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Journal of Clinical Laser Medicine & Surgery Volume 11, Number 5, 199.3 Mary Ann Liebert, Inc., Publishers pp.233-241 Second Generation Photodynamic Agents: A Review By ETHAN D. STERNBERG. PH .D. and DAVID DOLPHIN, PH.D. ABSTRACT Over the last decade, laser treatment of neoplastic diseases bas become routine. The ability of these light~ induced therapies to effect positive results is increased with the utilization of photosensitizing dyes. The approval of Pbotofrin® in Canada as a first generation photodynamic therapeutic agent for the treatment of some forms of bladder cancer is being followed by the development of other agents with improved properties. At this time a number of second generation photosensitizing dyes are under study in phase IIII clinical trials. A review of the status of these trials along with mechanistic aspects is reviewed in this article. In addition, a review of the status of lasers to be utilized for photodynamic therapy gives some indication of which instru· ments could be considered for this therapy in the future. INTRODUCTION P hotodynamic therapy (PDT) is a therapeutic modality that utilizes a photosensitizer and visible light to create a roxic environment to biological systems. PDT is utJIized in fields ranging from cancer therapy to viral inactivation of solid tumors and the treatment of leukemia (Fig. 1). The historical perspec~ tive of this methodology in the field of cancer is not within the domain of this review but has been reviewed elsewhere.! We will endeavor to enlighten the reader on the mechanistic aspects of the phototoxic effect, show how these relate to different therapeutic protocols, and review some aspects of recent clini~ cal trials for both the first and second generation compounds being studied at II number of clinical sites throughout the world. Photofrin® is the first generation compound for PDT and has become most familiar to the readers of thi~ joumal. 2 It consists of an oligomeric mixture of hematoporphyrin, a nonmetallaled derivative of the porphyrin found in hemoglobin (Fig. 2). This drug has undergone clinical trials throughout the world and hs!. been found to be effective in treating a number of cancers. ~ Trials for the therapy have been instituted in a number of hospi~ tals throughout the world. Quadra Logic Technologies in partnership with Lederle Laboratories has recently filed a new drug application (NOA) in a number of countries including Japan, Belgium, and Denmark and will have filed in the rest of Europe and the United States by the time of pUblication of this review. A Notice of CompHance from the Canadian Health Protection Branch was received in April J993 for Ihe lise of Photofrin® in the treatmenf of superficial bladder cancer. This was a very important landmark since it was the first approval for PDT anywhere in the world. Photofrin® is activated at 630 nm using an argon pumped dye laser. This wavelength is chosen since the strong absorptions due to oxy· and deoxyhemoglohin in the visible region become weaker than (he longest wavelength absorption band of Photof4 rin®. As the absorption of the natural chromophores (principally the hemoglobins) decreases then the effective depth of penetration increases with increasing wavelength (Fig. 3). In addition to absorption, scattering of photons also limits the effective penetration depth. In practice. light at 690 nm travels twice as deep as that at 630 nm. On moving from 690 nm into the near infrared (~OO "m) only an additional JO% effective depth of penetration is achieved. Clearly then, second generation photosensitizers will be most effective when they absorb and can be activated at wavelenglhs longer than that used to activate Photofrin e (630 om). In addition. successful second generation photosensitizers must exhibit Jess prolonged skin photosensitivity than seen with Photofrin®. where a patient's skin may remain photosensitive to strong sunlight for up to 4-6 weeks. s The side effects, small in comparison fo those seen with chemotherapy or radiation therapy, could limit the use of Photofrin® in non~life threatening indications. Department of Chemisb'y, University of British Columbia, Vancouver, B.C., Canada and Quadra Logic Technologies Inc., Vancouver, British Columbia, Canada. 