Patent Description:
Bacterial infections are in most instances successfully treated by administration of antibiotics to patients in need thereof. However, due to careless or thoughtless use of powerful antibiotics, many pathological germs become resistant against antibiotics over time. One threatening example is Staphyloccocus aureus. In particular in hospitals this bacterium is of relevance. So-called Methicillin Resistant S. Aureus (MRSA) strains jeopardize patient's survival in hospitals, in particular after surgery.

Vaccination is considered to be a very effective method of preventing infectious diseases in human and veterinary health care. Vaccination is the administration of immungenically effective amounts of antigenic material (the vaccine) to produce immunity to a disease/disease-causing pathogenic agent. Vaccines have contributed to the eradication of smallpox, the near eradication of polio, and the control of a variety of diseases, including rubella, measles, mumps, chickenpox, typhoid fever.

Before "the genomic era", vaccines were based on killed or live attenuated, microorganisms, or parts purified from them. Subunit vaccines are considered as a modern upgrade of these types of vaccine, as the subunit vaccines contain one or more protective antigens, which are more or less the weak spot of the pathogen. Hence, in order to develop subunit vaccines, it is critical to identify the proteins, which are important for inducing protection and to eliminate others.

An antigen is said to be protective if it is able to induce protection from subsequent challenge by a disease-causing infectious agent in an appropriate animal model following immunization.

The empirical approach to subunit vaccine development, which includes several steps, begins with pathogen cultivation, followed by purification into components, and then testing of antigens for protection. Apart from being time and labour consuming, this approach has several limitations that can lead to failure. It is not possible to develop vaccines using this approach for microorganisms, which cannot easily be cultured and only allows for the identification of the antigens, which can be obtained in sufficient quantities. The empirical approach has a tendency to focus on the most abundant proteins, which in some cases are not immuno-protective. In other cases, the antigen expressed during in vivo infection is not expressed during in vitro cultivation. Furthermore, antigen discovery by use of the empirical approach demands an extreme amount of proteins in order to discover the protective antigens, which are like finding needles in the haystack. This renders it a very expensive approach, and it limits the vaccine development around diseases, which is caused by pathogens with a large genome or disease areas, which perform badly in a cost-effective perspective.

It is an object of embodiments of the invention to provide S. aureus derived antigenic polypeptides that may serve as constituents in vaccines against S. aureus infections. It is also an object to provide nucleic acids, vectors, transformed cells, and vaccine compositions for therapy and diagnosis with relevance for S.

It has been found by the present inventor(s) that S. aureus, in particular drug resistant S. aureus, expresses a number of putatively surface exposed proteins which are candidates as vaccine targets as well as candidates as immunizing agents for preparation of antibodies that target S.

So, in a first aspect the present invention relates to a polypeptide for use as a medicament comprising.

In a second aspect, the invention relates to an isolated nucleic acid fragment for use as a medicament, wherein the nucleic acid fragment encodes a polypeptide defined in respect of the first aspect of the invention.

In a third aspect, the invention relates to a vector for use as a medicament, comprising the nucleic acid defined in respect of the second aspect of the invention, such as a cloning vector or an expression vector.

In fourth aspect, the invention relates to a cell for use as a medicament, wherein the cell is transformed so as to carry the vectordefined in respect of the third aspect of the invention.

In a fifth aspect, the invention relates to a pharmaceutical composition comprising a polypeptide defined in respect of the first aspect of the invention, a nucleic acid fragment defined in respect of the second aspect of the invention, a vector defined in respect of the third aspect of the invention, or a transformed cell defined in respect of the fourth aspect of the invention, and further comprising an immunological adjuvant and a pharmaceutically acceptable carrier, vehicle or diluent.

The term "polypeptide" is in the present context intended to mean both short peptides of from <NUM> to <NUM> amino acid residues, oligopeptides of from <NUM> to <NUM> amino acid residues, and polypeptides of more than <NUM> amino acid residues. Further-more, the term is also intended to include proteins, i.e. functional biomolecules comprising at least one polypeptide; when comprising at least two polypeptides, these may form complexes, be covalently linked, or may be non-covalently linked. The polypeptide (s) in a protein can be glycosylated and/or lipidated and/or comprise prosthetic groups.

The term "subsequence" means any consecutive stretch of at least <NUM> amino acids or, when relevant, of at least <NUM> nucleotides, derived directly from a naturally occurring amino acid sequence or nucleic acid sequence, respectively.

The term "amino acid sequence" s the order in which amino acid residues, connected by peptide bonds, lie in the chain in peptides and proteins.

The term "adjuvant" has its usual meaning in the art of vaccine technology, i.e. a substance or a composition of matter which is <NUM>) not in itself capable of mounting a specific immune response against the immunogen of the vaccine, but which is <NUM>) nevertheless capable of enhancing the immune response against the immunogen. Or, in other words, vaccination with the adjuvant alone does not provide an immune response against the immunogen, vaccination with the immunogen may or may not give rise to an immune response against the immunogen, but the combined vaccination with immunogen and adjuvant induces an immune response against the immunogen which is stronger than that induced by the immunogen alone.

"Sequence identity" is in the context of the present invention determined by comparing <NUM> optimally aligned sequences of equal length (e.g. DNA, RNA or amino acid)according to the following formula: (Nref - Ndif)·<NUM>/Nref , wherein Nref is the number of residues in one of the <NUM> sequences and Ndif is the number of residues which are non-identical in the two sequences when they are aligned over their entire lengths and in the same direction. So, two sequences <NUM>'-ATTCGGAACC-<NUM>' and <NUM>'- ATACGGGACC-<NUM>' will provide the sequence identity <NUM>% (Nref=<NUM> and Ndif=<NUM>).

An "assembly of amino acids" means two or more amino acids bound together by physical or chemical means.

The "3D conformation" is the <NUM> dimensional structure of a biomolecule such as a protein. In monomeric polypeptides/proteins, the 3D conformation is also termed "the tertiary structure" and denotes the relative locations in <NUM> dimensional space of the amino acid residues forming the polypeptide.

"An immunogenic carrier" is a molecule or moiety to which an immunogen or a hapten can be coupled in order to enhance or enable the elicitation of an immune response against the immunogen/hapten. Immunogenic carriers are in classical cases relatively large molecules (such as tetanus toxoid, KLH, diphtheria toxoid etc.) which can be fused or conjugated to an immunogen/hapten, which is not sufficiently immunogenic in its own right - typically, the immunogenic carrier is capable of eliciting a strong T-helper lymphocyte response against the combined substance constituted by the immunogen and the immunogenic carrier, and this in turn provides for improved responses against the immungon by B-lymphocytes and cytotoxic lymphocytes. More recently, the large carrier molecules have to a certain extent been substituted by so-called promiscuous T-helper epitopes, i.e. shorter peptides that are recognized by a large fraction of HLA haplotypes in a population, and which elicit T-helper lymphocyte responses.

A "T-helper lymphocyte response" is an immune response elicited on the basis of a peptide, which is able to bind to an MHC class II molecule (e.g. an HLA class II molecule) in an antigen-presenting cell and which stimulates T-helper lymphocytes in an animal species as a consequence of T-cell receptor recognition of the complex between the peptide and the MHC Class II molecule.

An "immunogen" is a substance of matter which is capable of inducing an adaptive immune response in a host, whose immune system is confronted with the immunogen. As such, immunogens are a subset of the larger genus "antigens", which are substances that can be recognized specifically by the immune system (e.g. when bound by antibodies or, alternatively, when fragments of the are antigens bound to MHC molecules are being recognized by T-cell receptors) but which are not necessarily capaple of inducing immunity - an antigen is, however, always capable of eliciting immunity, meaning that a host that has an established memory immunity against the antigen will mount a specific immune response against the antigen.

A "hapten" is a small molecule, which can neither induce or elicit an immune response, but if conjugated to an immunogenic carrier, antibodies or TCRs that recognize the hapten can be induced upon confrontation of the immune system with the hapten carrier conjugate.

An "adaptive immune response" is an immune response in response to confrontation with an antigen or immunogen, where the immune response is specific for antigenc determinants of the antigen/immunogen - examples of adaptive immune responses are induction of antigen specific antibody production or antigen specific induction/activation of T helper lymphocytes or cytotoxic lymphocytes.

