Showing papers on "Protoporphyrin IX published in 2021"

Protoporphyrin IX and verteporfin potently inhibit SARS-CoV-2 infection in vitro and in a mouse model expressing human ACE2.

Abstract: The SARS-CoV-2 infection is spreading rapidly worldwide. Efficacious antiviral therapeutics against SARS-CoV-2 is urgently needed. Here, we discovered that protoporphyrin IX (PpIX) and verteporfin, two FDA-approved drugs, completely inhibited the cytopathic effect produced by SARS-CoV-2 infection at 1.25 µmol/Land 0.31 µmol/L respectively, and their EC50 values of reduction of viral RNA were at nanomolar concentrations. The selectivity indices of PpIX and verteporfin were 952.74 and 368.93, respectively, suggesting broad margin of safety. Importantly, PpIX and verteporfin prevented SARS-CoV-2 infection in mice adenovirally transduced with human ACE2. The compounds, sharing a porphyrin ring structure, were shown to bind viral receptor ACE2 and interfere with the interaction between ACE2 and the receptor-binding domain of viral S protein. Our study suggests that PpIX and verteporfin are potent antiviral agents against SARS-CoV-2 infection and sheds new light on developing novel chemoprophylaxis and chemotherapy against SARS-CoV-2.

Photophysical Properties of Protoporphyrin IX, Pyropheophorbide-a and Photofrin ® in Different Conditions.

Abstract: Photodynamic therapy (PDT) is an innovative treatment of malignant or diseased tissues The effectiveness of PDT depends on light dosimetry, oxygen availability, and properties of the photosensitizer (PS) Depending on the medium, photophysical properties of the PS can change leading to increase or decrease in fluorescence emission and formation of reactive oxygen species (ROS) especially singlet oxygen (1O2) In this study, the influence of solvent polarity, viscosity, concentration, temperature, and pH medium on the photophysical properties of protoporphyrin IX, pyropheophorbide-a, and Photofrin® were investigated by UV-visible absorption, fluorescence emission, singlet oxygen emission, and time-resolved fluorescence spectroscopies

Molecular and Metabolic Mechanisms Underlying Selective 5-Aminolevulinic Acid-Induced Fluorescence in Gliomas.

Abstract: 5-aminolevulinic acid (5-ALA) is a porphyrin precursor in the heme synthesis pathway When supplied exogenously, certain cancers consume 5-ALA and convert it to the fluorogenic metabolite protoporphyrin IX (PpIX), causing tumor-specific tissue fluorescence Preoperative administration of 5-ALA is used to aid neurosurgical resection of high-grade gliomas such as glioblastoma, allowing for increased extent of resection and progression free survival for these patients A subset of gliomas, especially low-grade tumors, do not accumulate PpIX intracellularly or readily fluoresce upon 5-ALA administration, making gross total resection difficult to achieve in diffuse lesions We review existing literature on 5-ALA metabolism and PpIX accumulation to explore potential mechanisms of 5-ALA-induced glioma tissue fluorescence Targeting the heme synthesis pathway and understanding its dysregulation in malignant tissues could aid the development of adjunct therapies to increase intraoperative fluorescence after 5-ALA treatment

Bilin-dependent regulation of chlorophyll biosynthesis by GUN4

Abstract: Biosyntheses of chlorophyll and heme in oxygenic phototrophs share a common trunk pathway that diverges with insertion of magnesium or iron into the last common intermediate, protoporphyrin IX. Since both tetrapyrroles are pro-oxidants, it is essential that their metabolism is tightly regulated. Here, we establish that heme-derived linear tetrapyrroles (bilins) function to stimulate the enzymatic activity of magnesium chelatase (MgCh) via their interaction with GENOMES UNCOUPLED 4 (GUN4) in the model green alga Chlamydomonas reinhardtii. A key tetrapyrrole-binding component of MgCh found in all oxygenic photosynthetic species, CrGUN4, also stabilizes the bilin-dependent accumulation of protoporphyrin IX-binding CrCHLH1 subunit of MgCh in light-grown C. reinhardtii cells by preventing its photooxidative inactivation. Exogenous application of biliverdin IXα reverses the loss of CrCHLH1 in the bilin-deficient heme oxygenase (hmox1) mutant, but not in the gun4 mutant. We propose that these dual regulatory roles of GUN4:bilin complexes are responsible for the retention of bilin biosynthesis in all photosynthetic eukaryotes, which sustains chlorophyll biosynthesis in an illuminated oxic environment.

