What question did this study set out to answer?

This research aims to evaluate the effectiveness of methionine restriction as an immunotherapeutic approach for osteosarcoma.

March 1, 2026Open Access

Methionine Restriction Alone Induces T-cell-mediated Immunotherapy of Osteosarcoma in a Syngeneic Mouse Model

Key Points

This research aims to evaluate the effectiveness of methionine restriction as an immunotherapeutic approach for osteosarcoma.
Used K7M2 murine and 143B human osteosarcoma cell lines.
Determined cell viability and EC50 of methionine with WST-8 reagent.
Performed Western immunoblotting to assess PD-L1 expression after methionine restriction.
Implanted K7M2 cells in BALB/c mice and T-cell-deficient mice to investigate tumor growth and T-cell infiltration.
Analyzed tumor-infiltrating lymphocytes via immunohistochemistry.
EC50 values for K7M2 and 143B were 14.18 μM and 20.85 μM, showing strong methionine dependence.
MR treatment reduced PD-L1 expression in both K7M2 and 143B cells compared to controls.
MR diet significantly inhibited K7M2 tumor growth in BALB/c mice but not in T-cell-deficient mice.
MR diet enhanced CD8-positive T-cell infiltration into K7M2 tumors in BALB/c mice.

Abstract

Background/Aim: Osteosarcoma is the most common malignant bone tumor in pediatric and young adult patients. Osteosarcoma is also refractory to immune checkpoint inhibitors (ICIs). It has been recently demonstrated that methionine restriction (MR) increases the response to ICIs in melanoma and colon cancer. The present study aimed to determine whether MR alone can be an immunotherapeutic for osteosarcoma. Materials and Methods: K7M2 murine osteosarcoma cells and 143B human osteosarcoma cells were used for the present study. Cell viability and the half-maximal effective concentration (EC50) of methionine for K7M2 and 143B were determined with the WST-8 cell-viability reagent. Western immunoblotting was used to compare programmed cell death receptor ligand 1 (PD-L1) expression in K7M2 and 143B cells treated with and without MR. K7M2 cells were subcutaneously implanted in immunocompetent BALB/c mice and T-cell-deficient nude (nu/nu) mice to determine the efficacy of an MR diet on tumor growth and enhancing CD8-positive T-cell tumor infiltration in BALB/c mice. Tumor-infiltrating lymphocytes in the tumor of BALB/c mice were determined with immunohistochemistry. Results: The EC50 values of methionine for K7M2 and 143B were 14.18 μM and 20.85 μM, respectively. Both cell lines had a strong dependence on methionine at the concentration range of 4 to 32 μM. MR using methionine-depleted medium in vitro decreased PD-L1 expression in 143B and K7M2, compared to untreated control cells (pppConclusion: MR alone is a potential immunotherapeutic for osteosarcoma. The present results suggest MR is a T-cell stimulant and not a cause of T-cell exhaustion.

Mark Helpful

Bookmark

Relay

View Full Paper