Characterization of XAGE‐1b, a short major transcript of cancer/testis‐associated gene XAGE‐1, induced in melanoma metastasis

Abstract

Suppression subtractive hybridization, comparing mRNA expression profiles of common nevocellular nevi and melanoma metastases, was used to identify potential markers of melanoma progression. From the metastases we isolated XAGE-1b, a 470 bp transcript of the XAGE-1 gene. In general, expression of XAGE-1b was much more prominent than expression of the longer XAGE-1 transcript, isolated from Ewing's sarcoma. The XAGE-1b open-reading frame codes for a putative protein of 81 amino acids, harboring a functional bipartite nuclear localization signal and a C-terminal acidic transcription-activation-like domain. On the nucleotide level, XAGE-1b has a 50% homology with members of the GAGE family. However, homology between the corresponding proteins is weak. Expression of XAGE-1b in normal tissues was mainly restricted to testis, while placenta and brain were sporadically positive. In human tumor cell lines as well as in human tumor lesions, expression was most frequently found in melanocytic tumors and Ewing's sarcoma. In the different stages of melanocytic tumor progression, expression was exclusively seen in melanoma metastases (38%; n = 61), while all tested common and atypical nevi (n = 10) as well as primary melanomas (n = 8) were negative. Upregulation of expression after treatment with demethylating agent 5-aza-2'-deoxycytidine was detected in 1 of 4 human melanoma cell lines tested. The XAGE-1 gene consists of 4 exons and is located on chromosome Xp11.21-Xp11.22. After transfection into COS cells, the corresponding protein can direct the coupled fluorescent protein to the nucleus, showing a distinct speckled staining aspect. Our data imply the nuclear cancer/testis-associated XAGE-1b to be a marker for late melanocytic tumor progression.

XAGE‐1 mRNA Expression in Prostate Cancer and Antibody Response in Patients

“…NY-ESO-1 mRNA expression was correlated with tumor grade of transitional cell carcinoma (TCC), with the expression in 14/32 (44%) G3 TCCs, in contrast to 6/30 (20%) G1 and G2 TCCs (14). With regard to XAGE-1b, metastatic melanoma expressed the mRNA more frequently than did primary melanoma, suggesting its relation to the malignant potential in melanoma (22). In this study, we showed that stage D prostate cancer expressed XAGE-1b mRNA more frequently than stage C prostate cancer.…”

“…It was reported that XAGE-1 mRNA was expressed in Ewing's sarcoma, lung cancer and metastatic melanoma (15,22,23). Recent analysis using a cancer-profiling array showed that XAGE-1 mRNA was expressed in various types of cancer such as breast, lung and prostate cancer (7).…”

“…Four XAGE-1 transcript variants, XAGE-1a, b, c, and d, have been discovered and shown to be expressed in metastatic melanoma and Ewing's sarcoma (15,22,23). By recombinant expression cloning analysis (SEREX), we recently identified XAGE-1b as a dominant antigen recognized by serum from a lung adenocarcinoma patient using an autologous tumor cell line as a source of a cDNA library (1).…”

Activation of human cancer/testis antigen gene, XAGE‐1, in tumor cells is correlated with CpG island hypomethylation

“…Alternative splicing produces 4 different transcripts of XAGE-1 gene in Ewing's sarcoma. 4,5,8 In our study, we provide experimental evidence that the XAGE-1 promoter activity, which is induced by using the Sp1, is com- pletely blocked by heavy methylation of the XAGE-1 CpG island in cancer cell lines. The evidence of XAGE-1 expression, induced by hypomethylation, was provided by the re-expression of XAGE-1, after treatment with 5 0 -aza-deoxycytidine, in cancer cell lines not expressing XAGE-1.…”

“…3,4 Like most cancer/testis antigen family members, the XAGE-1 gene is located on the X-chromosome and is highly expressed in germ cells of the testis but not in other normal tissues. 5,6 Previous studies have shown that XAGE-1 is frequently expressed in Ewing's sarcomas and alveolar rhabdomyosarcomas, 7,8 suggesting that expression of this gene follows a pattern described for other cancer/testis antigen-associated genes.…”