Human T-cell leukemia virus type 1 associated with small cell lung cancer

A patient with small cell lung cancer (SCLC) whose serum contained high levels of soluble interleukin-2 receptors is reported. Soluble interleukin-2 receptors in the supernatant of cultured SCLC cells obtained from the patient's pleural effusion while he had malignant pleuritis, increased almost linearly from the time of cell seeding. The expression of interleukin-2 receptors (Tac) on the SCLC cells were demonstrated by an immunofluorescence study. However, other lymphocytic markers, including OKT 11, OKT 4, OKT 8, B 1, and B 4, were not found on the cells with the exception of the natural killer cell marker, NKH-1. Southern blot analysis indicated the rearrangement of the T-cell receptor of the cancer cells. Moreover, monoclonal integration of human T-cell leukemia virus type 1 (HTLV-1) provirus in DNA from the cancer cells was also demonstrated. These observations suggest that some SCLC in HTLV 1 endemic areas are associated with HTLV-1.     

Chemosensitivity testing of human lung cancer cell lines using the MTT assay

Thirty human lung cancer cell lines were tested for chemosensitivity using the semi-automated, non-clonogenic MTT assay. The tumour cell lines came from three major categories of patients: untreated small cell lung cancer (SCLC); SCLC relapsing on chemotherapy; and non-SCLC predominantly from untreated patients. From these data IC50 values were derived for each drug in each cell line. While some inter-experimental variability was observed, the rank order of chemosensitivity of each cell line within this panel was significantly correlated between experiments. These results show that tumour cell lines derived from untreated small cell lung cancer patients were the most chemosensitive for adriamycin, melphalan, vincristine and VP16 compared to the other cell types. In addition, untreated SCLC was more sensitive than non-SCLC to BCNU and cis-platin, while vincristine was the only drug to which treated SCLC was more sensitive compared to the non-SCLC lines. In contrast, no significant differences between the lung cancer types were observed for vinblastine. Thus, this panel of lung cancer cells exhibited a drug sensitivity profile paralleling that observed in clinical practice. These results suggest that this lung cancer cell line panel in combination with a relatively simple but reproducible chemosensitivity assay, such as the MTT assay, has potential for the testing of drug combinations and evaluating new anti-cancer agents in vitro.     

Experience with induced hypertension chemotherapy for lung cancer

Induced hypertension chemotherapy (IHC) using angiotensin II was applied for patients with lung cancer who had not been treated previously, and the results compared with those of preceding conventional chemotherapy as a sequential control. Twenty-nine patients with non-small cell lung cancer (non-SCLC) were treated with MTX and MMC. Response rate among evaluable cases was 23.1% (3/13) for conventional chemotherapy and 18.2% (2/11) for IHC. Twenty-eight patients with small cell lung cancer (SCLC) were treated with VCR, CPA and ACNU. Among evaluable cases, both chemotherapy groups with and without IHC showed the same response rate, 66.7% (8/12). With respect to response rate, there were no differences between conventional chemotherapy and IHC for non-SCLC or SCLC.     

Immunophenotype of small cell lung carcinoma. Expression of NKH-1 and transferrin receptor and absence of most myeloid antigens

NKH-1 is a monoclonal antibody that reacts with human natural killer (NK) cells and neural tissue. Because other monoclonal antibodies reacting with NK cells have been found on small cell lung carcinoma (SCLC), frozen tissue sections of 22 lung tumors including nine SCLC, two bronchial carcinoids, and 11 non-SCLC were tested for the presence of NKH-1 antigen by a sensitive alkaline phosphatase/anti-alkaline phosphatase technique. The labeling reactions of NKH-1 in frozen tissue sections were compared with reactions of a panel of 21 other monoclonal antibodies against NK cells, leukocyte antigens, cytokeratins, or nonlineage specific antigens. The antibody NKH-1 reacted strongly and diffusely with all of the SCLC and bronchial carcinoids but with none of the non-SCLC. NKH-1 also strongly labeled peripheral nerves in tissues adjacent to tumor. Two antibodies to cytokeratins reacted with all of the tumors and outlined tumor cells well, distinguishing them from surrounding stromal cells and leukocytes. OKT9, an antibody against transferrin receptor labeled all SCLC and eight of 11 non-SCLC but did not react with bronchial carcinoid. The antibodies Leu-M1, OKT10, Leu-7, and My4 reacted with 67%, 33%, 22%, and 11%, respectively, of the SCLC tested. The remaining 14 antibodies, including several with leukocyte specificity, labeled neither SCLC nor bronchial carcinoid. Thus, SCLC has a distinct immunophenotype (NKH-1 positive, keratin positive, and transferrin receptor positive), which may be helpful distinguishing this tumor from other tumors of lung including non-SCLC. SCLC infrequently expresses other leukocyte-associated antigens.     

