A solid-phase, enzyme-linked immunosorbent assay for a human lung tumor-associated antigen

A solid-phase, enzyme-linked immunosorbent assay (ELISA) for a human lung tumor-associated antigen (LTA) was based on immobilized LTA that was detected with the use of an antiserum raised in a goat against a highly purified antigen preparation. Bound goat antibodies were detected in a series of steps that included incubation with a) biotinylated rabbit antibodies to goat immunoglobulins, b) glucose oxidase conjugated to avidin, and c) peroxidase and the substrates glucose and 2,2'-azino-di-(3-ethylbenzthiazoline sulfonic acid). The absorbance of the final product was measured at 405 nm, and its formation was dependent on substrate incubation time and antibody concentration. The antigen was immobilized and highly purified, and the goat antiserum was bound to and eluted from an immobilized crude antigen column before use. The ELISA could detect less than 1 ng antigen and was able to discriminate extracts of normal lung tissue from those of lung tumor. As was found earlier with a radioimmunoassay for the same antigen, normal human serum could inhibit in the ELISA but only when used at high concentration, indicating levels of antigen or antigen-like activity in the 100-200 ng/ml range. With the use of this assay, 3 lung cancer patients were monitored 6-12 months prior to death. In all 3 patients, LTA levels rose dramatically 2-4 months before the patients died; in 2 patients the levels exceeded 3,000 ng/ml just before death. In contrast, in 2 of these patients, carcinoembryonic antigen levels remained essentially unchanged, with no more than a twofold increase prior to death.     

Inhibition of experimental pulmonary metastasis of mouse colon adenocarcinoma 26 sublines by a sialic acid:nucleoside conjugate having sialyltransferase inhibiting activity

The total and sialidase-releasable sialic acid contents of murine colon adenocarcinoma 26 sublines of high (NL-17) and low (NL-44) metastatic potential were found to be positively correlated with their ability to undergo metastasis. Furthermore, sialyltransferase activity of intact NL-17 cells was higher than that of NL-44 cells. These findings suggest that sialic acid on the cell surface may play a role in the metastasis of these cells. Therefore, the effect of a sialyltransferase inhibitor, 5-fluoro-2',3'-isopropylidene-5'-O-(4-N-acetyl-2,4-dideoxy-3,6,7,8-tetra -O -acetyl-1-methoxycarbonyl-D-glycero-alpha-D-galactooctapyranosyl)u ridine (Kl-8110), on the experimental lung metastasis of NL-17 or NL-44 cells was examined. Kl-8110 inhibited the transfer of sialic acid to its endogenous acceptor in NL-17 and NL-44 cells. NL-17 or NL-44 cells were injected into the tail veins of mice, and the metastasis-inhibiting activity of Kl-8110 was evaluated on the basis of both the lung weight and the number of pulmonary surface nodules about 3 wk after the tumor cell injection and of the survival ratio of mice inoculated with the tumor cells. Pretreatment of tumor cells with Kl-8110 together with i.v. injection of Kl-8110 caused significant inhibition of pulmonary metastasis of both NL-17 and NL-44 cells. Inhibition of metastasis and prolongation of survival were also observed on i.v. injection of Kl-8110 without pretreatment of the tumor cells with Kl-8110, but the degree of inhibition was lower than that in the case of the two treatments together. Kl-8110 itself had neither cytostatic nor cytotoxic effects on NL-17 and NL-44 but reduced the retention of tumor cells in the lungs. This antimetastatic effect of Kl-8110 may be due to modification of the tumor cell surface resulting from inhibition of sialyltransferase by Kl-8110. In addition, a beta-linked sialic acid:nucleoside conjugate (Kl-8111) and an equimolar mixture of Kl-8110 and Kl-8111 (Kl-414) also inhibited the metastatic ability of NL cells to the same extent as Kl-8110 did.     

