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Oncogene amplification of C-Myc and N-Myc in cell lines established from patients with small cell lung cancer (SCLC) is associated with shortened survival
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Amplified Variant SCLC Lines. Johns~::, B.E., Battey, J.F., Carney, D., Minna, J.D. NCI-Navy Medical Oncology Branch. COP/DCT/NCI, Bethesda, MD 20814. Human SCLC cell iines which exhibit amplification and expression of the c-myc proto-oncogene (variant phenotype) have a variety of biologic differences when compared to SCLC lines without c-myc amplification (classic phenotype) including: large cell loose morphology, shorter doubling time, higher cloning effiency, and lower dopa decarboxylase (DDC) production. To determine the relationship between increased c-myc gene expression and phenotype, a normal c-myc gene in the plasmid diagrammed below was transfected into a classic cloned SCLC line, H209. Eco R1 Eco R1 Bam H1 Bam HI Individual 12.5 kb c-m~c [ ~ 3.4kb neo I pBR 322 clones with the stably integrated plasmid were selected with G-418. The parent (H209), a c - w e amplified line (N417), and clones were analyzed for: c - w c and neo copy number and expression, doubling time (D.T.), cloning efficiency (C.F.), and DDC production. Cell line C-r~c# Neo # C-myc RNA O.T. C.E. DDC H209 1 0 1 52 hrs .33% 56 N417 60 0 60 26 hrs 8.4% <.1 209 A 1 1 1 52 hrs .23% 50 209 B 2 1 5 36 hrs .34% 55 209 C 6 6 10 26 hrs 1.8% 37
Clone A maintained the spherical morphology of H209 while clones B & C developed a loose morphology resembling N417. We conclude that c-myc gene can be introduced and expressed to varying degrees in SCLC. C-myc expression in a dose related fashion leads to a shortened doubling time, increased cloning efficiency, and a loosened morphology but dopa decarboxylase production is not dramatically lowered.
0ncogene Amplification of C-Myc and N-Myc in Cell Lines Established from Patients with Small Cell Lung Cancer (SCLC) is Associated with Shortened Survival. Johnson, B.E., Nau, M.N., Gazdar, A.F., Carney, D.N., Oie, H.K., Minna, J.D., Ihde, D.C..NCI-Navy Medical Oncology Branch, National Cancer Institute and Naval Hospital, Bethesda, MD 20814. U.S.A. To determine if c-myc and n-myc amplification in cell lines established from patients with SCLC influences survival, the clinical course of patients from whom cell lines were obtained was reviewed. DNA was prepared from 32 cell lines established between 1977-1984 from different SCLC patients, and the number of c-myc and n-myc gene copies was determined. In 20 patients the cell line was established at the time of tumor progression. Eight of 20 lines (40%) had 4-60
fold amplification of c-myc (5) or n-myc (3). The cell lines were started a median of 46 weeks (range 18-109) from the initiation of chemotherapy. Median survival from initiation of therapy for patients whose cells had myc amplification vs. those who did not was 33 weeks (c-myc), 40 weeks (c + n-myc) (range 20-70) vs. 53 weeks (range 32112) (p<0.05). Median survival from time of obtaining tumor for culture was only 4 weeks (range 0-25) and did not differ in amplified and unamplified patients. The two groups did not differ significantly in pretreatment performance status, stage, or therapy. In addition, DNA from 12 cell lines established from extensive stage patients at diagnosis was evaluated for myc gene copy number. Two of 12 (17%) were amplified, 1 c-myc and 1 n-myc. These patients survived i01 and 17 weeks respectively compared to a median survival of 15 weeks (range 0-55) in unamplified patients. We conclude treated patients whose SCLC cell lines have amplification of the c-myc or n-myc oncogene have shorter survival not explicable by differences in standard prognostic factors. Myc gene amplification appears more common in treated than in untreated patients.
c-sis Expression and Simultaneous Production of Platelet-Derived Growth Factor (PDGF) in a H ~ a n Lung Cancer Cell Line. Betsholtz, C., Bywater, M., Westermark, B., Bergh, J. Department of Pathology, University of Uppsala, Sjukhusvagen i0, S-751 85 Uppsala, Sweden. A cell line was derived from the pleural effusion of a haman large cell lung carcinoma. The primary bronchial and mediastinal biopsies demonstrated a poorly differentiated lung cancer of large cell type, surrounded by a prominent stroma reaction. Electronmicroscopy of biopsy cells and the established cell line revealed cellular junctions and tonofilaments. The intermediate filaments were, demonstrated by immunoblotting- and immunofluorescence techniques, to be of both cytokeratin- and neurofilament type. PDGF production by the cell line was ~emonstrated with a radio-receptor assay and by 3-S-cysteine labelling demonstrating PDGF specific bands at MW:s 31000 unreduced and 16000 after reduction. No PDGF cell surface receptors could, however, be detected. Northern blot analysis of poly (A) RNA demonstrated high levels of c-sis transcripts. (25 fold the level in U-20S osteosarcoma cells). Notably, the corresponding hybridisation to DNA only demonstrated a single gene copy of c-si___~s. We have thus demonstrated an abnormal expression of c-si___ss,previously only supposed to be found in gliomas and sarcomas, in a human lung cancer cell line. In tumor cell systems lacking PDGF receptors, the synthesis and release of PDGF-Iike substances might exclusively affect the growth properties of surrounding normal tissues. Indeed, the tumor from which this lung cancer cell line was derived, demonstrated a marked stroma reaction.
