Immune marker expression of irradiated mesothelioma cell lines

Our plain-English summary of Immune marker expression of irradiated mesothelioma cell lines, by F. Chang, S. Keam, T.S. Hoang, et al (Frontiers in Oncology, 2022).

The study

• Authors: F. Chang, S. Keam, T.S. Hoang, J. Creaney, S. Gill, A.K. Nowak, M. Ebert, A.M. Cook
• Journal: Frontiers in Oncology
• Published: 2022;12:1020493
• Cited by: 4 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Background: Though immune checkpoint inhibition has recently shown encouraging clinical efficacy in mesothelioma, most patients do not respond. Combining immune checkpoint inhibition with radiotherapy presents an attractive option for improving treatment responses owing to the various immunomodulatory effects of radiation on tumors. However, the ideal dosing and scheduling of combined treatment remains elusive, as it is poorly studied in mesothelioma. The present study characterizes the dose- and time-dependent changes to expression of various immune markers and cytokines important to antitumor responses following irradiation of mesothelioma cell lines.

Methods: Two murine (AB1, AE17) and two human (BYE, JU77) mesothelioma cell lines were treated with titrated gamma-radiation doses (1-8 Gy) and the expression of MHC class-I, MHC class-II and PD-L1 was measured over a series of post-irradiation timepoints (1-72 hours) by flow cytometry. Levels of cytokines IL-1α, IL-1β, IL-6, IL-10, IL-12p70, IL-17A, IL-23, IL-27, MCP-1, IFN-β, IFN-γ, TNF-α, and GM-CSF were measured by multiplex immunoassay in murine cell lines following 8 Gy radiation.

Results: Following irradiation, a dose-dependent upregulation of MHC-I and PD-L1 was observed on three of the four cell lines studied to varying extents. For all cell lines, the increase in marker expression was most pronounced 72 hours after radiation. At this timepoint, increases in levels of cytokines IFN-β, MCP-1 and IL-6 were observed following irradiation with 8 Gy in AB1 but not AE17, reflecting patterns in marker expression.

Conclusions: Overall, this study establishes the dose- and time-dependent changes in immune marker expression of commonly studied mesothelioma cell lines following radiation and will inform future study into optimal dosing and scheduling of combined radiotherapy and immune checkpoint inhibition for mesothelioma.

Copyright © 2022 Chang, Keam, Hoang, Creaney, Gill, Nowak, Ebert and Cook.

In plain English

This laboratory study used mesothelioma cells grown in culture. It did not study people or asbestos exposure. Researchers in Perth, Australia, gave two mouse mesothelioma cell lines (AB1, AE17) and two human lines (BYE, JU77, grown from patients' pleural fluid) single gamma-radiation doses of 1 to 8 Gy. They measured three immune-related surface markers (MHC-I, MHC-II and PD-L1) from 1 to 72 hours later. They also measured inflammatory cytokines in the mouse lines 72 hours after 8 Gy. Radiation increased MHC-I and PD-L1 on AB1, AE17 and BYE cells, more at higher doses and mainly at 72 hours. In AB1, 72 hours after 4 Gy, 67% of cells were PD-L1-positive, compared with 11.5% in unirradiated controls. Radiation did not substantially change MHC-I or significantly change PD-L1 on JU77, and it induced MHC-II on no line. Increases in IFN-beta, MCP-1 and IL-6 in AB1 were not statistically significant. The authors concluded that the findings will inform future study of combining radiotherapy with immune checkpoint inhibitor drugs. They noted that only single radiation doses were tested.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory (in vitro) experiment on cultured mesothelioma cell lines, with each irradiation done in triplicate
• Who was studied: Four mesothelioma cell lines: two murine (AB1, AE17) and two human (BYE10412, called BYE, and JU77). The human lines were generated from patient malignant pleural effusions and obtained from the National Centre for Asbestos Related Disease biobank in Perth, Australia.
• Exposure: Single doses of gamma radiation (1, 2, 4, 6 and 8 Gy) from a caesium-137 source. Cells were assessed at 1, 6, 24, 48 and 72 hours afterwards. Asbestos fibers were not an exposure in this study.
• Main results:
  • Before irradiation, AB1 cells were 95% positive for MHC-I and 21% positive for PD-L1. AE17 cells had negligible levels (2.6% and 1.2%).
  • In AB1 cells, MHC-I level (median fluorescence intensity) rose 2.5-fold 72 hours after 4 Gy (p = 0.0345). This was not significantly different from the effect of 8 Gy.
  • In AE17 cells, 8 Gy was needed to increase MHC-I. At 72 hours the percentage of MHC-I-positive cells was 1.91% in unirradiated controls versus 9.15% after 8 Gy (p = 0.0015), which the authors called only slightly higher.
  • In human BYE cells, 72 hours after irradiation, MHC-I-positive cells reached 35.0% after 6 Gy and 42.9% after 8 Gy (both p less than 0.0001). There was no significant difference between 6 and 8 Gy.
  • Radiation did not substantially alter MHC-I on the human JU77 line. It did not induce MHC-II on any of the four cell lines at any dose.
  • In AB1 cells 72 hours after 4 Gy, PD-L1-positive cells were 67% versus 11.5% in unirradiated controls (p less than 0.0001). PD-L1 level rose 3.3-fold after 4 Gy, which was not significantly different from the 3.9-fold rise after 8 Gy.
  • In AE17 cells, 1 to 6 Gy did not significantly alter PD-L1, but PD-L1 was higher after 8 Gy (p less than 0.05), only at 72 hours. BYE cells showed a similar PD-L1 increase after doses of 6 Gy and above. Radiation did not significantly alter PD-L1 on JU77.
  • Where marker changes were seen, they occurred maximally, and in most cases only, at the 72-hour timepoint.
  • In the mouse lines, 10 of the 13 cytokines measured were not detected before or after radiation. In AB1, IFN-beta, MCP-1 and IL-6 increased after 8 Gy, but the increase was not statistically significant. These cytokines were not upregulated in AE17.
• The authors' conclusion: The authors concluded that the study establishes dose- and time-dependent changes in immune marker expression of commonly studied mesothelioma cell lines after radiation. They said this will inform future study into the optimal dosing and scheduling of combined radiotherapy and immune checkpoint inhibition for mesothelioma.
• Limitations the authors state:
  • Only single radiation doses were used, although radiotherapy in the clinic is commonly fractionated (given in multiple doses).
  • The experiments were in cells in culture. The authors noted that effects in living tumors (in vivo) will be affected by interactions within the wider tumor environment.
  • Other cytokines not measured in this study may also influence immune responses in mesothelioma.
  • Measurements stopped at 72 hours, so it is not known whether the marker changes last longer or are transient.

Topics

Medical terms in this study, explained in our Lexicon: Mesothelioma, Cytokine, PD-L1.

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