Abstract 628: Imipramine induced apoptosis and inhibits invasion via suppression of EGFR/ERK/NF-kappa B activation in non-small cells lung cancer
Authors: Yueh Po-Fu , chiang I-Tsang , Hsu Fei-Ting 
Published: September 2020

Imipramine was known as an anti-depression agent; interestingly, recent studies indicated the potential of various tumors growth inhibition. However, the anticancer mechanism of imipramine on non-small cell lung cancer (NSCLC) is remaining ambiguous. Thus, the aim of the study was to determent the effects and underlying mechanism of imipramine on human NSCLC CL1-5/F4 cells. MTT assays were used for testing the cell viability of imipramine. The activation and expression of apoptosis, anti-apoptosis, DNA damage, cellular calcium, and metastasis-associated proteins were observed by flow cytometry, immunofluorescence stain, comet assay and Western blotting assay. The inhibition of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) after imipramine treatment was performed by NF-κB reporter gene assay and immunofluorescence nuclear translocation assay on CL1-5/F4/NF-κB/luc2 and CL1-5/F4 wild type cells. Moreover, the ability of migration and invasion after imipramine treatment were also tested by trans-well assay and wound healing assay. In animal experiment, CL1-5/F4/NF-κB/luc2 cells were subcutaneously injection into nude mice right flask and treated with imipramine by oral. In addition, the standard treatment erlotinib was also used as a positive control to compare treatment efficacy of imipramine. In vivo NF-κB activation was investigated by optical image. Mice were sacrificed and tumors were extracted for size and IHC staining evaluation on day 14. Our results demonstrated that imipramine significantly reduced cell viability, cell migration and invasion. Reporter gene assay and Western blotting revealed imipramine may decrease NF-κB activation through inhibition of EGFR/ERK/NF-κB pathway. Imipramine also diminished the phosphorylation of ERK, the expression of anti-apoptotic (c-FLIP and XIAP), angiogenesis (VEGF), and metastasis-associated proteins (MMP-9). Additionally, the activity of PARP-1 that induced by imipramine was associated with DNA damage. Furthermore, imipramine was also triggered extrinsic (Fas/FasL, and caspase-8), intrinsic [mitochondrial membrane potential (ΔΨm)] apoptosis pathways, and the accumulation of cellular calcium. Tumor growth and NF-κB activation were both suppressed by imipramine treatment. Tumors IHC stain showed p-EGFR, p-ERK and p-NF-κB expressions level were reduced in imipramine treatment group as compared to vehicle. Most importantly, although body weight was remained unchanged in three treatment group, the cellular necrosis nucleus was found in erlotinib treated liver pathology H&E stain which was not found in control and imipramine groups. Taken together, these results suggested that imipramine can induce NSCLC apoptosis by activating extrinsic/intrinsic apoptosis mechanisms, DNA damage effect and the accumulation of cellular calcium concentration. In addition, imipramine may also suppress tumor proliferation, invasion and migration ability. Most importantly, imipramine diminished tumor progression via inactivation of EGFR/ERK/NF-κB signaling on NSCLC in vitro and in vivo.

Citation Format: Po-Fu Yueh, I-Tsang chiang, Fei-Ting Hsu. Imipramine induced apoptosis and inhibits invasion via suppression of EGFR/ERK/NF-kappa B activation in non-small cells lung cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 628.