RET Is a Heat Shock Protein 90 (HSP90) Client Protein and Is Knocked Down upon HSP90 Pharmacological Block

Context: Mutations of the RET receptor tyrosine kinase are associated to multiple endocrine neoplasia type 2 (MEN2) and sporadic medullary thyroid carcinoma (MTC). The heat shock protein (HSP) 90 chaperone is required for folding and stability of several kinases. HSP90 is specifically inhibited by 17-allyl-amino-17-demethoxygeldanamycin (17-AAG). Objective: Our aim was to investigate whether RET protein half-life depends on HSP90 and to dissect the molecular pathway responsible for the degradation of RET upon HSP90 inhibition by 17-AAG. Design: 17-AAG effects were studied in RAT1 fibroblasts exogenously expressing MEN2-associated RET mutants and human MTC-derived cell lines. Results: 17-AAG induced a 26S proteasome-dependent degradation of wild-type RET and MEN2-associated RET mutants. The compound hampered HSP90/RET interaction and stabilized RET binding to HSP70, leading to the recruitment of the HSP70-associated E3 ligase C-terminus of Hsc70-interacting protein. In turn, C-terminus of Hsc70-interacting protein polyubiquitinated RET, promoting its proteasomal degradation. 17-AAG blocked RET downstream effectors and RET-dependent transcriptional activation of gene promoters. In human MTC cells carrying oncogenic RET mutants, HSP90 inhibition induced receptor degradation and signaling hindrance leading to cell cycle arrest. Conclusion: RET and MEN2-associated RET mutants rely on HSP90 for protein stability, and HSP90 blockade by 17-AAG promotes RET degradation. (J Clin Endocrinol Metab 95: 3552-3557, 2010)