Tumor-associated antigens in female genital tract cancers

The immunologic reactivity of glycoprotein antigens extractable from individual, histologically different ovarian and uterine cancers was studied taking into account their relationship with carcinoembryonic antigen (CEA), nonspecific cross-reacting antigen (NCA), alpha-feto-protein (AFP), and alpha-1-antichymotrypsin. All studies were performed using specific immune sera against perchloric acid (PCA) extracts of ovarian mucinous cystadenocarcinoma (anti-PCA-CaOm) and cervical squamous cell carcinoma (anti-PCA-CaCx), and antisera against the reference antigens mentioned above. A considerable antigenic heterogeneity and the existence of several immunologically related antigenic systems were found: 1) CEA-like antigens; 2) NCA-type antigens; 3) an antigen different from CEA and NCA present in ovarian mucinous adenocarcinomas and often cross-reacting, but not identical with respective antigens of uterine body and cervical carcinomas; 4) an antigen reacting with anti-alpha-1-anti-chymotrypsin serum; and 5) an antigen reacting with anti-AFP serum.     

Antigenic heterogeneity of human lung cancers

Antigenic reactivity of 35 perchloric acid (PCA) extracts of different histologic human lung cancer tissues was studied--in comparison with the reactivity of the carcinoembryonic antigen (CEA), the nonspecific cross-reacting antigen (NCA), and alpha-1-antichymotrypsin--with the use of specific immune sera against PCA extracts of lung squamous cell carcinoma, and anti-CEA, anti-NCA, and anti-alpha-1-antichymotrypsin sera. The following antigenic systems were found in lung cancers: a) antigens specific for most squamous cell cancers and adenocarcinomas, which are undetectable in small cell cancers; b) NCA-type antigens; c) CEA-like antigens; and d) the antigen responsible for alpha-1-antichymotrypsin reactivity. A considerable antigenic heterogeneity among lung cancers indicates the necessity for precise histopathologic verification of individual lung cancer cases before commencement of immunologic studies and purification of antigens specific for lung cancers.     

Density distribution and antigenic characterization of cell subpopulations isolated from cystic fluids of ovarian serous neoplasms

The immunological reactivity of serous ovarian tumor cells was evaluated, taking into account their density and morphological features. Discontinuous density gradient centrifugation was applied to fractionate cell subpopulations from cystic fluids of patients with ovarian serous carcinomas, cystadenomas and benign serous cysts. For phenotypic characterization of tumor cell subpopulations the immune sera against perchloric acid extracts of ovarian serous (anti-PCA-CaOs) and mucinous (anti-PCA-CaOm) carcinomas were used. The expression of carcinoembryonic antigen (CEA) and nonspecific cross-reacting antigen (NCA) on tumor cell fractions was also checked. Some relationship between immunological reactivity, cell morphology and cell density was found; however, individual patient-to-patient variations in cellular composition and antigenic expression were also observed. The presence of ovarian serous carcinoma-associated antigen (CaOs-Ag) was related mainly to frankly malignant cells. Neither CEA nor NCA were constant and characteristic markers for serous ovarian neoplasms. These neoplasms were unreactive with anti-PCA-CaOm serum. Our results indicate the possibility of correlation between morphological features, density distribution and immunological phenotypes of ovarian tumor cell populations.     

Studies on circulating antibody against carcinoembryonic antigen (CEA) and CEA-like antigen in cancer patients

Characteristics of auto-antibodies for carcinoembryonic antigen (CEA) detected in sera from 3 cancer patients (2 colorectal and 1 breast cancer) were examined. The antibodies belonged to polyclonal immunoglobulin G (IgG). The binding of auto-antibodies with the labeled CEA was inhibited by not only the unlabeled CEA but also NCA-2 (feces and meconium). However, no binding with NCA was observed. Among these auto-antibodies the antibody directed against blood group Lewis determinants which are known to be present in many purified CEA preparations was not found. Previously we had suggested that CEA, NCA-2 and NCA may contain immune determinant in common with alpha 1-acid glycoprotein (AG). These auto-antibodies showed significantly enhanced reactivity for the labeled CEA preparation after purification by anti-AG affinity chromatography in spite of no immunological reaction with AG. These results suggest that auto-antibodies are raised against the common antigenic determinants of both CEA and NCA-2 which do not exist in NCA. These antibodies might be directed to common amino acid sequence shared by CEA and NCA-2, though not excluding the carbohydrate moiety. We surveyed about 500,000 cancer patients but could find only 3 patients who showed a difference in the values of CEA by the indirect and direct method. Thus, the existence of this type auto-antibody to CEA in cancer patients is a rare phenomenon.     

