Bevacizumab induces ferroptosis and enhances CD8+ T cell immune activity in liver cancer via modulating HAT1 and increasing IL-9.

Bevacizumab is a recombinant humanized monoclonal immunoglobulin (Ig) G1 antibody of VEGF, and inhibits angiogenesis and tumor growth in hepatocellular carcinoma (HCC). Ferroptosis, a new form of regulated cell death function independently of the apoptotic machinery, has been accepted as an attractive target for pharmacological intervention; the ferroptosis pathway can enhance cell immune activity of anti-PD1 immunotherapy in HCC. In this study we investigated whether and how bevacizumab regulated ferroptosis and immune activity in liver cancer. Firstly, we performed RNA-sequencing in bevacizumab-treated human liver cancer cell line HepG2 cells, and found that bevacizumab significantly altered the expression of a number of genes including VEGF, PI3K, HAT1, SLC7A11 and IL-9 in liver cancer, bevacizumab upregulated 37 ferroptosis-related drivers, and downregulated 17 ferroptosis-related suppressors in particular. We demonstrated that bevacizumab triggered ferroptosis in liver cancer cells by driving VEGF/PI3K/HAT1/SLC7A11 axis. Clinical data confirmed that the expression levels of VEGF were positively associated with those of PI3K, HAT1 and SLC7A11 in HCC tissues. Meanwhile, we found that bevacizumab enhanced immune cell activity in tumor immune-microenvironment. We identified that HAT1 up-regulated miR-143 targeting IL-9 mRNA 3'UTR in liver cancer cells; bevacizumab treatment resulted in the increase of IL-9 levels and its secretion via VEGF/PI3K/HAT1/miR-143/IL-9 axis, which led to the inhibition of tumor growth in vivo through increasing the release of IL-2 and Granzyme B from activated CD8+ T cells. We conclude that in addition to inhibiting angiogenesis, bevacizumab induces ferroptosis and enhances CD8+ T cell immune activity in liver cancer. This study provides new insight into the mechanisms by which bevacizumab synergistically modulates ferroptosis and CD8+ T cell immune activity in liver cancer.

Inhibition of USP7 enhances CD8+ T cell activity in liver cancer by suppressing PRDM1-mediated FGL1 upregulation.

Lymphocyte activation gene 3 (LAG3), an immune checkpoint molecule expressed on activated T cells, functions as a negative regulator of immune responses. Persistent antigen exposure in the tumor microenvironment results in sustained LAG3 expression on T cells, contributing to T cell dysfunction. Fibrinogen-like protein 1 (FGL1) has been identified as a major ligand of LAG3, and FGL1/LAG3 interaction forms a novel immune checkpoint pathway that results in tumor immune evasion. In addition, ubiquitin-specific peptidase 7 (USP7) plays a crucial role in cancer development. In this study we investigated the role of USP7 in modulation of FGL1-mediated liver cancer immune evasion. We showed that knockdown of USP7 or treatment with USP7 inhibitor P5091 suppressed liver cancer growth by promoting CD8+ T cell activity in Hepa1-6 xenograft mice and in HepG2 or Huh7 cells co-cultured with T cells, whereas USP7 overexpression produced the opposite effect. We found that USP7 upregulated FGL1 in HepG2 and Huh7 cells by deubiquitination of transcriptional factor PR domain zinc finger protein 1 (PRDM1), which transcriptionally activated FGL1, and attenuated the CD8+ T cell activity, leading to the liver cancer growth. Interestingly, USP7 could be transcriptionally stimulated by PRDM1 as well in a positive feedback loop. P5091, an inhibitor of USP7, was able to downregulate FGL1 expression, thus enhancing CD8+ T cell activity. In an immunocompetent liver cancer mouse model, the dual blockade of USP7 and LAG3 resulted in a superior antitumor activity compared with anti-LAG3 therapy alone. We conclude that USP7 diminishes CD8+ T cell activity by a USP7/PRDM1 positive feedback loop on FGL1 production in liver cancer; USP7 might be a promising target for liver cancer immunotherapy.

