Photodynamic Therapy of Cancer
`
`Photodynamic Therapy of Cancer: An Update
`
`Patrizia Agostinis, PhD1; Kristian Berg, PhD2; Keith A. Cengel, MD, PhD3; Thomas H. Foster, PhD4; Albert W. Girotti, PhD5;
`Sandra O. Gollnick, PhD6; Stephen M. Hahn, MD, PhD7; Michael R. Hamblin, PhD8,9,10; Asta Juzeniene, PhD11; David Kessel, PhD12;
`Mladen Korbelik, PhD13; Johan Moan, PhD14,15; Pawel Mroz, MD, PhD16,17; Dominika Nowis, MD, PhD18; Jacques Piette, PhD19;
`Brian C. Wilson, PhD20; Jakub Golab, MD, PhD21,22
`
`Abstract
`
`Photodynamic therapy (PDT) is a clinically approved, minimally invasive therapeutic procedure that can exert a selective
`cytotoxic activity toward malignant cells. The procedure involves administration of a photosensitizing agent followed by
`irradiation at a wavelength corresponding to an absorbance band of the sensitizer. In the presence of oxygen, a series
`of events lead to direct tumor cell death, damage to the microvasculature, and induction of a local inflammatory
`reaction. Clinical studies revealed that PDT can be curative, particularly in early stage tumors. It can prolong survival in
`patients with inoperable cancers and significantly improve quality of life. Minimal normal tissue toxicity, negligible
`systemic effects, greatly reduced long-term morbidity, lack of intrinsic or acquired resistance mechanisms, and excel-
`lent cosmetic as well as organ function-sparing effects of this treatment make it a valuable therapeutic option for com-
`bination treatments. With a number of recent technological improvements, PDT has the potential to become integrated
`into the mainstream of cancer treatment. CA Cancer J Clin 2011;61:250-281. VC 2011 American Cancer Society, Inc.
`
`Introduction
`
`Despite progress in basic research that has given us a better understanding of tumor biology and led to the
`design of new generations of targeted drugs, recent large clinical trials for cancer, with some notable excep-
`tions, have been able to detect only small differences in treatment outcomes.1,2 Moreover, the number of
`
`1Professor and Head of the Department of Molecular Cell Biology, Cell Death Research and Therapy Laboratory, Catholic University of Leuven, Leuven, Belgium;
`2Professor and Head of the Department of Radiation Biology, Institute for Cancer Research, The Norwegian Radium Hospital, Oslo University Hospital, Oslo,
`Norway; 3Assistant Professor of Radiation Oncology at the Department of Radiation Oncology, University of Pennsylvania, Philadelphia, PA; 4Professor of
`Imaging Sciences, Department of Imaging Sciences, University of Rochester, Rochester, NY; 5Professor of Biochemistry at the Department of Biochemistry,
`Medical College of Wisconsin, Milwaukee, WI; 6Professor of Oncology, Department of Cell Stress Biology, Roswell Park Cancer Institute, Buffalo, NY; 7Henry K.
`Pancoast Professor and Chairman of the Department of Radiation Oncology, University of Pennsylvania, Philadelphia, PA; 8Principal Investigator, Wellman
`Center for Photomedicine, Massachusetts General Hospital, Boston, MA; 9Associate Professor of Dermatology, Department of Dermatology, Harvard Medical
`School, Boston, MA; 10Associate Member of the Affiliated Faculty, Harvard-Massachusetts Institute of Technology Division of Health Sciences and Technology,
`Cambridge, MA; 11Postdoctoral Fellow at the Department of Radiation Biology, Institute for Cancer Research, The Norwegian Radium Hospital, Oslo University
`Hospital, Oslo, Norway; 12Professor of Pharmacology, Department of Pharmacology, Wayne State University School of Medicine, Detroit, MI; 13Distinguished
`Scientist, Integrative Oncology Department, British Columbia Cancer Agency, Vancouver, British Columbia, Canada; 14Senior Researcher at the Department of
`Radiation Biology, Institute for Cancer Research, The Norwegian Radium Hospital, Oslo University Hospital, Oslo, Norway; 15Professor of Physics, Group of
`Plasma and Space Physics, Institute of Physics, University of Oslo, Oslo, Norway; 16Assistant in Immunology, Wellman Center for Photomedicine, Massachusetts
`General Hospital, Boston, MA; 17Instructor in Dermatology, Department of Dermatology, Harvard Medical School, Boston, MA; 18Assistant Professor at the
`Department of Immunology, Center of Biostructure Research, Medical University of Warsaw, Warsaw, Poland; 19Director of GIGA-Research, Laboratory of
`Virology and Immunology, Professor at the University of Lie`ge, Lie`ge, Belgium; 20Head of the Division of Biophysics and Imaging, Ontario Cancer Institute,
`University of Toronto, Toronto, Ontario, Canada; 21Professor of Immunology and Head of the Department of Immunology, Center of Biostructure Research,
`Medical University of Warsaw, Warsaw, Poland; 22Professor of Immunology, Institute of Physical Chemistry, Polish Academy of Sciences, Warsaw, Poland.