Monkey antisera with increased specificity to carcino-embryonic antigen (CEA).

Three Macaca irus monkeys were immunized with purified human CEA. One received unmodified CEA and two were immunized with haptenated (4-hydroxy-5-iodo-3-nitrophenyl acetyl) CEA. All three monkeys formed precipitating antibodies to CEA. The antisera did not precipitate the CEA-related normal glycoprotein antigen (NCA). In contrast, CEA-immunized rabbits, sheep and goats invariably form antibodies against NCA. Radio-immunoassays showed the monkey antisera to contain trace amounts of antibodies to NCA but the titers were about 10-1,000 times lower than those in rabbit and sheep sera with comparable anti-CEA titers. These results show that monkeys produce antibodies against CEA, but respond weakly to a normal, CEA-related antigen. The weak response to NCA suggests that monkeys may possess an NCA-related antigen. Antisera lacking reactivity to normal CEA-related antigens may be helpful in establishing a more specific diagnostic test for CEA.     

Production kinetics and immunochemical properties of carcinoembryonic antigen and nonspecific cross-reacting antigen synthesized by various human tumor cell lines

The production kinetics and immunochemical properties of carcinoembryonic antigen (CEA) and nonspecific cross-reacting antigen (NCA) in various human tumor cell lines were studied. By radioimmunoassay (RIA), five CEA-producing tumor cell lines tested--2 derived from colonic (M7609 and CCK-81), one from pancreatic (QGP-1) and 2 from lung (HLC-1 and KNS-62) carcinomas--were found to produce NCA simultaneously. The cellular contents of CEA and NCA and the amounts of both antigens released into the culture medium were highly variable among the cell lines. It was a distinct contrast that one cell line (CCK-81) released very large amounts of CEA and NCA into the medium while having the smallest amounts of both antigens in the cells, whereas the others contained much larger amounts of the antigens in the cells as compared with the amounts released into the medium. For most of the cell lines, the production of both CEA and NCA increased in the stationary phase of growth as compared with the exponential phase. The production kinetics of both CEA and NCA appeared to be parallel with each other in all the cell lines, though the amount ratio of CEA to NCA produced was variable. By means of a double immunodiffusion test with polyclonal antibodies, antigenic uniformity with no unique organ-specificity was confirmed for all the CEA preparations from spent media of the cell lines, though some differences in the sugar moiety of CEA were detected by RIA using monoclonal antibodies. No antigenic differences among NCA preparations were observed. Upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), molecular heterogeneity was observed among CEA or NCA preparations isolated from cell lysates.     

Purification and characterization of carcinoembryonic antigen-related antigens in normal adult feces

We tried to purify carcinoembryonic antigen (CEA)-related antigens in normal human feces and found that, besides nonspecific cross-reacting antigen (fecal NCA), there are three other molecular species of CEA-related antigens which have been inclusively called normal fecal antigen (NFA). These were designated normal fecal antigen 1 (NFA-1), normal fecal antigen 2 (NFA-2), and normal fecal cross-reacting antigen (NFCA), respectively. Among these antigens, NFA-1, NFA-2, and fecal NCA were isolated in pure form. NFCA has not yet been obtained in pure form but was identified as an antigen different from three other antigens. All antigens were glycoprotein in nature and migrated electrophoretically in the beta region. Their molecular weights were estimated to be 20,000 to 30,000 for NFA-1, 160,000 to 170,000 for NFA-2, and 80,000 to 90,000 for NCA, respectively. NFA-2 had amino acid and carbohydrate compositions similar to those of CEA. The results obtained by immunodiffusion analyses of antigenic determinants indicate that CEA and NFA-2 can be divided into four antigenic moieties: the first one is distinctive for CEA (cancer determinant) or NFA-2 (NFA-2-distinctive determinant); the second one corresponds to NFA-1 (NFA-1 determinant); and the remaining two moieties are present on NFCA, one of which is characteristic for NFCA (NFCA determinant) and the other of which is shared with NCA (NCA-common determinant).     

