Immunological approaches for early cancer detection.
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Animals↗
Biomedical subjects
D M Goldenberg
Publications and source records attributed to D M Goldenberg.
A murine monoclonal antibody raised against human non-small cell carcinoma of the lung.
Murine monoclonal antibodies (MAbs) reactive with human non-small cell carcinoma of the lung (NSCCL) were produced following immunization with a membrane preparation of Calu-3, a human adenocarcinoma of the lung cell line grown in nude mice. Positive hybrids unreactive with normal liver membranes and human peripheral white blood cells were selected for further testing. One MAb (RS5-4H6) recognized an antigen expressed in a variety of lung and other carcinoma cell lines as detected by flow cytometry. Immunohistology showed a selectivity for normal and neoplastic lung epithelium, as well as other cancers. The antigen was detected by immunohistology in 87% of tumor specimens derived from the lung, breast, colon, kidney, and ovary. Most other normal human tissues were not stained but occasional cells of the stomach, salivary and other glands, as well as kidney tubules were reactive. This MAb is an IgG1. Western blot analysis indicated that the antibody reacts selectively with an antigen greater than 300 kD. The antigen is resistant to neuraminidase treatment and periodate oxidation, but sensitive to pronase treatment, suggesting that the epitope is peptide in nature. This antibody may be potentially useful as a targeting agent for radioimmunodetection and immunoconjugate therapy.
Adenocarcinoma↗
Purification and characterization of carcinoembryonic antigen from GW-39, a xenografted human colonic tumor system.
Carcinoembryonic antigen (CEA) was purified from GW-39 human tumor xenografts in hamsters by immunoaffinity chromatography. Binding of the antigen to immobilized monoclonal antibody provided a high degree of purification of CEA in a single step. A recovery of 79% and a 750-fold purification were obtained. The purified CEA has a molecular size of 180 kilodaltons, an isoelectric point of 4.4, and a specific activity of 0.94. About 73% of the radiolabeled GW-39 CEA reacted with goat anti-CEA serum. The amino acid and carbohydrate compositions of the GW-39 CEA were comparable to those of CEA preparations from other sources.
Amino Acids↗
"Sandwich"-type immunoassay of carcinoembryonic antigen in patients receiving murine monoclonal antibodies for diagnosis and therapy.
Measurements of carcinoembryonic antigen (CEA) in blood increased dramatically in some patients who were receiving injections of monoclonal antibody. CEA titers were measured with a monoclonal antibody-based double-determinant enzyme immunoassay in which untreated plasma specimens were diluted with an equal volume of buffer containing mouse serum. Increasing CEA titers were accompanied by the appearance and coincident increase in titers of human antibody against mouse Ig (HAMA). Adsorption of these sera with solid-phase anti-human IgG or Protein A restored antigen titers to pretreatment values; evidently the serum factor eliciting false-positive CEA titers was most probably HAMA. Neither addition of undiluted mouse serum to the assay mixture nor pretreatment by heating plasma specimens to 70 degrees C effectively abolished HAMA interference. By contrast, protein precipitation with polyethylene glycol (130 g/L) or heating plasma samples to 90 degrees C eliminated false-positive titers caused by HAMA, but did not reduce authentic CEA titers.
Animals↗
Targeting of cancer with radiolabeled antibodies. Prospects for imaging and therapy.
This article reviews the current status, including problems and some proposed solutions, of the targeting of cancer with radioactive antibodies for use in antibody imaging (radioimmunodetection) and radioimmunotherapy. The problems and results are similar for purified polyclonal and murine monoclonal antibodies. Foremost among the problems are the low accretion of antibody (0.01% to 0.001% of injected dose per gram) in tumor and the nonspecific deposition of radiometals that are more ideal for imaging or therapy after antibody conjugation. Despite these limitations, radioimmunodetection appears to be a safe and useful method, even at this early stage of development. Sensitivity, specificity, and accuracy rates of 80% to 90% have been achieved in some studies involving radioiodine labels and polyclonal or monoclonal antibodies, whereas lower percentages have been achieved with indium 111 or technetium Tc 99m radioconjugates. Even at a usual tumor resolution of 1.5 to 2.0 cm, occult cancers have been disclosed by radioimmunodetection when missed by traditional detection measures, including computed tomography and magnetic resonance imaging. Radioimmunotherapy is somewhat less developed as a treatment modality, but encouraging remissions have been observed, thus stimulating further active pursuit of this technology. These targeting results have been achieved with antibodies that are not truly cancer specific, but only exploit quantitative differences in antigen expression between tumor and adjacent normal tissues. Circulating target tumor antigens do not appear to prevent successful tumor targeting of the radioactive antibodies.
