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The formation of complexes of β-formyl-heptaethylporphyrin and β-(2-formylvinyl)-heptaethylporphyrin in addition to the expected mesoformylporphyrin was detected when formylating the platinum complex of actaethylporphyrin by the Wilsmeier reaction. A possible mechanism is proposed for the unusual formation of similar compounds.

Interaction of the water-soluble platinum and palladium coproporphyrins with cationic carriers, DEAE-dextran and CTAB, was investigated in aqueous solution by optical spectroscopy including pH-dependences of the absorption and emission spectra. In addition, solid-state coatings of the complexes were prepared on solid supports and their phosphorescent properties were studied. Contrary to the pure dyes, in the dry and solid state, the complexes strongly phosphoresce in air, are weakly quenchable by air oxygen, but are sensitive to the water content in their environment. Their applicability to quenched-phosphorescence sensing was discussed and compared to that of the phosphorescent polymer film compositions and Langmuir-Blodgett coatings.

The chemical synthesis of six lipophilic conjugates of chlorins was carried out, in which lipophilic fragment (either hexadecyl- or cholest-5-en-3β-yloxyethyl-) bound to 13(1)-, 15(2)-, 17(3)-positions of macrocycle by formation of related carboxamides. Structure of synthesized conjugates was studied by spectral methods and molecular modeling. Lipophilic conjugates of chlorins, being mixed with egg yolk phosphatidyl choline, formed mixed micelles stable in aqueous media under physiological conditions. Mixed micelles of conjugates with phosphatidyl choline differing in stoichiometric compositions were prepared and characterized by absorption spectra, electron microscopy and laser scattering. These micelles were found to bind and internalized by human breast carcinoma MCF-7 cells. The presented data reveal that modification of macrocycle with lipophilic substituents, solubilization of obtained conjugates in aqueous medium as mixed micelles with phospholipids, and transfer of mixed micelles to cells is simple approach for targeting of chlorin derivatives, which apparently may be used in photodynamic therapy.

Vilsmeier formylation of the oporphyrin-I tetraethyl ester Ni-complex (III) followed by reduction of the intermediary imine salt (IV) with NaBH4 gave the o-dimethylaminomethyl-coproporphyrin-I Ni-complex (V). Heating the latter in methyl iodide gave 65% of the ethane- bis-porphyrin (I), which has been characterized by NMR-spectroscopy and a molecular and crystal structure investigation. The crystal structure shows an extended structure of the bisporphyrin with a parallel arrangement of the porphyrin subunits. Crystal data: I - C90H106N8Ni2O16. Triclinic, a = 12.319(3) Å. b = 13.297(3) Å, c = 14.444(3) Å, α = 105.56(2)°, β = 100.82(2)°, γ = 112.98(2)°, V = 1981.0(7) Å3, Cu Kα radiation, λ = 1.54178 Å, space group P1̄, Z = 1, R = 0.07.

A set of luminescent dyes, namely the porphyrin-ketones and their zinc(II), platinum(II) and palladium(II) complexes, are described. The probes are derived from a partially oxidized porphyrin ring and have been studied by absorption and luminescence spectroscopy including time-resolved measurements. The metal-free porphyrin-ketones display strong and pH-dependent fluorescence due to a protolytic equilibrium that exists between the free base and the dication. The zinc(II) complexes also give strong fluorescence along with a weak long-decay emission at room temperature which are attributed to delayed fluorescence and phosphorescence, respectively. Platinum(II) and palladium(II) complexes exhibit room-temperature phosphorescence without any detectable fluorescence. Compared to existing porphyrins, the new dyes display a considerably longwave-shifted luminescence along with substantially improved (photo)chemical stability. They are considered to be promising probes for very near-infrared luminescence studies, and to be particularly useful for applications such as labelling of biomolecules, pH-sensing, and detection of quenching species such as molecular oxygen.

Reaction of various trimethyl(porphyrinylmethyl) ammonium iodides (generated in situ from the corresponding meso -dimethylaminomethylporphyrins and iodomethane) with triphenylphosphine afforded triphenylporphyrinylmethylphosphonium iodides which, when subjected to mild basic hydrolysis, gave meso -methylporphyrins in high yields.

