Alan R. Morgan

SITES and EFFICACY OF PHOTODAMAGE BY TIN ETIOPURPURIN in vitro USING DIFFERENT DELIVERY SYSTEMS

Photosensitization by tin etiopurpurin (SnET2) was determined in cell culture using sensitizer dissolved in ethanol or solubilized via three different delivery systems: Cremophor EL, γ‐cyclodextrin or Molecusol (a more water‐soluble cyclodextrin derivative). Sensitizer uptake was substantially more efficient when the delivery systems were employed, in terms of intracellular level needed to lethally photosensitize cells. We observed photodamage at both membrane and mitochondrial loci, but the former was better correlated with loss of viability.

METALLOPURPURINS and LIGHT: EFFECT ON TRANSPLANTABLE RAT BLADDER TUMORS and MURINE SKIN

Purpurins are modified chlorins with photodynamic properties. Their strong absorption in the red region of the visible spectrum makes them candidates for use in photodynamic cancer therapy. A series of metal derivatives of the free base purpurins have been synthesized and shown to cause tumor necrosis in transplantable tumors when exposed to visible light. In the following set of experiments, the effects of two metallo‐derivatives (tin and zinc) of two purpurins, octaethylpurpurin (NT2) and etiopurpurin (ET2), and light on the N‐{4‐(5‐nitro‐2‐furyl)‐2‐thiazolyl}formamide transplantable tumors in Fischer CDF(F344)/CrlBr rats were studied. The photodynamic activity was assessed by a short term assay using tumor dry weight 12 days after purpurin‐PDT as a criterion of response. From these experiments it appears that SnET2>SnNT2>ZnET2>ZnNT2 in photodynamic activity. SnET2 was further characterized by attempting to determine the time interval after systemic injection at which maximum therapeutic effect occurred. These studies shown that 24 h after metallopurpurin injection was the optimum time for treatment of tumors with visible light. In a final set of experiments, the effect of solar light on the skin of hairless mice injected with SnET2 was found to be much less injurious than with hematoporphyrin derivative.

A Dose Response Analysis of Purpurin Derivatives Used as Photosensitizers for the Photodynamic Treatment of Transplantable Fanft Induced Urothelial Tumors

Rats engrafted with transplantable N-[4-(5-nitro-2-furyl)-2-thiazolyl] formamide (FANFT) induced urothelial tumors were treated with purpurin derivatives and red light (greater than 590 nm., 360 joules/cm.2). Treated and control tumors were harvested 12 days after treatment and dry weights compared. At doses of 5.0 micrograms./gm. and 2.5 micrograms./gm. body weight, the purpurins designated NT1 and NT2, when combined with light, caused statistically significant (p less than 0.02) tumor regression when compared to light shielded controls. At 1.0 micrograms./gm. body weight, NT1 and light also induced significant tumor regression (p less than 0.02). Purpurins, which have strong absorption bands above 650 nm. and can be synthesized with a high degree of purity, appear to have potential as photosensitizers for photodynamic therapy.

COPPER BENZOCHLORIN, A NOVEL PHOTOSENSITIZER FOR PHOTODYNAMIC THERAPY: EFFECTS ON A TRANSPLANTABLE UROTHELIAL TUMOR

Abstract— An iminum salt of octaethylbenzochlorin with copper in the aromatic ring, CDSI, was tested for its tumoricidal effects on theA–27 N‐[4‐(5‐notro‐2‐furyl)‐2‐thiazoly] formamide tumor line. CDSI was found to be an effective photosensitiz4r in vivo when used in combination with either a xenon arc lamp or a pulsed alexandrite laser. Hemodynamically, CDSI and light caused a rapid decrease in tumor blood flow. skin photosensitization was found to be minimal when drug‐injected mice werwe illuminated in a solar simulator.

Observations on the synthesis and in vivo photodynamic activity of some benzochlorins

Abstract— An improved synthesis of benzochlorins is reported. Demetallation of the meso‐hydroxymethylvinyl derivative of octaethylporphyrin, followed by treatment with sulfuric acid results in cyclization to generate the corresponding octaethylbenzochlorin in high yield. Prolonged treatment with acid generates the sulfonated derivative. These sensitizers were shown to be efficient photodynamic agents in vivo. Animals bearing a transplanted N‐{4‐(5‐nitro‐2‐furyl)‐2‐thiazolyl}formamide induced urothelial tumor were treated with either the benzochlorin or its sulfonated derivative. Irradiation of tumors 24 h later resulted in a significant tumoricidal effect in a short term assay. We conclude that benzochlorins warrant further examination as potential agents for use in photodynamic therapy.

