Photodynamic Inactivation of Human Coronaviruses.

Photodynamic inactivation (PDI) employs a photosensitizer, light, and oxygen to create a local burst of reactive oxygen species (ROS) that can inactivate microorganisms. The botanical extract PhytoQuinTM is a powerful photosensitizer with antimicrobial properties. We previously demonstrated that photoactivated PhytoQuin also has antiviral properties against herpes simplex viruses and adenoviruses in a dose-dependent manner across a broad range of sub-cytotoxic concentrations. Here, we report that human coronaviruses (HCoVs) are also susceptible to photodynamic inactivation. Photoactivated-PhytoQuin inhibited the replication of the alphacoronavirus HCoV-229E and the betacoronavirus HCoV-OC43 in cultured cells across a range of sub-cytotoxic doses. This antiviral effect was light-dependent, as we observed minimal antiviral effect of PhytoQuin in the absence of photoactivation. Using RNase protection assays, we observed that PDI disrupted HCoV particle integrity allowing for the digestion of viral RNA by exogenous ribonucleases. Using lentiviruses pseudotyped with the SARS-CoV-2 Spike (S) protein, we once again observed a strong, light-dependent antiviral effect of PhytoQuin, which prevented S-mediated entry into human cells. We also observed that PhytoQuin PDI altered S protein electrophoretic mobility. The PhytoQuin constituent emodin displayed equivalent light-dependent antiviral activity to PhytoQuin in matched-dose experiments, indicating that it plays a central role in PhytoQuin PDI against CoVs. Together, these findings demonstrate that HCoV lipid envelopes and proteins are damaged by PhytoQuin PDI and expands the list of susceptible viruses.

Enhanced Production and Anticancer Properties of Photoactivated Perylenequinones.

Hypocrellins and hypomycins are naturally occurring fungal perylenequinones with potential photodynamic activity against cancer and microbial diseases. This project pursued three lines of research. First, the production of perylenequinones was enhanced by investigating the effect of culture medium and light exposure on their biosynthesis. Solid-fermentation cultures on rice medium allowed for enhanced production of hypocrellins as compared to Cheerios or oatmeal medium. Alternatively, increased production of hypomycins, which are structurally related to the hypocrellins, was observed on oatmeal medium. In both cases, light exposure was an essential factor for the enhanced biosynthesis. In addition, this led to the discovery of two new perylenequinones, ent-shiraiachrome A (5) and hypomycin E (8), which were elucidated based on spectroscopic data. Finally, the photocytotoxic effects of both classes of compounds were evaluated against human skin melanoma, with EC50 values at nanomolar levels for hypocrellins and micromolar levels for hypomycins. In contrast, both classes of compounds showed reduced dark toxicity (EC50 values >100 µM), demonstrating promising phototherapeutic indices.

Breaking the barrier: an osmium photosensitizer with unprecedented hypoxic phototoxicity for real world photodynamic therapy.

Hypoxia presents a two-fold challenge in the treatment of cancer, as low oxygen conditions induce biological changes that make malignant tissues simultaneously more aggressive and less susceptible to standard chemotherapy. This paper reports the first metal-based photosensitizer that approaches the ideal properties for a phototherapy agent. The Os(phen)2-based scaffold was combined with a series of IP-nT ligands, where phen = 1,10-phenanthroline and IP-nT = imidazo[4,5-f][1,10]phenanthroline tethered to n = 0-4 thiophene rings. Os-4T (n = 4) emerged as the most promising complex in the series, with picomolar activity and a phototherapeutic index (PI) exceeding 106 in normoxia. The photosensitizer exhibited an unprecedented PI > 90 (EC50 = 0.651 µM) in hypoxia (1% O2) with visible and green light, and a PI > 70 with red light. Os-4T was also active with 733 nm near-infrared light (EC50 = 0.803 µM, PI = 77) under normoxia. Both computation and spectroscopic studies confirmed a switch in the nature of the lowest-lying triplet excited state from triplet metal-to-ligand charge transfer (3MLCT) to intraligand charge transfer (3ILCT) at n = 3, with a lower energy and longer lifetime for n = 4. All compounds in the series were relatively nontoxic in the dark but became increasingly phototoxic with additional thiophenes. These normoxic and hypoxic activities are the largest reported to date, demonstrating the utility of osmium for phototherapy applications. Moreover, Os-4T had a maximum tolerated dose (MTD) in mice that was >200 mg kg-1, which positions this photosensitizer as an excellent candidate for in vivo applications.

