Differential Expression of the MT1 E Gene in Estrogen-Receptor-Positive and-Negative Human Breast Cancer Cell Lines

The goal of this study was to determine which of the 10 functional metallothionein (MT) genes are expressed in four human breast cancer celi lines and whether expression varies among the cell lines. Using reverse transcription polymerase chain reaction (RTPCR) technology, it was shown that there was no expression of mRNA for the MT-1A, MT-1B, MT-1F, MT-1G, MT-1H, MT-3, and MT-4 genes in any of the four cell lines. All four cell lines were shown to express mRNA for the MT-2A and MT-1X genes. The expression level ofmRNA for the MT-2A gene demonstrated modest differences among the cell lines, whereas expression of the MT-1X gene was consistent. In contrast, mRNA for the MT-1E gene was expressed in only two of the four celi lines and expression correlated to the estrogen receptor status of the cell lines. The two estrogen-receptor-positive cell lines showed no mRNA expression for the MT-1E gene. In the two estrogen-receptor-negative cell lines, mRNA expression for the MT-1E gene was elevated with expression levels similar to the housekeeping gene, glyceraldehyde-3-phosphate dehydrogenase. The cellular content ofMT protein was also shown to be elevated in the estrogen-receptor-negative cell lines that express MT-1E mRNA. These results suggest a possible relationship between estrogen receptor status and MT-1E gene expression in human breast cancer. (AmJ Pathol 1998, 152.23-27) paraffin-embedded primary breast carcinomas. MT overexpression was found in the invasive components of 7 of 32 pTl and 17 of 28 pT2 invasive ductal carcinomas, whereas all 26 invasive lobular carcinomas yielded weak or negative results. Fourteen of seventeen pT2 and two of seven pTl invasive ductal carcinomas with MT overexpression developed metastases during follow-up with poor prognostic outcome. In contrast, only 3 of 11 pT2 and none of the 25 pTl cases without MT overexpression had a poor clinical course. The difference was shown to have a high level of significance. This initial study concluded that MT overexpression was associated with a poor prognosis, particularly in pT2 invasive ductal breast carcinomas. These findings regarding MT overexpression in ductal breast cancer have been confirmed and extended by subsequent investigations.2 7 Although these studies demonstrate the potential use of MT as a prognostic marker, further extension to the level of gene expression is limited by the fact that the MT antibody reveals the overall expression of a family of genes rather than a specific single gene product. In humans, the MTs are encoded by a family of genes located at 16q13 consisting of 10 functional MT isoforms (MT-1A, MT-1B, MT-1E, MT-1F, MT-1G, MT-1H, MT-1X, MT-2A, MT-3, and MT-4) and 7 nonfunctional MT isoforms (MT-1C, MT-1D, MT-11, MT-1J, MT-1K, MT-1L, and MT2B).e-l2 This raises the possibility that the prognostic significance of MT expression in human ductal breast cancer might be further enhanced if specific MT gene expression patterns were known. However, the nucleotide bases comprising the amino acid coding regions of all of the MT genes are highly conserved, and it is unlikely that specific antibodies can be made to the individual isoforms. An alternative approach, using reverse transcription polymerase chain reaction technology (RT-