Synthesis of carboranyl porphyrins: Potential drugs for boron neutron capture therapy

Abstract Amphiphilic carboranyl porphyrins were synthesised starting from commercially available hematoporphyprin for their potential use as boron neutron capture therapy agents.

An in vitro study of boronated porphyrins for potential use in boron neutron capture therapy

Abstract A series of boronated hematoporphyrin analogs differing in lipophilicity and in the length of the di-ether side chains, and their zinc derivatives were evaluated for use in boron neutron capture therapy. The porphyrins displayed no inherent cytotoxicity towards V79 cells at a concentration of 300 μM. Cellular uptake varied as a function of lipophilicity and metallation.

Tin (IV) Etiopurpurin Dichloride: An Alternative To DHE?

Etiopurpurin is a photosensitizer developed from etioporphyrin I, which we have shown to be useful in PDT. Incorporation of tin into the macrocycle gives a derivative which appears to be more effective both in vitro and in vivo studies on tumor cytotoxicity, than the parent purpurin. In vitro studies have shown not only that tin etiopurpurin dichloride (SnET2C12) is an efficient photosensitizer, but also that its action spectrum appears to be consistent with both the absorption and emission spectra. In vivo studies include a histological examination of tumors treated with SnET2C12 and PDT which suggests that disruption of the tumor vasculature plays an active role in tumor necrosis; a twelve day dose response study to determine effective dose; a thirty day study to determine tumor regrowth; a study of normal tissue response to PDT and a study to determine the optimum time for phototherapy after injection of the drug. For these studies, the FANFT induced urothelial tumor line was used, although we have recently initiated studies on the R3327 AT rat anaplastic adenocarcinoma, a tumor which appears to be more resistant to PDT.

PHOTOSENSITIZATION WITH ETIOBENZOCHLORINS AND OCTAETHYLBENZOCHLORINS

The photophysical and photobiological properties of a series of etiobenzochlorins were evaluated in cell culture using murine leukemia L1210 cells. In the series of agents tested, the chlorin‐(mono)sulfonate was the most efficacious, the tin chlorin somewhat less so and the tin chlorin‐sulfonate much less active. The parent chlorin was essentially inactive at the limit of solubility. Photodamage was assessed by measuring alterations in surface hydro‐phobicity (via a two‐phase partitioning procedure), amino acid transport and membrane potential. Additional information was provided from fluorescence microscopy, which was used to identify sites of sensitizer binding and effects of photodamage on the binding patterns of fluorescent probes specific for mitochondria, lysosomes and plasma membranes. Effects of photodamage on fluorescence lifetime distribution of the membrane probe trimethylamino‐diphenyl hexatriene were examined. The data obtained were consistent with localization of the parent etiobenzochlorin and tin derivative at lysosomal loci. the chlorin‐sulfonate at plasma and mitochondrial membranes and tin‐sulfonate at the cell surface.

Purpurins: Improved Photosensitizers For Photodynamic Therapy

Two new groups of photosensitizers, purpurins and metallopurpurins were tested for photodynamic activity when combined with visible light (>590nm), in the treatment of transplantable urothelial tumors (FANFT induced) in Fischer 344 rats. In preliminary studies, animals were injected with 10 mg/kg body weight (b.w.) of the free base purpurins (NT1, NT2, NT2H2, GG1, ET2, ET2H2 and JP1) and treated with 360J/cm2 of red light. Tumors were excised, fixed in formalin and stained for light microscopic examination. Purpurin based photodynamic therapy (PDT) was found to cause extensive hemorrhagic tumor necrosis. Control tumors shielded from the light showed no histological changes. These experiments were repeated using the same experimental design for the metallopurpurins (ZnET2, SnET2, ZnNT2H2, SnNT2H2, AgNT2H2 and ZnNT1). These compounds were also found to cause extensive hemorrhagic necrosis. These studies were followed by a dose response analysis in which groups of animals were treated with decreasing doses of drug while the light dose was kept constant. Response was determined by tumor dry weight, 12 days after completion of PDT. For the free base purpurins, drug doses as low as 1.0 mg/kg b.w. caused significant tumor regression while seleted metallopurpurins caused tumor regression at doses as low as 0.5 mg/kg b.w. In a series of related experiments the effect of purpurins on tumor blood flow was investigated since it has previously been shown that tumor blood flow is disrupted during porphyrin and phthalocyanine phototherapy. By using the radioactive microsphere technique, it was shown that both the free base purpurins and metallopurpurins caused a rapid decrease in tumor blood flow.