Bis[pyrrolyl Ru(ii)] triads: a new class of photosensitizers for metal-organic photodynamic therapy.

A new family of ten dinuclear Ru(ii) complexes based on the bis[pyrrolyl Ru(ii)] triad scaffold, where two Ru(bpy)2 centers are separated by a variety of organic linkers, was prepared to evaluate the influence of the organic chromophore on the spectroscopic and in vitro photodynamic therapy (PDT) properties of the compounds. The bis[pyrrolyl Ru(ii)] triads absorbed strongly throughout the visible region, with several members having molar extinction coefficients (ε) ≥ 104 at 600-620 nm and longer. Phosphorescence quantum yields (Φ p) were generally less than 0.1% and in some cases undetectable. The singlet oxygen quantum yields (Φ Δ) ranged from 5% to 77% and generally correlated with their photocytotoxicities toward human leukemia (HL-60) cells regardless of the wavelength of light used. Dark cytotoxicities varied ten-fold, with EC50 values in the range of 10-100 µM and phototherapeutic indices (PIs) as large as 5400 and 260 with broadband visible (28 J cm-2, 7.8 mW cm-2) and 625 nm red (100 J cm-2, 42 mW cm-2) light, respectively. The bis[pyrrolyl Ru(ii)] triad with a pyrenyl linker (5h) was especially potent, with an EC50 value of 1 nM and PI > 27 000 with visible light and subnanomolar activity with 625 nm light (100 J cm-2, 28 mW cm-2). The lead compound 5h was also tested in a tumor spheroid assay using the HL60 cell line and exhibited greater photocytotoxicity in this more resistant model (EC50 = 60 nM and PI > 1200 with 625 nm light) despite a lower dark cytotoxicity. The in vitro PDT effects of 5h extended to bacteria, where submicromolar EC50 values and PIs >300 against S. mutans and S. aureus were obtained with visible light. This activity was attenuated with 625 nm red light, but PIs were still near 50. The ligand-localized 3ππ* state contributed by the pyrenyl linker of 5h likely plays a key role in its phototoxic effects toward cancer cells and bacteria.

Near-infrared absorbing Ru(ii) complexes act as immunoprotective photodynamic therapy (PDT) agents against aggressive melanoma.

Mounting evidence over the past 20 years suggests that photodynamic therapy (PDT), an anticancer modality known mostly as a local treatment, has the capacity to invoke a systemic antitumor immune response, leading to protection against tumor recurrence. For aggressive cancers such as melanoma, where chemotherapy and radiotherapy are ineffective, immunomodulating PDT as an adjuvant to surgery is of interest. Towards the development of specialized photosensitizers (PSs) for treating pigmented melanomas, nine new near-infrared (NIR) absorbing PSs based on a Ru(ii) tris-heteroleptic scaffold [Ru(NNN)(NN)(L)]Cl n , were explored. Compounds 2, 6, and 9 exhibited high potency toward melanoma cells, with visible EC50 values as low as 0.292-0.602 µM and PIs as high as 156-360. Single-micromolar phototoxicity was obtained with NIR-light (733 nm) with PIs up to 71. The common feature of these lead NIR PSs was an accessible low-energy triplet intraligand (3IL) excited state for high singlet oxygen (1O2) quantum yields (69-93%), which was only possible when the photosensitizing 3IL states were lower in energy than the lowest triplet metal-to-ligand charge transfer (3MLCT) excited states that typically govern Ru(ii) polypyridyl photophysics. PDT treatment with 2 elicited a pro-inflammatory response alongside immunogenic cell death in mouse B16F10 melanoma cells and proved safe for in vivo administration (maximum tolerated dose = 50 mg kg-1). Female and male mice vaccinated with B16F10 cells that were PDT-treated with 2 and challenged with live B16F10 cells exhibited 80 and 55% protection from tumor growth, respectively, leading to significantly improved survival and excellent hazard ratios of ≤0.2.

Strained, Photoejecting Ru(II) Complexes that are Cytotoxic Under Hypoxic Conditions.

