1. RET Proto-Oncogene of Multiple Endocrine Neoplasia type 2(MEN2)

2. What is RET?  Codes for a RTK  On chromosome 10  21 exons  “Rearranged during Transfection” Alberti et al., Journal of Cellular Physiology. 195: 168-186 (2003)

3. RET protein function  signaling to a variety of pathways, most notably the Ras signaling pathway and the phosphatidylinositol-3 kinase pathway.  The ultimate biological effects: morphological changes in the cytoskeleton, cell scattering, proliferation, and differentiation. Alberti et al., Journal of Cellular Physiology. 195: 168-186 (2003)

4. RET protein structure  N-terminal signal peptide  Cadherin domain  Cysteine rich region  TK domain  C-terminal; different isoforms for different functions Alberti et al., Journal of Cellular Physiology. 195: 168-186 (2003)

5. How is RET normally activated?  Ligands: GDNF – glial derived neutrotrophic factor NTN – neurturin PSP – persephin Artemin  Co-receptors GFR 1, 2, 3 and 4 Alberti et al., Journal of Cellular Physiology. 195: 168-186 (2003)

6. Ligand Binding

7. How is RET normally activated? 1) RTK signalling induced by ligand oligomerization 2) Tyrosine autophosphorylation 3) Ca +2 -dependent extracellular cadherin domain Manie et al. http://tig.trends.comhttp://tig.trends.com

8. Ligand binding Manie et al. http://tig.trends.comhttp://tig.trends.com

9. RET activating pathways

10. Normal RET functions  TK receptor for GDNF, NTN, PSP and Artemin  Expressed in Neural crest Urogenital precursor cells  Required for Kidney morphogenesis Neuron maturation Spermatogonia maturation Alberti et al., Journal of Cellular Physiology. 195: 168-186 (2003)

11. Knockout Mutation

12. RET-null mice  Transgenic mice without functional RET or its co-receptors showed defective enteric growth and ureteric budding Manie et al. http://tig.trends.comhttp://tig.trends.com

13. Kidney morphogenesis Manie et al. http://tig.trends.comhttp://tig.trends.com

14. Recent Finding: RET apoptotic activity  RET to induce apoptosis in the absence of GDNF  Is ligand-independent  Molecular mechanisms unknown Manie et al. http://tig.trends.comhttp://tig.trends.com

15. Summary: Normal RET activation 1) Calcium-dependent ligand binding 2) Autophosphorylation of Tyrosine residues 3) Activation of proliferation pathways 4) Cell growth and differentiation

16. Carcinomas Caused by RET mutations  Papillary Thyroid Carcinoma (PTC)  Medullary Thyroid Carcinomas (MTC) Multiple Endocrine Neoplasia type 2 (MEN2) A,B Famillilar Medullary Thyroid Carcinoma (FMTC)  Hirschsprung disease Manie et al. http://tig.trends.comhttp://tig.trends.com

17. Mutations Manie et al. http://tig.trends.comhttp://tig.trends.com

18. Carcinomas

19. RET in PTC  Rearrangements of RET tyrosine kinase coding region with unrelated genes = PTC

20. Medullary Thyroid Carcinomas (MTCs)  Facts Thyroid C cells are derived from neural crest 75% are sporadic Remainding are inherited forms of MEN2  Activating mutations of RET Alberti et al., Journal of Cellular Physiology. 195: 168-186 (2003)

21. MEN2A  Mutations in cysteine-rich extracellular domain of RET 80% of these mutations are exchange of a cysteine for arginine Leaves unpaired cysteine residue, loss of intramolecular dissulfide bond, and an available intermolecular dissulfide bond with other mutant RET receptors. Alberti et al., Journal of Cellular Physiology. 195: 168-186 (2003)

22. Disulfide bond interactions Alberti et al., Journal of Cellular Physiology. 195: 168-186 (2003)

24. MEN2B  95% are mutations in M918T (exon 16) Usually paternal  5% mutations in RET TK domain Changes substrate binding, RET phosphorylation of proteins, and RET autophosphorylation Alberti et al., Journal of Cellular Physiology. 195: 168-186 (2003)

25. MEN2 Treatment  Good prognosis with early detection  Thryoidectomy early Removal of thyroid gland  RET oncoprotein inhibitors ZD6474 Pyrazolo-pyrimidine (PP2)

26. ZD6474: Oral inhibitor  In vivo Blocked phosphorylation of PTC3 and MEN2B oncoproteins Blocked EGF receptor Reverted some PTC cells http://cancerres.aacrjournals.org/cgi/content/full/62/24/7284

27. PP2  Blocked kinase activity of PTC1 oncoproteins  Prevented serium growth of PTC cells  Hopeful for MTC therapy http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=12679489&dopt=Abstract

28. Sources  Alberti et al., Journal of Cellular Physiology. 195: 168- 186 (2003)  Manie et al. http://tig.trends.com http://tig.trends.com  http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retr ieve&db=PubMed&list_uids=12679489&dopt=Abstract http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retr ieve&db=PubMed&list_uids=12679489&dopt=Abstract  http://cancerres.aacrjournals.org/cgi/content/full/64/2 4/7284.com http://cancerres.aacrjournals.org/cgi/content/full/64/2 4/7284.com  http://www.ohiolink.edu/etd/send- pdf.cgi?osu970163275