Deoxycytidine Kinase and Cytosine Nucleoside Deaminase Activities in Synchronized Cultures of Normal Rat Kidney Cells1 Charnwingwan2andtak W

Home , Deoxycytidine

(CANCERRESEARCH38,2768-2772,September1978] 0008-5472/78/0038-0000$02.00 Deoxycytidine Kinase and Cytosine Nucleoside Deaminase Activities in Synchronized Cultures of Normal Rat Kidney Cells1 CharnWingWan2andTak W. Mak Ontario Cancer Institute and Department of Medical Biophysics, university of Toronto, 500 Sherbourne Street, Toronto, Ontario, Canada M4X 1K9

ABSTRACT increased levels of cytidmnedeaminase in HeLa cell treated with ara-C. Other studies (1, 10, 23, 24, 34, 38, 40), however, Previouswork has suggestedthat 1-fi-D-arabinofurano have shown that the levels of kinase were important in sylcytosine5'-triphosphateis the active metaboliteof 1- determining the sensitivity of cells to ara-C treatment. On @-o-arabinofuranosylcytosine.Theamountof 1-/3-o-arabi the other hand it has been suggested that the nofuranosylcytosine5'-trlphosphate formed in tissues kinase:deaminase ratio could be indicative of the response has been shownto be influencedby the relativelevels of to treatment with ara-C (16, 18). These different observa deoxycytidine kinase and cytosine deaminase. In this tions are probably due to the wide variations in the kinases studywe have measuredthe intracellularlevels of deox and deaminase activities, which are known to exist among ycytidine kinase and cytosine deaminase activities in various normal and malignant tissues or cells (4, 8, 11, 18, synchronizedcultures of normal rat kidney cells. The 33). deoxycytidinekinase activftywas found to be cell cycle Since the action of ara-C is to inhibit DNA synthesis, it is relatedwith a minorpeak of activityin early G1phaseand believed to be a cell cycle-dependent drug (20, 22, 42). a major peak of activity in middleand late S phase. The Karon and Shirakawa (20, 23) and Young and Fischer (42) cytosinedeaminase activity was also found to be cycle have also determined that the action of ara-C is specifically dependent with a peak of activity at G1phase and another in the S phase of the cell cycle. at S phaseof the cell cycle. Similarresultswere obtained The purpose of this study was to measure the intracellular when cytosinedeaminase activitieswere measuredwith levels of both deoxycytidmne kinases and cytosine nucleo cytidine, deoxycytidine, or 1-$-D-arabinofuranosylcyto side deaminases in synchronized cultures of NRK cells and sine as substrate.Presentstudiesalso confirmedearlier to determine the sensitivity of these synchronized cells to studies by other workers that the main effect of 1-,3-D- ara-C. arabinofuranosylcytosineisin the late S phaseof the cell cycle. MATERIALSAND METHODS Materials. [â€˜4C]ara-C(NSC63878; specific activity, 56 INTRODUCTION mCi/mmol) was supplied by the Drug Development Branch, ara-C3 is a potent inhibitor of cellular proliferation and Cancer