Immunophenotypic analysis of colorectal carcinomas with monoclonal antibodies 47D10 and anti-carcinoembryonic antigen

A series of 80 colorectal adenocarcinomas were analyzed immunohistochemically for the antigen recognized by a new monoclonal antibody (MCA) 47D10. These antigens are part of a complex family of substances similar to, yet distinct from carcinoembryonic antigen (CEA), and are termed nonspecific cross-reacting antigens (NCA). Formalin-fixed paraffin-embedded sections of colorectal adenocarcinomas were evaluated for the expression of these antigens and compared to the expression of CEA. Our study shows that 83.8% of the cases were positively stained for NCA while 91.3% were positive for CEA. Both antigens were coexpressed in 80% of the cases. No correlation was found between MCA 47D10 immunoreactivity and tumor grade, stage, size or location within the colon. In 25 cases, the benign colonic mucosa adjacent to the carcinoma stained positively with MCA 47D10. Normal colon does not express NCA as recognized by MCA 47D10, except in rare cells. Forty-eight of these cases had serum available for study. Both NCA and CEA were determined in these serum samples. Forty-two of these sera demonstrated elevated CEA levels, whereas only 8 showed increased levels of the 47D10 antigen(s). These findings suggest that the gene product(s) recognized by MCA 47D10 can be independently expressed or, more commonly, coexpressed with CEA in these tissues.     

Immunocytochemical evaluation of human esophageal neoplasms and preneoplastic lesions for beta-chorionic gonadotropin, placental lactogen, alpha-fetoprotein, carcinoembryonic antigen, and nonspecific cross-reacting antigen

Human esophageal neoplasms were studied in comparison to normal, uninvolved, and preneoplastic human esophageal epithelium for the presence of human chorionic gonadotropin (HCG), human placental lactogen (HPL), alpha-fetoprotein (AFP), carcinoembryonic antigen (CEA), and nonspecific cross-reacting antigen (NCA) using the unlabeled antibody peroxidase-antiperoxidase technique. HCG immunoreactivity was identified in 10 of 33 squamous cell carcinomas (33%), in 1 of 6 adenocarcinomas (17%), and 1 of 6 preneoplastic esophageal lesions (17%); while 9 of 33 squamous cell carcinomas (33%) and 1 of 6 adenocarcinomas (17%) contained immunoreactive AFP. Immunoreactive HPL was detected in 6 of 33 squamous cell carcinomas (20%), but in none of the adenocarcinomas. Neither AFP nor HPL immunoreactivity was identified in the 6 hyperplastic lesions which were studied. When stained with an antiserum that was able to detect both CEA and NCA, 27 of 33 squamous cell tumors (82%) and 6 of 6 adenocarcinomas (100%) showed positive immunostaining reactions. Of these, 8 squamous cell carcinomas and 1 adenocarcinoma were subsequently shown to contain only NCA immunoreactivity, while 19 squamous cell carcinomas and 5 adenocarcinomas contained both NCA and CEA immunoreactivity. NCA immunoreactivity alone was identified in 3 of 6 preneoplastic lesions and NCA and CEA immunoreactivity in 1 of 6 preneoplastic lesions. None of the markers was detected in 8 specimens of normal esophageal epithelium which were studied as controls, nor in 6 specimens of uninvolved esophageal epithelium obtained from patients with esophageal cancer. Most tumors expressed 2 or 3 markers, and some tumors were identified which expressed up to 4 of the 5 markers investigated. Only 3 tumors failed to express any of the markers studied. No association was found between the degree of tumor differentiation and presence or absence of HCG immunoreactivity. However, HPL immunoreactivity was more common in poorly differentiated squamous cell carcinomas. In contrast, immunoreactive AFP was more common in well-differentiated squamous cell carcinomas than in other tumor types. Similarly, both CEA and NCA were more frequently expressed in well-differentiated squamous cell carcinomas, adenosquamous carcinomas, and adenocarcinomas than in less differentiated tumors. Our results suggest that HCG, HPL, AFP, CEA, and NCA are tumor-associated antigens in esophageal cancer. Therefore, they could be of value in screening tests for esophageal neoplasms and could be useful in subclassification of esophageal neoplasms.     

