COMBINATION HIV VACCINE

CROSS REFERENCE TO RELATED APPLICATION

[0001] This application claims the priority benefit of U.S. Provisional Application No. 63/380,113, filed October 19, 2022, which is incorporated herein by reference in its entirety.

REFERENCE TO SEQUENCE LISTING SUBMITTED ELECTRONICALLY

[0002] The content of the electronically submitted sequence listing (Name: 3834_011PC02_SequenceListing_ST26.xml; Size: 31,247 bytes; and Date of Creation: October 13, 2023) is herein incorporated by reference in its entirety.

BACKGROUND

[0003] HIV was first identified and described as the cause of AIDS four decades ago. Since then, 79.3 million people have become infected with HIV, from which more than 36 million people have already died from AIDS-related illnesses. Nowadays 37.7 million people globally are living with HIV (People Living With HIV, PLWH), with 1.5 million people newly infected in 2020 (Joint United Nations Programme on HIV/AIDS (UNAIDS), Data Book of 2020, https://www.unaids.org/sites/default/files/media_asset/2020_ aids-data-book_en.pdf). The development of effective antiretroviral therapies (ART) represented a turning point in the control of HIV transmission, as well as in improving the quality of life of people living with HIV. However, only 27.5 million people could access ART in 2020, meaning that more than 10 million people remained untreated and in a potential risk of suffering from other infections, as well as acting as vectors of HIV transmission.

[0004] With the current treatment protocols, the use of combined ART is indicated upon HIV diagnosis. This raises a huge challenge not only for health systems, but also for PLWH, as ART is a lifetime of daily medication that may extend in some cases beyond 60 years. This has several implications including, but not limited to 1) the emergence of long-term adverse effects of medications; 2) adherence fatigue resulting in development of antiretroviral resistance, risk to infect sexual partners due to an inadequately suppressed virus, associated poor health outcomes and increased resource utilization, among others; 3) costs related to routine clinical monitoring and investigations; and 4) challenging high costs to patients or health systems associated with long-term chronic medication supply (prescription fees and patient out-of-pocket costs, wholesalers and pharmacy margins, etc).

[0005] In this context, prevention of HIV infection as well as a therapeutic treatment for HIV has become a challenging major unmet need for society.

[0006] Many strategies to prevent and cure HIV have already been assessed. These include the use of chemical HIV replication inhibitors, latency reversal agents and cellular immunity therapies elicited by vaccines, soluble native-like HIV Env trimers (SOSIPs) and/or broadly neutralizing antibodies (bNAbs). Yet none of them have per se succeeded in the complete control and eradication of HIV from infected people. Thus, it is most likely that a combination of different approaches might be the preferred scheme leading to a final effective therapy to cure HIV.

[0007] T-cell vaccines are an essential component of a cure-eradication strategy. In addition to the huge efficient development of ART and despite decades of effort, prophylaxis of HIV- 1 transmission has been less successful as the development of an efficient HIV-1 vaccine has not been achieved yet. The high protein sequence variability among HIV strains combined with effective features of the HIV-1 proteins that decrease the neutralizing potency of antibodies are effective tools to evade the immune response. Thus, the development of potent immunogens able to effectively induce antiviral-specific T-cell responses is crucial.

[0008] The HTI immunogen is a fusion protein constructed from 16 segments of HIV-1, each between 11 and 78 amino acids in length, and encoding critical epitopes of the viral proteins Gag (45%), Pol (44%), Vif (8%) and Nef (3%). (Mothe, B. et al., J. Transl. Med. 13:6Q (2015); US Pat. No. 9,988,425 (herein incorporated by reference in its entirety)). These epitopes were identified as viral targets associated with relative HIV-1 viral control in a comprehensive screening of large cohorts of clade B and C HIV- 1 -infected participants. This vaccine design differs from previous designs based on full protein sequences, very short and conserved segments of the virus, conserved CD8 T-cell epitopes or other studies not based on relevant clinical data. The design of the HTI immunogen is disclosed in Mothe, B. et al., J. Transl. Med. 13:60 (2015) and U.S. Pat. No. 9,988,425) (both of which are incorporated herein by reference in their entirety) and efficacy of the HTI immunogen to control HIV replication after cART interruption has been demonstrated (Bailon, L. et al. A placebo-controlled ATI trial of HTI vaccines in early treated HIV infection. Virtual Conference on Retroviruses and Opportunistic Infections, abstract 161LB, 2021.).

[0009] The “HIVACAT T-cell immunogen” (HTI) was designed to cover T-cell targets, against which T-cell responses are predominantly observed in HIV- 1 -infected individuals with low HIV-1 viral loads. This immunogen has been manufactured as naked DNA, as well as introduced into viral vaccine vectors. The HTI plasmid DNA contains the expression-optimized HTI open reading frame inserted into a pCMVkan vector comprising a plasmid backbone optimized for growth in bacteria, the human cytomegalovirus (CMV) promoter without any introns, the optimized HTI gene, the bovine growth hormone (BGH) polyadenylation side, and the kanamycin resistance gene. The HTI gene contains the human GM-CSF signal peptide at the N terminus as a means to enhance translocation into the Endoplasmic Reticulum. A FLAG-tag was added at the C-terminus for expression analysis in transfected HEK293 cells. Mothe, B. et al., J. Trans. Med. 13:60 (2015) (hereby incorporated by reference in its entirety)

[0010] A recombinant MVA expressing the HTI gene has been generated. (Letourneau, S. etal., PLoS One 2e984 (2007); Nicola, JP et al., Gene Ther. 77: 1068-1080 (2004) (both of which are hereby incorporated by reference in their entirety). Briefly, chicken embryo fibroblast (CEF) cells grown in Dulbecco’s Modified Eagle’s Medium supplemented with 10% FBS, penicillin/streptomycin and glutamine (DMEM 10) were infected with parental MVA and transfected using Superfectine (Qiagen) with DNA.HTI. These cells were MoFlo single cell sorted into 96-well plates and these were used to culture recombinant virus upon addition of fresh CEF. Those wells containing suitably infected cells were harvested and screened by PCR to confirm identity and test purity. Plaque picking was performed using CEF until the culture was free of parental virus, as determined by PCR, after which a master virus stock was grown, purified on a 36% sucrose cushion, tittered and stored at -80C until use.

[0011] The chimpanzee adenovirus vector ChAdOxl is derived from ChAdV isolate Y25 of group E adenoviruses and preexisting antibodies to group E are rare in human populations. Its genome modifications include removal of the El, E3, and a substitution of simian region E4 with the human adenovirus 5 E4 orf4 and orf6/7 genes. For the generation of recombinant ChAdOxls, the tHIVconsv5 and tHIVconsv6 genes were subcloned under the control of the human cytomegalovirus immediate early promoter into plasmid pENTR4_Mono and inserted at the El locus of the ChAdOxl genome by GalK recombineering. Recombinant ChAdOxl vaccines were rescued by transfection of HEK293A T-Rex cells using linearized plasmid and grown in suspension culture of HEK 293 cells. The presence of the transgene and absence of contaminating empty parental adenovirus was confirmed by PCR. The virus was titred to obtain infectious units per milliliter, assayed by spectrophotometry to quantify the number of virus particles per milliliter and stored at -80 °C until use. (Ondondo, B. et al., Mol. Ther. 24(4/832-842 (2016) (hereby incorporated by reference in its entirety). A chimpanzee virus containing a sequence encoding the HTI immunogen, ChAdOxl. HTI, has been developed. (Bailon, L. et al., “A placebo-controlled ATI trial of HTI vaccines in early treated HIV infections”, Virtual Conference on Retroviruses and Opportunistic Infections, Abstract 161LB (2021); Bailon, L. et al., Nat. Med. (2022) (in press)).

[0012] An immunization regimen consisting of three injections of DNA encoding HTI followed by a boost with modified vaccinia Ankara (MVA)-vectored HTI has been shown in C57BL/6 mice to result in the induction of HIV- 1 specific T-cell responses to most antigen regions included in its design. High magnitudes of HIV-1 specific T-cells were also induced in macaques following three injections of DNA.HTI and two injections of MVA.HTI. It is well known that delivering a boosting injection of an immunogen using a viral vector such as MVA can boost the response of DNA immunization and increase the magnitude of responses. Such heterologous prime-boost regimens have been used for a number of immunogens in the context of HIV.

[0013] A randomized, single-center, double-blind and placebo-controlled trial to evaluate the safety, tolerability, immunogenicity and antiviral effect of a therapeutic HTI vaccine - after discontinuation of ART in 45 early-treated people living with HIV has been conducted. The vaccine used in the study included a DNA vector (DNA.HTI “D”), a Modified Vaccinia Ankara virus vector (MVA.HTI “M”) and a chimpanzee adenoviral vaccine vector (ChAdOxl .HTI “C”). Participants were randomized (2: 1) to receive heterologous prime-boost vaccination regimens consisting of DDDMM followed byh CCM, or matched placebo (P), followed by a 24-week ATI. Immunizations were well- tolerated with neither unexpected nor serious AEs. (See NCT03204617 at https://clinicaltrials.gov/ct2/show/NCT03204617).

[0014] Of the few proteins that compose the HIV genome, the highly glycosylated envelope protein (Env) is the only HIV protein on the surface of infected cells and virions, becoming the sole target of host antibodies that mediate virus neutralization, antibody-dependent cellular toxicity, or cellular neutralizing responses against infected cells. The mature Env protein is a homotrimer consisting of three non-covalently associated gpl20-gp41 heterodimers configuring a functional spike on the surface of the virion. This complex acts as an entry mechanism on which HIV-1 completely depends to enter cells. Antibody binding to the Env protein can lead to HIV-1 neutralization. However, host cellular immunity fails in creating neutralizing antibodies due to the highly glycosylated status of the Env protein, the high variability among different HIV strains and the different conformational changes that hide gpl20 antibody epitopes. In this context, the development of a combined therapy to address these issues would increase the prospects of obtaining an optimal anti-HIV therapy.