Chemotherapy National Services Center, National DNA synthesis in mammalian cells (6, 7, 14, 19, 25, 32, 42). Cancer Institute. The sample was diluted with unlabeled It has also been demonstrated to be effective in the treat ara-C to give the final specific activity of 0.53 mCi/mmol. [2- ment of various cancers and to be a potent agent in the I @C]Deoxycytidmne (specific activity, 29.7 mCi/mmol) was therapy of acute myeloblastic leukemia (5, 17, 27). purchased from New England Nuclear, Boston, Mass. The The mechanism of action of ara-C has been well studied radioactive compound was diluted with unlabeled deoxy and is currently believed to be one or a combination of the cytidmne to give a final specific activity of 1 mCi/mmol. following mechanisms: (a) inhibition of cellular DNA polym [â€œCjCytidine(specific activity, 500 mCi/mmol) was pur erases by ara-CTP (13, 15, 26, 30); and (b) incorporation chased from Amersham/Seanle Corp., Chicago, III. and was into cellular DNA (15, 31). diluted to give a final specific activity of 0.5 mCi/mmol. The active compound ara-CTP is formed intracellularly by Pyruvate kinase, unlabeled ara-C, deoxycytidine, cytidine, sequential action of nucleoside and nucleotide kinases (34, and phosphoenolpyruvate were purchased from Sigma 35). However, the effective concentration of the active Chemical Co., St. Louis, Mo. compound ara-CTP is also affected by the level of intracel Cells and Culture Methods. NRK cells were obtained lular cytidine deaminase, which converts ana-C to the inert from Dr. A. Bernstein (Ontario Cancer Institute, Toronto, metabolite 1-/3-D-arabinofuranosyluracil (6, 9). Canada). Cells were grown in culture in a-MEM with aspar Stewart and Burke (37) found higher levels of cytidmne agine, 50 @g/ml,and supplemented with 10% FCS (Flow deaminase in leukocytes from leukemic patients whose Laboratories, Rockville, Md.) in 75-sq cm Falcon tissue disease did not respond to ara-C than in cells from patients culture flasks. The cultures were maintained at 37Â°in sensitive to the agent, and Meyers et a!. (29) have described humidified atmosphere supplemented with 5% CO2. Cell SynchronizationProcedure. NRK cells were syn chronized by a modified serum deprivation method previ I This work is supported by a grant from the Medical Research Council of ously described by Farmilo and Stanners (12). In short 2.5 Canada. 2 Recipient of a studentship from the National Cancer Institute of Canada. x 10@cells in 25 ml of a-MEM supplemented with 0.75% 3 The abbreviations used are: ara-C, 1-$-D-arabinofuranosylcytosine; ara FCS were seeded in a 150-sq cm tissue culture flask. After CTP,1-@-D-arabinofuranosylcytosine5'-triphosphate;NRK,normalrat kid nay; a-MEM, minimal essential medium (Eagle's); FCS, fetal calf serum. 3 days at this low serum concentration, the medium was Received November 29, 1977; accepted June 5, 1978. replaced by fresh a-medium containing 10% FCS. The

