title: The marine triterpene glycoside frondoside A induces p53-independent apoptosis and inhibits autophagy in urothelial carcinoma cells.
abstract: Advanced urothelial carcinomas represent a considerable clinical challenge as they are difficult to treat. Platinum-based combination regimens obtain response rates ranging from 40 to 70% in first-line therapy of advanced urothelial carcinoma. In the majority of cases, however, the duration of these responses is limited, and when progression occurs, the outcome is generally poor. Therefore, novel therapeutic strategies are urgently needed. The purpose of the current research is to investigate the anticancer effects and the mode of action of the marine triterpene glycoside frondoside A in p53-wild type and p53-deficient human urothelial carcinoma cells.
  
  
  
  Activity of frondoside A was examined in the human urothelial carcinoma cell lines RT112, RT4, HT-1197, TCC-SUP, T-24, and 486p. Effects of frondoside A on cell viability, either alone or in combination with standard cytotoxic agents were investigated, and synergistic effects were analyzed. Pro-apoptotic activity was assessed by Western blotting and FACS, alone and in combination with a caspases-inhibitor. The impact of functional p53 was investigated by siRNA gene silencing and the p53 inhibitor pifithrin-α. Effects on autophagy were studied using LC3B-I/II and SQSTM/p62 as markers. The unpaired Student's t-test was used for comparison of the data sets.
  
  
  
  Frondoside A shows high cytotoxicity in urothelial carcinoma cells with IC50sranging from 0.55 to 2.33 μM while higher concentrations of cisplatin are required for comparable effects (IC50= 2.03 ~ 5.88 μM). Induction of apoptosis by frondoside A was associated with the regulation of several pro-apoptotic factors, like caspase-3, -8, and -9, PARP, Bax, p21, DNA fragmentation, and externalization of phosphatidylserine. Remarkably, inhibition of p53 by gene silencing or pifithrin-α pretreatment, as well as caspase inhibition, did not suppress apoptotic activity of frondoside A, while cisplatin activity, in contrast, was significantly decreased. Frondoside A inhibited pro-survival autophagy, a known mechanism of drug resistance in urothelial carcinoma and showed synergistic activity with cisplatin and gemcitabine.
  
  
  
