Identification of conditioned medium proteins from human cancer cells adapted to serum-free conditions

Recent studies have shown that human cancer cell lines can be adapted to grow in serum-free, unsupplemented RPMI-1640 (RO) medium. We have developed similar techniques to rapidly identify proteins of interest in serum-free conditioned medium (CM) of human lung cancer cell lines. Classic and variant small cell lung cancer (SCLC) lines were adapted to growth in RO medium. CM from each line was concentrated and fractionated on an anion-exchange column of a fast protein liquid chromatography system. Concentrates of each fraction were loaded onto lanes of minigels of an automated electrophoresis system. Analysis of the chromatograms reveals peaks seen only in CM of the classic SCLC lines. Electrophoretic analysis of the fractions containing these peaks reveal protein bands distinguishing between the subtypes of human SCLC. One protein was purified to homogeneity with subsequent reversed-phase chromatography and identified by protein microsequencing as histone H2B. These automated techniques have general use in the rapid identification of CM proteins associated with the differentiation or progression of the many types of neoplastic cells which can be adapted to growth in RO medium.     

Interaction of recombinant human tumor necrosis factor and etoposide in human lung cancer cell lines

Studies have suggested that recombinant tumor necrosis factor-alpha (TNF-alpha) may potentiate the killing of murine tumor cells by drugs targeted at DNA topoisomerase II. We have examined the combined cytotoxic effects of the topoisomerase-targeted drug etoposide and TNF in small cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC) cell lines using clonogenic assays and a novel flow cytometry technique relying on differential uptake of fluorescein diacetate (FDA) and propidium iodide (PI) by viable and nonviable cells. Good correlation of IC50 determinations for etoposide were noted between clonogenic assays and the FDA/PI technique for both classic and variant SCLC cell lines. The effects of etoposide on the classic SCLC line H209 were potentiated by TNF with a decrease in the IC50 from 3.3 microM to 1.0 microM as determined by FDA/PI. Tumor necrosis factor alone had little effect on the growth or cloning efficiency of H209 cells. Tumor necrosis factor alone stimulated the growth and cloning of variant SCLC line N417, but the cytotoxicity of etoposide was not potentiated by TNF in N417 cells. Tumor necrosis factor alone inhibited the growth and cloning of the NSCLC line H125 but exerted a marked protective effect against higher concentrations of etoposide. It appears that the interaction of TNF with etoposide varies between cell lines and between subclasses of human lung cancer.     

Different phenotypic variants of the mouse B cell tumor A20/2J are selected by antigen- and mitogen-triggered cytotoxicity of L3T4-positive, I-A-restricted T cell clones

The L3T4+, Lyt-2-, cloned BALB/c T cell lines 5.9.24 and 5.8.6 are cytotoxic for the BALB/c B cell tumor line A20/2J. The T cell cytotoxicity against A20/2J cells could be triggered either by the specific antigen ovalbumin (OVA), which is recognized by the T cell clones in association with I-Ad determinants, or by the T cell mitogens Con A and rabbit anti-mouse brain (RaMBr) antiserum. Repeated exposure of A20/2J cells to 5.9.24 and 5.8.6 T cell cytotoxicity selected variant cell lines that had developed resistance to cytotoxicity. The variant lines could be classified into four different variant phenotypes of which three were stably maintained in vitro. The type of variant obtained appeared to be related to the nature of the ligand used to trigger T cell cytotoxicity during selection. Cytotoxicity triggered by the antigen OVA generated type 1 variants that expressed abnormally low levels of I-Ad determinants at the cell surface. Type 1 variants were resistant to OVA-triggered 5.9.24 T cell cytotoxicity, but were fully susceptible to cytotoxicity triggered by Con A or RaMBr antiserum. RaMBr-triggered cytotoxicity generated two unique types of variant cell lines: type 3 variants that were deficient in cell surface Fc receptors and resistant to 5.9.24 cytotoxicity only when triggered by RaMBr antiserum, and type 4 variants that were resistant to cytotoxicity triggered by all three ligands. One type 4 variant, the IC-1 cell line, appeared to be resistant to soluble cytotoxic factors released by 5.9.24 T cells after activation by antigen. All of these variant lines retained sensitivity to cytotoxicity by classic Lyt-2+ cytotoxic T lymphocytes (CTL), a finding that indicates that L3T4a+ T cells and Lyt-2+ CTL use different molecules to attack their target cells. The variant phenotypes were inherited by clones derived from the original cell lines. Because the variants were generated without mutagenesis, they are thought to have been derived by the immunoselection of pre-existing variant cells that arose spontaneously in the parental A20/2J cell line. It is postulated that inheritable variation of A20/2J cells may represent changes that normally occur during B cell differentiation in response to T cell signals. The variant A20/2J cell lines described here provide material for the investigation of B cell surface structures that may regulate T-B cell interactions.     

