Role of BH3-Domain Proteins in the Effector Phase of RA

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DESCRIPTION (provided by applicant): Rheumatoid arthritis (RA) is an autoimmune disease characterized by hyperplasia of the synovial lining, inflammation, and destruction of cartilage and bone. In RA, the balance between anti- and pro-apoptotic members of the Bcl-2 family may be shifted towards survival. While we demonstrated that the anti-apoptotic proteins, Bcl-2 and Mcl-1 are increased in RA synovial tissue as compared to controls, no study has examined the therapeutic potential of antagonizing Bcl-2 anti-apoptotic members in arthritis. The study of deficiencies in anti-apoptotic members of the Bcl-2 family is complicated by embryonic lethality or early post-natal death. In contrast, mice deficient in pro-apoptotic Bcl-2 members such as Bak, Bax, and Bim survive and reach adulthood. Since Bim is a critical activator of apoptosis by virtue of its sequestration of Bcl-2/Mcl- 1 and/or its activation of Bak and Bax, it is a potential target for treatment of RA. We demonstrated that expression of the apoptotic initiator Bim is markedly reduced in synovial tissue from RA compared to controls. Further, mice lacking Bim but not the downstream effectors Bak or Bax develop a more severe form of inflammatory arthritis. This exacerbated disease in Bim-/- mice is associated with decreased apoptosis, increased expression of pro-inflammatory molecules, and more macrophages in pannus. Bim-/- macrophages display elevated levels of IL-6, TNF1, and IL-12, enhanced expression of CD40 and CD69, and increased and sustained level of active p38 in response to stimulation with LPS. Based on these data, we hypothesize that the ratio of Bim to Bcl-2 and/or Mcl-1 serves as molecular rheostat that determines the extent of hyperplasia and activation of macrophages in the joint. We will use a pharmacological approach, a whole animal approach, and a cell-specific approach to identify how deficiency in Bim exacerbates inflammatory arthritis. These studies will potentially lead to novel therapeutic approaches to RA. A.1. Examine the effect of altering the association of Bcl-2 family members in the development of inflammatory arthritis. A.2. Determine the effect of loss of function in Bak, Bax, or Bcl-2 on the development of inflammatory arthritis in Bim-/- mice. A.3. Determine the effect of targeted deletion of Bim in monocytes and macrophages on the development of inflammatory arthritis. A.4. Determine the mechanism of Bim-mediated suppression of macrophage activation. The regulation of cell death and growth is vital for maintaining a balance in the human body. However, during the initiation and/or progression of the autoimmune disease, rheumatoid arthritis (RA), this balance is disrupted. In RA there is an increase in cellular growth and a concomitant decrease in cell death leading to an abnormal increase in the tissue that attaches to the cartilage/bone junction, the synovial lining. During RA, the synovial lining invades and destroys the adjacent cartilage and bone. Analysis of tissue from joints of patients with RA revealed that the number of macrophages, correlated with a worse prognosis. We demonstrated that the death signaling cascade mediated by the pro-death protein Bim is dysfunctional in macrophages from patients with RA. Additionally, we have shown that mice lacking Bim in all cell types develop a worse form of arthritis and that the macrophages from these mice are highly activated, meaning they produce significant amounts of deleterious factors that exacerbate the inflammation. We have developed a potential therapeutic molecule that consists of a portion of Bim, termed BH3 domain. We have now demonstrated that systemic delivery of BH3 peptides to mice prevents the development of inflammatory arthritis with no toxicity to the mice. These studies are the first to show the efficacy of BH3 peptides as a potential molecule for treatment of RA. Our immediate goal is to develop the BH3 peptide therapy as a front line treatment for RA and other rheumatic diseases.