Anti-HIV vaccine antibodies with reduced polyreactivity

Abstract

To provide broad and potent antibodies against Human Immunodeficiency Virus ("HIV").SOLUTION: This disclosure provides novel broadly neutralizing anti-HIV antibodies and antigen-binding fragments thereof. The disclosed anti-HIV antibodies exhibited improved biophysical properties, e.g, reduced polyreactivity, prolonged half-life, while retaining broad and potent neutralization activity. The anti-HIVbNAb variants as disclosed constitute a novel therapeutic strategy for treating and / or preventing HIV infection.SELECTED DRAWING: None

Description

[Technical field]

[0001] (CROSS REFERENCE TO RELATED APPLICATIONS) This application claims priority under 35 U.S.C. §119(e) to U.S. Provisional Patent Application No. 62 / 871,393, filed July 8, 2019. The aforementioned application is hereby incorporated by reference in its entirety.

[0002] (Statement regarding federally sponsored research and development) This invention was made with Government support under P01AI100148 and R01AI29784 awarded by the National Institutes of Health (NIH). The Government has certain rights in this invention.

[0003] (Technical field) The present invention relates to broad spectrum and potent antibodies against the Human Immunodeficiency Virus ("HIV"). [Background technology]

[0004] HIV causes acquired immune deficiency syndrome (AIDS), a human condition characterized by clinical features such as a wasting syndrome, degeneration of the central nervous system, and profound immunosuppression leading to life-threatening opportunistic infections and malignancies. Since its discovery in 1981, HIV type 1 (HIV-1) has been responsible for at least 25 million deaths worldwide. Even if HIV infections were to decline by 2.5% per year, it is projected that 20-60 million people would become infected over the next 20 years. There is a need for therapeutic agents and methods for the treatment or inhibition of HIV infection.

[0005] Broadly neutralizing antibodies (bNAbs) against HIV are being developed as potential therapeutics for the treatment and / or prevention of HIV infection. Many bNAbs have been isolated and characterized from human donors. Modifications have been made to some of these bNAbs to improve breadth and / or potency of neutralization. One such engineered bNAb is NIH45-46, which has the heavy chain mutation G54W (NIH45-46 G54W). Although NIH45-46 G54W has a desirable neutralization profile, it has biophysical properties such as polyreactive / nonspecific binding, a short in vivo half-life, and a tendency to aggregate, which hinder its development as a human therapeutic. There are various known approaches to improve the biophysical properties of antibodies, but it can be difficult to improve these properties without sacrificing much of the antibody's neutralizing activity. Summary of the Invention

[0006] In one aspect, the disclosure provides novel, isolated, broadly neutralizing, anti-HIV antibodies, or antigen-binding portions thereof, with improved biophysical properties, such as reduced polyreactivity. The isolated, anti-HIV antibodies, or antigen-binding portions thereof, comprise one or more heavy chain substitutions at one or more residues selected from the group consisting of S5, S21, G54, G55, V57, T68, T70, V73, S75, F79, S82, D85, V89, Y97, and P112, where the numbering of residues is indicated by Kabat numbering, and have a similarity to SEQ ID NO:1 by at least 75% (e.g., 80%, 85%, a heavy chain having a heavy chain amino acid sequence identical (90%, 95%, 96%, 97%, 98%, 99% or greater) to SEQ ID NO:2; or one or more light chain substitutions at one or more residues selected from the group consisting of T5, S7, T10, S12, S14, T18, I20, S22, R24, Q27, S28, S30, R59, S61, S63, W65, D68, N70, S72, S74, and S78; light chains having a light chain amino acid sequence that is at least 75% (e.g., 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or more) identical to the light chain.

[0007] In some embodiments, the isolated anti-HIV antibody or antigen-binding portion thereof has a heavy chain polypeptide sequence that is at least 75% (e.g., 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or more) identical to SEQ ID NO:1, and the heavy chain amino acid sequence comprises one or more heavy chain substitutions at one or more residues selected from the group consisting of S5, S21, G54, G55, V57, T68, T70, V73, S75, F79, S82, D85, V89, Y97, and P112. and a light chain having a light chain amino acid sequence at least 75% (e.g., 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or more) identical to SEQ ID NO:2, wherein the light chain amino acid sequence includes one or more light chain substitutions at one or more residues selected from the group consisting of T5, S7, T10, S12, S14, T18, I20, S22, R24, Q27, S28, S30, R59, S61, S63, W65, D68, N70, S72, S74, and S78.

[0008] In some embodiments, the one or more heavy chain substitutions are: an S5D or S5E substitution, or a conservative substitution of Asp or Glu at residue S5; an S21D or S21E substitution, or a conservative substitution of Asp or Glu at residue S21; a G54W substitution, or a conservative substitution of Trp at residue G54; a G55D or G55E substitution, or a conservative substitution of Asp or Glu at residue G55; a V57D or V57E substitution, or a conservative substitution of Asp or Glu at residue V57; a T68D or T68E substitution, or a conservative substitution of Asp or Glu at residue T68; a T70D or T70E substitution, or a conservative substitution of Asp or Glu at residue T70; a V73D or V73E substitution, or a conservative substitution of Asp or Glu at residue V73. a conservative substitution of Asp or Glu at residue S75; a substitution of S75D or S75E, or a conservative substitution of Asp or Glu at residue S75; a substitution of F79D, F79E, F79Y or F79H, or a conservative substitution of Asp, Glu, Tyr or His at residue F79; a substitution of S82D or S82E, or a conservative substitution of Asp or Glu at residue S82; a substitution of D85E, or a conservative substitution of Asp at residue D85; a substitution of V89D or V89E, or a conservative substitution of Asp or Glu at residue V89; a substitution of Y97D or Y97E, or a conservative substitution of Asp or Glu at residue Y97; a substitution of P112D or P112E, or a conservative substitution of Asp or Glu at residue P112; or a combination thereof.

[0009] In some embodiments, the one or more light chain substitutions are a T5D or T5E substitution, or a conservative substitution of Asp or Glu at residue T5; a S7D or S7E substitution, or a conservative substitution of Asp or Glu at residue S7; a T10D or T10E substitution, or a conservative substitution of Asp or Glu at residue T10; a S12D or S12E substitution, or a conservative substitution of Asp or Glu at residue S12; a S14D or S14E substitution, or a conservative substitution of Asp or Glu at residue S14; a T18D or T18E substitution, or a conservative substitution of Asp or Glu at residue T18; is a substitution of I20E, or a conservative substitution of Asp or Glu at residue I20; a substitution of S22D or S22E, or a conservative substitution of Asp or Glu at residue S22; a substitution of R24D or R24E, or a conservative substitution of Asp or Glu at residue R24; a substitution of Q27D or Q27E, or a conservative substitution of Asp or Glu at residue Q27; a substitution of S28Y or S28H, or a conservative substitution of Tyr or His at residue S28; a substitution of S30D or S30E, or a conservative substitution of Asp or Glu at residue S30; a substitution of R59D or R59E, or a conservative substitution of Asp or Glu at residue R59. a conservative substitution of u; a substitution of S61D or S61E, or a conservative substitution of Asp or Glu at residue S61; a substitution of S63D or S63E, or a conservative substitution of Asp or Glu at residue S63; a substitution of W65D, W65E, W65Y or W65H, or a conservative substitution of Asp, Glu, Tyr or His at residue W65; a substitution of D68E, or a conservative substitution of Asp at residue D68; a substitution of N70D or N70E, or a conservative substitution of Asp or Glu at residue N70; a substitution of S72D or S72E, or a conservative substitution of Asp or Glu at residue S72; a substitution of S74D or S74E, or a conservative substitution of Asp or Glu at residue S74; a substitution of S78D or S78E, or a conservative substitution of Asp or Glu at residue S78; or a combination thereof.

[0010] In some embodiments, the one or more heavy chain substitutions further comprise the following substitutions: M428L and N434S.

[0011] In some embodiments, the light chain amino acid sequence comprises one or more of the following light chain substitutions: (i) Q27E, S28H, and S30D; (ii) Q27E, S28H, S30D, and S74T; (iii) Q27E, S28H, S30D, S74T, M428L, and N434S; (iv) Q27E, S28Y, and S30D; (v) Q27D and S28H; (vi) S28Y; (vii) T5D, T10D, S12D, S14D, I20D, and S22D; or (viii) S61D, S63D, W65D, N70D, S72D, and S74D.

[0012] In some embodiments, the heavy chain amino acid sequence comprises one or more heavy chain substitutions comprising G54W and T68D; and the light chain amino acid sequence comprises one or more light chain substitutions: (a) Q27E, S28H, S30D, and S74T; (b) Q27E, S28H, S30D, S74T, M428L, and N434S; (c) Q27E, S28Y, and S30D; (d) Q27D and S28H; or (e) S28Y.

[0013] In some embodiments, the heavy chain amino acid sequence comprises one or more heavy chain substitutions including G54W; and the light chain amino acid sequence comprises one or more light chain substitutions: (a) Q27E, S28H, and S30D; (b) Q27E, S28Y, and S30D; or (c) Q27D and S28H.

[0014] In some embodiments, the heavy chain amino acid sequence comprises the amino acid sequences of SEQ ID NO: 3 to SEQ ID NO: 35. In some embodiments, the light chain amino acid sequence comprises the amino acid sequences of SEQ ID NO: 2 and SEQ ID NO: 36 to SEQ ID NO: 80.

[0015] In some embodiments, the isolated anti-HIV antibody comprises CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 comprising the respective sequences set forth in the set of CDR sequences of SEQ ID NOs: 81, 84, 87, 90, 93, and 96; SEQ ID NOs: 81, 84, 87, 99, 93, and 96; SEQ ID NOs: 81, 84, 87, 100, 93, and 96; SEQ ID NOs: 81, 84, 87, 101, 93, and 96; or SEQ ID NOs: 81, 84, 87, 102, 93, and 96; (as set forth in Kabat numbering); SEQ ID NOs: 82, 85, 88, 91, 94, and 97 (as set forth in IMGT numbering); or SEQ ID NOs: 83, 86, 89, 92, 95, and 98 (as set forth in Chothia numbering).

[0016] In some embodiments, the heavy chain amino acid sequences and the light chain amino acid sequences are SEQ ID NOs: 3 and 36; SEQ ID NOs: 3 and 38; SEQ ID NOs: 3 and 39; SEQ ID NOs: 4 and 2 SEQ ID NOs:4 and 38; SEQ ID NOs:4 and 39; SEQ ID NOs:4 and 41; and each of the amino acid sequences set forth in the set of sequences selected from the group consisting of SEQ ID NOs:34 and 37.

[0017] In some embodiments, the isolated anti-HIV antibody is a bispecific antibody comprising a first antigen-binding arm that binds a first antigen and a second antigen-binding arm that binds a second antigen, wherein the first and second antigens are different and the first antigen-binding arm comprises the heavy and light chain amino acid sequences set forth above. In some embodiments, the second antigen is selected from the group consisting of CD3, FcγRI (CD64), FcγRII (CD32), FcγRIII (CD16); CD89, CCR5, CD4, CD8, CD28, CD137, CTLA-4, gp41, killer cell immunoglobulin-like receptor, three Ig domains, and long cytoplasmic tail 1 (KIR3DL1), ... Killer cell immunoglobulin-like receptor, two Ig domains and long cytoplasmic tail 1 (KIR2DL1), Killer cell immunoglobulin-like receptor, two Ig domains and long cytoplasmic tail 3 (KIR2DL3), Killer cell lectin-like receptor Cl (KLRC1), Killer cell lectin-like receptor C2 (KLRC2), Killer cell lectin-like receptor C3 (KLRC3), Killer cell lectin-like receptor C4 (KLRC4), Killer cell lectin-like receptor D1 (KLRD1), Killer cell lectin-like receptor KLRK1 (KLRK1), natural cytotoxicity-inducing receptor 3 (NCR3 or NKp30), natural cytotoxicity-inducing receptor 2 (NCR2 or NK-p44), natural cytotoxicity-inducing receptor 1 (NCR1 or NK-p46), CD226 (DNAM-1), cytotoxic and regulatory T cell molecule (CRTAM or CD355), signaling lymphocyte activation molecule family member 1 (SLAMF1), CD48 (SLAMF2), lymphocyte antigen 9 (LY9 or SLAMF3), CD2 44 (2B4 or SLAMF4), CD84 (SLAMF5), SLAM family member 6 (SLAMF6 or NTB-A), SLAM family member 7 (SLAMF7 or CRACC), CD27 (TNFRSF7), semaphorin 4D (SEMA4D or CD100), CD160 (NK1), an immune checkpoint protein (e.g., PD-1, PD-L1, CTLA-4), and the second epitope of gp120.

[0018] The disclosure also provides (i) an isolated nucleic acid comprising a sequence encoding the CDR, heavy chain variable region, or light chain variable region, or an antigen-binding portion thereof, of the aforementioned anti-HIV antibody, (ii) a vector comprising the aforementioned nucleic acid, and (iii) a cultured cell comprising the aforementioned vector.

[0019] Also within the scope of this disclosure is a method for producing an anti-HIV antibody or fragment thereof, comprising (a) obtaining the cultured cells described above, (b) culturing the cells in a medium under conditions that allow expression of the polypeptide encoded by the vector and assembly of an antibody or fragment thereof, and (c) purifying the antibody or fragment from the cultured cells or the medium of the cells.

[0020] In another aspect, the present disclosure also provides a pharmaceutical composition comprising: (i) at least one anti-HIV antibody or antigen-binding portion thereof, nucleic acid, or vector as described above; and (ii) a pharma- ceutically acceptable carrier.

[0021] In some embodiments, the pharmaceutical composition further comprises a second therapeutic agent. In some embodiments, the second therapeutic agent comprises an antiviral agent or one or more additional antibodies. In some embodiments, the one or more additional antibodies comprise a second anti-HIV antibody (e.g., an isolated anti-HIV bNAb disclosed) or an antigen-binding portion thereof, or a third antibody that binds to a third antigen. In some embodiments, the third antigen is CD3, FcγRI (CD64), FcγRII (CD32), FcγRII I (CD16); CD89, CCR5, CD4, CD8, CD28, CD137, CTLA-4, gp41, killer cell immunoglobulin-like receptor, three Ig domains, and long cytoplasmic tail 1 (KIR3DL1), killer cell immunoglobulin-like receptor, three Ig domains, and long cytoplasmic tail 1 (KIR3DL1), killer cell immunoglobulin-like receptor, two Ig domains, and long cytoplasmic tail 1 (KIR2DL1), killer cell Killer cell immunoglobulin-like receptor, two Ig domains and long cytoplasmic tail 3 (KIR2DL3), killer cell lectin-like receptor Cl (KLRC1), killer cell lectin-like receptor C2 (KLRC2), killer cell lectin-like receptor C3 (KLRC3), killer cell lectin-like receptor C4 (KLRC4), killer cell lectin-like receptor D1 (KLRD1), killer cell lectin-like receptor K1 (KLRK1), natural cytotoxicity-inducing receptor 3 (NCR3 or NKp30), natural cytotoxicity-inducing receptor 2 (NCR2 or NK-p44), natural cytotoxicity-inducing receptor 1 (NCR1 or NK-p46), CD226 (DNAM-1), cytotoxic and regulatory T cell molecule (CRTAM or CD355), signaling lymphocyte activation molecule family member 1 (SLAMF1), CD48 (SLAMF2), lymphocyte antigen 9 (LY9 or SLAMF3), CD244 (2B4 or SLAMF 4), CD84 (SLAMF5), SLAM family member 6 (SLAMF6 or NTB-A), SLAM family member 7 (SLAMF7 or CRACC), CD27 (TNFRSF7), semaphorin 4D (SEMA4D or CD100), CD160 (NK1), an immune checkpoint protein (e.g., PD-1, PD-L1, CTLA-4), and a second epitope of gp120.

[0022] In some embodiments, the antiviral agent is selected from the group consisting of a non-nucleoside reverse transcriptase inhibitor, a protease inhibitor, an entry or fusion inhibitor, and an integrase inhibitor.

[0023] In yet another aspect, the disclosure provides a method of preventing or treating an HIV infection or an HIV-related disease, the method comprising: (a) identifying a patient in need of said prevention or treatment, and (b) administering to said patient a first therapeutic agent comprising a therapeutically effective amount of at least one anti-HIV antibody or antigen-binding portion thereof, as described above.

[0024] In some embodiments, the method further comprises administering a second therapeutic agent. In some embodiments, the second therapeutic agent comprises an antiviral agent or one or more additional antibodies. In some embodiments, the one or more additional antibodies comprise a second anti-HIV antibody (e.g., an isolated anti-HIV bNAb disclosed) or an antigen-binding portion thereof, or a third antibody that binds to a third antigen. In some embodiments, the third antigen is CD3, FcγRI (CD64), FcγRII (CD32), FcγRIII (CD16); CD89, CCR5, CD4, CD8, CD28, CD137, CTLA-4, gp41, killer cell immunoglobulin-like receptor, three Ig domains and a long cytoplasmic tail 1 (KIR3DL1), killer cell immunoglobulin-like receptor, three Ig domains and a long cytoplasmic tail 1 (KIR3DL1), killer cell immunoglobulin-like receptor, two Ig domains and a long cytoplasmic tail 1 (KIR2DL1), killer cell immunoglobulin-like receptor, two Ig domains and a long cytoplasmic tail 3 (KIR2DL3), killer cell lectin-like Receptor Cl (KLRC1), killer cell lectin-like receptor C2 (KLRC2), killer cell lectin-like receptor C3 (KLRC3), killer cell lectin-like receptor C4 (KLRC4), killer cell lectin-like receptor D1 (KLRD1), killer cell lectin-like receptor K1 (KLRK1), natural cytotoxicity-inducing receptor 3 (NCR3 or NKp30), natural cytotoxicity-inducing receptor 2 (NCR2 or NK-p44), natural cytotoxicity-inducing receptor 1 (NCR1 or NK-p46), CD226 (DNAM-1), cytotoxic and regulatory T cell molecule (CRTAM or CD355), signaling lymphocyte activation molecule family member 1 (SLAMF1), CD48 (SLAMF2), lymphocyte and a second epitope of gp120.

[0025] In some embodiments, the first therapeutic agent or the second therapeutic agent is administered to the patient intratumorally, intravenously, subcutaneously, intraosseously, orally, transdermally, or sublingually. In some embodiments, the first therapeutic agent is administered to the patient before, after, or simultaneously with the second therapeutic agent.

[0026] In other aspects, the present disclosure further provides a kit comprising a pharma- ceutically effective amount of a pharma- ceutically acceptable dosage unit of at least one isolated anti-HIV antibody, or antigen-binding portion thereof, as described above.

[0027] In some embodiments, the kit further comprises a pharma- ceutically effective amount of a pharma- ceutically acceptable dosage unit of an anti-HIV agent. The two pharma-ceutically acceptable dosage units may be in the form of a single pharma-ceutically acceptable dosage unit. In some embodiments, the anti-HIV agent is selected from the group consisting of a non-nucleoside reverse transcriptase inhibitor, a protease inhibitor, an entry or fusion inhibitor, and an integrase inhibitor.

[0028] The foregoing summary is not intended to define all aspects of the disclosure, and additional aspects are described in other sections, such as the detailed description below. It is understood that the entire document is intended to relate as a unified disclosure, and that all combinations of features described herein are contemplated, even if the combinations of features are not found together in the same sentence, paragraph, or section of this document. Other features and advantages of the present invention will become apparent from the detailed description that follows. However, it should be understood that the detailed description and specific examples, while showing specific embodiments of the disclosure, are given by way of illustration only, since various changes and modifications within the spirit and scope of the disclosure will become apparent to those skilled in the art from this detailed description. [Brief description of the drawings]

[0029] [Figure 1] FIG. 1 is a graph showing HIV-1 neutralization coverage curves of several anti-HIV bNAbs, including the novel mutant HC1 / SAP10. [Diagram 2] FIG. 2 is a graph showing the polyreactivity of several anti-HIV bNAbs measured using a baculovirus-based assay as described in Hotzel, I., et al., 2012, mAbs 4, 753-760. [Diagram 3] FIG. 3 is a series of graphs showing that HC1 / SAP10 has similar biodistribution and half-life in mice as 3BNC117, a bNAb with good half-life and tissue distribution. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0030] Detailed Description of the Invention The present disclosure is based, at least in part, on the unexpected discovery that the disclosed variants of anti-HIV bNAb NIH45-46 exhibit improved biophysical properties while retaining broad and potent neutralizing activity. The disclosed anti-HIV bNAb variants and antigen-binding fragments constitute a novel therapeutic strategy for treating and / or preventing HIV infection.

[0031] A. Novel broadly neutralizing anti-HIV antibodies a.Antibodies The invention disclosed herein includes novel isolated anti-HIV broadly neutralizing antibodies or antigen-binding portions thereof, which have improved biophysical properties, such as reduced polyreactivity and extended half-life.

[0032] In some embodiments, the isolated anti-HIV antibody, or antigen-binding portion thereof, comprises one or more heavy chain substitutions at one or more residues selected from the group consisting of S5, S21, G54, G55, V57, T68, T70, V73, S75, F79, S82, D85, V89, Y97, and P112, where residue numbering is as indicated by Kabat numbering, and is at least 75% (e.g., 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or more) identical to SEQ ID NO:1. a heavy chain having a heavy chain amino acid sequence; or a light chain having one or more light chain substitutions at one or more residues selected from the group consisting of T5, S7, T10, S12, S14, T18, I20, S22, R24, Q27, S28, S30, R59, S61, S63, W65, D68, N70, S72, S74, and S78, and having a light chain amino acid sequence that is at least 75% (e.g., 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or more) identical to SEQ ID NO:2.

