Investigation of BRAF Mutations by Pyrosequencing in Lymphomas
Javier A Laurini, Patricia Aoun, Javeed Iqbal, John Chan, Timothy C Greiner. University of Nebraska Medical Center, Omaha, NE
Background: Recently the presence of the BRAF mutation V600E c.1799T>A has been described in 100% of cases of hairy cell leukemia (Tiacci et al;NEJM 2011), but not in 7 other common subtypes of B-cell lymphomas. That study utilized Sanger sequencing to identify the BRAF mutations. However, peripheral T-cell lymphomas (PTCL), nodal marginal zone lymphomas (NMZL), extranodal marginal zone lymphomas MALT type (MALT), and post transplant lymphoproliferative disorders (PTLD) have not been studied.
Design: We have designed a sensitive BRAF pyrosequencing assay for the V600E mutation that is applicable to DNA from fresh and paraffin-embedded tissue. The assay, which produces a 151bp product, has a sensitivity of 5% using the HT-29-cell line. DNA was extracted from rapidly frozen tissue on 34 cases of uncommon B and T-cell lymphomas. The B-cell cases were selected to have greater than 70% tumor cells by morphologic assessment. For PTCL, the cases were selected to have a clonal T-cell receptor gamma gene rearrangement that comprised greater than 70% of the electropherogram compared to background polyclonal T-cells. Cases of melanoma, colon carcinoma and hairy cell leukemia were used as positive controls for the assay.
Results: No BRAF V600E mutations were identified in 8 cases of monomorphous PTLD, 10 cases of PTCL, 8 cases of MALT, and 8 cases of NMZL.
Conclusions: The BRAF V600E mutation has not been identified in uncommon subtypes of B and T cell lymphoma, further suggesting that BRAF mutations are restricted to hairy cell leukemia.
Monday, March 19, 2012 1:00 PM
Poster Session II # 221, Monday Afternoon