Papillomavirus polyprotein constructs

Details Papillomavirus polyprotein constructs Papillomavirus polyprotein constructs

1998-04-21

2002-04-02

Salimi, Ali R. (Department: 1648)

Drug, bio-affecting and body treating compositions

Antigen, epitope, or other immunospecific immunoeffector

Fusion protein or fusion polypeptide

C424S199100, C424S204100, C424S186100, C435S320100, C435S235100, C435S069100, C435S069700, C536S023720, C530S350000

Reexamination Certificate

active

06365160

ABSTRACT:

FIELD OF THE INVENTION
This invention relates to polyprotein constructs and in particular polyprotein constructs comprising a plurality of papillomavirus (PV) amino acid sequences which may be used in compositions for eliciting an immune response against PV, and particularly human papillomavirus (HPV), in a host animal.
BACKGROUND OF THE INVENTION
Papillomaviruses induce benign hyperproliferative lesions in humans and in many animal species, some of which undergo malignant conversion. The biology of papillomavirus infection is summarised in a review by J. P. Sundberg, entitled “Papillomavirus Infections in Animals” In “Papillomaviruses and Human Disease” edited by K. Syrjanen, L. Gissmann and L. G. Koss, Springer Verlag (1987).
Papillomaviruses are a family of small DNA viruses encoding up to eight early (E1, E2, E3, E4, E5, E6, E7 and E8) and two late genes (L1 and L2). These viruses have been classified in several distinct groups such as HPV which are differentiated into types 1 to ~70 depending upon DNA sequence homology. A clinicopathological grouping of HPV and the malignant potential of the lesions with which they are most frequently associated are summarised in “Papillomaviruses and Human Cancer” by H. Pfister, CRC Press, Inc. (1990). For example, HPV type 1 (HPV-1) is present in plantar warts, HPV-6 or HPV-11 are associated with condylomata acuminata (anogenital warts), and HPV-16 or HPV-18 are common in pre-malignant and malignant lesions of the cervical squamous epithelium.
The immunological approach to the prevention of HPV disease requires a thorough analysis of the viral proteins against which humoral and cellular immune responses are mounted during and after infection. However, despite recent limited success (Kreider et al., 1986,
J. Virol.,
59, 369; Sterling et al., 1990,
J. Virol.,
64, 6305; Meyers et al., 1992,
Science,
257, 971; Dollard et al., 1992,
Genes and Development,
6, 1131), papillomaviruses are notoriously refractory to growth in cultured cells (Teichaman and LaPorta, 1987 In “The Papovaviridae”, Vol 2 edited by N. P. Salzman and P.M. Howley, p.109). As a consequence, the lack of viral reagents has delayed the analysis of the immune response to PV infection.
The recent advent of recombinant expression systems in vitro has allowed the production of viral proteins encoded by both early and late genes in relatively large amounts and in a purified form (Tindle et al., 1990,
J. Gen. Virol.,
71, 1347; Jarrett et al., 1991,
Virology,
184, 33; Ghim et al., 1992,
Virology,
190, 548; Stacey et al., 1991,
J. Gen. Virol.,
73, 2337). These systems have, for the first time, allowed the analysis of the host immune response to these viral proteins.
Interest in immune responses to the non-structural early open reading frame (ORF) proteins of HPV has centred on HPV-16 E7 because of an apparent association between serum antibodies to this protein and cervical cancer (for a review, see “Immune Response to Human Papillomaviruses and the Prospects of Human Papillomavirus-Specific Immunisation” by Tindle and Frazer In “Human Pathogenic Papillomaviruses” edited by H. zur Hausen, Current Topics in Microbiology Immunology, 186, Springer-Verlag, Berlin, 1994).
The immune responses to other HPV early ORF proteins have also been investigated including HPV-16 E6 (Stacey et al., 1992,
J. Gen. Virol.,
73, 2337; Bleul et al., 1991,
J. Clin. Microbiol.,
29, 1579; Dillner, 1990,
Int. J. Cancer,
46, 703; and Müller et al., 1992,
Virology,
187, 508), HPV-16 E2 (Dillner et al., 1989
Proc.Natl. Acad. Sci.USA,
86, 3838; Dillner, 1990, supra; Lehtinen et al., 1992,
J. Med. Virol.,
37, 180; Mann et al., 1990,
Cancer Res.,
50, 7815; and Jenison et al., 1990,
J. Infect. Dis.,
162, 60) and HPV-16 E4 (Köchel et al., 1991,
Int. J. Cancer,
48, 682; Jochmus-Kudielka et al., 1989,
