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Immune responses to RHAMM in patients with acute myeloid leukemia after chemotherapy and allogeneic stem cell transplantation.

Casalegno-Garduño R, Meier C, Schmitt A, Spitschak A, Hilgendorf I, Rohde S, Hirt C, Freund M, Pützer BM, Schmitt M - Clin. Dev. Immunol. (2012)

Bottom Line: Leukemic blasts overexpress immunogenic antigens, so-called leukemia-associated antigens like the receptor for hyaluronan acid-mediated motility (RHAMM).Results were correlated with the clinical outcome of patients.Immunotherapies like peptide vaccination or adoptive transfer of RHAMM-specific T cells might improve the immune response and the outcome of AML/MDS patients.

View Article: PubMed Central - PubMed

Affiliation: Department of Internal Medicine III, University of Rostock, 18057 Rostock, Germany.

ABSTRACT
Leukemic blasts overexpress immunogenic antigens, so-called leukemia-associated antigens like the receptor for hyaluronan acid-mediated motility (RHAMM). Persistent RHAMM expression and decreasing CD8+ T-cell responses to RHAMM in the framework of allogeneic stem cell transplantation or chemotherapy alone might indicate the immune escape of leukemia cells. In the present study, we analyzed the expression of RHAMM in 48 patients suffering from acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS). Furthermore, we correlated transcripts with the clinical course of the disease before and after treatment. Real-time quantitative reverse transcriptase polymerase chain reaction was performed from RNA of peripheral blood mononuclear cells. T cell responses against RHAMM were assessed by tetramer staining (flow cytometry) and enzyme-linked immunospot (ELISPOT) assays. Results were correlated with the clinical outcome of patients. The results of the present study showed that almost 60% of the patients were RHAMM positive; specific T-cells recognizing RHAMM could be detected, but they were nonfunctional in terms of interferon gamma or granzyme B release as demonstrated by ELISPOT assays. Immunotherapies like peptide vaccination or adoptive transfer of RHAMM-specific T cells might improve the immune response and the outcome of AML/MDS patients.

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Longitudinal study of RHAMM-specific CTLs in a patient with AML that received allo-SCT. PBMCs were collected from an AML patient at different time points before or after allo-SCT as indicated and subjected to MLPC. CD8+ T cells showed RHAMM-specific release of IFN-γ (a) and granzyme B (b) at one time point (30 days prior to transplant) when the patient was in CR. This active T-cell population was lost after allo-SCT and not reconstituted when the patient relapsed. (c) CD8+ T cells were stimulated with RHAMM peptide and stained with respective tetramer. Frequencies of RHAMM-specific CTLs vanished over the time, as detected by flow cytometry. Reported frequencies correspond to the gate of CD3+CD8+ T cells (upper numbers) and to all cells in the lymphocyte gate (lower numbers).
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fig3: Longitudinal study of RHAMM-specific CTLs in a patient with AML that received allo-SCT. PBMCs were collected from an AML patient at different time points before or after allo-SCT as indicated and subjected to MLPC. CD8+ T cells showed RHAMM-specific release of IFN-γ (a) and granzyme B (b) at one time point (30 days prior to transplant) when the patient was in CR. This active T-cell population was lost after allo-SCT and not reconstituted when the patient relapsed. (c) CD8+ T cells were stimulated with RHAMM peptide and stained with respective tetramer. Frequencies of RHAMM-specific CTLs vanished over the time, as detected by flow cytometry. Reported frequencies correspond to the gate of CD3+CD8+ T cells (upper numbers) and to all cells in the lymphocyte gate (lower numbers).

Mentions: Longitudinal studies of RHAMM-specific CD8+ T cells were only possible with samples of ten patients who overexpressed RHAMM (29/48 patients) and were HLA-A2 positive, as all peptides used in this study were HLA-A2 restricted. The frequency of RHAMM-specific CD8+ T cells in the peripheral blood was measured by flow cytometry during the course of the disease. Furthermore their activation status and potential to kill RHAMM+ malignant cells was assessed by secretion of IFN-γ and granzyme B, respectively. As displayed in Figure 3(c), RHAMM-specific CTLs were detected at a frequency of 1.24% up to 5.62% of all CD8+ T cells, and 0.03% up to 1.14% of all cells in the lymphocyte gate. In ELISPOT assays a general activity of CD8+ T cells (Figures 3(a) and 3(b)) was detected at the time of diagnosis (Dx) and in CR. Thirty days prior to allo-SCT a stronger release of granzyme B by RHAMM-R3-stimulated CD8+ T cells than by unstimulated T cells was detected. After allo-SCT a general silencing of T cells as for release of both IFN-γ and granzyme B was detectable (Figures 3(a) and 3(b)). In accordance with these findings flow cytometry analysis revealed that the number of RHAMM-tetramer+ CD8+ T cells vanished over the time.


