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Clonogenic assay with established human tumour xenografts

correlation of in vitro to in vivo activity as a basis for anticancer drug discovery
  • H.H Fiebig
    Correspondence
    Corresponding author. Tel.: +49-761-51559-14; fax: +49-761-51559-55
    Affiliations
    Institute for Experimental Oncology, Oncotest GmbH, Am Flughafen 12–4, D-79108 Freiburg, Germany

    Tumor Biology Center, At the University of Freiburg, Clinic for Medical Oncology, Breisacher Str. 117, D-79106 Freiburg, Germany
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  • A Maier
    Affiliations
    Institute for Experimental Oncology, Oncotest GmbH, Am Flughafen 12–4, D-79108 Freiburg, Germany
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  • A.M Burger
    Affiliations
    Tumor Biology Center, At the University of Freiburg, Clinic for Medical Oncology, Breisacher Str. 117, D-79106 Freiburg, Germany
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      Abstract

      Pluripotent cells can be grown in clonogenic assays. The tumour stem-cell fraction, which accounts for <0.4% of the total cells, and which is considered the most relevant cell type in the development of metastases and recurrences, is able to divide and to form colonies in a semisolid matrix (agar or methylcellulose). Major applications of the tumour clonogenic assay (TCA) are chemosensitivity testing of tumours and xenografts, and for assessments within drug discovery programmes. Of critical relevance for the usefulness of the TCA is whether it can predict sensitivity or resistance towards clinically used agents. When we compared the response of human tumours established as xenografts in nude mice in the TCA in vitro to that of the clinical response, 62% of the comparisons for drug sensitivity, and 92% of the comparisons for drug resistance were correct. The same percentage of true/false observations was found when tumours were tested after serial passage in nude mice in the TCA in vitro and their response compared to in vivo activity in corresponding xenografts (60% and 90%, respectively). The highest correct predictive values were, however, found when the clinical response of tumours was compared to their explants established in the nude mouse and treated in vivo. Of 80 comparisons performed, we observed a correct prediction for tumour resistance in 97% and for tumour sensitivity in 90%. In our opinion, the TCA with established human tumour xenografts has an important role in current drug discovery strategies. We therefore included the TCA as secondary assay in our approach to anticancer drug discovery and found that a number of novel agents were active; these are now in advanced preclinical development or clinical trials. Thus, the tumour clonogenic assay has proven predictive value in the chemosensitivity testing of standard and experimental anticancer drugs.

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      References

        • Hamburger A.W
        • Salmon S.E
        Primary bioassay of human tumor stem cells.
        Science. 1977; 197: 461-463
        • Von Hoff D.D
        Human tumor cloning assays.
        Cancer Metastasis Rev. 1988; 7: 357-371
        • Scholz C.C
        • Berger D.P
        • Winterhalter B.R
        • Henss H
        • Fiebig H.H
        Correlation of drug response in patients and in the clonogenic assay using solid human tumor xenografts.
        Eur. J. Cancer. 1990; 26: 901-905
        • Engelholm S.A
        • Vindelv L.L
        • Spang-Thomsen M
        • et al.
        Genetic instability of cell lines derived from a single human small-cell carcinoma of the lung.
        Eur. J. Cancer Clin. Oncol. 1985; 21: 815-824
        • Ferguson P.J
        • Cheng Y.C
        Phenotypic instability of drug sensitivity in a human colon carcinoma cell line.
        Cancer Res. 1989; 49: 1148-1153
        • Smith A
        • van Haaften Day C
        • Russell P
        Sequential cytogenetic studies in an ovarian cancer cell line.
        Cancer Genet. Cytogenet. 1989; 38: 13-24
        • Fiebig H.H
        • Schmid J.R
        • Bieser W
        • Henss H
        • Löhr G.W
        Colony assay with human tumor xenografts, murine tumors and human bone marrow. Potential for anticancer drug development.
        Eur. J. Cancer Clin. Oncol. 1987; 23: 937-948
        • Salmon S.E
        Human tumor colony assay and chemosensitivity testing.
        Cancer Treat Rep. 1984; 68: 117-125
        • Schlunk T
        • Schleyer M
        The influence of culture conditions on the production of colony-stimulating activity by human placenta.
        Exp. Hematol. 1980; 8: 179-184
        • Gallacher L
        • Murdoch B
        • Wu D
        • Karanu F
        • Fellows F.N
        • Bhatia M
        Identification of novel circulating human embryonic blood stem cells.
        Blood. 2000; 96: 1740-1747
        • Gallacher L
        • Murdoch B
        • Wu D
        • Karanu F.N
        • Keeney M
        • Bhatia M
        Isolation and characterization of human CD34Lin and CD34+Lin hematopoietic stem cells using cell surface markers AC133 and CD7.
        Blood. 2000; 95: 2813-2820
        • Xu R
        • Reems J.A
        Umbilical cord blood progeny cells that retain a CD34+ phenotype after ex vivo expansion have less engraftment potential than unexpanded CD34+ cells.
        Transfusion. 2001; 41: 213-218
      1. Fiebig HH, Berger DP, Dengler WA, Wallbrecher E, Winterhalter BR. Combined in vitro/in vivo test procedure with human tumor xenografts for new drug development. In Fiebig HH, Berger DP, eds. Immunodeficient Mice in Oncology. Contrib Oncol Basel, Karger, 1992, 42, 321–351.

