CTC and ctDNA in MRD Testing

By Priyanka Varma


Nov. 3rd, 2022

Minimal Residual Disease (MRD) testing can be done by detecting circulating tumor cells (CTC) and circulating tumor DNA (ctDNA).

CTCs have prognostic and predictive capabilities in monitoring early stages of cancer. Since the concentration of CTCs is extremely low (1 CTC in the background of millions to billions of blood cells), the methods employed to detect them are required to be highly sensitive. The current methodologies employ either physical attributes or expression of cell-surface based markers [1]. CTCs are also believed to attribute to early dissemination of the cancer [2].  CTC detection is done by utilizing either enrichment method like detection of immunophenotypic characteristics of circulating cells, like Epithelial Cell Adhesion Molecule (EpCAM) and cytokeratin (with absence of leukocyte markers)that mark the origin of the tumor, or by cell size or density [3, 4]. EpCAM is also called the “universal” epithelial marker of cancers. Its expression varies among cancer types and is shown to have strong expression in Breast and Prostate cancers, along with pancreatic, colorectal, and hepatocellular cancers. EpCAM positive CTCs predict early distant metastasis and overall, a poorer survival rate and prognosis. The drawback of using EpCAM as a biomarker is that it cannot detect cancers which are EpCAM negative like neurogenic cancers or cancers where CTCs are undergoing epithelial-mesenchymal transition (EMT) as EpCAM is downregulated during EMT [5].

ctDNA has also been recently employed to understand the tumor dynamics, detect MRD, assess the response/ resistance to therapy and monitor for any recurrence. ctDNA is secreted from either apoptotic tumor cells or from living cancer cells. Digital droplet PCR, Multiplex PCR, Next Generation Sequencing (NGS), Mass-Spectrometry assays have been used for detecting ctDNA [1, 2, 3]. There are two approaches to detect ctDNA for MRD testing, “Targeted” and “Untargeted”. Both approaches have their own pros and cons. While targeted MRD testing by NGS like Q-PCR, BEAMing, Safe-SeqS, CAPP-Seq, and TAmSeq can detect known tumor-specific mutations with fast, cost effective and accurate (high sensitivity and specificity) results, the underlying disadvantage is that knowledge of the entire genome is required as a prerequisite. On the other hand, untargeted testing by detecting Copy NumberAberrations (CNA) or point mutations by Whole Genome Sequencing (WGS), or Whole Exome Sequencing (WES) can identify new mutations without requiring a prior knowledge of the whole genome. However, untargeted testing needs a larger concentration of ctDNA and has a low sensitivity (5-10%) of detection [2].


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2.      Elazezy M, Joosse SA. Techniques of using circulating tumor DNA as a liquid biopsy component in cancer management. Comput Struct Biotechnol J. 2018 Oct 9;16:370-378. doi: 10.1016/j.csbj.2018.10.002.PMID: 30364656; PMCID: PMC6197739.

3.      Murray NP. Biomarkers detecting minimal residual disease in solid tumors: what do they mean in the clinical management of patients? Biomark Med. 2019 Dec;13(18):1535-1538. doi: 10.2217/bmm-2019-0401.Epub 2019 Oct 21. PMID: 31631673.

4.       Cescon DW, Kalinsky K, Parsons HA, Smith KL, Spears PA, Thomas A, Zhao F, DeMichele A. Therapeutic Targeting of Minimal Residual Disease to Prevent Late Recurrence in Hormone-Receptor Positive Breast Cancer: Challenges and New Approaches. Front Oncol. 2022 Feb10;11:667397. doi: 10.3389/fonc.2021.667397. PMID: 35223447; PMCID: PMC8867255.

5.      Lin D, Shen L, Luo M, Zhang K, Li J, Yang Q, Zhu F, Zhou D, Zheng S, Chen Y, Zhou J. Circulating tumor cells: biology and clinical significance. Signal Transduct Target Ther. 2021 Nov 22;6(1):404. doi:10.1038/s41392-021-00817-8. PMID: 34803167; PMCID: PMC8606574.

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