[0033] In some embodiments, the isolated anti-HIV antibody or antigen-binding portion thereof has a heavy chain polypeptide sequence that is at least 75% (e.g., 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or more) identical to SEQ ID NO:1, and the heavy chain amino acid sequence comprises one or more heavy chain substitutions at one or more residues selected from the group consisting of S5, S21, G54, G55, V57, T68, T70, V73, S75, F79, S82, D85, V89, Y97, and P112. and a light chain having a light chain amino acid sequence at least 75% (e.g., 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or more) identical to SEQ ID NO:2, wherein the light chain amino acid sequence includes one or more light chain substitutions at one or more residues selected from the group consisting of T5, S7, T10, S12, S14, T18, I20, S22, R24, Q27, S28, S30, R59, S61, S63, W65, D68, N70, S72, S74, and S78.

[0034] In some embodiments, the one or more heavy chain substitutions are: an S5D or S5E substitution, or a conservative substitution of Asp or Glu at residue S5; an S21D or S21E substitution, or a conservative substitution of Asp or Glu at residue S21; a G54W substitution, or a conservative substitution of Trp at residue G54; a G55D or G55E substitution, or a conservative substitution of Asp or Glu at residue G55; a V57D or V57E substitution, or a conservative substitution of Asp or Glu at residue V57; a T68D or T68E substitution, or a conservative substitution of Asp or Glu at residue T68; a T70D or T70E substitution, or a conservative substitution of Asp or Glu at residue T70; a V73D or V73E substitution, or a conservative substitution of Asp or Glu at residue V73. a conservative substitution of Asp or Glu at residue S75; a substitution of S75D or S75E, or a conservative substitution of Asp or Glu at residue S75; a substitution of F79D, F79E, F79Y or F79H, or a conservative substitution of Asp, Glu, Tyr or His at residue F79; a substitution of S82D or S82E, or a conservative substitution of Asp or Glu at residue S82; a substitution of D85E, or a conservative substitution of Asp at residue D85; a substitution of V89D or V89E, or a conservative substitution of Asp or Glu at residue V89; a substitution of Y97D or Y97E, or a conservative substitution of Asp or Glu at residue Y97; a substitution of P112D or P112E, or a conservative substitution of Asp or Glu at residue P112; or a combination thereof.

[0035] In some embodiments, the one or more light chain substitutions are: T5D or T5E, or a conservative substitution of Asp or Glu at residue T5; S7D or S7E, or a conservative substitution of Asp or Glu at residue S7; T10D or T10E, or a conservative substitution of Asp or Glu at residue T10; S12D or S12E, or a conservative substitution of Asp or Glu at residue S12; S14D or S14E, or a conservative substitution of Asp or Glu at residue S14. a substitution of T18D or T18E, or a conservative substitution of Asp or Glu at residue T18; a substitution of I20D or I20E, or a conservative substitution of Asp or Glu at residue I20; a substitution of S22D or S22E, or a conservative substitution of Asp or Glu at residue S22; a substitution of R24D or R24E, or a conservative substitution of Asp or Glu at residue R24; a substitution of Q27D or Q27E, or a conservative substitution of Asp or Glu at residue Q27; a substitution of S28Y or S28H, or a conservative substitution of Asp or Glu at residue a conservative substitution of Tyr or His at S28; a substitution of S30D or S30E, or a conservative substitution of Asp or Glu at residue S30; a substitution of R59D or R59E, or a conservative substitution of Asp or Glu at residue R59; a substitution of S61D or S61E, or a conservative substitution of Asp or Glu at residue S61; a substitution of S63D or S63E, or a conservative substitution of Asp or Glu at residue S63; a substitution of W65D, W65E, W65Y, or W65H, or a conservative substitution of Asp, G at residue W65 a conservative substitution of lu, Tyr or His; a D68E substitution, or a conservative substitution of Asp at residue D68; a N70D or N70E substitution, or a conservative substitution of Asp or Glu at residue N70; a S72D or S72E substitution, or a conservative substitution of Asp or Glu at residue S72; a S74D or S74E substitution, or a conservative substitution of Asp or Glu at residue S74; a S78D or S78E substitution, or a conservative substitution of Asp or Glu at residue S78; or a combination thereof.

[0036] In some embodiments, the one or more heavy chain substitutions further comprise the following substitutions: M428L and N434S.

[0037] In some embodiments, the light chain amino acid sequence comprises one or more of the following light chain substitutions: (i) Q27E, S28H, and S30D; (ii) Q27E, S28H, S30D, and S74T; (iii) Q27E, S28H, S30D, S74T, M428L, and N434S; (iv) Q27E, S28Y, and S30D; (v) Q27D and S28H; (vi) S28Y; (vii) T5D, T10D, S12D, S14D, I20D, and S22D; or (viii) S61D, S63D, W65D, N70D, S72D, and S74D.

[0038] In some embodiments, the heavy chain amino acid sequence comprises one or more heavy chain substitutions comprising G54W and T68D; and the light chain amino acid sequence comprises one or more light chain substitutions: (a) Q27E, S28H, S30D, and S74T; (b) Q27E, S28H, S30D, S74T, M428L, and N434S; (c) Q27E, S28Y, and S30D; (d) Q27D and S28H; or (e) S28Y.

[0039] In some embodiments, the heavy chain amino acid sequence comprises one or more heavy chain substitutions including G54W; and the light chain amino acid sequence comprises one or more light chain substitutions: (a) Q27E, S28H, and S30D; (b) Q27E, S28Y, and S30D; or (c) Q27D and S28H.

[0040] In some embodiments, the heavy chain amino acid sequence comprises the amino acid sequence of SEQ ID NO: 1 or SEQ ID NO: 3. In some embodiments, the light chain amino acid sequence comprises the amino acid sequences of SEQ ID NO: 2 and SEQ ID NOs: 4-10.

[0041] In some embodiments, the heavy chain amino acid sequence comprises the amino acid sequences of SEQ ID NO: 3 to SEQ ID NO: 35. In some embodiments, the light chain amino acid sequence comprises the amino acid sequences of SEQ ID NO: 2 and SEQ ID NO: 36 to SEQ ID NO: 80.

[0042] In some embodiments, the isolated anti-HIV antibody comprises CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 comprising the respective sequences set forth in the set of CDR sequences of SEQ ID NOs: 81, 84, 87, 90, 93, and 96; SEQ ID NOs: 81, 84, 87, 99, 93, and 96; SEQ ID NOs: 81, 84, 87, 100, 93, and 96; SEQ ID NOs: 81, 84, 87, 101, 93, and 96; or SEQ ID NOs: 81, 84, 87, 102, 93, and 96; (as set forth in Kabat numbering); SEQ ID NOs: 82, 85, 88, 91, 94, and 97 (as set forth in IMGT numbering); or SEQ ID NOs: 83, 86, 89, 92, 95, and 98 (as set forth in Chothia numbering).

[0043] In some embodiments, the heavy chain amino acid sequences and the light chain amino acid sequences comprise the respective amino acid sequences set forth in a set of sequences selected from the group consisting of SEQ ID NOs: 3 and 36; SEQ ID NOs: 3 and 38; SEQ ID NOs: 3 and 39; SEQ ID NOs: 4 and 2; SEQ ID NOs: 4 and 38; SEQ ID NOs: 4 and 39; SEQ ID NOs: 4 and 41; and SEQ ID NOs: 34 and 37.

[0044] [Table 1-1]

[0045] [Table 1-2]

[0046] [Table 1-3]

[0047]

Table 1-4

[0048]

Table 1-5

[0049]

Table 1-6

[0050]

Table 1-7

[0051]

Table 1-8

[0052]

Table 1-9

[0053]

Table 1-10

[0054]

Table 1-11

[0055]

Table 1-12

[0056]

Table 1-13

[0057] [Table 1-14]

[0058] [Table 1-15]

[0059] [Table 1-16]

[0060] [Table 1-17]

[0061] [Table 1-18]

[0062] [Table 1-19]

[0063] [Table 1-20]

[0064] In some embodiments, the antibody or antigen-binding fragment thereof further comprises a variant Fc constant region. In some embodiments, the antibody is a monoclonal antibody. In some embodiments, the antibody is a chimeric antibody, a humanized antibody, or a humanized monoclonal antibody. The antibody is a single chain antibody, a monovalent antibody, a single chain variable fragment (scFv), an scFv-Fc fusion, a minibody, a Fab, or a F(ab')2 fragment.

[0065] In some embodiments, the antibody or antigen-binding fragment thereof may be detectably labeled or conjugated to a toxin, therapeutic agent, polymer (e.g., polyethylene glycol (PEG)), receptor, enzyme, or receptor ligand. For example, an antibody of the invention may be conjugated to a toxin (e.g., tetanus toxin).

[0066] In other examples, the antibodies of the present invention may be conjugated to a detectable tag. Such antibodies may be used in diagnostic assays to determine whether an animal, such as a human, is infected with HIV-1. Examples of detectable tags include fluorescent proteins (i.e., green fluorescent protein, red fluorescent protein, yellow fluorescent protein), fluorescent markers (i.e., fluorescein isothiocyanate, rhodamine, Texas Red), radioactive labels (i.e., 3H, 32P, 125I), enzymes (i.e., β-galactosidase, horseradish peroxidase, β-glucuronidase, alkaline phosphatase), or affinity tags (i.e., avidin, biotin, streptavidin, Fc). Methods for conjugating antibodies to detectable tags are known in the art. Harlow et al., Antibodies: A Laboratory Manual, page 319 (Cold Spring Harbor Pub. 1988).

[0067] b. Fragment In some embodiments, the antibodies provided herein are antibody fragments. Antibody fragments include, but are not limited to, Fab, Fab', Fab'-SH, F(ab')2, Fv, and single-chain Fv (scFv) fragments, as well as other fragments described below, such as diabodies, triabodies, tetrabodies, and single domain antibodies. For a review of specific antibody fragments, see Hudson et al., Nat. Med. 9:129-134 (2003). For a review of scFv fragments, see, e.g., Pluckthun, in The Pharmacology of Monoclonal Antibodies, 1999, 14:131-134 (2003). dies, vol. 113, Rosenburg and Moore eds., (Springer-Verlag, New York), pp. 269-315 (1994); see also WO 93 / 16185; and U.S. Patent Nos. 5,571,894 and 5,587,458. For a discussion of Fab and F(ab')2 fragments that contain salvage receptor binding epitope residues and have increased in vivo half-lives, see U.S. Patent No. 5,869,046.

[0068] Diabodies are antibody fragments that have two bivalent or bispecific antigen-binding sites. See, e.g., EP 404,097; WO 1993 / 01161; Hudson et al., Nat. Med. 9:129-134 (2003); and Hollinger et al., Proc. Natl. Acad. Sci. USA 90: 6444-6448 (1993). Triabodies and tetrabodies are also described in Hudson et al., Nat. Med. 9:129-134 (2003).

[0069] Single domain antibodies are antibody fragments that contain all or a portion of the heavy chain variable region or all or a portion of the light chain variable region of an antibody. In some embodiments, single domain antibodies are human single domain antibodies (DOMANTIS, Inc., Waltham, Mass.; see, e.g., U.S. Patent No. 6,248,516).

[0070] Antibody fragments can be produced by a variety of techniques, including, but not limited to, proteolytic digestion of intact antibodies and production by recombinant host cells (e.g., E. coli or phages), as described herein.

[0071] c. Chimeric and humanized antibodies In some embodiments, the antibody provided herein is a chimeric antibody. Certain chimeric antibodies are described, for example, in U.S. Patent No. 4,816,567; and Morrison et al., Proc. Natl. Acad. Sci. USA, 81:6851-6855 (1984). In one example, a chimeric antibody comprises a non-human variable region (e.g., a variable region derived from a non-human primate, such as a mouse, rat, hamster, rabbit, or monkey) and a human constant region. In a further example, a chimeric antibody is a "class-switched" antibody whose class or subclass is changed from that of the parent antibody. Chimeric antibodies include antigen-binding fragments thereof.

[0072] In some embodiments, the chimeric antibody is a humanized antibody. Typically, a non-human antibody is humanized to reduce immunogenicity to humans while retaining the specificity and affinity of the parent non-human antibody. Usually, a humanized antibody comprises one or more variable regions in which the HVR, e.g., CDR (or a portion thereof) is derived from a non-human antibody and the FR (or a portion thereof) is derived from a human antibody sequence. The humanized antibody may also comprise at least a portion of a human constant region. In some embodiments, some FR residues of the humanized antibody are replaced with the corresponding residues from the non-human antibody (e.g., the antibody from which the HVR residues are derived), e.g., to restore or improve the specificity or affinity of the antibody.

[0073] Humanized antibodies and methods for producing them are reviewed, e.g., in Almagro and Fransson, Front. Biosci. 13:1619-1633 (2008), and further described, e.g., in Riechmann et al., Nature 332:323-329 (1988); Queen et al., Proc. Nat'l Acad. Sci. USA 86:10029-10033 (1989); U.S. Patent Nos. 5,821,337, 7,527,791, 6,982,321, and 7,087,409; Kashmiri et al., Met. Methods 36:25-34 (2005) (describing specificity determining region (SDR) grafting); Padlan, Mol. Immunol. 28:489-498 (1991) (describing "resurfacing"); Dall'Acqua et al., Methods 36:43-60 (2005) (describing "FR shuffling"); and Osbourn et al., Methods 36:61-68 (2005) and Klimka et al., Br. J. Cancer, 83:252-260 (2000) (describing a "guided selection" approach to FR shuffling).

[0074] Human framework regions that can be used for humanization include framework regions selected using the "best fit" method (see, e.g., Sims et al. J. Immunol. 151:2296 (1993)); framework regions derived from the consensus sequence of human antibodies of a particular subgroup of light or heavy chain variable regions (see, e.g., Carter et al. Proc. Natl. Acad. Sci. USA, 89:4285 (1992); and Presta et al. J. Immunol., 151:2623 (1993); human mature (somatically mutated) framework regions or human germline framework regions (see, e.g., Almagro and Fransson, Front. Biosci. 13:1619-1633 (2008)); as well as framework regions obtained from screening of FR libraries (see, e.g., Baca et al., J. Biol. Chem. 272:10678-10684 (1997) and Rosok et al., J. Biol. Chem. 271:22611-22618 (1996)).

[0075] D. Human antibodies In some embodiments, the antibody provided herein is a human antibody. Human antibodies can be produced using various techniques known in the art or using the techniques described herein. Human antibodies are generally described in van Dijk and van de Winkel, Curr. Opin. Pharmacol. 5: 368-74 (2001) and Lonberg, Curr. Opin. Immunol. 20:450-459 (2008).

[0076] Human antibodies can be prepared by administering immunogens to intact human antibodies or transgenic animals that have been modified to produce intact antibodies with human variable regions in response to antigen challenge. Such animals typically contain all or part of a human immunoglobulin locus that replaces the endogenous immunoglobulin locus or is present extrachromosomally or randomly integrated into the animal's chromosomes. In such transgenic mice, the endogenous immunoglobulin locus is usually inactivated. For a review of methods for obtaining human antibodies from transgenic animals, see Lonberg, Nat. Biotech. 23:1117-1125 (2005). See, further, for example, U.S. Patent Nos. 6,075,181 and 6,150,584, which describe the XENOMOUSE technology, U.S. Patent No. 5,770,429, which describes the HUMAB technology, U.S. Patent No. 7,041,870, which describes the KM MOUSE technology, and U.S. Provisional Patent Application No. 2007 / 0061900, which describes the VELOCIMOUSE technology. The human variable regions from intact antibodies produced by such animals can be further modified, for example, by combining with a different human constant region.

[0077] Human antibodies can also be produced by hybridoma-based methods. Human myeloma and mouse-human heteromyeloma cell lines for the production of human monoclonal antibodies have been described. (See, e.g., Kozbor J. Immunol., 133: 3001 (1984); Brodeur et al., Monoclonal Antibody Production Techniques and Applications, pp. 51-63 (Marcel Dekker, Inc., New York, 1987); and Boerner et al., J. Immunol., 147: 86 (1991). Human antibodies produced via human B cell hybridoma technology are also described in Li et al., Proc. Natl. Acad. Sci. USA, 103:3557-3562 (2006). Additional methods include those described, for example, in U.S. Pat. No. 7,189,826 (which describes the production of monoclonal human IgM antibodies from hybridoma cell lines), and Ni, Xiandai Mianyixue, 26(4):265-268 (2006) (which describes human-human hybridomas). Human hybridoma technology (trioma technology) is described in Vollmers and Brandlein, Histology and Histopathology, 20(3):927-937 (2005), and Vollmers and Brandlein, Methods and Findings in Experimental and Clinical Pharmacology, 27(3):185-91 (2005).

[0078] Human antibodies can also be produced by isolating Fv clone variable region sequences selected from a human-derived phage display library. Such variable region sequences can then be combined with the desired human constant regions. Techniques for selecting human antibodies from an antibody library are described below.

[0079] The antibodies of the present invention can be isolated by screening combinatorial libraries for antibodies with the desired activity or activities. For example, various methods are known in the art for generating phage display libraries and screening such libraries for antibodies with the desired binding properties. Such methods are described, for example, in Hoogenboom et al., in Methods in Molecular Biology 178:1-37 (O'Brien et al., ed., Human Press, Totowa, NJ, 2001) and further discussed, for example, in the McCafferty et al., Nature 348:552-554; Clackson et al., Nature 352: 624-628 (1991); Marks et al., J. Mol. Biol. 222: 581-597 (1992); Marks and Bradbury, in Methods in Molecular Biology 248:161-175 (Lo, ed., Human Press, Totowa, NJ, 2003); Sidhu et al., J. Mol. Biol. 338(2): 299-310 (2004); Lee et al., J. Mol. Biol. 340(5): 1073-1093 (2004); Fellouse, Proc. Natl. Acad. Sci. USA 101(34): 12467-12472 (2004); and Lee et al., J. Immunol. Methods 284(1-2): 119-132 (2004).

[0080] In certain phage display methods, repertoires of VH and VL genes can be separately cloned by polymerase chain reaction (PCR), randomly recombined in phage libraries, and then screened for antigen-binding phages, as described in Winter et al., Ann. Rev. Immunol., 12: 433-455 (1994). Phages are usually expressed as scFv fragments. The antibody fragments are shown as either fragments or Fab fragments. Libraries from immune sources provide high affinity antibodies to immunogens without the need to construct hybridomas. Alternatively, naive repertoires can be cloned (e.g., from humans) to provide a single source of antibodies to a wide range of non-self and self antigens without immunization, as described in Griffiths et al., EMBO J, 12: 725-734 (1993). Finally, naive libraries can also be synthetically produced by cloning unrearranged V gene segments from stem cells and using PCR primers containing random sequences to encode highly variable CDR3 regions and achieve rearrangement in vitro, as described in Hoogenboom and Winter, J. Mol. Biol., 227: 381-388 (1992). Patent publications describing human antibody phage libraries include, for example, U.S. Patent No. 5,750,373, and U.S. Provisional Patent Applications Nos. 2005 / 0079574, 2005 / 0119455, 2005 / 0266000, 2007 / 0117126, 2007 / 0160598, 2007 / 0237764, 2007 / 0292936, and 2009 / 0002360. Antibodies or antibody fragments isolated from a human antibody library are considered human antibodies or human antibody fragments herein.

[0081] e. Mutants In some embodiments, amino acid sequence variants of the antibodies provided herein are contemplated. For example, it may be desirable to improve the binding affinity and / or other biological properties of the antibody (e.g., reducing polyreactivity, increasing half-life, etc.). Amino acid sequence variants of the antibody can be prepared by introducing appropriate modifications into the nucleotide sequence encoding the antibody or by peptide synthesis. Such modifications include, for example, deletions from, and / or insertions and / or substitutions of residues within the amino acid sequence of the antibody. Any combination of deletions, insertions, and substitutions can be made to arrive at the final construct, provided that the final construct has the desired properties, e.g., antigen binding.

[0082] Substitution, insertion, and deletion mutants In some embodiments, antibody variants are provided that have one or more amino acid substitutions. The sites of substitution mutagenesis include HVR and FR. Conservative substitutions are defined herein. Amino acid substitutions can be introduced into the antibody of interest, and the products are screened for desired activity, such as retaining / improving antigen binding, reducing immunogenicity, or improving antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC).

[0083] Thus, the antibodies of the present invention may contain one or more conservative modifications of the CDRs, heavy chain variable regions, or light variable regions described herein. A conservative modification or functional equivalent of a peptide, polypeptide, or protein disclosed in the present invention refers to a polypeptide derivative of the peptide, polypeptide, or protein, e.g., a protein having one or more point mutations, insertions, deletions, truncations, fusion proteins, or combinations thereof. It substantially retains the activity of the parent peptide, polypeptide, or protein (such as those disclosed in the present invention). Typically, a conservative modification or equivalent function is at least 60% (e.g., any number between 60% and 100%, e.g., 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, and 99%) identical to the parent. Thus, antibodies having heavy chain or light variable regions and mutated regions having one or more point mutations, insertions, deletions, truncations, fusion proteins, or combinations thereof are within the scope of the present invention.

[0084] As used herein, the percent homology between two amino acid sequences is equivalent to the percent identity between two sequences.The percentage of identity between two sequences is a function of the number of identical positions shared by the sequences, taking into account the number of gaps and the length of each gap that need to be introduced for optimal alignment of the two sequences (i.e., % homology=number of identical positions / total number of positions×100).As described in the following non-limiting examples, the comparison of sequences and the determination of the percent identity between two sequences can be achieved using mathematical algorithms.

[0085] The percent identity between two amino acid sequences can be determined using the algorithm of E. Meyers and W. Miller (Comput. Appl. Biosci., 4:11-17 (1988)) as incorporated into the ALIGN program (version 2.0) using a weight residual table of PAM120, a gap length penalty of 12, and a gap penalty of 4. Additionally, the percent identity between two amino acid sequences can be determined using the algorithm of Needleman and Wunsch (J. Mol. Biol. 48:444-453 (1988)) as incorporated into the GAP program of the GCG software package (available at www.gcg.com) using either a Blossum 62 matrix or a PAM250 matrix, and gap weights of 16, 14, 12, 10, 8, 6, or 4, and length weights of 1, 2, 3, 4, 5, or 6. (1970)) algorithm.

