Down-regulation of circITCH promotes osteosarcoma development and resistance to doxorubicin via the miR-524/RASSF6 axis

J Gene Med. 2021 Oct;23(10):e3373. doi: 10.1002/jgm.3373. Epub 2021 Jul 20.


Background: Osteosarcoma (OS) is a malignant bone cancer, in which circular RNAs (circRNAs) act as important modulators. The present study aimed to explore the functional role of circRNA itchy E3 ubiquitin protein ligase (circITCH) in the development and doxorubicin (DXR) resistance of OS and the possible mechanistic pathway.

Methods: A quantitative real-time polymerase chain reaction or western blot assays were exploited to analyze the expression of circITCH, miR-524 and Ras association domain family member 6 (RASSF6). Cell viability and half-maximal inhibitory concentration (IC50 ) value of DXR were monitored using a cell counting kit-8 assay. Cell migration, invasion and apoptosis were determined via a transwell assay and flow cytometry. The target interaction among circITCH, miR-524 and RASSF6 was validated by dual-luciferase reporter and RNA immunoprecipitation assays. A xenograft model of MG-63/DXR cells stably expressing circITCH in nude mice was established for assessing the role of circITCH in vivo.

Results: Down-regulation of circITCH and RASSF6, as well as the up-regulation of miR-524, was revealed in OS by investigating 40 paired OS tissue and normal tissue samples. Overexpression of circITCH lowered the cell viability, IC50 value of DXR, migration and invasion, whereas it facilitated apoptosis of OS cells. circITCH sponged miR-524 to up-regulate RASSF6, causing OS progression inhibition and DXR resistance reduction. Additionally, circITCH up-regulation reduced tumor growth in vivo.

Conclusions: Transduction with circITCH represses OS progression and promotes DXR sensitivity by the miR-524/RASSF6 axis, providing a new perspective for therapeutic intervention.

Keywords: RASSF6; circITCH; doxorubicin resistance; miR-524; osteosarcoma.

MeSH terms

  • Animals
  • Apoptosis / genetics
  • Apoptosis Regulatory Proteins / genetics*
  • Bone Neoplasms / genetics*
  • Cell Line, Tumor
  • Cell Movement / genetics
  • Cell Proliferation / genetics
  • Cell Survival / genetics
  • Down-Regulation / genetics*
  • Doxorubicin / pharmacology
  • Drug Resistance, Neoplasm / genetics*
  • Gene Expression Regulation, Neoplastic / genetics
  • Humans
  • Male
  • Mice
  • Mice, Inbred BALB C
  • Mice, Nude
  • MicroRNAs / genetics*
  • Osteosarcoma / genetics*
  • RNA, Circular / genetics*
  • Up-Regulation / genetics


  • Apoptosis Regulatory Proteins
  • MIRN-524 microRNA, human
  • MicroRNAs
  • RASSF6 protein, human
  • RNA, Circular
  • Doxorubicin