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Obaid G, Celli JP, Broekgaarden M, Bulin AL, Uusimaa P, Pogue B, Hasan T, Huang HC. Engineering photodynamics for treatment, priming and imaging. NATURE REVIEWS BIOENGINEERING 2024; 2:752-769. [PMID: 39927170 PMCID: PMC11801064 DOI: 10.1038/s44222-024-00196-z] [Citation(s) in RCA: 11] [Impact Index Per Article: 11.0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Accepted: 05/07/2024] [Indexed: 02/11/2025]
Abstract
Photodynamic therapy (PDT) is a photochemistry-based treatment approach that relies on the activation of photosensitizers by light to locally generate reactive oxygen species that induce cellular cytotoxicity, in particular for the treatment of tumours. The cytotoxic effects of PDT are depth-limited owing to light penetration limits in tissue. However, photodynamic priming (PDP), which inherently occurs during PDT, can prime the tissue microenvironment to adjuvant therapies beyond the direct PDT ablative zone. In this Review, we discuss the underlying mechanisms of PDT and PDP, and their application to the treatment of cancer, outlining how PDP can permeabilize the tumour vasculature, overcome biological barriers, modulate multidrug resistance, enhance immune responses, increase tumour permeability and enable the photochemical release of drugs. We further examine the molecular engineering of photosensitizers to improve their pharmacodynamic and pharmacokinetic properties, increase their molecular specificity and allow image guidance of PDT, and investigate engineered cellular models for the design and optimization of PDT and PDP. Finally, we discuss alternative activation sources, including ultrasound, X-rays and self-illuminating compounds, and outline key barriers to the clinical translation of PDT and PDP.
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Affiliation(s)
- Girgis Obaid
- Department of Bioengineering, University of Texas at Dallas, Richardson, TX, USA
| | - Jonathan P. Celli
- Department of Physics, University of Massachusetts Boston, Boston, MA, USA
| | - Mans Broekgaarden
- Grenoble Alpes University, INSERM U1209, CNRS UMR5309, Institute for Advanced Biosciences, Grenoble, France
| | - Anne-Laure Bulin
- Grenoble Alpes University, INSERM U1209, CNRS UMR5309, Institute for Advanced Biosciences, Grenoble, France
| | | | - Brian Pogue
- Department of Medical Physics, University of Wisconsin School of Medicine, Madison, WI, USA
| | - Tayyaba Hasan
- Wellman Center for Photomedicine, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA
| | - Huang-Chiao Huang
- Fischell Department of Bioengineering, University of Maryland, College Park, MD, USA
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Anti-Hypoxia Nanoplatforms for Enhanced Photosensitizer Uptake and Photodynamic Therapy Effects in Cancer Cells. Int J Mol Sci 2023; 24:ijms24032656. [PMID: 36768975 PMCID: PMC9916860 DOI: 10.3390/ijms24032656] [Citation(s) in RCA: 7] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/04/2022] [Revised: 01/18/2023] [Accepted: 01/25/2023] [Indexed: 02/01/2023] Open
Abstract
Photodynamic therapy (PDT) holds great promise in cancer eradication due to its target selectivity, non-invasiveness, and low systemic toxicity. However, due to the hypoxic nature of many native tumors, PDT is frequently limited in its therapeutic effect. Additionally, oxygen consumption during PDT may exacerbate the tumor's hypoxic condition, which stimulates tumor proliferation, metastasis, and invasion, resulting in poor treatment outcomes. Therefore, various strategies have been developed to combat hypoxia in PDT, such as oxygen carriers, reactive oxygen supplements, and the modulation of tumor microenvironments. However, most PDT-related studies are still conducted on two-dimensional (2D) cell cultures, which fail to accurately reflect tissue complexity. Thus, three-dimensional (3D) cell cultures are ideal models for drug screening, disease simulation and targeted cancer therapy, since they accurately replicate the tumor tissue architecture and microenvironment. This review summarizes recent advances in the development of strategies to overcome tumor hypoxia for enhanced PDT efficiency, with a particular focus on nanoparticle-based photosensitizer (PS) delivery systems, as well as the advantages of 3D cell cultures.
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Xie J, Wang Y, Choi W, Jangili P, Ge Y, Xu Y, Kang J, Liu L, Zhang B, Xie Z, He J, Xie N, Nie G, Zhang H, Kim JS. Overcoming barriers in photodynamic therapy harnessing nano-formulation strategies. Chem Soc Rev 2021; 50:9152-9201. [PMID: 34223847 DOI: 10.1039/d0cs01370f] [Citation(s) in RCA: 265] [Impact Index Per Article: 66.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
Photodynamic therapy (PDT) has been extensively investigated for decades for tumor treatment because of its non-invasiveness, spatiotemporal selectivity, lower side-effects, and immune activation ability. It can be a promising treatment modality in several medical fields, including oncology, immunology, urology, dermatology, ophthalmology, cardiology, pneumology, and dentistry. Nevertheless, the clinical application of PDT is largely restricted by the drawbacks of traditional photosensitizers, limited tissue penetrability of light, inefficient induction of tumor cell death, tumor resistance to the therapy, and the severe pain induced by the therapy. Recently, various photosensitizer formulations and therapy strategies have been developed to overcome these barriers. Significantly, the introduction of nanomaterials in PDT, as carriers or photosensitizers, may overcome the drawbacks of traditional photosensitizers. Based on this, nanocomposites excited by various light sources are applied in the PDT of deep-seated tumors. Modulation of cell death pathways with co-delivered reagents promotes PDT induced tumor cell death. Relief of tumor resistance to PDT with combined therapy strategies further promotes tumor inhibition. Also, the optimization of photosensitizer formulations and therapy procedures reduces pain in PDT. Here, a systematic summary of recent advances in the fabrication of photosensitizers and the design of therapy strategies to overcome barriers in PDT is presented. Several aspects important for the clinical application of PDT in cancer treatment are also discussed.
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Affiliation(s)
- Jianlei Xie
- Key Laboratory of Optoelectronic Devices and Systems of Ministry of Education and Guangdong Province, Institute of Microscale Optoelectronics, and Otolaryngology Department and Biobank of the First Affiliated Hospital, Shenzhen Second People's Hospital, Health Science Center, Shenzhen University, Shenzhen 518060, P. R. China.
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Gunaydin G, Gedik ME, Ayan S. Photodynamic Therapy-Current Limitations and Novel Approaches. Front Chem 2021; 9:691697. [PMID: 34178948 PMCID: PMC8223074 DOI: 10.3389/fchem.2021.691697] [Citation(s) in RCA: 291] [Impact Index Per Article: 72.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/08/2021] [Accepted: 05/14/2021] [Indexed: 12/17/2022] Open
Abstract
Photodynamic therapy (PDT) mostly relies on the generation of singlet oxygen, via the excitation of a photosensitizer, so that target tumor cells can be destroyed. PDT can be applied in the settings of several malignant diseases. In fact, the earliest preclinical applications date back to 1900’s. Dougherty reported the treatment of skin tumors by PDT in 1978. Several further studies around 1980 demonstrated the effectiveness of PDT. Thus, the technique has attracted the attention of numerous researchers since then. Hematoporphyrin derivative received the FDA approval as a clinical application of PDT in 1995. We have indeed witnessed a considerable progress in the field over the last century. Given the fact that PDT has a favorable adverse event profile and can enhance anti-tumor immune responses as well as demonstrating minimally invasive characteristics, it is disappointing that PDT is not broadly utilized in the clinical setting for the treatment of malignant and/or non-malignant diseases. Several issues still hinder the development of PDT, such as those related with light, tissue oxygenation and inherent properties of the photosensitizers. Various photosensitizers have been designed/synthesized in order to overcome the limitations. In this Review, we provide a general overview of the mechanisms of action in terms of PDT in cancer, including the effects on immune system and vasculature as well as mechanisms related with tumor cell destruction. We will also briefly mention the application of PDT for non-malignant diseases. The current limitations of PDT utilization in cancer will be reviewed, since identifying problems associated with design/synthesis of photosensitizers as well as application of light and tissue oxygenation might pave the way for more effective PDT approaches. Furthermore, novel promising approaches to improve outcome in PDT such as selectivity, bioengineering, subcellular/organelle targeting, etc. will also be discussed in detail, since the potential of pioneering and exceptional approaches that aim to overcome the limitations and reveal the full potential of PDT in terms of clinical translation are undoubtedly exciting. A better understanding of novel concepts in the field (e.g. enhanced, two-stage, fractional PDT) will most likely prove to be very useful for pursuing and improving effective PDT strategies.
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Affiliation(s)
- Gurcan Gunaydin
- Department of Basic Oncology, Hacettepe University Cancer Institute, Sihhiye, Ankara, Turkey
| | - M Emre Gedik
- Department of Basic Oncology, Hacettepe University Cancer Institute, Sihhiye, Ankara, Turkey
| | - Seylan Ayan
- Department of Chemistry, Bilkent University, Ankara, Turkey
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Pola M, Kolarova H, Ruzicka J, Zholobenko A, Modriansky M, Mosinger J, Bajgar R. Effects of zinc porphyrin and zinc phthalocyanine derivatives in photodynamic anticancer therapy under different partial pressures of oxygen in vitro. Invest New Drugs 2020; 39:89-97. [PMID: 32833137 DOI: 10.1007/s10637-020-00990-7] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/03/2020] [Accepted: 08/18/2020] [Indexed: 01/05/2023]
Abstract
Photodynamic therapy (PDT) is gradually becoming an alternative method in the treatment of several diseases. Here, we investigated the role of oxygen in photodynamically treated cervical cancer cells (HeLa). The effect of PDT on HeLa cells was assessed by exposing cultured cells to disulphonated zinc phthalocyanine (ZnPcS2) and tetrasulphonated zinc tetraphenylporphyrin (ZnTPPS4). Fluorescence microscopy revealed their different localizations within the cells. ZnTPPS4 seems to be mostly limited to the cytosol and lysosomes, whereas ZnPcS2 is most likely predominantly attached to membrane structures, including plasmalemma and the mitochondrial membrane. Phototoxicity assays of PDT-treated cells carried out under different partial pressures of oxygen showed dose-dependent responses. Interestingly, ZnPcS2 was also photodynamically effective at a minimal level of oxygen, under a nitrogen atmosphere. On the other hand, hyperbaric oxygenation did not lead to a higher PDT efficiency of either photosensitizer. Although both photosensitizers can induce a significant drop in mitochondrial membrane potential, ZnPcS2 has a markedly higher effect on mitochondrial respiration that was completely blocked after two short light cycles. In conclusion, our observations suggest that PDT can be effective even in hypoxic conditions if a suitable sensitizer is chosen, such as ZnPcS2, which can inhibit mitochondrial respiration.
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Affiliation(s)
- Martin Pola
- Department of Medical Biophysics, Faculty of Medicine and Dentistry, Palacky University in Olomouc, Hnevotinska 3, 775 15, Olomouc, Czech Republic
| | - Hana Kolarova
- Department of Medical Biophysics, Faculty of Medicine and Dentistry, Palacky University in Olomouc, Hnevotinska 3, 775 15, Olomouc, Czech Republic
| | - Jiri Ruzicka
- Department of Medical Biophysics, Faculty of Medicine and Dentistry, Palacky University in Olomouc, Hnevotinska 3, 775 15, Olomouc, Czech Republic
| | - Aleksey Zholobenko
- Department of Medical Chemistry and Biochemistry, Faculty of Medicine and Dentistry, Palacky University in Olomouc, Hnevotinska 3, 775 15, Olomouc, Czech Republic
| | - Martin Modriansky
- Department of Medical Chemistry and Biochemistry, Faculty of Medicine and Dentistry, Palacky University in Olomouc, Hnevotinska 3, 775 15, Olomouc, Czech Republic
| | - Jiri Mosinger
- Department of Inorganic Chemistry, Faculty of Science, Charles University, Hlavova 2030, 128 43, Prague 2, Czech Republic.,Institute of Inorganic Chemistry of the Czech Academy of Sciences, v.v.i., Husinec-Rez 1001, 250 68, Rez, Czech Republic
| | - Robert Bajgar
- Department of Medical Biophysics, Faculty of Medicine and Dentistry, Palacky University in Olomouc, Hnevotinska 3, 775 15, Olomouc, Czech Republic. .,Institute of Molecular and Translational Medicine, Faculty of Medicine and Dentistry, Palacky University in Olomouc, Hnevotinska 3, 775 15, Olomouc, Czech Republic.
