| Authors | Amit Roshan, Fazlur R. Talukdar, Angela An, Emma-Jane Ditter, Ze Zhou, Paulius D. Mennea, Linfang Wu, Maria C. Neofytou, Anita Balakrishnan, Claire M. Connell, Gabriel Funingana, Giovanni Codacci-Pisanelli, Girma Mulisa, Tamrat Abebe, Constanza Linossi, Hui Zhao, Wendy N. Cooper, Huiqi Yang, Rajesh Jena, James D. Brenton, Pippa Corrie, Rebecca C. Fitzgerald, Simon Pacey, Richard Baird and Nitzan Rosenfeld |
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| Abstract | Background:
Proof-of-principle studies have identified circulating tumor DNA (ctDNA) in small volumes of whole blood, even dried blood spots (DBS) collected by finger-prick sampling from patients with advanced cancer. This minimally invasive sampling method offers ease, frequency and low cost of collection, simplified shipping and long-term room temperature stability. Adopting DBS for ctDNA collection and whole genome sequencing (WGS) analysis may support novel clinical trial design and increase real-world utility of liquid biopsies, especially where traditional phlebotomy is a barrier. Methods:
We collected sample quartets comprising serial DBS, and matching plasma, tumor, and buffy coat from ∼125 patients with multiple cancer types including oesophageal, lung, head & neck, melanoma, ovarian, colorectal and pancreatic cancer. Collections were in diverse settings at hospitals in the UK and Ethiopia, and by patients at home in the UK. DNA isolated from DBS was size-selected using bead-based enrichment to remove genomic DNA. Libraries were generated from DBS size-selected DNA, and DNA extracted from plasma, tumor and buffy coat samples had paired-end WGS at depth of ∼1X, 10X, and 50X respectively. Using bioinformatic pipelines for tumor-informed and tumor-agnostic detection, WGS data was analyzed for somatic copy number aberrations (SCNAs), fragment length, and tumor-specific single nucleotide variants (SNVs). Results:
We detected SNVs and SCNAs in DBS reflective of genomic changes in matching tumor DNA and plasma, from patients with Stage II-IV cancers. We observed concordant changes to the levels of those SNVs and SCNAs in plasma and DBS samples collected before and after systemic treatment compared with clinical and radiological measures. With tumor-informed analysis, utilizing patient-specific lists of 5, 930-51, 927 SNVs identified from tumor DNA, limit of detection for ctDNA was 34 ppm in plasma, and 95 ppm in DBS, comparable to commercially available assays for ctDNA. DBS collected in hospital by healthcare professionals, remotely in Ethiopia and at-home by patients in the UK with transportation at room temperature showed ctDNA can be extracted for analysis from diverse settings. Conclusion:
Our data highlights the promising role of DBS in advancing non-invasive cancer diagnostics and monitoring. The versatility of DBS in capturing ctDNA from various cancer types, coupled with its accessibility for collecting samples, is a significant step forward in blood-based cancer detection. This offers potential to enhance cancer monitoring and early detection strategies in both tumor-informed and tumor-agnostic settings. DBS is a cost-effective, user-friendly sampling strategy allowing at-home collection, enabling large-scale research and development programs needed to bring biomarker methods into widespread use. |
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