`
`Corresponding author: Jakub Golab, MD, PhD, Department of Immunology, Center of Biostructure Research, Medical University of Warsaw, 1a Banacha St, F
`Building, 02-097 Warsaw, Poland; jakub.golab@wum.edu.pl
`
`Some of the figures were produced with the help of Abhishek Garg using Servier Medical Art (available at www.servier.com) for which we would like to
`acknowledge Servier.
`
`DISCLOSURES: Supported by the Fund for Scientific Research (FWO)-Flanders (Belgium)
`(grant numbers G.0661.09 and G.0728.10), the Interuniversity
`Attraction Pole IAP6/18 of the Belgian Federal Government, and the Catholic University of Leuven (OT/06/49 and GOA/11/009) (to P.A.); National Institutes of
`Health (NIH) grant CA-087971 (to K.A.C. and S.M.H.); NIH grants CA72630, CA70823, and HL85677 (to A.W.G.); NIH grants CA55791 and CA98156 (to S.O.G.);
`NIH grants CA68409 and CA122093 (to T.H.F.); NIH grants AI050875 and CA083882 (to M.R.H.); and the European Regional Development Fund through
`Innovative Economy grant POIG.01.01.02-00-008/08 (to J.G.). Dr. Kessel’s research has been supported by NIH grants since 1980, predominantly by CA23378.
`Dr. Juzeniene’ research has been supported by the Norwegian Cancer Society. Dr. Mroz was partly supported by a Genzyme-Partners Translational Research
`Grant. Dr. Golab is a recipient of the Mistrz Award from the Foundation for Polish Science and a member of the TEAM Programme cofinanced by the Foundation
`for Polish Science and the European Union European Regional Development Fund.
`
`VC 2011 American Cancer Society, Inc. doi:10.3322/caac.20114.
`
`Available online at http://cacancerjournal.org
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`TABLE 1. Glossary of Specialty Terms
`
`SPECIALTY TERM
`
`Chaperone
`
`Damage-associated molecular patterns (DAMPs)
`
`Fluence rate
`
`Fluorescence-guided resection
`
`Ground state
`
`Immunocompromised mice
`
`Immunocompetent mice
`
`Intersystem crossing
`
`Macromolecular therapeutics
`
`Major histocompatibility complex class I molecules
`
`Naı¨ve mice
`
`Pathogen-associated molecular patterns (PAMPs)
`
`Pattern-recognition receptors
`
`Photosensitizer
`
`Singlet oxygen (1O2)
`
`Triplet state
`
`CA CANCER J CLIN 2011;61:250–281
`
`DEFINITION
`
`A protein that participates in the folding of newly synthesized or
`unfolded proteins into a particular 3-dimensional conformation.
`
`Intracellular proteins that, when released outside the cell after its injury,
`can initiate or sustain an immune response in the noninfectious inflammatory response.
`
`The number of particles that intersect a unit area in a given amount of time
`(typically measured in watts per m2).
`
`A technique to enhance contrast of viable tumor borders that uses fluorescence emission
`from tissue. Fluorescence can be enhanced by the addition of exogenous chromophores
`(such as photosensitizers) with specific absorption and fluorescence properties.
`
`A state of elementary particles with the least possible energy in a physical system.
`This is the usual (singlet) state of most molecules. One of the exceptions includes oxygen,
`which in its ground state is a triplet and can be converted to a higher
`energy state of singlet oxygen during photodynamic therapy.
