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Theranostics 2020; 10(11):5107-5119. doi:10.7150/thno.44705 This issue Cite

Research Paper

Combinative treatment of β-elemene and cetuximab is sensitive to KRAS mutant colorectal cancer cells by inducing ferroptosis and inhibiting epithelial-mesenchymal transformation

Peng Chen^1,2,3*, Xuejie Li^4*, Ruonan Zhang^2,3, Shuiping Liu^2,3, Yu Xiang^2,3, Mingming Zhang^2,3, Xiaying Chen^2,3, Ting Pan^2,3, Lili Yan^2,3, Jiao Feng^2,3, Ting Duan^2,3, Da Wang⁶, Bi Chen^2,3, Ting Jin^2,3, Wengang Wang^2,3, Liuxi Chen^2,3, Xingxing Huang^2,3, Wenzheng Zhang^2,3, Yitian Sun^2,3, Guohua Li^2,3, Lingpan Kong^2,3, Xiaohui Chen⁵, Yongqiang Li^2,3, Zuyi Yang⁵, Qin Zhang^2,3, Lvjia Zhuo^2,3, Xinbing Sui^2,3✉, Tian Xie^1,2,3✉

1. Institute of Chinese Materia Medica, Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
2. Holistic Integrative Pharmacy Institutes and Department of Medical Oncology, The Affiliated Hospital of Hangzhou Normal University, College of Medicine, Hangzhou Normal University, Hangzhou, Zhejiang, China
3. Key Laboratory of Elemene Class Anti-cancer Chinese Medicine of Zhejiang Province, Hangzhou Normal University, Hangzhou, Zhejiang, China
4. Department of Pathology, The First Affiliated Hospital of Medical School of Zhejiang University, Hangzhou, China
5. Department of Hematology and Oncology, the Affiliated Hospital of Hangzhou Normal University, College of Medicine, Hangzhou Normal University, Hangzhou, Zhejiang, China
6. Department of Colorectal Surgery, The Second Affiliated Hospital of Zhejiang University School of Medicine, Hangzhou, China
* These authors contributed equally to this work

✉ Corresponding authors: Tian Xie, Email: xbsedu.cn or Xinbing Sui, Email: hzzjuedu.cn

Abstract

Background and Purpose: RAS mutations limit the effectiveness of anti-epidermal growth factor receptor (EGFR) monoclonal antibodies in combination with chemotherapy for metastatic colorectal cancer (mCRC) patients. Therefore, new cell death forms have focused on identifying indirect targets to inhibit Ras-induced oncogenesis. Recently, emerging evidence has shown the potential of triggering ferroptosis for cancer therapy, particularly for eradicating aggressive malignancies that are resistant to traditional therapies.

Methods: KRAS mutant CRC cell HCT116 and Lovo were treated with cetuximab and β-elemene, a bioactive compound isolated from Chinese herb Curcumae Rhizoma. Ferroptosis and epithelial-mesenchymal transformation (EMT) were detected in vitro and in vivo. Orthotopic CRC animal model were established and the tumor growth was monitored by IVIS bioluminescence imaging. Tumor tissues were collected to determine ferroptosis effect and the expression of EMT markers after the treatment.

Results: CCK-8 assay showed that synergetic effect was obtained when 125 µg/ml β-elemene was combined with 25 µg/ml cetuximab in KRAS mutant CRC cells. AV/PI staining suggested a non-apoptotic mode of cell death after the treatment with β-elemene and cetuximab. In vitro, β-elemene in combination with cetuximab was shown to induce iron-dependent reactive oxygen species (ROS) accumulation, glutathione (GSH) depletion, lipid peroxidation, upregulation of HO-1 and transferrin, and downregulation of negative regulatory proteins for ferroptosis (GPX4, SLC7A11, FTH1, glutaminase, and SLC40A1) in KRAS mutant CRC cells. Meanwhile, combinative treatment of β-elemene and cetuximab inhibited cell migration and decreased the expression of mesenchymal markers (Vimentin, N-cadherin, Slug, Snail and MMP-9), but promoted the expression of epithelial marker E-cadherin. Moreover, ferroptosis inhibitors but not other cell death suppressors abrogated the effect of β-elemene in combination with cetuximab on KRAS mutant CRC cells. In vivo, co-treatment with β-elemene and cetuximab inhibited KRAS mutant tumor growth and lymph nodes metastases.

Conclusions: Our data for the first time suggest that the natural product β-elemene is a new ferroptosis inducer and combinative treatment of β-elemene and cetuximab is sensitive to KRAS mutant CRC cells by inducing ferroptosis and inhibiting EMT, which will hopefully provide a prospective strategy for CRC patients with RAS mutations.

Keywords: β-elemene, KRAS mutation, colorectal cancer, ferroptosis, epithelial-mesenchymal transformation

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