Induction of intercellular adhesion molecule 1 on small cell lung carcinoma cell lines by gamma-interferon enhances spontaneous and bispecific anti-CD3 x antitumor antibody-directed lymphokine activated killer cell cytotoxicity.

The interaction between LFA-1 and its natural ligand, ICAM-1, plays an important role in leukocyte adhesion and signal transduction. LFA-1-mediated T-cell adhesion is generally activated by CD3-mediated signal in association with T-cell receptor-mediated recognition of the antigen/major histocompatibility complex on antigen-presenting cells. In the present study, we compared spontaneous or bispecific antibody (BsAb)-directed LAK cell cytotoxicity against ICAM-1+ or ICAM-1- small cell lung cancer (SCLC) cell lines. gamma-Interferon (IFN-gamma)-induced ICAM-1 expression on ICAM-1- SCLC cell lines, and susceptibility to LAK cells was increased simultaneously. Increased cytolysis of the IFN-gamma-treated SCLC was inhibited by an anti-ICAM-1 monoclonal antibody (mAb). Furthermore, LAK cell cytotoxicity directed by BsAb, which was composed of OKT3 and anti-SCLC mAb, was also increased by the IFN-gamma treatment of SCLC, and this increase was inhibited by an anti-ICAM-1 mAb but not by anti-Class I or anti-CD2 mAb. These results suggest that a prior administration of IFN-gamma would enhance the efficacy of the following specific targeting therapy utilizing BsAb and LAK cells by up-regulating the ICAM-1 expression on tumor target cells. The combinational use of IFN-gamma and anti-CD3 x anti-tumor BsAb might be a promising way of enhancing LAK cell-mediated adoptive immunotherapy in small cell lung cancer patients.     

Biological and Clinical Implication of Neuron-Specific Enolase and Creatine Kinase BB in Small Cell Lung Cancer

The specificity of neuron-specific enolase (NSE) and creatinekinase BB (CK-BB) for small cell lung cancer (SCLC) was determinedby biological and immunohistochemical procedures in lung cancertissues and cultured cell lines. Average values of extractableNSE and CK-BB of SCLC tissues were significantly higher thanthose of non-SCLC and normal lung tissues. A large amount ofNSE and CK-BB was demonstrated in SCLC cell lines. Immunohistochemical examination showed positive staining forNSE and CK-BB in most cases of SCLC and in a few cases of non-SCLC.From these data NSE and CK-BB should be considered to be highlyspecific for SCLC. In a clinical study serum values exceeding 10 ng/ml for NSEand 1.5 ng/ml for CK-BB were set as positive for the enzymes.Positive rates in SCLC were 71.4% for NSE and 65.3% for CK-BB,which were significantly higher than those in non-SCLC. Allpositive cases were in an advanced stage. Consecutive dailyNSE determinations during induction chemotherapy showed transientelevation immediately after the initiation of drug administration(tumor lysis syndrome), followed by a decline to the normalrange in responders. This phe nomenon seems to indicate tumorsensitivity to cytotoxic drugs. NSE positive non-SCLC was assensitive to cytotoxic drugs as SCLC. These findings indicatethat lung cancer with elevated serum NSE and CK-BB levels atdiagnosis should be strongly suspected of being SCLC in theadvanced stage.     

Coexpression of ganglioside antigen Fuc-GM1, neural-cell adhesion molecule, carcinoembryonic antigen, and carbohydrate tumor-associated antigen CA 50 in lung cancer.

