Hypoxia, a hallmark of many solid tumors, is linked to increased cancer aggressiveness, metastasis, and resistance to conventional therapies, leading to poor patient outcomes. This challenges the efficiency of photodynamic therapy (PDT), which relies on the generation of cytotoxic reactive oxygen species (ROS) through the irradiation of a photosensitizer (PS), a process partially dependent on oxygen levels. In this work, we introduce a novel family of potent PSs based on ruthenium(II) polypyridyl complexes with 2,2'-bipyridyl ligands derived from COUPY coumarins, termed COUBPYs. Ru-COUBPY complexes exhibit outstanding in vitro cytotoxicity against CT-26 cancer cells when irradiated with light within the phototherapeutic window, achieving nanomolar potency in both normoxic and hypoxic conditions while remaining nontoxic in the dark, leading to impressive phototoxic indices (>30,000). Their ability to generate both Type I and Type II ROS underpins their exceptional PDT efficiency. The lead compound of this study, SCV49, shows a favorable in vivo pharmacokinetic profile, excellent toxicological tolerability, and potent tumor growth inhibition in mice bearing subcutaneous CT-26 tumors at doses as low as 3 mg/kg upon irradiation with deep-red light (660 nm). These results allow us to propose SCV49 as a strong candidate for further preclinical development, particularly for treating large hypoxic solid tumors.

Metal–coumarin photosensitizers enhance ROS generation and phototoxicity, offering promising solutions for cancer phototherapy.

Despite the potential of photodynamic therapy (PDT) in cancer treatment, the development of efficient and photostable photosensitizing molecules that operate at long wavelengths of light has become a major hurdle. Here, we report for the first time an Ir(III)-phthalocyanine conjugate (Ir-ZnPc) as a novel photosensitizer for high-efficiency synergistic PDT treatment that takes advantage of the long-wavelength excitation and near infrared (NIR) emission of the phthalocyanine scaffold and the known photostability and high phototoxicity of cyclometalated Ir(III) complexes. In order to increase water solubility and cell membrane permeability, the conjugate and parent zinc phthalocyanine (ZnPc) were encapsulated in amphoteric redox-responsive polyurethane-polyurea hybrid nanocapsules (Ir-ZnPc-NCs and ZnPc-NCs, respectively). Photobiological evaluations revealed that the encapsulated Ir-ZnPc conjugate achieved high photocytotoxicity in both normoxic and hypoxic conditions under 630 nm light irradiation, which can be attributed to dual Type I and Type II reactive oxygen species (ROS) photogeneration. Interestingly, PDT treatments with Ir-ZnPc-NCs and ZnPc-NCs significantly inhibited the growth of three-dimensional (3D) multicellular tumor spheroids. Overall, the nanoencapsulation of Zn phthalocyanines conjugated to cyclometalated Ir(III) complexes provides a new strategy for obtaining photostable and biocompatible red-light-activated nano-PDT agents with efficient performance under challenging hypoxic environments, thus offering new therapeutic opportunities for cancer treatment.

Light‐activated treatments, such as photodynamic therapy (PDT), provide temporal and spatial control over a specific cytotoxic response by exploiting toxicity differences between irradiated and dark conditions. In this work, a novel strategy for developing near infrared (NIR)‐activatable Ru(II) polypyridyl‐based photosensitizers (PSs) was successfully developed through the incorporation of symmetric heptamethine cyanine dyes in the metal complex via a phenanthrimidazole ligand. Owing to their strong absorption in the NIR region, the PSs could be efficiently photoactivated with highly penetrating NIR light (770 nm), leading to high photocytotoxicities towards several cancer cell lines under both normoxic and hypoxic conditions. Notably, our lead PS (Ru‐Cyn‐1), which accumulated in the mitochondria, exhibited a good photocytotoxic activity under challenging low‐oxygen concentration (2 % O₂) upon NIR light irradiation conditions (770 nm), owing to a combination of type I and II PDT mechanisms. The fact that the PS Protoporphyrin IX (PpIX), the metabolite of the clinically approved 5‐ALA PS, was found inactive under the same challenging conditions positions Ru‐Cyn‐1 complex as a promising PDT agent for the treatment of deep‐seated hypoxic tumours.

Despite the potential of photodynamic therapy (PDT), this oxygen-dependent oncological treatment is greatly restricted in the clinic by the well-known hypoxic feature of solid tumors. Here we provide new insights on the development of PDT agents based on conjugates between COUPY fluorophores and cyclometalated iridium(III) complexes with the aim of overcoming this limitation. The structural modifications carried out within the metal core of Ir(III)-COUPY conjugates, based on the incorporation of trifluorobenzyl groups at the cyclometalating ligands, allowed efficient explotation of Type I PDT mechanisms while retaining operativity at long-wavelength visible light, which facilitates deeper tissue penetration compared with short wavelengths. Photobiological evaluation revealed that Ir(III)‑COUPY conjugate 3c achieved potent photocytotoxicity towards cisplatin-resistant ovarian (A2780cis) and mammary (EO771) cancer cell lines, efficiently photogenerated Type I and Type II ROS and photoinduced apoptotic cell death using red light irradiation (620 nm). Importantly, this Ir(III)-COUPY conjugate retained such potent photoactivity under low-oxygen environment conditions (2% O2), delivering equipotent photocytotoxicity towards normoxic and hypoxic adherent cancer cells. Compound 3c was found highly phototoxic against EO771 multicellular tumor spheroids and showed no signs of toxicity or adverse effects in mice, which could facilitate in vivo phototherapeutic applications. Taken together, this study demonstrated that the conjugation between COUPY dyes and rationally-designed Ir(III) complexes represents a frontier strategy for the development of new red light‑activated photosensitizers capable of operating under hypoxia, holding promise to achieve satisfactory anticancer PDT effects.

Photodynamic therapy (PDT) is a promising cancer treatment modality, offering precise spatial and temporal control of drug activation using light. However, clinical translation of current photosensitizers (PSs) is limited by inefficient activation at wavelengths within the phototherapeutic window, especially in the deep-red and near-infrared (NIR) region. NIR light provides advantages such as reduced absorption by endogenous chromophores, minimized tissue photodamage, and improved tissue penetration, highlighting the need for PSs to be activatable in this range. Herein, we report a novel series of ruthenium(II) polypyridyl complexes (Ru4-7) featuring π-extended COUBPY ligands, designed via a vinylogation strategy and synthesized through an innovative postcoordination ligand assembly approach. This structural modification enhances molar absorptivity and red-shifts the absorption bands well into the NIR region without substantially compromising photostability. Complexes Ru4-7 efficiently generate both Type I and Type II reactive oxygen species, and their photodynamic activity, combined with preferential mitochondrial accumulation, leads to potent nanomolar phototoxicity against CT-26 colorectal cancer cells under deep-red and NIR irradiation, even under hypoxia. Notably, the lead complex Ru6 demonstrated strong in vivo phototoxicity in mice bearing subcutaneous CT-26 tumors, achieving significant tumor growth inhibition upon irradiation with 660 and 780 nm light. Ru6 thus represents one of the first Ru(II) polypyridyl complexes to exhibit robust in vivo PDT antitumor activity under one-photon NIR activation. Its broad wavelength activation profile further underscores its potential versatility for treating tumors of varying size and anatomical location depending on specific light penetration requirements. These findings mark a promising step toward next-generation PSs for treating deep-seated and hypoxic tumors.