PF366 CXCR5+PD‐1+ CD8+ T‐CELLS IN HUMAN BLOOD AND LYMPH NODES
Authors: Hofland T. , Remmerswaal E.B. , Bemelman F. , Levin M.‐D. , Kater A.P. , Tonino S.H. 
Published: June 2019

Background:Re‐invigorating exhausted T‐cells through immune checkpoint blockade (ICB) has shown promising activity in several malignancies. However, response rates vary between malignancies, and it is unclear which parameters predict response to ICB. Recently, a subset of CXCR5+PD‐1+ CD8+ T‐cells has been identified in mice. This memory‐like population appears to be crucial for the expansion of CD8+ T‐cells during ICB, and is characterized by expression of the transcription factor Tcf1. CXCR5+ CD8+ T‐cells have also been identified in humans, but it's currently unknown whether this distinctive CD8+ T cell subset has a similar function as described in mice, and plays an equally important role during ICB.Aims:To investigate the human CXCR5+PD‐1+ CD8+ T‐cell subset, we studied peripheral blood and lymph node samples derived from healthy donors and patients with a variety of B cell malignancies.Methods:PB and LN samples were analyzed by flow cytometry.Results:The frequency of CXCR5+PD‐1+ CD8+ T‐cells was higher in lymph nodes. CXCR5+PD‐1+ CD8+ T‐cells resemble antigen‐experienced memory‐like cells, with high expression of KLRG1, CD127, granzyme K, and Eomes, and low expression of effector molecules granzyme B and T‐bet. They express high levels of Tcf1, in accordance with the phenotype described in mice. CXCR5+PD‐1+ CD8+ T‐cells were present within the EBV‐specific T‐cell population, especially directed against epitopes expressed during viral latency. In line with mice data, we could not detect CXCR5+PD‐1+ CD8+ T‐cells within the FLU‐specific T‐cell compartment, suggesting that chronic antigen stimulation is required for this population to develop. On a functional level, CXCR5+PD‐1+ CD8+ T cells produce effector cytokines IL‐2, IFNγ and TNFα after stimulation, in line with their memory‐like phenotype. CXCR5+PD‐1+ CD8+ T cells proliferate and differentiate into effector CD8+ T cells, with higher expression of granzyme B and loss of Tcf1.We assessed the role of CXCR5+PD‐1+ CD8+ T cells in ICB by analyzing their phenotype in samples derived from patients with various B cell malignancies with differential clinical response rates to ICB. In chronic lymphocytic leukemia (CLL), a disease with no objective response to ICB, CXCR5+PD‐1+ CD8+ T cells express lower levels of Tcf1 and markers compatible with an advanced differentiation state, and express lower levels of PD‐1. We did not observe these changes in CXCR5+PD‐1+ CD8+ T cells in B cell malignancies with better response rates to ICB, like DLBCL, multiple myeloma, and follicular lymphoma. Currently, we perform functional experiments in the context of checkpoint blockade to verify whether this changed phenotype is related to the disappointing response towards ICB in CLL. To explore the potential role of CXCR5+PD‐1+ CD8+ T cells for ICB therapy further, we are analyzing peripheral blood samples derived from cancer patients who receive ICB therapy while also studying the localization and frequency of CXCR5+PD‐1+ CD8+ T‐cells in relation to tumor cells within a spectrum of malignant tissues by confocal microscopy.Summary/Conclusion:In conclusion, we find that on a phenotypical and functional level, human CXCR5+PD‐1+ CD8+ T‐cells closely resemble this subset in mice. In CLL, in which ICB therapy does not lead to clinical responses, CXCR5+PD‐1+ CD8+ T‐cells have an altered phenotype which might explain the lack of response to ICB. Furthermore, if human CXCR5+PD‐1+ CD8+ T‐cells indeed play a similar important role in the response to ICB therapy as in mice, they could potentially serve as a biomarker for clinical response in human malignancies.