Antibodies, Monoclonal↗
Radioimmunotherapy of the GW-39 human colonic tumor xenograft with 131I-labeled murine monoclonal antibody to carcinoembryonic antigen.
We have investigated the therapeutic efficacy of a single injection of 131I-labeled murine mouse monoclonal antibody (NP-4) against carcinoembryonic antigen using the human colonic tumor xenograft, GW-39, grown in the cheek pouches of adult hamsters. Therapeutic efficacy was dependent on the dose of radioactivity, the specificity of the antibody for the tumor, and the size of the tumor when the radioantibody was administered. A dose of 1 mCi of 131I-labeled NP-4 given 1 day after tumor transplantation completely inhibited the growth of 6 of 11 tumors over a 12-week period, and histological evidence indicated that viable tumor was absent in the tissue remaining at the injection site. Lower doses (0.5 mCi) of 131I-labeled NP-4 inhibited tumor growth over 90% in comparison to untreated animals, but the tumors eventually resumed growth. Delaying the administration of radioantibody for 4 or 7 days after tumor transplantation significantly reduced the therapeutic efficacy. Although the same dose of 131I-labeled irrelevant immunoglobulin G also inhibited tumor growth, 131I-labeled NP-4 was generally 2-3 times more effective in reducing tumor growth than was the control IgG. There was a 13% loss in body weight within 7 days after treatment with 1 mCi, but all the animals regained their weight by day 14, indicating that the level of radioactivity was tolerated well. Dosimetric calculations predicted that over 14 days a dose of nearly 2400 rads was delivered to the tumors with 131I-labeled NP-4. These results confirm our previous studies that 131I-labeled antibody can effectively inhibit tumor growth, but suggest that radioantibody therapy is most effectively administered when there is a low tumor burden.
Animals↗
Antibody protein dose and radioimmunodetection of GW-39 human colon tumor xenografts.
This study investigates the influence of antibody protein dose on the radioimmunodetection of a CEA-producing, human colonic tumor xenograft (GW-39) using affinity-purified goat anti-carcinoembryonic antigen (CEA) antibody and a murine monoclonal anti-CEA antibody (NP-2). Hamsters bearing GW-39 tumors were given an equal mixture of 131I-labelled antibody and 125I-labelled irrelevant IgG at doses varying from 0.01 to 1.0 mg (0.1 to 10 mg/kg body weight) for each antibody preparation. No differences were found in the percentage of antibody in the tumor or the clearance rate from the blood as the antibody dose was increased. The concentration of antibody in the tumor increased proportionally with the antibody dose and maintained a linear relationship with increasing dose, indicating that antigenic sites in the tumor were not saturated with antibody. The concentration of irrelevant IgG in the tumor also increased proportionally as the dose was increased, but the concentration of irrelevant IgG in the tumor was less than the antibody and was removed more rapidly from the tumor than the antibody. By considering the amount of irrelevant IgG in the tumor as a measure of the amount of antibody non-specifically bound, we determined that there was no change in the amount of antibody in the tumor between days 3 and 7, and the amount of antibody increased uniformly for both the tumor and non-tumor tissues, resulting in no improvement in the tumor/non-tumor ratios with increasing antibody dose. External scintigraphy verified that the dose of the antibody did not influence tumor imaging. Thus, tumor accretion of antibody in this model is not dependent principally on the antibody protein dose, and other factors such as accessibility to antigen and antibody metabolism may play important roles in regulating the uptake of antibody in tumor.
Animals↗
Relationship of radioantibody localization and cell viability in a xenografted human cancer model as measured by whole-body autoradiography.