2768 CANCER RESEARCHVOL. 38

Downloaded from cancerres.aacrjournals.org on September 25, 2021. © 1978 American Association for Cancer Research. Kinaseand Deaminasein Synchronized NRK Cells addition of the serum stimulates entry of the cells into the formed after 8 days of incubation at 37Â°in a humidified G, phaseofthecellcycle(39). atmosphere with 5% CO2. DNA Synthesis.The rate of DNA synthesisafterserum stimulation was determined by incorporation of RESULTS [3H]thymidmneinto acid-insoluble material. For pulsing a 20- l.Ll aliquot of [3H]thymidine (47 Ci/mmol; Amersham/Searle) SynchronizationofNRK Cells. NRKcellswere synchro was addedto 1.6 ml of medium(30-mmx 10-mm Falcon nized by serum deprivation as described in â€œMaterialsand Petni dish) for 30 mm. The cells were then washed 3 times Methods.â€•The degree of synchrony of these cells was with 5 ml cold phosphate-buffered saline (0.02 M measured by determining the rates of DNA synthesis with Na@HPO4:0.14MNaCI, pH 7.2), trypsinized, and precipitated [3H]thymidmne at various times after addition of serum. As with 10% cold tnichloroacetic acid. The precipitate was can be seen in Chart 1, the addition of serum resulted in a collected on Millipore membrane filters and washed 3 times peak of DNA synthesis 9 to 20 hr later. The number of cells with 30 ml of 10% trichloroacetic acid, washed once with 10 per plate was also determined to assess the synchrony of ml of cold ethanol (95%), and dried. Radioactivity was the cultures. As illustrated in Chart 1, the number of cells monitored in 10-mi Econofluor (New England Nuclear, per plate remained fairly constant for the first 14 hr, after Boston, Mass.) in a Beckman LS-230 scintillation spectnom which it increased gradually to approximately double the eter. initial cell number by 25 hr. The DNA synthesis and the cell PreparationofCell Extracts.Extractsofcellsuspensions doubling profiles are highly reproducible from experiment for enzymatic analysis were prepared as described by Ho to experiment. A lag time of 13 to 15 hr of DNA synthesis (18). In short, cells were trypsmnized and washed twice with and doubling time of 24 to 25 hr were seen in 8 independent ice-cold 0.9% NaCI solution. The cells were then resus experiments. pended at 2 x 10. cells/mI in cold sucrose (0.25 M) and Deoxycytidine Kinase Activity in Synchronized NRK sonically disrupted for 30 sec in a MSE sonic oscillator. The Cells. The level of deoxycytidmne kinase as a function of extract was then centrifuged for 1 hr at 130,000 x g in a 50 time after serum stimulation was measured. At various Ti rotor at 4Â°,and the supennatant was removed for enzy times after the addition of serum, cells were trypsinized, matic analysis of the kinase and deaminase. Protein con extracts were prepared, and the deoxycytidmne kinase activ centrations were determined by the method of Lowry et a!. ities were assayed . The results of 3 independent expeni (28). ments are illustrated in Chart 2. The enzymatic activity is DeaminaseEnzymeAssays.Cytidinenucleosidedeami cell cycle related with a reproducible minor peak of activity nase enzyme activities were measured by a modification of in early G, (about 5 hr after the addition of serum) and a the method described by Ho (18). The reaction mixture major peak of activity in middle and late S (between 15 and contained Tnis buffer, pH 8.0, 40 @.&mol;[â€˜4C]deoxycytidmne, 20 hr). [â€˜@C]cytidine,or[1@C]ara-C,60 nmol; and cell-free superna Cytosine Nucleoside Deaminase Activity in Synchro tant to give a total volume of 100 @I.Thereaction mixture nized NRK Cells. The intracellularconcentrationof the was incubated at 37Â°for 60 mm, and the reaction was active compound ara-CTP found inside a cell is known to terminated by putting the mixture into boiling water for 2 be affected by the cellular cytosine nucleoside deaminase, mm. The precipitate was removed by centnifugation at 1000 x g for 10 mm. Fifty @Iofthesupernatant fluid were spotted on Whatman No. 1 filter paper and subjected to ascending chromatography for 18 hr with a solvent system consisting of isopropyl alcohol:water:ethyl acetate (22.5:12.5:65). After the chromatogram was dried, the spots corresponding to the appropriate compounds were cut out. The paper was suspended in 10 ml of Econofluor, and the radioactivity was measured. Deoxycytidine Kinase Assay. A modified method of Ho (17) was used. The reaction mixture contained Tnis buffer, pH 8.0, 40 @moI;Mg,ATP, 4 @mol;phosphoenolpyruvate, 2 @mol;phosphenolpyruvate kinase, 6 units; [14C]deoxy cytidine, 60 nmol; and the cell-free supernatant to a final volume of 100 @l.Thereaction mixture was incubated at

370 for 60 mm. This method of separating and identifying the radioactive peaks (deoxycytidine and phosphorylated deoxycytidmne) was similar to those described for the deaminase assay. Plating Efficiencies.The survivalof NRK cells after ex 0 0 20 28 Time otter odd.t.on of serum (hr) posure to ara-C was estimated by the ability of the cells to Chart1. DNAsynthesisandcellnumbersofNRKcultureswithtimeafter form a colony. Cells were trypsinized and counted by serum addition. NRK cells were seeded at low FCS level (0.75%) for 3 days. hemacytometer, and 200 cells were plated in each Petni The medium was then replaced by fresh medium containing 10% serum. The dish containing 5 ml a-MEM plus 10% FCS. The number of rate of DNA synthesis was determined by incorporation of (3Hjthymidine for 30 mm and cell number per plate was determined by the use of a hemacy surviving cells was estimated by the number of colonies tometer. All values were determined in quadruplicate samples.

