BR102014011184A2 - immunogenic composition, methods of treatment or prophylaxis of infection, to reduce the risk of HIV transmission, and to produce a viral composition, kit, and vector - Google Patents

Abstract

immunogenic composition, methods of treating or prophylaxis of infection, to reduce the risk of HIV transmission, and to produce a viral composition, kit, and vector. The present invention relates to particular immunogenic compositions comprising a gp120-related polypeptide and a adjuvant, wherein the adjuvant comprises a saponin and a lipopolysaccharide. such compositions are substantially free of a neftat-related polypeptide, comprise from 10-40ug of a lipopolysaccharide and from 10-40ug of an immunologically active saponin fraction derived from quillaja saponaria molina presented in the form of a liposome or have a concentration of sodium chloride 130 mm or less.

Description

"IMMUNOGENIC COMPOSITION, METHODS OF TREATMENT OR PROPHYLAXIS OF INFECTION, TO REDUCE THE RISK OF HIV TRANSMISSION, AND TO PRODUCE A COMPOSITION, KIT, AND, VECTOR VIR AL" FIELD OF THE INVENTION

The present invention relates to immunogenic compositions comprising a gp120 related polypeptide and an adjuvant, wherein the adjuvant comprises a saponin and a lipopolysaccharide. Methods for preparing such immunogenic compositions and related kits are also provided.

BACKGROUND OF THE INVENTION

[0002] HIV is the leading cause of Acquired Immunodeficiency Syndrome (AIDS), which is considered as one of the major health problems in the world. There were approximately 34 million people living with HIV in 2011 (WHO HIV / AIDS Fact Report Number 360, June 2013) and over 4 million new infections occur each year. HIV has cost more than 25 million lives over the last three decades. Although antiretroviral therapy is prolonging the lives of many people infected with HIV, effective antiretroviral therapy requires strict adherence to multiple, often complex, non-curing drug regimens. New infections are far outnumbering those able to be treated through current global financial efforts. There is a need for a vaccine to prevent new infections, but developing a safe and effective HIV vaccine poses a significant challenge.

Two types of HIV have been characterized: HI-1 and HIV-2. HIV-1 is highly virulent and infectious and is the cause of most HIV infections worldwide, while HIV-2 has lower virulence and / and infectivity and is largely confined to West Africa (Gilbert et al., Stat In Med. 22 (4): 573-593 (2003) and Reeves and Doms J Gen Vir 83: 12531265 (2002)). There are many genetically distinct subtypes (also known as HIV-1 clades) and the amino acid sequences of only envelope glycoproteins (gp120 and gp41) can range from 25-30% between subtypes (Kalish et al., AIDS 9: 851-857 (1995)). The genetic diversity of HIV-1 along with the high mutation rate are major obstacles to the development of HIV-1 vaccines.

Although extensive research around the world has been conducted to produce a vaccine, much work is still needed.

The HIV gp120 envelope glycoprotein and other HIV1 gp120 proteins is the viral protein that is used for attachment of a host cell. This fixation is mediated by helper T cell and macrophage gp120 binding surface molecules, including one of two CCR-5 or CXCR-4 chemokine receptors. The gp120 protein is first expressed as a larger precursor molecule (gp160), which is then post-translationally cleaved to give gp120 and gp41. The gp120 protein is retained on the virion surface, non-covalently associated with the gp41 molecule, which is inserted into the viral membrane. Three non-covalently associated envelope glycoprotein heterodimers of gp120 and gp41 form the trimeric Env peak found on the surface of HIV-1.

Non-envelope proteins of HIV-1 have been described and include, for example, internal structural proteins, such as other Env gene products (such as gpl60 and gp41), gag and pol gene products (such as MA, CA, SP1, NC, SP2 and P6, and RT, RNase H, IN and RP, respectively) and other nonstructural proteins such as Rev, Nef, Vif, Vpr, Vpu and Tat (Greene et al., New Eng J Med , 324, 5, 308 et seq (1991) and Bryant et al. (Ed. Pizzo), Pediatr Infect Dis J, 11.5, 390 et seq (1992)). PREVIOUS FIELD WORKS

[0007] gpl20 was one of the first targets for HIV vaccine research, and was found to be useful as an antigenic component in vaccines designed to elicit cell-mediated immune responses. The gp120 protein contains epitopes that are recognized by cytotoxic T lymphocytes (CTL). These effector cells are capable of eliminating virus-infected cells and therefore constitute an antiviral immune mechanism. Some epitopes of CTL appear to be relatively conserved between different strains of HIV. However, standard CTL-based vaccines do not protect against infection in animal models of HIV (Amara et al., Science 292 (5514): 69-74 (2001)). In addition, CTL-based vaccines studied so far in humans have not induced these intended responses in all recipients (Goepfert et al., J Infect Dis 192 (7): 1249-1259 (2005)). These CTL-based vaccines were unable to protect against infection or impact viral load after infection (the “HVTN -505 study”).

The gpl20 protein is a major target of neutralizing antibodies (Pantophlet et al., Annu Rev Immunol 24: 739-769 (2006)) and neutralizing antibodies have previously been thought to be a potential protective correlate (Bruck et al. , Vaccine 12 (12): 1141-1148 (1994) and Plotkin, Pediatric Infect Dis 20: 63-75 (2001)).

One region of the gp120 protein, in particular the "V3 loop", is targeted by neutralizing antibodies. Antibodies present in the immune serum of infected individuals bind to V3 loop peptides (Spenlehauer et al, J Vir, 72 (12): 98559864 (1998)). Therefore, a lot of research work has focused on the V3 loop of the gpl20 protein. However, the V3 loop is unfortunately very variable and highly strain specific (Vaine et al., PLoS One, 5 (11): E13916, (2010), Jones et al., J Infect Dis, 179: 558-566 (1999). ) and McCormack et al., Vaccine 18 (13): 1166-1177 (2000)).

It has been shown that vaccination with gpl20 does not always induce neutralizing antibodies against HIV and SHIV, and when neutralizing antibodies are induced, these antibodies rarely confer protection against divergent viruses (Voss et al., J Vir 77 (2): 1049-1058 (2003), Plotkin, Pediatric Infect Dis 20: 63-75 (2001), Wren and Kent, Hum Vacc 7 (4): 466-473 (2011)).

Use of recombinant HIV-1 NefTat, gpl20w6iD and SIV Nef proteins formulated with the AS02A adjuvant system (50ug QS-21 and 50ug 3D-MPL in an oil-in-water emulsion) has been shown to protect against AIDS in a SHIV rhesus monkey animal model system when animals were challenged with SIV / HIV strain SHIV89.6p. However, HIV-1 gpl20W61D formulated alone with AS02A did not provide protection. The lack of protection after immunization with gp120W61D formulated alone with AS02A was suggested to be related to the heterologous nature of the challenge virus (20.2% sequence difference for gp120), since HIV-1 formulated in gp120W61D AS02A had previously been shown to induce sterile immunity against the SHIVW61D challenge (homologous (Voss et al, J Virol 77 (2): 1049-1058 (2003) and Mooij et al., AIDS 12: F15-F22 (1998)).

NefTat and gpl20w6iD formulated in AS02A were administered to HIV seronegative subjects in a safe and immunogenicity study. However, the neutralizing antibodies elicited by the vaccine showed poor subtype cross-reactivity (Leroux-Roels et al., Vaccine 28: 7016-7024 (2010) - the “PRO HIV-002” experiment). In addition, gpl20 had a negative impact on antibody and T-cell responses (geometric mean antibody titer and lymphocyte proliferation) when comparing administration of NefTat and gpl20 against NefTat alone (Goepfert et al, Vaccine 25: 510- 518 (2007) - the “study HVTN-041”). Thus, it was found that (i) a vaccine comprising gp120 with an adjuvant comprising QS-21 and 3D-MPL had poor subtype cross-reactivity and (ii) it was found that gp120 could have a detrimental impact on previously accepted potential markers. for efficacy for HIV vaccines containing other antigens.

V

In light of the above illustrated findings, the use of gp120 as a vaccine antigen to elicit a humoral response (particularly when administered with an adjuvant comprising QS-21 and 3D-MPL) was thought to be of limited use for a vaccine. broadly protective and, as a result, interest in this protein declined.

The RV144 HIV-1 vaccine trial was the first to show evidence of protection against HIV-1 infection, with an estimated vaccine efficacy of 31.2% (Rerks-Ngarm et al, N Engl J Med 361 : 2209-2220 (2009)). The protocol consisted of four ALVAC-HIV initiation injections (vCP1521), and two AIDS VAX B / E booster injections. ALVAC-HIV was administered at baseline (day 0), 4 weeks, 12 weeks and 24 weeks. AIDS VAX B / E was administered at weeks 12 and 24. ALVAC-HIV (vCP1521) is a recombinant canarypox viral vector containing the HIV-1 subtype gp120 (CRF01__AE) strain 92TH023, bound to the transmembrane anchoring moiety. gp41 (carrying a deletion in the immunodominant region) of the HIV-1 subtype B strain LAI. The vector also contained HIVLAI gag and pol genes. AIDS VAX B / E is a recombinant HIV-1 subtype B preparation MN gp120, CM244 subtype E and A244 gp120 and an aluminum adjuvant.

In order to identify correlates of risk of HIV-1 infection in RV144, plasma samples were analyzed from study participants with RV144 (Haynes et al., N Engl J Med 366: 1275-1286 (2012)). ).

Tests were performed on samples obtained two weeks after the final immunization from 41 vaccinated individuals who became infected and 205 uninfected vaccines. These tests examined the role of T-cell, IgG antibody, and IgA antibody responses in modulating the risk of infection. The levels of gp70-VlV2-specific antibodies (a scaffold protein) carrying the first and second gp70-fused murine leukemia virus-gp20 fusion regions-Pinter et al, Vaccine 16 ( 19): 1803-1811. (1998)) were correlated with a lower risk of infection and the binding of plasma IgA antibodies to envelope proteins correlated with a higher risk of infection. In addition, once the analysis was performed, RV144-induced V2 antibodies were found to cross-react with multiple subtypes of HIV-1 (Zolla-Pazner et al., PlosOne, 8 (1): E53629 ( 2013)). Therefore, for the above reasons, it was concluded that vaccines that induce higher levels of VIV2 antibodies may have improved efficacy against HIV-1 infection.

In order to produce an HIV vaccine it is therefore highly desirable to identify a composition capable of eliciting a high level of gp120 VIV2 region specific antibodies.

To summarize, previous work in the field has shown that: • gpl20 in AS02A adjuvant (QS-21 and 3D-MPL oil-in-water emulsion) does not provide protection against heterologous virus challenge in a primate model, • gpl20 may have a detrimental impact on vaccine efficacy markers and antibodies against the gp120 V1V2 region are a protective correlate for HIV-1.

The immunogenic compositions of the present invention may have one or more of the following advantages compared to prior art compositions: (i) achieve a stronger humoral immune response, for example, a higher serum titer of antibodies binding to g120 V1V2 region, (ii) achieve a stronger cellular immune response, for example cytokine proliferation and release by polyfunctional T cells, (iii) achieve a broader humoral immune response, for example, achieve an antibody titer linking the gp120 V1V2 region in a larger proportion of vaccinated individuals, (iv) achieving a broader cellular immune response, for example, proliferation and release of cytokines by polyfunctional T cells in a larger proportion of vaccinated individuals, (v) achieving a stronger humoral immune response against a particular subtype of HIV-1, (vi) achieve a stronger cellular immune response against a specific subtype (vii) require fewer components, (vii) require non-living components, (viii) involve a simpler dosing regimen, (ix) be simpler to produce, (x) be more easily stored, ( xi) be useful in treating or preventing HIV-1 infection, (xii) being useful in treating or preventing HIV-1 infection through a first HIV-1 subtype when the gp120-related polypeptide of the composition is derived from a second HIV-1 subtype, (xiii) achieve a more durable immune response, for example, based on response magnitude and / or respondent rates, (xiv) achieve greater viral load reduction, (xv) induce a higher level of protection against infection.

SUMMARY OF THE INVENTION

Compositions containing either (a) gpl20w6iD and AS01B or (b) gpl20w6iD> AS01B and NefTat were administered to mice and analyzed the serological response of mice.

Surprisingly, mice administered with gpl20w6iD and AS01B (without NefTat) were found to have a higher mean level of anti-VlV2 antibodies in their serum compared to mice administered with gpl20w6iD, AS01B and NefTat.

The claimed immunogenic compositions can be expected to elicit a high level of anti-VlV2 antibodies, and have utility in the prophylaxis of HIV infection.

The present invention provides an immunogenic composition which is in the form of a human dose comprising a gp120 related polypeptide and an adjuvant, wherein the adjuvant comprises between 10-40 µg of a lipopolysaccharide and between 10-40ug of a fraction of Immunologically active saponin derived from the Quillaja Saponaria Molina bark presented as a liposome.

Also provided is an immunogenic composition comprising a gp120 related polypeptide and an adjuvant, wherein the adjuvant comprises a lipopolysaccharide and an immunologically active saponin fraction derived from the Quillaja Saponaria Molina bark in the form of a liposome wherein: ( i) the conductivity of a composition is 13 mS / cm or less; and / or (ii) the salt concentration in said composition is 130 mM or less; and / or (iii) the sodium chloride concentration in said composition is 130 mM or less.

Further provided is an immunogenic composition comprising a gp120 related polypeptide and an adjuvant, wherein the adjuvant comprises a lipopolysaccharide and an immunologically active saponin fraction derived from the Quillaja Saponaria Molina bark in which the liposome is present. The composition is substantially free of a NefTat related polypeptide wherein the NefTat related polypeptide is a polypeptide consisting of SEQ ID NO: 4.

DESCRIPTION OF THE FIGURES

Figure 1: Comparison of mouse anti-VlV2 serology SEQUENCE DESCRIPTION SEQ ID NO: 1 gpl20W61D SEQ ID NO: 2 Nef SEQ ID NO: 3 Tat SEQ ID NO: 4 NefTat SEQ ID NO: 5 gpl20ZM, 8 SEQ ID NO: 6 polynucleotide sequence encoding gpl20ZMi8 SEQ ID NO: 7 native gpl20ZMi8 SEQ ID NO: 8 polynucleotide sequence encoding gpl20ZMi8 DETAILED DESCRIPTION OF THE INVENTION Polypeptides [00027] As used herein, the term 'a gpl20-related polypeptide' refers to to a polypeptide comprising the V1V2 region of SEQ ID NO: 1 or to an immunogenic derivative or fragment of the V1V2 region of SEQ ID NO: 1, or to a polypeptide comprising the V1V2 region of SEQ ID NO: 5 or to an immunogenic derivative or The V1V2 region fragment of SEQ ID NO: 5. Suitably, the gp120 related polypeptide refers to a polypeptide consisting of the V1V2 region of SEQ ID NO: 1 or an immunogenic derivative or fragment of the V1V2 region of SEQ ID NO: 1 , or to a poly peptide consisting of the VIV2 region of SEQ ID NO: 5 or an immunogenic derivative or fragment of the VIV2 region of SEQID NO: 5.

The VIV2 region of a gp120 related polypeptide is the residue segment defined by two particular cysteine residues that form a disulfide bridge in the duplicate state of the polypeptide. The VIV2 region itself excludes these cysteine residues. An example of the VIV2 region in the case of the gpl20W61D polypeptide (SEQ ID NO: 1) is the residue segment from residues 90 to 184. The person skilled in the art will note that the position of these cysteine residues and thus the position of the VIV2 region within a given gp120 related polypeptide may vary. This is illustrated by another gp120 related polypeptide, gp120ZMi8 (SEQ ID NO: 5) wherein the V1V2 region is the residue segment from residue 90 to 172.