Membrane antigen in small cell carcinoma of the lung defined by monoclonal antibody SM1

Mouse myeloma cells were fused with spleen cells from BALB/c mice immunized with a cell line derived from human small cell carcinoma (SCC) of the lung. The cloned hybridoma SM1 produced antibody that was reactive with the surface membrane of SCC cell lines and SCC tumors but not with the membrane of several non-SCC cell lines and tumors. SM1 ascites fluid was used to screen for reactivity of the antibody with other human cancer cell lines, tumor tissues, and normal tissues. SM1 antibody was found to be unreactive with neuroblastoma, adrenal carcinoma, melanoma, and bronchial carcinoid. Reactivity was detected with some breast carcinoma cell lines but not with breast cancer tissue specimens. In the same individual, the antibody was reactive with SCC lung tumor and SCC metastatic to the liver but not with normal tissues, including bronchus, lung parenchyma, liver, kidney, and brain. Human erythrocytes and marrow cells were also unreactive. Since SM1 detects an antigen that is present in greatest amounts on the surface membrane of SCC of the lung, this antibody may be useful in tracing the lineage patterns of human lung cancers.     

人肺癌高转移动物模型的筛选及其细胞系的建立

目的:建立人肺癌小鼠高转移模型及高转移细胞系,同时观察相关生物学特性,为肺癌转移机制和防治等研究提供有用的实验工具。方法:切除首代小鼠移植瘤,以延长动物生存时间而获得转移灶,从第二代起采用肺转移灶→皮下移植→肺转移灶→皮下移植的体内循环筛选方法建立NOD/SCID小鼠人肺癌细胞SPC-A-1皮下移植瘤高转移模型,并进行肿瘤的生长和转移情况、组织病理学观察,同时建立相应的高转移细胞系,进行各种相关生物学特性观察。结果:第一代移植瘤切除后转移率达66.7%,通过4代体内反复筛选建立了100%肺转移NOD/SCID小鼠模型及相应高转移细胞系,细胞生长行为和染色体分析等生物学特性观察表明该细胞系保持了原有的人肺腺癌的生物学特性。结论:应用体内筛选的方法成功建立了人肺癌皮下移植瘤高转移模型及高转移细胞系,为肺癌防治研究及抗转移实验治疗提供了理想的动物模型。     

Identification of a surface antigen, common to murine lung and Lewis lung carcinoma, by monoclonal antibody

Lewis lung carcinoma (3LL) is a spontaneous murine lung carcinoma which preferentially metastasizes into the lung. Monoclonal antibodies (MAb) against 3LL cells were prepared by fusing spleen cells from rats immunized with 3LL and the P3-NSI/I-Ag4 (NSI) plasmacytoma. Two hybridomas were selected which secreted antibodies capable of preferential binding to 3LL cells and not to other malignant cells. Immunohistochemical staining of sections prepared from various mouse organs with one of the MAbs (22.2), revealed exclusive staining of the lung tissue. The data suggest that a lung-specific, 3 LL-associated antigen, is detected by the 22.2 MAb on 3LL cells. The 22.2 antigen is a surface molecule, and is stable following trypsin treatment and fixation by glutaraldehyde or picric acid. Cells that were prepared from a subcutaneous tumor or from lung metastases express high amounts of 22.2 antigen compared to cells of the culture 3LL line (3LLC). Upon reinoculation of 3LLC cells into mice there was a gradual increase in 22.2 antigen expression. It was possible to select from the heterogeneous 3LLC cell population several clones which express high amounts of antigen. These clones differed also in their metastatic patterns. However, the metastatic capacity of these clones did not correlate with expression of the 22.2 antigen. Thus, it appears that expression of the 22.2 antigen is not a necessary condition for formation of lung metastases by 3LL.     