The simultaneous distribution of monoclonal 131I-labeled anti-carcinoembryonic antigen (CEA) immunoglobulin (IgG) (NP-2) or 131I-labeled irrelevant myeloma IgG (Ag8) and [3H]thymidine was studied in hamsters bearing transplants of the GW-39 human colon carcinoma by qualitative double-tracer whole-body autoradiography. Autoradiography showed that large solid GW-39 tumors are characterized by heterogeneity of radioantibody retention and uneven [3H]thymidine accumulation, reflecting zonal variations in antibody reactivity and tumor cell proliferation, respectively. The autoradiographic images showed that both 131I-labeled-monoclonal antibody and control 131I-labeled IgG targeted nonproliferating tumor zones, suggesting a mechanism of nonspecific tumor uptake of radioantibodies in these areas. Absence of tumor center labeling with [3H]thymidine, associated with cellular necrosis, was confirmed by histology and microautoradiography in separate animal studies. In confirmation of earlier reports, 131I-labeled anti-CEA monoclonal antibody gave higher tumor-to-non-tumor labeling patterns than did control 131I-labeled IgG, at both 3 and 7 days following treatment. Immunohistochemical localization of CEA in GW-39 tumors with necrotic centers showed the presence of CEA in nonviable cells, but CEA antigen concentrations were diminished as compared to cells located in the tumor's periphery. The results indicate that double-tracer whole-body autoradiography is well suited for studying the kinetics of radioantibody localization in relation to regional tumor cell viability.
Animals↗
Current status of cancer imaging with radiolabeled antibodies.
This editorial reviews the development, current status, and future prospects of cancer imaging with radioactive antibodies, termed radioimmunodetection (RAID). There has been a slow and steady development of this field for more than 35 years, with more recent activity and progress resulting from the identification of human tumor-associated antibodies and suitable human tumor xenograft models, the demonstration that circulating antigens do not prevent radioantibody localization in tumor, the development of computer-assisted and biological methods for reducing non-target background radioactivity, and the advent of hybridoma-produced monoclonal antibodies. At the present time, tumor sites in the range of 1.5 to 2.0 cm can be imaged, with the best resolution of 0.4-0.5 cm being reported with new chelates of 99mTc. A number of factors, including character of the radioantibody and its bioavailability, the tumor antigen site and bioavailability, the character of the radiolabel, and the target tumor's size, location and vascularization, contribute to the sensitivity, specificity, accuracy, and resolution of the method. Already at this early stage of development, RAID has been shown to have, in certain tumor types and with particular antibody and imaging systems, an accuracy of tumor site detection of over 90%, with the disclosure of occult lesions. Carefully designed prospective trials are needed to fully assess the role of this new modality in the management of cancer patients, particularly in early detection of primary and recurrent tumors.
Antibodies, Monoclonal↗
Imaging of primary and metastatic liver cancer with 131I monoclonal and polyclonal antibodies against alphafetoprotein.
Thirteen patients with a history of confirmed liver carcinoma were given either I131 goat polyclonal or murine monoclonal antibodies against alpha-fetoprotein (AFP), and then scanned with a gamma camera. In order to reduce background, nontarget activity, especially in the liver, blood pool, and reticuloendothelial system, 99mTc imaging agents were used for tumor image enhancement by computer-assisted subtraction. A sensitivity of 91% for the primary site, 50% for the lungs (33% for the chest area), and 75% for the abdomen and pelvis was achieved, with a specificity of 100%, 94%, and 100% for these sites, respectively. The accuracy was determined to be 93% for the liver, 86% for the lungs (77% for the chest), and 85% for the abdominal and pelvic area, resulting in an overall accuracy rate for imaging primary and metastatic hepatocellular cancer of 84% (90% if bone metastases are excluded). In two of the 13 patients, lesions that had been missed by conventional liver scintigraphy and transmission computed tomography (CT) were first shown by radioimmunodetection (RAID).
Abdominal Neoplasms↗
Representation of epitopes on colon-specific antigen-p defined by monoclonal antibodies.
