1. Field of the Invention
The present invention relates to a novel antitumor protein hybrid, for example, cytotoic against mouse tumors, and a process for the preparation thereof. More particularly, the present invention relates to a novel protein hybrid, which, having a moiety which is substantially the fragment Fab of an antitumor immunoglobulin and a moiety which is the subunit A of plant toxin ricin, is specifically useful as a remedy for malignant tumor e.g., in mice, and a process for the preparation of the same.
2. Description of the Prior Art
As for the remedies for malignant tumor or cancer (antitumor agents) many drugs have hitherto been known; however, these drugs have a disadvantage that they can not be administered enough to destroy tumor cells, because they have a toxic effect not only upon tumor cells but also upon normal cells to a considerable degree. Several attempts have been made to overcome this disadvantage by combining an antitumor agent or a protein toxin having cytotoxicity with a specific carrier in order to have them selectively absorbed by tumor cells. There exists an antitumor antibody (antitumor immunoglobulin), though very small in amount, in the blood of a cancer patient or on the surface of tumor cells. An antitumor antibody can also be obtained by immunizing an animal with the tumor cells and absorbing the obtained antiserum with the human normal cells. Antitumor antibodies, whether autochthonus, allogeneic, or xenogeneic, are not always capable of displaying a cytotoxic effect against tumor cells; however, they have a common nature of combining with tumor cells with an extremely high selectivity. Antitumor antibodies, therefore, have been used as a carrier to have an antitumor agent or a protein toxin absorbed by tumor cells selectively.
For instance, Japanese Patent Application Laid-open No. 144723/76 discloses, an an antitumor drug, a conjugate of antibody and antitumor drug in which such antitumor drug as daunomycin, etc. is bound covalently with Fab' dimer of antitumor immunoglobulin. This conjugate is superior in that it carries the antitumor drug selectively to the target tumor cells; however, since an antitumor drug itself such as daunomycin, etc. bound with the antibody (Fab' dimer) still exerts cytotoxic effects not only against tumor cells but also against normal cells, it is not satisfactory in view of destroying tumor cells only, and its cytotoxicity itself is not always sufficient either.
Studies have also been made to use diphtheria toxin, which is one of the protein toxins having much stronger toxicity, in the place of an antitumor drug.
For instance, F. L. Moolten et al. report that they prepared a conjugate by conjugating rabbit anti-SV40 antibody to a diphtheria toxin with glutaraldehyde as a coupling agent and were able to protect hamsters challenged with SV40-transformed 3T3 cells from developing tumors by administering the conjugate to hamsters (Journal of the National Cancer Institute, Vol. 55, pp. 473-477, 1975).
P. E. Thorpe et al. report that the conjugate prepared by coupling diphtheria toxin to antilymphocytic antibody by means of chlorambucil greatly reduced the protein synthesis of human lymphoblastoid cells, CLA4. (Nature, vol. 271, pp. 752-754, 1978). The results of these studies show that a conjugate of diphtheria toxin and antibody displays toxicity against the tumor cells selectively. However, these conjugates, when used as an antitumor drug, are believed to have some disadvantages as cited below. First, xenogenic antibody (immunoglobulin) has a strong antigenicity in the human body and induces the formation of anti-xenogeneic immunoglobulic antibody which deactivates the antitumor activity and further causes an anaphylaxis shock. The second of the disadvantages is that the nonspecific toxicity of diphtheria toxin is not nullified. More particularly, the object of these methods is to conjugate diphtheria toxin on the surface of tumor cells by the aid of antitumor antibody; however, since the conjugate contains the whole molecule of diphtheria toxin in its composition, it tends to bind with normal cell surface receptors for diphtheria toxin and display cytotoxicity against the normal cells. Thirdly comes the disadvantage which is found in the method of cross-linking the antibody with the diphtheria toxin. Many of the cross-linking agents such as glutaraldehyde, toluene diisocyanate, diethyl malonimidate, chlorambucil, etc. effect cross-linking not only between the antibody and the toxin but also between antibody and antibody, and toxin and toxin, and moreover, they effect the formation of intramolecule bonds in the antibody and in the toxin molecule, thus causing the formation of undesirable products and decrease or loss of the antitumor activity.