Aristolochic acids-hijacked p53 promotes liver cancer cell growth by inhibiting ferroptosis.

Aristolochic acids (AAs) have been identified as a significant risk factor for hepatocellular carcinoma (HCC). Ferroptosis is a type of regulated cell death involved in the tumor development. In this study, we investigated the molecular mechanisms by which AAs enhanced the growth of HCC. By conducting bioinformatics and RNA-Seq analyses, we found that AAs were closely correlated with ferroptosis. The physical interaction between p53 and AAs in HepG2 cells was validated by bioinformatics analysis and SPR assays with the binding pocket sites containing Pro92, Arg174, Asp207, Phe212, and His214 of p53. Based on the binding pocket that interacts with AAs, we designed a mutant and performed RNA-Seq profiling. Interestingly, we found that the binding pocket was responsible for ferroptosis, GADD45A, NRF2, and SLC7A11. Functionally, the interaction disturbed the binding of p53 to the promoter of GADD45A or NRF2, attenuating the role of p53 in enhancing GADD45A and suppressing NRF2; the mutant did not exhibit the same effects. Consequently, this event down-regulated GADD45A and up-regulated NRF2, ultimately inhibiting ferroptosis, suggesting that AAs hijacked p53 to down-regulate GADD45A and up-regulate NRF2 in HepG2 cells. Thus, AAs treatment resulted in the inhibition of ferroptosis via the p53/GADD45A/NRF2/SLC7A11 axis, which led to the enhancement of tumor growth. In conclusion, AAs-hijacked p53 restrains ferroptosis through the GADD45A/NRF2/SLC7A11 axis to enhance tumor growth. Our findings provide an underlying mechanism by which AAs enhance HCC and new insights into p53 in liver cancer. Therapeutically, the oncogene NRF2 is a promising target for liver cancer.

Aspirin modulates succinylation of PGAM1K99 to restrict the glycolysis through NF-κB/HAT1/PGAM1 signaling in liver cancer.

Aspirin as a chemopreventive agent is able to restrict the tumor growth. Phosphoglycerate mutase 1 (PGAM1) is a key enzyme of glycolysis, playing an important role in the development of cancer. However, the underlying mechanism by which aspirin inhibits the proliferation of cancer cells is poorly understood. This study aims to identify the effects of aspirin on modulating PGAM1 enzymatic activities in liver cancer. Here, we found that aspirin attenuated the PGAM1 succinylation to suppress the PGAM1 enzymatic activities and glycolysis in hepatoma cells. Mechanically, aspirin remarkably reduced the global succinylation levels of hepatoma cells, including the PGAM1 succinylation, which led to the block of conversion from 3-phosphoglycerate (3-PG) to 2-phosphoglycerate (2-PG) in cells. Interestingly, RNA-seq analysis identified that aspirin could significantly decrease the levels of histone acetyltransferase 1 (HAT1), a writer of PGAM1 succinylation, in liver cancer. As a target of aspirin, NF-κB p65 could effectively up-regulate the expression of HAT1 in the system, resulting in the increase of PGAM1 enzymatic activities. Moreover, we observed that the PGAM1-K99R mutant failed to rescue the aspirin-induced inhibition of PGAM1 activities, glycolysis, and proliferation of hepatoma cells relative to PGAM1-WT. Functionally, aspirin down-regulated HAT1 and decreased the PGAM1 succinylation levels in the tumor tissues from mice treated with aspirin in vivo. Thus, we conclude that aspirin modulates PGAM1K99 succinylation to restrict the PGAM1 activities and glycolysis through NF-κB p65/HAT1/PGAM1 signaling in liver cancer. Our finding provides new insights into the mechanism by which aspirin inhibits glycolysis in hepatocellular carcinoma.

Aspirin triggers ferroptosis in hepatocellular carcinoma cells through restricting NF-κB p65-activated SLC7A11 transcription.

