Diagnostic mutation testing in situ in routine FFPE tissue sections for treatment prediction in clinical oncology
(English)Manuscript (preprint) (Other academic)
Activating mutations in the KRAS gene are present in different cancer types and are strongly associated with resistance to epidermal growth factor receptor (EGFR) inhibitor therapy. Hence there is a requirement for sensitive KRAS mutation analysis to determine the most suitable treatment for the patients. Also, little is known about the impact of tumor heterogeneity with regard to KRAS mutation status in different sub-clones during tumorigenesis, and if this is important for treatment response and prognosis. To improve the diagnostic accuracy, we developed an RNA-based genotyping assay that targets KRAS-mutations in codon 12 and 13 in situ on tissue samples by the use of multiple mutation specific padlock probes and rolling-circle amplification. Thus, the distribution of wild-type (green rolling-circle products) and mutated (red rolling-circle products) KRAS alleles can be determined for single cancer cells in different parts of a heterogeneous tumor without the use of microdissection. We demonstrate reliable detection of KRAS point mutations on cytologic tumor imprints as well as on fresh frozen and formalin-fixed paraffin-embedded tissue sections from colorectal and lung cancer. This in situ method offers single cell mutation detection for diagnostics and holds great promise as a tool to investigate the role of oncogenic mutations in complex tumor tissues.
mutation, KRAS, padlock probes, rolling circle amplification, in situ, FFPE, touch imprints, cancer, diagnostics
Medical Genetics Medical Genetics Cancer and Oncology
Research subject Molecular Medicine; Oncology
IdentifiersURN: urn:nbn:se:uu:diva-149822OAI: oai:DiVA.org:uu-149822DiVA: diva2:405867