Signatures of CD8+ T cell dysfunction in AML patients and their reversibility with response to chemotherapy
Hanna A. Knaus, Sofia Berglund, Hubert Hackl, Amanda L. Blackford, Joshua F. Zeidner, Raúl Montiel-Esparza, Rupkatha Mukhopadhyay, Katrina Vanura, Bruce R. Blazar, Judith E. Karp, Leo Luznik, Ivana Gojo
Hanna A. Knaus, Sofia Berglund, Hubert Hackl, Amanda L. Blackford, Joshua F. Zeidner, Raúl Montiel-Esparza, Rupkatha Mukhopadhyay, Katrina Vanura, Bruce R. Blazar, Judith E. Karp, Leo Luznik, Ivana Gojo
View: Text | PDF
Clinical Research and Public Health Hematology Immunology

Signatures of CD8+ T cell dysfunction in AML patients and their reversibility with response to chemotherapy

Abstract

BACKGROUND. Our understanding of phenotypic and functional signatures of CD8+ T cell dysfunction in acute myeloid leukemia (AML) is limited. Deciphering these deranged T cell functional states and how they are impacted by induction chemotherapy is essential for incorporation of novel immune-based strategies to restore and maintain antileukemia immunity. METHODS. We utilized high-dimensional immunophenotyping, gene expression, and functional studies to characterize peripheral blood and bone marrow CD8+ T cells in 72 AML patients at diagnosis and after induction chemotherapy. RESULTS. Our data suggest that multiple aspects of deranged T cell function are operative in AML at diagnosis, with exhaustion and senescence being the dominant processes. Following treatment, the phenotypic and transcriptional profile of CD8+ T cells diverged between responders and nonresponders. Response to therapy correlated with upregulation of costimulatory, and downregulation of apoptotic and inhibitory, T cell signaling pathways, indicative of restoration of T cell function. In functional studies, AML blasts directly altered CD8+ T cell viability, expansion, co-signaling and senescence marker expression. This CD8+ T cell dysfunction was in part reversible upon PD-1 blockade or OX40 costimulation in vitro. CONCLUSION. Our findings highlight the uniqueness of AML in sculpting CD8+ T cell responses and the plasticity of their signatures upon chemotherapy response, providing a compelling rationale for integration of novel immunotherapies to augment antileukemia immunity. FUNDING. This work was supported by the Leukemia & Lymphoma Society grant no. 6449-13; NIH grants UM1-CA186691 and R01-HL110907-01; the American Society for Blood and Marrow Transplantation New Investigator Award/Gabrielle’s Angel Foundation; the Vienna Fund for Innovative Cancer Research; and by fellowships from the Wenner-Gren Foundation and the Swedish Society for Medical Research.

Authors

Hanna A. Knaus, Sofia Berglund, Hubert Hackl, Amanda L. Blackford, Joshua F. Zeidner, Raúl Montiel-Esparza, Rupkatha Mukhopadhyay, Katrina Vanura, Bruce R. Blazar, Judith E. Karp, Leo Luznik, Ivana Gojo

×

Figure 3

AML blasts alter viability and the expression of co-signaling molecules on CD8+ T cells.

Options: View larger image (or click on image) Download as PowerPoint
AML blasts alter viability and the expression of co-signaling molecules ...
Sorted PB T cells from AML patients (n = 20) and HCs (n = 18) were stimulated with anti–CD3/CD28 beads alone or with IL-2 (50 IU/ml) for 3 days. Stimulated T cells were either cultured alone or with autologous blasts (T cells + blasts; allogeneic blasts for HCs) in a 1:10 T cell/blast ratio. (A–C) Forest plots represent the ratio of log-scale mean percentages of CD8+ T cells expressing each marker as assessed by flow cytometry on day +3, comparing (A) AML patients to HCs, (B) CD8+ T cells cultured with or without IL-2, and (C) CD8+ T cells cultured with or without blasts. P values for the ratios being different from 1.0 are shown to the right and were calculated using (A) standard and (B and C) mixed-effects linear regression models. *P < 0.05 for the effect of (B) IL-2 and (C) blasts between the AML patients and HC. (D) CITRUS analysis of flow cytometry data on day +3 for AML patients and HCs. Depicted are cell abundances in clusters identified by CITRUS, stratified by the presence or absence of blasts. P values were calculated using Mann–Whitney U or Wilcoxon’s signed-rank test as appropriate. (E) bh-SNE map of PB CD8+ T cells expressing ICOS and OX40 upon 3-day stimulation in the presence or absence of blasts, both for HCs and AML patients. Each point in the bh-SNE map represents an individual cell, and the cells are colored according to the intensity of expression of the individual marker as indicated on the color scale. CIMminer software was used to summarize the MFI of ICOS and OX40 expression in heatmaps. P values were calculated using paired t test. (F) Fold expansion of AML (n = 10) and HC (n = 8) CD3+ T cells on day +6 upon culture with blasts and stimulation with anti–PD-1 mAb and OX40 ligand. Dashed horizontal line represents cell count on day 0. P values were calculated using mixed-effects linear regression model for the log-scale fold induction as a function of cell source (AML patients and HCs) and stimulator. (G) bh-SNE maps of Ki67 distribution on stimulated HC and AML PB CD8+ T cells upon culture alone, or with blasts ± anti–PD-1 mAb or ± OX40 ligand. The heatmap summarizes MFI of Ki67 expression. (H) Anti-CD33/CD3 BiTE–mediated cytotoxicity of CD8+CD57+ or CD8+CD57– T cells towards primary AML blasts (E/T ratio 1:1). Primary patient samples (n = 5) were cultured with BiTE or control BiTE (c-BiTE) for 48 hours. Experiments were run in duplicate and P values were calculated using paired t test.