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Effects of Ruxolitinib on Immune Checkpoint Molecule Expression in JAK2 V617F-Positive Cells
Background:
This study aimed to explore the clinical significance of ruxolitinib and its effects on the proliferation and apoptosis of human erythroleukemia (HEL) cells and the expression of immune checkpoint molecules programmed death-1 (PD-1), programmed death-ligand 1 (PD-L1), and regulatory T cells (Tregs) in HEL cells and JAK2 V617F-positive patients with myeloproliferative neoplasms (MPNs).
Methods:
JAK2 V617F-positive patients with MPNs admitted to the Baoding No. 1 Hospital from January 2016 to September 2023 were recruited, including 30 patients for the newly diagnosed group and 10 for the treatment group. Additionally, 15 healthy volunteers were selected as the control group. JAK2 V617F mutation was detected by using fluorescence quantitative PCR, and the expression levels of phosphorylated JAK2 (p-JAK2), PD-1, and PD-L1 in fresh bone marrow were examined by immunohistochemistry. HEL cells were treated with ruxolitinib at different concentrations (0, 50, 100, 250, 500, and 1,000 nmol/L). Cell viability was detected by CCK-8 assay. The mRNA expression levels of JAK2, PD-1, and PD-L1 were determined by using fluorescence quantitative PCR. The protein expression of p-JAK2 was detected by Western blot and those of PD-1 and PD-L1 were evaluated by flow cytometry. The expression of PD-1, PD-L1, and Tregs after the 48-hour co-culture of primary bone marrow cells and HEL cells were also analyzed by flow cytometry.
Results:
In the newly diagnosed group, the bone marrow myeloid cells highly expressed p-JAK2, PD-1, and PD-L1. The Tregs expression in their peripheral blood increased and was significantly higher than those in the treatment and control groups (all p < 0.05). Ruxolitinib at different concentrations could inhibit the proliferation of HEL cells and was positively correlated with treatment time and dose. Additionally, ruxolitinib could reduce p-JAK2, PD-1, and PD-L1 expression in HEL cells and Tregs expression.
Conclusions:
Ruxolitinib reduces the expression of p-JAK2, PD-1, and PD-L1 in JAK2 V617F-positive cells by specifically inhibiting the JAK2 signaling pathway, thereby suppressing the progression of MPNs.
Insights
Ruxolitinib effectively inhibits proliferation and reduces immune checkpoint molecule expression in JAK2 V617F-positive myeloproliferative neoplasms. This targeted therapy suppresses programmed death-1 (PD-1) and programmed death-ligand 1 (PD-L1) in cancer cells and regulatory T cells (Tregs).
Area of Science:
- Hematology
- Oncology
- Immunology
Background:
- Myeloproliferative neoplasms (MPNs) are characterized by the JAK2 V617F mutation.
- Immune checkpoint molecules like programmed death-1 (PD-1) and programmed death-ligand 1 (PD-L1) play a role in MPN pathogenesis.
- Regulatory T cells (Tregs) are implicated in immune suppression within the tumor microenvironment.
Purpose of the Study:
- To investigate the clinical significance of ruxolitinib in MPNs.
- To evaluate the effects of ruxolitinib on human erythroleukemia (HEL) cell proliferation and apoptosis.
- To assess ruxolitinib's impact on PD-1, PD-L1, and Tregs expression in HEL cells and MPN patients.
Main Methods:
- Recruited JAK2 V617F-positive MPN patients and healthy volunteers.
- Detected JAK2 V617F mutation and measured p-JAK2, PD-1, and PD-L1 expression in bone marrow.
- Treated HEL cells with ruxolitinib and analyzed cell viability, mRNA, and protein expression of key molecules.
Main Results:
- Newly diagnosed MPN patients showed high expression of p-JAK2, PD-1, and PD-L1, with increased Tregs.
- Ruxolitinib significantly inhibited HEL cell proliferation in a dose- and time-dependent manner.
- Ruxolitinib treatment reduced p-JAK2, PD-1, PD-L1 expression in HEL cells, and Tregs levels.
Conclusions:
- Ruxolitinib effectively inhibits JAK2 signaling, reducing p-JAK2, PD-1, and PD-L1 expression in JAK2 V617F-positive cells.
- This targeted inhibition suppresses MPN progression by modulating key immune checkpoint molecules and Tregs.
- Ruxolitinib demonstrates significant clinical potential for treating JAK2 V617F-positive MPNs.
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