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c-MYC amplification and maligantn behaviour of small cell lung cancer (SCLC)
38
role for these MoAbs in vivo. Production of Monoclonal Antibodies (MoAbs) with Specificity for Small Cell Lung Cancer (SCLC) Following in Vitro In~nunisation with I m u n e Complex (IC)-Derived Nbterial. Wulfrank, D.A., Reeve, J.G., Bleehen, N.M. MRC Clinical Oncology and Radiotherapeutics Unit, MRC Centre, Cambridge, U.K. ICs present in the pleural effusions of a number of lung cancer patients have been shown to contain lung carcinoma associated antibodies and antigens capable of eliciting tumour directed heterologous antisera (Cancer Res., 42, 292, 1982). We describe the production of MoAbs having reactivity with SCLC following in vitro immunisation with IC-derived material isolated from the SCLC associated pleural effusion. Isolation of IC-like material was achieved by polyethylene glycol precipitation and Protein A affinity chromatography. The IgG containing complexes were then dissociated by ion-exchange chromatography to yield antibody (Ab)- and antigen (Ag)- containing fractions. Using a solid phase radioimmunowassay the Ab-containing preparation was screened against a panel of lung and non-lung tumour cell lines and was found to contain IgG having reactivity with SCLC and non-small cell lung cancer (NSCLC) but not with most non-pulmonary tumours. In vitro immunisation with Ab- and Ag-containing fractions led to the production of MoAbs B7/6 and GII/7 respectively. Both MoAbs reacted strongly with SCLC and showed little or no reactivity with NSCLC or with other tumour type s. IC-derived material would seem a suitable alternative to conventional immunogens for the production of tumour specific MoAbs.
representing I0 patients. One patient, unresponsive to chemotherapy, showed a low degree of c-myc amplification in the tumour and a very high in the derived cell line. None of the other 4 tumours showed c-myc amplification, neither did the cell lines derived from 2 of them. One of the patients whose tumours had no amplification was unresponsive, another responsive to chemotherapy. In vivo sensitivity could not be assessed in the remaining two patients. From 8 patients SCLC-derived cell-lines were obtained, c-Myc was amplified in 4 lines. Two of these, as well as one of the lines without amplification, had numerous double minutes (DMs). A possible correlation between the observed unresponsiveness to chemotherapy and the occurrence of DMs could be studied in 4 cases: in 3 cases lines appeared to contain numerous DMs, in 1 case there occurred only occasionally a few of them. In conclusion, our preliminary results do not lend support to the idea that c-myc amplification would indicate variant forms of SCLC. Cytotoxic Activity of HL~aan Tumor Necrosis Factor (human TNF) on Lung Cancer Cells in Vitro. Tani, G., Sugiura, M., Suetsugu, S., Umeda, H. Fujita-Gakuen Health University Hospital, Toyoake, Aichi 470-11, Japan. Not much is known about the effect of human TNF so a study to compare the relative effectiveness of human and rabbit TNF was undertaken. The human TNF was obtained through the intraperitoneal injecting of OK-432 to patients with peritonitis carcinomatosa. The dosage was 2KE on days 1,3,5 and 7, with a final dosage of 5KE on day 12. Twenty four hours after the final injection ascites were collected. The supernatant from this collected ascites was found to possess antitumor properties on human and mouse cultured cells but none on normal or otherwise non-transformed cell. It was seen to cause hemorrhage necrosis in hetero-transplanted human tumor cells. This action was not due to interferons or lymphokines, suggesting that it was human TNF. Percent I n h l b l t l o n control
c-MYC Amplification and Maligantn Behaviour ~f Small Cell Lung ~ancer (SCLC). 2 Buys , C.H.~.M., Osinga,. J''ide Leij , L., Post~us , P.E., Mooibroek , H., Scheffer-, H. 1. Departments of Human Genetics. 2. Climical Immunology, and 3. Pulmonary Diseases, State University of Groningen, Groningen, The Netherlands. In vitro SCLC can be divided into classic and variant forms, the latter representing the in vitro correlate of highly malignant, therapy-resistant SCLC. The recent suggestion that c-myc DNA a~mplification would play a role in the malignant behaviour of SCLC prompted us to study c-myc amplification in 5 SCLC tumours and 8 SCLC-derived cell lines,
of 3H-thymldlne uptake asciten
Before 0K-432 L-929
100
-27.6
A.S. (Small cell ca.)
i00 i00 i00
-19.1 41.I
K.O. T.O.