A new class of dyes, platinum(II) and palladium(II) complexes of the porphyrin ketones (or oxochlorins), exhibiting strong phosphorescence at room temperature is described. Several representative compounds were prepared and studied by spectral luminescence methods in solution. Compared to the related porphyrin and chlorin complexes, the new dyes display high photochemical stability, long wave spectral characteristics, and good compatibility with semiconductor optoelectronics (e.g., excitation by light-emitting diodes). These properties make the new dyes promising for a number of relevant applications, such as quenched phosphorescence sensing and phosphorescence probing (e.g., in binding assays). Analytical application of the porphyrin ketone complexes to phosphorescence lifetime-based sensing of molecular oxygen is described. Platinum(II) octaethylporphine ketone was dissolved in a polystyrene layer to give an oxygen-sensitive film. Oxygen measurements were performed with a prototype fiber-optic instrument based on solid-state components, such as light-emitting diodes and photodiodes. The instrument measured phosphorescence lifetime via changes in phase shift. The phosphorescence lifetime was determined to change from about 61.4 μs at zero oxygen to 16.3 μs in air (210 hPa of oxygen) at 22 °C. The analytically useful range of the sensor was 0-210 hPa of oxygen partial pressure, with a detection limit of 1.5 hPa. The precision of the device was 1.0 hPa at 210 hPa of oxygen and 0.5 hPa at zero oxygen.

Vilsmeier reaction of mono-Ni 1,2-ethanebis(octacthylporphyrin) favoured 10-substitution. Functional group intcrconvcrsions led to pure 10-dimethylaminomethyl, -azomethine, -methoxymethyl, and -formyl derivatives. Demetallation gave a range of labile 10-substiluted free base dinners. Oxidation of the mono-Ni compounds in AcOH gave the corresponding trans-ethylene mono-Ni derivatives, while the free bases were all converted to 10'-formyl-1,2-cthylenebis(octaethylporphyrin) under the same conditions.

Phosphorescent polymer film compositions and phosphorescent Langmuir-Blodgett coatings based on platinum and palladium porphyrins were tested in the presence of nitrogen oxides (NOx). Dynamic quenching of phosphorescence by NOx was observed. It was accompanied by chemical interaction of NOx with the dye molecules which caused persistent decrease of the phosphorescence signal and irreversible degradation of the sensor. Two modes of NOx quantitation based on measurement of either phosphorescence lifetime or time drift of intensity were examined. For the phosphorescent polymer film compositions the analytical range of NOx was found to be 0-3.0% ( m m ) with a detection limit of 0.1%. For the LB-film coatings the sensitivity for NOx was about five times better: the range was 0-1.0% ( m m ) with a detection limit of 0.02%. Although for both systems the analytical range of NOx covered was poor for practical purposes, the data obtained are useful for evaluation of possible interferences by NOx in other applications, such as in oxygen and SO2 sensors. The possibilities of the realization of a dual-sensing concept with a single phosphorescent probe is discussed with respect to simultaneous measurement of NOx oxygen using polymer film compositions and NOx SO 2 using Langmuir-Blodgett coatings.

We describe a novel technique for heme removal and replacement in the heme domain of P450BM-3 (BMP). The method was applied to obtain the aluminum-protoporphyrin IX (Al-PP) substituted derivative of BMP (Al-BMP). The overall yield of the purified Al-BMP was about 15% as related to the initial amount of the hemeprotein. Al-BMP possesses extensive fluorescence in the 550-650 nm region with excitation in the porphyrin absorbance bands. The protein was shown to bind substrates of P450BM-3 (palmitic, arachidonic, and cis-parinaric acids) with affinities similar to those of the native enzyme (3-6 µM). However, the substrate-induced changes in fluorescence of Al-PP reveal the existence of a second, low-affinity substrate-binding site, which cannot be detected by the spin shift in the native, heme-containing BMP. Using fluorescence resonance energy transfer, we have demonstrated that Al-BMP forms a complex with the flavoprotein domain of P450BM-3 labeled with 7-ethylamino-3-(4'-maleimidylphenyl)-4-methylcoumarin maleimide, revealing the affinity similar to that of native BMP (Kd = 5 µM at 0.06 M ionic strength). Therefore, aluminum-substituted BMP may serve as a valuable tool in studies on the mechanisms of interactions of P450s with their substrates and protein partners.

