Abstract:Objective To explore the mechanism of secondary resistance to imatinib(IM) in gastrointestinal stromal tumors(GIST). Methods Ten IM-resistant and ten IM-sensitive GIST patients admitted to the First Affiliated Hospital with Nanjing Medical University from December 2018 to April 2021 were selected, and their tumor tissues were obtained by surgery or needle biopsy. The IM-resistant strain GIST-T1R of GIST was constructed by gradient addition using purchased IM-sensitive parental cells GIST-T1S. Real-time quantitative polymerase chain reaction, immunohistochemistry and western blot were used to detect the expression of erythropoietin-producing human hepatocellular （EPH) B2 in IM-sensitive or IM-resistant cells and tissues at RNA and protein levels, respectively. After knockdown or overexpression of EPHB2 in GIST cell lines, CCK-8 assay was used to detect cell viability to determine the half inhibitory concentration(IC₅₀) of IM on cells, and clone formation assay was used to detect the effect of EPHB2 on cell proliferation ability. Results EPHB2 was significantly overexpressed in GIST tissues of patients with secondary IM-resistance(P＜0.01).Compared with GIST-T1S cells, EPHB2 RNA and protein levels were significantly up-regulated in GIST-T1R cells(P＜0.01). In GIST-T1S cells, overexpressing EPHB2 significantly increased the IC₅₀ of IM on cells, and knocking down of EPHB2 significantly decreased the IC₅₀ of IM on cells(P＜0.01). Conclusion EPHB2 mediates IM-resistance of GIST cells and promotes their malignant proliferation.