Colon-specific antigen-p (CSAp) is a large molecular-sized protein restricted to normal and neoplastic gastrointestinal tissues and to some mucinous ovarian tumors. Murine monoclonal antibodies (MAbs) were raised against CSAp that was affinity purified with goat polyclonal antibodies from GW-39 human colonic carcinoma xenografts or against the CSAp-producing colon cancer cell line SW-948. Two of the MAbs, designated Mu-2 and Mu-4, recognized a CSAp determinant containing sialic acid, and this epitope was also expressed on bovine submaxillary mucin (BSM). Blocking experiments demonstrated that the Mu-2 and Mu-4 MAbs recognized different determinants. A third MAb, Mu-3, did not cross-react with BSM, but unlike Mu-2 and Mu-4, it did react with human saliva. Reactivity of Mu-3 with saliva did not correlate with major blood group and Lewis-related secretory blood group substances in saliva. This reactivity was not related to sialylated Lewis activity. The fourth antibody, Mu-1, appeared to react with a conformational determinant since its epitope was destroyed by heat treatment or thiol reduction. Enzyme immunoassays have demonstrated that all four epitopes may be expressed on one molecular species.
Animals↗
Anti-antibody enhancement of iodine-131 anti-CEA radioimmunodetection in experimental and clinical studies.
Imaging of tumors with radiolabeled antibodies, especially when located in the blood-rich visceral organs, may be improved through administration of a second antibody directed against the primary tumor-associated antibody. In hamsters bearing a human colonic carcinoma xenograft producing carcinoembryonic antigen (CEA), we injected donkey anti-goat IgG 24 hr after administration of 131I-labeled goat anti-CEA IgG and achieved enhanced tumor imaging 24-48 hr later, with a significant relative decrease of radioactivity in blood and all major organs except the spleen. In seven of nine patients, this method of anti-antibody clearance of nontargeted radioactive murine monoclonal antibodies revealed sites of cancer, including liver metastases. Characterization of radioactivity in the plasma before and after administration of the second antibody confirmed that complexes were quickly formed between primary and secondary antibodies, and imaging of the patients revealed a rapid uptake of radioactivity in the liver at 2 hr that dissipated within 24 hr. Radioactivity in the spleen gradually increased over time. The method of anti-antibody immunological enhancement of cancer imaging is feasible and may reveal tumor sites missed by conventional imaging.
Animals↗
Advances in cancer: antibody targeting of cancer--prospects for detection and therapy.
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Antibodies, Monoclonal↗
Preliminary findings in the evaluation of hepatic malignancies by radioimmunodetection, X-ray computed tomography, and magnetic resonance imaging.
In 12 consecutive patients with suspected metastatic carcinoma of the liver, we evaluated the sensitivity of radiolabeled antibodies to tumor antigens, magnetic resonance imaging, and X-ray computed tomography imaging in the detection of hepatic malignancies. Studies were performed with 131I labeled antibodies to CEA and/or CSAp; polyclonal, monoclonal and F(ab')2 antibodies were used. Nontarget radioactivity was diminished by administration of 99mTc reagents simulating nontumor distribution and use of a computer subtraction method. In nine patients with confirmed liver neoplasms, radioimmunodetection disclosed the foci of hepatic malignancies. In three patients with suspected liver neoplasms, the antibody studies were positive, but at this time have not been confirmed. X-ray computed tomography each disclosed mass lesions in five patients and magnetic resonance in three. These findings suggest that radioimmunodetection provides greater accuracy in the detection and localization of cancer than other diagnostic modalities currently used.
Adenocarcinoma↗
A human colon cancer metastasis model for radioimmunodetection.
