PTEN is a dual phosphatase tumor suppressor, which is mutated in Cowden Syndrome (CS), characterized by high risk of breast and thyroid cancer, Bannayan-Riley-Ruvalcaba syndrome (BRRS), a rare developmental disorder, and altered in many types of sporadic cancers. Inspite of the growing importance of alternative splicing, there is very limited information on its role in PTEN and associated cancers. We identified 8 novel splice variants: 3 retained various lengths of intron 3 (3a, 3b, 3c); 3 retained various regions of intron 5 (5a, 5b, 5c); and 2 excluded part of exon 5 (delE5) and all of exon 6 (delE6), respectively. To ascertain the role of these variants in sporadic breast cancer, we utilized the rapid-scan panel of cDNA from 12 normal and 12 tumor tissues. While PTEN FL levels were reduced in 10 of 12 breast cancers, variants 3b, 3c and 5c were not expressed in 7, 6 and 4 of 12 breast cancers, respectively and reduced in the remainder. On the other hand, variant 5b seemed to be overexpressed in all the breast cancer tissues. To ascertain the role of these variants in heritable breast cancer, we analyzed 16 CS/BRRS patients with no PTEN mutation and 8 normal controls. Variant 5a was relatively under-expressed and 3a over-expressed in the germline of all CS/BRRS individuals compared to normal controls. We then functionally analyzed these variants by measuring PTEN’s downstream readouts, ie, the effect of each variant on the cyclinD1 promoter and phosphorylation of Akt (P-Akt). Variant 5a and delE6 behaved functionally like FL-PTEN by decreasing P-Akt and CyclinD1 promoter activity, while the variants 5b and 5c had the opposite effect on the CyclinD1 promoter activity. These functional consequences such as 5a behaving like FL corroborates our observation that 5a is underexpressed in CS/BRRS compared to controls. Similarly, our demonstration that variant 5b behaves functionally to counter PTEN’s action corroborates our observation that 5b is over-expressed in sporadic breast cancers. Interestingly, overexpression of p53 resulted in reduced expression of 3a, 3b and 5a and induction of 3c, 5b and 5c. Thus, the expression of these variants seems to be under the regulation of p53, a transcription factor of PTEN. Overall, our observations suggest that differential expression of PTEN and its splice variants could play a role in the pathogenesis of sporadic breast cancer and CS/BRRS, perhaps by functionally aiding or counteracting FL-PTEN downstream function, and may lend a novel way of making a rapid molecular diagnosis of CS/BRRS without mutation analysis.

[Proc Amer Assoc Cancer Res, Volume 47, 2006]