Antimicrobial photodynamic therapy (aPDT) is an evolving treatment strategy against human pathogenic microbes such as the Candida species, including the emerging pathogen C. auris. Using a modified... Show moreAntimicrobial photodynamic therapy (aPDT) is an evolving treatment strategy against human pathogenic microbes such as the Candida species, including the emerging pathogen C. auris. Using a modified EUCAST protocol, the light-enhanced antifungal activity of the natural compound parietin was explored. The photoactivity was evaluated against three separate strains of five yeasts, and its molecular mode of action was analysed via several techniques, i.e., cellular uptake, reactive electrophilic species (RES), and singlet oxygen yield. Under experimental conditions (λ = 428 nm, H = 30 J/cm2, PI = 30 min), microbial growth was inhibited by more than 90% at parietin concentrations as low as c = 0.156 mg/L (0.55 µM) for C. tropicalis and Cryptococcus neoformans, c = 0.313 mg/L (1.10 µM) for C. auris, c = 0.625 mg/L (2.20 µM) for C. glabrata, and c = 1.250 mg/L (4.40 µM) for C. albicans. Mode-of-action analysis demonstrated fungicidal activity. Parietin targets the cell membrane and induces cell death via ROS-mediated lipid peroxidation after light irradiation. In summary, parietin exhibits light-enhanced fungicidal activity against all Candida species tested (including C. auris) and Cryptococcus neoformans, covering three of the four critical threats on the WHO's most recent fungal priority list. Show less
To investigate the potential of tumor-targeting photoactivated chemotherapy, a chiral ruthenium-based anticancer warhead, Λ/Δ-[Ru(Ph2phen)2(OH2)2]2+, was conjugated to the RGD-containing Ac-MRGDH... Show moreTo investigate the potential of tumor-targeting photoactivated chemotherapy, a chiral ruthenium-based anticancer warhead, Λ/Δ-[Ru(Ph2phen)2(OH2)2]2+, was conjugated to the RGD-containing Ac-MRGDH-NH2 peptide by direct coordination of the M and H residues to the metal. This design afforded two diastereoisomers of a cyclic metallopeptide, Λ-[1]Cl2 and Δ-[1]Cl2. In the dark, the ruthenium-chelating peptide had a triple action. First, it prevented other biomolecules from coordinating with the metal center. Second, its hydrophilicity made [1]Cl2 amphiphilic so that it self-assembled in culture medium into nanoparticles. Third, it acted as a tumor-targeting motif by strongly binding to the integrin (Kd = 0.061 μM for the binding of Λ-[1]Cl2 to αIIbβ3), which resulted in the receptor-mediated uptake of the conjugate in vitro. Phototoxicity studies in two-dimensional (2D) monolayers of A549, U87MG, and PC-3 human cancer cell lines and U87MG three-dimensional (3D) tumor spheroids showed that the two isomers of [1]Cl2 were strongly phototoxic, with photoindexes up to 17. Mechanistic studies indicated that such phototoxicity was due to a combination of photodynamic therapy (PDT) and photoactivated chemotherapy (PACT) effects, resulting from both reactive oxygen species generation and peptide photosubstitution. Finally, in vivo studies in a subcutaneous U87MG glioblastoma mice model showed that [1]Cl2 efficiently accumulated in the tumor 12 h after injection, where green light irradiation generated a stronger tumoricidal effect than a nontargeted analogue ruthenium complex [2]Cl2. Considering the absence of systemic toxicity for the treated mice, these results demonstrate the high potential of light-sensitive integrin-targeted ruthenium-based anticancer compounds for the treatment of brain cancer in vivo. Show less
Geest, E.P. van; Götzfried, S. K.; Klein, D. M.; Salitra, N.; Popal, S.; Husiev, Y.; ... ; Bonnet, S. 2022