A "protective, adaptive immune response" is an antigen-specific immune response induced in a subject as a reaction to immunization (artificial or natural) with an antigen, where the immune response is capable of protecting the subject against subsequent challenges with the antigen or a pathology-related agent that includes the antigen. Typically, prophylactic vaccination aims at establishing a protective adaptive immune response against one or several pathogens.

"Stimulation of the immune system" means that a substance or composition of matter exhibits a general, non-specific immunostimulatory effect. A number of adjuvants and putative adjuvants (such as certain cytokines) share the ability to stimulate the immune system. The result of using an immunostimulating agent is an increased "alertness" of the immune system meaning that simultaneous or subsequent immunization with an immunogen induces a significantly more effective immune response compared to isolated use of the immunogen.

Hybridization under "stringent conditions" is herein defined as hybridization performed under conditions by which a probe will hybridize to its target sequence, to a detectably greater degree than to other sequences. Stringent conditions are target-sequence-dependent and will differ depending on the structure of the polynucleotide. By controlling the stringency of the hybridization and/or washing conditions, target sequences can be identified which are <NUM>% complementary to a probe (homologous probing). Alternatively, stringency conditions can be adjusted to allow some mismatching in sequences so that lower degrees of similarity are detected (heterologous probing). Specificity is typically the function of post-hybridization washes, the critical factors being the ionic strength and temperature of the final wash solution. Generally, stringent wash temperature conditions are selected to be about <NUM> to about <NUM> lower than the melting point (Tm) for the specific sequence at a defined ionic strength and pH. The melting point, or denaturation, of DNA occurs over a narrow temperature range and represents the disruption of the double helix into its complementary single strands. The process is described by the temperature of the midpoint of transition, Tm, which is also called the melting temperature. Formulas are available in the art for the determination of melting temperatures.

The term "animal" is in the present context in general intended to denote an animal species (preferably mammalian), such as Homo sapiens, Canis domesticus, etc. and not just one single animal. However, the term also denotes a population of such an animal species, since it is important that the individuals immunized according to the method of the invention substantially all will mount an immune response against the immunogen of the present invention.

As used herein, the term "antibody"refers to a polypeptide or group of polypeptides composed of at least one antibody combining site. An "antibody combining site" is the three-dimensional binding space with an internal surface shape and charge distribution complementary to the features of an epitope of an antigen, which allows a binding of the antibody with the antigen. "Antibody"includes, for example, vertebrate antibodies, hybrid antibodies, chimeric antibodies, humanised antibodies, altered antibodies, univalent antibodies, Fab proteins, and single domain antibodies.

"Specific binding" denotes binding between two substances which goes beyond binding of either substance to randomly chosen substances and also goes beyond simple association between substances that tend to aggregate because they share the same overall hydrophobicity or hydrophilicity. As such, specific binding usually involves a combination of electrostatic and other interactions between two conformationally complementary areas on the two substances, meaning that the substances can "recognize" each other in a complex mixture.

The term "vector" is used to refer to a carrier nucleic acid molecule into which a heterologous nucleic acid sequence can be inserted for introduction into a cell where it can be replicated and expressed. The term further denotes certain biological vehicles useful for the same purpose, e.g. viral vectors and phage - both these infectious agents are capable of introducing a heterelogous nucleic acid sequence.

The term "expression vector" refers to a vector containing a nucleic acid sequence coding for at least part of a gene product capable of being transcribed. In some cases, when the transcription product is an mRNA molecule, this is in trun translated into a protein, polypeptide, or peptide.

In some embodiments the at least <NUM> contiguous amino acids referred to in option b) in the definition of the first aspect of the invention constitute at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, at least or exactly or at most <NUM>, and at least or exactly or at most <NUM> contiguous amino acid residues.

In some embodiments, the polypeptide of the invention also has a sequence identity with the amino acid sequence of a) defined above of at least <NUM>%, at least <NUM>%, at least <NUM>%, at least <NUM>%, at least <NUM>%, at least <NUM>%, at least <NUM>%, at least <NUM>%, and at least <NUM>%.

Similarly, the polypeptide of the invention in some embodiments also has a sequence identity with the amino acid sequence of b) defined above of at least <NUM>%, at least <NUM>%, at least <NUM>%, at least <NUM>%, at least <NUM>%, at least <NUM>%, at least <NUM>%, at least <NUM>%, and at least <NUM>%.

In the embodiments defined by option b) above, the polypeptide for the use of the invention is also one that has at least or exactly or at most <NUM> contiguous amino acid residues defined for option b) above and also has its N-terminal amino acid residue corresponding to any one of amino acid residues <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>,<NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, and <NUM> in SEQ ID NO: <NUM>, if the length of the at least or exactly or at most <NUM> amino acid residues so permit - if the length of the at least or exactly or at most <NUM> amino acids is higher than <NUM>, the N-terminal first residue will not be higher numbered than N-L+<NUM>, where N is the number of amino acid residues of SEQ ID NO: <NUM> and L is the number of amino acids defined for option b.

The polypeptide for the use of the invention is in certain embodiments also fused or conjugated to an immunogenic carrier molecule; or, phrased otherwise, the polypeptide for the use of the invention also includes such an immunogenic carrier molecule in addition to the material derived from SEQ ID NO. The immunogenic carrier molecule is typically a polypeptide that induces T-helper lymphocyte responses in a majority of humans, such as immunogenic carrier proteins selected from the group consisting of keyhole limpet hemocyanino or a fragment thereof, tetanus toxoid or a fragment thereof, dipththeria toxoid or a fragment thereof. Other suitable carrier molecules are discussed infra. One further fusion partner, which is preferably incorporated is a "His tag", i.e. a stretch of amino acids, which is rich or only consists of histidinyl residues so as to facilitate protein purification.

In preferred embodiments, the polypeptide for the use of the invention detailed above is capable of inducing an adaptive immune response against the polypeptide in a mammal, in particular in a human being. Preferably, the adaptive immune response is a protective adaptive immune response against infection with S. aureus, in particular multi-resistant S. The polypeptide may in these cases induce a humeral and/or a cellular immune response.

SEQ ID NOs: <NUM>-<NUM> include antigenic determinants (epitopes) that are as such recognized by antibodies and/or when bound to MHC molecules by T-cell receptors. For the purposes of the present invention, B-cell epitopes (i.e. antibody binding epitopes) are of particular relevance.

It is relatively uncomplicated to identify linear B-cell epitopes - one very simple approach entails that antibodies raised agains S. aureus or S. aureus derived proteins disclosed herein are tested for binding to overlapping oligomeric peptides derived from any one of SEQ ID NO: <NUM>-<NUM>. Thereby, the regions of the S. aureus polypeptide which are responsible for or contribute to binding to the antibodies can be identified.

Alternatively, or additionally, one can produce mutated versions of the polypeptides for the use of the invention, e.g. versions where each single non-alanine residue in SEQ ID NO. : <NUM> are point mutated to alanine - this method also assists in identifying complex assembled B-cell epitopes; this is the case when binding of the same antibody is modified by exchanging amino acids in different areas of the full-length polypeptide.

Also, in silico methods for B-cell epitope prediction can be employed: useful state-of-the-art systems for β-turn prediction is provided in <NPL>; prediction of linear B-cell epitopes, cf: <NPL>; predictionof solvent exposed amino acids: <NPL>.

The nucleic acid fragment for the use of the invention referred to above is preferably is a DNA fragment (such as SEQ ID NO: <NUM>) or an RNA fragment (such as SEQ ID NOs <NUM>).

It will be understood that the nucleic acid fragments of the invention may be used for both production, carrier and vaccine purposes - the latter will require that the sequences are included in expression vectors that may lead to production of immunogenic proteins in the host animal receiving the vector.

