The FDA's perspective on the evaluation of tumor marker tests.

Cancer marker tests are often proposed for three intended uses: screening, diagnosis, and monitoring. For each intended use, performance characteristics need to be well defined. The utility of a marker in a given setting depends heavily on two predominant performance characteristics--sensitivity and specificity. These parameters must be established with respect to the intended clinical use of the marker. The value of the marker in a particular situation also depends on the effectiveness of therapy for the malignancy. In reviewing a cancer marker test, the US Food and Drug Administration focuses on both the proposed intended use statement and the clinical utility of the marker. The sponsor is expected to provide specific claims data in support of the safety and effectiveness of the device through well-designed and -executed clinical studies. Several cancer markers are already available. In the future, new markers are anticipated that may greatly expand the range of usefulness in cancer diagnosis screening and monitoring.

Appraisal and coping in the process of patient change during short-term psychotherapy.

The process of patient change during short-term dynamic therapy was followed in eight patients by repeatedly measuring their appraisal and coping with significant people and issues in their lives. Patients' ratings of their appraisal and their coping with each of these variables were compared to a line of optimal functioning, which describes the ideal appraisal/coping relationship. Two groups of patients were identified who must carry out different corrections on these measures during therapy in order to approximate the optimal functioning line. The appraisal/coping relationship measure appears promising as a way to reflect patient change during therapy.

Circulating IgA immune complexes in head and neck cancer, nasopharyngeal carcinoma, lung cancer, and colon cancer.

A double antibody enzyme-linked immunosorbent assay (ELISA) was developed to quantitate circulating immune complexed IgA (IgA IC) in human serum. The serum panel for this study consisted of normal blood donors, benign surgery (BS), head and neck cancer (HN), nasopharyngeal carcinoma (NPC), lung cancer (LC), and colon cancer (CC) patients. Immune complexes (IC) were isolated from these sera by precipitation with 3.5% polyethylene glycol (PEG), washed and then redissolved in 0.1 M phosphate-buffered saline pH 7.2. The amount of IgA IC present were then quantified using the double antibody IgA ELISA. This assay was found to be both sensitive (26.0 ng/ml) and reproducible (intra-assay coefficient of variation 4.0%). The mean IgA IC for each cancer group tested (HN = 11.38 +/- 12.54 micrograms/ml; NPC = 13.36 +/- 17.56 micrograms/ml; LC = 17.39 +/- 13.04 micrograms/ml; CC = 26.50 +/- 4.60 micrograms/ml) were significantly elevated (P = 0.001) over both the normals (5.12 +/- 4.09 micrograms/ml) and the benign surgery controls (5.92 +/- 5.04 micrograms/ml). In addition to providing a new tumor marker the presence of high levels of IgA IC in cancer patients could provide a source of tumor-specific antibody as well as antigen and provide reagents to study immune regulation in cancer patients.

Immune complexes, serum proteins, cell-mediated immunity, and immune regulation in patients with squamous cell carcinoma of the head and neck.

A collaborative study of the humoral and cellular immune status of patients with carcinoma of the Head and Neck (H&N) was conducted at the West Virginia University (WVU) hospital. In addition, blind-coded serum panels were supplied on H&N cancer patients being treated at the National Cancer Institute (NCI). Serum protein analysis of the WVU study groups revealed that at the pretreatment sampling, the alpha-1 acid glycoprotein (AGP), total complement, and IgA levels were significantly elevated. The AGP levels and total complement levels declined to normal levels in the post-treatment period, whereas the IgA levels remained elevated throughout the entire observation period. Levels of serum immune complexes (SIC) were measured in both the WVU and NCI H&N cancer populations using the polyethylene glycol (PEG) precipitation method. In both survey populations all cancer groups had significantly elevated levels of SIC when compared to any of the control populations. The SIC levels never returned to comparative normal values even in cases after successful treatment. A subpopulation of the WVU-H&N cancer study group underwent a short course of intravenous hyperalimentation prior to their treatment regimen. These patients demonstrated a transient decrease in their SIC levels as well as a concomitant increase in their in vitro cell-mediated immune (CMI) correlates. The analysis of in vitro CMI correlates of the WVU study group using both polyclonal mitogens and specific antigens demonstrated a significant depression in these parameters pretreatment and post-treatment. In addition, it was observed that the time course for elevation of selected serum proteins (i.e., IgA and SIC) correlated with concomitant drops in CMI activity. Investigations were also conducted into the effects of immune complex-rich serum fractions upon selected in vitro CMI correlates. Significant blockage of a normal donor leukocyte migration-inhibition assay was demonstrated. Also, a similar inhibition of the ability of normal human lymphocytes to form high affinity rosettes was accomplished with serum from H&N cancer patients.

