Photodynamic Therapy Experiments Research Articles

Theranostic dye entrapped in an optimized blended-polymer matrix for effective photodynamic inactivation of diseased cells.

Despite the wide range of treatment options available for cancer therapy, including chemotherapy, radiation therapy, and surgical procedures, each of these treatments has a different side-effect profile and leaves the patient with no option but to choose. Due to their insensitivity and nonspecificity, conventional treatments damage normal cells together with cancer cells. In recent years, a significant amount of attention has been focused on photodynamic therapy (PDT) as a treatment for cancer and drug-resistant microbes. An activated photosensitizer is used as a part of the procedure along with oxygen molecules and a specific wavelength of light belonging to the visible or NIR spectral zone. A light-sensitive laser dye, rhodamine 6G (R6G), was used in the present study as a photosensitizer, taking a challenge to improve the aqueous solubility and ROS quantum yield using optimum concentration (160mg/ml) of chitosan-alginate (Cs-Alg) blended polymeric nanoformulations. As evidenced by steady-state spectrophotometric and fluorometric measurements, ROS quantum yield increases three-fold over aqueous solution along with solubility gaining that was validated by PDT experiment using human epithelial carcinoma (KB) cell line. Phantom optical imaging was taken using the IVIS imaging system to establish the formulations as a fluorescence-based optical contrast agent, and zebrafish embryos were used to establish their safe in vivo use. The release profile of R6G was fitted using kinetic models, which followed the Non-Fickian kinetic profile. In conclusion, we recommend the formulations as a potential theranostic agent that will aid in PDT-based therapy in conjunction with optical imaging-based diagnosis.

Molecular engineering of a spheroid-penetrating phage nanovector for photodynamic treatment of colon cancer cells

Photodynamic therapy (PDT) represents an emerging strategy to treat various malignancies, including colorectal cancer (CC), the third most common cancer type. This work presents an engineered M13 phage retargeted towards CC cells through pentavalent display of a disulfide-constrained peptide nonamer. The M13CC nanovector was conjugated with the photosensitizer Rose Bengal (RB), and the photodynamic anticancer effects of the resulting M13CC-RB bioconjugate were investigated on CC cells. We show that upon irradiation M13CC-RB is able to impair CC cell viability, and that this effect depends on i) photosensitizer concentration and ii) targeting efficiency towards CC cell lines, proving the specificity of the vector compared to unmodified M13 phage. We also demonstrate that M13CC-RB enhances generation and intracellular accumulation of reactive oxygen species (ROS) triggering CC cell death. To further investigate the anticancer potential of M13CC-RB, we performed PDT experiments on 3D CC spheroids, proving, for the first time, the ability of engineered M13 phage conjugates to deeply penetrate multicellular spheroids. Moreover, significant photodynamic effects, including spheroid disruption and cytotoxicity, were readily triggered at picomolar concentrations of the phage vector. Taken together, our results promote engineered M13 phages as promising nanovector platform for targeted photosensitization, paving the way to novel adjuvant approaches to fight CC malignancies.

Constructing efficient and photostable photosensitizer with aggregation-induced emission by introducing highly electronegative nitrogen atom for photodynamic therapy

Photodynamic therapy (PDT) has drawn considerable attention owning to many unique advantages, such as precise controllability, noninvasiveness, high efficiency and spatiotemporal selectivity. However, most of the currently utilized photosensitizers (PSs) have drawbacks of weak light absorption for excitation and low reactive oxygen species (ROS) generation ability. In this contribution, through introducing highly electronegative nitrogen atom, two photostable photosensitizers are designed to display high absorption coefficient and efficient ROS generation ability. The yield BTPTT-CNTPE nanoparticles show good performance of excellent photostability, high ROS generation efficiency, negligible dark toxicity and high absorption coefficient. Subsequent PDT experiments both in vitro and in vivo prove that BTPTT-CNTPE nanoparticles are promising candidates for PDT anticancer therapy.

On the need for standardized reporting of photophysical parameters of in vitro photodynamic therapy studies.

