Formalin-Fixed, Paraffin-Embedded-Targeted Locus Capture A Next-Generation Sequencing Technology for Accurate DNA-Based Gene Fusion Detection in Bone and Soft Tissue Tumors

Chromosomal rearrangements are important drivers in cancer, and their robust detection is essential for diagnosis, prognosis, and treatment selection, particularly for bone and soft tissue tumors. Current diagnostic methods are hindered by limitations, including difficulties with multiplexing targets and poor quality of RNA. A novel targeted DNA-based next-generation sequencing method, formalin-fixed, paraffinembedded-targeted locus capture (FFPE-TLC), has shown advantages over current diagnostic methods when applied on FFPE lymphomas, including the ability to detect novel rearrangements. We evaluated the utility of FFPE-TLC in bone and soft tissue tumor diagnostics. FFPE-TLC sequencing was successfully applied on noncalcified and decalcified FFPE samples (n = 44) and control samples (n = 19). In total, 58 rearrangements were identified in 40 FFPE tumor samples, including three previously negative samples, and none was identified in the FFPE control samples. In all five discordant cases, FFPE-TLC could identify gene fusions where other methods had failed due to either detection limits or poor sample quality. FFPE-TLC achieved a high specificity and sensitivity (no false positives and negatives). These results indicate that FFPE-TLC is applicable in cancer diagnostics to simultaneously analyze many genes for their involvement in gene fusions. Similar to the observation in lymphomas, FFPE-TLC is a good DNA-based alternative to the conventional methods for detection of rearrangements in bone and soft tissue tumors. (J Mol Diagn 2023, 25: 758-770; https://doi.org/10.1016/j.jmoldx.2023.06.012)

Automated summary

Chromosomal rearrangements play a crucial role in cancer development, and their accurate detection is important for the diagnosis, prognosis, and treatment selection of bone and soft tissue tumors. Current diagnostic methods are limited by challenges in amplifying multiple targets and poor RNA quality. This study evaluated a novel DNA-based sequencing method called FFPE-TLC and found that it showed high specificity and sensitivity in identifying gene fusions in bone and soft tissue tumors, even in cases where other methods failed due to detection limits or sample quality issues.