Cancer cells preferentially utilize glycolysis for ATP production even in aerobic conditions (the Warburg effect) and adapt mitochondrial processes to their specific needs. Recent studies indicate that altered mitochondrial activities in cancer represent an actionable target for therapy. We previously showed that salt 1-3C, a quinoxaline unit (with cytotoxic activity) incorporated into a meso-substituted pentamethinium salt (with mitochondrial selectivity and fluorescence properties), displayed potent cytotoxic effects in vitro and in vivo, without significant toxic effects to normal tissues. Here, we investigated the cytotoxic mechanism of salt 1-3C compared to its analogue, salt 1-8C, with an extended side carbon chain. Live cell imaging demonstrated that salt 1-3C, but not 1-8C, is rapidly incorporated into mitochondria, correlating with increased cytotoxicity of salt 1-3C. The accumulation in mitochondria led to their fragmentation and loss of function, accompanied by increased autophagy/mitophagy. Salt 1-3C preferentially activated AMP-activated kinase and inhibited mammalian target of rapamycin (mTOR) signaling pathways, sensors of cellular metabolism, but did not induce apoptosis. These data indicate that salt 1-3C cytotoxicity involves mitochondrial perturbation and disintegration, and such compounds are promising candidates for targeting mitochondria as a weak spot of cancer.

BIOCEV 1st Faculty of Medicine Charles University Průmyslová 595 252 50 Vestec Czech Republic

Department of Analytical Chemistry University of Chemistry and Technology Prague Technická 5 166 28 Prague 6 Czech Republic

Department of Pediatrics and Adolescent Medicine 1st Faculty of Medicine Charles University Prague Kateřinská 32 121 08 Prague 2 Czech Republic

General University Hospital U nemocnice 2 128 08 Prague 2 Czech Republic

Regional Centre for Applied Molecular Oncology Masaryk Memorial Cancer Institute 656 53 Brno Czech Republic

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Saraste M. Oxidative phosphorylation at the fin de siecle. Science. 1999;283:1488–1493. doi: 10.1126/science.283.5407.1488. PubMed DOI

Pavlova N.N., Thompson C.B. The Emerging Hallmarks of Cancer Metabolism. Cell Metab. 2016;23:27–47. doi: 10.1016/j.cmet.2015.12.006. PubMed DOI PMC

Wallace D.C. Mitochondria and cancer. Nat. Rev. Cancer. 2012;12:685–698. doi: 10.1038/nrc3365. PubMed DOI PMC

Wang J.Y. Regulation of cell death by the Abl tyrosine kinase. Oncogene. 2000;19:5643–5650. doi: 10.1038/sj.onc.1203878. PubMed DOI

Mazel S., Burtrum D., Petrie H.T. Regulation of cell division cycle progression by bcl-2 expression: A potential mechanism for inhibition of programmed cell death. J. Exp. Med. 1996;183:2219–2226. doi: 10.1084/jem.183.5.2219. PubMed DOI PMC

Li G., Lin Q., Sun L., Feng C., Zhang P., Yu B., Chen Y., Wen Y., Wang H., Ji L., et al. A mitochondrial targeted two-photon iridium(III) phosphorescent probe for selective detection of hypochlorite in live cells and in vivo. Biomaterials. 2015;53:285–295. doi: 10.1016/j.biomaterials.2015.02.106. PubMed DOI

Weinberg S.E., Chandel N.S. Targeting mitochondria metabolism for cancer therapy. Nat. Chem. Biol. 2015;11:9–15. doi: 10.1038/nchembio.1712. PubMed DOI PMC

Zielonka J., Joseph J., Sikora A., Hardy M., Ouari O., Vasquez-Vivar J., Cheng G., Lopez M., Kalyanaraman B. Mitochondria-Targeted Triphenylphosphonium-Based Compounds: Syntheses, Mechanisms of Action, and Therapeutic and Diagnostic Applications. Chem. Rev. 2017;117:10043–10120. doi: 10.1021/acs.chemrev.7b00042. PubMed DOI PMC