[0015] SOSIPs are stabilized HIV-1 envelope glycoproteins (Env) that resemble the native Env and are used in vaccination strategies aimed at inducing broadly neutralizing antibodies (bNAbs) . The SOSIP design of trimers is now a widely used immunogen design platform. SOSIP trimers adopt a native-like conformation, in that they resemble the form of trimer that is found on infectious HIV-1 virions and present multiple epitopes for broadly neutralizing antibodies (bNAbs). The latter property is relevant to the longterm goal of inducing this type of antibody via a suitably designed immunization regimen. In practice, SOSIP trimers, like all other designs of HIV-1 Env protein, require delivery as an admixture with an adjuvant to boost their immunogenicity (Fischer B, et al, PLoS One (2020); 15(5): e0233577) (hereby incorporated by reference in their entirety) These soluble trimers can induce the production of neutralizing antibodies against HIV-1 which, in turn, block the entrance of the viral particle to the cell preventing infection, as well as help to eliminate HIV-1 viral particles from blood, preventing transmission. Soluble gpl40 (sgpl40) SOSIP.664 Env trimers are stabilized by a disulfide bond linking gpl20 and gp41 (SOS) and an I559P (IP) change in gp41, which also bears a truncation of the gp41 ectodomain at residue 664 (Sanders, R, et al, PLOS Pathogens (2013); https://doi.org/10.1371/journal.ppat.1003618) (hereby incorporated by reference in their entirety). The first ever designed SOSIP, BG505 SOSIP.664, was described as consistently inducing neutralizing antibodies against neutralization-resistant viruses in animals. However, due to the high variability of the Env sequences, it is believed that the use of consensus sequences to design SOSIPs might benefit the acquisition of broad neutralization breadth against specific strains. Thus, the native-like Env trimer ConM SOSIP.v7 gpl40 was developed upon the BG505 SOSIP.664 prototype, but based on a consensus sequence of all HIV-1 envelope sequences in group M. (Sliepen, K. et al., Nat. Comm. 70:2355-2371 (2019) (hereby incorporated by reference in its entirety. )Two ongoing clinical trials are testing the safety and immunogenicity of the ConM SOSIP. v7 gpl40 vaccine, given alone or in combination with other envelope immunogens in HIVnegative populations, using native-like Env trimer ConM SOSIP. v7 gpl40 adjuvanted with monophosphoryl lipid A (MPLA) liposome (Sliepen et al. Nat. Comm. 10: 2355- 2371, 2019). However, there remains a need for a combination therapy that targets an envelope immunogen and other HIV proteins, such as gag, pol, vif, and nef.

SUMMARY OF THE INVENTION

[0016] In one embodiment, the present disclosure provides a method of treating or preventing an human immunodeficiency virus (HIV) infection in a human in need thereof suffering from HIV infection or at risk of developing HIV infection by administering to said human the HTI immunogen in combination with at least one antibody inducing stable soluble HIV-1 enveloped glycoprotein trimer mimic in an amount sufficient to induce an immune response against HIV in said human. The following detailed description is given by way of example, and is not intended to limit the invention solely to the specific embodiments described.

[0017] In some aspects, the present disclosure provides a method of treating or preventing an HIV infection in a human in need thereof suffering from HIV infection or at risk of developing HIV infection, the method comprising administering to the human a therapeutically effective amount of the HTI immunogen and a HIV-1 envelope glycoprotein trimer mimic (SOSIP).

[0018] In some aspects, the HTI immunogen is administered as plasmid DNA (DNA.HTI or D), In some aspects, the HTI immunogen is administered as a modified vaccinia virus Ankara (MV A) vector (MVA.HTI or M). In some aspects, the HTI immunogen is administered as a Chimpanzee adenovirus vector (C).

[0019] In some aspects, the HTI immunogen is administered in a regimen that comprises three DNA.HTI administrations followed by two HTLMVA administrations in the sequence DDD-MM. In some aspects, the HTI immunogen administration regimen comprises one chimpanzee adenovirus. HTI administration followed by one MVA.HTI administration in the sequence CM. In some aspects, the HTI immunogen administration regimen comprises two chimpanzee adenovirus. HTI administrations followed by two MVA.HTI administrations (CCMM). In some aspects, the HTI immunogen administration regimen comprises two chimpanzee adenovirus. HTI administrations followed by one MVA.HTI administration (CCM). In some aspects, the HTI immunogen administration regimen comprises three DNA.HTI administrations followed by two MVA.HTI administrations followed by two chimpanzee adenovirus. HTI administrations followed by one MVA.HTI administration (DDD-MM-CCM).

[0020] In another embodiment, the present disclosure provides a method comprising (a) administering to the subject 1 to 5 administrations of an immunogenically effective amount of a first suitable vector encoding an immunogenic polypeptide; and (b) administering to the subject 1 to 5 administrations of an immunogenically effective amount of a second suitable vector encoding the immunogenic polypeptide; wherein the immunogenic polypeptide comprises:

[0021] a sequence having at least 90% sequence identity to SEQ ID NO: 1;

[0022] a sequence having at least 90% sequence identity to SEQ ID NO: 2;

[0023] a sequence having at least 90% sequence identity to SEQ ID NO: 3;

[0024] a sequence having at least 90% sequence identity to SEQ ID NO: 4;

[0025] a sequence having at least 90% sequence identity to SEQ ID NO: 5;

[0026] a sequence having at least 90% sequence identity to SEQ ID NO: 6;

[0027] a sequence having at least 90% sequence identity to SEQ ID NO: 7;

[0028] a sequence having at least 90% sequence identity to SEQ ID NO: 8;

[0029] a sequence having at least 90% sequence identity to SEQ ID NO: 9;

[0030] a sequence having at least 90% sequence identity to SEQ ID NO: 10;

[0031] a sequence having at least 90% sequence identity to SEQ ID NO: 11;

[0032] a sequence having at least 90% sequence identity to SEQ ID NO: 12; [0033] a sequence having at least 90% sequence identity to SEQ ID NO: 13;

[0034] a sequence having at least 90% sequence identity to SEQ ID NO: 14;

[0035] a sequence having at least 90% sequence identity to SEQ ID NO: 15; and

[0036] a sequence having at least 90% sequence identity to SEQ ID NO: 16;

[0037] wherein at least two of (a) to (p) are joined by a single, dual, or triple alanine amino acid linker, wherein the linker results in the formation of an AAA sequence in the junction region between adjoining sequences, and wherein the sequence of each of (a) to (p) is 11-85 amino acids in length.

[0038] In another embodiment, the present disclosure further provides a method comprising administration of the HTI immunogen in combination with at least one stable soluble HIV-1 envelope glycoprotein trimer mimic, such as ConM SOSIP.v7 gpl40 trimer. In some embodiments, the HTI vaccine and ConM SOSIP.v7 gpl40 trimer are administered in a single dose. In some embodiments, the HTI vaccine and ConM SOSIP.v7 gpl40 trimer are administered in multiple doses. In some embodiments, the HTI vaccine and ConM SOSIP.v7 gpl40 trimer are prepared, purified and formulated for immunization in a human. In some embodiments, the HTI vaccine and ConM SOSIP.v7 gpl40 trimer are formulated in one composition. In some embodiments, the HTI vaccine and ConM SOSIP.v7 gpl40 trimer are formulated in separate compositions. In another particularly advantageous embodiment, the trimer protein is formulated for immunization in a human to contain an adjuvant. A number of adjuvants are well known to those investigating vaccines but could include TLR agonist, in particular TLR4 agonists, in particular MPLA, but are not limited to those containing MPLA.

[0039] In some aspects, the present disclosure further provides a method comprising administering the HTI immunogen in combination with at least one stable soluble HIV-1 envelope glycoprotein trimer and at least one broadly neutralizing antibody against HIV (i.e., monoclonal antibodies that are able to neutralize more than 90% of circulating HIV strains). In some aspects, the at least one broad neutralizing antibody is specific for an HIV envelope protein. In some aspects, the at least one broad neutralizing antibody is specific for a HIV envelope trimer. In some aspects, the at least one broadly neutralizing antibody binds gpl20 and gp41 envelope glycoproteins. In some aspects, the gpl20 and gp41 targeting antibody is an antibody against the CD4 binding site. In some aspects, the broad neutralizing antibody is the long-lasting variant VRC07-523-LS, that targets the HIV CD4 binding site of gpl20 (see, e.g., Rudicellet al. J. Virology 88: 12669-, 2014; International Appl. No. PCT/US2012/068827, incorporated herein by reference in their entirety).

[0040] A method comprising administering to a human the HTI immunogen , and at least one stable soluble HIV-1 envelope glycoprotein trimer mimic, in particular ConM SOSIP.v7 gpl40 trimer and at least one broad neutralizing antibody as described herein is further provided as in the example.

BRIEF DESCRIPTION OF THE FIGURES

[0041] FIG. 1 : Depicts treatment protocol of a HIV vaccine comprising HTI immunogen in conjunction with SOSIPs. “C” is ChAdOx.1 HTI; “S” is ConM SOSIP.v7gpl40; and “M” is MVA.HTI. ATI is analytical treatment interruption. The regimen is CSSMS at the intervals noted.

DETAILED DESCRIPTION

I. GENERAL

[0042] The present disclosure provides methods, compositions, and kits for the treatment or prevention of an HIV infection in a human having or at risk of having HIV, comprising the combination of the HTI immunogen and at least one stable soluble HIV-1 envelope glycoprotein trimer mimic, such as ConM SOSIP.v7 gpl40 trimer. In some aspects, the HTI immunogen and ConM SOSIP.v7 gpl40 trimer are co-adjuvated with a TLR4 agonist. In some aspects, the TLR4 agonist is monophosphoryl lipid A (MPLA). In some aspects, the HIV-HTI vaccine and the ConM SOSIP.v7 gpl40 trimer are combined with at least one broad neutralizing antibody. In some aspects, the at least one broadly neutralizing antibody binds gpl20 and gp41 envelope glycoproteins.

II. DEFINITIONS

[0043] As used herein, “HTI immunogen” is meant an immunogenic fusion protein constructed from 16 segments of HIV- 1, each between 11 and 78 amino acids in length, and encoding critical epitopes of the viral proteins Gag (45%), Pol (44%), Vif (8%) and Nef (3%) as described in Mothe, B. et al., J. Transl. Med. 13:6Q (2015) and US Pat. No. 9,988,425 (each of which is incorporated herein by reference in its entirety).

[0044] As used herein, “SOSIP” means an antibody-inducing, stable, soluble HIV-1 envelope glycoprotein trimer mimic. For example the SOSIP can be Con M, which is based on a consensus of the consensus sequence of each clade in group M. In some aspects, the ConM SOSIP is ConM SOSIP. v7ConM SOSIP. v7 gpl40 trimer. See Sliepen, K. et al., Nat. Comm. 70:2355-2371 (2019) (hereby incorporated by reference in its entirety.)

[0045] As used herein, “ChAdOxl.HTI” refers to chimpanzee adenovirus vector containing a sequence encoding the HTI immunogen.

[0046] As used herein, “MVA.HTI” refers to a MVA vector containing a sequence encoding the HTI immunogen.