The mass spectral behavior of 1, 2-di (mesoetioporphyr inyl-1)- and 1, 2-di (mesooctaethylporphyrlnyl) ethanes and ethylenes, their Cu and Ni complexes, as well as Ni Schiff bases of 1, 2-di (mesooctaethylporphyrinyl)ethane complexes, under ionization by electron impact and bombardment by accelerated atoms, was studied. Peaks of molecular ions, for which the basic decomposition pathways were established on the basis of an analysis of the parent and daughter ions, were observed in the electron impact mass spectra of all these compounds.

A facile aerobic oxidation of 1,2-bis(meso-octaethylporphyrinyl)ethane and other similar dimers to the corresponding trans-1,2-bis(meso-porphyrinyl)ethenes in several organic acids has been studied by UV-vis and 1H NMR spectroscopy and the general mechanism of the oxidation has been proposed. The key step of the oxidation is monoprotonation of the two porphyrin rings in dimer followed by two electron oxidation.

X-Ray analysis of trans-1,2-bis{meso-[nickel(II)octaethylporphyrinyl]} ethene has revealed that the compound has nonplanar structure and the nickel-coordination planes of porphyrin rings are close ...

A new cis-form of ethylene-bridged octaethylporphyrin dimer 3a has been obtained by isomerization of the corresponding trans-isomer 2a . On the basis of spectral data the structure of 3a was assigned to be close to a “face-to-face” orientation. Reflecting the peculiar structure of 3a UV-vis and 1 H NMR spectra are remarkably distinguished from those of ordinary porphyrins.

Research on cell-penetrating peptides for the intracellular delivery of porphyrin compounds has mainly focused on the use of trans-activator of transcription (TAT)-derived peptides and, to a lesser extent, on proline-rich peptides and phosphorescent metalloporphyrins. In this article, we describe a novel phosphorescent oxygen-sensitive probe for intracellular use which comprises a bactenecin 7 peptide fragment (15-24) conjugated with the uncharged monofunctional derivative of Pt(II) coproporphyrin I (PEPP0). This probe provides efficient loading of various mammalian cells, including PC12, HCT116, SH-SY5Y and HeLa, via cell-type-dependent uptake mechanisms. The conjugate displays a similar distribution in cytoplasm and mitochondria which allows local oxygen levels to be monitored. Respiratory responses of PC12 cells loaded with the conjugate, measured on a time-resolved fluorescent reader, showed significant cell deoxygenation in response to uncoupling by carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone and external hypoxia. Treatment with mitochondrial inhibitors led to a decrease in cell deoxygenation. Although the biophysical properties of this conjugate are similar to those of the phosphorescent intracellular oxygen-sensitive probes described previously, it possesses a number of advantages, including ease of synthesis, high loading efficiency and reliability in physiological experiments with cells. This intracellular probe can be employed for the measurement of intracellular O(2) levels in samples containing mammalian cells using the phosphorescence quenching technique. In addition, the responses to metabolic stimuli can be assessed in a wide range of cells, as can the levels of relative cell oxygenation under external hypoxia.

A new cis-form of ethylene-bridged octaethylporphyrin dimer 3a has been obtained by isomerization of the corresponding trans-isomer 2a . On the basis of spectral data the structure of 3a was assigned to be close to a “face-to-face” orientation. Reflecting the peculiar structure of 3a UV-vis and 1 H NMR spectra are remarkably distinguished from those of ordinary porphyrins.