[0086] Additionally or alternatively, the protein sequences of the present invention can be further used as a "query sequence" to perform searches against public databases, for example to identify related sequences. Such searches can be performed using the XBLAST program (version 2.0) of Altschul, et al. (1990) J. Mol. Biol. 215:403-10. BLAST protein searches can be performed with the XBLAST program, score=50, wordlength=3, to obtain amino acid sequences homologous to the antibody molecules of the present invention. To obtain gapped alignments for comparison purposes, gapped BLAST can be used as described in Altschul et al., (1997) Nucleic Acids Res. 25(17):3389-3402. When using BLAST and gapped BLAST programs, the default parameters of the respective programs (e.g., XBLAST and NBLAST) can be used. (See www.ncbi.nlm.nih.gov).

[0087] As used herein, the term "conservative modification" or "conservative substitution" refers to an amino acid modification that does not significantly affect or change the binding properties of an antibody containing an amino acid sequence. Such conservative modifications include amino acid substitutions, additions, and deletions. Modifications can be introduced into the antibody of the present invention by standard techniques known in the art, such as site-directed mutagenesis and PCR-mediated mutagenesis. Conservative amino acid substitutions are those in which an amino acid residue is replaced with an amino acid residue having a similar side chain. Families of amino acid residues having similar side chains have been defined in the art. These families include: (i) amino acids with basic side chains (e.g., lysine, arginine, histidine), (ii) acidic side chains (e.g., aspartic acid, glutamic acid), (iii) uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine, tryptophan), (iv) nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine), (v) beta-branched side chains (e.g., threonine, valine, isoleucine), and (vi) aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine).

[0088] Alternatively, see Lehninger [Biochemistry, Second Edition] tion; Worth Publishers, Inc. NY, NY (1 975), pp. 71-77], conservative amino acids may be grouped as follows:

[0089] [Table 2]

[0090] As yet another alternative, exemplary conservative substitutions are shown below:

[0091] [Table 3]

[0092] Conservative substitution of an existing substitution refers to conservative substitution of the substituted residue. For example, conservative substitution of Trp at residue G54 (or position 54) refers to conservative substitution of Trp at position 54, i.e., Tyr (Y). Similarly, conservative substitution of Tyr at residue S30 (or position 30) refers to conservative substitution of Tyr at position 30, i.e., Trp, Phe, Thr, Ser.

[0093] Non-conservative substitutions entail the exchange of a member of one of these classes for another class.

[0094] Exemplary substitutional variants are affinity matured antibodies, which may be conveniently produced using phage display-based affinity maturation techniques, for example, as described in Hoogenboom et al., in Methods in Molecular Biology 178:1-37 (O'Brien et al., ed., Human Press, Totowa, NJ, (2001). Amino acid sequence insertions include amino- and / or carboxyl-terminal fusions of lengths ranging from one residue to polypeptides containing 100 or more residues, as well as intrasequence insertions of single or multiple amino acid residues. An example of a terminal insertion is an antibody with an N-terminal methionyl residue. Other insertional variants of the antibody molecule include the fusion to the N- or C-terminus of the antibody to an enzyme (e.g., in the case of ADEPT) or a polypeptide which increases the serum half-life of the antibody. Some examples include:

[0095] Glycosylation variants In some embodiments, the antibodies provided herein are altered to increase or decrease the extent to which the antibody is glycosylated. Addition or deletion of glycosylation sites to an antibody can be conveniently accomplished by altering the amino acid sequence such that one or more glycosylation sites are created or removed.

[0096] For example, aglycosylated antibodies can be produced (i.e., the antibody lacks glycosylation). Glycosylation can be altered, for example, to increase the affinity of the antibody for the antigen. Such carbohydrate modifications can be accomplished, for example, by altering one or more sites of glycosylation within the antibody sequence. For example, one or more amino acid substitutions can be made that result in the elimination of one or more variable region framework glycosylation sites, thereby eliminating glycosylation at that site. Such glycosylation can increase the affinity of the antibody for the antigen. Such approaches are described in further detail in U.S. Pat. Nos. 5,714,350 and 6,350,861 by Co et al. Glycosylation of the constant region on N297 can be inhibited by mutating the N297 residue to another residue, e.g., N297A, and / or by mutating the adjacent amino acid, e.g., 298, thereby reducing glycosylation on N297.

[0097] Additionally or alternatively, antibodies may be produced that have an altered type of glycosylation, such as hypofucosylated antibodies with reduced amounts of fucosyl residues or antibodies with increased bisecting GlcNac structures. Such altered glycosylation patterns have been demonstrated to enhance the ADCC ability of antibodies. Such carbohydrate modifications may be achieved, for example, by expressing the antibody in a host cell with altered glycosylation machinery. Cells with altered glycosylation machinery have been described in the art and may be used as host cells to express the recombinant antibodies described herein, thereby producing antibodies with altered glycosylation. For example, EP 1,176,195 by Hanai et al. describes a cell line with a functionally disrupted FUT8 gene encoding a fucosyltransferase such that antibodies expressed in such cell lines exhibit hypofucosylation. PCT application WO 2006 / 023366 by Presta describes a method for the production of a fucosylated antibody comprising the steps of: 03 / 035835 describes that Led3 cells, a mutant of a Chinese hamster ovary cell line, have a reduced ability to attach fucose to Asn(297)-linked carbohydrates, which also leads to hypofucosylation of antibodies expressed in the host cells (see also Shields, RL et al. (2002) J. Biol. Chem. 277:26733-26740). Umana et al., PCT application WO 99 / 54342 describes that cell lines are engineered to express glycoprotein-modifying glycosyltransferases (e.g., beta(1,4)-N-acetylglucosaminyltransferase III (GnTIII)) and that antibodies expressed in the engineered cell lines exhibit increased bisecting GlcNac structures, resulting in increased ADCC activity of the antibodies (see further Umana et al. (1999) Nat. Biotech. 17: 176-180).

[0098] Fc Region Mutations The variable regions of the antibodies described herein can be linked (e.g., covalently bound or fused) to Fc, e.g., IgG1 Fc, IgG2 Fc, IgG3 Fc, or IgG4 Fc, which can be of any allotype or isoallotype, e.g., for IgG1: Glm, Glml(a), Glm2(x), Glm3(f), Glml7(z); for IgG2: G2m, G2m23(n); for IgG3: G3m, G3m21(gl), G3m28(g5), G3ml l(b0), G3m5(bl), G3ml3(b3), G3ml4(b4), G3ml0(b5), G3ml5( s), G3ml6(t), G3m6(c3), G3m24(c5), G3m26(u), G3m27(v); for K: Km, Kml, Km2, Km3, (see, e.g., Jefferies et al. (2009) mAbs 1: 1). In some embodiments, the antibody variable regions described herein can be linked to Fc that binds to one or more activating Fc receptors (FcγI, Fcγlla, or FcγIIIa), thereby stimulating ADCC and causing T cell depletion. In some embodiments, the antibody variable regions described herein are linked to Fc that causes depletion.

[0099] In some embodiments, the antibody variable regions described herein may be linked to an Fc that includes one or more modifications, typically to alter one or more functional properties of the antibody, such as serum half-life, complement binding, Fc receptor binding, and / or antigen-dependent cellular cytotoxicity. Additionally, the antibodies described herein may be modified chemically (e.g., one or more chemical moieties may be attached to the antibody) or to alter its glycosylation, to alter one or more functional properties of the antibody. The numbering of the residues in the Fc region is that of the EU index of Kabat.

[0100] The Fc region comprises a domain derived from the constant region of an immunoglobulin, preferably a human immunoglobulin, including fragments, analogs, mutants, variants or derivatives of the constant region. Suitable immunoglobulins include IgG1, IgG2, IgG3, IgG4, and other classes, such as IgA, IgD, IgE, and IgM. The constant region of the immunoglobulin is defined as a natural or synthetically produced polypeptide that is homologous to the C-terminal region of an immunoglobulin and may comprise a CH1 region, a hinge, a CH2 region, a CH3 region, or a CH4 region, either individually or in combination. In some embodiments, the antibody of the present invention has an Fc region other than wild-type IgA1. The antibody may have an Fc region from IgG (e.g., IgG1, IgG2, IgG3, and IgG4) or from other classes, such as IgA2, IgD, IgE, and IgM. The Fc may be a variant of IgA1.

[0101] The constant region of the immunoglobulin is responsible for many important antibody functions, such as Fc receptor (FcR) binding and complement fixation. There are five major classes of heavy chain constant regions, classified as IgA, IgG, IgD, IgE, and IgM, each with a characteristic effector function designated by its isotype. For example, IgG is divided into four subclasses known as IgG1, IgG2, IgG3, and IgG4.

[0102] Ig molecules interact with multiple classes of cell receptors. For example, IgG molecules interact with three classes of Fcγ receptors (FcγR), namely FcγRI, FcγRII, and FcγRIIL, which are specific for IgG class antibodies. It has been reported that the important sequences for IgG binding to FcγR receptors are located in the CH2 and CH3 regions. The serum half-life of an antibody is influenced by its ability to bind to FcR.

[0103] In some embodiments, the Fc region is a variant Fc region, such as an Fc sequence that has been modified (e.g., by amino acid substitution, deletion, and / or insertion) relative to a parent Fc sequence (e.g., an unmodified Fc polypeptide that is subsequently modified to generate the variant) to provide desired structural features and / or biological activity. For example, alterations may be made to the Fc region to generate Fc variants that have (a) increased or decreased ADCC, (b) increased or decreased CDC, (c) increased or decreased affinity for Clq, and / or (d) increased or decreased affinity for Fc receptors compared to the parent Fc. Such Fc region variants will typically include at least one amino acid modification in the Fc region. Combining amino acid modifications is believed to be particularly desirable. For example, the variant Fc region may comprise 2, 3, 4, 5, etc. substitutions therein, e.g., at the specific Fc region positions identified herein.

[0104] The variant Fc region may also include sequence changes in which amino acids involved in disulfide bond formation are removed or replaced with other amino acids. Such removal may avoid reactions with other cysteine-containing proteins present in the host cell used to produce the antibodies described herein. Even when cysteine ​​residues are removed, the single-chain Fc region may form a non-covalently bound dimeric Fc region. In some embodiments, the Fc region may be modified to increase compatibility with a selected host cell. For example, the PA sequence near the N-terminus of a typical native Fc region may be removed, which may be recognized by E. coli digestive enzymes such as proline iminopeptidase. In other embodiments, one or more glycosylation sites within the Fc region may be removed. Residues that are normally glycosylated (e.g., asparagine) may trigger a cytolytic response. Such residues may be removed or replaced with non-glycosylated residues (e.g., alanine). In other embodiments, sites involved in interactions with complement, such as the Clq binding site, may be removed from the Fc region. For example, the EKK sequence of human IgG1 may be removed or replaced. In some embodiments, sites that affect binding to Fc receptors, preferably sites other than the salvage receptor binding site, may be removed. In other embodiments, the Fc region may be modified to remove ADCC sites. ADCC sites are known in the art; see, for example, Molec. Immunol. 29 (5): 633-9 (1992) for IgG1 ADCC sites. Particular examples of Fc regions are disclosed, for example, in WO 97 / 34631 and WO 96 / 32478.

[0105] In one embodiment, the Fc hinge region is modified such that the number of cysteine ​​residues in the hinge region is altered, e.g., increased or decreased. This approach is further described in U.S. Patent No. 5,677,425 by Bodmer et al. The number of cysteine ​​residues in the Fc hinge region is altered, e.g., to facilitate assembly of the light and heavy chains or to increase or decrease the stability of the antibody. In one embodiment, the Fc hinge region of the antibody is mutated to reduce the biological half-life of the antibody. More specifically, one or more amino acid mutations are introduced into the CH2-CH3 region interface region of the Fc hinge fragment to render the antibody weaker in Staphylococcus protein A (SpA) binding compared to native Fc hinge region SpA binding. This approach is further described in U.S. Patent No. 6,165,745 by Ward et al.

[0106] In yet another embodiment, the Fc region is modified by replacing at least one amino acid residue with a different amino acid residue to modify the effector function of the antibody. For example, one or more amino acids selected from amino acid residues 234, 235, 236, 237, 297, 318, 320 and 322 may be replaced with a different amino acid residue so that the antibody has a modified affinity for the effector ligand but retains the antigen-binding ability of the parent antibody. The effector ligand with altered affinity may be, for example, an Fc receptor or the CI component of complement. This approach is described in more detail in U.S. Patent Nos. 5,624,821 and 5,648,260 by Winter et al.

[0107] In other examples, one or more amino acids selected from amino acid residues 329, 331, and 322 may be replaced with a different amino acid residue such that the antibody has altered Clq binding or reduced or abolished CDC. This approach is described in further detail in U.S. Patent No. 6,194,551 by Idusogie et al.

[0108] In other examples, one or more amino acid residues within amino acid positions 231 and 239 are mutated. The antibody may be modified to bind complement, thereby altering its ability to fix complement. This approach is described in detail in PCT application WO 94 / 29351 by Bodmer et al.

[0109] In yet other examples, the Fc region can be modified to increase ADCC and / or to increase affinity for Fcγ receptors by modifying one or more amino acids at the following positions: 234, 235, 236, 238, 239, 240, 241, 243, 244, 245, 247, 248, 249, 252, 254, 255, 256, 258, 262, 263, 264, 265, 267, 268, 269, 270, 272, 276, 278, 280, 283 , 285, 286, 289, 290, 292, 293, 294, 295, 296, 298, 299, 301, 303, 305, 307, 309, 312, 313, 315, 320, 322, 324, 325, 326, 327, 329, 330, 331, 332, 333, 334, 335, 337, 338, 340, 360, 373, 376, 378, 382, ​​388, 389, 398, 414, 416, 419, 430, 433, 434, 435, 436, 437, 438 or 439. Exemplary substitutions include 236A, 239D, 239E, 268D, 267E, 268E, 268F, 324T, 332D, and 332E. Exemplary mutations include 239D / 332E, 236A / 332E, 236A / 239D / 332E, 268F / 324T, 267E / 268F, 267E / 324T, and 267E / 268F / 324T. Other modifications to enhance FcγR and complement interactions include, but are not limited to, the following substitutions: 298A, 333A, 334A, 326A, 247I, 339D, 339Q, 280H, 290S, 298D, 298V, 243L, 292P, 300L, 396L, 305I, and 396L. These and other modifications are reviewed in Strohl, 2009, Current Opinion in Biotechnology 20:685-691.

[0110] Fc modifications that increase binding to Fcγ receptors include modifications of one or more of the following amino acids in the Fc region: 238, 239, 248, 249, 252, 254, 255, 256, 258, 265, 267, 268, 269, 270, 272, 279, 280, 283, 285, 298, 289, 290, 292, 293, 294, 295, 296, 298, 301, 303, 305, 307, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 358, 359, 360, 361, 362, 363, 364, 365, 366, 367, 368, 369, 270, 272, 279, 280, 283, 285, 298, 2 , 315, 324, 327, 329, 330, 335, 337, 3338, 340, 360, 373, 376, 379, 382, ​​388, 389, 398, 414, 416, 419, 430, 434, 435, 437, 438 or 439, where the numbering of the residues in the Fc region is that of the EU index as in Kabat (WO 00 / 42072).

[0111] Other Fc modifications that can be made to the Fc are to reduce or eliminate binding to FcγR and / or complement proteins, thereby reducing or eliminating Fc-mediated effector functions such as ADCC, antibody-dependent cellular cytotoxicity (ADCP), and CDC. Exemplary modifications include, but are not limited to, substitutions, insertions, and deletions at positions 234, 235, 236, 237, 267, 269, 325, and 328, numbered according to the EU index. Exemplary substitutions include, but are not limited to, 234G, 235G, 236R, 237K, 267R, 269R, 325L, and 328R. Fc variants can include 236R / 328R. Other modifications to reduce FcγR and complement interactions include substitutions at 297A, 234A, 235A, 237A, 318A, 228P, 236E, 268Q, 309L, 330S, 331S, 220S, 226S, 229S, 238S, 233P, and 234V, as well as modifications to reduce FcγR and complement interactions by mutational or enzymatic means, or in organisms such as bacteria that do not glycosylate proteins. These and other modifications are reviewed in Strohl, 2009, Current Opinion in Biotechnology 20:685-691.

[0112] Optionally, the Fc region may include non-naturally occurring amino acid residues at additional and / or alternative positions known to those of skill in the art (e.g., U.S. Patent Application Nos. 5,624,821; 6,277,375; 6,737,056; 6,194,551; 7,317,091; 8,101,720; WO00 / 42072; WO01 / 58957; WO02 / 06919; WO04 / 016750; WO04 / 029207; WO04 / 035752; WO04 / 074455; WO04 / 099249; WO04 / 063351; WO05 / 070963; (see WO05 / 040217, WO05 / 092925, and WO06 / 020114).

[0113] Fc variants that enhance affinity to the inhibitory receptor FcγRIIb can also be used. Such variants can provide Fc fusion proteins with immunomodulatory activity associated with FcγRIIb cells, including, for example, B cells and monocytes. In some embodiments, the Fc variants provide selectively enhanced affinity to FcγRIIb compared to one or more activating receptors. Modifications for altering binding to FcγRIIb include one or more modifications at positions selected from the group consisting of 234, 235, 236, 237, 239, 266, 267, 268, 325, 326, 327, 328, and 332 according to the EU index. Exemplary substitutions for enhancing FcγRIIb affinity include, but are not limited to, 234D, 234E, 234F, 234W, 235D, 235F, 235R, 235Y, 236D, 236N, 237D, 237N, 239D, 239E, 266M, 267D, 267E, 268D, 268E, 327D, 327E, 328F, 328W, 328Y, and 332E. Exemplary substitutions include 235Y, 236D, 239D, 266M, 267E, 268D, 268E, 328F, 328W, and 328Y. Other Fc mutants for enhancing binding to FcγRllb include 235Y / 267E, 236D / 267E, 239D / 268D, 239D / 267E, 267E / 268D, 267E / 268E, and 267E / 328F.

[0114] The affinity and binding properties of an Fc region for its ligand can be determined by a variety of in vitro assay methods (biochemical or immunological based assays) known in the art, including but not limited to equilibrium methods (e.g., ELISA, or radioimmunoassay), or kinetics (e.g., BIACORE analysis), as well as other methods such as indirect binding assays, competitive inhibition assays, fluorescence resonance energy transfer (FRET), gel electrophoresis and chromatography (e.g., gel filtration). These and other methods may utilize one or more labels of the component being examined and / or employ a variety of detection methods, including but not limited to chromogenic, fluorescent, luminescent, or isotopic labels. A detailed description of binding affinity and kinetics can be found in Paul, WE, ed., Fundamental Immunology, 4th Ed., Lippincott-Raven, Philadelphia (1999), which focuses on the interaction of antibodies and immunogens.

[0115] In some embodiments, the antibody can be modified to extend its biological half-life. Various approaches are possible. For example, this can be done by increasing the binding affinity of Fc region to FcRn. For example, as described in U.S. Patent No. 6,277,375, one or more of the following residues can be mutated: 252, 254, 256, 433, 435, 436. Specific exemplary substitutions include one or more of the following: T252L, T254S, and / or T256F. Alternatively, to increase biological half-life, the antibody may be modified in the CH1 or CL region to contain a salvage receptor binding epitope obtained from two loops in the CH2 region of the Fc region of IgG, as described by Presta et al. in U.S. Patent Nos. 5,869,046 and 6,121,022. Other exemplary mutations that increase binding to FcRn and / or improve pharmacokinetic properties include substitutions at positions 259, 308, 428, and 434, such as, for example, 259I, 308F, 428L, 428M, 434S, 434H, 434F, 434Y, and 434M. Other mutants that increase Fc binding to FcRn include 250E, 250Q, 428L, 428F, 250Q / 428L (Hinton et al., 2004, J. Biol. Chem. 279(8): 6213-6216, Hinton et al. 2006 Journal of Immunology 176:346-356), 256A, 272A, 286A, 305A, 307A, 307Q, 311A, 312A, 376A, 378Q, 380A, 382A, 434A (Shields et al, Journal of Biological Chemistry, 2001, 276(9):6591-6604), 252F, 252T, 252Y, 252W, 254T, 256S, 256R, 256Q, 256E, 256D, 256T, 309P, 311S, 433 R, 433S, 433I, 433P, 433Q, 434H, 434F, 434Y, 252Y / 254T / 256E, 433K / 434F / 436H, 308T / 309P / 311S(Dall Acqua et al. Journal of Immunology, 2002, 169:5171-5180, Dall'Acqua et al., 2006, Journal of Biological Chemistry 281:23514-23524). Other modifications to modulate FcRn binding are described in Yeung et al., 2010, J Immunol, 182:7663-7671. In some embodiments, hybrid IgG isotypes with specific biological properties may be used.For example, IgG1 / IgG3 hybrid variants can be constructed by substituting amino acids from IgG3 for IgG1 positions in the CH2 and / or CH3 regions at positions where the two isotypes differ. Thus, hybrid variant IgG antibodies can be constructed that contain one or more substitutions, for example, 274Q, 276K, 300F, 339T, 356E, 358M, 384S, 392N, 397M, 422I, 435R, and 436F. In other embodiments described herein, IgG1 / IgG2 hybrid variants can be constructed by substituting amino acids from IgG1 for IgG2 positions in the CH2 and / or CH3 regions at positions where the two isotypes differ. Thus, hybrid variant IgG antibodies can be constructed with chats that contain one or more substitutions, for example, one or more of the following amino acid substitutions: 233E, 234L, 235L, 236G (referring to the insertion of glycine at position 236), and 321h.