With the aid of specific monoclonal antibodies, tumor tissues from 68 patients with lung cancer were examined for their expression of two small cell lung carcinoma (SCLC) antigens, Fuc-GM1 (fucosyl GM1; IV2FucII3NeuAc GgOse4) and neural-cell adhesion molecule (NCAM), and two broader tumor antigens, carcinoembryonic antigen (CEA) and carbohydrate cancer-associated antigen CA 50. Expression of Fuc-GM1 was seen in 75% and NCAM in 78% of the SCLC specimens, but also in 12 and 20% of non-SCLC. Either or both of these antigens were expressed in more than 90% of SCLC and in 25% of non-SCLC. CEA was found in more than 80% of SCLC and non-SCLC. Expression of CA 50 was seen in 65-68% of non-SCLC and SCLC, showing preference for SCLC and lung adenocarcinoma. In SCLC, cellular expression of Fuc-GM1 was generally seen together with NCAM and CA 50, but rarely with CEA. There was considerable inter- and intratumor heterogeneity in the expression of all four antigens. The results suggest that CEA is the antigen of choice for the detection of lung cancer regardless of histotype. In combined analysis of CEA, CA 50, Fuc-GM1 and NCAM, two patterns of antigen expression were recognized that appear to discriminate between SCLC and non-SCLC tumors, respectively. A considerable fraction of SCLC and non-SCLC tumors, however, exhibited similar patterns of antigen expression. The biological and clinical significance of these observations remains to be investigated.     

Determination of various tumor markers, with special reference to neuron-specific enolase in lung cancer

Serum neuron-specific enolase (NSE) was measured in 23 patients with small cell lung cancer (SCLC) and 184 patients with non-small cell lung cancer (non-SCLC), both of which were untreated. Increased levels of serum NSE were observed in 82.6% (19/23) of SCLC, whereas 9.8% (18/184) of non-SCLC had positive results showing an overall positive rate of 17.9% (37/207) in lung cancer cases. In addition, the elevation of serum NSE levels in non-SCLC patients seemed to suggest poor prognosis. Elevated serum NSE levels returned to normal with either surgical resection of the tumor or response to chemotherapy, after which serum NSE levels were again raised to levels higher than the previous ones in cases of relapse or progression. The evaluation of serum NSE may be a useful marker for both diagnosis and monitoring of responsiveness to therapy as well as for recognition of relapse and progression in SCLC. Identification of NSE as assessed by immunohistochemical procedure employing the ABC method on formalin-fixed paraffin-embedded tissue sections in lung cancer cases of each histological type, showed that some materials from non-SCLC cases were positively stained despite the presence of normal serum NSE levels, and did not always parallel the serum levels. Among other various tumor markers determined, serum CA 19-9 had a relatively high positive rate of 38.2% (42/110) in adenocarcinoma of the lung.     

Loss of expression of death-inducing signaling complex (DISC) components in lung cancer cell lines and the influence of MYC amplification

We have previously reported that the key apoptosis related gene caspase 8 (CASP8) is frequently silenced in small cell lung cancer (SCLC) tumors and cell lines usually, but not always, by aberrant promoter methylation. Because CASP8 is a key component of the death-inducing signaling complex (DISC) when specific death receptors (including DR4, DR5, FAS) are activated by their specific ligands (TRAIL/FASL), we examined expression of the components of the DISC complex in lung cancer cell lines. MYC family members are frequently amplified (MYC+ve) in SCLC, and MYC is a potent inducer of apoptosis. We examined 34 SCLC lines (12 of which were MYC+ve) and 22 NSCLC lines. CASP8 gene expression was frequently lost (79%) at message and protein levels in SCLC but not in non-SCLC (NSCLC). MYC amplification was present in 45% of SCLC cell lines, which had lost CASP8 expression, but not in any of the CASP8 positive lines. The frequency of CASP8 loss was significantly higher in MYC+ve SCLC compared to MYC-ve SCLC or in NSCLC. Analyses of other DISC components showed significantly higher rates of loss of expression of CASP10, DR5, FAS and FASL in SCLC compared to NSCLC. The loss of expression of proapoptotic DISC components was significantly higher in MYC+ve SCLC cell lines and these lines were completely resistant to TRAIL. Expression of CASP10 (a caspase closely related to CASP8) was frequently absent at the protein level in both SCLC and NSCLC lines. Expression of c-FLIP (proteolytically inactive homolog of CASP8) was inversely related to expression of CASP8. Our major conclusions are: (a) The death receptor pathway is differently inactivated at multiple levels in lung cancer cell lines; and (b) MYC amplification in SCLC is associated with inactivation of most components of the DISC complex, with resistance to TRAIL and with expression of c-FLIP. These findings may have considerable clinical and therapeutic implications.     