Several new derivatives of the phosphorescent Pt(II)-coproporphyrin (PtCP) were evaluated with respect to the sensing of intracellular oxygen by phosphorescence quenching. Despite the more favorable molecular charge compared to PtCP, self-loading into mammalian cells was rather inefficient for all the dyes, while cell loading by facilitated transport using transfection reagents produced promising results. The PtCP-NH(2) derivative, which gave best loading efficiency and S/N ratio, was investigated in detail including the optimisation of loading conditions, studies of sub-cellular localization, cytotoxicity, oxygen sensitivity and long-term signal stability. Being spectrally similar to the macromolecular MitoXpress™ probe currently used in this application, the PtCP-NH(2) demonstrated higher loading efficiency and phosphorescent signals, suitability for several problematic cell lines and a slightly increased lifetime scale for the physiological range (0-200 μM O(2)). In physiological experiments with different cell types, mitochondrial uncouplers and inhibitors performed on a time-resolved fluorescence plate reader, this probe produced the anticipated profiles of intracellular oxygen concentration and responses to cell stimulation. Therefore, PtCP-NH(2) represents a convenient probe for the experiments and applications in which monitoring of cellular oxygen levels is required.

Probing of molecular oxygen in mammalian cells is important for the analysis of mitochondrial function, metabolic responses, and energetic status of the cells. We describe a new panel of intracellular O 2 -sensitive probes based on phosphorescent porphyrin dyes conjugated to cell-penetrating peptides. The probes comprising the uncharged derivatives of Pt(II)-coproporphyrin I covalently linked to positively charged TAT-derived peptides are shown to effectively load live mammalian cells without any transfection reagents. The probes work well with all cell types tested, show similar subcellular localization, and produce characteristic responses to cell stimulation with mitochondrial uncouplers and inhibitors. They provide a simple and versatile tool for O 2 monitoring in live cells and in tissue, and an alternative to the existing O 2 probes which require facilitated transport into the cell.

Diquinone derivatives of hematoporphyrin IX with different structures of the quinone fragments were synthesized by the method of mixed anhydrides. The compounds obtained were investigated by UV, IR, PMR, and fluorescence spectroscopy. Pronounced quenching of the fluorescence of porphyrin in the porphyrin quinones, which depends on the acceptor properties of the quinones and the spatial orientation of the donor and acceptor, was observed.

The synthesis of a diquinone derivative of the amino-containing analog of hematoporphyrin IX, obtained by the sequential modification of the tetramethyl hematoporphyrin IX, was accomplished.

A simple, minimally invasive methodology for the analysis of intracellular oxygen in populations of live mammalian cells is described. Loading of the cells with the phosphorescent O(2)-sensing probe, MitoXpress, is achieved by passive liposomal transfer or facilitated endocytosis, followed by monitoring in standard microwell plates on a time-resolved fluorescent reader. Phosphorescence lifetime measurements provide accurate, real-time, quantitative assessment of average oxygen levels in resting cells and their alterations in response to stimulation. Analytical performance of the method is examined, optimized, and then demonstrated with different suspension and adherent cell lines including Jurkat, PC12, A549, HeLa, SH-SY5Y, and C2C12, by monitoring responses to mitochondrial uncouplers, inhibitors, plasma membrane depolarization, and Ca(2+) effectors. The assay provides relevant, information-rich data on cellular function and metabolism. It allows monitoring of small, rapid, and transient changes in cell respiration and screening of new chemical entities.

[structure: see text] A racemic mixture of the ethane-bridged bis(zinc octaethylchlorin) was successfully resolved for the first time to yield two enantiomers that exhibit substantial CD signals in the regions of chlorin B and Q transitions. The absolute configuration of the corresponding enantiomers was assigned and the origin of its high optical activity was rationalized through a combined spectral, crystallographic, and theoretical analysis.

Phosphorescent platinum(II) coproporphyrin label (PtCP) is evaluated for the detection of cellular proteases by time-resolved fluorescence in homogeneous format. An octameric peptide containing the recognition motif for the caspase-3 enzyme was dual labeled with a new maleimide derivative of PtCP and with the dark quencher dabcyl. Following photophysical characterization, the quenched substrate was employed in cleavage assays for caspase-3 using Jurkat and HL60 cell lines treated with proapoptotic stimuli performed on a commercial plate reader. Dose-response and time course assays for the drug camptothecin were obtained for comparison with conventional fluorometric detection.

