E6 binding proteins

Details E6 binding proteins E6 binding proteins

1999-08-26

2001-10-02

Salimi, Ali R. (Department: 1648)

Drug, bio-affecting and body treating compositions

Antigen, epitope, or other immunospecific immunoeffector

Virus or component thereof

C514S012200, C530S300000, C530S350000, C536S023720, C435S005000, C435S007230, C435S007600

Reexamination Certificate

active

06296853

ABSTRACT:

GOVERNMENT FUNDING
Work described herein was supported in part by funding from the National Institute of Health. The United States Government has certain rights in the invention.
BACKGROUND OF THE INVENTION
Papillomaviruses (PV) have been linked to widespread, serious human diseases, especially carcinomas of the genital and oral mucosa. It is estimated that there are currently somewhere in the neighborhood of tens of millions of women who suffer from human papilloma virus (HPV) infection of the genital tract. Many of these women eventually develop cancer of the cervix. For example, it has been estimated that about twenty percent (20%) of all cancer deaths in women worldwide are from cancers which are associated with HPV. It has also been estimated that 90% of all cervical cancer is linked to HPV.
Papillomaviruses induce benign, dysplastic and malignant hyperproliferations of skin or mucosal epithelium (see, for example, Mansur and Androphy, (1993)
Biochim Biophys Acta
1155:323-345; Pfister (1984)
Rev. Physiol. Biochem. Pharmacol.
99:111-181; and Broker et al. (1986)
Cancer Cells
4:17-36, for reviews of the molecular, cellular, and clinical aspects of the papillomaviruses). Almost 70 human papillomavirus types have been identified, and different papillomavirus types are known to cause distinct diseases, Pfister, (1987)
Adv. Cancer Res.,
48:113-147, Syrjanen, (1984)
Obstet. Gynecol. Survey
39:252-265. Human papillomaviruses (HPVs) are a heterogeneous group of DNA tumor viruses associated with hyperplastic (warts, condylomata), pre-malignant and malignant lesions (carcinomas) of squamous epithelium. For example, HPV types 1 and 2 cause common warts, and types 6 and 11 cause warts of the external genitalia, anus and cervix. HPV, types 16, 18, 31 and 33 have been isolated from the majority of cervical cancers with HPV-16 present in about 50 percent of all cervical cancers. These HPV's are referred to as “high risk”. While HPV 6 and 11 are the most common isolates for cervical warts, these infections rarely progress to invasive cancer, and therefore these HPV's are referred to as “low risk”.
Studies of viral gene expression in carcinomas suggest the importance of two HPV encoded proteins, E6 and E7, in malignant development and these proteins have been shown to encode transforming and immortalizing activities. The two proteins show some functional resemblance to the transforming proteins of other small DNA tumor viruses such as adenovirus and SV40. E7 shares functional and structural features with the adenovirus E1A proteins. Like Ad E1A and the large T proteins of the polyomaviruses, E7 can complex pRB. Likewise, the E6 oncoprotein encoded by the “high risk” HPV's can form a complex with p53. In vitro, E6 promotes the degradation of p53 and this degradation involves the ubiquitin-dependent protease system. The selective degradation of cellular negative regulatory proteins such as p53 regulatory functions provides an explanation of the action for dominant acting oncoproteins. The relevance of the inactivation of the normal functions of pRB and p53 in human cervical carcinogenesis has recently been demonstrated by the analysis of these two genes and their products in a series of HPV-positive and HPV-negative cell lines. These studies support the notion that the inactivation of the normal functions of the tumor suppressor proteins pRB and p53 are important steps in human cervical carcinogenesis, either by mutation or through complex formation with HPV E6 and E7 oncoproteins.
SUMMARY OF THE INVENTION
The present invention relates to the discovery in eukaryotic cells, particularly human cells, of novel protein-protein interactions between the papillomavirus transforming protein E6 and certain cellular proteins, referred to hereinafter as “E6-binding proteins” or “E6-BP”.
In general, the invention features a E6-BP
SD-7
polypeptide, preferably a substantially pure preparation of an E6-BP
SD-7
polypeptide, or a recombinant E6-BP
SD-7
