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Studies of a tumor-associated antigen, COX-1, recognized by a monoclonal antibody
1Department of Obstetrics and Gynecology, University of British Columbia, Vancouver, Canada.
Abstract:
Monoclonal antibodies against an ovarian tumor cell line, OC-3-VGH, were generated using modified hybridoma technology. Among the seven that were selected for their high specificity and affinity to ovarian cancer cells and low cross-reactivity to most normal human tissues, RP 215 was shown to react specifically with a tumor-associated antigen, COX-1, from certain ovarian/cervical cancer cell lines. By Western blot assay, COX-1 was shown to have a subunit molecular mass of about 60 kDa and exist as an aggregate in the native state. COX-1 could also be detected in the shed medium of certain cultured tumor cells. A solid-phase sandwich enzyme-immunoassay procedure was designed for quantitative determinations of COX-1 in the shed medium or in patients' sera using RP 215 for both well-coating and the signal detection. Highly purified COX-1 was obtained from the shed medium of cultured OC-3-VGH tumor cells mainly by hydroxyapatite and immunoaffinity chromatography with RP 215 as the affinity ligand. At neutral pH, purified COX-1 also exists as an aggregate and is relatively stable at temperatures below 50 degrees C. Its immunoactivity was found to decrease with time in the presence of trypsin. However, the immunoactivity of COX-1 was not affected upon incubation with carbohydrate-digestive enzymes or concanavalin A and only partially inactivated in the presence of NaIO4 or iodoacetamide. Treatments of COX-1 with dithiothreitol and guanidine thiocyanate resulted in a complete loss of activity. Furthermore, rabbit antisera raised against purified COX-1 exhibited similar immunospecificity to that of RP 215. The results of this study suggest that COX-1 is a glycoprotein consisting of a 60 kDa subunit, which is recognized by RP 215 through its peptide determinant. Preliminary retrospective clinical studies were performed to assess the utility of a COX-1 enzyme immunoassay kit for detection and monitoring of patients with ovarian and cervical cancers.
Insights
Researchers developed a new method to detect COX-1, a tumor antigen, using monoclonal antibody RP 215. This advance aids in the detection and monitoring of ovarian and cervical cancers.
Area of Science:
- Immunology
- Biochemistry
- Oncology
Background:
- Monoclonal antibodies are crucial tools in cancer research.
- Ovarian and cervical cancers require sensitive diagnostic markers.
Purpose of the Study:
- To generate and characterize monoclonal antibodies against ovarian cancer.
- To identify and investigate a specific tumor-associated antigen, COX-1.
- To develop a diagnostic assay for ovarian and cervical cancers.
Main Methods:
- Modified hybridoma technology for antibody generation.
- Western blot and immunoassays for antigen characterization.
- Hydroxyapatite and immunoaffinity chromatography for purification.
- Enzyme-immunoassay development for quantitative detection.
Main Results:
- Monoclonal antibody RP 215 specifically targets COX-1, a 60 kDa glycoprotein antigen.
- COX-1 is found in ovarian/cervical cancer cells and shed medium.
- A sensitive enzyme immunoassay for COX-1 was developed.
- Preliminary clinical studies show potential for cancer detection and monitoring.
Conclusions:
- COX-1 is a viable tumor marker for ovarian and cervical cancers.
- The developed immunoassay shows promise for clinical application.
- Further studies are warranted to validate the diagnostic utility of COX-1.