`
`Animals having an immune system that has been impaired by genetic modification,
`disease, or treatment.
`
`Animals having an intact (ie, normally functioning) immune system.
`
`A radiationless process in which a singlet excited electronic state makes a transition to
`a triplet excited state.
`
`Proteins such as antibodies and growth factors for cell surface targeting, peptides
`and mRNA for cancer vaccination, and nucleotides for gene delivery and silencing as well as
`drug moieties such as polymers and nanoparticles for the delivery of therapeutics.
`
`Transmembrane glycoproteins that bind short 8-11 amino acid long peptides
`recognized by T-cell receptors.
`
`Nonimmunized animals (ie, those that were not previously exposed to a particular
`antigen [such as tumor-associated antigen]).
`
`Evolutionary conserved microbial molecules that are not normally produced by
`mammalian cells and are often common to whole classes of micro-organisms. PAMPs are
`recognized by pattern-recognition receptors.
`
`Receptors for detection of DAMPs and PAMPs, initiating signaling cascades that trigger
`innate immune response.
`
`A light-absorbing compound that initiates a photochemical or photophysical reaction.
`
`An excited or energized form of molecular oxygen characterized by the opposite spin of a
`pair of electrons that is less stable and more reactive than the normal triplet oxygen (O2).
`
`A state of a molecule or a free radical in which there are 2 unpaired electrons.
`
`Ubiquitin-proteasome system
`
`The major intracellular pathway for protein degradation.
`
`new clinically approved drugs is disappointingly
`low.3 These sobering facts indicate that to make
`further progress, it is necessary to put an emphasis
`on other existing but still underappreciated thera-
`peutic approaches. Photodynamic therapy (PDT)
`has the potential to meet many currently unmet
`medical needs. Although still emerging,
`it
`is
`already
`a
`successful
`and
`clinically
`approved
`therapeutic modality used for the management of
`neoplastic and nonmalignant diseases. PDT was
`the first drug-device combination approved by the
`US Food and Drug Administration (FDA) almost
`2 decades ago, but even so remains underutilized
`clinically.
`
`PDT consists of 3 essential components: photo-
`sensitizer (PS) (see Table 1 for the definitions of spe-
`cialty terms), light, and oxygen.4,5 None of these is
`individually toxic, but together they initiate a photo-
`chemical reaction that culminates in the generation
`of a highly reactive product termed singlet oxygen
`(1O2) (Table 1). The latter can rapidly cause signifi-
`cant toxicity leading to cell death via apoptosis or ne-
`crosis. Antitumor effects of PDT derive from 3
`inter-related mechanisms: direct cytotoxic effects on
`tumor cells, damage to the tumor vasculature, and
`induction of a robust inflammatory reaction that can
`lead to the development of systemic immunity. The
`relative contribution of these mechanisms depends to
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`Photodynamic Therapy of Cancer
`
`FIGURE 1. The Principles of Photodynamic Therapy (PDT). A photosensitizer (PS) is administered systemically or topically. After a period of systemic PS
`distribution it selectively accumulates in the tumor. Irradiation activates the PS and in the presence of molecular oxygen triggers a photochemical reaction
`that culminates in the production of singlet oxygen (1O2). Irreparable damage to cellular macromolecules leads to tumor cell death via an apoptotic, necrotic,
`or autophagic mechanism, accompanied by induction of an acute local
`inflammatory reaction that participates in the removal of dead cells, restoration of
`normal tissue homeostasis, and, sometimes, in the development of systemic immunity.
`
`a large extent on the type and dose of PS used, the
`time between PS administration and light exposure,
`total light dose and its fluence rate (Table 1), tumor
`oxygen concentration, and perhaps other still poorly
`recognized variables. Therefore, determination of
`optimal conditions for using PDT requires a coordi-
`nated interdisciplinary effort. This
`review will
`address the most important biological and physico-
`chemical aspects of PDT, summarize its clinical
`status, and provide an outlook for its potential future
`development.