Suitably, the term 'a gp120 related polypeptide' refers to a polypeptide comprising SEQ ID NO: 1 or an immunogenic derivative or fragment of SEQ ID NO: 1, or a polypeptide comprising SEQ ID NO: 5 or an immunogenic derivative or fragment of SEQ ID NO: 5. Suitably, the gp120 related polypeptide refers to a polypeptide consisting of SEQ ID NO: 1 or an immunogenic derivative or fragment of SEQ ID NO: 1, or a polypeptide consisting of SEQ ID NO: 5 or an immunogenic derivative or fragment of SEQ ID NO: 5.

As used herein, the term "derivative" refers to a polypeptide that is modified with respect to the reference sequence. Immunogenic derivatives are sufficiently similar to the reference sequence to remain capable of eliciting an immune response against the V1V2 region of the reference sequence. Suitably, immunogenic derivatives are sufficiently similar to the reference sequence to retain other key immunogenic properties of the reference sequence as preventing the introduction of immunodominant epitopes. A derivative may, for example, comprise a modified version of the reference sequence or alternatively may consist of a modified version of the reference sequence.

Suitably, the gp120-related polypeptide comprises or consists of an HIV-1 or HIV-2 derived gp120 polypeptide, suitably from HIV-1 Μ, N, O or P groups, suitably from subtype A Group B, C, D, E, F, G, Η, I, J or K Suitably, the gp120-related polypeptide comprises or consists of an HIV-1-derived gp120 polypeptide, suitably from the M-group. more suitably subtype B. For example, the gp120-related polypeptide comprises or consists of the gp120 polypeptide from an HIV-1 or HIV-2, suitably from an HIV-1 Μ, N, O or P group, suitably. of a subtype A, B, C, D, E, F, G, Η, I, J or K of HIV-1 group M. Suitably, the gp120 related polypeptide comprises or consists of the gp120 polypeptide from HIV-1, appropriately from HIV-1 group M, more appropriately from HIV-1 subtype M group.

The skilled artisan will recognize that individual substitutions, deletions or additions to the gp120 related polypeptide that alter, add or delete a single amino acid or a small percentage of amino acids is an “immunogenic derivative” where the alteration (s) (s) results in the substitution / deletion / addition of residues that do not substantially impact immunogenic function. By "do not substantially impact immunogenic function" is meant that at least 50%, appropriately at least 75%, and appropriately at least 90% reference sequence activity in any antibody level assay binding the gp120 V1V2 region (e.g. ELISA) produced at a given time point after immunization with the gp120 related polypeptide.

Conservative substitution tables giving functionally similar amino acids are well known in the art. In general, these conservative substitutions will be within one of the amino acid groupings specified below, although in some circumstances other substitutions may be possible without substantially affecting the immunogenic properties of the antigen. The following eight groups each contain amino acids that are typically conservative substitutions for one another: 1) Alanine (A), Glycine (G); s s 2) Aspartic acid (D), Glutamic acid (E); 3) Asparagine (N), Glutamine (Q); 4) Arginine (R), Lysine (K); 5) Isoleucine (I), Leucine (L), Methionine (M), Valine (V); 6) Phenylalanine (F), Tyrosine (I), Tryptophan (W); 7) Serine (S), Threonine (T); and 8) Cysteine (C), Methionine (M) (see for example Creighton, Proteins 1984).

Suitably such substitutions do not occur in the region of an epitope, and thus have no significant impact on the immunogenic properties of the antigen. Suitably an immunogenic derivative will contain substitutions of up to 20 residues (e.g. up to 5 residues) with respect to the reference sequence.

Immunogenic derivatives may also include those in which additional amino acids are inserted compared to the reference sequence. Appropriately such insertions do not occur in the region of an epitope, and do not have a significant impact on the immunogenic properties of the antigen. Suitably an immunogenic derivative will contain additions of up to 5 residues (e.g. 1 or 2 residues) at 0-5 locations (e.g. 0-2 locations) with respect to the reference sequence. An example of insertions includes a short segment of histidine residues (e.g. 6 residues) to aid purification of the antigen in question.

Immunogenic derivatives include those in which amino acids have been deleted compared to the reference sequence. Suitably such deletions do not occur in the region of an epitope, and thus do not have a significant impact on the immunogenic properties of the antigen. Suitably an immunogenic derivative will contain deletions of up to 5 residues (e.g. 1 or 2 residues) at 0-5 locations (e.g. 0-2 locations) with respect to the reference sequence.

Suitably the gp120 related polypeptide will comprise as consisting of an immunogenic derivative of the VIV2 region of SEQ ID NO: 1 having a small number of deletions, insertions and / or substitutions as a derivative of the V1V2 region of SEQ ID NO: 1 having deletions of up to 5 residues (eg 1 or 2 residues) at 0-5 locations (eg 0-2 locations), insertions of up to 5 residues (eg 1 or 2 residues) at 0-5 five locations (e.g. 0-12 locations) and substitutions of up to 20 residues (e.g. up to 5 residues).

Alternatively, the gp120 related polypeptide will comprise, as consisting of, an immunogenic derivative of the V1V2 region of SEQ ID NO: 5 having a small number of deletions, insertions and / or substitutions, such as a derivative of the V1V2 region of SEQ. ID NO: 5 having deletions of up to 5 residues (eg 1 or 2 residues) at 0-5 locations (eg 0-2 locations), insertions of up to 5 residues (eg 1 or 2 residues) at 0-5 five locations (eg 0-2 locations) and substitutions of up to 20 residues (eg up to 5 residues).

Suitably the gp120 related polypeptide will comprise, as consisting of, an immunogenic derivative of SEQ ID NO: 1 having a small number of deletions, insertions and / or substitutions, such as a derivative of SEQ ID NO: 1 having deletions of up to 5 residues at 0-5 locations, inserts of up to 5 residues at 0-5 five locations and substitutions of up to 20 residues.

Alternatively, the gp120 related polypeptide will comprise, as consisting of, an immunogenic derivative of SEQ ID NO: 5 having a small number of deletions, insertions and / or substitutions, such as a derivative of SEQ ID NO: 5 having deletions. up to 5 residues at 0-5 locations, inserts of up to 5 residues at 0-5 five locations and replacements of up to 20 residues.

Suitably, the gp120 related polypeptide comprises a polypeptide with at least 70% identity, more appropriately at least 80% identity, more appropriately at least 85% identity, more appropriately at least 90% identity, more appropriately at least 95% identity, more appropriately at least 98% identity, more appropriately at least 99% identity with the V1V2 region of SEQ ID NO: 1. Suitably the gp120 related polypeptide comprises the V1V2 region of SEQ ID NO: 1.

Suitably, the gp120 related polypeptide consists of a polypeptide with at least 70% identity, suitably at least 80% identity, more appropriately at least 85% identity, more appropriately at least 90% identity, more appropriately at least 95% identity, more appropriately at least 98% identity, more appropriately at least 99% identity with the VIV2 region of SEQ ID NO: 1. Suitably, the gp120 related polypeptide consists of the VIV2 region of SEQ ID NO: 1.

Alternatively, the gp120 related polypeptide comprises a polypeptide with at least 70% identity, more appropriately at least 80% identity, more appropriately at least 85% identity, more appropriately at least 90% identity, more appropriately at least 95% identity, more appropriately at least 98% identity, more appropriately at least 99% identity with the V1V2 region of SEQ ID NO: 5. Suitably the gp120 related polypeptide comprises the V1V2 region of SEQ ID NO: 5.

Suitably, the gp120 related polypeptide consists of a polypeptide with at least 70% identity, suitably at least 80% identity, more appropriately at least 85% identity, more appropriately at least 90% identity, more appropriately at least 95% identity, more appropriately at least 98% identity, more appropriately at least 99% identity with the V1V2 region of SEQ ID NO: 5. Suitably the gp120 related polypeptide consists of the V1V2 region of SEQ ID NO: 5.

Suitably, the gp120 related polypeptide comprises a polypeptide with at least 70% identity, more appropriately at least 80% identity, more appropriately at least 85% identity, more appropriately at least 90% identity, more appropriately at least 95% identity, more appropriately at least 98% identity, more appropriately at least 99% identity with SEQ ID NO: 1. Suitably, the gp120 related polypeptide comprises SEQ ID NO: 1.

Suitably, the gp120 related polypeptide consists of a polypeptide with at least 70% identity, suitably at least 80% identity, more appropriately at least 85% identity, more appropriately at least 90% identity, more appropriately at least 95% identity, more appropriately at least 98% identity, more appropriately at least 99% identity with SEQ ID NO: 1. Suitably, the gp120 related polypeptide consists of SEQ ID NO: 1.

Alternatively, the gp120 related polypeptide comprises a polypeptide with at least 70% identity, more appropriately at least 80% identity, more appropriately at least 85% identity, more appropriately at least 90% identity, more appropriately at least 95% identity, more appropriately at least 98% identity, more appropriately at least 99% identity with SEQ ID NO: 5. Suitably, the gp120 related polypeptide comprises SEQ ID NO: 5.

Suitably, the gp120 related polypeptide consists of a polypeptide with at least 70% identity, suitably at least 80% identity, more appropriately at least 85% identity, more appropriately at least 90% identity, more appropriately at least 95% identity, more appropriately at least 98% identity, more appropriately at least 99% identity with SEQ ID NO: 5. Suitably, the gp120 related polypeptide consists of SEQ ID NO: 5.

The terms "identical" or "% identity," in the context of two or more polypeptide sequences, refer to two or more sequences or subsequences that are the same or have a specified percentage of amino acid residues that are the same. over a specified region when compared and aligned to a maximum match over a comparison window, or region designated as measured using one of the sequence comparison algorithms or by manual alignment and visual inspection This definition also refers to test sequence compliance . Optionally, the identity exists over a region that is at least 300 amino acids in length, such as at least 400 amino acids or at least 500 amino acids. Most appropriately, the comparison is performed over a window corresponding to the full length of the reference sequence (as opposed to the derived sequence).

For sequence comparison, a sequence acts as the reference sequence to which test sequences are compared. When using a sequence comparison algorithm, test and reference sequences are entered as input into a computer, subsequence coordinates are assigned, if necessary, and sequence algorithm program parameters are assigned. Default program parameters can be used, or alternate parameters can be assigned. The sequence comparison algorithm then calculates the percent sequence identities for the test sequences with respect to the reference sequence, based on program parameters.

A "comparison window" as used herein refers to a segment in which the sequence can be compared to a reference sequence of the same number of contiguous positions after the two sequences are optimally aligned. Sequence alignment methods for comparison are well known in the art. Optimal alignment of sequences for comparison can be conducted, for example, by the local homology algorithm of Smith & Waterman, Adv. Appl. Math 2: 482 (1981), by the homology alignment algorithm of Needleman & Wunsch, J. Mol. Biol. 48: 443 (1970), by searching Pearson & Lipman's similarity method, Proc. Nat'l. Acad. Know. USA 85: 2444 (1988), by computerized implementations of these algorithms (GAP, BESTFIT, FASTA, and TFASTA according to the Wisconsin Genetics Software Package, Genetics Computer Group, 575 Dr. Science, Madison, WI), or by manual alignment. visual inspection (see, for example, Curr Pro Mol Biol (Ausubel et al., eds. 1995 supplement)).

An example of an algorithm that is suitable for determining percent sequence identity and sequence similarity includes the BLAST and BLAST 2.0 algorithms, which are described in Altschul et al., Nuc. Acids Res. 25: 3389-3402 (1977) and Altschul et al., J. Mol. Biol. 215: 403-410 (1990), respectively. Software for performing BLAST analyzes is publicly accessible through the National Center for Biotechnology Information (website at www.ncbi.nlm.nih.gov/). This algorithm involves first identifying high score sequence pairs (HSPs) by identifying short-length words W in the query sequence that either match or meet the positive value threshold T score when aligned with a word of the same length in a database string. T is referred to as the neighborhood word score threshold (Altschul et al., Supra). These initial neighborhood word tips act as seeds for starting searches to find longer HSPs containing them. These word tips are extended in both directions along each sequence as far as the cumulative alignment score can be increased. Cumulative scores are calculated using, for nucleotide sequences, the parameters M (reward score for a matched residue pair; always> 0) and N (penalty score for mismatched residues; always <0). For amino acid sequences, a score matrix is used to calculate the cumulative score. Word cue extensions in each direction are stopped when: the cumulative alignment score falls by the amount X from its maximum obtained value; the cumulative score drops to zero or below due to the accumulation of one or more negative score residue alignments; or the end of both sequences is reached. The W, T, and X parameters of the BLAST algorithm determine the sensitivity and speed of alignment. The BLASTN program (for nucleotide sequences) defaults to a word length (W) of 11, an expectation (E) or 10, M = 5, N = -4, and a comparison of both filaments. For amino acid sequences, the BLASTP program uses, as defaults, a word length of 3, expectation (E) of 10, and the BLOSUM62 score matrix (see Henikoff & Henikoff, Proc. Natl. Acad. Sci. USA 89: 10915 (1989)) alignments (B) of 50, expectation (E) of 10, M = 5, N = -4, and a comparison of both filaments.

The BLAST algorithm also performs a statistical analysis of similarity between two sequences (see, for example, Karlin & Altschul, Proc. Nat'l. Acad. Sci. USA 90: 5873-5787 (1993)). A measure of similarity given by the BLAST algorithm is the least sum probability (P (N)), which provides an indication of the probability by which a match between two nucleotide or amino acid sequences would occur by chance. For example, a nucleic acid is considered similar to a reference sequence if the probability of summation lower in a test nucleic acid comparison to the reference nucleic acid is less than about 0.2, more suitably less than about of 0.01, and the most suitably less than about 0.001.

An "immunogenic fragment" will contain a contiguous amino acid sequence from the gp120 related polypeptide of which it is a fragment. Suitably, the fragment contains at least 15 to 50 amino acids, at least 51 to 150, at least 300 amino acids, at least 350, at least 400, or at least 450 contiguous amino acids of SEQ.

Alternatively, the fragment contains at least 15 to 50 amino acids, at least 51 to 150 amino acids, at least 300, at least 350, at least 400, or at least 450 contiguous amino acids of SEQ ID NO: 5.

The immunogenic fragment will remain capable of eliciting an immune response against the V1V2 region of the reference sequence. By "remaining capable of eliciting an immune response against the reference sequence V1V2 region" means that at least 50%, appropriately at least 75%, and appropriately at least 90% reference sequence activity in an antibody level test binding to gp120 V1V2 region (e.g. ELISA) produced at a given time point after immunization with the gp120 related polypeptide.

The gp120 related polypeptide may, for example, contain 2000 amino acid residues or less, such as 1500 amino acid residues or less, in particular 1000 amino acid residues or less, especially 800 amino acid residues or less.

Fusion proteins (also known as chimeric proteins) are proteins created by covalently bonding two or more polypeptide sequences that are not joined in nature, such as by peptide bonding. For example, the gp120 related polypeptide may be provided in the form of the fusion protein also comprising a second HIV antigen.

The dose of a gp120 related polypeptide is suitably capable of producing an adequate immune response in a human while having an acceptable reacogenicity profile.