MicroRNA expression and clinical outcome of small cell lung cancer

The role of microRNAs in small-cell lung carcinoma (SCLC) is largely unknown. miR-34a is known as a p53 regulated tumor suppressor microRNA in many cancer types. However, its therapeutic implication has never been studied in SCLC, a cancer type with frequent dysfunction of p53. We investigated the expression of a panel of 7 microRNAs (miR-21, miR-29b, miR-34a/b/c, miR-155, and let-7a) in 31 SCLC tumors, 14 SCLC cell lines, and 26 NSCLC cell lines. We observed significantly lower miR-21, miR-29b, and miR-34a expression in SCLC cell lines than in NSCLC cell lines. The expression of the 7 microRNAs was unrelated to SCLC patients' clinical characteristics and was neither prognostic in term of overall survival or progression-free survival nor predictive of treatment response. Overexpression or downregulation of miR-34a did not influence SCLC cell viability. The expression of these 7 microRNAs also did not predict in vitro sensitivity to cisplatin or etoposide in SCLC cell lines. Overexpression or downregulation of miR-34a did not influence sensitivity to cisplatin or etoposide in SCLC cell lines. In contrast to downregulation of the miR-34a target genes cMET and Axl by overexpression of miR-34a in NSCLC cell lines, the intrinsic expression of cMET and Axl was low in SCLC cell lines and was not influenced by overexpression of miR-34a. Our results suggest that the expression of the 7 selected microRNAs are not prognostic in SCLC patients, and miR-34a is unrelated to the malignant behavior of SCLC cells and is unlikely to be a therapeutic target.     

Neuroendocrine-specific protein (NSP)-reticulons as independent markers for non-small cell lung cancer with neuroendocrine differentiation

Neuroendocrine-specific protein (NSP)-reticulons have recently been discovered and were shown to exhibit a restricted, neuroendocrine/neural-specific expression pattern. These protein aggregates are anchored to the membranes of the endoplasmic reticulum and occur in small cell lung cancer (SCLC), but not in typical non-SCLC. In the current study we have examined the occurrence of NSP-reticulons in non-SCLC cell lines known to express neuroendocrine features (non-SCLC-NE). NSP-reticulon expression was observed in all three non-SCLC-NE cell lines studied, albeit with variable intensity and in varying numbers of cells. Western blot analysis confirmed the presence of NSP-reticulon expression in these non-SCLC-NE cell lines, and showed that they were predominantly of the NSP-A type. When compared to conventional neuroendocrine markers, NSP-reticulons revealed a distinct staining profile, showing only partial overlap with the other markers. The non-SCLC-NE cell lines combined these neuroendocrine characteristics with some features of non-SCLC. We conclude that NSP-reticulon expression is restricted to lung carcinoma cells with a neuroendocrine phenotype and predict that these constituents may become clinically relevant markers for the detection of neuroendocrine differentiation in solid tumours. Accepted: 27 March 1997     

Proteomic analysis of anaplastic lymphoma cell lines: identification of potential tumour markers

Anaplastic large-cell lymphomas (ALCL) are high grade lymphomas of T or null phenotype often associated with the t(2;5) translocation leading to the expression of a chimeric protein consisting of the N-terminal portion of nucleophosmin (NPM) and the intracellular domain of the anaplastic lymphoma kinase (ALK). Although ALCL are recognized as distinct clinical, biological and cytogenetic entities, heterogeneities persist in this group of tumours, which exhibit a broad spectrum of morphological features. Particularly, the common type tumour consisting in large cells contrast with the small cell variant that is sometimes associated with a leukemic phase. The ALK-negative ALCL is often associated with a poor prognosis. Here, we investigated the proteome of these subtypes of tumours using patient-derived cell lines. We compared the proteome of the cytosolic fraction of NPM-ALK-positive versus NPM-ALK-negative cells on one hand, and the proteome of common cell type versus small cell variant on the other hand. The identification of a set of proteins differentially expressed in the subtypes of ALCL points to new diagnosis/prognosis markers. This study also provides interesting information on the molecular mechanisms responsible for the different subtypes of ALCL.     

