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Administration of antisense oligodeoxyribonucleotides against mRNA of CD80 or CD86 prolongs survival of cardiac allografts by inhibition of CTL activity
Administration of Antisense Oligodeoxyribonucleotides Against mRNA of CD80 or CD86 Prolongs Survival of Cardiac Allografts by Inhibition of CTL Activity S. Qian, Z. Wang, W. Li, Z. Chen, H. Zhang, T. Vickers, J.J. Fung, and L. Lu
A
CUTE rejection of organ allografts is T cell mediated, and initiated by alloantigen presentation by antigen presenting cells (APC). The interaction of APC and T cells not only assists in presentation of foreign antigens but also provides additional signals for T cell activation through costimulatory molecules (CM) such as CD80 and CD86.1,2 A blockade of CD80 or CD86 signaling induces antigen– specific T cell hyporesponsiveness or tolerance.3,4 We designed antisense phosphorothioate–modified oligodeoxyribonucleotides (ODN) that were resistant to nucleases and degraded slowly in vivo5 to specifically target mRNA of mouse CD80 or CD86 and examined their effect on cardiac allograft survival. MATERIALS AND METHODS Antisense ODN designed to specifically target mRNA of mouse CD80 or CD86 (ISIS Pharmaceuticals, Carlsbad, CA) were delivered by continuous release from an Alzet osmotic pump (Alza, Palo Alto, Calif) implanted in the abdominal cavity of C3H (H2k) recipients following transplantation of a B10 (H2b) vascularized heart allograft. The animals in the control groups were given scrambled control ODN or were untreated. Graft infiltrating cells, spleen cells and serum were collected from animals on day 7 posttransplantation and evaluated for donor-specific CTL activity in a 4 hour 51Cr release assay using EL4 (H2b) cells (ATCC, Rockville, Md) as donor MHC specific targets. Titers of donor– specific antibody were determined in a complement-dependent cytotoxicity (CDC) assay using B10 spleen cells as donor–specific targets.
RESULTS AND DISCUSSION
Normal C3H recipients rejected B10 heart allografts at a median survival time (MST) of 12 days (n ⫽ 7). A 14-day administration of anti–CD80 ODN at 12.5 mg/kg per day prolonged survival of cardiac allografts to an MST of 48 days (n ⫽ 4, P ⬍ .01, compared with untreated controls). MST of cardiac allografts in anti–CD86 ODN-treated group was prolonged to 20 days (n ⫽ 4, P ⬍ .05 compared with untreated controls). Both scrambled ODN controls for anti–CD80 and –CD86 failed to prolong allograft survival (MST 12 and 12.5 days, respectively, n ⫽ 4, P ⬎ .05
© 2001 by Elsevier Science Inc. 655 Avenue of the Americas, New York, NY 10010
compared to untreated controls). Antisense ODN targeting CD80 appeared more effective in preventing allograft rejection than that targeting CD86. Graft infiltrating cells freshly isolated from anti-CD80 –treated recipients exhibited decreased donor-specific CTL activity compared to those cells isolated from animals treated with scrambled ODN (22 ⫾ 4.7 vs 35.9 ⫾ 0.7%, at E/T ratio ⫽ 100/1, P ⬍ .05). Anti-CD86 ODN treatment also significantly suppressed donor–specific CTL activity in graft infiltrating cells (6.7 ⫾ 2.6 vs 32.8 ⫾ 4.9% in scrambled ODN controls, at E/T ratio ⫽ 100/1, P ⬍ .01). Administration of anti-CD80 or -CD86 ODN did not influence the titers of CDC donor–specific cytotoxic antibodies in serum. These data suggest that treatment with anti-CD80 or -CD86 ODN markedly inhibits the cellular response of allograft recipients resulting in prolongation of allograft survival. AntiCD80 ODN appears more immunosuppressive than antiCD86 ODN in vivo. Treatment with antisense ODN specifically targeting mRNA of CM on APC may conceivably be a unique approach to preventing acute rejection of allografts. REFERENCES 1. Harding FA, McArthur JG, Gross JA, et al: Nature 356:607, 1992 2. Coulombe M, Yang H, Guerder S, et al: J Immunol 157:4790, 1996 3. Pearson TC, Alexander DZ, Winn KJ, et al: Transplantation 57:1701, 1994 4. Sayegh MH, Akalin E, Hancock WW, et al: J Exp Med 181:1869, 1995 5. Crooke RM, Graham MJ, Cooke ME, et al: J Pharm Exp Ther 275:462, 1995
From the Thomas E. Starzl Transplantation Institute, University of Pittsburgh Medical Center (S.Q., Z.W., W.L., Z.C., J.J.F., L.L.), Pittsburgh, PA; and ISIS Pharmaceuticals (H.Z., T.V.), Carlsbad, CA, USA. Address reprint requests to S. Qian, E1540, BST, University of Pittsburgh Medical Center, 200 Lothrop St, Pittsburgh, PA 15213, USA.