Enhanced lipid metabolism induces the sensitivity of dormant cancer cells to 5-aminolevulinic acid-based photodynamic therapy.

Abstract: Cancer can develop into a recurrent metastatic disease with latency periods of years to decades. Dormant cancer cells, which represent a major cause of recurrent cancer, are relatively insensitive to most chemotherapeutic drugs and radiation. We previously demonstrated that cancer cells exhibited dormancy in a cell density-dependent manner. Dormant cancer cells exhibited increased porphyrin metabolism and sensitivity to 5-aminolevulinic acid-based photodynamic therapy (ALA-PDT). However, the metabolic changes in dormant cancer cells or the factors that enhance porphyrin metabolism have not been fully clarified. In this study, we revealed that lipid metabolism was increased in dormant cancer cells, leading to ALA-PDT sensitivity. We performed microarray analysis in non-dormant and dormant cancer cells and revealed that lipid metabolism was remarkably enhanced in dormant cancer cells. In addition, triacsin C, a potent inhibitor of acyl-CoA synthetases (ACSs), reduced protoporphyrin IX (PpIX) accumulation and decreased ALA-PDT sensitivity. We demonstrated that lipid metabolism including ACS expression was positively associated with PpIX accumulation. This research suggested that the enhancement of lipid metabolism in cancer cells induces PpIX accumulation and ALA-PDT sensitivity.

Dissolving microneedles containing aminolevulinic acid improves protoporphyrin IX distribution

Abstract: One important limitation of topical photodynamic therapy (PDT) is the limited tissue penetration of precursors. Microneedles (MNs) are minimally invasive devices used to promote intradermal drug delivery. Dissolving MNs contain drug-associated to polymer blends, dissolving after insertion into skin, allowing drug release. This study comprises development and characterization of a pyramidal model of dissolving MNs (500 μm) prepared with 5% wt/wt aminolevulinic acid and 20% wt/wt Gantrez AN-139 in aqueous blend. Protoporphyrin IX formation and distribution were evaluated in tumor mice model by using fluorescence widefield imaging, spectroscopy, and confocal microscopy. MNs demonstrated excellent mechanical resistance penetrating about 250 μm with minor size alteration in vitro, and fluorescence intensity was 5-times higher at 0.5 mm on average compared to cream in vivo (being 10 ± 5 a.u. for MNs and 2.4 ± 0.8 a.u. for cream). Dissolving MNs have overcome topical cream application, being extremely promising especially for thicker skin lesions treatment using PDT.

Fluorescence Lifetime Imaging and Spectroscopic Co-Validation for Protoporphyrin IX-Guided Tumor Visualization in Neurosurgery.

Abstract: Maximal safe resection is a key strategy for improving patient prognosis in the management of brain tumors. Intraoperative fluorescence guidance has emerged as a standard in the surgery of high-grade gliomas. The administration of 5-aminolevulinic acid prior to surgery induces tumor-specific accumulation of protoporphyrin IX, which emits red fluorescence under blue-light illumination. The technology, however, is substantially limited for low-grade gliomas and weakly tumor-infiltrated brain, where low protoporphyrin IX concentrations are outweighed by tissue autofluorescence. In this context, fluorescence lifetime imaging has shown promise to distinguish spectrally overlapping fluorophores. We integrated frequency-domain fluorescence lifetime imaging in a surgical microscope and combined it with spatially registered fluorescence spectroscopy, which can be considered a research benchmark for sensitive protoporphyrin IX detection. Fluorescence lifetime maps and spectra were acquired for a representative set of fresh ex-vivo brain tumor specimens (low-grade gliomas n = 15, high-grade gliomas n = 80, meningiomas n = 41, and metastases n = 35). Combining the fluorescence lifetime with fluorescence spectra unveiled how weak protoporphyrin IX accumulations increased the lifetime respective to tissue autofluorescence. Infiltration zones (4.1ns ± 1.8ns, p = 0.017) and core tumor areas (4.8ns ± 1.3ns, p = 0.040) of low-grade gliomas were significantly distinguishable from non-pathologic tissue (1.6ns ± 0.5ns). Similarly, fluorescence lifetimes for infiltrated and reactive tissue as well as necrotic and core tumor areas were increased for high-grade gliomas and metastasis. Meningioma tumor specimens showed strongly increased lifetimes (12.2ns ± 2.5ns, p = 0.005). Our results emphasize the potential of fluorescence lifetime imaging to optimize maximal safe resection in brain tumors in future and highlight its potential toward clinical translation.