Serum ferritin as a tumor marker in patients with squamous cell carcinoma of the head and neck.

A double antibody enzyme immunoassay was used to measure serum ferritin levels in several different control and tumor-bearing populations collected from two institutions. The control groups consisted of normal volunteers, smokers, and Latter Day Saints. No statistically significant differences were noted in ferritin levels between pairs of these groups. Differences were noted among the normal groups when separated on the basis of age and sex, with higher ferritin levels in individuals older than 32 years of age and in men. By one-way analysis of variance, most control groups and subgroups were shown to have significantly lower levels (P less than 0.05) than the head and neck cancer group, with the exception of male smokers, who had levels comparable to male head and neck cancer patients. Serum ferritin levels were higher in head and neck cancer patients than in controls; however, there was no difference when compared with patients with other types of solid malignancies or when considering the anatomic site of the head and neck lesion. Ferritin levels were significantly (P less than 0.05) higher in patients with advanced (Stages III and IV) cancer than in those individuals with Stage I or II cancer. In patients with no evidence of clinical disease 5 years after treatment, the ferritin level had essentially returned to normal. In a group of head and neck cancer patients followed longitudinally, a significant decline in ferritin levels (P less than 0.05) was seen by 5 months after the completion of successful treatment. Furthermore, ferritin levels showed a tendency to increase or remain at high levels in patients with a poor prognosis and to decrease in those patients with a favorable prognosis, giving support to the contention that ferritin may prove to be a valuable tumor marker in head and neck cancer.

Increased serum antibacterial activity after turpentine-induced acute inflammation.

An acute inflammatory response was induced in rabbits by an intramuscular injection of turpentine. After this injection serial serum samples were obtained and serum haptoglobin levels were monitored. In addition, increased antibacterial activity was measured using a viable plate and photometric growth assay. As early as 4 hrs after turpentine challenge an increase was noted in serum antibacterial activity towards Staphylococcus aureus. At 48 hrs pronounced killing activity was demonstrated against S. aureus and S. epidermidis but not against Escherichia coli or other gram negative bacilli. By five days another serum bactericidal activity was present against Bacillus subtilis. Enhanced serum antibacterial activity persisted for 2 weeks. Partial characterization of these factors indicates that they are probably beta lysins.

Soluble tumor-associated markers in lung cancer extracts.

Three lung tumor-associated markers (LTAM), previously identified as a Cohn Fraction IV alpha-globulin (LTAM-1), ferritin (LTAM-2) and lactoferrin (LTAM-3) were separated by a combination of ion exchange, dye-affinity and molecular sieve chromatography. They were further purified by polyacrylamide gel electrophoresis and antisera were raised. Analysis of human extracts by immunodiffusion showed that 80% of lung tumor extracts were positive for all three markers. Similarly, 70% of extracts from other tumors wre positive for LTAM 1, but only 10% of these extracts were positive for LTAM 2 and 3. Variation in concentration of LTAM 2 and 3 among several extracts was determined by a quantitative enzyme immunoassay. Analysis of a select group of extracts for carcinoembryonic antigen (CEA), alpha-fetoprotein and beta 2-microglobulin showed 50% of these extracts to have markedly elevated levels of CEA. The results suggest that ferritin, lactoferrin and CEA offer promise as markers for lung cancer.

A human tumour-associated membrane antigen from squamous-cell carcinoma of the lung.

Primary human squamous-cell lung tumours were disrupted using mechanical high-speed homogenization. Crude membranes were isolated by differential centrifugation at low speeds followed by 100,000 g centrifugation. Such membrane preparations were extracted with Triton-X-100 and passed over a DEAE cellulose column; the DEAE unbound fraction and the bound fraction eluted with 0·4M NaCl were used to immunize rabbits. We present here only data on a single lung-tumour-associated membrane antigen (TAMA) found in the unbound DEAE cellulose column eluates. This new Triton-X-100 extractable antigen is termed lung TAMA-1.A further purification of the antigen was achieved using two methods. The first used G-200 molecular-seive chromatography and this revealed lung TAMA-1 to have a mol. wt of 200,000. A second method used sucrose density-gradient isoelectric focusing, and the antigen had an isoelectric point of ∼3·0.Several important properties of the lung TAMA-1 were determined. The antigen has a cathodal gamma-type electrophoretic mobility at pH 7·6. The antigen was not detected in any normal human adult or foetal tissue extracts tested. It did not crossreact immunochemically with CEA, AFP or ß2 microglobulin. Lung TAMA-1 was detected in 80% of lung tumour Triton-X-100 extracts tested by counterimmuno-electrophoresis (CIEP) but was undetectable in breast or colon carcinoma extracts. Low frequency sonication did not deleteriously affect lung TAMA-1, but, 3M KCl eliminated its immunologic reactivity in CIEP. Finally, preliminary data were obtained using immunohistochemistry to localize in vivo lung TAMA-1 production.

