Oral Presentations - Pathology and Laboratory Medicine - University ...

ResidentAbstract # 10Tony NgTony Ng 1,2 , Matthew Robertson 1 , Hao Zhu3, William Einhorn 3 , George Daley 3 ,Poul Sorensen 1,21Department of Molecular Oncology, BC Cancer Research Centre, 2 Department of Pathologyand Laboratory Medicine, University of British Columbia, Vancouver, BC and 3 Children’sHospital Boston, Harvard Medical School, Boston, MAidentification and characterization of a novel TCF12-LIN28B fusion in sporadic Wilms’ tumorBackround/ObjectivesWilms’ tumor (WT) is the most common form of pediatric renal cancer; however, the genetics underlying thesporadic form of WT remain largely unknown. We recently characterized chromosome 6q21 breakpoints in twoindependent sporadic WT cases with t(6;15) translocations in an otherwise cytogenetically normal background. Inboth cases, we mapped the 6q21 breakpoints to an intergenic region between the HACE1 and LIN28B genes; thechromosome 15 breakpoint mapped to the TCF12 gene. LIN28B has been shown to have oncogenic propertiesthrough its ability to bind let-7 microRNAs (miRNAs) and reverse let-7 inhibition of multiple oncogenic pathwayssuch as Ras and c-Myc. Furthermore, 6q21 rearrangements in the region of the LIN28B locus may be recurrent inWT based on multiple case reports in WT. We hypothesized that these t(6;15) translocations lead to the generation ofa fusion transcript between TCF12 and LIN28B with oncogenic properties.MethodsRT-PCR and sequencing was used to identify a putative TCF12-LIN28B fusion in WT samples. This fusion wascloned and expressed in NIH/3T3 and HEK293T cell lines for biochemical studies, including assays of let-7 levels,levels of let-7 targets, and transformation assays.ResultsAnalysis of the index t(6;15) WT case by RT-PCR confirmed expression of a TCF12-LIN28B fusion transcript.Cloning and sequencing of the coding region of the full-length transcript showed fusion of exons 1-5 of TCF12to exons 2-4 of LIN28B. RT-PCR for the fusion in the second t(6;15) case also demonstrated expression of thesame fusion transcript, but was not detected in a small series of 10 other WT cases. When TCF12-LIN28B wasoverexpressed in NIH/3T3 cells, multiple let-7 miRNA family members were suppressed to a level similar to thatseen with wild-type LIN28B overexpression. To establish the transformation ability of this fusion, we used the HEK-293T human embryonic kidney cell line to serve as a more relevant WT model. Again, akin to that seen with wildtypeLIN28B, overexpression of TCF12-LIN28B in HEK-293T indeed promoted anchorage-independent growthin the soft agar colony assay. Furthermore, multiple let-7 targets are elevated in TCF12-LIN28B overexpressing cellscompared to control cells at the protein level, including K-Ras and c-Myc. Paradoxically, the mitogenic response ofboth TCF12-LIN28B and LIN28B overexpressing cells to serum stimulation are reduced, based on levels of pERKand pMEK, and pAkt to a lesser extent, as readouts of Ras pathway signalling.ConclusionWe identified a novel TCF12-LIN28B fusion gene in WT in the context of two cases bearing t(6;15) translocations.TCF12-LIN28B appears to possess oncogenic properties similar to wild-type LIN28B through their ability to modifylet-7 miRNA levels and their downstream targets. Further studies are needed to determine the incidence of this fusionin WT and other cancers, and to elucidate the mechanisms by which it promotes WT tumorigenesis.20 2 0 1 0 * O r a l P r e s e n t a t i o n s