Key transporters leading to specific protoporphyrin IX accumulation in cancer cell following administration of aminolevulinic acid in photodynamic therapy/diagnosis.

Abstract: The administration of aminolevulinic acid allow the formation and accumulation of protoporphyrin IX specifically in cancer cells, which then lead to photocytotoxicity following light irradiation. This compound, when accumulated at high levels, could also be used in cancer diagnosis as it would emit red fluorescence when being light irradiated. The concentration of protoporphyrin IX is pivotal in ensuring the effectiveness of the therapy. Studies have been carried out and showed the importance of various transporters in regulating the amount of these substrates by controlling the transport of various related metabolites in and out of the cell. There are many transporters involved and their expression levels are dependent on various factors, such as oxygen availability and iron ions. It is also important to note that these transporters may also have different expression levels depending on their organ. Understanding the mechanisms and the roles of these transporters are essential to ensure maximum accumulation of protoporphyrin IX, leading to higher efficiency in photodynamic therapy/diagnosis. In this review, we would like to discuss the roles of various transporters in protoporphyrin IX accumulation and how their involvement directly affect cancerous microenvironment.

Stimulation and homogenization of the protoporphyrin IX endogenous production by photobiomodulation to increase the potency of photodynamic therapy.

Abstract: Protoporphyrin IX (PpIX) is produced in the mitochondria and used as fluorescent contrast agent or photosensitizer after exogenous 5-aminolevulinic acid (ALA) delivery in cancer photodynamic detection and therapy (PDT). Although routinely used in the clinics, the stimulated production of PpIX is often insufficient and/or heterogeneous within the lesions, thereby limiting the PDT performances. Since photobiomodulation, which is based on the illumination of the tissues with sub-thermal radiometric conditions in the red or near-infrared, is known to stimulate the cell metabolism, we have optimized these conditions in vitro. Some of them lead to the homogenization and strong stimulation of the PpIX endogenous production. Interestingly, combined sequentially, PBM enhanced significantly the potency of PpIX-based PDT in vitro and in vivo in tumors grown on the chicken embryo chorioallantoic membrane. These results are in excellent agreement with other assays based on measurements of the cell survival/death, the production of reactive oxygen species, including singlet oxygen, and the mitochondrial membrane potential.

Photodynamic therapy of cutaneous T-cell lymphoma cell lines mediated by 5-aminolevulinic acid and derivatives.

Abstract: The delta-amino acid 5-aminolevulinic acid (ALA), is the precursor of the endogenous photosensitiser Protoporphyrin IX (PpIX), and is currently approved for Photodynamic Therapy (PDT) of certain superficial cancers. However, ALA-PDT is not very effective in diseases in which T-cells play a significant role. Cutaneous T-cell lymphomas (CTCL) is a group of non-Hodgkin malignant diseases, which includes mycosis fungoides (MF) and Sezary syndrome (SS). In previous work, we have designed new ALA esters synthesised by three-component Passerini reactions, and some of them showed higher performance as compared to ALA. This work aimed to determine the efficacy as pro-photosensitisers of five new ALA esters of 2-hydroxy-N-arylacetamides (1f, 1 g, 1 h, 1i and 1 k) of higher lipophilicity than ALA in Myla cells of MF and HuT-78 cells of SS. We have also tested its effectiveness against ALA and the already marketed ALA methyl ester (Me-ALA) and ALA hexyl ester (He-ALA). Both cell Myla and SS cells were effectively and equally photoinactivated by ALA-PDT. Besides, the concentration of ALA required to induce half the maximal porphyrin synthesis was 209 μM for Myla and 169 μM for HuT-78 cells. As a criterion of efficacy, we calculated the concentration of the ALA derivatives necessary to induce half the plateau porphyrin values obtained from ALA. These values were achieved at concentrations 4 and 12 times lower compared to ALA, according to the derivative used. For He-ALA, concentrations were 24 to 25 times lower than required for ALA for inducing comparable porphyrin synthesis in both CTCL cells. The light doses for inducing 50% of cell death (LD50) for He-ALA, 1f, 1 g, 1 h and 1i were around 18 and 25 J/cm2 for Myla and HuT-78 cells respectively, after exposure to 0.05 mM concentrations of the compounds. On the other hand, the LD50s for the compound 1 k were 40 and 57 J/cm2 for Myla and HuT-78, respectively. In contrast, 0.05 mM of ALA and Me-ALA did not provoke photokilling since the concentration employed was far below the porphyrin saturation point for these compounds. Our results suggest the potential use of ALA derivatives for topical application in PDT treatment of MF and extracorporeal PDT for the depletion of activated T-cells in SS.

