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T cells affect the prognosis of myelofibrosis
tinib treatment have been associated with improved infection- free survival.39 However, while neoplastic myeloid cells are thought to be the main target of JAK inhibitors, ruxolitinib also exerts a potent effect on non-malignant immune cells. The pres- ent study is the first to demonstrate that long-term treatment with ruxolitinib repolarizes activated T cells in MF patients. This finding is in agreement with previous in vitro studies demonstrat- ing decreased cytokine production in T cells from ruxolitinib- treated MF patients,11 and decreased activation, proliferation and function of T cells from normal individuals.9 In addition, our observation of decreased CD4+ cell subsets as a result of ruxoli- tinib therapy confirms the findings of a previous study that showed JAK inhibitor-induced decreases in the number and function of helper T cells.10 Similar suppressive effects of ruxoli- tinib have been observed in NK cells from MF patients.40
Importantly, we observed increased PD1+ fractions among all assessed subsets, indicating that T cells are functionally exhaust- ed in MF. These findings are in agreement with a recent study that showed increased PDL1 expression on myeloid cells from patients with JAK2-mutated myeloproliferative neoplasms.19 Our findings also corroborate PD1 expression patterns previous- ly reported in circulating CD4+ and CD8+ cells of patients with MF.20 In the aforementioned study, however, lack of any T-cell- specific markers in the gating strategy employed makes the reported results difficult to interpret. In our study, we specifical- ly analyzed CD45+/CD3+/αβ+ T cells and assessed PD1-express- ing cell fractions across the different activation subsets. T-cell exhaustion is typically manifested by a progressive defect in production of interferon-γ, IL-2, and tumor necrosis factor; T cells incapable of releasing these cytokines have been implicated in promoting the differentiation of monocytes into fibrocytes.41 It remains to be established how T-cell dysfunction affects the population of neoplastic fibrocytes, which induce BM fibrosis in MF.42 Conversely, there was no significant difference in the expression of either PD1 in T cells or PDL1 in blasts of patients with newly diagnosed acute myeloid leukemia, chronic myelomonocytic leukemia or myelodysplastic syndromes,37,43,44 suggesting that the neoplastic clone in MF exerts stronger immunogenicity with a significantly dysfunctional capacity as compared with other myeloid mallignancies.
In our cohort of MF patients, monocytosis and thrombo- cythopenia were associated with a predominantly CD8+ T-cell phenotype. In addition, high levels of CD8+ cells and increased PD1+ fractions within the CD8 compartment correlated with disease progression and poor outcome. Although, we did not
observe that ruxolitinib altered the percentage of PD1+ T cells, the survival of ruxolitinib-treated patients with low PD1 levels was significantly improved, suggesting that T-cell dysfunction is associated with a poor response to treatment with ruxolitinib. Remarkably, like in MF, in acute lymphoblastic leukemia, a dis- ease in which the JAK-STAT pathway is often constitutively activated,45-47 low numbers of PD1+ T cells predicted an improved treatment outcome. Because the spleen is a T-cell reservoir, and spleen size correlated with PD1+ fractions, where- as a low CD8+/PD1+ T-cell percent correlated with a favorable response to ruxolitinib treatment, it is likely that T-cell exhaus- tion plays a role in the pathogenesis of MF and the response to JAK-inhibitor treatment.
Collectively, our data suggest that both cytotoxic and helper T cells in MF are overly activated and harbor increased PD1+ frac- tions. Long-term JAK inhibition reverses terminal T-cell overac- tivation; nonetheless, high levels of PD1-expressing CD8+ T cells result in poor survival. A further in-depth analysis of the innate immune system, including the heterogeneous T-cell populations and their interaction with the MF neoplastic myeloid cells, is warranted.
Disclosures
SV receives research funding from Incyte Corporation, Wilmington, DE, USA. The remaining authors declare that they have no competing financial interests.
Contributions
IV analyzed and interpreted data, performed the statistical analyses, created the figures, and wrote the manuscript; SP ana- lyzed and interpreted data; SP and TM carried out the experi- ments; GMNG performed the statistical analyses; SV directed the project, supervised the study, and treated the patients included in the study; and ZE conceived, designed and supervised the study, interpreted data, and wrote the manuscript. All authors provided critical feedback and helped to develop the final manuscript.
Acknowledgments
The authors thank the Hanns A. Pielenz Foundation for financial support. The authors acknowledge Helen T. Chifotides for scientific editing assistance. This work was performed in part in the Flow Cytometry and Cellular Imaging Core Facility and used the Biostatistics Resource Group; both are supported by the National Cancer Institute, National Institutes of Health under award number P30 CA016672.
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