`
`Basic Components of PDT
`
`PDT is a 2-stage procedure. After the administra-
`tion of a light-sensitive PS, tumor loci are irradiated
`with a light of appropriate wavelength. The latter
`can be delivered to virtually any organ in the body by
`means of flexible fiber-optic devices (Fig. 1). Selec-
`tivity is derived from both the ability of useful PSs to
`localize in neoplastic lesions and the precise delivery
`of light to the treated sites. Paradoxically, the highly
`localized nature of PDT is one of
`its current
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`CA CANCER J CLIN 2011;61:250–281
`
`ineffective
`is
`limitations, because the treatment
`against metastatic lesions, which are the most fre-
`quent cause of death in cancer patients. Ongoing
`research is focused on finding optimal PDT condi-
`tions to induce systemic immunity that might, at
`least to some extent, obviate this limitation in the
`future. PDT can be used either before or after chem-
`otherapy, radiotherapy, or surgery without compro-
`mising these therapeutic modalities. None of the
`clinically approved PSs accumulate in cell nuclei,
`limiting DNA damage that could be carcinogenic or
`lead to the development of resistant clones. More-
`over, the adverse effects of chemotherapy or radia-
`tion are absent. Radioresistance or chemoresistance
`do not affect sensitivity to PDT. Excellent cosmetic
`outcomes make PDT suitable for patients with skin
`cancers. There are no significant changes in tissue
`temperature, and the preservation of connective tis-
`sue leads to minimal fibrosis, allowing retention of
`functional anatomy and mechanical integrity of hol-
`low organs undergoing PDT. Selected patients with
`inoperable tumors, who have exhausted other treat-
`ment options, can also achieve improvement in qual-
`ity of life with PDT. Finally, many PDT procedures
`can be performed in an outpatient or ambulatory set-
`ting, thereby not only alleviating costs, but also mak-
`ing the treatment patient-friendly. The only adverse
`effects of PDT relate to pain during some treatment
`protocols and a persistent skin photosensitization
`that has been circumvented by the newer agents.
`
`Photosensitizers
`Most of the PSs used in cancer therapy are based on
`a tetrapyrrole structure, similar to that of the proto-
`porphyrin contained in hemoglobin. An ideal PS
`agent should be a single pure compound to allow
`quality control analysis with low manufacturing costs
`and good stability in storage. It should have a high
`absorption peak between 600 and 800 nanometers
`(nm)
`(red to deep red), because absorption of
`photons with wavelengths longer than 800 nm does
`not provide enough energy to excite oxygen to its
`singlet state and to form a substantial yield of reactive
`oxygen species. Because the penetration of light into
`tissue increases with its wavelength, agents with
`strong absorbance in the deep red such as chlorins,
`bacteriochlorins, and phthalocyanines offer improve-
`ment in tumor control. It should have no dark
`
`toxicity and relatively rapid clearance from normal
`tissues, thereby minimizing phototoxic side effects.
`Other pertinent desirable properties of PS agents
`have been summarized elsewhere.6 Although the
`interval between drug administration and irradiation
`is usually long, so that the sensitizer is given sufficient
`time to diffuse from normal tissues, reports now
`suggest that the tumor response may be sometimes
`better when light is delivered at a shorter drug-light
`interval when PS is still present in the blood vessels,
`thus producing marked vascular damage.7 Some
`reports
`suggest
`that a pronounced inflammatory
`response and necrotic cell death after illumination are
`important in the immune-stimulating function of
`PDT, whereas others suggest that PSs that produce
`more apoptosis and less inflammation are suitable for
`applications such as brain tumors, where swelling is
`undesirable. Recent findings show that certain PDT-
`induced apoptotic cell death mechanisms are highly
`immunogenic and capable of driving antitumor
`immunity as well.8 Finally,
`the light-mediated
`destruction of the PS known as photobleaching was
`previously thought to be undesirable, but some reports
`suggest that this property may make light dosimetry
`less critical because overtreatment is avoided when
`the PS is destroyed during the illumination.9
`The first PS to be clinically employed for cancer
`therapy was a water-soluble mixture of porphyrins
`called hematoporphyrin derivative (HPD), a purified
`form of which, porfimer
`sodium,
`later became
`known as Photofrin. Although porfimer sodium is
`still the most widely employed PS, the product has
`some disadvantages,
`including a long-lasting skin
`photosensitivity and a relatively low absorbance at
`630 nm. Although a photodynamic effect can be
`produced with porfimer sodium, efficacy would be
`improved by red-shifting the red absorbance band
`and increasing the absorbance at the longer wave-
`lengths. There has been a major effort among
`medicinal chemists to discover second-generation
`PSs, and several hundred compounds have been
`proposed as potentially useful for anticancer PDT.
