Abstract
It is generally assumed that drug concentration does not change significantly under cell culture conditions. Nevertheless, most of the therapeutic trials in acute leukemia that were based on in vitro drug sensitivity assays of patient samples have been disappointing. In order to show possible pitfalls of unphysiological alterations in vitro we investigated concentration versus time curves, metabolism and effects on the culture media for some antineoplastic drugs.
Oxazaphosphorines and cytarabine were incubated in RPMI and in established cell lines and measured by HPLC. HPLC also served to measure enzyme activity and levels of related amino acids at various concentrations of asparaginase, ammonia release was photometrically determined. Etoposide was monitored by HPLC relative to different contents of FCS in RPMI.
All oxazaphosphorines showed a rapid decrease of in vitro activity down to about 10% within 4–6 h, and 2% within 72 h. The level of cytarabine, when incubated in RPMI, was stable over 24h, and no change was seen with K562, while a rapid decrease to below 50% occurred within 6h in the presence of HL 60 and BLIN. 2 U/L of asparaginase led to asparagine depletion of the medium within 4h, while 200 U/L were associated with a preferential increase of glutamic acid and ammonia. Further, there was evidence of instability by rapid adsorption to plastic surfaces (paclitaxel) or isomerisation (etoposide) in RPMI with low FCS content.
The instability of drugs in vitro is attributed to a variety of different factors: i.e. physico-chemical instability results in inactivation of oxazaphosphorines, cytarabine dissappears by cellular metabolism without saturation depending on the cell-line. Epiphenomena like adsorption and isomerisation in vitro are unphysiological.
Results of drug sensitivity assays should be interpreted with great caution.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Preview
Unable to display preview. Download preview PDF.
Similar content being viewed by others
References
Pieters R., Loonen A.H:, Huismans D.R., Broekema G.J., Dirven M.W.J., Heyenbrok M.W., Hählen K., Veerman AJ.P. In vitro drug sensitivity of cells from children with leukemia using the MTT assay with improved culture conditions. Blood 1990, 76, 2327–2336
Pieters R., Kaspers G.J.L., van Wering E.R., Huismans D.R., Loonen A.H:, Hählen K., Veerman A.J.P. Cellular drug resistance profiles that might explain the prognostic value of immunophenotype and age in childhood acute lymphoblastic leukemia. Leukemia 1993, 7, 392–397
Weisenthal, L.M., Lippmann, M.E.: Clonogenic and nonclonogenic in vitro chemosensitivity assays. Cancer Treat Rep 1993 69:615–624
Selby, P, Buick R.N., Tannock I.A.A., A critical appraisal of the “human tumor-stem cell assay” N Engl J Med 1983, 308:129–134
Boos J., Küpker F., Blaschke G, Jürgens H. Trofosfamide metabolism in different species-ifosfamide is the predominant metabolite. Cancer Chemother Pharmacol 1993, 33:71–76
Wörmann B, Anderson JM, Liberty JA, Gajl-Peczalska K, Brunning RD, Silbermann TL, Arthur DC, LeBien TW. Establishement of a leukemic cell modell for studying human pre-B to B cell differentiation. J Immunol 1989, 142:110–117
Kaspers, G.J.L., Veerman A.J.P., Pieters R., Broekema G.J., Huismans D.R., Kazmier K.M., Loonen A.H., Rottier M.A.A., Van Zantwijk C.H., Hählen K., Van Wering E.R. Mononuclear cells contaminating acute lymphoblastic leukaemic samples tested for cellular drug resistance using the methyl-thiazol-tetrazolium assay. Br J Cancer, 1994, 70:1047–1052
Woollard, G.A. An unusual column effect during analysis of 5-fluorocytosine by high-performance liquid chromatography. J of Chromatogr 1986, 368:162–163
Boos J, Werber G, Ahlke E, Schultze-Westhoff P, Nowak-Göttl U, Würthwein G, Verspohl EJ, Ritter J, Jürgens H. Monitoring of asparaginase activities and asparagine levels in children on different asparaginase preparations. Eur J Cancer 1996, 32:1544–1550
Heinzel G, Woloszcak R and Thomann P. Dr. Karl Thomae GmbH, Schering AG, Gödecke AG, Stuttgart;Jena; New York G. Fischer, 1993
Liliemark, E., Petterson B, Peterson C., Liliemark J. High-performance liquid chromatography with fluorometric detection for monitoring of etoposide and its cis-isomer in plasma and leukaemic cells. J of chromatography B: biomedical applications 1995, 669:311–317