In vitro dose escalation experiments are one of the first gatekeepers in therapeutic evaluation and development. This also holds for evaluating novel photosensitizers (PS) and Photodynamic Therapy (PDT) co-therapies as needed to provide dose response guidelines before engaging in further pre-clinical studies. The dose needed to achieve 50% cell kill (LD50) is a standard metric to report the potency of a therapeutic agents that is widely accepted for single-drug therapies. In reporting results of PDT experiments, which involve delivery of both drug and light, it is inherently more complicated to identify such a convenient dose response metric that actually captures the larger space of treatment parameters. In addition to ubiquitous sources of biological variability that apply broadly in biomedical research, PDT treatment efficacy is determined by multiple key parameters that may or may not have been documented, including PS concentration and light fluence, where the latter is itself a function of the spectral properties of the light source used (often not described), not to mention dose rate, fractionation and other parameters that potentially vary between individual studies. It is impossible to compare results between two study when, for example one reports LD50 PS concentration without providing essential light dosimetry details. Motivated by this challenge in comparing outcomes and establishing reproducibility of in vitro PDT studies, we endeavored to perform a meta-analysis of the reporting of PDT results by converting, where possible, the disparately reported experimental details into a consistent metric that could be used to compare across studies. In this context we adopt here the number of photons absorbed by photosensitizers per unit volume to affect a 50% decline in cell survival as a standardized metric. By choosing this metric one can acknowledge the quantum-based generation of cytotoxins. While this metric does not cover every possible source of variability between any two studies, for a PS with known optical properties, this does encapsulate PS concentration as well as irradiance and spectral properties of light delivered. For the sake of focus we adopt this approach for study of reported results with two photosensitizers, Protoporphyrin IX, either synthesized in the cells by aminolevulinic acid or administered exogenously, and Chlorin e6. A literature search was performed to identify in vitro studies with these two photosensitizers and collect necessary information to calculate the absorbed photon LD50 threshold for each study. Only approximately 1/10 of the manuscripts reporting on in vitro studies provide the minimum required information to calculate the threshold values. While the majority of the determined threshold values are within a factor of 10, the range of threshold values spanned close to 7 orders of magnitude for both photosensitizers. To contrast with single-agent therapies, a similar exercise was performed for chemotherapeutic drugs targeting cellular mitosis or tyrosine kinase inhibitors resulted in an LD50 or IC50 range of 1-2 orders of magnitude, with LD50 or IC50 values for a single cell line being within a factor of 5. This review underscores challenges in the reporting of in vitro PDT efficacy. In many cases it takes considerable effort to extract the necessary methodology information to make meaningful comparison between PDT studies. Only when results between studies can be compared is it possible to begin to assess reproducibility which, as shown here, can be a major issue. Hence, guidelines need to be developed and enforced through the peer review process for meaningful reporting of preclinical PDT results in order for the most promising sensitizers and co-therapies to be identified and translated into the clinic.

An open-source LED array illumination system for automated multiwell plate cell culture photodynamic therapy experiments

Photodynamic therapy (PDT) research would benefit from an automated, low-cost, and easy-to-use cell culture light treatment setup capable of illuminating multiple well replicates within standard multiwell plate formats. We present an LED-array suitable for performing high-throughput cell culture PDT experiments. The setup features a water-cooling loop to keep the LED-array temperature nearly constant, thus stabilizing the output power and spectrum. The setup also features two custom-made actuator arms, in combination with a pulse-width-modulation (PWM) technique, to achieve programmable and automatic light exposures for PDT. The setup operates at ~ 690 nm (676–702 nm, spectral output full-width half-maximum) and the array module can be readily adapted to other LED wavelengths. This system provides an illumination field with adjustable irradiance up to 400 mW/cm2 with relatively high spectral and power stability comparing with previously reported LED-based setups. The light doses provided by the LED array were validated with comparison to traditional laser PDT. This open-source illumination platform (including the detailed technical description, fabrication protocols, and parts list provided here) helps to make custom light sources more accessible and of practical use for photomedicine research.

A Novel PSMA-Targeted Probe for NIRF-Guided Surgery and Photodynamic Therapy: Synthesis and Preclinical Validation.