[0116] Additionally, the binding sites of human IgG1 for FcγR1, FcγRII, FcγRIII, and FcRn have been mapped and mutants with improved binding have been described (see Shields, RL et al. (2001) J. Biol. Chem. 276:6591-6604). Specific mutations at positions 256, 290, 298, 333, 334, and 339 have been shown to improve binding to FcγRIII. Additionally, the following combination mutations have been shown to improve FcγRIII binding: T256A / S298A, S298A / E333A, S298A / K224A, and S298A / E333A / K334A, which have been shown to exhibit enhanced FcγRIIIa binding and ADCC activity (Shields et al., 2001). Other IgG1 mutants with strongly enhanced binding to FcγRIIIa have been identified, including mutants with S239D / I332E and S239D / I332E / A330L mutations that showed the greatest increase in affinity for FcγRIIIa, reduced FcγRIIb binding, and potent cytotoxic activity in cynomolgus monkeys (Lazar et al., 2013). et al., 2006). Introduction of triple mutations into antibodies such as alemtuzumab (CD52-specific), trastuzumab (HER2 / neu-specific), rituximab (CD20-specific), and cetuximab (EGFR-specific) translated into greatly enhanced ADCC activity in vitro, and the S239D / I332E mutation was shown to enhance their ability to deplete monkey B cells (Lazar et al., 2006). Furthermore, IgG1 mutants containing L235V, F243L, R292P, Y300L, and P396L mutations were identified that showed enhanced binding to FcγRIIIa and concomitantly enhanced ADCC activity in transgenic mice expressing human FcγRIIIa in models of B cell malignancies and breast cancer (Stavenhagen et al., 2007; Nordstrom et al., 2011). Other Fc mutations that can be used include: S298A / E333A / L334A, S239D / I332E, S239D / I332E / A330L, L235V / F243L / R292P / Y300L / P396L, and M428L / N434S.

[0117] In some embodiments, an Fc with reduced binding to FcγR is selected. An exemplary Fc, for example an IgG1 Fc with reduced FcγR binding, contains the following three amino acid substitutions: L234A, L235E, and G237A.

[0118] In some embodiments, an Fc with reduced complement binding is selected. An exemplary Fc with reduced complement binding, e.g., an IgG1 Fc, has two amino acid substitutions: A330S and P331S.

[0119] In some embodiments, the Fc is selected to have substantially no effector function, i.e., reduced binding to FcγR and reduced complement fixation. An exemplary effector-less Fc, e.g., an IgG1 Fc, contains the following five mutations: L234A, L235E, G237A, A330S, and P331S.

[0120] When an IgG4 constant region is used, it is usually preferable to include the substitution S228P, which mimics the hinge sequence of IgG1, thereby stabilizing the IgG4 molecule.

[0121] Fc mutations that extend serum half-life In some embodiments, the Fc region or Fc domain of the anti-HIV gp120 directed antibody comprises amino acid modifications that promote the serum half-life of the anti-binding molecule. Mutations that increase the half-life of antibodies have been reported. In one embodiment, the Fc region or Fc domain of one or both of the CD3-targeting heavy chain and the HIV antigen-targeting heavy chain comprises a methionine to tyrosine substitution at position 252 (EU numbering), a serine to threonine substitution at position 254 (EU numbering), and a threonine to glutamic acid substitution at position 256 (EU numbering). See, for example, U.S. Patent No. 7,658,921. This type of variant, termed "YTE variants", exhibits a 4-fold longer half-life compared to the wild-type version of the same antibody (Dall'Acqua, et al., J Biol Chem, 281: 23514-24 (2006); Robbie, et al., Antimicrob Agents Chemotherap., 57(12):6147-6153 (2013)). In certain embodiments, the Fc region or domain of one or both of the CD3-targeting heavy chain and the HIV antigen-targeting heavy chain comprises an IgG constant region comprising one, two, three or more amino acid substitutions at amino acid residues 251-257, 285-290, 308-314, 385-389, and 428-436 (EU numbering). Alternatively, M428L and N434S ("LS") substitutions may extend the pharmacokinetic half-life of the multispecific antigen binding molecule. In other embodiments, the Fc region or Fc domain of one or both of the CD3-targeting heavy chain and the HIV antigen-targeting heavy chain comprises M428L and and N434S (EU numbering) substitutions. In another embodiment, the Fc region or Fc domain of one or both of the CD3-targeting heavy chain and the HIV antigen-targeting heavy chain comprises a T250Q and M428L (EU numbering) mutation. In another embodiment, the Fc region or Fc domain of one or both of the CD3-targeting heavy chain and the HIV antigen-targeting heavy chain comprises a H433K and N434F (EU numbering) mutation.

[0122] Fc mutations that enhance effector activity In some embodiments, the Fc region or Fc domain of the anti-HIV gp120-directed antibody comprises amino acid modifications that increase post-translational and / or effector activity, e.g., improved FcγIIIa binding and increased antibody-dependent cellular cytotoxicity (ADCC). In some embodiments, the Fc region or Fc domain of the anti-HIV gp120-directed antibody comprises a DE modification in the Fc region (i.e., S239D and I332E according to EU numbering). In some embodiments, the Fc region or Fc domain of the anti-HIV gp120-directed antibody comprises a DEL modification in the Fc region (i.e., S239D, I332E, and A330L according to EU numbering). In some embodiments, the Fc region or Fc domain of the anti-HIV gp120-directed antibody comprises a DEA modification in the Fc region (i.e., S239D, I332E, and G236A according to EU numbering). In some embodiments, the Fc region or Fc domain of the anti-HIV gp120 directed antibody comprises DEAL modifications in the Fc region (i.e., S239D, I332E, G236A, and A330L according to EU numbering).See, for example, U.S. Patent Nos. 7,317,091; 7,662,925; 8,039,592; 8,093,357; 8,093,359; 8,383,109; 8,388,955; 8,735,545; 8,858,937; 8,937,158; 9,040,041; 9,353,187; 10,184,000; and 10,584,176. Additional amino acid modifications that enhance effector activity, e.g., improved FcγIIIa binding and increased antibody-dependent cellular cytotoxicity (ADCC), include, but are not limited to, F243L / R292P / Y300L / V305I / P396L; S298A / E333A / K334A; or L234Y / L235Q / G236W / S239M / H268D / D270E / S298A on the first Fc region, and D270E / K326D / A330M / K334E on the second Fc region (EU numbering).Amino acid mutations that increase C1q binding and complement-dependent cytotoxicity (CDC) include, but are not limited to, S267E / H268F / S324T or K326W / E333S (EU numbering). Mutations in the Fc region that enhance effector activity are described, for example, in Wang, et al., Protein Cell (2018) 9(1): 63-73; and Saunders, Front Imm. Reviewed in Unol. (2019) 10:1296.

[0123] In other embodiments, the anti-HIV gp120 directed antibody or antigen-binding fragment thereof has modified glycosylation, which may be introduced, for example, post-translationally or by genetic engineering. In some embodiments, the anti-HIV gp120 directed antibody or antigen-binding fragment thereof is afucosylated, for example, at glycosylation sites present in the antibody or antigen-binding fragment thereof. Most approved monoclonal antibodies are of the IgG1 isotype, with two N-linked biantennary complex type oligosaccharides attached to the Fc region. The Fc region exerts the effector function of ADCC through interaction with leukocyte receptors of the FcγR family. Defucosylated monoclonal antibodies are monoclonal antibodies that have been designed such that the oligosaccharides of the Fc region of the antibody do not have fucose sugar units.

[0124] f. multivalent antibodies In one embodiment, the antibodies of the present invention can be monovalent or multivalent (e.g., bivalent, trivalent, etc.). As used herein, the term "valency" refers to the number of potential target binding sites associated with an antibody. Each target binding site specifically binds to one target molecule, or to a specific position or locus on a target molecule. When an antibody is monovalent, each binding site on the molecule specifically binds to a single antigen position or epitope. When an antibody contains multiple target binding sites (multivalent), each target binding site can specifically bind to the same or different molecules (e.g., different ligands or different antigens, or different epitopes or positions on the same antigen). See, e.g., USPN 2009 / 0130105. In each case, at least one of the binding sites includes an epitope, motif, or domain associated with a DLL3 isoform.

[0125] In one embodiment, the antibody is a bispecific antibody, in which the two chains have different specificities, as described in Millstein et al., 1983, Nature, 305:537-539. Other embodiments include antibodies with additional specificities, such as trispecific antibodies. Other more sophisticated interchangeable multispecific constructs and methods for their production are described in USPN 2009 / 0155255, and WO 94 / 04690; Suresh et al., 1986, Methods in Enzymology, 121:210; and WO96 / 27011.

[0126] As noted above, a multivalent antibody may immunospecifically bind to different epitopes of a target molecule of interest, or may immunospecifically bind to both the target molecule and a heterologous epitope, such as a heterologous polypeptide or solid support material, hi some embodiments, the multivalent antibody may include a bispecific antibody or a trispecific antibody.

[0127] Bispecific antibodies are antibodies with binding specificities for at least two different epitopes. Exemplary bispecific antibodies may bind to two different epitopes of a single antigen. Other such antibodies may combine a first antigen binding site with a binding site for a second antigen. Alternatively, to focus and localize cellular defense mechanisms to infected cells, the anti-HIV arm may be combined with an arm that binds to a trigger molecule on white blood cells, such as a T cell receptor molecule (e.g., CD3), or an Fc receptor for IgG (Fc gamma R), such as Fc gamma RI (CD64), Fc gamma RII (CD32), and Fc gamma RIII (CD16). Bispecific antibodies may be used to localize cytotoxic agents to infected cells. Bispecific antibodies may be prepared as full-length antibodies or antibody fragments (e.g., F(ab')2 bispecific antibodies). For example, WO96 / 16673 describes a bispecific anti-ErbB2 / anti-Fc gamma RIII antibody, and U.S. Patent No. 5,837,234 discloses a bispecific anti-ErbB2 / anti-Fc gamma RI antibody. For example, a bispecific anti-ErbB2 / Fc alpha antibody is reported in WO98 / 02463; and U.S. Patent No. 5,821,337 teaches a bispecific anti-ErbB2 / anti-CD3 antibody. Further, see, for example, Mouquet et al., Polyreactivity Increases The Apparent Affinity Of Anti-HIV Antibodies By Heteroligation. NATUR E. 467, 591-5 (2010).

[0128] Bispecific antibodies also include cross-linked or "heteroconjugate" antibodies. For example, one of the antibodies of the heteroconjugate can be bound to avidin and the other to biotin. Such antibodies have been proposed, for example, to target immune system cells to unwanted cells (U.S. Pat. No. 4,676,980) and for the treatment of HIV infection (WO 91 / 00360, WO 92 / 200373, and EP 03089). Heteroconjugate antibodies can be produced using any convenient cross-linking method. Suitable cross-linking agents are known in the art and are described in U.S. Pat. No. 4,676,980, along with a number of cross-linking techniques. This is disclosed in US Pat. No. 4,676,980.

[0129] Methods for producing bispecific antibodies are known in the art. Traditional production of full-length bispecific antibodies is based on the co-expression of two immunoglobulin heavy-light chain pairs, where the two chains have different specificities (see, e.g., Millstein et al., Nature, 305:537-539 (1983)). Similar procedures are disclosed, for example, in WO 93 / 08829, Traunecker et al., EMBO J., 10:3655-3659 (1991), and further in Mouquet et al., Polyreactivity Increases The See Apparent Affinity Of Anti-HIV Antibodies By Heteroligation. NATURE. 467, 591-5 (2010).

[0130] Alternatively, an antibody variable region with the desired binding specificity (antibody-antigen binding site) is fused to an immunoglobulin constant region sequence. The fusion is with an Ig heavy chain constant region, including at least a portion of the hinge, CH2, and CH3 regions. According to some embodiments, the first heavy chain constant region (CH1), including the site required for light chain binding, is present in at least one of the fusions. DNA encoding the immunoglobulin heavy chain fusions, and optionally the immunoglobulin light chain, are inserted into separate expression vectors and co-transfected into a suitable host cell. If an unequal ratio of the three polypeptide chains used in the construction provides the optimal yield of the desired bispecific antibody, this provides greater flexibility in adjusting the mutual ratio of the three polypeptide fragments in embodiments. However, if the expression of at least two polypeptide chains in equal ratios results in high yields, or if the ratio does not significantly affect the yield of the desired chain combination, the coding sequences for two or all three polypeptide chains may be inserted into a single expression vector.

[0131] Techniques for producing bispecific antibodies from antibody fragments are also described in the literature. For example, bispecific antibodies can be prepared using chemical linkage. For example, Brennan et al., Science, 229: 81 (1985) describes a procedure in which intact antibodies are proteolytically cleaved to produce F(ab')2 fragments. These fragments are reduced in the presence of the dithiol complexing agent sodium arsenite to stabilize vicinal dithiols and prevent intermolecular disulfide formation. The Fab' fragments produced are then converted to thionitrobenzoic acid (TNB) derivatives. One of the Fab'-TNB derivatives is then reconverted to a Fab'-thiol by reduction with mercaptoethylamine and mixed with an equimolar amount of the other Fab'-TNB derivative to form the bispecific antibody. The bispecific antibody produced can be used as an agent for selective immobilization of enzymes.

[0132] Other modifications of the antibody are contemplated herein. For example, the antibody can be linked to one of a variety of non-proteinaceous polymers, such as polyethylene glycol, polypropylene glycol, polyoxyalkylene, or copolymers of polyethylene glycol and polypropylene glycol. The antibody can also be encapsulated in microcapsules (e.g., hydroxymethylcellulose or gelatin-microcapsules, and poly-(methylmethacylate) microcapsules, respectively), colloidal drug delivery systems (e.g., liposomes, albumin microspheres, microemulsions, nanoparticles, and nanocapsules), or macroemulsions, prepared, for example, by coacervation techniques or interfacial polymerization. Such techniques are disclosed, for example, in Remington's Pharmaceutical Sciences, 16th edition, Oslo, A., Ed., (1980).

[0133] In some embodiments, a bispecific antibody of the invention comprises a first antigen-binding arm that binds a first antigen (e.g., gp120) and a second antigen-binding arm that binds a second antigen, wherein said first and second antigens are different.

[0134] In some embodiments, the first antigen-binding arm comprises: (a) VH CDR1-3 and VL CDR1-3 set forth in SEQ ID NOs:11-16, respectively; or (b) HC and LC set forth in SEQ ID NO:3 and SEQ ID NO:2; SEQ ID NO:3 and SEQ ID NO:6; SEQ ID NO:3 and SEQ ID NO:7; SEQ ID NO:3 and SEQ ID NO:8; SEQ ID NO:3 and SEQ ID NO:9; SEQ ID NO:3 and SEQ ID NO:10; SEQ ID NO:1 and SEQ ID NO:5; SEQ ID NO:1 and SEQ ID NO:8; or SEQ ID NO:1 and SEQ ID NO:9, respectively.

[0135] In some embodiments, the second antigen is CD3, FcγRI (CD64), FcγRII (CD32), FcγRIII (CD16); CD89, CCR5, CD4, CD8, CD28, CD137, CTLA-4, gp41, killer cell immunoglobulin-like receptor, three Ig domains, and long cytoplasmic tail 1 (KIR3DL1), killer cell immunoglobulin-like receptor, three Ig domains, and long cytoplasmic tail 1 (KIR3DL1), killer cell immunoglobulin Killer cell immunoglobulin-like receptor, two Ig domains and long cytoplasmic tail 1 (KIR2DL1), Killer cell immunoglobulin-like receptor, two Ig domains and long cytoplasmic tail 3 (KIR2DL3), Killer cell lectin-like receptor Cl (KLRC1), Killer cell lectin-like receptor C2 (KLRC2), Killer cell lectin-like receptor C3 (KLRC3), Killer cell lectin-like receptor C4 (KLRC4), Killer cell lectin-like receptor D1 (KLRD1), Killer cell lectin-like receptor KLRK1 (KLRK1), natural cytotoxicity-inducing receptor 3 (NCR3 or NKp30), natural cytotoxicity-inducing receptor 2 (NCR2 or NK-p44), natural cytotoxicity-inducing receptor 1 (NCR1 or NK-p46), CD226 (DNAM-1), cytotoxic and regulatory T cell molecule (CRTAM or CD355), signaling lymphocyte activation molecule family member 1 (SLAMF1), CD48 (SLAMF2), lymphocyte antigen 9 (LY9 or SLAMF3), CD2 44 (2B4 or SLAMF4), CD84 (SLAMF5), SLAM family member 6 (SLAMF6 or NTB-A), SLAM family member 7 (SLAMF7 or CRACC), CD27 (TNFRSF7), semaphorin 4D (SEMA4D or CD100), CD160 (NK1), an immune checkpoint protein (e.g., PD-1, PD-L1, CTLA-4), and the second epitope of gp120.

[0136] g.Antibody derivatives The antibodies provided herein can be further modified to contain additional nonproteinaceous moieties that are known in the art and readily available. Moieties suitable for derivatization of the antibodies include, but are not limited to, water soluble polymers.

[0137] Non-limiting examples of water-soluble polymers include, but are not limited to, PEG, ethylene glycol / propylene glycol copolymers, carboxymethylcellulose, dextran, polyvinyl alcohol, polyvinylpyrrolidone, poly-1,3-dioxolane, poly-1,3,6-trioxane, ethylene / maleic anhydride copolymers, polyamino acids (either homopolymers or random copolymers), dextran or poly(n-vinylpyrrolidone), polyethylene glycol, propylene glycol homopolymers, and polypropylene oxide / ethylene oxide copolymers, polyoxyethylated polyols (e.g., glycerol), polyvinyl alcohol, and mixtures thereof. Polyethylene glycol propionaldehyde has advantages in manufacturing because it is stable in water. The polymers can be of any molecular weight and can be branched or unbranched. The number of polymers attached to the antibody can vary, and when multiple polymers are attached, they can be the same molecule or different molecules. In general, the number and / or type of polymers used for derivatization can be determined based on considerations including, but not limited to, the particular property or function of the antibody to be improved, whether the antibody derivative will be used in therapy under defined conditions, etc.

[0138] In other embodiments, conjugates of antibodies and non-proteinaceous moieties are provided that can be selectively heated by exposure to radiation. In one embodiment, the non-proteinaceous moiety is a carbon nanotube (Kam et al., Proc. Natl. Acad. Sci. USA 102: 11600-11605 (2005)). The radiation can be of any wavelength, including but not limited to, that is not harmful to normal cells but heats the non-proteinaceous moiety to a temperature that kills cells in the vicinity of the antibody-non-proteinaceous moiety.

[0139] Another modification of the antibodies described herein is PEGylation. Antibodies may be PEGylated, for example, to increase the biological (e.g., serum) half-life of the antibody. To PEGylate an antibody, said antibody, or a fragment thereof, it is reacted with PEG, such as a reactive ester or aldehyde derivative of PEG, under conditions that typically result in one or more PEG groups being attached to the antibody or antibody fragment. Preferably, said PEGylation is carried out via an acylation reaction or an alkylation reaction with a reactive PEG molecule (or a similar reactive water-soluble polymer). As used herein, the term "polyethylene glycol" is intended to encompass any of the forms of PEG that have been used to derivatize other proteins, such as mono(CI-CIO)alkoxy- or aryloxy-polyethylene glycol or polyethylene glycol-maleimide. In some embodiments, said antibody to be PEGylated is a non-glycosylated antibody. Methods for PEGylating proteins are known in the art and may be applied to the antibodies described herein. See, for example, EP 0 154 316 by Nishimura et al. and EP 0401384 by Ishikawa et al.

[0140] The present invention also encompasses a human monoclonal antibody as described herein conjugated to a therapeutic agent, polymer, detectable label, or enzyme. In one embodiment, the therapeutic agent is a cytotoxic drug. In one embodiment, the polymer is PEG.

[0141] h. Nucleic acids, expression cassettes, and vectors The present invention provides isolated nucleic acid segments encoding the polypeptides, peptide fragments, and binding proteins of the present invention. The nucleic acid segments of the present invention also include segments that code for the same amino acid due to the degeneracy of the genetic code. For example, the amino acid threonine is coded by ACU, ACC, ACA, and ACG, and is therefore degenerate. The present invention is intended to include all variations of polynucleotide segments that code for the same amino acid. Such mutations are known in the art (Watson et al., Molecular Biology of the Gene, Benjamin Cummings 1987). Mutations also include alterations of nucleic acid segments to code for conservative amino acid changes, such as the substitution of leucine for isoleucine. Such mutations are also known in the art. Thus, the genes and nucleotide sequences of the present invention include both naturally occurring sequences and variants.

[0142] The nucleic acid segments of the present invention may be contained within a vector. Vectors may include, but are not limited to, double-stranded or single-stranded, linear or circular plasmids, phagemids, F factors, viruses, cosmids, or phages, which may or may not be self-transmissible or mobilizable. The vectors may also be capable of being transferred to a cell by integration into the cell genome. The vector may be capable of transforming a prokaryotic or eukaryotic host by being present extrachromosomally (eg, an autonomously replicating plasmid having an origin of replication).

[0143] Preferably, the nucleic acid segment in the vector is under the control of and operably linked to a suitable promoter or other regulatory element for transcription in vitro or in a host cell, such as a eukaryotic cell, or in a microorganism, such as a bacterium. The vector may be a shuttle vector that functions in multiple hosts. The vector may also be a cloning vector, which typically contains one or a few restriction endonuclease recognition sites into which a foreign DNA sequence may be inserted in a determinable manner. Such insertion may occur without losing the essential biological function of the cloning vector. The cloning vector may also contain a marker gene suitable for use in identifying and selecting cells transformed with the cloning vector. Examples of marker genes are tetracycline resistance or ampicillin resistance. Many cloning vectors are commercially available (Stratagene, New England Biolabs, Clonetech).

