Construction of glycosylation high fidelity human lung organoids

• 1、Jiangnan University-School of bioengineering,wuxi 214122
• 2、Institute of Process Engineering (IPE),Chinese Academy of Sciences,beijing 100190

Abstract：Lung cancer, as the malignant tumor with the highest mortality rate worldwide, among which lung adenocarcinoma is the most common subtype of non-small cell lung cancer and the most widely studied subtype of lung cancer with somatic gene aberrations. Tumor organoids derived from adult stem cells, as an in vitro model with a relatively high success rate, retain to some extent the genes and tissue characteristics of the original tissue. However, as a glycosylation modification closely related to tumor development, its similarity with the original tissue is currently unclear. This study mainly used a combination of transcriptomics, proteomics, and other omics analysis methods to investigate the similarities and differences between organoid gene expression and protein glycosylation modification at the molecular level and the original tissue. Purpose: To clarify the degree of preservation of gene expression and glycosylation modifications in the original tissue of adult stem cell-derived organoids during 3D culture, and to guide the establishment of tumor organoids that can better maintain the original tissue morphology and molecular characterization of tumors. Method: This study utilized multiple omics joint analysis methods such as deep transcriptomics, TMT labeled quantitative proteomics, and glycation modification prediction tool GlycoMaple analysis to perform morphological characterization, tumor marker identification, gene expression, protein level, and glycation modification level analysis on organoids and primitive tissues. Results: Morphological examination confirmed that lung adenocarcinoma like organs have a cystic structure and identified their key molecular pathological features (Napsin A+, CK7+, TTF1+, P40-) and cellular subtypes of the lung and trachea (basal cells, ciliated cells, goblet cells). Through multi omics joint analysis of organoids and primitive tissues (transcriptomics and proteomics), it was demonstrated that organoids are consistent with primitive tissues at the molecular level, with transcriptional similarity greater than 80%, 48% of sugar chains showing no significant differences, and 91% of high transcriptional proteins in primitive tissues consistent with organoids. The differences are mainly reflected in the absence of immune microenvironment, the orientation of glycoschain synthesis in organoids is high fucose and sialylation, and the high transcriptional ratio glycoproteins unique to organoids are mainly involved in the activation of the classic tumor pathway PI3K/AKT and cell adhesion. The results of this study demonstrate that organoids are a high fidelity 3D culture model that can simulate glycosylation modifications in primitive tissues and have higher purity characteristics than primitive tissues.

Keywords： Proteomic Glycosylation Organoid Multiomics