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Dual role of IFNγ in reprogramming the undifferentiated pleomorphic sarcoma cell line JBT19 towards cytotoxic chemotherapy and antitumor immunity

  • Authors:
    • Pavla Taborska
    • Dmitry Stakheev
    • Daniel Smrz
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    Affiliations: Department of Immunology, Second Faculty of Medicine, Charles University and University Hospital Motol, 150 06 Prague, Czech Republic
    Copyright: © Taborska et al. This is an open access article distributed under the terms of Creative Commons Attribution License.
  • Article Number: 78
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    Published online on: December 16, 2025
       https://doi.org/10.3892/ol.2025.15431
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Abstract

Soft tissue sarcomas are therapeutically challenging. Among soft tissue sarcoma subtypes, undifferentiated pleomorphic sarcoma (UPS) exhibits one of the most pronounced disparities between its comparatively higher responsiveness to immunotherapy and its limited responsiveness to conventional cytotoxic chemotherapy. The interplay between immunotherapy and cytotoxic chemotherapy is still largely unknown. Interferon‑γ (IFNγ) is a key player in antitumor immunity and contributes to the modulation of the tumor microenvironment, which impacts both immune and cancer cells. The mechanism by which this interplay can affect cancer cell chemosensitivity and immune sensitivity is difficult to predict. The present study aimed to investigate the interplay of IFNγ signaling in the UPS cell line JBT19. It was identified that IFNγ treatment significantly decreased the proliferation of JBT19 cells and increased the surface expression levels of cluster of differentiation (CD)44, CD47, CD95 (Fas), major histocompatibility complex (MHC)‑I and programmed death‑ligand 1 (PD‑L1). In addition, IFNγ strongly upregulated surface expression levels of MHC‑II and converted JBT19 cells into docetaxel‑resistant cells. The IFNγ‑induced changes were sustained but reversible after 3 weeks of cell culture without IFNγ. Regardless of IFNγ treatment, JBT19 cells could elicit and amplify the adaptive immune response in vitro. The in vitro JBT19‑reactive lymphocytes effectively eliminated both IFNγ‑treated and non‑treated JBT19 cells, thus overcoming IFNγ‑induced chemoresistance. To the best of our knowledge, the present study demonstrated a dual role of IFNγ towards cancer cell chemoresistance and immunostimulatory potential for the first time. The present study findings may have potential implications for combining immunotherapy with cytotoxic chemotherapy in cancer treatment in the future.
View Figures

Figure 1

Impact of IFNγ on the in vitro
proliferation of JBT19 cells and their expression levels of MHC-II,
CD44, CD47 and CD95 (Fas), and their sensitivity to docetaxel. (A)
Gating strategy of flow cytometry data. (B) Extracellular MHC-II
staining of 3-day treated JBT19 cells with the indicated
concentrations of IFNγ. (C) Cell number fold increase in
vehicle-treated JBT19 cells (No treatment, left panel) or 7-day
pre-treated and then cultured with IFNγ (200 ng/ml; IFNγ, right
panel). The evaluated are represented as means ± SEM. *P<0.05,
**P<0.01, ***P<0.001 and ****P<0.0001; n=3 independent
experiments; one-way ANOVA with Tukey's post hoc test). (D)
Calculated proliferation curves and pertinent doubling times from
the data shown in panel C. (E) Extracellular staining of 7-day
vehicle-treated (No treatment) or 7-day IFNγ-treated (IFNγ, 200
ng/ml) JBT19 cells. The cells were stained with specific antibodies
against CD44, CD47, CD95 (Fas) or FAP. In the top panels,
representative histograms are shown and the control (No stain) for
individual fluorochromes refers to staining with vehicle alone. In
the bottom panels, the evaluated data presented as means ± SEM are
shown and statistically significant differences between the groups
are indicated. **P<0.01, ***P<0.001 and ****P<0.0001. MTT
assay of (F) vehicle-treated JBT19 cells (No treatment), or (G and
H) 7-day IFNγ-treated (IFNγ, 200 ng/ml) JBT19 cells (7 days IFNγ)
exposed to docetaxel at the indicated concentrations for 3 days.
The calculated IC50 from three independent experiments
is shown. MFI, mean fluorescence intensity; MHC, major
histocompatibility complex; nM, nanomolar; FAP, fibroblast
activation protein.

