Theranostics 2021; 11(8):3964-3980. doi:10.7150/thno.54824

Research Paper

Downregulation of death receptor 4 is tightly associated with positive response of EGFR mutant lung cancer to EGFR-targeted therapy and improved prognosis

Shuo Zhang1, Zhen Chen1, Puyu Shi1, Songqing Fan2, Yong He3, Qiming Wang4, Yixiang Li1, Suresh S. Ramalingam1, Taofeek K. Owonikoko1, Shi-Yong Sun1✉

1. Department of Hematology and Medical Oncology, Emory University School of Medicine and Winship Cancer Institute, Atlanta, Georgia, USA.
2. Department of Pathology, The Second Xiangya Hospital, Central South University, Changsha, Hunan, China.
3. Department of Respiratory Disease, Daping Hospital, Chongqing, China.
4. Department of Internal Medicine, The Affiliated Cancer Hospital of Zhengzhou University, Henan Cancer Hospital, Zhengzhou, Henan, China.

This is an open access article distributed under the terms of the Creative Commons Attribution License ( See for full terms and conditions.
Zhang S, Chen Z, Shi P, Fan S, He Y, Wang Q, Li Y, Ramalingam SS, Owonikoko TK, Sun SY. Downregulation of death receptor 4 is tightly associated with positive response of EGFR mutant lung cancer to EGFR-targeted therapy and improved prognosis. Theranostics 2021; 11(8):3964-3980. doi:10.7150/thno.54824. Available from

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Death receptor 4 (DR4), a cell surface receptor, mediates apoptosis or induces inflammatory cytokine secretion upon binding to its ligand depending on cell contexts. Its prognostic impact in lung cancer and connection between EGFR-targeted therapy and DR4 modulation has not been reported and thus was the focus of this study.

Methods: Intracellular protein alterations were measured by Western blotting. Cell surface protein was detected with antibody staining and flow cytometry. mRNA expression was monitored with qRT-PCR. Gene transactivation was analyzed with promoter reporter assay. Drug dynamic effects in vivo were evaluated using xenografts. Gene modulations were achieved with gene overexpression and knockdown. Proteins in human archived tissues were stained with immunohistochemistry.

Results: EGFR inhibitors (e.g., osimertinib) decreased DR4 levels only in EGFR mutant NSCLC cells and tumors, being tightly associated with induction of apoptosis. This modulation was lost once cells became resistant to these inhibitors. Increased levels of DR4 were detected in cell lines with acquired osimertinib resistance and in NSCLC tissues relapsed from EGFR-targeted therapy. DR4 knockdown induced apoptosis and augmented apoptosis when combined with osimertinib in both sensitive and resistant cell lines, whereas enforced DR4 expression significantly attenuated osimertinib-induced apoptosis. Mechanistically, osimertinib induced MARCH8-mediated DR4 proteasomal degradation and suppressed MEK/ERK/AP-1-dependent DR4 transcription, resulting in DR4 downregulation. Moreover, we found that DR4 positive expression in human lung adenocarcinoma was significantly associated with poor patient survival.

Conclusions: Collectively, we suggest that DR4 downregulation is coupled to therapeutic efficacy of EGFR-targeted therapy and predicts improved prognosis, revealing a previously undiscovered connection between EGFR-targeted therapy and DR4 modulation.

Keywords: EGFR inhibitors, osimertinib, death receptor 4, apoptosis, acquired resistance