[0047] As used herein, "administering" is meant a method of giving a dosage of a pharmaceutical composition (e.g., a composition of the invention, such as any one of the vaccines of the first or fourth aspects, the compositions of the third aspect, the nucleic acid molecules of the fifth aspect, and/or the vectors of the sixth aspect) to a subject. The compositions utilized in the methods described herein can be administered, for example, intramuscularly, intravenously, intradermally, percutaneously, intraarterially, intraperitoneally, intralesionally, intracranially, intraarticularly, intraprostatically, intrapleurally, intratracheally, intranasally, intravitreally, intravaginally, intrarectally, topically, intratumorally, peritoneally, subcutaneously, subconjunctival, intravesicularlly, mucosally, intrapericardially, intraumbilically, intraocularly, orally, topically, locally, by inhalation, by injection, by infusion, by continuous infusion, by localized perfusion bathing target cells directly, by catheter, by lavage, by gavage, in cremes, or in lipid compositions. The preferred method of administration can vary depending on various factors (e.g., the components of the composition being administered and the seventy of the condition being treated).

[0048] As used herein, the term " reducing" with respect to HIV refers to a reduction or decrease of an HIV-mediated activity (e.g., infection, fusion {e.g., target cell entry and/or syncytia formation), viral spread, etc.) and/or a decrease in viral titer. HIV-mediated activity and/or HIV titer may be decreased by 5%, 0%, 15%, 20%, 25%. 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 81% , 82%. 83%. 84%, 85%, 86%, 87%, 88%, 89%, 90%, 9 1%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99. 1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, 99.9% or more compared to that of a control subject (e.g., an untreated subject or a subject treated with a placebo).

[0049] The term "vaccine," as used herein, is defined as material used to provoke an immune response (e.g., the production of neutralizing anti-HIV antisera). Administration of the vaccine to a subject may confer at least partial immunity against HIV infection. In one aspect, the vaccine comprises the HTI immunogen.

[0050] The term "virus," as used herein, is defined as an infectious agent that is unable to grow or reproduce outside a host cell and that infects mammals (e.g., humans) or birds. HIV is a representative virus.

[0051] By "adenovirus" is meant a medium-sized (90-1 00 nm), non-enveloped icosahedral virus that includes a capsid and a double-stranded linear DNA genome. The adenovirus can be a naturally occurring, but isolated, adenovirus (e.g., sAd4287, sAd431 0A, or sAd4312) or a recombinant adenovirus (e.g., replication-defective or replication competent sAc>4287, sAd431 0A, or sAd431 2 or a chimeric variant thereof).

[0052] By "human immunodeficiency virus" or "HIV" is meant a virus of the genus Lentivirinae, part of the family of Retroviridae, and includes, but is not limited to, HIV type 1 (HIV-1) and HIV type 2 (HIV-2), two species of HIV that infect humans.

[0053] By "immune response" is meant any response to an antigen or antigenic determinant by the immune system of a subject (e.g., a human). Exemplary immune responses include humoral immune responses (e.g., production of antigen-specific antibodies e.g., neutralizing antibodies (NAbs)) and cellmediated immune responses (e.g, lymphocyte proliferation).

[0054] By “bNAb” is meant a broad neutralizing antibody that is an antibody that neutralizes more than one HIV strain.

[0055] Throughout this specification and claims, the word "comprise," or variations such as "comprises" or "comprising," will be understood to imply the inclusion of a stated integer or group of integers but not the exclusion of any other integer or group of integers.

[0056] The functional equivalent or fragment of the functional equivalent, in the context of a protein, may have one or more conservative amino acid substitutions. The term “conservative amino acid substitution” refers to substitution of an amino acid for another amino acid that has similar properties as the original amino acid. The groups of conservative amino acids are as follows:

Group Name of the amino acids

Aliphatic Gly, Ala, Vai, Leu, He

Hydroxyl or Sulfhydryl/Selenium-containing Ser, Cys, Thr, Met

Cyclic Pro

Aromatic Phe, Tyr, Trp

Basic His, Lys, Arg

Acidic and their Amide Asp, Glu, Asn, Gin

[0057] Conservative substitutions may be introduced in any position of a predetermined peptide or fragment thereof. It may however also be desirable to introduce nonconservative substitutions, particularly, but not limited to, a non-conservative substitution in any one or more positions. A non-conservative substitution leading to the formation of a functionally equivalent fragment of the peptide would for example differ substantially in polarity, in electric charge, and/or in steric bulk while maintaining the functionality of the derivative or variant fragment.

[0058] By "sequence identity" or ' sequence similarity" is meant that the identity or similarity between two or more amino acid sequences, or two or more nucleotide sequences, s expressed in terms of the identity or similarity between the sequences. Sequence identity can be measured in terms of ' percentage (%) identity," wherein the higher the percentage, the more identity shared between the sequences. Sequence similarity can be measured In terms of percentage similarity (which takes into account conservative amino acid substitutions); the nigher the percentage, the more similarity shared between the sequences. Homologs or orthologs of nucleic acid or amino acid sequences possess a relatively high degree of sequence identity/ similarity when aligned using standard methods. Sequence identity may be measured using sequence analysis software on the default setting (e.g., Sequence Analysis Software Package of the Genetics Computer Group, University of Wisconsin Biotechnology Center, 17 10 University Avenue, Madison, W1 53705). Such software may match similar sequences by assigning degrees of homology to various substitutions, deletions, and other modifications.

[0059] “Percentage of sequence identity” is determined by comparing two optimally aligned sequences over a comparison window, wherein the portion of the polynucleotide or polypeptide sequence in the comparison window may have additions or deletions (i.e., gaps) as compared to the reference sequence (which does not have additions or deletions) for optimal alignment of the two sequences. In some cases, the percentage can be calculated by determining the number of positions at which the identical nucleic acid base or amino acid residue occurs in both sequences to yield the number of matched positions, dividing the number of matched positions by the total number of positions in the window of comparison and multiplying the result by 100 to yield the percentage of sequence identity.

[0060] “Identical” or percent “identity” in the context of two or more nucleic acid or polypeptide sequences, refer to two or more sequences or subsequences that are the same or have a specified percentage of amino acid residues or nucleotides that are the same (e.g., 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 98%, or 99% identity over a specified region, e.g., the entire polypeptide sequences or individual domains of the polypeptides), when compared and aligned for maximum correspondence over a comparison window or designated region as measured using one of the following sequence comparison algorithms or by manual alignment and visual inspection. Such sequences are then said to be “substantially identical.” This definition also refers to the complement of a test sequence.

[0061] "Nucleic acid" or "polynucleotide," as used interchangeably herein, refer to polymers of nucleotides of any length, and include DNA and RNA. The nucleotides can be deoxyribonucleotides, ribonucleotides, modified nucleotides or bases, and/or their analogs, or any substrate that can be incorporated into a polymer by DNA or RNA polymerase, or by a synthetic reaction. A polynucleotide may comprise modified nucleotides, such as methylated nucleotides and their analogs. If present, modification to the nucleotide structure may be imparted before or after assembly of the polymer. The sequence of nucleotides may be interrupted by non-nucleotide components. A polynucleotide may be further modified after synthesis, such as by conjugation with a label. Other types of modifications include, for example, "caps," substitution of one or more of the naturally occurring nucleotides with an analog, internucleotide modifications such as, for example, those with uncharged linkages (e.g., methyl phosphonates, phosphotriesters, phosphoamidates, carbamates, etc.) and with charged linkages (e.g. , phosphorothioates, phosphorodithioates, etc.), those containing pendant moieties, such as, for example, proteins (e.g. , nucleases, toxins, antibodies, signal peptides, poly-L-lysine, etc.), those with intercalators (e.g. acridine, psoralen, etc.), those containing chelators (e.g., meiais, radioactive metals boron, oxidative metals, etc.), those containing alkylators, those with modified linkages (e.g., alpha anomeric nucleic acids etc.), as well as unmodified forms of the polynucleotide(s). Further, any of the hydroxyl groups ordinarily present in the sugars may be replaced, for example, by phosphonate groups, phosphate groups protected by standard protecting groups, or activated to prepare additional linkages to additional nucleotides, or may be conjugated to solid or semi-solid supports. The 5 ' and 3' terminal OH can be phosphorylated or substituted with amines or organic capping group moieties of from 1 to 20 carbon atoms. Other hydroxyls may also be derivatized to standard protecting groups. Polynucleotides can also contain analogous forms of ribose or deoxyribose sugars that are generally known in the art, including, for example, 2'-0-methyl-, 2'-0-allyl, 2'-fluoro- or 2'-azido-ribose, carbocyclic sugar analogs, alpha-anomeric sugars, epimeric sugars such as arabinose, xyloses or lyxoses, pyranose sugars, furanose sugars, sedoheptuloses, acyclic analogs and a basic nucleoside analogs such as methyl riboside. One or more phosphodiester linkages may be replaced by alternative linking groups. These alternative linking groups include, but are not limited to, embodiments wherein phosphate is replaced by P(0)S("thioate"), P(S)S ("dithioate"), "(0)NR 2 ("amidate"), P(0)R, P(0)OR', CO or CH 2 ("formacetal"), in which each R or R' is independently H or substituted or unsubstituted alkyl ( 1-20 C) optionally containing an ether (-0-) linkage, aryl, alkenyl, cycloalkyl, cycloalkenyl or araldyl. Not all linkages in a polynucleotide need be identical. The preceding description applies to all polynucleotides referred to herein, including RNA, such as mRNA, saRNI, circular RNA, siRNA, and DNA.

[0062] A DNA sequence that “encodes” a particular type of RNA is a DNA nucleic acid sequence that can be transcribed into any type of RNA. A DNA polynucleotide may encode an RNA (mRNA) that is translated into protein, or a DNA polynucleotide may encode an RNA that is not translated into protein (e.g., tRNA, rRNA, saRNA, cRNA, gRNA or others; also referred to herein as “non-coding” RNA or “ncRNA”). A “protein coding sequence or a sequence that encodes a particular protein or polypeptide, is a nucleic acid sequence that is transcribed into mRNA (in the case of DNA) and is translated (in the case of mRNA) into a polypeptide in vitro or in vivo when placed under the control of appropriate regulatory sequences.

[0063] By "recombinant," with respect to a composition (e.g., a vector of the invention, such as an adenovirus or poxvirus vector), is meant a composition that has been manipulated in vitro (e.g., using standard cloning techniques) to introduce changes (e.g., changes to the composition, e.g. , adenovirus or poxvirus genome of an adenovirus or poxvirus vector, respectively) that enable binding to or containment of a therapeutic agent and/or that promote the introduction of a therapeutic agent into a subject (e.g., a human) or a host cell. The recombinant composition of the invention may therefore be an adenoviral or poxviral transport vector (e.g., a replication-defective adenoviral or poxviral vector) for delivery of one or more of the stabilized Env polypeptide trimers of the invention.

