Correlative insights into luteolin regulating the macrophage-p53-CCNB1/CDK1 axis associated with cell-cycle control in hepatocellular carcinoma

The polarization of macrophages and the deactivation of oncogenic pathways within the tumor microenvironment are directly linked to the evolution of hepatocellular carcinoma (HCC), a relatively frequent cancer in the world. Though the exact mechanism is yet unknown, tumor-associated macrophages (TAMs) may alter p53 activity by secreting cytokines. In contrast, the (tumor protein p53, p53) signaling pathway is frequently functionally compromised in HCC as a result of mutation or epigenetic silencing. It is yet unknown if the natural substance luteolin restores p53 function by controlling macrophage polarization, despite its multi-targeted anti-tumor potential.

To clarify the interventional role of luteolin in the evolution of HCC and to uncover the mechanism by which macrophage polarization influences this process via the p53 signaling pathway.

The functions of pivotal genes (Cyclin B1,CCNB1), (Cyclin-dependent kinase 1, CDK1), p53 and macrophage polarization-associated factors (Interleukin 6, IL-6), (tumor necrosis factor-α, TNF-α), (Interleukin 10, IL-10), (Transforming growth factor beta 1, TGF-β1) were experimentally verified in conjunction with bioinformatics analysis (screening of luteolin targets for intersection with HCC differential genes and pivotal genes), transcriptome enrichment analysis, immune infiltration assessment, and single-cell sequencing techniques.

20 hub genes were screened, 5760 HCC differential genes were found, and 104 luteolin action targets were found. According to enrichment analysis, luteolin inhibited HCC via controlling the metabolism of ROS, extracellular matrix catabolism, and cell cycle pathways (p53, cellular senescence). The presence of M2-type TAMs in the tumor core was confirmed by single-cell sequencing, and their secretion of TGF-β and IL-10 led to aberrant activation of CCNB1/CDK1 and cell cycle dysregulation by promoting the ubiquitylation degradation of p53 and inhibiting its nuclear translocation. Luteolin reversed M2-type macrophage polarization, down-regulated IL-10/TGF-β, and was associated with restoration of p53 function via ATM-dependent p53 phosphorylation. This event may induce G2/M-phase arrest and cellular senescence, as well as suppress the activity of the CCNB1/CDK1 complex.

Our correlative results suggest that luteolin may reprogram macrophage polarization (inhibiting M2-type and encouraging M1-type), which is associated with relieving IL-10/TGF-β1-mediated suppression of p53 signaling and subsequent inhibition of CCNB1/CDK1-driven cell cycle progression. Nevertheless, the causal direction between macrophage polarization status and p53 function, the primary cellular target of luteolin, protein-level phenotypic validation of macrophage polarization, p53-dependency of CCNB1/CDK1 inhibition, and direct experimental proof for immune escape remain to be fully elucidated. This study highlights the multifaceted impacts of natural medicines in controlling tumor microenvironment and genome stability, and it offers a novel target for combined immune-metabolic therapy of HCC.