A series of strained Ru(II) complexes were studied for potential anticancer activity in hypoxic tissues. The complexes were constructed with methylated ligands that were photolabile and an imidizo[4,5-f][1,10]phenanthroline ligand that contained an appended aromatic group to potentially allow for contributions of ligand-centered excited states. A systematic variation of the size and energy of the aromatic group was performed using systems containing 1-4 fused rings, and the photochemical and photobiological behaviors of all complexes were assessed. The structure and nature of the aromatic group had a subtle impact on photochemistry, altering environmental sensitivity, and had a significant impact on cellular cytotoxicity and photobiology. Up to 5-fold differences in cytotoxicity were observed in the absence of light activation; this rose to 50-fold differences upon exposure to 453 nm light. Most significantly, one complex retained activity under conditions with 1% O2 , which is used to induce hypoxic changes. This system exhibited a photocytotoxicity index (PI) of 15, which is in marked contrast to most other Ru(II) complexes, including those designed for O2 -independent mechanisms of action.

Synthesis and Characterization of Ru(II) Complexes with π-Expansive Imidazophen Ligands for the Photokilling of Human Melanoma Cells.

Ru(II) complexes were synthesized with π-expanding (phenyl, fluorenyl, phenanthrenyl, naphthalen-1-yl, naphthalene-2-yl, anthryl and pyrenyl groups) attached at a 1H-imidazo[4,5-f][1,10]phenanthroline ligand and 4,4'-dimethyl-2,2'-bipyridine (4,4'-dmb) coligands. These Ru(II) complexes were characterized by 1D and 2D NMR, and mass spectroscopy, and studied for visible light and dark toxicity to human malignant melanoma SK-MEL-28 cells. In the SK-MEL-28 cells, the Ru(II) complexes are highly phototoxic (EC50  = 0.2-0.5 µm) and have low dark toxicity (EC50  = 58-230 µm). The highest phototherapeutic index (PI) of the series was found with the Ru(II) complex bearing the 2-(pyren-1-yl)-1H-imidazo[4,5-f][1,10]phenanthroline ligand. This high PI is in part attributed to the π-rich character added by the pyrenyl group, and a possible low-lying and longer-lived 3 IL state due to equilibration with the 3 MLCT state. While this pyrenyl Ru(II) complex possessed a relatively high quantum yield for singlet oxygen formation (Φ∆  = 0.84), contributions from type-I processes (oxygen radicals and radical ions) are competitive with the type-II (1 O2 ) process based on effects of added sodium azide and solvent deuteration.

A New Class of Homoleptic and Heteroleptic Bis(terpyridine) Iridium(III) Complexes with Strong Photodynamic Therapy Effects.

Six homo- or heteroleptic tricationic Ir(R1-tpy)(R2-tpy)3+ complexes (Ir1-Ir6, R1/R2 = Ph, 4'-N(CH3)2Ph, pyren-1-yl, or 4'-{2-[2-(2-methoxyethoxy)ethoxy]ethoxy}Ph, tpy = 2,2';6',2"-terpyridine) were synthesized and tested for photodynamic therapy (PDT) effects. The ground- and excited-state characteristics of these complexes were studied systematically via spectroscopic methods and quantum chemistry calculations. All complexes possessed intraligand charge transfer (1ILCT) / metal-to-ligand charge transfer (1MLCT) dominated transition(s) in their low-energy absorption bands, which red-shifted with the increased electron-releasing strength of the R1/R2 substituent. Five of the complexes exhibited ligand-centered 3 π,π*/3ILCT/3MLCT emission. With a stronger electron-releasing R1/R2 substituent, the degree of charge transfer contribution increased, leading to a decrease of the emission quantum yield. When the 4'-N(CH3)2Ph substituent was introduced on both tpy ligands, the emission of Ir3 was completely quenched. Our study on the transient absorption of these complexes demonstrated that they all possessed broadband triplet excited-state absorption in the 400-800 nm region. Pyrenyl substitution of one or more tpy ligands, as in Ir4 and Ir5, increased the lifetimes of the lowest triplet excited state and the singlet oxygen (1O2) production efficiencies. Ir1-Ir5 were nontoxic toward SK-MEL-28 cells, with photocytotoxicities that varied from 0.18 to 153 µM. Among them, Ir4 had the highest 1O2 quantum yield (0.81) in cell-free conditions, showing the largest photocytotoxicity against SK-MEL-28 cells for Ir(III) PSs to date, and was the most efficient generator of reactive oxygen species (ROS) in vitro. Ir4 possessed a very large phototherapeutic index (PI = dark EC50 / light EC50) of >1657, the largest reported for an Ir(III) complex photosensitizer upon broadband visible light (400-700 nm) activation. Ir4 also exhibited a very strong PDT effect toward MCF-7 breast cancer cells and its xenograft tumor model. Upon 450-nm light activation, Ir4 dramatically inhibited the xenograft tumor growth and exhibited negligible side effects upon PDT treatment.