Inhibition of spontaneous and experimental metastasis by a new derivative of camptothecin,CPT-11, in mice

Summary The antimetastatic effect of a new water-soluble derivative of camptothecin, 7-ethyl-10-(4-(1-piperidino)-1-piperidino) carbonyloxy-camptothecin (CPT-11), were examined in several metastatic murine tumor systems. Intravenous (i.v.) injection of CPT-11 into BALB/c mice inhibited lung metastasis by i.v. inoculated, metastatic colonic adenocarcinoma 26 (C26) cells, C26NL-17, in BALB/c mice. This treatment was also effective in C57BL/6 mice against lung metastasis by i.v. inoculated B16-F10 and B16-BL6 cells, highly metastatic variants of the B16 melanoma. Furthermore, intraperitoneal (i.p.) injection of CPT-11 significantly inhibited the growth of C26NL-22 cells, a highly metastatic variant of C26, inoculated subcutaneously (s.c.) into the left front footpads of BALB/c mice. Also, i.p. or i.v. injection of CPT-11 effectively inhibited the growth of 3LL tumors inoculated s.c. into the hind footpads of C57BL/6 mice. Moreover, following s.c. inoculation of either C26NL-22 or 3LL cells, combined surgical excision of the primary tumor and either i.p. or i. v. CPT-11 injections given before or after surgery markedly inhibited the formation of pulmonary metastases. These results show that a new derivative of camptothecin, CPT-11, has a potent inhibitory effect against both spontaneous and experimental lung metastasis.     

Experimental studies with the use of metastatic clones isolated from mouse tumors

Low and high metastatic clones were isolated from mouse colon 26 by repeated cloning of tumor cells in the lung. These clone cells showed an interesting metastatic capacities, low and high lung metastasis. The results indicate that original tumor consisted of various and heterogeneous cell clones having different metastatic capacities. Using highly metastatic clones, NL-17 and B16-F10 and B16-BL6, adriamycin and 5-fluorouracil were found similarly effective to both parental and cloned tumor lines. Although there are several reports to indicate heterogeneous drug sensitivity of metastatic clones from the parental line, present study indicated the similar drug sensitivity. Therefore, heterogeneity of drug sensitivity is scheduled to check with different antitumor drugs.     

Suppression of experimental lung colonization of a metastatic variant of murine colon adenocarcinoma 26 by a monoclonal antibody 8F11 inhibiting tumor cell-induced platelet aggregation

We have previously established and characterized two monoclonal antibodies, 8F11 and 20A11, that recognize an Mr 44,000 membrane glycoprotein of metastatic murine colon 26 cells. Both monoclonal antibodies inhibit platelet aggregation induced by the tumor cells in vitro. In this report, the inhibitory effect of 8F11 on lung colonization of i.v.-inoculated tumor cells was examined. The i.v. administration of 8F11 suppressed lung colonization of NL-17, a highly metastatic variant of colon 26. Inhibition of NL-17 lung colonization by 8F11 was dose dependent with a maximum of 80% inhibition at a dose of 800 micrograms 8F11/mouse. 8F11 did not inhibit metastases at doses lower than 100 micrograms/mouse. Inhibition of pulmonary metastases by 8F11 was greatest when the antibody was administered 2 h before tumor inoculation. The effect was diminished when the antibody was given 2 h after tumor inoculation. The pulmonary retention of i.v.-inoculated radiolabeled NL-17 cells was decreased by 8F11. F(ab')2 fragments of 8F11 also effectively inhibited lung colonization by NL-17 cells, suggesting that mechanisms unrelated to immune-mediated destruction are involved. These results indicate that the monoclonal antibody 8F11 suppresses the lung colonization of NL-17 cells by interfering with the initial arrest of tumor cells in the lung vasculature through the inhibition of tumor cell-platelet interaction.     

