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Material, methods and results In rat recipients of kidney allografts where tolerance was induced by costimulation blockade, we observed a 2-fold decrease of plasma CCL5 levels as compared to control syngeneic grafted animals (0.60 ± 0.17 vs. 1.17 ± 0.48 ng/ml; p

Methods and results Here we evaluated in a non-human primates this “Treg sparing strategy” with FR104, a novel monovalent humanized and pegylated Fab’ anti-CD28 antibody fragment. PK/PD studies in monkeys revealed that FR104 presented an elimination half-life of 8 days and 100% target saturation over at least a month after a single iv injection of 5 mg/kg. FR104 was next evaluated in a baboon kidney allograft model at the dose of 5 mg/kg at day 0, 4, 14 and then every two-week until 3 months. Monotherapy modestly but significantly prolonged allograft survival (MST: 18.5 days for monotherapy vs 6 days for untreated recipients). FR104 synergized with low doses tacrolimus (lowTac, trough: 5-10 ng/ml; MST >100 days for FR104/ lowTac vs. 15 days for lowTac alone) as well as with calcineurin-free regimens: therapeutic doses of MMF or rapamycin (day 0-90) with 1 mg/kg of corticosteroids from day 0-14 (MST >100 days for FR104 + MMF/Rapa vs. 18/15 days for MMF/Rapa alone). Flow cytometry analyses indicated that blood Treg cells of the natural and inducible types were preserved in FR104/MMF or FR104/lowTAC bitherapies and accumulated in FR104 monotherapy and in FR104/Rapa bitherapy, whereas Treg cells were lowered by MMF and lowTac monotherapies. Histology also revealed that CTLA4+ and Foxp3+ T lymphocytes were accumulated into the graft of FR104 treated recipients.

Background We recently reported that in a rat major histocompatibility complex (MHC) mismatched heart allograft model, treatment with CD40Ig, a chimeric molecule that blocks CD40L, leads to indefinite allograft survival mediated by CD8CD45RC Tregs [1]. Although essential, the exact role of TCR/MHC/peptide interaction in Treg activity is still unknown. We therefore characterize the allogeneic peptide(s) recognized and the TCR usage of the CD8 CD45RC Tregs.

Material and methods A lewis 1W rat heart is grafted in a MHC-mismatched Lewis 1A rat and infected with 2.10pi of adenovirus recombinant for CD40Ig molecule (AdCD40Ig) the day of the graft. Tregs, effector CD4CD25 T lymphocytes (TL), and plasmacytoide dendritic cells (pDC) from spleen are sorted by FACS Aria for in vitro tests. For in vivo studies, 4,5.10vg of FGL2-recombinant adenovirus associated virus (AAVFGL2) are intramuscularly or intravenously injected in recipients 30 days before the graft. Splenocytes are transferred to sublethaly irradiated rats by i.v injection the day before the graft.

Serum levels of soluble Flt1 (sFlt1), a potent inhibitor of VEGF, are significantly increased during the acute phase of PR3-AAV (2592 ± 11347 pg/ml) and MPO-AAV (476 ± 4258 pg/ml) compared to controls (118 ± 269 pg/ml). sFlt1 levels decreased during disease remission but remained increased compared to controls in patients with PR3-AAV. sFlt1 serum levels correlated with serum levels of C5a, an anaphylatoxin released following complement activation. Serum from patients with acute PR3-AAV induced the release of sFlt1 by human monocytes in vitro, but failed to induce a similar effect on endothelial cells. Pre-treatment of monocytes with an anti-C5a receptor blocking antibody attenuated sFlt1 release. Serum from patients with acute AAV induced a disruption of blood flow in the chicken chorioallantoic membrane assay and this effect was prevented by incubating patients’ serum with an excess of human VEGF.

Background/objectives T cells play probably a major role in the physiopathology of Multiple Sclerosis (MS). One way to study T cell responses in MS is to investigate the T cell receptor (TCR) structure in patients. This method is based on the analysis of the hypervariable region of the TCR named CDR3, responsible of the specific recognition of the antigen. To date, we know that the alterations of T cell repertoire in circulating lymphocytes are more pronounced in MS patients than in healthy controls. Moreover, several teams showed that the infiltrating T cells of the Central Nervous System (CNS) of MS patients present an oligoclonal repartition. So, it would be essential to know if correspondence exists between the alterations of the repertoire in the blood or Cerebrospinal Fluid (CSF) and in the CNS.

Methods and results Here we evaluated FR104, a novel antagonist pegylated Fab’ anti-CD28 monovalent antibody, in xenogeneic graft-versus-host disease (GVHD) induced in NOD/ SCID mice infused with human PBMC and compared efficacy with CTLA4Ig molecules. In vitro, FR104 and CTLA4-Ig (LEA29Y) dose-dependently prevented human T cell proliferation to a similar extent in mixed lymphocytes reactions with EC50 at 0.16 and 0.18 micrograms/ml, respectively. IL-2 secretion after stimulation with SEE superantigens was also inhibited in a similar manner. NOD/SCID mice adoptively transferred with human PBMC presented > 80% engraftment after a week and developed a florid GVHD in the third week due to the proliferation of xenogeneic human T cells and their infiltration into the liver, gut, lung and skin, eventually leading to weight loss and sacrifice. Treatment of recipient mice with 5 mg/Kg FR104 biweekly from day 0 to day 25 reduced target organs infiltration by human T cells and completely prevented weight loss and mortality (p < 0.0001). Mice continued gaining weight and did not succumb even a month after treatment withdrawal. Flow cytometry analysis 2 month post engraftment revealed that FR104 treated mice displayed low level of human cells engraftment (