`Table 2 displays the most promising PSs that have
`been used clinically for
`cancer PDT (whether
`approved or in trials). The discovery that 5-aminole-
`vulinic acid (ALA) was a biosynthetic precursor of the
`PS protoporphyrin IX10 has led to many applications
`in which ALA or ALA esters can be topically applied
`or administered orally. These are considered to be
`‘‘prodrugs,’’ needing to be converted to protoporphyrin
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`Photodynamic Therapy of Cancer
`
`TABLE 2. Clinically Applied Photosensitizers
`
`PHOTOSENSITIZER
`
`STRUCTURE
`
`Porfimer sodium (Photofrin) (HPD)
`
`Porphyrin
`
`ALA
`
`ALA esters
`
`Temoporfin (Foscan) (mTHPC)
`
`Verteporfin
`
`HPPH
`
`SnEt2 (Purlytin)
`
`Talaporfin (LS11, MACE, NPe6)
`
`Ce6-PVP (Fotolon), Ce6 derivatives
`(Radachlorin, Photodithazine)
`
`Porphyrin
`precursor
`
`Porphyrin
`precursor
`
`Chlorine
`
`Chlorine
`
`Chlorin
`
`Chlorin
`
`Chlorin
`
`Chlorin
`
`Silicon phthalocyanine (Pc4)
`
`Phthalocyanine
`
`Padoporfin (TOOKAD)
`
`Bacteriochlorin
`
`Motexafin lutetium (Lutex)
`
`Texaphyrin
`
`WAVELENGTH,
`nm
`
`APPROVED
`
`TRIALS
`
`CANCER TYPES
`
`630
`
`635
`
`635
`
`652
`
`690
`
`665
`
`660
`
`660
`
`660
`
`675
`
`762
`
`732
`
`Worldwide
`
`Worldwide
`
`Europe
`
`Lung, esophagus, bile duct, bladder, brain, ovarian
`
`Skin, bladder, brain, esophagus
`
`Skin, bladder
`
`Europe
`
`United States
`
`Head and neck, lung, brain, skin, bile duct
`
`Worldwide
`(AMD)
`
`United
`Kingdom
`
`Ophthalmic, pancreatic, skin
`
`United States
`
`Head and neck, esophagus, lung
`
`United States
`
`Skin, breast
`
`United States
`
`Liver, colon, brain
`
`Belarus, Russia
`
`Nasopharyngeal, sarcoma, brain
`
`United States
`
`Cutaneous T-cell lymphoma
`
`United States
`
`Prostate
`
`United States
`
`Breast
`
`Abbreviations: ALA, 5-aminolevulinic acid; AMD, age-related macular degeneration; Ce6-PVP, chlorin e6-polyvinypyrrolidone; HPD, hematoporphyrin derivative;
`HPPH, 2- (1-hexyloxyethyl)-2-devinyl pyropheophorbide-a; MACE, mono-(L)-aspartylchlorin-e6; mTHPC, m-tetrahydroxyphenylchlorin; nm indicates nanometers;
`SnEt2, tin ethyl etiopurpurin.
`
`to be active PSs. Many hypotheses have been proposed
`to account
`for the tumor-localizing properties in
`PDT.11 These include the preponderance of leaky and
`tortuous tumor blood vessels due to neovascularization
`and the absence of lymphatic drainage known as the
`enhanced permeability and retention effect.12 Some of
`the most effective compounds bind preferentially to
`low-density lipoprotein (LDL), suggesting that upreg-
`ulated LDL receptors found on tumor cells could be
`important.13
`There have been targeting studies in which PSs
`are covalently attached to various molecules that have
`some affinity for neoplasia or to receptors expressed
`on specific tumors.14 The intention is to rely on the
`ability of the targeting vehicle to control localization
`factors so that the PS can be chosen based on its
`photochemical properties. These vehicles include
`monoclonal antibodies, antibody fragments, peptides,
`proteins (such as transferrin, epidermal growth factor
`and insulin), LDL, various carbohydrates, somatosta-
`tin, folic acid, and many others.
`
`Light Sources
`Blue light penetrates least efficiently through tissue,
`whereas red and infrared radiations penetrate more
`deeply (Fig. 2). The region between 600 and 1200 nm
`
`tissue.
