Induction of catalytic activity in proteins by lyophilization in the presence of a transition state analogue.

The induction of catalytic activity in proteins by lyophilization in the presence of a transition state analogue (biomolecular imprinting) has been attempted. It was shown that proteins which were freeze-dried with n-isopropyl-4-nitrobenzyl-amine (a transition state analogue for the reaction of dehydrofluorination of 4-fluoro-4-[p-nitrophenyl] butan-2-one) displayed higher beta-elimination activity as compared to their-non-imprinted counterparts. It was also found that native bovine serum albumin has a high dehydrofluorination activity towards the above substrate with kinetic parameters rather similar to those of a catalytic antibody prepared by Shokat et al. (1989). A comparison of the kinetic parameters determined in this study with those obtained for analogous catalytic antibodies and imprinted polymers was made.

Preliminary studies for an immunotherapeutic approach to the treatment of human myeloma using chimeric anti-CD38 antibody.

Multiple myeloma is a disease in which conventional chemotherapy has only limited value, but which may be ideal for treatment with passive antibody against a suitable cell surface antigen on the neoplastic plasma cell. The CD38 antigen is known to be present on the majority of neoplastic plasma cells, and this was confirmed by detailed examination of bone marrow aspirates from three patients. Strong expression of CD38 was confined to cells which, by the criteria of light-scattering profiles and possession of cytoplasmic Ig, were plasma cells. The vast majority of neoplastic plasma cells appeared to be involved. Using a cell line as a model, it was found that the CD38 antigen acts as a target for a chimeric antibody prepared from the antibody OKT10. The chimeric antibody consists of the Fab portion of the mouse monoclonal antibody linked by a stable thioether bond to an Fc molecule derived from human IgG1, thereby forming mouse Fab-human Fc. In contrast to the parent antibody, the chimeric molecule mediates antibody-dependent cellular cytotoxicity (ADCC) very efficiently with human blood mononuclear effector cells, and is effective at low concentration. Also, even though the CD38 antigen is present on natural killer cells, there appears to be little deleterious action of the antibody on effector cell function. The antibody also failed to affect the growth of progenitor cells of the granulocyte/macrophage or erythroid lineages present in normal bone marrows, despite the suspicion that these cells express the antigen. Other advantages of the CD38 molecule are that it is not found in the serum of patients with myeloma, and it does not appear to modulate in vitro. Fourteen patients with florid myeloma and on various chemotherapeutic regimes had an undiminished capacity to mediate ADCC with the chimeric antibody, when compared with normal individuals. The maintenance of ADCC activity, coupled with the known suppression of the antibody response in these patients, augers well for treatment with chimeric antibody.

A chimeric antibody with dual Fc regions (bisFabFc) prepared by manipulations at the IgG hinge.

A new chimeric antibody for therapeutic use in human cancer is described. First the derivative FabFc was prepared by linking Fab' gamma from monoclonal antibody to Fc gamma from human normal IgG1. The bismaleimide linking agent forms a thioether bond with an SH group released by reduction of SS bonds in the hinge of each constituent. It follows that one of the original two SS bonds in the Fc hinge still has both its S atoms free, and this bond is reformed by thiol-disulphide interchange. The lone free SH in the Fc hinge can now be used to join two FabFc molecules through a similar bismaleimide linker to yield bisFabFc. As regards antibody activity against target cells, bisFabFc can be univalent, bivalent, or bispecific. Its juxtaposed dual Fc regions are designed to promote cooperative binding of effectors, and bisFabFc is indeed notably more powerful than its parent FabFc molecules in promoting complement lysis and antibody-dependent cellular cytotoxicity. However it is not possible at present to distinguish the separate contributions of Fc architecture, antibody affinity and other factors towards this improvement. In the present state of development a variety of FabFc against a given neoplasm may be prepared in high yield from mouse IgG1 and IgG2a antibodies, and when convenient dimerized to bisFabFc in any combination of specificities.