Inhibition of mitogen‐activated protein kinase pathway can induce upregulation of human leukocyte antigen class I without PD‐L1‐upregulation in contrast to interferon‐γ treatment

Abstract

Recently, we reported that human leukocyte antigen (HLA) class I expression is predominantly regulated by the mitogen-activated protein kinase (MAPK) pathway as one of the oncogenic regulations of HLA class I expression. In the present study, we examined mechanisms of how HLA class I and PD-L1 are regulated by MAPK inhibitors and interferon-γ (IFN-γ). Furthermore, we evaluated the expression of major signal transduction molecules by Western blot and anti-tumor CTL activity by a cytotoxic assay when HLA class I and PD-L1 were modulated by MAPK inhibitors and/or IFN-γ. As a result, we confirmed, as a more general phenomenon, that the inhibition of MAPK could upregulate HLA class I expression in a panel of human solid tumors (n = 26). Of note, we showed that MAPK inhibitors act on the upregulation of HLA class I expression through a different pathway from IFN-γ; there was an additive effect in the upregulation of HLA class I when treated with the combination of MAPK inhibitors and IFN-γ, and there was no overlapping activation of JAK2/STAT1 and Erk1/2 molecules when treated with either IFN-γ or MAPK inhibitors. Furthermore, we showed that IFN-γ–treatment impaired the tumor-specific CTL activity due to the upregulation of PD-L1 in spite of the upregulation of HLA class I, while MAPK inhibitors can augment the tumor-specific CTL activity due to the upregulated HLA class I without PD-L1 alterations. In conclusion, in addition to the original anti-proliferative activity, MAPK inhibitors may work toward the enhancement of T-cell-mediated anti-tumor immunity through the upregulation of HLA class I without the upregulation of PD-L1.

PD‐L1 expression is mainly regulated by interferon gamma associated with JAK‐STAT pathway in gastric cancer

“…To evaluate the effect of the PD‐1/PD‐L1 interaction on anti‐tumor CTL activity, HLA‐A24‐positive, LY6K‐positive tumor cells, namely MKN7 and TE1, were pre‐treated with or without IFN‐γ for 48 hour. The cells were then treated with anti‐PD‐L1 mAb or isotype control for 1 hour and subjected to cytotoxic assay.…”

“…An isotype‐matched immunoglobulin served as a negative control, and staining was detected using an LSRII flow cytometer (Becton Dickinson). Dead and/or apoptotic cells were excluded using Annexin‐V and 7‐Aminoactinomycin D. The relative mean fluorescence intensity (rMFI) was calculated according to the formula: [(MFI with specific mAb − MFI with isotype mAb)/MFI with isotype mAb]/[(MFI with specific mAb of control treatment − MFI with isotype mAb of control treatment)/MFI with isotype mAb of control treatment] …”

“…An HRP‐linked anti‐rabbit IgG (Cell Signaling Technology) was used as secondary antibody. All samples were prepared and stained with antibodies as previously described, and blots were visualized by ECL Prime (Amersham Pharmacia, Uppsala, Sweden) and with a Konica SRX‐101A film processor (JZ Imaging & Consulting, Willoughby, OH, USA).…”

“…Cytotoxicity was measured using a calcein‐release assay, as previously described . Briefly, target cells were incubated with 5 μmol L −1 of calcein‐AM (Dojindo Laboratories, Kumamoto, Japan) for 30 minute at 37°C and 5% CO 2 .…”

IFN-γ-mediated hematopoietic cell destruction in murine models of immune-mediated bone marrow failure

“…36 IFN-g sensitizes target cells to killing by diverse mechanisms, including modulation of HLA expression, 62,63 production of nitric oxide, 3 activation of caspase death pathways, [64][65][66] and induction of Fas expression. [44][45][46]67 We previously reported that Fas/FasL is the major cell death pathway for marrow destruction For personal use only.…”