`is often called the optical window of
`However, light up to only approximately 800 nm can
`generate 1O2, because longer wavelengths have
`insufficient energy to initiate a photodynamic reac-
`tion.15 No single light source is ideal for all PDT
`indications, even with the same PS. The choice of
`light source should therefore be based on PS absorp-
`tion (fluorescence excitation and action spectra), dis-
`ease (location, size of lesions, accessibility, and tissue
`characteristics), cost, and size. The clinical efficacy
`of PDT is dependent on complex dosimetry: total
`light dose, light exposure time, and light delivery
`mode (single vs fractionated or even metronomic).
`The fluence rate also affects PDT response.16 Inte-
`grated systems that measure the light distribution
`and fluence rate either interstitially or on the surface
`of the tissues being treated are so far used only in
`experimental studies.
`Both lasers and incandescent light sources have
`been used for PDT and show similar efficacies.17
`Unlike the large and inefficient pumped dye lasers,
`diode lasers are small and cost-effective, are simple
`to install, and have automated dosimetry and calibra-
`tion features and a longer operational
`life. Such
`lasers are now being specifically designed for PDT.
`Light-emitting diodes (LEDs) are alternative light
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`CA CANCER J CLIN 2011;61:250–281
`
`FIGURE 2. Light Propagation Through the Tissues.
`
`in being a triplet in its
`ground state. This step
`leads to the formation
`of 1O2, and the reac-
`tion is referred to as a
`Type II process.23 A
`Type
`I
`process
`can
`also occur whereby the
`PS reacts directly with
`an organic molecule in
`a
`cellular microenvir-
`onment,
`acquiring
`a
`hydrogen atom or elec-
`tron to form a radical.
`Subsequent
`autoxida-
`tion of the reduced PS
`produces a superoxide
`
`anion
`radical
`(O
`2 ).
`or
`Dismutation
`one-
`electron
`reduction
`of
`
`O
`gives
`hydrogen
`2
`peroxide (H2O2), which
`in turn can undergo
`one-electron
`reduction
`to a powerful and virtually indiscriminate oxidant
`
`hydroxyl
`radical
`(HO
`). Reactive oxygen species
`(ROS) generation via Type II chemistry is mecha-
`nistically much simpler than via Type I, and most
`PSs are believed to operate via a Type II rather
`than Type I mechanism.
`
`Mechanisms of PDT-Mediated Cytotoxicity
`The lifetime of 1O2 is very short (approximately
`10-320 nanoseconds),
`limiting its diffusion to
`only approximately 10 nm to 55 nm in cells.24
`Thus, photodynamic damage will occur very close
`to the intracellular location of the PS.25 Porfimer
`sodium is a complex mixture of porphyrin ethers
`with variable localization patterns mostly associ-
`ated with lipid membranes. Of
`the other PS
`agents in current use, the mono-L-aspartyl chlorin
`e6
`(NPe6,
`talaporfin)
`targets
`lysosomes;
`the
`benzoporphyrin derivative (BPD)
`targets mito-
`chondria; m-tetrahydroxyphenylchlorin (mTHPC,
`temeporfin) has been reported to target mitochon-
`dria, endoplasmic reticulum (ER), or both; and
`the phthalocyanine Pc4 has a broad spectrum of
`affinity, although mitochondria are reported to be
`a primary target.6 Other agents that have been
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`sources with relatively narrow spectral bandwidths
`and high fluence rates.18,19 Lasers can be coupled
`into fibers with diffusing tips to treat tumors in the
`urinary bladder and the digestive tract. Inflatable
`balloons, covered on the inside with a strongly scat-
`tering material and formed to fit an organ, are also
`commercially available.20 It
`is quite feasible to
`implant a light source in solid organs deep in the
`body under image guidance. The choice of optimal
`combinations of PSs, light sources, and treatment
`parameters is crucial for successful PDT.21,22
`
`Photophysics and Photochemistry
`Most PSs in their ground (ie, singlet) state (Table 1)
`have 2 electrons with opposite spins located in an
`energetically most
`favorable molecular orbital.