233 Sternberg and Dolpbin 234 A c D FIG. J. Feline T cells were treated with 2 ~gJml of BPDMA and .20 J/cm 2 of light. (A) Normal cells. (8) Initial damage on close inspection shows small holes. (C) These holes increase in size after time. (D) Cell membranes arc complclcly nlptured. Reprinted with permission from North cl a1. 49 Blood Cells 18: 129-[40. THE PHOTODYNAMIC EFJ:1'"ECT The processes hy which light creates a photoloxic effect in the presence of a dye are still under debate . There are two categories of energy transfer that may occur.t. The first step for both of these is absorption of light by a dye. Typically the dye is and n= an extended aromatic system such as rnerocyanine, methylene hlue, or a variety of porphyrin derivatives (Fig. 4) . An excited stale of the dye is generated that can interact with a biomolecule via an ~Iectron transfer mechanism reSUlting in the destruction of the hiomolecule and the bleaching of the dye (Fig. 5) . This is known as a Type I photo process and the foonation of radicals during such proce.c;ses may result in additional damage due to ~ub,<;equent radical chain or oxidation reactions . A more 550nm 630nm 700nm FIG. 3. FIG. 2. Photofrin® is a purified fraction of oligomers of he- matoporphyrin. 800nm 0-7 Relative effective light dose versus wavelength (nm). Light pencrratcs tissue better in region!>. near the inti·ared. Only a moot!rate increase is found ill going from 700 to 800 nm. Second Generation Photodynamic Agents 23S ~ CH,3SCH2CH2cHCR ~HR 102 FIG. 4. (top) Methyle~ blue; (middle) merocyanine; (/JOItom) tetraphenylporphyrin. interesting process occu~ when the energetically exciled chromophore. after imersystem crossing from the singlet excited state 10 the triplet slale, interacts with ground state triplet oxygen. This interaction results in the fonnation of singlet oxygen e00 and the dye returns to it." original singlet ground state ready to absorb another photon and to initiate another catalytic cycle and the fOllTlation of more singlet oxygen. Although singlet oKygen has a short lifetime (approximately 6 ~sec in water and a little longer in cell membranes), it has the ability to readily react with a wide variety of biomolecules as shown in Fig. 6. The~ latter reactions can cause the destruction of vari.~ ous biological components and shut down numerous biological processes. Singlet oxygen is generated catalytically and the rltle of singlet oxygen generation depends on some of the crileria described below. In the case of several phototoxins the quantum yield for singlet oxygen production is around 0.7. 7 Three criteria determine the efficacy of a sensitizing dye, in 11 non- + hv , -¢: Sensitizer 9 I e ~ 2 10 2 biological system. The molecular extinction coefficient (E.) measures the ability of a sensitizer to absorb light falling on it at a specific wavelength effectively. The quantum yield of bleach~ ing of the pholo~ensitizer detemlines its lifetime when undergo~ ing a Type I photo process, and the quantum yield for singlet oxygen production measures the ratio for the number of photons absorbed to the number of singlet oxygen molecules produced. These three criteria are relatively easy values to measure, how~ ever. relating these parameters to in vitro and especially in vivo cyloloxicity is difficult. The environment of a phototoxin not only determines which biological systems nre most disrupted by PDT but also determines the efficiency of the sensitizer, becau~ of photobleach~ iog and the photo properties of the tissue. This synergism complicates the biologjcal studies in that it is possible that the site of the greatest concentration of the dye may not define the site of cell death. Thus the development of dyes for PDT differs from the classical development of pharmaceutical compounds where one hopes to find a drug that interacts with a specific binding site and then further modify the drug to interact more effectively with that binding site. 8 + e •e Sensitizer e FIG. 6. Biomolecules such as cholesterol (A). methionine (8), guanidine (C), and histidine (D) give a number of products ranging from hydropcroxides to sulfoxides. .61 Sensitizer .3 Sens itizer· 30 • Type I Type II FIG. S. Two types of