Mlh1 And Braf Status Analysis In Metastatic Germ Cell Tumors

e15092 Background: Germ cell tumors (GCTs) are the most common malignancy in young men. Recently have been proposed Mismatch repair (MMR) deficiency, microsatellite instability (MSI) and BRAF mutations to be related with cisplatin (CDDP) resistance in nonseminomatous (NSE) GCTs. On the other hand, we wanted to use our orthotopic nude mice GCTs implantation model for microsatellite instability (MSI) study. Methods: A retrospective cohort of 75 male patients with metastatic germ cell tumors treated at the Catalan Cancer Institute from August 1989 to November 2004 was chosen to generate a tissue microarray. We identified he presence of the MMR factor MLH1 by immunohistochemistry (IHC) using the G168-728 antibody. We performed a double check for BRAF mutations in paraffin embedded tissue using restriction fragment length polymorphism (RFLP) with the Spel restriction enzyme; and single strand conformation polymorphism (SSCP) analyses. SK-Mel28 and RKO cell lines were used as positive controls for BRAF mutations in homozygosis and heterozygosis respectively. We have generated resitance to CDDP in vivo in five tumors by exposition to repetitive cycles (1 yolk-sac; 1 choriocarcinoma; 2 embrional carcinoma; 1 mix tumor). Initial tumors and their respective CDDP resistant induced tumors were checked for MSI using the Promega commercial kit that include BAT26, BAT25, NR21, NR24 and MONO27 markers. Results: Median age was 28 years (16-56). 16% were seminoma and 84% NSE. One third of patients were refractory to first line CDDP based chemotherapy. All samples stained positive for MLH1 and we did not identify any BRAF mutation in sensitive patients neither in refractory ones. None of the xenografted tumor did show MSI in the locus analyzed in the original and the CDDP refractory in vivo induced tumors. Conclusions: In our series MLH1 inactivation and BRAF mutations do not seems to be involved in CDDP resistance. We are analyzing MSH2 and MSH6 expression by IHC as well as checking refractory tumors for BRAF mutations with a third technique using a multiplex SNaPshot. No significant financial relationships to disclose.