Photosubstitutionally active ruthenium complexes show high potential as prodrugs for the photoactivated chemotherapy (PACT) treatment of tumors. One of the problems in PACT is that the localization... Show morePhotosubstitutionally active ruthenium complexes show high potential as prodrugs for the photoactivated chemotherapy (PACT) treatment of tumors. One of the problems in PACT is that the localization of the ruthenium compound is hard to trace. Here, a ruthenium PACT prodrug, [Ru(3)(biq)(STF-31)](PF6 )2 (where 3 = 3-(([2,2':6',2″-ter- pyridin]-4'-yloxy)propyl-4-(pyren-1-yl)butanoate) and biq = 2,2'-biquinoline), has been prepared, in which a pyrene tracker is attached via an ester bond. The proximity between the fluorophore and the ruthenium center leads to fluorescence quenching. Upon intracellular hydrolysis of the ester linkage, however, the fluorescence of the pyrene moiety is recovered, thus demonstrating prodrug cellular uptake. Further light irradiation of this molecule liberates by photosubstitution STF-31, a known cytotoxic nicotinamide phosphoribosyltransferase (NAMPT) inhibitor, as well as singlet oxygen via excitation of the free pyrene chromophore. The dark and light cytotoxicity of the prodrug, embedded in liposomes, as well as the appearance of blue emission upon uptake, were evaluated in A375 human skin melanoma cells. The cytotoxicity of the liposome-embedded prodrug was indeed increased by light irradiation. This work realizes an in vitro proof-of-concept of the lock-and-kill principle, which may ultimately be used to design strategies aimed at knowing where and when light irradiation should be realized in vivo. Show less
In vivo data are rare but essential for establishing the clinical potential of ruthenium-based photoactivated chemotherapy (PACT) compounds, a new family of phototherapeutic drugs that are... Show moreIn vivo data are rare but essential for establishing the clinical potential of ruthenium-based photoactivated chemotherapy (PACT) compounds, a new family of phototherapeutic drugs that are activated via ligand photosubstitution. Here a novel trisheteroleptic ruthenium complex [Ru(dpp)(bpy)(mtmp)](PF6)2 ([2](PF6)2, dpp = 4,7-diphenyl-1,10-phenanthroline, bpy = 2,2'-bipyridine, mtmp = 2-methylthiomethylpyridine) was synthesized and its light-activated anticancer properties were validated in cancer cell monolayers, 3D tumor spheroids, and in embryonic zebrafish cancer models. Upon green light irradiation, the non-toxic mtmp ligand is selectively cleaved off, thereby releasing a phototoxic ruthenium-based photoproduct capable notably of binding to nuclear DNA and triggering DNA damage and apoptosis within 24-48 h. In vitro, fifteen minutes of green light irradiation (21 mW cm-2, 19 J cm-2, 520 nm) were sufficient to generate high phototherapeutic indexes (PI) for this compound in a range of cancer cell lines including lung (A549), prostate (PC3Pro4), conjunctival melanoma (CRMM1, CRMM2, CM2005.1) and uveal melanoma (OMM1, OMM2.5, Mel270) cancer cell lines. The therapeutic potential of [2](PF6)2 was further evaluated in zebrafish embryo ectopic (PC3Pro4) or orthotopic (CRMM1, CRMM2) tumour models. The ectopic model consisted of red fluorescent PC3Pro4-mCherry cells injected intravenously (IV) into zebrafish, that formed perivascular metastatic lesions at the posterior ventral end of caudal hematopoietic tissue (CHT). By contrast, in the orthotopic model, CRMM1- and CRMM2-mCherry cells were injected behind the eye where they developed primary lesions. The maximally-tolerated dose (MTD) of [2](PF6)2 was first determined for three different modes of compound administration: (i) incubating the fish in prodrug-containing water (WA); (ii) injecting the prodrug intravenously (IV) into the fish; or (iii) injecting the prodrug retro-orbitally (RO) into the fish. To test the anticancer efficiency of [2](PF6)2, the embryos were treated 24 h after engraftment at the MTD. Optimally, four consecutive PACT treatments were performed on engrafted embryos using 60 min drug-to-light intervals and 