Deciphering tumor-specific T cells in Acute Myeloid Leukemia – DTSTAML

This project aims to analyze the composition, antigenic specificity, and function of CD8+ T lymphocytes in the peripheral blood and bone marrow of AML patients. For phenotyping, we use mass cytometry (CyTOF), which allows simultaneous characterization of a large number of surface and intracellular markers, identifying distinct and functional cell subpopulations, such as CD69+ cells enriched in the bone marrow. Antigen specificity is assessed using MHC I tetramer staining, which precisely detects CD8+ T cells recognizing viral or tumor antigens, including those derived from EBV and CMV. To study cellular function, we combine cytokine production analyses, particularly IL-8, with cytotoxicity and granzyme K secretion assays to understand the role of these cells in inflammation and AML progression. Finally, ex vivo cell cultures are established to reproduce the bone marrow microenvironment and test the effect of these lymphocytes on leukemic cells, allowing assessment of their direct impact on cell survival and modulation of inflammation.

In contrast to what is observed in many solid tumors, where CD8+ T lymphocytes often display an “exhausted” phenotype that limits their anti-tumor activity, this study did not identify exhausted CD8+ T cells in patients with acute myeloid leukemia (AML). Instead, a distinct population of functional CD8+ T cells expressing the activation marker CD69 was specifically enriched in the bone marrow. These cells exhibit an active phenotype and appear capable of responding to immune signals, suggesting they are not functionally inert. Interestingly, this population does not primarily target leukemic cells but rather recognizes non-tumor antigens, including epitopes derived from common viruses such as Epstein–Barr virus (EBV) and cytomegalovirus (CMV). This type of T cell, referred to as a “bystander,” seems to play an indirect role in the bone marrow microenvironment rather than directly eliminating cancer cells.

A key feature of this observation is the high expression of granzyme K by these bystander CD8+ T cells. Granzyme K is a cytotoxic molecule that, in this context, does not induce leukemic cell death. Instead, it promotes the secretion of pro-inflammatory cytokines, particularly IL-8, which is known to play a deleterious role in AML pathophysiology. IL-8 contributes to creating a pro-inflammatory microenvironment in the bone marrow, potentially supporting leukemic cell survival and disease progression. Thus, rather than functioning as anti-tumor effectors, these CD8+ T cells contribute to an inflammatory state that may exacerbate the severity and aggressiveness of AML.

These findings reveal a paradoxical mechanism in which CD8+ T lymphocytes, traditionally considered protective in the fight against cancer, can actually support a harmful pro-inflammatory microenvironment. Understanding this phenomenon provides a new perspective for developing therapeutic strategies aimed at targeting this cell population or modulating granzyme K activity, with the goal of reducing deleterious inflammation and limiting AML progression. By exploring these avenues, it becomes possible to envision interventions that restore a beneficial immune balance, potentially transforming the immune system from a disease-promoting factor into an ally for the patient.

Acute Myeloid Leukemias (AML) are a heterogeneous group of blood cancers. Recently, the presence of neoantigens tumor-specific T cells has been reported, opening a new area of research in AML. Our project will explore whether these T cells response can be broadly identified in AML using advances methodologies, to determine which AML patients could be a good candidate for immunotherapies targeting T cells. To this end, two strategies will be used: 1- Identification and characterization of tumor-specific T cells using high throughput screening approaches for tumor epitopes (i.e. neoantigens, tumor-associated antigens). 2- Identification of tumor-specific T cells through their TCR repertoire using a multi-omics approach. These two strategies will be applied in blood and bone marrow of AML patients at diagnosis, and after treatment (remission and/or relapse). This project will allow us to better understand tumor-specific T cells response in AML patients (e.g. shared tumor epitopes and tumor specific TCR, phenotypic characterisation of tumor-specific T cells) and determined if innovative therapeutic approach could be developed in AML treatment(e.g. TCR transgenic T cells, neoantigens vaccination).