Study validates immune protein upregulation in cancer-immune cell co-culture using MRM technique
image: Targeted phase proteomics by multiple reaction monitoring showing upregulation of immune activated proteins in immune cells in Gipie-silenced ACC -immune co-culture model. (A) Granzyme A, (C) CD48, (E) Granzyme B, (G) HLA class I, antigen B, (I) HLA class I, antigen A; normalised relative protein expression graphs done with Graph Pad prism 9.4.1 showing t-test (Wilcoxon matched-pairs signed rank test) comparison data of immune proteins of NK cells from UM-HACC-2A co-culture model compared to NK cells from Gipie-silenced UM-HACC-2A co-culture model. ∗P≤ 0.05, ∗∗∗P≤ 0.0001. n = 24 biological replicates at different time points. (B, D, F, H, J). MRM ion chromatogram peaks of analytes. All peaks: Gaussian smooth width 0, retention time half window 30 s, noise percentage 40 %, baseline sub window 2 min, and peak splitting 2 points. (B) Granzyme A, +2, GVTSFGLENK. All three transitions (y8, y6, y5) shown. Expected retention time 32.70 min, (D) CD48, +2, LDPQSGALYISK. All three transitions (y10, y9, y8) shown. Expected retention time 32.70 min. (F) Granzyme B, +2, GDSGGPLVCNK. All three transitions (y8, y7, y6) shown. Expected retention time 36.49 min. (H) HLA class I, antigen B, +2, AYLEGECVEWLR. All three transitions (y9, y8, y6) shown. Expected retention time 47.60 min, (J) HLA class I, antigen A, +2, FIAVGYVDDTQFVR. All three transitions (y10, y8, y7) shown. Expected retention time 48.85 min.
Credit: Rajdeep Chakraborty, Thiri Zaw.
Summary:
A recent study published in LabMed Discovery utilized Multiple Reaction Monitoring (MRM), a targeted mass spectrometry technique, to validate the upregulation of immune-activated proteins in a 3D co-culture model of adenoid cystic carcinoma (ACC) and immune cells. The research focused on the effects of silencing the protein Gipie in UM-HACC-2A cancer cells, which led to significant increases in immune-related proteins such as Granzyme A, Granzyme B, CD48, and HLA class I antigens. These findings confirm enhanced immune cell cytotoxicity against cancer cells. The study highlights MRM as a reliable alternative to antibody-based methods for protein validation, offering high sensitivity and reproducibility. The work underscores the potential of MRM in translational biomedical research, particularly for preclinical studies involving novel drug testing and personalized therapies.
Key Findings:
Silencing Gipie in ACC cells upregulated immune proteins in co-cultured NK-92 cells.
MRM validated differential expression of proteins linked to immune activation.
The technique demonstrated consistency and reproducibility, supporting its use in proteomic research.
The study provides a foundation for further exploration of immune evasion mechanisms in cancer.
Implications:
The research advances understanding of immune responses in cancer and showcases MRM's utility in validating proteomic data, paving the way for future therapeutic developments.