2. Academic Validation
  3. Mechanism of d-Glucaro-1,4-lactone enhancing the anticancer efficacy of lenvatinib via the IFN-γ-STAT3-PD-L1 signaling pathway in hepatocellular carcinoma

Mechanism of d-Glucaro-1,4-lactone enhancing the anticancer efficacy of lenvatinib via the IFN-γ-STAT3-PD-L1 signaling pathway in hepatocellular carcinoma

  • Phytomedicine. 2025 Dec 31:150:157760. doi: 10.1016/j.phymed.2025.157760.
Yufeng Deng 1,  Qi Hu 2,  Zhiying Song 3,  Mengyuan Nie 4,  Zhiguo Wu 5,  Yongrong Lei 6,  Meng Liu 6,  Dujuan Zhan 6,  Baogang Xie 7
Affiliations

Affiliations

  • 1 Modern Industrial College of Traditional Chinese Medicine and Health, Lishui University, Lishui, 323000, China; Medical College of Jiaxing University, Key Laboratory of Medical Electronics and Digital Health of Zhejiang Province, Jiaxing University, Jiaxing, China; School of Pharmacy, Jiangxi Medical College, Nanchang University, Nanchang, China; Department of Pharmacy, Guangdong Provincial People's Hospital (Guangdong Academy of Medical Sciences), Southern Medical University, Guangzhou, China.
  • 2 Modern Industrial College of Traditional Chinese Medicine and Health, Lishui University, Lishui, 323000, China; Medical College of Jiaxing University, Key Laboratory of Medical Electronics and Digital Health of Zhejiang Province, Jiaxing University, Jiaxing, China; School of Pharmacy, Jiangxi Medical College, Nanchang University, Nanchang, China.
  • 3 Modern Industrial College of Traditional Chinese Medicine and Health, Lishui University, Lishui, 323000, China; Medical College of Jiaxing University, Key Laboratory of Medical Electronics and Digital Health of Zhejiang Province, Jiaxing University, Jiaxing, China; Department of Infectious Diseases, The Second Affiliated Hospital of Nanchang University, Nanchang 330006.
  • 4 Modern Industrial College of Traditional Chinese Medicine and Health, Lishui University, Lishui, 323000, China; School of Pharmacy, Jiangxi Medical College, Nanchang University, Nanchang, China.
  • 5 Department of Infectious Diseases, The Second Affiliated Hospital of Nanchang University, Nanchang 330006.
  • 6 Modern Industrial College of Traditional Chinese Medicine and Health, Lishui University, Lishui, 323000, China.
  • 7 Modern Industrial College of Traditional Chinese Medicine and Health, Lishui University, Lishui, 323000, China; Medical College of Jiaxing University, Key Laboratory of Medical Electronics and Digital Health of Zhejiang Province, Jiaxing University, Jiaxing, China. Electronic address: [email protected].
PMID: 41485289 DOI: 10.1016/j.phymed.2025.157760
Abstract

Objective: Hepatocellular carcinoma (HCC) remains a leading cause of Cancer mortality. Targeted therapies like lenvatinib face limitations due to resistance and immunosuppression. d-Glucaro-1,4-lactone (1,4-GL), a bioactive natural compound with anti-HCC activity, holds unexplored synergistic potential with lenvatinib. This study investigates the combinatorial efficacy of 1,4-GL and lenvatinib against HCC and elucidates the underlying mechanisms.

Methods: Efficacy was evaluated in H22 tumor-bearing mice treated with lenvatinib (20 mg/kg/day) ± 1,4-GL (25, 50, 100 mg/kg/day). Tumor inhibition, serum cytokines (TNF-α, IFN-γ, AFP), oxidative stress markers (MDA, SOD), liver Enzymes (ALT, AST), and splenic T-cell subsets (flow cytometry) were assessed. Histopathology was analyzed via H&E staining. Lenvatinib pharmacokinetics (serum, liver, tumor) with/without 1,4-GL was determined in mice and SD rats using UPLC-MS/MS. In vitro, combinatorial effects on proliferation (CCK-8), migration (scratch assay), and clonogenicity (colony formation) were tested in Huh7 and HepG2 cells. STAT3 phosphorylation and PD-L1 expression (protein: Western blot; mRNA: RT-qPCR) were analyzed. PD-L1 induction was achieved using IFN-γ (20 ng/ml).

Results: The medium-dose combination (LMGL) achieved 89.97% tumor inhibition in H22 mice, significantly reducing tumor mass by 75% vs. lenvatinib alone (0.21±0.18 g vs. 0.84±0.79 g, p < 0.01). H&E staining revealed extensive necrosis in combination groups. Serum TNF-α and IFN-γ levels significantly increased (p < 0.001), while AFP approached baseline. Flow cytometry showed an elevated CD4⁺/CD8⁺ T-cell ratio. Oxidative stress was modulated (reduced MDA and elevated SOD, p < 0.01). Pharmacokinetic studies revealed no significant differences in lenvatinib exposure or steady-state concentrations (p > 0.05) with 1,4-GL co-administration. In vitro, 1,4-GL significantly reduced lenvatinib's IC50 (Huh7: 18.54 μmol/l; HepG2: 30.34 μmol/l). Combinatorial treatment markedly downregulated PD-L1 protein/mRNA and inhibited STAT3 phosphorylation (p < 0.05). Scratch and colony formation assays indicated synergy stemmed from immunomodulation and signaling inhibition, not enhanced anti-migration/proliferation.

Conclusion: 1,4-GL potently synergizes with lenvatinib against HCC by enhancing oxidative stress, reconstituting antitumor immunity, and suppressing the STAT3/PD-L1 signaling axis, without perturbing lenvatinib pharmacokinetics. This defines a novel phytochemical-based combinatorial strategy with significant clinical translation potential for HCC therapy.

Keywords

Hepatocellular carcinoma; Lenvatinib; PD-L1; Pharmacokinetics; STAT3; Synergism; Tumor microenvironment; d-Glucaro-1,4-lactone.

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