Suitably the immunogenic composition comprises from about 1 to 200 µg of gp120 related polypeptide, suitably from 1 to 100 µg, suitably from 2 to 50 µg, such as from 3 to 30 µg, in particular from 5 to 15 µg or 16 to 25 µg, 9 to 11 µg or 319 to 21 µg, most suitably 10 µg or 20 µg.

Suitably, the immunogenic composition is provided in a volume that is suitable for administration to a human as a single dose. The volume that can be administered to a human is dependent on the method, route and / or location of administration. In one embodiment the human dose is between 0.1 and 1 ml, more suitably between 0.3 and 0.75 ml, such as between 0.45 and 0.55 ml, in particular 0.5 ml. Volumes of from 0.1 to 1 ml are particularly suitable for administration via routes such as subcutaneous and in particular intramuscular release. In another embodiment the human dose is between 0.05 and 0.2 ml, suitably between 0.075 and 0.15 ml, in particular 0.1 ml. Volumes of between 0.05 and 0.2 ml are particularly suitable for administration via routes in which a limited volume may be administered as intradermal release.

A gp120 polypeptide may be prepared by methods described in the art or methods analogous thereto.

As used herein, the term "a Nef-related polypeptide" refers to the polypeptide provided in SEQ ID NO: 2, or an immunogenic derivative or fragment thereof.

As used herein, the term "Tat related polypeptide" refers to a polypeptide provided in SEQ ID NO: 3, or an immunogenic derivative or fragment thereof.

As used herein, the term 'a Neitat-related polypeptide' refers to the polypeptide provided in SEQ ID NO: 4, or an immunogenic derivative or fragment thereof.

Suitably the Nef-related polypeptide will comprise, as consisting of, an immunogenic derivative of SEQ ID NO: 2, optionally having a small number of deletions, insertions and / or substitutions. An example is a derivative of SEQ ID NO: 2 having deletions of up to 5 residues at 0-5 locations, insertions of up to 5 residues at 0-5 locations and substitutions of up to 20 residues.

Suitably the Tat related polypeptide will comprise, as consisting of, an immunogenic derivative of SEQ ID NO: 3, optionally having a small number of deletions, insertions and / or substitutions. An example is a derivative of SEQ ID NO: 3 having deletions of up to 5 residues at 0-5 locations, insertions of up to 5 residues at 0-5 locations and substitutions of up to 20 residues.

Suitably the NefTat related polypeptide will comprise, as consisting of, an immunogenic derivative of SEQID NO: 4, optionally having a small number of deletions, insertions and / or substitutions. An example is a derivative of SEQ ID NO: 4 having deletions of up to 5 residues at 0-5 locations, insertions of up to 5 residues at 0-5 locations and substitutions of up to 20 residues.

Suitably, the Nef-related polypeptide comprises SEQ ID NO: 2. Suitably the Nef-related polypeptide comprises a polypeptide with at least 99% identity, appropriately at least 98% identity, more appropriately at least 95% identity, more appropriately. at least 90% identity, more appropriately at least 85% identity, more appropriately at least 80% identity, more appropriately at least 75% identity, more appropriately at least 70% identity with SEQ ID NO: 2.

Suitably, the Tat-related polypeptide comprises SEQ ID NO: 3. Suitably the Tat-related polypeptide comprises a polypeptide with at least 99% identity, appropriately at least 98% identity, more appropriately at least 95% identity, more appropriately at least 90% identity, more appropriately at least 85% identity, more appropriately at least 80% identity, more appropriately at least 75% identity, more appropriately at least 70% identity with SEQ ID NO: 3.

Suitably the NefTat-related polypeptide comprises SEQ ID NO: 4. Suitably the NefTat-related polypeptide comprises a polypeptide with at least 99% identity, appropriately at least 98% identity, more appropriately at least 95% identity, more appropriately at least 90% identity, more appropriately at least 85% identity, more appropriately at least 80% identity, more appropriately at least 75% identity, more appropriately at least 70% identity with SEQ ID NO: 4.

Suitably, the Nef-related polypeptide consists of SEQ ID NO: 2. Suitably the Nef-related polypeptide consists of a polypeptide with at least 99% identity, appropriately at least 98% identity, more appropriately at least 95% identity, more appropriately at least 90% identity, more appropriately at least 85% identity, more appropriately at least 80% identity, more appropriately at least 75% identity, more appropriately at least 70% identity with SEQ ID NO: 2.

Suitably, the Tat-related polypeptide consists of SEQ ID NO: 3. Suitably the Tat-related polypeptide consists of a polypeptide with at least 99% identity, suitably at least 98% identity, more appropriately at least 95% identity, more appropriately at least 90% identity, more appropriately at least 85% identity, more appropriately at least 80% identity, more appropriately at least 75% identity, more appropriately at least 70% identity with SEQ ID NO: 3.

Suitably, the NefTat-related polypeptide consists of SEQ ID NO: 4. Suitably the NefTat-related polypeptide consists of a polypeptide with at least 99% identity, appropriately at least 98% identity, more appropriately at least 95% identity, more appropriately at least 90% identity, more appropriately at least 85% identity, more appropriately at least 80% identity, more appropriately at least 75% identity, more appropriately at least 70% identity with SEQ ID NO: 4.

In one embodiment a Nef-related polypeptide is a NefTat-related polypeptide. In another embodiment a Tat related polypeptide is a NefTat related polypeptide.

Particular derivatives of a Nef-related polypeptide, Tat-related polypeptide or NefTat-related polypeptide include those with additional N-terminus His residues (for example a six-His residues polyhistidine tag, which may be used for nickel affinity purification).

Suitably the compositions of the present invention are substantially free of a Nef-related polypeptide. Suitably, the compositions of the present invention contain a ratio of a Nef-related polypeptide related polypeptide: gp 120 of less than 1:20, suitably less than 1:25, suitably less than 1:50, suitably less than 1: 100, suitably less than 1: 200, suitably less than 1: 500, more suitably less than 1: 1000 by weight.

Suitably the compositions of the present invention contain a Nef-related polypeptide at a level of less than 5ug, suitably less than 4ug, suitably less than 3ug, suitably less than 2ug, suitably less than lug, suitably less. than 0.5ug, suitably less than 0.2ug, suitably less than 0.1ug, appropriately less than 0.05ug, more suitably less than 0.0lug per human dose.

Most suitably the compositions of the present invention are free of a Nef related polypeptide.

Suitably the compositions of the present invention are substantially free of a Tat related polypeptide.

Suitably, the compositions of the present invention contain a ratio of the Tat: gp120 related polypeptide related polypeptide of less than 1:20, suitably less than 1:25, suitably less than 1:50, suitably less than 1 : 100, suitably less than 1: 200, suitably less than 1: 500, more suitably less than 1: 1000 by weight.

Suitably the compositions of the present invention contain a Tat related polypeptide at a level of less than 5ug, suitably less than 4ug, suitably less than 3ug, suitably less than 2ug, suitably less than lug, suitably less. than 0.5ug, suitably less than 0.2ug, suitably less than 0.1ug, appropriately less than 0.05ug, more suitably less than 0.0lug per human dose.

Most suitably the compositions of the present invention are free of a Tat related polypeptide.

Suitably the compositions of the present invention are substantially free of a NefTat related polypeptide. Suitably, the compositions of the present invention contain a ratio of NefTat-related polypeptide related polypeptide: gp120 of less than 1:20, suitably less than 1:25, suitably less than 1:50, suitably less than 1 : 100, suitably less than 1: 200, suitably less than 1: 500, more suitably less than 1: 1000 by weight.

Suitably the compositions of the present invention contain the NefTat related polypeptide at a level of less than 5ug, suitably less than 4ug, suitably less than 3ug, suitably less than 2ug, suitably less than lug, suitably less than 0.5ug, suitably less than 0.2ug, suitably less than 0.1ug, appropriately less than 0.05ug, more suitably less than 0.0lug per human dose.

Most suitably the compositions of the present invention are free of a NefTat related polypeptide.

The presence of other HIV proteins may increase the effectiveness of the composition of the invention. Suitably the composition of the invention may comprise additional proteins which may further be Env gene products (such as gp160 and gp41 or gp120 related polypeptide) gag and pol gene products (such as MA, CA, SP1, NC, SP2 and P6; and RT, RNase H, IN and PR, respectively) and other structural proteins such as Rev, Vif, Vpr and Vpu (or immunogenic derivatives or fragments thereof).

The HIV gag gene encodes a p55 precursor protein that can spontaneously assemble into immature virus-like particles (VLPs). The precursor is proteolytically cleaved into the major structural proteins CA (capsid) and MA (matrix), and various minor proteins. Both p55 precursor protein and its major derivatives CA and MA may be considered as appropriate vaccine antigens which may increase the efficacy of the composition of the invention. The p55 precursor and CA capsid protein may be used as VLPs or as monomeric proteins. Suitably the composition of the present invention may comprise one or more of these proteins.

Suitably, the composition of the present invention may comprise a polynucleotide encoding a polypeptide described above including, for example, a gp120 related polypeptide. The polynucleotide may be in the form of plasmid DNA or in the form of the recombinant living vector.

In one embodiment, the composition of the present invention does not comprise additional HIV antigens. In a second embodiment of the invention the composition comprises 1-5 additional HIV antigens, such as 1 or 2 additional HIV antigens. Additional HIV antigens may be provided in the form of proteins or polynucleotides encoding proteins.

Suitably, the immunogenic composition comprises a total of from about 1 to 500 µg of antigenic material, suitably from 1 to 200 µg, such as from 5 to 100 µg, most suitably from 5 to 50 µg.

Suitably, the polypeptides and polynucleotides used in the present invention are isolated. An "isolated" polypeptide or polynucleotide is one that is removed from its natural environment. For example, a naturally occurring protein is isolated if it is separated from some or all of the coexisting materials in the natural system. Preferably, such polypeptides are at least about 90% pure, more suitably at least about 95% pure and most suitably at least about 99% pure.

A polynucleotide is considered to be isolated if, for example, it is cloned into a vector that is not part of its natural environment.

Adjuvant Saponins The immunogenic composition of the invention comprises an immunologically active saponin moiety ("a saponin") as an adjuvant or as a component of an adjuvant. A particularly suitable saponin for use in the present invention is Quil A and derivatives thereof. Quil A is a saponin preparation isolated from the South American tree Quillaja Saponaria Molina and was first described by Dalsgaard et al. In 1974 ("Saponin adjuvants", Archiv. Für die gesamte Virusesforschung, Vol. 44, Springer Verlag, Berlin, p243-254) as having an adjuvant activity. Purified Quil A fragments were isolated by HPLC which retains adjuvant activity without the toxicity associated with Quil A (US5604106), for example QS-7 and QS-21 (also known as QA7 and QA21). QS-21 is a natural saponin derivative of the Quillaja saponaria Molina bark, which induces CD8 + cytotoxic T cells (CTLs), Th1 cells and a predominant IgG2a antibody response and is the preferred saponin in the present invention.

In an appropriate form of the present invention, the saponin adjuvant within an immunogenic composition is a derivative of saponaria molina kil A, suitably an immunologically active fraction of Quil A, such as QS-7 or QS-21, suitably QS- 21 In one embodiment the compositions of the invention contain the immunologically active saponin fraction in a substantially pure form. Suitably the compositions of the invention contain QS-21 in substantially pure form, that is, QS-21 is at least 90% pure, for example at least 95% pure, or at least 98% pure.

In a specific embodiment, QS-21 is provided in a less reacogenic composition, where it is quenched with an exogenous sterol such as cholesterol. Several particular forms of less reacogenic compositions in which the lytic activity of QS-21 is extinguished with an exogenous cholesterol exist. In a specific embodiment, the saponin / sterol is in the form of a liposome structure (US6846489, Example 1). In this embodiment the liposomes suitably contain a neutral lipid, for example phosphatidylcholine, which is suitably non-crystalline at room temperature, for example egg yolk phosphatidylcholine, dioleoyl phosphatidylcholine (DOPC) or dilauryl phosphatidylcholine. Liposomes may also contain a charged lipid that increases the stability of the QS-21 liposome structure for liposomes composed of saturated lipids. In such cases, the amount of lipid loaded is suitably 1-20 wt%, suitably 5-10 wt%. The sterol to phospholipid ratio is 1-50% (mol / mol), suitably 20-25%.

Suitable sterols include beta-sitosterol, stigmasterol, ergosterol, ergocalciferol and cholesterol. In a particular embodiment, the adjuvant composition comprises cholesterol as sterol.

These sterols are well known in the art, for example, cholesterol is described in the Merck Index, 11a. Ed. Pag. 341, as a naturally occurring sterol found in animal fat.

The sterol according to the invention is taken to mean an exogenous sterol, that is, a sterol that is not endogenous to the organism from which an antigen preparation is withdrawn but is added to the antigen preparation or thereafter at the time. of the formulation. Typically, sterol may be added during subsequent formulation of the antigen preparation with the saponin adjuvant, using, for example, saponin in the form in which its lytic activity is quenched with sterol. Suitably exogenous sterol is associated with saponin adjuvant as described in US6846489.

When the active saponin fraction is QS-21, the ratio of QS-21: sterol will typically be in the order of 1: 100 to 1: 1 (weight / weight), suitably 1:10 to 1: 1 ( weight / weight), and suitably 1: 5 to 1: 1 (weight / weight). Suitably an excess sterol is present, the ratio of QS-21: sterol being at least 1: 2 (w / w). In one embodiment, the ratio of QS-21: sterol is 1: 5 (weight / weight).

Other saponins that have been described in the literature include Escina, which was described in the Merck index (12th ed: entry 3737) as a mixture of saponins occurring in the horse chestnut seed, Lat: Aesculus hippocastanum. Their isolation is described by chromatography and purification (Fiedler, Arzneimittel-Forsch. 4, 213 (1953)), and by ion exchange resins (Erbring et al., US 3,238,190). The escine fractions were purified and shown to be biologically active (Yoshikawa M, et al. (Chem Pharm Bull (Tokyo) 1996 44 (8): 1454-1464)). Sapoalbin from Gypsophilla struthium (R. Vochten et al., 1968, J Pharm Belg, 42, 213-226) have also been described with respect to the production of ISCOM for example. Another usable saponin is that derived from the plant Gyophilla struthium.

Suitably, the total amount of saponin in an immunogenic composition of the present invention, particularly in a human dose of the immunogenic composition of the present invention is between 1-100ug.

In one embodiment, an immunogenic composition comprising QS-21 is provided at a level around 50 µg, for example, between 38-100 µg, suitably between 40-75 µg or between 45-60 µg, most suitably 49-51, most appropriately 50 µg.

In another embodiment, an immunogenic composition comprising QS-21 is provided at a level of about 25 µg, for example, between 10-37 µg, suitably between 15-30 µg or 20-27 µg. , more appropriately 24-26, more appropriately 25 µg.

In another embodiment, there is provided an immunogenic composition in a volume that is appropriate for a human dose whose human dose of the immunogenic composition comprises QS-21 at a level of about 50 µg, for example, between 38-40 ° C. 100 µg, suitably between 40-75 µg or between 45-60 µg, more appropriately 49-51, most appropriately 50 µg.

In another embodiment, there is provided an immunogenic composition in a volume that is appropriate for a human dose whose human dose of the immunogenic composition comprises QS-21 at a level around 25 µg, for example, between 10-25 ° C. 37 µg, suitably between 15-30 µg or between 20-27 µg, more suitably 24-26, more appropriately 25 µg.