Johnson L.V., Walsh M.L., Chen L.B. Localization of mitochondria in living cells with rhodamine 123. Proc. Natl. Acad. Sci. USA. 1980;77:990–994. doi: 10.1073/pnas.77.2.990. PubMed DOI PMC

Heller A., Brockhoff G., Goepferich A. Targeting drugs to mitochondria. Eur. J. Pharm. Biopharm. 2012;82:1–18. doi: 10.1016/j.ejpb.2012.05.014. PubMed DOI

Onoe S., Temma T., Shimizu Y., Ono M., Saji H. Investigation of cyanine dyes for in vivo optical imaging of altered mitochondrial membrane potential in tumors. Cancer Med. 2014;3:775–786. doi: 10.1002/cam4.252. PubMed DOI PMC

Briza T., Kejik Z., Cisarova I., Kralova J., Martasek P., Kral V. Optical sensing of sulfate by polymethinium salt receptors: Colorimetric sensor for heparin. Chem. Commun. 2008:1901–1903. doi: 10.1039/b718492a. PubMed DOI

Rimpelova S., Briza T., Kralova J., Zaruba K., Kejik Z., Cisarova I., Martasek P., Ruml T., Kral V. Rational design of chemical ligands for selective mitochondrial targeting. Bioconjugate Chem. 2013;24:1445–1454. doi: 10.1021/bc400291f. PubMed DOI

Briza T., Kralova J., Dolensky B., Rimpelova S., Kejik Z., Ruml T., Hajduch M., Dzubak P., Mikula I., Martasek P., et al. Striking antitumor activity of a methinium system with incorporated quinoxaline unit obtained by spontaneous cyclization. Chembiochem. 2015;16:555–558. doi: 10.1002/cbic.201402662. PubMed DOI

Yang N.J., Hinner M.J. Getting across the cell membrane: An overview for small molecules, peptides, and proteins. Methods Mol Biol. 2015;1266:29–53. doi: 10.1007/978-1-4939-2272-7_3. PubMed DOI PMC

Okatsu K., Saisho K., Shimanuki M., Nakada K., Shitara H., Sou Y.S., Kimura M., Sato S., Hattori N., Komatsu M., et al. p62/SQSTM1 cooperates with Parkin for perinuclear clustering of depolarized mitochondria. Genes Cells. 2010;15:887–900. doi: 10.1111/j.1365-2443.2010.01426.x. PubMed DOI PMC

Tung Y.T., Hsu W.M., Lee H., Huang W.P., Liao Y.F. The evolutionarily conserved interaction between LC3 and p62 selectively mediates autophagy-dependent degradation of mutant huntingtin. Cell. Mol. Neurobiol. 2010;30:795–806. doi: 10.1007/s10571-010-9507-y. PubMed DOI

Wang Y., Xu W., Yan Z., Zhao W., Mi J., Li J., Yan H. Metformin induces autophagy and G0/G1 phase cell cycle arrest in myeloma by targeting the AMPK/mTORC1 and mTORC2 pathways. J Exp Clin Cancer Res. 2018;37:63. doi: 10.1186/s13046-018-0731-5. PubMed DOI PMC

Yoo S.M., Jung Y.K. A Molecular Approach to Mitophagy and Mitochondrial Dynamics. Mol. Cells. 2018;41:18–26. doi: 10.14348/molcells.2018.2277. PubMed DOI PMC

Perfettini J.L., Roumier T., Kroemer G. Mitochondrial fusion and fission in the control of apoptosis. Trends Cell Biol. 2005;15:179–183. doi: 10.1016/j.tcb.2005.02.005. PubMed DOI

Arnoult D. Mitochondrial fragmentation in apoptosis. Trends Cell Biol. 2007;17:6–12. doi: 10.1016/j.tcb.2006.11.001. PubMed DOI

