Sarcoma is a rare disease affecting both bone and connective tissue and with over 100 pathologic entities, diagnosis can be difficult. Approximately one third of sarcoma patients however are characterised by recurrent genetic alterations mainly chromosomal translocations, which result in fusion genes, and gene amplifications. Fusion genes are highly useful diagnostic markers and are currently identified in the clinical setting using FISH/RT-PCR. Conversely, conclusive results cannot be achieved in up to 25% of fusion positive patients using these methods. Next generation sequencing (NGS) is a promising tool that can improve soft-tissue sarcoma diagnosis.
A novel sarcoma-specific NGS gene capture panel containing probes for 90 fusion genes and 7 genes with frequent copy number changes was designed, optimised and validated in order to improve fusion-positive sarcoma diagnosis. Genomic DNA was extracted from 92 well-characterised FFPE samples and libraries prepared using the KAPA Hyperplus kit (Roche) and hybridised using the SeqCap Hybridisation kit (Roche) and sequenced on a NextSeq platform (Illumina).
Sarcoma specific translocations or gene amplifications were identified in 92 out of 94 cases giving the targeted gene panel a sensitivity of approximately 98% and specificity of 100% (Table).Table:
1717P Sarcoma tumour types and number of cases with structural variants or copy number variations detected
|Soft Tissue Tumour Type||No. of cases||SV/CNV detected|
|Desmoplastic Small Round Cell Tumour (DSRCT)||2||2|
|Inflammatory Myofibroblastic Tumour||2||2|
|Clear Cell Sarcoma||4||4|
|Low-grade Fibromyxoid Sarcoma||3||3|
|Alveolar Soft Part Cell Sarcoma||1||1|
|Solitary Fibrous Tumour||2||2|
|Aneurysmal Bone Cyst||1||1|
|Extraskeletal Myxoid Chondrosarcoma||5||5|
|Dermatofibrosarcoma Protuberans (DFSP)||3||3|
To the best of our knowledge this is the first sarcoma-specific NGS panel validated in FFPE-derived genomic DNA. It is at least as sensitive and specific as the combination of FISH and RT-PCR and can be implemented in routine clinical diagnostic workflows. We are currently investigating whether fusion genes can be detected in plasma using ctDNA from 14 patients and results will be reported at the conference.
Clinical trial identification
Legal entity responsible for the study
David Gonzalez de Castro.
All authors have declared no conflicts of interest.