A number of studies have shown that aspirin, as commonly prescribed drug, prevents the development of hepatocellular carcinoma (HCC). Ferroptosis as a dynamic tumor suppressor plays a vital role in hepatocarcinogenesis. In this study we investigated whether aspirin affected ferroptosis in liver cancer cells. RNA-seq analysis revealed that aspirin up-regulated 4 ferroptosis-related drivers and down-regulated 5 ferroptosis-related suppressors in aspirin-treated HepG2 cells. Treatment with aspirin (4 mM) induced remarkable ferroptosis in HepG2 and Huh7 cells, which was enhanced by the ferroptosis inducer erastin (10 µM). We demonstrated that NF-κB p65 restricted ferroptosis in HepG2 and Huh7 cells through directly binding to the core region of SLC7A11 promoter and activating the transcription of ferroptosis inhibitor SLC7A11, whereas aspirin induced ferroptosis through inhibiting NF-κB p65-activated SLC7A11 transcription. Overexpression of p65 rescued HepG2 and Huh7 cells from aspirin-induced ferroptosis. HCC patients with high expression levels of SLC7A11 and p65 presented lower survival rate. Functionally, NF-κB p65 blocked the aspirin-induced ferroptosis in vitro and in vivo, which was attenuated by erastin. We conclude that aspirin triggers ferroptosis by restricting NF-κB-activated SLC7A11 transcription to suppress the growth of HCC. These results provide a new insight into the mechanism by which aspirin regulates ferroptosis in hepatocarcinogenesis. A combination of aspirin and ferroptosis inducer may provide a potential strategy for the treatment of HCC in clinic.

Heat shock protein family A member 8 serving as a co-activator of transcriptional factor ETV4 up-regulates PHLDA2 to promote the growth of liver cancer.

Heat shock protein family A member 8 (HSPA8) participates in the folding or degradation of misfolded proteins under stress and plays critical roles in cancer. In this study, we investigated the function of HSPA8 in the development of liver cancer. By analyzing the TCGA transcriptome dataset, we found that HSPA8 was upregulated in 134 clinical liver cancer tissue samples, and positively correlated with poor prognosis. IHC staining showed the nuclear and cytoplasmic localization of HSPA8 in liver cancer cells. Knockdown of HSPA8 resulted in a decrease in the proliferation of HepG2 and Huh-7 cells. ChIP-seq and RNA-seq analysis revealed that HSPA8 bound to the promoter of pleckstrin homology-like domain family A member 2 (PHLDA2) and regulated its expression. The transcription factor ETV4 in HepG2 cells activated PHLDA2 transcription. HSPA8 and ETV4 could interact with each other in the cells and colocalize in the nucleus. From a functional perspective, we demonstrated that HSPA8 upregulated PHDLA2 through the coactivating transcription factor ETV4 to enhance the growth of liver cancer in vitro and in vivo. From a therapeutic perspective, we identified both HSPA8 and PHDLA2 as novel targets in the treatment of HCC. In conclusion, this study demonstrates that HSPA8 serves as a coactivator of ETV4 and upregulates PHLDA2, leading to the growth of HCC, and is a potential therapeutic target in HCC treatment.

High-Symmetry Co/Ni Triazine Polycarboxylate Diverse Frameworks Constructed by Mx(COO)y Building Blocks: Characterization and Catalytic Performance Evaluation of p-Nitrophenol.