reactions can occur from the photoex.cited state of a sensitizer 5 . They may react directly with a substrate via electron transfer Hnd bleach (Type 1). or undergo intersystem crossing, react with oxygen, and generate singlet oxygen. PHOTOTOXJNS IN BIOLOGY Many photodynamic agents have been investigated for their abilities to induce a phototox.ic effect. Compounds that are near or currently in clinical trials range from a totally synthetic tetrahydroxytetraphcnylchlorin (1) developed by Bonnell and Bercnbawn9 (tnd zinc phalocyanine (3) being used by ClBA~ GcigyJO to the modified n~tural product monoaspartylchlorin eo (4) being investigated by Nippon Petrochemical, II tin etiopurpunn (2) synthesized by Morgan et aI., 12 and benzoporphyrin derivative monoacid (BPOMA) (5), which we at QLT and Sternberg and Dolphin 236 CI HO~"''' " CO:;!CH.:s "~ 01< 1 3 "' 'iHH H H 01< !:02h CO',zH FIG. 7. 4 700 Various photosensitizers in, or approaching clinical FIG. 8. Four analogues of sulfonated phthalocyanines are strucLurdlly similar but have significant differences in phototoxicity. (A) X = (H.H,H,SO,H); (B) X = (H,H,SOJH,SO)H); (C) X = (H,SO,\H,S03H,SO.,H); (D) X = (SO)H,SO.,H, trials. SO]H,SOlH). Ledene Laborutories are developing l3 (Fig. 7). All of these compounds have absorption maxima beyond 650 nm and ex.~ tinction coefficient at their maximal long wavelength abso~ tion at least IO-foJd over that of Photofrin® at 630 om. Figure 7 illustrates the near red absorption maxima of several of these newer photosensitizers. Two sites of destrucLion are generally acknowledged to be important to the in vitro damage 10 cells. TIle cell membrane is susceptible to a number of as~ults by singlet oxygen. 14 Proteins within the membrane, on PDT treatment. may be modified and have their ability to channel ions disrupted. Although lipid peroxidation can occur routinely on PDT the changes in membrane permeability in models such as red hlood cells seem to be associated with damage to proteins . l~ Memhranes within the cell such as those associated with the Iysosomes have also been suggested as sites of cell destruction. A further site of destruction is associated with mitochondria . 16 Isolated mitochondria have sbown that photosensitization by Photofrin® created a loss of oxidative phosphorylation that could be traced to oxidation of thiols, in carrier proteins, by singlet oxygen. 17 The modit1cation of other enzymatic processes has also been observed in vitro. For instance, aldehyde-3-phosphate dehydrogenase has been shown to lose the integrity of the SH sites in the active center after PDT. 18 Changes in activity. conformation, and fluores(''ence along with an increase toward protease susceptability are some examples of such destmction in this cxtensi vely studied enzyme. Magnetic resonance imaging (MRI) has shown a dramatic drop in ATP levels immediately after PDT,I? probably as a result of cell repair mechanisms being activated . Even nuclear enzymes are affected by PDT. 20 In the calSC of DNA repair, photodynamic treatment of L929 cells generates nuclease activity that is nonnally controlled by ADP-ribosylation. The effect of direct single-strand damage to DNA has been noted with select photosensitizers in some systems. Recent studies suggest a dominant fonn of destruction associated with the DNA may correspond to interruption of microtubulin formation by a low-density lipoprotein delivered photosensiLizer such as with Pholofrin®. 21 The site at which tumor damage occurs is related not only to where the sensiti7.er causes damage in the cell but also to the morphology of the tumor. (t has heen observed depending on the hydrophilicity of the photosensiti7..er, that a higher concentration can be found around the tumor as compared 10 that of surrounding tissue. 22 This increase in concentration may ~ accounted for by either specific upwkc by a lipoprotein-associated mechanism as ~een with other drugs or by simple physical morphology of leaky Lumor vasculature and poor lymphatic drainage. Richter et al. have shown that low-density lipoprotein (LDL) bound BPDMA reduces the amount of drug in the skin while destruction of the tumor remains the same. 2 .l II is known that LDL binding sites are higher on cancer cells and it is a topic of intense interest in the field or drug delivery. 