Oral administration of OK-432 (picibanil). (8th report) clinical application: its immunomodulatory effects on the patients with gastrointestinal cancers

A streptococcal preparation, OK-432 at a dose of 5 KE was orally administered to the patients with gastric or colorectal cancer for 7 approximately 14 days before operation, and its immunomodulatory effects on peripheral blood lymphocytes (PBL), regional lymph node lymphocytes (RLNL) and tumor infiltrating lymphocytes (TIL) were assessed. OK-432 treated group included 5 gastric and 6 colorectal cancers, and control group included 6 gastric and 8 colorectal cancers. After oral administration of OK-432 the proportion of Leu 7+ and Leu 11+ cells in PBL increased, and NK cell activity of PBL also was augmented. The proportion of OKT8+ cells increased in PBL and those of OKT3+ cells and OKT8+ cells decreased in RLNL after oral administration of OK-432. The responsiveness of TIL to autologous tumor extracts in the presence of interleukin-2 was enhanced in oral OK-432 group. These results indicate that oral OK-432 affects on NK and T cells and augments the antitumor immunity of the patients with gastrointestinal cancer.     

Expression of placental growth factor gene in lung cancer.

Differences in the gene expression profiles in small cell lung cancers (SCLC) and non-small cell lung cancers (NSCLC) may explain their different clinical characteristics. The aims of this study were (1) to identify genes differentially expressed in SCLC and NSCLC using mRNA differential display, and (2) to determine the clinical relevance of such genes in lung cancer. RNA differential display using three SCLC and six non-SCLC cell lines was used to identify a differentially expressed gene. Differential expression of the gene was confirmed in additional lung cancer cell lines using RT-PCR. Immunohistochemical staining for the gene product was performed on paraffin-embedded tissue from lung cancer patients. We examined the relationship between the expression of the gene and clinical parameters, including disease stage, response to treatment and survival time. The placental growth factor (PGF) gene was identified as preferentially expressed in SCLC compared with NSCLC cell lines using mRNA differential display. Further analysis of 45 lung cancer cell lines using RT-PCR showed that the placental growth factor (PGF) gene was expressed in nine of 13 SCLC cell lines (69%) and five of 32 NSCLC cell lines (15.6%) (p < 0.001, Fisher's exact test). Immunohistochemistry using anti-PGF antibody on the paraffin blocks from lung cancer patients showed that PGF expression was significantly higher in SCLC than NSCLC tissue sections (32 vs. 5.6%, p = 0.041, Fisher's exact test). Expression of PGF protein did not correlate with disease stage, response to treatment or survival time in SCLC patients. The present study suggests there is higher expression of PGF in SCLC compared to NSCLC. It may be that higher expression of the angiogenic factor PGF contributes to differences between the progression of SCLC and NSCLC, especially in regard to the nature of SCLC metastasis.     

The Release of Chromogranin A and B Like Activity from Human Lung Cancer Cell Lines: A Potential Marker for a Subset of Small Cell Lung Cancer

Human small cell lung cancer (SCLC) is in vivo and in vitro characterized by a heterogeneous spectrum of neuroendocrine markers. the non-SCLC group is deprived of these markers, or expresses them in low quantities. in this paper we report on the release to the culture medium of neuroendocrine associated proteins, chromogranin A and B like activity (CABLA). the culture medium from three out of five SCLC cell lines and in onehive non-SCLC cell line contained significant levels of CABLA. Normal diploid foreskin fibroblasts and a histiocytic lymphoma cell line were deprived of CABLA production. the presence of CABLA in both SCLC and non-SCLC further stress their common histogenetic origin. the CABLA values were partly unrelated to other neuroendocrine markers. Determinations of CABLA could thus be a potential and valuable marker for a subset of SCLC.     

