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Clinical Chemistry 47: 505-512, 2001;
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(Clinical Chemistry. 2001;47:505-512.)
© 2001 American Association for Clinical Chemistry, Inc.

Articles

Molecular Strategy for Detecting Metastatic Cancers with Use of Multiple Tumor-specific MAGE-A Genes

Isao Miyashiro1, Christine Kuo1, Kelly Huynh1, Aritoshi Iida1, Donald Morton2, Anton Bilchik3, Armando Giuliano4 and Dave S.B. Hoon1,a

1 Department of Molecular Oncology,
2 John Wayne Cancer Clinic,
3 Division Gastrointestinal Surgery, and
4 Joyce Eisenberg Keefer Breast Center, John Wayne Cancer Institute, Santa Monica, CA 90404.
a Address correspondence to this author at: Department of Molecular Oncology, John Wayne Cancer Institute, 2200 Santa Monica Blvd., Santa Monica, CA 90404. Fax 310-449-5282; e-mail hoon{at}jwci.org.

Background: The human melanoma-associated antigen family A (MAGE-A) has high specificity and expression in various malignancies, but individual family members are expressed at low frequency in any one particular type of cancer. We therefore developed a method to detect mRNAs from multiple MAGE-A genes in a single reaction.

Methods: Universal MAGE-A (uMAGE-A) primers and probe were designed to reverse-transcribe, amplify, and detect by electrochemiluminescence (ECL) MAGE-A mRNAs on the Origen Analyzer. The assay was performed on total RNA of melanoma (n = 9 cell lines and 24 tumors), breast cancer (n = 7 and 26), and colorectal cancer (CRC; n = 5 and 12). We also evaluated blood from melanoma (n = 50), breast cancer (n = 16), and CRC (n = 21) patients.

Results: The uMAGE-A mRNA was detectable in 0.01–1 ng of cell line RNA. The identity of the uMAGE-A cDNA products was confirmed by sequencing and polyacrylamide gel electrophoresis. The uMAGE-A assay increased detection of melanoma, breast cancer, and CRC tumor by 13%, 31%, and 25%, respectively, compared with a MAGE-A1 assay, and by 17%, 19%, and 25%, respectively, compared with a MAGE-A3 assay. The uMAGE-A assay detected circulating tumor cells in the blood of melanoma (24%), breast cancer (25%), and CRC (29%) patients.

Conclusions: The uMAGE-A reverse transcription-PCR/ECL assay provides a practical and sensitive approach for detection of various metastatic cancers in tissues and blood.




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