The dose of QS-21 is suitably capable of enhancing an immune response to an antigen in a human. In particular an appropriate amount of QS-21 is that which enhances the immunological potential of the composition compared to the non-adjuvanted composition, or compared to an adjuvanted composition with another amount of QS-21, while being acceptable from a reacogenicity profile.

Lipopolysaccharide Adjuvants [000107] Lipopolysaccharide (LPS) is the major surface molecule of and occurs exclusively in the outer shale of the outer membrane of gram-negative bacteria. LPS prevents the destruction of bacteria by serum supplements and phagocytic cells and are involved in adhesion for colonization. LPS are a group of structurally related complex molecules of approximately 10,000 Daltons in size and consist of three covalently linked regions: (i) an O-specific polysaccharide chain (O-antigen) in the outer region (ii) a central oligosaccharide region of (iii) lipid nucleus A. The innermost region that serves as a hydrophobic anchor, it comprises glucosamine disaccharide units that carry long chain fatty acids.

The biological activities of LPS, such as lethal toxicity, pyrogenicity, and adjuvanticity, have been shown to be related to the lipid A moiety. In contrast, immunogenicity is associated with the O-specific polysaccharide component (O-antigen). Both LPS and lipid A have long been known as adjuvant effects, but the high toxicity of these molecules has excluded their use in vaccine formulations. Significant effort was thus made to reduce the toxicity of LPS or lipid A while maintaining its adjuvanticity.

The Salmonella Minnesota R595 mutant was isolated in 1966 from a parental (mild) strain culture (Luderitz et al. 1966 Ann N Y Acad Sci 133: 349-374). Selected colonies were screened for their susceptibility to lysis by a phage panel, and only those colonies that exhibited a narrow range of sensitivity (susceptible to only one or two phages) were selected for another study. This effort has led to the isolation of the deep rough mutant strain that is defective in LPS biosynthesis and referred to as S. minnesota R595.

Compared to other LPS, those produced by the S. minnesota R595 mutant have a relatively simple structure. (i) they do not contain O-specific region - a feature that is responsible for shifting from wild-type mild phenotype to rough mutant phenotype and results in a loss of virulence (ii) a very short core region - this feature increases Strain susceptibility to a variety of chemicals (iii) Lipid A moiety is highly acylated with up to 7 fatty acids.

[000111] 4'-Monophosphoryl lipid A (MPL), which can be obtained by hydrolysis of LPS acid extracted from a deep rough mutant strain of gram-negative bacteria, retains the properties of the LPS adjuvant while demonstrating a reduced toxicity. by a factor of more than 1000 (as measured by lethal dose in chick embryo eggs) (Johnson et al. 1987 Rev Infect Dis 9 Suppl: S512-S516). LPS is typically refluxed in moderate strength mineral acid solution (e.g. 0.1 M HCl) over a period of approximately 30 minutes. This process results in dephosphorylation at position 1, and decarburization at position 6 ', giving MPL.

[000112] 3-O-Deacylated Monophosphoryl lipid A (3D-MPL), which can be obtained by mild alkaline hydrolysis of MPL, has even reduced toxicity while again maintaining adjuvanticity, see US4912094 (Ribi Immunochemicals). Alkaline hydrolysis is typically performed in organic solvent as a chloroform / methanol mixture by saturation with a weak base aqueous solution such as 0.5 M sodium carbonate at pH 10.5. Other information about the preparation of 3D-MPL is available from, for example, US4912094 (Corixa Corporation).

The composition further comprises an additional adjuvant which is a lipopolysaccharide, suitably a non-toxic lipid A derivative, particularly monophosphoryl lipid A or more particularly 3-deacylated monophosphorylipid A (3D-MPL).

[000114] 3D-MPL is sold under the name MPL by GlaxoSmithKline Biologicals N.A. and is referred to throughout the document as MPL or 3D-MPL. see, for example, US4436727; US4877611; US4866034 and US4912094. 3D-MPL may be produced according to methods described in US4912094. Chemically it is a mixture of 3-deacylated monophosphoryl lipid A with 3, 4, 5 or 6 acylated chains. Suitably in the compositions of the present invention small particle 3D-MPL is used. Small particle 3D-MPL has a particle size so that it can be sterile filtered through a 0.22um filter. These preparations are described in US5776468.

Suitably, the total amount of lipopolysaccharide in an immunogenic composition of the present invention, particularly in a human dose of the immunogenic composition of the present invention, is between 1-100ug.

In one embodiment, an immunogenic composition comprising 3D-MPL is provided at a level of about 50 µg, for example, between 38-100 µg, suitably between 40-75 µg or 45-60 µg, most suitably 49-51, most appropriately 50 µg.

In another embodiment, an immunogenic composition comprising 3D-MPL is provided at a level around 25 µg, for example, between 10-37 µg, suitably between 15-30 µg or 20-27 µg, more suitably 24-26, more appropriately 25 µg.

In another embodiment, there is provided an immunogenic composition in a volume that is appropriate for a human dose whose human dose of the immunogenic composition comprises 3D-MPL at a level around 50 µg, for example, between 38-40 ° C. 100 µg, suitably between 4075 µg or between 45-60 µg, more appropriately 49-51, most appropriately 50 µg.

In another embodiment, there is provided an immunogenic composition in a volume that is appropriate for a human dose whose human dose of the immunogenic composition comprises 3D-MPL at a level around 25 µg, for example, between 10-30 ° C. 37 µg, suitably between 15-30 µg or between 20-27 µg, more suitably 24-26, more appropriately 25 µg.

Suitable compositions of the invention are those wherein liposomes are initially prepared without MPL (as described in US6846489), and MPL is then added, suitably as small particles below 100 nm or particles that are susceptible to sterile filtration through a 0.22 µm membrane. MPL thus is not contained within a gallbladder membrane (known as MPL outside). Compositions where MPL is contained within the gallbladder membrane (known as MPL within) also form an aspect of the invention. The antigen may be contained within a gallbladder membrane or contained outside a gallbladder membrane. Suitably soluble antigens are outside and hydrophobic or lipid antigens are either contained within or outside the membrane.

[000121] The invention comprises both lipopolysaccharide and immunologically active saponin. In a specific embodiment of the invention the lipopolysaccharide is 3D-MPL and the immunologically active saponin is QS-21. In one embodiment of the invention, the composition comprises a lipopolysaccharide and immunologically active saponin in a liposome formulation. Suitably in one form of these embodiments, the composition comprises 3D-MPL and QS-21, with optionally a sterol which is suitably cholesterol.

In another embodiment of the invention, the adjuvant composition comprises, in a liposome formulation, lipopolysaccharide and immunologically active saponin in combination with one or more other immunostimulants or adjuvants. Suitably in one embodiment of this embodiment the lipopolysaccharide is 3D-MPL and the immunologically active saponin is QS-21.

In a specific embodiment, QS-21 and 3D-MPL are present in a weight ratio between 1: 2 to 2: 1. Suitably QS-21 and 3D-MPL are present in the same final amount per human dose of the immunogenic composition. In one aspect of this embodiment, a human dose of immunogenic composition comprises a final level of 50 µg 3D-MPL and 50 µg QS-21. In another aspect, a human dose of immunogenic composition comprises a final level of 25 µg 3D-MPL and 25 µg QS-21. In another embodiment, a human dose of immunogenic composition comprises a final 10ug level of MPL and QS-21.

Vaccine preparation is generally described in New Trends and Developments in Vaccines, edited by Voller et al., University Park Press, Baltimore, Maryland, U.S.A. 1978. Encapsulation within liposomes is described, for example, in US4235877. Conjugation of proteins to macromolecules is described, for example, in US4372945 and US4474757. Salt Concentration [000125] Some antigens are sensitive to the presence of salts. Without being bound by theory, it is believed that these antigens are detrimentally impacted by a phenomenon known as “salinization” which can be defined as protein precipitation from its solution by interaction with salts such as sodium chloride. . These antigens aggregate and precipitate to a sodium chloride concentration as low as 150 mM. Accordingly, the stability of immunogenic compositions comprising a gp120-related polypeptide may be enhanced by a reduction in sodium chloride concentration.

The immunogenic compositions of the invention will suitably be aqueous preparations.

Thus, the present invention provides an immunogenic composition comprising a gp120 related polypeptide, wherein the conductivity of the composition is 13 mS / cm or less. In particular, the present invention provides immunogenic compositions wherein the conductivity of the immunogenic composition is 12 mS / cm or less, for example 10 mS / cm or less, 8 mS / cm or less, 6 mS / cm or less, 5 mS / cm or less, 4 mS / cm or less, or 3 mS / cm or less.

In a particular embodiment the conductivity of the immunogenic composition is 2.5 mS / cm or less, as 2.25 mS / cm or less, or 2.0 mS / cm or less. In another specific embodiment the conductivity of the immunogenic composition is 1.5 to 2.5 mS / cm. Further provided is an immunogenic composition comprising a gp120 related polypeptide, wherein the concentration of salts in said composition is 130 mM or less. In particular, the present invention provides immunogenic compositions wherein the salt concentration in said composition is 100 mM or less, for example 90 mM or less, 80 mM or less, 70 mM or less, 60 mM or less, 50 mM or less. smaller, or 40 mM or smaller. In a particular embodiment the concentration of salts in said composition is 35 mM or less, such as 30 mM or less, or 25 mM or less. In another specific embodiment the concentration of salts in said composition is 20 to 40 mM, such as 25 to 35 mM.

The present invention also provides an immunogenic composition comprising a gp120 related polypeptide, wherein the concentration of sodium chloride in said composition is 130 mM or less. In particular, the present invention provides immunogenic compositions wherein the sodium chloride concentration is 100 mM or less, for example 90 mM or less, 80 mM or less, 70 mM or less, 60 mM or less, 50 mM or less. , 40 mM or less, 30 mM or less, 20 mM or less or 15 mM or less. In a particular embodiment the sodium chloride concentration in an immunogenic composition is 10 mM or less, such as 7.5 mM or less. Suitably the concentration of sodium chloride in an immunogenic composition is at or below 5 mM. In another specific embodiment, the immunogenic composition is essentially free of sodium chloride. By essentially free means that the sodium chloride concentration is at or near zero mM (such as 3 mM or less, 2 mM or less or 1 mM or less).

Suitably, the concentration of CaC12 in immunogenic compositions will be 40 mM or less, 30 mM or less, 20 mM or less, 15 mM or less or 10 mM or less.

Suitably, the concentration of MgSO4 in an immunogenic composition will be 80 mM or less, 60 mM or less, 40 mM or less, 30 mM or less, 20 mM or less or 10 mM or less.

Suitably, the total concentration of NH +, Mg2 + and Ca2 + ions in immunogenic compositions will be 80 mM or less, 60 mM or less, 40 mM or less, 30 mM or less, 20 mM or less or 10 mM or less. It is well known that for parenteral administration the solutions should have a pharmaceutically acceptable osmolality to avoid cell distortion or lysis. A pharmaceutically acceptable osmolality will generally mean that the solutions will have an osmolality that is approximately isotonic or mildly hypertonic. Suitably the immunogenic compositions of the present invention will have an osmolality in the range of 250 to 750 mOsm / kg, for example, the osmolality may be in the range of 250 to 550 mOsm / kg, as in the range of 280 to 500 mOsm / kg.

Osmolality can be measured according to techniques known in the art, such as using a commercially available osmometer, for example, the 2020 Advanced® model available from Advanced Instruments Inc. (USA).

An "isotonicity agent" is a compound that is physiologically tolerated and gives an appropriate tonicity to a formulation to prevent liquid water flow through the cell membranes that are in contact with the formulation.

In a particular embodiment there are provided immunogenic compositions further comprising a nonionic tonicity agent. A nonionic tonicity agent for use in an immunogenic composition will need to be itself pharmaceutically acceptable, e.g. suitable for use in humans, as well as compatible with the gp120-related antigen and further compatible with other components such as immunostimulants (s). ).

In one embodiment of the present invention, suitable nonionic tonicity agents are polyols, sugars (in particular sucrose, fructose, dextrose or glucose) or amino acids such as glycine. In one embodiment the polyol is a sugar alcohol, especially a C 3-6 sugar alcohol. Exemplary sugar alcohols include glycerol, erythritol, treitol, arabitol, xylitol, ribitol, sorbitol, mannitol, dulcitol and iditol. In a specific example of this embodiment, a suitable nonionic tonicity agent is sorbitol. One skilled in the art will recognize that an appropriate osmolality can be achieved by using a mixture of different tonicity agents. In a particular embodiment of the invention the nonionic tonicity agent in the compositions of the invention incorporates sucrose and / or sorbitol.

In one embodiment, an appropriate concentration of polyol within the immunogenic composition is between about 2.5 and about 15% (w / v), in particular between about 2.5 and about 10% ( w / v), for example from about 3 to about 7% (w / v), such as from about 4 to about 6% (w / v). In a specific example of this embodiment, the polyol is sorbitol.

In another embodiment, the immunogenic composition comprises sucrose and sorbitol. In such circumstances the immunogenic composition may suitably contain from about 2.5 to about 15% (w / v) sucrose and from about 2.5 to about 15% (w / v) sorbitol, in particular between about 2.5 to about 10% (w / v) sucrose and about 2.5 to about 10% (w / v) sorbitol, for example about 3 to about 7% ( (w / v) sucrose and from about 3 to about 7% (w / v) sorbitol, such as from about 4 to about 6% (w / v) sucrose and from about 4 to about 6 % (w / v) sorbitol.

The pH of the immunogenic compositions should be appropriate for parenteral administration. Typically the pH will be in the range from 6.0 to 9.0. Suitably the pH will be in the range of 7.0 to 9.0, especially 7.25 to 8.75, such as 7.5 to 8.5, in particular pH 7.75 to 8.25. A pH of about 8.0 is of particular interest.

The pH may be controlled by the use of buffers, including, for example, tris or phosphate buffers.

The conductivity of an immunogenic composition of the invention may be measured using techniques known in the art, for example using a dedicated conductivity or other instrument with the ability to measure conductivity. A suitable instrument is Zetasizer Nano ZS from Malvem Instruments (UK).

The skilled artisan can readily test the concentration of both sodium (Na +) and chloride (C1-) ions using known techniques and kits. For example, sodium can be determined using a kit such as Biosupply's Sodium Enzymatic Assay Kit (Catalog Number: BQ011EAEL). Chloride can be determined using a kit such as Chloride Enzymatic Assay Kit (Catalog Number: BQ006EAEL) from Biosupply.

[000145] Immunogenic properties of the immunogenic composition of the present invention [000146] In the present invention the immunogenic composition is suitably capable of inducing a humoral response in a mammal, such as a human, administered with the immunogenic composition.

Humoral responses can be detected using an appropriate antibody-based test. For example, the presence or absence in serum of an immunoglobulin G (IgG) antibody response to a gp120-related polypeptide can be analyzed using ELISA. Induction of humoral responses as IgG antibodies, suitably IgG antibodies binding the gp120 VIV2 region, indicates the immunogenicity of the immunogenic compositions of the invention.

In another embodiment, the immunogenic composition is capable of inducing an enhanced CD4 T cell immune response.

By "enhanced CD4 T cell immune response" means that a larger CD4 response is obtained in a mammal, such as a human, upon administration of the adjuvanted immunogenic composition.