One important issue in radioimmunodetection is how well the current methods can locate and disclose small metastatic foci in visceral sites. We have developed a human colonic tumor metastasis model by surgically implanting GW-39 tumor cells in the liver of unconditioned hamsters. Tumors were produced in 71 of 73 animals and were macroscopically apparent within 1 wk. In addition, multiple nodular lung metastases of GW-39 were found in about 80% of the animals given implants of tumor in the liver, but implantation of tumor in the spleen failed to show lung metastases even after 4 wk. Hamsters bearing GW-39 liver and cheek pouch grafts or normal hamsters were given injections of a mixture of 131I-labeled anti-carcinoembryonic antigen antibody and 125I-labeled irrelevant immunoglobulin G. After 7 days, tumor was localized by external scintigraphy without subtraction techniques in both the liver and cheek pouch, but even in animals with extensive lung metastases we failed to unequivocally detect tumor in the lungs by external imaging or by comparing tissue counting data from uninvolved and tumor-bearing lungs. However, whole-body autoradiography confirmed specific localization of anti-carcinoembryonic antigen antibody in the tumors at all sites indicating that tissue counting and external imaging were not sensitive enough to reveal micrometastatic tumors. Thus, the current methods used for this model appear to be useful for further investigation of the radioimaging of tumors growing in visceral organs.
Animals↗
Quantitative whole-body autoradiography of radiolabeled antibody distribution in a xenografted human cancer model.
Quantitative whole-body autoradiography (WBAR) was used to study the biodistribution of goat anti-carcinoembryonic antigen and normal goat IgG, each labeled with 125I, in hamsters bearing the carcinoembryonic antigen-producing GW-39 human colonic carcinoma xenograft. Comparisons between computer-assisted videodensitometric profiles of WBARs and tissue radioactivity counts were made at 1, 3, and 7 days following administration of the radiolabeled IgGs. The results indicated that maximal tumor accretion of the radiolabeled antibody and normal IgG occurred within 1-3 days, with a marked selective accretion of antibody in the tumor being evidenced at 3-7 days because of clearance of normal IgG. Radioactivity derived from antibody IgG showed 6.5 to 118.7 times that found in other tissues, as measured by videodensitometry, whereas organ radioactivity counting revealed ratios of only 6.7 to 29.6. Specificity of tumor-cell accretion of the radiolabeled antibody was confirmed by microscopic autoradiography, showing intense labeling of the proliferating perimeters of GW-39 tumors. WBAR was found to have a resolution of 0.10 to 0.25 mm in 100-g hamsters, which appears to be greater than the resolving power of external body imaging by gamma camera scintigraphy. These studies suggest the use of WBAR and microautoradiography to complement external imaging methods for the analysis of antibody distribution and localization in cancer radioimmunodetection models.
Animals↗
Diagnosis and treatment of neoplasms with radionuclide-labeled antibodies.
The in vivo use of radionuclide-labeled antibodies to several types of tumor-associated antigens or products has proven to be a reliable method for defining primary, recurrent, metastatic, and occult carcinoma. Specificity for the radioimmunodetection procedure is high; however, the sensitivity varies depending on the size and site of the lesions. The relative roles of polyclonal- and monoclonal-type antibodies are still to be defined, since at this stage of development little difference can be demonstrated in the detection of tumor with either one. The introduction of emission computed tomography should simplify the imaging techniques appreciably and improve the resolution between nontarget and target radioactivity. Satisfactory radionuclide labels with photon energies more compatible with current imaging equipment will enhance resolution and sensitivity.
Animals↗
Neutron-capture therapy of human cancer: in vivo results on tumor localization of boron-10-labeled antibodies to carcinoembryonic antigen in the GW-39 tumor model system.
Antibody against carcinoembryonic antigen (CEA) was conjugated with p-[1,2-dicarba-closo-[1-3H]do-decaboran(12)-2-yl] benzenediazonium ion by an azo-coupling reaction, resulting in 30 boron atoms per IgG molecule with no loss of antibody protein. Antibody immunoreactivity was not appreciably affected by this conjugation and was stable after incubation in vitro in hamster plasma for 24 hr. The efficacy of the boron-conjugated anti-CEA IgG for localizing selectively in CEA-containing human colonic carcinomas propagated in the hind leg musculature of hamsters was evaluated by labeling the antibodies with 131I and determining distribution of the radioactivity in vivo. The results show that the boron-conjugated antibodies retain selective localization in the tumors, thus indicating their suitability for transporting boron-10 to tumors for use in neutron-capture therapy of cancer.
Antibodies, Neoplasm↗