Insulin-like growth factor binding protein expression in human small cell lung cancer cell lines

Insulin-like growth factor binding proteins (IGF-BP) are secreted by several human small cell lung cancer cell lines (SCLC). In order to identify the IGF-BPs from SCLC cell lines the RNA from 10 different SCLC cell lines was analyzed by Northern blot analysis with the probes for three different IGF-BPs, IGFBP-1, IGFBP-2, and IGFBP-3. No hybridization signal could be detected with the probes encoding for IGFBP-1 and IGFBP-3. The hybridization with different IGFBP-2-specific oligodeoxynucleotide probes and with the corresponding full-length cDNA showed that all SCLC cell lines which secreted IGF-BPs express IGFBP-2.     

Differences in expression of pro-caspases in small cell and non-small cell lung carcinoma.

Expression of several molecular determinants of apoptosis was analyzed in 10 untreated small cell (SCLC) and 6 untreated non-small cell (NSCLC) lung carcinoma cell lines. Although SCLC lines were more prone to spontaneous apoptosis compared with NSCLC lines, the former showed higher Bcl-2 expression and a higher Bcl-2/Bax ratio. In order to understand this apparent contradiction, the expression of pro-caspases as well as calpain was analyzed in these cell lines at the protein and mRNA levels. No differences in protein level of pro-caspases-2, -3, -7, and -9 and of calpain were detected between the SCLC and the NSCLC lines, but a striking difference in pro-caspase-8 expression was noted. All 6 NSCLC, but only 2 of the 10 SCLC lines, expressed pro-caspase-8 protein. Further experiments using the RNase protection assay indicated that the lack of pro-caspase-8 expression at the mRNA level was characteristic for SCLC. Using the same experimental approach, we found that SCLC cell lines in addition to pro-caspase-8 were deficient in mRNA expression of pro-caspases-1, -4, and -10, suggesting a different caspase-activating cascade in SCLC compared with NSCLC. This first systematic characterization of pro-caspase expression in lung cancer surprisingly showed that SCLC, which are more prone to undergo spontaneous apoptosis, are deficient in several pro-caspases and have a high Bcl-2/Bax ratio. Thus, the propensity of SCLC cells to undergo apoptosis cannot be explained only by the expression of factors involved in regulation or execution of apoptosis.     

SCLC-J1, a novel small cell lung cancer cell line

Small cell lung cancer (SCLC) is a type of high-grade neuroendocrine carcinoma. It initially responds to chemotherapy but rapidly becomes chemoresistant and it is highly proliferative. The prognosis in SCLC is poor. We have established a novel SCLC cell line, SCLC-J1, from a malignant pleural effusion in a patient with advanced SCLC. SCLC-J1 cells express ganglioside GD2, CD276, and Delta-like protein 3. RB1 is lost.These features of the new SCLC cell line may be useful in understanding the cellular and molecular biology of SCLC and in designing better treatment.     

Oncogene amplification and chromosomal abnormalities in small cell lung cancer

Twelve cell lines isolated from patients with small cell lung cancer have been studied for amplification of the three characterised members of the myc proto-oncogene family (c-myc, N-myc, and L-myc) and for abnormalities of chromosome 3. Ten of these lines were being studied for the first time. Ten of the 12 small cell lung cancer cell lines had amplification of one member of the myc proto-oncogene family. Amplification of c-myc was observed in only one small cell lung line--a "morphological variant". One "classic" small cell lung cancer line expressed c-myc but had no obvious amplification of the gene. N-myc and L-myc were more commonly amplified than c-myc. Chromosomal abnormalities (mainly deletions) in chromosome 3 were observed in all small cell lung carcinoma cell lines examined. When the small cell lung carcinoma lines were grouped according to "classic" or "variant" characteristics, it was found that the "classics" had deletions of the short arm of chromosome 3, whereas the "biochemical variants" had deletions of the long arm of chromosome 3. The extent of the deletions varied between cell lines. For the deletion in the short arm of chromosome 3 the minimum common region of overlap was assigned to bands 3p23-3p24.     