Immune monitoring protocol for patients with carcinoma of the head and neck. Preliminary report.

Numerous investigators have observed a depression of cell-mediated immunity in patients with carcinoma of the head and neck using a variety of in vitro and in vivo assays. This report presents the data obtained when a group of head and neck cancer patients were evaluated for reactivity in an in vitro lymphocyte blastogenesis assay using polyclonal mitogens and specific antigens, numbers of peripheral blood T-lymphocytes, and levels of circulating immune complexes. Such an immunological monitoring protocol revealed a depressed reactivity of the cancer patients in the lymphocyte blastogenesis assay when compared to normal age-matched controls. We also observed that 75% of these patients had circulating soluble immune complexes in their sera before and after therapy. These preliminary data indicate that further research is needed to examine the potential role of soluble immune complexes in modulating the host's immune response.

Soluble immune complexes in sera from head and neck cancer patients: a preliminary report.

With the recent demonstration of circulating immune complexes in a variety of malignant and nonmalignant diseases, we have examined the sera of head and neck cancer patients for evidence of soluble immune compleses. Using the Raji, cell test, we have shown that immune complexes are present in over 80% of the cancer sera examined as compared to less than 10% of normal control sera, and that these complexes persist following treatment of the patients by surgery or radiation therapy. These complexes may be acting as blocking factors which would account for the anergic state of these patients.

Isolation and identification of human lung tumor-associated antigens.

Three human lung tumor-associated antigens (TAA's) have been identified in soluble and membrane-solubilized extracts of human squamous cell lung carcinoma with the use of antisera raised in rabbits. The antigens were identified and partially characterized by means of an agarose adsorption technique. These antigens, termed lung TAA's 1,2, and 3, are all soluble in 50% ammonium sulfate, are antigenically distinct, and do not cross-react with carcinoembryonic antigen or alpha-fetoprotein. Lung TAA's 1 and 2 are oncofetal antigens demonstrable in soluble extracts from 24-week-old but not from 26-week-old fetal lungs. Rabbit antibodies to these lung TAA's were not adsorbed by types A, B, and O human red blood cells, serum proteins as well as soluble or insoluble lung preparations. Of several commercial antisera to human proteins, none cross-reacted with lung TTA 1, but anti-human liver ferritin cross-reacted with lung TAA 2, and anti-human lactoferrin cross-reacted with lung TAA 3. Lung TAA 1 was partially adsorbed and cross-reacted with certain normal serum or plasma preparations used and appears to be a normal serum protein in Cohn Fraction IV-4. Lung TAA 2 and 3 appear only in lung tumor-soluble extracts, whereas the lung TAA 1 was demonstrable in soluble extracts of breast, colon, cervical and head and neck carcinoma. All may be tumor markers of value in immunodiagnosis.

Chronic serous otitis media: an immune complex disease.

In characterizing the immunochemical and biochemical nature of middle ear effusions (MEE) of chronic serous otitis media (SOM), we have previously shown that MEE contain all major immunoglobulin classes, increased levels of several lysosomal enzymes, decreased total complement, but increased levels of C3. This report extends these observations by showing that MEE also contains C3 proactivator (C3PA) that can be activated by those organisms involved in acute otitis media, which confirms a functional alternate complement pathway. Furthermore, we have demonstrated the presence of soluble immune complexes in MEE using the Raji cell test. These data further support an immune complex mediated etiopathogenesis for chronic SOM.

Single-tube mixed agglutination test for the detection of staphylococcal protein A.

A simple, rapid mixed agglutination test using sheep erythrocytes (SRBC) sensitized with rabbit hemolysin and intact viable staphylococci is described for the detection of bound staphylococcal protein A. Soluble protein A was heat extracted from 50 clinical isolates as well as the Cowan I and Wood 46 strains of Staphylococcus aureus and titered by a hemagglutination test using sensitized SRBC and dilutions of soluble protein A. Protein A could be detected in all of these supernatants including that of S. aureus Wood 46, a strain generally considered to be protein A negative. These organisms were later retested by the mixed agglutination test and even those staphylococcal isolates expressing very low heat-extractable soluble protein A concentrations (1:2 titers) were positive, confirming the sensitivity of the test. In a screen of clinical isolates, only 4 of 235 (1.8%) coagulase-positive isolates were negative in the mixed agglutination test. Of 25 coagulase-negative isolates, none yielded a positive reaction.