Vectors for the use of the invention fall into several categories discussed infra. One preferred vector for the use of the invention comprises in operable linkage and in the <NUM>'-<NUM>' direction, an expression control region comprising an enhancer/promoter for driving expression of the nucleic acid fragment defined for option i) above, optionally a signal peptide coding sequence, a nucleotide sequence defined for option i), and optionally a terminator. Hence, such a vector constitutes an expression vector useful for effecting production in cells of the polypeptide of the invention. Since the polypeptides for the use of the invention are bacterial of orgin, recombinant production is conveniently effected in bacterial host cells, so here it is preferred that the expression control region drives expression in prokaryotic cell such as a bacterium, e.g. in E coli. However, if the vector is to drive expression in mammalian cell (as would be the case for a DNA vaccine vector), the expression control region should be adapted to this particular use.

At any rate, certain vectors for the use of the invention are capable of autonomous replication.

Also, the vector for the use of the invention may be one that is capable of being integrated into the genome of a host cell - this is particularly useful if the vector is used in the production of stably transformed cells, where the progeny will also include the genetic information introduced via the vector. Alternatively, vectors incapable of being integrated into the genome of a mammalian host cell are useful in e.g. DNA vaccination.

Typically, the vector for the use of the invention is selected from the group consisting of a virus, such as a attenuated virus (which may in itself be useful as a vaccine agent), a bacteriophage, a plasmid, a minichromosome, and a cosmid.

Particularly interesting vectors are viral vectors (in particular those useful as vaccine agents). These may be selected from the group consisting of a retrovirus vector, such as a lentivirus vector, an adenovirus vector, an adeno-associated virus vector, and a pox virus vector. Certain pox virus vectors are preferred, in particular vaccinia virus vectors. A particluarly preferred vaccinia virus vector is a modifed vaccinia Ankara (MVA) vector.

A more detailed discussion of vectors for the use of the invention is provided in the following: Polypeptides for the use of the invention may be encoded by a nucleic acid molecule comprised in a vector. A nucleic acid sequence can be "heterologous," which means that it is in a context foreign to the cell in which the vector is being introduced, which includes a sequence homologous to a sequence in the cell but in a position within the host cell where it is ordinarily not found. Vectors include naked DNAs, RNAs, plasmids, cosmids, viruses (bacteriophage, animal viruses, and plant viruses), and artificial chromosomes (e.g., YACs). One of skill in the art would be well equipped to construct a vector through standard recombinant techniques (for example Sambrook et al, <NUM>; Ausubel et al, <NUM>). In addition to encoding the polypeptides of this invention, a vector of the present invention may encode polypeptide sequences such as a tag or immunogenicity enhancing peptide (e.g. an immunogenic carrier or a fusion partner that stimulates the immune system, such as a cytokine or active fragment thereof). Useful vectors encoding such fusion proteins include pIN vectors (Inouye et al, <NUM>), vectors encoding a stretch of histidines, and pGEX vectors, for use in generating glutathione S-transferase (GST) soluble fusion proteins for later purification and separation or cleavage.

Vectors for the use of the invention may be used in a host cell to produce a polypeptide of the invention that may subsequently be purified for administration to a subject or the vector may be purified for direct administration to a subject for expression of the protein in the subject (as is the case when administering a nucleic acid vaccine).

Expression vectors can contain a variety of "control sequences," which refer to nucleic acid sequences necessary for the transcription and possibly translation of an operably linked coding sequence in a particular host organism. In addition to control sequences that govern transcription and translation, vectors and expression vectors may contain nucleic acid sequences that serve other functions as well and are described infra.

A "promoter" is a control sequence. The promoter is typically a region of a nucleic acid sequence at which initiation and rate of transcription are controlled. It may contain genetic elements at which regulatory proteins and molecules may bind such as RNA polymerase and other transcription factors. The phrases "operatively positioned," "operatively linked," "under control," and "under transcriptional control" mean that a promoter is in a correct functional location and/or orientation in relation to a nucleic acid sequence to control transcriptional initiation and expression of that sequence. A promoter may or may not be used in conjunction with an "enhancer," which refers to a cis-acting regulatory sequence involved in the transcriptional activation of a nucleic acid sequence.

A promoter may be one naturally associated with a gene or sequence, as may be obtained by isolating the <NUM>' non-coding sequences located upstream of the coding segment or exon. Such a promoter can be referred to as "endogenous. " Similarly, an enhancer may be one naturally associated with a nucleic acid sequence, located either downstream or upstream of that sequence. Alternatively, certain advantages will be gained by positioning the coding nucleic acid segment under the control of a recombinant or heterologous promoter, which refers to a promoter that is not normally associated with a nucleic acid sequence in its natural environment. A recombinant or heterologous enhancer refers also to an enhancer not normally associated with a nucleic acid sequence in its natural state. Such promoters or enhancers may include promoters or enhancers of other genes, and promoters or enhancers isolated from any other prokaryotic, viral, or eukaryotic cell, and promoters or enhancers not "naturally occurring," i.e., containing different elements of different transcriptional regulatory regions, and/or mutations that alter expression. In addition to producing nucleic acid sequences of promoters and enhancers synthetically, sequences may be produced using recombinant cloning and/or nucleic acid amplification technology, including PCR™, in connection with the compositions disclosed herein (see <CIT>, <CIT>).

Naturally, it may be important to employ a promoter and/or enhancer that effectively direct(s) the expression of the DNA segment in the cell type or organism chosen for expression. Those of skill in the art of molecular biology generally know the use of promoters, enhancers, and cell type combinations for protein expression (see Sambrook et al, <NUM>).

The promoters employed may be constitutive, tissue-specific, or inducible and in certain embodiments may direct high level expression of the introduced DNA segment under specified conditions, such as large-scale production of recombinant proteins or peptides.

Examples of inducible elements, which are regions of a nucleic acid sequence that can be activated in response to a specific stimulus, include but are not limited to Immunoglobulin Heavy Chain (Banerji et al, <NUM>; Gilles et al, <NUM>; Grosschedl et al, <NUM>; Atchinson et al, <NUM>, <NUM>; toiler et al, <NUM>; Weinberger et al, <NUM>; Kiledjian et al, <NUM>; Porton et al; <NUM>), Immunoglobulin Light Chain (Queen et al, <NUM>; Picard et al, <NUM>), T Cell Receptor (Luria et al, <NUM>; Winoto et al, <NUM>; Redondo et al; <NUM>), HLA DQa and/or DQβ (Sullivan et al, <NUM>), β-Interferon (Goodbourn et al, <NUM>; Fujita et al, <NUM>; Goodbourn et al, <NUM>), Interleukin-<NUM> (Greene et al, <NUM>), Interleukin-<NUM> Receptor (Greene et al, <NUM>; Lin et al, <NUM>), MHC Class II <NUM> (Koch et al, <NUM>), MHC Class II HLA-DRa (Sherman et al, <NUM>), β-Actin (Kawamoto et al, <NUM>; Ng et al; <NUM>), Muscle Creatine Kinase (MCK) (Jaynes et al, <NUM>; Horlick et al, <NUM>; Johnson et al, <NUM>), Prealbumin (Transthyretin) (Costa et al, <NUM>), Elastase I (Omitz et al, <NUM>), Metallothionein (MTII) (Karin et al, <NUM>; Culotta et al, <NUM>), Collagenase (Pinkert et al, <NUM>; Angel et al, <NUM>), Albumin (Pinkert et al, <NUM>; Tranche et al, <NUM>, <NUM>),a-Fetoprotein (Godbout et al, <NUM>; Campere et al, <NUM>), γ-Globin (Bodine et al, <NUM>; Perez-Stable et al, <NUM>),β- Globin (Trudel et al, <NUM>), c-fos (Cohen et al, <NUM>), c-HA-ras (Triesman, <NUM>; Deschamps et al, <NUM>), Insulin (Edlund et al, <NUM>), Neural Cell Adhesion Molecule (NCAM) (Hirsh et al, <NUM>),al-Antitrypain (Larimer et al, <NUM>), H2B (TH2B) Histone (Hwang et al, <NUM>), Mouse and/or Type I Collagen (Ripe et al, <NUM>), Glucose-Regulated Proteins (GRP94 and GRP78) (Chang et al, <NUM>), Rat Growth Hormone (Larsen et al, <NUM>), Human Serum Amyloid A (SAA) (Edbrooke et al, <NUM>), Troponin I (TN I) (Yutzey et al, <NUM>), Platelet-Derived Growth Factor (PDGF) (Pech et al, <NUM>), Duchenne Muscular Dystrophy (Klamut et al, <NUM>), SV40 (Banerji et al, <NUM>; Moreau et al, <NUM> ; Sleigh et al, <NUM>; Firak et al, <NUM>; Herr et al, <NUM>; Imbra et al, <NUM>; Kadesch et al, <NUM>; Wang et al, <NUM>; Ondek et al, <NUM>; Kuhl et al, <NUM>; Schaffner et al, <NUM>), Polyoma (Swartzendruber et al, <NUM>; Vasseur et al, <NUM>; Katinka et al, <NUM>, <NUM>; Tyndell et al, <NUM> ; Dandolo et al, <NUM>; de Villiers et al, <NUM>; Hen et al, <NUM>; Satake et al, <NUM>; Campbell et al, <NUM>), Retroviruses (Kriegler et al, <NUM>, <NUM>; Levinson et al, <NUM>; Kriegler et al, <NUM>, 1984a, b, <NUM>; Bosze et al, <NUM>; Miksicek et al, <NUM>; Celander et al, <NUM>; Thiesen et al, <NUM>; Celander et al, <NUM>; Choi et al, <NUM>; Reisman et al, <NUM>), Papilloma Virus (Campo et al, <NUM>; Lusky et al, <NUM>; Spandidos and Wilkie, <NUM>; Spalholz et al, <NUM>; Lusky et al, <NUM>; Cripe et al, <NUM>; Gloss et al, <NUM>; Hirochika et al, <NUM>; Stephens et al, <NUM>), Hepatitis B Virus (Bulla et al, <NUM>; Jameel et al, <NUM>; Shaul et al, <NUM>; Spandau et al, <NUM>; Vannice et al, <NUM>), Human Immunodeficiency Virus (Muesing et al, <NUM>; Hauber et al, <NUM>; Jakobovits et al, <NUM>; Feng et al, <NUM>; Takebe et al, <NUM>; Rosen et al, <NUM>; Berkhout et al, <NUM>; Laspia et al, <NUM>; Sharp et al, <NUM>; Braddock et al, <NUM>), Cytomegalovirus (CMV) IE (Weber et al, <NUM>; Boshart et al, <NUM>; Foecking et al, <NUM>), Gibbon Ape Leukemia Virus (Holbrook et al, <NUM>; Quinn et al, <NUM>).