Establishment of cell lines from serial samples from two patients with lung tumours before and after chemotherapy

Six cell lines were derived from pleural effusions of two lung cancer patients and established in vitro in our laboratory. Cell line AE1 was obtained from a small cell lung cancer (SCLC) before the patient had received any chemotherapy; the other lines (AE2 and AE3) were established from tumour recurrences in the same patient after therapy. Cell lines DG1 and DG2 were derived from specimens of an untreated non-small cell lung cancer (NSCLC), while cell line DG3 originated from pleural effusions recurring in the same patient after therapy. The results of the present study show that: (a) the SCLC lines AE1, AE2 and AE3 are heterogeneous in their biological characteristics and in their chemosensitivity patterns. In particular lines AE2 and AE3 are less responsive to cis-Platinum (DDP) and Adriamycin (ADM) than line AE1, so that they may reflect resistant subpopulations existing within the original tumour, selected following therapy with these drugs. In contrast, however, line AE1 proved more resistant to Vepesid (VP16) than lines AE2 and AE3. (b) The three NSCLC lines are similar in various biological features as well as in their chemosensitivity to DDP and Vinblastine (VBL).Abbreviations NSCLC Non Small Cell Lung Cancer - SCLC Small Cell Lung Cancer Recipient of a Fellowship of the Italian Association for Cancer Research     

Columnar cell variant of papillary thyroid carcinoma. Report of a case with cytologic findings

BACKGROUND: The columnar and tall cell variants of papillary thyroid carcinoma (PTC) are uncommon variants and have generally been regarded as more aggressive forms in comparison to the more common classic papillary and follicular subtypes. Cytologic diagnosis of these rare variants is elusive since the characteristic nuclear features of the usual papillary thyroid carcinoma are very often absent or inconspicuous. We present a case of the columnar cell variant of PTC in a young woman that demonstrates the diagnostic challenge. CASE: A 24-year-old woman presented with a solitary, 3-cm mass in the left aspect of the thyroid. The aspirate consisted of a moderately cellular sampling of sheets, papillary clusters and microfollicles of cells with oval nuclei and uniform, finely granular chromatin. These cells were arranged in a peudostratified manner around well-defined fibrovascular cores. There were no intranuclear inclusions or well-defined nuclear grooves in the cells of the aspirate. There was also absence of colloid despite the presence of well-formed follicles. The resected thyroid revealed a columnar cell variant of PTC. CONCLUSION: The cytologic features of columnar cell-type PTC are at variance with those of classic PTC and are elusive in fine needle aspiration cytology. It is the lack of classic cytologic features of PTC that is distinctly apparent, yet it is the monomorphism of cells in the aspirate, their papillary configuration and their pseudostratification in well-formed fibrovascular cores that are the keys to the diagnosis. Immunohistochemical staining to rule out other thyroid neoplasms can be performed to aid in the diagnosis.     

Lung carcinoid cell lines have bombesin-like peptides and EGF receptors

The biochemical properties of lung cancer cell lines were investigated. Bombesin-like peptides were present in three small cell lung cancer (SCLC) cell lines examined and three of four lung carcinoids but not in five non-small cell lung cancer (NSCLC) cell lines. Therefore SCLC and some lung carcinoids, but not NSCLC, are enriched in neuroendocrine properties. In contrast, 125I-EGF bound with high affinity to all five NSCLC cell lines and three of four lung carcinoids but not to the three SCLC cell lines examined. For lung carcinoid cell line NCI-H727, 125I-EGF bound with high affinity (Kd = 6 nM) to a single class of sites (Bmax = 110,000/cell). The 125I-EGF bound was rapidly internalized at 37 degrees C but not 4 degrees C. Using Western blot techniques and antiphosphotyrosine antibodies, EGF induced phosphorylation of a major 170 Kd protein. Using immunoprecipitation techniques and anti-EGF receptor antibodies a major 170 Kd protein was labeled. These data indicate that biologically active EGF receptors are present on NSCLC and lung carcinoid cell lines.     

Differential expression of the LAMB3 and LAMC2 genes between small cell and non-small cell lung carcinomas

To identify genes differentially expressed between small cell lung carcinoma (SCLC) cells and non-SCLC cells, mRNA differential display was applied to 3 SCLC cell lines and 6 non-SCLC cell lines. The LAMB3 gene was identified as being expressed only in non-SCLC cells and not in SCLC cells. The LAMB3 gene encodes the laminin beta3 chain, which is a unique component of laminin-5. Laminin-5 is a heterotrimer protein consisting of the alpha3, beta3, and gamma2 chains, and another unique component of laminin-5 is the gamma2 chain encoded by the LAMC2 gene. RT-PCR analysis of the LAMB3 and LAMC2 genes in 45 lung cancer cell lines revealed that both the LAMB3 and LAMC2 genes were co-expressed in 21 of 32 non-SCLC cell lines (66%) but only in one of 13 SCLC cell lines (8%). Coexpression of the LAMB3 and LAMC2 genes was also observed in all 4 cases of primary non-SCLC cells examined but not in the corresponding non-cancerous lung cells. Since alpha6beta4 integrin, the specific laminin-5 binding receptor, is known to be expressed only in non-SCLC cells and not in SCLC cells, it was indicated that laminin-5 is a critical microenvironmental factor for the growth of non-SCLC cells but not of SCLC cells. The differences in the expression of integrins and laminins would be critical factors to distinguish SCLC and non-SCLC cells, and such differences might be associated with the unique biological properties of SCLC cells, including metastatic potential and drug sensitivity.     

