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Research
Chun-Chia Cheng
Chang Gung University
Corresponding Author
ORCiD: https://orcid.org/0000-0002-6455-1798
License:
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Background
Tumor cells progress to evade immunological attacks and prohibit activity of CD8+ T cells. Irradiation damages tumor cells and augments tumor immunotherapy in clinical application. However, the detail mechanism remains elusive. We aimed to uncover the mechanism of irradiation augmenting cytotoxic CD8+ T cells to suppress tumor progression in non-small-cell lung cancer (NSCLC).
Methods
EGFR-positive NSCLC cell lines were co-cultured with isolated PBMCs from healthy volunteers, cell viability and apoptosis were measured. RNAseq was used to screen the IFNγ-mediated gene expression in A549 cells. Irradiation was used to augment PBMCs-mediated anti-tumor effect and the irradiation effect to IFNγ-mediated gene expression was investigated using qPCR and Western blots.
Results
Co-culture of tumor cells stimulates increase of granzyme B and IFNγ in CD8+ T, but A549 exhibits resistance against CD8+ T cytotoxicity. Irradiation inhibits A549 proliferation and enhances apoptosis, augmenting PBMCs-mediated cytotoxicity against A549. IFNγ simultaneously increased phosphorylation on STAT1 and STAT3 in EGFR-positive lung cancer, resulting in overexpression of PD-L1. In RNAseq analysis, MCL1 was identified and increased by IFNγ-STAT3 axis in A549 cells, we found that irradiation specifically inhibits phosphorylation on STAT1 and STAT3 in IFNr-treated A549, resulting in reductions of PD-L1 and MCL1. Moreover, knockdowns of STAT3 and MCL1 increased PBMCs against irradiated A549 cells.
Conclusion
This study demonstrated that A549 expressed MCL1 against CD8+ T cell-mediated apoptosis. In addition, we found that irradiation suppressed STAT3 phosphorylation and IFNγ-mediated PD-L1 and MCL1 expression, revealing a potential mechanism of irradiation augmenting immune surveillance.
Keywords
Non-small-cell lung cancer, STAT1, STAT3, PD-L1, MCL1, CD8+ T cells, irradiation, radiotherapy
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A new preprint design is coming!
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This is a preprint, a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Research
Chun-Chia Cheng
Chang Gung University
Corresponding Author
ORCiD: https://orcid.org/0000-0002-6455-1798
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Posted
Version 1
Posted 11 May, 2021
No community comments so far
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Background
Tumor cells progress to evade immunological attacks and prohibit activity of CD8+ T cells. Irradiation damages tumor cells and augments tumor immunotherapy in clinical application. However, the detail mechanism remains elusive. We aimed to uncover the mechanism of irradiation augmenting cytotoxic CD8+ T cells to suppress tumor progression in non-small-cell lung cancer (NSCLC).
Methods
EGFR-positive NSCLC cell lines were co-cultured with isolated PBMCs from healthy volunteers, cell viability and apoptosis were measured. RNAseq was used to screen the IFNγ-mediated gene expression in A549 cells. Irradiation was used to augment PBMCs-mediated anti-tumor effect and the irradiation effect to IFNγ-mediated gene expression was investigated using qPCR and Western blots.
Results
Co-culture of tumor cells stimulates increase of granzyme B and IFNγ in CD8+ T, but A549 exhibits resistance against CD8+ T cytotoxicity. Irradiation inhibits A549 proliferation and enhances apoptosis, augmenting PBMCs-mediated cytotoxicity against A549. IFNγ simultaneously increased phosphorylation on STAT1 and STAT3 in EGFR-positive lung cancer, resulting in overexpression of PD-L1. In RNAseq analysis, MCL1 was identified and increased by IFNγ-STAT3 axis in A549 cells, we found that irradiation specifically inhibits phosphorylation on STAT1 and STAT3 in IFNr-treated A549, resulting in reductions of PD-L1 and MCL1. Moreover, knockdowns of STAT3 and MCL1 increased PBMCs against irradiated A549 cells.
Conclusion
This study demonstrated that A549 expressed MCL1 against CD8+ T cell-mediated apoptosis. In addition, we found that irradiation suppressed STAT3 phosphorylation and IFNγ-mediated PD-L1 and MCL1 expression, revealing a potential mechanism of irradiation augmenting immune surveillance.
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7
This is a list of supplementary files associated with this preprint. Click to download.
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