Three novel mouse monoclonal antibodies, OM-A, OM-B, and OM-C, reactive with mucinous type ovarian tumors

Three mouse IgG1 monoclonal antibodies (MoAbs) reactive predominantly with cytoplasmic antigens of mucinous type ovarian tumors were produced. OM-A MoAb was reactive with 11 of 14 mucinous type and two of three endometrioid type ovarian tumors, although only a minor population of the tumor cells was positive in the latter type. This MoAb was not reactive with serous type, clear cell type, or other types of ovarian tumors, nor with various types of uterine carcinoma. Normal adult and fetal tissues of female genital organs were not positive with this MoAb. Among nongynecological carcinomas, three of six metastatic tumors to the ovary from the gastrointestinal tract, one of five gastric carcinomas, and one of eight lung adenocarcinomas were positive. As for normal adult and fetal tissues of nongynecological origin, epithelium of the normal stomach, small bowel, and bronchus as well as epithelium of fetal small and large bowel and secretory products were weakly positive. Thus, this MoAb showed a selected specificity against mucinous and endometrioid types of ovarian tumors. OM-B MoAb showed a broader specificity than OM-A, reacting with all mucinous type, two of three endometrioid type, and three of 16 serous type ovarian tumors, but not with clear cell type tumors. Adenoma type, but not squamous type, cervical carcinomas and one-half of endometrial carcinomas were positive. This antigen is present in cervical mucosa, but not in ovary or endometrium. OM-C MoAb showed a specificity similar to, but broader than that of, OM-B; i.e., 11 of 14 mucinous type, two of three endometrioid type, nine of 16 serous type, and one of nine clear cell type ovarian tumors were positive. It is reactive with adenoma type uterine carcinoma and normal mucosa of the uterine cervix and with normal surface epithelium of the oviduct. Among nongynecological tumors, OM-B antigen was present in metastatic tumors to the ovary as well as in gastric and pancreatic carcinomas, while OM-C was in metastatic tumors to the ovary and gastric and colonic carcinomas. Thus, the serological analysis showed that these three MoAbs showed selective specificities to mucinous and endometrioid types of ovarian tumors. Preliminary characterization of these three OM antigens suggested that these are distinct from carcinoembryonic antigen or ABH blood group-related antigens.     

Characterization of carcinoembryonic antigen-specific monoclonal antibodies and specific carcinoembryonic antigen assay in sera of patients

Six murine monoclonal antibodies (mAbs) reactive with carcinoembryonic antigen (CEA) were prepared. Four of these, CA204, CA205, CA206 and CA208, were specific to purified CEA whereas the other two, NA201 and NA203, were also reactive with the non-specific cross-reacting antigen (NCA). These mAbs were all IgG1, except one IgG2a (CA206), with high affinities for CEA. NA203, CA204 and CA208 appear to define three different epitopes on the CEA molecule as determined by competitive binding assay. These mAbs also reacted with CEA-producing cells. The treatment of the cells with tunicamycin did not affect the binding of the mAbs to CEA-producing cells. None of the above mAbs bound to CEA-related antigens, NCA-2, alpha 1-acid glycoprotein, or blood group antigens. The combination of CA208 (solid phase) and CA204 (tracer) was used to construct a sensitive CEA-specific sandwich ELISA to detect CEA in the sera of patients with various malignant and non-malignant diseases. Particularly when CEA values were low in sera from non-cancerous patients, the above mAbs sandwich assay showed reduced background reactivity with NCA-like substances and permitted the detection of CEA at a level as low as 1 ng/ml.     

Immunohistochemical localization of carcinoembryonic antigen (CEA), CEA-S, and nonspecific cross-reacting antigen (NCA) in carcinomas of lung

Antisera to carcinoembryonic antigen (CEA), to a physicochemical subset of CEA, namely CEA-S, and to nonspecific cross-reacting antigen (NCA) were used for the immunohistochemical localization of these antigens in human bronchogenic carcinomas using a triple layer immunoperoxidase technique. The study is based on an analysis of tumors from 130 patients. CEA, CEA-S, and NCA were all identified in the membrane and/or cytoplasm of neoplastic cells, and a good correlation between the antigens was observed in a majority of tumors. The presence or absence of these tumor-associated glycoproteins appeared to be correlated with the histologic type of the tumors, especially in small cell anaplastic carcinoma and adenocarcinoma, and the degree of histologic differentiation of adenocarcinomas correlated positively with these tumor-associated antigens. Data from this group of patients suggest that analysis of tissue CEA at the time of biopsy or surgical resection may facilitate a more objective interpretation of serial plasma CEA assays.     

