Restoration of Suppressive Activity of Regulatory T-Cells from Crohn's Disease Patients Following Ex Vivo Expansion with Rapamycin. One Step Closer to Novel Cell Based Therapy?

Cell-based therapy with CD4⁺CD25^hiCD127^loFOXP3⁺ regulatory T cells (Tregs) is conceptually attractive to treat chronic mucosal inflammation in Crohn’s disease (CD).   We recently showed that Tregs expanded (with rapamycin) from FACS-sorted CD PB CD4⁺CD25^hiCD127^loCD45RA⁺ precursors, yield an epigenetically stable FOXP3⁺ cell population that is resistant to pro-inflammatory cytokine expression and homes to human gut in a murine human xenograft model.  However, CD4⁺CD25^lo-int conventional T cells (Tcons) from inflamed CD mucosa are resistant to Treg-mediated suppression in vitro.  To determine if culture with rapamycin enhanced the suppressive ability of CD45RA⁺ Tregs from CD PB, Tcon suppression by expanded and freshly-isolated Tregs from the same donor was compared.  Expansion significantly improved the suppressive ability of CD45RA⁺ Tregs (97.5%±1.5% vs. 55.9%±10.8% at 4:1 Tcon:Treg ratio;<0.05,n=3).  Next, LPMCs and MLN mononuclear cells (MLNMCs) were obtained from inflamed CD resection specimens.  Expanded CD45RA⁺ Tregs significantly suppressed activation of MLN and LP CD3⁺ cells (CD154 expression at 7h), and significantly impaired MLN CD3⁺ proliferation (CFSE dilution at 96h).  These data show that in vitro expansion of CD PB CD45RA⁺ Tregs enhances their suppressive ability and that these cells may modulate immune responses in niches relevant to the pathogenesis of CD, supporting their use in forthcoming clinical trials.