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Antioxidant therapy maximizes tolerance induction to islet allografts

Carmen Fotino¹, R. Damaris Molano¹, Elsie Zahr-Akrawi¹, Judith Molina¹, Maite Lopez-Cabezas¹, Giulia Merizzi³, Antonio Soleti³, Luca Inverardi^1,2,4, Camillo Ricordi^1,2,4,5,6, Antonello Pileggi^1,4,5,6

¹Diabetes Research Institute, University of Miami; ²Medicine, University of Miami Miller School of Medicine; ⁴Microbiology & Immunology, University of Miami Miller School of Medicine; ⁵Surgery, University of Miami Miller School of Medicine; ⁶Biomedical Engineering, University of Miami, Miami, FL, United States; ³Medestea Research, Turin, Italy

We evaluated the effect of the combination of antioxidant drugs with blockade of Lymphocyte Function Associated Antigen-1 (LFA-1) on the survival of fully MHC-mismatched islet allografts.

Streptozotocin-induced diabetic C56BL/6 (H-2^b) mice received DBA/2 (H-2^d) islets under the kidney capsule. Anti-LFA-1 antibody (KBA clone; 100ug daily for 1 week) and/or 15-day antioxidant treatment with 15mg/kg/day of Decanedioic-acid-bis(1-hydroxy-2,2,6,6-tetramethylpiperidin-4-yl)diester-dihydrochloride (Iacvita, IAC, Medestea) were given intraperitoneally. Nonfasting glycemic values were monitored after transplant.

Untreated mice rejected with a median survival time (MST) of 15 (range 7-24) days (n=12). IAC treatment alone resulted in a MST of 17 (12-19) days (n=4; p=N.S.). Anti-LFA-1 alone resulted in long-term (>100 days) survival in 62.5% of the recipients (n=8; p=0.03 vs. controls). Combinatorial anti-LFA-1+IAC resulted in 100% long-term (>100 days) graft survival (n=3; p<0.014 vs. control; p<0.03 vs. IAC). Re-challenge with donor-specific islets without further treatment after nephrectomy of graft-bearing kidney showed that 50% of anti-LFA-1 treated mice (n=2) rejected on 60 days, while all anti-LFA-1+IAC treated animals (n=3) accepted donor-specific islets (>100 days). Administration of anti-CD25 antibody associated with a reduction of Treg cell numbers without resulting in islet rejection. Challenge with donor-specific and third-party (C3H, H2 ^k) skins resulted in rejection of the grafts without loss of islet function.

Our data indicates that peri-transplant LFA-1 blockade results in long-term survival of islet allografts in only a proportion of recipients. A synergistic effect of the combinatorial treatment with LFA-1 blockade and IAC, an antioxidant drug, was observed in this model with all animals maintaining a functional graft long-term and accepting indefinitely donor-specific islets, while able to reject donor-specific and third-party skins. This data suggests the achievement of operational tolerance in this model. The use of antioxidants, such as IAC, may represent a viable strategy to enhance the success of tolerance inducing protocols.