[0064] “Vector,” “expression vector,” or “construct” is a nucleic acid used to introduce heterologous nucleic acids into a cell that has regulatory elements to provide expression of the heterologous nucleic acids in the cell. Vectors include but are not limited to plasmid, minicircles, yeast, and viral genomes. In some embodiments, the vectors are plasmid, minicircles, yeast, or viral genomes. In some embodiments, the vector is a viral vector. As used herein, the term "vector" is meant to include, but is not limited to, a virus (e.g., adenovirus or poxvirus), naked DNA, RNA oligonucleotide, cationic lipid (e.g., liposome), cationic polymer (e.g., polysome), virosome, nanoparticle, or dentrimer. By "adenovirus vector" is meant a composition that includes one or more genes (non- structural or structural), or fragments thereof, from an adenoviral species (e.g. , adenovirus serotype 11 (Adi 1), adenovirus serotype 15 (Adi 5), adenovirus serotype 24 (Ad24), adenovirus serotype 26 (Ad26), adenovirus serotype 34 (Ad34), adenovirus serotype 35 (Ad35), adenovirus serotype 48 (Ad48), adenovirus serotype 49 (Ad49), adenovirus serotype 50 (Ad50), Pan9 (AdC68), or a chimeric variant thereof (e.g. , adenovirus serotype 5 HVR48 (Ad5HVR48))) that may be used to transmit one or more heterologous genes (e.g. , one or more of the optimized gpl40 polypeptides of the invention) from a viral or non-viral source to a subject or a host. The nucleic acid material of the viral vector may be encapsulated, e.g., in a lipid membrane or by structural proteins (e.g., capsid proteins), that may include one or more viral polypeptides (e.g., an envelope glycoprotein). The virai vector can be used to infect cells of a subject, which, in turn, promotes the translation of the heterologous gene(s) of the viral vector into a protein product (e.g., one or more of the gp 0 Env polypeptides described herein, such that a stabilized trimer of the invention is formed).

[0065] Treatment” or “treat” or “treating” as used herein refers to an approach for obtaining beneficial or desired results. For purposes of the present disclosure, beneficial or desired results include, but are not limited to, alleviation of a symptom and/or dimini shment of the extent of a symptom and/or preventing a worsening of a symptom associated with a disease or condition. In one embodiment, “treatment” or “treating” includes one or more of the following: a) inhibiting the disease or condition (e.g., decreasing one or more symptoms resulting from the disease or condition, and/or diminishing the extent of the disease or condition); b) slowing or arresting the development of one or more symptoms associated with the disease or condition (e.g., stabilizing the disease or condition, delaying the worsening or progression of the disease or condition); and c) relieving the disease or condition, e.g., causing the regression of clinical symptoms, ameliorating the disease state, delaying the progression of the disease, increasing the quality of life, and/or prolonging survival.

[0066] “Therapeutically effective amount" or “effective amount” as used herein refers to an amount that is effective to elicit the desired biological or medical response, including the amount of an agent, or an HIV vaccine, that, when administered to a patient for treating a disease, is sufficient to effect such treatment for the disease. The effective amount will vary depending on the agent, the disease, and its severity and the age, weight, etc., of the patient to be treated. The effective amount can include a range of amounts. As is understood in the art, an effective amount may be in one or more doses, i.e., a single dose or multiple doses may be required to achieve the desired treatment endpoint. An effective amount may be considered in the context of administering one or more therapeutic agents, and a single agent may be considered to be given in an effective amount if, in conjunction with one or more other agents, a desirable or beneficial result may be or is achieved. Suitable doses of any co-administered agents may optionally be lowered due to the combined action (e.g., additive or synergistic effects) of the agents.

[0067] “Delaying” as used herein refers to development of a disease or condition means to defer, hinder, slow, retard, stabilize and/or postpone development of the disease or condition. This delay can be of varying lengths of time, depending on the history of the disease and/or individual being treated. As is evident to one skilled in the art, a sufficient or significant delay can, in effect, encompass prevention, in that the individual does not develop the disease or condition.

[0068] “Prevent” or “prevention” or “preventing” as used herein refers to a regimen that protects against the onset of the disease or disorder such that the clinical symptoms of the disease do not develop. Thus, “prevention” relates to administration of a therapy (e.g., administration of a therapeutic substance) to a patient before signs of the disease are detectable in the patient (e.g., administration of a therapeutic substance to a patient in the absence of detectable infectious agent (e.g., virus) in the patient). The patient may be an individual at risk of developing the disease or disorder, such as an individual who has one or more risk factors known to be associated with development or onset of the disease or disorder. Thus, in certain embodiments, the term “preventing HIV infection” refers to administering an anti-HIV therapeutic substance to a patient who does not have a detectable HIV infection. It is understood that the patient for anti-HIV preventative therapy may be an individual at risk of contracting the HIV virus. It is also understood that prevention does not require a 100% success rate. In some instances, prevention may be understood as a reduction of the risk of infection, but not a complete elimination in the occurrence of an infection.

[0069] “At risk human” as used herein refers to a person who is at risk of developing a condition to be treated. A person “at risk” may or may not have detectable disease or condition, and may or may not have displayed detectable disease prior to the treatment of methods described herein. “At risk” denotes that a person has one or more risk factors, which are measurable parameters that correlate with development of a disease or condition and are known in the art. A person having one or more of these risk factors has a higher probability of developing the disease or condition than a person without these risk factor(s).

[0070] “Viral infection” describes a diseased state in which a virus invades healthy cells, uses the cell's reproductive machinery to multiply or replicate and ultimately lyse the cell resulting in cell death, release of viral particles and the infection of other cells by the newly produced progeny viruses. Latent infection by certain viruses, e.g., HIV, is also a possible result of viral infection. [0071] “ART” as used herein refers to anti-retroviral therapy. Generally, the term refers to combinations of anti-retroviral medications used to treat human viral infections, including HIV infections. Combinations and regimens can include multiple, often three or more, drugs such as nucleoside reverse transcriptase inhibitors (NRTIs), non-nucleoside reverse transcriptase inhibitors (NNRTIs), protease inhibitors (Pls), fusion inhibitors, CCR5 agonists, and/or integrase inhibitors.

[0072] “Viral load” and “HIV viral load” refer to the level of HIV detectable in the blood of an HIV infected human. It can be calculated by estimating the amount of virus in an involved bodily fluid. For example, it can be given in HIV RNA copies per milliliter of blood or blood plasma. An “undetectable” HIV viral load comprises a condition in which HIV RNA copies are not routinely detected by standard viral load tests, e.g., less than 50 copies HIV RNA per milliliter of blood or blood plasma.

[0073] “Viremia” refers to the measurable presence of virus or viral particles in circulation in a virally infected human. Transient viremia refers to a brief, transitory, or temporary increase in the measurable presence of virus or viral particles in circulation in a virally infected human. An example of transient HIV viremia includes a period in which the HIV-1 RNA level in the blood or plasma of an HIV infected human which has been maintained for a period of time at a concentration of less than 50 copies of HIV-1 RNA per mL briefly, transitorily, or temporarily rises to a concentration of greater than 50 copies/mL, such as from 50 to 2,000 copies/mL.

[0074] As used herein, the term "envelope glycoprotein" refers, but is not limited to, the glycoprotein that is expressed on the surface of the envelope of HIV virions and the surface of the plasma membrane of HIV infected cells. The e gene encodes gpl 60, which is proteolytically cleaved into the gpl20 and gp41 Envelope (Env) proteins. Gpl20 binds to the CD4 receptor on a target cell that has such a receptor, such as, e.g., a T-helper cell. Gp41 is non-covalently bound to gpl20, and provides the second step by which HIV enters the cell. It is originally buried within the viral envelope, but when gpl20 binds to a CD4 receptor, gpl 20 changes its conformation causing gp41 to become exposed, where it can assist in fusion with the host cell.

[0075] As used herein, the term "stabilized polypeptide trimer" refers, but is not limited to, an oligomer that includes a protein and/or polypeptide sequence that increases the stability (e.g., via the presence of one or more oligomerization domains) of the trimeric structure (e.g., reduces dissociation of a trimer into monomeric units). In particular, the stabilized polypeptide trimer is composed of three mosaic Env proteins (e.g., Envl, Env2, and/or Env3), three clade C Env proteins, or a combination of one or more mosaic Env proteins and one or more clade C Env proteins, in which at least one Env protein includes an oligomerization domain. An "oligomerization domain" refers, but is not limited to, a polypeptide sequence that can be used to increase the stability of an oligomeric envelope protein such as, e.g., to increase the stability of a HIV gpl 40 trimer. Oligomerization domains can be used to increase the stability of homooligomeric polypeptides as well as heterooligomeric polypeptides. Oligomerization domains are well known in the art, and include "trimerization domains." A trimerization domain refers to an oligomerization domain that stabilizes trimeric polypeptides (e.g., trimers consisting of one or more of the gpl 40 polypeptides of the invention). Examples of trimerization domains include, but are not limited to, the T4- fibritin "foldon" trimerization domain; the coiled-coil trimerization domain derived from GCN4 (Yang e ai. (2002) J. Virol. 76:4634); and the catalytic subunit of E. coli aspartate transcarbamoylase as a trimer tag (Chen et al. (2004) J. Virol. 78:4508).

[0076] By "having substantially the sequence of' with respect to constructs of the invention is meant having at least 99% sequence identity to a recited reference sequence (e.g., having no more than 7 amino acid residue differences, e.g., 1, 2, 3, 4, 5, or 6 amino acid residue differences (e.g., additions, deletions, or conservative amino acid substitutions), relative to a recited reference sequence).

III. VACCINES

[0077] HIV vaccines that specifically target regions on the Gag, Pol, Vif, and Nef proteins of the HIV virus are described herein. Such HIV vaccines can induce an immunological response to one or more HIV proteins and may either protect a human who does not have an HIV infection from contracting the virus or may have a therapeutic effect for persons infected with HIV or who later contract HIV. A vaccine generally comprises a delivery mechanism, e.g., a viral vector, and a package, such as an immunogenic composition or a nucleic acid encoding an immunogenic composition, designed to generate a desired immunological response. In some embodiments, the immunogenic composition comprises an immunogenic polypeptide that is an antigen capable of inducing an adaptive immune response, i.e., a humoral or cell-mediated immune response, when introduced in vivo.

