HPMA copolymer-anticancer drug-OV-TL16 antibody conjugates. 1. influence of the method of synthesis on the binding affinity to OVCAR-3 ovarian carcinoma cells in vitro.
The influence of different methods of binding the OV-TL16 antibody and its Fab' fragment to N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer--drug (adriamycin [ADR] or meso chlorin e6 mono(N-2-aminoethylamide) (Mce6)) conjugates on the affinity of conjugates to an ovarian carcinoma (OVCAR-3) cell associated antigen was investigated. The binding of the antibody to HPMA copolymer--drug (ADR or Mce6) conjugates via amino groups resulted in conjugates which were heterogeneous in their antigen binding. Coupling, the HPMA copolymer--Mce6 conjugate to the carbohydrate region of the antibody resulted in conjugates with a more homogeneous distribution of affinity constants than conjugates prepared by linking the antibody to the polymer via amino groups. However, both methods resulted in a decrease in the affinity constant compared to the native antibody. Conjugates prepared with the Fab' frgment of the OV-TL16 antibody demonstrated a more homogenous affinity than either conjugate prepared with the whole antibody. To verify the hypothesis that the changes in the binding affinity and homogeneity are a consequence of conformational changes in the antibody structure, a series of physiocochemical methods were employed to characterize the conjugates. The excitation energy transfer between OV-TL16 antibody and drugs (ADR and Mce6) and the spectral properties of Mce6 were used to monitor the interactions between the antibody and drugs. The quenching of the intrinsic fluorescence of the antibody was also employed to study its conformational changes. An attempt has been made to correlate the biorecognition at the cellular surface with the interactions of drug with the antibody molecule and with the changes in antibody conformation.
Omelyanenko
V
V
Department of Pharmaceutics, University of Utah, Salt Lake City, Utah 84112, USA.
Kopecková
P
P
Gentry
C
C
Shiah
J G
JG
Kopecek
J
J
D000918:Antibody Specificity; D000936:Antigen-Antibody Complex; D001665:Binding Sites; D002277:Carcinoma / Q000188:drug therapy / Q000378:metabolism* / Q000473:pathology; D004337:Drug Carriers; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006801:Humans; D006825:Hybridomas; D007140:Immunoglobulin Fab Fragments / Q000378:metabolism / Q000627:therapeutic use*; D007457:Iodine Radioisotopes; D007553:Isotope Labeling; D008689:Methacrylates / Q000138:chemical synthesis / Q000378:metabolism / Q000627:therapeutic use*; D010051:Ovarian Neoplasms / Q000188:drug therapy / Q000378:metabolism* / Q000473:pathology; D011108:Polymers; D013050:Spectrometry, Fluorescence; D014407:Tumor Cells, Cultured
D000936:Antigen-Antibody Complex; D004337:Drug Carriers; D007140:Immunoglobulin Fab Fragments; D007457:Iodine Radioisotopes; D008689:Methacrylates; D011108:Polymers; C032802:hydroxypropyl methacrylate