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Falk-Mahapatra R, Gollnick SO. Photodynamic Therapy and Immunity: An Update. Photochem Photobiol 2020; 96:550-559. [PMID: 32128821 DOI: 10.1111/php.13253] [Citation(s) in RCA: 115] [Impact Index Per Article: 23.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/15/2019] [Accepted: 02/04/2020] [Indexed: 12/18/2022]
Abstract
Dr. Thomas Dougherty and his Oncology Foundation of Buffalo were the first to support my (S.O.G.) research into the effects of photodynamic therapy (PDT) on the host immune system. The small grant I was awarded in 2002 launched my career as an independent researcher; at the time, there were few studies on the importance of the immune response on the efficacy of PDT and no studies demonstrating the ability of PDT to enhance antitumor immunity. Over the last decades, the interest in PDT as an enhancer of antitumor immunity and our understanding of the mechanisms by which PDT enhances antitumor immunity have dramatically increased. In this review article, we look back on the studies that laid the foundation for our understanding and provide an update on current advances and therapies that take advantage of PDT enhancement of immunity.
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Affiliation(s)
| | - Sandra O Gollnick
- Department of Immunology, Roswell Park Comprehensive Cancer Center, Buffalo, NY.,Department of Cell Stress Biology, Roswell Park Comprehensive Cancer Center, Buffalo, NY
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Liu CG, Han YH, Kankala RK, Wang SB, Chen AZ. Subcellular Performance of Nanoparticles in Cancer Therapy. Int J Nanomedicine 2020; 15:675-704. [PMID: 32103936 PMCID: PMC7008395 DOI: 10.2147/ijn.s226186] [Citation(s) in RCA: 88] [Impact Index Per Article: 17.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/06/2019] [Accepted: 12/16/2019] [Indexed: 12/12/2022] Open
Abstract
With the advent of nanotechnology, various modes of traditional treatment strategies have been transformed extensively owing to the advantageous morphological, physiochemical, and functional attributes of nano-sized materials, which are of particular interest in diverse biomedical applications, such as diagnostics, sensing, imaging, and drug delivery. Despite their success in delivering therapeutic agents, several traditional nanocarriers often end up with deprived selectivity and undesired therapeutic outcome, which significantly limit their clinical applicability. Further advancements in terms of improved selectivity to exhibit desired therapeutic outcome toward ablating cancer cells have been predominantly made focusing on the precise entry of nanoparticles into tumor cells via targeting ligands, and subsequent delivery of therapeutic cargo in response to specific biological or external stimuli. However, there is enough room intracellularly, where diverse small-sized nanomaterials can accumulate and significantly exert potentially specific mechanisms of antitumor effects toward activation of precise cancer cell death pathways that can be explored. In this review, we aim to summarize the intracellular pathways of nanoparticles, highlighting the principles and state of their destructive effects in the subcellular structures as well as the current limitations of conventional therapeutic approaches. Next, we give an overview of subcellular performances and the fate of internalized nanoparticles under various organelle circumstances, particularly endosome or lysosome, mitochondria, nucleus, endoplasmic reticulum, and Golgi apparatus, by comprehensively emphasizing the unique mechanisms with a series of interesting reports. Moreover, intracellular transformation of the internalized nanoparticles, prominent outcome and potential affluence of these interdependent subcellular components in cancer therapy are emphasized. Finally, we conclude with perspectives with a focus on the contemporary challenges in their clinical applicability.
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Affiliation(s)
- Chen-Guang Liu
- Institute of Biomaterials and Tissue Engineering, Huaqiao University, Xiamen, Fujian361021, People’s Republic of China
- College of Chemical Engineering, Huaqiao University, Xiamen, Fujian361021, People’s Republic of China
| | - Ya-Hui Han
- Institute of Biomaterials and Tissue Engineering, Huaqiao University, Xiamen, Fujian361021, People’s Republic of China
- College of Chemical Engineering, Huaqiao University, Xiamen, Fujian361021, People’s Republic of China
| | - Ranjith Kumar Kankala
- Institute of Biomaterials and Tissue Engineering, Huaqiao University, Xiamen, Fujian361021, People’s Republic of China
- College of Chemical Engineering, Huaqiao University, Xiamen, Fujian361021, People’s Republic of China
- Fujian Provincial Key Laboratory of Biochemical Technology (Huaqiao University), Xiamen, Fujian361021, People’s Republic of China
| | - Shi-Bin Wang
- Institute of Biomaterials and Tissue Engineering, Huaqiao University, Xiamen, Fujian361021, People’s Republic of China
- College of Chemical Engineering, Huaqiao University, Xiamen, Fujian361021, People’s Republic of China
- Fujian Provincial Key Laboratory of Biochemical Technology (Huaqiao University), Xiamen, Fujian361021, People’s Republic of China
| | - Ai-Zheng Chen
- Institute of Biomaterials and Tissue Engineering, Huaqiao University, Xiamen, Fujian361021, People’s Republic of China
- College of Chemical Engineering, Huaqiao University, Xiamen, Fujian361021, People’s Republic of China
- Fujian Provincial Key Laboratory of Biochemical Technology (Huaqiao University), Xiamen, Fujian361021, People’s Republic of China
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Qi S, Guo L, Yan S, Lee RJ, Yu S, Chen S. Hypocrellin A-based photodynamic action induces apoptosis in A549 cells through ROS-mediated mitochondrial signaling pathway. Acta Pharm Sin B 2019; 9:279-293. [PMID: 30972277 PMCID: PMC6437636 DOI: 10.1016/j.apsb.2018.12.004] [Citation(s) in RCA: 77] [Impact Index Per Article: 12.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/17/2018] [Revised: 11/15/2018] [Accepted: 11/28/2018] [Indexed: 12/31/2022] Open
Abstract
Over recent decades, many studies have reported that hypocrellin A (HA) can eliminate cancer cells with proper irradiation in several cancer cell lines. However, the precise molecular mechanism underlying its anticancer effect has not been fully defined. HA-mediated cytotoxicity and apoptosis in human lung adenocarcinoma A549 cells were evaluated after photodynamic therapy (PDT). A temporal quantitative proteomics approach by isobaric tag for relative and absolute quantitation (iTRAQ) 2D liquid chromatography with tandem mass spectrometric (LC–MS/MS) was introduced to help clarify molecular cytotoxic mechanisms and identify candidate targets of HA-induced apoptotic cell death. Specific caspase inhibitors were used to further elucidate the molecular pathway underlying apoptosis in PDT-treated A549 cells. Finally, down-stream apoptosis-related protein was evaluated. Apoptosis induced by HA was associated with cell shrinkage, externalization of cell membrane phosphatidylserine, DNA fragmentation, and mitochondrial disruption, which were preceded by increased intracellular reactive oxygen species (ROS) generations. Further studies showed that PDT treatment with 0.08 µmol/L HA resulted in mitochondrial disruption, pronounced release of cytochrome c, and activation of caspase-3, -9, and -7. Together, HA may be a possible therapeutic agent directed toward mitochondria and a promising photodynamic anticancer candidate for further evaluation.
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Key Words
- ACN, acetonitrile
- CLSM, confocal laser scanning confocal microscopy
- DCFH-DA, 2′,7′-dichlorofuorescin diacetate
- DMEM, Dulbecco׳s modified Eagle׳s medium
- Dox, doxorubicin
- ECL, enhanced chemiluminescence
- FCCP, carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone
- FDR, false discovery rate
- GO, gene ontology
- HA, hypocrellin A
- HRP, horseradish peroxidase
- Hypocrellin A
- IAA, iodoacetamide
- IKK, IκB kinase complex
- JC-1, 5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethyl-benzimidazolcarbocyanine iodide
- LC–MS/MS
- MMP, mitochondrial membrane potential
- MPT, mitochondrial permeability transition
- NAC, N-acetyl-l-cysteine
- OCR, oxygen consumption rate
- PDT, photodynamic therapy
- PI, propidium iodide
- PS, photosensitizer
- Photodynamic therapy
- Proteomic
- ROS, reactive oxygen species
- Reactive oxygen species
- SCX, strong cation exchange
- TCM, traditional Chinese medicinal
- TEM, transmission electron microscope
- TFA, trifluoroacetic acid
- UA, urea
- iTRAQ
- iTRAQ, isobaric tag for relative and absolute quantitation
- z-IETD-fmk, z-Ile-Glu-Asp-fluoromethylketone
- z-LEHD-fmk, z-Leu-Glu(OMe)-His-Asp(OMe)-fluoromethylketone
- z-VAD-fmk, z-Val-Ala-Asp-fluoromethylketone
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Affiliation(s)
- Shanshan Qi
- Jiangsu Province Key Laboratory for Microbes and Functional Genomics, College of Life Sciences, Nanjing Normal University, Nanjing 210023, China
- College of Pharmacy, the Ohio State University, Columbus, OH 43210, USA
| | - Lingyuan Guo
- Jiangsu Province Key Laboratory for Microbes and Functional Genomics, College of Life Sciences, Nanjing Normal University, Nanjing 210023, China
| | - Shuzhen Yan
- Jiangsu Province Key Laboratory for Microbes and Functional Genomics, College of Life Sciences, Nanjing Normal University, Nanjing 210023, China
| | - Robert J. Lee
- College of Pharmacy, the Ohio State University, Columbus, OH 43210, USA
| | - Shuqin Yu
- Jiangsu Province Key Laboratory for Molecular and Medical Biotechnology, College of Life Sciences, Nanjing Normal University, Nanjing 210023, China
- Corresponding author. Tel./fax: +86 25 8559 1050.
| | - Shuanglin Chen
- Jiangsu Province Key Laboratory for Microbes and Functional Genomics, College of Life Sciences, Nanjing Normal University, Nanjing 210023, China
- Corresponding author. Tel.: +86 25 8589 1265.
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Kessel D, Oleinick NL. Cell Death Pathways Associated with Photodynamic Therapy: An Update. Photochem Photobiol 2018; 94:213-218. [PMID: 29143339 DOI: 10.1111/php.12857] [Citation(s) in RCA: 149] [Impact Index Per Article: 21.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/07/2017] [Accepted: 10/18/2017] [Indexed: 12/13/2022]
Abstract
Photodynamic therapy (PDT) has the potential to make a significant impact on cancer treatment. PDT can sensitize malignant tissues to light, leading to a highly selective effect if an appropriate light dose can be delivered. Variations in light distribution and drug delivery, along with impaired efficacy in hypoxic regions, can reduce the overall tumor response. There is also evidence that malignant cells surviving PDT may become more aggressive than the initial tumor population. Promotion of more effective direct tumor eradication is therefore an important goal. While a list of properties for the "ideal" photosensitizing agent often includes formulation, pharmacologic and photophysical elements, we propose that subcellular targeting is also an important consideration. Perspectives relating to optimizing PDT efficacy are offered here. These relate to death pathways initiated by photodamage to particular subcellular organelles.