Photophysical Properties and Photobiological Activities of Ruthenium(II) Complexes Bearing π-Expansive Cyclometalating Ligands with Thienyl Groups.

A new family of cyclometalated ruthenium(II) complexes [Ru(N^N)2(C^N)]+ derived from the π-extended benzo[h]imidazo[4,5-f]quinolone ligand appended with thienyl groups (n = 1-4, compounds 1-4) was prepared and its members were characterized for their chemical, photophysical, and photobiological properties. The lipophilicities of 1-4, determined as octanol-water partition coefficients (log Po/w), were positive and increased with the number of thienyl units. The absorption and emission bands of the C^N compounds were red-shifted by up to 200 nm relative to the analogous Ru(II) diimine systems. All of the complexes exhibited dual emission with the intraligand fluorescence (1IL, C^N-based) shifting to lower energies with increasing n and the metal-to-ligand charge transfer phosphorescence (3MLCT, N^N-based) remaining unchanged. Compounds 1-3 exhibited excited state absorption (ESA) profiles consistent with lowest-lying 3MLCT states when probed by nanosecond transient absorption (TA) spectroscopy with 532 nm excitation and had contributions from 1IL(C^N) states with 355 nm excitation. These assignments were supported by the lifetimes observed (<10 ns for the 1IL states and around 20 ns for the 3MLCT states) as well as a noticeable ESA for 3 with 355 nm excitation that did not occur with 532 nm excitation. Compound 4 was the only member of the family with two 3MLCT emissive lifetimes (15, 110 ns), and the TA spectra collected with both 355 and 532 nm excitation was assigned to the 3IL state, which was corroborated by its 4-6 µs lifetime. The ESA for 4 had a rise time of approximately 10 ns and an initial decay of 110 ns, which suggests a possible 3MLCT-3IL excited state equilibrium that results in delayed emission from the 3MLCT state. Compound 4 was nontoxic toward human skin melanoma cells (SKMEL28) in the dark (EC50 = >300 µM); 1-3 were cytotoxic and yielded EC50 values between 1 and 20 µM. The photocytotoxicites with visible light ranged from 87 nM with a phototherapeutic index (PI) of 13 for 1 to approximately 1 µM (PI = >267) for 4. With red light, EC50 values varied from 270 nM (PI = 21) for 3 to 12 µM for 4 (PI = >25). The larger PIs for 4, especially with visible light, were attributed to the much lower dark cytotoxicity for this compound. Because the dark cytotoxicity contributes substantially to the observed photocytotoxicity for 1-3, it was not possible to assess whether the 3IL state of 4 led to a much more potent phototoxic mechanism in the absence of dark toxicity. There was no stark contrast in cellular uptake and accumulation by laser scanning confocal and differential interference contrast microscopy to explain the large differences in dark toxicities between 1-3 and 4. Nevertheless, the study highlights a new family of Ru(II) C^N complexes where π-conjugation beyond a certain point results in low dark cytotoxicity with high photocytotoxicity, opposing the notion that cyclometalated Ru(II) systems are too toxic to be phototherapeutic agents.

Neutral iridium(iii) complexes bearing BODIPY-substituted N-heterocyclic carbene (NHC) ligands: synthesis, photophysics, in vitro theranostic photodynamic therapy, and antimicrobial activity.