Suen D.F., Norris K.L., Youle R.J. Mitochondrial dynamics and apoptosis. Genes Dev. 2008;22:1577–1590. doi: 10.1101/gad.1658508. PubMed DOI PMC

Scorrano L., Ashiya M., Buttle K., Weiler S., Oakes S.A., Mannella C.A., Korsmeyer S.J. A distinct pathway remodels mitochondrial cristae and mobilizes cytochrome c during apoptosis. Dev. Cell. 2002;2:55–67. doi: 10.1016/S1534-5807(01)00116-2. PubMed DOI

Kisfalvi K., Eibl G., Sinnett-Smith J., Rozengurt E. Metformin disrupts crosstalk between G protein-coupled receptor and insulin receptor signaling systems and inhibits pancreatic cancer growth. Cancer Res. 2009;69:6539–6545. doi: 10.1158/0008-5472.CAN-09-0418. PubMed DOI PMC

Bartolome A., Garcia-Aguilar A., Asahara S.I., Kido Y., Guillen C., Pajvani U.B., Benito M. MTORC1 Regulates both General Autophagy and Mitophagy Induction after Oxidative Phosphorylation Uncoupling. Mol. Cell. Biol. 2017:e00441-17. doi: 10.1128/MCB.00441-17. PubMed DOI PMC

Ma X.M., Blenis J. Molecular mechanisms of mTOR-mediated translational control. Nat. Rev. Mol. Cell Biol. 2009;10:307–318. doi: 10.1038/nrm2672. PubMed DOI

Dranka B.P., Benavides G.A., Diers A.R., Giordano S., Zelickson B.R., Reily C., Zou L., Chatham J.C., Hill B.G., Zhang J., et al. Assessing bioenergetic function in response to oxidative stress by metabolic profiling. Free Radic. Biol. Med. 2011;51:1621–1635. doi: 10.1016/j.freeradbiomed.2011.08.005. PubMed DOI PMC

Ross M.F., Kelso G.F., Blaikie F.H., James A.M., Cocheme H.M., Filipovska A., Da Ros T., Hurd T.R., Smith R.A., Murphy M.P. Lipophilic triphenylphosphonium cations as tools in mitochondrial bioenergetics and free radical biology. Biochem. Biokhimiia. 2005;70:222–230. doi: 10.1007/s10541-005-0104-5. PubMed DOI

Maiuri M.C., Zalckvar E., Kimchi A., Kroemer G. Self-eating and self-killing: Crosstalk between autophagy and apoptosis. Nat. Rev. Mol. Cell Biol. 2007;8:741–752. doi: 10.1038/nrm2239. PubMed DOI

Chen Q., Kang J., Fu C. The independence of and associations among apoptosis, autophagy, and necrosis. Signal Transduct. Target. Ther. 2018;3:18. doi: 10.1038/s41392-018-0018-5. PubMed DOI PMC

Kimura S., Noda T., Yoshimori T. Dynein-dependent movement of autophagosomes mediates efficient encounters with lysosomes. Cell Struct. Funct. 2008;33:109–122. doi: 10.1247/csf.08005. PubMed DOI

Vander Heiden M.G., Cantley L.C., Thompson C.B. Understanding the Warburg effect: The metabolic requirements of cell proliferation. Science. 2009;324:1029–1033. doi: 10.1126/science.1160809. PubMed DOI PMC

Muller P., Ceskova P., Vojtesek B. Hsp90 is essential for restoring cellular functions of temperature-sensitive p53 mutant protein but not for stabilization and activation of wild-type p53: Implications for cancer therapy. J. Biol. Chem. 2005;280:6682–6691. doi: 10.1074/jbc.M412767200. PubMed DOI

New-Generation Heterocyclic Bis-Pentamethinium Salts as Potential Cytostatic Drugs with Dual IL-6R and Mitochondria-Targeting Activity

Influence of fluorophore and linker length on the localization and trafficking of fluorescent sterol probes