Sindermann, H, Peukert M, Hilgard P. Bone marrow purging with mafosfamide-A critical survey. Blut 1989, 59:432–441
Pieters, R., Loonen A.H:, Huismans D.R., Broekema G.J., Dirven M.W.J., Heyenbrok M.W., Hählen K., Veerman AJ.P. In vitro drug sensitivity of cells from children with leukemia using the MTT assay with improved culture conditions. Blood, 1990, 76, 2327–2336
Capizzi R. L., Bertino J.R., Skeel R.T., Creasey W.A., Zanes R., Olayon C., Peterson R.G., Handschumacher R.E. L-Asparaginase: clinical, biochemical, pharmacological and immunological studies. Ann Intern Med 1971, 74, 893–904
Cooney DA, Handschumacher RE L-asparaginase and L-asparagine metabolism. Annu Rev Pharmacol 1970, 10, 421–440
Kaspers G.J.L., Pieters R., Van Zantwijk C.H., De Laat P.A.J.M., De Waal F.C., Van Wering, Veerman AJ.P. In vitro drug sensitivity of normal peripheral blood lymphocytes and childhood leukaemic cells from bone marrow and peripheral blood. Br J Cancer 1991, 64, 469–474
Asselin, BL, Ryan D, Frantz CN, Bernai SD, Leavitt P, Sallan SE and Cohen HJ. In vitro and in vivo killing of acute lymphoblastic leukemia cells by Asparaginase. Cancer Res 1989, 49, 4363–4371
Ertel IJ, Nesbit ME, Hammond D, Weiner J and Sather H. Effective dose of Asparaginase for induction of remission in previously treated children with acute lymphoblastic leukemia: a report from Childrens Cancer Study Group Cancer Res. 1979, 39, 3893–3896
Mader R.M., Steger, GG, Moser K, Rainer H., Krenmayr P., Dittrich C. Instability of the anticancer agent etoposide under in vitro culture conditions, Cancer Chemother Pharmacol 1991, 27, 354–360
Yang L.Y., Drewinko B. Cytotoxic efficacy of reconstituted and stored antitumor agents. Cancer Res 1985, 45, 1511–1515
Evans W.E., Sinkule J.A:, Crom W.R., Dow L.W., Look A.T., Rivera G. Pharmacokinetics of teniposide (VM 26) and etoposide (VP 16) in children with cancer. Cancer Chemother Pharmacol 1982, 7, 147–150
Ludwig R., Alberts D.S., Chemical and biological stability of anticancer drugs used in a human tumor clonogenic assay. Cancer Chemother Pharmacol 1984, 12, 142–145
Lanvers C, Jürgens H, Blaschke G., Boos J. Non-enzymatic isomerisation of retinoic acids during incubation in RPMI 1640 medium. poster presented at 9th NCI-EORTC Symposium on New Drugs in Cancer Therapy, 1996
Song, D., Hsu, L.-F., Jessie, L.-S. Binding of taxol to plastic and glass containers and protein under in vitro conditions, J Pharm Sci 1996, 85, 29–31
Liebmann JE, Cook, J.A., Lipschultz C., Teague D., Fisher J., Mitchell JB. Cytotoxic studies of paclitaxel (Taxol) in human tumour cell lines. Br J Cancer 1993, 68, 1104–1109
Ross, D.D., Wooten P.J., Tong Y., Cornblatt B., Levy C., Sridhara R., Lee E.J., Schiffer C.A. Synergistic reversal of multidrug-resistance phenotype in acute myeloid leukemia cells by cyclosporin A and cremophor EL. Blood 1994, 83, 1337–1347
Dudeja P.K., Anderson, K.M., Harris J.S., Buckingham L., Coon J.S. Reversal of multidrug resistance phenotype by surfactants: relationship to membrane lipid fluidity. Arch Biochem Biophys 1995, 319, 309–315
Fulda S., Honer, M., Menke-Moellers, I, Berthold, F. Antiproliferative potential of cytostatic drugs on neuroblastoma cells in vitro. Eur J Cancer 1995, 31, 616–621
Beijnen JH, Wiese G, Underberg WJ Aspects of the chemical stability of Doxorubicin and seven ohter anthracyclines in acidic solution. Pharm Weekbl Sci 1985, 7, 109–116
Bachur NR, Gee M, Daunorubicin metabolism by rat tissue preparations. J Pharmacol Exp Ther 1971, 177, 567–572
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 1999 Springer Science+Business Media New York
About this chapter
Cite this chapter
Wagner, A., Hempel, G., Gumbinger, H.G., Jürgens, H., Boos, J. (1999). Pharmacokinetics of Anticancer Drugs In Vitro. In: Kaspers, G.J.L., Pieters, R., Veerman, A.J.P. (eds) Drug Resistance in Leukemia and Lymphoma III. Advances in Experimental Medicine and Biology, vol 457. Springer, Boston, MA. https://doi.org/10.1007/978-1-4615-4811-9_42
Download citation
DOI: https://doi.org/10.1007/978-1-4615-4811-9_42
Publisher Name: Springer, Boston, MA
Print ISBN: 978-1-4613-7180-9
Online ISBN: 978-1-4615-4811-9
eBook Packages: Springer Book Archive