Intermediate filament and cross-linked envelope expression in human lung tumor cell lines

Human lung tumor cell lines established from the major histological types of lung cancer were examined by immunofluorescent staining techniques for their patterns of intermediate filament (keratin, vimentin, and neurofilament triplet protein) expression. All cell lines examined, both small cell lung carcinoma (SCLC) and non-SCLC (squamous cell carcinoma, adenocarcinoma, large cell carcinoma, and mesothelioma) contained keratin, consistent with their epithelial derivation. These lung carcinoma cell lines also expressed vimentin, the characteristic intermediate filament of mesenchymal cells in vivo. In light of the proposed neuroectodermal origin of SCLC, cell lines were also studied for neurofilament expression. Two of four SCLC tumor cell lines, as well as non-SCLC cell lines, showed no reactivity with antibodies to neurofilament triplet protein. Two of the SCLC cell lines stained weakly with anti-neurofilament antibody. Examination of specific keratin patterns in human lung tumor cell lines by selective immunoprecipitation with keratin antiserum and sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated that small-sized keratin proteins (Mr 44,000 to 52,000) were present in cell lines derived from SCLC and non-SCLC types of lung cancer. Tumor cell lines exhibiting squamous differentiation by light microscopic criteria (i.e., intracellular keratin, intercellular bridging, "pearl" formation, and/or individual cell keratinization) also displayed a preponderance of intermediate-sized keratins (Mr 57,000 and 59,000) and exhibited another feature of terminal keratinocyte differentiation (cross-linked envelope formation). Mesothelioma cell lines had varying keratin profiles. The presence of keratin proteins in all SCLC cell lines examined argues against a neuroectodermal origin for these tumors and is consistent with the notion that these tumors arise from a common bronchial "stem cell," similar to that from which other types of bronchogenic carcinomas arise.     

Transitions between lung cancer phenotypes--implications for tumor progression.

Progression from a treatment-sensitive to a treatment-resistant tumor state is a virtually universal phenomenon in patients with small-cell lung carcinoma (SCLC). In such individuals, this tumor progression may involve transitions from a SCLC to a non-SCLC lung cancer phenotype. We are investigating the cell and molecular biology aspects of these transitions and have derived a cell culture model of one such change, oncogene-induced transition of SCLC to the large-cell undifferentiated lung cancer phenotype. Here we discuss the potential implication of this model for understanding the cell lineage and molecular events regulating normal bronchial epithelial cell differentiation and their relationships to the histogenesis and behavior of lung cancers.     

Phenotypic heterogeneity studied by immunohistochemistry and aneuploidy in non-small cell lung cancers

Non-small cell lung cancers (non-SCLC) differ from small cell lung cancers (SCLC) by many clinical features and prognosis. However, recent studies suggest that lung cancer heterogeneity frequently leads to the association of SCLC and non-SCLC in the same tumor. This phenotypic heterogeneity can be analyzed by immunohistochemistry using monoclonal antibodies (Mab) raised against differentiation related antigens. It may have clinical relevance inasmuch as the diversification of malignant cells is a well-known factor of tumor progression and may be due to chromosomal instability because inappropriate gene expression leads to the formation of antigens unrelated to cell lineage. Chromosomal instability in cancer leads to aneuploidy detectable by cell DNA content analysis. In a prospective study, we analyzed, in parallel, the expression of neuroendocrine related antigens by immunohistochemistry and the cell DNA content in frozen specimens from 40 patients who underwent complete surgical resection of primary non-SCLC in an attempt (a) to characterize the phenotypic heterogeneity and (b) to determine whether this heterogeneity is correlated with aneuploidy and clinical staging. Three Mabs were used in association as a marker of neuroendocrine antigen expression (S-L 11.14, MOC-1, and NE-25); reactivity of these Mabs in 9 SCLC and 3 lung carcinoid tissue sections was used as positive control. All SCLC and 2 of 3 lung carcinoids tested were homogeneously positive with Mabs S-L 11.14, MOC-1, and NE-25; 13 of 40 non-SCLC were homogeneously positive and 11 additional specimens focally positive with Mabs S-L 11.14, MOC-1, and NE-25. The frequency of this abnormal phenotype was significantly higher in poorly differentiated squamous cell carcinomas (chi 2 10.08; P less than 0.005), in clinical stage III non-SCLC (chi 2 5.93; P less than 0.02), and in tumors involving mediastinal lymph nodes (chi 2 5; P less than 0.03). The percentage of cells in the modal DNA of G0-G1 phase was significantly lower in non-SCLC homogeneously positive with Mabs S-L 11.14, MOC-1, and NE-25 [27.4 +/- 10.3% (SD)] in comparison with non-SCLC negative with these same Mabs [56.8 +/- 21.3%; P less than 0.01, Mann-Whitney U test]. We conclude that (a) mixed SCLC-non-SCLC differentiation is frequent and can be assessed by immunohistochemistry, (b) neuroendocrine differentiation in non-SCLC is mainly observed in poorly differentiated tumors and in advanced clinical stages, and that (c) this heterotopic phenotype is correlated with aneuploidy and has clinical implications.     