[0078] Any viral vector capable of introducing the desired package into the body to prompt an adaptive response can be used in the presently described methods, compositions, and/or kits. In some embodiments, the viral vector comprises a live vector vaccine, an inactivated vaccine, or a modified envelope vaccine. In some embodiments, the viral vector comprises an Adenoviridae, Poxviridae, Herpesviridae, Adeno-associated virus, cytomegalovirus, carynpox, rubella poliovirus, Venezuelan equine encephalitis virus, lentivirus, or Sendai viral vector. In some embodiments, the viral vector comprises an Adenoviridae or a Poxviridae viral vector. In some embodiments, the viral vector comprises a poxvirus viral vector, e.g., a modified vaccinia virus Ankara (MV A) vector such as the MVA.HTI vector. An exemplary MVA vector is described in Barouch, D. H. et al. Cell 2013, 155(3), 531-539 (incorporated herein by reference in its entirety). In some embodiments, the viral vector comprises an adenovirus viral vector, such as a chimpanzee adenovirus, e.g., a replication-defective chimpanzee adenovirus. Exemplary chimpanzee adenovirus vectors have been described, e.g., in US Patent No. 9,714,435 (incorporated herein by reference in its entirely). In one embodiment, the chimpanzee adenovirus vector is ChAdOxl.HTI.

[0079] International Publication WO 2013/110818 and US Patent No. 9988425 (each of which is herein incorporated by reference in its entirety) describe immunogens for HIV vaccination. Sixteen regions in the Gag, Pol, Vif, and Nef proteins of the HIV-1 virus were relatively conserved and were targeted by HIV patients having a reduced viral load of <5000 copies of HIV-1 RNA per mL. Hancock, G. et al. PLOS Pathogens 2015, 11(2), el004658; Mothe, B. et al. J. Translational Med. 2015, 13, 60. These regions of HIV proteins formed the basis of an immunogen for therapeutic vaccination of HIV. The following table summarizes the regions of HIV-1 targeted by the immunogens:

[0080] The HIV numbering is as described in Korber, B. T. et al. (1998) Numbering positions in HIV relative to HXB2CG. In: Korber, C. K., Foley, B., Hahn, B., McCutchan, F., Mellors, J. and Sodroski, J (eds). Human Retroviruses and AIDS 1998. Theoretical Biology and Biophysics Group, Los Alamos National Laboratory, Los Alamos, NM, pp. Ill- 102-111 (incorporated by reference herein in its entirety).

[0081] In some embodiments, the HIV vaccine comprises a virus comprising an immunogenic polypeptide, or a nucleic acid encoding an immunogenic polypeptide, wherein the immunogenic polypeptide comprises:

[0082] a sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 1;

[0083] a sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 2;

[0084] a sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 3;

[0085] a sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 4; [0086] a sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 5;

[0087] a sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 6;

[0088] a sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 7;

[0089] a sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 8;

[0090] a sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 9;

[0091] a sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 10;

[0092] a sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 11;

[0093] a sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 12;

[0094] a sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 13;

[0095] a sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 14;

[0096] a sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 15; and

[0097] a sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 16;

[0098] wherein at least two of (a) to (p) are joined by a single, dual, or triple alanine amino acid linker, wherein the linker results in the formation of an AAA sequence in the junction region between adjoining sequences, and wherein the sequence of each of (a) to (p) is 11-85, e.g., from 11 to 82, from 11 to 80, or from 11 to 78, amino acids in length. In some embodiments, the immunogenic polypeptide comprises a sequence having amino acid sequences with no more than 1, 2, or 3 substitutions in any one of SEQ ID NOS: 1- 16. In some embodiments, the immunogenic polypeptide comprises a sequence having amino acid sequences according to SEQ ID NOS: 1-16. [0099] In some embodiments, the immunogenic polypeptide comprises an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 17 or 18. In some embodiments, the immunogenic polypeptide comprises an amino acid sequence according to SEQ ID NO: 17 or 18.

[0100] The immunogenic polypeptide can be encoded by any suitable nucleic acid sequence. In some embodiments, the nucleic acid encoding the immunogenic polypeptide comprises a nucleic acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 19 or 20. In some embodiments, the nucleic acid encoding the immunogenic polypeptide comprises a nucleic acid sequence according to SEQ ID NO: 19 or 20.

[0101] In some embodiments, the HIV vaccine comprises a modified vaccinia virus Ankara (MV A) comprising a nucleic acid encoding an immunogenic polypeptide having an amino acid sequence according to SEQ ID NO: 19 or 20. In some embodiments, the HIV vaccine comprises a replication-defective chimpanzee adenovirus comprising a nucleic acid encoding an immunogenic polypeptide having an amino acid sequence according to SEQ ID NO: 19 or 20.

STABLE SOLUBLE HIV-1 ENVELOPE GLYCOPROTEIN TRIMERS (SOSIPs)

[0102] In some aspects, the HIV vaccine comprises a stable soluble HIV envelope glycoprotein trimer (SOSIP). In some aspects, the SOSIP is Con M that is based on a consensus of the consensus sequence of each clade in group M. In some aspects, the ConM SOSIP is ConM SOSIP.v7 (Sliepen, K. etal., Nat. Comm. 10:2355 (2019) (incorporated herein by reference in its entirety). In some embodiments, the ConM SOSIP comprises a TPA signal. In some aspects, the ConM SOSIP comprises a gpl20- gp41Ecro disulfide bond (A501C-T605C). In some aspects, the ConM SOSIP comprises a trimer-stabilizing I559P mutation in gp41Ecro. In some aspects, the ConM SOSIP further comprises trimer-stabilizing mutations H66R and A316W. In some aspects, the ConM SOSIP further comprises improved trimerization mutations 1535M and Q543N. In some aspects, the ConM SOSIP comprises an improved furin cleavage site (R6) comprising an REKR to RRRRRR mutation. In some aspects, the ConM SOSIP comprises a stop codon after gp41Ecro residue 664. In some aspects, the ConM SOSIP comprises an extra intermolecular disulfide bond A73C-A561C in gp41. In some aspects, the ConM SOSIP further comprises mutations E47D, N49E, V65K, E106T, I165L, G429R, K500R and a Q at position 432. In some aspects, the ConM SOSIP construct is codon-optimized. In some aspects, the ConM SOSIP is ConM SOSIP.v7 as disclosed in Sliepen et al., Nature Communications 10: 2355-2371, 2019 (incorporated herein by reference in its entirety). The HIV numbering is as described in Sliepen et al.

METHODS

[0103] In general terms, the present invention is directed to a method of treating or preventing an HIV infection or a disease associated with an HIV infection in a subject having or at risk of having an HIV infection, comprising administering a chimpanzee adenovirus vector comprising the HTI immunogen in combination with a Modified Vaccinia Ankara virus vector comprising the HTI immunogen and an SOSIP. In one embodiment, the chimpanzee adenovirus vector is ChAdOxl.HTI. In one embodiment, the MVA vector is MVA.HTI. In one embodiment, the SOSIP is ConM SOSIP, such as ConM SOSIP.v7. In one embodiment, ChAdOxl.HTI is administered to the subject followed by ConM SOSIP followed by MVA.HTI. In a particular embodiment, ChAdOxl.HTI is administered in a single dose, followed by at least two doses of ConM SOSIP and at least one dose of MVA.HTI. In a particular embodiment, ChAdOxl.HTI is administered to the subject at week 0, ConM SOSIP. v7 is administered at weeks 4, 12 and 28, and MVA.HTI is administered at week 22. In one embodiment, the subject is also administered MPLA liposomes as adjuvant. In one embodiment, the subject has previously been treated with ART prior to receiving the first dose of HTI immunogen. In one embodiment, the subject is HIV positive. In one embodiment, the HTI vaccine is administered intramuscularly. In one embodiment, the ChAdOxl.HTI is administered at a dose of 5 x 10 ¹⁰ viral particles. In one embodiment, the ConM SOSIP.v7 is administered at a dose of 100 ug. In one embodiment, the MVA.HTI is administered at a dose of 2 x 10 ⁸ pfu. In one embodiment, the MPLA liposomes are administered at a dose of 500 ug. In one embodiment, the subject is further administered a TLR4 agonist or TLR7 agonist in combination with the HTI vaccine regimen above. In one embodiment, the TLR7 agonist is as disclosed in WO 2020/237027, the entire contents of which is hereby incorporated by reference.

[0104] As will be appreciated by those skilled in the art, when treating a viral infection such as HIV, such treatment may be characterized in a variety of ways and measured by a variety of endpoints. The scope of the present disclosure is intended to encompass all such characterizations.

[0105] In some embodiments, the HTI immunogen and SOSIP can be administered through a heterologous prime-boost vaccination that includes different components and vectors, which can be selected from nucleic acids (for example, DNA and RNA vectors), viral vectors (for example, poxvirus, adenovirus, lentivirus, arenavirus and others), bacterial vectors, polypeptides, or antibodies. The aim of the sequential administration of the therapeutic vaccines is to achieve a so-called "functional cure", in which HIV-infected participants could prevent viral infection or control viral replication in the absence of anti-retroviral treatment.

[0106] In some embodiments, the immunogenic polypeptide comprises a sequence having amino acid sequences with no more than 1, 2, or 3 substitutions in any one of SEQ ID NOs: 1-16. In some embodiments, the immunogenic polypeptide comprises a sequence having amino acid sequences according to SEQ ID NOs: 1-16.

[0107] In some embodiments, the immunogenic polypeptide comprises an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 17 orl8. In some embodiments, the immunogenic polypeptide comprises an amino acid sequence according to SEQ ID NO: 17 or 18.

[0108] In some embodiments, the immunogenic polypeptide is encoded by any suitable nucleic acid sequence. In some embodiments, the immunogenic polypeptide is encoded by a nucleic acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 19 or 20. In some embodiments, the immunogenic polypeptide is encoded by a nucleic acid sequence of SEQ ID NO: 19 or 20. In some embodiments, the nucleic acid encodes an immunogenic polypeptide comprising SEQ ID NO: 19 or 20. In some embodiments, the nucleic acid is contained in a viral vector (e.g., a MVA or ChAd vector) or a nucleic acid vector.

[0109] In other embodiments, the immunogenic polypeptide comprises SEQ ID NOs:l- 16. In other embodiments, the immunogenic polypeptide comprises the sequence of SEQ ID NOs: 1-16 or a variant or fragment thereof. In some embodiments, the variant has a length of at least 8 amino acids, and does not comprise any sequence stretches derived from the HIV genome of a length of 8 or more amino acids other than an amino acid sequence according to any of SEQ ID NOs: 1-16 or the variant thereof. In other embodiments, the variant is equivalent to its related sequence and derives from a different HIV strain or is an artificial HIV sequence. Equivalent in this respect means different in one or more amino acid residues, but corresponding to the same sequence (e.g., determined by the position in the genome or sequence similarity). In other words, in one embodiment, the variant is a "naturally occurring variant", which refers to nucleic acid sequences derived from an HIV genome of a presently or formerly circulating virus and can be identified from existing databases (e.g., GenBank and Los Alamos sequence databases). The sequence of circulating viruses can also be determined by molecular biology methodologies. See Brown T, "Gene Cloning" (Chapman & Hall, London, GB, 1995); Watson et al., "Recombinant DNA", 2nd Ed. (Scientific American Books, New York, N.Y., US, 1992); Sambrook et al., "Molecular Cloning. A Laboratory Manual" (Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., US, 1989). In some embodiments, a variant of any of SEQ ID NOs: 1-16 has an amino acid sequence identity of at least 70%, at least 80%, at least 90%, at least 95%, at least 98%, or at least 99% to its corresponding (i.e., SEQ ID NOs: 1-16). Examples of algorithms suitable for determining percent sequence identity and sequence similarity are BLAST and BLAST 2.0 algorithms. Altschul et al., Nuc. Acids Res. 1977; 25:3389-3402 and Altschul et al., J. Mol. Biol. 1990; 215:403-410. The BLAST and BLAST 2.0 programs can be used to determine percent sequence identity for the nucleic acids and proteins of the invention. Software for performing BLAST analyses is publicly available through the National Center for Biotechnology Information. See http://blast.ncbi.nlm.nih.gov/blast.cgi, January 2012.

