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Archive > Volume 31 Issue 8 > 2024,31(8):777-785. DOI:10.3872/j.issn.1007-385X.2024.08.005 Prev Next
Basic Research
Preparation of targeting CPI-444-loaded nanoparticles and investigation of its effects on T cell activity and anti-tumor response
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Abstract:
Objective：To prepare and characterize CD8 antibody-conjugated CPI-444 (C) -loaded nanoparticles (CNP/α CD8) and investigate their effects on CD8+ T cell activation, proliferation, and anti-tumor activity. Methods：Nanoparticles encapsulating the adenosine A2A receptor(A2AR) antagonist CPI-444 (C) or the fluorescent dye Coumarin-6 (C6) were prepared using the double emulsion solvent evaporation method and EDC/NHS chemistry for antibody conjugation, resulting in CNP/αCD8 and C6NP/αCD8. The morphology and size of the nanoparticles were characterized by scanning electron microscopy and NanoPlus particle size analyzer. Drug loading and release profiles were determined using liquid chromatography-tandem mass spectrometry (LC-MS/MS) and centrifugation. The internalization of C6NP/αCD8 by CD8+ T cells were examined by flow cytometry and fluorescence microscopy. The effects of CNP/αCD8 on the proliferation, activation, cytotoxicity, and tumor killing ability of CD8+ T cells were examined by flow cytometry, ELISA, and lactate dehydrogenase (LDH) assay. Results: The CNP/αCD8 nanoparticles were spherical with an average diameter of about 150 nm, effectively encapsulating CPI-444 and conjugating CD8 antibodies, with a drug encapsulation efficiency and a CD8 antibody conjugation efficiency of approximately 60% and 53.4%, respectively. The nanoparticles exhibited good stability and were efficiently internalized by CD8+ T cells, inhibiting A2AR expression. Biological function assays showed that CNP/αCD8 enhanced CD8+ T cell proliferation, promoted T cell activation, cytokine secretion, granzyme B, and perforin production, and improved the tumorkilling ability of CD8+ T cells. Conclusion：CNP/αCD8 nanoparticles can significantly enhance the immune functions of CD8+ T cells, likely by inhibiting A2AR expression.
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