EPHB2 Alterations in Prostate Carcinoma: Evaluation of Mutational Status in a Large Series of Prostate Carcinoma Tissues and Cell Lines
A Andea, L Chen, Y Xiao, W Gerald. UAB, Birmingham, AL; MSKCC, New York, NY
Background: Ephrin receptor B2 (EPHB2) is part of the ephrin receptor family which is the largest member of the tyrosine kinase group. EPHB2 gene is located on 1p36.12 and is composed of 17 exons. Recent work has uncovered that, in prostate carcinoma (PCa), EPHB2 gene is the site of frequent mutations and that the gene locus is often deleted, suggesting a possible tumor suppressor role for this gene. In the current study we proposed to evaluate the presence of DNA alterations in EPHB2 gene in a large series of PCa cases, PCa cell lines and PCa xenografts.
Design: The study included a total of 69 cases of human PCa specimens, 6 PCa cell lines (CWR22, DU145, LnCap, LnCap104S, PC3 and Vcap) and 9 PCa xenografts (LAPC3, 4, 9, 14, 17, 18, 23, 25 and LuCap35). Fresh tissue was collected from radical prostatectomy specimens and metastatic samples and snap frozen in liquid nitrogen. DNA was extracted following microscopically controlled dissection of the frozen tissue blocks to enrich the tumor content. Sequence analysis of the coding and exon-flanking intronic regions of EPHB2 was performed using standard techniques (Agencourt Biosciences).
Results: A total number of 2856 sequence electropherograms were generated and analyzed. We identified 61 different point mutations in the EPHB2 gene. A majority of them involved introns (41) or untranslated regions (3). Five intronic mutations involved the intron/exon boundary region defined as occurring within 20 bp from the end of the exon. A total of 18 coding region mutations were found (11 silent, 6 missense and 1 nonsense). The missense mutations involved the ephrin receptor ligand binding domain (c.63A>T/ p.20Glu>Asp -1 case), the fibronectin type III domain (c.1081A>G/ p361Ile>Val -1 case and c.1273G>A/ p.425Ala>Thr -1 case) and the tyrosine kinase domain (c.1952T>C/ p.651Val>Ala -1 case, c.2035G>A/ p.679Asp>Asn -2 cases and c.2623G>A/ p.875Gln>Ser -1 case). The nonsense mutation involved the tyrosine kinase domain of DU145 cell line (c.2623G>A/ p.723Gln>X). Out of the 7 non-silent mutations, 3 were reported previously and 4 represent novel mutations of the EPHB2 gene (c.63A>T, c.1081A>G, c.1273G>A and c.2623G>A).
Conclusions: Our study successfully identified 7 non-silent mutations of the EPHB2 tyrosine kinase gene occurring in prostate cancer, 4 of which were not described previously. Functional studies to determine the biological significance of these mutations are needed in order to assess the impact of EPHB2 in prostate carcinogenesis.
Category: Genitourinary (including renal tumors)
Wednesday, March 11, 2009 9:30 AM
Poster Session V # 96, Wednesday Morning