Comparison of the chemosensitivity of human neoplastic tissues between succinate dehydrogenase inhibition test and ATP assay

Comparison of the chemosensitivity of human neoplastic tissues between succinate dehydrogenase inhibition test and ATP assay

Clkcu Chimica Elsevier Acra, 166 (1987) 107-109 107 CCA 03817 Letter to the editor Comparison of the chemosensitivity of human neoplastic tissues...

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Clkcu Chimica Elsevier

Acra, 166 (1987) 107-109

107

CCA 03817

Letter to the editor

Comparison of the chemosensitivity of human neoplastic tissues between succinate dehydrogenase inhibition test and ATP assay Dear Editors, To evaluate the succinate dehydrogenase inhibition (SDI) test [1,2] for in vitro clinical testing, we compared the chemosensitivity between the SD1 test and the ATP assay. We found that the SD activity remaining in the dead cells must be taken into consideration for the chemosensitive prediction using the SD1 test, and that the ATP assay had a higher sensitivity than the SD1 test for predicting cell viability [3]. The SD1 test was done as described [2-51. Thirty-three human tumor tissues were cut with scissors and passed through a no. 32 mesh in minimal essential medium, plated in each of 35 mm plastic dishes, incubated at 37°C in a humidified 5% CO, atmosphere for 3 days, exposed to carboquone, adriamycin, mitomycin C, aclacinomycin A, cisplatin or 5-fluorouracil at the peak plasma concentration (1 x PPC) and the ten times peak plasma concentration (10 X PPC) [5,6]. Three(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl 2H tetrazolium bromide (MTT) [7] was used as a hydrogen acceptor for SD activity. The absorbance of formazan (optical density: o.d.), formed from MTT was measured at 565 nm. The tumor fragments with an o.d. of 0.5 on day 0 were plated in separate dishes. The ATP level was measured using the luciferin-luciferase method [8,9]. The tumor fragments were exposed to the drug for 3 days and boiled for 3 min to release the ATP into the extracellular medium [lo]. The SD activity and the ATP level were presented per milligram protein, determined using Bio-Rad protein assay, and the chemosensitivity was estimated by the percentage of these levels of the drug-treated cells, compared to that of control cells. The ATP level decreased more than the SD activity, with each concentration of the drugs (p < 0.001, Student’s t test) (Table I). Positive correlations (r = 0.659-0.884, p < 0.01) were noted between them, with each concentration of the 6 drugs. Moreover, a good correlation was evident between the SD activity at 10 X PPC and the ATP level at 1 X PPC (r = 0.788-0.863, p < 0.01). In the ATP assay, the chemosensitivity could be considered at 1 x PPC of all drugs tested. The ATP level is more sensitive than the SD activity for predicting cell viability. When the cell died, the respiratory activity ceased and a rapid and a irreversible ATP depression followed [8,9]. The positive correlation, however, between the chemosensitivity results of the two assays showed that the SD1 test and 0009-8981/87/%03.50

0 1987 Elsevier Science Publishers

B.V. (Biomedical

Division)

108 TABLE

I

Percentages of the SD activity and the ATP level of drug-treated cells, exposed to 6 different antitumor drugs: carboquone (CQ), adriamycin (ADM), mitomycin C (MMC), aclacinomycin A (ACR), cisplatin (DDP), S-fluorouracil (5-FU), compared to that of control cells in 33 tumor tissues Drug

SD activity 1XPPC

CQ ADM MMC ACR DDP 5-FU

90.74 f 90.65 f 89.55 f 90.24 f 88.89* 92.93 f

8.86 B 7.31 9.00 7.45 8.73 6.34

ATP level 10 x PPC

1XPPC

49.43 f 24.81 59.22 k22.52 55.26 k-24.63 54.73 + 23.47 51.52i24.26 66.89 f 19.36

51.08 69.86 56.28 58.03 65.48 70.07

f + f f + f

10 x PPC 22.98 20.96 25.90 20.08 24.11 19.14

13.95 30.03 16.90 25.40 28.50 41.82

+ + f f f f

14.47 20.76 16.43 20.75 21.21 22.79

a Meanfsn. the ATP assay are complementary. The two assays are simple, rapid, and as the SD1 test is less expensive than the ATP assay, the former will be useful for selecting sensitive drugs to treat individual cancer patients. Acknowledgements

This work was supported by a grant from the Fukuoka Cancer Society. We thank K. Miyamoto and K. Fukuchi for technical assistance and M. Ohara for reviewing the manuscript. Hideaki Anai Yoshihiko Maehara Hiroki Kusumoto Keizo Sugimachi

a, a, a, a*b

a Cancer Center of Kyushu University Hospital, b Second Department of Surgery, Faculty of IMedicine, Kyushu University, Fukuoka, Japan

References 1 Kondo T. Prediction of response of tumor and host to cancer chemotherapy. Nat1 Cancer Inst Monogr 1971;34:251-256. 2 Maehara Y, Anai H, Tamada R, Sugimachi K. Partial hepatectomy alters sensitivity of rat hepatocytes to 5-fluorouracil. Cancer Lett 1986;31:227-233.

Correspondence to: Dr. Yoshihiko Maehara, Cancer Center of Kyushu University Medicine, Kyushu University, 3-l-l Maidashi, Higashi-ku, Fukuoka 812, Japan.

Hospital,

Faculty

of

109 3 Maehara Y, Anai H, Tamada R, Sugimachi K. The ATP assay is more sensitive than the succinate dehydrogenase inhibition test for predicting cell viability. Em J Cancer Clin Oncol 1987;23:in press. 4 Maehara Y, Anai H, Kusumoto H, Sugimachi K. Poorly differentiated human gastric carcinoma is more sensitive to antitumor drugs than is well differentiated carcinoma. Eur J Surg Oncol 1987;13:in press. 5 Anai H, Maehara Y, Kusumoto H, Sugimachi K. Comparison between succinate dehydrogenase inhibition test and subrenal capsule assay for chemosensitivity testing. Oncology 1987;44:in press. 6 Anai H, Maehara Y, Kusumoto H, Masuda H, Miyamoto K, Fukuchi K, Tamada R, Sugimachi K. In vitro chemosensitivity of various human tumors evaluated by the succinate dehydrogenase inhibition test. Jpn J Cancer Chemother 1986;13:2544-2548. 7 Mosmann T. Rapid calorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J Immunol Methods 1983;65:55-63. 8 Kangas L, Grijnroos M, Nieminen A-L. Bioluminescence of cellular ATP: a new method for evaluating cytotoxic agents in vitro. Med Biol 1984;62:338-343. 9 Kuzmits R, Aiginger P, Frass M, Schopf G, Rumpold H, Schwarz HP, Mtiller MM. Influence of cytostatics on ATP-levels of leukemic cells. Adv Exp Med Biol 1984;165:383-388. 10 Wieb WJ, Bancroft K. Use of the adenylate energy charge ratio to measure growth state of natural microbial communities. Proc Nat1 Acad Sci USA 1975;72:2112-2115.