Inducible Elements include, but are not limited to MT II - Phorbol Ester (TFA)/Heavy metals (Palmiter et al, <NUM>; Haslinger et al, <NUM>; Searle et al, <NUM>; Stuart et al, <NUM>; Imagawa et al, <NUM>, Karin et al, <NUM>; Angel et al, 1987b; McNeall et al, <NUM>); MMTV (mouse mammary tumor virus) - Glucocorticoids (Huang et al, <NUM>; Lee et al, <NUM>; Majors et al, <NUM>; Chandler et al, <NUM>; Lee et al, <NUM>; Ponta et al, <NUM>; Sakai et al, <NUM>);β-Interferon - poly(rl)x/poly(rc) (Tavernier et al, <NUM>); Adenovirus <NUM> E2 - ElA (Imperiale et al, <NUM>); Collagenase - Phorbol Ester (TPA) (Angel et al, 1987a); Stromelysin - Phorbol Ester (TPA) (Angel et al, 1987b); SV40 - Phorbol Ester (TPA) (Angel et al, 1987b); Murine MX Gene - Interferon, Newcastle Disease Virus (Hug et al, <NUM>); GRP78 Gene - A23187 (Resendez et al, <NUM>);a-<NUM>-Macroglobulin - IL-<NUM> (Kunz et al, <NUM>); Vimentin - Serum (Rittling et al, <NUM>); MHC Class I Gene H-2κb - Interferon (Blanar et al, <NUM>); HSP70 - E1A/SV40 Large T Antigen (Taylor et al, <NUM>, 1990a, 1990b); Proliferin - Phorbol Ester/TPA (Mordacq et al, <NUM>); Tumor Necrosis Factor - PMA (Hensel et al, <NUM>); and Thyroid Stimulating Hormonea Gene - Thyroid Hormone (Chatterjee et al, <NUM>).

Also contemplated as useful in the present invention are the dectin-<NUM> and dectin-<NUM> promoters. Additionally any promoter/enhancer combination (as per the Eukaryotic Promoter Data Base EPDB) could also be used to drive expression of structural genes encoding oligosaccharide processing enzymes, protein folding accessory proteins, selectable marker proteins or a heterologous protein of interest.

The particular promoter that is employed to control the expression of peptide or protein encoding polynucleotide for the use of the invention is not believed to be critical, so long as it is capable of expressing the polynucleotide in a targeted cell, preferably a bacterial cell. Where a human cell is targeted, it is preferable to position the polynucleotide coding region adjacent to and under the control of a promoter that is capable of being expressed in a human cell. Generally speaking, such a promoter might include either a bacterial, human or viral promoter.

In various embodiments, the human cytomegalovirus (CMV) immediate early gene promoter, the SV40 early promoter, and the Rous sarcoma virus long terminal repeat can be used to obtain high level expression of a related polynucleotide to this invention. The use of other viral or mammalian cellular or bacterial phage promoters, which are well known in the art, to achieve expression of polynucleotides is contemplated as well.

In embodiments in which a vector is administered to a subject for expression of the protein, it is contemplated that a desirable promoter for use with the vector is one that is not down-regulated by cytokines or one that is strong enough that even if down-regulated, it produces an effective amount of the protein/polypeptide of the current invention in a subject to elicit an immune response. Non-limiting examples of these are CMV IE and RSV LTR. In other embodiments, a promoter that is up-regulated in the presence of cytokines is employed. The MHC I promoter increases expression in the presence of IFN-y.

Tissue specific promoters can be used, particularly if expression is in cells in which expression of an antigen is desirable, such as dendritic cells or macrophages. The mammalian MHC I and MHC II promoters are examples of such tissue-specific promoters. Initiation Signals and Internal Ribosome Binding Sites (IRES).

A specific initiation signal also may be required for efficient translation of coding sequences. These signals include the ATG initiation codon or adjacent sequences. Exogenous translational control signals, including the ATG initiation codon, may need to be provided. One of ordinary skill in the art would readily be capable of determining this and providing the necessary signals. It is well known that the initiation codon must be "in-frame" with the reading frame of the desired coding sequence to ensure translation of the entire insert. The exogenous translational control signals and initiation codons can be either natural or synthetic and may be operable in bacteria or mammalian cells. The efficiency of expression may be enhanced by the inclusion of appropriate transcription enhancer elements.

In certain embodiments of the invention, the use of internal ribosome entry sites (IRES) elements are used to create multigene, or polycistronic, messages. IRES elements are able to bypass the ribosome scanning model of <NUM>' methylated Cap dependent translation and begin translation at internal sites (Pelletier and Sonenberg, <NUM>). IRES elements from two members of the picornavirus family (polio and encephalomyocarditis) have been described (Pelletier and Sonenberg, <NUM>), as well an IRES from a mammalian message (Macejak and Sarnow, <NUM>). IRES elements can be linked to heterologous open reading frames. Multiple open reading frames can be transcribed together, each separated by an IRES, creating polycistronic messages. By virtue of the IRES element, each open reading frame is accessible to ribosomes for efficient translation. Multiple genes can be efficiently expressed using a single promoter/enhancer to transcribe a single message (see <CIT> and <CIT>).

Vectors can include a multiple cloning site (MCS), which is a nucleic acid region that contains multiple restriction enzyme sites, any of which can be used in conjunction with standard recombinant technology to digest the vector. (See Carbonelli et al, <NUM>, Levenson et al, <NUM>, and Cocea, <NUM>). Frequently, a vector is linearized or fragmented using a restriction enzyme that cuts within the MCS to enable exogenous sequences to be ligated to the vector. Techniques involving restriction enzymes and ligation reactions are well known to those of skill in the art of recombinant technology.

