Oncology 36: 242-244 (1979)
Correlation between Tumour Size, Metastatic Spread and Galactosyl Transferase Activity in Cyclophosphamide-Treated Mice Bearing the Lewis Lung Carcinoma I. D. Capet, M. Jenner, M. H. Pinnock, It. M. Dorrell, D. C. Payne and D. C. Williams Research Department, Marie Curie Memorial Foundation, The Chart, Oxtcd, Surrey
Key Words. Galactosyl transferase • Metastasis •Growth •Lewis lung carcinoma • Cyclophosphamide • Mice
Introduction The inherent toxicity of cancer chemotherapeutic agents has stimulated research into markers which could be used to monitor tumour status. Antigenic markers such as carcinogenic embryonic antigen and a-fetoprotein, which are proving useful for cancer screening and detection, are of limited value in deter mination of tumour status [1], Recently, determina tion of the levels of certain key enzymes associated with tumour growth have differentiated between some ma lignant and non-malignant experimental tumours [2, 3]. The Lewis lung carcinoma in mice is frequently used as an experimental model of a metastasizing tumour and can be controlled chemotherapeutically [4, 5]. In this experiment the level of galactosyl transferase, which has been examined as a possible tumour marker [3, 6], was measured in mice receiving cyclophosphamide so as to correlate the enzyme activity with tumour growth.
Materials and Methods Chemicals. Uridine diphospho-D-1U-IJC] galactose (specific ac tivity 200 Ci/mol) was purchased from the Radiochemical Centre. Amcrsham, Bucks. Non-labelled UDP galactose, ATP. dithiothrcitol. fetuin and phosphotungstic acid were obtained from the Sigma Chem ical Company, Poole, Dorset and cyclophosphamide from Koch-Light
Laboratories, Colnbrook. Bucks. All other reagents were of the purest grade available and supplied by Fisons, Loughborough, Leics. Animals and Treatment. 100 male C57/BL/10ScSn/Ola mice of body weight 20 ± 2 g were purchased from Olac, Bicester. Oxon, and maintained on PRM diet (Dixons, Ware, Herts.). The mice were injected intramuscularly in one flank with I x It)6 freshly prepared Lewis lung carcinoma cells suspended in phosphatebuffered saline (0.1 ml) which had been maintained by intramuscular passage in the syngeneic strain every 14 days. The animals were then randomly divided into four groups and mice in three of the groups received cyclophosphamide by intraperitoneal injection on alternate days, beginning five days after inoculation of the tumour cells. Cyclo phosphamide was administered to each group at one of the three dose levels indicated in table 1, control animals were injected with saline. 15 days after inoculation of the tumour the mice were sacrificed by cervical dislocation and the normal and tumour-bearing thighs removed and weighed. The lungs of each mouse were removed, fixed in Bouin’s solution and the number of tumour foci estimated manually. Enzyme Incubations. The normal and tumour-bearing thighs were homogenized separately in Tris buffer. Galactosyl transferase was assayed by the method of I’att and Grimes |7| with the modifica tion of Chatterjee and Kim |3], using saturating concentrations of agalactofctuin prepared in the manner of Spiro [8, 9] as exogenous acceptor. Protein was determined by the method of Lowry et at. 110]. Radiochemical Analysis. The reaction mixture from each incubate was filtered through 0.20 ftm Millipore filters and the associated radioactivity in the filters determined in lnstagel® by counting for 2 cycles in a Packard 2650 liquid scintillation spectrometer'. Counting efficiency was determined by the twin-channel ratio method. Statistical Analysis. The statistical significance between test and control groups was determined by the Student's t test. The difference between means was considered significant when p
Correlation between Tumour Size, Metastatic Spread and Galactosyl Transferase Activity in Cyclophosphamide-Treated Mice Bearing the Lewis Lung Carcinoma I. D. Capet, M. Jenner, M. H. Pinnock, It. M. Dorrell, D. C. Payne and D. C. Williams Research Department, Marie Curie Memorial Foundation, The Chart, Oxtcd, Surrey
Key Words. Galactosyl transferase • Metastasis •Growth •Lewis lung carcinoma • Cyclophosphamide • Mice
Introduction The inherent toxicity of cancer chemotherapeutic agents has stimulated research into markers which could be used to monitor tumour status. Antigenic markers such as carcinogenic embryonic antigen and a-fetoprotein, which are proving useful for cancer screening and detection, are of limited value in deter mination of tumour status [1], Recently, determina tion of the levels of certain key enzymes associated with tumour growth have differentiated between some ma lignant and non-malignant experimental tumours [2, 3]. The Lewis lung carcinoma in mice is frequently used as an experimental model of a metastasizing tumour and can be controlled chemotherapeutically [4, 5]. In this experiment the level of galactosyl transferase, which has been examined as a possible tumour marker [3, 6], was measured in mice receiving cyclophosphamide so as to correlate the enzyme activity with tumour growth.
Materials and Methods Chemicals. Uridine diphospho-D-1U-IJC] galactose (specific ac tivity 200 Ci/mol) was purchased from the Radiochemical Centre. Amcrsham, Bucks. Non-labelled UDP galactose, ATP. dithiothrcitol. fetuin and phosphotungstic acid were obtained from the Sigma Chem ical Company, Poole, Dorset and cyclophosphamide from Koch-Light
Laboratories, Colnbrook. Bucks. All other reagents were of the purest grade available and supplied by Fisons, Loughborough, Leics. Animals and Treatment. 100 male C57/BL/10ScSn/Ola mice of body weight 20 ± 2 g were purchased from Olac, Bicester. Oxon, and maintained on PRM diet (Dixons, Ware, Herts.). The mice were injected intramuscularly in one flank with I x It)6 freshly prepared Lewis lung carcinoma cells suspended in phosphatebuffered saline (0.1 ml) which had been maintained by intramuscular passage in the syngeneic strain every 14 days. The animals were then randomly divided into four groups and mice in three of the groups received cyclophosphamide by intraperitoneal injection on alternate days, beginning five days after inoculation of the tumour cells. Cyclo phosphamide was administered to each group at one of the three dose levels indicated in table 1, control animals were injected with saline. 15 days after inoculation of the tumour the mice were sacrificed by cervical dislocation and the normal and tumour-bearing thighs removed and weighed. The lungs of each mouse were removed, fixed in Bouin’s solution and the number of tumour foci estimated manually. Enzyme Incubations. The normal and tumour-bearing thighs were homogenized separately in Tris buffer. Galactosyl transferase was assayed by the method of I’att and Grimes |7| with the modifica tion of Chatterjee and Kim |3], using saturating concentrations of agalactofctuin prepared in the manner of Spiro [8, 9] as exogenous acceptor. Protein was determined by the method of Lowry et at. 110]. Radiochemical Analysis. The reaction mixture from each incubate was filtered through 0.20 ftm Millipore filters and the associated radioactivity in the filters determined in lnstagel® by counting for 2 cycles in a Packard 2650 liquid scintillation spectrometer'. Counting efficiency was determined by the twin-channel ratio method. Statistical Analysis. The statistical significance between test and control groups was determined by the Student's t test. The difference between means was considered significant when p
