HMGB1 as a Key Mediator in Malignant Mesothelioma and a Potential Target for Asbestos-Related Cancer Therapy

Abstract

Malignant mesothelioma (MM) is a highly aggressive cancer strongly associated with asbestos exposure, and accumulating evidence suggests that high mobility group box 1 (HMGB1) plays a central role in its pathogenesis. Our in vitro and in vivo experiments revealed that HMGB1 was highly expressed in MM. Both genetic and pharmacological inhibition of HMGB1 markedly suppressed MM cell viability, migration, and invasion, while inducing G1-phase cell cycle arrest and enhancing apoptosis. Interestingly, the inhibition of Toll-like receptor 4 (TLR4), achieved through both siRNA and TAK-242 treatment, not only suppressed tumor-promoting signals but also reduced HMGB1 expression, suggesting a self-amplifying HMGB1-TLR4 loop. Mechanistically, in vitro experiments indicated that suppression of HMGB1 and TLR4 was associated with decreased activation of NF-κB, AKT, and ERK pathways, which are involved in regulating MM cell survival and motility. In xenograft models, treatment with ethyl pyruvate (EP) and TAK-242 significantly suppressed tumor growth and HMGB1 expression, reinforcing their therapeutic potential. Given HMGB1's influence on both tumor cell behavior and the immune microenvironment, targeting the HMGB1-TLR4 axis may not only provide a novel therapeutic strategy for MM but also offer insights into the mechanisms underlying asbestos-induced tumorigenesis, potentially guiding future prevention and intervention strategies in asbestos-exposed populations.

Keywords: HMGB1; TLR4; asbestos-related tumors; malignant mesothelioma; tumor progression.

Figure 1

Elevated HMGB1 Expression in MM cell lines and xenograft models. (A) Relative HMGB1 mRNA levels in MM cell lines (NCI-H2452 and MSTO-211H) compared to normal mesothelial cells (MeT-5A), determined by quantitative real-time PCR. (B) Representative Western blot images showing HMGB1 protein expression in MM cell lines (NCI-H2452 and MSTO-211H) and normal mesothelial cells (MeT-5A). (C) Representative images of tumors from the MM xenograft model established by subcutaneous injection of MSTO-211H cells into BALB/c nude mice. (D) Plasma HMGB1 levels measured by ELISA in xenograft model mice. **, p < 0.01; ***, p < 0.001.

Figure 2

Effect of EP treatment on MM cell migration and invasion. Scratch wound healing assays showing the migration of MSTO-211H (A) and NCI-H2452 cells (B) treated with 0, 5, and 10 mM EP at 0, 24, 48, and 72 h. Representative images and quantification of the Transwell assays assessing invasion in MSTO-211H (C) and NCI-H2452 cells (D) treated with 0 and 10 mM EP. ** p < 0.01, *** p < 0.001 vs. 0 mM EP group; ## p < 0.01, ### p < 0.001 vs. 5 mM EP group.

Figure 3

Effect of HMGB1 silencing on MM cell migration and invasion. Scratch wound healing assays showing the migration of MSTO-211H (A) and NCI-H2452 cells (B) treated with blank control, siRNA control, and HMGB1 siRNA at 0, 24, 48, and 72 h. Representative images and quantification of Transwell assays assessing invasion in MSTO-211H and NCI-H2452 cells (C) treated with blank control, siRNA control, and HMGB1 siRNA. * p < 0.05, ** p < 0.01, *** p < 0.001 vs. black group; ## p < 0.01, ### p < 0.001 vs. siRNA control group.

Figure 4

Effect of HMGB1 silencing and EP treatment on cell cycle arrest and apoptosis. Comparison of apoptosis in MSTO-211H (A) and NCI-H2452 (B) cells treated with blank control, siRNA control, and HMGB1 siRNA. Cell cycle distribution in MSTO-211H (C) and NCI-H2452 (D) cells treated with 0, 2.5, 5, and 10 mM EP. Apoptosis analysis of MSTO-211H (E) and NCI-H2452 (F) cells treated with 0, 2.5, 5, 10, and 20 mM EP. * p < 0.05, ** p < 0.01, *** p < 0.001 vs. control group; # p < 0.05, ## p < 0.01 vs. siRNA control group.

Figure 5

Effect of TAK-242 and TLR4 siRNA treatment on HMGB1-TLR4 signaling pathway in MM cells. The mRNA expression of key molecules in the HMGB1-TLR4 signaling axis in MSTO-211H (A) and NCI-H2452 (B) cells treated with 0 and 100 μM TAK-242. The mRNA expression of key molecules in the HMGB1-TLR4 signaling axis in MSTO-211H (C) and NCI-H2452 (D) cells treated with TLR4 siRNA. * p < 0.05, ** p < 0.01, *** p < 0.001 vs. control group; # p < 0.05, ## p < 0.01, ### p < 0.001 vs. siRNA control group.

Figure 6

Effect of EP and TAK-242 treatment on MM xenograft model in vivo. (A) Representative images of mice and tumors after treatment treated with PBS, EP, or TAK-242. (B) Changes in body weight of mice over time after PBS, EP, or TAK-242 treatment. (C) Tumor volume measurements over time in mice treated with PBS, EP, or TAK-242. (D) Tumor weights in mice treated with PBS, EP, or TAK-242. (E) H&E staining of tumor tissues from PBS, EP, and TAK-242 treated groups. (F) Immunohistochemical staining showing HMGB1 expression in tumor tissues from the three treatment groups. (G) Plasma HMGB1 levels measured by ELISA in the three treatment groups. * p < 0.05.