24 Formulation of a drug may 00 important in optimizing (his increalie in drug localization perhaps by an LDL delivery system. In the case of BPDMA, the fonnulation that is utilized for clinical trials are unilamellar vesicles (Iiposomes) to help solubilize the hydrophobic drug and direct it to its lipoprotein-dominated delivery mechanism . 2~ Lipo~ome..~ are alw being investigated for use in the delivery of zinc phthalocyan;ne,2(, a compound that normally ha~ extremely poor 501ubility even in organic solvents. The water-soluble NEP6 also accumulates in tumors although it is readily taken up in the blood by ~erum albumin. 27 Perhap~ even in th is case, initi al delivery of the drug is dominated by the lipoprotein fraction of Mood even though a majority of il hinds to the serum albumin. It may nor only be the actuat destruction of cancer cells that results in tumor ablation during PDT but also that of the micro(neo)vasculature. 28 Some control over the biological behavior of the photosensitizer associated with its structure has been observed. A series of modified aluminum phthalocyanines (Fig. 8) in which the parent molecule has sulfonation ratios ranging from 1 to 4 has shown that the more lipophilic monosulfonated compounds are marginally more effective than the tetrasulfonatcd compound in terms of absolute phototoxicity in vivo .29 However, in vitro experiments show that the very hy~ drophilic telrasulfonated material has poor photosensitizing abilities. Some property of the ;,~ vivo delivery system to the vasculature has increased the more hydrophobic compound's efficacy. Such possible vasculature destmction has been dem* onstrated by a numbcrofrcsearchers . Morgan et aJ. have shown that rddioactive microsphcres are held up to a greater extent in tumors after PDT. JO MRI has been used to show that the relax* ation coefficients (T r T 2) of water around the tumor RfC dntmatically changed soon after PDT.3' A new technique thaI utilizes laser doppler mt!asurements can generate information much like that given by MRI about blood flow of a recently treated tu- mor.32 237 Second Generation Photodynamic Agents Perhaps the most interesting :1!>pect of PDT is associaLed with (he immunological effects of posttreatment. Most of the significant studies in this field have been carried out with HpD. a slightly less pure [Offil of Photofrin®, which was initially developed in the 1950s by Cohen and Schwartz as a radio protectorl sensilizer. 3J In the dark HpD has been observed by Canti et al. to cause increase in bone marrow and spleen hypercellularity in mice. 34 This compound has been used to rescue mice from nu.liation therapy for leukemia. Photodynamic therapy in mice in contrast has shown immunosuppression as indicated by sensitivity 10 dinitronuorobenzene. :l.~ Observation of patients after !>uccessful treatment for bladder cancer showed an increase in chronic inflammatory cells. J6 Further observations have shown a relationship between the cytokine~, interleukin-l~, interleukin-2, and tumor necrosis factor-lX concentrations and the level of PDT trealrnent 1n patients. More direct evidence of cytokine release was found hy Evans el al. who observed "tumor necrosis factor" release by macrophages after PDT.37 Bellnier has used combination therapy of PDT and tumor necrosis factor to increase tumor response ..1R Thromboxane has also been found 10 be released shortly after PDT. V) It is also suspected that nitric oxide, better known as endothelium releasing factor (EDRF), which is related to guanylate cyclase, may be a consequence of macroph age ac (i vation. 40 Th is is not surpris i ng in that at least in the case of Photofrin®, a minor constituent of the drug is protoporphyrin IX, a cofactor in the cascade in nitric oxide formation. LIGHT DELIVERY DEVICES In the Ia.'