A total of 20% to 50% of prostate cancer (PCa) patients leave the surgery room with positive tumour margins. The intraoperative combination of fluorescence guided surgery (FGS) and photodynamic therapy (PDT) may be very helpful for improving tumour margin delineation and cancer therapy. PSMA is a transmembrane protein overexpressed in 90-100% of PCa cells. The goal of this work is the development of a PSMA-targeted Near InfraRed Fluorescent probe to offer the surgeon a valuable intraoperative tool for allowing a complete tumour removal, implemented with the possibility of using PDT to kill the eventual not resected cancer cells. PSMA-617 binding motif was conjugated to IRDye700DX-NHS and the conjugation did not affect the photophysical characteristics of the fluorophore. The affinity of IRDye700DX-PSMA-617 towards PCa cells followed the order of their PSMA expression, i.e., PC3-PIP > LNCaP > PC3, PC3-FLU. NIRF imaging showed a significant PC3-PIP tumour uptake after the injection of 1 or 5 nmol with a maximum tumour-to-muscle ratio (ca. 60) observed for both doses 24 h post-injection. Importantly, urine, healthy prostate, and the bladder were not fluorescent at 24 h post-injection. Flow cytometry and confocal images highlighted a co-localization of PSMA+ cells with IRDye700DX-PSMA uptake. Very interestingly, ex vivo analysis on a tumour specimen highlighted a significant PSMA expression by tumour-associated macrophages, likely attributable to extracellular vesicles secreted by the PSMA(+) tumour cells. FGS proved that IRDye700DX-PSMA was able to easily delineate tumour margins. PDT experiments showed a concentration-dependent decrease in cell viability (from 75% at 10 nM to 12% at 500 nM), whereas controls did not show any cytotoxicity. PC3-PIP tumour-bearing mice subjected to photodynamic therapy showed a delayed tumour growth. In conclusion, a novel PSMA-targeted NIRF dye with dual imaging-PDT capabilities was synthesized and displayed superior specificity compared to other small PSMA targeted molecules.

Indomethacin-based near-infrared photosensitizer for targeted photodynamic cancer therapy

Near-IR fluorescent sensitizers based on heptamethine cyanine (Cy820 and Cy820–IMC) were synthesized and their abilities to target and abolish tumor cells via photodynamic therapy (PDT) were explored. Some hepthamethine cyanine dyes can be transported into cancer cells via the organic anion transporting polypeptides (OATPs). In this study, we aimed to enhance the target ability of the sensitizer by conjugation Cy820 with indomethacin, a non-steroidal anti-inflammatory drug (NSAID), to obtain Cy820–IMC that aimed to target cyclooxygenase–2 (COX–2) which overexpresses in cancer cells. The results showed that Cy820–IMC internalized the cancer cells faster than Cy820 which was verified to be related to COX–2 level and OATPs. Based on PDT experiments, Cy820–IMC has higher photocytotoxicity index than Cy820, >7.13 and 4.90, respectively, implying that Cy820–IMC showed better PDT property than Cy820. However, Cy820 exhibits slightly higher normal-to-cancer cell toxicity ratio than Cy820–IMC, 6.58 and 3.63, respectively. Overall, Cy820–IMC has superior cancer targetability and enhanced photocytoxicity. These characteristics can be further improved towards clinically approved sensitizers for PDT.

Orange Trunk Waste-Based Lignin Nanoparticles Encapsulating Curcumin as a Photodynamic Therapy Agent against Liver Cancer

Orange wood biomass is an interesting lignocellulosic material, but its lignin has been little explored. As a structural macromolecule present in the plant cell wall, lignin has been used in nanoencapsulation and nanomedicine, standing out as a good drug carrier. In this study, lignin was extracted by soda pulping process and used to produce nanoparticles for curcumin encapsulation and subsequent application in photodynamic therapy (PDT). The nanoparticles were synthesized by the solvent shifting process and characterized with UV–vis, fluorescence, dynamic light scattering (DLS), ζ-potential, and scanning electron microscopy (SEM). The lignin was soluble in organic solvent and presented an extraction yield of 20.6 ± 2%. The Fourier transform infrared (FTIR) spectroscopy and NMR signals showed that the lignin presents hardwood-type characteristics. The lignin nanoparticles were easily obtained and presented average sizes of 284 ± 60, 287 ± 60, and 502 ± 100 nm for pure lignin, 1% curcumin, and 5% curcumin nanoparticles, respectively. They exhibited a negative ζ-potential of about −42 ± 5 mV. The formulations were evaluated for 30 days and remained stable in phosphate-buffered saline (PBS). The curcumin encapsulation yields were high, above 90% in all formulations. Scanning electron microscopy (SEM) images showed that the nanoparticles are spherical and homogeneous. Studies using hepatocarcinoma (HTC) and HepaRG cells showed that the lignin nanoparticles were nontoxic within 4 h of incubation, and for nanoparticles containing 5% curcumin, toxicity was observed upon application of light in PDT experiments.