The synthesis, photophysics, and photobiological activities of a series of novel neutral heteroleptic cyclometalated iridium(iii) complexes incorporating boron dipyrromethene (BODIPY) substituted N-heterocyclic carbene (NHC) ligands (Ir1-Ir5) are reported. The effect of the substitution position of BODIPY on the NHC ligands, either on C4 of the phenyl ring (Ir1-Ir3) or C5 of the benzimidazole unit (Ir4 and Ir5), and its linker type (single or triple bond) on the photophysical properties was studied. Ir1-Ir5 exhibited BODIPY-localized intense 1IL (intraligand transition)/1MLCT (metal-to-ligand charge transfer) absorption at 530-543 nm and 1,3IL/1,3CT (charge transfer) emission at 582-610 nm. The nanosecond transient absorption results revealed that the lowest triplet excited states of these complexes were the BODIPY-localized 3π,π* states. Complexes Ir4 and Ir5 exhibited blue-shifted 1IL absorption and 1,3IL/1,3CT emission bands compared to the corresponding absorption and emission bands in complexes Ir1 and Ir3. However, replacing the methyl substituents on N3 of benzimidazole in complexes Ir1 and Ir4 with oligoether substituents in Ir3 and Ir5, respectively, did not impact the energies of the low-energy absorption and emission bands in the corresponding complexes. Water-soluble complexes Ir3 and Ir5 have been explored as photosensitizers for in vitro photodynamic therapy (PDT) effects toward human SKMEL28 melanoma cells. Ir3 showed no dark cytotoxicity (EC50 > 300 µM) but good photocytotoxic activity (9.66 ± 0.28 µM), whereas Ir5 exhibited a higher dark cytotoxicity (20.2 ± 1.26 µM) and excellent photocytotoxicity (0.15 ± 0.01 µM). The phototherapeutic indices with visible light (400-700 nm) activation were >31 for Ir3 and 135 for Ir5. Ir3 and Ir5 displayed 1O2 quantum yields of 38% and 22% in CH3CN, respectively, upon 450 nm excitation. Ir5 was more effective at generating reactive oxygen species (ROS) in vitro. Ir5 was also active against Staphylococcus aureus upon visible light activation, with a phototherapeutic index of >15 and EC50 value of 6.67 µM. These photobiological activities demonstrated that these neutral Ir(iii) complexes are promising in vitro PDT reagents, and substitution at C5 on the benzimidazole group of the NHC ligand was superior to C4 substitution on the phenyl ring.

Predictive Strength of Photophysical Measurements for in Vitro Photobiological Activity in a Series of Ru(II) Polypyridyl Complexes Derived from π-Extended Ligands.

This study investigates the correlation between photocytotoxicity and the prolonged excited-state lifetimes exhibited by certain Ru(II) polypyridyl photosensitizers comprised of π-expansive ligands. The eight metal complexes selected for this study differ markedly in their triplet state configurations and lifetimes. Human melanoma SKMEL28 and human leukemia HL60 cells were used as in vitro models to test photocytotoxicity induced by the compounds when activated by either broadband visible or monochromatic red light. The photocytotoxicities of the metal complexes investigated varied over 2 orders of magnitude and were positively correlated with their excited-state lifetimes. The complexes with the longest excited-state lifetimes, contributed by low-lying 3IL states, were the most phototoxic toward cancer cells under all conditions.

Heteroleptic Ir(III)N6 Complexes with Long-Lived Triplet Excited States and in Vitro Photobiological Activities.

A series of cationic heteroleptic iridium(III) complexes bearing tris-diimine ligands [Ir(phen)2(R-phen)]3+ (R-phen = phenanthroline (1), 3,8-diphenylphenanthroline (2), 3,8-dipyrenylphenanthroline (3), 3-phenylphenanthroline (4), 3-pyrenylphenanthroline (5), and 3,8-diphenylethynylphenanthroline (6)) were synthesized and characterized. These complexes possessed phen ligand-localized 1π,π* transitions below 300 nm, and charge transfer (1CT) and/or 1π,π* transitions between 300 and 520 nm. In 1, 2, 4, and 6, the low-energy bands were mixed 1CT/1π,π*. However, the increased π-donating ability of the pyrenyl substituent(s) in 3 and 5 split the low-energy bands into a pyrene-based 1π,π* transition at 300-380 nm and an intraligand charge transfer (1ILCT) transition at 380-520 nm. All complexes were emissive at room temperature in CH3CN, but the parentage of the emitting state varied depending on the R substituent(s). Complex 1 exhibited predominantly phen ligand-localized 3π,π* emission mixed with metal-to-ligand charge transfer (3MLCT) character, whereas the emission of 2, 4, and 6 was predominantly from the excited-state with 3π,π*/3ILCT/3MLCT character. The emission from 3 and 5 was dominated by pyrene-based 3π,π* states mixed with 3ILCT character. The different natures of the lowest triplet excited states were also reflected by the different spectral features and lifetimes of the triplet transient absorption of these complexes. Complexes 3 and 5 had singlet oxygen quantum yields as high as 81 and 72%, respectively. Both gave submicromolar phototoxicities toward cancer cells (SK-MEL-28 human melanoma) and bacteria ( S. aureus and S. mutans) with visible-light activation (and marginal to no photobiological activity with red light). Their visible-light phototherapeutic indices (PIs) toward SK-MEL-28 cells were 248 for 3 and >435 for 5; PIs were lower in bacteria (≤62) because of their inherent antimicrobial activities. Both complexes were shown to produce substantial amounts of intracellular reactive oxygen species (ROS), which may account for their photobiological activities.

