Development of an enzyme-linked immunosorbent assay (ELISA) for antitumor agent MKT 077

A sensitive enzyme-linked immunosorbent assay (ELISA) for a novel antitumor agent MKT 077, which is currently subjected to investigation in a Phase I clinical trial, has been developed and applied to the evaluation of plasma levels in a dog. MKT 077 was structurally modified by the attachment of the 4-carboxylbutyl group at the pyridinium ring through which it was subsequently coupled with carrier protein, KLH. This conjugate was used for producing antibodies against MKT 077 in rabbits. The antibodies thus obtained were characterized with respect to their cross reactivity toward MKT 077-related compounds and found to recognize benzothiazole and 4-oxothiazolidine moieties. Two types of assay methods, antigen- and antibody-immobilized methods, using the antibodies, were compared for their ability to detect MKT 077. The heterologous antibody-immobilized method, which had the lower detection limit, was chosen for use in assays. This method enabled the detection of as little as 20 pg of MKT 077 in phosphate-buffered saline, with satisfactory precision. The plasma levels of MKT 077 in a dog were determined by use of the proposed method.