        • Maurer H.R
        • Ali-Osman F
        Tumor stem cell cloning in agar-containing capillaries.
        Naturwissenschaften. 1981; 68: 381-383
        • Hanauske A.R
        • Hanauske U
        • Buchok J
        • Von Hoff D.D
        Recombinant human transforming growth factor-alpha stimulates in vitro colony formation of fresh human tumor specimens.
        Int. J. Cell Cloning. 1988; 6: 221-229
        • Bertelsen C.A
        • Sondak V.K
        • Mann B.D
        • Korn E.L
        • Kern D.H
        Chemosensitivity testing of human solid tumors. A review of 1582 assays with 258 clinical correlations.
        Cancer. 1984; 53: 1240-1245
        • Cole S.P
        Rapid chemosensitivity testing of human lung tumor cells using the MTT assay.
        Cancer Chemother. Pharmacol. 1986; 17: 259-263
        • Carmichael J
        • DeGraff W.G
        • Gazdar A.F
        • Minna J.D
        • Mitchell J.B
        Evaluation of a tetrazolium-based semiautomated colometric assay.
        Cancer Res. 1987; 47: 936-942
        • Hoffman R.M
        Three-dimensional histoculture.
        Cancer Cells. 1991; 3: 86-92
        • Hoffman R.M
        In vitro sensitivity assays in cancer.
        J. Clin. Lab. Anal. 1991; 5: 133-143
        • Furukawa T
        • Kubota T
        • Hoffman R.M
        Clinical applications of the histoculture drug response assay.
        Clin. Cancer Res. 1995; 1: 305-311
        • Inaba M
        • Tashiro T
        • Sato S
        • Ohnishi Y
        • Tanisaka K
        • Kobayashi H
        In vitro-in vivo correlation in anticancer drug sensitivity test using AUC-based concentrations and collagen gel-droplet-embedded culture.
        Oncology. 1996; 53: 250-257
        • Takamura Y
        • Kobayashi H
        • Taguchi T
        • Motomura K
        • Inaji H
        • Noguchi S
        Prediction of chemotherapeutic response by collagen gel-droplet-embedded-culture drug sensitivity test in human breast cancers.
        Int. J. Cancer. 2002; 98: 450-455
        • Sevin B.U
        • Peng Z.L
        • Perras J.P
        • Ganjei P
        • Penalever M
        • Averette H.E
        Application of an ATP-bioluminescense assay in human tumor chemosensitivty testing.
        Gynecol. Oncol. 1988; 31: 191-204
        • Kurbacher C.M
        • Grecu O.M
        • et al.
        ATP chemosensitivity testing in ovarian and breast cancer.
        Recent Results Cancer Res. 2003; 161: 221-230
        • Mercer S.J
        • Somers S.S
        • Knight L.A
        • et al.
        Heterogeneity of chemosensitivity of esophageal and gastric carcinoma.
        Anticancer Drugs. 2003; 14: 397-403
        • Salmon S.E
        • Hamburger A.W
        • Soehnlein B
        • Durie B.G
        • Alberts D.S
        • Moon T.E
        Quantitation of differential sensitivity of human tumor stem cells to anticancer drugs.
        N. Engl. J. Med. 1978; 298: 1321-1327
        • Lieber M.M
        • Ames M.M
        • Powis G
        • Kovach J.S
        Anticancer drug testing in vitro.
        Life Sciences. 1981; 28: 287-293
        • Link K.H
        • Kornmann M
        • Leder G.H
        • et al.
        Regional chemotherapy directed by individual chemosensitivity testing in vitro.
        Clin. Cancer Res. 1996; 2: 1469-1474
        • Link K.H
        • Kornmann M
        • Butzer U
        • et al.
        Thymidylate synthase quantitation and in vitro chemosensitivity testing predicts responses and survival of patients with isolated nonresectable liver tumors receiving hepatic arterial infusion chemotherapy.
        Cancer. 2000; 89: 288-296
        • Boyd M.R
        Status of the NCI preclinical antitumor drug discovery screen.
        in: DeVita Jr., V.T Hellmann S Rosenberg S.A Cancer: Principles and Practice of Oncology, Updates, (vol 3). Lippincott, Philadelphia1989: 1-12
        • Grever M.R
        • Schepartz S.A
        • Chabner B.A
        The National Cancer Institute.
        Semin. Oncol. 1992; 19: 622-638
      2. Roth T, Dengler W, Fiebig HH. Human tumor cell lines demonstrating the characteristics of patient tumors as useful models for anticancer drug screening. Fiebig HH, Burger AM, eds. Relevance of Tumor Models for Anticancer Drug Development, Contrib Oncol. Basel, Karger, 1999, 54, 145–157.

