Diagnostic Utility of a Novel ETV6/NTRK3 RT-PCR Assay and Break-Apart ETV6 FISH Analysis in the Diagnosis of Congenital Fibrosarcoma and Cellular/Mixed Congenital Mesoblastic Nephroma
A Igbokwe, AJ Lazar, WL Wang, A Folpe, V Hernandez, J Hicks, D Lopez-Terrada. Baylor College of Medicine, Houston, TX; UT-MD Anderson Cancer Center, Houston, TX; Mayo Clinic, Rochester, MN
Background: Congenital Fibrosarcoma (CFS) and Cellular Congenital Mesoblastic Nephroma (CMN) are characterized by ETV6/NTRK3 gene fusions [t(12;15)(p13;q25)]. Both neoplasms occur chiefly in children; CFS (<2 yrs) and CMN (<6 months). As a diagnostic aid, and to determine prevalence of these translocations in CFS and cellular/mixed CMN, we developed novel RT-PCR and FISH-based approaches. This strategy is useful to: (1) test formalin-fixed, paraffin-embedded (FFPE) tissue specimens, (2) detect transcript variants in specimens with minute/poor quality RNA or small biopsies and (3) identify potential variant translocations or cryptic rearrangements of ETV6.
Design: A RT-PCR assay to detect ETV6-NTRK3 transcripts (variants 1/2) was designed and tested on morphologically classic cases of CFS (12) and cellular/mixed CMN (12) from patients aged 4 days-17 months and 3 differentials (1 lipofibromatosis, 1 fibrous hamartoma of infancy, 1 desmoid tumor). Unstained FFPE sections from 10 of 12 CFS and all 12 CMN samples were tested by commercial break-apart FISH ETV6 probe.
Results: An ETV6-NTRK3 transcript was detected in 6/12 CFS (50%) and 2/12 CMN (17%) including 1 cellular CMN and 1 mixed CMN. Primers designed to detect a theoretical ETV6-NTRK3 variant 3 transcript amplified a novel sequence with homology to ETV6 and truncated NTRK3 (1 case). 3' RACE failed to amplify cases negative for the ETV6-NTRK3 variants 1/2 or reveal new ETV6 fusion partners. The 3 differentials for CFS/CMN were negative for the fusion transcripts. FISH analysis (100 nuclei count) showed 100% correlation with RT-PCR results in cases with utilizable RNA and was also positive in 1 case with degraded RNA (non-diagnostic RT-PCR).
Conclusions: RT-PCR detection of ETV6-NTRK3 transcripts and ETV6 FISH are reliable diagnostic adjuncts for CFS and cellular or mixed CMN. Confirmatory ETV6 FISH analysis is valuable in cases with degraded or non-diagnostic RNA. This data suggests that the ETV6/NTRK3 transcript is more prevalent in CFS than cellular/mixed CMN. Since the ETV6 rearrangement and ETV6-NTRK3 fusion was not detected in a subset of cases, cryptic/variant translocations, alternative fusion partners or completely novel molecular derangements are relevant considerations, and are subjects of on-going investigation in our laboratory.
Category: Bone & Soft Tissue
Tuesday, March 23, 2010 9:30 AM
Poster Session III # 1, Tuesday Morning