SIO

transferrin synthesis by a small cell lung cancer line (NCI-HSIO) that survives in serum-free media without added transferrin. Immunoassays for human transferrin demonstrated that these cells contained im- munoreactive human transferrin. Immunofluorescence studies showed that the protein is expressed on the surface of cells, presumably bound to transfctin receptor. Media conditioned by NCI-H-510 cells support

proliferation of human leukemic cells that would not survive in media lacking transferrin. [“S]Methionine incorporation documented tram-

ferrin synthesis by NCI-HSlOcellsas wellasthreeothersmallce111ines. Transferrin synthesis by NCI-HSlO cells increased more than IO-fold when cells entered active phases of the cell cycle, and this increase was seen before large increases in transfetin-receptor expression. Further experiments examining the effects of agents that affect iron metabolism show that the addition of agents that affect iron metabolism show that the ad&ion of wansfcrrin-iron or hemin to the media is associated with a more rapid initial rate of proliferation and lower rates of transferrin synthesis than control cells. Gallium salts, which inhibit iron uptake, inhibited proliferation of these cells. If the cells recovered from this effect, aansfcrrin synthesis remained greatly increased compared to control. We conclude that transferrin synthesis by these malignantce11s is ultimately related to an iron requirement for cellular proliferation. 11

appears that this synthcsizcd transferrin acts as part of an important autocrinc mechanism permitting proliferation of these cells, and per- haps permitting tumor cell growth in vivo in areas not well vascularized.

Human lung cancer nodules in organotypic culture: No evidence of correlation between the antiproliferative effects of interferoas and the induction of 2’,_5’-oligoadenylate synthetase. Martyrc M-C, Beaupain R, Falcoff E. C/ 196 INSERM, Instilut Curie, F- 75231 Paris Cedex 05. Tumor Biol 1988;9:263-9.

Alveolar II pulmonary tumor cells (A.549), maintained in continuous tridimcnsional organotypic culture, were used in an attempt to investi- gate whether there could be a relationship of the 2’S’-oligioadenylate (2,SA)synthetasepathway totheantiproliferativeactivity ofinterferons (IFNs) in this particular tumor ccl1 model. IF?+_,, -8 and -gamma were used separately and in combinations. IFN-_, and -gamma demonstrated an inhibitory effect on the nodule growth, whereas IFN-I3 did not. Moreover, combinations of IFN-_, and -gamma resulted in a significant synergisticanriproliferativeactivity;lFN-OonlypotentiatedslighUythe effect of IFN-gamma. All three IFNs induced an increase in the 2,ZA synthetase activity, indicating a discrepancy with the pattern of anticel- lular activity. Furthermore, whereas the combination of IFN-, and - gamma resulted in a synergistic antiproliferative effect, no synergism was observed in the induction of the enzyme. These results show that there is a lack of correlation between the sensitivity or the resistance to IFNs of AS49 tumor nodules and the induction of the 2,5A synthetase activity.

Methylation status of epidermal growth factor receptor gene in lung carcinoma cells. Gamou S, Shimosato Y, Merlin0 GT, Shimizu N. Deparlment of Molecular Biology, Keio University School of Medicine, Shinjuku-ku, Tokyo 160. Jpn J Cancer Res, Gann 1988;79:989-95.

The methylation status of the epidermal growth factor receptor (EGFR) gene was compared in cell lines from four major types of lung carcinoma, small cell lung carcinoma (SCLC), large cell lung carci- noma, squamous cell carcinoma and adenocarcinoma, in order to examine whether DNA meth ylation is responsible for the suppression of EGFR gene in SCLC cells. Southern blot analysis revealed that the structural region of the EGFR gene is methylated in various degrees regardless of the expression of EGF receptor on the surface. An 8- kilobase EcoRl fragment which contains the EGFR gene promoter region is readily digested with various methylation-sensitive restriction enzymes in all types of cells, indicating that the EGFR gene 5’ region is

barely methylated. Thus, a mechanism other than DNA methylation appears to control EGFR gene expression and the lack of EGFR gene expression in SCLC cells may be caused by a paucity of some transcrip- tion regulatory factor(s).

Castrin-releasingpeptidegene-associatedpeptidesareexpressed in normal human fetal lung and small cell lung cancer: A novel peptide family found in man. CuttittaF,FedorkoJ,GuJ,Lebacq-VerheydenA-M,LinnoilaRl,Battey JF. Uniformed Services Universiry of the Health Sciences, Department of Defense, Division of Cancer Treatment. National Cancer Institute and National Naval Medical Center, Bethesda, MD 20814. J Clin Endocrinol Metab 1988;67:576-83.

Mammalian gastrin-releasing peptide (GRP) is found in cells of neuroendocrine and neural origin, and GRP mediates a variety of physiological and trophic responses when it binds to high affinity cell surface receptors on effector cells. Analysis of cDNA clones derived from prepro-GRP mRNAs predict the concurrent expression ofa unique series of peptide hormones, the GRP gene-associated peptides (GGAPs). AltemativeRNA splicing of the primary GRPgene transcript results in mRNAs that could encode 3 distinct forms of GGAPs. Using specific antisera directed against synthetic peptides representing par- tions of the predicted GGAPs, we found multiple GGAP forms in the endocrine cells of human fetal lung and in human small cell lung cancer cells (SCLC). Within a single pulmonary endocrine cell, at least 2 of these 3 predicted forms could be demonstrated using immunohisto- chemical techniques. In addition, concordant expression of GRP and GGAPs was found in 10 SCLC cell lines and 3 human SCLC tumors. These findings establish GGAPs as a novel peptide family in man and warrant further investigation into their potential role in normal and malignant growth.

Response of primary human lung carcinomas to autocrine growth factors produced by a lung carcinoma cell line. Siegfried JM, Owens SE. Carcinogenesis and Melabolism Section, Environmerual Health Research and Teaing. Inc., Research Triangle Park, NC 27709. Cancer Res 1988;48:4976-81.

Medium conditioned for 48 to 72 h by A549-1 lung carcinoma cells was used to culture primary solid lung tumors on feeder layers of inactivated Swiss 3T3 cells. Of 22 cases placed into culture, primary cultures of carcinoma cells were obtained in 20. Subcultures were obtained in 18 cases, and cell lines were established in nine cases. The neoplastic origin of the cultured cells was demonstrated by several criteria: tumorigenicity in athymic mice; anchorage-independent growth; expression of altered lactate dehydrogenase isoenzyme pro- tiles; and expression of the lung tumor marker pregnancy-specific glycoprotein 1. The epithelial nature of cultured carcinoma cells was demonstrated by expression of keratin. These characteristics were compared to normal epithclial cells established in culture from bron- chialexplantsfromthesamedonorsas tumortissue,orotherdonors.The growth-stimulating effect of conditioned medium toward primary or newly cultured tumor cells was quantitated by clonal assays in soft agar and in monolayer culture. Growth response in clonal assays of newly cultured carcinoma cells to the purified growth factors transforming growth factor-and insulin-likegrowthfactor 1, two knowncomponents of medium conditioned by A549-1 cells, was also demonstrated.

Effects of bombesin antagonists on the growth of small cell lung cancer cells in vitro. Layton JE, Scanlon DB, Soveny C, Morstyn G. Ludwig Insiitufe for CancerResearch,Melbourne TumourBiologyBranch,Melbourne.Vic. 3050. Cancer Res 1988;48:4783-9.

Small cell lung cancer (SCLC) produces several neuroendocrine peptides, including gasuin-releasing peptide (GRP), the mammalian