Immune responses to RHAMM in patients with acute myeloid leukemia after chemotherapy and allogeneic stem cell transplantation.

Casalegno-Garduño R, Meier C, Schmitt A, Spitschak A, Hilgendorf I, Rohde S, Hirt C, Freund M, Pützer BM, Schmitt M - Clin. Dev. Immunol. (2012)

Longitudinal study of RHAMM-specific CTLs in a patient with AML that received allo-SCT. PBMCs were collected from an AML patient at different time points before or after allo-SCT as indicated and subjected to MLPC. CD8+ T cells showed RHAMM-specific release of IFN-γ (a) and granzyme B (b) at one time point (30 days prior to transplant) when the patient was in CR. This active T-cell population was lost after allo-SCT and not reconstituted when the patient relapsed. (c) CD8+ T cells were stimulated with RHAMM peptide and stained with respective tetramer. Frequencies of RHAMM-specific CTLs vanished over the time, as detected by flow cytometry. Reported frequencies correspond to the gate of CD3+CD8+ T cells (upper numbers) and to all cells in the lymphocyte gate (lower numbers).
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3375151&req=5

fig3: Longitudinal study of RHAMM-specific CTLs in a patient with AML that received allo-SCT. PBMCs were collected from an AML patient at different time points before or after allo-SCT as indicated and subjected to MLPC. CD8+ T cells showed RHAMM-specific release of IFN-γ (a) and granzyme B (b) at one time point (30 days prior to transplant) when the patient was in CR. This active T-cell population was lost after allo-SCT and not reconstituted when the patient relapsed. (c) CD8+ T cells were stimulated with RHAMM peptide and stained with respective tetramer. Frequencies of RHAMM-specific CTLs vanished over the time, as detected by flow cytometry. Reported frequencies correspond to the gate of CD3+CD8+ T cells (upper numbers) and to all cells in the lymphocyte gate (lower numbers).
Mentions: Longitudinal studies of RHAMM-specific CD8+ T cells were only possible with samples of ten patients who overexpressed RHAMM (29/48 patients) and were HLA-A2 positive, as all peptides used in this study were HLA-A2 restricted. The frequency of RHAMM-specific CD8+ T cells in the peripheral blood was measured by flow cytometry during the course of the disease. Furthermore their activation status and potential to kill RHAMM+ malignant cells was assessed by secretion of IFN-γ and granzyme B, respectively. As displayed in Figure 3(c), RHAMM-specific CTLs were detected at a frequency of 1.24% up to 5.62% of all CD8+ T cells, and 0.03% up to 1.14% of all cells in the lymphocyte gate. In ELISPOT assays a general activity of CD8+ T cells (Figures 3(a) and 3(b)) was detected at the time of diagnosis (Dx) and in CR. Thirty days prior to allo-SCT a stronger release of granzyme B by RHAMM-R3-stimulated CD8+ T cells than by unstimulated T cells was detected. After allo-SCT a general silencing of T cells as for release of both IFN-γ and granzyme B was detectable (Figures 3(a) and 3(b)). In accordance with these findings flow cytometry analysis revealed that the number of RHAMM-tetramer+ CD8+ T cells vanished over the time.

Bottom Line: Leukemic blasts overexpress immunogenic antigens, so-called leukemia-associated antigens like the receptor for hyaluronan acid-mediated motility (RHAMM).Results were correlated with the clinical outcome of patients.Immunotherapies like peptide vaccination or adoptive transfer of RHAMM-specific T cells might improve the immune response and the outcome of AML/MDS patients.

View Article: PubMed Central - PubMed

Affiliation: Department of Internal Medicine III, University of Rostock, 18057 Rostock, Germany.

ABSTRACT
Leukemic blasts overexpress immunogenic antigens, so-called leukemia-associated antigens like the receptor for hyaluronan acid-mediated motility (RHAMM). Persistent RHAMM expression and decreasing CD8+ T-cell responses to RHAMM in the framework of allogeneic stem cell transplantation or chemotherapy alone might indicate the immune escape of leukemia cells. In the present study, we analyzed the expression of RHAMM in 48 patients suffering from acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS). Furthermore, we correlated transcripts with the clinical course of the disease before and after treatment. Real-time quantitative reverse transcriptase polymerase chain reaction was performed from RNA of peripheral blood mononuclear cells. T cell responses against RHAMM were assessed by tetramer staining (flow cytometry) and enzyme-linked immunospot (ELISPOT) assays. Results were correlated with the clinical outcome of patients. The results of the present study showed that almost 60% of the patients were RHAMM positive; specific T-cells recognizing RHAMM could be detected, but they were nonfunctional in terms of interferon gamma or granzyme B release as demonstrated by ELISPOT assays. Immunotherapies like peptide vaccination or adoptive transfer of RHAMM-specific T cells might improve the immune response and the outcome of AML/MDS patients.

Show MeSH
Related in: MedlinePlus