Characterization and purification of an immunosuppressive factor produced by a small cell lung cancer cell line.

The present study was undertaken to determine whether small cell lung cancer (SCLC) cell lines produce immunosuppressive factors and, if they do, to characterize the factors. The supernatants of SCLC cell lines, H69 and N857, inhibited not only the blastogenic response of human peripheral blood lymphocytes (PBL) to phytohemagglutinin or concanavalin A, but also the cytotoxic activity of lymphokine-activated killer cells. Neither was inhibited by supernatants from non-SCLC cell lines PC9, QG56, and A549. The immunosuppressive activity of H69 supernatant was stable upon heating to 56 degrees C for 60 min, but labile when heated to 70 degrees C for 10 min. The activity was abolished after dialysis at pH 2.0 or pH 11.0, but not at pH 4.5 or pH 9.0. Digestion with trypsin or proteinase eliminated the immunosuppressive activity, whereas treatment with neuraminidase, mixed glycosidase, DNase or RNase had no effect, suggesting that the immunosuppressive activity in H69 supernatant is due to a protein factor. This H69-derived immunosuppressive factor was isolated by ion exchange chromatography using a gradient of 0.04 to 0.08 M NaCl solution. Gel filtration and sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed the factor to have molecular weights of 98 kD and 102 kD, respectively. These results suggest that SCLC cells produce a potent immunosuppressive factor which may account for the immune deficiency in SCLC patients.     

Electron microscopy in small cell lung carcinomas: clinical correlation

In order to investigate the relationship of subcellular differentiation of small cell lung carcinomas (SCLC) and clinical response, we reviewed the electron microscopic (EM) features of tumor biopsy specimens from 33 patients with SCLC diagnosed by light microscopy (LM). These tumors were divided by EM into four groups according to the ultrastructural features. Group I (13 patients) had tumors with only neurosecretory granules on EM. Group II (11 patients) had tumors with neurosecretory granules and other subcellular features of non-SCLC. Group III (five patients) had tumors that lacked neurosecretory granules but contained subcellular features of non-SCLC. Group IV (four patients) had tumors that lacked all of these features. The complete and partial response rate to systemic chemotherapy with or without radiation therapy was 88% in the total population studied. The response rates were not statistically different in any of the four groups based on EM findings. The results of this study suggest that the LM diagnosis of SCLC alone adequately identifies lung cancer patients with a high response rate to systemic therapy.     

肿瘤浸润淋巴细胞在小细胞肺癌中的研究进展

小细胞肺癌（small cell lung cancer,SCLC）是一种预后极差的肺癌亚型,虽前期对放化疗敏感,但多数患者会在短期内因肿瘤复发、转移而死亡。探寻有效的、能指导预后及预测疗效的生物标志,以达到SCLC患者治疗的个体化和最优化是肿瘤研究的热门领域之一。目前有大量文献报道肿瘤浸润淋巴细胞（tumor-infiltrating lymphocytes,TILs）与较多类型癌种的（包括非小细胞肺癌、乳腺癌、结直肠癌、黑色素瘤等）预后及预测免疫治疗疗效的关系,本文将肿瘤浸润淋巴细胞在SCLC预后及疗效预测方面的作用作一综述。