In particular but not exclusively, said 'enhanced CD4 T-cell immune response' is obtained in a nonimmunologically initiated patient, that is, a patient who is seronegative for HIV.

Enhanced CD4 T cell immune response (which may be provided by "polyfunctional" T cells) can be assessed by measuring the number of cells producing any of the following immune markers: • CD4 T cells expressing at least one marker immune cells • cells producing at least two different immune markers (eg CD40L, IL-2 and / or IFN-gamma, TNF-alpha) • cells producing at least CD40L and another immune marker (eg IL-2, TNF-gamma, and / or IFN-gamma) • cells producing at least IL-2 and another immune marker (eg CD40L, TNF-alpha and / or IFN-gamma) • cells producing at least IFN-gamma and other immune marker (eg IL -2, TNF-alpha and / or CD40L) • cells producing at least TNF-alpha and another immune marker (eg IL-2, CD40L and / or IFN-gamma) [000152] A CD4 T cell immune response will be present improved when cells producing any of the above immune markers are at a higher q amount after administration. Typically at least one, suitably two of the six conditions mentioned herein above will be met. In a particular embodiment, cells producing all four immune markers will be present in greater amount.

The enhanced CD4 T cell immune response conferred by the gp120 composition of the present invention can ideally be obtained after a single administration.

In another embodiment, administration of said immunogenic composition induces an improved memory cell B response in a mammal, such as a human, administered the immunogenic composition. An improved memory B response is intended to mean increased frequency of peripheral blood B lymphocytes capable of differentiation into plasma cells secreting antibodies upon antigen encounter, as measured by in vitro differentiation stimulation.

In a specific embodiment, administration of said immunogenic composition induces at least two of the following responses: (i) an enhanced CD4 T cell immune response, (ii) an improved B memory cell response, (iii) ) an improved humoral response against at least one of the antigen (s) component (s) or antigenic composition compared to the immune response obtained with other compositions.

The magnitude of an immune response may also be expressed as the titer (or concentration) of antigen-specific antibodies induced by the immunogenic composition as determined by an appropriate serological test. The large T cell response can be expressed as the frequency (or number of antigen-specific cells induced by an immunogenic composition among the total T cell population, which can be monitored by cytokine production.

Suitably the composition of the present invention elicits an immune response capable of cross-reactivity. Cross-reactivity is taken herein to mean the ability of immune responses induced by an immunogenic composition of the HIV-1 invention from subtypes that are not represented in an immunogenic composition. For example, an immunogenic composition of the invention comprising a gp120-related polypeptide comprising a subtype B HIV-1 strain is considered cross-reactive if the HIV-specific immune response, such as HIV-specific antibody or CD4 + T cell response ( in particular an antibody response in the gp120 V1V2 loop) induced by a composition reacted with one or more different strains of HIV-1 not in the composition, for example with a strain of HIV-1 from a different subtype of subtype. B. Suitably, cross-reactivity will be to one strain of HIV-1 from a different subtype, in particular to one strain of HIV-1 from a different group.

Suitably, the observed cross reactivity level is up to 10%, up to 15%, up to 20%, up to 25%, up to 35%, up to 40%, up to 45%, up to 50%, up to 55%, up to 60%, up to 65% to 70%, up to 80%, up to 90% or up to 100% of immunogenic composition-induced antigen-specific cells among the total T-cell population or specific antibody titer (or concentration) for antigen induced by immunogenic composition.

When measuring cross-reactivity in terms of the percentage of respondents to strains of HIV-1 of different subtypes, the number or percentage of vaccinated individuals showing a positive response on an immunoassay after subsequent challenge can be measured. The respondent can respond to one or more epitopes on an antigen. A respondent may also respond to one or more polypeptides in an immunogenic composition of the invention and / or one or more antigens in an immunogenic composition of the invention.

Immunological tests such as serological tests that can be used to analyze the percentage of respondents or the magnitude of an immune response are known in the art. Examples of such tests are known to the person skilled in the art.

Suitably, the observed cross-reactivity level is up to 10%, up to 15%, up to 25%, up to 30%, up to 35%, up to 40%, up to 45%, up to 50%, up to 55%, up to 60%, up to 65% up to 70%, up to 80%, up to 90% or up to 100% of individuals in a sample are respondents.

[000162] In one embodiment, the immunogenic composition of the invention is for use in eliciting high and long-term numbers of HIV-1 specific antibodies in an uninfected individual.

[000163] In another embodiment, the immunogenic composition of the invention is for use in eliciting high and long-term numbers of HIV-1 specific antibodies in an individual at risk of infection with an HIV-1 strain from one or more clades other than one or more HIV-1 clades from which the gp120 related polypeptide in an immunogenic composition is derived.

[000164] In one embodiment, the immunogenic composition of the invention is for use in controlling or reducing viremia in an HIV-infected individual.

Suitably, after administration of the composition, the individual's viral load remains below 100,000 copies / ml for at least four months after administration. In another embodiment, the subject's viral load remains below 100,000 copies / ml serum for at least six months, at least twelve months, at least eighteen months, at least two years, at least three years, at least four years. , at least five years, at least six years, at least seven years, at least eight years, at least nine years, or at least ten years. In another embodiment, the subject maintains a viral load below 50,000 copies / ml, below 10,000 copies / ml, below 5000 copies / ml, below 1000 copies / ml, or below 500 copies / ml. Suitably, viral load is maintained or reduced for at least six months, at least twelve months, at least eighteen months, at least two years, at least three years, at least four years, at least five years, at least six years, at least at least seven years, at least eight years, at least nine years, or at least ten years after administration of the composition.

Suitably, administration of the inventive composition results in a durable response. A durable response is, for example, the ability to detect, in a subject's serum, IgG antibody capable of binding to the V1V2 region of the gp120-related polypeptide of the composition for at least 24 weeks, at least 48 weeks, at least 72 weeks. , at least 96 weeks, at least two years, at least three years, at least four years, at least five years, at least six years, at least seven years, at least eight years, at least nine years, or at least ten years. years after the single administration of the composition, or the first administration of the composition in a course of repeated administrations to the subject, to the subject. Suitably, antibody levels will be detected at a level of at least 5%, more appropriately at least 10% and in particular at least 20% of the serum titer two weeks after the first administration. Suitably the antibody will be detectable in at least 50% of individuals, more appropriately at least 60% of individuals and in particular at least 75%.

Suitably, a durable response is, for example, the ability to detect, in an individual's serum, IgG antibody by binding to the gp120-related polypeptide V1V2 region of the composition for at least 2 weeks, at least 6 months, for at least 6 months. at least 12 months, at least 18 months, at least two years, at least three years, at least four years, at least five years, at least six years, at least seven years, at least eight years, at least nine years, or at least ten years after the final administration of the composition in a course of repeated administrations to the individual. Suitably, antibody levels will be detected at a level of at least 5%, more appropriately at least 10% and in particular at least 20% of the serum titer two weeks after final administration. Suitably the antibody will be detectable in at least 50% of individuals, more appropriately at least 60% of individuals and in particular at least 75%.

Suitably, the present invention is capable of achieving a more durable immune response based on respondent rates. Suitably up to 10% up to 15% up to 20% up to 25% up to 30% up to 35% up to 40% up to 45% up to 50% up to 55% up to 60% up to 65% up to Up to 70%, up to 80%, up to 90% or up to 100% of vaccinated individuals achieve an increased humoral response as an increased serum level of IgG antibody by binding to the gp120-related polypeptide V1V2 region of the composition.

Vaccination means The immunogenic compositions of the invention may be administered by any appropriate release route, such as intradermal, mucosal, for example intranasal, oral, intramuscular or subcutaneous. Other routes of release are well known in the art. The intramuscular release route is preferred for an immunogenic composition.

[000170] Intradermal release is another appropriate route. Any suitable device may be used for intradermal delivery, for example, short needle devices such as those described in US4886499. Intradermal vaccines may also be administered by devices that limit the extent of effective needle penetration into the skin. Also suitable are jet injection devices which release liquid vaccines into the dermis via a liquid jet injector that punctures the stratum corneum and produces a jet that reaches the dermis. Jet injection devices are described, for example, in US5480381. Also suitable are ballistic powder / particle devices that use compressed gas to accelerate a powdered vaccine through the outer layers of the skin for the dermis. In addition, conventional syringes may be used in the classic mantoux method of intradermal administration.

Another appropriate route of administration is the subcutaneous route. Any suitable device may be used for subcutaneous release, for example, classic needle. Suitably, a needleless jet injector service is used, such as that published in US6623446. More suitably said device is preloaded with the liquid vaccine formulation.

Alternatively, the vaccine is administered intranasally.

Typically, the vaccine is administered locally in the nasopharyngeal area, without being properly inhaled into the lungs. It is desirable to use an intranasal delivery device that releases the vaccine formulation into the nasopharyngeal area without or substantially without entering the lungs.

In a specific aspect of the present invention, the immunogenic composition may be given intramuscularly for the first administration, and a booster composition may be administered via a different route, for example intradermal, subcutaneous or intranasal.

In one aspect of the invention a schedule for vaccination with the immunogenic composition may comprise sequential, simultaneous "prime-boost" administration of the immunogenic composition and DNA encoding any of the above-mentioned proteins.

In one embodiment the schedule for vaccination with the immunogenic composition consists of three to five (e.g. table) administrations of the immunogenic composition to an individual over a period of four to eight (e.g. five to seven) months. Suitably, the schedule for vaccination with the immunogenic composition consists of four administrations of the immunogenic composition to an individual over a period of five to seven months.

Suitably, the gp120 related polypeptide of the invention may be replaced by a polynucleotide encoding the gp120 related polypeptide of the invention. Suitably, the polypeptide is codon optimized.

DNA may be released as plasmid DNA or as a recombinant live vector, for example, a poxvirus vector or any other appropriate live vector such as retrovirus, lentivirus, adenovirus, adeno-associated virus and modified vaccinia ankara virus ( MVA). Suitably, an adenovirus is used. Alternatively a canarypox virus can be used.

[000178] The immunogenic composition may be injected once or several times followed by one or more DNA administrations. DNA may be used first for one or more administrations followed by one or more immunizations with the immunogenic composition. Alternatively, the immunogenic composition may be injected once or several times in conjunction with DNA administrations.

Antiretrovirals [000179] HIV is a retrovirus. The conversion of your RNA into DNA is achieved through the action of the reverse transcriptase enzyme. Compounds that inhibit this reverse transcriptase function inhibit HIV replication in infected cells. Drugs incorporating these compounds are usable for the prevention or treatment of HIV infection in humans and may be used in conjunction with a composition of the present invention.

The composition of the present invention may be administered in conjunction with (ie before, during or after administration of) antiretroviral therapy (ART) as nucleoside or non-nucleoside reverse transcriptase inhibitors, protease inhibitors, fusion inhibitors, inhibitors maturation inhibitors, cellular inhibitors, and integrase filament transfer inhibitors.

Antiretroviral drugs include lamivudine and zidovudine, emtricitabine (FTC), zidovudine (ZDV), azidothymidine (AZT), lamivudine (3TC), zalcitabine, dideoxycytidine (ddC), tenofovir fumarate (tenofovir) tyropyridine (ddC) , and (d4T), abacavir sulfate (ABC), etravirine, delavirdine (DLV), efavirenz (EFV), nevirapine (NVP), amprenavir (APV), tipranavir (TPV), indinavir (IDV), saquinavir, saquinavir mesylate (SQV), lopinavir (LPV), ritonavir (RTV), calcium fosamprenavir (FOS-APV), ritonavir, RTV, darunavir, atazanavir sulfate (ATV), nelfinavir mesylate (NFV), enfuvirtide, T-20, maraviroc, dolutegravir and raltegravir. ART drugs may also include antibodies such as ibalizumab, HIV targeting proteins or cellular proteins associated with disease progression. Also included are immune-based therapies such as IL-2, IL-12 and alpha-epibromide. Each of these drugs may be administered alone or in combination with any other ART drug. Information about ART drugs and their administration can be found in various pharmacopoeias such as the United States Pharmacopoeia (USP) or accessed online, such as at www.aidsmeds.com (accessed September 5, 2013). Drug trade names and combinations These drugs include Atripla (efavirenz, emtricitabine and tenofovir disoproxil fumarate), Complera (emtricitabine, rilpivirine, and tenofovir disoproxyl fumarate), Stribild (elvitegravir, cobicistat, emtricitabine, emtricitabine, tenofovir fumarate, tenofovir (emtricitabine, FTC), Epivir (lamivudine, 3TC), Epzicom (abacavir and lamivudine), Hivid (zalcitabine, dideoxycytidine, ddC), Retro vir (zidovudine, azidothymine, AZT, ZDV), Trizivir (abacavudine, zidov) , Truvada (tenofovir disoproxil fumarate and emtricitabine), Videx EC (enteric coated didanosine, ddl EC), Videx (didanosine, dideoxyinosine, ddl), Viread (tenofovir disoproxyl fumarate) TDF), Z erit (stavudine, d4T), Ziagen (abacavir sulfate, ABC), rilpivirine, etravirine, Rescriptor (delavirdine, DLV), sustiva (efavirenz, EFV), Viramune (nevirapine, NVP), Agenerase (amprenavir, APV), Aptivir (tipran) , TPV), saquinavir, Invirase (saquinavir mesylate, SQV), Kaletra (lopinavir and ritonavir, LPV / RTV), Lexiva (Fosamprenavir Calcium, FOS-APV), Norvir (ritonavir, RTV), Darunavir, Reiataz (sulphate) , ATV), Viracept (nelfinavir mesylate, NFV), Fuzeon (enfuvirtide, T-20), maraviroc, raltegravir, dolutegravir.

[000182] In one embodiment, the immunogenic composition is administered to a patient who is also taking antiretrovirals. In another embodiment, the immunogenic composition is administered to a patient who has previously taken antiretrovirals. In another embodiment, the immunogenic composition is administered to a non-taking patient who has not previously taken antiretrovirals.

[000183] In one embodiment, the immunogenic composition is administered to a patient having a CD4 count of 200 or more, appropriately 500 or more, most appropriately between 500-1700 cells per cubic millimeter of blood. In another embodiment, the immunogenic composition is administered to a patient having a nadir CD4 count of 200 or more, more appropriately 500 or more cells per cubic millimeter of blood.

[000184] In one embodiment the immunogenic composition is administered to an individual, such as a human, who is not infected with HIV. In one embodiment the immunogenic composition is administered to an individual, such as a human, who is infected with HIV.

In one aspect of the invention, a gp120 related protein is provided for use in the manufacture of an immunogenic composition as described herein. As for the prophylaxis of HIV infection, alternatively, the composition may be intended for the treatment of HIV infection.

In another aspect there is provided a method for the prophylaxis of HIV infection by administering an immunogenic composition as described herein to a mammal, such as a human. This administration may reduce the risk of HIV infection and / or the severity of HIV infection.

In another aspect there is provided a method for treating HIV infection by administering an immunogenic composition as described herein to an HIV infected mammal, such as a human. This administration may reduce the severity of HIV infection.

In another aspect, a method is provided for reducing the risk of HIV transmission from an HIV-infected individual to a partner of said HIV-infected individual by administering an immunogenic composition as described herein to the infected individual. for HIV.

An immunogenic composition as described herein for use in the prophylaxis of HIV infection is also provided.

An immunogenic composition as described herein for use in the treatment of HIV infection is also provided.

