The HLA-G Knockout NCI-H1975 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population designed to disrupt the HLA-G gene in human NCI-H1975 lung adenocarcinoma cells. This polyclonal pool provides a loss-of-function model of the non-classical MHC class I immune checkpoint ligand HLA-G, enabling investigation of its role in immune evasion. The knockout abrogates HLA-G-mediated inhibitory signaling, restoring anti-tumor immune responses in co-culture systems.
The parental NCI-H1975 cell line is a human non-small cell lung carcinoma (NSCLC) epithelial model harboring an activating EGFR L858R mutation. This well-characterized line is extensively used to study oncogenic signaling, drug resistance, and tumor biology. When combined with HLA-G disruption, these cells offer a relevant platform to examine the interplay between EGFR-driven oncogenesis and immune checkpoint pathways in lung adenocarcinoma.
HLA-G exerts immunosuppressive functions by binding inhibitory receptors LILRB1 (ILT2), LILRB2 (ILT4), and KIR2DL4 on NK cells, T cells, and myeloid cells. Ligand engagement induces ITIM-mediated recruitment and activation of phosphatases SHP-1 (PTPN6) and SHP-2 (PTPN11). These phosphatases dephosphorylate ZAP70, inhibit NF-??B, and promote IL-10 secretion, collectively suppressing cytotoxicity and inflammatory cytokine production. HLA-G expression is upregulated by IL-10, IFN-??, TGF-??, HIF1A, and CREB1, and is linked to the JAK-STAT pathway. Thus, the knockout eliminates these downstream inhibitory events.
In NCI-H1975 cells, HLA-G contributes to immune escape by protecting tumor cells from NK cell and CTL-mediated lysis. CRISPR-mediated disruption of HLA-G is expected to reverse this protection, rendering the cells sensitive to immune attack. This makes the model valuable for dissecting tumor-intrinsic immune checkpoint mechanisms and for evaluating therapeutic strategies targeting the HLA-G/ILT axis, particularly in the context of EGFR-mutant NSCLC.
Research applications include NK cell and T cell co-culture assays to assess restored cytotoxicity, proliferation, and cytokine output (e.g., IFN-??, TNF-??). The cells are suitable for screening HLA-G-targeted biologics, profiling immune gene expression via RNA-seq, and biochemical validation using Western blotting, flow cytometry, and RT-qPCR. The polyclonal format ensures experimental reproducibility while avoiding clonal artifacts. For additional details or technical inquiries, please contact Ascent Research.
