Claims
- 1. A process for producing a pH-sensitive immunoconjugate between an antibody reactive with a tumor-associated antigen and a chemotherapeutic agent comprising:
- a) attaching aconitic anhydride to an available amino group of a chemotherapeutic agent to form a first intermediate with a free carboxyl group;
- b) reacting said free carboxyl group of said first intermediate with the amino group of a lysine containing amino acid spacer molecule using a first activating reagent to form a chemotherapeutic agent-spacer-reagent compound;
- c) reacting said chemotherapeutic agent-spacer reagent compound with a second activating reagent to form modified chemotherapeutic agent-spacer-reagent compound;
- d) reacting the lysine group of an non-intervalizing antibody using a thiolating reagent to form modified antibody; and
- e) reacting said modified chemotherapeutic agent-spacer reagent compound with said modified antibody to form an immunoconjugate comprising a chemotherapeutic agent coupled by a spacer molecule to an antibody,
- whereby said immunoconjugate dissociates in low pH tumor tissue, releasing said chemotherapeutic agent in said tumor tissue.
- 2. The process according to claim 1 wherein said spacer molecule is poly-L-Lysine.
- 3. The process according to claim 2 wherein said first activating agent used to form the chemotherapeutic agent-spacer-reagent compound is
- 1-ethyl-3-(dimethylaminopropyl)carbodiimide hydrochloride.
- 4. The process according to claim 2 wherein said second activating reagent is maleiimide reagent.
- 5. The process according to claim 4 wherein said maleiimide reagent is
- sulfosuccinimidyl-4-(N-maleimidomethyl)cyclohexane-1-carboxylate.
- 6. The process according to claim 2 wherein said thiolating reagent is S-acetylmercaptosuccinicanhydride.
- 7. The process according to claim 1, wherein said chemotherapeutic agent is selected from the group consisting of Daunomycin, Mitomycin C, Adriamycin and Methotrexate.
- 8. The process according to claim 7 wherein said chemotherapeutic agent is Daunomycin.
- 9. The process according to claim 7, wherein said antibody is L6 monoclonal antibody.
- 10. The process according to claim 7, wherein said antibody is an antibody which is not internalized by tumor cells.
CROSS REFERENCE TO RELATED APPLICATION
This is a divisional of U.S. patent application Ser. No. 047,161, filed May 12, 1987, now U.S. Pat. No. 4,997,913 issued Mar. 5, 1991, which is a continuation-in-part of U.S. patent application Ser. No. 880,674, filed June 30, 1986 now abandoned.
US Referenced Citations (1)
Number |
Name |
Date |
Kind |
4631190 |
Shen et al. |
Dec 1986 |
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Foreign Referenced Citations (1)
Number |
Date |
Country |
1079268 |
Jun 1980 |
CAX |
Non-Patent Literature Citations (3)
Entry |
Blair et al (1983) J. Immunol. Methods 59: 129-143. |
Gallego et al. (1984) Int. J. Cancer 33:737-744. |
Jung et al. (1981) Biochem. Biophys. Res. Commun. 101(2):599-606. |
Divisions (1)
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Number |
Date |
Country |
Parent |
47161 |
May 1987 |
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Continuation in Parts (1)
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Number |
Date |
Country |
Parent |
880674 |
Jun 1986 |
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