Most transcribed eukaryotic RNA molecules will undergo RNA splicing to remove introns from the primary transcripts. If relevant in the context of vectors of the present invention, vectors containing genomic eukaryotic sequences may require donor and/or acceptor splicing sites to ensure proper processing of the transcript for protein expression. (See Chandler et al, <NUM>).

The vectors or constructs for the use of the present invention will generally comprise at least one termination signal. A "termination signal" or "terminator" is comprised of the DNA sequences involved in specific termination of an RNA transcript by an RNA polymerase. Thus, in certain embodiments a termination signal that ends the production of an RNA transcript is contemplated. A terminator may be necessary in vivo to achieve desirable message levels.

In eukaryotic systems, the terminator region may also comprise specific DNA sequences that permit site-specific cleavage of the new transcript so as to expose a polyadenylation site. This signals a specialized endogenous polymerase to add a stretch of about <NUM> A residues (poly A) to the <NUM>' end of the transcript. RNA molecules modified with this polyA tail appear to more stable and are translated more efficiently. Thus, in other embodiments involving eukaryotes, it is preferred that that terminator comprises a signal for the cleavage of the RNA, and it is more preferred that the terminator signal promotes polyadenylation of the message.

Terminators contemplated for use in the invention include any known terminator of transcription described herein or known to one of ordinary skill in the art, including but not limited to, for example, the bovine growth hormone terminator or viral termination sequences, such as the SV40 terminator. In certain embodiments, the termination signal may be a lack of transcribable or translatable sequence, such as due to a sequence truncation.

In expression, particularly eukaryotic expression (as is relevant in nucleic acid vaccination), one will typically include a polyadenylation signal to effect proper polyadenylation of the transcript. The nature of the polyadenylation signal is not believed to be crucial to the successful practice of the invention, and/or any such sequence may be employed. Preferred embodiments include the SV40 polyadenylation signal and/or the bovine growth hormone polyadenylation signal, convenient and/or known to function well in various target cells. Polyadenylation may increase the stability of the transcript or may facilitate cytoplasmic transport.

In order to propagate a vector in a host cell, it may contain one or more origins of replication sites (often termed "on"), which is a specific nucleic acid sequence at which replication is initiated. Alternatively an autonomously replicating sequence (ARS) can be employed if the host cell is yeast.

In certain embodiments of the invention, cells containing a nucleic acid construct of the present invention may be identified in vitro or in vivo by encoding a screenable or selectable marker in the expression vector. When transcribed and translated, a marker confers an identifiable change to the cell permitting easy identification of cells containing the expression vector. Generally, a selectable marker is one that confers a property that allows for selection. A positive selectable marker is one in which the presence of the marker allows for its selection, while a negative selectable marker is one in which its presence prevents its selection. An example of a positive selectable marker is a drug resistance marker.

Usually the inclusion of a drug selection marker aids in the cloning and identification of transformants, for example, markers that confer resistance to neomycin, puromycin, hygromycin, DHFR, GPT, zeocin or histidinol are useful selectable markers. In addition to markers conferring a phenotype that allows for the discrimination of transformants based on the implementation of conditions, other types of markers including screenable markers such as GFP for colorimetric analysis. Alternatively, screenable enzymes such as herpes simplex virus thymidine kinase (tk) or chloramphenicol acetyltransferase (CAT) may be utilized. One of skill in the art would also know how to employ immunologic markers that can be used in conjunction with FACS analysis. The marker used is not believed to be important, so long as it is capable of being expressed simultaneously with the nucleic acid encoding a protein of the invention. Further examples of selectable and screenable markers are well known to one of skill in the art.

Transformed cells for the use of the invention are useful as organisms for producing the polypeptide of the invention, but also as simple "containers" of nucleic acids and vectors of the invention.

Certain transformed cells of the invention are capable of replicating the nucleic acid fragment defined in respect of the second aspect of the invention. Preferred transformed cells of the invention are capable of expressing the nucleic acid fragment defined in respect of the second aspect of the invention.

For recombinant production it is convenient, but not a prerequisite that the transformed cell is prokaryotic, such as a bacterium, but generally both prokaryotic cells and eukaryotic cells may be used.

Suitable prokaryotic cells are bacterial cells selected from the group consisting of Escherichia (such as E. ), Bacillus [e.g. Bacillus subtilis], Salmonella, and Mycobacterium [preferably non-pathogenic, e.g. M. bovis BCG].

Eukaryotic cells can be in the form of yeasts (such as Saccharomyces cerevisiae) and protozoans. Alternatively, the transformed eukaryotic cells are derived from a multicellular organism such as a fungus, an insect cell, a plant cell, or a mammalian cell.

For production purposes, it is advantageous that the transformed cell is stably transformed by having the nucleic acid defined above stably integrated into its genome, and in certain embodiments it is also preferred that the transformed cell secretes or carries on its surface the polypeptide of the invention, since this facilitates recovery of the polypeptides produced. A particular version is a bacterium and secretion of the polypeptide of the invention is into the periplasmic space.

Further details on cells and cell lines are presented in the following:
Suitable cells for recombinant nucleic acid expression of the nucleic acid fragments of the present invention are prokaryotes and eukaryotes. Examples of prokaryotic cells include E. coli; members of the Staphylococcus genus, such as S. epidermidis; members of the Lactobacillus genus, such as L. plantarum; members of the Lactococcus genus, such as L. lactis; members of the Bacillus genus, such as B. subtilis; members of the Corynebacterium genus such as C. glutamicum; and members of the Pseudomonas genus such as Ps. fluorescens. Examples of eukaryotic cells include mammalian cells; insect cells; yeast cells such as members of the Saccharomyces genus (e.g. S. cerevisiae), members of the Pichia genus (e.g. P. pastoris), members of the Hansenula genus (e.g. H. polymorpha), members of the Kluyveromyces genus (e.g. K. lactis or K. fragilis) and members of the Schizosaccharomyces genus (e.g. S.

Techniques for recombinant gene production, introduction into a cell, and recombinant gene expression are well known in the art. Examples of such techniques are provided in references such as <NPL>, and <NPL>.

As used herein, the terms "cell," "cell line," and "cell culture" may be used interchangeably. All of these terms also include their progeny, which is any and all subsequent generations. It is understood that all progeny may not be identical due to deliberate or inadvertent mutations. In the context of expressing a heterologous nucleic acid sequence, "host cell" refers to a prokaryotic or eukaryotic cell, and it includes any transformable organism that is capable of replicating a vector or expressing a heterologous gene encoded by a vector. A host cell can, and has been, used as a recipient for vectors or viruses. A host cell may be "transfected" or "transformed," which refers to a process by which exogenous nucleic acid, such as a recombinant protein-encoding sequence, is transferred or introduced into the host cell. A transformed cell includes the primary subject cell and its progeny.

Host cells may be derived from prokaryotes or eukaryotes, including bacteria, yeast cells, insect cells, and mammalian cells for replication of the vector or expression of part or all of the nucleic acid sequence(s). Numerous cell lines and cultures are available for use as a host cell, and they can be obtained through the American Type Culture Collection (ATCC), which is an organization that serves as an archive for living cultures and genetic materials (www. org) or from other depository institutions such as Deutsche Sammlung vor Micrroorganismen und Zellkulturen (DSM). An appropriate host can be determined by one of skill in the art based on the vector backbone and the desired result. A plasmid or cosmid, for example, can be introduced into a prokaryote host cell for replication of many vectors or expression of encoded proteins. Bacterial cells used as host cells for vector replication and/or expression include Staphylococcus strains, DH5a, JMl <NUM>, and KC8, as well as a number of commercially available bacterial hosts such as SURE(R) Competent Cells and SOLOP ACK(TM) Gold Cells (STRATAGENE®, La Jolla, CA). Alternatively, bacterial cells such as E. coli LE392 could be used as host cells for phage viruses. Appropriate yeast cells include Saccharomyces cerevisiae, Saccharomyces pombe, and Pichia pastoris.

Examples of eukaryotic host cells for replication and/or expression of a vector include HeLa, NIH3T3, Jurkat, <NUM>, Cos, CHO, Saos, and PC12. Many host cells from various cell types and organisms are available and would be known to one of skill in the art. Similarly, a viral vector may be used in conjunction with either a eukaryotic or prokaryotic host cell, particularly one that is permissive for replication or expression of the vector.