[0144] The nucleic acid segment of the present invention can also be inserted into an expression vector.Usually, an expression vector contains the following prokaryotic DNA elements: a bacterial origin of replication and an antibiotic resistance gene coding for providing the amplification and selection of the expression vector in a bacterial host; a regulatory element such as a promoter that controls the initiation of transcription; a DNA element that controls the processing of the transcript such as an intron, or a transcription termination / polyadenylation sequence.

[0145] Methods for introducing nucleic acid segments into vectors are available in the art (Sambrook et al., Molecular Cloning: A Laboratory Manual, 3rd edition, Cold Spring Harbor Press, Cold Spring Harbor, NY (2001)). Briefly, the vector into which the nucleic acid segment is to be inserted is treated with one or more restriction enzymes (restriction endonucleases) to produce a linearized vector with blunt ends, "sticky ends" with 5' or 3' overhangs, or any combination of the above. The vector may also be treated with a restriction enzyme followed by other modifying enzymes such as polymerases, exonucleases, phosphatases, or kinases to create a linearized vector with properties useful for ligating the nucleic acid segment into the vector. The nucleic acid segment to be inserted into the vector is treated with one or more restriction enzymes to produce a linearized segment with blunt ends, "sticky ends" with 5' or 3' overhangs, or any combination of the above. The nucleic acid segment may also be treated with a restriction enzyme followed by other DNA modifying enzymes. Such DNA modifying enzymes include, but are not limited to, polymerases, exonucleases, phosphatases, or kinases to generate a nucleic acid segment with properties useful for ligating the nucleic acid segment into said vector.

[0146] The treated vector and nucleic acid segment are then ligated together to form a construct containing the nucleic acid segment according to methods available in the art (Sambrook et al., Molecular Cloning: A Laboratory Manual, 3rd edition, Cold Spring Harbor Press, Cold Spring Harbor, NY (2001)). Briefly, the treated nucleic acid fragment and the treated vector are combined in the presence of a suitable buffer and ligase. The mixture is then incubated under suitable conditions to allow the ligase to link the nucleic acid fragment to the vector.

[0147] The present invention also provides expression cassettes comprising nucleic acid sequences capable of directing the expression of particular nucleic acid segments of the present invention, either in vitro or in a host cell. The ment can be inserted into an expression cassette so that an antisense message is produced. The expression cassette is a separable unit such that the expression cassette is in linear form and can be functional for in vitro transcription and translation assays. Materials and procedures for carrying out these assays are commercially available from Promega (Madison, Wisconsin). For example, an in vitro transcript can be produced by placing a nucleic acid sequence under the control of a T7 promoter and then using T7 RNA polymerase to produce an in vitro transcript. The transcript can then be translated in vitro by using rabbit reticulocyte lysate. Alternatively, the expression cassette can be incorporated into a vector that allows for replication and amplification of the expression cassette in a host cell, or also for in vitro transcription and translation of a nucleic acid segment.

[0148] Such an expression cassette may include one or more restriction sites that allow the placement of the nucleic acid segment under the control of regulatory sequences. The expression cassette may also include a termination signal operably linked to the nucleic acid segment, and regulatory sequences necessary for proper translation of the nucleic acid segment. An expression cassette containing the nucleic acid segment may be chimeric, meaning that at least one of its components is heterologous with respect to at least one of the other components. The expression cassette may also be naturally occurring but obtained in a recombinant form useful for heterologous expression. Expression of the nucleic acid segment in the expression cassette may be under the control of a constitutive or inducible promoter that initiates transcription only when the host cell is exposed to a particular external stimulus. The expression cassette may include, in the 5' to 3' direction of transcription, a transcription and translation initiation region, a nucleic acid segment, and a transcription and translation termination region that functions in vivo and / or in vitro. The termination region may be native to the transcription initiation region, native to the nucleic acid segment, or may be derived from another source.

[0149] The regulatory sequences may be polynucleotide sequences located upstream (5' non-coding sequences), within, or downstream (3' non-coding sequences) of a coding sequence that affect the transcription, RNA processing or stability, or translation of the associated coding sequence. Regulatory sequences may include, but are not limited to, enhancers, promoters, repressor binding sites, translation leader sequences, introns, and polyadenylation signal sequences. They may include sequences that may be natural and synthetic sequences, as well as combinations of synthetic and natural sequences. Regulatory sequences are not limited to promoters, but some useful regulatory sequences include constitutive promoters, inducible promoters, regulated promoters, tissue-specific promoters, viral promoters, and synthetic promoters.

[0150] A promoter is a nucleotide sequence that controls the expression of a coding sequence by providing recognition of RNA polymerase and other factors required for proper transcription. Promoters include minimal promoters that are composed of only all the basic elements required for transcription initiation, such as a TATA box and / or an initiator, which is a short DNA sequence composed of a TATA box, and other sequences that are useful for identifying the site of transcription initiation, to which regulatory elements are added for the control of expression. Promoters can be entirely derived from natural genes, or can be composed of different elements derived from different promoters found in nature, or can be composed of synthetic DNA segments. Promoters can include DNA sequences that are involved in the binding of protein factors that control the effectiveness of transcription initiation in response to physiological or developmental conditions.

[0151] The present invention also provides a construct comprising a vector and an expression cassette. The vector may be selected from, but is not limited to, any of the vectors described above. The expression cassette may be inserted into the vector via methods known in the art and described above (Sambrook et al., Molecular Cloning: A Laboratory Manual, 3rd edition, Cold Spring Harbor Laboratory Press, 2003). rbor Press, Cold Spring Harbor, NY (2001). In one embodiment, the regulatory sequences of the expression cassette may be derived from a source other than the vector into which the expression cassette is inserted. In other embodiments, a construct comprising a vector and an expression cassette is formed upon insertion of a nucleic acid segment of the invention into a vector that itself contains regulatory sequences. Thus, an expression cassette is formed upon insertion of a nucleic acid segment into a vector. Vectors containing regulatory sequences are commercially available and methods for their use are known in the art (Clonetech, Promega, Stratagene).

[0152] In other aspects, the disclosure also provides (i) a nucleic acid molecule encoding a polypeptide chain of the antibody or antigen-binding fragment thereof described above; (ii) a vector comprising the nucleic acid molecule as described; and (iii) a cultured host cell comprising the vector as described. Also provided is a method for producing a polypeptide, comprising: (a) obtaining a cultured host cell as described; (b) culturing the cultured host cell in a medium under conditions that permit expression of a polypeptide encoded by the vector and assembly of an antibody or fragment thereof; and (c) purifying the antibody or fragment from the cultured cell or the medium of the cell.

[0153] i. Manufacturing method Antibodies may be produced using recombinant methods and compositions, for example, as described in U.S. Pat. No. 4,816,567. In one embodiment, an isolated nucleic acid is provided that encodes an antibody described herein. Such a nucleic acid may encode an amino acid sequence comprising the VL and / or an amino acid sequence comprising the VH of said antibody (e.g., the light and / or heavy chains of the antibody). In a further embodiment, one or more vectors (e.g., expression vectors) comprising such a nucleic acid are provided. In a further embodiment, a host cell comprising such a nucleic acid is provided. In one such embodiment, the host cell comprises (e.g., is transformed with): (1) a vector comprising a nucleic acid encoding an amino acid sequence comprising the VL of the antibody and an amino acid sequence comprising the VH of the antibody, or (2) a first vector comprising a nucleic acid encoding an amino acid sequence comprising the VL of the antibody, and a second vector comprising a nucleic acid encoding an amino acid sequence comprising the VH of the antibody. In one embodiment, the host cell is eukaryotic, such as a Chinese Hamster Ovary (CHO) cell or a lymphoid cell (e.g., a Y0 cell, an NS0 cell, an Sp20 cell). In one embodiment, a method of making an antibody is provided, the method comprising culturing a host cell (or host cell culture medium) containing nucleic acid encoding the antibody under conditions suitable for expression of the antibody, as described above, and optionally recovering the antibody from the host cell.

[0154] For recombinant production of an antibody, nucleic acid encoding the antibody is isolated and inserted into one or more vectors for further cloning and / or expression in host cells, e.g., as described above. Such nucleic acid can be readily isolated and sequenced using conventional procedures (e.g., by using oligonucleotide probes capable of specifically binding to genes encoding the heavy and light chains of the antibody).

[0155] Suitable host cells for cloning or expressing antibody-encoding vectors include prokaryotic or eukaryotic cells as described herein. For example, antibodies can be produced in bacteria, particularly where glycosylation and Fc effector functions are not required. For expression of antibody fragments and polypeptides in bacteria, see, e.g., U.S. Pat. Nos. 5,648,237, 5,789,199, and 5,840,523 (also see Charlton, Methods in Molecular Biology, Vol. 248 (BKC Lo, ed., Humana Press, Totowa, N.Y.), which describes the expression of antibody fragments in E. coli. J., 2003), pp. 245-254. After expression, the antibody can be isolated from the bacterial cell paste in a soluble fraction and further purified.

[0156] In addition to prokaryotes, eukaryotic microbes such as filamentous fungi or yeast are suitable cloning or expression hosts for antibody-encoding vectors, including fungal and yeast strains in which the glycosylation pathway has been "humanized" to produce antibodies with partially or fully human glycosylation patterns. See Gerngross, Nat. Biotech. 22:1409-1414 (2004), and Li et al., Nat. Biotech. 24:210-215 (2006).

[0157] Host cells suitable for the expression of glycosylated antibodies are also derived from multicellular organisms (invertebrates and vertebrates). Examples of invertebrate cells include plant and insect cells. Many baculovirus strains have been identified that can be used in combination with insect cells, particularly for the transfection of Spodoptera afrugiperda cells.

[0158] Plant cell cultures can also be used as hosts. See, for example, U.S. Patent Nos. 5,959,177, 6,040,498, 6,420,548, 7,125,978, and 6,417,429 (describing the PLANTIBODIES technology for producing antibodies in transgenic plants). .

[0159] Vertebrate cells may also be used as hosts, for example mammalian cell lines adapted to grow in suspension may be useful. Other examples of useful mammalian host cell lines are monkey kidney CV1 line transformed by SV40 (COS-7); human embryonic kidney lines (e.g., 293 or 293 cells as described in Graham et al., J. Gen Virol. 36:59 (1977)); baby hamster kidney cells (BHK); mouse Sertoli cells (e.g., TM4 cells as described in Mather, Biol. Reprod. 23:243-251 (1980)); monkey kidney cells (CV1); African green monkey kidney cells (VERO-76); human cervical carcinoma cells (HELA); dog kidney cells (MDCK; buffalo rat liver cells (BRL 3A); human lung cells (W138); human hepatocytes (Hep G2); mouse mammary tumor (MMT 060562); e.g., Mather et al., Annals NY Acad. Sci. 383:44-68 (1982); MRC5 cells; and FS4 cells. Other useful mammalian host cell lines include CHO cells, such as DHFR-CHO cells (Urlaub et al., Proc. Natl. Acad. Sci. USA 77:4216 (1980)); and myeloma cell lines, such as Y0, NS0, and Sp2 / 0. For reviews of specific mammalian host cell lines suitable for antibody production, see, for example, Yazaki and Wu, Methods in Molecular Biology, Vol. 248 (BKC Lo, ed., Humana Press, Totowa, NJ), pp. 255-268 (2003).

[0160] B. Compositions and Formulations In another aspect, the present disclosure also provides a pharmaceutical composition comprising: (i) at least one anti-HIV antibody or antigen-binding portion thereof, nucleic acid, or vector as described above; and (ii) a pharma- ceutically acceptable carrier.

[0161] In some embodiments, the pharmaceutical composition further comprises a second therapeutic agent. In some embodiments, the second therapeutic agent comprises an antiviral agent or one or more additional antibodies. In some embodiments, the one or more additional antibodies are a second anti-HIV The antibody (e.g., an isolated anti-HIV bNAb disclosed herein) or antigen-binding portion thereof, or a third antibody that binds to a third antigen. In some embodiments, the third antigen is selected from the group consisting of CD3, FcγRI (CD64), FcγRII (CD32), FcγRIII (CD16); CD89, CCR5, CD4, CD8, CD28, CD137, CTLA-4, gp41, killer cell immunoglobulin-like receptor, three Ig domains, and long cytoplasmic tail 1 (KIR3DL1), ... Killer cell immunoglobulin-like receptor, two Ig domains and long cytoplasmic tail 1 (KIR2DL1), Killer cell immunoglobulin-like receptor, two Ig domains and long cytoplasmic tail 3 (KIR2DL3), Killer cell lectin-like receptor Cl (KLRC1), Killer cell lectin-like receptor C2 (KLRC2), Killer cell lectin-like receptor C3 (KLRC3), Killer cell lectin-like receptor C4 (KLRC4), Killer cell lectin-like receptor D1 (KLRD1), Killer cell lectin-like receptor KLRK1 (KLRK1), natural cytotoxicity-inducing receptor 3 (NCR3 or NKp30), natural cytotoxicity-inducing receptor 2 (NCR2 or NK-p44), natural cytotoxicity-inducing receptor 1 (NCR1 or NK-p46), CD226 (DNAM-1), cytotoxic and regulatory T cell molecule (CRTAM or CD355), signaling lymphocyte activation molecule family member 1 (SLAMF1), CD48 (SLAMF2), lymphocyte antigen 9 (LY9 or SLAMF3), CD2 44 (2B4 or SLAMF4), CD84 (SLAMF5), SLAM family member 6 (SLAMF6 or NTB-A), SLAM family member 7 (SLAMF7 or CRACC), CD27 (TNFRSF7), semaphorin 4D (SEMA4D or CD100), CD160 (NK1), an immune checkpoint protein (e.g., PD-1, PD-L1, CTLA-4), and the second epitope of gp120.

[0162] In some embodiments, the antiviral agent is selected from the group consisting of a non-nucleoside reverse transcriptase inhibitor, a protease inhibitor, an entry or fusion inhibitor, and an integrase inhibitor.

[0163] Pharmaceutical compositions of the invention can also be administered as vaccines (e.g., as AAV-based vaccines) or in combination therapy with, for example, other immunostimulants, antivirals, or vaccines, etc. In some embodiments, the composition comprises an antibody of the invention in a concentration of at least 1 mg / ml, 5 mg / ml, 10 mg / ml, 50 mg / ml, 100 mg / ml, 150 mg / ml, 200 mg / ml, 1-300 mg / ml, or 100-300 mg / ml.

[0164] In some embodiments, the second therapeutic agent comprises an anti-inflammatory drug or an anti-viral compound. In some embodiments, the anti-viral compound comprises a nucleoside analog, a peptoid, an oligopeptide, a polypeptide, a protease inhibitor, a 3C-like protease inhibitor, a papain-like protease inhibitor, or an inhibitor of RNA-dependent RNA polymerase. In some embodiments, the anti-viral compound may include acyclovir, ganciclovir, vidarabine, foscarnet, cidofovir, amantadine, ribavirin, trifluorothymidine, zidovudine, didanosine, zalcitabine, or an interferon. In some embodiments, the interferon is interferon-α or interferon-β.

[0165] Use of the pharmaceutical composition in the preparation of a medicament for the diagnosis, prevention, treatment, or a combination thereof, of a condition resulting from HIV-1 infection is also within the scope of the disclosure.

[0166] The pharmaceutical composition may include any number of excipients. Excipients that may be used include carriers, surfactants, thickeners or emulsifiers, solid binders, dispersing or suspending aids, solubilizers, and the like. The selection and use of suitable additives may include additives, colorants, flavoring agents, coatings, disintegrants, lubricants, sweeteners, preservatives, isotonicity agents, and combinations thereof. The selection and use of suitable additives is described in detail in Gennaro, ed., Remington: The Science of Drug Delivery, the disclosure of which is incorporated herein by reference. Taught in Science and Practice of Pharmacy, 20th Ed. (Lippincott Williams & Wilkins 2003).

[0167] Preferably, the pharmaceutical composition is suitable for intravenous, intramuscular, subcutaneous, parenteral, spinal or epidermal administration (e.g., by injection or infusion). Depending on the route of administration, the active compound may be coated with a material to protect it from the action of acids and other natural conditions that may inactivate it. The phrase "parenteral administration" as used herein refers to modes of administration other than enteral and topical administration, usually by injection, including, but not limited to, intravenous, intramuscular, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal, epidural and intrasternal injection and infusion. Alternatively, the antibody of the present invention described herein may be administered via a parenteral route, such as a topical, epidermal or mucosal route of administration, e.g., intranasal, oral, vaginal, rectal, sublingual or topical.

[0168] The pharmaceutical composition of the present invention can be prepared in many forms, including tablets, hard or soft gelatin capsules, aqueous solutions, suspensions, liposomes, and other sustained release formulations such as molded polymer gels.Oral dosage forms can be formulated so that the antibody is released in the intestine after passing through the stomach.Such formulations are described in U.S. Patent No. 6,306,434 and the references contained therein.

[0169] Oral liquid pharmaceutical compositions may be in the form of, for example, aqueous or oily suspensions, solutions, emulsions, syrups or elixirs, or may be presented as a dry product for constitution with water or other suitable vehicle before use. Such liquid pharmaceutical compositions may contain conventional additives such as suspending agents, emulsifying agents, non-aqueous vehicles (which may include edible oils), or preservatives.

[0170] The antibody can be formulated for parenteral administration (e.g., by injection, e.g., bolus injection or continuous infusion) and can be presented in unit dosage form in ampoules, pre-filled syringes, small volume infusion containers, or multi-dose containers with added preservatives. The pharmaceutical composition can take the form of a suspension, solution, or emulsion in oily or aqueous vehicles, and can contain formulating agents such as suspending agents, stabilizers, and / or dispersing agents. Pharmaceutical compositions suitable for rectal administration can be prepared as unit dosage suppositories. Suitable carriers include physiological saline and other materials commonly used in the art.

[0171] For administration by inhalation, the antibodies may be conveniently delivered from an insufflator, nebulizer, or pressurized pack, or other convenient means for delivering an aerosol spray. Pressurized packs may contain a suitable propellant, such as dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide, or other suitable gas. In the case of a pressurized aerosol, the dosage unit may be determined by providing a valve to deliver a metered amount.

[0172] Alternatively, for administration by inhalation or insufflation, the antibodies may take the form of a dry powder composition, e.g., a powder mix with the modulator and a suitable powder base, such as lactose or starch. The powder composition may be presented in unit dosage form, for example, in capsules or cartridges, or, for example, gelatin or blister packs from which the powder may be administered with the aid of an inhaler or insufflator. For intranasal administration, the antibodies may be administered via a liquid spray, such as a plastic bottle atomizer.

[0173] The pharmaceutical composition of the present invention may also contain other ingredients such as flavorings, colorings, antibacterial agents, or preservatives. It will be understood that the amount of antibody required for therapeutic use will vary not only with the specific carrier selected, but also with the route of administration, the nature of the condition being treated, and the age and condition of the patient. Ultimately, the attending medical provider can determine the appropriate dosage. Furthermore, the pharmaceutical composition may be formulated as a single unit dosage form.

[0174] The pharmaceutical compositions of the present invention may be in the form of a sterile aqueous solution or dispersion. It may also be formulated as a microemulsion, liposome, or other ordered structure suitable to high drug concentration.

[0175] The antibodies of the present invention described herein may be administered as sustained release formulations, in which case less frequent administration is required. Dosage and frequency of administration may vary depending on the half-life of the antibody in the patient. Human antibodies usually exhibit the longest half-life, followed by humanized antibodies, chimeric antibodies, and non-human antibodies. Dosage and frequency of administration may vary depending on whether the treatment is preventive or therapeutic. In preventive applications, relatively low doses are administered at relatively infrequent intervals over a long period of time. Some patients continue to receive treatment for the rest of their lives. In therapeutic applications, relatively high doses may be required at relatively short intervals until the progression of the disease is reduced or terminated, preferably until the patient shows partial or complete improvement of the symptoms of the disease. The patient may then be administered a prophylactic regime.

[0176] The amount of active ingredient that can be combined with a carrier material to produce a single dosage form varies depending on the subject being treated and the particular mode of administration, and is usually that amount of the composition that produces a therapeutic effect. Usually, out of 100%, this amount will range from about 0.01% to about 99% of the active ingredient, preferably about 0.1% to about 70% of the active ingredient, and most preferably about 1% to about 30% of the active ingredient, in combination with a pharma- ceutically acceptable carrier.

[0177] The pharmaceutical composition may be a controlled release formulation, such as an implant, a transdermal patch, and a microencapsulated delivery system. Biodegradable, biocompatible polymers may be used, such as ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoesters, and polylactic acid. See, for example, Sustained and Controlled Release Drug Delivery Systems, J.R. Robinson, ed., Marcel Dekker, Inc., New York, See 1978.

[0178] Therapeutic compositions may be administered via medical devices such as: (1) needleless hypodermic injection devices (e.g., US 5,399,163; 5,383,851; 5,312,335; 5,064,413; 4,941,880; 4,790,824; and 4,596,556); (2) microinfusion pumps (US 4,487,603); (3) transdermal devices (US 4,486,194); (4) infusion devices (US 4,447,233 and 4,447,224); and (5) osmotic devices (US 4,439,196 and 4,475,196), the disclosures of which are incorporated herein by reference.

[0179] In some embodiments, the human monoclonal antibodies of the present invention described herein can be formulated to ensure proper distribution in vivo. For example, to ensure that the therapeutic compounds of the present invention cross the blood-brain barrier, they can be formulated into liposomes, which can further include targeting moieties to enhance selective delivery to specific cells or organs. See, for example, US 4,522,811; 5,374,548; 5,416,016; and 5,399,331; VV Ranade (1989) Clin. Pharmacol. 29:685; Umezawa et al., (1988) B iochem. Biophys. Res. Commun. 153:1038; Bloeman et al. (1995) FEBS Lett. 357:140; M. Owais et al. (1995) Antimicrob. Agents Chemother. 39:180; Briscoe et al. (1995) Am. Physiol. 1233:134; Schreier et al. (1994). Biol. Chem. 269:9090; Keinanen and Laukkanen (1994) FEBS Lett. 346:123; and Killion and Fidler (1994) Immunomethods 4:273.