[0110] In some embodiments, the immunogenic polypeptide comprises at least two, at least three, or at least four sequences selected from SEQ ID NOs: 1-16 or variants thereof, wherein when the immunogen comprises only two, three, or four sequences selected from SEQ ID NOs: 1-16, then not all of these sequences are selected from the group consisting of SEQ ID NOs:3, 5, 6 and 16. In another embodiment, said immunogen has an amino acid sequence comprising at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten sequences selected from SEQ ID NOs: 1-16 or variants thereof, wherein when the immunogen comprises only two, three, four, five, six, seven, eight, nine or ten sequences selected from the group consisting of SEQ ID N0s:l-16, then not all of these sequences are selected from the group consisting of SEQ ID NOs: 1-16.

[OHl] In another embodiment, the variant or fragment has a length of 8 to 40 amino acids, for example, 11 to 27 amino acids. In some embodiments, the variant or fragment does not comprise an amino acid linker adjoining any of SEQ ID NOs: 1-16. In some embodiments, the C-terminal amino acid of said variant or fragment is neither G, P, E, D, Q, N, T, S, nor C.

[0112] In some embodiments, the variant or fragment is combined with or fused to a heat shock protein, for example, HsplO, Hsp20, Hsp30, Hsp40, Hsp60, Hsp70, Hsp90, gp96, or HsplOO.

[0113] In another embodiment, the immunogenic polypeptide further comprises a secretion signal peptide at the N-terminus. In some embodiments, the signal peptide enhances secretion of the immunogen from cells expressing the immunogen. In some embodiments, the signal peptide is derived from tPA (tissue plasminogen activator), for example, followed by an arginine to increase stability. The sequence of the tPA signal peptide is, for example, MDAMKRGLCCVLLLCGAVFVSA (SEQ ID NO:21) or MDAMKRGLCCVLLLCGAVFVSAR (SEQ ID NO:22).

[0114] In some embodiments, at least two sequences of the immunogenic polypeptide are adjoined by an amino acid linker. In some embodiments, the linker has the amino acid sequence A, AA or AAA. In some embodiments, the C-terminal residue of the sequence located N-terminally with respect to the linker or the N-terminal residue of the sequence located C-terminally is an alanine residue, the linker can be shortened so that an AAA sequence is formed in the junction region between adjoining sequences. Thus, in some embodiments, if the C-terminal residue of the sequence located N-terminally with respect to the linker is an alanine or if the N-terminal residue of the sequence located C- terminally with respect to the linker is alanine, the linker has the sequence AA. In another embodiment, if the C-terminal residue of the sequence located N-terminally with respect to the linker and the N-terminal residue of the sequence located C-terminally with respect to the linker are both alanine, then the linker has the sequence A.

[0115] In some aspects, the methods further comprise administering a stable HIV-1 envelope glycoprotein trimer. [0116] In some aspects, the stable soluble HIV-1 envelope glycoprotein trimer comprises mutations to increase stability and improve functionality.

[0117] In some aspects, the HIV vaccine comprises a stable soluble HIV envelope glycoprotein trimer (SOSIP). In some aspects, the SOSIP is Con M that is based on a consensus sequence of each clade in group M. In some aspects, the ConM SOSIP comprises a TPA signal. In some aspects, the ConM SOSIP comprises a gpl20- gp41ECTO disulfide bond (A501C-T605C). In some aspects, the ConM SOSIP comprises a trimer-stabilizing I559P mutation in gp41ECTO. In some aspects, the ConM SOSIP further comprises trimer-stabilizing mutations H66R and A316W. In some aspects, the ConM SOSIP further comprises improved trimerization mutations 1535M and Q543N). In some aspects, the ConM SOSIP comprises an improved furin cleavage site (R6) comprising an REKR to RRRRRR mutation. In some aspects, the ConM SOSPI comprises a stop codon after gp41ECTO residue 664. In some aspects, the ConM SOSIP comprises an extra intermolecular disulfide bond A73C-A561C in gp41. In some aspects, the ConM SOSIP further comprises mutations E47D, N49E, V65K, E106T, I165L, G429R, K500R and a Q at position 432. In some aspects, the ConM SOSIP construct is codon-optimized.

[0118] In some aspects, the HTI vaccine, the ConM SOSIP. v7 gpl40 trimer and, optionally, the broad neutralizing antibody are administered in a single dose. In some embodiments, the HTI vaccine, the ConM SOSIP. v7 gpl40 trimer and, optionally, the broad neutralizing antibody are administered in multiple doses. In some embodiments, the HTI vaccine, ConM SOSIP.v7 gpl40 trimer and, optionally, the broad neutralizing antibody are prepared, purified and formulated for immunization in a human. In some embodiments, the HTI vaccine, ConM SOSIP. v7 gpl40 trimer and, optionally, the broad neutralizing antibody are formulated in one composition. In some embodiments, the HTI vaccine, ConM SOSIP.v7 gpl40 trimer vaccine and, optionally, the broad neutralizing antibody are formulated in separate compositions. In some aspects, the HTI vaccine, ConM SOSIP.v7 gpl40 trimer and, optionally, the broad neutralizing antibody are formulated for immunization in a human to contain an adjuvant. A number of adjuvants are well known to those investigating vaccines. In some aspects, the adjuvant comprises a TLR agonist. In some aspects, the adjuvant comprises a TLR4 agonist. In some aspects, the adjuvant comprises MPLA. In some aspects, additional adjuvants are used with the HTI vaccine, ConM SO SIP trimer and, optionally, the broad neutralizing antibody according to the invention. In some aspects, the HTI vaccine, ConM SOSIP trimer and, optionally, the broad neutralizing antibody are administered with an interval between the administration of the HTI vaccine and Con M SOSIP trimer and the broad neutralizing antibody. In some aspects, the time interval is between 1 hour and 4 weeks.

[0119] In some aspects, the HTI vaccine and the ConM SOSIP. v7 gpl40 trimer are administered with a broad neutralizing antibody.

[0120] In some aspects, the HTI vaccine and the ConM SOSIP trimer are formulated in one composition and the broad neutralizing antibody is formulated in a further separate composition. In some aspects, the HTI vaccine, the ConM SOSIP trimer and the broad neutralizing antibody are formulated in separate compositions.

[0121] In some aspects, the HTI vaccine and the ConM SOSIP trimer are administered together and the broad neutralizing antibody is administered with a time interval of at least 1 h. In some aspects, the HTI vaccine, the ConM SOSIP trimer and the broad neutralizing antibody are administered in separate compositions.

[0122] In some aspects, the HTI vaccine, the ConM SOSIP trimer and the broad neutralizing antibody are administered at the same time. In some aspects, the HTI vaccine, the ConM SOSIP trimer and the broad neutralizing antibody are administered at the same time but at different locations. In some aspects, the HTI vaccine, the ConM SOSIP trimer and the broad neutralizing antibody are administered at different time points. In some aspects, the HTI vaccine and the ConM SOSIP trimer are administered first and the broad neutralizing antibody is administered thereafter in a time interval from about 1 h to about 4 weeks or any time interval therebetween.

Sequences

[0123] In addition to sequences disclosed elsewhere in the present disclosures, the following sequences are provided as they are mentioned or used in various exemplary embodiments of the disclosures, which are provided for the purpose of illustration.

[0124] Although the foregoing disclosure has been described in some detail by way of illustration and Example for purposes of clarity of understanding, one of skill in the art will appreciate that certain changes and modifications may be practiced within the scope of the appended claims. In addition, each reference provided herein is incorporated by reference in its entirety to the same extent as if each reference was individually incorporated by reference. Where a conflict exists between the instant application and a reference provided herein, the instant application shall dominate.

Additional Dosing and Dosing Regimens

[0125] In some embodiments, the method of the present invention comprises (a) administering to the subject 1 to 10 (i.e., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) administrations of a vector (e.g., DNA vector or viral vector) encoding the HTI immunogen, and (b) administering to the subject 1 to 10 (i.e., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) administrations of a recombinant HIV-1 Env ectodomain trimer stabilized in a prefusion mature closed conformation.

[0126] In some embodiments, the method comprises (a) administering to the subject 1 to 10 (i.e., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) administrations of a viral vector encoding the HTI immunogen, followed by 1 to 10 (i.e., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) administrations of a recombinant HIV-1 Env ectodomain trimer stabilized in a prefusion mature closed conformation; and (b) administering to the subject 1 to 10 (i.e., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) administrations of a second viral vector encoding the HTI immunogen followed by 1 to 10 (i.e., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) administrations of a recombinant HIV-1 Env ectodomain trimer stabilized in a prefusion mature closed conformation.

[0127] In some embodiments, the method comprises (a) administering to the subject 1 to 10 (i.e., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) administrations of a viral vector encoding the HTI immunogen, followed by 1 to 10 (i.e., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) administrations of a recombinant HIV-1 Env ectodomain trimer stabilized in a prefusion mature closed conformation and 1 to 10 (i.e., 1, 2, 3, 4, 5, 6, 7,8, 9, or 10) administrations of a broad neutralizing antibody; and (b) administering to the subject 1 to 10 (i.e., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) administrations of a second viral vector encoding the HTI immunogen followed by 1 to 10 (i.e., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10) administrations of a recombinant HIV-1 Env ectodomain trimer stabilized in a prefusion mature closed conformation and 1 to 10 (i.e., 1, 2, 3, 4, 5, 6, 7,8, 9, or 10) administrations of a broadly neutralizing antibody.

[0128] In some embodiments, the method comprises (a) administering to the subject 1 to 4 administrations of a vector (e.g., a DNA vector or viral vector) encoding the HTI immunogen followed by 1 to 4 administrations of a recombinant HIV-1 Env ectodomain trimer stabilized in a prefusion mature closed conformation; and (b) administering to the subject 1 to 4 administrations of a second viral vector encoding the HTI immunogen followed by 1 to 4 administrations of a recombinant HIV-1 Env ectodomain trimer stabilized in a prefusion mature closed conformation.