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Affiliation(s)
- David Kessel
- Wayne State University School of Medicine, Detroit, MI
| | - Nancy L Oleinick
- Case Western Reserve University School of Medicine and the Case Comprehensive Cancer Center, Cleveland, OH
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Kraus D, Palasuberniam P, Chen B. Targeting Phosphatidylinositol 3-Kinase Signaling Pathway for Therapeutic Enhancement of Vascular-Targeted Photodynamic Therapy. Mol Cancer Ther 2017; 16:2422-2431. [PMID: 28835385 DOI: 10.1158/1535-7163.mct-17-0326] [Citation(s) in RCA: 17] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/12/2017] [Revised: 07/10/2017] [Accepted: 08/07/2017] [Indexed: 11/16/2022]
Abstract
Vascular-targeted photodynamic therapy (PDT) selectively disrupts vascular function by inducing oxidative damages to the vasculature, particularly endothelial cells. Although effective tumor eradication and excellent safety profile are well demonstrated in both preclinical and clinical studies, incomplete vascular shutdown and angiogenesis are known to cause tumor recurrence after vascular-targeted PDT. We have explored therapeutic enhancement of vascular-targeted PDT with PI3K signaling pathway inhibitors because the activation of PI3K pathway was involved in promoting endothelial cell survival and proliferation after PDT. Here, three clinically relevant small-molecule inhibitors (BYL719, BKM120, and BEZ235) of the PI3K pathway were evaluated in combination with verteporfin-PDT. Although all three inhibitors were able to synergistically enhance PDT response in endothelial cells, PDT combined with dual PI3K/mTOR inhibitor BEZ235 exhibited the strongest synergism, followed in order by combinations with pan-PI3K inhibitor BKM120 and p110α isoform-selective inhibitor BYL719. Combination treatments of PDT and BEZ235 exhibited a cooperative inhibition of antiapoptotic Bcl-2 family protein Mcl-1 and induced more cell apoptosis than each treatment alone. In addition to increasing treatment lethality, BEZ235 combined with PDT effectively inhibited PI3K pathway activation and consequent endothelial cell proliferation after PDT alone, leading to a sustained growth inhibition. In the PC-3 prostate tumor model, combination treatments improved treatment outcomes by turning a temporary tumor regrowth delay induced by PDT alone to a more long-lasting treatment response. Our study strongly supports the combination of vascular-targeted PDT and PI3K pathway inhibitors, particularly mTOR inhibitors, for therapeutic enhancement. Mol Cancer Ther; 16(11); 2422-31. ©2017 AACR.
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Affiliation(s)
- Daniel Kraus
- Department of Pharmaceutical Sciences, Philadelphia College of Pharmacy, University of the Sciences, Philadelphia, Pennsylvania
| | - Pratheeba Palasuberniam
- Department of Pharmaceutical Sciences, Philadelphia College of Pharmacy, University of the Sciences, Philadelphia, Pennsylvania
| | - Bin Chen
- Department of Pharmaceutical Sciences, Philadelphia College of Pharmacy, University of the Sciences, Philadelphia, Pennsylvania. .,Department of Radiation Oncology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania
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11
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Gómez-Sintes R, Ledesma MD, Boya P. Lysosomal cell death mechanisms in aging. Ageing Res Rev 2016; 32:150-168. [PMID: 26947122 DOI: 10.1016/j.arr.2016.02.009] [Citation(s) in RCA: 140] [Impact Index Per Article: 15.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/28/2015] [Revised: 02/22/2016] [Accepted: 02/29/2016] [Indexed: 12/14/2022]
Abstract
Lysosomes are degradative organelles essential for cell homeostasis that regulate a variety of processes, from calcium signaling and nutrient responses to autophagic degradation of intracellular components. Lysosomal cell death is mediated by the lethal effects of cathepsins, which are released into the cytoplasm following lysosomal damage. This process of lysosomal membrane permeabilization and cathepsin release is observed in several physiopathological conditions and plays a role in tissue remodeling, the immune response to intracellular pathogens and neurodegenerative diseases. Many evidences indicate that aging strongly influences lysosomal activity by altering the physical and chemical properties of these organelles, rendering them more sensitive to stress. In this review we focus on how aging alters lysosomal function and increases cell sensitivity to lysosomal membrane permeabilization and lysosomal cell death, both in physiological conditions and age-related pathologies.
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Affiliation(s)
- Raquel Gómez-Sintes
- Department of Cellular and Molecular Biology, Centro de Investigaciones Biologicas, CIB-CSIC, C/Ramiro de Maeztu 9, 28040 Madrid, Spain
| | - María Dolores Ledesma
- Department of Molecular Neurobiology, Centro Biologia Molecular Severo Ochoa, CSIC-UAM, C/Nicolás Cabrera 1, 28049 Madrid, Spain
| | - Patricia Boya
- Department of Cellular and Molecular Biology, Centro de Investigaciones Biologicas, CIB-CSIC, C/Ramiro de Maeztu 9, 28040 Madrid, Spain.
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Soler DC, Ohtola J, Sugiyama H, Rodriguez ME, Han L, Oleinick NL, Lam M, Baron ED, Cooper KD, McCormick TS. Activated T cells exhibit increased uptake of silicon phthalocyanine Pc 4 and increased susceptibility to Pc 4-photodynamic therapy-mediated cell death. Photochem Photobiol Sci 2016; 15:822-31. [PMID: 27161819 DOI: 10.1039/c6pp00058d] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/21/2022]
Abstract
Photodynamic therapy (PDT) is an emerging treatment for malignant and inflammatory dermal disorders. Photoirradiation of the silicon phthalocyanine (Pc) 4 photosensitizer with red light generates singlet oxygen and other reactive oxygen species to induce cell death. We previously reported that Pc 4-PDT elicited cell death in lymphoid-derived (Jurkat) and epithelial-derived (A431) cell lines in vitro, and furthermore that Jurkat cells were more sensitive than A431 cells to treatment. In this study, we examined the effectiveness of Pc 4-PDT on primary human CD3(+) T cells in vitro. Fluorometric analyses of lysed T cells confirmed the dose-dependent uptake of Pc 4 in non-stimulated and stimulated T cells. Flow cytometric analyses measuring annexin V and propidium iodide (PI) demonstrated a dose-dependent increase of T cell apoptosis (6.6-59.9%) at Pc 4 doses ranging from 0-300 nM. Following T cell stimulation through the T cell receptor using a combination of anti-CD3 and anti-CD28 antibodies, activated T cells exhibited increased susceptibility to Pc 4-PDT-induced apoptosis (10.6-81.2%) as determined by Pc 4 fluorescence in each cell, in both non-stimulated and stimulated T cells, Pc 4 uptake increased with Pc 4 dose up to 300 nM as assessed by flow cytometry. The mean fluorescence intensity (MFI) of Pc 4 uptake measured in stimulated T cells was significantly increased over the uptake of resting T cells at each dose of Pc 4 tested (50, 100, 150 and 300 nM, p < 0.001 between 50 and 150 nM, n = 8). Treg uptake was diminished relative to other T cells. Cutaneous T cell lymphoma (CTCL) T cells appeared to take up somewhat more Pc 4 than normal resting T cells at 100 and 150 nm Pc 4. Confocal imaging revealed that Pc 4 localized in cytoplasmic organelles, with approximately half of the Pc 4 co-localized with mitochondria in T cells. Thus, Pc 4-PDT exerts an enhanced apoptotic effect on activated CD3(+) T cells that may be exploited in targeting T cell-mediated skin diseases, such as cutaneous T cell lymphoma (CTCL) or psoriasis.
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Affiliation(s)
- David C Soler
- Department of Dermatology, Case Western Reserve University, University Hospitals Case Medical Center, Cleveland, OH, USA.
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13
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Bacellar IOL, Tsubone TM, Pavani C, Baptista MS. Photodynamic Efficiency: From Molecular Photochemistry to Cell Death. Int J Mol Sci 2015; 16:20523-59. [PMID: 26334268 PMCID: PMC4613217 DOI: 10.3390/ijms160920523] [Citation(s) in RCA: 260] [Impact Index Per Article: 26.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/28/2015] [Revised: 08/18/2015] [Accepted: 08/24/2015] [Indexed: 12/11/2022] Open
Abstract
Photodynamic therapy (PDT) is a clinical modality used to treat cancer and infectious diseases. The main agent is the photosensitizer (PS), which is excited by light and converted to a triplet excited state. This latter species leads to the formation of singlet oxygen and radicals that oxidize biomolecules. The main motivation for this review is to suggest alternatives for achieving high-efficiency PDT protocols, by taking advantage of knowledge on the chemical and biological processes taking place during and after photosensitization. We defend that in order to obtain specific mechanisms of cell death and maximize PDT efficiency, PSes should oxidize specific molecular targets. We consider the role of subcellular localization, how PS photochemistry and photophysics can change according to its nanoenvironment, and how can all these trigger specific cell death mechanisms. We propose that in order to develop PSes that will cause a breakthrough enhancement in the efficiency of PDT, researchers should first consider tissue and intracellular localization, instead of trying to maximize singlet oxygen quantum yields in in vitro tests. In addition to this, we also indicate many open questions and challenges remaining in this field, hoping to encourage future research.
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Affiliation(s)
- Isabel O L Bacellar
- Instituto de Química, Universidade de São Paulo, São Paulo 05508-900, Brazil.
| | - Tayana M Tsubone
- Instituto de Química, Universidade de São Paulo, São Paulo 05508-900, Brazil.
| | - Christiane Pavani
- Programa de Pós Graduação em Biofotônica Aplicada às Ciências da Saúde, Universidade Nove de Julho, São Paulo 01504-001, Brazil.
| | - Mauricio S Baptista
- Instituto de Química, Universidade de São Paulo, São Paulo 05508-900, Brazil.
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14
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Master A, Malamas A, Solanki R, Clausen DM, Eiseman JL, Sen Gupta A. A cell-targeted photodynamic nanomedicine strategy for head and neck cancers. Mol Pharm 2013; 10:1988-97. [PMID: 23531079 DOI: 10.1021/mp400007k] [Citation(s) in RCA: 51] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
Photodynamic therapy (PDT) holds great promise for the treatment of head and neck (H&N) carcinomas where repeated loco-regional therapy often becomes necessary due to the highly aggressive and recurrent nature of the cancers. While interstitial light delivery technologies are being refined for PDT of H&N and other cancers, a parallel clinically relevant research area is the formulation of photosensitizers in nanovehicles that allow systemic administration yet preferential enhanced uptake in the tumor. This approach can render dual-selectivity of PDT, by harnessing both the drug and the light delivery within the tumor. To this end, we report on a cell-targeted nanomedicine approach for the photosensitizer silicon phthalocyanine-4 (Pc 4), by packaging it within polymeric micelles that are surface-decorated with GE11-peptides to promote enhanced cell-selective binding and receptor-mediated internalization in EGFR-overexpressing H&N cancer cells. Using fluorescence spectroscopy and confocal microscopy, we demonstrate in vitro that the EGFR-targeted Pc 4-nanoformulation undergoes faster and higher uptake in EGFR-overexpressing H&N SCC-15 cells. We further demonstrate that this enhanced Pc 4 uptake results in significant cell-killing and drastically reduced post-PDT clonogenicity. Building on this in vitro data, we demonstrate that the EGFR-targeted Pc 4-nanoformulation results in significant intratumoral drug uptake and subsequent enhanced PDT response, in vivo, in SCC-15 xenografts in mice. Altogether our results show significant promise toward a cell-targeted photodynamic nanomedicine for effective treatment of H&N carcinomas.
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Affiliation(s)
- Alyssa Master
- Department of Biomedical Engineering, Case Western Reserve University, 2071 Martin Luther King Blvd, Cleveland, Ohio 44106, United States
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15
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Song J, Chen Q, Xing D. Enhanced apoptotic effects by downregulating Mcl-1: evidence for the improvement of photodynamic therapy with Celecoxib. Exp Cell Res 2013; 319:1491-504. [PMID: 23524145 DOI: 10.1016/j.yexcr.2013.03.012] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/31/2012] [Revised: 03/05/2013] [Accepted: 03/09/2013] [Indexed: 12/23/2022]
Abstract
Tumor cells exposed to sub-lethal photodynamic therapy (PDT) cause cellular rescue responses that lead to resistance to the therapy, including expression of angiogenic factors and survival molecules. However, the mechanisms contributing to the resistance are yet to be fully understood. Here, we show for the first time that Mcl-1, an anti-apoptotic protein, plays an important role in protecting cells from PDT-induced apoptosis. In contrast to the reduction in the anti-apoptotic proteins Bcl-2 and Bcl-xl, sub-lethal PDT induces an increase in Mcl-1 expression. Silencing Mcl-1 sensitizes tumor cells to PDT-induced apoptosis, and ectopic expression of Mcl-1 significantly delays Bax translocation to mitochondria and inhibits caspase-3 activity following PDT. Mcl-1 expression is associated closely with activated AKT signaling following PDT. AKT can regulate Mcl-1 expression through GSK-3β and NF-κB at the protein and transcriptional levels, respectively. Inhibition of AKT by Wortmannin or siRNA significantly reduces the levels of Mcl-1 mRNA and protein and enhances PDT-induced apoptosis. Treatment with Celecoxib, a non-steroidal anti-inflammatory drug (NSAID), is shown to downregulate Mcl-1 expression, and enhances PDT-induced apoptosis both in vitro and in vivo. This down-regulation is closely related to the inhibition effect of Celecoxib on the AKT/GSK-3β pathway, and was blocked upon addition of GSK-3β inhibitor LiCl or the proteasome inhibitor MG132. These results suggest that Mcl-1 is a potential target for improving the antitumor efficiency of PDT. A loss in Mcl-1 by inhibiting AKT promotes PDT-induced apoptosis through the mitochondrial pathway. This also provides a novel rationale for utilizing Celecoxib to improve the efficacy of PDT.
