Ex vivo characterization of polyclonal memory CD8+ T-cell responses to PRAME-specific peptides in patients with acute lymphoblastic leukemia and acute and chronic myeloid leukemia - PubMed (original) (raw)

Ex vivo characterization of polyclonal memory CD8+ T-cell responses to PRAME-specific peptides in patients with acute lymphoblastic leukemia and acute and chronic myeloid leukemia

Katayoun Rezvani et al. Blood. 2009.

Abstract

Preferentially expressed antigen of melanoma (PRAME) is aberrantly expressed in hematologic malignancies and may be a useful target for immunotherapy in leukemia. To determine whether PRAME is naturally immunogenic, we studied CD8(+) T-cell responses to 4 HLA-A*0201-restricted PRAME-derived epitopes (PRA100, PRA142, PRA300, PRA425) in HLA-A*0201-positive patients with acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), chronic myeloid leukemia (CML), and healthy donors. CD8(+) T cells recognizing PRAME peptides could be detected ex vivo in 4 of 10 ALL, 6 of 10 AML, 3 of 10 CML patients, and 3 of 10 donors by HLA-A2 tetramer analysis and flow cytometry for intracellular interferon-gamma. The frequency of PRAME-specific CD8(+) T cells was greater in patients with AML, CML, and ALL than healthy controls. All peptides were immunogenic in patients, while responses were only detected to PRA300 in donors. High PRAME expression in patient peripheral blood mononuclear cells was associated with responses to greater than or equal to 2 PRAME epitopes compared with low PRAME expression levels (4/7 vs 0/23, P = .001), suggesting a PRAME-driven T-cell response. PRAME-specific T cells were readily expanded in short-term cultures in donors and patients. These results provide evidence for spontaneous T cell reactivity against multiple epitopes of PRAME in ALL, AML, and CML. The potential for developing PRAME as a target for immunotherapy in leukemia deserves further exploration.

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Figures

Figure 1

CD8+ T-cell responses defined by analysis of intracellular cytokine production. (A) AML patient 3. (B) CML patient 17. (C) ALL patient 23. (D) Healthy donor 34. PBMCs (106) were loaded with test peptides, and intracellular cytokine analysis for IFN-γ was performed as described in “Methods.” Plots are gated on CD3+ T cells. Numbers in the upper right quadrant represent the percentage of IFN-γ–producing CD8+ T cells.

Figure 2

Relationship between PRAME gene expression and the number of PRAME epitopes recognized by CD8+ T cells in patients with AML, CML, ALL, and donors. Bars represent median values.

Figure 3

CD8+ T-cell response to PRAME in patients and donors. (A) Cumulative CD8+ T-cell response to stimulation with all 4 PRAME peptides, PRA100, PRA142, PRA300, and PRA425, in patients with myeloid leukemia (AML and CML), patients with ALL, and healthy donors. CD8+ T-cell responses to stimulation with the HLA-A*0201–restricted peptides: (B) PRA100, (C) PRA142, (D) PRA300, and (E) PRA425 in patients with leukemia (AML, ALL, and CML) compared with healthy donors. Values represent frequencies of PRAME-specific CD8+ T cells. (F) High- and low-avidity CD8+ T-cell responses determined by sensitivity to peptide concentration. Stimulation of PBMCs with 0.1 and 10 μM of PRA100, PRA142, PRA300, and PRA425 determined high- and low-avidity responses, respectively. Results shown are the ratios of high- to low-avidity CD8+ T-cell responses, calculated for patients with PRAME greater than 0.001 (○) and for patients with PRAME less than 0.001 (●). Ratios were obtained by the following calculation: IFN-γ plus CD8+ T cell (%) with 0.1 μM peptide/IFN-γ plus CD8+ T cell (%) with 10 μM peptide. Bars represent the median high/low-avidity ratio for each peptide.

Figure 4

Characterization of PRAME-specific and CMV-specific CD8+ T-cell functionality in patients with leukemia. Shown are representative data of the PRAME- and CMV-specific CD8+ T-cell response from patient 9, after a 5-hour in vitro stimulation. See “Methods” for a detailed explanation of the procedure.

Figure 5

Phenotypic characterization of tetramer-positive CD3+CD8+ T cells. CMVpp65495/HLA-A*0201 CD8+ T cells were used as a positive control and HLA-A2 null as negative control. (A) Peptide/HLA-A2 tetramer analysis of PBMCs was performed by 6-color flow cytometry in 2 patients. (B) CD45RO and CD27 phenotype of CD3+ CD8+ T-cell gated tetramer-positive lymphocytes on samples from patient 25.

References

1. Ikeda H, Lethe B, Lehmann F, et al. Characterization of an antigen that is recognized on a melanoma showing partial HLA loss by CTL expressing an NK inhibitory receptor. Immunity. 1997;6:199–208. - PubMed
1. van Baren N, Chambost H, Ferrant A, et al. PRAME, a gene encoding an antigen recognized on a human melanoma by cytolytic T cells, is expressed in acute leukaemia cells. Br J Haematol. 1998;102:1376–1379. - PubMed
1. van't Veer LJ, Dai H, van de Vijver MJ, et al. Gene expression profiling predicts clinical outcome of breast cancer. Nature. 2002;415:530–536. - PubMed
1. Boon K, Edwards JB, Siu IM, et al. Comparison of medulloblastoma and normal neural transcriptomes identifies a restricted set of activated genes. Oncogene. 2003;22:7687–7694. - PubMed
1. Epping MT, Wang L, Edel MJ, et al. The human tumor antigen PRAME is a dominant repressor of retinoic acid receptor signaling. Cell. 2005;122:835–847. - PubMed

Ex vivo characterization of polyclonal memory CD8+ T-cell responses to PRAME-specific peptides in patients with acute lymphoblastic leukemia and acute and chronic myeloid leukemia - PubMed (original) (raw)