Single-cell EMT-related transcriptional analysis revealed intra-cluster heterogeneity of tumor cell clusters in epithelial ovarian cancer ascites

Abstract

Malignant ascites of epithelial ovarian cancer is a metastatic tumor microenvironment in which large amounts of disseminated single cells (DSCs) and disseminated tumor cell clusters (DTCCs) are commonly observed. The tumor cell clusters are known to be more aggressive than individual tumor cells in cancer metastasis; however, little is known about the mechanism. Applying single-cell epithelial-to-mesenchymal transition (EMT)-related transcriptional analysis in 120 DSCs and 195 intra-cluster cells from 27 DTCCs, we demonstrated that DTCCs were heterogeneous cellular units comprised of epithelial tumor cells, leukocytes, and cancer-associated fibroblasts (CAFs). Through the analysis of intra-DTCC heterogeneity, we identified that CAFs induced EMT of tumor cells via TGFβ signaling within the DTCC microenvironment. The activation of EMT program, in particular the upregulation of ZEB2, enabled the acquisition of additional chemoresistance and metastasis abilities of the intra-DTCC tumor cells, which resulted in the aggressiveness of DTCCs.

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Fig. 1: Study workflow and the characterization of ascitic cellular components.
Fig. 2: Unsupervised classifications of DSCs.
Fig. 3: The identity of EpCAM−/CD45− subpopulation is CAFs.
Fig. 4: Intra-cluster heterogeneity analysis of DTCCs.
Fig. 5: Intra-DTCC CAFs promote the progression of intra-DTCC tumor cells via TGFβ signaling.
Fig. 6: ZEB2 is the most significantly upregulated EMT signature gene in intra-DTCC tumor cells.
Fig. 7: A schematic model illustrates the mechanism of DTCC aggressiveness.

Data availability

The single-cell qPCR data generated in this study have been deposited in the NCBI Gene Expression Omnibus (GEO) under accession code GSE126192.

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Acknowledgements

We thank Maggie K.S. Tang from the School of Biological Sciences, The University of Hong Kong for assistance in delivering ascites samples and ovarian cancer cell lines. This work was supported by grants from the General Research Fund (CityU_11303815) and the Collaborative Research Fund (CRF/1013-15G) of Hong Kong Research Grant Council, the Guangdong Frontier and Key Technology Development Fund (2017B020226001) of Guangdong Province, PR China, and the Knowledge Innovation Program (JCYJ20150601102053070) of Shenzhen Municipality, PR China.

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TK designed the study, carried out the experiments, acquired and analyzed the single-cell data, interpreted the data, and drafted the paper. WW did the public data analysis and drafted the related computation methods. PPI provided clinical samples. SZ designed the ZEB2 functional study. ASW provided the cell lines and gave suggestions to improve the study. XW contributed to the supervision of bioinformatic computation. MY supervised the project, guided the study concept and design, revised the paper, and provided funding and other supports for the project.

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Correspondence to Mengsu Yang.

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Kan, T., Wang, W., Ip, P.P. et al. Single-cell EMT-related transcriptional analysis revealed intra-cluster heterogeneity of tumor cell clusters in epithelial ovarian cancer ascites. Oncogene 39, 4227–4240 (2020). https://doi.org/10.1038/s41388-020-1288-2

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