Non‐myeloablative mixed chimerism approaches and tolerance, a split decision

Abstract

Stable mixed chimerism has been considered the most robust tolerance strategy. However, rejection of solid donor tissues by chimeras has been observed, a state termed split tolerance. Since new non-myeloablative mixed chimerism approaches are being actively pursued, we sought to determine whether they lead to full tolerance or split tolerance and to define the mechanisms involved. Fully mismatched mixed chimeras generated by induction with various lymphocyte-depleting antibodies along with either low-dose irradiation or busulfan and temporary sirolimus, maintained stable mixed chimerism but nevertheless rejected donor skin grafts. Generation of stable mixed chimerism using antibody targeting CD40L, but not depleting antibodies to CD4 and CD8, could prevent split tolerance when skin grafts were given together with donor bone marrow. Minor antigen matching abrogated the ability of effector T cells to reject donor skin grafts. A CFSE killing assay indicated that chimeras were both directly and indirectly tolerant of donor hematopoietic cell antigens, suggesting that minor mismatches triggered a tissue-specific response. Thus, split tolerance due to tissuerestricted polymorphic antigens prevents full tolerance in a number of nonmyeloablative mixed chimerism protocols and a 'tolerizing' agent is required to overcome split tolerance. A model of the requirements for split tolerance is presented.Supporting information for this article is available at http://www.wiley-vch.de/contents/jc_2040/2007/36938_s.pdf IntroductionMuch of the effort to develop donor-specific transplantation tolerance has been focused on inducing peripheral tolerance through costimulation blockade. While initially promising, with more extensive tests of this approach the success has been somewhat limited, particularly when translated to larger animal models and to the clinic [1]. In hindsight this may not be surprising given that tolerance naturally occurs primarily in the thymus and only secondarily in the periphery [2]. In contrast, the approach of generating hematopoietic chimerism via bone marrow transplantation (BMT) takes advantage of the thymic central tolerance mechanisms, and is considered the most robust method of inducing donor-specific tolerance [3][4][5][6]. The chimer- ism approach is limited clinically by the harsh recipient conditioning needed to establish chimerism and the possibility of graft-vs.-host disease [7]. More recently less toxic strategies have been developed that establish mixed allogeneic chimerism, where substantial levels of donor and recipient hematopoietic cells co-exist in the recipient, and have raised hope that robust transplantation tolerance will soon be routine clinically [6]. Mixed chimerism has been considered to induce tolerance to all other donor tissues. If true, mixed chimerism could be a solution for both solid organ transplantation and cellular transplants, such as allogeneic islets used to treat type-1 diabetes. However, studies in full chimeras, where virtually all hematopoietic cells in the recipien...

Development of Either Split Tolerance or Robust Tolerance along with Humoral Tolerance to Donor and Third-Party Alloantigens in Nonmyeloablative Mixed Chimeras

“…However, it is clear that even with chimerism, whether induced naturally or otherwise (6 -11), it is possible for donor tissues to be rejected (known as "split tolerance"). We recently confirmed and extended this observation to murine chimeras generated by more than one clinically relevant protocol (12). Importantly, split tolerance appears to be possible in mixed chimeras (10 -12) that are more clinically favored than full chimeras.…”

“…Importantly, split tolerance appears to be possible in mixed chimeras (10 -12) that are more clinically favored than full chimeras. Most past studies demonstrated split tolerance in chimeras that maintained donor hematopoietic cells but rejected donor skin transplants, the cause of which was likely immunity toward polymorphic tissue-specific Ags expressed by donor skin but absent from their bone marrow cells (10,(12)(13)(14)(15)(16)(17)(18). Although split tolerance involving tissues other than skin has not often been reported in murine chimeras, in a canine model split tolerance was observed in which chimeric recipients rejected donor hearts (19).…”

Establishment of Transplantation Tolerance via Minimal Conditioning in Aged Recipients

“…In a previous study, C57BL/6 mice treated with a similar protocol using 20 mg/kg Bu, T cell-depleting antibodies and transplanted with full-mismatched BALB/c WBM, followed by 14-day SIR immunosuppression, showed stable chimerism but nevertheless rejected donor skin grafts (37). Using a more clinically realistic donor-recipient strain combination, we found that decreasing the Bu dose to 10 mg/kg and increasing the SIR course to 28 days could efficiently establish long-term allogeneic tolerance to donor skin graft and hematopoietic antigens in 100% of mice.…”

“…Bu is typically administered at a dose of 20 mg/kg or higher, with higher levels of chimerism corresponding to dosage increases (6,36,37), as mice are relatively more resistant to Bu than humans. We found that 10 mg/kg Bu was sufficient to deplete LSK progenitors without severely affecting WBM cell numbers.…”

Deletional and regulatory mechanisms coalesce to drive transplantation tolerance through mixed chimerism

“…Increased regulatory activity might thus also provide a potential solution for the phenomenon of "split tolerance" seen in some chimeras, in which some types of grafts (e.g., BM) are accepted while other types (e.g., skin or heart) are rejected [20,72]. Tissuespecific minor antigens that have been identified to cause split tolerance in several models [73,74], are now known to be amenable to active regulation [20,41]. Thus, deliberately shifting the contributing mechanisms toward regulation indeed appears beneficial for the robustness of the state of tolerance achieved through mixed chimerism and increases the efficacy of such protocols.…”