This invention relates to identification of a new common antigen of colorectal and mucinous ovarian tumors (COTA) and to a process for isolating the same. This invention also relates to clinical application of COTA and anti-COTA antibodies in detection and diagnosis of colorectal and ovarian tumors.
Presently, known and well studied antigens associated with colon cancer include CSAp, CEA and Ca 19-9. The term CEA is conventionally recognized as representing "Carcimoembryonic antigen" which was originally described as being present in intodermal derived tumors of the gastrointestinal tract [Gold et al., J. Exp. Med. 121:439-462 (1965)]. The term CSAp is conventionally recognized as representing "colon specific antigen (protein)" which was originally described as a colon cancer associated antigen and subsequently detected in some ovarian cancers [Pant et al., Immun. Comm. 6:411-421 (1977)]. The term Ca 19-9 is conventionally recognized as representing "Carbohydrate antigen 19-9" and was originally found as a colorectal carcinoma-specific antigen [Koprowski et al., Somatic Cell Genetics 5:957-972 (1979)]. It is not practical to diagnose colorectal cancers by assaying for CSAp, since CSAp is only elevated in the plasma of about 61% of advanced colorectal cancer patients. Also, the assay for CEA in diagnosis of colorectal cancers is not definitive since CEA is an antigen marker for a wide variety of cancers, only one of which is colon cancer. Similarly, the diagnosis of colorectal cancer based on Ca 19-9 assay is not definitive as Ca 19-9 is elevated in greater percentage in the plasma of pancreatic cancer patients and is elevated in only about 58% of advanced untreated colorectal cancer patients.
Reference to mucins in the neoplasms of colon have suggested chemical and immunological differences between normal and neoplastic colonic mucoproteins (Gold et al, Nature 255, 85-87) (1975). Unfortunately, these studies which describe a normal and tumor derived mucoprotein antigen designated CMA were conducted with unabsorbed xenogenic antiserum and require substantial validation with application of more specific antibodies before a definitive statement can be made on tumor specificity of tumor derived CMA. It has been observed that an antiserum to mucin isolated from benign mucinous cytadenomas from human ovaries has antigenic reactivity towards mucosal extracts of colorectal cancers, although was absent in extracts from normal tissue and tumors of the ovaries, stomach, lung and breast. (McNeil et al, Cancer Res. 29, 1535-1540). McNeil et al postulated that ovarian mucin contained antigenic material common to "altered" mucin found in colon cancer tissue. In another report (Narin et al, Brit. J. Cancer 25, 276-279) an antigen common to normal colon. and ovarian cystadenoma was described leading authors to conclude that ovarian mucinous cystadenomas were derived from multipotent intestinal cells and could arise by unilateral intestinal development of a teratoma.
A composition containing a common antigen for malignant human colorectal and mucinous ovarian tumors has been disclosed by Pant et al at the Oncodevelopmental Biology and Medicine XI annual meeting, Stockholm, Sweden (1983). This antigen was obtained by first forming an antibody against human colon cancer tissue extract by immunizing a goat with the extract. The goat sera containing the antibodies was absorbed with extracts of human tissues, plasma and with CEA. The resultant antiserum contained the antibody to colorectal and ovarian tumor antigen (COTA). However, the COTA antibody containing serum was not purified further.
Although the anti-COTA antibodies were specific for COTA, the antiserum also contained an excess of dissolved normal tissue antigens, normal plasma, and CEA.
In order to prepare specific anti-COTA antibodies without having dissolved normal antigens, plasma and CEA in it, goat serum containing anti-COTA antibodies was first precipitated by conventional 50% and then by 33% ammonium sulfate method. The immunoglobulins (after dissolving and dialysis in phosphate buffered saline) were concentrated to half the original volume. The preparation was then passed through an affinity column (cyanogen bromide activated Sepharose 4B) made of a mixture of normal human colon, lung, liver, spleen, kidney and plasma. The unadsorbed material was concentrated to half the original volume and then passed through an affinity support made of CEA. The unadsorbed material was concentrated to give a concentration of 2 mg/ml solution and contained specific anti-COTA reactivity.
In order to recover COTA, colon cancer homogenate and/or mucinous ovarian cyst fluid was fractionated on Sephadex G-200. COTA appeared in the void volume from Sephadex G-200 column. This material containing COTA was further fractionated on Sepharose 4B column wherefrom COTA was eluted at 3-6 million molecular weight fraction. This COTA preparation was not adequately pure so that it could be used to raise specific anti-COTA antibodies in the animals since the preparation still contained other tissue antigens present in the original colon cancer homogenate or in the mucinous ovarian cyst fluid.
Accordingly, it would be highly desirable to provide a means for obtaining a COTA which is pure so that it could be utilized to produce specific antibodies to COTA. These antibodies, in turn, could be utilized to recover more highly purified COTA and could be used in development of diagnostic tests for detection and diagnosis of colorectal and ovarian cancers. Presently available COTA is not sufficiently pure for raising specific anti-COTA antibodies nor the antibodies are without dissolved normal tissue antigens, plasma and CEA so as to render them useful to correspondingly recover COTA or to utilize them in development of reliable diagnostic tests.