( A d e n o c a r c *noma ) ( A d e n o c a r c lnomD )
-30.4
After 0K-432 37.5
12.5 66.5 -19.0
Furthermore, when compared to the action of rabbit TNF it was seen that the anti-tumor effects were the same or particularly for L-929 cell, stronger. We therefore concluded that human TNF may have wider ranging anti-tumor properties than rabbit TNF, i.e., that human TNF may destroy tumor cells that rabbit TNF has no effect upon. Squamous Cell TAA Epitope D36h6: Enzyme In~nunoassay of Serial Sera From Patients on Specific Active I m u n o t h e r a p y . Hollinshead, A., Stewart, T.H.M. George Washington Univ. Med. Cre. Washington D.C. & Ottawa Univ.
role for these MoAbs in vivo. Production of Monoclonal Antibodies (MoAbs) with Specificity for Small Cell Lung Cancer (SCLC) Following in Vitro In~nunisation with I m u n e Complex (IC)-Derived Nbterial. Wulfrank, D.A., Reeve, J.G., Bleehen, N.M. MRC Clinical Oncology and Radiotherapeutics Unit, MRC Centre, Cambridge, U.K. ICs present in the pleural effusions of a number of lung cancer patients have been shown to contain lung carcinoma associated antibodies and antigens capable of eliciting tumour directed heterologous antisera (Cancer Res., 42, 292, 1982). We describe the production of MoAbs having reactivity with SCLC following in vitro immunisation with IC-derived material isolated from the SCLC associated pleural effusion. Isolation of IC-like material was achieved by polyethylene glycol precipitation and Protein A affinity chromatography. The IgG containing complexes were then dissociated by ion-exchange chromatography to yield antibody (Ab)- and antigen (Ag)- containing fractions. Using a solid phase radioimmunowassay the Ab-containing preparation was screened against a panel of lung and non-lung tumour cell lines and was found to contain IgG having reactivity with SCLC and non-small cell lung cancer (NSCLC) but not with most non-pulmonary tumours. In vitro immunisation with Ab- and Ag-containing fractions led to the production of MoAbs B7/6 and GII/7 respectively. Both MoAbs reacted strongly with SCLC and showed little or no reactivity with NSCLC or with other tumour type s. IC-derived material would seem a suitable alternative to conventional immunogens for the production of tumour specific MoAbs.
representing I0 patients. One patient, unresponsive to chemotherapy, showed a low degree of c-myc amplification in the tumour and a very high in the derived cell line. None of the other 4 tumours showed c-myc amplification, neither did the cell lines derived from 2 of them. One of the patients whose tumours had no amplification was unresponsive, another responsive to chemotherapy. In vivo sensitivity could not be assessed in the remaining two patients. From 8 patients SCLC-derived cell-lines were obtained, c-Myc was amplified in 4 lines. Two of these, as well as one of the lines without amplification, had numerous double minutes (DMs). A possible correlation between the observed unresponsiveness to chemotherapy and the occurrence of DMs could be studied in 4 cases: in 3 cases lines appeared to contain numerous DMs, in 1 case there occurred only occasionally a few of them. In conclusion, our preliminary results do not lend support to the idea that c-myc amplification would indicate variant forms of SCLC. Cytotoxic Activity of HL~aan Tumor Necrosis Factor (human TNF) on Lung Cancer Cells in Vitro. Tani, G., Sugiura, M., Suetsugu, S., Umeda, H. Fujita-Gakuen Health University Hospital, Toyoake, Aichi 470-11, Japan. Not much is known about the effect of human TNF so a study to compare the relative effectiveness of human and rabbit TNF was undertaken. The human TNF was obtained through the intraperitoneal injecting of OK-432 to patients with peritonitis carcinomatosa. The dosage was 2KE on days 1,3,5 and 7, with a final dosage of 5KE on day 12. Twenty four hours after the final injection ascites were collected. The supernatant from this collected ascites was found to possess antitumor properties on human and mouse cultured cells but none on normal or otherwise non-transformed cell. It was seen to cause hemorrhage necrosis in hetero-transplanted human tumor cells. This action was not due to interferons or lymphokines, suggesting that it was human TNF. Percent I n h l b l t l o n control
c-MYC Amplification and Maligantn Behaviour ~f Small Cell Lung ~ancer (SCLC). 2 Buys , C.H.~.M., Osinga,. J''ide Leij , L., Post~us , P.E., Mooibroek , H., Scheffer-, H. 1. Departments of Human Genetics. 2. Climical Immunology, and 3. Pulmonary Diseases, State University of Groningen, Groningen, The Netherlands. In vitro SCLC can be divided into classic and variant forms, the latter representing the in vitro correlate of highly malignant, therapy-resistant SCLC. The recent suggestion that c-myc DNA a~mplification would play a role in the malignant behaviour of SCLC prompted us to study c-myc amplification in 5 SCLC tumours and 8 SCLC-derived cell lines,
of 3H-thymldlne uptake asciten
Before 0K-432 L-929
100
-27.6
A.S. (Small cell ca.)
i00 i00 i00
-19.1 41.I
K.O. T.O.
( A d e n o c a r c *noma ) ( A d e n o c a r c lnomD )
-30.4
After 0K-432 37.5
12.5 66.5 -19.0
Furthermore, when compared to the action of rabbit TNF it was seen that the anti-tumor effects were the same or particularly for L-929 cell, stronger. We therefore concluded that human TNF may have wider ranging anti-tumor properties than rabbit TNF, i.e., that human TNF may destroy tumor cells that rabbit TNF has no effect upon. Squamous Cell TAA Epitope D36h6: Enzyme In~nunoassay of Serial Sera From Patients on Specific Active I m u n o t h e r a p y . Hollinshead, A., Stewart, T.H.M. George Washington Univ. Med. Cre. Washington D.C. & Ottawa Univ.