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[Assessment of P70S6 kinase phosphorylation after liver transplantation by phospho-flow cytometry]
1Department of Emergency Medicine, Beijing Chaoyang Hospital, Capital Medical University, Beijing 100020, China.
Objective:
To explore the role of P70S6 kinase phosphorylation as a downstream of mammalian target of rapamycin (mTOR) pathway in CD3 positive cells of liver transplant patients.
Methods:
A total of 84 liver transplant recipients were recruited from our hospital and divided into 3 treatment groups of sirolimus (n = 26), tacrolimus (n = 35) and cyclosporine (n = 23). The P70S6 kinase phosphorylation of CD3 positive cell of patients and healthy control (HC) were analyzed by phospho-flow cytometry. A correlation analysis between P70S6 kinase phosphorylation and sirolimus trough level was performed. Intra-individual variability and inter-individual variability of P70S6 kinase phosphorylation were measured.
Results:
The P70S6 kinase phosphorylation in HC showed a low degree of intra-individual variability (3.5% to 5.6%) while the inter-individual variability between different healthy volunteers was higher (18.9% to 22.5%). The P70S6 kinase phosphorylation in patients treated with sirolimus (28.9 ± 10.5) was significantly lower than in HC (57.2 ± 8.4, P < 0.001), tacrolimus (42.5 ± 14.1, P < 0.001) or cyclosporine treated ones (51.4 ± 10.9, P < 0.001). The P70S6 kinase phosphorylation in HC (57.2 ± 8.4) was significantly higher than in tacrolimus (42.5 ± 14.1, P < 0.01) or cyclosporine treated patients (51.4 ± 10.9, P < 0.05). No correlation existed between P70S6 kinase phosphorylation and trough level of sirolimus (r = -0.18, P = 0.39).
Conclusion:
Phospho-flow cytometry assay has been established for determining the degree of mTOR inhibition by assessing P70S6 kinase phosphorylation. Quantification of P70S6 kinase phosphorylation may play an adjunct role in pharmacodynamically guiding an individualized mTOR inhibitor based immunosuppression.
Insights
P70S6 kinase phosphorylation in liver transplant patients varied by immunosuppressant. Sirolimus significantly reduced phosphorylation, suggesting its potent mTOR pathway inhibition, while tacrolimus and cyclosporine showed less impact. This assay can guide personalized immunosuppression.
Area of Science:
- Immunology
- Pharmacology
- Transplantation
Background:
- The mammalian target of rapamycin (mTOR) pathway is crucial in regulating immune cell function.
- Assessing downstream markers like P70S6 kinase phosphorylation can evaluate mTOR inhibition by immunosuppressants.
- Understanding these pathways is vital for optimizing immunosuppression in liver transplant recipients.
Purpose of the Study:
- To investigate the role of P70S6 kinase phosphorylation as a downstream marker of the mTOR pathway in CD3+ cells from liver transplant patients.
- To compare P70S6 kinase phosphorylation levels across different immunosuppressive drug regimens (sirolimus, tacrolimus, cyclosporine).
Main Methods:
- Phospho-flow cytometry was used to analyze P70S6 kinase phosphorylation in CD3+ cells from 84 liver transplant recipients and healthy controls.
- Patients were grouped based on their immunosuppressive therapy: sirolimus, tacrolimus, or cyclosporine.
- Correlation analysis was performed between P70S6 kinase phosphorylation and sirolimus trough levels.
Main Results:
- P70S6 kinase phosphorylation exhibited low intra-individual variability but higher inter-individual variability in healthy controls.
- Sirolimus treatment significantly reduced P70S6 kinase phosphorylation compared to healthy controls, tacrolimus, and cyclosporine.
- Healthy controls showed higher P70S6 kinase phosphorylation than patients treated with tacrolimus or cyclosporine.
- No significant correlation was found between P70S6 kinase phosphorylation and sirolimus trough levels.
Conclusions:
- A phospho-flow cytometry assay for P70S6 kinase phosphorylation effectively measures mTOR inhibition.
- Quantifying P70S6 kinase phosphorylation may aid in the pharmacodynamic guidance of individualized mTOR inhibitor-based immunosuppression.
- This method offers potential for optimizing immunosuppressive therapy in liver transplant patients.
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