        • Sausville E.A
        • Feigal E
        Evolving approaches to cancer drug discovery and development at the National Cancer Institute.
        Ann. Oncol. 1999; 10: 1287-1292
        • Von Hoff D.D
        In vitro predictive testing.
        Int. J. Cell Cloning. 1987; 5: 179-190
        • Von Hoff D.D
        He's not going to talk about in vitro predictive assays again, is he?.
        J. Natl. Cancer Inst. 1990; 82: 96-101
        • Von Hoff D.D
        • Sandbach J.F
        • Clark G.M
        • et al.
        Selection of cancer chemotherapy for a patient by an in vitro assay versus a clincian.
        J. Natl. Cancer Inst. 1990; 82: 110-116
        • Kern D.H
        • Weisenthal L.M
        Highly specific prediction of antieoplastic drug resistance with an in vitro assay using suprapharmacologic drug exposures.
        J. Natl. Cancer Inst. 1990; 82: 582-588
        • Shoemaker R.H
        • Wolpert-DeFilippes M.K
        • Kern D.H
        • et al.
        Application of a human tumor colony-forming assay to new drug screening.
        Cancer Res. 1985; 45: 2145-2153
        • Carney D.N
        • Winkler C.F
        In vitro assays of chemotherapeutic sensitivity.
        in: De Vita V.T Hellmann S.H Rosenberg S.A Important advances in oncology. Lippincott, Philadelphia1985: 78-103
        • Berger D.P
        • Fiebig H.H
        • Winterhalter B.R
        • Scholz C
        • Löhr G.W
        Quality control measures in the clonogenic assay.
        Proc. Am. Ass. Cancer Res. 1989; 30 (abstr 2427): 610
        • Berger D.P
        • Henss H
        • Winterhalter B.R
        • Fiebig H.H
        The clonogenic assay with human tumor xenografts.
        Ann. Oncol. 1990; 1: 333-341
        • Weisenthal L.M
        • Lippmann E.M
        Clonogenic and nonclonogenic in vitro assays.
        Cancer Treat. Rep. 1985; 69: 615-632
        • Thomson S.P
        • Moon T.E
        • Meyskens F.L
        Kinetics of clonogenic melanoma cell proliferation and the limits of growth within a bilayer agar system.
        J. Cell. Physiol. 1984; 121: 114-124
        • Fiebig H.H
        • Neumann H
        • Henss H
        Development of three human small cell lung cancer models in nude mice.
        Recent Results Cancer Res. 1985; 97: 77-86
        • Fortmeyer H.P
        • Bastert G
        Breeding and maintenance of nu/nu mice and rnu/rnu rats.
        in: Bastert G Fortmeyer H.P Schmidt-Mathiesen H Thymus Aplastic Nude Mice and Rats in Clinical Oncology. Fischer, Stuttgart/New York1981: 25-38
        • Alley M.C
        • Uhi C.B
        • Lieber M.M
        Improved detection of drug cytotoxicity in the soft agar colony formation assay through use of a metabolizable tetrazolium salt.
        Life Sci. 1982; 31: 3071-3078
        • Fiebig H.H
        Comparison of tumor response in nude mice and in the patients.
        in: Winograd B Pinedo H Human Tumor Xenografts in Anticancer Drug Development. Springer, Berlin1988: 25-30
        • Berger D.P
        • Winterhalter B.R
        • Fiebig H.H
        Chemotherapy.
        in: Boven E Winograd W The Nude Mouse in Oncology Research. CRC Press, Amsterdam1991: 165-184
      3. Berger DP, Fiebig HH, Winterhalter BR. Establishment and characterization of human tumor xenograft models in nude mice. In. Fiebig HH, Berger DP, eds. Immunodeficient Mice in Oncology. Contrib Oncol Basel, Karger, 1992, 42, 23–46.