Chlorin e6 and its derivatives are promising sensitizers for photodynamic therapy (PDT). In order to compare the photodynamic effects of 8 novel derivatives of chlorin e6 and to explore some mechanisms of their effects at the cellular level, we studied PDT-induced changes in bioelectric activity of crayfish mechanoreceptor neuron that was used as a sensitive experimental model. Neurons were insensitive to red laser irradiation (632.8 nm; 0.3 W/cm2) or to photosensitizers alone, but changed firing rate and died under the photodynamic effect of nanomolar concentrations of sensitizers. The dynamics of neuron responses depended on photosensitizer type and concentration. The dependence of neuron lifetime on photosensitizer concentration allowed comparing efficiencies of different photosensitizers. Radachlorin was the most potent photosensitizer comparable with mTHPC. High photodynamic efficiency of some chlorin e6 derivatives was related to weak dependence of neuron lifetime on sensitizer concentration, indicating to the initiation of 2–3 secondary processes such as free radical membrane damage by one absorbed photon. Photodynamic efficiency of sensitizers depended on amphiphilicity influencing their intracellular localization.

Syntheses of octaethylporphine-diketone (OEPDK) and its platinum(II) and palladium(II) complexes (PtOEPDK, PdOEPDK) were optimized, and the dyes were isolated in a pure form in preparative quantities. They were characterized by the NMR, UV–VIS absorption and emission spectroscopy. Electronic spectra of these dyes (absorption and luminescence) were investigated in detail, and compared to corresponding porphyrins and porphyrin-monoketones. OEPDK showed a strong fluorescence at about 700 nm, while PtOEPDK and PdOEPDK showed very weak room-temperature phosphorescence in the region of 850–1100 nm and practically no fluorescence. Protonation mechanisms were studied for these dyes. Protonation at sites other than pyrrole nitrogen atoms was shown to occur, corresponding protomeric spectral forms are presented. The possibilities of the use of porphyrin-diketones as longwave fluorescent and phosphorescent probes are discussed.

Five novel photosensitizers for photodynamic tumor therapy have been offered as potential drug substances for the prospective clinical usage. A complex of pre-clinic biological studies has been performed, unambiguously demonstrating low systemic toxicity and high photodynamic potential of the substances proposed. The one named 'Photodithazine/Photochlorin I' has already successfully passed a number of clinical test on patients having immergent vital indications.

In 1998 a new second-generation photosensitizer was introduced in Russia - a chlorin e 6 derivative named Photodithazine - water-soluble preparation with absorption peak at 668 nm. This novel preparation has passed pre-clinical in vitro and in vivo studies. In 1999 a limited series of clinical testing of photodynamic therapy with photodithazine has been performed in the State Research Center for Laser Medicine of Russian Ministry of Public Health. In comparison to hematoporphyrin derivative (Photofrin, Photosan, Photoheme etc.) and aluminum phthalocyanine (Photosense) used in clinical practice, Photodithazine has a number of special properties. Most significant of them is rapid tissue accumulation and clearance from the organism within 48 - 72 hours. Photodithazine was used as a photosensitizer for treatment of 44 patients with malignant tumors of skin, oral cavity, larynx, bronchi, esophagus, stomach, vulva and other locations. The dose of photodithazine ranged from 0,8 to 1,2 mg/kg of body weight. Laser irradiation has been performed on a solid-state laser device Poljus-2 with wavelength 670 nm and output power 1000 mW. Energy density ranged from 100 to 400 J/cm 2 . Laser irradiation of the inner organ was performed via flexible endoscope. PDT with Photodithazine was not accompanied with systemic and local complications. The treatment was well-tolerated by the patients, including those with severe accompanying diseases, There were no complications associated with possible high skin photosensitivity in all treated group. Therapeutic effect took place in all cases, including complete tumor regression in 22 cases (53,7%) and partial regression in 19 cases (46,3%). By present time clinical PDT with Photodithazine is in the initial state. Photodithazine-based PDT appears to be a very promising technique, but analysis of long-term follow-up results is necessary to confirm its therapeutic efficacy. Further research is required to estimate adequate doses of drug and light for various types of tumors.

