Wilms tumor is genetically heterogeneous, and until recently only one Wilms tumor gene was known,WT1at 11p13. However,WT1is altered in only ˜20% of Wilms tumors. Recently a novel gene,WTXat Xq11.1, was reported to be mutated in Wilms tumors. No overlap between tumors with mutations inWTXandWT1was noted, suggesting thatWT1andWTXmutations could account for the genetic basis of roughly half of Wilms tumors. To assess the frequency ofWTXmutations and their relationship toWT1mutations in a larger (n= 125) panel of Wilms tumors which had been thoroughly assessed for mutations inWT1, we conducted a complete mutational analysis ofWTXthat included sequencing of the entire coding region and quantitative PCR to identify deletions of theWTXgene. Twenty-three (18.4%) tumors carried a total of 24WTXmutations, a lowerWTXmutation frequency than that previously observed. Surprisingly, we observed an equivalent frequency ofWTXmutations in tumors with mutations in either or bothWT1andCTNNB1(20.0%) and tumors with no mutation in eitherWT1orCTNNB1(17.5%).WTXhas been reported to play a role in the WNT/β-catenin signaling pathway, and, interestingly,WTXdeletion/truncation mutations appeared to be rare in tumors carrying exon 3 mutations ofCTNNB1, encoding β-catenin. Our findings indicate thatWT1andWTXmutations occur with similar frequency, that they partially overlap in Wilms tumors, and that mutations inWT1,WTX, andCTNNB1underlie the genetic basis of about one-third of Wilms tumors. © 2008 Wiley-Liss, Inc.