Clonal Deletion and Anergy Play Dominant Role To Achieve Immune Tolerance After Reduced Intensity Unrelated Donor Cord Blood Transplantation (UCBT)

Abstract

The mechanism of immune tolerance after successful hematopoietic stem cell transplantation is not fully understood. Both central (clonal deletion) and peripheral (anergy, Treg, Tr1) mechanisms are conceivable and may even coexist. In this study, we set up serial experiments to test for the presence and relative contribution of these mechanisms after HLA-mismatched UCBT in the GvH direction. Nine patients (age 1m to 9y) were transplanted and enrolled on a clinical trial (NCT01852370). Five immunocompetent patients were off immunosuppression therapy (IST) with no GVHD, while 4 had either limited or recently resolved GvHD and were still on IST when studied. Donor T cell chimerism was a median 97%. Purified T cells responses to host APCs were measured by MLR (3HTdR) and CTL reaction (Europium assay), along with cytokine secretion profiling (Bioplex). Host-reactive T cell clones were tracked with TCRVβ repertoire by ImmunoSEQ® (Adaptive Biotech). In tolerant patients, there was no significant MLR or CTL response towards host APC. Similarly, cytokine profiles revealed non-responsiveness while T cells responded vigorously to 3rd party APC by all assays. Recipient-specific T cell clones that were identified from infused UCB stimulated by host APC pre-UCBT became undetectable. There was no indication of Treg or Tr1 cell involvement in sustaining tolerance as deletion by IL-2 immunotoxin or IL-10R blockade was unable to “break” tolerance. However, low dose IL-2 (5U/mL) was able to restore some proliferation (S.I.=19; p=0.05) and IL-13, IFNγ, TNFα secretion, but in lesser degree in comparison to 3rd party responses (S.I.=112). In summary, clonal deletion and anergy contribute to immune tolerance after UCBT with deletion being dominant.