Global InformationHUMARA assay information
HUMARA assay is one of the most widely used methods to determine the clonal origin of a tumor.[1][2] The method is based on X chromosome inactivation and it takes advantage of the different methylation status of the gene HUMARA (short for human androgen receptor) located on the X chromosome. Considering the fact that once one X chromosome is inactivated in a cell,[3] all other cells derived from it will have the same X chromosome inactivated, this approach becomes a tool to differentiate a monoclonal population from a polyclonal one in a female tissue. The HUMARA gene, in particular, has three important features that make it highly convenient for the purpose:
- The gene is located on the X chromosome and it goes through inactivation by methylation in normal embryogenesis of a female infant.[3] Because most genes on the X chromosome undergo inactivation, this feature is important.
- Human androgen receptor gene alleles have varying numbers of CAG repeats.[4] Thus, when DNA from a healthy female tissue is amplified by polymerase chain reaction (PCR) for a specific region of the gene, two separated bands can be seen on the gel.
- The region that is amplified by PCR also has certain base orders that make it susceptible to be digested by HpaII (or HhaI) enzyme when it is not methylated.[4] This detail gives the opportunity to researchers to differentiate a methylated allele from the unmethylated allele.
Due to these qualities of the HUMARA gene, clonal origin of any tissue from a female mammalian organism can be determined.
- ^ Parsons, Barbara L. (2008-10-01). "Many different tumor types have polyclonal tumor origin: evidence and implications". Mutation Research. 659 (3): 232–247. doi:10.1016/j.mrrev.2008.05.004. ISSN 0027-5107. PMID 18614394.
- ^ Comertpay, Sabahattin; Pastorino, Sandra; Tanji, Mika; Mezzapelle, Rosanna; Strianese, Oriana; Napolitano, Andrea; Baumann, Francine; Weigel, Tracey; Friedberg, Joseph (2014-01-01). "Evaluation of clonal origin of malignant mesothelioma". Journal of Translational Medicine. 12: 301. doi:10.1186/s12967-014-0301-3. ISSN 1479-5876. PMC 4255423. PMID 25471750.
- ^ a b Gartler, S. M.; Riggs, A. D. (1983-01-01). "Mammalian X-chromosome inactivation". Annual Review of Genetics. 17: 155–190. doi:10.1146/annurev.ge.17.120183.001103. ISSN 0066-4197. PMID 6364959.
- ^ a b Allen, R. C.; Zoghbi, H. Y.; Moseley, A. B.; Rosenblatt, H. M.; Belmont, J. W. (1992-12-01). "Methylation of HpaII and HhaI sites near the polymorphic CAG repeat in the human androgen-receptor gene correlates with X chromosome inactivation". American Journal of Human Genetics. 51 (6): 1229–1239. ISSN 0002-9297. PMC 1682906. PMID 1281384.