[0180] In some embodiments, the first dose may be followed by a second or multiple subsequent doses of the antibody or antigen-binding fragment thereof, which may be about the same or less than the first dose, and the subsequent doses are separated by at least 1 to 3 days; at least 1 week, at least 2 weeks; at least 3 weeks; at least 4 weeks; at least 5 weeks; at least 6 weeks; at least 7 weeks; at least 8 weeks; at least 9 weeks; at least 10 weeks; at least 12 weeks; or at least 14 weeks.

[0181] Various delivery systems are known and can be used to administer the pharmaceutical compositions of the present invention, e.g., liposomes, microparticles, microcapsules, encapsulation into recombinant cells capable of expressing mutant viruses, receptor-mediated endocytosis (see, e.g., Wu et al. (1987) J. Biol. Chem. 262:4429-4432). Methods of introduction include, but are not limited to, intradermal, transdermal, intramuscular, intraperitoneal, intravenous, subcutaneous, intranasal, epidural, and oral routes. The compositions can be administered by any convenient route, e.g., infusion or bolus injection, absorption through epithelial or mucocutaneous linings (e.g., oral mucosa, rectal and intestinal mucosa, etc.), and can be administered together with other biologically active agents. Administration can be systemic or local. The pharmaceutical compositions can also be carried in vesicles, particularly liposomes (see, e.g., Langer (1990) Science 249: 1527-1533).

[0182] The use of nanoparticles to deliver the antibodies of the invention is also contemplated herein. Antibody-conjugated nanoparticles can be used for both therapeutic and diagnostic applications. Antibody-conjugated nanoparticles and methods of preparation and use are described in Arruebo, M., et al., incorporated herein by reference. al. 2009 ("Antibody-conjugated nanoparticles for biomedical applications" in J. Nanomat. Volume 2009, Article ID 439389). Nanoparticles can be developed and conjugated to antibodies contained in pharmaceutical compositions to target cells. Nanoparticles for drug delivery are also described, for example, in US 8257740, or US 8246995, each of which is incorporated herein in its entirety.

[0183] In certain conditions, the pharmaceutical composition may be delivered in a controlled release system. In one embodiment, a pump may be used. In another embodiment, a polymeric material may be used. In yet another embodiment, the controlled release system may be placed close to the target of the composition, thus requiring only a fraction of the systemic dose.

[0184] The injectable preparations may include dosage forms for intravenous, subcutaneous, intradermal, intracranial, intraperitoneal and intramuscular injections, drip infusions, etc. These injectable preparations may be prepared by known methods. For example, the injectable preparations may be prepared by dissolving, suspending or emulsifying the antibody or its salt in a sterile aqueous or oily medium conventionally used for injections. Aqueous media for injection include, for example, saline, glucose and other liquids. Examples of the isotonic solution include an isotonic solution containing an auxiliary agent such as ethanol, polyalcohol (e.g., propylene glycol, polyethylene glycol), nonionic surfactants [e.g., polysorbate 80, HCO-50 (polyoxyethylene (50 mol) adduct of hydrogenated castor oil)], etc., which may be used in combination with a suitable solubilizer such as polysorbate 80, HCO-50 (polyoxyethylene (50 mol) adduct of hydrogenated castor oil)]. As the oil medium, for example, sesame oil, soybean oil, etc. may be used in combination with a solubilizer such as benzyl benzoate, benzyl alcohol, etc. The injection thus prepared is preferably filled into a suitable ampoule.

[0185] The pharmaceutical composition of the present invention can be delivered subcutaneously or intravenously using a standard needle and syringe. Furthermore, for subcutaneous delivery, a pen delivery device has easy application in delivering the pharmaceutical composition of the present invention. Such a pen delivery device can be reusable or disposable. A reusable pen delivery device typically utilizes a replaceable cartridge containing the pharmaceutical composition. Once all the pharmaceutical composition in the cartridge has been administered and the cartridge is empty, the empty cartridge can be easily discarded and replaced with a new cartridge containing the pharmaceutical composition. The pen delivery device can then be reused. A disposable pen delivery device does not have a replaceable cartridge. Rather, the disposable pen delivery device is pre-filled with the pharmaceutical composition held in a reservoir within the device. Once the reservoir is emptied of the pharmaceutical composition, the entire device is discarded.

[0186] Numerous reusable pen and autoinjector delivery devices have application in the subcutaneous delivery of the pharmaceutical compositions of the present invention. Examples include the AUTOPEN™ (Owen Mumford, Inc., Woodstock, UK), the DISETRONIC™ pen (Disetronic Medical Systems, Burghdorf, Switzerland), the HUMALOG MIX 75 / 25™ pen, the HUMALOG™ pen, the HUMALIN 70 / 30™ pen (Eli Lilly and Co., Indianapolis, IN), the NOVOPEN™ I, II, and III (Novo Nordisk, Copenhagen, Denmark), NOVOPEN JUNIOR™ (Novo Nordisk, Copenhagen, Denmark), BD™ pens (Becton Dickinson, Franklin Lakes, NJ), OPTIPEN™, OPTIPEN PRO™, OPTIPEN STARLET™, and OPTICLIK™ (Sanofi-Aventis, Frankfurt, Germany). Examples of disposable pen delivery devices having utility for subcutaneous delivery of the pharmaceutical compositions of the present invention include, but are not limited to, the SOLOSTAR™ pen (Sanofi- Aventis), FLEXPEN™ (Novo Nordisk), and KWIKPEN™ (Eli Lilly), the SURECLICK™ autoinjector (Amgen, Thousand Oaks, CA), PENLET™ (Haselmeier, Stuttgart, Germany), EPIPEN (Dey, LP), and HUMIRA™ pen (Abbott Labs, Abbott Park, IL), to name just a few.

[0187] Advantageously, the pharmaceutical composition for oral or parenteral use is prepared in a dosage form of a unit dose suitable for the dosage of the active ingredient. Such dosage forms in a unit dose include, for example, tablets, pills, capsules, injections (ampoules), suppositories, etc. The amount of antibody contained is usually about 5 mg to about 500 mg per dosage form in a unit dose; particularly in the form of injection, the antibody is preferably contained about 5 mg to about 300 mg, and in other dosage forms, about 10 mg to about 300 mg.

[0188] C. Methods and Uses a. Treatment method According to another embodiment, the present invention provides a method for treating a mammal infected with a viral (e.g., HIV) infection, comprising administering to the mammal a pharmaceutical composition comprising an anti-HIV antibody disclosed herein. According to one embodiment, the method for treating a mammal infected with HIV comprises administering to the mammal a pharmaceutical composition comprising an antibody or fragment thereof of the present invention. The composition of the present invention may comprise two or more antibodies (e.g., a plurality of antibodies or a pool of antibodies) having the disclosed characteristics. It may also be used in combination with other HIV neutralizing antibodies known in the art, such as GS-9722 (elipovimab), PGT-121, PGT-121.66, PGT-121.414, PGT-122, PGT-123, PGT-124, PGT-125, PGT-126, PGT-128, PGT-130, PGT-133, PGT-134, PGT-135, PGT-136, PGT-137, PGT-138, PGT-139, PGT-140, PGT-141, PGT-142, PGT-143, PGT-144, PGT-145, PGT-146, PGT-147, PGT-148, PGT-149 ...9, PGT-149, PGT-149, PGT-149, PGT-149, PGT-149, PGT-149, PGT-149, PGT-149, PGT-149, PGT-149, PGT- GT-134, PGT-135, PGT-136, PGT-137, PGT-138, PGT-139, 10-1074, 10-1074-J, VRC24, 2 G12, BG18, 354BG8, 354BG18, 354BG42, 354BG33, 354BG129, 354BG188, 354BG411, 354BG These may include, but are not limited to, 426, DH270.1, DH270.6, PGDM12, VRC41.01, PGDM21, PCDN-33A, BF520.1, and VRC29.03 (all of which bind to the V3 glycan region), 2F5, 4E10, M66.6, CAP206-CH12, 10E8, 10E8v4, 10E8-5R-100cF, DH511.11P, 7b2, and LN01 (all of which bind to the MPER of gp41); PG9, PG16, CH01-04 (all of which bind to V1V2-glycans), 2G12 (binds to ectodomain glycans), VRC01, PG9, and b12.

[0189] In some embodiments, the disclosure provides a method of preventing or treating an HIV infection or an HIV-related disease, the method comprising: (a) identifying a patient in need of said prevention or treatment, and (b) administering to said patient a first therapeutic agent comprising a therapeutically effective amount of at least one anti-HIV antibody or antigen-binding portion thereof, as described above.

[0190] In some embodiments, the method further comprises administering a second therapeutic agent. In some embodiments, the second therapeutic agent comprises an antiviral agent or one or more additional antibodies. In some embodiments, the one or more additional antibodies comprise a second anti-HIV antibody (e.g., an isolated anti-HIV bNAb disclosed) or an antigen-binding portion thereof, or a third antibody that binds to a third antigen. In some embodiments, the third antigen is CD3, FcγRI (CD64), FcγRII (CD32), FcγRIII (CD16); CD89, CCR5, CD4, CD8, CD28, CD137, CTLA-4, gp41, killer cell immunoglobulin-like receptor, three Ig domains and long cytoplasmic tail 1 (KIR3DL1), killer cell immunoglobulin-like receptor, three Ig domains and long cytoplasmic tail 1 (KIR3DL1), killer cell immunoglobulin-like receptor, two Ig domains and long cytoplasmic tail 1 (KIR2DL1), killer cell immunoglobulin-like receptor, two Ig domains and long cytoplasmic tail 3 (KIR2DL3), killer cell lectin-like receptor Cl (KLRC1), killer cell lectin-like receptor C2 (KLRC2), killer cell Killer cell lectin-like receptor C3 (KLRC3), killer cell lectin-like receptor C4 (KLRC4), killer cell lectin-like receptor D1 (KLRD1), killer cell lectin-like receptor K1 (KLRK1), natural cytotoxicity-inducing receptor 3 (NCR3 or NKp30), natural cytotoxicity-inducing receptor 2 (NCR2 or NK-p44), natural cytotoxicity-inducing receptor 1 (NCR1 or NK-p46), CD226 (DNAM-1), cytotoxic and regulatory T cell molecule (CRTAM or CD355), signaling lymphocyte activation molecule family member 1 (SLAMF1), CD48 (SLAMF2), lymphocyte antigen 9 (LY9 or SLAMF3), CD244 (2B4 or SLAMF4), CD84 (SLAMF5), SLAM family member 6 (SLAMF6 or NTB- A), SLAM family member 7 (SLAMF7 or CRACC), CD27 (TNFRSF7), semaphorin 4D (SEMA4D or CD100), CD160 (NK1), an immune checkpoint protein (e.g., PD-1, PD-L1, CTLA-4), and a second epitope of gp120.

[0191] In some embodiments, the antiviral agent is selected from the group consisting of a non-nucleoside reverse transcriptase inhibitor, a protease inhibitor, an entry or fusion inhibitor, and an integrase inhibitor.

[0192] In some embodiments, the first or second therapeutic agent is administered to the patient intratumorally, intravenously, subcutaneously, intraosseously, orally, transdermally, or sublingually. In some embodiments, the first therapeutic agent is administered to the patient before, after, or simultaneously with the second therapeutic agent. In some embodiments, the antibody or antigen-binding fragment thereof is administered prophylactically or therapeutically.

[0193] Passive immunization has been proven to be an effective and safe strategy for the prevention and treatment of viral diseases. (See, for example, Keller et al., Clin. Microbiol. Rev. 13:602-14 (2000); Casadevall, Nat. Biotechnol. 20:114 (2002); Shibata et al., Nat. Med. 5:204-10 (1999); and Igarashi et al., Nat. Med. 5:211-16 (1999), each of which is incorporated herein by reference). Passive immunization using human monoclonal antibodies provides an immediate therapeutic strategy for the emergency prevention and treatment of HIV. Subjects at risk of HIV-related disease or disorder include patients who have come into contact with infected individuals or who have been exposed to HIV in other ways. Prophylactic administration can be performed before the symptoms characteristic of HIV-related disease or disorder appear, thereby preventing or slowing the progression of the disease or disorder.

[0194] For in vivo treatment of human and non-human patients, the patient is administered or provided with a formulation comprising the HIV antibody of the present invention. When used for in vivo treatment, the antibody of the present invention is administered to the patient in a therapeutically effective amount (i.e., an amount that eliminates or reduces the viral load of the patient). The antibody is administered to the human patient according to known methods, such as intravenous administration, for example, by intramuscular, intraperitoneal, intraspinal, subcutaneous, intra-articular, intrasynovial, intrathecal, oral, topical, or inhalation routes, as a bolus or by continuous infusion over a period of time. The antibody may be administered parenterally or intravenously, if possible, to the target cell site. In some embodiments, the antibody is administered by intravenous or subcutaneous administration. The therapeutic composition of the present invention may be administered to the patient or subject systemically, parenterally, or locally. The above parameters for evaluating the success and improvement of disease treatment are easily measurable by routine procedures familiar to physicians.

[0195] For parenteral administration, the antibody may be formulated in a unit dose injectable form (solution, suspension, emulsion) in combination with a pharma- ceutically acceptable parenteral vehicle. Examples of such vehicles include, but are not limited to, water, saline, Ringer's solution, dextrose solution, and 5% human serum albumin. Non-aqueous vehicles include, but are not limited to, fixed oils and ethyl oleate. Liposomes may be used as carriers. The vehicle may contain small amounts of additives, such as substances that enhance isotonicity and chemical stability, e.g., buffers and preservatives. The antibody may be formulated in such vehicles at a concentration of about 1 mg / ml to 10 mg / ml.

[0196] The dosage and dosing regimen will depend on the nature of the infection, e.g., its therapeutic index, the patient, and the The amount of antibody administered depends on a variety of factors, which are readily determined by a physician, such as the patient's medical history. Typically, a therapeutically effective amount of the antibody is administered to the patient. In some embodiments, the amount of antibody administered ranges from about 0.1 mg / kg to about 50 mg / kg of the patient's body weight. Depending on the type and severity of the infection, about 0.1 mg / kg to about 50 mg / kg of body weight (e.g., about 0.1 to 15 mg / kg / dose) of the antibody is an initial candidate dose for administration to the patient, whether by one or more separate administrations or by continuous infusion, for example. The progress of this treatment is easily monitored by conventional methods and assays and based on criteria known to the physician or one of skill in the art. The above parameters for assessing the success and improvement of the treatment of the disease are readily measurable by routine procedures familiar to physicians.

[0197] Other therapeutic regimens may be combined with the administration of the HIV antibody of the present invention. The combined administration includes simultaneous administration using separate or single formulations, and sequential administration in any order, preferably with a period during which both (or all) active agents simultaneously exert their biological activity. Such combined therapy may result in synergistic therapeutic effects. The above parameters for evaluating the success and improvement of disease treatment are easily measurable by routine procedures familiar to physicians.

[0198] b. Methods for reducing viral replication Further provided is a method for reducing the increase in HIV viral titer, viral replication, viral growth or the amount of HIV viral proteins in a subject. According to another embodiment, the method comprises administering to a subject an amount of an HIV antibody effective to reduce the increase in HIV titer, viral replication or the amount of HIV proteins of one or more HIV strains or isolates in the subject.

[0199] According to other embodiments, the present invention provides a method for reducing viral replication or spread of HIV infection to additional host cells or tissues comprising contacting a mammalian cell with an antibody or portion thereof that binds to an antigenic epitope, e.g., an antigenic epitope on gp120.

[0200] C. Combination therapy (1) Combination therapy with two or more anti-HIV antibodies In some embodiments, the disclosure provides a method for treating or preventing HIV infection in a human subject having or at risk of having HIV infection. The method comprises administering to the human subject a therapeutically effective amount of an antibody or antigen-binding fragment, or pharmaceutical composition thereof, as disclosed herein, in combination with a therapeutically effective amount of one or more (e.g., one, two, three, one or two, or one to three) additional therapeutic agents. In one embodiment, a method is provided for treating HIV infection in a human subject suffering from or at risk of suffering from an infection, the method comprises administering to the human subject a therapeutically effective amount of one or more antibodies, or pharma- ceutical acceptable salts thereof, as disclosed herein, in combination with a therapeutically effective amount of one or more (e.g., one, two, three, one or two, or one to three) additional therapeutic agents.

[0201] Antibody Combination Therapy In some embodiments, the disclosed antibodies or antigen-binding fragments thereof are co-administered with a second anti-HIV antibody. In some embodiments, the antibodies or antigen-binding fragments thereof are co-administered with a second anti-HIV antibody that binds to an epitope or region of gp120 selected from the group consisting of: (i) the third variable loop (V3) and / or the high mannose patch containing the N332 oligomannose glycan; (ii) the second variable loop (V2) and / or the apex of the Env trimer; (iii) the CD4 binding site (CD4b); (iv) the gp120 / gp41 interface; or (v) gp120. The above-mentioned epitopes or regions of gp120 bound by broadly neutralizing antibodies are described, for example, in McCoy, Retrovirology (2018) 15:70; Sok and Burton, Nat Immunol. 2018 19(11):1179-1188; Possas, et al., Expert Opin Ther Pat. 2018 Jul;28(7):551-560; and Stephenson and Barouch, Curr HIV / AIDS Rep (2016) 13:31-37, which are incorporated by reference in their entirety for all purposes.

[0202] In some embodiments, the combination therapy involves the co-administration of an antibody or antigen-binding fragment thereof with another anti-HIV broadly neutralizing antibody or bNAb (i.e., a neutralizing antibody that neutralizes multiple HIV-1 virus strains). A variety of bNAbs are known in the art and can be used as combination therapeutic agents. Additional exemplary bNAbs of use include those that comprise a VH and a VL that bind to or compete with an epitope or region of gp120 selected from the group consisting of: (i) the third variable loop (V3) and / or the high mannose patch containing the N332 oligomannose glycan; (ii) the second variable loop (V2) and / or the apex of the Env trimer; (iii) the CD4 binding site (CD4b); (iv) the gp120 / gp41 interface; or (v) the silent face of gp120.Exemplary bNAbs for use in anti-HIV antibody combination therapy include GS-9722 (elipovimab), PGT-121, PGT-121.66, PGT-121.414, PGT-122, PGT-123, PGT-124, PGT-125, PGT-126, PGT-128, PGT-130, PGT-133, PGT-134, PGT-135, PGT-136, PGT-137 , PGT-138, PGT-139, 10-1074, 10-1074-J, VRC24, 2G12, BG18, 354BG8, 354BG18, 354BG42, 354BG33, 354B G129, 354BG188, 354BG411, 354BG426, DH270.1, DH270.6, PGDM12, VRC41.01, PGDM21, PCDN-33A, BF520.1 and VRC29.03 (all of which bind to the V3 glycan region), 2F5, 4E10, M66.6, CAP206-CH12, 10E8, 10E8v4, 10E8-5R-100cF, DH511.11P, 7b2, and LN01 (all of which bind to the MPER of gp41); PG9, PG16, CH01-04 (all of which bind to V1V2-glycans), 2G12 (which binds to glycans in the outer domain); b12, F105, VRC01, VRC07, VRC07-523, VRC03, VRC06, and VRC06b01. VRC08, VRC0801, NIH45-46, GS-9723, GS-5423, 3BNC117, 3BNC60, VRC-PG04, PGV04; CH103, 44-VRC13.01, 1NC9, 12A12, N6, N6LS (VRC-HIVMAB091-00-AB), N49-P7, NC-Cow1, IOMA, CH235 and CH235.12, which contain a VH and VL that bind or compete with N49P6, N49P7, N49P11, N49P9 and N60P25 (all of which bind to the CD4 binding site).

[0203] In some embodiments, the combination therapy includes N332 oligomannose glycan and is selected from the group consisting of GS-9722 (elipovimab), PGT-121, PGT-121.66, PGT-121.414, PGT-122, PGT-123, PGT-124, PGT-125, PGT-126, PGT-128, PGT-130, PGT-133, PGT-134, PGT-135, PGT-136, PGT-137, PGT-138, PGT-139, PGT-200, PGT-201, PGT-202, PGT-203, PGT-204, PGT-205, PGT-206, PGT-207, PGT-208, PGT-209, PGT-300, PGT-310, PGT-311, PGT-312, PGT-313, PGT-314, PGT-315, PGT-316, PGT-317, PGT-318, PGT-319, PGT-320, PGT-321, PGT-322, PGT-323, PGT-324, PGT-325, PGT-326, PGT-328, PGT-329, PGT-400, PGT-410, PGT-421, PGT-422, PGT-423, PGT-424, PGT-425, PGT-426, PGT-427, PGT-428, PGT-430, PGT-431, PGT-432, PGT-433, PGT-434, PGT-435, PGT-436, PGT-437, PGT-438, PGT GT-137, PGT-138, PGT-139, 10-1074, 10-1074-J, VRC24, 2G12, BG18, 354BG8, 354BG18, 354BG42, 354BG33, 35 4BG129, 354BG188, 354BG411, 354BG426, DH270.1, DH270.6, PGDM12, VRC41.01, PGDM21, PCDN-33A, BF520.1 and and VRC29.03. Additional broadly neutralizing antibodies that may be used in the second antigen-binding domain of the multispecific antigen-binding molecules described herein that bind to gp120 in the third variable loop (V3) and / or the high mannose patch, including the N332 oligomannose glycan, and that compete with or include the VH and VL regions from an antibody selected from the group consisting of: WO 2012 / 030904; WO 2014 / 032661; WO 2015 / 023631; WO 2015 / 023662; WO 2015 / 023671; WO 2015 / 023681; WO 2015 / 023697; WO 2015 / 023698; WO 2015 / 02369 ... 63059;WO 2016 / 149698;WO 2017 / 106346;WO 2018 / 075564, WO 2018 / 125813; WO 2018 / 23714 8, WO 2019 / 226829, WO 2020 / 023827, WO2020 / 056145, and Kerwin, et al., J Pharm Sci. 2020 Jan;109(1):233-246.

