Further characterization of the first seminoma cell line TCam‐2

Abstract

Testicular germ cell tumors of adolescents and adults (TGCTs) can be classified into seminomatous and nonseminomatous tumors. Various nonseminomatous cell lines, predominantly embryonal carcinoma, have been established and proven to be valuable for pathobiological and clinical studies. So far, no cell lines have been derived from seminoma which constitutes more than 50% of invasive TGCTs. Such a cell line is essential for experimental investigation of biological characteristics of the cell of origin of TGCTs, i.e., carcinoma in situ of the testis, which shows characteristics of a seminoma cell. Before a cell line can be used as model, it must be verified regarding its origin and characteristics. Therefore, a multidisciplinary approach was undertaken on TCam-2 cells, supposedly the first seminoma cell line. Fluorescence in situ hybridization, array comparative genomic hybridization, and spectral karyotyping demonstrated an aneuploid DNA content, with gain of 12p, characteristic for TGCTs. Genome wide mRNA and microRNA expression profiling supported the seminoma origin, in line with the biallelic expression of imprinted genes IGF2/H19 and associated demethylation of the imprinting control region. Moreover, the presence of specific markers, demonstrated by immunohistochemistry, including (wild type) KIT, stem cell factor, placental alkaline phosphatase, OCT3/4 (also demonstrated by a specific Q-PCR) and NANOG, and the absence of CD30, SSX2-4, and SOX2, confirms that TCam-2 is a seminoma cell line. Although mutations in oncogenes and tumor suppressor genes are rather rare in TGCTs, TCam-2 had a mutated BRAF gene (V600E), which likely explains the fact that these cells could be propagated in vitro. In conclusion, TCam-2 is the first well-characterized seminoma-derived cell line, with an exceptional mutation, rarely found in TGCTs.

GPR30 is overexpressed in post-puberal testicular germ cell tumors

“…GPR30 expression has also been analyzed in GC1 and TCam-2 cell lines, respectively derived from immortalized type B murine spermatogonia and human seminoma. [19][20][21] As shown in Figure 3A and B the GPR30 protein was detected in the cytoplasm and along plasma membrane of GC1 cells, with a diffuse expression, whereas in the seminoma cell line TCam-2 the protein was confined in specific compartments, probably into the Golgi (Fig. 3C and D).…”

NANOG promoter methylation and expression correlation during normal and malignant human germ cell development

“…Alternatively, the data for JKT-1, together with experiments from us and others, further indicate that JKT-1 does not (or no longer) display seminoma-like characteristics. 25,26 To address the question whether NANOG expression correlates to methylation of the CpGs in the NRR in GCTs and derived cell lines, we performed qRT-PCR analysis. NANOG expression levels were high in seminomas and embryonal carcinomas, while low and absent in teratomas, yolk sac tumors, choriocarcinomas and mixed non-seminomas (Fig.…”

A PCR/restriction digestion assay for the detection of the transcript variants 1 and 2 of POU5F1

“…Thus, modification of the original protocol (Liedtke et al, 2007) may be required. Another RT-PCR methodology for transcript variant 1 of POU5F1 has recently been reported (de Jong et al, 2008). The forward primer was placed in the beginning of the sequence with accession number NM_002701 and the reverse at the junction of exons 1 and 2 close to that used by Liedtke et al (2007).…”