SEPTEMBER 1978 2769

which is capable of converting ara-C to the inert metabolite 1-f3-D-arabinofuranosyluracil (6, 9). The intracellular level of deoxycytidine deaminase was also measured in these syn chronized cells. The results of 2 independent experiments are illustrated in Chart 3. There are 2 peaks of enzyme activities, one occurring in G1phase and the other at the S phase of the cell cycle. Similar experiments were also I performed with cytidine or ara-C as substrates. The results of these studies, illustrated in Chart 4, were similar to those with deoxycytidine as substrate, suggesting that similar if @03I not identical enzymes are responsible for the deamination of these compounds. Effectof ara-C on the Survivalof SynchronizedCultures of NRKcells. Theeffectof ana-Conthesurvivalofsynchro nized cultures of NRK cells was also determined. Cells were synchronized by serum deprivation. At different times after the addition of serum, cells were treated with 5 x 10@ M of ara-C for 2 hr, and the survival of the NRK cells was determined by measurement of their plating efficiency. The results are illustrated in Chart 5. ara-C has little or no effect on the survival of the cells when the drug was present during the first 16 hr. The main effect of ara-C was during the S phase of the cell cycle, between 16 and 22 hr after the Time after oddition of serum (hr) addition of serum. The plating efficiency of untreated NRK Chart 3. Levels of deoxycytidine deaminase activities in synchronized cells is approximately 50% of input cells. Although the cell cultures of NRK cells. Cells were synchronized, extracts were prepared, and nucleoside deaminase activities were measured as described in â€˜Materials cycle time varies slightly from experiment to experiment, and Methodsâ€•withdeoxycytidineassubstrate.Resultsareshownof 2 the main killing effect of the ara-C was always observed independentexperiments. after the peak of DNA synthesis.

. Cytidine

@ @A1TYT'\@

@ 0.2

@ 0.I

â€˜i@ I E@

â€¢Câ€¢@\ @4 0.2 Aro - C S

i\ â€¢/\ @@â€˜\ 0. I \5@5/5/ v:31 â€˜I 0 5 10 5 20 25 30 Time (hr) Chart4. Levelsof nucleosidedeaminaseactivities in synchronizedcul 3 tures of NRK cells with cytidine, deoxycytidine, or ara-C as substrate. See legendtoChart3.

2 â€¢@ /â€˜ \ @@@ â€¢â€˜â€¢@@â€¢:5 25@30 DISCUSSION

Time after additionof serum (hr) In an effort to understand more clearly the metabolism of Chart 2. Levels of deoxycytidine kinase activities in synchronized cultures ara-C, we have studied the levels of deoxycytidine kinase of NRK cells. Cells were synchronized as described in â€œMaterialsand and cytosine nucleoside deaminase in synchronized cells. Methodsâ€•.At various times after addition of serum, cells were trypsinized anddeoxycytidinekinaseactivitiesweremeasuredasdescribedin â€œMaterialsThere are 2 peaks of enzymatic activities for the deoxycyti and Methodsâ€•.Results are shown of 3 independent experiments. dine kinase, one in early G, phase and another in S phase.

