healthy cfDNA. The capture of Point-n-Seq CRC panel is highly efficient resulting in high uniformity (94% > 0.5X, 100% >0.2X) and on-target rate (>80%). For 20ng cfDNA input, more than 1000 deduped informative reads were obtained for each marker on average, despite the high GC content (> 80%). The output of informative reads was linear to the cfDNA input ranging from 1ng to 40ng. In titration studies, 0.6pg (0.2X genome equivalent) methylated DNA in 20ng cfDNA (0.003%) was reliably detected over cfDNA background. In a pilot clinical study using plasma samples from patients with colorectal adenocarcinoma - early stage (I, n=7; II, n=7), late stage (III, n=11; IV, n=3), and control individuals (n=105), the average fractions of methylated signal are 0.0034%, 0.013%, 0.09%, 0.17%, 0.29% for control, stage I, II, III, IV accordingly. The methylation fraction of stage I samples is significantly different from the control group (P Citation Format: Grace Zhao, Yun Bao, Heng Wang, Bin Zheng, Jianmin Wang, Shengrong Lin. Innovative targeted methylation sequencing detects tumor signal as low as 0.003% in early-stage cancer plasma [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr LB-290. |
| Databáze: | OpenAIRE |
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