The TLE genes constitute a family of important transcriptional co-repressors involved in many cellular processes. We found evidence of alternatively spliced mRNAs for human TLE1-4 containing premature stop codons, thus encoding putative shortened proteins. Microarray experiments and Real-time RT-PCR assays showed that alternatively spliced isoforms of TLE1, TLE2 an d TLE3 were preferentially expressed in prostate in comparison to liver and kidney tissues. We identified by orientation-specific R T-PCR an antisense partially intronic non-coding RNA that overlaps a novel exon of the TLE3 gene, raising the possibility of regulation of alternative splicing by this non-coding transcript. The alternatively spliced isoform of TLE3 was up-regulated (6- to 17-fo ld) in prostate tumors in comparison to matched non-tumor adjacent tissue from 7 out of 11 (64%) patients and in four prostate tumor cell lines in comparison to a normal prostate cell line. These results demonstrate that different isoforms of TLE genes are commonly transcribed in human tissues and suggest that TLE3 could be involved in prostate cancer development.