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Affiliation(s)
- Jiaxing Song
- MOE Key Laboratory of Laser Life Science & Institute of Laser Life Science, College of Biophotonics, South China Normal University, Guangzhou 510631, China
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16
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Yoo JO, Ha KS. New insights into the mechanisms for photodynamic therapy-induced cancer cell death. INTERNATIONAL REVIEW OF CELL AND MOLECULAR BIOLOGY 2012; 295:139-74. [PMID: 22449489 DOI: 10.1016/b978-0-12-394306-4.00010-1] [Citation(s) in RCA: 108] [Impact Index Per Article: 8.3] [Reference Citation Analysis] [Abstract] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
Photodynamic therapy (PDT) is a promising therapeutic modality for cancer treatment; however, a more detailed understanding is needed to improve the clinical use of this therapy. PDT induces cancer cell death by apoptosis, necrosis, and autophagy, and these mechanisms can be concurrently occurred. PDT destroys cancer cells by inducing apoptosis through diverse signaling pathways coupled with Bcl-2 family members, caspases, and apopotosis-inducing factor. When the apoptotic pathway is unavailable, PDT can cause cancer cell death through induction of a necrotic or autophagic mechanism. Autophagy is occurred in a Bax-independent manner and can be stimulated in parallel with apoptosis. PDT directly destroys cancer cells by inducing either apoptotic or necrotic death. PDT also can induce autophagy as a death or a survival mechanism. These mechanisms are dependent on a variety of parameters including the nature of the photosensitizer, PDT dose, and cell genotype. Understanding the complex cross talk between these pathways may improve the effectiveness of PDT. Here, we discuss the interplay between these mechanisms based on recent evidence and suggest prospects with regard to advances in PDT.
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Affiliation(s)
- Je-Ok Yoo
- Department of Molecular and Cellular Biochemistry, Kangwon National University School of Medicine, Chuncheon, Kangwon-do, South Korea
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17
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Autophagy Contributes to the Death/Survival Balance in Cancer PhotoDynamic Therapy. Cells 2012; 1:464-91. [PMID: 24710486 PMCID: PMC3901101 DOI: 10.3390/cells1030464] [Citation(s) in RCA: 50] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/15/2012] [Revised: 07/09/2012] [Accepted: 07/19/2012] [Indexed: 12/19/2022] Open
Abstract
Autophagy is an important cellular program with a “double face” role, since it promotes either cell survival or cell death, also in cancer therapies. Its survival role occurs by recycling cell components during starvation or removing stressed organelles; when damage becomes extensive, autophagy provides another programmed cell death pathway, known as Autophagic Cell Death (ACD). The induction of autophagy is a common outcome in PhotoDynamic Therapy (PDT), a two-step process involving the irradiation of photosensitizer (PS)-loaded cancer cells. Upon tissue oxygen interaction, PS provokes immediate and direct Reactive Oxygen Species (ROS)-induced damage to Endoplasmic Reticulum (ER), mitochondria, plasma membrane, and/or lysosomes. The main biological effects carried out in cancer PDT are direct cytotoxicity to tumor cells, vasculature damage and induction of inflammatory reactions stimulating immunological responses. The question about the role of autophagy in PDT and its putative immunological impact is hotly controversial and largely studied in recent times. This review deals with the induction of autophagy in PDT protocols and its dual role, also considering its interrelationship with apoptosis, the preferential cell death program triggered in the photodynamic process.
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18
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Lam M, Lee Y, Deng M, Hsia AH, Morrissey KA, Yan C, Azzizudin K, Oleinick NL, McCormick TS, Cooper KD, Baron ED. Photodynamic therapy with the silicon phthalocyanine pc 4 induces apoptosis in mycosis fungoides and sezary syndrome. Adv Hematol 2010; 2010:896161. [PMID: 21197103 PMCID: PMC3004392 DOI: 10.1155/2010/896161] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/06/2010] [Revised: 09/02/2010] [Accepted: 10/14/2010] [Indexed: 11/18/2022] Open
Abstract
Our current focus on the effects of Photodynamic Therapy (PDT) using silicon phthalocyanine Pc 4 photosensitizer on malignant T lymphocytes arose due to preclinical observations that Jurkat cells, common surrogate for human T cell lymphoma, were more sensitive to Pc 4-PDT-induced killing than epidermoid carcinoma A431 cells. Mycosis fungoides (MF) as well as Sezary syndrome (SS) are variants of cutaneous T-cell lymphoma (CTCL) in which malignant T-cells invade the epidermis. In this study, we investigated the cytotoxicity of Pc 4-PDT in peripheral blood cells obtained from patients with SS and in skin biopsies of patients with MF. Our data suggest that Pc 4-PDT preferentially induces apoptosis of CD4(+)CD7(-) malignant T-lymphocytes in the blood relative to CD11b(+) monocytes and nonmalignant T-cells. In vivo Pc 4-PDT of MF skin also photodamages the antiapoptotic protein Bcl-2.
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Affiliation(s)
- Minh Lam
- Department of Dermatology, University Hospitals of Cleveland, Case Western Reserve University, 11100 Euclid Avenue, Lakeside 3500, Cleveland, OH 44106-5028, USA
| | - YooJin Lee
- Department of Dermatology, University Hospitals of Cleveland, Case Western Reserve University, 11100 Euclid Avenue, Lakeside 3500, Cleveland, OH 44106-5028, USA
| | - Min Deng
- Department of Dermatology, University Hospitals of Cleveland, Case Western Reserve University, 11100 Euclid Avenue, Lakeside 3500, Cleveland, OH 44106-5028, USA
| | - Andrew H. Hsia
- Department of Dermatology, University Hospitals of Cleveland, Case Western Reserve University, 11100 Euclid Avenue, Lakeside 3500, Cleveland, OH 44106-5028, USA
| | - Kelly A. Morrissey
- Department of Dermatology, University Hospitals of Cleveland, Case Western Reserve University, 11100 Euclid Avenue, Lakeside 3500, Cleveland, OH 44106-5028, USA
| | - Chunlin Yan
- Department of Dermatology, University Hospitals of Cleveland, Case Western Reserve University, 11100 Euclid Avenue, Lakeside 3500, Cleveland, OH 44106-5028, USA
- Department of Dermatology, Huashan Hospital, Fudan University, Shanghai, China
| | - Kashif Azzizudin
- Department of Radiation Oncology, Case Western Reserve University, 10900 Euclid Ave., Cleveland, OH 44106-4942, USA
| | - Nancy L. Oleinick
- Department of Radiation Oncology, Case Western Reserve University, 10900 Euclid Ave., Cleveland, OH 44106-4942, USA
| | - Thomas S. McCormick
- Department of Dermatology, University Hospitals of Cleveland, Case Western Reserve University, 11100 Euclid Avenue, Lakeside 3500, Cleveland, OH 44106-5028, USA
| | - Kevin D. Cooper
- Department of Dermatology, University Hospitals of Cleveland, Case Western Reserve University, 11100 Euclid Avenue, Lakeside 3500, Cleveland, OH 44106-5028, USA
- Dermatology Department, Cleveland Veterans Affairs Medical Center, Cleveland, OH 44106, USA
| | - Elma D. Baron
- Department of Dermatology, University Hospitals of Cleveland, Case Western Reserve University, 11100 Euclid Avenue, Lakeside 3500, Cleveland, OH 44106-5028, USA
- Dermatology Department, Cleveland Veterans Affairs Medical Center, Cleveland, OH 44106, USA
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19
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Photodynamic properties of ZnTPPS4, ClAlPcS2 and ALA in human melanoma G361 cells. Toxicol In Vitro 2010; 24:286-91. [DOI: 10.1016/j.tiv.2009.08.015] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/07/2009] [Revised: 08/13/2009] [Accepted: 08/25/2009] [Indexed: 12/15/2022]
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20
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Bonchev D, Thomas S, Apte A, Kier LB. Cellular automata modelling of biomolecular networks dynamics. SAR AND QSAR IN ENVIRONMENTAL RESEARCH 2010; 21:77-102. [PMID: 20373215 DOI: 10.1080/10629360903568580] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/29/2023]
Abstract
The modelling of biological systems dynamics is traditionally performed by ordinary differential equations (ODEs). When dealing with intracellular networks of genes, proteins and metabolites, however, this approach is hindered by network complexity and the lack of experimental kinetic parameters. This opened the field for other modelling techniques, such as cellular automata (CA) and agent-based modelling (ABM). This article reviews this emerging field of studies on network dynamics in molecular biology. The basics of the CA technique are discussed along with an extensive list of related software and websites. The application of CA to networks of biochemical reactions is exemplified in detail by the case studies of the mitogen-activated protein kinase (MAPK) signalling pathway, the FAS-ligand (FASL)-induced and Bcl-2-related apoptosis. The potential of the CA method to model basic pathways patterns, to identify ways to control pathway dynamics and to help in generating strategies to fight with cancer is demonstrated. The different line of CA applications presented includes the search for the best-performing network motifs, an analysis of importance for effective intracellular signalling and pathway cross-talk.
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Affiliation(s)
- D Bonchev
- Department of Mathematics and Applied Mathematics, Virginia Commonwealth University, Center for the Study of Biological Complexity, Richmond, Virginia, USA.
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21
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Abstract
Photodynamic therapy (PDT) is the term used to describe the irradiation of photosensitized cells or tissue with phototoxic consequences. This process can result in the rapid initiation of not only apoptosis, an irreversible death pathway, but also autophagy. The procedures described here are designed to characterize the correlation between the PDT dose vs. survival of cells in vitro, the apoptotic effects of photodamage, and the extent of an autophagic response. These are assessed by clonogenic assays, observation of condensed chromatin characteristic of apoptosis, activation of "executioner" caspases, and the autophagic flux as indicated by comparing accumulation of the LC3-II protein under conditions where processing of autophagosomes is retarded vs. is not retarded.
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22
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Reiners JJ, Agostinis P, Berg K, Oleinick NL, Kessel D. Assessing autophagy in the context of photodynamic therapy. Autophagy 2010; 6:7-18. [PMID: 19855190 DOI: 10.4161/auto.6.1.10220] [Citation(s) in RCA: 166] [Impact Index Per Article: 11.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/08/2023] Open
Abstract
Photodynamic therapy (PDT) is a procedure that has applications in the selective eradication of neoplasia where sites of malignant lesions are clearly delineated. It is a two-step process whereby cells are first sensitized to light and then photoirradiated. This results in the formation of singlet molecular oxygen and other reactive oxygen species that can cause photodamage at sites where the photosensitizing agent has localized. Photosensitizers found to be clinically useful show affinity for the endoplasmic reticulum (ER), mitochondria, lysosomes, or combinations of these sites. The induction of apoptosis and/or autophagy in photosensitized cells is a common outcome of PDT. This report explores the following issues: (1) Does the induction of autophagy in PDT protocols occur independent of, or in association with, apoptosis? (2) Does the resulting autophagy play a prosurvival or prodeath role? (3) Do photosensitizers damage/inactivate specific proteins that are components of, or that modulate the autophagic process? (4) Can an autophagic response be mounted in cells in which lysosomes are specifically photodamaged? In brief, autophagy can occur independently of apoptosis in PDT protocols, and appears to play a prosurvival role in apoptosis competent cells, and a prodeath role in apoptosis incompetent cells. Mitochondrial and ER-localized sensitizers cause selective photodamage to some (i.e., Bcl-2, Bcl-x(L), mTOR) proteins involved in the apoptotic/autophagic process. Finally, an aborted autophagic response occurs in cells with photodamaged lysosomes. Whereas autophagosomes form, digestion of their cargo is compromised because of the absence of functional lysosomes.
