By: Jubair Aziz, Jing Nan Qiao, Bita Janzadeh Sirolimus By: Jubair Aziz, Jing Nan Qiao, Bita Janzadeh PHM142 Fall 2017 Instructor: Dr. Jeffrey Henderson

What is Sirolimus? a.k.a. Rapamycin immunosuppressant agent used to prevent organ transplant rejection Product of streptomyces hygroscopicus FDA approved as an anti-rejection drug for kidney transplants in 1997 site of action: mTOR (mechanistic target of rapamycin)

mechanistic target of rapamycin (mTOR) 289-kD Ser/Thr protein kinase Mediates various cell processes, including cell growth, cell proliferation, protein synthesis, transcription, and more Sirolimus inhibits function of mTOR → immune cells don’t proliferate

Allograft Rejection MHC (major histocompatibility complex) molecules bind to graft and display antigens to T cell receptors → recognize as foreign → T cell activation Via autocrine signalling, IL-2 (cytokine signalling molecule) produced → activation/proliferation of lymphocytes → transplanted tissue destroyed Sirolimus inhibits function of mTOR → halts IL-2 mediated T cell activation and proliferation → suppressed immune system, tissue not rejected

Mechanism of Action Sirolimus binds to a class of protein called FK Binding Protein. The sirolimus:FKPB12 complex act on mTOR, which inhibits mTOR signalling pathways

mTOR and S6K1 mTOR activates S6K1, which phosphorylates the S6 protein of the 40s subunit of ribosome Activation of S6 dramatically increases the translation of a class of polypyrimidine mRNA. Examples of protein coded include eEF2 (eukaryotic elongation factor 2) involved in translation. Blocking mTOR will block protein translation Decreasing synthesis of cell-cycle proteins halts cell cycles and prevents IL 2 mediated T cell proliferation.

mTOR and 4EBP1 When eIF4E binding protein 1 is phosphorylated, it dissociates from eIF4E and allows for translation Initiation. Active mTOR complex phosphorylates eIF4E binding protein 1. Rapamycin inhibits the action of mTOR, therefore inhibiting phosphorylation, preventing translation initiation.

⇆ mTOR and the cell cycle Inhibiting protein translation leads to shortage of proteins at G1 phase Cells fail to progress to S phase ⇆

Effects of Sirolimus mTOR pathway inhibition leads to suppression of T cells Suppression of CD4 effector T cells, promotion of Tregs → Suppression in lymphocyte function mTOR pathway is common to multiple cell lines → Proliferation of other cells Sirolimus binding does not affect calcineurin Relative lack of nephrotoxicity Enhances toxicity of calcineurin inhibitors

Side Effects Hyperlipidemia Myelosuppression Impaired wound healing Proteinuria

A promising drug Promising drug for transplantation, anticancer and novel vaccination strategies

Summary Slide Sirolimus (aka rapamycin) is an immunosuppressant agent used to prevent organ transplant rejection Inhibits the function of mTOR, a protein controls the growth/proliferation of various cells halts IL-2 mediated T cell activation and proliferation through mTOR Sirolimus:FKBP12 acts on mTOR Sirolimus prevents activation of S6K1 and phosphorylation of S6 protein in the 40S subunit, inhibiting translation initiation. Sirolimus inhibits phosphorylation of 4EBP and prevents eIF4E release from 4EBP,also inhibiting translation initiation. Sirolimus stops the cell at the G1 phase and makes it unable to proceed to S phase mTOR pathway inhibition leads to suppression of T cells Sirolimus does not affect calcineurin, but enhances nephrotoxicity of calcineurin inhibitors A balance needs to be found between immunologic tolerance to organ allograft and immune response against malignant cancers

References Sehgal, S. (2003). Sirolimus: its discovery, biological properties, and mechanism of action. Transplantation Proceedings, 35(3), pp.S7-S14. Augustine, J., Bodziak, K. and Hricik, D. (2007). Use of Sirolimus in Solid Organ Transplantation. Drugs, 67(3), pp.369-391. Geissler, E., Schlitt, H. and Thomas, G. (2008). mTOR, Cancer and Transplantation. American Journal of Transplantation, 8(11), pp.2212-2218. Tsang, C.K., Qi H., Liu L.F., Zheng X.F. (2007). Targeting mammalian target of rapamycin (mTOR) for health and diseases. Drug Discovery Today. 12 pp.112–124. Säemann M.D., Haidinger M., Hecking M., Hörl W.H., Weichhart T. (2009). The multifunctional role of mTOR in innate immunity: implications for transplant immunity. Am J Transplant. 9(12), pp.2655–61. Garret et al. (2013). First Canadian Edition. Nelson education p. 1036.