Celastrol acts synergistically with afatinib to suppress non‐small cell lung cancer cell proliferation by inducing paraptosis

• Published in: Journal of cellular physiology Vol. 236; no. 6; pp. 4538 - 4554
• Main Authors: Dai, Chun‐Hau, Zhu, Li‐Rong, Wang, Yi, Tang, Xing‐Ping, Du, Yong‐Jie, Chen, Yong‐Chang, Li, Jian
• Format: Journal Article
• Language: English
• Published: United States Wiley Subscription Services, Inc 01.06.2021
• Subjects: afatinib; Afatinib - pharmacology; Animals; Anticancer properties; Antineoplastic Combined Chemotherapy Protocols - pharmacology; Antitumor activity; Apoptosis; Autophagy; Calcium (mitochondrial); Calcium - metabolism; Calcium ions; Carcinoma, Non-Small-Cell Lung - drug therapy; Carcinoma, Non-Small-Cell Lung - metabolism; Carcinoma, Non-Small-Cell Lung - pathology; celastrol; Cell death; Cell Death - drug effects; Cell Line, Tumor; Cell proliferation; Cell Proliferation - drug effects; Cholecystokinin; Drug Resistance, Neoplasm; Drug Synergism; Electron microscopy; Endoplasmic reticulum; Endoplasmic Reticulum Stress - drug effects; Epidermal growth factor; epidermal growth factor receptor (EGFR)‐tyrosine kinase inhibitor (TKI); Epidermal growth factor receptors; Growth factors; Humans; Immunofluorescence; In vivo methods and tests; Kinases; Lung cancer; Lung Neoplasms - drug therapy; Lung Neoplasms - metabolism; Lung Neoplasms - pathology; Male; Mice; Mice, Inbred BALB C; Mice, Nude; Mitochondria; Mitochondria - drug effects; Mitochondria - metabolism; Mitochondria - pathology; Mutation; Non-small cell lung carcinoma; non‐small cell lung cancer; paraptosis; Pentacyclic Triterpenes - pharmacology; Phagocytosis; Protein folding; Protein-tyrosine kinase; Reactive oxygen species; Reactive Oxygen Species - metabolism; resistance; Small cell lung carcinoma; Survival; Synergistic effect; Tumors; Tyrosine; Unfolded Protein Response - drug effects; Xenograft Model Antitumor Assays; Xenografts; Xenotransplantation; non-small cell lung cancer; celastrol; afatinib; epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitor (TKI); paraptosis; resistance
• ISSN: 0021-9541; 1097-4652
• DOI: 10.1002/jcp.30172

Non‐small cell lung cancer (NSCLC) with wild‐type epidermal growth factor receptor (EGFR) is intrinsic resistance to EGFR‐tyrosine kinase inhibitors (TKIs), such as afatinib. Celastrol, a natural compound with antitumor activity, was reported to induce paraptosis in cancer cells. In this study, intrinsic EGFR‐TKI‐resistant NSCLC cell lines H23 (EGFR wild‐type and KRAS mutation) and H292 (EGFR wild‐type and overexpression) were used to test whether celastrol could overcome primary afatinib resistance through paraptosis induction. The synergistic effect of celastrol and afatinib on survival inhibition of the NSCLC cells was evaluated by CCK‐8 assay and isobologram analysis. The paraptosis and its modulation were assessed by light and electron microscopy, Western blot analysis, and immunofluorescence. Xenografts models were established to investigate the inhibitory effect of celastrol plus afatinib on the growth of the NSCLC tumors in vivo. Results showed that celastrol acted synergistically with afatinib to suppress the survival of H23 and H292 cells by inducing paraptosis characterized by extensive cytoplasmic vacuolation. This process was independent of apoptosis and not associated with autophagy induction. Afatinib plus celastrol‐induced cytoplasmic vacuolation was preceded by endoplasmic reticulum stress and unfolded protein response. Accumulation of intracellular reactive oxygen species and mitochondrial Ca2+ overload may be initiating factors of celastrol/afatinib‐induced paraptosis and subsequent cell death. Furthermore, Celastrol and afatinib synergistically suppressed the growth of H23 cell xenograft tumors in vivo. The data indicate that a combination of afatinib and celastrol may be a promising therapeutic strategy to surmount intrinsic afatinib resistance in NSCLC cells. Graphical Celastrol acted synergistically with afatinib to suppress the survival of non‐small cell lung cancer cells by inducing paraptosis. Afatinib plus celastrol‐induced paraptosis was mediated by endoplasmic reticulum stress and unfolded protein response. Accumulation of intracellular reactive oxygen species and mitochondrial Ca2+ overload induced by celastrol/afatinib may be an initiating factor of paraptosis and subsequent cell death.