Photodynamic and Sonodynamic Therapy with Protoporphyrin IX: In Vitro and In Vivo Studies.

Abstract: Sono-photodynamic therapy is a promising anticancer technique based on the combination of sonodynamic and photodynamic therapy to improve the cancer treatment effectiveness. This study was aimed at analyzing the effects of the sono-photodynamic (SPD) activity on protoporphyrin IX (PpIX) solution and PpIX-loaded rat liver. In vitro, PpIX 5 μM solutions were irradiated with light (635 nm, 30-50 mW/cm2), ultrasound (1 MHz, 1-2 W/cm2) and both. The PpIX absorption spectra recorded over exposure time revealed that the PpIX decay rate induced by SPD activity (combined irradiation) was approximately the sum of those induced by photodynamic and sonodynamic activity. In vivo, rats were intraperitoneally injected with 5-aminolevulinic acid at the dose of 500 mg/kg weight. After 3 h of injection, the PpIX-loaded livers were irradiated with light (635 nm, 180 ± 9 J/cm2), ultrasound (1.0 MHz, 770 ± 40 J/cm2) and both using a single probe capable of illuminating and sonicating the liver simultaneously. After 30 h, the liver damage induced by each protocol was analyzed histologically. It was found that a greater necrosis depth was induced by the SPD activity. These results suggest that the SPD activity could improve the PpIX decay rate and have greater scope than photodynamic or sonodynamic activity. Further studies should be performed to gain a better understanding of this protocol.

Biochemistry, Heme Synthesis

Abstract: Heme is a porphyrin ring complexed with ferrous iron and protoporphyrin IX. Heme is an essential prosthetic group in proteins that is necessary as a subcellular compartment to perform diverse biological functions like hemoglobin and myoglobin. [1] Other enzymes which use heme as a prosthetic group includes cytochromes of the electron transport chain, catalase, and nitric oxide synthase. The major tissues for heme synthesis are bone marrow by erythrocytes and the liver by hepatocytes.

Characterization of autofluorescence and quantitative protoporphyrin IX biomarkers for optical spectroscopy-guided glioma surgery.

Abstract: 5-Aminolevulinic acid (5-ALA)-mediated fluorescence does not effectively depict low grade gliomas (LGG) or the infiltrative tumor portion of high-grade gliomas (HGG). While spectroscopy improves sensitivity and precision, this is currently limited by autofluorescence and a second protoporphyrin IX (PpIX) fluorescence state at 620 nm. We investigated the autofluorescence to better characterize the present spectra and thus increase PpIX quantification precision and sensitivity. This study included 128 patients undergoing surgery for malignant glioma. 5-ALA (Gliolan) was administered before anesthesia, and fluorescence was measured using a hyperspectral device. It was found that all 2692 measured spectra consisted of contributions from 620 to 634 nm PpIX, NADH, lipofuscin, and flavins. The basis spectra were characterized and their use in spectral unmixing led to 82.4% lower fitting error for weakly fluorescing areas (p < 0.001), and 92.3% fewer false positive tumor identifications in control measurements (p = 0.0065) compared to previous works. They also decreased the PpIX620 contribution, thus halving the mean Ratio620/634 (p < 0.001). The ratio was approximately 0 for HGGs and increasing for LGGs, as demonstrated previously. Additionally, the Ratio620/634, the MIB-1/Ki-67 proliferation index, and the PpIX peak blue-shift were found to be significantly related to WHO grade, fluorescence visibility, and PpIX contribution (p < 0.001), and the value of these three as quantitative biomarkers is discussed.

