Stem cell studies of human malignant brain tumors

Part 2: Proliferation kinetics of brain-tumor cells in vitro in early-passage cultures

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✓ The proliferation kinetics were studied in early-passage cultures of cells from 13 human malignant brain tumors and two specimens of normal brain under conditions similar to those used in clonogenic cell-survival studies. Autoradiography was performed in all but four cases to estimate the fraction of cells actively replicating deoxyribonucleic acid (DNA), the approximate cell cycle time, and the effect of low-dose tritiated thymidine on cell proliferation. The mean tumor cell doubling time (TD) was 53 hours for five glioblastomas, 46 hours for two ependymomas, and 83 hours for two medulloblastomas. A gliosarcoma grew fastest (TD = 22 hours) in culture and a pilocytic astrocytoma grew slowest (TD = 144 hours). The approximate cell cycle time ranged from 1 to 2.5 days for all tumors tested. This suggests that chemotherapeutic agents that predominantly kill proliferating cells should be administered in vitro for at least 2 to 2.5 days to achieve maximum cell kill.

The approximate growth fraction ranged from 0.65 to 0.96 for all tumors except for the two medulloblastomas and the pilocytic astrocytoma, which had growth fractions of 0.34 and 0.35, respectively. Most laboratories investigating the chemosensitivity of primary or early-passage human tumor cells require that 40% to 70% of cells be killed to consider a drug active in vitro. The results of this study suggest that the cell-cycle-specific agents cannot achieve a high enough cell kill to be considered active for some tumors that grow slowly in culture. An estimate of the in vitro growth rate is necessary to reliably interpret cell-survival results with such agents.

Tritiated thymidine appeared to slow cell proliferation in some of the cultures, presumably as a result of radiation-induced DNA damage caused by tritium that had been incorporated into DNA. The degree to which cell growth was slowed in individual tumors correlated with the patient's clinical response to radiation therapy and postoperative survival time.

Article Information

Dr. Rosenblum is the recipient of Teacher-Investigator Development Award 1K-07NS 00604 NSPA from the National Institute of Neurological and Communicative Disorders and Stroke.

Address for Dr. Pertuiset: Clinique Neurologique, Division MAZARIN, Hôpital de la Salpetriere, 47 Boulevard de l'Hôpital, 75651 Paris CEDEX 13, France.

Address reprint requests to: Mark L. Rosenblum, M.D., c/o The Editorial Office, Department of Neurological Surgery, 1360 Ninth Avenue, Suite 210, San Francisco, California 94122.

© AANS, except where prohibited by US copyright law.

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Figures

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    Fraction of tritium-labeled cells present in cultures of a gliosarcoma (GS), a glioblastoma (GM), and a medulloblastoma (Med) up to 4 days after exposure to tritiated thymidine (3H-TdR) at a concentration of 0.09 µCi/ml. The labeled fraction of cells was least for the putatively more radiation-sensitive medulloblastoma.

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    The number of cells 3 days (triangle) and 6 days (circles) after planting 4 × 105 SF 209 glioblastoma cells in 25-sq cm culture flasks. When tritiated thymidine (3H-TdR) was not added to the flasks on Day 3, the number of cells increased by Day 6. However, a 3-day exposure to 3H-TdR decreased cell proliferation in a dose-related manner. The error bars represent the standard deviation (SD).

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