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Direct effects of the small molecule PD‑L1 inhibitor BMS‑202 on A375 melanoma cells: Anti‑tumor activity accompanied by increased mitochondrial function

  • Authors:
    • Nieng Zhang
    • Fenglan Feng
    • Ruonan Dang
    • Xiaoqing Zhao
    • Xingrong Wang
    • Yuqi  Yang
    • Jinjin Deng
    • Yujie Wang
    • Zhuofan  Wen
    • Wei  Meng
    • Xinglan Huang
    • Shunying  Zhang
    • Yuqiong Deng
    • Caifeng  Huang
    • Peng Yan
    • Zhongrong Liu
    • Xiping Cheng
  • View Affiliations / Copyright

    Affiliations: Department of Dermatology, The First Affiliated Hospital of Guangzhou Medical University, School of First Clinical Medicine, Guangzhou Medical University, Guangzhou, Guangdong 510120, P.R. China, The Key Laboratory of Advanced Interdisciplinary Studies, The First Affiliated Hospital of Guangzhou Medical University, Guangzhou Medical University, Guangzhou, Guangdong 510120, P.R. China, Department of Chinese Medicine, The First Affiliated Hospital of Guangzhou Medical University, School of First Clinical Medicine, Guangzhou Medical University, Guangzhou, Guangdong 510120, P.R. China, Department of Traditional Chinese Medicine, Institute of Integrated Chinese and Western Medicine of Guangzhou Medical University, Guangzhou, Guangdong 510182, P.R. China, Department of Obstetrics and Gynecology, The First Affiliated Hospital of Guangzhou Medical University, Guangzhou, Guangdong 510120, P.R. China, Department of Dermatology, Guangzhou Twelfth People's Hospital, Guangzhou, Guangdong 510620, P.R China, Department of Dermatology, Songshan Lake Central Hospital of Dongguan City of Southern Medical University, Dongguan, Guangdong 523000, P.R. China, Department of Dermatology, Panyu Maternal and Child Care Service Centre of Guangzhou, Guangzhou, Guangdong 511400, P.R. China, Department of Critical Care Medicine, Guangzhou First People's Hospital, South China University of Technology, Guangzhou, Guangdong 510180, P.R. China
    Copyright: © Zhang et al. This is an open access article distributed under the terms of Creative Commons Attribution License.
  • Article Number: 242
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    Published online on: July 7, 2025
       https://doi.org/10.3892/mmr.2025.13607
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Abstract

The aim of the present study was to investigate the direct effects of BMS‑202 on melanoma cells. The small molecule programmed cell death ligand 1 (PD‑L1) inhibitor BMS‑202 was used to treat A375 melanoma cells. The cell distribution of BMS‑202 was examined using low‑power and high‑resolution confocal microscopy, focusing on its localization in mitochondria. The impact of BMS‑202 on mitochondrial gene expression levels, the activity of respiratory chain complexes, and the levels of reactive oxygen species and apoptosis‑related genes, including Bax, Bcl‑2, PARP and caspase‑3, were assessed by quantitative PCR and western blotting. Additionally, tumor cell viability, proliferation, migration and invasion were evaluated in vitro, with in vivo experiments conducted through the construction of tumor‑bearing mouse models and Ki‑67 immunohistochemical staining to validate tumor proliferation. The function of mitochondria was inhibited using a pyruvate carrier inhibitor to examine how this affected the action of BMS‑202. The results revealed that BMS‑202 can inhibit tumor cell function and promote apoptosis. Furthermore, BMS‑202 was shown to enter the mitochondria where it may bind to PD‑L1 and improve mitochondrial function. By inhibiting mitochondrial function, the antitumor effects of BMS‑202 can be enhanced. Overall, the present study provides information on the potential antitumor mechanisms of BMS‑202 as well as a theoretical basis for its application in melanoma therapy.
View Figures

Figure 1

BMS-202 inhibits A375 cell viability,
migration and invasion, and induces apoptosis. (A) Chemical
structure of BMS-202. (B) Cell Counting Kit-8 assay of cell
viability and determine the IC50 of BMS-202. (C)
Assessment of PD-L1 expression by western blotting. (D) Assessment
of cell migration. (E) Evaluation of cell invasion. (F) Reverse
transcription-quantitative PCR of mRNA expression levels of the
pro-apoptotic protein Bax and the anti-apoptotic protein Bcl-2. (G)
Western blotting of the protein expression levels of Bax and Bcl-2.
*P<0.05, **P<0.01, ***P<0.001. Ctrl, control.