Some vectors may employ control sequences that allow it to be replicated and/or expressed in both prokaryotic and eukaryotic cells. One of skill in the art would further understand the conditions under which to incubate all of the above described host cells to maintain them and to permit replication of a vector. Also understood and known are techniques and conditions that would allow large-scale production of vectors, as well as production of the nucleic acids encoded by vectors and their cognate polypeptides, proteins, or peptides.

Numerous expression systems exist that comprise at least a part or all of the compositions discussed above. Prokaryote- and/or eukaryote-based systems can be employed for use with the present invention to produce nucleic acid sequences, or their cognate polypeptides, proteins and peptides. Many such systems are commercially and widely available.

The insect cell/baculovirus system can produce a high level of protein expression of a heterologous nucleic acid segment, such as described in <CIT>, <CIT>, and which can be bought, for example, under the name MAXBAC® <NUM> from INVITROGEN® and BACPACK™ Baculovirus expression system from CLONTECH®.

In addition to the disclosed expression systems of the invention, other examples of expression systems include STRATAGENE®'s COMPLETE CONTROL™ Inducible Mammalian Expression System, which involves a synthetic ecdysone-inducible receptor, or its pET Expression System, an E. coli expression system. Another example of an inducible expression system is available from INVITROGEN® , which carries the T-REX™ (tetracycline-regulated expression) System, an inducible mammalian expression system that uses the full-length CMV promoter. INVITROGEN® also provides a yeast expression system called the Pichia methanolica Expression System, which is designed for high-level production of recombinant proteins in the methylotrophic yeast Pichia methanolica. One of skill in the art would know how to express a vector, such as an expression construct, to produce a nucleic acid sequence or its cognate polypeptide, protein, or peptide.

Nucleic acids used as a template for amplification may be isolated from cells, tissues or other samples according to standard methodologies (Sambrook et al, <NUM>). In certain embodiments, analysis is performed on whole cell or tissue homogenates or biological fluid samples without substantial purification of the template nucleic acid. The nucleic acid may be genomic DNA or fractionated or whole cell RNA. Where RNA is used, it may be desired to first convert the RNA to a complementary DNA.

The term "primer," as used herein, is meant to encompass any nucleic acid that is capable of priming the synthesis of a nascent nucleic acid in a template-dependent process. Typically, primers are oligonucleotides from ten to twenty and/or thirty base pairs in length, but longer sequences can be employed. Primers may be provided in double-stranded and/or single-stranded form, although the single-stranded form is preferred.

Pairs of primers designed to selectively hybridize to nucleic acids corresponding to sequences of genes identified herein are contacted with the template nucleic acid under conditions that permit selective hybridization. Depending upon the desired application, high stringency hybridization conditions may be selected that will only allow hybridization to sequences that are completely complementary to the primers. In other embodiments, hybridization may occur under reduced stringency to allow for amplification of nucleic acids containing one or more mismatches with the primer sequences. Once hybridized, the template-primer complex is contacted with one or more enzymes that facilitate template-dependent nucleic acid synthesis. Multiple rounds of amplification, also referred to as "cycles," are conducted until a sufficient amount of amplification product is produced.

The amplification product may be detected or quantified. In certain applications, the detection may be performed by visual means. Alternatively, the detection may involve indirect identification of the product via chemiluminescence, radioactive scintigraphy of incorporated radiolabel or fluorescent label or even via a system using electrical and/or thermal impulse signals (Bellus, <NUM>).

A number of template dependent processes are available to amplify the oligonucleotide sequences present in a given template sample. One of the best known amplification methods is the polymerase chain reaction (referred to as PCR(TM)) which is described in detail in <CIT>, <CIT> and <CIT>, and in Innis et al.

Alternative methods for amplification of target nucleic acid sequences that may be used in the practice of the present invention are disclosed in <CIT>, <CIT>, <CIT>,<CIT>, <CIT>, <CIT>, <CIT>,<CIT>,<CIT>,<CIT>,<CIT>, <CIT>, <CIT>,<CIT>,<CIT> and <CIT>, <CIT>, and in <CIT>.

Suitable methods for nucleic acid delivery to effect expression of compositions of the present invention are believed to include virtually any method by which a nucleic acid (e.g., DNA, including viral and nonviral vectors) can be introduced into a cell, a tissue or an organism, as described herein or as would be known to one of ordinary skill in the art. Such methods include, but are not limited to, direct delivery of DNA such as by injection (<CIT>,<CIT>, <CIT>, <CIT>, <CIT>,<CIT>,<CIT>, <CIT> and <CIT>), including microinjection (Harland and Weintraub, <NUM>; <CIT>); by electroporation (<CIT>); by calcium phosphate precipitation (Graham and Van Der Eb, <NUM>; Chen and Okayama, <NUM>; Rippe et al. , <NUM>); by using DEAE dextran followed by polyethylene glycol (Gopal, <NUM>); by direct sonic loading (Fechheimer et al, <NUM>); by liposome mediated transfection (Nicolau and Sene, <NUM>; Fraley et al, <NUM>; Nicolau et al, <NUM>; Wong et al, <NUM>; Kaneda et al, <NUM>; Kato et al, <NUM>); by microprojectile bombardment (<CIT> and <CIT>; <CIT>; <CIT>,<CIT>,<CIT>, <CIT> and <CIT>); by agitation with silicon carbide fibers (Kaeppler et al, <NUM>; <CIT> and <CIT>) by Agrobacterium mediated transformation (<CIT> and <CIT>); or by PEG mediated transformation of protoplasts (Omirulleh et al, <NUM>; <CIT> and <CIT>); by desiccation/inhibition mediated DNA uptake (Potrykus et al, <NUM>). Through the application of techniques such as these, organelle(s), cell(s), tissue(s) or organism(s) may be stably or transiently transformed.

Pharmaceutical compositions, in particular vaccines, according to the invention may either be prophylactic (ie. to prevent infection) or therapeutic (ie, to treat disease after infection).

Such vaccines comprise immunising antigen(s), immunogen(s), polypeptide(s), protein(s) or nucleic acid(s), usually in combination with "pharmaceutically acceptable carriers", which include any carrier that does not itself induce the production of antibodies harmful to the individual receiving the composition.

In some embodiments of the invention, the pharmaceutical compositions such as vaccines include merely one single antigen, immunogen, polypeptide, protein, nucleic acid or vector of the invention, but in other embodiments, the pharmaceutical compositions comprise "cocktails" of the antigens or of the immunogens or of the polypeptides or of the protein or of the nucleic acids or of the vectors of the invention.

In particularly interesting embodiments, the pharmaceutical composition is an MVA vector mentioned herein, which encodes and can effect expression of at least <NUM> nucleic acid fragments of the invention.

Another interesting embodiment of a pharmaceutical composition comprises RNA as the active principle, i.e. at least one mRNA encoding a polypeptide of the invention.

An embodiment of a pharmaceutical composition of the invention comprises Y or at least Y or at most Y distinct polypeptides dsclosed herein, wherein one is derived from SEQ ID NO: <NUM>, and where each of said Y or at least Y or at most Y distinct polypeptides comprises an immunogenic amino acid sequence present in or derived from any one of SEQ ID NOs: <NUM>-<NUM> and wherein said Y or at least Y or at most Y distinct polypeptides together comprise immunogenic amino acid sequences present in or derived from Y or at least Y or at most Y of SEQ ID NOs. <NUM>-<NUM>, wherein Y is an integer selected from <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, and <NUM>.

Another embodiment of the pharmaceutical composition of the invention comprises Z or at least Z or at most Z distinct nucleic acid molecules (such as DNA and RNA) each encoding a polypeptide of the invention, of which one is as defined above for the second aspect of the invention, where each of said Z or at least Z or at most Z distinct nucleic acid molecules encodes an immunogenic amino acid sequence present in or derived from any one of SEQ ID NOs: <NUM>-<NUM> and wherein said at Z or least Z distinct nucleic acid molecules together encode immunogenic amino acid sequences present in or derived from Z or at least Z or at most Z of SEQ ID NOs. <NUM>-<NUM>, wherein Z is an integer selected from <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, and <NUM>.

