Graft-versus-Leukemia Antigen CML66 Elicits Coordinated B-Cell and T-Cell Immunity after Donor Lymphocyte Infusion

Abstract

Purpose: The target antigens of graft-versus-leukemia that are tumor associated are incompletely characterized.Experimental Design: We examined responses developing against CML66, an immunogenic antigen preferentially expressed in myeloid progenitor cells identified from a patient with chronic myelogenous leukemia who attained long-lived remission following CD4+ donor lymphocyte infusion (DLI).Results: From this patient, CML66-reactive CD8+ T-cell clones were detected against an endogenously presented HLA-B*4403-restricted epitope (HDVDALLW). Neither CML66-specific antibody nor T-cell responses were detectable in peripheral blood before DLI. However, by 1 month after DLI, CD8+ T cells were present in peripheral blood and at 10-fold higher frequency in marrow. Subsequently, plasma antibody to CML66 developed in association with disease remission. Donor-derived CML66-reactive T cells were detected at low levels in vivo in marrow before DLI by ELISpot and by a nested PCR-based assay to detect clonotypic T-cell receptor sequences but not in blood of the patient pre-DLI nor of the graft donor.Conclusions: CD4+ DLI results in rapid expansion of preexisting marrow-resident leukemia-specific donor CD8+ T cells, followed by a cascade of antigen-specific immune responses detectable in blood. Our single-antigen analysis thus shows that durable posttransplant tumor immunity is directed in part against nonpolymorphic overexpressed leukemia antigens that elicit coordinated cellular and humoral immunity.

Autologous CLL cell vaccination early after transplant induces leukemia-specific T cells

“…Recipient PHA T cell blasts were generated by exposing recipient pre-HSCT PBMCs to 1% PHA (Life Technologies) in IMDM supplemented with 10% human AB serum and 10 mM HEPES (Life Technologies) for 72 hours and by subsequent expansion of the cells in the presence of 100 U/ml IL-2 (R&D Systems). Recipient fibroblast lines were generated by the Cell Culture Core lab of the Harvard Skin Disease Research Center from 5-mmdiameter skin punch biopsies and subsequently cultured in a 1:1 mixture of M199 and M106 medium containing 15% FBS, 10 ng/ml EGF, and 0.4 μg/ml hydrocortisone (EMD Millipore), as described (15).…”

“…IFN-γ secretion was detected using capture (15 μg/ml) and detection (1 μg/ml) antibodies (Mabtech AB) and imaged on an ELISpot reader (ImmunoSpot Series Analyzer; Cellular Technology). For some study subjects, bulk CD8 + T cells were cloned by limiting dilution on feeder cells (irradiated allogeneic PBMCs [and 5:1 autologous EBV cells (15) in the case of T cells obtained from Patient 5] with 100 U/ml recombinant human IL-2 and 1% PHA) after confirming their CLL reactivity by IFN-γ ELIspot assays as described above. Following expansion, we performed ELISpot assays to test the clones (10 4 cells per well) for reactivity against irradiated CD40L-activated CLL B cells, PHA T cell blasts, or fibroblasts (5 × 10 5 cells per well).…”

“…The presence and expansion of antigen-specific CD8 + T cells have been associated with tumor regression following allo-HSCT or donor lymphocyte infusion (DLI) (13,15,25). We sought to determine whether tumor-specific CD8 + T cell reactivity was enhanced by autologous tumor cell vaccination, beyond the effects of allo-HSCT alone.…”

“…We and others have demonstrated that GvL responses are initiated and sustained by the development of coordinated cellular and humoral immunity against tumor antigens and are not limited to a sole alloantigen response (12)(13)(14)(15). These studies have further suggested that individual patients have unique profiles of immunogenic tumor antigens, likely reflecting the heterogeneity of the genetic alterations found in tumor cells from different patients as well as the diversity of HLA (12)(13)(14)(15). Based on these principles, vaccination with autologous, irradiated leukemia cells is an attractive approach to expand leukemia-reactive T cells, since this cancer vaccine formulation reliably includes personal tumor antigens and can potentially elicit polyclonal CD4 + and CD8 + antitumor T cell responses (16).…”

New strategies of DLI in the management of relapse of hematological malignancies after allogeneic hematopoietic SCT

“…There is evidence that CD8-depleted (CD4+) DLI can lead to a rapid expansion of pre-existing marrow-resident leukemia-specific CD8+ T cells, followed by a cascade of peripheral B-and T-antigenspecific immune responses. 35 These infiltrating T cells in the site of disease are very critical to tumor control. 36,37 Bachireddy et al /kg CD4+ T cells as DLI.…”