Nuclear reactivity for estrogen. A possible diagnostic tool in differentiating colon cancer metastases in the ovary from primary mucinous ovarian tumors

Tissue distribution of carcinoembryonic antigen (CEA) and estrogen was analyzed in eight primary ovarian mucinous carcinomas, three primary mucinous colonic cancers, and four ovarian metastatic lesions from colonic and pancreatic adenocarcinomas. Primary ovarian mucinous carcinomas and metastatic colonic carcinomas had similar localization of CEA within the epithelial cytoplasm. Estrogen localization, in contrast, was shown within the epithelial nuclei of seven of eight primary ovarian mucinous tumors; whereas the nuclei of metastatic and primary mucinous colonic carcinomas did not have intranuclear estrogen. The cytoplasm and the stroma of both primary ovarian and metastatic colonic carcinomas stained for estrogen.     

Selection of carbohydrate antigens in human epithelial ovarian cancers as targets for immunotherapy: serous and mucinous tumors exhibit distinctive patterns of expression

Expression of blood group-related carbohydrate antigens was examined in frozen sections from a series of ovarian carcinomas of different histological types using an indirect immunoperoxidase technique. Antigenic specificities belonging to the O(H) and Lewis blood group families (H-1, H-2, Le(a), sLe(a), Le(x), sLe(x), Le(b) and Le(y)) or the mucin-core family (Tn, sTn and TF) were studied. A distinct difference in antigen expression between mucinous and other ovarian carcinomas (serous and endometrioid) was observed. Specifically, mucinous tumors tended to express sTn, Le(a) and sLe(a) strongly and homogeneously, whereas serous and endometrioid tumors rarely expressed these specificities and, in contrast, expressed Le(y) and H type 2 antigen strongly. When expressed in serous tumors, sTn was usually distributed in a heterogeneous pattern, whereas sTn expression in mucinous tumors was much more homogeneous. The distribution of Le(y) in serous tumors was noticeably homogeneous. H-1, Le(x), sLe(x), Le(b), TF and Tn specificities were rarely expressed in any type of ovarian carcinoma. Our results provide further support for the different histogenesis of mucinous and non-mucinous tumors and indicate alternative differentiation pathways for the 3 pathological subtypes of ovarian tumor. They also provide the basis for the choice of carbohydrate antigens for active and passive immunotherapy of ovarian carcinomas.     

Human ovarian cancer xenografts in nude mice: characterization and analysis of antigen expression

We have characterized 13 different human ovarian cancer xenografts grown subcutaneously in nude mice. The tumor lines represented 5 histological subtypes: serous (4), mucinous (4), clear-cell carcinoma (1), carcinosarcoma (1) and undifferentiated (3). The specific histology and the degree of differentiation resembled those of the original patients' tumors and were maintained upon serial transfer. Volume doubling times of the xenografts ranged from 3.5 to 15 days. The xenografts were also analyzed for their antigen expression using 20 monoclonal antibodies (MAbs) reactive with 15 tumor-associated antigens. Immunohistochemical examination of tissue sections showed a positive reaction pattern with MAbs 115D8, 140C1, 139H2, 175C5, HMFG1 and HMFG2, each recognizing episialin, as well as with MAbs AUA1, 358.4.32 and 199-157 in xenografts of the serous, mucinous and clear-cell carcinoma subtype. MAb OC125 was reactive with xenografts of the serous subtypes. Other antibodies, such as 494 and OV-TL3 infrequently demonstrated positive reactions. Reactivity of all MAbs was low in the carcinosarcoma and undifferentiated tumor lines. With the exception of AUA1, 495 and 126E5, all MAbs revealed a heterogeneous staining pattern. MAbs against episialin and OC125 predominantly stained the apical site of the tumor cells. Strongest reactivity with almost all histological subtypes was observed with MAbs 115D8, 140C1, 139H2 and AUA1. In cases where we were able to compare the patients' tumor tissue with the respective xenografts, retention of antigen expression was demonstrated in each instance. Release of tumor-associated antigens was shown for CA125 in 2 serous-tumor lines, for CA15.3 in 1 serous-tumor line, and for CEA in 3 lines of the mucinous subtype. This panel of human tumor xenografts could be a valuable tool to determine the potential usefulness of MAb-guided therapy in ovarian cancer.     