polypeptide. In preferred embodiments: the polypeptide has biological activity, e.g., it specifically binds a papillomavirus E6 protein; the polypeptide has an amino acid sequence at least 60%, 70%, 80%, 90% or 95% homologous to the amino acid sequence in SEQ ID No: 8; the polypeptide has an amino acid sequence essentially the same as the amino acid sequence in SEQ ID No: 8; the polypeptide is at least 5, 10, 20, 25, 30, 40, 50, 100, or 150 amino acids in length; the polypeptide comprises at least 5, preferably at least 10, more preferably at least 20, 25, 30, 40, more preferably at least 50, 100, or 150 contiguous amino acids from SEQ ID No: 8; the E6-BP
SD-7
polypeptide is either, an agonist or an antagonist of a biological activity of an E6-BP, e.g., of the regulation of cell proliferation; the polypeptide includes an E6-binding motif corresponding to Ala 194-Asp 218 of SEQ. ID No. 8.
In preferred embodiments the invention includes E6-binding proteins with antagonistic activity, and which preferably are capable of: suppressing tumor growth, e.g. in a tumor cell in which endogenous E6-BP is misexpressed; suppressing growth of papillomavirus-infected cells, e.g. HPV-infected cells; blocking or inducing apoptosis; inhibiting growth of a papillomavirus-infected cell, e.g. an HPV-infected cell, e.g. a high-risk HPV infected cell, e.g. and HPV-16, -18, -31, or -33 infected cell, e.g. a bovine papillomavirus (BPV)-infected cell; inhibiting infection of a cell by a papillomavirus, e.g. an HPV, e.g. a high-risk HPV, e.g. and HPV-16, -18, -31, or -33, e.g. a bovine papillomavirus (BPV); inhibiting transformation of a cell by a papillomavirus, e.g. an HPV, e.g. a high-risk HPV, e.g. and HPV-16, -18, -31, or -33, e.g. a bovine papillomavirus; or inhibiting immortalization of a cell, e.g. a human cell, by a papillomavirus, e.g. an HPV, e.g. a high-risk HPV, e.g. and HPV-16, -18, -31, or -33, e.g. a bovine papillomavirus. In preferred embodiments, the antagonist is a fragment of the full-length SD-7 protein, which fragment, for example, retains the ability to bind E6 and competitively inhibits binding of the full-length SD-7 protein. For example, fragments containing the E6-binding motif corresponding to about Ala 194-Asp 218 can be provided as antagonists of the full-length protein.
In a preferred embodiment, a peptide having at least one biological activity of the subject E6-BP
SD-7
polypeptide may differ in amino acid sequence from the sequence in SEQ ID No: 8, but such differences result in a modified protein which functions in the same or similar manner as the native E6-binding protein or which has the same or similar characteristics of the native E6-binding protein.
In yet other preferred embodiments, E6-binding protein is a recombinant fusion protein which includes a second polypeptide portion, e.g., a second polypeptide having an amino acid sequence unrelated to a protein represented by one of SEQ ID Nos: 8-14, e.g. the second polypeptide portion is glutathione-S-transferase, e.g. the second polypeptide portion is a DNA binding domain, e.g. the second polypeptide portion is a polymerase activating domain, e.g. the fusion protein is functional in a two-hybrid assay.
In preferred embodiments of a protein homologous to SEQ ID No: 8, the protein has a molecular weight of approximately 50 kilodaltons, e.g. in the range of 45-55 kD, e.g. in the range of 48-52 kD.
In preferred embodiments: the peptide includes at least 1, 2, 3, or 5, and preferably 10, 20, and 30, amino acid residues from residues 1-133 of Sequence ID No: 8.
Yet another aspect of the present invention concerns an immunogen comprising an E6-BP polypeptide in an immunogenic preparation, the immunogen being capable of eliciting an immune response specific for said E6-BP polypeptide; e.g. a humoral response, e.g. an antibody response; e.g. a cellular response. In preferred embodiments, the immunogen comprising an antigenic determinant, e.g. a unique determinant, from a protein represented by SEQ ID No: 8.
A still further aspect of the present invention features an antibody pre

Affiliated with

Also associated with

Rating

E6 binding proteins does not yet have a rating. At this time, there are no reviews or comments for this patent.

If you have personal experience with E6 binding proteins, we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and E6 binding proteins will most certainly appreciate the feedback.

Rate now

Comments { 0 }

Profile ID: LFUS-PAI-O-2588580