The invention is illustrated by way of the following clauses: Clause 1. An immunogenic composition comprising a gp120 related polypeptide and an adjuvant, wherein the adjuvant comprises a lipopolysaccharide and an immunologically active saponin fraction derived from the Quillaja Saponaria Molina bark. presented in the form of a liposome and wherein the composition is substantially free of a Neitat related polypeptide wherein the NefTat related polypeptide is a polypeptide consisting of SEQ ID NO: 4.

Clause 2. An immunogenic composition which is in the form of a human dose comprising a gp120 related polypeptide and an adjuvant, wherein the adjuvant comprises from 10-40ug of a lipopolysaccharide and from 10-40ug of an immunologically active saponin fraction derived from Quillaja Saponaria Molina bark presented in the form of a liposome.

Clause 3. An immunogenic composition comprising a gp120 related polypeptide and an adjuvant, wherein the adjuvant comprises a lipopolysaccharide and an immunologically active saponin fraction derived from the Quillaja Saponaria Molina bark in the form of a liposome wherein: (i) the conductivity of the composition is 13 mS / cm or less; and / or (ii) the salt concentration in said composition is 130 mM or less; and / or (iii) the sodium chloride concentration in said composition is 130 mM or less.

Clause 4. The immunogenic composition according to clause 2 or 3, which is substantially free of a NefTat-related polypeptide wherein the NefTat-related polypeptide is a polypeptide consisting of SEQID NO: 4.

Clause 5. The immunogenic composition according to clause 1 or 4, which contains a NefTat: gp120-related polypeptide-related polypeptide ratio of less than 1:20.

Clause 6. The immunogenic composition according to clause 1 or 4, which contains less than 1ug of NefTat-related polypeptide.

Clause 7. The immunogenic composition according to clause 5 or 6 wherein the composition is free of NefTat-related polypeptide.

Clause 8. The immunogenic composition according to any one of clauses 1 or 4 to 7 wherein the NefTat-related polypeptide comprises a polypeptide with at least 99% identity to SEQ ID NO: 4.

Clause 9. The immunogenic composition according to clause 8 wherein the NefTat-related polypeptide comprises a polypeptide with at least 90% identity to SEQ ID NO: 4.

Clause 10. The immunogenic composition according to clause 9 wherein the NefTat-related polypeptide comprises a polypeptide with at least 80% identity to SEQ ID NO: 4.

Clause 11. The immunogenic composition according to clause 10 wherein the NefTat-related polypeptide comprises a polypeptide with at least 70% identity to SEQ ID NO: 4.

Clause 12. The immunogenic composition according to either clause 1 or 4, which contains a Nef-related polypeptide: gp120-related polypeptide ratio of less than 1:20.

Clause 13. The immunogenic composition according to clause 1 or 4, which contains less than 1 µg of Nef-related polypeptide.

Clause 14. The immunogenic composition according to clause 12 or 13 wherein the composition is free of Nef-related polypeptide.

Clause 15. The immunogenic composition according to any of clause 1,4oul2al4 wherein the Nef-related polypeptide comprises a polypeptide with at least 99% identity to SEQID NO: 2.

Clause 16. The immunogenic composition according to clause 15 wherein the Nef-related polypeptide comprises a polypeptide with at least 90% identity to SEQ ID NO: 2.

Clause 17. The immunogenic composition according to clause 16 wherein the Nef-related polypeptide comprises a polypeptide with at least 80% identity to SEQ ID NO: 2.

Clause 18. The immunogenic composition according to clause 17 wherein the Nef-related polypeptide comprises a polypeptide with at least 70% identity to SEQ ID NO: 2.

Clause 19. The immunogenic composition according to clause 1 or 4, which contains a Tat: gpl20-related Polypeptide-related polypeptide ratio of less than 1:20.

Clause 20. The immunogenic composition according to clause 1 or 4, which contains less than lug of Tat related polypeptide.

Clause 21. The immunogenic composition according to clause 19 or 20 wherein the composition is free of Tat related polypeptide.

Clause 22. The immunogenic composition according to any of clause 1,4oul9a21 wherein the Tat-related polypeptide comprises a polypeptide with at least 99% identity to SEQ ID NO: 3.

Clause 23. The immunogenic composition according to clause 22 wherein the Tat-related polypeptide comprises a polypeptide with at least 90% identity to SEQ ID NO: 3.

Clause 24. The immunogenic composition according to clause 23 wherein the Tat-related polypeptide comprises a polypeptide with at least 80% identity to SEQ ID NO: 3.

Clause 25. The immunogenic composition according to clause 24 wherein the Tat related polypeptide comprises a polypeptide with at least 70% identity to SEQ ID NO: 3.

Clause 26. The immunogenic composition according to any one of clauses 1 or 3 to 25 wherein the lipopolysaccharide is present at a level of between 10-100ug.

Clause 27. The immunogenic composition according to clause 26 wherein the lipopolysaccharide is present at a level of between 15-80ug.

Clause 28. The immunogenic composition according to clause 27 wherein the lipopolysaccharide is present at a level of between 20-65ug.

Clause 29. The immunogenic composition according to clause 28 wherein the lipopolysaccharide is present at a level of between 30-60ug.

Clause 30. The immunogenic composition according to clause 29 wherein the lipopolysaccharide is present at a level of between 45-55ug.

Clause 31. The immunogenic composition according to clause 30 wherein the lipopolysaccharide is present at a level of 50ug.

Clause 32. The immunogenic composition according to any one of clauses 1 or 3 to 31 wherein the saponin is present at a level of between 10-100ug.

Clause 33. The immunogenic composition according to clause 32 wherein the saponin is present at a level of between 15-80ug.

Clause 34. The immunogenic composition according to clause 33 wherein the saponin is present at a level of between 20-65ug.

Clause 35. The immunogenic composition according to clause 34 wherein the saponin is present at a level of between 30-60ug.

Clause 36. The immunogenic composition according to clause 35 wherein the saponin is present at a level of between 45-55ug.

Clause 37. The immunogenic composition according to clause 36 wherein the saponin is present at a level of 50ug.

Clause 38. The immunogenic composition according to any one of clauses 1 to 25 wherein the lipopolysaccharide is present at a level of between 15-35ug.

Clause 39. The immunogenic composition according to clause 38 wherein the lipopolysaccharide is present at a level of between 20-30ug.

Clause 40. The immunogenic composition according to clause 39 wherein the lipopolysaccharide is present at a level of 25ug.

Clause 41. The immunogenic composition according to any one of clauses 1 to 31 wherein the saponin is present at a level of between 15-35ug.

Clause 42. The immunogenic composition according to clause 41 wherein the saponin is present at a level of between 20-30ug.

Clause 43. The immunogenic composition according to clause 42 wherein the saponin is present at a level of 25ug.

Clause 44. The immunogenic composition according to any one of clauses 1 to 43 wherein the lipopolysaccharide is 3D-MPL.

Clause 45. The immunogenic composition according to any of clauses 1 to 44 wherein the saponin is QS-21.

Clause 46. The immunogenic composition according to any one of clauses 1, 2 or 4 to 45 wherein: (i) the conductivity of the composition is 13 mS / cm or less; and / or (ii) the salt concentration in said composition is 130 mM or less; and / or (iii) the sodium chloride concentration in said composition is 130 mM or less.

Clause 47. The immunogenic composition according to clause 46 wherein the conductivity of the composition is 13 mS / cm or less.

Clause 48. The immunogenic composition according to clause 47 wherein the conductivity of the composition is 6 mS / cm or less.

Clause 49. The immunogenic composition according to clause 48 wherein the conductivity of the composition is 1.5 to 2.5 mS / cm.

Clause 50. The immunogenic composition according to clause 46 wherein the concentration of salts in said composition is 130 mM or less.

Clause 51. The immunogenic composition according to clause 50 wherein the concentration of salts in said composition is 60 mM or less.

Clause 52. The immunogenic composition according to clause 51 wherein the concentration of salts in said composition is 20 to 40 mM.

Clause 53. The immunogenic composition according to clause 46 wherein the concentration of sodium chloride in said composition is 130 mM or less.

Clause 54. The immunogenic composition according to clause 53 wherein the concentration of sodium chloride in said composition is 60 mM or less.

Clause 55. The immunogenic composition according to clause 54 wherein the concentration of sodium chloride in said composition is 10 mM or less.

Clause 56. The immunogenic composition according to any of clauses 1 to 55, wherein the concentration of CaC12 in an immunogenic composition is 30 mM or less.

Clause 57. The immunogenic composition according to any of clauses 1 to 56, wherein the concentration of MgSO4 in an immunogenic composition is 60 mM or less.

Clause 58? The immunogenic composition according to any one of clauses 1 to 57, the total concentration of NH +, Mg2 + and Ca2 + ions is 40 mM or less.

Clause 59. The immunogenic composition according to any one of clauses 1 to 58 which is an aqueous solution.

Clause 60. The immunogenic composition according to any one of clauses 1 to 59 which is a single human dose.

Clause 61. The immunogenic composition according to any one of clauses 1 to 60 wherein the human dose is between 0.1 and 1 ml.

Clause 62. The immunogenic composition according to clause 61 wherein the human dose is between 0.3 and 0.75 ml.

Clause 63. The immunogenic composition according to clause 62 wherein the human dose is 0.5 ml.

Clause 64. The immunogenic composition according to any one of clauses 1 to 60 wherein the human dose is between 0.05 and 0.2 ml.

Clause 65. The immunogenic composition according to any of clauses 1 to 64 wherein the osmolality is in the range of 250 to 750 mOsm / kg.

Clause 66. The immunogenic composition according to clause 65 wherein the osmolality is in the range of 250 to 550 mOsm / kg.

Clause 67. The immunogenic composition according to any one of clauses 1 to 66, comprising a nonionic tonicity agent.

Clause 68. The immunogenic composition according to clause 67, wherein the nonionic tonicity agent is a polyol.

Clause 69. The immunogenic composition according to clause 68, wherein the polyol is sorbitol.

Clause 70. The immunogenic composition according to clause 69, wherein the concentration of sorbitol is between about 4 and about 6% (w / v).

Clause 71. The immunogenic composition according to any one of clauses 1 to 70, wherein the sucrose concentration is between about 4 and about 6% (w / v).

Clause 72. The immunogenic composition according to any one of clauses 1 to 71 wherein the pH is in the range of 6.0 to 9.0.

Clause 73. The immunogenic composition according to any one of clauses 1 to 72 wherein the composition comprises the gp120 related polypeptide at a level of about 1 to 100 µg.

Clause 74. The immunogenic composition according to clause 73 wherein the composition comprises the gp120 related polypeptide at a level of 3 to 30 µg.

Clause 75. The immunogenic composition according to clause 74 wherein the composition comprises the gp120 related polypeptide at a level of 5 to 15 µg.

Clause 76. The immunogenic composition according to clause 74 wherein the composition comprises the gp120 related polypeptide at a level of 16 to 25 µg.

Clause 77. The immunogenic composition according to any one of clauses 1 to 76 wherein the composition comprises 1-5 additional HIV antigens.

Clause 78. The immunogenic composition according to clause 77 wherein the composition comprises additional HIV antigens selected from the list consisting of gpl60, gp41, MA, CA, SP1, NC, SP2, P6, RT, RNase H, IN, PR , Rev, Vif, Vpr and Vpu.

Clause 79. The immunogenic composition according to any one of clauses 1 to 78 wherein the composition comprises a total of from 1 to 500 µg of antigenic material.

Clause 80. The composition according to any one of clauses 1 to 79 wherein the gp120 related polypeptide comprises an HIV-1 derived gp120 polypeptide.

Clause 81. The composition according to clause 80 wherein the gp120 related polypeptide comprises an HIV-1 gppo M-derived gp120 polypeptide.

Clause 82. The composition according to clause 81 wherein the gp120-related polypeptide comprises an HIV-1 subtype G-gp M-derived gp120 polypeptide.

Clause 83. The composition according to clause 82 wherein the gp120-related polypeptide comprises a gp1 M-derived polypeptide from HIV-1 subtype B.

Clause 84. The composition according to any one of clauses 1 to 81 and 83 wherein the gp120 related polypeptide comprises a polypeptide with at least 70% identity to the V1V2 region of SEQ ID NO: 1.

Clause 85. The composition according to clause 84 wherein the gp120 related polypeptide comprises a polypeptide with at least 80% identity to the VIV2 region of SEQ ID NO: 1.

Clause 86. The composition according to clause 85 wherein the gp120 related polypeptide comprises a polypeptide with at least 90% identity to the VIV2 region of SEQ ID NO: 1.

Clause 87. The composition according to clause 86 wherein the gp120 related polypeptide comprises a polypeptide with at least 95% identity to the VIV2 region of SEQ 2ID NO: 1.

Clause 88. The composition according to clause 87 wherein the gp120 related polypeptide comprises a polypeptide with at least 98% identity to the VIV2 region of SEQ ID NO: 1.

Clause 89. The composition according to clause 88 wherein the gp120 related polypeptide comprises a polypeptide with at least 99% identity to the VIV2 region of SEQ ID NO: 1.

Clause 90. The composition according to clause 89 wherein the gp120-related polypeptide comprises V1V2 region of SEQ ID NO: 1.

Clause 91. The composition according to any one of clauses 1 to 81 and 83 to 90 wherein the gp120 related polypeptide consists of the polypeptide with at least 70% identity to the V1V2 region of SEQ ID NO: 1.

Clause 92. The composition according to clause 91 wherein the gp120-related polypeptide consists of a polypeptide with at least 80% identity to the VIV2 region of SEQ ID NO: 1.

Clause 93. The composition according to clause 92 wherein the gp120-related polypeptide consists of a polypeptide with at least 90% identity to the VIV2 region of SEQ ID NO: 1.

Clause 94. The composition according to clause 93 wherein the gp120 related polypeptide consists of a polypeptide with at least 95% identity to the VIV2 region of SEQ ID NO: 1.

Clause 95. The composition according to clause 94 wherein the gp120 related polypeptide consists of a polypeptide with at least 98% identity to the VIV2 region of SEQ ID NO: 1.

Clause 96. The composition according to clause 95 wherein the gp120-related polypeptide consists of a polypeptide with at least 99% identity to the VIV2 region of SEQ ID NO: 1.

Clause 97. The composition according to clause 96 wherein the gp120 related polypeptide consists of the V1V2 region of SEQ ID NO: 1.

Clause 98. The composition according to any one of clauses 1 to 81 and 83 to 97 wherein the gp120 related polypeptide comprises a polypeptide with at least 70% identity to SEQ ID NO: 1.

Clause 99. The composition according to clause 98 wherein the gp120 related polypeptide comprises a polypeptide with at least 80% identity to SEQ ID NO: 1.

Clause 100. The composition according to clause 99 wherein the gp120 related polypeptide comprises a polypeptide with at least 90% identity to SEQ ID NO: 1.

Clause 101. The composition according to clause 100 wherein the gp120 related polypeptide comprises a polypeptide with at least 95% identity to SEQ ID NO: 1.

Clause 102. The composition according to clause 101 wherein the gp120 related polypeptide comprises a polypeptide with at least 98% identity to SEQ ID NO: 1.

Clause 103. The composition according to clause 102 wherein the gp120 related polypeptide comprises a polypeptide with at least 99% identity to SEQ ID NO: 1.

Clause 104. The composition according to clause 103 wherein the gp120 related polypeptide comprises SEQ ID NO: 1.