Expression of tumor antigen correlated with metastatic potential of Lewis lung carcinoma and B16 melanoma clones in mice

Expression of a tumor-associated antigen, recognized by a monoclonal antibody (MoAb 135-13C) to lung carcinoma cells, has been studied in cloned Lewis lung carcinoma (3LL) and in B16 melanoma (F1 and F10) tumor lines endowed with different metastatic potentials. MoAb 135-13C recognizes a protein complex (tumor-specific Mr 180,000 protein) that appears on the cell surface of several murine lung carcinomas but is not detected on normal cells in culture. Standard metastatic variants of B16 melanoma (F1 and F10) and two variant sublines of 3LL (M1087 and BM21548) together with the parental line of 3LL have been used for these experiments. The two cloned variant lines derived from 3LL have been shown to retain high (M1087) and low (BM21548) metastatic phenotypes during in vivo passaging. We found that all three cell lines of 3LL bind monoclonal antibody specifically, but one cell variant with higher metastatic potential shows a higher capacity to bind MoAb 135-13C than did the other variant. Similarly we found that B16 F10 cells bind higher amounts of MoAb 135-13C than did B16 F1 cells. In addition the analysis of the amounts of MoAb 135-13C bound to the cell surface of several other in vitro and in vivo tumor lines with different metastatic capacity demonstrates that all tumor lines which express high ability to colonize to the lung also express, on the cell surface, higher amounts of tumor-specific Mr 180,000 protein. The sodium dodecyl sulfate-polyacrylamide gel electrophoresis autoradiograms of immunoprecipitates from cell lysates of 3LL and B16 tumor lines demonstrate that MoAb 135-13C specifically precipitated three proteins banding at molecular weights of 204,000, 134,000, and 116,000. We conclude that MoAb 135-13C recognizes a surface protein complex which is present in higher amounts in 3LL and B16 cells which possess higher capacity to metastasize to the lung.     

Characterization of a tumor cell surface protein with heterologous antisera to a spontaneous BALB/c lung carcinoma.

Line 1, a spontaneous alveolar carcinoma from a BALB/c mouse, is highly metastatic and weakly antigenic in the syngeneic host. Sera and enriched antibody preparations were made specific for line 1 cells by in vitro and in vivo absorptions. By lactoperoxidase-catalyzed radioiodination of cell surface protein followed by precipitation with specific antibodies, a protein with a molecular weight of about 180,000 (designated TSP-180) was identified that was present on line 1 cells but not on normal lung cells or Moloney sarcoma tumor cells. Studies of competition for immune precipitation of TSP-180 by unlabeled protein preparations from various sources indicate that (a) TSP-180 is present in tumor cells of various culture passage levels, (b) tumor cells grown i.m. also contain TSP-180, and (c) normal lung proteins compete weakly, and only at very high protein concentrations. A protein similar to TSP-180 was detected on Madison 109 carcinoma and on three other lung carcinomas adapted to culture. Experiments with specific antisera show that TSP-180 is not lactoperoxidase, fetal bovine serum protein, large external transformation-sensitive protein, or an antigen related to murine leukemia virus proteins.     

Insulin and insulin-like growth factor 1 stimulate proliferation of metastatic variants of colon carcinoma 26

The proliferation rate of malignant cells in vivo is one of the important factors which affect the formation of tumor metastasis. A highly metastatic variant of mouse colon adenocarcinoma 26 (NL-17) grew more rapidly than a low-metastatic variant (NL-44) both in vitro and in vivo. The effect of growth factors on the proliferation of NL-17 and NL-44 cells was examined in serum-free medium. Among growth factors examined, human insulin and insulin-like growth factor 1 (IGF-1), which were produced by gene engineering techniques, stimulated the growth of metastatic NL-17 and NL-44 cells as determined by thymidine incorporation and cell counts. DNA synthesis and cell proliferation of the high-metastatic NL-17 was stimulated to a greater extent by insulin and IGF-1 than those of the low-metastatic NL-44. These findings suggest that circulating growth factors could enhance the formation of tumor metastasis. Scatchard analysis of [125I]IGF-1 binding to NL-17 and NL-44 showed that each cell line had an almost equal number of IGF-1 receptors (1.37 x 10(5)/cell and 1.26 x 10(5)/cell, respectively), which had similar dissociation constants (8.94 x 10(-10) M and 9.54 x 10(-10) M, respectively). Since the number and affinity of IGF-1 receptors are equivalent between low- and high-metastatic cells, the intracellular events which result in the cell growth after binding of IGF-1 may differ between NL-17 and NL-44 cells.     

