PD-L1– and calcitriol-dependent liposomal antigen-specific regulation of systemic inflammatory autoimmune disease
Ryan Galea, Hendrik J. Nel, Meghna Talekar, Xiao Liu, Joshua D. Ooi, Megan Huynh, Sara Hadjigol, Kate J. Robson, Yi Tian Ting, Suzanne Cole, Karyn Cochlin, Shannon Hitchcock, Bijun Zeng, Suman Yekollu, Martine Boks, Natalie Goh, Helen Roberts, Jamie Rossjohn, Hugh H. Reid, Ben J. Boyd, Ravi Malaviya, David J. Shealy, Daniel G. Baker, Loui Madakamutil, A. Richard Kitching, Brendan J. O’Sullivan, Ranjeny Thomas
Ryan Galea, Hendrik J. Nel, Meghna Talekar, Xiao Liu, Joshua D. Ooi, Megan Huynh, Sara Hadjigol, Kate J. Robson, Yi Tian Ting, Suzanne Cole, Karyn Cochlin, Shannon Hitchcock, Bijun Zeng, Suman Yekollu, Martine Boks, Natalie Goh, Helen Roberts, Jamie Rossjohn, Hugh H. Reid, Ben J. Boyd, Ravi Malaviya, David J. Shealy, Daniel G. Baker, Loui Madakamutil, A. Richard Kitching, Brendan J. O’Sullivan, Ranjeny Thomas
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Research Article Nephrology

PD-L1– and calcitriol-dependent liposomal antigen-specific regulation of systemic inflammatory autoimmune disease

Abstract

Autoimmune diseases resulting from MHC class II–restricted autoantigen-specific T cell immunity include the systemic inflammatory autoimmune conditions rheumatoid arthritis and vasculitis. While currently treated with broad-acting immunosuppressive drugs, a preferable strategy is to regulate antigen-specific effector T cells (Teffs) to restore tolerance by exploiting DC antigen presentation. We targeted draining lymph node (dLN) phagocytic DCs using liposomes encapsulating 1α,25-dihydroxyvitamin D3 (calcitriol) and antigenic peptide to elucidate mechanisms of tolerance used by DCs and responding T cells under resting and immunized conditions. PD-L1 expression was upregulated in dLNs of immunized relative to naive mice. Subcutaneous administration of liposomes encapsulating OVA323–339 and calcitriol targeted dLN PD-L1hi DCs of immunized mice and reduced their MHC class II expression. OVA323–339/calcitriol liposomes suppressed expansion, differentiation, and function of Teffs and induced Foxp3+ and IL-10+ peripheral Tregs in an antigen-specific manner, which was dependent on PD-L1. Peptide/calcitriol liposomes modulated CD40 expression by human DCs and promoted Treg induction in vitro. Liposomes encapsulating calcitriol and disease-associated peptides suppressed the severity of rheumatoid arthritis and Goodpasture’s vasculitis models with suppression of antigen-specific memory T cell differentiation and function. Accordingly, peptide/calcitriol liposomes leverage DC PD-L1 for antigen-specific T cell regulation and induce antigen-specific tolerance in inflammatory autoimmune diseases.

Authors

Ryan Galea, Hendrik J. Nel, Meghna Talekar, Xiao Liu, Joshua D. Ooi, Megan Huynh, Sara Hadjigol, Kate J. Robson, Yi Tian Ting, Suzanne Cole, Karyn Cochlin, Shannon Hitchcock, Bijun Zeng, Suman Yekollu, Martine Boks, Natalie Goh, Helen Roberts, Jamie Rossjohn, Hugh H. Reid, Ben J. Boyd, Ravi Malaviya, David J. Shealy, Daniel G. Baker, Loui Madakamutil, A. Richard Kitching, Brendan J. O’Sullivan, Ranjeny Thomas

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Figure 3

Induction of antigen-specific pTregs, prolonged suppression of antigen-specific immunity, and infectious tolerance after s.c. injection of calcitriol/OVA323–339 liposomes.

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Induction of antigen-specific pTregs, prolonged suppression of antigen-s...
(A) DO11.10 T cells were transferred to BALB/c hosts. Mice were administered liposomes s.c. once on day 0 or twice on days 0 and 6. Splenocytes were analyzed on day 6 (1 injection) or 10 (2 injections). (B) After adoptive transfer of DO11.10 T cells, mice were administered liposomes or the same concentrations of nonencapsulated OVA323–339 and calcitriol s.c. on days 0 and 6; splenocytes were analyzed on day 12. For intracellular (i.c.) IFN-γ analysis, splenocytes were restimulated overnight with OVA323–339. (C) Thee days after s.c OVA/QuilA priming, liposomes were administered s.c. After 24 hours, sorted dLN MHCII+CD11c+ DCs, MHCII+CD19+ B cells, and CD45– stromal cells were incubated with DO11.10 CD4+ T cells in the presence of IL-2. In vitro positive control DO11.10 CD4+ T cells were stimulated with anti-CD3 and anti-CD28 in presence of IL-2, TGF-β, anti-IL-4, and anti-IFN-γ. CD25+Foxp3+ Tregs were enumerated 5 days later. (D) After DO11.10 T cell transfer, mice were immunized with OVA/QuilA (day 0). Liposomes were administered s.c. on days 0 and 7. Splenocytes were analyzed on day 12 and after overnight OVA323–339 stimulation for i.c. IFN-γ. (E) 5 × 106 DO11.10 in vitro–generated memory T cells were adoptively transferred to BALB/c hosts, which were administered liposomes s.c. on days 0 and 7. Mice were immunized with OVA/QuilA on day 10 and splenocytes were analyzed on day 17. (F) After DO11.10 T cell transfer, mice were primed with OVA/QuilA (day 0). Liposomes were administered on days 0 and 7. Splenocytes were analyzed on day 12 or day 42 and after overnight OVA323–339 stimulation for i.c. IFN-γ. Mice analyzed later were boosted with OVA/QuilA on day 35. (G) After adoptive transfer of 5 × 106 DO11.10 T cells, BALB/c hosts were administered calcitriol/OVA323–339 liposomes twice s.c. into the tail base, 1 week apart. A second cohort of 5 × 106 CFSE-labeled DO11.10 T cells was adoptively transferred, and mice were immunized with OVA/QuilA on day 10. Splenocytes were analyzed on day 15. For i.c. IFN-γ and IL-10 analysis, splenocytes were restimulated overnight with OVA323–339. n = 6–8 per group, representative of 2 replicates. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001. ANOVA with Tukey’s multiple comparison test (A–C and E) and t tests (D, F, and G).