Synthesis of a porphyrin-labelled carboranyl phosphate diester: a potential new drug for boron neutron capture therapy of cancer.

A boron-rich, water-soluble porphyrin conjugate was synthesized by coupling of two carboranyl alcohols with 2-chlorophenoxyphosphorus dichloride, followed by conjugation to an amine-functionalized tetraphenyl-porphyrin via an amide linkage.

Localization and photodynamic efficacy of two cationic porphyrins varying in charge distributions.

Two meso-tetraphenylporphyrin derivatives bearing adjacent: 5,10-di[4-(N-trimethylaminophenyl)-15,20-diphenylporphyrin (DADP-a) or opposite: 5,15-di[4-(N-trimethylaminophenyl)-10,20-diphenylporphyrin (DADP-o) cationic-N-(CH3)3+ groups on two of the para-phenyl positions were examined with regard to photodynamic properties as a function of charge distribution. The two adjacent positive charges in the DADP-a structure result in a molecular distortion (asymmetry), likely from electrostatic repulsion. This could be responsible for the unusual interaction of this compound with some solvents and detergent micelles. In contrast, DADP-o is a much more symmetric molecule. In a cellular environment, fluorescence spectra of the two agents were essentially identical. Subcellular localization played a major role in photodynamic efficacy. DADP-a localized in mitochondria, and irradiation of photosensitized cells (640-650 nm) resulted in a rapid loss of the mitochondrial membrane potential (delta(psi)m), usually a prelude to apoptotic cell death. In contrast, DADP-o localized in lysosomes, and extensive lysosomal photodamage was observed after irradiation. Both steady-state accumulation levels and absorbance spectra favored DADP-o, but the light dose required for a 90% cell kill was two-fold greater for DADP-o than for DADP-a, at a constant extracellular sensitizer concentration. These data indicate that, on a photons/cell basis, DADP-a was five-fold more efficacious. Fluorescence emission spectra in different solvents and detergents demonstrated a tendency for DADP-a association. We interpret these results to indicate partition of both drugs to membrane loci, with mitochondriabeing the more lethal site for photodamage.

Effect of overall charge and charge distribution on cellular uptake, distribution and phototoxicity of cationic porphyrins in HEp2 cells.

Five cationic porphyrins bearing one to four -N(CH(3))(3)(+) groups linked to the p-phenyl positions of 5,10,15,20-tetraphenylporphyrin (TPP) were synthesized in order to study the effect of overall charge and its distribution on the cellular uptake, phototoxicity and intracellular localization using human carcinoma HEp2 cells. The di-cationic porphyrins DADP-o and DADP-a accumulated the most within cells and preferentially localize within vesicular compartments and in mitochondria. Of these two only DADP-a was phototoxic to the cells (IC(50)=3 microM at 1 J/cm(2)). The mono-cationic porphyrin MAP was found to be the most phototoxic of the series, and it localized mainly in lipid membranes, including the plasma membrane, ER, mitochondria, and Golgi. Both the tri-cationic porphyrin TRAP and the tetra-cationic porphyrin TEAP localized subcellularly mainly in the mitochondria, but of the two only TEAP showed moderate phototoxicity (IC(50)=8 microM at 1 J/cm(2)). Our results suggest that MAP is the most promising PDT photosensitizer, and that both DADP-o and TRAP might find application as transport vehicles for therapeutics into cells.

Synthesis and cellular studies of a carboranylchlorin for the PDT and BNCT of tumors.

The syntheses of closo- and nido-carboranylchlorins 4 and 5 from a known carboranylporphyrin are described. Water-soluble nido-carboranylporphyrin 5 was found to have very low dark cytotoxicity (IC50 > 500 microM using a MTT-based assay) but to be toxic in the presence of red light (IC50 = 80 microM at 0.55 J/cm2 light dose). Under the same experimental conditions, carboranylchlorin 5 was taken up by human glioma T98G cells to a significantly higher extent than chlorin e6, a chlorophyll degradation product. The preferred sites of subcellular localization of carboranylchlorin 5 were found to be the cell lysosomes. Our results suggest that carboranylchlorin 5 is a promising new dual sensitizer for the PDT and BNCT treatment of tumors.

Synthesis and cellular studies of an octa-anionic 5,10,15,20-tetra[3,5-(nido-carboranylmethyl)phenyl]porphyrin (H(2)OCP) for application in BNCT.

The total synthesis of 5,10,15,20-tetra[3,5-(carboranylmethyl)phenyl]porphyrins 2-5 containing 36-43% boron by weight are reported. All compounds were characterized by spectroscopic methods and, in the case of 2, by X-ray crystallography. The water-soluble nido-carboranylporphyrin 5 (H(2)OCP) was found to have low dark toxicity toward V79 lung fibroblasts (CS(50) > or = 250 microM), to be readily taken up by human glioblastoma T98G cells in culture and to localize subcellularly preferentially in the cell lysosomes. In comparison with a known tetra(nido-carboranyl)porphyrin (6), H(2)OCP (5) is taken up slower and to a lower extent by T98G cells, possibly as a result of its higher hydrophilic character. The metal-free H(2)OCP (5) was also found to accumulate to a higher extent in T98G cells compared with its zinc(II) complex analog 4. Our studies show that carboranylporphyrins bearing eight nido-carborane cages can still accumulate intracellularly and have low dark toxicity toward cells in culture, and therefore might have promise for application in BNCT.