Molecular Engineering of Efficient Singlet Oxygen Generators with Near‐Infrared AIE Features for Mitochondrial Targeted Photodynamic Therapy

AbstractAggregation‐caused fluorescence quenching with insufficient production of reactive oxygen species (ROS) has limited the application of photosensitizers (PSs) in fluorescence‐imaging‐guided photodynamic therapy (PDT). Aggregation‐induced emission PSs (AIE‐PSs) exhibit enhanced fluorescence intensity and a high efficiency of ROS generation in the aggregation state, which provides an opportunity to solve the above problems. Herein, a series of AIE‐PSs are successfully designed and synthesized by adjusting the D–A intensity through molecular engineering. The photophysical properties and theoretical calculations prove that the synergistic effect of 3,4‐ethylenedioxythiophene and quinolinium increases the intramolecular charge transfer effect (ICT) of the whole molecule and promotes the intersystem crossing (ISC) from the lowest excited singlet state (S1) to the lowest triplet state (T1). Among these AIE‐PSs, the optimal AIE‐PS (TPA‐DT‐Qy) exhibits the highest generation yield of 1O2 (5.3‐fold of Rose Bengal). Further PDT experiments show that the TPA‐DT‐Qy has a highly efficient photodynamic ablation of breast cancer cells (MCF‐7 and MDA‐MB‐231) under white light irradiation. Moreover, the photodynamic antibacterial study indicates that TPA‐DT‐Qy has the discrimination and excellent photodynamic inactivation of S. aureus. This work provides a feasible strategy for the molecular engineering of novel AIE‐PSs to improve the development of fluorescence‐imaging‐guided PDT.

NIR Fluorescent Imaging and Photodynamic Therapy with a Novel Theranostic Phospholipid Probe for Triple-Negative Breast Cancer Cells.

New exogenous probes are needed for both imaging diagnostics and therapeutics. Here, we introduce a novel nanocomposite near-infrared (NIR) fluorescent imaging probe and test its potency as a photosensitizing agent for photodynamic therapy (PDT) against triple-negative breast cancer cells. The active component in the nanocomposite is a small molecule, pyropheophorbide a-phosphatidylethanolamine-QSY21 (Pyro-PtdEtn-QSY), which is imbedded into lipid nanoparticles for transport in the body. The probe targets abnormal choline metabolism in cancer cells; specifically, the overexpression of phosphatidylcholine-specific phospholipase C (PC-PLC) in breast, prostate, and ovarian cancers. Pyro-PtdEtn-QSY consists of a NIR fluorophore and a quencher, attached to a PtdEtn moiety. It is selectively activated by PC-PLC resulting in enhanced fluorescence in cancer cells compared to normal cells. In our in vitro investigation, four breast cancer cell lines showed higher probe activation levels than noncancerous control cells, immortalized human mammary gland cells, and normal human T cells. Moreover, the ability of this nanocomposite to function as a sensitizer in PDT experiments on MDA-MB-231 cells suggests that the probe is promising as a theranostic agent.

The construction of supramolecular and hybrid Ag-AgCl nanoparticles with photodynamic therapy action on the base of tetraundecylсalix[4]resorcinarene-mPEG conjugate

The development of new nanomaterials with therapeutic potential is a modern task due to their outstanding properties and promising effectiveness. Here the usage of acylhydrazone bond-based сalix[4]resorcinarene-mPEG conjugate (C11-mPEG) in the formation of both supramolecular and hybrid nanoparticles with in vitro photodynamic therapy (PDT) activity is described. C11-mPEG is hydrolyzed with a decrease in pH of solution; the conjugate and its degradation product have the low hemolytic activity and low cytotoxicity against normal cells. The self-association of the conjugate was studied in the aqueous solutions with pH 7.4 and 5. C11-mPEG forms self-associates that serve as pH-sensitive supramolecular nanocontainers for photosensitizer Methylene Blue (MB). In the in vitro PDT experiment the stronger decrease of tumor cells (M-Hela cells) viability in the presence of encapsulated MB is observed in comparison with free MB. Also C11-mPEG acts as a stabilizing agent for Ag@AgCl nanoparticles, which were obtained with 5/1 and 2/1 Ag+/C11-mPEG molar ratio and different reducing agents (sodium borohydride and hydrazine hydrate). The stronger cytotoxicity of hybrid NPs against tumor cells in comparison with normal was shown. Further it was found that the irradiation of the hybrid nanoparticles at 630 nm leads to the sharp increase in their cytotoxicity against M-Hela cells. The high level of ROS formation in the cells in the presence of both supramolecular and hybrid NPs under in vitro PDT experiment was determined.