Transition Metal Complexes and Photodynamic Therapy from a Tumor-Centered Approach: Challenges, Opportunities, and Highlights from the Development of TLD1433.

Transition metal complexes are of increasing interest as photosensitizers in photodynamic therapy (PDT) and, more recently, for photochemotherapy (PCT). In recent years, Ru(II) polypyridyl complexes have emerged as promising systems for both PDT and PCT. Their rich photochemical and photophysical properties derive from a variety of excited-state electronic configurations accessible with visible and near-infrared light, and these properties can be exploited for both energy- and electron-transfer processes that can yield highly potent oxygen-dependent and/or oxygen-independent photobiological activity. Selected examples highlight the use of rational design in coordination chemistry to control the lowest-energy triplet excited-state configurations for eliciting a particular type of photoreactivity for PDT and/or PCT effects. These principles are also discussed in the context of the development of TLD1433, the first Ru(II)-based photosensitizer for PDT to enter a human clinical trial. The design of TLD1433 arose from a tumor-centered approach, as part of a complete PDT package that includes the light component and the protocol for treating non-muscle invasive bladder cancer. Briefly, this review summarizes the challenges to bringing PDT into mainstream cancer therapy. It considers the chemical and photophysical solutions that transition metal complexes offer, and it puts into context the multidisciplinary effort needed to bring a new drug to clinical trial.

Synthesis, Characterization and Photobiological Studies of Ru(II) Dyads Derived from α-Oligothiophene Derivatives of 1,10-Phenanthroline.

Three new bis(2,2'-bipyridine)-heteroleptic Ru(II) dyads incorporating thienyl groups (n = 1-3, compounds 1, 2 and 3, respectively) appended to 1,10-phenanthroline were synthesized and characterized to investigate the impact of n on the photophysical and photobiological properties within the series. All three complexes showed unstructured emission near 618 nm from a triplet metal-to-ligand charge transfer (3 MLCT) state with a lifetime (τem ) of approximately 1 µs. Transient absorption measurements revealed an additional excited state that was nonemissive and long-lived (τTA  = 43 µs for 2 and 27 µs for 3), assigned as a triplet intraligand (3 IL) state that was accessible only in 2 and 3. All three complexes were strong singlet oxygen (1 O2 ) sensitizers, with quantum yields (Φ∆ ) for 2 and 3 being the largest (74-78%), and all three were photocytotoxic to cancer cells with visible light activation in the order: 3 > 2 > 1. Cell-free DNA photodamage followed the same trend, where potency increased with decreasing 3 IL energy. Compounds 2 and 3 also showed in vitro photobiological effects with red light (625 nm), where their molar absorptivities were <100 m-1  cm-1 . These findings highlight that Ru(II) dyads derived from α-oligothiophenes directly appended to 1,10-phenanthroline-namely 2 and 3-possess low-lying 3 IL states that are highly photosensitizing, and they may therefore be of interest for photobiological applications such as photodynamic therapy (PDT).

Photodynamic Inactivation of Herpes Simplex Viruses.