`Absorption of light leads to a transfer of one electron
`to a higher energy orbital (Fig. 3). This excited PS is
`very unstable and emits this excess energy as fluores-
`cence and/or heat. Alternatively, an excited PS may
`undergo an intersystem crossing (Table 1) to form a
`more stable triplet state (Table 1) with inverted spin
`of one electron. The PS in triplet state can either
`decay radiationlessly to the ground state or transfer
`its energy to molecular oxygen (O2), which is unique
`
`

`

`Photodynamic Therapy of Cancer
`
`FIGURE 3. Photosensitization Processes Illustrated by a Modified Jablonski Diagram. Light exposure takes a
`photosensitizer molecule from the ground singlet state (S0) to an excited singlet state (S1). The molecule in S1
`may undergo intersystem crossing to an excited triplet state (T1) and then either form radicals via a Type I
`reaction or, more likely, transfer its energy to molecular oxygen (3O2) and form singlet oxygen (1O2), which is
`the major cytotoxic agent involved in photodynamic therapy. ns indicates nanoseconds; ls, microseconds; nm,
`nanometers; eV, electron volts.
`
`Ca2þ
`overload could pro-
`mote mitochondrial per-
`meability
`transition,
`an
`event that may favor ne-
`crotic rather than apoptotic
`phototoxicity.26,35
`cells
`of
`Photodamage
`can also lead to the stimu-
`lation of macroautophagy
`(hereafter
`referred to as
`autophagy).36,37 This is a
`lysosomal pathway for the
`degradation and recycling
`of intracellular proteins and
`organelles. Autophagy can
`be stimulated by various stress signals including
`oxidative stress.38 This process can have both a
`cytoprotective and a prodeath role after cancer
`chemotherapies, including those involving ROS as
`primary damaging agents.38 Recent studies delineate
`autophagy as a mechanism to preserve cell viability
`after photodynamic injury.37 PSs that photodamage
`the
`lysosomal
`compartment may
`compromise
`completion of
`the autophagic process,
`causing
`incomplete clearance of
`the autophagic
`cargo.
`Accumulation
`of ROS-damaged
`cytoplasmic
`components may then potentiate phototoxicity in
`apoptosis-competent cells.37 A better understanding
`of the interplay between autophagy, apoptosis, and
`necrosis and how these processes lead to improved
`tumor response will be a requisite to devise better
`therapeutic strategies in PDT.
`
`Cytoprotective Mechanisms
`Numerous publications have reported cytoprotective
`mechanisms that cancer cells exploit to avoid the
`cytotoxic effects of PDT.26 The first mechanism
`identified was based on the large variation observed
`in the level of antioxidant molecules expressed in
`cancer cells.39 Both water-soluble antioxidants (eg,
`some amino acids, glutathione [GSH], or vitamin
`C) and lipid-soluble antioxidants (eg, vitamin E) are
`present at variable levels in many cancer cell types,
`explaining the large variation in PDT sensitivity.40
`A second mechanism is associated with expression in
`cancer cells of enzymes that can detoxify ROS.
`Although there is no specific cellular enzyme that
`can directly detoxify 1O2, enzymes involved in other
`ROS metabolism can influence the cytotoxic effect
`
`targets. Specific
`developed can have multiple
`patterns of localization may vary also among dif-
`ferent cell types.
`PDT can evoke the 3 main cell death pathways:
`apoptotic, necrotic, and autophagy-associated cell
`death (Fig. 4). Apoptosis is a generally major cell
`death modality
`in cells
`responding to PDT.
`Mitochondria outer membrane permeabilization
`(MOMP) after photodynamic injury is controlled by
`Bcl-2 family members and thought to be largely
`p53-independent.26 With mitochondria-associated
`PSs, photodamage to membrane-bound Bcl-227-29
`can be a permissive signal for MOMP and the
`subsequent release of caspase activators such as cyto-
`chrome c and Smac/DIABLO, or other proapop-
`totic molecules, including apoptosis-inducing factor
`(AIF).26 Lysosomal membrane rupture and leakage
`of cathepsins from photo-oxidized lysosomes30,31
`induces Bid cleavage and MOMP.31
`Phototoxicity is not propagated only through cas-
`pase signaling but involves other proteases, such as
`calpains, as well as nonapoptotic pathways.26 Typi-
`cally,
`inhibition or genetic deficiency of caspases
`only delays phototoxicity or shifts the cell death mo-
`dality toward necrotic cell death.32 Recent evidence
`suggests indeed that certain forms of necrosis can be
`propagated through signal transduction pathways.33
`The molecular mechanisms underlying programmed
`necrosis are still elusive, but certain events including
`activation of receptor interacting protein 1 (RIP1)
`kinase, excessive mitochondrial ROS production,
`lysosomal damage, and intracellular Ca2þ
`overload
`are recurrently involved.33,34 Severe inner mito-
`chondria membrane photodamage or intracellular
`
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`
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`CA CANCER J CLIN 2011;61:250–281
`
`FIGURE 4. Three Major Cell Death Morphotypes and Their Immunological Profiles. Apoptosis is morphologically
`characterized by chromatin condensation, cleavage of chromosomal DNA into internucleosomal fragments, cell
`shrinkage, membrane blebbing, and the formation of apoptotic bodies without plasma membrane breakdown.
