Abstracts/Lung Cancer 13 (1995) 185-232 195

Multiple forms of cathepsin B in human lung cancer Krepela E, Prochazka J, Mynarikova H, Karova B, Polak J, Cermak J et al. Dept. Molecular/CeNular Pneumologv, Institute Chest Diseases, Budinova 67, 180 71 Prague 8. Int J Cancer 1995;61:44-53.

In this study we have examined, by means of isoelectric focusing (IEF) in native polyacrylamide gel and contact-print fluorescence zymography, whether human lung carcinomas and the lung parenchyma contain di&rent pools of multiple charge forms of the cysteine pmteinase cathepsin B. The isoelectric point @I) patterns of cathepsin B from lung carcinoma and matched lung were similar, particularly with regard to 2 major intermediate acidic enzyme p1 forms designated as I and II (PI(app)) of 5.10 and 4.93 in tumors, and 5.11 and 4.94 in lungs, respectively). The slightly acidic cathepsin B p1 forms @I(app)) 5.47- 5.19) in squamous-cell lung carcinoma (SQCLC) were significantly more numemus than such enzyme p1 forms in lungs. The numbers of the highly acidic cathepsin B p1 forms (PI(app)) 4.82-4.33) were significantly higher in SQCLC and lung adenocarcinoma (ACL) than in matched lung. The activity distribution percentage in the set of highly acidic cathepsin B p1 forms was signiticantly higher in SQCLC and ACL than in matched lung. We also observed that cathepsin B from SQCLC and matched lung was fully recoverable by IEF from inhibition by leupeptin. Using the cysteine-proteinase-specific inactivator E-64, we revealed by IEF that some cathepsin B isoforms (charge forms) from SQCLC were more resistant to inactivation by this compound than the corresponding enzyme isoforms from lungs. After IEF, the enzyme isoforms apparently lost their resistance to E-64. Our results indicate that the pool of multiple charge forms of cathepsin B in SQCLC and ACL is different from that in the lung. and also that there may be an increased level of loose complexes between cathepsin B and some proteins or polypeptides in SQCLC compared to the lung.

Establishment and characterixation of a new human lung poorly differentiated adenocarcinoma cell line, GLL-1, producing carcinoembryonic antigen (CEA) and CA19-9 Mitsuhashi N, Takahashi T, Sakurai H, Yamakawa M, Hasegawa M, Furuta M. et al. Dept. Radiology/Radiation Oncology. Gunma Universi@ School Medicine. 3-39-22 Showa-machi Maebashi, Gunma 371. Lung Cancer (Ireland) 1995;12: 13-24.

A new human lung poorly differentiated adenocarcinoma cell line producing carcinoembryonic antigen (CEA) and CAI9-9, designated GLL-1, was established from a serial xenograft established in our laboratory. In this paper, we investigated the characteristics of GLL-I cells. The cell line grew in a monolayer and had a polygonal epithehoid appearance in phase-contrast microscopy. The doubling time of GLL-I cells during exponential growth was approximately 28 h in RPMI-1640 with 10% fetal calf serum. Positive reactions for CEA and for CA19-9 were seen within the cytoplasm of GLL-I cells by immunohistochemical staining. In log phase, the fractions for GdG,, S and G,M were 76.2%, 20.0% and 3.8% respectively. Chromosome analysis revealed that GLL- 1 cells contained the modal number of 72 chromosomes (range 66-97). GLL-1 cells exhibited numerous numerical and st~ctural abnormahties. The Do value for radiation sensitivity, calculated by taking the reciprocals of the slope of the radiation dose response curve, was 109 cGy. The extrapolation number (n) for the radiation response. curve was 1.57. Tumor take was observed in nude mice inoculated subcutaneously in the back with 2 x 10’ GLL-1 cells. The histopathological features of the tumor induced by inoculation of GLL-I cells were also the same as the original lung cancer.