In vitro and in vivo biological evaluation of a novel drug substance 'photodithazine' has been performed. In vitro photocytotoxicity (EC50) was 1 (mu) M together with some cytotoxicity. In vivo acute toxicity has been found to be 170 mg/kg for KD10, 175 mg/kg for LD16, 197 mg/kg for LD50, 220 mg/kg for LD84 following 1 percent aqueous solution i.v. injection. Pharmacokinetics and biodistribution studies have been done using the same mice bearing inoculated under the skin of the flanks T36 embryocarcinomas injected i.v. with 40 mg/kg dose of the drug. Maximal tumor and most organs' uptake was attained 1 h.p.i., however, the drug's level in the organs rapidly decreasing to zero with the best tumor/muscle ratios over 10 by 5 h.p.i. 'Photodithazine' has been found to possess rapid clearance from the organism: 94 percent elimination 24 h.p.i. and 98 percent to 48 h.p.i. PDT has been performed in vivo involving 21-23 g A/Snell mice with the same type 0.9-1 g tumors injected 40 mg/kg drug i.p., with 670 nm light being delivered 5-6 h p.i. at different doses. The best irradiation dose has been found to be 170 J/cm2 with a sound necrotic effect and 1-3 week remission. Thus, 'Photodithazine' represents a potent photosensitizer for PDT.© (2000) COPYRIGHT SPIE--The International Society for Optical Engineering. Downloading of the abstract is permitted for personal use only.

Deuteroporphyrin IX derivatives are prospective PDT porphyrin photosensitizers (PS). The photodynamic effects of 6 new amphiphilic deuteroporphyrin derivatives with different hydrophobicity, as well as effects of known photosensitizers Photoheme and Photosens (used for comparison) on the firing of isolated crayfish mechanoreceptor neuron have been studied. After 30 min photosensitization, neurons were irradiated with He-Ne laser (632.8 nm, 0.3 W/cm2), and changes in neuron firing frequency were recorded. It has been shown that neuron firing is very sensitive to photodynamic impact and can serve as a sensitive indicator of cell photodamage.

The deuteroporphyrin-IX derivative Dimegin [2,4-di-(alpha-methoxyethyl)-deuteroporphyrin IX] was investigated with respect to cellular uptake, intracellular localization and cell survival following photodynamic treatment in human cell lines derived from the skin (SCL1 and SCL2, squamous cell carcinoma; HaCaT keratinocytes; N1 fibroblasts). Using flow cytometry, we determined the cellular fluorescence as a marker of the uptake of Dimegin after incubation for 24 h. The intracellular localization of Dimegin was analysed using fluorescence microscopy and co-staining with fluorescent dyes specific for cell organelles. Following irradiation with an incoherent light source (580-740 nm) using a light dose of 24 J/cm2, phototoxicity was determined by means of trypan blue dye exclusion, MTT assays and growth curves. The relative Dimegin fluorescence of the different cell lines declined as follows: SCL1 > HaCaT > N1 > SCL2. Intracellular localization of Dimegin was found in the mitochondria. For all cell lines Dimegin concentrations above 15 microM yielded a significant phototoxic effect. The EC50 for SCL1 cells was 8.9 +/- 2.0 microM Dimegin. The EC50 for the cell lines increased as follows: SCL1 < HaCaT < N1 < SCL2, thus correlating with the cellular fluorescence of Dimegin. The results of the MTT assay were confirmed by trypan blue dye exclusion assay and growth curves. In conclusion, the study shows that Dimegin is an effective photosensitizer with a rapid mechanism of action in vitro, resulting in an immediate loss of plasma membrane integrity following irradiation.

We report on a study on the protonation of representative porphyrins, metalloporphyrins, porphyrin-ketones and a Pd-porphyrin Schiff base in aqueous solution and when dissolved in plasticized PVC membranes. The respective protolytic forms were characterized by absorption and emission spectra as well as apparent p K a values. It is shown that the protonation of porphyrins dissolved in PVC membranes is much more difficult than in solution, resulting in an extraordinarily large decrease in the apparent p K a values which can be as large as 5 units when compared to the corresponding data for the porphyrins in solution. In certain cases, no protonation at all occurred within the pH 2–12 range. At the same time, N-MeEP which is much more basic (its intrinsic p K being 11.2) can be protonated in such membranes into the monocation at weakly acidic pH, but the apparent p K a also is lowered by about 6 units. On addition of a tetraphenylborate anion to the PVC membranes, protonation was found to proceed much easier. Both the monocation and dication was identified in case for the porphyrins, while for the prophyrin-ketones protonation into dication took place, and N-methyl-etioprophyrin was present as the monocation over the whole pH 2–12 range. Membranes composed of solutions of porphyrin-ketones or the Pd-porphyrin Schiff base in plasticized PVC were identified as being useful for purposes of continuously sensing pH in the physiological range. Their response time, calibration curves, signal stability and effects of buffer ionic strength and temperature were investigated in more detail.