[0204] In some embodiments, the combination therapy includes any of the following: b12, F105, VRC01, VRC07, VRC07-523, VRC03, VRC06, VRC06b01, VRC06b01, VRC07-523, VRC0 ... and antibodies that compete with or comprise the VH and VL regions of an antibody selected from the group consisting of VRC08, VRC0801, NIH45-46, 3BNC117, GS-9723, GS-5423, 3BNC60, VRC-PG04, PGV04; CH103, 44-VRC13.01, 1NC9, 12A12, N6, N49-P7, NC-Cow1, IOMA, CH235, and CH235.12, N49P6, N49P7, N49P11, N49P9, and N60P25. Additional broadly neutralizing antibodies that bind to gp120 in the CD4 binding site (CD4b) and may be used in the second antigen-binding domain of the multispecific antigen-binding molecules described herein are described, for example, in WO 2011 / 038290; WO 2012 / 158948; WO 2013 / 016468; WO 2013 / 192589; WO 2013 / 086533; WO 2015 / 128846; WO 2016 / 149698. ; WO 2016 / 149695; WO 2018 / 075564; WO 2018 / 125813; WO 2018 / 237357, and U.S. Pat. No. 9,493,544. 9, and 9,879,068. Additional antibodies that bind to epitopes or regions of gp120 on CD4b that can be administered in combination with or simultaneously with the present antibodies are described, for example, in Schommers, et al., Cell (2020) , 14:131-135, 2011. 180: 471-489;Freund, et al., Sci Transl. Med (2017) 9: eaal2144; Diskin, et al., J Exp Med (2013) 210: 1235-49; and Scheid, et al., Science (2011) 333: 1633-1637. The foregoing publications are incorporated herein by reference in their entireties for all purposes.

[0205] In some embodiments, the combination therapy includes an antibody that binds to an epitope or region of gp120 in the second variable loop (V2) and / or the Env trimer apex and competes with or includes the CDRs and / or VH and VL regions of an antibody selected from the group consisting of PG9, PG16, PGC14, PGG14, PGT-142, PGT-143, PGT-144, PGT-145, CH01, CH59, PGDM1400, CAP256, CAP256-VRC26.08, CAP256-VRC26.09, CAP256-VRC26.25, PCT64-24E, and VRC38.01.

[0206] In some embodiments, the combination therapy includes an antibody that binds to an epitope or region of gp120 at the gp120 / gp41 interface and competes with the CDRs and / or VH and VL regions from an antibody selected from the group consisting of PGT-151, CAP248-2B, 35O22, 8ANC195, ACS202, VRC34, and VRC34.01. Examples of antibodies include antibodies that are or contain the antibodies.

[0207] In some embodiments, the combination therapy includes an antibody that binds to an epitope or region of the gp120 silent face and competes with or includes a second VH and VL region of the antibody VRC-PG05.

[0208] In some embodiments, the combination therapy includes an antibody that binds to an epitope or region of gp41 in the membrane proximal region (MPER) and competes with or includes a second VH and VL region of an antibody selected from the group consisting of 10E8, 10E8v4, 10E8-5R-100cF, 4E10, DH511.11P, 2F5, 7b2, and LN01. In some embodiments, the combination therapy includes an antibody that binds to an epitope or region of KLIC ("KLIC"), an invariant site of the transmembrane protein gp41, and competes with or includes a second VH and VL region of Clone3 human monoclonal antibody (Cl3hmAb) (Protheragen). See, e.g., Vanini, et al., AIDS. (1993) 7(2):167-74.

[0209] In some embodiments, the combination therapy includes an antibody that binds to an epitope or region of a gp41 fusion peptide and competes with or includes a second VH and VL region from an antibody selected from the group consisting of VRC34 and ACS202.

[0210] In some embodiments, the combination therapy includes a multispecific, e.g., bispecific or trispecific, antibody that binds to an HIV antigen. Examples of HIV bispecific and trispecific antibodies include MGD014, B12BiTe, BiIA-SG, TMB bispecific, SAR-441236, VRC-01 / PGDM-1400 / 10E8v4, 10E8.4 / iMab, and 10E8v4 / PGT121-VRC01.

[0211] Prior to administration, bNAbs can be improved to have enhanced drug-like properties, reduced immunogenicity, enhanced ADCC, and suitable pharmacokinetic properties. Such antibodies have been shown to bind to HIV envelope glycoproteins expressed on the surface of virions or infected cells, mediating both direct neutralization of the virus and potent NK, monocyte, and PBMC killing of these cells. This property allows the antibodies to treat HIV infection by neutralizing the virus and kill and eliminate latently HIV-infected cells in infected individuals, leading to a sterilizing treatment of HIV.

[0212] In various embodiments, all of the antibodies administered in the combination anti-HIV antibody therapy may possess Fc and / or post-translational modifications that extend serum half-life and / or enhance effector activity, as described above.

[0213] In various embodiments, the antibody or antigen-binding fragment, and optionally the combined bNAb, can be delivered in vivo and expressed in vivo, for example, from administered mRNA or engineered B cells. Examples of bNAbs delivered in vivo include AAV8-VRC07; mRNA encoding the anti-HIV antibody VRC01; and engineered B cells encoding 3BNC117 (Hartweger et al., J. Exp. Med. 2019, 1301).

[0214] (2) Combination therapy with other anti-HIV drugs In some embodiments, a method is provided for treating or preventing HIV infection in a human suffering from or at risk of infection, comprising administering to a human a therapeutic agent comprising one or more (e.g., one, two, three, one or two, or one to three) of the therapeutic agents disclosed herein. administering to the human a therapeutically effective amount of an antibody or antigen-binding fragment in combination with a therapeutically effective amount of an additional therapeutic agent. In one embodiment, a method for treating HIV infection in a human suffering from or at risk of infection is provided, comprising administering to the human a therapeutically effective amount of an antibody or antigen-binding fragment in combination with a therapeutically effective amount of one or more (e.g., 1, 2, 3, 1 or 2, or 1-3) additional therapeutic agents as disclosed herein.

[0215] In one embodiment, a pharmaceutical composition is provided that includes an antibody or antigen-binding fragment as disclosed herein in combination with one or more (e.g., 1, 2, 3, 1 or 2, or 1-3) additional therapeutic agents and a pharma- ceutically acceptable carrier, diluent, or excipient.

[0216] In some embodiments, methods are provided for treating HIV infection, comprising administering to a patient in need thereof a therapeutically effective amount of an antibody or antigen-binding fragment thereof, as described herein, in combination with a therapeutically effective amount of one or more additional therapeutic agents suitable for treating HIV infection.

[0217] In some embodiments, the antibody or antigen-binding fragment thereof is combined with one, two, three, four, or more additional therapeutic agents. In some embodiments, the antibody or antigen-binding fragment thereof is combined with two additional therapeutic agents. In other embodiments, the antibody or antigen-binding fragment thereof is combined with three additional therapeutic agents. In further embodiments, the antibody or antigen-binding fragment thereof is combined with four additional therapeutic agents. The one, two, three, four, or more additional therapeutic agents may be different therapeutic agents selected from the same class of therapeutic agents (e.g., one or more anti-HIV broadly neutralizing antibodies) and / or they may be selected from different classes of therapeutic agents.

[0218] Administration of HIV combination therapy In some embodiments, the antibody or antigen-binding fragment thereof is co-administered with one or more additional therapeutic agents, as described herein. Co-administration of the antibody or antigen-binding fragment disclosed herein with one or more additional therapeutic agents generally refers to simultaneous or sequential administration of the antibody or antigen-binding fragment disclosed herein and one or more additional therapeutic agents, such that a therapeutically effective amount of both the antibody or antigen-binding fragment disclosed herein and one or more additional therapeutic agents are present in the patient's body. When administered sequentially, the combination can be administered in two or more doses.

[0219] Concurrent administration includes simultaneous administration of the antibody or antigen-binding fragment thereof before or after administration of a unit dose of one or more additional therapeutic agents, as well as administration of a unit dose, as described herein. For example, an antibody or antigen-binding fragment thereof described herein may be administered within seconds, minutes, hours, or days of administration of one or more additional therapeutic agents. In some embodiments, a unit dose of an antibody or antigen-binding fragment disclosed herein is administered first, followed by a unit dose of one or more additional therapeutic agents within seconds, minutes, hours, or days. Alternatively, a unit dose of one or more additional therapeutic agents is administered first, followed by a unit dose of an antibody or antigen-binding fragment disclosed herein within seconds, minutes, hours, or days. In other embodiments, a unit dose of an antibody or antigen-binding fragment disclosed herein is administered first, followed by a unit dose of one or more additional therapeutic agents within a few hours (e.g., 1-12 hours, 1-24 hours, 1-36 hours, 1-48 hours, 1-60 hours, 1-72 hours). In yet other embodiments, one or more additional therapeutic agents A unit dose of the therapeutic agent is administered first, followed several hours (e.g., 1-12 hours, 1-24 hours, 1-36 hours, 1-48 hours, 1-60 hours, 1-72 hours) later by a unit dose of an antibody or antigen-binding fragment disclosed herein.

[0220] In some embodiments, the antibodies or antigen-binding fragments disclosed herein are combined with one or more additional therapeutic agents in a single dosage form for simultaneous administration to a patient, e.g., as a solid, liquid or suspension dosage form for oral, intravenous, intramuscular or subcutaneous administration.

[0221] In some embodiments, the antibody or antigen-binding fragment is formulated as a liquid solution or suspension, which may optionally include one or more other compounds useful in the treatment of HIV, hi some embodiments, the liquid solution or suspension may include other active ingredients for the treatment of HIV, such as HIV protease inhibitors, HIV non-nucleoside or non-nucleotide inhibitors of reverse transcriptase, HIV nucleoside or nucleotide inhibitors of reverse transcriptase, HIV integrase inhibitors, HIV non-catalytic site (or allosteric) integrase inhibitors, pharmacokinetic enhancers, and combinations thereof.

[0222] In some embodiments, such liquid solutions or suspensions are suitable for dosing or administration intervals of once a day, once a week (i.e., QW), once every two weeks (i.e., once every other week or once every two weeks, or Q2W), once a month (i.e., QM), or once every two months (i.e., once every other month or once every two months, or Q2M). In some embodiments, the antibody or antigen-binding fragment is administered once a day, once a week (i.e., QW), once every two weeks (i.e., once every other week or once every two weeks, or Q2W), once a month (i.e., QM), once a two-month dose (i.e., once every other month or once every two months, or Q2M), once every three months (i.e., Q3M), once every four months (i.e., Q4M).

[0223] HIV combination therapy In the above embodiments, the additional therapeutic agent can be an anti-HIV agent. HIV protease inhibitors, HIV non-nucleoside or non-nucleotide inhibitors of reverse transcriptase, HIV nucleoside or nucleotide inhibitors of reverse transcriptase, HIV integrase inhibitors, HIV non-catalytic site (or allosteric) integrase inhibitors, HIV entry inhibitors, HIV maturation inhibitors, HIV capsid inhibitors, HIV Tat or Rev inhibitors, immunomodulators (e.g., immunostimulants), immunotherapeutics, immunomodulators, immunotherapeutics, antibody-drug conjugates, gene modifiers, gene editors (CRISPR / Cas9, zinc finger nucleases, homing nucleases, synthetic nucleases, TALENs, etc.), cell therapy (chimeric antigen receptor T cells, CAR-T, and engineered T cell receptors, TCR-T, autologous T cell therapy, etc.), latency reversers, compounds targeting the HIV capsid, immune-based therapies, phosphatidylinositol 3-kinase (PI3K) inhibitors, HIV antibodies, bispecific antibodies and "antibody-like" therapeutic proteins, HIV p17 matrix protein inhibitors, IL-13 antagonists, peptidyl prolyl cis-trans isomerase A modulators, protein disulfide isomerase inhibitors, complement C5a receptor antagonists, DNA methyltransferase inhibitors, fatty acid synthase inhibitors, HIV vif gene modulators, Vif dimerization antagonists, HIV-1 viral infectivity factor inhibitors, TAT protein inhibitors, HIV-1 Nef modulators (e.g., Nef inhibitors), Hck tyrosine kinase modulators, mixed lineage kinase-3 (MLK-3) inhibitors, HIV-1 splicing inhibitors, Rev protein inhibitors, integrin antagonists, nuclear protein inhibitors, splicing factor modulators, COMM domain-containing protein 1 modulators, HIV ribonuclease H inhibitors, retrocyclin modulators, CDK-4 inhibitors, CDK-6 inhibitors, CDK-9 inhibitors, dendritic I CAM-3 grab nonintegrin 1 inhibitors, HIV GAG protein inhibitors, HIV POL protein inhibitors, complement factor H modulators, ubiquitin ligase inhibitors, deoxycytidine kinase inhibitors, cyclin-dependent kinase inhibitors, proprotein convertase PC9 stimulators, ATP-dependent RNA helicase DDX3X inhibitors, reverse transcriptase priming complex inhibitors, G6PD and NADH-oxidase inhibitors, mTOR complex 1 inhibitors, mTOR complex 2 inhibitors, P-glycoprotein modulators, TAT protein inhibitors, prolylendopeptidase inhibitors, phospholipase A2 inhibitors, pharmacokinetic enhancers, HIV gene therapy, TNFα ligand inhibitors, IFN antagonists, HIV vaccines, and combinations thereof.

[0224] In some embodiments, the additional therapeutic agent is selected from the group consisting of HIV combination medications, other drugs treating HIV, HIV protease inhibitors, HIV reverse transcriptase inhibitors, HIV integrase inhibitors, HIV non-catalytic site (or allosteric) integrase inhibitors, HIV entry (fusion) inhibitors, HIV maturation inhibitors, latency reversing agents, HIV capsid inhibitors, HIV Tat or Rev inhibitors, immunomodulatory agents (e.g., immunostimulants), immunotherapeutic agents, immune-based therapies, PI3K inhibitors, HIV antibodies, and bispecific antibodies, and "antibody-like" therapeutic proteins, and combinations thereof.

[0225] HIV combination drugs In some embodiments, the antibodies or antigen-binding fragments described herein are combined with one, two, three, four, or more additional anti-HIV therapeutic agents. Examples of anti-HIV therapeutic agents that may be combined include ATRIPLA® (efavirenz, tenofovir disoproxil fumarate, and emtricitabine); COMPLERA® (EVIPLERA®; rilpivirine, tenofovir disoproxil fumarate, and emtricitabine); STRIBILD® (elvitegravir, cobicistat, tenofovir disoproxil fumarate, and emtricitabine); TRUVADA® (tenofovir disoproxil fumarate and emtricitabine; TDF+FTC; DESCOVY® (tenofovir alafenamide and emtricitabine); ODEFSEY® (tenofovir alafenamide, emtricitabine, and rilpivirine); GENVOYA® (tenofovir alafenamide, emtricitabine, cobicistat, and elvitegravir); BIKTARVY (bictegravir + emtricitabine + tenofovir alafenamide), Adefovir; adefovir dipivoxil; cobicistat; emtricitabine; tenofovir; tenofovir alafenamide and elvitegravir; tenofovir disoproxil; tenofovir disoproxil fumarate; tenofovir alafenamide; tenofovir alafenamide hemifumarate; TRIUMEQ® (dolutegravir, abacavir, and lamivudine); dolutegravir, abacavir sulfate, and lamivudine.Raltegravir; PEGylated Raltegravir; Raltegravir and Lamivudine; Maraviroc; Tenofovir + Emtricitabine + Maraviroc, Enfuvirtide; ALUVIA® (KALETRA®; Lopinavir and Ritonavir); COMBIVIR® (Zidovudine and Lamivudine; AZT+3TC); EPZICOM® (LIVEXA®; Abacavir Sulfate and Lamivudine; ABC+3TC); TRIZIVIR® (Abacavir Sulfate, Zidovudine, and Lamivudine; ABC+AZT+3TC); Atazanavir and Cobicistat; Atazanavir sulfate and cobicistat; Atazanavir sulfate and ritonavir; Darunavir; Darunavir and cobicistat; Dolutegravir and rilpivirine; Dolutegravir and rilpivirine hydrochloride; Dolutegravir, abacavir sulfate, and lamivudine; Lamivudine, nevirapine, and zidovudine; Raltegravir and lamivudine; Doravirine, lamivudine, and tenofovir disoproxil fumarate; Doravirine, lamivudine, and tenofovir disoproxil; Dolutegravir + lamivudine, lamivudine + abacavir + zidovudine, lamivudine +. Abacavir, Lamivudine + Tenofovir Disoproxil Fumarate, Lamivudine + Zidovudine + Nevirapine, Lopinavir + Ritonavir, Lopinavir + Ritonavir + Abacavir + Lamivudine, Lopinavir + Ritonavir + Zidovudine + Lamivudine, Tenofovir + Lamivudine, and Tenofovir Disoproxil Fumarate + Emtricitabine + Rilpivirine Hydrochloride, Lopinavir, Ritonavir, Zidovudine and Lamivudine; Cabotegravir + rilpivirine;Elpida (elsulfavirine; VM-1500; VM-1500A);Rilpivirine;Rilpivirine hydrochloride;Atazanavir sulfate and cobicistat;Atazanavir and cobicistat;Darunavir and cobicistat;Atazanavir;Atazanavir sulfate;Dolutegravir;Elvitegravir;Ritonavir;Atazanavir sulfate and ritonavir;Darunavir;Ramivir These include, but are not limited to, vudine; prolastin; fosamprenavir; fosamprenavir calcium efavirenz; efavirenz, lamivudine, and emtricitabine; etravirine; nelfinavir; nelfinavir mesylate; interferon; didanosine; stavudine; indinavir; indinavir sulfate; tenofovir and lamivudine; zidovudine; nevirapine; saquinavir; saquinavir mesylate; aldesleukin; zalcitabine; tipranavir; amprenavir; delavirdine; delavirdine mesylate; Radha-108 (Receptor); lamivudine and tenofovir disoproxil fumarate; efavirenz, lamivudine, and tenofovir disoproxil fumarate; phosphazide; lamivudine, nevirapine, and zidovudine; abacavir; and abacavir sulfate.

[0226] Other HIV drugs Examples of other drugs for treating HIV that may be combined with the agents of the present disclosure include aspergillin C, acemannan, alisporivir, BanLec, deferiprone, gamimune, metenkephalin, naltrexone, prolastin, REP 9, RPI-MN, VSSP, H1 virus, SB-728-T, 1,5-dicaffeoylquinic acid, rHIV7-shl-TAR-CCR5RZ, AAV-eCD4-Ig gene therapy, MazF gene therapy, BlockAide, Bevirimat derivatives, ABX-464, AG-1105, APH-0812, bryostatin analogs, BIT-225, CYT-107, CS-TATI-1, fluoro-beta-D-arabinose nucleic acid (FANA) modified antisense oligonucleotides, FX-101, Griffithsin, HGTV-43, HPH-116, HS-10234, hydroxychloroquine, IMB-10035, IMO-3100, IND-02, JL-18008, LADAVRU, MK-1376, MK-2048, MK-4250, MK-8507, MK-8558, MK-8591 (Islatravil), NOV-205, OB-002H, ODE-Bn-TFV, M1-TFV, PA-1050040 (PA-040), PC-707, PGN-007, QF-036, S-648414, These include SCY-635, SB-9200, SCB-719, TR-452, TEV-90110, TEV-90112, TEV-90111, TEV-90113, RN-18, DIACC-1010, Fasnall, Immuglo, 2-CLIPS peptide, HRF-4467, thrombospondin analogs, TBL-1004HI, VG-1177, xl-081, rfhSP-D, [18F]-MC-225, URMC-099-C, RES-529, and VIR-576.

[0227] HIV protease inhibitors In some embodiments, the antibodies or antigen-binding fragments described herein are combined with HIV protease inhibitors. Examples of HIV protease inhibitors include amprenavir, atazanavir, brecanavir, darunavir, fosamprenavir, fosamprenavir calcium, indinavir, indinavir sulfate, lopinavir, nelfinavir, nelfinavir mesylate, ritonavir, saquinavir, saquinavir mesylate, tipranavir, AEBL-2, DG-17, GS-1156, TMB-657 (PPL-100), T-169, BL-008, MK-8122, TMB-607, GRL-02031, and and TMC-310911.

[0228] HIV RNase H inhibitors In some embodiments, the antibodies or antigen-binding fragments described herein are combined with an HIV RNase H inhibitor. An example of an HIV RNase H inhibitor that can be combined includes NSC-727447.

[0229] HIV Nef inhibitors In some embodiments, the antibodies or antigen-binding fragments described herein are combined with an HIV Nef inhibitor. An example of an HIV Nef inhibitor that can be combined includes FP-1.

[0230] HIV reverse transcriptase inhibitors In some embodiments, the antibody or antigen-binding fragment described herein is combined with a non-nucleoside or non-nucleotide inhibitor. Examples of HIV non-nucleoside or non-nucleotide inhibitors of reverse transcriptase include dapivirine, delavirdine, delavirdine mesylate, doravirine, efavirenz, etravirine, lentinan, nevirapine, rilpivirine, ACC-007, ACC-008, AIC-292, F-18, KM-023, PC-1005, VM-1500A-LAI, PF-3450074, ersulfavirine (sustained release oral, HIV infection), ersulfavirine (long-acting injectable nanosuspension, HIV infection), and ersulfavirine (VM-1500).

