Antibody mediated rejection (AMR) is one of the most important barriers to improving long-term transplant outcomes. Traditional therapies for AMR do not deplete the source of antibody production such as the mature plasma cells. Bortezomib is the only plasma cell-targeted therapy used in humans with AMR. Bortezomib inhibits the constitutive proteasomes (c-20S), cellular proteases that degrade polyubiquitinated proteins and regulate many cell functions. However, c-20S are ubiquitously expressed in tissues and its inhibition leads to apoptotic cell death, which increases the toxicity of inhibition, limiting the use of bortezomib. However, immunoproteasomes (IP or i-20S) are primarily expressed in immune cells. c-20S and i-20S differ only in 3 proteolytic β subunits. We show here that memory B cells and plasma cells increase their expression of i-20S in the spleen and bone marrow in mice with a heart allograft. We hypothesized that i-20S inhibition would target B cell activation, memory B cells and plasma cells without the toxicity of inhibiting c-20S.

We designed and synthesized a novel, specific, non-covalent, small molecule inhibitor (DPLG3) of the IP β5i subunit. DPLG3 inhibited mouse i-20S with IC50 of 9.4 nM and 1500-fold selectivity over mouse c-20S, and inhibited human i-20S with an IC50 of 4.5 nM and 7000-fold selectivity over human c-20S.

C57BL/6 recipients of BALB/c hearts treated with low dose sCTLA4Ig (250 [micro]g on day 2) and 2 weeks injections of DPLG3 (25 mg/kg/day) exhibited indefinite heart survival prolongation compared to control (CTLA4Ig + vehicle) (MST >100 and 30 days respectively, n=6/group, p<0.05).

We found a significant reduction in activated B cells, memory B cells and plasma cells in the spleen and the bone marrow of DPLG3-treated mice compared to vehicle (p<0.05, n=6-8 mice/group). DPLG3 treatment induced significant suppression of donor specific antibodies measured in the serum of allograft recipients (p<0.05, n=6-8 mice/group).

In summary, we designed a novel and safe plasma cell targeted inhibitor that could improve the treatment of AMR in transplant recipients.

CITATION INFORMATION: Mihali A, Sula Karreci E, Kurdi A, Riella L, Kawano Y, Fan H, Singh P, Ghobrial I, Nathan C, Lin G, Azzi J. Designing and Synthesizing a Novel Highly Selective Immunoproteasome Inhibitor That Promotes Long Term Heart Allograft Acceptance While Reducing Plasma Cells and Donor Specific Antibodies. Am J Transplant. 2016;16 (suppl 3).

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