2770 CANCER RESEARCH VOL. 38

too REFERENCES

1. Barranco, S. C., Ho., D. H. W., Drewinko, B., Romsdahl, M. M., and 80 Humphrey, R. M. Differential Sensitivity of Human Melanoma Cells Grown in Vitro to Arabinosylcytosine. Cancer Res., 32: 2733-2736, 1972. 2. Benedict,W. F., Harris,N., and Karon,M. Kineticsof 1-@-D-Arabinofur @60 anosylcytosine-induced Chromosome Breaks. Cancer Res.. 30: 2477- ,@ 2483, 1970. 3. Bray, G. and Brent, T. P. Deoxyribonucleoside 5'-Triphosphate Pool ,@ 40 Fluctuations during the Mammalian Cell Cycle. Biochim. Biophys. Acta, 269: 184-191, 1972. 4. Camiener, G. W., and Smith, C. G. Studies of the Enzymatic Deamination 20 of Cytosine Arabinoside. I. Enzyme Distribution and Species Specificity. Biochem.Pharmacol.,14:1405-1406,1965. 5. Chabner, B. A., Myers, C. E., Coleman, C. N., and Johns, D. G. The 0 Clinical Pharmacology of Antineoplastic Agents (Part 2). New EngI. J. Med., 292: 1159-1168, 1975. Time after addition ofserum (hr) 6. Chu, M. Y., and Fischer,G. A. A ProposedMechanismof Action of 1-13- Chart 5. The effect of ara-Con the survivalof synchronizedNRKcells. D-ArabinofuranosyI-Cytosir@eas an Inhibitor of Growth Leukemic Cells. NRKcells releasedinto cell division by serumadditionwereexposedto 5 x Biochem. Pharmacol., 11: 423-430, 1962. 10 N ara-Cfor 2-hr pulsesand washed3 times.Thesurvivalof thesecells 7. Cohen, S. S. Introduction to Biochemistry of D-Arabinosyl Nucleosides. wasestimatedfrom their platingefficiencies(seeâ€˜MaterialsandMethodsâ€•). Progr. Nucleic Acid Mol. Biol., 5: 1-88, 1966. The plating efficiencyof untreatedNRKcells is approximately50%of input 8. Coleman, C. N., Stoller, R. G., Draka, J. C., and Chabner, B. A. Deoxycytidine Kinase: Properties of the Enzyme from Human Leukemic cell numbers. Granulocyte. Blood, 46: 791-803, 1975. 9. Creasey, W. A., Papac, R. J., Markiw, M. E., Cababresi, P., and Welch, A. It is possible that the first peak of kinase activity is D. Biochemical and Pharamacologicab Studies With 1-13-D-Arabinofura nosylcytosine In Man. Biochem. Pharmacol., 15: 1417-1428, 1966. responsible for the increased pool sizes of the deoxynibo 10. Drahovsky, D., and Kresis, W. Studies of Drug Resistance. II. Kinase nucleotide triphosphate at late G1 (1, 41) and that the Patternsin P 815NeoplasmsSensitiveand Resistantto 1-f3-D-Arabino second peak is responsible for the continuing expansion of furanosylcytosine. Biochem. Pharmacol.. 19: 940â€”944.1969. 11. Durham, J. P., and Ives, D. H. Deoxycytidine Kinase. I. Distribution in the pools of the deoxyribonucleotide triphosphate in the S Normaland NeoplasticTissuesand Interrelationshipsof Deoxycytidine phase (3, 36, 41). and 1-19-D-ArabinofuranosylcytosinePhosphorylation.Mob.Pharmacol., The levels of cytosine nucleoside deaminase are also 5:358-375,1969. 12. Farmilo,A. J., and Stanners,C. P. Mutant of VSVWhich Allows DNA found to have 2 peaks with one in middle G, phase and Synthesis and Division in Cells Synthesizing Viral RNA. J. Virol., 10: 605- another broad peak in S phase. The significance of these 611.1973. 13. Furth, J. J., and Cohen, S. 5. Inhibition of Mammalian DNA Pobymerase peaks of activity is not known. They may be required for the by the 5'-Triphosphateof 1-fl-D-Arabinofuranosylcytosineandthe 5'- synthesis of precursors of RNA and DNA. Although it has Triphosphate of 9-13-o-Arabinofuranosyladenine. Cancer Res., 28: 2061- been suggested that the intracellular concentrations of the 2067,1968. 