JNCI,
81, 1698; and Barber et al., 1992,
Cancer Immunol. Immunother.,
35, 33). However, comparison of these studies reveals a lack of correlation between the results of the various assays which have been used in assessing HPV early ORF protein reactivity in serum (Tindle and Frazer, 1994, supra).
In addition, antibodies to other HPV early ORF proteins have not yet been sought with sufficient rigour in large enough numbers of patients to determine their utility as disease markers or as indicators of HPV protein immunogenicity following HPV infection.
A problem associated with immunising animals with preparations of individual PV proteins is that most of these proteins are comparatively small and might therefore not comprise many reactive epitopes. In addition, immunodominance of particular B or T cell epitopes within a single PV protein would vary presumably between animals of different major histocompatibility (MHC) backgrounds. To this end, the efficacy of such immunogens, in respect of eliciting an immune response against PV, might be expected to differ between animals of diverse MHC background.
In addition, there is surprisingly little knowledge regarding which PV proteins are expressed by infected cells at various stages of differentiation, and hence it is not possible to predict which proteins will be responsible for defining appropriate immunological targets.
The present invention provides a polyprotein construct comprising a plurality of PV early ORF proteins in one fused or linked construct to improve the efficacy of immune stimulation against PV infection and to avoid the need to define specific immunological targets.
SUMMARY OF THE INVENTION
In one aspect, the present invention provides as an isolated product, a polyprotein construct comprising at least two amino acid sequences fused directly or indirectly together, each of said sequences being the sequence of an early open reading frame (ORF) protein of papillomavirus (PV) or an immunogenic variant or fragment thereof, and at least one of said sequences being other than the E6 or E7 protein sequence or an immunogenic variant or fragment thereof.
In yet another aspect, the present invention provides a composition for eliciting a humoral and/or cellular immune response against PV in a host animal, said composition comprising an immunologically effective amount of a construct as described above, together with a pharmaceutically acceptable carrier and/or diluent.
In yet another aspect, this invention provides a method for eliciting a humoral and/or cellular response against PV in a host animal, which method comprises administering to the host animal an immunologically effective amount of a polyprotein construct as described above. In a related aspect, the invention also extends to use of such a polyprotein construct in eliciting an immune response against PV in a host animal. Preferably, the host animal is a human, however the host animal may also be a non-human mammal.
The present invention also extends to a nucleic acid molecule which encodes a polypeptide construct as broadly described above. Such a nucleic acid molecule may be delivered to a host animal in a nucleic acid vaccine composition with a pharmaceutically acceptable carrier and/or diluent, for expression of the encoded polyprotein construct in vivo in a host animal. Alternatively, the nucleic acid molecule may be included in a recombinant DNA molecule comprising an expression control sequence operatively linked to the nucleic acid molecule.
Throughout this specification and the claims which follow, unless the context requires otherwise, the word “comprise”, or variations such as “comprises” or “comprising”, will be understood to imply the inclusion of a stated integer or group or integers but not the exclusion of any other integer or group of integers.”
DETAILED DESCRIPTION OF THE INVENTION
The term “polyprotein construct” as used herein is used to describe a protein construct made up of individual proteins that have been joined together in a sequence whereby they retain their original relevant biological activities.
The term “isolated” as used herein denotes that the polyprotein construct has undergone at least one purification or isolation step, and preferably is

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