>t several years QLT and Lederle have set up total laser systems that meet international clinical standards. Among the areas to review were the safety. equivalency of clinical lasers to a reference standard, compatibility of existing fiber optics and power meters, verification of manufacturers specifications, conformance with international regulatory require~ lnents, and suitability for the clinical setting. This i:s an ongoing process. For PDT many types of lasers have been evaluated. The mosl common type in use is the argon pumped dye laser (APDL). This has been used exclusively in the QLT/Lcdcrlc Photofrin~ Phase III clinica.l trials in North America. Recently, APDLs such as the Lambda Plus from Coherent Medical have been engineered specifically for PDT with Photofrin® as the toxin. Il produces upward of 1.9 W from the tip of the fiber at 630 nm, which is more than enough power [or most clinical requirements. Other lasers that have been considered for use with Photofrin® are (he copper vapor pumped dye lasers and the gold vapor lasers. These pulse systems suffer from several problems such as lack of technical compatibility for the clinical setting, developing light dosage standards to relate to the APDLs. and high peak pulse energy output lhat lend~ to degrade the optical fibers. The utilization of a KTP Y AG to pump a dye in the region of 630 nm has been achieved and an ex.ample of such a system has been made by Laserscope. At the primary output of 1066 nm this laser can be utilized for general Y AG surgical applications. When the output in frequency doubled to 532!lm it can pump a dye laser 10 a number of standard frequencies. This laser provides a high frequency pulse of -- J0 kHz with a relatively low flower output of I kW, which should prove to maintain fiber integrity. The last ~everal years have seen the development of {l new type of laser that has the potential to be comp8ct, require little or no cooling, and should work directly from a wall plug. These solid state diode lasers have shown that clinically significant output (-- 3 W) can be achieved at 630 nm and thus may be considered in the future for use with Photofrin®. 41 For second generation compounds the diode laser. KTP Y AG pumped dye lasers. and the standard APD laser are being reviewed. Clinical trials at present are concentrating on utilizing research type APDL. However, as the second generdtion drugs move through clinical trials it becomes mon: likely that a diode laser designed for the drug's specific wavelength will be developed. In the case of psoriasis or cutaneous cancers it may be practical to u~e nonlascr light sources such liS arc lamps. fluorescent tubes, and light emitting diodes since the light does not have to be concentrated into a small a!C<t. These light sources will require all the clinical equivalency t.1udies, which have been nccessary for the lasers. At present PDT requires the utilization of a laser couploo to a fiber optic system incorporated info an endoscope. Fiber optic systems have been developed with specific treatments in mind. In the instance of cutaneous cancer il GRIN lens is utiliz,ed to spread the light to a cone. Thi.s allows light dosages to be easily calculated by taking into account the transmitted power from the tip, the time, and the distance from the tip to the treatment field. Bladder cancer requires the incorporation of a sphere diffuser into the above system. Newer systems have utilize.d a balloon catheter to center the fiber as the balloon expands the bladder to be roughly spherical. This, in combination with an intrabladder detector, should ensure constant and even light dosimetry. 42 Cylindrical geometries that are found in the esophetgus or the bronchi can be treated by fibers developed with cylindrical output. A fiber with a cylindrical output tip of up to 2.5 cm in length i!. passed into the treatment area and placed parallel to the tumor. Improvements to !hi~ system include the development of a balloon catheter with detector probes on the wall. In combination with this a face of the balloon has been darkened generating a lantern effect. In the case of a very large tumor the fiber can be placed directly into it. 43 CLINICAL TRIALS The details and complexity of preclillical studies and clinical trials are even more complicated for PDT than for a "traditional" drug since both drug and light are involved. Toxicity must examine the traditional "dark" effects as well as those generated on illumination. For the most part none of the drugs described above ha<; been repolted to have any Inherent dark toxicity in doses ranging much higher (X 10-X 1(0) than the therapeutic dose. After stability and