Figure 2

IFNγ in JBT19 cells changes the cell
cycle, does not induce apoptosis and increases STAT1 expression
without inducing the surface expression levels of MDR-1 or
enhancing the expression levels of Bcl-2. (A) Gating strategy for
cell cycle analysis by flow cytometry using propidium iodide
staining. (B) Cell cycle evaluation in vehicle-treated (No
treatment) or 7-day treated (7 d) JBT19 cells with IFNγ (200
ng/ml). The data indicate the proportions of JBT19 cells in the
G1, S, G2 and S + G2 phases.
**P<0.01. n=7. (C) Gating strategy for annexin V analysis by
flow cytometry. (D) Proportions of annexin V− cells. (E)
Gating strategy of flow cytometry data from intracellular staining.
Staining with isotype controls or specific antibodies against (F)
MDR-1 (extracellular staining), (G) Bcl-2 (intracellular staining)
or (H) STAT1 (intracellular staining). The graphs demonstrate the
evaluation of mean fluorescence intensity (MFI) staining. n=3
independent experiments. PD-L1, programmed cell death-ligand 1;
MHC, major histocompatibility complex; nM, nanomolar.

Figure 3

In JBT19 cells, IFNγ decreases the
intracellular expression levels of Ki-67, increases the
extracellular expression levels of PD-L1 and MHC-I and enhances the
expression levels of MHC-II. (A) Gating strategy of flow cytometry
data. (B) Intracellular staining with anti-Ki-67-specific antibody
in vehicle-treated (No treatment), 2-day treated (2 d) or 7-day
treated (7 d) JBT19 cells with IFNγ (200 ng/ml). Alternatively, the
cells were treated for 2 days with IFNγ (200 ng/ml) and the
cytokine was then extensively removed, while the cells were
cultured in vehicle alone for additional 5 days (2d + 5d rev). As
control (No stain) for individual fluorochromes, staining with
vehicle alone was used. (C) Evaluation of the MFI staining in panel
B. (D) Histograms of extracellular staining with anti-PD-L1-,
MHC-I- or MHC-II-specific antibodies in samples treated as in panel
B. The control (No stain) for individual fluorochromes was staining
with vehicle alone. *P<0.05 and **P<0.01 and ****P<0.0001;
C, n=3 (Ki-67) independent experiments. D, n=6. (E) Extracellular
MHC-II staining of 14-day vehicle-treated (No treatment), 7-day or
14-day IFNγ-treated (IFNγ, 200 ng/ml) JBT19 cells. (F)
Extracellular MHC-II staining of 48-h vehicle-treated or 6-, 24- or
48-h IFNγ-treated (IFNγ, 200 ng/ml) JBT19 cells. *P<0.05,
**P<0.01, ***P<0.001 and ****P<0.0001; E, n=3 independent
experiments. F, n=4 independent experiments. Panels E and F,
one-way ANOVA with Tukey's post hoc test. MTT assay of (G)
vehicle-treated JBT19 cells or (H and I) JBT19 cells treated for 6
h with IFNγ (200 ng/ml). Next, the treated JBT19 cells were rinsed
and exposed to docetaxel at the indicated concentrations for 3 days
in the (G and H) absence (no treatment + 3 d no treatment, 6 h
IFNγ+3 d no treatment) or (I) presence of IFNγ (200 ng/ml). The
calculated IC50 from n=3 independent experiments is
shown. IFNγ, interferon γ; MFI, mean fluorescence intensity; d,
days; PD-L1, programmed cell death-ligand 1; MHC, major
histocompatibility complex; nM, nanomolar.