Small cell lung cancer,an epithelial to mesenchymal transition (EMT)-like cancer: significance of inactive Notch signaling and expression of achaete-scute complex homologue 1

Small cell lung cancer (SCLC) is one of the most malignant neoplasms in common human cancers. The tumor is composed of small immature-looking cells with a round or fusiform shape, which possesses weak adhesion features among them, suggesting that SCLC shows the morphological characteristics of epithelial to mesenchymal transition (EMT). SCLC is characterized by high metastatic and recurrent rates, sensitivity to the initial chemotherapy, and easy acquirement of chemoresistance afterwards. These characters may be related to the EMT phenotype of SCLC. Notch signaling is an important signaling pathway, and could have roles in regulating neuroendocrine differentiation, proliferation, cell adhesion, EMT, and chemoresistance. Notch1 is usually absent in SCLC in vivo, but could appear after chemotherapy. Notch1 can enhance cell adhesion by induction of E-cadherin in SCLC, which indicates mesenchymal to epithelial transition. On the other hand, achaete-scute complex homologue 1 (ASCL1), negatively regulated by Notch signaling, is a lineage-specific gene of SCLC, and functions to promote neuroendocrine differentiation as well as EMT. ASCL1-transfected adenocarcinoma cell lines induced neuroendocrine phenotypes and lost epithelial cell features. SCLC is characterized by neuroendocrine differentiation and EMT-like features, which could be produced by inactive Notch signaling and ASCL1 expression. In addition, chemical and radiation treatments can activate Notch signaling, which suppress neuroendocrine differentiation and induces chemoradioresistance, accompanied by secession from EMT. Thus, the status of Notch signaling and ASCL1 expression may determine the cell behaviors of SCLC partly through modifying EMT phenotypes.     

Small cell lung cancer: Circulating tumor cells of extended stage patients express a mesenchymal-epithelial transition phenotype

Small cell lung cancer (SCLC) is distinguished by aggressive growth, early dissemination and a poor prognosis at advanced stage. The remarkably high count of circulating tumor cells (CTCs) of SCLC allowed for the establishment of permanent CTC cultures at our institution for the first time. CTCs are assumed to have characteristics of cancer stem cells (CSCs) and an epithelial-mesenchymal transition (EMT) phenotype, but extravasation of tumors at distal sites is marked by epithelial features. Two SCLC CTC cell lines, namely BHGc7 and BHGc10, as well as SCLC cell lines derived from primary tumors and metastases were analyzed for the expression of pluripotent stem cell markers and growth factors. Expression of E-cadherin and β-Catenin were determined by flow cytometry. Stem cell-associated markers SOX17, α-fetoprotein, OCT-3/4, KDR, Otx2, GATA-4, Nanog, HCG, TP63 and Goosecoid were not expressed in the 2 CTC lines. In contrast, high expression was found for HNF-3β/FOXA2, SOX2, PDX-1/IPF1 and E-cadherin. E-cadherin expression was restricted to the 2 CTCs and 2 cell lines derived from pleural effusion (SCLC26A) and bone metastases (NCI-H526), respectively. Thus, these SCLC CTCs established from extended disease SCLC patients lack expression of stem cell markers which suppress the epithelial phenotype. Instead they express high levels of E-cadherin consistent with a mesenchymal-epithelial transition (MET or EMrT) and form large tumorospheres possibly in response to the selection pressure of first-line chemotherapy. HNF-3β/FOXA2 and PDX-1/IPF1 expression seem to be related to growth factor dependence on insulin/IGF-1 receptors and IGF-binding proteins.     