Herpes simplex virus (HSV) infections can be treated with direct acting antivirals like acyclovir and foscarnet, but long-term use can lead to drug resistance, which motivates research into broadly-acting antivirals that can provide a greater genetic barrier to resistance. Photodynamic inactivation (PDI) employs a photosensitizer, light, and oxygen to create a local burst of reactive oxygen species that inactivate microorganisms. The botanical plant extract OrthoquinTM is a powerful photosensitizer with antimicrobial properties. Here we report that Orthoquin also has antiviral properties. Photoactivated Orthoquin inhibited herpes simplex virus type 1 (HSV-1) and herpes simplex virus type 2 (HSV-2) infection of target cells in a dose-dependent manner across a broad range of sub-cytotoxic concentrations. HSV inactivation required direct contact between Orthoquin and the inoculum, whereas pre-treatment of target cells had no effect. Orthoquin did not cause appreciable damage to viral capsids or premature release of viral genomes, as measured by qPCR for the HSV-1 genome. By contrast, immunoblotting for HSV-1 antigens in purified virion preparations suggested that higher doses of Orthoquin had a physical impact on certain HSV-1 proteins that altered protein mobility or antigen detection. Orthoquin PDI also inhibited the non-enveloped adenovirus (AdV) in a dose-dependent manner, whereas Orthoquin-mediated inhibition of the enveloped vesicular stomatitis virus (VSV) was light-independent. Together, these findings suggest that the broad antiviral effects of Orthoquin-mediated PDI may stem from damage to viral attachment proteins.

Photophysical and Photobiological Properties of Dinuclear Iridium(III) Bis-tridentate Complexes.

A series of cationic dinuclear iridium(III) complexes (Ir1-Ir5) bearing terpyridine-capped fluorenyl bridging ligands and different polypyridyl or cyclometalating terminal tridentate ligands were synthesized, characterized, and evaluated for their photophysical and photobiological activities. The influence of the bridging and terminal ligands on the photophysical properties of the complexes was investigated by UV-vis absorption, emission, and transient absorption spectroscopy and simulated by TDDFT calculations. All of the complexes displayed strong bridging-ligand localized visible 1π,π* absorption and red- or near-infrared phosphorescence as well as broad triplet excited-state absorption across both visible and NIR wavelengths. These triplet states were assigned as predominantly 3π,π* for Ir1 (τ = 3.1 µs) and Ir4 (τ = 48 µs) and 3CT (charge transfer) for Ir2, Ir3, and Ir5 (τ = 1.7-2.7 µs). Complexes Ir1-Ir5 acted as in vitro photodynamic therapy (PDT) agents toward human SK-MEL-28 melanoma cells when activated with visible light, with submicromolar photocytotoxicity and phototherapeutic indices ranging from 20 to almost 300. The in vitro PDT effects with visible light did not correlate with singlet oxygen (1O2) quantum yields or DNA photocleaving capacity probed under cell-free conditions. All of the Ir(III) complexes phosphoresced brightly when associated with compromised cells (with or without light treatment) and exhibited photoactivated cellular uptake, highlighting the theranostic potential of this new class of Ir(III) complex photosensitizers.

Cyclometalated Ruthenium(II) Complexes Derived from α-Oligothiophenes as Highly Selective Cytotoxic or Photocytotoxic Agents.

The photophysical and photobiological properties of a new class of cyclometalated ruthenium(II) compounds incorporating π-extended benzo[ h]imidazo[4,5- f]quinoline (IBQ) cyclometalating ligands (C^N) bearing thienyl rings ( n = 1-4, compounds 1-4) were investigated. Their octanol-water partition coefficients (log Po/w) were positive and increased with n. Their absorption and emission energies were red-shifted substantially compared to the analogous Ru(II) diimine (N^N) complexes. They displayed C^N-based intraligand (IL) fluorescence and triplet excited-state absorption that shifted to longer wavelengths with increasing n and N^N-based metal-to-ligand charge transfer (MLCT) phosphorescence that was independent of n. Their photoluminescence lifetimes (τem) ranged from 4-10 ns for 1IL states and 12-18 ns for 3MLCT states. Transient absorption lifetimes (τTA) were 5-8 µs with 355 nm excitation, ascribed to 3IL states that became inaccessible for 1-3 with 532 nm excitation (1-3, τTA = 16-17 ns); the 3IL of 4 only was accessible by lower energy excitation, τTA = 3.8 µs. Complex 4 was nontoxic (EC50 > 300 µM) to SK-MEL-28 melanoma cells and CCD1064-Sk normal skin fibroblasts in the dark, while 3 was selectively cytotoxic to melanoma (EC50= 5.1 µM) only. Compounds 1 and 2 were selective for melanoma cells in the dark, with submicromolar potencies (EC50 = 350-500 nM) and selectivity factors (SFs) around 50. The photocytotoxicities of compounds 1-4 toward melanoma cells were similar, but only compounds 3 and 4 displayed significant phototherapeutic indices (PIs; 3, 43; 4, >1100). The larger cytotoxicities for compounds 1 and 2 were attributed to increased cellular uptake and nuclear accumulation, and possibly related to the DNA-aggregating properties of all four compounds as demonstrated by cell-free gel mobility-shift assays. Together, these results demonstrate a new class of thiophene-containing Ru(II) cyclometalated compounds that contain both highly selective chemotherapeutic agents and extremely potent photocytotoxic agents.