Clause 105. The composition according to any one of clauses 1 to 81 and 83 to 104 wherein gp120-related polypeptide consists of a polypeptide with at least 70% identity to SEQ ID NO: 1.

Clause 106. The composition according to clause 105 wherein the gp120-related polypeptide consists of a polypeptide with at least 80% identity to SEQ ID NO: 1.

Clause 107. The composition according to clause 106 wherein the gp120 related polypeptide consists of a polypeptide with at least 90% identity to SEQ ID NO: 1.

Clause 108. The composition according to clause 107 wherein the gp120 related polypeptide consists of a polypeptide with at least 95% identity to SEQ ID NO: 1.

Clause 109. The composition according to clause 108 wherein the gp120-related polypeptide consists of a polypeptide with at least 98% identity to SEQ ID NO: 1.

Clause 110. The composition according to clause 109 wherein the gp120-related polypeptide consists of a polypeptide with at least 99% identity to SEQ ID NO: 1.

Clause 111. The composition according to clause 110 wherein the gp120 related polypeptide consists of SEQ ID NO: 1.

Clause 112. The composition according to any one of clauses 1 to 82 wherein the gp120 related polypeptide comprises a polypeptide with at least 70% identity to the V1V2 region of SEQ ID NO: 5.

Clause 113. The composition according to clause 112 wherein the gp120 related polypeptide comprises a polypeptide with at least 80% identity to the VIV2 region of SEQ ID NO: 5.

Clause 114. The composition according to clause 113 wherein the gp120 related polypeptide comprises a polypeptide with at least 90% identity to the VIV2 region of SEQ ID NO: 5.

Clause 115. The composition according to clause 114 wherein the gp120 related polypeptide comprises a polypeptide with at least 95% identity to the VIV2 region of SEQ ID NO: 5.

Clause 116. The composition according to clause 115 wherein the gp120 related polypeptide comprises a polypeptide with at least 98% identity to the VIV2 region of SEQ ID NO: 5.

Clause 117. The composition according to clause 116 wherein the gp120 related polypeptide comprises a polypeptide with at least 99% identity to the VIV2 region of SEQ ID NO: 5.

Clause 118. The composition according to clause 117 wherein the gp120 related polypeptide comprises the V1V2 region of SEQ ID NO: 5.

Clause 119. The composition according to any one of clauses Ia82ell2all8 wherein the gp120 related polypeptide consists of a polypeptide with at least 70% identity to the V1V2 region of SEQ ID NO: 5.

Clause 120. The composition according to clause 119 wherein the gp120-related polypeptide consists of a polypeptide with at least 80% identity to the VIV2 region of SEQ ID NO: 5.

Clause 121. The composition according to clause 120 wherein the gp120-related polypeptide consists of a polypeptide with at least 90% identity to the VIV2 region of SEQ ID NO: 5.

Clause 122. The composition according to clause 121 wherein the gp120-related polypeptide consists of a polypeptide with at least 95% identity to the VIV2 region of SEQ ID NO: 5.

Clause 123. The composition according to clause 122 wherein the gp120-related polypeptide consists of a polypeptide with at least 98% identity to the VIV2 region of SEQ ID NO: 5.

Clause 124. The composition according to clause 123 wherein the gp120-related polypeptide consists of a polypeptide with at least 99% identity to the VIV2 region of SEQ ID NO: 5.

Clause 125. The composition according to clause 124 wherein the gp120 related polypeptide consists of the V1V2 region of SEQ ID NO: 5.

Clause 126. The composition according to any one of clauses 1 to 82 and 112 to 125 wherein the gp120 related polypeptide comprises a polypeptide with at least 70% identity to SEQ ID NO: 5.

Clause 127. The composition according to clause 126 wherein the gp120 related polypeptide comprises a polypeptide with at least 80% identity to SEQ ID NO: 5.

Clause 128. The composition according to clause 127 wherein the gp120 related polypeptide comprises a polypeptide with at least 90% identity to SEQ ID NO: 5.

Clause 129. The composition according to clause 128 wherein the gp120 related polypeptide comprises a polypeptide with at least 95% identity to SEQ ID NO: 5.

Clause 130. The composition according to clause 129 wherein the gp120 related polypeptide comprises a polypeptide with at least 98% identity to SEQ ID NO: 5.

Clause 131. The composition according to clause 130 wherein the gp120 related polypeptide comprises a polypeptide with at least 99% identity to SEQ ID NO: 5.

Clause 132. The composition according to clause 131 wherein the gp120 related polypeptide comprises SEQ ID NO: 5.

Clause 133. The composition according to any one of clauses 1 to 82 and 112 to 132 wherein the gp120 related polypeptide consists of a polypeptide with at least 70% identity to SEQ ID NO: 5.

Clause 134. The composition according to clause 133 wherein the gp120-related polypeptide consists of a polypeptide with at least 80% identity to SEQ ID NO: 5.

Clause 135. The composition according to clause 134 wherein the gp120-related polypeptide consists of a polypeptide with at least 90% identity to SEQ ID NO: 5.

Clause 136. The composition according to clause 135 wherein the gp120 related polypeptide consists of a polypeptide with at least 95% identity to SEQ ID NO: 5.

Clause 137. The composition according to clause 136 wherein the gp120-related polypeptide consists of a polypeptide with at least 98% identity to SEQ ID NO: 5.

Clause 138. The composition according to clause 137 wherein the gp120 related polypeptide consists of a polypeptide with at least 99% identity to SEQ ID NO: 5.

Clause 139. The composition according to clause 138 wherein the gp120 related polypeptide consists of SEQ ID NO: 5.

Clause 140. The immunogenic composition according to any one of clauses 1 to 139 wherein the composition further comprises a sterol, wherein the saponin: sterol ratio is 1: 1 to 1: 100 weight / weight.

Clause 141. The immunogenic composition according to clause 140 wherein the saponin: sterol ratio is 1: 1 to 1: 5 weight / weight.

Clause 142. The immunogenic composition according to clause 140 or 141 wherein said sterol is cholesterol.

Clause 143. The immunogenic composition according to any one of clauses 1 to 142, which is provided in a multiple dose presentation.

Clause 144. The immunogenic composition according to any one of clauses 1 to 143, which is provided in a presentation containing an average of 1-50% to allow for a waste during administration.

Clause 145. The immunogenic composition according to any one of clauses 1 to 144 for use as a medicament.

Clause 146. The immunogenic composition according to clause 145 for use in the treatment or prevention of HIV-1 from group Μ, N, O or P.

Clause 147. The immunogenic composition according to clause 146 for use in the treatment or prevention of group M of HIV-1 subtype A, B, C, D, E, F, G, Η, I, J or K.

Clause 148. The immunogenic composition according to any one of clauses 1 to 147, wherein the gp120-related polypeptide is derived from a first HIV-1 subtype, for use in the treatment or prevention of HIV-1 infection for a second HIV-1 subtype.

Clause 149. The immunogenic composition according to any one of clauses 1 to 148, wherein the gp120-related polypeptide is derived from a first HIV-1 subtype, for use in the treatment or prevention of HIV-1 infection per second. HIV-1 subtype, wherein the first and second HIV-1 subtypes have different native polypeptide gp120 sequences.

Clause 150. The immunogenic composition according to clause 149, wherein the first subtype of HIV-1 is selected from the list consisting of A, B, C, D, E, F, G, I, I, J or K; group M of HIV-1.

Clause 151. The immunogenic composition of any one of clauses 1 to 150 for use in inducing the humoral immune response against HIV-1 strains of one or more different subtypes of the HIV-1 subtype of which the gpl20-related polypeptide of the composition is derivative.

Clause 152. The immunogenic composition according to any one of clauses 1 to 151 for use in eliciting antibodies against the HIV-1 gp120 V1V2 loop.

Clause 153. A method of treating or prophylaxis of HIV-1 infection comprising the step of administering a composition according to any one of clause 1 to 152 to an individual.

Clause 154. The method of treatment or prophylaxis of HIV-1 infection according to clause 153 further comprising concomitant administration of an antiretroviral drug.

Clause 155. The method of treatment or prophylaxis of HIV-1 infection according to clause 153 or 154 wherein the IgG antibody capable of binding to the gp120-related polypeptide V1V2 region of the composition is detectable in a subject's serum by at least 24 weeks after the single administration of the composition or the first administration of the composition in a course of repeated administrations to the subject.

Clause 156. The method of treatment or prophylaxis of HIV-1 infection according to clause 155 wherein the IgG antibody is detectable at least 48 weeks after the single administration of the composition or the first administration of the composition in a course of repeated administrations. for the individual.

Clause 157. The method of treatment or prophylaxis of HIV-1 infection according to clause 156 wherein the IgG antibody is detectable at least 72 weeks after the single administration of the composition or the first administration of the composition in a course of repeated administrations. for the individual.

Clause 158. The method of treatment or prophylaxis of HIV-1 infection according to clause 157 wherein the IgG antibody is detectable at least 96 weeks after the single administration of the composition or the first administration of the composition in a course of repeated administrations. for the individual.

Clause 159. The method of treatment or prophylaxis of HIV-1 infection according to clause 153 or 154 wherein IgG antibody capable of binding to the gp120-related polypeptide V1V2 region of the composition is detectable in a subject's serum by at least 2 weeks after final administration of the composition in a course of repeated administrations to the subject.

Clause 160. The method of treatment or prophylaxis of HIV-1 infection according to clause 159 wherein the IgG antibody is detectable at least 6 months after the final administration of the composition in a course of repeated administrations to the subject.

Clause 161. The method of treatment or prophylaxis of HIV-1 infection according to clause 160 wherein the IgG antibody is detectable at least 12 months after the final administration of the composition in a course of repeated administrations to the subject.

Clause 162. The method of treatment or prophylaxis of HIV-1 infection according to clause 161 wherein the IgG antibody is detectable at least 18 months after the final administration of the composition in a course of repeated administrations to the subject.

Clause 163. The method of treatment or prophylaxis of HIV-1 infection according to any of clauses 155 to 158 wherein antibody levels are detectable in at least 5% of the serum titer two weeks after the single administration.

Clause 164. The method of treatment or prophylaxis of HIV-1 infection according to any of clauses 155 to 162 wherein antibody levels are detectable in at least 5% of the serum titer two weeks after the first administration.

Clause 165. The method of treatment or prophylaxis of HIV-1 infection according to any of clauses 159 to 162 wherein antibody levels are detectable in at least 5% of the serum titer two weeks after final administration.

Clause 166. The method of treatment or prophylaxis according to any of clauses 155 to 165 wherein the antibody will be detectable in at least 50% of subjects administered the composition.

Clause 167. The method of treatment or prophylaxis according to clause 166 wherein the antibody will be detectable in at least 80% of subjects administered the composition.

Clause 168. The method of treatment or prophylaxis according to any of clauses 153 to 167 wherein a polynucleotide encoding a gp120 related polypeptide is administered to the subject and subsequently the composition according to any of clauses 1 to 152 is administered to the subject. individual.

Clause 169. The method of treatment or prophylaxis according to any one of clause 153 to 167 wherein the composition according to any one of clause 1 to 152 is administered to the subject and subsequently a polynucleotide encoding a gp120 related polypeptide is administered to the subject. individual.

Clause 170. A method of reducing the risk of HIV transmission from an HIV-infected individual to a partner of said HIV-infected individual comprising the step of administering the immunogenic composition according to any of clauses 1 to 152 to the individual. infected with HIV.

Clause 171. A method of producing a composition according to any one of clauses 1 to 152, comprising adding one to gp120-derived polypeptide to a lipopolysaccharide and an immunologically active saponin fraction derived from the Quillaja Saponaria Molina bark presented as a liposome.

Clause 172. A kit for producing an immunogenic composition according to any one of clauses 1 to 152, comprising a first container and a second container, wherein the first container comprises a gp120-derived polypeptide and the second container comprises a lipopolysaccharide and a immunologically active saponin fraction derived from the Quillaja Saponaria Molina bark presented as a liposome.

Clause 173. The kit according to clause 172, wherein the second container comprises an aqueous solution.

Clause 174. A viral vector comprising a polynucleotide encoding a polypeptide comprising the gpl20W61D polypeptide of SEQ ID NO: 1.

Clause 175. The viral vector of clause 174 comprising a polynucleotide encoding a polypeptide consisting of gp120W61D polypeptide of SEQ ID NO: 1.

Clause 176. A viral vector comprising a polynucleotide encoding a polypeptide comprising the gp120ZMi8 polypeptide of SEQ ID NO: 5.

Clause 177. The viral vector of clause 176 comprising a polynucleotide encoding a polypeptide consisting of gp120ZMi8 polypeptide of SEQ ID NO: 5.

Clause 178. The viral vector of any one of clause 174 to 177 which is an adenovirus, canarypox virus or MVA virus.

[000192] The present invention will now be further described by way of non-limiting examples.

EXAMPLES

Example 1: Comparison of serology Method [000193] A first composition containing 2ug gpl20w6iD and 50ul AS01B and a second composition containing 2ug gpl20w6iD, 50ul AS01B and 2ug NefTat were prepared. At three time points (days 0, 14 and 28), the first composition was administered intramuscularly (IM) to a first group of 25 CB6F1 mice and at the same three time points the second composition was administered IM to a second group of 25 mice. CB6F1 mice.

The titer of anti-VlV2 antibodies in mouse sera in each group was compared at 14 days after the second administration and at 14 days after the third administration. To assess the amount of anti-VlV2 IgG antibodies present in each serum sample, the HIV-1 subtype B case A2 gpl20 scaffolding V1V2 region over a murine gp70 protein was used in an Igtot ELISA binding antibody assay. During the development of the tests, CH58 (the anti-V2 monoclonal antibody used in the RV144 test) was used as a positive control to successfully detect the binding of serum Igtot antibodies to VIV2 with scaffolding. In addition, the gp70 protein (not bound to a VIV2 region) was used as a negative control to assess specificity during test development.

Results At both time points tested, a trend towards an anti-VlV2 antibody response was observed in serum samples from mice that had been administered the non-NefTat-containing composition as compared to serum samples from mice that had been administered with the NefTat containing composition. This was particularly noticeable 14 days after the third administration (see Figure 1).

All references given in this application, including patents and patent applications, are incorporated herein by reference to the greatest extent possible.

Throughout the report and the claims that follow, unless the context otherwise requires, the word "understand" and variations such as "understand" and "understanding" shall be understood to imply the inclusion of an integer, step, whole number group or step groups described, but not to the exclusion of any other integer, step, whole number group, or step groups.