How Much Protoporphyrin IX Must Be Activated to Obtain Full Efficacy of Methyl Aminolevulinate Photodynamic Therapy? Implication for Treatment Modifications

Abstract: Photodynamic therapy (PDT) with methyl aminolevulinate (MAL) is a popular treatment for actinic keratoses (AK), and several PDT treatment modalities with similar cure rates are in use. The effect relies on the activation of protoporphyrin IX (PpIX) in premalignant cells. This study aimed to measure PpIX during each treatment modality to determine the minimal PpIX activation and shortest exposure time for optimal cure rate. In four different treatment modalities, we established the PpIX formation up to three hours after MAL application without illumination and measured the speed of PpIX photoactivation during 9 min of red light (37 J/cm2). The level of PpIX three hours after MAL application was set to 100 PpIX units. In comparison, 85 PpIX units were formed during daylight PDT, 57 PpIX units during pulse PDT, and 52 PpIX units without any curettage prior to MAL. The activation of 50 PpIX units should, therefore, be enough to obtain a full effect on AK. Further, red light illumination may be shortened from 9 min to 1–2 min. The results indicate that PDT can be performed successfully with half the illumination time used in daylight PDT today and with one fourth of the illumination time used in classical PDT.

Light-Activated Protoporphyrin IX-Based Polysilsesquioxane Nanoparticles Induce Ferroptosis in Melanoma Cells.

Abstract: The use of nanoparticle-based materials to improve the efficacy of photodynamic therapy (PDT) to treat cancer has been a burgeoning field of research in recent years. Polysilsesquioxane (PSilQ) nanoparticles with remarkable features, such as high loading of photosensitizers, biodegradability, surface tunability, and biocompatibility, have been used for the treatment of cancer in vitro and in vivo using PDT. The PSilQ platform typically shows an enhanced PDT performance following a cell death mechanism similar to the parent photosensitizer. Ferroptosis is a new cell death mechanism recently associated with PDT that has not been investigated using PSilQ nanoparticles. Herein, we synthesized a protoporphyrin IX (PpIX)-based PSilQ platform (PpIX-PSilQ NPs) to study the cell death pathways, with special focus on ferroptosis, during PDT in vitro. Our data obtained from different assays that analyzed Annexin V binding, glutathione peroxidase activity, and lipid peroxidation demonstrate that the cell death in PDT using PpIX-PSilQ NPs is regulated by apoptosis and ferroptosis. These results can provide alternative approaches in designing PDT strategies to enhance therapeutic response in conditions stymied by apoptosis resistance.

Influence of dexamethasone on visible 5-ALA fluorescence and quantitative protoporphyrin IX accumulation measured by fluorescence lifetime imaging in glioblastomas: is pretreatment obligatory before fluorescence-guided surgery?

Abstract: Objective Fluorescence-guided surgery using 5-aminolevulinic acid (5-ALA) is nowadays widely applied for improved resection of glioblastomas (GBMs). Initially, pretreatment with dexamethasone was considered to be essential for optimal fluorescence effect. However, recent studies reported comparably high rates of visible fluorescence in GBMs despite absence of dexamethasone pretreatment. Recently, the authors proposed fluorescence lifetime imaging (FLIM) for the quantitative analysis of 5-ALA-induced protoporphyrin IX (PpIX) accumulation. The aim of this study was thus to investigate the influence of dexamethasone on visible fluorescence and quantitative PpIX accumulation. Methods The authors prospectively analyzed the presence of visible fluorescence during surgery in a cohort of patients with GBMs. In this study, patients received dexamethasone preoperatively only if clinically indicated. One representative tumor sample was collected from each GBM, and PpIX accumulation was analyzed ex vivo by FLIM. The visible fluorescence status and mean FLIM values were correlated with preoperative intake of dexamethasone. Results In total, two subgroups with (n = 27) and without (n = 20) pretreatment with dexamethasone were analyzed. All patients showed visible fluorescence independent from preoperative dexamethasone intake. Furthermore, the authors did not find a statistically significant difference in the mean FLIM values between patients with and without dexamethasone pretreatment (p = 0.097). Conclusions In this first study to date, the authors found no significant influence of dexamethasone pretreatment on either visible 5-ALA fluorescence during GBM surgery or PpIX accumulation based on FLIM. According to these preliminary data, the authors recommend administering dexamethasone prior to fluorescence-guided surgery of GBMs only when clinically indicated.