toxicity of the appropriate drug fonnulation are addressed, il is necessary in a Phase I clinical trial to investigate drug dosage, time of irradiation after injection, pharmacokinetics, human drug toxicity, and, most importantly, the extent of skin photosensitivity during and after 238 treatment. The development of most standard therapeutic agents generally does not have 10 correlate drug dosage to skin toxicity and light activation of drug. For these reasons initial clinical trials in PDT have been directed toward investigating the use of the drug for treatment of skin cancers. In these studies a comhined review of toxicity toward the skin while deriving information about the light doses necessary to damage the tumor can be detennined. Of course such infonnation gives an indication of the efficacy toward the IUmor, which is not generally the goal of initial clinical trials. A profile of drugs in Phase I investigations 10 date has shown that after initial injection the drug takes some hours to accumulate in the tumor and the skin. One hopes to irra<.liate at some time after the injection so that therapy can proceed on the same day. Time of accumulation into the tumor as compared to the skin or other tissue may not be ideal for all the new drugs. In the case of Photofrin® a dosage of approximately 2.5 mg/kg i~ st(tndard .44 Irradiation occurs some 4H to 72 hr after injection at a time when the drug has a higher concentration in tumor compared to that in surrounding tissue. A typical light dosage from the APDL is 150-250 J/cm 2 for treatment of a lung tumor. In the case of bladder cancer the light is diffused over the entire surface of the organ to give a light dosage of 150 J/cm 2 •4~ With the above dosage of drug, skin photosensitivity may last about 4~ weeks. Most photosensitizers are also good fluorophores. This property may allow the clinician to eventually determine the level of drug in both the skin and tumor before and after irradiation, which could be useful both for delennining the site of treatment and for calculating the appropriate light dosage during therapy. Four second generation drugs are currently in Phase Tclinical trials. These are BPDMA, with which the authors are most familiar, monoaspactylchlorin e 6 • mesQ-tetra(m-hydroxyphenyl)chlorin (mTHPC), and tin eliopurpurin. Since much of the preclinical and clinical research is of a proprietory nalure, information on the progress of Ihese drugs toward a submission of a new drug application (NDA) is rather limited. None exists, as of yel, for the tin etiopurpurin. There is a pllblished Hccounl on the treatment of 4 patients using mTHPC and it is known that additional patients have been enrolled in the study. 46 mTHPC is somewhat hydrophilic and rapi<.lly accumulates in malignant mesothelioma. The drug is delivered in a mixture of polyethylene glycol, ethanol, and water and is formulated just prior to injection. The ratio of drug in tumor as compared to surrounding muscle was determined to be 7: 1 and 14: 1 relative to skin. These ratios are higher than the 2-4: 1 found for Photofrin® in human..I\. The circulating concentration of the drug 48 hr after injection was found to be 0.15 mg/ml. while the half-life of the drug in circulation was approximately 12 hr. Dosages ranged from 0.075 to 0.3 mg/kg with a light irradiation of 10 J/cm 2 at 650 nrn. At the lower dosage 50% necrosis of the lumor was shown while the next higher dosage showed 80% necrosi~ and the final dosage 100% necrosis to a depth of 1 em. Derailed histological profiles of the tumor and surrounding tissues described the extent and area of lissue damage. Description of skin photosensitivity studies was sparce and the studies appeared not to be carried out under controlled conditions with a solar simulator. However, some anecdotaJ data were noted. Skin photosensitivity appeared to begin some time after injection (3-10 days); it does appear, however, thai Sternberg Bnd Dolphin significant clinical photosensitivity has been observed in some patients up to 10 days after treatment, which may place limitations on the use of this drug in PDT. Phase I clinical trials for the water~soluble monoClspartylchlorin C6 began at the same time as for BPDMA and were examincd against the same skin tumors described below. 