Figure 4

IFNγ-induced phenotypic remodeling
and docetaxel resistance are sustained for an extended time but are
reversible. (A) Schematic representation of the times of the
experiment. (B) JBT19 cells were 7-day treated with IFNγ (200
ng/ml). The cells were then passaged and cultured for 9 days in the
presence (IFNγ; dark pink) or absence (Rev0-9; No treatment, light
pink) of IFNγ (200 ng/ml) and cell number fold increase was
determined. (C) The Rev0-9 sample of JBT19 cells from panel B
(Rev9-24, light pink) or JBT19 cells not treated with IFNγ (No
treatment, light blue) were passaged and cultured in the absence of
IFNγ for 15 days and the cell number fold increase was determined.
(D) Rev9-24 sample of JBT19 cells from C (Rev24, light pink) and
vehicle-treated sample of JBT19 cells from C (No treatment, light
blue) were passaged and cultured for 12 days. Next, the cell number
fold increase was determined at the indicated days and
proliferation curves and pertinent doubling times were calculated.
(E) Evaluation of the mean fluorescence intensities of
extracellular staining with anti-PD-L1-, MHC-I- or MHC-II-specific
antibodies in the Rev24 sample of JBT19 cells at day 0 from panel D
(Rev24, light pink) and vehicle-treated sample of JBT19 cells at
day 0 (No treatment 2, light blue). (F) Results of MTT assay of No
treatment 2 (blue) and Rev24 (red) samples at day 0 from panel D.
Cells were exposed to docetaxel at the indicated concentrations for
3 days. The calculated IC50 are shown (n=3 independent
experiments). MFI, mean fluorescence intensity (arbitrary units);
IFNγ, interferon γ; nM, nanomolar; PD-L1, programmed cell
death-ligand 1; MHC, major histocompatibility complex.

Figure 5

IFNγ-induced chemoresistance is not
JBT19 cell-restricted and can be mitigated by mitomycin C or
doxorubicin in JBT19 cells. (A) MTT assay of vehicle-treated (No
treatment) or 7-day IFNγ-treated (IFNγ, 200 ng/ml) PC-3 cells
exposed to docetaxel at the indicated concentrations for 3 days.
MTT assay of vehicle-treated (No treatment) or 7-day IFNγ-treated
(IFNγ, 200 ng/ml) JBT19 cells exposed to (B) mitomycin C or (C)
doxorubicin at the indicated concentrations for 3 days. The
calculated IC50 from three independent experiments are
shown. IFNγ, interferon γ; NA, not available; nM, nanomolar.

Figure 6

Reactivity of JBT19- or γJBT19 (7-day
IFNγ-treated)-primed and enriched healthy donor lymphocytes to
JBT19 or γJBT19 cells. (A) Lymphocytes of healthy donors were
stimulated (primed) with non-treated (JBT19-primed) or 7-day
IFNγ-treated (γJBT19-primed) JBT19 cells. The cell cultures were
stimulated with vehicle alone (No stim), non-treated (JBT19-stim)
or 7-day IFNγ-treated (γJBT19-stim) JBT19 cells and the lymphocyte
reactivity was then determined by intracellular staining of IFNγ
and TNFα and flow cytometry analysis. (B and C) The frequencies of
TNFα- and/or IFNγ-producing CD4+ or CD8+ T
cells in the gated CD4+ or CD8+ T cell
(CD3+) populations are shown. Evaluated frequencies of
TNFα+-(left panels), IFNγ+-(middle panels)
and TNFα+/IFNγ+-(right panels) producing
JBT19- or γJBT19-primed and enriched healthy donors (B)
CD4+ or (C) CD8+ T-cell populations after
stimulation with non-treated (JBT19 stim) or 7-day IFNγ-treated
(γJBT19-stim) JBT19 cells. In panels B and C, the statistically
significant differences between JBT19- and γJBT19-primed cells were
determined. *P<0.05; n=4 healthy donors; paired two-tailed
Student's t-test. IFNγ, interferon γ; stim, stimulation.

Figure 7

Reactivity of REP-expanded JBT19- or
γJBT19 (7-day IFNγ-treated)-primed and enriched healthy donor
lymphocytes to JBT19 or γJBT19 cells. (A) REP-expanded cell
cultures were stimulated and lymphocyte reactivity was determined
by intracellular staining of IFNγ and TNFα and flow cytometry
analysis. The calculated frequencies of TNFα+-(left
panels), IFNγ+-(middle panels) and
TNFα+/IFNγ+-(right panels) producing JBT19-
or γJBT19-primed and enriched healthy donors (B) CD4+ or
(C) CD8+ T-cell populations in the REP-expanded cell
cultures stimulated with non-treated (JBT19 stim) or 7-day
IFNγ-treated (γJBT19-stim) JBT19 cells are shown. In panels A and
B, the statistically significant differences between JBT19- and
γJBT19-primed cells were determined (*P<0.05, **P<0.01; n=4
healthy donors; paired two-tailed Student's t-test). (C and D) The
cells in panels A and B were pretreated or not for 30 min with 20
µg/ml anti-MHC-I or anti-MHC-II blocking antibodies in RPMI 1640
medium and then stimulated as shown in panels A and B in the
presence or absence of 10 µg/ml blocking antibodies. Data are
presented as means ± SEM. The statistically significant differences
between the treatment groups are indicated (*P<0.05, **P<0.01
and ***P<0.001; n=3 independent experiments; one-way ANOVA with
Tukey's post hoc test). IFNγ, interferon γ; REP, rapid expansion
protocol; MHC, major histocompatibility complex.

