Efficient killing of CD22+ tumor cells by a humanized diabody-RNase fusion protein.

Abstract

We report on the generation of a dimeric immunoenzyme capable of simultaneously delivering two ribonuclease (RNase) effector domains on one molecule to CD22(+) tumor cells. As targeting moiety a diabody derived from the previously humanized scFv SGIII with grafted specificity of the murine anti-CD22 mAb RFB4 was constructed. Further engineering the interface of this construct (V(L)36(Leu-->Tyr)) resulted in a highly robust bivalent molecule that retained the same high affinity as the murine mAb RFB4 (K(D)=0.2 nM). A dimeric immunoenzyme comprising this diabody and Rana pipiens liver ribonuclease I (rapLRI) was generated, expressed as soluble protein in bacteria, and purified to homogeneity. The dimeric fusion protein killed several CD22(+) tumor cell lines with high efficacy (IC(50)=3-20 nM) and exhibited 9- to 48-fold stronger cytotoxicity than a monovalent rapLRI-scFv counterpart. Our results demonstrate that engineering of dimeric antibody-ribonuclease fusion proteins can markedly enhance their biological efficacy.

• Authors: Jürgen Krauss, Michaela A E Arndt, Bang K Vu, Dianne L Newton, Siegfried Seeber, Susanna M Rybak
• Journal: Biochemical and biophysical research communications (Biochem Biophys Res Commun) Vol. 331 Issue 2 Pg. 595-602 (Jun 03 2005) ISSN: 0006-291X [Print] United States
• PMID: 15850802 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't, Research Support, U.S. Gov't, P.H.S.)

Chemical References

• Antibodies, Monoclonal
• Antigens, CD
• Antigens, Differentiation, B-Lymphocyte
• CD22 protein, human
• Cd22 protein, mouse
• Cell Adhesion Molecules
• Immunoglobulin Variable Region
• Lectins
• Recombinant Fusion Proteins
• Sialic Acid Binding Ig-like Lectin 2
• Ribonucleases

Topics

• Animals
• Antibodies, Monoclonal (chemistry, genetics, immunology, isolation & purification)
• Antibody Specificity
• Antigens, CD (immunology, metabolism)
• Antigens, Differentiation, B-Lymphocyte (immunology, metabolism)
• Cell Adhesion Molecules (immunology, metabolism)
• Cell Death
• Cell Line, Tumor
• Cell Survival
• Cytotoxicity, Immunologic
• Dimerization
• Humans
• Immunoglobulin Variable Region (chemistry, genetics, immunology)
• Inhibitory Concentration 50
• Lectins (immunology, metabolism)
• Mice
• Neoplasms (immunology, pathology)
• Protein Structure, Quaternary
• Recombinant Fusion Proteins (chemistry, immunology, isolation & purification, metabolism)
• Ribonucleases (chemistry, genetics, isolation & purification, metabolism)
• Sensitivity and Specificity
• Sialic Acid Binding Ig-like Lectin 2