        • Fiebig H.H
        • Burger A.M
        Human tumor xenografts and explants.
        in: Teicher B.A Tumor Models in Cancer Research. Humana Press, Totowa2001: 113-137
        • Selby P
        • Buick R.N
        • Tannock I
        A critical appraisal of the “human tumor stem cell assay”.
        N. Engl. J. Med. 1983; 308: 129-134
      4. Phillips RM. In vitro models of solid-tumour biology and drug delivery: Implications for and applications to target-oriented screening for novel anticancer drugs. Fiebig HH, Burger AM, eds. Relevance of Tumor Models for Anticancer Drug Development. Contrib Oncol Basel, Karger, 1999, 54, 67–80.

        • Phillips R.M
        Three dimensional cellular models and hypoxia selective toxins.
        Onkologie. 2003; 26 (abstr L24): 390
        • Fruehauf J.P
        • Bosanquet A.G
        In vitro determination of drug response.
        in: DeVita V.T Hellman S Rosenberg S.A Cancer, Principals and Practice of Oncology, PPO updates, vol. 12. Lippincott, Philadelphia1993: 1-16
        • Fiebig H.H
        • Berger D.P
        Preclinical phase II trials.
        in: Boven E Winograd W The Nude Mouse in Oncology Research. CRC Press, Amsterdam1991: 317-326
        • Berger D.P
        • Herbstritt L
        • Dengler W
        • Marmé D
        • Mertelsmann R
        • Fiebig H.H
        Vascular endothelial growth factor (VEGF) mRNA expression in human tumor models of different histologies.
        Ann. Oncol. 1995; 6: 817-825
        • Burger A.M
        • Fiebig H.H
        Screening using animal systems.
        in: Baguley B.C Kerr D.J Anticancer Drug Development. Academic Press, San Diego2001: 285-297
      5. Burger AM, Fiebig HH. Preclinical Screening for New Anticancer Agents. Figg WD, McLeod H, eds. Handbook of Anticancer Pharmacokinetics and Pharmacodynamics, Humana Press (in press).

        • Kononen J
        • Bubendorf L
        • Kallioniemi A
        • et al.
        Tissue microarrays for high-throughput molecular profiling of tumor specimens.
        Nat. Med. 1998; 4: 844-847
      6. Mirlacher M, Storz M. Gewebe-Chips für die molekulare Untersuchung von Tumoren. Labmed 2000, September, 293–297.

        • Nocito A
        • Kononen J
        • Kallioniemi O.P
        • Sauter G
        Tissue microarrays (TMAs) for high-throughput molecular pathology research.
        Int. J. Cancer. 2001; 94: 1-5
        • Wirth J.G
        • Schandelmaier K
        • Tetling E
        • Burger A.M
        • Fiebig H.H
        Xenograft tissue microarrays.
        Onkologie. 2003; 26 (abstr L20): 388
        • Kelter G
        • Maier A
        • Metz T
        • Fiebig H.H
        A cellular high throughput screen for anticancer drug discovery.
        Onkologie. 2003; 26 (abstr L22): 389
      7. Plowmann J, Camalier R, Alley M, Sausville E, Schepartz S. US-NCI Testing procedures. In Fiebig HH, Burger AM, eds. Relevance of Tumor Models for Anticancer Drug Development. Contrib Oncol Basel, Karger, 1999, 54, 121–135.

