Astragalus injection inhibits the growth of osteosarcoma by activating cytotoxic T lymphocyte and targeting CTSL

Abstract

Ethnopharmacological relevance

Astragalus membranaceus, commonly known as Huangqi in China, is a traditional herbal medicine that has attracted significant attention for its immunomodulatory effects. It has been widely studied in various clinical contexts, including cancer treatment. Astragalus injection (HQI) is clinically used for treating myocarditis and cardiac insufficiency. However, its potential therapeutic effects on osteosarcoma, a highly aggressive bone tumor, remain largely unexplored.

Aim of the study

The aim of this study was to investigate the potential therapeutic effects of HQI on osteosarcoma and to elucidate its underlying mechanisms of action. Specifically, we aimed to determine whether HQI could inhibit osteosarcoma growth in vivo, identify its key active components and molecular targets, and explore its immunomodulatory effects on the tumor microenvironment.

Materials and methods

Mice with osteosarcoma were treated with HQI, and tumor growth was monitored. The number of CD8⁺ T cells in spleen was assessed using flow cytometry. High-performance liquid chromatography coupled with electrospray ionization time-of-flight mass spectrometry (HPLC-ESI-TOF-MS/MS) was used to identify the active ingredients of HQI that entered the peripheral blood of treated mice. Network pharmacology and weighted gene co-expression network analysis (WGCNA) were employed to identify key molecular targets of HQI in osteosarcoma inhibition. In vitro cell assays were conducted to evaluate the effects of HQI and its active components on osteosarcoma cell viability. Molecular docking studies were performed to identify the binding affinity of key active components to the identified molecular targets. The expression of Cathepsin L (CTSL) and the activation of cytotoxic T lymphocytes were assessed in vivo and in vitro to elucidate the primary mechanism of action of HQI and its active component calycosin 7-O-β-D-glucoside (CG).

Results

Our study found that HQI significantly suppresses osteosarcoma growth in vivo by increasing the number of CD8⁺ T cells, without causing significant toxic side effects. Eight active ingredients entered the peripheral blood of mice through HPLC-ESI-TOF-MS/MS detection. The network pharmacology and WGCNA revealed that CTSL was a key target of HQI in osteosarcoma inhibition. Cell assays and molecular docking identified CG as the key active component of HQI to inhibit the activity of osteosarcoma cells, capable of binding to CTSL. In vivo, CG activates cytotoxic T lymphocytes and inhibits CTSL expression, thereby exerting its anti-osteosarcoma effects.

Conclusion

Our study demonstrated that HQI, particularly its active component CG, holds potential as a therapeutic agent for osteosarcoma. The primary mechanism underlying its anti-osteosarcoma effects involves modulating the immune response and targeting CTSL. These findings provide a scientific basis for the development of HQI as a novel immunomodulatory therapy for osteosarcoma.