A phase I/II minor histocompatibility antigen-loaded dendritic cell vaccination trial to safely improve the efficacy of donor lymphocyte infusions in myeloma.

Allogeneic stem cell transplantation (allo-SCT) with or without donor lymphocyte infusions (DLI) is the only curative option for several hematological malignancies. Unfortunately, allo-SCT is often associated with GvHD, and patients often relapse. We therefore aim to improve the graft-versus-tumor effect, without increasing the risk of GvHD, by targeting hematopoietic lineage-restricted and tumor-associated minor histocompatibility antigens using peptide-loaded dendritic cell (DC) vaccinations. In the present multicenter study, we report the feasibility, safety and efficacy of this concept. We treated nine multiple myeloma patients with persistent or relapsed disease after allo-SCT and a previous DLI, with donor monocyte-derived mHag-peptide-loaded DC vaccinations combined with a second DLI. Vaccinations were well tolerated and no occurrence of GvHD was observed. In five out of nine patients, we were able to show the induction of mHag-specific CD8+ T cells in peripheral blood. Five out of nine patients, of which four developed mHag-specific T cells, showed stable disease (SD) for 3.5-10 months. This study shows that mHag-based donor monocyte-derived DC vaccination combined with DLI is safe, feasible and capable of inducing objective mHag-specific T-cell responses. Future research should focus on further improvement of the vaccination strategy, toward translating the observed T-cell responses into robust clinical responses.

Induction of multiple myeloma-reactive T cells during post-transplantation immunotherapy with donor lymphocytes and recipient DCs.

Recently, we demonstrated that reduced intensity conditioning (RIC) followed by partial T-cell-depleted SCT creates a platform for inducing the graft-versus-myeloma effect by adjuvant immunotherapy. Here, we evaluated mHA-specific T-cell responses in a multiple myeloma (MM) patient who was treated with RIC-SCT followed by donor lymphocyte infusion (DLI) and subsequent recipient DC vaccination. We isolated a mHA-specific CTL clone with the capacity to target MM tumor cells from this patient experiencing long-term CR. This CTL clone recognizes an HLA-A3-restricted mHA and mediates killing of both primary MM cells and the MM-cell line U266, while BM-derived fibroblasts are not recognized. CTL-specific T-cell receptor (TCR) transcripts could be detected by quantitative PCR analysis in both peripheral blood and BM during tumor remission. Interestingly, a strong increase of CTL-specific TCR transcripts at the BM tumor site was observed following DLI and recipient DC vaccination, while the TCR signal in peripheral blood decreased. These findings illustrate that the approach of partial T-cell-depleted RIC-SCT followed by post-transplantation immunotherapy induces mHA-specific T-cell responses targeting MM tumor cells.

TCR gamma delta cytotoxic T lymphocytes expressing the killer cell-inhibitory receptor p58.2 (CD158b) selectively lyse acute myeloid leukemia cells.

Cytotoxic T lymphocytes (CTL) are thought to play an important role in the graft-versus-leukemia (GVL) response. Unfortunately, GVL reactivity is often associated with life-threatening graft-versus-host disease (GVHD). Characterization of CTL that selectively attack leukemic cells but not normal cells may lead to the development of adjuvant immunotherapy that separates GVL from GVHD. Here, we describe TCR gamma delta (V gamma 9/V delta 1) CTL, isolated from the peripheral blood of an AML patient after stem cell transplantation (SCT), that very efficiently lysed freshly isolated acute myeloid leukemia (AML) cells and AML cell lines. Interestingly, HLA-matched non-malignant hematopoietic cells were not killed. We revealed that the killer cell-inhibitory receptor (KIR) p58.2 (CD158b) specific for group 2 HLA-C molecules negatively regulates the cytotoxic effector function displayed by these TCR gamma delta CTL. First, an antibody against HLA-C enhances lysis of non-malignant cells. Secondly, stable transfection of HLA-Cw*0304 into the class I-negative cell line 721.221 inhibited lysis. Finally, engagement of p58.2 by antibodies immobilized on Fc gamma R-expressing murine P815 cells inhibits CD3- and TCR gamma delta-directed lysis. Compared to non-malignant hematopoietic cells, AML cells express much lower levels of MHC class I molecules making them susceptible to lysis by p58.2(+) TCR gamma delta CTL. Such KIR-regulated CTL reactivity may have a role in the GVL response without affecting normal tissues of the host and leading to GVHD.

A human minor histocompatibility antigen specific for B cell acute lymphoblastic leukemia.

Human minor histocompatibility antigens (mHags) play an important role in the induction of cytotoxic T lymphocyte (CTL) reactivity against leukemia after human histocompatibility leukocyte antigen (HLA)-identical allogeneic bone marrow transplantation (BMT). As most mHags are not leukemia specific but are also expressed by normal tissues, antileukemia reactivity is often associated with life-threatening graft-versus-host disease (GVHD). Here, we describe a novel mHag, HB-1, that elicits donor-derived CTL reactivity in a B cell acute lymphoblastic leukemia (B-ALL) patient treated by HLA-matched BMT. We identified the gene encoding the antigenic peptide recognized by HB-1-specific CTLs. Interestingly, expression of the HB-1 gene was only observed in B-ALL cells and Epstein-Barr virus-transformed B cells. The HB-1 gene-encoded peptide EEKRGSLHVW is recognized by the CTL in association with HLA-B44. Further analysis reveals that a polymorphism in the HB-1 gene generates a single amino acid exchange from His to Tyr at position 8 within this peptide. This amino acid substitution is critical for recognition by HB-1-specific CTLs. The restricted expression of the polymorphic HB-1 Ag by B-ALL cells and the ability to generate HB-1-specific CTLs in vitro using peptide-loaded dendritic cells offer novel opportunities to specifically target the immune system against B-ALL without the risk of evoking GVHD.

Recognition of a B cell leukemia-associated minor histocompatibility antigen by CTL.

CTL directed against minor histocompatibility Ags (mHag) play a major role in antileukemia reactivity after HLA-identical bone marrow transplantation. Some of these mHag are restricted to hemopoietic cells, others show a broad tissue expression. Therefore, antileukemia reactivity is often associated with graft-vs-host disease. Here, we report the identification of a B cell leukemia-associated mHag, HB-1, recognized by a CD8+ CTL clone derived from peripheral blood of an acute lymphoblastic B cell leukemia patient who has been treated by HLA-matched bone marrow transplantation. Interestingly, the CTL clone that recognizes HB-1 exhibits specific cytotoxicity toward leukemic as well as EBV-transformed B cells, but not against untransformed B cells. Moreover, the CTL clone does not lyse PHA-stimulated T cell blasts, monocytes, and fibroblasts, indicating that HB-1 is mainly expressed by transformed B cells. Further analysis reveals that HB-1 is restricted by HLA-B44 (both B*4402 and B*4403) and that 28% of HLA-B44-positive individuals express HB-1. These findings demonstrate that leukemia-associated mHag with a restricted tissue distribution, such as HB-1, elicit CTL reactivity in vivo. These Ags are of potential use in immunotherapy against leukemia because they generate antileukemia reactivity that is not associated with graft-vs-host disease.