Preparation of zinc phthalocyanine-loaded amphiphilic phosphonium chitosan nanomicelles for enhancement of photodynamic therapy efficacy

To increase the solubility and the encapsulation of zinc phthalocyanine (ZnPc) photosensitizer for photodynamic therapy (PDT), a positively charged amphiphilic phosphonium chitosan nanomicelle with multi-benzene structure was developed, and its application to PDT was explored. N-acetyl-l-phenylalanine-(4-carboxybutyl) triphenylphosphonium bromide chitosan (CTPB-CS-NAP), a chitosan derivative with tunable amphiphilicity, was synthesized first. ZnPc was encapsulated in CTPB-CS-NAP at the critical micelle concentration (CMC) of 4.898 mg/L by a hydrophobic self-assembly method to form ZnPc-loaded nanomicelles (ZnPc@CTPB-CS-NAP). The method gives the highest encapsulation efficiency and drug loading of 89.4 % and 22.3 %, respectively. ZnPc@CTPB-CS-NAP is stably dispersed in aqueous solution and shows the average particle size of 103±5 nm. PDT experiments suggest the phototoxicity of ZnPc@CTPB-CS-NAP is much higher than that of ZnPc, but no obvious dark cytotoxicity is observed. Our study has provided a new strategy for improving the photodynamic therapy efficacy of hydrophobic photosensitizer by the encapsulation with chitosan derivative carriers.

Systematic Review and Meta-Analysis of In Vitro Anti-Human Cancer Experiments Investigating the Use of 5-Aminolevulinic Acid (5-ALA) for Photodynamic Therapy.

5-Aminolevulinic acid (5-ALA) is an amino acid derivative and a precursor of protoporphyrin IX (PpIX). The photophysical feature of PpIX is clinically used in photodynamic diagnosis (PDD) and photodynamic therapy (PDT). These clinical applications are potentially based on in vitro cell culture experiments. Thus, conducting a systematic review and meta-analysis of in vitro 5-ALA PDT experiments is meaningful and may provide opportunities to consider future perspectives in this field. We conducted a systematic literature search in PubMed to summarize the in vitro 5-ALA PDT experiments and calculated the effectiveness of 5-ALA PDT for several cancer cell types. In total, 412 articles were identified, and 77 were extracted based on our inclusion criteria. The calculated effectiveness of 5-ALA PDT was statistically analyzed, which revealed a tendency of cancer-classification-dependent sensitivity to 5-ALA PDT, and stomach cancer was significantly more sensitive to 5-ALA PDT compared with cancers of different origins. Based on our analysis, we suggest a standardized in vitro experimental protocol for 5-ALA PDT.

Enhancement of Phthalocyanine Mediated Photodynamic Therapy by Catechin on Lung Cancer Cells.

Worldwide, lung cancer remains one of the leading cancers with increasing mortality rates. Though chemotherapy for lung cancer is effective, it is always accompanied by unavoidable and grave side effects. Photodynamic therapy (PDT), using novel photosensitizers, is an advanced treatment method with relatively few side effects. Plant products are emerging as potent photosensitizers (PSs). The dose-dependent effect of Catechin (CA) (20–100 µM) on cellular morphological changes, cell viability, cytotoxicity, proliferation, DNA damage and apoptosis were studied on A549 adenocarcinoma alveolar basal epithelial cells. The effect of CA, along with Zinc phthalocyanine PS at 680 nm and 5 J/cm2 fluency was also studied. As the doses of CA increased, the results showed a pattern of increased cytotoxicity, accompanied by decreased cell viability and proliferation in A549 cells. Also, at 52 µM (IC50), CA in combination with PS significantly increased the cytotoxicity, DNA damage, and apoptosis, as compared to control and PS alone, treated cells in PDT experiments. These findings leave a possible thread that CA can be used in the application of phyto-photodynamic therapy of cancer in future.

