ABSTRACT
Cancer immunotherapy using T cells redirected with chimeric antigen receptor (CAR) has shown a lot of promise. We have established a single-chain antibody (scFv) generation system in which scFv library-expressing CAR-T cells can be screened appropriately based on their antitumor functions. A variable region library containing the variable and J regions of the human immunoglobulin light or heavy chain was fused with the variable region of a heavy or light chain encoded by an existing tumor-specific antibody to generate a new scFv library. Then, scFv library-expressing CAR-T cells were generated and stimulated with target cells to concentrate the antigen-specific population. Using this system, target-specific recognition of CAR-T cells appeared to be finely tuned by selecting a new variable region. Importantly, we have demonstrated that the newly optimized scFv-expressing CAR-T cells had better proliferation capacity and durable phenotypes, enabling superior reactivity against advanced tumors in vivo in comparison with the original CAR-T cells. Therefore, the optimization of an scFv is needed to maximize the in vivo antitumor functions of CAR-T cells. This system may allow us to adjust an immunological synapse formed by an scFv expressed by CAR-T cells and a target antigen, representing an ideal form of CAR-T-cell immunotherapy.
Subject(s)
Immunoglobulin Heavy Chains/metabolism , Immunoglobulin Light Chains/metabolism , Immunoglobulin Variable Region/metabolism , Immunotherapy, Adoptive , Lymphoma/therapy , Receptors, Chimeric Antigen/metabolism , Single-Chain Antibodies/metabolism , T-Lymphocytes/transplantation , Animals , Female , Humans , Immunoglobulin Heavy Chains/genetics , Immunoglobulin Light Chains/genetics , Immunoglobulin Variable Region/genetics , Immunological Synapses , Jurkat Cells , K562 Cells , Lymphoma/genetics , Lymphoma/immunology , Lymphoma/metabolism , Mice, Inbred NOD , Mice, SCID , Receptors, Chimeric Antigen/genetics , Single-Chain Antibodies/genetics , T-Lymphocytes/immunology , T-Lymphocytes/metabolism , Tumor Burden , Xenograft Model Antitumor AssaysABSTRACT
Contamination of surface water by antibacterial pharmaceuticals (antibacterials) from clinical settings may affect aquatic organisms, plants growth, and environmental floral bacteria. One of the methods to decrease the contamination is inactivation of antibacterials before being discharged to the sewage system. Recently, we reported the novel method based on electrolysis for detoxifying wastewater containing antineoplastics. In the present study, to clarify whether the electrolysis method is applicable to the inactivation of antibacterials, we electrolyzed solutions of 10 groups of individual antibacterials including amikacin sulfate (AMK) and a mixture (MIX) of some commercial antibacterials commonly prescribed at hospitals, and measured their antibacterial activities. AMK was inactivated in its antibacterial activities and its concentration decreased by electrolysis in a time-dependent manner. Eighty to ninety-nine percent of almost all antibacterials and MIX were inactivated within 6h of electrolysis. Additionally, cytotoxicity was not detected in any of the electrolyzed solutions of antibacterials and MIX by the Molt-4-based cytotoxicity test.
Subject(s)
Anti-Bacterial Agents/chemistry , Electrolysis , Environmental Pollutants/chemistry , Waste Disposal, Fluid/methods , Anti-Bacterial Agents/pharmacology , Cell Line , Cell Survival/drug effects , Environmental Pollutants/toxicity , Escherichia coli/drug effects , Hospitals , Humans , Sewage/chemistry , Staphylococcus aureus/drug effectsABSTRACT
Wastewater from clinical institutions contains a considerable amount of toxic substances. Among the toxic substances, antineoplastics may induce carcinogenesis, teratogenesis, and the emergence of mutant microorganisms in the environment. Although the incineration or chemical treatments of disposed antineoplastics are recommended, a high energy during incineration and a careful quality control during chemical treatment are required. In this study, we determined the conditions for the electrolytic treatment of an antineoplastic, epirubicin hydrochloride (EH), using two platinum electrodes with a constant current of 100 mA. We analyzed the cytotoxicity, mutagenicity and antibacterial activity of electrolyzed EH and compared them with those of unelectrolyzed EH. Nearly 100% cytotoxicity, mutagenicity and antibacterial activity were eliminated and HPLC did not detect an EH molecule, in the case of electrolysis for 6 h. We also examined the biological cytotoxicities of electrolyzed irinotecan hydrochloride, vincristine sulfate, mitomycin C, paclitaxel, methotrexate and cisplatin, and found that 72.1-99.999% toxicity was eliminated by electrolysis under the same conditions. The biological toxicity of a mixture of these drugs was determined to be decreased by approximately 99% by electrolysis under the same conditions.