Azacitidine augments expansion of regulatory T cells after allogeneic stem cell transplantation in patients with acute myeloid leukemia (AML).

Strategies that augment a GVL effect without increasing the risk of GVHD are required to improve the outcome after allogeneic stem cell transplantation (SCT). Azacitidine (AZA) up-regulates the expression of tumor Ags on leukemic blasts in vitro and expands the numbers of immunomodulatory T regulatory cells (Tregs) in animal models. Reasoning that AZA might selectively augment a GVL effect, we studied the immunologic sequelae of AZA administration after allogeneic SCT. Twenty-seven patients who had undergone a reduced intensity allogeneic transplantation for acute myeloid leukemia were treated with monthly courses of AZA, and CD8(+) T-cell responses to candidate tumor Ags and circulating Tregs were measured. AZA after transplantation was well tolerated, and its administration was associated with a low incidence of GVHD. Administration of AZA increased the number of Tregs within the first 3 months after transplantation compared with a control population (P = .0127). AZA administration also induced a cytotoxic CD8(+) T-cell response to several tumor Ags, including melanoma-associated Ag 1, B melanoma antigen 1, and Wilm tumor Ag 1. These data support the further examination of AZA after transplantation as a mechanism of augmenting a GVL effect without a concomitant increase in GVHD.

Dominant responses with conservation of T-cell receptor usage in the CD8+ T-cell recognition of a cancer testis antigen peptide presented through HLA-Cw7 in patients with multiple myeloma.

Cancer testis antigens exhibit physiological expression within germ cells and are frequently expressed in malignant tissue. Interestingly, immunological tolerance to cancer testis proteins does not appear to be established, and the expression of CTAg proteins within malignant cells can therefore lead to induction of cellular and humoral immunity. A considerable body of evidence now indicates that CD8-specific immunity plays an important role in the control of cancer cell growth, and a number of vaccine studies are in progress to boost CTAg-specific cellular immune responses. We have previously identified CTAg-specific immune responses in patients with multiple myeloma and reported that recognition of the MAGE-A1(289-298) peptide, which is described as being restricted by HLA-B*0702, was the most frequent response seen with our peptide panel. Here, we studied seven CD8+ T-cell clones specific for this peptide which were isolated from three patients with myeloma at several time-points. The affinity of peptide recognition was high with 50% maximal interferon-γ production observed at a peptide concentration of 10(-10) M and variation of only one order of magnitude between the affinities of the clones. Importantly, all the clones were able to recognise and kill multiple myeloma cell lines. Interestingly, one patient did not express HLA-B*0702, but three clones from this patient recognised the MAGE-A1(289-298) peptide on a lymphoblastoid cell line (LCLs) expressing HLA-Cw7, and we now show evidence that the MAGE-A1(289-298) peptide is expressed and recognised through Cw7. The T-cell receptor gene usage was determined in five clones and showed conserved features in both the α and the ß chain genes indicating correlation between T-cell receptor usage and peptide specificity of cancer testis antigen-specific T-cell clones.

Differential pattern of CD4+ and CD8+ T-cell immunity to MAGE-A1/A2/A3 in patients with monoclonal gammopathy of undetermined significance (MGUS) and multiple myeloma.

The factors that determine progression from monoclonal gammopathy of undetermined significance (MGUS) to multiple myeloma are unclear but may include the breakdown of immune surveillance. Cancer testis antigens (CTAgs) are expressed by the majority of myelomas and MGUS tumors and are a potential immune target. We have characterized CD4(+) and CD8(+) T-cell immune responses to MAGE-A1/A2/A3 in these patients. CD4(+) T-cell immunity to MAGE proteins is stronger and more frequent in MGUS compared with myeloma with a predominantly CD45RA(-)CCR7(-) effector memory profile and cytotoxicity against MAGE-positive cell lines. In contrast CD8(+) T-cell immune responses were present almost exclusively in patients with multiple myeloma, correlating with disease, with a CD45RA(+)CCR7(-) memory phenotype, localizing poorly to the bone marrow but were able to lyse myeloma cell lines in vitro. This suggests that the CD4(+) CTAg-specific immune response may play a role in controlling tumor growth, whereas the efficacy of the CD8(+) T-cell response appears to be limited in vivo. Despite this, patients with evidence of a CTAg-specific immune response had a 53% reduction in mortality over a median follow-up of 4 years. These findings have important implications for clinical approaches to CTAg-specific immunotherapy in patients with cancer.

CD8(+) T-cell immunity against cancer-testis antigens develops following allogeneic stem cell transplantation and reveals a potential mechanism for the graft-versus-leukemia effect.

BACKGROUND: Allogeneic stem cell transplantation is associated with a powerful 'graft-versus-leukemia' effect that is generally considered to result from an alloreactive T-cell immune response. However, disease remission can also be observed after syngeneic transplantation and we investigated whether a T-cell immune response to cancer-testis antigens can be detected in patients in the post-transplant period. DESIGN AND METHODS: The T-cell immune response against cancer-testis antigens was studied in a cohort of 41 patients who underwent allogeneic stem cell transplantation for the management of acute myeloid leukemia or multiple myeloma. The cytokine secretion assay was combined with magnetic selection to allow detection of an interferon-gamma-secreting T-cell response to a panel of cancer-testis antigen peptides. RESULTS: A cancer-testis antigen-specific CD8(+) T-cell immune response was observed in the peripheral blood of five patients with an average magnitude of 0.045% of the CD8(+) T-cell repertoire. Four of these patients had undergone reduced intensity conditioning transplantation with alemtuzumab for the treatment of acute myeloid leukemia and three remain in long-term remission. T-cell immunity was focused against peptides derived from MAGE proteins and was markedly increased within the bone marrow. CONCLUSIONS: Functional cancer-testis antigen-specific CD8(+) T-cell immune responses develop in the early period following reduced intensity allogeneic stem cell transplantation and are preferentially localized to bone marrow. These immune responses are likely to contribute to the cellular basis of the graft-versus-leukemia effect.

Neoplastic plasma cells generate an inflammatory environment within bone marrow and markedly alter the distribution of T cells between lymphoid compartments.

Monoclonal gammopathy of undetermined significance (MGUS) and multiple myeloma (MM) are characterised by the accumulation of malignant plasma cells within bone marrow and lead to a range of abnormalities in the peripheral blood T cell repertoire. We investigated the level of inflammatory chemokines within the bone marrow and blood of patients with MGUS and MM and related this to the pattern of chemokine receptor expression on T cells in both compartments.The expression of a wide range of chemokine ligands for CXCR3 and CCR4 was markedly increased within the bone marrow of patients with MGUS and MM compared to healthy donors. The most marked effects were seen for CCL4 and CXCL9 which were increased by 4 and 6 fold respectively in the bone marrow of patients with myeloma. The expression of CXCR3 and CCR4, the major TH1 and TH2-associated chemokine receptors, was increased substantially on T cells within the bone marrow of patients whereas the percentage of CXCR3-expressing T cells within blood was correspondingly decreased. The presence of even small numbers of neoplastic plasma cells or associated stroma can therefore generate an inflammatory chemokine tumour microenvironment. This leads to the selective recruitment or retention of specific T cell subsets which is likely to underlie many of the features regarding the peripheral T cell repertoire in myeloma and may also contribute to the immune suppression associated with this disease. This local inflammatory reaction may represent a tumour-specific immune response or may itself play an important role in tumour progression and as such may offers a potential novel target for therapeutic intervention.