Abstract
The possibility of selective killing of transformed cells in a mixed population of untransformed and transformed cells was examined using a cell culture system of rat 3Y1 fibroblasts (parental 3Y1 cells, 3Y1 cells transformed with either SV40, polyoma virus, Rous avian sarcoma virus, E1A gene of adenovirus type 12, or H‐v‐ras oncogene). The principle of the selective killing is as follows. Under suboptimal culture conditions, untransformed cells are inhibited from progressing through G1 phase and retain viability, while transformed cells are not arrested. When DNA synthesis is inhibited for a long period, both types of cells in S phase die. Therefore, if we administer inhibitors of G1 progression and of DNA synthesis simultaneously to a cell population consisting of untransformed and transformed cells, most untransformed cells are arrested in G1 phase, retaining viability, while transformed cells leak from the G1 phase, cease DNA synthesis, and gradually die. The present study shows that all types of transformants in stationary‐phase cultures (consisting of cells mainly with a G1 DNA content) were killed to higher extents compared with untransformed cells, during incubation at lower cell densities with a combination of cycloheximide (G1 inhibitor) and aphidicolin (DNA‐synthesis inhibitor). However, cycloheximide reduced the killing effect of aphidicolin by changing the irreversible DNA‐synthesis inhibition to a reversible inhibition. The availability of G1 inhibitors that do not interfere with the irreversibility of inhibition of DNA synthesis is required for the treatment of cancer based on this idea.
Keywords: Transformed cells, Cell killing, Cell cycle, Cycloheximide, Aphidicolin
Full Text
The Full Text of this article is available as a PDF (387.2 KB).
REFERENCES
- 1. ) Baserga , R. “ The Biology of Cell Reproduction ” ( 1985. ). Harvard University Press; , Cambridge , Massachusetts . [Google Scholar]
- 2. ) Campisi , J. , Medrano , E. E. , Morreo , G. and Pardee , A. B.Restriction point control of cell growth by a labile protein: evidence for increased stability in transformed cells . Proc. Natl. Acad. Sci. USA , 79 , 436 – 440 ( 1982. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
- 3. ) Okuda , A. and Kimura , G.Non‐specific elongation of cell cycle phases by cycloheximide in rat 3 Y1 cells, and specific reduction of G1 phase elongation by simian virus 40 large T antigen . J.Cell Sci. , 91 , 295 – 302 ( 1988. ). [DOI] [PubMed] [Google Scholar]
- 4. ) Pfeiffer , S. E. and Tolmach , L. J.Inhibition of DNA synthesis in HeLa cells by hydroxyurea . Cancer Res. , 27 , 124 – 129 ( 1967. ). [PubMed] [Google Scholar]
- 5. ) Kim , J. H. , Gelbard , A. S. and Perez , A. G.Action of hydroxyurea on the nucleic acid metabolism and viability of HeLa cells . Cancer Res. , 27 , 1301 – 1305 ( 1967. ). [PubMed] [Google Scholar]
- 6. ) Ikegami , S. , Taguchi , T. , Ohashi , M.Oguro , M. , Nagano , H. and Mano , Y.Aphidicolm prevents mitotic cell division by interfering with the activity of DNA polymerase‐α . Nature , 275 , 458 – 460 ( 1978. ). [DOI] [PubMed] [Google Scholar]
- 7. ) Kimura , G. , Itagaki , A. and Summers , J.Rat cell line 3Y1 and its virogenic polyoma‐ and SV40‐transformed derivatives . Int. J. Cancer , 15 , 694 – 706 ( 1975. ). [DOI] [PubMed] [Google Scholar]
- 8. ) Zaitsu , H. , Tanaka , H. , Mitsudomi , T. , Matsuzaki , A. , Ohtsu , M. and Kimura , G.Differences in proliferation properties among sublines of rat 3Y1 fibroblasts transformed by various agents in vitro . Biomed. Res. , 9 , 181 – 197 ( 1988. ). [Google Scholar]
- 9. ) Shimura , H. , Ohtsu , M. , Matsuzaki , A. , Mitsudomi , T. , Onodera , K. and Kimura , G.Selective cytotoxicity of phospholipids and diacylglycerols to rat 3Y1 fibroblasts transformed by adenovirus type 12 or its E1A gene . Cancer Res. , 48 , 578 – 583 ( 1988. ). [PubMed] [Google Scholar]
- 10. ) Okuda , A. , Shimura , H. and Kimura , G.Abortive transformation of rat 3Y1 cells by simian virus 40: viral function overcoming inhibition of cellular proliferation under various conditions of culture . Virology , 133 , 35 – 45 ( 1984. ). [DOI] [PubMed] [Google Scholar]
- 11. ) Okuda , A. and Kimura , G.Serum stimulation of DNA synthesis in rat 3Y1 cells: dependence on cell density, serum concentration, and ratio of cell number to medium volume . Exp. Cell Res. , 111 , 55 – 62 ( 1978. ). [DOI] [PubMed] [Google Scholar]
- 12. ) Okuda , A. and Kimura , G.Serum‐dependent control of entry into S phase of next generation in rat 3Y1 fibroblasts: effect of large T antigen of simian virus 40 . Exp. Cell Res. , 163 , 127 – 134 ( 1986. ). [DOI] [PubMed] [Google Scholar]
- 13. ) Bhuyan , B. K. and Fraser , T. J.Antagonism between DNA synthesis inhibitors and protein synthesis inhibitors in mammalian cell cultures . Cancer Res. , 34 , 778 – 782 ( 1974. ). [PubMed] [Google Scholar]
- 14. ) Johnston , R. N. , Feder , J. , Hill , A. B. , Sherwood , S. W. and Schimke , R.T.Transient inhibition of DNA synthesis results in increased dihydrofolate reductase synthesis and subsequent increased DNA content per cell . Mol. Cell. Biol. , 6 , 3373 – 3381 ( 1986. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
- 15. ) Sherwood , S. W. , Schumacher , R. I. and Schimke , R. T.Effect of cycloheximide on development of methotrexate resistance of Chinese hamster ovary cells treated with inhibitors of DNA synthesis . Mol. Cell. Biol. , 8 , 2822 – 2827 ( 1988. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
- 16. ) Van Putten , L. M.Are cell kinetic data relevant for the design of tumour chemotherapy schedules ? Cell Tissue Kinet. , 7 , 493 – 504 ( 1974. ). [DOI] [PubMed] [Google Scholar]
- 17. ) Pardee , A. B. and James , L. J.Selective killing of transformed baby hamster kidney (BHK) cells . Proc. Natl. Acad. Sci. USA , 72 , 4994 – 4998 ( 1975. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
- 18. ) Rozengurt , E. and Po , C. C.Selective cytotoxicity for transformed 3T3 cells . Nature , 261 , 701 – 702 ( 1976. ). [DOI] [PubMed] [Google Scholar]