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Affiliation(s)
- John J Reiners
- Institute of Environmental Health Sciences, Wayne State University, Detroit, MI, USA
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23
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Belcher J, Sansone S, Fernandez NF, Haskins WE, Brancaleon L. Photoinduced unfolding of beta-lactoglobulin mediated by a water-soluble porphyrin. J Phys Chem B 2009; 113:6020-30. [PMID: 19351165 PMCID: PMC2735475 DOI: 10.1021/jp900957d] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/20/2023]
Abstract
We investigated the effects that the irradiation of a tetra-anionic porphyrin (mesotetrakis(sulfonatophenyl)porphyrin) noncovalently bound to beta-lactoglobulin (BLG) produces on the conformation of the protein. Although BLG is not a potential target for the biomedical applications of porphyrins, it is a useful model for investigating the effects of photoactive ligands on small globular proteins. We show in this paper that irradiation causes a large unfolding of the protein and that the conformational change is not mediated by the formation of reactive oxygen species. Instead, our data are consistent with an electron-transfer mechanism that is capable of triggering structural changes in the protein and causes the Trp19 residue to undergo chemical modifications to form a derivative of kynurenine. This demonstrates that protein unfolding is prompted by a type-III photosensitizing mechanisms. Type-III mechanisms have been suggested previously, but they have been largely neglected as useful mediators of biomolecular damage. Our study demonstrates that porphyrins can be used as mediators of localized protein conformational changes and that the biomedical applications as well as the mechanistic details of electron transfer between exogenous ligands and proteins merit further investigation.
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Affiliation(s)
- John Belcher
- Department of Physics and Astronomy, University of Texas at San Antonio, San Antonio, TX, USA
| | - Samuel Sansone
- Department of Physics and Astronomy, University of Texas at San Antonio, San Antonio, TX, USA
| | - Nicholas F. Fernandez
- Department of Physics and Astronomy, University of Texas at San Antonio, San Antonio, TX, USA
| | - William E. Haskins
- Department of Biology, University of Texas at San Antonio, San Antonio, TX, USA
| | - Lorenzo Brancaleon
- Department of Physics and Astronomy, University of Texas at San Antonio, San Antonio, TX, USA
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24
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Abstract
Mitochondrial outer membrane permeabilization (MOMP) constitutes one of the major checkpoint(s) of apoptotic and necrotic cell death. Recently, the permeabilization of yet another organelle, the lysosome, has been shown to initiate a cell death pathway, in specific circumstances. Lysosomal membrane permeabilization (LMP) causes the release of cathepsins and other hydrolases from the lysosomal lumen to the cytosol. LMP is induced by a plethora of distinct stimuli including reactive oxygen species, lysosomotropic compounds with detergent activity, as well as some endogenous cell death effectors such as Bax. LMP is a potentially lethal event because the ectopic presence of lysosomal proteases in the cytosol causes digestion of vital proteins and the activation of additional hydrolases including caspases. This latter process is usually mediated indirectly, through a cascade in which LMP causes the proteolytic activation of Bid (which is cleaved by the two lysosomal cathepsins B and D), which then induces MOMP, resulting in cytochrome c release and apoptosome-dependent caspase activation. However, massive LMP often results in cell death without caspase activation; this cell death may adopt a subapoptotic or necrotic appearance. The regulation of LMP is perturbed in cancer cells, suggesting that specific strategies for LMP induction might lead to novel therapeutic avenues.
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Affiliation(s)
- P Boya
- 3D Lab (Development, Differentiation and Degeneration), Department of Cellular and Molecular Physiopathology, Centro de Investigaciones Biológicas, CSIC, Madrid, Spain.
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25
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Fernandez NF, Sansone S, Mazzini A, Brancaleon L. Irradiation of the porphyrin causes unfolding of the protein in the protoporphyrin IX/beta-lactoglobulin noncovalent complex. J Phys Chem B 2008; 112:7592-600. [PMID: 18517238 PMCID: PMC3514890 DOI: 10.1021/jp710249d] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Abstract
Porphyrins such as protoporphyrin IX (PPIX) are known to occasionally cause conformational changes in proteins for which they are specific ligands. It has also been established that irradiation of porphyrins noncovalently intercalated between bases or bound to one of the grooves can cause conformational effects on DNA. Conversely, there is no evidence reported in the literature of conformational changes caused by noncovalently bound PPIX to globular proteins for which the porphyrin is not a specific ligand. This study shows that the irradiation of the porphyrin in the PPIX/lactoglobulin noncovalent complex indeed causes a local and limited (approximately 7%) unfolding of the protein near the location of Trp19. This event causes the intrinsic fluorescence spectrum of the protein to shift to the red by 2 nm and the average decay lifetime to lengthen by approximately 0.5 ns. The unfolding of lactoglobulin occurs only at pH >7 because of the increased instability of the protein at alkaline pH. The photoinduced unfolding does not depend on the presence of O2 in solution; therefore, it is not mediated by formation of singlet oxygen and is likely the result of electron transfer between the porphyrin and amino acid residues.
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26
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Kim J, Rodriguez ME, Guo M, Kenney ME, Oleinick NL, Anderson VE. Oxidative modification of cytochrome c by singlet oxygen. Free Radic Biol Med 2008; 44:1700-11. [PMID: 18242196 PMCID: PMC2424268 DOI: 10.1016/j.freeradbiomed.2007.12.031] [Citation(s) in RCA: 50] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/10/2007] [Revised: 12/06/2007] [Accepted: 12/18/2007] [Indexed: 11/15/2022]
Abstract
Singlet oxygen ((1)O(2)) is a reactive oxygen species that may be generated in biological systems. Photodynamic therapy generates (1)O(2) by photoexcitation of sensitizers resulting in intracellular oxidative stress and induction of apoptosis. (1)O(2) oxidizes amino acid side chains of proteins and inactivates enzymes when generated in vitro. Among proteogenic amino acids, His, Tyr, Met, Cys, and Trp are known to be oxidized by (1)O(2) at physiological pH. However, there is a lack of direct evidence of oxidation of proteins by (1)O(2). Because (1)O(2) is difficult to detect in cells, identifying oxidized cellular products uniquely derived from (1)O(2) could serve as a marker of its presence. In the present study, (1)O(2) reactions with model peptides analyzed by tandem mass spectrometry provide insight into the mass of prominent adducts formed with the reactive amino acids. Analysis by MALDI-TOF and tandem mass spectrometry of peptides of cytochrome c exposed to (1)O(2) generated by photoexcitation of the phthalocyanine Pc 4 showed unique oxidation products, which might be used as markers of the presence of (1)O(2) in the mitochondrial intermembrane space. Differences in the elemental composition of the oxidized amino acid residues observed with cytochrome c and the model peptides suggest that the protein environment can affect the oxidation pathway.
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Affiliation(s)
- Junhwan Kim
- Department of Biochemistry, School of Medicine, Case Western Reserve University, Cleveland, OH, USA
| | - Myriam E. Rodriguez
- Department of Radiation Oncology, School of Medicine, Case Western Reserve University/University Hospitals of Cleveland, Cleveland, OH 44106, USA
| | - Ming Guo
- Department of Chemistry, Case Western Reserve University, Cleveland, OH, USA
| | - Malcolm E. Kenney
- Department of Chemistry, Case Western Reserve University, Cleveland, OH, USA
- Case Comprehensive Cancer Center, Case Western Reserve University, Cleveland, OH, USA
| | - Nancy L. Oleinick
- Department of Biochemistry, School of Medicine, Case Western Reserve University, Cleveland, OH, USA
- Department of Radiation Oncology, School of Medicine, Case Western Reserve University/University Hospitals of Cleveland, Cleveland, OH 44106, USA
- Case Comprehensive Cancer Center, Case Western Reserve University, Cleveland, OH, USA
| | - Vernon E. Anderson
- Department of Biochemistry, School of Medicine, Case Western Reserve University, Cleveland, OH, USA
- Department of Chemistry, Case Western Reserve University, Cleveland, OH, USA
- Case Comprehensive Cancer Center, Case Western Reserve University, Cleveland, OH, USA
- Corresponding author: 10900 Euclid Avenue, Cleveland, OH 44106-4935, Ph: (216) 368-2599, fax (216) 368 3419, E-mail
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27
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Chiu SM, Xue LY, Azizuddin K, Oleinick NL. Photodynamic therapy-induced death of HCT 116 cells: Apoptosis with or without Bax expression. Apoptosis 2008; 10:1357-68. [PMID: 16215676 DOI: 10.1007/s10495-005-2217-0] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
Cell death following photodynamic therapy (PDT) with the photosensitizer Pc 4 involves the intrinsic pathway of apoptosis. To evaluate the importance of Bax in apoptosis after PDT, we compared the PDT responses of Bax-proficient (Bax(+/-)) and Bax knock-out (BaxKO) HCT116 human colon cancer cells. PDT induced a slow apoptotic process in HCT Bax(+/-) cells following a long delay in the activation of Bax and release of cytochrome c from mitochondria. Although cytochrome c was not released from mitochondria following PDT in BaxKO cells, an alternative mechanism of caspase-dependent apoptosis with extensive chromatin and DNA degradation was found in these cells. This alternative process was less efficient and slower than the normal apoptotic process observed in Bax(+/-) cells. Early events upon PDT, such as the loss of mitochondrial membrane potential, photodamage to Bcl-2, and activation of p38 MAP kinase, were observed in both HCT116 cell lines. In spite of differences in the efficiency and mode of apoptosis induced by PDT in the Bax(+/-) and BaxKO cells, they were found to be equally sensitive to killing by PDT, as determined by loss of clonogenicity. Thus, for Pc 4-PDT, the commitment to cell death occurs prior to and independent of Bax activation, but the process of cellular disassembly differs in Bax-expressing vs. non-expressing cells.
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Affiliation(s)
- S-M Chiu
- Department of Radiation Oncology and The Case Comprehensive Cancer Center, School of Medicine, Case Western Reserve University, Cleveland, OH 44106, USA
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Ke MS, Xue LY, Feyes DK, Azizuddin K, Baron ED, McCormick TS, Mukhtar H, Panneerselvam A, Schluchter MD, Cooper KD, Oleinick NL, Stevens SR. Apoptosis mechanisms related to the increased sensitivity of Jurkat T-cells vs A431 epidermoid cells to photodynamic therapy with the phthalocyanine Pc 4. Photochem Photobiol 2008; 84:407-14. [PMID: 18221452 DOI: 10.1111/j.1751-1097.2007.00278.x] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/28/2022]
Abstract
To examine the clinical applicability of Pc 4, a promising second-generation photosensitizer, for the photodynamic treatment of lymphocyte-mediated skin diseases, we studied the A431 and Jurkat cell lines, commonly used as surrogates for human keratinocyte-derived carcinomas and lymphocytes, respectively. As revealed by ethyl acetate extraction and absorption spectrophotometry, uptake of Pc 4 into the two cell lines was linear with Pc 4 concentration and similar on a per cell basis but greater in Jurkat cells on a per mass basis. Flow cytometry showed that uptake was linear at low doses; variations in the dose-response for uptake measured by fluorescence supported differential aggregation of Pc 4 in the two cell types. As detected by confocal microscopy, Pc 4 localized to mitochondria and endoplasmic reticulum in both cell lines. Jurkat cells were much more sensitive to the lethal effects of phthalocyanine photodynamic therapy (Pc 4-PDT) than were A431 cells, as measured by a tetrazolium dye reduction assay, and more readily underwent morphological apoptosis. In a search for molecular factors to explain the greater photosensitivity of Jurkat cells, the fate of important Bcl-2 family members was monitored. Jurkat cells were more sensitive to the induction of immediate photodamage to Bcl-2, but the difference was insufficient to account fully for their greater sensitivity. The antiapoptotic protein Mcl-1 was extensively cleaved in a dose- and caspase-dependent manner in Jurkat, but not in A431, cells exposed to Pc 4-PDT. Thus, the greater killing by Pc 4-PDT in Jurkat compared with A431 cells correlated with greater Bcl-2 photodamage and more strongly to the more extensive Mcl-1 degradation. Pc 4-PDT may offer therapeutic advantages in targeting inflammatory cells over normal keratinocytes in the treatment of T-cell-mediated skin diseases, such as cutaneous lymphomas, dermatitis, lichenoid tissue reactions and psoriasis, and it will be instructive to evaluate the role of Bcl-2 family proteins, especially Mcl-1, in the therapeutic response.
