Sensitizers For Photodynamic Therapy Research Articles

ChemInform Abstract: New Sensitizers for Photodynamic Therapy: Controlled Synthesis of Purpurins and Their Effect on Normal Tissue.

AbstractThe nickel porphyrinaldehydes (Ia) undergo Wittig olefination with the ester (II) to give the acrylic esters (Ib) which are transformed into the demetalated porphyrins (III).

PHOTOPROPERTIES OF ALKOXY‐SUBSTITUTED PHTHALOCYANINES WITH DEEP‐RED OPTICAL ABSORBANCE

Triplet-state properties of 1,4,8,11,15,18,22,25-octa-n-butoxyphthalocyanine and its zinc derivative were determined for the first time. The T1 state of the metal-free phthalocyanine was characterized by a short lifetime (tau T = 17 microseconds) and low quantum yield (phi T = 0.095), and quenching of the triplet by O2 occurred with a bimolecular rate constant (kT sigma = 1.3 x 10(8) M-1 s-1) that is indicative of an endogonic reaction. The zinc complex (ZnPc(OBu)8) was markedly better as a triplet photosensitizer with respect to both tau T (60 microseconds) and phi T (0.5). Quenching by O2 produced singlet oxygen with nearly 100% efficiency, and kT sigma (1.7 x 10(9) M-1s-1) was close to the spin-statistical diffusion-controlled limit. Phosphorescence measurements showed the energy of the T1 state of ZnPc(OBu)8 to be 100 kJ/mol, which is 6 kJ/mol above the 1 delta g state of O2. These photoproperties, together with Q-band absorption maxima in the mid-700 nm range indicate that metal-centered 1,4,8,11,15,18,22,25-octaalkoxyphthalocyanines have excellent potential as sensitizers in photodynamic therapy.

In vitro photodynamic therapy of human lung cancer

Photodynamic therapy (PDT) using a dihematoporphyrin ether (DHE) sensitizes malignant cells to damage by 630-nm light. This study investigated in vitro PDT sensitivity of human lung cancer cells (A549) and those factors which influence cell survival as determined by the colony formation assay. After incubation for 2, 4, or 6 hr with [DHE] of 2.5, 25, or 50 μg/ml, A549 received red light at dose rates of 0.27 or 0.09 mW/cm 2 and energies of 0–250 mJ/cm 2. Neither 630-nm light alone nor DHE alone affected cell survival. A dose rate of 0.27 mW/cm 2 required less energy than 0.09 mW/cm 2 for 90% cytotoxicity (180 mJ/cm 2 vs 250 mJ/cm 2, P < 0.05). The energy required for 90% cytotoxicity with 25 μg/ml [DHE] was dependent on DHE incubation time (2 hr, 90% cytotoxicity not reached; 4 hr, 116 mJ/cm 2; 6 hr, 69 mJ/cm 2; P < 0.05). In contrast, cellular [DHE] as measured by fluorescence, plateaued after 2 hr of incubation. Fluorescence microscopy revealed a time-dependent redistribution of fluorescence from the cell membrane to perinuclear and intracytoplasmic organelles. A 99% cytotoxicity required significantly less energy as [DHE] was increased (2.5 μg/ml, no cytotoxicity; 25 μg/ml, 243 mJ/cm 2; 50 μg/ml, 111 mJ/cm 2; P < 0.05). Intracellular [DHE] was directly dependent on the incubating media [DHE] (2.5 μg/ml, 0.09 ± 0.01 μg/10 6 cells; 25 μg/ml, 0.80 ± 0.07 μg/10 6 cells; 50 μg/ml, 1.31 ± 0.11 μg/10 6 cells; P < 0.05). PDT cytotoxicity was inversely proportional to concentration of serum in the DHE media. These data illustrate that lung cancer in vitro is sensitive to PDT and is influenced by dose rate, energy input, and DHE environmental manipulations. These factors may be important in increasing the efficiency of PDT of thoracic malignancies in vivo.

New sensitizers for photodynamic therapy: controlled synthesis of purpurins and their effect on normal tissue.

Purpurins are a class of porphyrin derivative that have been shown to have good in vivo cytotoxicity to N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide (FANFT) induced rat bladder tumors (AY-27) implanted into Fisher 344 rats. The synthesis of purpurins from etioporphyrin I and coproporphyrin I proceeds in high yield and with a high degree of regioselectivity. Product formation can be rationalized in terms of relief of steric strain about the periphery of the purpurin macrocycle. The effect of therapeutic light doses using the rat footpad model suggests that, at therapeutic sensitizer doses, normal tissue damage is within acceptable limits, particularly for metalated purpurins.

