DNA fragment encoding tumor cell growth inhibitors

Details DNA fragment encoding tumor cell growth inhibitors DNA fragment encoding tumor cell growth inhibitors

1994-05-25

1996-06-04

Wax, Robert A.

Organic compounds -- part of the class 532-570 series

Organic compounds

Carbohydrates or derivatives

536 231, 530324, 435 691, 4351723, 4353201, 4352523, C12N 1528, C12N 1519, C12N 1512

Patent

active

055233910

DESCRIPTION:

BRIEF SUMMARY
TECHNICAL FIELD

The present invention relates to DNA fragments encoding novel tumor cell growth inhibitors. More particularly, the present invention relates to DNA fragments encoding novel tumor cell growth factors which can be obtained from the culture supernatant of 3T3 cell-derived cell line and exhibit an activity of inhibiting the growth of tumor cells.


BACKGROUND ART

Synthetic drugs such as chemotherapeutic agents or immunotherapeutic agents have been heretofore widely used as anti-tumor agents. However, these drugs generally encounter problems that their specificity is low and side-effects are serious. On the other hand, many tumor cell growth inhibitors have been found in tissue culture cells. These inhibitors could be such anti-tumor agents that would be highly specific and would have minimized side-effects. Representative examples of such inhibitors are interferon, lymphotoxin and tumor necrosis factor (TNF). Recently, a tumor cell cytotoxic factor obtained from human fibroblast and a tumor cell growth inhibitor obtained from human lung cancer cells are reported in Japanese Patent KOKAI Nos. 1-148197 and 1-187094, respectively.
Some cell growth inhibitors are isolated also from the fibroblastic 3T3 cell line established from the cells obtained from Swiss fetal mice. For example, Natraj et al. has reported that a growth inhibitor was obtained from the cell surface of 3T3 cells in the stationary phase, cf., Proc. Natl. Acad. Sci. U.S.A., 75, 6115-6119 (1978). Harel et al. has reported that a growth inhibitor having a molecular weight of 40 kDa was obtained from the culture supernatant of 3T3 cells, see J. Cell. Physiol., 119, 101-106 (1984), ibid., 123, 139-143 (1985). However, these growth inhibitors all fail to show any significant inhibitory action against tumor cells, as is known in the art.
The present inventors previously succeeded in isolating, from the culture supernatant of 3T3 cell-derived cell line, novel tumor cell growth inhibitors having an activity of inhibiting the growth of tumor cells, which was filed as Japanese Patent Application No. 3-11950.
The tumor cell growth inhibitors exhibit a potent growth inhibition activity against human promyelogenous leukemia cells or human uterine cervical cancer-derived cells and are expected to be effective for the treatment of cancer.
For use as new carcinostatic agents, it is required to supply the tumor cell growth inhibitors in a sufficient amount. It is thus desired to develop a method for production available with industrial advantages.


DISCLOSURE OF INVENTION

The present inventors have brought attention to recombinant DNA technique applicable to the process for production of the tumor cell growth inhibitors in an industrially efficient way, and made investigations on cloning of cDNA encoding the tumor cell growth inhibitors. Succeeded by recombinant DNA technique in obtaining DNA fragments encoding the inhibitors which can be used for production of the inhibitors, the present invention has been accomplished.
That is, the present invention relates to DNA fragments encoding the tumor cell growth inhibitors, which has nucleotide sequence shown by formula (1): ##STR2## wherein X represents TTC TTT CTA or TTC.


BRIEF DESCRIPTION OF DRAWINGS

FIG. 1 is a graph showing elution profile of phenyl 5PW-RP reversed phase HPLC of tumor cell growth inhibitor P-1, which is the subject of the present invention.
FIG. 2 is a graph showing elution profile of phenyl 5PW-RP reversed phase HPLC of tumor cell growth inhibitor P-2 which is the subject of the present invention.
FIG. 3 shows oligonucleotides (1) through (11) used as primers in the PCR method.
FIG. 4 briefly shows the amplified site of DNA fragment amplified by the PCR method.
FIG. 5 is an outline of the DNA fragment obtained by the PCR method.
FIG. 6 shows a nucleotide sequence of the DNA fragment encoding a part of P-1and the translated amino acid sequence.
FIG. 7 shows a nucleotide sequence of the DNA fragment encoding the C-terminal region of P-1 and the translated amino acid sequence.
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REFERENCES:
patent: 4675285 (1987-06-01), Clark et al.
patent: 5281520 (1994-01-01), O'Hara et al.
patent: 5384394 (1995-01-01), Komurasaki et al.
C. C. Lee et al. Science 239:1288-1291 (Mar. 1988).
J. M. Wozney Meth. in Enzymol. 182:738-751 (1990).
J. Cell Physiol., vol. 119, 1984, pp. 101-106 Harel et al.

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