90 min green light irradiation (21 mW cm-2, 114 J cm-2, 520 nm). Most importantly, this PACT protocol was not toxic to the zebrafish. In the ectopic prostate tumour models, where [2](PF6)2 showed the highest photoindex in vitro (PI > 31), the PACT treatment did not significantly diminish the growth of primary lesions, while in both conjunctival melanoma orthotopic tumour models, where [2](PF6)2 showed more modest photoindexes (PI ∼ 9), retro-orbitally administered PACT treatment significantly inhibited growth of the engrafted tumors. Overall, this study represents the first demonstration in zebrafish cancer models of the clinical potential of ruthenium-based PACT, here against conjunctival melanoma. Show less
In vivo data are rare but essential for establishing the clinical potential of ruthenium-based photoactivated chemotherapy (PACT) compounds, a new family of phototherapeutic drugs that are... Show moreIn vivo data are rare but essential for establishing the clinical potential of ruthenium-based photoactivated chemotherapy (PACT) compounds, a new family of phototherapeutic drugs that are activated via ligand photosubstitution. Here a novel trisheteroleptic ruthenium complex [Ru(dpp)(bpy)(mtmp)](PF6)(2) ([2](PF6)(2), dpp = 4,7-diphenyl-1,10-phenanthroline, bpy = 2,2'-bipyridine, mtmp = 2-methylthiomethylpyridine) was synthesized and its light-activated anticancer properties were validated in cancer cell monolayers, 3D tumor spheroids, and in embryonic zebrafish cancer models. Upon green light irradiation, the non-toxic mtmp ligand is selectively cleaved off, thereby releasing a phototoxic ruthenium-based photoproduct capable notably of binding to nuclear DNA and triggering DNA damage and apoptosis within 24-48 h. In vitro, fifteen minutes of green light irradiation (21 mW cm(-2), 19 J cm(-2), 520 nm) were sufficient to generate high phototherapeutic indexes (PI) for this compound in a range of cancer cell lines including lung (A549), prostate (PC3Pro4), conjunctival melanoma (CRMM1, CRMM2, CM2005.1) and uveal melanoma (OMM1, OMM2.5, Me1270) cancer cell lines. The therapeutic potential of [2](PF6)(2) was further evaluated in zebrafish embryo ectopic (PC3Pro4) or orthotopic (CRMM1, CRMM2) tumour models. The ectopic model consisted of red fluorescent PC3Pro4-mCherry cells injected intravenously (IV) into zebrafish, that formed perivascular metastatic lesions at the posterior ventral end of caudal hematopoietic tissue (CHT). By contrast, in the orthotopic model, CRMM1- and CRMM2-mCherry cells were injected behind the eye where they developed primary lesions. The maximally-tolerated dose (MTD) of [2](PF6)(2) was first determined for three different modes of compound administration: (i) incubating the fish in prodrug-containing water (WA); (ii) injecting the prodrug intravenously (I into the fish; or (iii) injecting the prodrug retro-orbitally (RO) into the fish. To test the anticancer efficiency of [2](PF6)(2), the embryos were treated 24 h after engraftment at the MTD. Optimally, four consecutive PACT treatments were performed on engrafted embryos using 60 min drug-to-light intervals and 90 min green light irradiation (21 mW cm(-2), 114 J cm(-2), 520 nm). Most importantly, this PACT protocol was not toxic to the zebrafish. In the ectopic prostate tumour models, where [2](PF6)(2) showed the highest photoindex in vitro (PI > 31), the PACT treatment did not significantly diminish the growth of primary lesions, while in both conjunctival melanoma orthotopic tumour models, where [2](PF6)(2) showed more modest photoindexes (PI similar to 9), retro-orbitally administered PACT treatment significantly inhibited growth of the engrafted tumors. Overall, this study represents the first demonstration in zebrafish cancer models of the clinical potential of ruthenium-based PACT, here against conjunctival melanoma. Show less