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Affiliation(s)
- Malcolm S Ke
- Department of Dermatology, University Hospitals Case Medical Center and Case Western Reserve University, Cleveland, OH, USA
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Wang KKH, Wilson JD, Kenney ME, Mitra S, Foster TH. Irradiation-induced enhancement of Pc 4 fluorescence and changes in light scattering are potential dosimeters for Pc 4-PDT. Photochem Photobiol 2008; 83:1056-62. [PMID: 17880500 DOI: 10.1111/j.1751-1097.2007.00128.x] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/27/2022]
Abstract
Phthalocyanine 4 (Pc 4) is a promising photosensitizer currently in clinical trials. Photobiological responses to Pc 4 photodynamic therapy (Pc 4-PDT) have been characterized extensively, but relatively little has been done to evaluate dose metrics for this sensitizer. We describe an irradiation-induced increase in fluorescence in tumor cell monolayers. This increase is due solely to enhanced fluorescence from Pc 4, as confirmed by confocal spectroscopy. In EMT6 cells incubated with 250 nM Pc 4 for 24 h, the maximum increase in fluorescence is approximately 3.7-fold above baseline levels. This increase occurs over a range of fluences, 0.05-0.6 J cm(-2), where clonogenic survival decreases by 3 orders of magnitude. Light scattering measurements performed on similarly treated EMT6 cells in suspension suggested a Pc 4-PDT-mediated mitochondrial swelling of approximately 13% at 0.6 J cm(-2), where fluorescence enhancement saturates under these treatment conditions. Fluorescence imaging and light scattering experiments performed at a five-fold lower Pc 4 incubation concentration revealed a reduced fluorescence enhancement at a five-fold higher fluence, which produced comparable mitochondrial swelling. Taken together, these data suggest that Pc 4 is initially aggregated at high local concentration in mitochondria and that irradiation relaxes the quenching of Pc 4 fluorescence through a mechanism that may involve mitochondrial swelling.
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Affiliation(s)
- Ken Kang-Hsin Wang
- Department of Physics and Astronomy, University of Rochester, Rochester, NY, USA
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Cittelly DM, Nesic-Taylor O, Perez-Polo JR. Phosphorylation of Bcl-xL after spinal cord injury. J Neurosci Res 2007; 85:1894-911. [PMID: 17551978 DOI: 10.1002/jnr.21313] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/11/2022]
Abstract
Spinal cord injury (SCI)-induced functional impairment results from secondary apoptosis regulated in part by SCI-induced decreases in the antiapoptotic protein Bcl-x(L). We assessed the role that Bcl-x(L) subcellular rerouting and posttranslational phosphorylation play in Bcl-x(L) decreases in a contusion model of rat SCI. Immunohistochemical analysis showed the presence of Bcl-x(L) in neurons and oligodendrocytes, but not in astrocytes and microglia, whereas phosphorylated Bcl-x(L) (P-ser(62)-Bcl-x(L)) was present only in neurons. Western blot analyses showed Bcl-x(L) present in mitochondria, endoplasmic reticulum, nuclei, and cytosolic extracts, whereas P-ser(62)-Bcl-x(L) was restricted to organelles. During the first 24 hr after SCI, Bcl-x(L) levels decreased in all fractions but with a different time course, suggesting an independent regulation of Bcl-x(L) shuttling from the cytosol to each compartment after SCI. SCI did not affect P-ser(62)-Bcl-x(L) levels in organelles. However, P-ser(62)-Bcl-x(L), which was not detected in the cytosolic fraction of uninjured spinal cord, appeared in the cytosol as early as 15 min postcontusion, suggesting a role for phosphorylation in SCI-induced Bcl-x(L)-decreases. Using an in vitro model, we observed a correlation between levels of cytosolic phosphorylated Bcl-x(L) and neuronal apoptosis, supporting the hypothesis that Bcl-x(L) phosphorylation is proapoptotic. Activated microglia/macrophages robustly expressed Bcl-x(L) 7 days after SCI, and a subpopulation showing nuclear condensation also expressed P-ser(62)-Bcl-x(L). Therefore, phosphorylation of Bcl-x(L) may have opposite effects in injured spinal cords: 1) it may decrease levels of the antiapoptotic Bcl-x(L) in neurons contributing to neuronal death, and 2) it may promote apoptosis in activated microglia/macrophages, thus curtailing the inflammatory cascades associated with SCI.
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Affiliation(s)
- Diana M Cittelly
- Neuroscience and Cell Biology Department, University of Texas Medical Branch, Galveston, Texas 77555-1072, USA
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Henderson BW, Daroqui C, Tracy E, Vaughan LA, Loewen GM, Cooper MT, Baumann H. Cross-linking of signal transducer and activator of transcription 3--a molecular marker for the photodynamic reaction in cells and tumors. Clin Cancer Res 2007; 13:3156-63. [PMID: 17545518 DOI: 10.1158/1078-0432.ccr-06-2950] [Citation(s) in RCA: 35] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
PURPOSE Photodynamic therapy (PDT) depends on the delivery of a photosensitizer to the target tissue that, under light exposure, produces singlet oxygen and other reactive oxygen species, which in turn cause the death of the treated cell. This study establishes a quantitative marker for the photoreaction that will predict the outcome of PDT. EXPERIMENTAL DESIGN Cells in tissue culture, murine s.c. tumors, and endobronchial carcinomas in patients were treated with PDT, and the noncleavable cross-linking of the latent signal transducer and activator of transcription 3 (STAT3) was determined. RESULTS Murine and human cancer cell lines reacted to PDT by an immediate covalent cross-linking of STAT3 to homodimeric and other complexes. The magnitude of this effect was strictly a function of the PDT reaction that is determined by the photosensitizer concentration and light dose. The cross-link reaction of STAT3 was proportional to the subsequent cytotoxic outcome of PDT. An equivalent photoreaction as detected in vitro occurred in tumors treated in situ with PDT. The light dose-dependent STAT3 cross-linking indicated the relative effectiveness of PDT as a function of the distance of the tissue to the treating laser light source. Absence of cross-links correlated with treatment failure. CONCLUSIONS The data suggest that the relative amount of cross-linked STAT3 predicts the probability for beneficial outcome, whereas absence of cross-links predicts treatment failure. Determination of STAT3 cross-links after PDT might be clinically useful for early assessment of PDT response.
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Affiliation(s)
- Barbara W Henderson
- Department of Cell Stress Biology, Roswell Park Cancer Institute, Buffalo, New York 14263, USA
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32
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Buytaert E, Dewaele M, Agostinis P. Molecular effectors of multiple cell death pathways initiated by photodynamic therapy. Biochim Biophys Acta Rev Cancer 2007; 1776:86-107. [PMID: 17693025 DOI: 10.1016/j.bbcan.2007.07.001] [Citation(s) in RCA: 288] [Impact Index Per Article: 16.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/01/2007] [Revised: 06/27/2007] [Accepted: 07/01/2007] [Indexed: 12/12/2022]
Abstract
Photodynamic therapy (PDT) is a recently developed anticancer modality utilizing the generation of singlet oxygen and other reactive oxygen species, through visible light irradiation of a photosensitive dye accumulated in the cancerous tissue. Multiple signaling cascades are concomitantly activated in cancer cells exposed to the photodynamic stress and depending on the subcellular localization of the damaging ROS, these signals are transduced into adaptive or cell death responses. Recent evidence indicates that PDT can kill cancer cells directly by the efficient induction of apoptotic as well as non-apoptotic cell death pathways. The identification of the molecular effectors regulating the cross-talk between apoptosis and other major cell death subroutines (e.g. necrosis, autophagic cell death) is an area of intense research in cancer therapy. Signaling molecules modulating the induction of different cell death pathways can become useful targets to induce or increase photokilling in cancer cells harboring defects in apoptotic pathways, which is a crucial step in carcinogenesis and therapy resistance. This review highlights recent developments aimed at deciphering the molecular interplay between cell death pathways as well as their possible therapeutic exploitation in photosensitized cells.
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Affiliation(s)
- Esther Buytaert
- Department of Molecular and Cell Biology, Faculty of Medicine, Catholic University of Leuven, Campus Gasthuisberg, Herestraat 49, B-3000, Leuven Belgium
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Miller JD, Baron ED, Scull H, Hsia A, Berlin JC, McCormick T, Colussi V, Kenney ME, Cooper KD, Oleinick NL. Photodynamic therapy with the phthalocyanine photosensitizer Pc 4: the case experience with preclinical mechanistic and early clinical-translational studies. Toxicol Appl Pharmacol 2007; 224:290-9. [PMID: 17397888 PMCID: PMC2128784 DOI: 10.1016/j.taap.2007.01.025] [Citation(s) in RCA: 165] [Impact Index Per Article: 9.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/04/2006] [Revised: 01/06/2007] [Accepted: 01/10/2007] [Indexed: 12/01/2022]
Abstract
Photodynamic therapy (PDT) is emerging as a promising non-invasive treatment for cancers. PDT involves either local or systemic administration of a photosensitizing drug, which preferentially localizes within the tumor, followed by illumination of the involved organ with light, usually from a laser source. Here, we provide a selective overview of our experience with PDT at Case Western Reserve University, specifically with the silicon phthalocyanine photosensitizer Pc 4. We first review our in vitro studies evaluating the mechanism of cell killing by Pc 4-PDT. Then we briefly describe our clinical experience in a Phase I trial of Pc 4-PDT and our preliminary translational studies evaluating the mechanisms behind tumor responses. Preclinical work identified (a) cardiolipin and the anti-apoptotic proteins Bcl-2 and Bcl-xL as targets of Pc 4-PDT, (b) the intrinsic pathway of apoptosis, with the key participation of caspase-3, as a central response of many human cancer cells to Pc 4-PDT, (c) signaling pathways that could modify apoptosis, and (d) a formulation by which Pc 4 could be applied topically to human skin and penetrate at least through the basal layer of the epidermis. Clinical-translational studies enabled us to develop an immunohistochemical assay for caspase-3 activation, using biopsies from patients treated with topical Pc 4 in a Phase I PDT trial for cutaneous T-cell lymphoma. Results suggest that this assay may be used as an early biomarker of clinical response.
