Amino acid chloramines (AA-Cl) are reactive compounds generated at the sites of inflammation by the reaction of hypochlorous acid with amino acids. It has been thought that only small uncharged chloramines such as ammonia chloramine can cross the cell membrane. Consequently, it was believed that AA-Cl do not contribute much to inflammation induced cellular damage. Our work has shown that chloramines of proline, arginine, and glycine are membrane permeable and can reach the mammalian cell nucleus and damage the proliferating cell nuclear antigen (PCNA), a major component of DNA replication and repair machinery. The PCNA damage included covalent crosslinking of PCNA subunits, peptide backbone cleavage, and oxidation of methionine residues. Our work also have revealed that cysteine 148, located at the PCNA subunits interface, is required for PCNA crosslinking by AA-Cl. Our results suggested intersubunits sulfonamide formation as a mechanism of crosslinking. We concluded that some AA-Cl generated at the inflammation sites can penetrate into the surrounding normal tissues and modify critical nuclear proteins. Such protein modification may contribute to a variety of inflammation induced diseases.
Photodynamic therapy (PDT) employs visible light and photosensitizers (PS) to cure cancer. After PS administration, light is focused locally on the tumor tissue, activating the PS which in turn destroys the irradiated tumor cells. A common observation is the shrinkage of distant un-irradiated tumors after PDT of a single tumor. This systemic response against tumors is thought to be immunological. The possible role of photodynamic protein modification in such a global immune response has been underexplored. We hypothesized that photodynamic modification to tumor cell proteins might act as an immunological adjuvant, stimulating an immune response against tumor specific proteins. Photo-adducting of the PS to tumor specific proteins could be one molecular mechanism. We have investigated photo-addition of acridines PS to mammalian cellular proteins. Our results have shown that acridine PS photo-adducted to wide range of cellular proteins and the adduct is formed through oxidized histidine residues. We conclude that photodynamic treatment can modify cellular proteins via photo-addition of the PS. Photo-addition of PS to normal cellular protein may generate potentially immunogenic haptens. Moreover, photo-addition of PS to the normally low immunogenic tumor specific proteins may enhance their immunogenicity, inducing a robust systemic immune response. Such immune response can help in the eradication of both locally irradiated and distant, un-irradiated tumors of the same type.
Camptothecins, a class of anticancer drugs, kills mammalian cells by stabilizing the topoisomerase I-DNA strand passing intermediate which in turn is converted to lethal double strand DNA breaks. Camptothecin-stabilized topoisomerase I-DNA cleavage intermediates in mammalian cells are uniquely modified by ubiquitin-family proteins. The structure, composition, and function of these ubiquitin-family modifications are poorly understood. We have analyzed these ubiquitin-family modifications in human breast cancer cells using mass spectrometry. Peptides shared by SUMO-2 and SUMO-3 were abundant, and a peptide unique to SUMO-2 was identified. Ubiquitin was also identified while no SUMO-1 peptide was detected.