Genetic mechanisms for the synthesis of fucosyl GM1 in small cell lung cancer cell lines

Fucosyl GM1 has been reported to be specifically expressed in small cell lung cancer (SCLC) cells. However, the genetic basis for the synthesis of fucosyl GM1 has not been investigated. We analyzed the glycosyltransferases responsible for the synthesis of fucosyl GM1 in SCLC cell lines. In four SCLC cell lines expressing fucosyl GM1, both FUT1 and FUT2 mRNAs were detected, indicating that either one or both of alpha1,2-fucosyltransferases may be involved in the expression of fucosyl GM1. However, three of these four lines contained function-loss mutations in the FUT2 coding region, suggesting that FUT1 is mainly involved in the alpha1,2-fucosylation of GM1. The expression levels of the GM1 synthase gene showed no correlation with those of fucosyl GM1, whereas the co-transfection of GM1 synthase cDNA with FUT1 or FUT2 into SK-LC-17 clearly enhanced the neo-expression of fucosyl GM1, indicating its essential role. In contrast, the co-transfection of GD3 synthase cDNA reduced the expression levels of fucosyl GM1 with FUT1 or FUT2. Consequently, FUT1 seems to mainly contribute to the expression of fucosyl GM1, although both FUT1 and FUT2 are capable of generating the antigen. These results should promote the functional analysis of fucosyl GM1 leading to the development of novel therapies for SCLC.     

Effects of bombesin on human small cell lung cancer cells: evidence for a subset of bombesin non-responsive cell lines

The effects of bombesin on three human small cell lung carcinoma cell (SCLC) lines (NCI-H69, NCI-H128, and NCI-H345) have been examined and compared to the effects of the peptide on the mouse fibroblast cell line Swiss 3T3, and the rat pituitary tumor cell line GH3W5. While all three SCLC lines expressed messenger RNA encoding pro-gastrin releasing peptide (GRP), only the NCI-H345 cells expressed detectable membrane receptors for GRP and responded to nanomolar concentrations of bombesin as shown by 125I-GRP binding, total inositol phosphate accumulation, and increased clonal growth in soft agarose. These data show that some SCLC lines are insensitive to bombesin and do not express detectable membrane receptors for GRP.     

Neurotensin may function as a regulatory peptide in small cell lung cancer.

Neurotensin (NT) has been postulated to act as a modulatory agent in the central nervous system. Besides its presence in mammalian brain, NT is produced by small cell carcinoma of the lung (SCLC) and cell lines derived from these tumors. Receptors have also been characterized in some SCLC cell lines leading to the suggestion that NT could regulate the growth of SCLC in an autocrine fashion similar to bombesin/GRP. Previously, we had reported that a 10 nM dose of NT and NT(8-13), but not NT(1-8), elevated cytosolic Ca2+, indicating that SCLC NT receptors may use Ca2+ as a second messenger. Using intact SCLC cells we report that time-course incubations with NT lead to the formation of the amino-terminal fragment NT(1-8) and small amounts of the C-terminal fragment NT(9-13). These fragments are formed by metalloendopeptidase 3.4.24.15 cleaving enzyme at the Arg8-Arg9 bond of NT. Significant levels of soluble 3.4.24.15 (10-17 nmoles/mg Pr-/min) are present in SCLC cell lines. Using the in vitro clonogenic assay we tested the effect of 0.5, 5.0 and 10.0 nM doses of NT, NT(1-8) and NT(8-13) on SCLC clonal growth. NT and the C-terminal fragment NT(8-13) stimulated colony formation whereas the N-terminal fragment did not. In summary, NT may function as a regulatory peptide in SCLC through the formation of peptide fragments.     

Bioluminescence-based high-throughput screen identifies pharmacological agents that target neurotransmitter signaling in small cell lung carcinoma

Background

Frontline treatment of small cell lung carcinoma (SCLC) relies heavily on chemotherapeutic agents and radiation therapy. Though SCLC patients respond well to initial cycles of chemotherapy, they eventually develop resistance. Identification of novel therapies against SCLC is therefore imperative.

Methods and Findings

We have designed a bioluminescence-based cell viability assay for high-throughput screening of anti-SCLC agents. The assay was first validated via standard pharmacological agents and RNA interference using two human SCLC cell lines. We then utilized the assay in a high-throughput screen using the LOPAC¹²⁸⁰ compound library. The screening identified several drugs that target classic cancer signaling pathways as well as neuroendocrine markers in SCLC. In particular, perturbation of dopaminergic and serotonergic signaling inhibits SCLC cell viability.

Conclusions

The convergence of our pharmacological data with key SCLC pathway components reiterates the importance of neurotransmitter signaling in SCLC etiology and points to possible leads for drug development.