The Preparation And Anticancer Activity Of Immunoconjugates Of Lidamycin And Monoclonal Antibody Or Its Fab' Fragment Against Type Ⅳ Collagenase

Selectively improving the curative effects and reducing the side reactions of chemotherapeutants have been the key projects in the field of cancer therapy and medicinal biotechnology. Immunoconjugates which can recognize tumour-associated antigens seem to be one of the strategies to solve the problem. In our lab we had ever coupled the monoclonal antibody 3G11 against type â…£ collagenase with the high potent antitumor antibiotic lidamycin and achieved prominent curative effects in mouse H22 hepatoma and C26 colon cancer transplatation tumor models. But, such conjugates were routinely prepared using techniques which employed random derivation of primary amines like α-amine on N-termini of LDM or ε-amine on the side chains and the yield was poor. High yield, site-specific coupling of LDM to antibody or its fragment is needed. Finding of proper linkers is critical for exerting the most efficiency of lidamycin in treatment. The studies focus on these fields.The hybridoma 3G11 cells were cultured and amplicated and then transplanted to BALB/c mouse abdominal cavity. Ascites fluids were collected 10 days latter. After caprylic acid/ammonium sulfate precipitation, the purity of 3G11 was 80%⁹0%. Furthermore, 3G11 was purified on Sepharose protein G column. The subclass of the mAb 3G11 was divided by ELISA assay and the result indicated that 3G11 belonged to IgG1 subclass.The purified 3G11 was digested by Ficin and the F(ab')2 was aquired after purified on sephadex G-150. The F(ab')₂ reserved highy yield and affinity. The affinity constants detected by ELISA were 1.3×10^-8 M^-1 and 2.7×10^-8 M^-1 for 3G11 and F(ab')2.We used wavelenghth scan to detect the difference between LDM and 3G11. The chromophore of LDM had a characteristic absorption peak at 341nm. The relationship between concentration and A₃₄₁ was C=1.4583×A₃₄₁+0.0068. Because the 3G11...