title: Macrolide Antibiotics Exhibit Cytotoxic Effect under Amino Acid-Depleted Culture Condition by Blocking Autophagy Flux in Head and Neck Squamous Cell Carcinoma Cell Lines.
abstract: Autophagy, a self-digestive system for cytoplasmic components, is required to maintain the amino acid pool for cellular homeostasis. We previously reported that the macrolide antibiotics azithromycin (AZM) and clarithromycin (CAM) have an inhibitory effect on autophagy flux, and they potently enhance the cytocidal effect of various anticancer reagents in vitro. This suggests that macrolide antibiotics can be used as an adjuvant for cancer chemotherapy. Since cancer cells require a larger metabolic demand than normal cells because of their exuberant growth, upregulated autophagy in tumor cells has now become the target for cancer therapy. In the present study, we examined whether macrolides exhibit cytotoxic effect under an amino acid-starving condition in head and neck squamous cancer cell lines such as CAL 27 and Detroit 562 as models of solid tumors with an upregulated autophagy in the central region owing to hypovascularity. AZM and CAM induced cell death under the amino acid-depleted (AAD) culture condition in these cell lines along with CHOP upregulation, although they showed no cytotoxicity under the complete culture medium. CHOP knockdown by siRNA in the CAL 27 cells significantly suppressed macrolide-induced cell death under the AAD culture condition. CHOP-/- murine embryonic fibroblast (MEF) cell lines also attenuated AZM-induced cell death compared with CHOP+/+ MEF cell lines. Using a tet-off atg5 MEF cell line, knockout of atg5, an essential gene for autophagy, also induced cell death and CHOP in the AAD culture medium but not in the complete culture medium. This suggest that macrolide-induced cell death via CHOP induction is dependent on autophagy inhibition. The cytotoxicity of macrolide with CHOP induction was completely cancelled by the addition of amino acids in the culture medium, indicating that the cytotoxicity is due to the insufficient amino acid pool. These data suggest the possibility of using macrolides for "tumor-starving therapy".
authors.affiliation: Department of Otolaryngology (Head and Neck Surgery), Tokyo Medical University, Tokyo, Japan.
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Hirasawa
authors.affiliation: Department of Biochemistry, Tokyo Medical University, Tokyo, Japan.
authors.forename: Shota
authors.initials: S
authors.lastname: Moriya
authors.affiliation: Department of Breast Surgery, Tokyo Medical University, Tokyo, Japan.
authors.forename: Kana
authors.initials: K
authors.lastname: Miyahara
authors.affiliation: Department of Biochemistry, Tokyo Medical University, Tokyo, Japan.
authors.forename: Hiromi
authors.initials: H
authors.lastname: Kazama
authors.affiliation: Department of Biochemistry, Tokyo Medical University, Tokyo, Japan.
authors.forename: Ayako
authors.initials: A
authors.lastname: Hirota
authors.affiliation: Department of Biochemistry, Tokyo Medical University, Tokyo, Japan.
authors.forename: Jun
authors.initials: J
authors.lastname: Takemura
authors.affiliation: Department of Biochemistry, Tokyo Medical University, Tokyo, Japan.
authors.forename: Akihisa
authors.initials: A
authors.lastname: Abe
authors.affiliation: Institute of Medical Science, Tokyo Medical University, Tokyo, Japan.
authors.forename: Masato
authors.initials: M
authors.lastname: Inazu
authors.affiliation: Department of Biochemistry, Tokyo Medical University, Tokyo, Japan.
authors.forename: Masaki
authors.initials: M
authors.lastname: Hiramoto
authors.affiliation: Department of Otolaryngology (Head and Neck Surgery), Tokyo Medical University, Tokyo, Japan.
authors.forename: Kiyoaki
authors.initials: K
authors.lastname: Tsukahara
authors.affiliation: Department of Biochemistry, Tokyo Medical University, Tokyo, Japan.
authors.forename: Keisuke
authors.identifier: http://orcid.org/0000-0002-8435-1304
authors.initials: K
authors.lastname: Miyazawa
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001343:Autophagy / Q000187:drug effects*; D017963:Azithromycin / Q000494:pharmacology*; D002294:Carcinoma, Squamous Cell / Q000473:pathology*; D016923:Cell Death / Q000187:drug effects*; D045744:Cell Line, Tumor; D017291:Clarithromycin / Q000494:pharmacology*; D006258:Head and Neck Neoplasms / Q000473:pathology*; D006801:Humans; D018942:Macrolides / Q000494:pharmacology*; D051379:Mice
chemical_list: D000900:Anti-Bacterial Agents; D018942:Macrolides; D017963:Azithromycin; D017291:Clarithromycin