Could a blood test offer clues to tumor-specific transcriptional activity?
Scientists from IMI’s CANCER-ID http://www.cancer-id.eu/ project have obtained unprecedented levels of information on genetic activity in cancerous tumors by analyzing circulating tumor DNA (ctDNA) taken from blood samples. Tumors may shed fragments of genetic material such as DNA into the blood and many research efforts have been made to develop the best ways of capturing and analyzing this material. The study, http://www.nature.com/ng/journal/vaop/ncurrent/full/ng.3648.html, published in Nature Genetics http://www.nature.com/ng/index.html last week, adds to our understanding of the genetics of cancer and may provide valuable information for patient stratification from minimally invasive liquid biopsies.
Previous methods for the analysis of ctDNA only allowed the detection of somatic alterations present in the tumor genome, such as mutations or copy number alterations. The data presented by Ulz et al. from Michael Speicher’s group at the Institute of Human Genetics at the Medical University of Graz (Austria, https://www.medunigraz.at/en/) also provides functional information about the tumor from the peripheral blood, i.e. which genes are expressed and which genes are not expressed. The approach yields additional information about a tumor genome, as it is now possible to monitor the expression status of genes in regions with copy number gains in tumors. Some of these genes are tumor driver genes and the knowledge that these genes are indeed expressed facilitates the identification of altered pathways in tumor cells.
The reduced nucleosome occupancy in the vicinity of the transcriptional start site (TSS) of expressed genes results in a significantly lower sequencing depth of ctDNA fragments (cf. Figure). Genes that are more abundantly expressed in tumors can therefore be monitored based on the relative representation of their TSS in ctDNA fragments. This can be used to monitor drug target expression, potentially signaling pathway activation and response to treatments as monitored by liquid biopsy, reducing the need for traditional tissue biopsies that may be difficult to impossible to access. Many targeted therapy strategies are based on interfering deregulated pathways in tumor cells, such as the EGFR/HER2-AKT/MAPK pathways. The newly reported findings facilitate the identification of disturbed pathways and may therefore improve the clinical management of patients with cancer and furthermore contribute to the development of new cancer treatments.
In addition, information about the predominantly expressed isoforms can be gathered using this new methodology. Such information may be useful for the identification of isoforms predominantly expressed in cancer, which may further contribute to the development of targeted therapies.
Relevance to CANCER-ID
One of the tumor indications investigated in the context of the IMI CANCER-ID consortium (www.cancer-id.eu) is treatment refractory HER2-positive metastatic breast cancer. In this tumor entity, focal high-level amplifications, such as the HER2 amplicon on chromosome 17q12, are of particular importance. It will now be possible to monitor the expression of the genes in these amplicons and to analyze their role in the development of therapy resistance. Our approach will also be of importance for the large indication in the focus of CANCER-ID, non-small cell lung carcinomas, as it may provide novel insights into recurrently gained genomic regions.
Read the Medical University of Graz’s press release https://www.medunigraz.at/fileadmin/medunigraz/oeffentlichkeitsarbeit/presse/2016/presse_info_ctDNA.pdf (in German)
The sequencing coverage of plasma DNA at promoter sites reflects nucleosome organization at transcription start sites (TSS; in the image at position 0, find further information in the article). In expressed genes, nucleosomes are removed at the start of transcription to create an NDR (nucleosome-depleted region) over the promoter, allowing transcription factors to bind. The reduction in nucleosome occupancy for highly expressed genes corresponded to decreased coverage at the TSS, which is flanked by wave-like patterns of plasma DNA coverage indicating the strong positioning of adjacent nucleosomes. This plasma DNA read depth pattern even reflects differently expressed genes as indicated by FPKMs (fragments per kilobase of mature transcript per million mapped reads) as the oscillating periodicity upstream and downstream of the TSS decreases in less expressed genes, reflecting the denser nucleosome packing (x axis, distance from the TSS; y axis, relative coverage reflecting nucleosome dyads).