`Typically, apoptotic cells release ‘‘find me’’ and ‘‘eat me’’ signals required for the clearance of the remaining corpses
`by phagocytic cells. At the biochemical level, apoptosis entails the activation of caspases, a highly conserved family of
`cysteine-dependent, aspartate-specific proteases. Necrosis is morphologically characterized by vacuolization of the
`cytoplasm and swelling and breakdown of the plasma membrane, resulting in an inflammatory reaction due to the
`release of cellular contents and proinflammatory molecules. Classically, necrosis is thought to be the result of
`pathological insults or to be caused by a bioenergetic catastrophe, adenosine triphosphate (ATP) depletion to a level
`incompatible with cell survival. The biochemistry of necrosis is characterized mostly in negative terms by the absence
`of caspase activation, cytochrome c release, and DNA oligonucleosomal fragmentation. Autophagy is characterized by
`a massive vacuolization of the cytoplasm. Autophagic cytoplasmic degradation requires the formation of a double-
`membrane structure called the autophagosome, which sequesters cytoplasmic components as well as organelles and
`traffics them to the lysosomes. Autophagosome-lysosome fusion results in the degradation of the cytoplasmic
`components by the lysosomal hydrolases. In adult organisms, autophagy functions as a self-digestion pathway
`promoting cell survival
`in an adverse environment and as a quality control mechanism by removing damaged
`organelles, toxic metabolites, or intracellular pathogens. DAMPs indicates damage-associated molecular patterns;
`HSPs, heat shock proteins; HMGB1, high-mobility group protein B1;
`IL,
`interleukin; ATP/MSU, adenosine
`triphosphate/monosodium urate.
`
`(HO-1) expression, and the
`mechanism is dependent on
`Nrf2 nuclear accumulation
`and
`on
`p38 mitogen-
`activated
`protein
`kinase
`(p38MAPK)
`and phospho-
`inositide 3-kinase
`(PI3K)
`activities. Because of
`the
`antioxidant activity of HO-
`1, it can be envisioned that
`Nrf2-
`dependent
`signal
`transduction
`can
`control
`cellular protection against
`PDT-mediated
`cytotoxic
`effects.
`PDT was found to induce
`expression of various heat
`shock proteins (HSPs) for
`which a protective role in
`PDT has been described.
`For example, transfection of
`tumor cells with the HSP27
`gene increased the survival
`of tumor cells after PDT.45
`Similarly, increased HSP60
`and HSP70 levels are inver-
`sely correlated with sensitiv-
`ity
`to the photodynamic
`treatment.46,47 The simplest
`explanation for these observations is the ability of
`HSPs to bind to oxidatively damaged proteins.
`Moreover, the intracellular function of HSPs is not
`only restricted to protein refolding. Many HSPs ‘‘cli-
`ent’’ proteins play a critical role in the regulation of
`prosurvival pathways. PDT also leads to increased
`ubiquitination of carbonylated proteins, thereby tag-
`ging them for degradation in proteasomes, which
`prevents the formation of toxic protein aggregates.48
`
`Antivascular Effects of PDT
`
`Photodynamic perturbation of tissue microcircula-
`tion was first reported in 1963.49 A study by Star
`et al50 utilized a window chamber to make direct
`observations of implanted mammary tumor and adja-
`cent normal tissue microcirculation in rats before,
`during, and at various times after PDT sensitized
`with HPD. An initial blanching and vas