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Affiliation(s)
- Janine D. Miller
- Department of Dermatology, Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
- The Case Skin Diseases Research Center, Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
- The Case Comprehensive Cancer Center Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
| | - Elma D. Baron
- Department of Dermatology, Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
- The Case Skin Diseases Research Center, Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
- The Case Comprehensive Cancer Center Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
- Louis-Stokes VA Medical Center 10701 East Boulevard Cleveland, OH 44106
| | - Heather Scull
- Department of Dermatology, Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
- The Case Skin Diseases Research Center, Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
- The Case Comprehensive Cancer Center Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
| | - Andrew Hsia
- Department of Dermatology, Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
| | - Jeffrey C. Berlin
- Department of Dermatology, Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
- Department of Chemistry, Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
| | - Thomas McCormick
- Department of Dermatology, Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
- The Case Skin Diseases Research Center, Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
| | - Valdir Colussi
- Department of Radiation Oncology, School of Medicine, Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
| | - Malcolm E. Kenney
- Department of Dermatology, Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
- The Case Comprehensive Cancer Center Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
| | - Kevin D. Cooper
- Department of Dermatology, Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
- The Case Skin Diseases Research Center, Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
- The Case Comprehensive Cancer Center Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
- Louis-Stokes VA Medical Center 10701 East Boulevard Cleveland, OH 44106
| | - Nancy L. Oleinick
- Department of Radiation Oncology, School of Medicine, Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
- The Case Skin Diseases Research Center, Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
- The Case Comprehensive Cancer Center Case Western Reserve University/University Hospitals of Cleveland Cleveland, OH 44106
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Mitsunaga M, Tsubota A, Nariai K, Namiki Y, Sumi M, Yoshikawa T, Fujise K. Early apoptosis and cell death induced by ATX-S10Na (II)-mediated photodynamic therapy are Bax- and p53-dependent in human colon cancer cells. World J Gastroenterol 2007; 13:692-8. [PMID: 17278191 PMCID: PMC4066001 DOI: 10.3748/wjg.v13.i5.692] [Citation(s) in RCA: 17] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To investigate the roles of Bax and p53 proteins in photosensitivity of human colon cancer cells by using lysosome-localizing photosensitizer, ATX-S10Na (II).
METHODS: HCT116 human colon cancer cells and Bax-null or p53-null isogenic derivatives were irradiated with a diode laser. Early apoptosis and cell death in response to photodynamic therapy were determined by MTT assays, annexin V assays, transmission electron microscopy assays, caspase assays and western blotting.
RESULTS: Induction of early apoptosis and cell death was Bax- and p53-dependent. Bax and p53 were required for caspase-dependent apoptosis. The levels of anti-apoptotic Bcl-2 family proteins, Bcl-2 and Bcl-xL, were decreased in Bax- and p53-independent manner.
CONCLUSION: Our results indicate that early apoptosis and cell death of human colon cancer cells induced by photodynamic therapy with lysosome-localizing photosensitizer ATX-S10Na (II) are mediated by p53-Bax network and low levels of Bcl-2 and Bcl-xL proteins. Our results might help in formulating new therapeutic approaches in photodynamic therapy.
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Affiliation(s)
- Makoto Mitsunaga
- Institute of Clinical Medicine and Research, Jikei University School of Medicine, 163-1 Kashiwa-shita, Kashiwa, Chiba 277-8567, Japan
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Rocha-Viegas L, Vicent GP, Barañao JL, Beato M, Pecci A. Glucocorticoids repress bcl-X expression in lymphoid cells by recruiting STAT5B to the P4 promoter. J Biol Chem 2006; 281:33959-70. [PMID: 16959781 DOI: 10.1074/jbc.m602408200] [Citation(s) in RCA: 17] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/22/2023] Open
Abstract
The bcl-X gene plays a critical role in apoptosis. Six different isoforms generated by tissue-specific promoter usage and alternative splicing were described. Some of them exert opposite effects on cell death. In mammary epithelial cells glucocorticoids induce bcl-X expression and increase the ratio bcl-X(L) (antiapoptotic)/bcl-X(S) (apoptotic) by activating P4 promoter, which contains two hormone response elements. Here we show that, on mouse thymocytes and T lymphocyte derivative S49 cells, glucocorticoids inhibited transcription from P4 and decreased the ratio bcl-X(L)/bcl-X(S) favoring apoptosis. Upon hormonal treatment, glucocorticoid receptor (GR), steroid receptor coactivator-1, and RNA polymerase II were transiently recruited to P4 promoter, whereas STAT5B was also recruited but remained bound. Concomitant with the release of GR, silencing mediator for retinoic acid receptor and thyroid hormone receptor and histone deacetylase 3 were recruited, histone H3 was deacetylated, and RNA polymerase II left the promoter. Inhibition of STAT5 activity reverted glucocorticoid repression to activation of transcription and was accompanied by stable recruitment of GR and RNA polymerase II to P4.
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Affiliation(s)
- Luciana Rocha-Viegas
- Departamento de Fisiología, Biología Molecular y Celular, Instituto de Fisiología, Biología Molecular y Neurociencias-CONICET, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Argentina
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Abstract
When the mitochondria and/or the endoplasmic reticulum were targeted by photodynamic therapy, photodamage to the anti-apoptotic protein Bcl-2 was observed. This led to an apoptotic outcome if that death pathway was available. Lysosomal photodamage ultimately resulted in activation of the pro-apoptotic protein Bid, also leading to apoptosis. Photodamage to the plasma membrane was associated with migration of sensitizers to the cytosol and procaspase photodamage, with apoptosis impaired. Where apoptosis was unavailable because of lack of necessary components of the program, an autophagic outcome has been observed. It is also clear that autophagy can occur along with apoptosis as a PDT response, and may play a role in immunologic responses to photodamaged tumor cells.
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Affiliation(s)
- David Kessel
- Departments of Pharmacology and Medicine, Wayne State University School of Medicine, Detroit MI 48201, USA
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Crescenzi E, Chiaviello A, Canti G, Reddi E, Veneziani BM, Palumbo G. Low doses of cisplatin or gemcitabine plus Photofrin/photodynamic therapy: Disjointed cell cycle phase-related activity accounts for synergistic outcome in metastatic non–small cell lung cancer cells (H1299). Mol Cancer Ther 2006; 5:776-85. [PMID: 16546993 DOI: 10.1158/1535-7163.mct-05-0425] [Citation(s) in RCA: 61] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
We compared the effects of monotherapy (photodynamic therapy or chemotherapy) versus combination therapy (photodynamic therapy plus a specific drug) on the non-small cell lung cancer cell line H1299. Our aim was to evaluate whether the additive/synergistic effects of combination treatment were such that the cytostatic dose could be reduced without affecting treatment efficacy. Photodynamic therapy was done by irradiating Photofrin-preloaded H1299 p53/p16-null cells with a halogen lamp equipped with a bandpass filter. The cytotoxic drugs used were cis-diammine-dichloroplatinum [II] (CDDP or cisplatin) and 2',2'-difluoro-2'-deoxycytidine (gemcitabine). Various treatment combinations yielded therapeutic effects (trypan blue dye exclusion test) ranging from additive to clearly synergistic, the most effective being a combination of photodynamic therapy and CDDP. To gain insight into the cellular response mechanisms underlying favorable outcomes, we analyzed the H1299 cell cycle profiles and the expression patterns of several key proteins after monotherapy. In our conditions, we found that photodynamic therapy with Photofrin targeted G0-G1 cells, thereby causing cells to accumulate in S phase. In contrast, low-dose CDDP killed cells in S phase, thereby causing an accumulation of G0-G1 cells (and increased p21 expression). Like photodynamic therapy, low-dose gemcitabine targeted G0-G1 cells, which caused a massive accumulation of cells in S phase (and increased cyclin A expression). Although we observed therapeutic reinforcement with both drugs and photodynamic therapy, reinforcement was more pronounced when the drug (CDDP) and photodynamic therapy exert disjointed phase-related cytotoxic activity. Thus, if photodynamic therapy is appropriately tuned, the dose of the cytostatic drug can be reduced without compromising the therapeutic response.
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Affiliation(s)
- Elvira Crescenzi
- Dipartimento di Biologia e Patologia Cellulare e Molecolare L. Califano, Università di Napoli Federico II, Via S. Pansini 5, 80131 Naples, Italy
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38
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George JE, Ahmad Y, Varghai D, Li X, Berlin J, Jackowe D, Jungermann M, Wolfe MS, Lilge L, Totonchi A, Morris RL, Peterson A, Lust WD, Kenney ME, Hoppel CL, Sun J, Oleinick NL, Dean D. Pc 4 photodynamic therapy of U87-derived human glioma in the nude rat. Lasers Surg Med 2006; 36:383-9. [PMID: 15965990 DOI: 10.1002/lsm.20185] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/11/2022]
Abstract
BACKGROUND AND OBJECTIVES As a potential therapy for malignant glioma, we tested the phthalocyanine photosensitizer Pc 4 for: (1) rapid clearance from the vasculature, (2) specificity for glioma, and (3) tumoricidal photosensitizing capability. STUDY DESIGN/MATERIALS AND METHODS Parenchymal injection of U87 cells into athymic rat brains (N = 100) was followed after 12 days by tail vein injection of 0.5 mg/kg Pc 4. After 1 day, the tumor was illuminated with either 5 (N = 11) or 30 (N = 16) J/cm(2) red light at 672 nm. Sacrifice was 1 day later. The brains from these 27 animals underwent H&E (necrosis) and TUNEL assay (apoptosis) histology. Pc 4 concentration of explanted brains and tumors (N = 16), and all blood samples (N = 52) were determined by HPLC-MS 1 day post Pc 4 administration. RESULTS Tumor-specific apoptosis was almost uniformly seen; however, necrosis was found mostly in the high-light-dose group. Pc 4 concentration in bulk tumor averaged 3.8 times greater than in normal brain. CONCLUSIONS These results warrant expanding this pre-clinical study to seek effective baseline Pc 4 drug- and light-doses and infusion-to-photoirradiation timing that would be necessary for a Pc 4-mediated PDT clinical trial for glioma patients.
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Affiliation(s)
- John E George
- Department of Surgery, Case Western Reserve University, 10900 Euclid Avenue, Cleveland, Ohio 44106, USA
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Abstract
Singlet oxygen (1O2) is unique amongst reactive oxygen species formed in cells in that it is an excited state molecule with an inherent upper lifetime of 4 micros in water. Whether the lifetime of 1O2 in cells is shortened by reactions with cellular molecules or reaches the inherent maximum value is still unclear. However, even with the maximum lifetime, the diffusion radius is only approximately 220 nm during three lifetimes (approximately 5% 1O2 remaining), much shorter than cellular dimensions indicating that the primary reactions of 1O2 will be subcellularly localized near the site of 1O2 formation. This fact has raised the question of whether spatially resolved cellular responses to 1O2 occur, i.e. whether responses can be initiated by generation and reaction of 1O2 at a particular subcellular location that would not have been produced by 1O2 generation at other subcellular sites. In this paper, we discuss examples of spatially resolved responses initiated by 1O2 as a function of distance from the site of generation of 1O2. Three levels are recognized, namely, a molecular level where the primary oxidation product directly modifies the behavior of a cell, an organelle level where the initial photo-oxidation products initiate mechanisms that are unique to the organelle and the cellular level where mediators diffuse from their site of formation to the target molecules that initiate the response. These examples indicate that, indeed, spatially resolved responses to 'O2 occur in cells.
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Affiliation(s)
- Robert W Redmond
- Wellman Center for Photomedicine, Harvard Medical School, Massachusetts General Hospital, Boston, MA, USA.
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Xue LY, Chiu SM, Oleinick NL. Differential responses of Mcl-1 in photosensitized epithelial vs lymphoid-derived human cancer cells. Oncogene 2005; 24:6987-92. [PMID: 16007152 DOI: 10.1038/sj.onc.1208837] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
The antiapoptotic Bcl-2-family proteins, Bcl-2 and Bcl-xL, are recognized phototargets of photodynamic therapy (PDT) with the mitochondrion-targeting phthalocyanine photosensitizer Pc 4. In the present study, we found that myeloid cell leukemia 1 (Mcl-1), another antiapoptotic member of the Bcl-2 family, was not photodamaged in Pc 4-PDT-treated human carcinoma cells MCF-7c3, MDA-MB468, DU145, and A431, although Mcl-1 turnover was observed after exposure of HeLa or MCF-7c3 cells to a supralethal dose of UVC. In contrast, when human lymphoma U937 and Jurkat cells were treated with Pc 4-PDT, staurosporine (STS) or UVC, Mcl-1 was cleaved to generate a 28-kDa fragment over a 2-4 h period. The cleavage of Mcl-1 was accompanied by the activation of caspases-3, -9, and -8. The broad-specificity caspase inhibitor z-VAD-fmk completely blocked Mcl-1 cleavage induced by PDT, STS or UVC, providing evidence for Mcl-1 as a substrate for caspases. Western blot analysis localized Mcl-1 to mitochondria, ER, and cytosol of both MCF-7c3 and U937 cells, suggesting that Mcl-1 protein, unlike Bcl-2 and Bcl-xL, is not a target for Pc 4-PDT, probably due to its localization to sites removed from those of Pc 4 binding. The 28-kDa cleaved fragment of Mcl-1, which has proapoptotic activity, was produced in PDT-treated lymphoid-derived cells, but not in cells of epithelial origin, suggesting that PDT-induced rapid and extensive apoptosis in lymphoma cells may result in part from the sensitivity of their Mcl-1 to caspase cleavage, removing an important negative control on apoptosis.