Polybiotinylated porphyrinoids and linear alkyl-dibiotins were used to describe accumulation behavior of avidin and POD-labelled avidin on tumor cells preincubated with biotinylated monoclonal antibodies. The accumulation behavior followed more a liner than a formerly postulated branched pattern. This allows for us to calculate necessary repeats of accumulation in order to acquire the necessary amount of sensitizer molecules per tumor cell at low concentration of tumor specific marker molecules at the cell surface.© (1996) COPYRIGHT SPIE--The International Society for Optical Engineering. Downloading of the abstract is permitted for personal use only.

The accumulation of fluorescent dyes and anticancer drugs in tumors is a fundamental requirement for tumor-diagnosis and therapy. We tested various photosensitizers on vascularized microtumors in the hen's egg test system according to their distribution and accumulation by in-situ videofluorescence microscopy and their influence on tumor blood flow by laser- Doppler spectroscopy. Only 2,4-di((alpha) -methoxyethyl)- hematoporphyrin showed strict tumor localization while diglucosamidyl-heptyl-chlorin labeled cell membranes exclusively. Results from laser-Doppler spectroscopy did not show any influence on tumor blood flow. Intelligent combining of the properties of these sensitizers with accumulation strategies should show in principle strict tumor localization without targeting of non-tumor tissues.© (1996) COPYRIGHT SPIE--The International Society for Optical Engineering. Downloading of the abstract is permitted for personal use only.

The photodynamic therapy is highly promising for its dual specificity, namely, for the site of accumulation of the phototoxic substance and for the site of action of laser. A drawback of the PDT technique is a low selectivity of the photosensitizers accumulation in tumor tissue. Therapeutic doses of photosensitizers are fairly high, that leads to unfavorable effects (toxicoses, lesions of normal skin, prolonged rehabilitation). This drawback can be overcome is photoimmunotoxins are used, i.e. conjugates of photosensitizers with antibodies to specific tumor markers. We tried to introduce a simple model for estimation of singlet oxygen effect on cells for the case of free photosensitizers, presented in outside solution, and in case of conjugated dye, bound by cell surface receptors. To achieve the same effect the required concentration of conjugated photosensitizer can be significantly decreased (50 to 100 fold) as compared with free dye. Methods for porphyrins and different phthalocyanines and naphthalocyanines conjugation with protein were developed. The use of bacteriophage T4 as a model for studying the action of photoimmunotoxines on virus, unambiguously demonstrated the applicability of this approach for targeted destruction of viral particles.

The kinetics of photosensitizer distribution and elimination have been studied using fluorescent methods in organs and tumors of A/Snell mice with embriocarcinoma inoculated into their thigh muscles for the porphyrin compound 2,4-di((alpha) -methoxyethyl)deuteroporphyrin-- IX (DMH, `Dimehin') and its complex with polysaccharide chitosan. DMH fluorescence differs in samples of liver and faeces which follows from the spectra comparison. DMH is metabolizable upon passing through liver into a form eliminated by the gastrointestinal tract as our pharmacokinetic data have shown. DMH has been found to be a short-term highly photodynamically efficient photosensitizer judging by combined analysis of our toxicological, pharmacokinetic and photodynamic research data. DMH-chitosan uptake and distribution studies have shown the complex's long-term persistence in blood circulation, high level accumulation in spleen and lungs, whereas there was no complex registered in tumors and other tissues following i.v. administration.© (1996) COPYRIGHT SPIE--The International Society for Optical Engineering. Downloading of the abstract is permitted for personal use only.