P064 IL-21 neutralization in experimental colitis: regulation of cytokines, chemokines and colonic transcripts

Abstract

Background: Interleukin 21 (IL-21), a member of the common gamma-chain family of cytokines, is secreted by activated T cells and NKT cells, and has a diverse range of immunomodulatory effects dependent upon the immunological context. IL-21 is produced in the inflamed intestine of patients with IBD, mostly by activated CD4+ helper T cells. Methods: Using a CD4+CD25 SCID adoptive transfer colitis model, we evaluated the effect of a murine monoclonal antibody against IL-21. Mice were treated with anti-IL-21 mAb 25mg/kg (i.p.) 3 × week in either a prophylactic (day 0 42) or a therapeutic (day 21 56) setup. Colon biopsies were analyzed using either luminex or deep sequencing technology. Mesenteric lymph node CD4+ T cells were isolated from mice with colitis and stimulated with different combinations of anti-CD3 and IL-21 or analyzed directly by intracellular FACS analysis. Results: IL-21 production by the transferred CD4+ T cells was found increased over time and correlated with clinical disease. Moreover, a significant reduction of clinical signs of colitis was shown in both anti-IL-21 mAb prophylactic and therapeutically treated mice, when compared to control Ig-treated mice. In anti-IL-21 mAb prophylactic treated mice, several cytokines (including TNF-a and IL-1b) were significantly down-regulated, revealed by deep sequencing analysis of colon biopsies. Additionally, in anti-IL-21 mAb treated mice, luminex analysis on colon biopsies showed a significant down regulation of CCL5, IL-17 and TNF-a. This is in agreement with an up-regulation of the same molecules when mesenteric lymph node CD4+ T cells isolated from mice with colitis were stimulated in vitro with anti-CD3 and IL-21. The percentage of IFN-g and IL-21 positive CD4+ T cells in mesenteric lymph nodes was also significantly down regulated in anti-IL-21 treated mice. Moreover, anti-IL-21 mAb treatment significantly down-regulated colon transcripts for metallo matrix proteases (MMPs), activation markers, adhesions molecules and chemokines (including MMP3, MMP9, CD44, Integrin a4b7, ICAM-1, CXCL13, CCL5, CCL20) and upregulated transcripts involved in mucosal barrier function (including Muc2, TFF3 and Occludin). Conclusions: These results suggest that IL-21 is a relevant target in experimental colitis regulating several pro-inflammatory cytokines and chemokines as well as key molecules in mucosal barrier function. Thus, blockade of this pathway may be of therapeutic benefit in patients with IBD.