[0129] In some embodiments, the method comprises (a) administering to the subject 1 to 4 administrations of a vector (e.g., a DNA vector or viral vector) encoding the HTI immunogen followed by 1 to 4 administrations of a recombinant HIV-1 Env ectodomain trimer stabilized in a prefusion mature closed conformation followed by 1 to 4 administrations of a broad neutralizing antibody; and (b) administering to the subject 1 to 4 administrations of a second viral vector encoding the HTI immunogen followed by 1 to 4 administrations of a recombinant HIV-1 Env ectodomain trimer stabilized in a prefusion mature closed conformation followed by 1 to 4 administrations of a broadly neutralizing antibody.

[0130] In some embodiments, the method comprises (a) administering to the subject 1 to 4 administrations of a viral vector encoding the HTI immunogen, followed by 1 to 4 administrations of a recombinant HIV-1 Env ectodomain trimer stabilized in a prefusion mature closed conformation; and (b) administering to the subject 1 to 4 administrations of a second viral vector encoding the HTI immunogen followed by 1 to 4 administrations of a recombinant HIV-1 Env ectodomain trimer stabilized in a prefusion mature closed conformation.

[0131] In some embodiments, the method comprises (a) administering to the subject 1 administrations of a viral vector encoding the HTI immunogen, followed by 2 administrations of a recombinant HIV-1 Env ectodomain trimer stabilized in a prefusion mature closed conformation; and (b) administering to the subject 1 administration of a second viral vector encoding the HTI immunogen followed by 1 administration of a recombinant HIV-1 Env ectodomain trimer stabilized in a prefusion mature closed conformation.

[0132] In some embodiments, the method comprises (a) administering to the subject (i) 1 administration of a first viral vector encoding the HTI; (ii) 2 administration of a recombinant HIV-1 Env ectodomain trimer stabilized in a prefusion mature closed conformation, each separated by a period of about 8 weeks, about 4 weeks after the administration of the first viral vector; and (b) administering to the subject (i); 1 administration of a second viral vector encoding the HTI; (ii) 1 administration of a recombinant HIV-1 Env ectodomain trimer stabilized in a prefusion mature closed conformation, about 6 weeks after the administration of the second viral vector. In some embodiments of such a method, the administrations of (a)(i) is at a dose of about 5xlO ¹⁰ viral particles, the administration of (a)(ii) is at a dose of about 100 ug; the administration of (b)(i) is at a dose of about 2xl0 ⁸ pfu, and the administration of (b)(ii) is at a dose of about 100 ug. In other embodiments, the first viral vector of (a)(i) is a chimpanzee adenoviral (ChAd) vector. In other embodiments, the second viral vector of (b)(i) is a Modified Vaccine virus Ankara (MV A) vector. In other embodiments, the recombinant HIV-1 Env ectodomain trimer stabilized in a prefusion mature closed conformation of (a)(ii) and (b)(ii) is the SOSIP immunogen ConM SOSIP.v7 gpl40 as in Slieppen et al. 2019 Nat. Com.

[0133] Immunogenic polypeptides such as the recombinant HIV-1 Env ectodomain trimer stabilized in a prefusion mature closed conformation and viral vectors encoding the same of the invention can be administered in a variety of manners, for example, via the mucosa, such as oral and nasal, pulmonary, intramuscular, subcutaneous or intradermal routes.

[0134] Immunogenic polypeptides such as the recombinant HIV-1 Env ectodomain trimer stabilized in a prefusion mature closed conformation and viral vectors encoding the same of the invention can also be administered in a pharmaceutical composition comprising a pharmaceutically acceptable carrier (also referred to herein as a vaccine or vaccine formulation). Examples of a pharmaceutically acceptable carrier include, but are not limited to, a solid, semisolid or liquid filler, diluent, encapsulating material or formulation auxiliary of any conventional type. Other suitable pharmaceutically acceptable carriers include, but are not limited to, water, dextrose, glycerol, saline, ethanol, and combinations thereof. In some embodiments, a pharmaceutically acceptable carrier can contain additional agents such as wetting or emulsifying agents, pH buffering agents, or adjuvants that enhance the effectiveness of the formulation.

[0135] In addition, aqueous compositions, such as those used to prepare HIV vaccine formulations, may be prepared in sterile form, and when intended for delivery by other than oral administration generally may be isotonic. All compositions may optionally contain excipients such as those set forth in the Rowe et al, Handbook of Pharmaceutical Excipients, 9 ^th edition, American Pharmacists Association, 2020. Excipients can include ascorbic acid and other antioxidants, chelating agents such as EDTA, carbohydrates such as dextrin, hydroxyalkylcellulose, hydroxyalkylmethylcellulose, stearic acid and the like.

[0136] In some embodiments, the composition comprises 5 * IO ¹⁰ viral particles (vp) in 0.5 mL formulation buffer of a viral vector encoding the HTI immunogen. In some embodiments, the composition comprises 5 x 1O ¹⁰ viral particles (vp) in 0.5 mL formulation buffer of a replication-defective chimpanzee adenovirus encoding the HTI immunogen. In some embodiments, the composition comprises 5 x 10 ¹⁰ viral particles (vp) in 0.5 mL formulation buffer of a ChAdOxl vector encoding the HTI immunogen comprising the sequence of SEQ ID NO: 17 or 18, where in the formulation buffer comprises 10 mM L-Histidine, 35 mM NaCl, 7.5 % (w/v) of sucrose, 1 mM MgC12, 0.1 mM EDTA disodium, 0.1% (w/v) Polysorbate-80, 0.5% (v/v) ethanol, and a pH of 6.6.

[0137] In some embodiments, the pharmaceutical composition comprises 2 X 10 ⁸ plaque forming units (PFU) of a viral vector encoding the HTI immunogen in 0.5 mL Tris buffer. In some embodiments, the pharmaceutical composition comprises 2 X 10 ⁸ plaque forming units (PFU) of an MVA vector encoding the HTI immunogen in 0.5 mL Tris buffer. In some embodiments, the pharmaceutical composition comprises 2 X 10 ⁸ plaque forming units (PFU) of an MVA vector comprising a nucleic acid encoding an immunogenic polypeptide having an amino acid sequence of SEQ ID NO: 17 or 18 in 0.5 mL Tris buffer. In some embodiments, the pharmaceutical composition comprises 2 X 10 ⁸ PFU of an MVA vector comprising a nucleic acid comprising the sequence of SEQ ID NO: 17 or 18 in 0.5 mL Tris buffer, wherein Tris buffer comprises 10 mM Tris HC1, pH 7.7 and 140 mM NaCl.

[0138] In some embodiments, the pharmaceutical composition comprises 100 ug of a recombinant HIV-1 Env ectodomain trimer stabilized in a prefusion mature closed conformation in 0.6 ml of formulation buffer. In some embodiments, the pharmaceutical composition comprises 100 ug of a recombinant HIV-1 Env ConM SOSIP.v7 gpl40 stabilized in a prefusion mature closed conformation in 0.6 ml of formulation buffer, wherein the formulation buffer contains Tris, NaCl Hydrochloric acid (25%). In some embodiments, the administration of the recombinant HIV-1 Env ConM SOSIP.v7 gpl40 stabilized in a prefusion mature closed conformation is co-adjuvated with 500 ug of MPLA liposomes in administration solution, wherein the administration solution is dimiristoyl phosphatidylcholine (DMPC), dimyristoyl phosphatidylglycerol (DMPG), cholesterol, sucrose, Na2HPO4*2H2O, KC1 and KH2PO4.

[0139] It should be understood that description herein related to the administration of an immunogenic polypeptide or nucleic acid encoding an immunogenic polypeptide also applies to administration of a pharmaceutical composition or vaccine containing the same.

[0140] The amount of the virus within a pharmaceutical composition can be measured by any means known in the art. The amount may be determined by bulk measurement of the number of viral particles (vp) within an amount of aqueous composition, e.g., by flow cytometry. Alternatively, the amount may be determined by the activity of the virus within the composition, e.g., by plaque assay. Plaque-based assays can be used to determine virus concentration in terms of infectious dose. Viral plaque assays determine the number of plaque forming units (pfu) in a virus sample, which can be used as a measure of virus quantity. Kaufmann et al. 2002; Methods in Microbiology Vol.32: Immunology of Infection. Academic Press. ISBN 0-12-521532-0.

[0141] In some embodiments, a viral vector (e.g., MVA or ChAd vector) encoding an immunogenic polypeptide of the present invention is administered at a dose of from about IxlO ⁷ plaque forming units (pfu) to about IxlO ⁹ pfu, for example, from about 5xl0 ⁷ pfu to about IxlO ⁹ pfu, from about IxlO ⁸ pfu to about IxlO ⁹ pfu, from about 5xl0 ⁸ pfu to about IxlO ⁹ pfu. In some embodiments, a viral vector encoding an immunogenic polypeptide of the present invention is administered at a dose of from about 5xl0 ⁷ pfu to about 5xl0 ⁸ pfu. In some embodiments, a viral vector encoding an immunogenic polypeptide of the present invention is administered at a dose of about 2.5xl0 ⁸ pfu. In some embodiments, a viral vector encoding an immunogenic polypeptide of the present invention is administered at a dose of about IxlO ⁷ pfu, about IxlO ⁸ pfu, about IxlO ⁹ pfu, about 5xl0 ⁷ pfu or about 5xl0 ⁸ pfu. [0142] In some embodiments, a viral vector (e.g., MVA or ChAd vector) encoding an immunogenic polypeptide of the present invention is administered at a dose of from about IxlO ⁹ viral particles and 5xl0 ⁿ viral particles, for example, from about 5xl0 ⁹ pfu to about 5xl0 ⁿ pfu, from about IxlO ¹⁰ pfu to about 5xl0 ⁿ pfu, from about 5xlO ¹⁰ pfu to about 5xl0 ⁿ pfu, or from about IxlO ¹¹ pfu to about 5xl0 ⁿ pfu. In some embodiments, a viral vector encoding an immunogenic polypeptide of the present invention is administered at a dose of from about IxlO ¹⁰ to about IxlO ¹¹ viral particles, for example, from about 5xlO ¹⁰ pfu to about IxlO ¹¹ pfu. In some embodiments, a viral vector encoding an immunogenic polypeptide of the present invention is administered at a dose of from about 5xlO ¹⁰ viral particles.

