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RON Receptor Tyrosine Kinase in Tumorigenic Stemness as a Therapeutic Target of Antibody-Drug Conjugates for Eradication of Triple-Negative Breast Cancer Stem Cells

Curr Cancer Drug Targets. 2022 Aug 25. doi: 10.2174/1568009622666220825115528. Online ahead of print.

ABSTRACT

BACKGROUND: Cancer stem-like cells in triple-negative breast cancer (TNBC-SLCs) are the tumorigenic core for malignancy. Aberrant expression of the RON receptor tyrosine kinase has been implication in TNBC tumorigenesis and malignancy.

OBJECTIVE: Here we identified the RON receptor as a pathogenic factor contributing to TNBC cell stemness and validated anti-RON antibody-drug conjugate Zt/g4-MMAE for eradication of RON-expressing TNBC-SLCs.

METHODS: Immunofluorescence and Western blotting were used for analyzing cellular marker expression. TNBC-SLCs were isolated by magnetic-immunofluorescence cell-sorting techniques. Spheroids were generated using the ultralow adhesion culture methods. Levels of TNBC-SLC chemosensitivity were determined by MTS assays. TNBC-SLC mediated tumor growth was determined in athymic nude mice. The effectiveness of Zt/g4-induced RON internalization was measured by immunofluorescence analysis. Efficacies of Zt/g4-MMAE in killing TNBC-SLCs in vitro and in eradicating TNBC-SLC-mediated tumors were determined in mouse models. All data were statistically analyzed using the GraphPad Prism 7 software.

RESULTS: Increased RON expression existed in TNBC-SLCs with CD44+/CD24- phenotypes and ALDH activities and facilitates epithelial to mesenchymal transition. RON-positive TNBC-SLCs had enhanced spheroid-formatting capability compared with RON-negative TNBC-SLCs, which was sensitive to small molecule kinase inhibitor BMS-777607. Increased RON expression also promoted TNBC-SLC chemoresistance and facilitated tumor growth at an accelerated rate. In vitro, Zt/g4-MMAE caused massive TNBC-SLC death with an average IC50 value of ~1.56 µg per/ml and impaired TNBC cell spheroid formation. In mice, Zt/g4-MMAE effectively inhibited and/or eradicated TNBC-SLC mediated tumors in a single agent regimen.

CONCLUSIONS: Sustained RON expression contributes to TNBC-SLC tumorigenesis. Zt/g4-MMAE is effective in vivo in killing TNBC-SLC-mediated xenograft tumors. Our findings highlight the feasibility of Zt/g4-MMAE for eradication of TNBC-SLCs in the future.

PMID:36028965 | DOI:10.2174/1568009622666220825115528

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