A different pattern of cytotoxic T lymphocyte recognition against primary and metastatic tumor cells in a patient with nonsmall cell lung carcinoma

BACKGROUND: Lung carcinoma represents the most frequent cause of cancer death worldwide because of tumor metastases. The objective of the current study was to analyze the immunologic response during the progress of lung carcinoma metastasis. METHODS: The authors established two tumor cell lines that were derived from primary and metastatic lesions in a patient with lung carcinoma (Patient G603). One cell line (G603L) was established from the primary lesion, and the other cell line (G603AD) was established from a metastatic lesion in the right adrenal gland 7 months after the patient underwent surgery for the primary lesion. Autologous regional lymph node lymphocytes were stimulated with CD80-transfected G603L cells, then cytotoxic T lymphocytes (CTLs) were induced against both lung carcinoma cell lines. RESULTS: Both G603L cells and G603AD cells expressed Class I human leukocyte antigen, intracellular cell adhesion molecule 1, and lymphocyte-associated antigen type 3 (LFA-3), but not Fas or Fas ligand on their surfaces. By stimulation with CD80-transfected G603L cells, 2 CTL clones (H2/17 and H2/36) were established from the bulk CTLs. CTL clone H2/17 lysed G603L cells but not G603AD cells, suggesting that the antigen recognized by CTL clone H2/17 was abrogated during the process of metastasis. In contrast, CTL clone H2/36 lysed both G603L cells and G603AD cells, indicating that the antigen recognized by CTL clone H2/36 was maintained in the tumor cells throughout tumor progression. CONCLUSIONS: The results demonstrated the possibility that some tumor-associated antigens may be abrogated during the process of metastasis, although others are maintained. The identification of these antigens will lead to a better understanding of their immunologic role during disease progression in patients with lung carcinoma.     

Murine monoclonal antibody LAM2 defines cell membrane determinant with preferential expression on human lung small-cell carcinoma and squamous-cell carcinomas

Murine monoclonal antibodies (MAbs) were generated against a human undifferentiated lung carcinoma cell line. The hybridoma designated LAM2 produced an IgM kappa MAb with reactivity to the cell membrane. Indirect immunofluorescence staining and radioimmunoassay showed LAM2 antibody to react preferentially with lung small-cell carcinoma (SCC) cell lines and squamous-cell carcinoma (SQC) cell lines. LAM2 antibody also stained primary cultures of normal bronchial epithelial cells, but was unreactive with human erythrocytes and nucleated marrow cells. Indirect immunofluorescent staining with LAM2 antibody was performed on frozen sections of human tumor tissues and normal tissues. LAM2 antibody stained all 8 SCC carcinomas, 4 of 5 SQC of the lung and head and neck region, and 2 or 4 lung large-cell carcinomas. No staining was seen on lung adenocarcinomas, breast carcinomas, ovarian carcinomas, renal cell carcinomas, colon carcinomas, or mesotheliomas. Staining was present on sections of normal bronchus, but not on normal lung parenchyma, liver, kidney, adrenal or skin. While LAM2 antibody was highly reactive with all SCC examined, its antigenic determinant was not expressed in other cell lines and tumors of presumed neuroectodermal origin, including neuroblastoma, melanoma, and bronchial carcinoid. Radioimmunoprecipitation showed the antigen defined by LAM2 antibody to have two major bands of approximate molecular weights of 45,000 and 125,000. The selective reactivity of LAM2 antibody with SCC and SQC, but not with most other tumor tissues and normal tissues, makes it a good candidate for use in clinical diagnosis and possibly serotherapy.     