A COMPARISON OF SERUM KINETICS AND TISSUE DISTRIBUTION OF PHOTOFRIN II FOLLOWING INTRAVENOUS AND INTRAPERITONEAL INJECTION IN THE MOUSE

Abstract Although hematoporphyrin derivative (HPD) and its ‘purers’ variety Photofrin II are the most widely used tissue sensitizers in both clinical and experimental photodynamic therapy (PDT), quantitative studies of tissue distribution have been few. We have extracted and measured Photofrin II in several organs of the normal mouse including those of relevance to urological practice. In view of the reported heterogeneities in the distribution within tissues of various cytotoxics when administered intraperitoneally. we have compared results for Photofrin II given by this route with those for intravenous injection. Although both routes of administration gave equally consistent results, differences in absolute tissue concentration as a function of time after injection were found for several but not all tissues. Furthermore, the porphyrin accumulated following intravenous administration seemed to contain more of the non‐polar photodynamically active component than that accumulated following the intraperitoneal route. We attempt to explain these differences by reference to published data on porphyrin binding to serum proteins.

Recovery of Chinese hamster cells following photosensitization by zinc tetrahydroxyphthalocyanine

The phthalocyanine dyes are attractive sensitizers for photodynamic therapy of cancer. The light fluence response curves for photocytotoxicity of zinc tetrahydroxyphthalocyanine were constructed using the colony-forming ability of Chinese hamster cells as an end-point. The survival curve of cells photosensitized to white light by this dye has a pronounced shoulder followed by an exponential decline. Postillumination hypertonic treatment (0.5 M NaCl for 20 min at 37 °C) enhanced log-phase killing, although to a lesser extent than after exposure to ionizing radiation. While such an enhancement usually indicates that the cells are able to repair potentially lethal damage, delayed trypsinization of photosensitized cells in plateau-phase failed to show a significant increase in cell survival. Thus, the repair of such a damage in plateau-phase is apparently absent. Experiments with split light fluence indicated that log-phase cells can repair sublethal damage during a 24 h interval, as evidenced by the reappearance of the shoulder on the split-dose survival curve.

Photobleaching of photofrin II as a means of eliminating skin photosensitivity.

AbstractOne of the effects of using Photofrin II as a sensitizer in clinical photodynamic therapy is skin photosensitivity. Avoiding the harmful effect of this photosensitivity typically requires the patient to remain indoors during daylight hours for 4 weeks post injection. Photosensitivity can last up to 8 weeks. A possible method of quickly eliminating skin photosensitivity is the use of photobleaching to eliminate the DHE (dihematoporphyrin ether, the active agent in Photofrin II) from the skin. The Photofrin II was used as supplied (Photomedica, Raritan, NJ) and will be referred to as DHE.We report a series of experiments using Swiss mice which demonstrate the ability to reduce the period of photosensitivity to 1 week. The Swiss mice received 5 mg kg‐1 DHE intraperitoneal. Beginning at 24 h post injection they were given daily increasing light doses starting with 18 J cm‐2 (at 630 nm). Only the left hind feet were treated with light. At 7 days post injection both hind feet were treated with 135 J cm‐2. The pretreated left foot showed no reaction to this treatment whereas the right foot reacted as strongly as control mice which had received no pretreatment with light. The control mice were also injected 7 days prior to treatment.As a check for tolerance animals received a second injection of 5 mg kg‐1 DHE intraperitoneally 96 h after the conclusion of the bleaching protocol. Twenty‐four h later the left hind foot (previously desensitized by light) reacted to 135 J cm‐2 as strongly as controls which had received the 5 mg kg‐1 of DHE but no pretreatment.

A test of different photosensitizers for photodynamic treatment of cancer in a murine tumor model.

Abstract The efficiency of different sensitizers for photodynamic therapy (PDT) was tested using a model system with a C3H mammary carcinoma growing subcutaneously on the dorsal side of mouse feet. Growth curves were constructed from which growth delay and doubling time in the regrowth phase were calculated. As PDT induced oedema in the mouse foot, this model system also allowed assessment of normal tissue response.The following sensitizers were tested: hematoporphyrin derivative (HpD), Photofrin II (PII), tetraphenylporphinetetrasulfonate (TPPS4), acridine orange (AO), phthalocyanine tetrasulfonate (PCTS), Al‐ and Zn‐phthalocyanine tetrasulfonate (A1PCTS and ZnPCTS). For tumor control, the following sensitizer efficiencies were found: PII &gt; HpD &gt; AIPCTS &gt; TPPS4 &gt;&gt;&gt; ZnPCTS, PCTS, AO. With regard to sensitizing normal‐tissue damage: PII &gt; AIPCTS, TPPS4 &gt; HpD, ZnPCTS, PCTS. The results suggest that AIPCTS should be further evaluated for use in PDT.

Effects of membrane physical parameters on hematoporphyrin-derivative binding to liposomes: a spectroscopic study.