Immunohistochemical analysis of pulmonary and pleural neoplasms using a monoclonal antibody (47D10) which reacts with nonspecific cross-reacting antigen

A series of 251 human pulmonary carcinomas were analyzed immunohistochemically for the antigens recognized by a new monoclonal antibody (MAb) 47D10. These antigens are part of a complex family of substances similar to, yet distinct from carcinoembryonic antigen (CEA), and are termed 'nonspecific cross-reacting antigens' (NCAs). The NCA epitope recognized by the MAb 47D10 is expressed on the cell surface and has previously been shown to be distinct from epitopes detected by several anti-CEA MAbs, as well as by MAbs 19-9 and Du-PAN-2. The NCA epitope recognized by MAb 47D10 is well preserved in formalin-fixed and paraffin-embedded tissues. Using immunohistochemistry, this epitope has been shown to have a limited biodistribution in normal tissues, and to be expressed by adenocarcinomas arising in the pancreas, colon, breast, ovary, prostate and lung. The frequency and pattern of NCA expression in human pulmonary neoplasms was found to correlate with the known distribution of CEA: and was often present in the non-small-cell carcinomas. In addition, the expression of CEA relative to NCA was evaluated in a select group of non-small-cell carcinoma cases using several anti-CEA MAbs, to directly compare the expression of CEA to NCA. In general, the NCA reaction pattern is more intense and expressed on more cells within the tumors than that of CEA expression.     

The clinical evaluation of sialyl Lewis Xi antigen in patients with gynecologic tumors

In order to estimate the clinical significance of sialyl Lewis Xi (SLXi) antigen, the antigen was measured with an "FH-6" Otsuka Kit in sera from patients with various gynecologic tumors and healthy women. The antigen in ovarian cyst fluids was also determined. Furthermore, serum SLXi antigen levels were serially followed up in the patients with elevated serum SLXi levels to evaluate the correlation between serum SLXi levels and the response to treatment. Results obtained were as follows. 1) Among the patients with uterine myoma, uterine malignancies and benign ovarian tumors, the incidence of elevated serum SLXi antigen levels was very low. 2) Among the patients with ovarian malignancies, serum SLXi antigen levels was significantly increased in the following order: clinical stage I (38%), stage II (50%) and stage III (65%). 3) The high SLXi value was observed in the cyst fluid from all ovarian mucinous adenomas and all ovarian cancers. In addition SLXi antigen level was significantly higher in the cyst fluid of mucinous adenocarcinomas than that of serous cystadenocarcinomas. 4) Serum SLXi values were correlated with the effect of treatment. Interestingly, the elevation of serum SLXi levels preceded the clinical detection of recurrence by 3 month in a case. Thus SLXi antigen appears to be a useful marker for monitoring of ovarian malignancies, especially of mucinous adenocarcinomas.     

Carcinoembryonic antigen gene family and its clinical application

Carcinoembryonic antigen (CEA) gene was cloned in 1987. Thereafter, the structures of non-specific cross-reacting antigen (NCA) and biliary glycoprotein I (BGPI) have also been clarified. These three antigens contain immunoglobulin-like domains in their basic structures and it could be possible that CEA gene was originated from this basic structure by internal gene multiplication. Each CEA and NCA has hydrophobic domain in the C-terminus consisting of 26 amino acids which is eliminated when it binds with membrane and is reconstituted by combining with phosphatidylinositol glycan, whereas BGPI contains transmembrane and cytoplasmic domains. It is of interest that CEA and NCA have been found to function as adhesion molecules. The structure and possible function of another CEA gene family, PS beta G, were also introduced in this short review.     

Plasma protein profiles and prognosis in gastric cancer.

In 104 patients with gastric cancer the serum proteins carcinoembryonic antigen (CEA), C-reactive protein (CRP), alpha 1-acid glycoprotein (AGP) (orosomucoid) and alpha 1-antichymotrypsin (ACT) were measured pre-operatively. The estimated median survival of patients with both raised CEA and ACT was only 5 weeks in contrast to 64 weeks for those with both proteins normal. An intermediary group with one of these proteins raised and the other normal had an estimated median survival of 15 weeks. Similar results were obtained by considering a combination of CEA with either AGP or CRP. For these data the results were not explicable in terms of associations between survival time and patient''s age, stage, operative procedure, histological classification or site of primary tumour.