Immunohistopathological characterization of spontaneous metastases in a human lung mucoepidermoid adenocarcinoma (HLMC) Xenograft

The most common clinical form of lung cancer is a disseminated disease with distant metastases; several years of cancer progression precede presentation, and this ultimately limits the efficacy of curative therapy. In this immunohistochemical study, we examined a mucinous adenocarcinoma cell line, maintained by xenogeneic transplantation, and a spontaneous metastatic variant which produces distant tumors (in liver, spleen and kidney). The aim was to investigate possible parameters which characterize the metastatic process. Histopathological comparison between the two subcutaneous transplanted tumor lines showed that both lines presented a similar cellular morphology, a different pattern of cellular growth and an increased vascularization in the metastatic line with respect to its parent. All the tumor sections expressed differential immune reactivity with monoclonal antibodies against Lewis y (MAb C14), sialyl-Lewis x (MAb SNH3) and Lewis x (MAb FH2) determinants. Neither expressed MUC 1 mucins detectable with monoclonal antibodies reactive with the mucin protein core (MAbs C595 and SM3) nor was carcinoembryonic antigen (MAb C365) expressed. Neoplastic cells were reactive with an anti-pan cytokeratin monoclonal antibody confirming their epithelial histogenesis. Our findings have been evaluated with respect to defining metastatic phenotypes in lung cancer by examination of distinct histopathological and immunological parameters.     

Stimulation of the Growth of Metastatic Clones of Mouse Colon Adenocarcinoma 26 in vitro by Platelet-derived Growth Factor

The induction of platelet aggregation by tumor cells was found to be an important determinant for the formation of metastasis of a highly metastatic variant of mouse colon adenocarcinoma 26. We found that the growth of highly metastatic clones, NL-17 and NL-33, of mouse colon 26 was well stimulated by platelet-derived growth factor (PDGF) and the stimulation was dependent on the concentration of the growth factor. The growth of weakly metastatic clones, NL-4 and NL-44, was stimulated marginally by PDGF, Other factors such as transforming growth factor β and epidermal growth factor did not stimulate the growth of metastatic clones. As the amounts of the receptor of PDGF, as determined by [¹²⁵I]PDGF binding and mRNA expression, were almost equal in NL-17 and NL-44 clones, the difference in growth potential of these clones after the treatment with PDGF could be explained by post-receptor mechanism(s). The present findings indicate that when tumor cells are arrested in a capillary through the formation of aggregates with platelets, PDGF might play an important role in the establishment of metastasis of mouse colon 26.     

Identification of a platelet-aggregating factor of murine colon adenocarcinoma 26: Mr 44,000 membrane protein as determined by monoclonal antibodies

The interaction between platelets and tumor cells plays an important role in the hematogenous spread of certain malignant cancers. We found that metastatic clones of murine colon adenocarcinoma 26 induced platelet aggregation in a membrane protein-dependent manner. Two monoclonal antibodies (mAbs) of IgG2a class were generated against a highly metastatic colon 26 clone, NL-17. These two mAbs, designated 8F11 and 20A11, showed a two-fold higher level of NL-17 binding than a low metastatic clone, NL-14, which possesses low platelet-aggregating ability. Both mAbs retarded platelet aggregation induced by NL-17. Western blot analysis showed that both mAbs recognized the same Mr 44,000 membrane protein as antigen under reducing conditions. Trypsin treatment of NL-17 diminished the ability of the cells to induce platelet-aggregation and resulted in a decrease in the reactivity of the cells to 8F11. These results suggest that the Mr 44,000 membrane protein recognized by the two mAbs is a platelet-aggregating factor of colon 26 cells.     

Purification and characterization of a human lung tumor-associated antigen.