Physical parameters of membrane bilayers were studied for their effect on the binding of hematoporphyrin derivative (Hpd), which is used as a sensitizer in photodynamic therapy of cancerous tissues. The purpose of this study was to clarify which parameters were relevant, under physiological conditions, to the selectivity of Hpd binding to cancer cells. Fluorescence spectroscopy was used to measure the relative partitioning of the dye between the lipid and aqueous media. Increasing the microviscosity of the liposomes' membranes by various bilayer additives results in a strong reduction of Hpd binding, to an extent independent of the specific additive. The effect of temperature near the physiological value as well as the effect of cross membrane potential are small. Surface potential does not affect the binding constant, indicating that the binding species does not carry a net electric charge.

A comparison of different photosensitizing dyes with respect to uptake C3H-tumors and tissues of mice

Nine dyes, all potential sensitizers for photodynamic cancer therapy (PDT), were injected in mice with C3H mammary carcinomas. Twenty-four hours later the animals were sacrificed and the dye concentrations in tumors and 9 other tissues were measured by means of spectrofluorimetry. The 9 dyes were: Photofrin II (PII), hematoporphyrin (HP)-di-hexyl-ether, PSD-007 (a sensitizer used in clinical trials in China), tetraphenyl porphine tetrasulfonate (TPPS 4), tetra(3-hydroxy phenyl)porphyrin (3THPP), aluminium phthalocyanine tetrasulfonate (AlPCTS), aluminium phthalocyanine (AlPC), chlorin e 6 (Chl e 6) and merocyanine 540 (MC 540). The porphyrin precursor δ-aminolevulinic acid was also tested and found to induce porphyrin fluorescence in tumors and some other tissues. The best tumorlocalizer of those tested was 3THPP. This drug also showed a favorable tissue distribution. The following dyes showed lower skin/tumor concentration ratios than PII (the most widely used dye for PDT): Chl e 6, PSD-007, HP-di-hexyl-ether and 3THPP. Low brain/tumor ratios were found for: PSD-007, HP-di-hexyl-ether, 3THPP, TPPS 4 and AlPCTS.

Photodynamic therapy of C3H mouse mammary carcinoma with haematoporphyrin di-ethers as sensitizers.

Haematoporphyrin di-ethers were synthesized and tested as sensitizers for photodynamic therapy of C3H/Tif mammary tumours in mice. Growth curves of the tumours were determined by measuring the tumour volume. The animals were given 25 mg porphyrins kg-1 body weight i.p. and 24 h later exposed to 135 J cm-2 of 630 nm light at a fluence rate of 150 mW cm-2. The sensitizing efficiency of the ethers was measured in terms of the increase in growth time of the treated tumours, as compared with that of untreated controls, needed to reach a volume 5 times larger than that at the time of the treatment. This sensitizing efficiency increased with decreasing polarity, i.e. in the order di-methyl ether, di-propyl ether, dibutyl ether and di-amyl ether. Haematoporphyrin di-amyl ether was more efficient than haematoporphyrin derivative and insignificantly less efficient than photofrin II (DHE). This was true for sensitization of both tumours and normal tissue.

Laser spectroscopy on organic dye in living cells.

Recent investigations done in our laboratory on living cells by means of laser spectroscopy are reviewed. The first two are concerned with Chlorella: Stokes and anti-Stokes resonance Raman scatterings from carotenoid are measured to show that the Boltzmann distribution holds even in living cells. The fluorescence decay characteristics of chlorophyll are also studied by time-correlated single-photon counting under very weak light excitation. The remaining investigations are concerned with lymphocyte and are diagnostically important: The mechanism of fluorescence depolarization of fluorescein, which is known to show a change in the degree of polarization after antigenic stimulation, is clarified by time-resolved technique. Spectral properies of hematoporphyrin derivative in the cell, known as a sensitizer for recent photodynamic therapy, are also presented. Finally, the lineshape of organic dyes in solution is discussed with stochastic theories.

Separation of different fractions of hematoporphyrin derivative by a two-phase extraction system.

Hematoporphyrin derivative (Hpd), prepared as described by Lipson et al. is the most widely used sensitizer in photodynamic cancer therapy (PDT). Hpd contains a number of porphyrins, some of which contribute minimally to its photodynamic action2. A somewhat purified version of Hpd, so-called photofrin II (PII), is also used in animal- and in clinical experiments3,4. PII is probably mainly composed of porphyrin dimers and/or oligomers linked together with ether- or ester- bonds, together with a variable proportion of hematoporphyrin (Hp), hydroxyethyl-vinyl-deuteroporphyrin (Hvd) and protoporphyrin (Pp). Porphyrins witji low activity may be removed from Hpd by means of gel chromatography5, dialysis or ultra-centrifugation. Below is presented a simple extraction procedure that leads to a product with properties similar to those of PII.