Suitable carriers are typically large, slowly metabolized macromolecules such as proteins, polysaccharides, polylactic acids, polyglycolic acids, polymeric amino acids, amino acid copolymers, lipid aggregates (such as oil droplets or liposomes), and inactive virus particles.

Such carriers are well known to those of ordinary skill in the art. Additionally, these carriers may function as immunostimulating agents ("adjuvants"). Furthermore, the antigen or immunogen may be conjugated to a bacterial toxoid, such as a toxoid from diphtheria, tetanus, cholera, H. pylori, etc. pathogen, cf. the description of immunogenic carriers supra.

The pharmaceutical compositions of the invention thus typically contain an immunological adjuvant, which is commonly an aluminium based adjuvant or one of the other adjuvants described in the following:
Preferred adjuvants to enhance effectiveness of the composition include, but are not limited to: (<NUM>) aluminum salts (alum), such as aluminum hydroxide, aluminum phosphate, aluminum sulfate, etc; (<NUM>) oil-in-water emulsion formulations (with or without other specific immunostimulating agents such as muramyl peptides (see below) or bacterial cell wall components), such as for example (a) MF59 (<CIT>; <NPL>), containing <NUM>% Squalene, <NUM>% Tween <NUM>, and <NUM>% Span <NUM> (optionally containing various amounts of MTP-PE (see below), although not required) formulated into submicron particles using a microfluidizer such as Model 110Y microfluidizer (Microfluidics, Newton, MA), (b) SAF, containing <NUM>% Squalane, <NUM>% Tween <NUM>, <NUM>% pluronic-blocked polymer L121, and thr-.

MDP (see below) either microfluidized into a submicron emulsion or vortexed to generate a larger particle size emulsion, and (c) Ribi adjuvant system (RAS), (Ribi Immunochem, Hamilton, MT) containing <NUM>% Squalene, <NUM>% Tween <NUM>, and one or more bacterial cell wall components from the group consisting of monophosphoryl lipid A (MPL), trehalose dimycolate (TDM), and cell wall skeleton (CWS), preferably MPL + CWS (DetoxTM) ; (<NUM>) saponin adjuvants such as Stimulon™ (Cambridge Bioscience, Worcester, MA) may be used or particles generated therefrom such as ISCOMs (immunostimulating complexes); (<NUM>) Complete Freund's Adjuvant (CFA) and Incomplete Freund's Adjuvant (IFA); (<NUM>) cytokines, such as interleukins (eg. IL-<NUM>, IL-<NUM>, IL-<NUM>, IL-<NUM>, IL-<NUM>, IL-<NUM>, IL-<NUM>, etc.), interferons (eg. gamma interferon), macrophage colony stimulating factor (M-CSF), tumor necrosis factor (TNF), etc.; and (<NUM>) other substances that act as immunostimulating agents to enhance the effectiveness of the composition. Alum and MF59™ adjuvants are preferred.

As mentioned above, muramyl peptides include, but are not limited to, N-acetyl-muramyl-L-threonyl-D-isoglutamine (thr-MDP), N-acetyl-normuramyl-L-alanyl-D-isoglutamine (nor-MDP), N-acetylmuramyl-L-alanyl-D-isoglutaminyl- L-alanine-<NUM>"-<NUM>'-dipalmitoyl-sn-glycero-<NUM>-hydroxyphosphoryloxy)-ethylamine (MTP-PE), etc..

The immunogenic compositions (eg. the immunising antigen or immunogen or polypeptide or protein or nucleic acid, pharmaceutically acceptable carrier, and adjuvant) typically will contain diluents, such as water, saline, glycerol, ethanol, etc. Additionally, auxiliary substances, such as wetting or emulsifying agents, pH buffering substances, and the like, may be present in such vehicles.

Typically, the immunogenic compositions are prepared as injectables, either as liquid solutions or suspensions; solid forms suitable for solution in, or suspension in, liquid vehicles prior to injection may also be prepared. The preparation also may be emulsified or encapsulated in liposomes for enhanced adjuvant effect, as discussed above under pharmaceutically acceptable carriers.

Immunogenic compositions used as vaccines comprise an immunologically effective amount of the antigenic or immunogenic polypeptides, as well as any other of the above-mentioned components, as needed. By "immunollogically effective amount", it is meant that the administration of that amount to an individual, either in a single dose or as part of a series, is effective for treatment or prevention. This amount varies depending upon the health and physical condition of the individual to be treated, the taxonomic group of individual to be treated (eg. nonhuma primate, primate, etc.), the capacity of the individual's immune system to synthesize antibodies or generally mount an immune response, the degree of protection desired, the formulation of the vaccine, the treating doctor's assessment of the medical situation, and other relevant factors. It is expected that the amount will fall in a relatively broad range that can be determined through routine trials. However, for the purposes of protein vaccination, the amount administered per immunization is typically in the range between <NUM>µg and <NUM> (however, oftn not higher than <NUM>,<NUM>µg), and very often in the range between <NUM> and <NUM>µg.

The immunogenic compositions are conventionally administered parenterally, eg, by injection, either subcutaneously, intramuscularly, or transdermally/transcutaneously (eg. Additional formulations suitable for other modes of administration include oral and pulmonary formulations, suppositories, and transdermal applications. In the case of nucleic acid vaccination, also the intravenous or intraarterial routes may be applicable.

Dosage treatment may be a single dose schedule or a multiple dose schedule. The vaccine may be administered in conjunction with other immunoregulatory agents.

As an alternative to protein-based vaccines, DNA vaccination (also termed nucleic acid vaccination or gene vaccination) may be used [eg. <NPL>; <NPL>; later herein].

A further aspect of the invention is as mentioned above the recognition that combination vaccines can be provided, wherein <NUM> or more antigens disclosed herein are combined to enhance the immune response by the vaccinated animal, including to optimize initial immune response and duration of immunity. For the purposes of this aspect of the invention, multiple antigenic fragments derived from the same, longer protein can also be used, such as the use of a combination of different lengths of polypeptide sequence fragments from one protein.

Also, embodiments of the invention relate to a composition (or the use as a vaccine thereof) comprising <NUM> distinct (i.e. non-identical) proteinaceaous immunogens disclosed herein wherein the first of said immunogens is SEQ ID NO: <NUM> or a variant or fragment thereof disclosed herein in combination with a proteinaceous immunogen selected from any one of SEQ ID NOs: <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, and <NUM> or in combination with a variant or fragment disclosed herein of any one of SEQ ID NOs: <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, and <NUM>.

Treatment methods enabled generally relate to induction of immunity and as such also entail methods that relate to treatment, prophylaxis and amelioration of disease.

When immunization methods entail that a polypeptide for the use of the invention or a composition comprising such a polypeptide is administered the animal (e.g. the human) typically receives between <NUM> and <NUM>,<NUM>µg of the polypeptide of the invention per administration.

In preferred embodiments, the immuniation scheme includes that the animal (e.g. the human) receives a priming administration and one or more booster administrations.

Preferred embodimentms comprise that the administration is for the purpose of inducing protective immunity against S. In this embodiment it is particularly preferred that the protective immunity is effective in reducing the risk of attracting infection with S. aureus or is effective in treating or ameliorating infection with S.

As mentioned herein, the preferred vaccines of the invention induce humoral immunity, so it is preferred that the administration is for the purpose of inducing antibodies specific for S. aureus and wherein said antibodies or B-lymphocytes producing said antibodies are subsequently recovered from the animal.

Immunization methods disclosed herein may also be useful in antibody production, so in other embodiments the administration is for the purpose of inducing antibodies specific for S. aureus and wherrein B-lymphocytes producing said antibodies are subsequently recovered from the animal and used for preparation of monoclonal antibodies.

Pharmaceutical compositions can as mentioned above comprise polypeptides or nucleic acids of the invention. The pharmaceutical compositions will comprise a therapeutically effective amount thereof.

