Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving nucleic acid
Patent
1996-01-19
1998-06-23
Wax, Robert A.
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving nucleic acid
4353201, 4352523, 43525433, 435325, 435366, 435 691, 4351723, 536 235, 530350, 5303879, 514 12, C12Q 168, C12N 1512, C12N 508, C12N 510, C12N 121, C12N 100, A61K 3817, C07K 14425, C07K 1618
Patent
active
057703663
DESCRIPTION:
BRIEF SUMMARY
FIELD OF THE INVENTION
The invention concerns a melanoma-inhibiting protein (MIA), a nucleic acid which codes for it, a process for the isolation and for the detection of this protein as well as its use for the production of a therapeutic agent.
BACKGROUND AND PRIOR ART
The regulation of cell growth is controlled by factors which act positively as well as negatively. The factors with a positive effect include the known growth factors such as e.g. epidermal growth factor (EGF), platelet derived growth factor (PDGF), insulin and somatomedins. The factors with a negative i.e. inhibitory activity include, in addition to TGF-.beta. which can act as a growth stimulator as well as a growth inhibitor (Roberts et al., Proc. Natl. Acad. Sci. 82 (1985), 119-123), the endogenous, tumour-inhibiting factors from colon carcinoma cells (Levine et al., Cancer Research 45 (1985), 2248-2254), melanomas (Bogdahn et al., Cancer Research 49 (1989), 5358-5363) as well as from healthy epithelial cells from the mammary glands of the rat (Ethier et al., J. Cell. Phys. 142 (1990), 15-20).
Disturbances of this regulatory system such as for example by overproduction of growth factors with a positive action or by a reduced dependence of mutated cells on these growth factors (Rodeck et al., International Journal of Cancer 40 (1987), 687-690) enable tumour cells to proliferate in an uncontrolled manner. The aforementioned tumour-inhibitory factors from various tumour tissues represent interesting compounds which may be able to intervene therapeutically in this impaired regulatory system. It must be possible to provide these factors in large amounts and in reproducible purity for such a therapeutic use. However, for most of these factors only enriched fractions from cell lysates have been described up to now which are not suitable for a therapeutic application due to their complex and sometimes unknown composition and the concomitant non-reproducibility of their production.
SUMMARY OF THE INVENTION
The invention is based on a new melanoma-inhibiting protein (denoted MIA protein, or MIA, in the following) which inhibits growth of the cell lines HTZ 19-dM and ATCC CRL 1424 and protein or for the protein with a N-terminal pre sequence, or by the genomic sequence shown in SEQ ID NO: 3, in SEQ ID NO: 1 or 3 or fragments of these DNA sequences in the DNA region which codes for the mature protein.
Growth inhibition is to be understood as an anti-proliferative activity. In this process the growth of the cells is considerably retarded by addition of the MIA protein to the culture medium. A suitable concentration for this is for example 0.1 .mu.g MIA protein/ml culture medium. Higher or lower concentrations of the MIA protein are, however, also suitable for growth inhibition which is observed to be higher or lower depending on the concentration.
The properties of such a protein are described by the inventors in Cancer Research 49 (1989), 5358-5363, Cancer Research 50 (1990), 6981-6986, Melanoma Research 2 (1992), 327-336. However, a process for the production of this protein which can be reproduced is not stated in these publications. The protein is obtainable from the human melanoma cell line HTZ 19-dM which was previously not yet available to the public. This cell line was derived from a metastizing malignant melanoma and cultured as a monolayer culture in a defined serum-free culture medium (50% Dulbecco's minimal essential medium, 50% F-12) containing 0.8 mmol/l L-glutamine, non-essential amino acids, 10 .mu.g/ml transferrin, 30 nmol/l sodium selenite and 4 .mu.g/ml gentamicin under standard culture conditions. The cell line was deposited at the "Deutsche Sammlung fur Mikroorganismen und Zellkulturen GmbH" in Braunschweig on the 23.06.93 (DSM ACC 2133). It is also a further subject matter of the invention. The protein according to the invention can be obtained from the culture supernatant of this cell line by gel chromatographic isolation of a protein fraction having a size of ca. 11 kD and subsequent purification of this fraction by mea
REFERENCES:
Apfel, R., et al., "Purification and analysis of growth regulating proteins, secreted by a human melanoma cell line", j. Cell.. Biochem., Suppl. 16B:149 (1992).
Apfel, R., et al., "Tumor progression associated proteins are secreted by a human malignant melanoma cell line", Proceedings of the American Association for Cancer Research vol. 33:71 (1992).
Blesch, A., et al., "Cloning of a novel malignant melanoma-derived growth regulatory protein, MIA", Cancer Res. 54:5695-5701 (1994).
Blesch, A., et al., "Effects of Melanoma Inhibitory Activity upon growth of various tumour cell lines in vitro", J. Cancer Res. Clin. Oncol. 120:Supl. R112 (1994).
Blesch, A., et al., "Expression and Function of a novel, potent malignant melanoma cell growth inhibitor (MIA", Proceedings of the American Association for Cancer Research 35:567 (3379A) (1994).
Blesch, A., et al., "Cloning of a novel malignant melanoma cell growth inhibitor (MIA) and expression pattern in neuroectoderm derived tumor cell lines", Journal of Neurooncology 21:51 (1994).
Bogdahn, U., et al., "Autocrine tumor growth inhibiting activity from human malignant melanoma", EMBL--Conference Oncogenes and Growth Control, Heidleberg, EMBO Verlag, p. 23 (1988).
Behl, C., et al., "Biological characterization of a novel neuroectodermal-derived tumour growth inhibitor, melanoma-inhibiting activity", Second Meeting of the European Neurological Society, Brighton, england, UK, Jun. 30-Jul. 5, 1990, J. Neurol. 237 (Suppl. 1):S4 (1990).
Bogdahn, U., et al., "Melanoma inhibitory activity (MIA)--a new potent tumour growth regulatory protein", J. Cancer Res. Clin. Oncol. 120:Supl. R113 (1994).
Bosserhoff, A.-K., et al., "Inhibition of melanoma cell proliferation and invasion by human and murine MIA (melanona inhibiting activity)", J. Invest. Dermatol. 103(3):430 (1994).
Bosserhoff, A.-K., et al., "Inhibition of melanoma cell invasion by human and murine MIA (melanoma inhibitory activity)", Proceedings of the AACR 35:253 (1994).
Bosserhoff, A.-K., et al., "Molecular cloning and characterization of a new protein with melanoma inhibiting activity (MIA)", Arch. of Dermatolog. Res. 286(3/4):213-214 (1994).
U. Bogdahn et al., Autocrine Tumor Cell Growth-inhibiting Activities from Human Malignant Melanoma, Cancer Research 49, 5358-53-63, Oct. 1, 1989.
R. Apfel et al., "Purification and analysis of growth regulating proteins secrted by a human melanoma cell line", Melanoma Research, vol. 2, 1992, pp. 327-336.
F.X. Welbach, et al., Melanoma-inhibiting Activity Inhibits Cell Proliferation by Prolongation of the S Phase and Arrest of Cells in the G2 Compartment, Cancer Reserach 50, 6981-6986, Nov. 1, 1990.
Bogdahn Ulrich
Burrner Reinhard
Kaluza Brigitte
Boehringer Mannheim GmbH
Bugaisky G. E.
Wax Robert A.
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