HLA-G gene editing in tumor cell lines as a novel tool in cancer therapies

PALMA MARÍA BELÉN; TRONIK-LE ROUX, DIANA; SEOANE ROCHA, CAMILA; IRIBARNE, AILEN; LA GRECA, ALEJANDRO; MORO, LUCÍA N.; DAOUYA, MARINA; PORAS, ISABELLE; CAROSELLA, EDGARDO D.; GARCÍA, MARCELA N.; MIRIUKA, SANTIAGO G.

Paris

Conferencia; 9th International Conference on HLA-G; 2022

CEA, DRF-Francois Jacob Institute SRHI, Saint-Louis Hospital

Cancer immunotherapies, based mainly on the blockade of immune-checkpoint (IC) molecules by anti-IC antibodies, offer new alternatives for the treatment of oncological diseases. However, a considerable proportion of patients remain unresponsive to these therapies and therefore novel clinical approaches and/or targets should be identified. In this context, the IC HLA-G has caused great interest since it is abnormally expressed in several tumors generating a tolerogenic microenvironment. The fact that Its expression is highly restricted under physiological conditions in tissues and organs makes HLA-G a promising target for the development of new immunotherapies, with relatively low chances of generating significant side effects. Here, we used CRISPR/Cas9 gene editing to block the expression of HLA-G in two tumor cell lines, which express high levels of this protein, named: renal cell carcinoma (RCC7) and choriocarcinoma (JEG-3). We design four different single guide-RNA targeting HLA-G exon 1 and 2. A downregulation of HLA-G was reached to different degrees in both tumor cell lines, but neither of these could generated a complete HLA-G knockout. This was only achieved in JEG-3 cells using all 4 sgRNAs simultaneously. This suggest that different isoforms might be expressed from the two translation start sites (located in exon 1 or 2). The NK degranulation assay showed that edited JEG-3 cells (HLA-G negatives) triggered a higher in vitro response of NK cells compared to wild type JEG-3 cells (HLA-G positives). Altogether, we demonstrated for the first time the downregulation of HLA-G through gene editing, with a concomitant effect on immune cell activation. This approach might constitute a novel cancer immunotherapy strategy.