Figure 2

BMS-202 enhances mitochondrial
respiratory chain complex expression levels and increases ATP
levels. (A) Reverse transcription-quantitative PCR analysis of the
mRNA expression levels of MT-CO1, MT-CO3 and MT-ND4. (B) ELISA of
MRCC I and IV levels. (C) ATP quantification assay. ns, no
significant difference; **P<0.01, ****P<0.0001. Ctrl,
control; MT-CO, mitochondrially encoded cytochrome c
oxidase; MT-ND4, mitochondrially encoded NADH dehydrogenase 4;
MRCC, Mitochondrial respiratory chain complex.

Figure 3

Localization of BMS-202 in cells. (A)
BMS-202 localization within cells observed under low magnification
(magnification, ×10). (B) Live-cell imaging using structured
illumination microscopy to examine the mitochondrial localization
of BMS-202 (magnification, ×100). (C) Immunofluorescence analysis
showing the binding of BMS-202 to PD-L1 (magnification, ×100).
PD-L1, programmed cell death ligand 1.

Figure 4

Effect of GW on the enhancement of
mitochondrial function induced by BMS-202. (A) Determination of the
IC50 of GW. (B) Reverse transcription-quantitative PCR
analysis of the mRNA expression levels of MT-CO1, MT-ND4 and
MT-ND5. (C) Western blotting was performed to assess the protein
expression levels of MT-CO1, MT-ND4 and MT-ND5. (D) ELISA
measurement of MRCC I and IV levels. (E) ATP level quantification.
(F) JC-1 fluorescence assay was performed to evaluate mitochondrial
membrane potential. ns, no significant difference; *P<0.05,
**P<0.01, ***P<0.001, ****P<0.0001. CCCP, carbonyl cyanide
m-chlorophenyl hydrazone; Ctrl, control; MT-CO, mitochondrially
encoded cytochrome c oxidase; MT-ND, mitochondrially encoded NADH
dehydrogenase; GW, GW604714X; MRCC, Mitochondrial Respiratory Chain
Complex.

Figure 5

GW inhibits changes in oxidative
stress levels induced by BMS-202. (A) Detection of Mn-SOD activity
in mitochondria. (B) Detection of MDA levels. (C) Detection of
Mt-ROS. ns, no significant difference; *P<0.05, **P<0.01,
***P<0.001, ****P<0.0001. SOD, superoxide dismutase; MDA,
malondialdehyde; Mt-ROS, mitochondrial reactive oxygen species. GW,
GW604714X.

Figure 6

Assessment of apoptosis levels. (A)
Changes in Bcl-2 and Bax mRNA expression levels were detected by
reverse transcription-quantitative PCR. (B) Changes in Bax, Bcl-2,
cleaved-caspase-3 and cleaved-PARP protein expression were detected
by western blotting. (C) Analysis of caspase-3 and caspase-8
expression by cellular immunofluorescence. (D) Flow cytometric
analysis of cell apoptosis. *P<0.05, **P<0.01, ***P<0.001,
****P<0.000. Ctrl, control; PARP, poly(ADP-ribose)
polymerase.

Figure 7

Changes in tumor cell function. (A)
Cell Counting Kit-8 assay was used to assess changes in cell
viability. (B) Cell migration assay was performed to assess changes
in migration. (C) Cell invasion assay was performed to assess
changes in invasion. (D) Clonogenic assay was used to detect cell
proliferation ability. ns, no significant difference; *P<0.05,
**P<0.01, ***P<0.001, ****P<0.0001. GW, GW604714X.