The order of which this description and claims form part may be used as the basis for priority over any subsequent order. The claims of such subsequent application may be directed to any aspect or combination of aspects described herein. They may take the form of product, composition, process claims and may include, by way of example, and without limitation, the following claims. SEQUENCE LISTING SEQ ID NO:. L-gpl20w n AEQLWVTVYYGVPVWKEATTTLFCASDAKAYDTEVHNVWATHACVPTDPNPQEVVLGNVTEYFNMWKNNM VDQMHEDIISLWDQSLKPCVKLTPLCVTLDCDDVNTTNSTTTTSNGWTGEIRKGEIKNCSFNITTSIRDK VQKEYALFYNLDVVPIDDDNATTKNKTTRNFRLIHCNSSVMTQACPKVSFEPIPIHYCAPAGFAILKCNN KTFDGKGLCTNVSTVQCTHGIRPVVSTQLLLNGSLAEEEVVIRSDNFMDNTKTIIVQLNESVAINCTRPN NNTRKGIHIGPGRAFYAARKIIGDIRQAHCNLSRAQWNNTLKQIVIKLREHFGNKTIKFNQSSGGDPEIV RHSFNCGGEFFYCDTTQLFNSTWNGTEGNNTEGNSTITLPCRIKQIINMWQEVGKAMYAPPIGGQIRCSS NITGLLLTRDGGTEGNGTENETEIFRPGGGDMRDNWRSELYKYKWKVEPLGVAPTRAKRRWQR V1V2 region underlined. SEQ ID NO: 2 - Nef MGGKWSKSSVVGWPTVRERMRRAEPAADGVGAASRDLEKHGAITSSNTAATNAACAWLEAQEEEEVGFPV TPQVPLRPMTYKAAVDLSHFLKEKGGLEGLIHSQRRQDILDLWIYHTQGYFPDWQNYTPGPGVRYPLTFG WCYKLVPVEPDKVEEANKGENTSLLHPVSLHGMDDPEREVLEWRFDSRLAFHHVARELHPEYFKNC SEQ ID NO: 3 - Tat EPVDPRLEPWRHPGSQPKTACTNCYCKKCCFHCQVCFITKALGISYGRKKRRQRRRPPQGSQTHQVSLSK

QPTSQSRGDPTGPKE SEQ ID NO: 4 - NefTat MGGKWSKSSWGWPTVRERMRRAEPAADGVGAASRDLEKHGAITSSNTAATNAACAWLEAQEEEEVGFPV TPQVPLRPMTYKAAVDLSHFLKEKGGLEGLIHSQRRQDILDLWIYHTQGYFPDWQNYTPGPGVRYPLTFG WCYKLVPVEPDKVEEANKGENTSLLHPVSLHGMDDPEREVLEWRFDSRLAFHHVARELHPEYFKNCTSE PVDPRLEPWKHPGSQPKTACTNCYCKKCCFHCQVCFITKALGISYGRKKRRQRRRPPQGSQTHQVSLSKQ - PTSQSRGDPTGPKE

Nef portion: amino acids 1-206 Tat portion: amino acids 209-293 (underline) Box: additional binding amino acids introduced into fusion protein SEQ ID NO: 5 - gp 120ZM18 GDNLWVTVYYGVPVWKEAKTTLFCASDAKAYEREVHNVWATHACVPTDPNPQEENMNG

VDQMHEDIIRLWDQSLKPCVKLTPLCVTLECGNVNVTHENSTKGEMKNCSFNATTELKDKKQRVYALFYK

LDIVPLNENNNSSEDSSEYRLINCiSITSAITQACPKVTLDPIPIHYCAPAGYAILKCNNKTFNGTGPCHNV

STVQCTHGIKPVVSTQLLLNGSLAEEEIIIRSENLTNNAKTIIVHLNESVEIVCTRPSNNTRKSIRIGPG

QAFYATGGIIGNIRQAHCNISKENWNKTLQKVGKKLAEHFPNKTIKFDQHSGGDLEITTHSFNCRGEFFY

CNTSNLFNSTYKPNDTNSTYNPNDTITLPCRIKQIINMWQGVGQAMYAPPIAGNITCKSNITGLLLTRDG

GSNDTTNTETFRPGGGDMRDNWRSELYKYKVVEIKTLGIAPTAAKRRVVETR underlined V1V2 region. SEQ ID NO: 6 - _gpl20ZM18 polynucleotide sequence encoding ggagacaacttgtgggtcacagtctattatggggtacctgtgtggaaagaagcaaaaactactttattctgtg catcagatgctaaagcatatgagagagaagtgcataatgtctgggctacacatgcctgtgtacccac agaccccaacccacaagaaatagttttgggaaatgtaacagaaaattttaacatgtggaaaaatgacatg gtggatcagatgcatgaggatataatcaggttatgggatcaaagcttaaagccatgtgtaaagttgaccc cactctgtgtcactttagaatgtggaaatgttaatgttacccatgagaatagcacgaagggggaaatgaa aaattgctctttcaatgcaaccacagaactaaaagataaaaaacagagagtgtatgcacttttttataaa cttgatatagtaccacttaatgagaataacaactctagtgaggactctagtgagtatagattaataaatt gtaatacctcagccataacacaagcctgtccaaaggtcactttggacccaattcctatacattattgtgc tccagctggatatgcgattctaaagtgtaataataagacattcaatgggacaggaccatgccataatgtc agcacagtacaatgtacacacggaatcaagccagtggtatcaactcaactactgttaaatggtagcctag cagaagaagagataataattaggtctgaaaatctaacaaacaatgccaaaacaataatagtacatcttaa tgaatctgtagaaattgtgtgtacaagacccagcaataatacaagaaaaagtataaggataggaccagga caagcattctatgcaacaggtggcataataggaaacataagacaagcacattgtaacattagtaaagagaact ggaat aaaactttacaaaaggtaggaaaaaaattagcagagcacttccctaataaaacaataaaatt tgaccaacactcaggaggggacctagaaattacaacacatagctttaattgtagaggagaatttttctat tgcaatacatcaaacctgtttaatagtacatataagcctaatgatacaaatagtacatataatcctaatg atacaatcacactcccatgcagaataaaacaaattataaacatgtggcagggggtaggacaagcaatgta tgcccctcccattgcaggaaacataacatgtaaatcaaatatcacaggactactattgacacgggatgga gggtcaaatgataccacaaacacagagacattcagacctggaggaggagatatgagggacaattggagaa gtgaactatataaatataaagtggtagaaattaaaacattgggcatagcacccactgcggcaaaaaggag agtggtggagacgagataa SEQ ID NO: 7 -GP 1 2.0zm 18nativo GDNLWVTVYYGVPVWKEAKTTLFCASDAKAYEREVHNVWATHACVPTDPNPQEIVLGNVTENFNMWKNDM VDQMHEDIIRLWDQSLKPCVKLTPLCVTLECGNVNVTHENSTKGEMKNCSFNATTELKDKKQRVYALFYK LDIVPLNENNNSSEDSSEYRLINCNTSAITQACPKVTLDPIPIHYCAPAGYAILKCNNKTFNGXGPCHNV STVQCTHGIKPVVSTQLLLNGSLAEEEIIIRSENLTNNAKTIIVHLNESVEIVCTRPSNNTRKSIRIGPG QAFYATGGIIGNIRQAHCNISKENWNKTLQKVGKKLAEHFPNKTIKFDQHSGGDLEITTHSFNCRGEFFY CNTSNLFNSTYKPNDTNSTYNPNDTITLPCRIKQIINMWQGVGQAMYAPPIAGNITCKSNITGLLLTRDG GSNDTTNTETFRPGGGDMRDNWRSELYKYKV VEIKPLGIAPTAAKRRVVETR

Gp! 20ZM18 native polypeptide sequence contains underlined proline instead of threonine. SEQ ID NO: 8 - polynucleotide sequence encoding gpl207.Mi 'native ggagacaacttgtgggtcacagtctattatggggtacctgtgtggaaagaagcaaaaactactttattct gtgcatcagatgctaaagcatatgagagagaagtgcataatgtctgggctacacatgcctgtgtacccac agaccccaacccacaagaaatagttttgggaaatgtaacagaaaattttaacatgtggaaaaatgacatg gtggatcagatgcatgaggatataatcaggttatgggatcaaagcttaaagccatgtgtaaagttgaccc cactctgtgtcactttagaatgtggaaatgttaatgttacccatgagaatagcacgaagggggaaatgaa aaattgctctttcaatgcaaccacagaactaaaagataaaaaacagagagtgtatgcacttttttataaa cttgatatagtaccacttaatgagaataacaactctagtgaggactctagtgagtatagattaataaatt gtaatacctcagccataacacaagcctgtccaaaggtcactttggaliccaattcctatacattattgtgc tccagctggatatgcgattctaaagtgtaataataagacattcaatgggacaggaccatgccataatgtc agcacagtacaatgtacacacggaatcaagccagtggtatcaactcaactactgttaaatggtagcctag cagaagaagagataataattaggtctgaaaatctaacaaacaatgccaaaacaataatagtacatcttaa tgaatctgtagaaattgtgtgtacaagacccagcaataatacaagaaaaagtataaggataggaccagga caagcattctatgcaacaggtggcataataggaaacataagacaagcacattgtaacattagtaaagaga actggaataaaactttacaaaaggtaggaaaaaaattagcagagcacttccctaataaaacaataaaatt tgaccaacactcaggaggggacctagaaattacaacacatagctttaattgtagaggagaatttttctat tgcaatacatcaaacctgtttaatagtacatataagcctaatgatacaaatagtacatataatcctaatg atacaatcacactcccatgcagaataaaacaaattataaacatgtggcagggggtaggacaagcaatgta tgcccctcccattgcaggaaacataacatgtaaatcaaatatcacaggactactattgacacgggatgga gggtcaaatgataccacaaacacagagacattcagacctggaggaggagatatgagggacaattggagaa gtgaactatataaatataaagtggtagaaattaaaccattgggcatagcacccactgcggcaaaaaggag agtggtggagacgagataa [000199] To produce the polynucleotide sequence gpl20ZMi8 from the native polynucleotide gpl sequence 20Z \ m: a) a silent point mutation is introduced (C instead of T at position 597 (underlined ). b) non-silent point mutation A instead of C is introduced at position 1426 (underline) by modifying native codon CCA (proline) by codon AC A (threonine).

Claims (32)

1. Immunogenic composition, characterized in that it comprises a gp120 related polypeptide and an adjuvant, wherein the adjuvant comprises a lipopolysaccharide and an immunologically active saponin fraction derived from the Quillaja Saponaria Molina bark in which the liposome is present. The composition is substantially free of a NefTat related polypeptide wherein the NefTat related polypeptide is a polypeptide consisting of SEQ ID NO: 4. Immunogenic composition, characterized in that it is in the form of a human dose comprising a gp120 related polypeptide and an adjuvant, wherein the adjuvant comprises between 10-40ug of a lipopolysaccharide and 10-40ug of an immunologically active saponin fraction derived from it. of the shell of Quillaja Saponaria Molina presented as a liposome. 3. Immunogenic composition, characterized in that it comprises a gp120 related polypeptide and an adjuvant, wherein the adjuvant comprises a lipopolysaccharide and an immunologically active saponin fraction derived from the Quillaja Saponaria Molina bark in the form of a liposome in which: ( i) the conductivity of a composition is 13 mS / cm or less; and / or (ii) the salt concentration in said composition is 130 mM or less; and / or (iii) the sodium chloride concentration in said composition is 130 mM or less. Immunogenic composition according to one of claims 2 or 3, characterized in that it is substantially free of a NefTat-related polypeptide wherein the NefTat-related polypeptide is a polypeptide consisting of SEQ ID NO: 4. Immunogenic composition according to any one of claims 1, 3 or 4, characterized in that the lipopolysaccharide is present at a level of between 10-100ug. Immunogenic composition according to any one of claims 1 or 3 to 5, characterized in that the saponin is present at a level of between 10-100ug. Immunogenic composition according to any one of claims 1 to 6, characterized in that the lipopolysaccharide is 3D-MPL. Immunogenic composition according to any one of claims 1 to 7, characterized in that the saponin is QS-21. Immunogenic composition according to any one of claims 1, 2 or 4 to 8, characterized in that: (i) the conductivity of a composition is 13 mS / cm or less; and / or (ii) the salt concentration in said composition is 130 mM or less; and / or (iii) the sodium chloride concentration in said composition is 130 mM or less. Immunogenic composition according to any one of claims 1 to 9, characterized in that it is an aqueous solution. Immunogenic composition according to any one of claims 1 to 10, characterized in that it is a single human dose. Immunogenic composition according to any one of claims 1 to 11, characterized in that the human dose is between 0.1 and 1 ml. Immunogenic composition according to any one of claims 1 to 12, characterized in that it comprises a nonionic tonicity agent. Immunogenic composition according to any one of claims 1 to 13, characterized in that the composition comprises the gp120 related polypeptide at a level of about 1 to 100 µg. Immunogenic composition according to any one of claims 1 to 14, characterized in that the composition comprises 1-5 additional HIV antigens. Composition according to any one of Claims 1 to 15, characterized in that the gp120-related polypeptide comprises a polypeptide with at least 70% identity to the VIV2 region of SEQ ID NO: 1. Composition according to any one of Claims 1 to 16, characterized in that the gp120-related polypeptide comprises a polypeptide of at least 70% identity to SEQ ID NO: 1. Composition according to any one of Claims 1 to 15, characterized in that the gp120-related polypeptide comprises a polypeptide with at least 70% identity to the VIV2 region of SEQ ID NO: 5. Composition according to any one of Claims 1 to 16, characterized in that the gp120-related polypeptide comprises a polypeptide of at least 70% identity to SEQ ID NO: 5. Immunogenic composition according to any one of claims 1 to 19, characterized in that the composition further comprises a sterol, wherein the saponin: sterol ratio is 1: 1 to 1: 100 weight / weight. Immunogenic composition according to any one of claims 1 to 20, characterized in that it is for use in the treatment or prevention of group Μ, N, O or P HIV-1. Immunogenic composition according to any one of claims 1 to 20, characterized in that it is for use in the treatment or prevention of HIV-1 group M, subtype A, B, C, D, E, F, G, Η, I, J or K. Immunogenic composition according to any one of claims 1 to 22, characterized in that the gp120-related polypeptide is derived from a first subtype of HIV-1 for use in the treatment or prevention of HIV-1 infection by a second HIV-1 subtype, wherein the first and second HIV-1 subtypes have different native gp120 polypeptide sequences. Immunogenic composition according to any one of claims 1 to 23, characterized in that it is for use to elicit antibodies against HIV-1 gp120 V1V2 loop. Immunogenic composition according to any one of claims 1 to 23, characterized in that it is for use in the treatment or prophylaxis of HIV infection wherein a polynucleotide encoding a gp120 related polypeptide is administered to the subject and subsequently the composition as defined. in any one of claims 1 to 24 is administered to the individual. Immunogenic composition according to any one of claims 1 to 23, characterized in that it is for use in the treatment or prophylaxis of HIV infection wherein the composition as defined in any one of claims 1 to 23 is administered to the subject and subsequently a polynucleotide encoding a gp120 related polypeptide is administered to the subject. A method of treating or prophylaxis of HIV-1 infection, comprising the step of administering the composition as defined in any one of claims 1 to 26 for an individual. A method for reducing the risk of HIV transmission from an HIV-infected individual to a partner of said HIV-infected individual, comprising the step of administering the immunogenic composition as defined in any one of claims 1 to 26 to said HIV-infected individual. A method for producing a composition as defined in any one of claims 1 to 26, comprising adding a gp120-derived polypeptide to a lipopolysaccharide and an immunologically active saponin fraction derived from the Quillaja Saponaria Molina bark presented as a a liposome. Kit for producing an immunogenic composition according to any one of claims 1 to 26, characterized in that it comprises a first container and a second container, wherein the first container comprises a gp120-derived polypeptide and the second container comprises a lipopolysaccharide. and an immunologically active saponin fraction derived from the Quillaja Saponaria Molina bark presented as a liposome. Viral vector, characterized in that it comprises a polynucleotide encoding a polypeptide comprising the gpl20W61D polypeptide of SEQ ID NO: 1. 32. Viral vector, characterized in that it comprises a polynucleotide encoding a polypeptide comprising the gpl20ZM18 polypeptide of SEQ ID NO: 5.