47 Drug doses were assigned in the range 0.5 to 2 mg/kg with concurrent increases of tight dosage from 12.5 to 200 J/cm 2 . As in the ca~e of BPD <.Irug dosages were limited to (he lower concentrations because of the initial positive responses found. Skin sensitivity in the lowl.!r dosages used (0 .5 mglkg) lasted up to 7 days. This drug has strong binding to serum albumin~ and it appears to circulate for a much longer lime than other PDT drugs; in fact the drug is readily detectahle 4 weeh after treatme-nt while 80% of the initial concentration is detected in the blood 10 days after injection. The second generation photosensitizer BPDMA is a semisynthetic porphyrin derivative that has been found to be effective in treating a number of cancers in vivo along with purging of viruses from blood:19 In the ongoing clinical trial information about the pharmacokinetics, skin photosensitivity along with tumor response has been reported. 50 The patients had a range of cancers Ihat included basal cell carcinoma, metastatic breast carcinoma, metastatic gRstrointestinal carcinoma, and metastatic amelanotic melanoma. Drug dosages ranged from 0.25 to 0.5 mg/mg in the patients who were infused 3 hr prior to irradiation while light dosages administered by a laser tuned to 690 nm ranged from 50 to 150 J/cm 2 for the low drug dosages to 50 J/cm 2 for the high drug dosage. Sixty-four cutanoous tumors were treated and followed for several months after treatment. Overal1 tumor response was 63%, while 100% response.~ were noled for the higher drug and light dosages . ~J Since the major purpose of the first generation trial was to judge skin photosensitivity the most extensive studies for UV N VIS light were carried out with the utilization of a filtered xenon arc lamp source onto normal skin soon after light treatment of the tumors while UVB light was generated from fluorescent sun lamps. Testing of UVB sensitivity showed no response for the carly patients, so these assays were discontinued. Marked skin photosensitivity to UV A/VIS light was noted early after treatment. On the day of treatment the challenged light dose was reduced for the higher drug dosages so that minimal erythema dose (MED) could be noted. Close examination of light exposure onto the patients ranging up to 9 days after treatment for high drug dosages shows u logarithmic loss of photosensitivity. An overall phannacokinetic profile of drug serum concentration is reviewed in Fig. 9. It is noted that the drop of all drug dosage levels down from the maximum concentration corresponded to -I hr. The relationship between these results and the corre~ sponding clearance of drug from sk.1n has yet to be investigated. All the drugs investigated to date have exhibited distinct clinical responses. The maximum effective drug do~e corresponding to minimum skin sensitivity has yet to be determined for any of these materials. However, general approaches toward optimizing such a differential can be applied to all of them. Wilson 5 and others have suggested that a minimum drug dose that would then be treated with a large light dose would illicit the best response over all clinical responses, especially if moderate bJeaching of the drug occurs. Several other approaches may be considered when optimizing light dosage for clinical 239 Second Generation Photodynamic Agents -:J' ',000 ~ Q> 5 500 200 ""§ 100 ~ u ~ 20 ~~ ~e: 0.. o 8. ~O 9. 10 5+----r--~----T---~--~ o 6 12 18 24 30 TIME (h) 10. FIG. 9. Post injection plasma concentration versus time profiles of BPDMA in blood with a doseage of 0.25 mglkg in humans. 11. respunses, which could include fractionation of the light dosage. 12. 13. ACKNOWLEDGMENTS We wish to thank our colleagues at Quadra Logic Te<:hnologies and Lederle Laboratories for their assi~tan{;e in preparing this manuscript, especially Michael Stonefield and Ron Caroll for their comments on light delivery devices. 14 . 15. REFERENCES I. Dougherty, T. S., Henderson, B. W., Schwartz. S .. 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(993). Pmc. SPIE. lnt. Soc. Opt. 1987, 147-151. 241 Address reprint requests to; Ethan D. Sternberg, Ph.D. Depanmenr oj Chemistry University of British Columbia 2036 Main Mall Vancouver, B.C .. Canada V6T IZI