P715: tox regulated cd8+ t lymphocytes exhausted in myelodysplastic syndromes patients through activating akt/pi3k/mtor pathway

Background: CD8+ T lymphocytes of myelodysplastic syndromes (MDS) patients are frequently exhausted, which plays a key role in the pathogenesis of MDS. Thymocyte selection-related high mobility box protein (TOX) programs CD8+ T lymphocyte exhaustion. However, the role of TOX in the CD8+ T lymphocytes of MDS patients is not clear. Aims: The aim of this study is to investigate the role of TOX in the CD8+ T lymphocytes of MDS patients. Methods: The expression of TOX, inhibitory receptors (PD-1, TIGIT, TIM3, LAG3, and CTLA4), and functional molecules (Perforin and Granzyme B) in peripheral blood T lymphocytes of MDS patients and normal controls was detected by flow cytometry. Lentiviral transduction for stable TOX-knockdown CD8+ T lymphocytes and small interfering RNA (siRNA) for TOX-knockdown Jurkat was used to investigate the relevant mechanisms. Results: The expression of TOX was enriched in CD3+CD8+ lymphocytes of MDS patients and increased gradually with the increase of risk degree (31.82% ± 12.20% in controls, 52.22% ± 15.61% in LR-MDS, 55.32% ± 19.56% in HR-MDS) (A). Moreover, the upregulation of TOX was associated with the upregulation of inhibitory receptors (PD-1, TIGIT, TIM3, LAG3, and CTLA4) and the downregulation of functional molecules such as perforin (42.27% ± 15.24% in controls, 42.50% ± 10.03% in LR-MDS, 31.20% ± 12.93% in HR-MDS) and granzyme B (35.27% ± 11.49% in controls, 32.45% ± 13.91% in LR-MDS, 27.99% ± 11.78% in HR-MDS) (B). These findings suggest that TOX plays a crucial role in the dysfunction of CD8+ T lymphocytes in MDS patients. To further investigate the role of TOX in CD8+ T lymphocytes, we knocked down TOX expression using lentiviral transduction and small interfering RNA (siRNA). We found that targeting TOX partially reversed the exhausted phenotype of CD8+ T lymphocytes and enhanced the lethality of SKM-1 cell lines (C). In addition, we found that knocking down the expression of TOX with siRNA in Jurkat cell lines improved cell proliferation activity, downregulated inhibitory receptors, and activated the AKT/PI3K/mTOR signaling pathway (D). Finally, we found that the upregulation of TOX was associated with shorter survival in MDS patients (23.9 months VS 34.5 months, HR = 3.989, P = 0.047), suggesting that TOX could be a potential prognostic biomarker for MDS. Summary/Conclusion: CD8+ T lymphocytes of myelodysplastic syndromes (MDS) patients are frequently exhausted, which plays a key role in the pathogenesis of MDS. Thymocyte selection-related high mobility box protein (TOX) programs CD8+ T lymphocyte exhaustion. However, the role of TOX in the CD8+ T lymphocytes of MDS patients is not clear.Keywords: Akt, Myelodysplastic syndrome, CD8 T cells