Abstract 1819: Intrinsic determinants of constitutive activity of exon-skipping variants of androgen receptor in castration resistant prostate cancer
Authors: Uo Takuma , Dvinge Heidi , Sprenger Cynthia C. , Bradley Robert K. , Nelson Peter S. , Plymate Stephen R. 
Published: July 2016

The C-terminally located ligand-binding domain (LBD) of androgen receptor (AR) folds into 12 helices (H1-H12) and its structural integrity permits a strict ligand-dependency of AR to act as a transcription factor. The two most common AR variants (ARVs) V7 and v567es lack LBD and are constitutively active and resistant to the forefront antiandrogen drugs. Moreover, the recent comprehensive splicing landscapes revealed that metastatic castration resistant prostate cancer (mCRPC) harbored multiple forms of ARVs, many of which have diverse patterns of inclusion/exclusion of exons (exon 4-8) corresponding to LBD to produce namely exon-skipping variants. The presence of these previously unreported ARVs adds another layer of complexity to AR signaling. Thus, it crucial to predict the functional outcomes associated with their expression of these ARVs in relation to mCRPC progression. To this end, we characterized these ARVs with respect to their subcellular localization and transactivation capability. Notably, v5es, which skips exon 5 (E5), displayed predominant nuclear localization and constitutive activity as reported for V7 and v567es. v6es, which includes E5 but excludes E6, is mainly localized in cytoplasm and transcriptionally inactive. Exclusion of E5 and 6 introduce premature stop codons in E6 and E7, respectively, which results in deletion of most of the LBD in v5es and v6es. On the other hand, v56es maintains the reading frame resulting in the inclusion of C-terminal half of LBD corresponding to E7 and 8, show nuclear and cytoplasmic distribution, and lack transactivation capability. Both active and inactive ARVs commonly possess E1 through E4. Accordingly, E5 appears to blunt transactivation capability of v6es while both/either of two exons E7 and 8 suppress the activity of v56es. To gain insight into what aspects are different between active and inactive LBD-less ARVs, we first generated and systematically analyzed a series of mutations with deletion of E5 in v6es toward E4-E5 junction. Among the 12 helices in LBD, H4 and H5 reside in E5. While the deletion mutants with intact H5 were similar to v6es in their properties, those with compromise or deletion of residues in H5 exhibited exclusive nuclear localization and constitutive activity. When inhibitory effects of E7 and E8 are considered, v56es without E7 and v56es was found to share similar properties. In contrast, v56es missing E8 was exclusively localized in nucleus and displayed significant but much lower transcriptional activity than that of v5es and the v56es derivative with simultaneous deletion of E7 and 8. These clearly suggest that multiple mechanisms exist to determine subcellular localizations and transactivation activities of ARVs. To develop the treatment based on AR expression profiles, we are continuing to maximize precise prediction of characteristics of ARVs that can singly or in aggregate contribute to mCRPC progression.

Citation Format: Takuma Uo, Heidi Dvinge, Cynthia C. Sprenger, Robert K. Bradley, Peter S. Nelson, Stephen R. Plymate. Intrinsic determinants of constitutive activity of exon-skipping variants of androgen receptor in castration resistant prostate cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1819.