Session Time: 2:30pm-4:00pm
Presentation Time: 3:42pm-3:54pm
Constitutive proteasomes (c-20S) are ubiquitously expressed cellular proteases that degrade polyubiquitinated proteins and regulate cell functions. Immunoproteasomes (IP or i-20S) are primarily expressed in immune cells. C-20S and i-20S differ only in the 3 proteolytic β subunits. We show here that in a murine heart transplant model, effector and memory T and B cells upregulate their expression of i-20S. We hypothesized that i-20S inhibition would target memory T and B cells without the toxicity of inhibiting c-20S.
We designed and synthesized a specific, small molecule inhibitor (DPLG3) of the i-20S β5i subunit. DPLG3 inhibited mouse and human i-20S with IC50 of 9.4 nM and 4.5 nM, respectively with 1500-fold and 7000-fold selectivity over mouse and human c-20S, respectively. C57BL/6 recipients of BALB/c hearts treated with low dose CTLA4Ig (250 [micro]g on day 2) and 14 injections of DPLG3 (25 mg/kg) exhibited indefinite heart survival compared to control (CTLA4Ig + vehicle) (MST >100 and 30 days respectively, n=6/group, p<0.05). DPLG3 significantly reduced effector CD4+ and CD8+ T cells (Teff). Furthermore, CD4 and CD8 Teff in DPLG3-treated mice acquired an exhaustion phenotype with an increased expression of PD1, TIM3 and LAG3, lower Th1 and Th17 markers, and lower GrB in CD8+ Teff (P<0.05, n=8 mice/group). RAG-/- mice recipients of BALB/c skin received 10 x 106 CD3 Foxp3-GFP negative cells and were treated with DPLG3 for 7d. DPLG3-treated mice showed >5-fold increase in induced Tregs in vivo. CD4 and CD8 Teff also exhibited more apoptosis in DPLG3-treated mice (p<0.05, n= 8 mice/group). Similar results were observed following the transfer of β5i-/- CD3 Foxp3-GFP negative cells. Whole RNA seq analysis and gene set enrichment analysis demonstrated upregulation of the STAT5 pathway. β5i-/- CD4 and CD8 T cells and DPLG3 treated T cells demonstrated upregulation of pSTAT5 and exhaustion markers compared to control (p<0.05, 3 separate experiments).
Our data reveals a critical role for the IP in protecting T cells against exhaustion and inhibiting induction of Tregs in response to alloimmune activation via inhibition of the JAK-STAT5 pathway.
CITATION INFORMATION: Routray S, Karreci E, Uehara M, Assaker J, Mihali A, Eskandari S, Riella L, Abdi R, Kawano Y, Nathan C, Lin G, Azzi J. Designing a Novel Selective Immunoproteasome Inhibitor Reveals a Critical Role of the IP and STAT5 Pathway in Regulating T Cell Exhaustion and Promoting Long Term Heart Allograft Acceptance. Am J Transplant. 2017;17 (suppl 3).
To cite this abstract in AMA style:Routray S, Karreci E, Uehara M, Assaker J, Mihali A, Eskandari S, Riella L, Abdi R, Kawano Y, Nathan C, Lin G, Azzi J. Designing a Novel Selective Immunoproteasome Inhibitor Reveals a Critical Role of the IP and STAT5 Pathway in Regulating T Cell Exhaustion and Promoting Long Term Heart Allograft Acceptance. [abstract]. Am J Transplant. 2017; 17 (suppl 3). https://atcmeetingabstracts.com/abstract/designing-a-novel-selective-immunoproteasome-inhibitor-reveals-a-critical-role-of-the-ip-and-stat5-pathway-in-regulating-t-cell-exhaustion-and-promoting-long-term-heart-allograft-acceptance/. Accessed May 8, 2021.
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