Tin-Doped Near-Infrared Persistent Luminescence Nanoparticles with Considerable Improvement of Biological Window Activation for Deep Tumor Photodynamic Therapy.

The photodynamic therapy (PDT) as a promising antitumor therapy technique is greatly hampered by the low tissue penetration of light and the photothermal effect of prolonged irradiation. Near-infrared (NIR) persistent luminescence nanoparticles (NPLNPs) possess the potential for application in next-generation PDT. However, owing to the low re-excitation efficiency of NPLNPs in deep tissue, the current PDT nanoplatform based on NPLNPs is faced with the disadvantage of decreased PDT efficiency induced by persistent luminescence (PersL) decay at the lesion site. Herein, NPLNPs, Zn1.3Ga1.4Sn0.3O4:Cr3+ (ZGS), with small particle sizes and excellent optical properties are synthesized via a simple acetylacetonate combustion method. The ZGS can be repeatedly excited by the biological window (659 nm) light to produce a strong NIR (700 nm) PersL. The response efficiency of ZGS to the wavelength in the biological window has been greatly improved by doping Sn4+ into the ZnGa2O4 matrix, which is 55 times higher than that of traditional ZnGa2O4:Cr3+. We further develop a PDT nanoplatform by modifying a photosensitizer on its surface. The PDT experiments show that the developed nanoplatform can achieve continuous and efficient tumor PDT with a depth of up to 3 cm by repeated excitation using a 659 nm LED. The NPLNPs largely solve the problem of the low re-excitation efficiency after NIR PersL decay of traditional NPLNPs in deep tissue applications and will further promote the application of NIR PLNPs in the biomedical field.

Photodynamic therapy in 2D and 3D human cervical carcinoma cell cultures employing LED light sources emitting at different wavelengths

Light of different wavelengths can be used to obtain a more profitable outcome of photodynamic therapy (PDT), according to the absorption bands of the photosensitizer (PS). Low-grade cervical intraepithelial neoplasias (CINs) are superficial lesions that can be treated with light of shorter wavelength than red because a large light penetration depth in tissue is not necessary. We report a comparative investigation performed to evaluate the efficacy of light-emitting diodes (LEDs) of different wavelengths in the photodynamic treatment applied to both 2D and 3D HeLa cell spheroid cultures. The spheroids are utilized as a PDT dosage model, and cell viability is evaluated at different sections of the spheroids by confocal microscopy. Cells incubated with m-tetrahydroxyphenyl chlorin are illuminated with LED systems working in the low fluence range, emitting in the violet (390–415 nm), blue (440–470 nm), red (620–645 nm) and deep red (640–670 nm) regions of the light spectrum at various exposures times (tI) comprised between 0.5 and 30 min. PDT experiments performed on both 2D and 3D cell cultures indicate that the PDT treatment outcome is more efficient with violet light followed by red light. Dynamic data from the front displacement velocity of large 2D-quasi-radial colonies generated from cell spheroids adhered to the Petri dish bottom as well as the evolution of the 3D growth give further insight about the effect of PDT at each condition. Results from 3D cultures indicate that the penetration of the violet light is appropriate to kill HeLa cells several layers below, showing cell damage and death not only in the outer rim of the illuminated spheroids, where a PS accumulation exists, but also in the more internal region. Results indicate that violet LED light could be useful to treat CINs involving superficial dysplasia.

Targeted photodynamic therapy treatment of in vitro A375 metastatic melanoma cells.

Metastatic Melanoma (MM) is a deadly form of skin cancer and many photodynamic therapy (PDT) studies have noted limitations in relation to effective photosensitizer (PS) drug uptake in tumors. The focus of this study was to develop a PS multicomponent nanoparticle drug conjugate carrier system which specifically targets MM cells via biomarkers to actively enhance PS delivery and so improve MM PDT. An antibody-metallated phthalocyanine-polyethylene glycol-gold nanoparticle drug conjugate, was successfully synthesized and characterized. PS active drug targeting PDT experiments at 673 nm were conducted within in vitro cultured MM. Results noted that this drug conjugate enhanced the PDT treatment of MM, through improved subcellular localization of the PS, as well as noted significantly improved cytotoxic and late apoptotic cellular death in cells. The results from this study demonstrate that through the bio-active antibody PS drug targeting of MM, the efficacy of PDT treatment for this cancer can be enhanced.