14. Graham, F. L., and Whitmore, G. G. The Effect of 1-f3-D-Arabinofurano active compound ara-CTP are affected by the cellular cyto sylcytosine on Growth, Viability and DNA Synthesis in Mouse L-celbs. sine nucleoside deaminase, it is not known whether the CancerRes.,30: 2627-2635,1970. enzyme responsible is a cytidine deaminase or deoxycyti 15. Graham,F. L., and Whitmore,G. F. Studies in Mouse-L-Cellsonthe Incorporation of 1-$-D-Arabinofuranosylcytosine 5'-Triphosphate. Can dine deaminase. We have also determined the levels of the car Rae., 30: 2636-2644, 1970. deaminase activities with either cytidine, deoxycytidmne, or 16. Hart,J. S., Ho,D. H., George,S. L., Salem,P., Gottlieb,J. A., and Frei, E., III. Cytokinetic and Molecular Pharmacology Studies of Arabinosyl ara-C as substrate, and similar results were obtained cytosine in Metastatic Melanoma. Cancer Rca., 32: 2711-2716, 1972. throughout the cell cycle, suggesting that similar if not 17. Henderson, E. S., and Burke, P. J. Clinical Experience with Cytosine identical enzymes are responsible for the deamination of Arabinoside. Proc. Am. Assoc. Cancer Res., 6: 26-37, 1965. 18. Ho,D. H. W. Distributionof Kinaseand Deaminaseof1-13-o-Arabinofur these compounds. anosylcytosine in Tissues of Man and Mouse. Cancer Res., 33: 2816- These data in the present study also confirm the early 2820, 1973. studies by others that the main effect of ara-C is in the S 19. Jones, P. A., Baker, M. S., and Benedict, W. F. The effect of 1-13-0- Arabinofuranosylcytosine on Call Viability, DNA Synthesis and Chroma phase of the cell cycle (22, 42) and that the peak of tid BreakageInSynchronizedHamsterFibrosarcomaCells.CancerRes., cytotoxicity occurred toward the latter half of the S phase 36: 3789â€”3797,1976. 20. Karen, M., Benedict, W. F., and Rucker, N. Mechanism of 1-13-D-Arabi (21). The basis of the S-phase cytotoxicity of ana-C is nofuranosylcytosine Induced Cell Lethality. Cancer Res., 32: 2612-2615, presumably due to the action of its phosphorylated deniva 1972. tive ara-CTP on the DNA polymenases, incorporation into 21. Karen,M., Momparler,R. L., and Benedict,W. F. RelevanceofMolecu lar Mechanismof Drug Action to Clinical Trials In Man: Combination DNA and chain termination, that resulted in the inhibition Therapy Model using Are-C and Aza-C. In: Pharmacological Basis of of DNA synthesis (13-15, 26, 30, 31). However, this is CancerChemotherapy,N. D. AndersonHospitaland Tumor Institute, probably not the complete explanation, since the inhibition pp. 329-339. Baftimore: Williams & Wilkins, 1975. 22. Karon, M., and Shirakawa, S. The Locus of Action of 1-$-D-Arabinosyl of DNA synthesis per se is not correlated with cell death cytosine in Cell Cycle. Cancer Res., 29: 687-696, 1969. (19-21). Two additional factors affecting the action of ara-C 23. Kessel, D., Hall, T. C., and Rosenthal, D. Uptake and Phosphorybation of CytosineArabinosideby Normaland LeukemicHumanBlood Cells in should be considered. A decreased ability to repair suble Vitro. Cancer Res., 29: 459-493, 1969. thaI chromatid breakage during late S phase (2, 20) and 24. Kessel,D., Hall,T. C., and Wodinsky,I. Transportand Phosphorylation the relative concentrations of intracellular dCTP and ara as Factors In the Antitumor Action of CytosineArabinoside.Science 156: 1240-1241, 1967. CTP mediated by the cell cycle may also be important. 25. Kim,J. H., and Eldinoff, M. L. Action of 1-fl-D-Arabinofuranosylcytosine on the NucleicAcid MetabolismandViabilityof HeLaCells.CancerRes., 25: 698-702, 1965. ACKNOWLEDGMENTS 26. Kimball,A. P., andWilson,M. I. Inhibitionof DNAPolymeraseby1-13-D- Arabinosylcytosine and Reversal of Inhibition by Deoxycytidine 5'-Tri We would like to thank the Drug DevelopmentBranch of the Cancer phosphate. Proc. Soc. Exptl. Bid. Med., 127: 429-432, 1968. Chemotherapy National Services Center, National Cancer Institute, for sup 27. Livingston, R. B., and Carter, S. K. Cytosine Arabinoside Clinical plying [â€˜C]ara-Cfor this study. Brochure. Cancer Chemotherapy Rept., 1:179-205,1968.

