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Fig. 2 | Molecular Cancer

Fig. 2

From: A clinician’s handbook for using ctDNA throughout the patient journey

Fig. 2

Choosing the right ctDNA assay based on sensitivity and breadth of genome coverage. A The number of detectable alterations critically depends on the selected cfDNA assay. The first row illustrates a DNA segment with various alterations (explained in the bottom legend). The second row (untargeted profiling) indicates the use of an “off the shelf” panel, which is capable of identifying a number of alterations, but as it represents a rather general assay, it may miss a considerable number of alterations (indicated by empty symbols). The third row (targeted profiling) indicates the use of a panel tailored for a specific tumor entity. For example, after screening of databases such as COSMIC and TCGA, panels can be designed that will identify specific alterations for this particular tumor entity with high likelihood. However, mutations “private”, i.e. unique, to the patient’s tumor will be missed. The fourth row (targeted, personalized profiling) indicates the use of a patient-specific multiplex assay, which was individually designed based on sequencing information from the primary tumor. In theory, all mutations from the primary tumor are detectable; however, new alterations that may have occurred at a later timepoint will be missed. The fifth row shows the use of whole-exome sequencing (WES) or whole-genome sequencing (WGS), which enables comprehensive coverage of all coding regions of the genome and, in case of WGS, also of all non-coding regions. The actual capability of detecting variants does not solely depend on the selected assay but also on other factors such as the ctDNA levels. B For a personalized approach, the tumor or a baseline plasma sample needs to be sequenced first. The observed mutations can then be leveraged for subsequent cfDNA analyses. The triangle in the center indicates the various breadth of such analyses. Advantages of analyzing only a single locus include low costs and easy interpretation without the necessity of sophisticated bioinformatics. However, sensitivity is limited, as sampling issues represent a significant confounding factor. In contrast, analyses of hundreds or thousands of targets requires some error-suppression means, i.e., bioinformatics tools. At the same time, the likelihood for the detection of evidence for the presence of ctDNA increases tremendously, making such approaches the most sensitive for MRD detection. In fact, while sequencing depth remains a critical factor for ctDNA detection, sequencing breadth may supplant the importance of high coverage analyses. C Some clinics may have access to their own academic or partner laboratory, which may develop and apply its own tests and address liquid biopsy related research questions. Alternatively, samples can be sent to a commercial end-to-end provider. Regardless which laboratory conducts the analyses, the aim is to provide the MTB with all relevant information at hand so that the best decisions can be made for patients

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