Study flowchart. (IMAGE)
Caption
Part I integrated single-cell transcriptomic and metabolomic profiling to reveal cellular interactions in the breast cancer tumor microenvironment, characterize metabolic programs across TAM subsets, and identify marked downregulation of the glutamine transporter SLC38A2 alongside extensive glutamine metabolic defects. These analyses revealed impaired glutamine utilization as a central metabolic vulnerability of tumor-associated macrophages. Part II engineered metabolically enhanced HER2-targeted CAR-macrophages by overexpressing SLC38A2. This part included CAR construct design; adenoviral transduction of macrophages; and validation of CAR and SLC38A2 expression via western blotting, qPCR, flow cytometry, and immunofluorescence. Part III assessed the functional consequences of SLC38A2 overexpression. In vitro assays demonstrated greater glutamine uptake and markedly greater phagocytosis of HER2+ tumor cells by SLC38A2-overexpressing CAR-Ms than conventional CAR-Ms. These enhanced functional activities translated to in vivo results indicating that SLC38A2/anti-HER2 CAR-M treatment significantly suppressed tumor growth in HER2+ tumor-bearing mice. Collectively, these findings established that metabolic engineering via SLC38A2 restored glutamine fitness and markedly augmented the antitumor efficacy of HER2-targeted CAR-macrophages. CAR-M, chimeric antigen receptor macrophage; TAM, tumor-associated macrophage; TME, tumor microenvironment; WB, western blotting; qPCR, quantitative PCR; FC, flow cytometry; IF, immunofluorescence.
Credit
Cancer Biology & Medicine
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CC BY-NC