Polyhedral Borane Anions: Investigation of the Mechanism of Retention
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The current therapies for cancer, such as radiation, surgery, and chemotherapy, are lacking in specificity and incomplete removal of the cancer tissues. Radiation and chemotherapy both result in the damage or destruction of normal cells. The incomplete removal of the cells by surgery leaves more cells to regenerate and the cells could possibly migrate to other regions of the body. These flaws are exacerbated in aggressive forms of cancer such as glioblastoma multiforme. Glioblastoma multiforme occurs in the brain, which profoundly affects the severity of destruction and or damage to the cells around the tumor. The ineffectiveness of these treatments, both adjunctively and synergistically, has lead to a need for a different method of treatment.
Boron neutron capture therapy (BNCT) is a binary cancer therapy believed to be an improved method to treat aggressive forms of cancer. The first step of the therapy requires the localization of the boron-10 isotope in the tumor cells. Once the boron-10 isotope is localized, the area is irradiated by neutrons, ultimately resulting in the formation of an 'a'-particle, a lithium particle, and a large amount of energy. The linear energy transfer from the fission process dissipates within the distance of one cell diameter and causes death of the cell where the boron-10 isotope was located. The development of BNCT has been limited by the lack of boron-containing compounds that are taken up and retained in the tumor cells.
One method that has been utilized to deliver boron-containing compounds specifically to the interior of the tumor cells is incorporation of the compounds into unilamellar liposomes. A variety of polyhedral borane anions have been encapsulated into unilamellar liposomes and evaluated in murine biodistribution experiments. Based on the results of the experiments, three mechanisms of retention have been proposed. The first mechanism is nucleophilic attack at the electron-deficient binding region of, for example, the [n-B20H18]2- ion. The second mechanism of retention is oxidation of the reduced anions, such as the [ae-B20H17NH3]3- ion, to the oxidized species, such as the [n- B20H17NH3]– ion, which is susceptible again to nucleophilic attack. The third mechanism involves thiol-containing ions, such as the [a2B20H17SH]4- ion, and is based on the proposed formation of a disulfide bond between the polyhedral borane anion and an intracellular protein sulfhydryl moiety on a cysteine residue.
The evaluation of the products formed from the reaction of three polyhedral borane anions, [a2-B20H17SH]-4, [n-B20H18]2-, and [ae-B20H17OH]4-, with select biomolecules was the focus of this research. The products of the reactions were evaluated using 1H and 13C NMR spectroscopy, COSY and HETCOR NMR experiments, electrospray ionization (ESI) mass spectrometry, matrix assisted laser desorption ionization time of flight (MALDI-TOF) mass spectrometry, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), and native PAGE. The 13C NMR spectrum of the product of the reaction between cysteine and the [a2-B20H17SH]-4 ion and the product of the reaction between reduced glutathione and the [a2-B20H17SH]-4 ion both exhibited a shift in the peak assigned to the ß-carbon of cysteine. The ESI-MS spectrum of the product formed from the reaction between reduced glutathione and the [a2-B20H17SH]-4 ion contained a new product peak which was attributed to the formation of a covalent bond between the two molecules. The SDS-PAGE and the native PAGE were used to evaluate the reaction of human serum albumin (HSA) with the polyhedral borane ions. The SDS-PAGE results were uninformative due to the association of the HSA with itself under the reaction conditions; however, the native PAGE yielded a difference in migration of the product formed from the reaction between HSA and the [a2-B20H17SH]-4 ion and the control HSA which was consistent with a strong interaction between the two molecules. The results from the MALDI-TOF experiment contained too much of the keratin contaminant and lacked the peaks characteristic of even the control HSA spectrum. The combination of the experiments provide significant evidence of the ability of the [a2-B20H17SH]-4 ion to form a covalent disulfide bond with biomolecules that contain a cysteine amino acid. The [n-B20H18]2- and [ae-B20H17OH]4- ions did not exhibit any results with cysteine, reduced glutathione, or HSA, which would be consistent with the formation of a covalent bond.