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Affiliation(s)
- Liang-yan Xue
- Department of Radiation Oncology and the Case Comprehensive Cancer Center, School of Medicine, Case Western Reserve University, 10900 Euclid Avenue, Cleveland, OH 44106-4942, USA
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Lilge L, Pomerleau-Dalcourt N, Douplik A, Selman SH, Keck RW, Szkudlarek M, Pestka M, Jankun J. Transperineal in vivo fluence-rate dosimetry in the canine prostate during SnET2-mediated PDT. Phys Med Biol 2005; 49:3209-25. [PMID: 15357193 DOI: 10.1088/0031-9155/49/14/014] [Citation(s) in RCA: 18] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022]
Abstract
Advances in photodynamic therapy (PDT) treatment for prostate cancer can be achieved either by improving selectivity of the photosensitizer towards prostate gland tissue or improving the dosimetry by means of individualized treatment planning using currently available photosensitizers. The latter approach requires the ability to measure, among other parameters, the fluence rate at different positions within the prostate and the ability to derive the tissue optical properties. Here fibre optic probes are presented capable of measuring the fluence rate throughout large tissue volumes and a method to derive the tissue optical properties for different volumes of the prostate. The responsivity of the sensors is sufficient to detect a fluence rate of 0.1 mW cm(-2). The effective attenuation coefficient in the canine prostate at 660 nm is higher at the capsule (2.15+/-0.19 cm(-1)) than in proximity of the urethra (1.84+/-0.36 cm(-1)). Significant spatial and temporal intra- and inter-canine variability in the tissue optical properties was noted, highlighting the need for individualized monitoring of the fluence rate for improved dosimetry.
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Affiliation(s)
- Lothar Lilge
- Ontario Cancer Institute, Princess Margaret Hospital, Toronto, ON, Canada.
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42
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Castelli M, Reiners JJ, Kessel D. A mechanism for the proapoptotic activity of ursodeoxycholic acid: effects on Bcl-2 conformation. Cell Death Differ 2005; 11:906-14. [PMID: 15258617 DOI: 10.1038/sj.cdd.4401433] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/08/2022] Open
Abstract
Ursodeoxycholic acid (UDCA), a relatively nontoxic bile acid, enhanced the apoptotic response of tumor cells to both photosensitizers that cause photodamage to Bcl-2 and to the nonpeptidic Bcl-2/Bcl-x(L) antagonist HA14-1. The latter agent binds to the surface pocket formed by the BH1, BH2 and BH3 domains of Bcl-2 and Bcl-x(L). Fluorescence polarization studies indicated that affinity of HA14-1 for Bcl-2 was enhanced in the presence of UDCA. Moreover, Bcl-2 photodamage was promoted by UDCA using a photosensitizing agent with affinity for the endoplasmic reticulum, a site of Bcl-2 localization. Fluorescence resonance energy transfer (FRET) studies revealed that the proximity of Bcl-2 to a hydrophobic photosensitizing agent embedded in liposomes was enhanced by UDCA. Since photodamage will occur only if a protein is in close contact with a photosensitizing agent, we propose that these findings support the hypothesis that UDCA causes a conformational change in Bcl-2, promoting HA14-1 binding and enhancing affinity for certain membrane-bound photosensitizers.
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Affiliation(s)
- M Castelli
- Cancer Biology Program, Wayne State University School of Medicine, Detroit, MI 48201, USA
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43
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Liu W, Oseroff AR, Baumann H. Photodynamic therapy causes cross-linking of signal transducer and activator of transcription proteins and attenuation of interleukin-6 cytokine responsiveness in epithelial cells. Cancer Res 2004; 64:6579-87. [PMID: 15374971 DOI: 10.1158/0008-5472.can-04-1580] [Citation(s) in RCA: 49] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/03/2023]
Abstract
Photodynamic therapy (PDT) is a local treatment of cancers. The principle of PDT is the production of reactive oxygen species, in particular singlet oxygen, by light activation of a photosensitizer introduced into the target cells. The direct photochemical and subsequent redox reactions can lead to cell death. This study sought to identify effects occurring during PDT and some of their consequences in surviving cells. Using epithelial cells in tissue culture and in tumors, several distinct PDT-mediated reactions were found, including global dephosphorylation of proteins, induced phosphorylation of a 71-kDa protein, initiation of cellular stress responses, structural modification and loss of epidermal growth factor receptor, and cross-linking of proteins. Specific covalent cross-linking of nonactivated signal transducer and activator of transcription (STAT)-3, and to a lesser extent of STAT1 and STAT4, correlated with PDT dose. Cross-linked STAT3 was primarily localized to the cytoplasm and failed to bind to DNA. The combination of STAT cross-linking and inactivation of receptor functions rendered PDT-treated cells refractory for at least 24 hours to interleukin-6 and oncostatin M, cytokines known to be elevated at site of tissue damage and inflammation. It is suggested that the loss of responsiveness to these inflammatory cytokines in the PDT-treated field assists tumor cells in evading the growth-suppressive activity of these mediators expected to be present at tissue sites after PDT.
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Affiliation(s)
- Weiguo Liu
- Department of Molecular and Cellular Biology, Roswell Park Cancer Institute, Buffalo, New York 14263, USA
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Almeida RD, Manadas BJ, Carvalho AP, Duarte CB. Intracellular signaling mechanisms in photodynamic therapy. Biochim Biophys Acta Rev Cancer 2004; 1704:59-86. [PMID: 15363861 DOI: 10.1016/j.bbcan.2004.05.003] [Citation(s) in RCA: 107] [Impact Index Per Article: 5.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/14/2004] [Revised: 05/26/2004] [Accepted: 05/28/2004] [Indexed: 12/28/2022]
Abstract
In photodynamic therapy (PDT) a sensitizer, light and oxygen are used to induce death of tumor cells and in the treatment of certain noncancerous conditions. Cell death in PDT may occur by apoptosis or by necrosis, depending on the sensitizer, on the PDT dose and on the cell genotype. Some sensitizers that have been used in PDT are accumulated in the mitochondria, and this may explain their efficiency in inducing apoptotic cell death, both in vitro and in vivo. In this review we will focus on the events that characterize apoptotic death in PDT and on the intracellular signaling events that are set in motion in photosensitized cells. Activation of phospholipases, changes in ceramide metabolism, a rise in the cytosolic free Ca2+ concentration, stimulation of nitric oxide synthase (NOS), changes in protein phosphorylation and alterations in the activity of transcription factors and on gene expression have all been observed in PDT-treated cells. Although many of these metabolic reactions contribute to the demise process, some of them may antagonize cell death. Understanding the signaling mechanisms in PDT may provide means to modulate the PDT effects at the molecular level and potentiate its antitumor effectiveness.
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Affiliation(s)
- Ramiro D Almeida
- Center for Neuroscience and Cell Biology, Department of Zoology, University of Coimbra, Coimbra, 3004-517 Portugal
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Abstract
Reactive oxidizing species (ROS), such as hydrogen peroxide, nitric oxide, superoxide anion, and singlet oxygen, can damage cells as well as initiate responses such as new gene expression. The cell response evoked is strongly dependent on several factors. The subcellular location for formation of an ROS may be especially important for a highly reactive ROS, because it diffuses only a very short distance before reacting with a cellular molecule. How does a short-lived, locally acting ROS trigger responses at distant subcellular sites? This issue is discussed in light of a study of gene expression initiated by singlet oxygen, a ROS that exists for less than 100 ns in cells. Singlet oxygen is generated after a photosensitizer absorbs light energy and transfers the energy to molecular oxygen. To assess the role of singlet oxygen in light-induced gene expression in Arabidopsis, a mutant was used that accumulates the photosensitizer protochlorphyllide in chloroplasts. Three mechanisms are discussed that may connect the site of singlet oxygen formation to the signal transduction components: (i) direct oxidation of a signaling component by singlet oxygen, (ii) formation of oxidation products near the sites of singlet oxygen formation that diffuse to and react with signaling components, and (iii) alteration of the redox balance of the cell to a more oxidized state such that a greater proportion of a signaling pathway component is oxidized.
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Affiliation(s)
- Irene E Kochevar
- Wellman Center for Photomedicine, Massachusetts General Hospital, Harvard Medical School, Boston, MA 02114, USA.
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Chiu SM, Xue LY, Usuda J, Azizuddin K, Oleinick NL. Bax is essential for mitochondrion-mediated apoptosis but not for cell death caused by photodynamic therapy. Br J Cancer 2003; 89:1590-7. [PMID: 14562036 PMCID: PMC2394333 DOI: 10.1038/sj.bjc.6601298] [Citation(s) in RCA: 58] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022] Open
Abstract
The role of Bax in the release of cytochrome c from mitochondria and the induction of apoptosis has been demonstrated in many systems. Using immunocytochemical staining, we observed that photodynamic therapy (PDT) with the photosensitiser Pc 4 induced Bax translocation from the cytosol to mitochondria, and the release of cytochrome c from mitochondria as early signalling for the intrinsic pathway of apoptosis in human breast cancer MCF-7c3 cells. To test the role of Bax in apoptosis, MCF-7c3 cells were treated with Bax antisense oligonucleotides, which resulted in as much as a 50% inhibition of PDT-induced apoptosis. In the second approach, Bax-negative human prostate cancer DU-145 cells were studied. Following PDT, the hallmarks of apoptosis, including the release of cytochrome c from mitochondria, loss of mitochondrial membrane potential, caspase activation, and chromatin condensation and fragmentation, were completely blocked in these cells. Restoration of Bax expression in DU-145 cells restored apoptosis, indicating that the resistance of DU-145 cells to PDT-induced apoptosis is due to the lack of Bax rather than to another defect in the apoptotic machinery. However, despite the inhibition of apoptosis, the Bax-negative DU-145 cells were as photosensitive as Bax-replete MCF-7c3 cells, as determined by clonogenic assay. Thus, for Pc 4-PDT, the commitment to cell death occurs prior to Bax activation.
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Affiliation(s)
- S-M Chiu
- Department of Radiation Oncology and The CWRU/Ireland Comprehensive Cancer Center, School of Medicine (BRB-324), Case Western Reserve University, 10900 Euclid Avenue, Cleveland, OH 44106-4942, USA
| | - L-Y Xue
- Department of Radiation Oncology and The CWRU/Ireland Comprehensive Cancer Center, School of Medicine (BRB-324), Case Western Reserve University, 10900 Euclid Avenue, Cleveland, OH 44106-4942, USA
| | - J Usuda
- Department of Radiation Oncology and The CWRU/Ireland Comprehensive Cancer Center, School of Medicine (BRB-324), Case Western Reserve University, 10900 Euclid Avenue, Cleveland, OH 44106-4942, USA
| | - K Azizuddin
- Department of Radiation Oncology and The CWRU/Ireland Comprehensive Cancer Center, School of Medicine (BRB-324), Case Western Reserve University, 10900 Euclid Avenue, Cleveland, OH 44106-4942, USA
| | - N L Oleinick
- Department of Radiation Oncology and The CWRU/Ireland Comprehensive Cancer Center, School of Medicine (BRB-324), Case Western Reserve University, 10900 Euclid Avenue, Cleveland, OH 44106-4942, USA
- Department of Radiation Oncology and The CWRU/Ireland Comprehensive Cancer Center, School of Medicine (BRB-324), Case Western Reserve University, 10900 Euclid Avenue, Cleveland, OH 44106-4942, USA. E-mail:
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