SEPTEMBER 1978 2771

28. Lowry,0. H., Rosebrough,N.J., Farr,A. L., and Randall,R.J. Protein 28: 793-801, 1968. Measurementwiththe Folin PhenolReagent.J. Biol. Chem.,193:265- 36. Skoog, K. L., Nordenskjold, B. A., and Bjursell, K. G. Deoxyribonucleo 275,1951. sidetriphosphate Pools and DNA Synthesis in Synchronized Hamster 29. Meyers, R., Malathi, V. G., Cox, R. P., and Silber, R. Studies on Cells. European J. Biochem. 33: 428-432, 1973. NucleosideDeaminase.J.Biol. Chem.,248:5909-5913,1973. 37. Stewart,C. D., and Burke, P. J. Cytidine Deaminaseandthe Develop 30. Momparler, R. L. Effect of Cytosine Arabinoside 5'-Triphosphate on ment of Resistance to Arabinosyb Cytosine. Nature New Blob., 233: 109- Mammalian DNA Polymerase. Blochem. Blophys. Rae. Commun., 34: 110,1971. 46@-471,1969. 38. Tattersall, M. H., Ganeshaguru, K., and Hoffbrand. A. V. Mechanisms of 31. Momparlr, R. L. KineticandTemplateStudieswith 1-$-D-Arabinofura Resistance of Human Acute Leukemia Cells to Cytosine Arabinoside. nosylcytosine 5'-Triphosphate and Mammalian Deoxyribonucleic Acid Brit. J. Haematol., 27: 39-46, 1974. Polymerase. Mob. Pharmacol., 8: 362-370, 1972. 39. Temin, H. M. Stimulation by Serum Multiplication of Stationary Chicken 32. Moore, E. C., and Cohen, 5. 5. Effects of Arabinonucleotideson Cells. J. Cellular Physiol., 78: 161-170. 1971. Ribonucleotide Reduction by an Enzyme System from Rat Tumor. J. 40. Uchida, K., and Kreis, W. Studeas on Drug Resistance. I. Distribution of Blob.Chem.,242:2116-2118,1967. 1-13-D-ArabinofuranosylCytosine,Cytidine and Deoxycytidinein Mice 33. Niho, Y., Till, J. E., and McCulboch, E. A. Effect of Arabinosylcytosine on Bearing Ara-C Sensitive and Resistant P815 Neoplasms. Biochem. Phar Granulopoietic Colony Formation by Marrow Cells from Leukemic and macol., 18: 1115-1128, 1969. Nonleukemic Patients. Exptl. Hemato., 4: 63-69, 1976. 41. Walters, R. A., Tobey, R. A., and Ratliff, R. L. Cell Cycle-Dependent 34. Schrecker, A. W. Metabolism of 1-@-D-arablnofuranosylcytosine in Leu Variations of Deoxyribonucleoside Triphosphate Pools In Chinese Ham kemia L1210 Nucleoside and Nucbeotide Kinases in Cell Free Extract. ster Cells. Biochim. Blophys. Acta, 319: 336-347, 1973. CancerRes.,30: 632-641,1970. 42. Young, R. S. K., and Fisher,G. A. TheAction of Arabinosylcytosineon 35. Schrecker, A. W., and Urshel, M. J. Metabolism of 1-@-o-Arabinofurano Synchronously Growing Population of Mammalian Cells. Biochem. Bio sylcytosine in Leukemia L1210 Studies with Intact Cells. Cancer Rca., phys.Rca.Commun.,32:23-29,1969.

2772 CANCER RESEARCHVOL. 38

Downloaded from cancerres.aacrjournals.org on September 25, 2021. © 1978 American Association for Cancer Research. Deoxycytidine Kinase and Cytosine Nucleoside Deaminase Activities in Synchronized Cultures of Normal Rat Kidney Cells

Charn Wing Wan and Tak W. Mak

Cancer Res 1978;38:2768-2772.

Updated version Access the most recent version of this article at: http://cancerres.aacrjournals.org/content/38/9/2768

E-mail alerts Sign up to receive free email-alerts related to this article or journal.

Reprints and To order reprints of this article or to subscribe to the journal, contact the AACR Publications Subscriptions Department at [email protected].

Permissions To request permission to re-use all or part of this article, use this link http://cancerres.aacrjournals.org/content/38/9/2768. Click on "Request Permissions" which will take you to the Copyright Clearance Center's (CCC) Rightslink site.