Monoclonal Antibodies to Lung TL~nor-Associated Antigens.
Amplified Variant SCLC Lines. Johns~::, B.E., Battey, J.F., Carney, D., Minna, J.D. NCI-Navy Medical Oncology Branch. COP/DCT/NCI, Bethesda, MD 20814. Human SCLC cell iines which exhibit amplification and expression of the c-myc proto-oncogene (variant phenotype) have a variety of biologic differences when compared to SCLC lines without c-myc amplification (classic phenotype) including: large cell loose morphology, shorter doubling time, higher cloning effiency, and lower dopa decarboxylase (DDC) production. To determine the relationship between increased c-myc gene expression and phenotype, a normal c-myc gene in the plasmid diagrammed below was transfected into a classic cloned SCLC line, H209. Eco R1 Eco R1 Bam H1 Bam HI Individual 12.5 kb c-m~c [ ~ 3.4kb neo I pBR 322 clones with the stably integrated plasmid were selected with G-418. The parent (H209), a c - w e amplified line (N417), and clones were analyzed for: c - w c and neo copy number and expression, doubling time (D.T.), cloning efficiency (C.F.), and DDC production. Cell line C-r~c# Neo # C-myc RNA O.T. C.E. DDC H209 1 0 1 52 hrs .33% 56 N417 60 0 60 26 hrs 8.4% <.1 209 A 1 1 1 52 hrs .23% 50 209 B 2 1 5 36 hrs .34% 55 209 C 6 6 10 26 hrs 1.8% 37
Clone A maintained the spherical morphology of H209 while clones B & C developed a loose morphology resembling N417. We conclude that c-myc gene can be introduced and expressed to varying degrees in SCLC. C-myc expression in a dose related fashion leads to a shortened doubling time, increased cloning efficiency, and a loosened morphology but dopa decarboxylase production is not dramatically lowered.
0ncogene Amplification of C-Myc and N-Myc in Cell Lines Established from Patients with Small Cell Lung Cancer (SCLC) is Associated with Shortened Survival. Johnson, B.E., Nau, M.N., Gazdar, A.F., Carney, D.N., Oie, H.K., Minna, J.D., Ihde, D.C..NCI-Navy Medical Oncology Branch, National Cancer Institute and Naval Hospital, Bethesda, MD 20814. U.S.A. To determine if c-myc and n-myc amplification in cell lines established from patients with SCLC influences survival, the clinical course of patients from whom cell lines were obtained was reviewed. DNA was prepared from 32 cell lines established between 1977-1984 from different SCLC patients, and the number of c-myc and n-myc gene copies was determined. In 20 patients the cell line was established at the time of tumor progression. Eight of 20 lines (40%) had 4-60
fold amplification of c-myc (5) or n-myc (3). The cell lines were started a median of 46 weeks (range 18-109) from the initiation of chemotherapy. Median survival from initiation of therapy for patients whose cells had myc amplification vs. those who did not was 33 weeks (c-myc), 40 weeks (c + n-myc) (range 20-70) vs. 53 weeks (range 32112) (p<0.05). Median survival from time of obtaining tumor for culture was only 4 weeks (range 0-25) and did not differ in amplified and unamplified patients. The two groups did not differ significantly in pretreatment performance status, stage, or therapy. In addition, DNA from 12 cell lines established from extensive stage patients at diagnosis was evaluated for myc gene copy number. Two of 12 (17%) were amplified, 1 c-myc and 1 n-myc. These patients survived i01 and 17 weeks respectively compared to a median survival of 15 weeks (range 0-55) in unamplified patients. We conclude treated patients whose SCLC cell lines have amplification of the c-myc or n-myc oncogene have shorter survival not explicable by differences in standard prognostic factors. Myc gene amplification appears more common in treated than in untreated patients.
c-sis Expression and Simultaneous Production of Platelet-Derived Growth Factor (PDGF) in a H ~ a n Lung Cancer Cell Line. Betsholtz, C., Bywater, M., Westermark, B., Bergh, J. Department of Pathology, University of Uppsala, Sjukhusvagen i0, S-751 85 Uppsala, Sweden. A cell line was derived from the pleural effusion of a haman large cell lung carcinoma. The primary bronchial and mediastinal biopsies demonstrated a poorly differentiated lung cancer of large cell type, surrounded by a prominent stroma reaction. Electronmicroscopy of biopsy cells and the established cell line revealed cellular junctions and tonofilaments. The intermediate filaments were, demonstrated by immunoblotting- and immunofluorescence techniques, to be of both cytokeratin- and neurofilament type. PDGF production by the cell line was ~emonstrated with a radio-receptor assay and by 3-S-cysteine labelling demonstrating PDGF specific bands at MW:s 31000 unreduced and 16000 after reduction. No PDGF cell surface receptors could, however, be detected. Northern blot analysis of poly (A) RNA demonstrated high levels of c-sis transcripts. (25 fold the level in U-20S osteosarcoma cells). Notably, the corresponding hybridisation to DNA only demonstrated a single gene copy of c-si___~s. We have thus demonstrated an abnormal expression of c-si___ss,previously only supposed to be found in gliomas and sarcomas, in a human lung cancer cell line. In tumor cell systems lacking PDGF receptors, the synthesis and release of PDGF-Iike substances might exclusively affect the growth properties of surrounding normal tissues. Indeed, the tumor from which this lung cancer cell line was derived, demonstrated a marked stroma reaction.
Monoclonal Antibodies to Lung TL~nor-Associated Antigens.