The term "therapeutically effective amount" or "prophylactically effective amount" as used herein refers to an amount of a therapeutic agent to treat, ameliorate, or prevent a desired disease or condition, or to exhibit a detectable therapeutic or preventative effect. The effect can be detected by, for example, chemical markers or antigen levels. Therapeutic effects also include reduction in physical symptoms, such as decreased body temperature. The precise effective amount for a subject will depend upon the subject's size and health, the nature and extent of the condition, and the therapeutics or combination of therapeutics selected for administration. Thus, it is not useful to specify an exact effective amount in advance. Reference is however made to the ranges for dosages of immunologically effective amounts of polypeptides, cf.

However, the effective amount for a given situation can be determined by routine experimentation and is within the judgement of the clinician.

For purposes of the present invention, an effective dose will be from about <NUM>/kg to <NUM>/kg or <NUM>/kg to about <NUM>/kg of the DNA constructs in the individual to which it is administered.

A pharmaceutical composition can also contain a pharmaceutically acceptable carrier. The term "pharmaceutically acceptable carrier" refers to a carrier for administration of a therapeutic agent, such as antibodies or a polypeptide, genes, and other therapeutic agents. The term refers to any pharmaceutical carrier that does not itself induce the production of antibodies harmful to the individual receiving the composition, and which may be administered without undue toxicity. Suitable carriers may be large, slowly metabolized macromolecules such as proteins, polysaccharides, polylactic acids, polyglycolic acids, polymeric amino acids, amino acid copolymers, and inactive virus particles. Such carriers are well known to those of ordinary skill in the art.

Pharmaceutically acceptable salts can be used therein, for example, mineral acid salts such as hydrochlorides, hydrobromides, phosphates, sulfates, and the like; and the salts of organic acids such as acetates, propionates, malonates, benzoates, and the like. A thorough discussion of pharmaceutically acceptable excipients is available in <NPL>).

Pharmaceutically acceptable carriers in therapeutic compositions may contain liquids such as water, saline, glycerol and ethanol. Additionally, auxiliary substances, such as wetting or emulsifying agents, pH buffering substances, and the like, may be present in such vehicles. Typically, the therapeutic compositions are prepared as injectables, either as liquid solutions or suspensions; solid forms suitable for solution in, or suspension in, liquid vehicles prior to injection may also be prepared. Liposomes are included within the definition of a pharmaceutically acceptable carrier.

<NUM> months old NMRI mice were used.

Groups of at least <NUM> mice were used for immunization. The mice were immunized <NUM> times (at day <NUM>, <NUM>, and <NUM>) prior to challenge infection. A control group was treated according to an identical protocol with the exception that an irrelevant protein antigen or phosphate buffered saline was used for immunization.

<NUM>µg protein (per mice) was mixed with <NUM>µl aluminum hydroxide (Alhydrogel <NUM>%, Brenntag) per <NUM> ug protein and incubated with end-over-end rotation for <NUM>. Freund's incomplete adjuvant (sigma) was added in the volume <NUM>:<NUM> and the mixture was vortexed thoroughly for <NUM> hour. This mixture was injected subcutaneously.

The mice were booser injected intraperitoneally with <NUM> weeks interval, using the same amount of protein mixed with aluminum hydroxide and physiological saline solution.

<NUM> days after the last immunization, a number of bacteria (<NUM> x <NUM><NUM> cells) corresponding to a predetermined LD<NUM> in the control group of mice was administered intraperitoneally to all mice.

The cells were handled cold and kept on ice until use. The stock solution of MRSA cells were thawed on ice and then the appropriate amount of cells was diluted in sterile physiological saline (total volume per mouse <NUM>µl).

The survival was surveilled twice daily in the first <NUM> hours after challenge and once daily in the subsequent <NUM> days. The mice were sacrificed if they showed signs of suffering. The mice were monitored with respect to loss of weight and body temperature using an implanted chip. The organs of the mice were used for determination of CFU counts for for SAR1402 (SEQ ID NO: <NUM>), SAR <NUM> (SEQ ID NO: <NUM>) and SAR2753 (SEQ ID NO: <NUM>).

The results from <NUM> polypeptide vaccinations are presented in the Figures.

<FIG> shows the survival curves for <NUM> mice immunized with full-length SAR1402 (SEQ ID NO: <NUM>) and <NUM> mice immunized with negative control. Survival in the vaccinated group at day <NUM> was <NUM>/<NUM> mice, whereas only <NUM>/<NUM> mice in the control group survived. The increased survival in the vaccinated group is statistically highly significant (P=<NUM> according to a Log-rank Mantel-Cox test).

<FIG> shows the survival curves for <NUM> mice immunized with amino acids <NUM>-<NUM> of SAR2496 (SEQ ID NO: <NUM>) and <NUM> mice immunized with negative control. Survival in the vaccinated group at day <NUM> was <NUM>/<NUM> mice, whereas only <NUM>/<NUM> mice in the control group survived. The increased survival in the vaccinated group is statistically highly significant (P=<NUM> according to a Log-rank Mantel-Cox test).

<FIG> shows the survival curves for <NUM> mice immunized with amino acids <NUM>-<NUM> of SAR2723 (SEQ ID NO: <NUM>) and <NUM> mice immunized with negative control. Survival in the vaccinated group at day <NUM> was <NUM>/<NUM> mice, whereas only <NUM>/<NUM> mice in the control group survived. The increased survival in the vaccinated group is statistically highly significant (P=<NUM> according to a Log-rank Mantel-Cox test).

<FIG> shows the survival curves for <NUM> mice immunized with amino acids <NUM>-<NUM> of SAR2753 (SEQ ID NO: <NUM>) and <NUM> mice immunized with negative control. Survival in the vaccinated group at day <NUM> was <NUM>/<NUM> mice, whereas only <NUM>/<NUM> mice in the control group survived. The increased survival in the vaccinated group is statistically highly significant (P=<NUM> according to a Log-rank Mantel-Cox test).

<FIG> shows the survival curves for <NUM> mice immunized with amino acids <NUM>-<NUM> of SAR2753 (SEQ ID NO: <NUM>) and <NUM> mice immunized with negative control (phosphate buffered saline, PBS). Survival in the vaccinated group at day <NUM> was <NUM>/<NUM> mice, whereas only <NUM>/<NUM> mice in the control group survived. The increased survival in the vaccinated group is statistically highly significant (P=<NUM> according to a Log-rank Mantel-Cox test).

<FIG> shows the survival curves for <NUM> mice immunized with a homologue of SAR2716 (USA300HOU_2637_28 <NUM> shown in SEQ ID NO: <NUM>) and <NUM> mice immunized with negative control (PBS). Survival in the vaccinated group at day <NUM> was <NUM>/<NUM> mice, whereas only <NUM>/<NUM> mice in the control group survived. The increased survival in the vaccinated group is statistically highly significant (P=<NUM> according to a Log-rank Mantel-Cox test). USA300HOU_2637_28 <NUM> has a sequence identity with SEQ ID NO: <NUM>, residues <NUM>-<NUM>, of <NUM>%.

<FIG> shows the survival curves for <NUM> mice immunized with amino acids <NUM>-<NUM> of SAR1795 (SEQ ID NO: <NUM>) and <NUM> mice immunized with negative control (PBS). Survival in the vaccinated group at day <NUM> was <NUM>/<NUM> mice, whereas only <NUM>/<NUM> mice in the control group survived. The increased survival in the vaccinated group is statistically highly significant (P=<NUM> according to a Log-rank Mantel-Cox test).

The sequence listing included sets forth the sequences of polypeptides and nucleic acids of the present invention. For easy reference, the sequences are presented in the following:.

The polypeptides of the present disclosure have the following amino acid sequences:.

The nucleic acid fragments of the present disclosure have the following sequences:.

The polypeptides of the present disclosure are also designated as follows herein:.

Claim 1:
A polypeptide for use as a medicament, said polypeptide comprising
a) an amino acid sequence SEQ ID NO: <NUM>, or
b) an amino acid sequence consisting of at least or exactly <NUM> contiguous amino acid residues of SEQ ID NOs: <NUM>, or
c) an amino acid sequence having a sequence identity of at least <NUM>% with the amino acid sequence of a), or
d) an amino acid sequence having a sequence identity of at least <NUM>% with the amino acid sequence of b)
said polypeptide being antigenic in a mammal.