0041-1345/01/$–see front matter PII S0041-1345(00)02137-0 551
Transplantation Proceedings, 33, 551 (2001)
A
CUTE rejection of organ allografts is T cell mediated, and initiated by alloantigen presentation by antigen presenting cells (APC). The interaction of APC and T cells not only assists in presentation of foreign antigens but also provides additional signals for T cell activation through costimulatory molecules (CM) such as CD80 and CD86.1,2 A blockade of CD80 or CD86 signaling induces antigen– specific T cell hyporesponsiveness or tolerance.3,4 We designed antisense phosphorothioate–modified oligodeoxyribonucleotides (ODN) that were resistant to nucleases and degraded slowly in vivo5 to specifically target mRNA of mouse CD80 or CD86 and examined their effect on cardiac allograft survival. MATERIALS AND METHODS Antisense ODN designed to specifically target mRNA of mouse CD80 or CD86 (ISIS Pharmaceuticals, Carlsbad, CA) were delivered by continuous release from an Alzet osmotic pump (Alza, Palo Alto, Calif) implanted in the abdominal cavity of C3H (H2k) recipients following transplantation of a B10 (H2b) vascularized heart allograft. The animals in the control groups were given scrambled control ODN or were untreated. Graft infiltrating cells, spleen cells and serum were collected from animals on day 7 posttransplantation and evaluated for donor-specific CTL activity in a 4 hour 51Cr release assay using EL4 (H2b) cells (ATCC, Rockville, Md) as donor MHC specific targets. Titers of donor– specific antibody were determined in a complement-dependent cytotoxicity (CDC) assay using B10 spleen cells as donor–specific targets.
RESULTS AND DISCUSSION
Normal C3H recipients rejected B10 heart allografts at a median survival time (MST) of 12 days (n ⫽ 7). A 14-day administration of anti–CD80 ODN at 12.5 mg/kg per day prolonged survival of cardiac allografts to an MST of 48 days (n ⫽ 4, P ⬍ .01, compared with untreated controls). MST of cardiac allografts in anti–CD86 ODN-treated group was prolonged to 20 days (n ⫽ 4, P ⬍ .05 compared with untreated controls). Both scrambled ODN controls for anti–CD80 and –CD86 failed to prolong allograft survival (MST 12 and 12.5 days, respectively, n ⫽ 4, P ⬎ .05
© 2001 by Elsevier Science Inc. 655 Avenue of the Americas, New York, NY 10010
compared to untreated controls). Antisense ODN targeting CD80 appeared more effective in preventing allograft rejection than that targeting CD86. Graft infiltrating cells freshly isolated from anti-CD80 –treated recipients exhibited decreased donor-specific CTL activity compared to those cells isolated from animals treated with scrambled ODN (22 ⫾ 4.7 vs 35.9 ⫾ 0.7%, at E/T ratio ⫽ 100/1, P ⬍ .05). Anti-CD86 ODN treatment also significantly suppressed donor–specific CTL activity in graft infiltrating cells (6.7 ⫾ 2.6 vs 32.8 ⫾ 4.9% in scrambled ODN controls, at E/T ratio ⫽ 100/1, P ⬍ .01). Administration of anti-CD80 or -CD86 ODN did not influence the titers of CDC donor–specific cytotoxic antibodies in serum. These data suggest that treatment with anti-CD80 or -CD86 ODN markedly inhibits the cellular response of allograft recipients resulting in prolongation of allograft survival. AntiCD80 ODN appears more immunosuppressive than antiCD86 ODN in vivo. Treatment with antisense ODN specifically targeting mRNA of CM on APC may conceivably be a unique approach to preventing acute rejection of allografts. REFERENCES 1. Harding FA, McArthur JG, Gross JA, et al: Nature 356:607, 1992 2. Coulombe M, Yang H, Guerder S, et al: J Immunol 157:4790, 1996 3. Pearson TC, Alexander DZ, Winn KJ, et al: Transplantation 57:1701, 1994 4. Sayegh MH, Akalin E, Hancock WW, et al: J Exp Med 181:1869, 1995 5. Crooke RM, Graham MJ, Cooke ME, et al: J Pharm Exp Ther 275:462, 1995
From the Thomas E. Starzl Transplantation Institute, University of Pittsburgh Medical Center (S.Q., Z.W., W.L., Z.C., J.J.F., L.L.), Pittsburgh, PA; and ISIS Pharmaceuticals (H.Z., T.V.), Carlsbad, CA, USA. Address reprint requests to S. Qian, E1540, BST, University of Pittsburgh Medical Center, 200 Lothrop St, Pittsburgh, PA 15213, USA.
0041-1345/01/$–see front matter PII S0041-1345(00)02137-0 551
Transplantation Proceedings, 33, 551 (2001)