A human lung tumor-associated antigen was purified from a saline extract of a lung adenocarcinoma. The antigen was demonstrated in extracts of lung adenocarcinoma. The antigen was demonstrated in extracts of lung tumors with the use of an absorbed antiserum by double-diffusion immunoprecipitation. The antiserum did not react with extracts of normal lung or other normal tissues, and the antigen was immunologically distinct from other tumor-associated antigens. Purification was achieved by antibody affinity chromatography and preparative polyacrylamide gel electrophoresis. Isolation procedures were monitored by immunoreactivity with absorbed monospecific antiserum. The antigen was labeled with 125I and judged homogeneous by 1) polyacrylamide gel elecrophoresis in detergent and nondetergent gels, 2) molecular sieve chromatography, 3) ion exchange chromatography, and 4) sucrose gradient sedimentation analysis. A molecular weight of 77,000 was calculated from the s20.w value of 4.24S and from the D20.w value of 5.0X10(-7) cm2/sec. Sodium dodecyl sulfate gel electrophoresis indicated a subunit molecular weight of 42,000. The Stokes radius of the antigen was 40 A and the frictional ratio was 1.42, indicating a nonspherical molecule. The purified radioiodinated antigen could be quantitatively precipitated with specific antiserum.     

Neuroendocrine differentiation antigen on human lung carcinoma and Kulchitski cells

In the normal lung, a subset of cells with a histological appearance consistent with that of Kulchitski cells are the only lung cells reacting with a monoclonal antibody (MOC-1) raised against a human small cell lung carcinoma-derived cell line. Outside the lung, a subset of normal endocrine cells (in the adrenal, thyroid, ovary, and pancreas) as well as neural cells (brain and peripheral Schwann cells) also express the antigen detected by MOC-1 (named MOC-1-related antigen). Some of these positively reacting cells are ectodermally derived, whereas others are of proven endodermal origin, indicating that the MOC-1-related antigen is not a cell lineage-specific antigen. Instead, the common expression of the antigen by cells with a neural, endocrine, or neuroendocrine function suggests that the antigen related to a neuroendocrine differentiation state of these cells. The presence of the MOC-1-related antigen on several non-lung tumors mostly paralleled its normal tissue distribution, indicating that the antigen is generally retained upon malignant transformation. In lung carcinoma, the antigen proves to be present on almost all small cell carcinomas tested. In addition, adenocarcinoma and mixed adenosquamous carcinoma could also express the antigen, whereas pure squamous cell carcinoma generally did not. This finding will be discussed in relation to a proposed "common stem cell" histogenesis of lung carcinoma.     

Expression of neural cell adhesion molecule-related sialoglycoprotein in small cell lung cancer and neuroblastoma cell lines H69 and CHP-212

Monoclonal antibodies (MAbs) 123C3 and 123A8 generated against a membrane preparation of a small cell lung carcinoma (SCLC) specimen recognize not only SCLC and bronchial carcinoids but also a significant portion of non-small cell lung carcinomas (non-SCLC) of various histological types. Together with 13 other monoclonal antibodies, which show preference for SCLC, they have been ranked as SCLC cluster 1 (SC-1) Mabs. In this study we show that SC-1 MAbs are directed against a restricted number of epitopes, and that SC-1 MAbs and a polyclonal antiserum directed against the neural cell adhesion molecule (NCAM) recognize identical glycoproteins, indicating that SC-1 antigens are closely related to or identical with NCAM. Long polysialic acid units composed of alpha-(2,8)-linked N-acetylneuraminic acid units, which in mammals are found exclusively on NCAM, were present on SC-1 antigens in SCLC. This provides further evidence that SC-1 MAbs recognize NCAM. The SC-1 antigens in the SCLC cell line H69 were present in two forms, NCAM-containing alpha-(2,8)-polysialic acid units identified by antiserum 735, the NCAM-H form, and the less sialylated NCAM-L form. The NCAM-H form consisted of diffusely migrating sialoglycoproteins with a molecular weight of 200,000-250,000, which resolved after neuraminidase treatment into two proteins with molecular weights of 140,000 and 180,000. Since the NCAM-H form is expressed in the lung tumor type with a poor prognosis, our results suggest that NCAM might be implicated in the invasive behavior of these NCAM-positive lung tumors.