The Genetic And Cell Free Dna Characteristics Of Soft Tissue Sarcomas

Soft tissue sarcomas (STSs) are a group of rare, malignant tumours with a relatively poor prognosis. Although they have been broadly classified as complex or simple based on their karyotype, only a few studies have investigated the genetic characteristics of STSs in detail. To further explore STS genomics we analysed a group of primary STSs using Illumina Next Generation Sequencing. This analysis revealed several characteristics of the analysed tumours including 1) a low single nucleotide variant and insertion/deletion mutational burden, 2) a high number of recurrent amplifications/deletions, 3) significant inter-tumoural heterogeneity regardless of histopathological classification and 4) complex genotypes in the vast majority of STSs analysed. Currently no circulating STS biomarkers exist. Short fragments of DNA termed cell free DNA (cfDNA) can be found in the bloodstream. Some cancer patient cfDNA is tumour derived (circulating tumour derived DNA / ctDNA) and in several malignancies this ctDNA appears to correlate with disease behaviour. Despite this, very few studies have investigated STS patient cfDNA/ctDNA. To address this paucity of work we next used quantitative PCR, semiconductor targeted NGS and digital droplet PCR to characterise the cfDNA/ctDNA characteristics of two groups of metastatic and non-metastatic STS patients. This analysis revealed elevated cfDNA levels in the metastatic patients, which weakly correlated with disease burden suggesting a potential diagnostic role. Overall ctDNA was also identified in 27% (non-metastatic) - 36% (metastatic) of the analysed patients suggesting either that 1) the experimental approach used was not specific enough to detect ctDNA in tumours as genetically heterogeneous as STSs, or that 2) not all STSs shed ctDNA. Moving forwards although this analysis highlights a potential role for the use of ctDNA profiling in STS patients, it has also identified several significant challenges that must be addressed before ctDNA can be proposed as a realistic source for novel biomarkers

Circulating tumour-derived DNA in metastatic soft tissue sarcoma.

Following treatment 40% of soft tissue sarcoma (STS) patients suffer disease recurrence. In certain cancers circulating cell free DNA (cfDNA) and circulating tumour-derived DNA (ctDNA) characteristics correlate closely with disease burden, making them exciting potential sources of biomarkers. Despite this, the circulating nucleic acid characteristics of only 2 STS patients have been reported to date. To address this we used an Ion AmpliSeq™ panel custom specifically designed for STS patients to conduct a genetic characterisation of plasma cfDNA, buffy coat (germline) DNA and where available Formalin-Fixed Paraffin-Embedded (FFPE) primary STS tissue DNA in a cohort of 11 metastatic STS patients. We found that total cfDNA levels were significantly elevated in the STS patients analysed, and weakly correlated with disease burden. Using our Ion AmpliSeq™ panel we also successfully detected ctDNA in 4/11 (36%) patients analysed with a wide variety of STS subtypes and disease burdens. This evidence included the presence of cancer associated TP53 / PIK3CA mutations in 2 patients' plasma and matched primary STS tumour tissue, and in the plasma alone for 2 patients. We also identified 2 potential examples of allelic loss of heterozygosity in an additional patient's STS DNA and cfDNA. This is the largest study performed characterising STS patient cfDNA/ctDNA and confirms that the field remains an attractive potential source of novel STS biomarkers. Further work is required to investigate the circulating nucleic acid characteristics of individual STS subtypes, and the potential prognostic or therapeutic roles that cfDNA/ctDNA may hold for patients with these complex tumours