The identification and characterisation of novel cancer-specific biomarkers
and/or targets is a key challenge for establishment of successful
diagnostic, prognostic and immunotherapeutic strategies. Cancer/testis (CT) genes are an attractive group of genes that encode proteins that are
restricted to the human testis and malignant tumours but are not expressed in healthysomatic cells. The testes are an immunologically privileged site;
therefore, CT genes represent important targets/biomarkers in the diagnosis
and treatment of cancers. Here, targeted genes identified from a literature search for meiosis-specific genes as well as from bioinformatics pipelines were analysed using RT-PCR in a range of normal and cancerous cell types.
Ten out of 24 genes were identified as promising CT genes as they were
expressed in cancer cells but not in normal tissues. A subsequent meta
-analysis of seven testis-restricted genes in a microarray data set also identified five genes that were up-regulated in clinical samples derived from patients. The use of hypomethylation (5-aza-2ˈ-deoxycytidine) and/or histone deacetylation (trichostatin; TSA) drugs further identified a new
class of CT genes with refractory transcriptional silencing. In contrast,
STRA8 and TDRD12 were transcriptionally regulated by DNA methylation and histone acetylation. Western blot analysis revealed the presence of PRDM9 protein in nearly all cancer cell lines; PRDM9 was localised in the nucleus in three cancer cell lines and in the cytoplasm in NT2 cells. Knockdown of
PRDM9 protein in HCT116 and SW480 cells reduced the survival of cancer cells, with a major effect seen with siRNA-7. This may indicate an oncogenic role for PRDM9 in cancer cells, this was explored further using an over expression system. Finally, PRDM7 and PRDM9 were overexpressed in
E. coli cells using a Glutathione S-transferase (GST) Gene Fusion System, but the proteins were foundmostly in the insoluble fraction and were difficult to purify.