Figure 8

Evaluation of tumor proliferation
in vivo. (A) Animal experimental procedure. (B)
Tumor-bearing mice. (C) Comparison of tumor sizes. (D) Weight
changes in tumor-bearing mice. (E) Comparison of tumor volume (F)
H&E staining (magnification, 20×). (G) Tumor proliferation was
assessed by immunohistochemical analysis of Ki-67 (magnification,
20×). (H) Fluorescence intensity of Ki-67. ns, no significant
difference; ****P<0.05; ****P<0.0001. Ctrl, control. GW,
GW604714X.
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Spandidos Publications style
Zhang N, Feng F, Dang R, Zhao X, Wang X, Yang Y, Deng J, Wang Y, Wen Z, Meng W, Meng W, et al: Direct effects of the small molecule PD‑L1 inhibitor BMS‑202 on A375 melanoma cells: Anti‑tumor activity accompanied by increased mitochondrial function. Mol Med Rep 32: 242, 2025.
APA
Zhang, N., Feng, F., Dang, R., Zhao, X., Wang, X., Yang, Y. ... Cheng, X. (2025). Direct effects of the small molecule PD‑L1 inhibitor BMS‑202 on A375 melanoma cells: Anti‑tumor activity accompanied by increased mitochondrial function. Molecular Medicine Reports, 32, 242. https://doi.org/10.3892/mmr.2025.13607
MLA
Zhang, N., Feng, F., Dang, R., Zhao, X., Wang, X., Yang, Y., Deng, J., Wang, Y., Wen, Z., Meng, W., Huang, X., Zhang, S., Deng, Y., Huang, C., Yan, P., Liu, Z., Cheng, X."Direct effects of the small molecule PD‑L1 inhibitor BMS‑202 on A375 melanoma cells: Anti‑tumor activity accompanied by increased mitochondrial function". Molecular Medicine Reports 32.3 (2025): 242.
Chicago
Zhang, N., Feng, F., Dang, R., Zhao, X., Wang, X., Yang, Y., Deng, J., Wang, Y., Wen, Z., Meng, W., Huang, X., Zhang, S., Deng, Y., Huang, C., Yan, P., Liu, Z., Cheng, X."Direct effects of the small molecule PD‑L1 inhibitor BMS‑202 on A375 melanoma cells: Anti‑tumor activity accompanied by increased mitochondrial function". Molecular Medicine Reports 32, no. 3 (2025): 242. https://doi.org/10.3892/mmr.2025.13607
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Spandidos Publications style
Zhang N, Feng F, Dang R, Zhao X, Wang X, Yang Y, Deng J, Wang Y, Wen Z, Meng W, Meng W, et al: Direct effects of the small molecule PD‑L1 inhibitor BMS‑202 on A375 melanoma cells: Anti‑tumor activity accompanied by increased mitochondrial function. Mol Med Rep 32: 242, 2025.
APA
Zhang, N., Feng, F., Dang, R., Zhao, X., Wang, X., Yang, Y. ... Cheng, X. (2025). Direct effects of the small molecule PD‑L1 inhibitor BMS‑202 on A375 melanoma cells: Anti‑tumor activity accompanied by increased mitochondrial function. Molecular Medicine Reports, 32, 242. https://doi.org/10.3892/mmr.2025.13607
MLA
Zhang, N., Feng, F., Dang, R., Zhao, X., Wang, X., Yang, Y., Deng, J., Wang, Y., Wen, Z., Meng, W., Huang, X., Zhang, S., Deng, Y., Huang, C., Yan, P., Liu, Z., Cheng, X."Direct effects of the small molecule PD‑L1 inhibitor BMS‑202 on A375 melanoma cells: Anti‑tumor activity accompanied by increased mitochondrial function". Molecular Medicine Reports 32.3 (2025): 242.
Chicago
Zhang, N., Feng, F., Dang, R., Zhao, X., Wang, X., Yang, Y., Deng, J., Wang, Y., Wen, Z., Meng, W., Huang, X., Zhang, S., Deng, Y., Huang, C., Yan, P., Liu, Z., Cheng, X."Direct effects of the small molecule PD‑L1 inhibitor BMS‑202 on A375 melanoma cells: Anti‑tumor activity accompanied by increased mitochondrial function". Molecular Medicine Reports 32, no. 3 (2025): 242. https://doi.org/10.3892/mmr.2025.13607
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