Figure 8

Cytotoxic impact of REP-expanded
JBT19- or γJBT19 (7-day IFNγ-treated)-primed and enriched healthy
donor lymphocytes on JBT19 and γJBT19 cells. (A) REP-expanded
JBT19-(JBT19-primed Ly) or γJBT19-(γJBT19-primed Ly) primed and
enriched healthy donor lymphocytes were cocultured with adherent
non-treated (JBT19-GFP) or 7-day IFNγ-treated (γJBT19-GFP)
JBT19-GFP cells for 3 days and images of BF and GFP fluorescence
were acquired using a fluorescence microscope at 10 X
magnification. (scale bar, 200 µm). JBT19-GFP or γJBT19-GFP cells
cultured without lymphocytes were used as a positive control (No
treatment). (B) The cytotoxic impact of the coculture on JBT19-GFP
cells was evaluated via the MFI of GFP fluorescence (JBT19-GFP MFI)
in the cell coculture, as described in the Materials and methods
section. (C) Cytotoxic impact of the coculture on γJBT19-GFP cells
was evaluated via the MFI of GFP fluorescence (γJBT19-GFP MFI) in
the cell coculture. **P<0.01, ****P<0.0001; n=4 healthy
donors; paired two-tailed Student's t-test). E; effector cells; T;
target cells; IFNγ, interferon γ; MFI, mean fluorescence intensity;
REP, rapid expansion protocol; BF, bright field; GFP, green
fluorescent protein; Ly, lymphocytes.
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Spandidos Publications style
Taborska P, Stakheev D and Smrz D: Dual role of IFN&gamma; in reprogramming the undifferentiated pleomorphic sarcoma cell line JBT19 towards cytotoxic chemotherapy and antitumor immunity. Oncol Lett 31: 78, 2026.
APA
Taborska, P., Stakheev, D., & Smrz, D. (2026). Dual role of IFN&gamma; in reprogramming the undifferentiated pleomorphic sarcoma cell line JBT19 towards cytotoxic chemotherapy and antitumor immunity. Oncology Letters, 31, 78. https://doi.org/10.3892/ol.2025.15431
MLA
Taborska, P., Stakheev, D., Smrz, D."Dual role of IFN&gamma; in reprogramming the undifferentiated pleomorphic sarcoma cell line JBT19 towards cytotoxic chemotherapy and antitumor immunity". Oncology Letters 31.2 (2026): 78.
Chicago
Taborska, P., Stakheev, D., Smrz, D."Dual role of IFN&gamma; in reprogramming the undifferentiated pleomorphic sarcoma cell line JBT19 towards cytotoxic chemotherapy and antitumor immunity". Oncology Letters 31, no. 2 (2026): 78. https://doi.org/10.3892/ol.2025.15431
Copy and paste a formatted citation
x
Spandidos Publications style
Taborska P, Stakheev D and Smrz D: Dual role of IFN&gamma; in reprogramming the undifferentiated pleomorphic sarcoma cell line JBT19 towards cytotoxic chemotherapy and antitumor immunity. Oncol Lett 31: 78, 2026.
APA
Taborska, P., Stakheev, D., & Smrz, D. (2026). Dual role of IFN&gamma; in reprogramming the undifferentiated pleomorphic sarcoma cell line JBT19 towards cytotoxic chemotherapy and antitumor immunity. Oncology Letters, 31, 78. https://doi.org/10.3892/ol.2025.15431
MLA
Taborska, P., Stakheev, D., Smrz, D."Dual role of IFN&gamma; in reprogramming the undifferentiated pleomorphic sarcoma cell line JBT19 towards cytotoxic chemotherapy and antitumor immunity". Oncology Letters 31.2 (2026): 78.
Chicago
Taborska, P., Stakheev, D., Smrz, D."Dual role of IFN&gamma; in reprogramming the undifferentiated pleomorphic sarcoma cell line JBT19 towards cytotoxic chemotherapy and antitumor immunity". Oncology Letters 31, no. 2 (2026): 78. https://doi.org/10.3892/ol.2025.15431
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