CLONAL ANALYSIS BY STUDY OF X-CHROMOSOME INACTIVATION IN FORMALIN-FIXED PARAFFIN-EMBEDDED TISSUE

Analysis of clonality by X chromosome inactivation has proven to be a powerful strategy in the study of neoplastic and preneoplastic disorde rs (P. J. Fialkow, Biochim. Biophys. Acta, 458: 283-321, 1976; B. Voge lstein et al., Cancer Res., 47: 4806-4813, 1987). Recently, the gene f or the androgen receptor has been shown to be a highly polymorphic loc us in which methylation of DNA correlates with inactivation of one or the other X homologue (R. C. Allen et al., Am. J. Hum. Genet., 51: 122 9-1239, 1992). Unlike other loci used for analysis of X inactivation, the methylation sites within the androgen receptor gene lie close to t he region of DNA containing the polymorphism. Consequently, it should be possible to use methylation-sensitive restriction enzymes and polym erase chain reaction to study differential methylation among alleles o f this gene in formalin-fixed and paraffin-embedded archival tissue sp ecimens. To investigate this question, we performed clonal analysis on a variety of randomly selected, formalin-fixed, paraffin-embedded tum or tissues from 15 women. Thirteen of the women were found to be heter ozygous for polymorphisms at the androgen receptor locus. Among these women, 11 tumors were clearly clonal in assays of methylation of the a ndrogen receptor gene. Interpretation of results for the remaining two cases was complicated because of a skewed pattern of X chromosome ina ctivation found in normal control tissues. We conclude that analysis o f methylation in the androgen receptor gene should allow study of clon ality in most formalin-fixed, paraffin-embedded tissue specimens from women, including small preneoplastic lesions or rare conditions for wh ich sufficient material is not available for analysis by other techniq ues.