Three new triazine compounds [Co1.5(H3TDPAT)(H2O)3]·6H2O (1), [Co2(TCPT)(µ2-H2O)2]·OH (2), and [Ni3(TCPT)]·3OH (3) were designed and synthesized via the reaction of the symmetrical triazine ligand connected by C-N-C and C-O-C bonds with triazine poly(carboxylic acid)s ligands as the side arms: H6TDPAT (H6TDPAT = 2,4,6-tris(3,5-dicarboxylphenylamino)-1,3,5-triazine) and H3TCPT (H3TCPT = 2,4,6-tris(4-carboxyphenoxy)-1,3,5-triazine) as well as the corresponding metal salts under the solvothermal condition. Three triazine polycarboxylate frameworks were characterized by elemental analysis, infrared spectroscopy, ultraviolet spectroscopy, thermogravimetric analysis, X-ray powder diffraction, and solid fluorescent spectra in detail. The structural analysis results showed that the three-dimensional porous cage framework of compound 1 was constructed by three different polyhedral cages connected with [Co(COO)4(H2O)2] building blocks. One of the compounds, 2, is formed by twin propeller Co2(µ2-H2O)(COO)3 building blocks connecting two-dimensional layers and the intermolecular π-π interactions involved the triazine rings between the layers. While the structure of compound 3 is similar to that of 2, assembly is by Ni(COO)3 building blocks and adjacent layers of the face-to-face π-π interaction between the triazine rings. In order to explore functional properties, the catalytic reduction of p-nitrophenol (PNP) of compounds 1-3 was investigated. They exhibit excellent catalytic activity of more than 95% for reduction of PNP with a dose of 2.5 mg of the compounds.

Dynamics of soil arthropod communities in the annual cultivated Gramineae grasslands in alpine region, Northwest Sichuan.

The dynamics of soil arthropod communities in annual monoculture grasslands is still unclear, which restricts the understanding of the degradation mechanism of cultivated grasslands. We cultivated two annual gramineae species, Lolium multiflorum and Avena sativa, separately in Hongyuan County, located on the eastern edge of the Qinghai-Tibet Plateau, in April 2019. We investigated soil arthropods, plant communities and soil properties in the cultivated grasslands and natural grassland in the late September every year from 2019 to 2022. The results showed that: 1) The taxonomic composition of soil arthropod communities differed significantly among three grasslands and sampling years. 2) There was no significant difference in the density, taxonomic richness, Shannon index and evenness index of soil arthropod communities among three grasslands. 3) The density of soil arthropod communities significantly fluctuated across years in three grasslands, and the taxonomic richness and Shannon index decreased significantly in the L. multiflorum and A. sativa grasslands, with the evenness index declining significantly only in the fourth year. The Shannon index fluctuated significantly and the evenness index varied little in natural grassland. 4) The above- and below-ground biomass, the contents of soil total P, total K and available N were the main factors influencing the taxonomic composition, density and diversity indices of soil arthropod communities. The results suggested that the cultivation of annual gramineae grasslands have significant effects on taxonomic composition, but not on density and diversity of soil arthropod communities, and those variables change significantly across different years.

[Effects of cultivating Coptis chinensis and Paris polyphylla on soil microarthropod communities.] / 种植黄连和重楼对小型土壤节肢动物群落的影响.

In recent years, the area of herbal medicine planting is rapidly increasing. The effects of planting herbal medicines on soil invertebrate communities are still unclear. To reveal the effects of planting different herbal medicines on the soil microarthropod communities, soil microarthropods in two fields of planting Coptis chinensis and Paris polyphylla for 3-year and 5-year, respectively, were investigated in Pengzhou, Chengdu in July 2020. A total of 526 individuals of soil microarthropods were recorded and classified into 4 classes, 17 orders, 69 families, and 98 genera or taxonomic groups. The communities were dominated by Isotoma, Piatynothrus, Folsomia, and Paranura. The community structure of soil microarthropods differed obviously among the two herbal medicine fields, with the main influencing taxonomic groups of Proisotoma, Ocesobates and Epicridae. The total taxonomic group richness of soil microarthropods were richer in C. chinensis field than P. polyphylla field. There was no significant difference in the abundance and diversity index between the two fields. With the increases of cultivating years, the abundance of soil microarthropods in C. chinensis field declined significantly, and Shannon index increased significantly in P. polyphylla field. The redundancy analysis showed that the community structure of soil microarthropods was mainly affected by soil available N, pH, total K, and available K. It suggested that the effects of cultivating herbal medicines on soil microarthropod communities differed between herbal medicine species. Therefore, we recommended to intercrop C. chinensis and P. polyphylla for maintaining the stability of soil microarthropod diversity and promoting ecosystem function.