[0143] The amount of immunogenic compound (e.g., HTI immunogen, or the recombinant HIV-1 Env ectodomain trimer or the broadly neutralizing antibody) delivered can vary, depending upon the intended use (preventive or therapeutic vaccination), and age and weight of the subject to be immunized, the vaccination protocol adopted (i.e., single administration versus repeated doses), the route of administration and the potency and dose of the adjuvant compound chosen. The amount can be ascertained by standard studies involving observation of appropriate immune responses in vaccinated subjects. In some embodiments, following an initial vaccination, composed itself by one or several doses, subjects can receive one or several booster immunizations adequately spaced.

[0144] In some embodiments, an immunogenic compound or composition is administered once, or can be administered repeatedly, for example, from about 1 and about 10 times, for example, from about 1 to about 9 times, from about 1 to about 8 times, from about 1 to about 7 times, from about 1 to about 6 times, from about 1 to about 5 times, from about 1 to about 4 times, from about 1 to about 3 times, from about 1 to about 2 times, from about 2 to about 9 times, from about 2 to about 8 times, from about 2 to about 7 times, from about 2 to about 6 times, from about 2 to about 5 times, from about 2 to about 4 times, from about 2 to about 3 times, from about 3 to about 9 times, from about 3 to about 8 times, from about 3 to about 7 times, from about 3 to about 6 times, from about 3 to about 5 times, from about 3 to about 4 times, from about 4 to about 9 times, from about 4 to about 8 times, from about 4 to about 7 times, from about 4 to about 6 times, or from about 4 to about 5 times. [0145] In some embodiments, an immunogenic compound or composition is administered at different intervals between doses of the same component or doses of different component. In some embodiments, the interval between doses is from about 1 week to about 24 months, for example, from about 2 weeks to about 24 months, from about 3 weeks to about 24 months, from about 4 weeks to about 24 months, from about 2 weeks to about 56 weeks, from about 4 weeks and about 12 weeks.

[0146] In other embodiments, each administration of the methods of the present invention is separated by a period of from about 15 days to about 18 months. In some embodiments, each administration of the methods of the present invention is separated by a period of from about 1 week to about 24 months. In some embodiments, each administration of the methods of the present invention is separated by a period of from about 2 weeks to about 56 weeks. In some embodiments, each administration of the methods of the present invention is separated by a period of from about 4 weeks to about 12 weeks. In some embodiments of the methods of the present invention, the administering of step (a) of the methods of the present invention is separated from the administering of step (b) by a period of from about 2 months to about 24 months. In some embodiments of the methods of the present invention, the administering of step (a) is separated from the administering of step (b) by a period of from about 3 months to about 18 months.

[0147] In some embodiments, the vaccine components of the present invention can be grouped in a priming phase and a subsequent one or multiple boosting phases. In some embodiments, the priming phase and the boosting phase can be separated by from about 2 weeks to about 20 weeks, for example, from about 5 weeks to about 14 weeks. In some embodiments, the subject will receive the immunogen compound or composition of the invention as different vaccine components in a prime-boost regime. In some embodiments, such a regimen is followed by dosing at regular intervals of from about 1 months to about 12 months for a period up to the remainder of the subject's life.

[0148] In some embodiments, the immunogenic compounds or compositions of the invention are used in any sequence, each component be used one or several times, in any order, and with any interval between doses. For example, a sequence is MVA.HTI + ConM SOSIP.v7 gpl40 (S) (vaccination sequence MS), each dose separated 6 weeks apart. In some embodiments, the sequence comprises a priming phase of ChAd.HTI (C) + S0SIP.v7 gpl40 (S), each dose administered at 4 weeks interval, followed by a S dose 8 weeks after the last S dose. Thus, the full sequence is: (1) Priming phase of CSS, followed 10 weeks after by a (2) boosting phase of MS.

[0149] In some embodiments, the sequence comprises a priming phase of C (5.OxlO ¹⁰ viral particles), at week 0 followed by two S administrations (100 ug) at week 4 and week 12; followed by a boosting phase of a dose of M (2xl0 ⁸ pfu) 10 weeks after the last S and a third dose of S 6 weeks after the M.

[0150] In some embodiments, the broadly neutralizing antibodies to gpl20 and gp41 envelope glycoproteins (Env) are antibodies against the CD4 binding site and in particular the VRC07-523-LS, that has been engineered to increase clinical protective efficacy in terms of neutralization potency and in vivo efficacy when administered in humans from 1 to 6 times and at a dose between lOOmg and 5g in a regimen described in Example II.

Kits

[0151] In some embodiments, the present invention relates to a kit comprising immunogenic polypeptide of the invention, or nucleic acid or vector encoding the same, or a pharmaceutical composition comprising the same, and instructions for using the same in a method of present invention described herein. In some embodiments, the kit comprises a packaging, such as glass, plastic (e.g., polyethylene, polypropylene, polycarbonate), bottles, vials, paper, or sachets for the components. In some embodiments, the instructions are in the form of printed material or in the form of an electronic support which can store the instructions, for example, electronic storage media (e.g., magnetic disks, tapes), or optical media (e.g., CD-ROM, DVD). The media can additionally or alternatively contain internet websites providing such instructions.

[0152] All publications, patents and patent applications mentioned in this application are herein incorporated in their entirety by reference into the specification, to the same extent as if each individual publication, patent or patent application was specifically and individually indicated to be incorporated herein by reference. In addition, citation or identification of any reference in this application shall not be construed as an admission that such reference is available as prior art to the present invention. To the extent that section headings are used, they should not be construed as necessarily limiting.

[0153] Embodiments of the present disclosure can be further defined by reference to the following non-limiting examples, which describe in detail preparation of some antibodies of the present disclosure and methods for using antibodies of the present disclosure. It will be apparent to those skilled in the art that many modifications, both to materials and methods, may be practiced without departing from the scope of the present disclosure.

[0154] It is to be appreciated that the Detailed Description section, and not the Summary and Abstract sections, is intended to be used to interpret the claims. The Summary and Abstract sections may set forth one or more but not all exemplary embodiments of the present invention as contemplated by the inventor(s), and thus, are not intended to limit the present invention and the appended claims in any way.

[0155] The present invention has been described above with the aid of functional building blocks illustrating the implementation of specified functions and relationships thereof. The boundaries of these functional building blocks have been arbitrarily defined herein for the convenience of the description. Alternate boundaries can be defined so long as the specified functions and relationships thereof are appropriately performed.

[0156] The foregoing description of the specific embodiments will so fully reveal the general nature of the invention that others can, by applying knowledge within the skill of the art, readily modify and/or adapt for various applications such specific embodiments, without undue experimentation, without departing from the general concept of the present invention. Therefore, such adaptations and modifications are intended to be within the meaning and range of equivalents of the disclosed embodiments, based on the teaching and guidance presented herein. It is to be understood that the phraseology or terminology herein is for the purpose of description and not of limitation, such that the terminology or phraseology of the present specification is to be interpreted by the skilled artisan in light of the teachings and guidance.

[0157] The breadth and scope of the present invention should not be limited by any of the above-described exemplary embodiments, but should be defined only in accordance with the following claims and their equivalents.

[0158] The claims in the instant application are different than those of the parent application or other related applications. The Applicant therefore rescinds any disclaimer of claim scope made in the parent application or any predecessor application in relation to the instant application. The Examiner is therefore advised that any such previous disclaimer and the cited references that it was made to avoid, may need to be revisited. Further, the Examiner is also reminded that any disclaimer made in the instant application should not be read into or against the parent application.

EXAMPLE

Example 1: HIV Treatment Protocol of a HIV Vaccine and SOSIPs

[0159] A single-site phase I, randomized, double-blind, placebo-controlled clinical trial to evaluate the safety, tolerability, immunogenicity and efficacy of a vaccine regimen of the HTI immunogen is conducted. In particular, the study involves the administration in combination of ChAdOxl.HTI with MVA.HTI and ConMSOSIP.v7 gpl40 adjuvanted with MPLA liposomes in 30 virologically suppressed ART -treated HIV-1 positive individuals. Participants are randomized to receive either a combined regimen with the HTI immunogen (CSSMS) or placeb (PPPPP) in a double-blinded fashion at a ratio of 2:1. The treatment group receives ChAdOx.1.HTI (“C”) at week 0 followed by ConM SOSIP.v7 (“S”) at weeks 4, 12 and 28 and MVA.HTI (“M”)at week 22 (i.e., CSSMS). The control group receives normal saline solution. Safety and immunogenicity of the vaccine regimen is measured up to week 30 (2 weeks after last ConMSOSIP.v7/placebo administration). At week 30, all participants undergo ATI of 24 weeks duration. At visit week 54, or before according to specified criteria, ART is resumed. Efficacy endpoints are measured at 12 and 24 weeks of ATI (study visits week 42 and 54). After ART resumption, participants are followed during an additional safety period of 12 weeks (study visit week 66, End-of-Study visit). Treatment allocation is blinded until week 54 (End-of-ATI visit). ChAdOxl.HTI is administered intramuscularly at a dose of 5xlO ¹⁰ Vp. ConM SOSIP.v7 is administered at a dose of 100 ug per administration adjuvanted with MPLA liposomes (500 ug). MVA.HTI is administered at a dose of 2xl0 ⁸ pfu. The HTI immunogen vaccine regimen with ChAdOxl.HTI, MVA.HTI and Recombinant HIV-1 Envelope ConM SOSIP.v7 gpl40 adjuvanted with MPLA liposomes (CSSMS) is shown to prevent/delay viral rebound, induce post-rebound viral control, and/or prevent or delay the need for resumption of ART during an analytical treatment interruption (ATI) of ART in virologically-suppressed ART -treated HIV-1 positive individuals. This clinical study demonstrates the safety of an ATI period after the combined T- and B-cell regimen with ChAdOxl.HTI, MVA.HTI and Recombinant HIV-1 Envelope ConM S0SIP.v7 gpl40 adjuvanted with MPLA liposomes (CSSMS) in virologically-suppressed ART -treated HIV-1 positive individuals. Figure 1 summarizes the overall design.

[0160] The following criteria for a participant to be enrolled in the study includes that the participant:

(1) is at least 18 years on the day of screening and no greater than 60 years on the day of the first IMP administration.

(1) Has confirmed HIV-1 infection.

(2) Has been virologically suppressed for at least 2 years prior to the screening visit, defined as maintained pVL <50 cop/ml allowing for isolated blips (non-consecutive 50-200 copies/mL)

(3) Is under the same ART regimen within at least 4 weeks prior to screening visit.

(3) Has stable CD4 count > 500 cells/mm ³ at the screening visit.

(4) Has nadir CD4 count > 350 cells/mm ³ . Lower counts at the moment of acute HIV-1 infection will be allowed only if appropriate immune recovery was followed after ART initiation and ART was not initiated within first 6 months after estimated time of HIV-1 acquisition.