Macrolides sensitize EGFR-TKI-induced non-apoptotic cell death via blocking autophagy flux in pancreatic cancer cell lines

Abstract

Pancreatic cancer is one of the most difficult types of cancer to treat because of its high mortality rate due to chemotherapy resistance. We previously reported that combined treatment with gefitinib (GEF) and clarithromycin (CAM) results in enhanced cytotoxicity of GEF along with endoplasmic reticulum (ER) stress loading in non-small cell lung cancer cell lines. An epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) such as GEF induces autophagy in a pro-survival role, whereas CAM inhibits autophagy flux in various cell lines. Pronounced GEF-induced cytotoxicity therefore appears to depend on the efficacy of autophagy inhibition. In the present study, we compared the effect on autophagy inhibition among such macrolides as CAM, azithromycin (AZM), and EM900, a novel 12-membered non-antibiotic macrolide. We then assessed the enhanced GEF-induced cytotoxic effect on pancreatic cancer cell lines BxPC-3 and PANC-1. Autophagy flux analysis indicated that AZM is the most effective autophagy inhibitor of the three macrolides. CAM exhibits an inhibitory effect but less than AZM and EM900. Notably, the enhancing effect of GEF-induced cytotoxicity by combining macrolides correlated well with their efficient autophagy inhibition. However, this pronounced cytotoxicity was not due to upregulation of apoptosis induction, but was at least partially mediated through necroptosis. Our data suggest the possibility of using macrolides as ‘chemosensitizers’ for EGFR-TKI therapy in pancreatic cancer patients to enhance non-apoptotic tumor cell death induction.

Macrolide Antibiotics Exhibit Cytotoxic Effect under Amino Acid-Depleted Culture Condition by Blocking Autophagy Flux in Head and Neck Squamous Cell Carcinoma Cell Lines

“…Worth noting was that the LC3B-II upregulation by CAM was transient, whereas the LC3B-II upregulation by AZM persisted and further increased during 60 min exposure. This may be because AZM is more potent than CAM in blocking autophagy flux as we previously described in pancreatic cell lines; however, other molecular mechanisms may be involved [13]. …”

“…Immunoblotting with anti-GAPDH mAb was used as an internal control. Cell lysate derived from PANC-1 cells treated with AZM for 24 hrs was used as a positive control [13]. (B) (upper panel) Immunoblotting of LC3 from the cell lysates of CAL 27 cells cultured in the complete culture medium or AAD culture medium containing 10% FBS with or without AZM (50 μM) and CAM (50 μM) for up to 60 min.…”

“…Immunoblotting with anti-GAPDH mAb was used as an internal control. Cell lysate derived from PANC-1 cells treated with AZM (50 μM) for 24 hrs was used as a positive control [13]. (lower panel) The ratios of LC3B-II/GAPDH were plotted.…”

“…We previously reported that clinically well-used macrolide antibiotics, such as azithromycin (AZM) and clarithromycin (CAM), block autophagy flux [12, 13]. In multiple myeloma and breast cancer cells, we found that apoptosis induction was enhanced by the proteasome inhibitor bortezomib and AZM or CAM combination treatment together with endoplasmic reticulum (ER)-stress loading [12, 14].…”

Is clarithromycin a potential treatment for cachexia in people with lung cancer? A feasibility study

“…For example, although azithromycin has no 'track record' in cachexia, it also has immunomodulatory properties [17,18], is used long-term [19,20], and has a lower risk of a drug-drug interaction, such that only 10 of our patients would have been excluded on this basis. A formal feasibility study is required to confirm improved recruitment and to explore other aspects of the study still untested in our patient group.…”