Near-infrared-emitting heteroleptic cationic iridium complexes derived from 2,3-diphenylbenzo[g]quinoxaline as in vitro theranostic photodynamic therapy agents.

Five heteroleptic cationic iridium complexes with a π-expansive cyclometalating 2,3-diphenylbenzo[g]quinoxaline (dpbq) ligand (C^N ligand) and different diimine ligands (N^N ligands) (i.e. 2,2'-bipyridine (bpy, 1), phenanthroline (phen, 2), 2-(2-pyridinyl)quinoline (pqu, 3), 2,2'-bisquinoline (bqu, 4), and 2-(quinolin-2-yl)quinoxaline (quqo, 5)) were synthesized and characterized. The lowest-energy singlet electronic transitions (S1 states) were mainly dpbq ligand-centred 1ILCT (intraligand charge transfer)/1MLCT (metal to ligand charge transfer) transitions mixed with some 1π,π* transitions for complexes 1-4 with increased contributions from 1LLCT (ligand to ligand charge transfer) in 3 and 4. For complex 5, the S1 state was switched to the 1LLCT/1MLCT transitions. All five complexes displayed weak near-infrared (NIR) phosphorescence, with maximal emission output spanning 700-1400 nm and quantum yields being on the order of 10-3. The triplet state absorptions of 1-4 all resembled that of the [Ir(dpbq)2Cl]2 dimer with lifetimes of ca. 400 ns, while the TA spectrum of 5 possessed the characteristics of both the quqo ligand and the [Ir(dpbq)2Cl]2 dimer with a bi-exponential decay of ca. 5 µs and 400 ns. While the photophysics of these complexes differ slightly, their theranostic photodynamic therapy (PDT) effects varied drastically. All of the complexes were biologically active toward melanoma cells. Complexes 2 and 3 were the most cytotoxic, with 230-340 nM activity and selectivity factors for melanoma cells over normal skin fibroblasts of 34 to 40 fold. Complexes 2, 3, and 5 became very potent cytotoxins with light activation, with EC50 values as low as 12-18 nM. This potent nanomolar light-triggered activity combined with a lower dark toxicity resulted in 5 having a phototherapeutic index (PI) margin of almost 275. The bpy coligand led to the least amount of dark toxicity of 1, while phen and pqu produced cytotoxic but selective complexes 2 and 3. The quqo coligand produced the most potent complex 5 for in vitro PDT, both in terms of photocytotoxicity and PI. All Ir(iii) complexes exhibited very bright NIR phosphorescence in melanoma cells. The wide range of cytotoxicity and photocytotoxicity effects within a relatively small class of complexes highlights the importance of the identity of the coligand in the biological activity of the π-expansive biscyclometalated Ir(iii) complexes, and their bright NIR emission in live cells demonstrates their potential as theranostic PDT agents.

Novel Osmium-based Coordination Complexes as Photosensitizers for Panchromatic Photodynamic Therapy.

Cancer remains a major global malaise requiring the advent of new, efficient and low-cost treatments. Photodynamic therapy, which combines a photosensitizer and photons to produce cytotoxic reactive oxygen species, has been established as an effective cancer treatment but has yet to become mainstream. One of the main limitations has been the paucity of photosensitizers that are effective over a wide range of wavelengths, can exert their cytotoxic effects in hypoxia, are easily synthesized and produce few if any side effects. To address these shortfalls, three new osmium-based photosensitizers (TLD1822, TLD1824 and TLD1829) were synthesized and their photophysical and photobiological attributes determined. These photosensitizers are panchromatic (i.e. black absorbers), activatable from 200 to 900 nm and have strong resistance to photobleaching. In vitro studies show photodynamic therapy efficacy with both red and near-infrared light in normoxic and hypoxic conditions, which translated to good in vivo efficacy of TLD1829 in a subcutaneous murine colon cancer model.