662 Targeting the α9-nAChR/PD-L1 axis in antiproliferative effects of Allium cepa L. var. proliferum Regel extract on melanoma cells

Abstract

Programmed cell death ligand-1 (PD-L1) is critical for melanoma development, progression and treatment. The α9 nicotinic acetylcholine receptor (nAChR) is shown to affect melanoma cell proliferation. Allium cepa L. var. proliferum Regel extract possesses anticancer properties. However, the mechanism of Allium cepa L. var. proliferum Regel extract in the antiproliferative effects of melanoma cells remains unclear. Using UPLC-ESI-MS/MS method, a total of 42 compounds were identified in Allium cepa L. var. proliferum Regel extract classified into five categories, including organosulfur compounds (962.20 ± 34.55 μg/g), polyphenols (100.40 ± 12.55 μg/g), flavonoids (58.36 ± 10.75 μg/g), organic acids (54.04 ± 2.69 μg/g), and alkaloids (15.08 ± 3.10 μg/g) where we found various bioactive components targeted to nAChRs. We analyzed the genetic dependency of α1-10 nAChRs from Achilles_CRISPR dataset (n=38) using CRISPR-Cas9 technology. α9-nAChR contributed significantly to the survival of melanoma cells (dependency scores < 0). We evaluated α1-10 nAChRs and PD-L1 mRNA expression from CCLE_Expression dataset (n=50). α9-nAChR was the only gene that strongly correlated to PD-L1 expression (r=0.6, p<0.001). There was also a strong correlation between α9-nAChR and PD-L1 expression (r=0.7, p<0.001) in the tissue microarray (n=192) using the IHC staining. Knockdown or overexpression of α9-nAChR significantly down- or upregulated the expression of PD-L1 via the transcription factor STAT3 binding to the PD-L1 promoter, respectively, and affected melanoma cell proliferation. Allium cepa L. var. proliferum Regel extract inhibited melanoma cell proliferation through inactivation of α9-nAChR/PD-L1 axis. Our results suggest that α9-nAChR/ PD-L1 axis regulates the cell proliferation of melanoma and Allium cepa L. var. proliferum Regel extract is considered as a potential candidate to prevent melanoma.