Cancer/Testis Antigen Profiling in Multiple Myeloma Define a Cohort of Patients with Poor Prognosis Regardless of Genetic Subtypes.

Cancer/testis antigens (CTAs), comprising the MAGE, CTAG, SSX and GAGE families, are expressed in different tumor types and restricted in normal tissues to germ cells of the testis. Many CTAs contain epitopes recognized by cytotoxic T cells and are potential targets for active immunotherapy. In MM, different CTAs have been shown to be expressed in cell lines and patient cells, and are associated with more proliferative and advance stage tumors. However, correlation with survival has never been reported. We undertook a global survey of CTA expression using gene expression profiling of CD138-enriched plasma-cell (PC) RNA in 15 normal, 101 MM, 22 MGUS and 24 SMM samples using the Affymetrix U133A chip (Affymetrix, Santa Clara, CA). 22 CTA genes were differentially expressed across the samples and unsupervised clustering using these genes clearly identifies a group of patients with high CTA expression. Most CTAs have significantly higher expression in MM as compared to SMM and especially MGUS, which has similar expression as normal PCs. As these antigens are not expressed in normal PCs, an upper limit of normal expression (ULN) was derived based on mean+3SD of expression in normal PCs for each CTA. A strict cut-off of normalized expression value greater than 2xULN was used to assign positive expression for each CTA genes. Using this method, the most commonly expressed CTAs are MAGE-A3 (73% of MM), MAGE-C1 (45%), MAGE-A6 (35%), SSX-1 (21%), GAGE-5 (18%) and NY-ESO-1 (14%). We then assigned patients with expression of more than 2 of these genes as having a CTA signature and correlated with clinical parameters. 45 patents (41 MM and 4 SMM) have the CTA signature. These patients have a significantly higher PCLI (median 0.2 v 0.7, p=0.0003). In fact, there is a significant correlation between the number of positive CTAs and PCLI (r=0.59, p<0.0001). Interestingly, there is no association between patients with a CTA signature and any of the recurrent genetic abnormalities (IgH translocations, chromosome 13 deletion and TC classes), yet MM patients with this signature have a significantly shorter survival (median 459 days vs not reached, log-rank p=0.017). A prognostic hierarchy exists amongst these CTAs. Only MAGE-A6, MAGE-A12, SSX-1 and SSX-5 expression has significant prognostic implications with the SSX family conferring the worse prognosis. Using a restricted panel of 3 CTAs (SSX-1, -5 and MAGE-A12), about 28% of MM patients expressing one or more of these genes can be identified with poor survival (median less than 1 year) that is independent of genetic subtypes. The genes differentially expressed between patients with and without the CTA signature are mainly involved in cell cycle regulation. Since the expression of this large set of CTAs appears to be under epigenetic control, we speculate that the same epigenetic processes that lead to dysregulation of the cell cycle also cause ectopic expression of CTA genes. As CTA expression is rare in MGUS, epigenetic processes may therefore also mediate transformation from MGUS to MM. These poor prognosis patients represent good candidates for novel therapies targeting the epigenetic machinery.