Inter-individual variability in human responsiveness to carcinogens, drugs and environmental chemicals may reside in the ability to metabolize these chemicals to detoxified or toxic products. It is necessary to define the role of individual cytochrome P450s in the metabolism of the vast number of compounds to which humans are exposed. The ideal and most precise method for this evaluation is with the use of inhibitory monoclonal antibodies to specific P450s which upon addition to tissue preparations inhibit the metabolism of the substrate. This inhibition defines the amount of metabolism catalyzed by a specific P450. The MAbs are precise reagents which can determine the activity of a single P450 towards a substrate in the presence of the tissue complement of P450s. Initially we have isolated inhibitory monoclonals and some non-inhibitory immunoblotting monoclonals which determine P450 amount to rat, mouse, rabbit and fish P450s. To date we have isolated 16 monoclonal antibodies that are highly specific towards rodent and fish P450s. P450s of rodents and humans may vary significantly in activity. We developed a program to isolate monoclonal antibodies that immunoblot and/or inhibit human P450s. This was made possible by the use of cDNAs for P450 expressed from vaccinia or baculovirus vectors. Individual P450s were used either as antigens or for screening the monoclonal antibodies. We have succeeded in isolating seven monoclonal antibodies to five different human cytochrome P450s. These are P450 3A4, 2E1, 1A1 2C8/9, and 2B6. This P450 3A4 metabolizes more than 70-80% of all known drugs, some carcinogens and steroids. The 2E1 metabolizes small molecules of environmental importance. The 1A1 metabolizes polycyclic hydrocarbons and 2C8/9 metabolizes a variety of both drugs and carcinogens. We have analyzed the immunoblot, specificity and inhibitory activity of the monoclonal antibodies which we produced in the previous years. These include P450 3A4, 2E1, 1A1, 2C8/9/19 and 2B6. This year we have successfully produced highly specific immunoblot and inhibitory monoclonal antibodies to the polymorphic P450s, 2D6, 2A6, 2C9 and to P450 1A2. The MAbs that we have isolated are the most powerful tools that determine in-situ the role of each P450 in drug and carcinogen metabolism. The MAb can determine drug-drug interactions, side effects, determine the individual sensitivity to the type of drug and the proper dosage. This work is of considerable importance in drug metabolism, carcinogenesis, and human polymorphisms.
