Phthalocyanines and Tetrapyrazinoporphyrazines with Two Cationic Donuts: High Photodynamic Activity as a Result of Rigid Spatial Arrangement of Peripheral Substituents Basma Ghazal, † Miloslav Machacek, ‡ Mona Abbas Shalaby, † Veronika Novakova, § Petr Zimcik,* ,∥ and Saad Makhseed* ,† † Department of Chemistry, Kuwait University, P.O. Box 5969, Safat, 13060, Kuwait ‡ Department of Biochemical Sciences, Faculty of Pharmacy in Hradec Kralove, Charles University, Heyrovskeho 1203, Hradec Kralove, 500 05, Czech Republic § Department of Biophysics and Physical Chemistry, Faculty of Pharmacy in Hradec Kralove, Charles University, Heyrovskeho 1203, Hradec Kralove, 500 05, Czech Republic ∥ Department of Pharmaceutical Chemistry and Pharmaceutical Analysis, Faculty of Pharmacy in Hradec Kralove, Charles University, Heyrovskeho 1203, Hradec Kralove, 500 05, Czech Republic * S Supporting Information ABSTRACT: High photodynamic activity was observed for hexadeca-cationic zinc, magnesium, and metal-free phthalo- cyanines (Pcs) and tetrapyrazinoporphyrazines with EC 50 values as low as 5 nM (MCF-7 cells) for the best compound; this activity was several times better than that of clinically established photosensitizers vertepor fi n, temopor fi n, S 3 AlOHPc, or protoporphyrin IX. This lead compound was characterized by low dark toxicity (TC 50 = 369 μM), high efficiency against other cell lines (HCT 116 and HeLa), and possible activation by light above 680 nm. The excellent photodynamic activity resulted from the rigid spatial arrange- ment of the quaternized triazole moieties above and below the Pc core, as confirmed by X-ray crystallography. The triazole moieties thus formed two “cationic donuts” that protected the hydrophobic core against aggregation in water. The lysosomes were found to be the site of subcellular localization and were consequently the primary targets of photodynamic injury, resulting in predominantly necrotic cell death. ■ INTRODUCTION Photodynamic therapy (PDT) is a relatively modern cancer treatment used as an alternative to chemotherapy, radiotherapy, and surgical intervention. PDT combines three potentially nontoxic components, light, oxygen, and a photosensitizer (PS), that are able to destroy target cells by the production of toxic species, the most important being singlet oxygen. 1 PSs from various structural groups have been approved for use in clinical practice or are under investigation in clinical trials. 2 The key photophysical characteristics of an efficient PS are good production of singlet oxygen and strong absorption of light in the optical window of tissues (or phototherapeutic window, i.e., 650−850 nm), where the penetration of light is the deepest. 3 From this point of view, phthalocyanines (Pcs), which typically possess very good singlet oxygen production and extinction coefficients over 2 × 10 5 M −1 cm −1 are interesting molecular structures. 4,5 Indeed, sulfonated hydroxyaluminum Pc with an average degree of sulfonation n = 3 (S 3 AlOHPc) has been approved in Russia since 2001. 5,6 On the other hand, the Pc core is planar and strongly hydrophobic; as a result, it tends to form strong aggregates in water and water-based media, which can substantially reduce or completely eliminate the photo- physical pathways important for PDT. Several strategies on how to resolve this obstacle have been developed by various research groups. The strategies include the use of surfactants/ cosolvents 7,8 or delivery systems 9−11 to dissolve the lipophilic compounds or substitution by highly hydrophilic substituents axially on the central metal 12,13 or on the Pc core. 14−17 Recently, we identified Pcs with rigid cationic substituents bearing quaternized imidazolyl moieties, which led to highly water-soluble Pcs that did not display any aggregation in water. 18 Applying the same substituents to tetrapyridoporphyr- azines resulted in a compound that belongs among the most potent PSs reported to date. 19 Considering the promising anticancer photodynamic activity of these PSs bearing imidazolyl moieties, we applied the same protocol to prepare triazole-containing Pc complexes and anticipated obtaining new Received: February 21, 2017 Published: May 30, 2017 Article pubs.acs.org/jmc © 2017 American Chemical Society 6060 DOI: 10.1021/acs.jmedchem.7b00272 J. Med. Chem. 2017, 60, 6060−6076