  A unique combination of properties makes marine compound frondoside A a promising candidate for the treatment of human urothelial carcinomas.
authors.affiliation: Laboratory of Experimental Oncology, Department of Oncology, Hematology and Bone Marrow Transplantation with Section Pneumology, University Medical Center Hamburg-Eppendorf, Martinistr. 52, Hamburg, 20246, Germany. dyshlovoy@gmail.com.|Laboratory of Marine Natural Products Chemistry, G.B. Elyakov Pacific Institute of Bioorganic Chemistry, Far-East Branch, Russian Academy of Sciences, Prospekt 100-let Vladivostoku 159, Vladivostok, 690022, Russian Federation. dyshlovoy@gmail.com.|School of Natural Sciences, Far Eastern Federal University, Sukhanova Street 8, Vladivostok, 690091, Russian Federation. dyshlovoy@gmail.com.
authors.forename: Sergey A
authors.initials: SA
authors.lastname: Dyshlovoy
authors.affiliation: Laboratory of Experimental Oncology, Department of Oncology, Hematology and Bone Marrow Transplantation with Section Pneumology, University Medical Center Hamburg-Eppendorf, Martinistr. 52, Hamburg, 20246, Germany.
authors.forename: Ramin
authors.initials: R
authors.lastname: Madanchi
authors.affiliation: Laboratory of Experimental Oncology, Department of Oncology, Hematology and Bone Marrow Transplantation with Section Pneumology, University Medical Center Hamburg-Eppendorf, Martinistr. 52, Hamburg, 20246, Germany.
authors.forename: Jessica
authors.initials: J
authors.lastname: Hauschild
authors.affiliation: Laboratory of Experimental Oncology, Department of Oncology, Hematology and Bone Marrow Transplantation with Section Pneumology, University Medical Center Hamburg-Eppendorf, Martinistr. 52, Hamburg, 20246, Germany.
authors.forename: Katharina
authors.initials: K
authors.lastname: Otte
authors.affiliation: Laboratory of Experimental Oncology, Department of Oncology, Hematology and Bone Marrow Transplantation with Section Pneumology, University Medical Center Hamburg-Eppendorf, Martinistr. 52, Hamburg, 20246, Germany.
authors.forename: Winfried H
authors.initials: WH
authors.lastname: Alsdorf
authors.affiliation: Institute of Anatomy and Experimental Morphology, University Cancer Center Hamburg, University Medical Center Hamburg-Eppendorf, Martinistr. 52, Hamburg, 20246, Germany.
authors.forename: Udo
authors.initials: U
authors.lastname: Schumacher
authors.affiliation: Laboratory of Marine Natural Products Chemistry, G.B. Elyakov Pacific Institute of Bioorganic Chemistry, Far-East Branch, Russian Academy of Sciences, Prospekt 100-let Vladivostoku 159, Vladivostok, 690022, Russian Federation.
authors.forename: Vladimir I
authors.initials: VI
authors.lastname: Kalinin
authors.affiliation: Laboratory of Marine Natural Products Chemistry, G.B. Elyakov Pacific Institute of Bioorganic Chemistry, Far-East Branch, Russian Academy of Sciences, Prospekt 100-let Vladivostoku 159, Vladivostok, 690022, Russian Federation.
authors.forename: Alexandra S
authors.initials: AS
authors.lastname: Silchenko
authors.affiliation: Laboratory of Experimental Oncology, Department of Oncology, Hematology and Bone Marrow Transplantation with Section Pneumology, University Medical Center Hamburg-Eppendorf, Martinistr. 52, Hamburg, 20246, Germany.|Laboratory of Marine Natural Products Chemistry, G.B. Elyakov Pacific Institute of Bioorganic Chemistry, Far-East Branch, Russian Academy of Sciences, Prospekt 100-let Vladivostoku 159, Vladivostok, 690022, Russian Federation.|School of Natural Sciences, Far Eastern Federal University, Sukhanova Street 8, Vladivostok, 690091, Russian Federation.
authors.forename: Sergey A
authors.initials: SA
authors.lastname: Avilov
authors.affiliation: Laboratory of Experimental Oncology, Department of Oncology, Hematology and Bone Marrow Transplantation with Section Pneumology, University Medical Center Hamburg-Eppendorf, Martinistr. 52, Hamburg, 20246, Germany.|Tumor and Breast Center ZeTuP St. Gallen, Rorschacher Strasse 150, St. Gallen, 9006, Switzerland.
authors.forename: Friedemann
authors.initials: F
authors.lastname: Honecker
authors.affiliation: Laboratory of Marine Natural Products Chemistry, G.B. Elyakov Pacific Institute of Bioorganic Chemistry, Far-East Branch, Russian Academy of Sciences, Prospekt 100-let Vladivostoku 159, Vladivostok, 690022, Russian Federation.
authors.forename: Valentin A
authors.initials: VA
authors.lastname: Stonik
authors.affiliation: Laboratory of Experimental Oncology, Department of Oncology, Hematology and Bone Marrow Transplantation with Section Pneumology, University Medical Center Hamburg-Eppendorf, Martinistr. 52, Hamburg, 20246, Germany.
authors.forename: Carsten
authors.initials: C
authors.lastname: Bokemeyer
authors.affiliation: Laboratory of Experimental Oncology, Department of Oncology, Hematology and Bone Marrow Transplantation with Section Pneumology, University Medical Center Hamburg-Eppendorf, Martinistr. 52, Hamburg, 20246, Germany.
authors.forename: Gunhild
authors.initials: G
authors.lastname: von Amsberg
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D001343:Autophagy / Q000187:drug effects*; D002295:Carcinoma, Transitional Cell / Q000188:drug therapy; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D002945:Cisplatin / Q000494:pharmacology; D003841:Deoxycytidine / Q000031:analogs & derivatives / Q000494:pharmacology; D004354:Drug Screening Assays, Antitumor; D004357:Drug Synergism; D006027:Glycosides / Q000494:pharmacology*; D006801:Humans; D012616:Sea Cucumbers / Q000737:chemistry; D014315:Triterpenes / Q000494:pharmacology*; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D001749:Urinary Bladder Neoplasms / Q000188:drug therapy; D000093542:Gemcitabine
chemical_list: D000970:Antineoplastic Agents; D006027:Glycosides; C495901:TP53 protein, human; D014315:Triterpenes; D016159:Tumor Suppressor Protein p53; C534797:frondoside A; D003841:Deoxycytidine; D002945:Cisplatin; D000093542:Gemcitabine
keywords: Apoptosis; Autophagy; Frondoside A; Marine natural compounds; Urothelial carcinoma; p53