Mitochondria-localized iridium(III) complexes with anthraquinone groups as effective photosensitizers for photodynamic therapy under hypoxia

Photodynamic therapy (PDT) is a potential way for the tumor treatment. However, it notably suffers the limitation of hypoxia in solid tumors. Thus, it is significant to develop effective photosensitizers which can exhibit excellent therapeutic performance under both normoxia and hypoxia. Herein, we reported four ionic iridium(III) complexes ( Ir1 – Ir4 ) with anthraquinone groups which can regulate their excited state energy levels effectively. Among them, the energy gap of Ir1 was between 1.63 and 2.21 eV, which can match well with that of O2, and the HOMO energy of Ir1 is less than −5.51 eV. Compared with Ir2 – Ir4 , the luminescent quantum efficiency of Ir1 was the highest. Particularly, Ir1 can specifically target the mitochondria of the tumor cells. Meanwhile, Ir1 showed high singlet oxygen quantum yields ( Φ Δ) in both solutions and living cells with low cytotoxicity. The results of PDT experiments revealed that Ir1 , as a photosensitizer, exhibited excellent therapeutic effect not only in normoxia but also in hypoxia condition. We believe that this work is meaningful for developing excellent PDT agents based on cyclometalated Ir(III) complexes via rational ligand modification.

A dual-targeted theranostic photosensitizer based on a TADF fluorescein derivative

Specific diagnosis and therapy of cancer is still a challenge in biomedical research. Photodynamic therapy (PDT) has emerged as a novel therapeutic modality for cancer treatment. However, the traditional PDT photosensitizers often exhibit low specific selectivity. In this study, we have reported a dual-targeted theranostic photosensitizer FL-RGD by covalently conjugating tumor marker cyclic arginine-glycine-aspartic acid tripeptide (RGD) and a fluorescein derivative FL which has a property of thermally activated delayed fluorescence (TADF) and a long triplet lifetime for efficient PDT. The FL-RGD can target tumor tissues and further locate lysosomes of tumor cells to concurrently achieve the cancers' specific diagnosis and efficient treatment. The mechanism of its highly efficient PDT was attributed to the damage of lysosome via 1O2. Besides, FL-RGD has the potential to be utilized in depth imaging and treatment by two-photon excitation. The actual diagnosis performance of FL-RGD was proved by fluorescence imaging of living cells and tumor bearing mice. The therapy performance was proved by MTT assays, fluorescence-activated cell sorting (FACS) analysis and PDT experiments on tumor bearing mice. The research obviously exhibited the potential of FL-RGD for tumor theranostics in vivo and in vitro.

An <em>In Vitro</em> Approach to Photodynamic Therapy

Photodynamic therapy (PDT) is a medical procedure that involves incubation of an exogenously applied photosensitizer (PS) followed by visible light photoactivation to induce cell apoptosis. The Federal Drug Administration has approved PDT for the treatment of actinic keratosis, and clinical guidelines recommend PDT as a treatment for certain non-melanoma skin cancers and acne vulgaris. PDT is an advantageous therapeutic modality as it is low cost, non-invasive, and associated with minimal adverse events and scaring. In the first step of PDT, a PS is applied and allowed to accumulate intracellularly. Subsequent light irradiation induces reactive oxygen species formation, which may ultimately lead to cell apoptosis, membrane disruption, mitochondrial damage, immune modulation, keratinocyte proliferation, and collagen turnover. Herein, we present an in vitro method to study PDT in an adherent cell line. This treatment protocol is designed to simulate PDT and may be adjusted to studying the use of PDT with various cell lines, photosensitizers, incubation temperatures, or photoactivation wavelengths. Squamous cell carcinoma cells were incubated with 0, 0.5, 1.0, and 2 mM 5-aminolevulinic acid (5-ALA) for 30 min and photoactivated with 417 nm blue light for 1,000 s. The primary outcome measure was apoptosis and necrosis, as measured by annexin-V and 7-aminoactinomycin D flow cytometry. There was a dose-dependent increase in cell apoptosis following thirty-minute incubation of 5-ALA. To achieve high inter-test validity, it is important to maintain consistent incubation and light parameters when performing in vitro PDT experiments. PDT is a useful clinical procedure and in vitro research may allow for the development of novel PSs, optimization of protocols, and new indications for PDT.