Alternative splicing of the neurofibromatosis I gene correlates with growth patterns and neuroendocrine properties of human small-cell lung-carcinoma cells Koh T, Yokota J, Gokawa K, Kina T, Koshimura K, Miwa S et al. Dept. Tumor Biology and Pathology Faculty o/Medicine, Kyoto Universi&, Yoshida-konoe, Sakyo-ku, Kpto 606. Int J Cancer 1995;60:843-7.

Iwo distinct transcripts, type I and type. II, of the neurofibromatosis I (NFI) gene are generated by alternative splicing in the region corresponding to the gene’s GTPase-activating protein-related domain (GRD). Relative expression levels of these 2 transcripts were previously correlated to neural differentiation. Since smaIl-cell lung carcinoma (SCLC) often exhibits neurcendoctine properties, we analyzed the typc- I to type-II mRNA ratio in 15 SCLC &I lines, using reverse tmnscriptase and polymerase chain reaction methods. The type-1 mRNA was predominant in 10 cell lines; 8 of them grew as floating aggregates in culture and had high Ldopa decarboxylase @DC) activity. The other 5 lines predominantly expressed type-11 mRNA, adhered to the culture substrate, and expressed low or undetectable levels of neural cell- adhesion molecule (NCAM) antigen and DDC activity. N2+, one of the subclones of NCI-N4 17 cells, exhibited a higher type-1 to type-II ratio after the cells had adhered to a laminin-coated plate and had emitted neurite-like processes. These findings provide evidence that alternative splicing patterns of NFI mRNA correlate with the mechanisms that regulate the growth patterns and neuroendocrine properties of SCLC cells in vitro.

Frequent loss of imprinting of the H19 gene is often associated with its overexpression in human lung cancers Kondo M, Suzuki H, Ueda R, Osada H, Takagi K, Takahashi T et al. Ltiboratoty of Chemotherapy, Aichi Cancer Center Research Inst.. Chikusa-ku, Nagoya 464. Oncogene 1995;lO: 1193-S.

Accumulating evidence suggests that deregulation of the insuhn- like growth factor II (IGFZ) and H19 genes at Ilpl5, due to loss of imprinting (LOI), plays a role in the oncogenesis of Wilms’ tumors. We previously reported LGI of IGF2 in carcinomas of the lung, a common cancer of adults. We show here that LO1 of HI9 is also a frequent event in lung cancer development, and correlated with hypomethylation of the promoter region. Furthermore, the present study also revealed that overexpression of H19 is often associated with LO1 of H 19 in lung cancers retaining both parental alleles, showing a contrast to LO1 in Wilms’ tumors. Interestingly, in contrast to frequent LO1 of the imprinted genes at lIpIS, LOI was not observed for the remaining gene known to be imprinted in man, SNRPN at 15q12.

Growth and biochemical effects of unsymmetrically substituted polyamine analogues in human lung tumor cells Casero RA Jr, Mank AR Saab NH, Wu R, Dyer WJ, Woster PM. Oncology Center, Johns Hopkins University School Med., 424 North Bond Street, Ballimore, MD 21231. Cancer Chemother Pharmacol 1995;36:69-74.

Three unsymmetricahy substituted polyamine analogues demonstrate significant and selective antitumor effects. Each of the analogues N’- ethyl-N”-propargyl-4,gdiazaundecane (PENSpm), Ni-ethyl-N”- (cyclobutyl)methyl-4,8-diazaundecane (CBENSpm), and N’ethyl-N”- (cyclopropyl)methyI-4,8-diar.aundecane (CPENSpm) is cytotoxic to a representative non-smallceI1 lung carcinoma line, NC1 H157, while being only growth-inhibitory to a representative small-cell-lung carcinoma line, NC1 H82. Cytotoxicity is accompanied by a significant increase in expression of the polyamine catabolic enzyme spermidine/ spermine N1-acetyhransferase (SSAT) at the levels ofactivity and stcady- state mRNA. These new analogues are signifmant both for their cell- type-specific activity and as synthetic prototypes for the addition of SSAT-activated functional groups.