        • Smith V
        • Sausville E.A
        • Camalier R.F
        • Fiebig H.H
        • Burger A.M
        17-DMA-Geldanamycin is a novel water-soluble, orally bioavailable Hsp90 inhibitor with potent in vitro and in vivo anticancer activity.
        Proc. Am. Assoc. Cancer Res. 2003; 44 (abstr #786): 153
        • Smith V
        • Sausville E
        • Fiebig H.H
        • Burger A.M
        Modulators of heat shock protein 90 –– targeting multiple signal transduction pathways.
        Onkologie. 2003; 26 (abstr L15): 386
        • Jones S
        • Winer E
        • Vogel C
        • et al.
        Randomized comparison of vinorelbine and melphalan in anthracycline-refractory advanced breast cancer.
        J. Clin. Oncol. 1995; 13: 2567-2574
        • Morris M
        • Brader K.R
        • Levenback C
        • Burke T.W
        • Atkinson E.N
        • Scott W.R
        • Gershenson D.M
        Phase II study of vinorelbine in advanced and recurrent squamous cell carcinoma of the cervix.
        J. Clin. Oncol. 1998; 16: 1094-1098
        • Pignata S
        • Silvestro G
        • Ferrari E
        • et al.
        Phase II study of cisplatin and vinorelbine as first-line chemotherapy in patients with carcinoma of the uterine cervix.
        J. Clin. Oncol. 1999; 17: 756-760
        • Maier A
        • Kelter G
        • Gianella-Borradori A
        • Fiebig H.H
        Antitumor activity of CYC202, a cyclin dependent kinase inhibitor, in human tumor xenografts in vitro.
        Proc. Am. Assoc. Cancer Res. 2003; 44 (abstr #R3119): 619
        • Burger A.M
        • Mengs U
        • Wetzel D
        • Schüler J.B
        • Fiebig H.H
        Antitumor activity of an aqueous mistletoe extract (AME) standardized for ML-1 in human tumor cell lines and xenografts in vitro.
        Drug Res. 2001; 51: 748-757
        • Burger A.M
        • Mengs U
        • Gerstmayer B
        • Weber K
        • Fiebig H.H
        • Lentzen H
        Recombinant mistletoe lectin (rML) is a potent anticancer agent in experimental murine and human tumor models in vivo.
        Proc. Clin. Cancer Res. 1999; 5: 160
        • Kurbacher C.M
        • Cree I.A
        • Bruckner H.W
        • et al.
        Use of an ex vivo ATP luminescence assay to direct chemotherapy for recurrent ovarian cancer.
        Anticancer Drugs. 1998; 9: 51-57
        • Sharma S
        • Neale M.H
        • Nicolantonio D.F
        • et al.
        Outcome of ATP-based tumor chemosensitivity assay directed chemotherapy in heavily pre-treated recurrent ovarian carcinoma.
        BMC Cancer. 2003; 3: 19-28
        • Tanigawa N
        • Mizuno Y
        • Hashimura T
        • et al.
        Comparison of drug sensitivity among tumor cells within a tumor, between primary tumor and metastases, and between different metastases in the human tumor colony-forming assay.
        Cancer Res. 1984; 44: 2309-2312
        • Von Hoff D.D
        • Clark G.M
        Drug sensitivity of primary versus metastasis.
        in: Salmon S.E Trent J.M Human Tumor Cloning. Grune & Stratton, Orlando1984: 183-195
        • Bertelsen C.A
        • Korn E.I
        • Morton D.L
        • Kern D.H
        Heterogeneity of human metastatic clones by in vitro chemosensitivity testing.
        Arch. Surg. 1983; 118: 1406-1409
        • Kern D.H
        Heterogeneity of drug resistance in human breast and ovarian cancers.
        Cancer J. Sci. Am. 1998; 4: 41-45
        • Drees M
        • Dengler W.A
        • Roth T
        • et al.
        Flavopiridol (L86-8275).
        Clin. Cancer Res. 1997; 3: 273-279
        • Fiebig H.H
        • Berger D.P
        • Dengler W.A
        • Drees M
        • Mayo J
        • Malspeis L
        • Grever M
        Cyanocyclin A and the quinocarmycin analog NSC 607097 demonstrate selectivity against melanoma xenografts in vitro and in vivo.
        Proc. Am. Assoc. Cancer Res. 1994; 35 (abstr #2794): 468
        • Burger A.M
        • Kaur G
        • Hollingshead M
        • et al.
        Antiproliferative activity in vitro and in vivo of the spicamycin analog KRN5500 with altered glycoprotein expression in vitro.
        Clin. Cancer Res. 1997; 3: 455-463
        • Winterhalter B.R
        • Berger D.P
        • Dengler W.A
        • Hendriks H.R
        • Mertelsmann R
        • Fiebig H.H
        High antitumor activity of rhizoxin in a combined in vitro and in vivo test procedure with human tumor xenografts.
        Proc. Am. Assoc. Cancer Res. 1993; 34 (abstr #2240): 376
        • Burger A.M
        • Mengs U
        • Schüler J.B
        • Zinke H
        • Lentzen H
        • Fiebig H.H
        Recombinant mistletoe lectin (rML) is a potent inhibitor of tumor cell growth in vitro and in vivo.
        Proc. Am. Assoc. Cancer Res. 1999; 40 (abstr #2639): 399