[0231] In some embodiments, the antibodies or antigen-binding fragments described herein are combined with an HIV nucleoside or nucleotide inhibitor. Examples of HIV nucleoside or nucleotide inhibitors of reverse transcriptase include adefovir, adefovir dipivoxil, azuvudine, emtricitabine, tenofovir, tenofovir alafenamide, tenofovir alafenamide fumarate, tenofovir alafenamide hemifumarate, tenofovir disoproxil, tenofovir disoproxil fumarate, tenofovir octadecyloxyethyl ester (AGX-1009), tenofovir disoproxil hemifumarate, VIDEX®, and VIDEXEC®. (didanosine, ddl), abacavir, abacavir sulfate, alovudine, apricitabine, censavudine, didanosine, elvucitabine, festinavir, fosalvudine tidoxyl, CMX-157, dapivirine, doravirine, etravirine, OCR-5753, tenofovir disoproxil orotate, tidoxylfodivudine, lamivudine, phosphazide, stavudine, zalcitabine, zidovudine, lobafovir etalafenamid (GS-9131), GS-9148, MK-8504, MK-8591, MK-858, and KP-1461.

[0232] HIV integrase inhibitors In some embodiments, the antibodies or antigen-binding fragments described herein are combined with HIV integrase inhibitors. Examples of HIV integrase inhibitors include elvitegravir, elvitegravir (slow-release microcapsules), curcumin, derivatives of curcumin, ticholic acid, derivatives of ticholic acid, 3,5-dicaffeoylquinic acid, derivatives of 3,5-dicaffeoylquinic acid, aurintricarboxylic acid, derivatives of aurintricarboxylic acid, caffeic acid phenethyl ester, derivatives of caffeic acid phenethyl ester, tyrphostin, derivatives of tyrphostin, quercetin, quercetin derivatives, raltegravir, PEGylated raltegravir, dolutegravir, JTK-351, bictegravir, AVX-15567, cabotegravir (long-acting injectable), diketoquinoline-4-1 derivatives, integrase-LEDGF inhibitors, ledgins, M-522, M-532, MK-0536, NSC-310217, NSC-371056, NSC-48240, NSC-642710, NSC-699171, NSC-699172, NSC-699 173, NSC-699174, stilbenedisulfonic acid, T-169, STP-0404, VM-3500, and cabotegravir.

[0233] In some embodiments, the antibodies or antigen-binding fragments described herein are combined with HIV non-catalytic or allosteric integrase inhibitors (NCINI). Examples of HIV non-catalytic or allosteric integrase inhibitors (NCINI) include CX-05045, CX-05168, and CX-14442.

[0234] HIV entry inhibitors In some embodiments, the antibodies or antigen-binding fragments described herein are combined with HIV entry inhibitors. Examples of HIV entry (fusion) inhibitors include AAR-501, LBT-5001, cenicriviroc, CCR5 inhibitors, gp41 inhibitors, CD4 attachment inhibitors, gp120 inhibitors, gp160 inhibitors, and CXCR4 inhibitors.

[0235] In some embodiments, the antibody or antigen-binding fragment described herein is combined with a CCR5 inhibitor. Examples of CCR5 inhibitors include aplaviroc, vicriviroc, maraviroc, maraviroc (long-acting injectable nanoemulsion), cenicriviroc, leronlimab (PRO-140), adaptavir (RAP-101), nifeviroc (TD-0232), anti-GP120 / CD4 or CCR5 bispecific antibody, B-07, MB-66, polypeptide C25P, TD-0680, thioraviroc, and vMIP (Haimipu).

[0236] In some embodiments, the antibody or antigen-binding fragment described herein is combined with a CXCR4 inhibitor. Examples of CXCR4 inhibitors include plerixafor, ALT-1188, N15 peptide, and vMIP (Haimipu).

[0237] In some embodiments, the antibodies or antigen-binding fragments described herein are combined with gp41 inhibitors. Examples of gp41 inhibitors include albuvirtide, enfuvirtide, griffithsin (gp41 / gp120 / gp160 inhibitor), BMS-986197, enfuvirtide biobetter, enfuvirtide biosimilar, HIV-1 fusion inhibitor (P26-Bapc), ITV-1, ITV-2, ITV-3, ITV-4, CPT-31, Cl3hmAb, PIE-12 trimer, and sifuvirtide.

[0238] In some embodiments, the antibodies or antigen-binding fragments described herein are combined with a CD4 adhesion inhibitor. Examples of CD4 adhesion inhibitors include ibalizumab and CADA analogs.

[0239] In some embodiments, the antibodies or antigen-binding fragments described herein are combined with gp120 inhibitors. Examples of gp120 inhibitors include anti-HIV microbicides, Radha-108 (receptor) 3B3-PE38, BanLec, bentonite-based nanomedicines, fostemsavir tromethamine, IQP-0831, VVX-004, and BMS-663068.

[0240] In some embodiments, the antibodies or antigen-binding fragments described herein are combined with a gp160 inhibitor. Examples of gp160 inhibitors that can be combined include fanquinolines.

[0241] HIV maturation inhibitors In some embodiments, the antibodies or antigen-binding fragments described herein are Combined with a V maturation inhibitor. Examples of HIV maturation inhibitors include BMS-955176, GSK-3640254, and GSK-2838232.

[0242] Latent antagonist In some embodiments, the antibodies or antigen-binding fragments described herein are combined with HIV latency reversing agents. Examples of latency reversing agents that may be combined with one or more multispecific antigen-binding molecules described herein include IL-15 receptor agonists (e.g., ALT-803; interleukin-15 / Fc fusion proteins (e.g., XmAb24306)); recombinant interleukin-15 (e.g., AM0015, NIZ-985); pegylated IL-15 (e.g., NKTR-255); toll-like receptor (TLR) agonists (including TLR7 agonists, e.g., GS-9620 and TLR8 agonists, e.g., GS-9688), histone deacetylase (HDAC) inhibitors, proteasome inhibitors such as Velcade, protein kinase C (PKC) activators, Smyd2 inhibitors, BET-bromodomes, and the like. These include BRD4 inhibitors, ionomycin, IAP antagonists (inhibitors of apoptotic proteins such as APG-1387, LBW-242, etc.), SMAC mimetics (such as ciapavir, TL32711, LCL161, GDC-0917, HGS1029, AT-406, etc.), Devio-1143, PMA, SAHA (suberanilohydroxamic acid, or suberoyl, anilide, and hydroxamic acid), NIZ-985, IL-15 modulating antibodies (including IL-15, IL-15 fusion proteins, and IL-15 receptor agonists, such as ALT-803), JQ1, ubiquitin inhibitors such as disulfiram, amphotericin B, and largazole analogs, APH-0812, and GSK-343. Examples of HDAC inhibitors include romidepsin, vorinostat, and panobinostat. Examples of PKC activators include indolactam, prostratin, ingenol B, and DAG-lactone.

[0243] Toll-like receptor (TLR) agonists In some embodiments, the antibodies or antigen-binding fragments described herein are combined with an agonist of a Toll-like receptor (TLR), such as an agonist of TLR1 (NCBI Gene ID: 7096), TLR2 (NCBI Gene ID: 7097), TLR3 (NCBI Gene ID: 7098), TLR4 (NCBI Gene ID: 7099), TLR5 (NCBI Gene ID: 7100), TLR6 (NCBI Gene ID: 10333), TLR7 (NCBI Gene ID: 51284), TLR8 (NCBI Gene ID: 51311), TLR9 (NCBI Gene ID: 54106), and / or TLR10 (NCBI Gene ID: 81793). Exemplary TLR7 agonists that may be co-administered or combined with one or more multispecific antigen binding molecules described herein include AL-034, DSP-0509, GS-9620 (vesatolimod), vesatolimod analogs, LHC-165, TMX-101 (imiquimod), GSK-2245035, resiquimod, DSR-6434, DSP-3025, IMO-4200, MCT-465, MEDI-9197, 3M-051, SB-9922, 3M- 052, Limtop, TMX-30X, TMX-202, RG-7863, RG-7854, RG-7795, as well as US20100143301 (Gilead Sciences), US20110098248 (Gilead Sciences, and US20090047249 (Gilead Sciences), US20140045849(Janssen), US20140073642(Janssen), WO2014 / 056953(Janssen), WO2014 / 076221(Ja nssen), WO2014 / 128189(Janssen), US20140350031(Janssen), WO2014 / 023813(Janssen), US20080234251(Array Biopharma), US20080306050(Array Biopharma), US20100029585(Ventirx Pharma), US20110092485(Ventirx Pharma), US20110118235(Vent TLR7 / TLR8 agonists that may be co-administered include, but are not limited to, compounds disclosed in US20120082658 (Ventirx Pharma), US20120219615 (Ventirx Pharma), US20140066432 (Ventirx Pharma), US20140088085 (Ventirx Pharma), US20140275167 (Novira Therapeutics), and US20130251673 (Novira Therapeutics). TLR7 / TLR8 agonists that may be co-administered include NKTR-262, telluritolimod, and BDB-001. Examples of TLR8 agonists that may be co-administered or combined with one or more multispecific antigen binding molecules described herein include E-6887, IMO-4200, IMO-8400, IMO-9200, MCT-465, MEDI-9197, motolimod, resiquimod, GS-9688, VTX-1463, VTX-763, 3M-051, 3M-052, and US2014 0045849(Janssen), US20140073642(Janssen), WO2014 / 056953(Janssen), WO2014 / 076221(Janssen), W O2014 / 128189(Janssen), US20140350031(Janssen), WO2014 / 023813(Janssen), US20080234251(Array US20140066432 (Ventirx Pharma), US20140088085 (Ventirx Pharma), US20140275167 (Novira Therapeutics), and compounds described in US20130251673 (Novira Therapeutics).Examples of TLR9 agonists that may be co-administered include, but are not limited to, AST-008, cobitolimod, CMP-001, IMO-2055, IMO-2125, ritenimod, MGN-1601, BB-001, BB-006, IMO-3100, IMO-8400, IR-103, IMO-9200, agatolimod, DIMS-9054, DV-1079, DV-1179, AZD-1419, lefitolimod (MGN-1703), CYT-003, CYT-003-QbG10, tilsotolimod, and PUL-042. Examples of TLR3 agonists include lintatorimod, polyICLC, RIBOXXON®, Apoxxim, RIBOXXIM®, IPH-33, MCT-465, MCT-475, and ND-1.1. Examples of TLR4 agonists include G-100 and GSK-1795091.

[0244] Histone deacetylase (HDAC) inhibitors In some embodiments, the antibodies or antigen-binding fragments described herein are combined with inhibitors of histone deacetylase, e.g., histone deacetylase 1, histone deacetylase 9 (HDAC9, HD7, HD7b, HD9, HDAC, HDAC7, HDAC7B, HDAC9B, HDAC9FL, HDRP, MITR; Gene ID: 9734). Examples of HDAC inhibitors include, but are not limited to, abexinostat, ACY-241, AR-42, BEBT-908, belinostat, CKD-581, CS-055 (HBI-8000), CT-101, CUDC-907 (fimepinostat), entinostat, gibinostat, mocetinostat, panobinostat, pracinostat, xinostat (JNJ-26481585), resminostat, licorinostat, romidepsin, SHP-141, TMB-ADC, valproic acid (VAL-001), vorinostat, tinostamustine, remetinostat, and entinostat.

[0245] Cyclin-dependent kinase (CDK) inhibitors or antagonists In some embodiments, the antibodies or antigen-binding fragments described herein are combined with an inhibitor or antagonist of a cyclin-dependent kinase (CDK), such as cyclin-dependent kinase 4 (CDK4; NCBI Gene ID: 1019), cyclin-dependent kinase 6 (CDK6; NCBI Gene ID: 1021), or cyclin-dependent kinase 9 (CDK9; NCBI Gene ID: 1025). In some embodiments, the CDK4 / CDK6 / CDK9 inhibitor or antagonist is selected from the group consisting of VS2-370.

[0246] Stimulator of interferon genes (STING) agonists In some embodiments, the antibodies or antigen-binding fragments described herein are combined with a stimulator of interferon genes (STING). In some embodiments, the STING receptor agonist or activator is selected from the group consisting of ADU-S100 (MIW-815), SB-11285, MK-1454, SR-8291, AdVCA0848, GSK-532, SYN-STING, MSA-1, SR-8291, 5,6-dimethylxanthenone-4-acetic acid (DMXAA), cyclic GAMP (cGAMP), and cyclic di-AMP.

[0247] RIG-I agonists In some embodiments, the antibodies or antigen-binding fragments described herein are combined with an agonist of DExD / H-box helicase 58 (DDX58; aka RIG-I, RIG1, RIGI, RLR-1, SGMRT2; NCBI Gene ID: 23586). In some embodiments, the agents described herein are combined with a RIG-I modulator, such as RGT-100, or a NOD2 modulator, such as SB-9200 (aka GS 9992; Inarigivir), and IR-103. An exemplary RIG-I agonist is KIN1148, described by Hemann, et al., J Immunol May 1, 2016, 196 (1 Supplement) 76.1. Additional RIG-I agonists are described, for example, in Elion, et al., Cancer Res. (2018) 78(21):6183-6195; and Liu, et al., J Virol. (2016) 90(20):9406-19. RIG-I agonists are commercially available, for example, from Invivogen (invivogen.com).

[0248] LAG-3 and TIM-3 inhibitors In some embodiments, the antibodies or antigen-binding fragments described herein are combined with anti-TIM-3 (also known as Hepatitis A Virus Cellular Receptor 2 antibody (HAVCR2; NCBI Gene ID: 84868)), such as TSR-022, LY-3321367, MBG-453, INCAGN-2390. In some embodiments, the antibodies or antigen-binding fragments described herein are combined with anti-LAG-3 (lymphocyte activation) (NCBI Gene ID: 3902) antibodies, such as leratolimab (ONO-4482), LAG-525, MK-4280, REGN-3767, INCAGN2385.

[0249] Capsid Inhibitors In some embodiments, the antibodies or antigen-binding fragments described herein are combined with a capsid inhibitor. Examples of capsid inhibitors that may be combined with the agents of the present disclosure include capsid polymerization inhibitors or capsid disrupting compounds, HIV nucleocapsid p7 (NCp7) inhibitors such as azodicarbonamide, HIV p24 capsid protein inhibitors, GS-6207 (lenacapavir), GS-CA1, AVI-621, AVI-101, AVI-201, AVI-301, and the AVI-CAN1-15 series, PF-3450074, and those described in International Patent Publication No. WO2019 / 087016. Compounds include:

[0250] Immune-Based Therapies In some embodiments, the antibodies or antigen-binding fragments described herein are combined with immune-based therapies. Examples of immune-based therapies include toll-like receptor (TLR) modulators, such as TLR1, TLR2, TLR3, TLR4, TLR5, TLR6, TLR7, TLR8, TLR9, TLR10, TLR11, TLR12, TLR13; programmed cell death protein 1 (PD-1) modulators; programmed death ligand 1 (PD-L1) modulators; IL-15 modulators (e.g., IL-15 receptor agonists (e.g., ALT-803; interleukin-15 / Fc fusion proteins (e.g., XmAb24306)); recombinant interleukin-15 (e.g., AM0015, NIZ-985); pegylated IL-15 (e.g., NKTR-255)); DermaVir; interleukin-7; plaquenil (hydroxychloroquine); interferon; proleukin (aldesleukin, IL-2); interferon alpha; interferon alpha-2b; interferon alpha-n3; pegylated interferon alpha; interferon gamma; hydroxyurea; mycophenolate mofetil (MPA) and its ester derivative mycophenolate mofetil (MMF); ribavirin; the polymer polyethyleneimine (PEI); gepon; IL-12; WF-10; VGV-1; MOR-22; BMS-936559; CYT-107, normferon, pegylated interferon alpha-2a, pegylated interferon alpha-2b, RPI-MN, STING modulators, RIG-I modulators, NOD2 modulators, SB-9200, and IR-103.

[0251] In some embodiments, the antibody or antigen-binding fragment described herein is combined with a TLR agonist. Examples of TLR agonists include, but are not limited to, vesatolimod (GS-9620), lefitolimod, tilsotolimod, lintatolimod, DSP-0509, AL-034, G-100, cobitolimod, AST-008, motolimod, GSK-1795091, GSK-2245035, VTX-1463, GS-9688, LHC-165, BDB-001, RG-7854, and tellulatorimod.

[0252] CD47-targeting agents In various embodiments, the antibody or antigen-binding fragment described herein is combined with an agent that interferes with the binding of CD47 to SIRPα, such as an agent that targets CD47 or an agent that targets SIRPα. In various embodiments, the antibody or antigen-binding fragment described herein is combined with an inhibitor of CD47 (IAP, MER6, OA3; NCBI gene ID: 961). Examples of CD47 inhibitors include, but are not limited to, anti-CD47 mAb (Vx-1004), anti-human CD47 mAb (CNTO-7108), CC-90002, CC-90002-ST-001, humanized anti-CD47 antibody (Hu5F9-G4; magrolimab), NI-1701, NI-1801, RCT-1938, and TTI-621. In some embodiments, the CD47 inhibitor is magrolimab.

[0253] Immune Checkpoint Receptor Protein Modulators In various embodiments, the antibodies or antigen-binding fragments described herein are combined with one or more blockers or inhibitors of inhibitory immune checkpoint proteins or receptors and / or one or more stimulatory agents, activators or agonists of one or more stimulatory immune checkpoint proteins or receptors. Blocking or inhibiting an inhibitory immune checkpoint may positively regulate T cell or NK cell activation and inhibit immune evasion of infected cells. Activation or stimulation of a stimulatory immune checkpoint may inhibit immune evasion of infected cells. In various embodiments, immune checkpoint proteins or receptors modulate T cell responses (e.g., as reviewed in Xu, et al., J Exp Clin Cancer Res. (2018) 37:110). In various embodiments, immune checkpoint proteins or receptors modulate NK cell responses (e.g., as reviewed in Davis, et al., Semin Immunol. (2017) 31:64-75, and Chioss, et al., J Exp Clin Cancer Res. (2017) 31:64-75). one, et al., Nat Rev Immunol. (2018) 18(11):671-688).

[0254] Examples of immune checkpoint proteins or receptors that may be combined with the antibodies or antigen-binding fragments described herein include CD27, CD70; CD40, CD40LG; CD47, CD48 (SLAMF2), transmembrane and immunoglobulin domain containing 2 (TMIGD2, CD28H), CD84 (LY9B, SLAMF5), CD96, CD160, MS4A1 (CD20), CD244 (SLAMF4); CD276 (B7H3); V-set domain containing T-cell activation inhibitor 1 (VTCN1, B7H4); V-set immunoregulatory receptor (VSIR, B7H5, VISTA); immunoglobulin superfamily member 11 (IGSF11, VSIG3); natural killer cell cytotoxicity receptor 3 ligand 1 (NCR3LG1, B7H6); HERV-H LTR-associated 2 (HHLA2, B7H7); inducible T cell costimulatory factor (ICOS, CD278); inducible T cell costimulatory ligand (ICOSLG, B7H2); TNF receptor superfamily member 4 (TNFRSF4, OX40); TNF superfamily member 4 (TNFSF4, OX40L); TNFRSF8 (CD30), TNFSF8 (CD30L); TNFRSF10A (CD261, DR4, TRAILR1), TNFRSF9 (CD137), TNFSF9 (CD137L); TNFRSF10B (CD262, DR5, TRAILR2), TNFRSF10 (TRAIL); TNFRSF14 (HVEM, CD270), TNFSF14 (HVEML); CD272 (B and T lymphocyte-associated (BTL A);TNFRSF17 (BCMA, CD269), TNFSF13B (BAFF);TNFRSF18 (GITR), TNFSF18 (GITRL);MHC class I polypeptide-related sequence A (MICA);MHC class I polypeptide-related sequence B (MICB);CD274 (CD274, PDL1, PD-L1);programmed cell death 1 (PDCD1, PD1, PD-1);cytotoxic T lymphocyte-associated protein 4 (CTLA4, CD152);CD80 (B7-1), CD28;nectin cell adhesion molecule 2 (NECTIN2, CD112);CD226 (DNAM-1);poliovirus receptor (PVR) cell adhesion molecule (PVR, CD155);PVR-related immunoglobulin domain containing (PVRIG, CD112R);T-cell immunoreceptor with Ig and ITIM domains (TIGIT); T-cell immunoglobulin and mucin domain containing 4 (TIMD4; TIM4); Hepatitis A virus cellular receptor 2 (HAVCR2, TIMD3, TIM3); Galectin 9 (LGALS9); Lymphocyte activation 3 (LAG3, CD223); Signaling lymphocyte activation molecule family member 1 (SLAMF1, SLAM, CD150); Lymphocyte antigen 9 (LY9, CD229, SLAMF3); SLAM family member 6 (SLAMF6, CD352); SLAM family member 7 (SLAMF7, CD319); UL16 binding protein 1 (ULBP1); UL16 binding protein 2 (ULBP2); UL16 binding protein 3 (ULBP3); Retinoic acid early transcript 1E (RAET1E; ULBP4); Retinoic acid early transcript 1G (RAET1G; ULBP5); Retinoic acid early transcript 1L (RAET1L; ULBP6; lymphocyte activation 3 (CD223); killer cell immunoglobulin like receptor, three Ig domains and long cytoplasmic tail 1 (KIR, CD158E1); killer cell lectin like receptor C1 (KLRC1, NKG2A, CD159A); killer cell lectin like receptor K1 (KLRK1, NKG2D, CD314); killer cell lectin-like receptor C2 (KLRC2, CD159c, NKG2C); killer cell lectin-like receptor C3 (KLRC3, NKG2E); killer cell lectin-like receptor; Killer cell immunoglobulin-like receptor, 2 Ig domains and long cytoplasmic tail 1 (KIR2DL1); killer cell immunoglobulin-like receptor, 2 Ig domains and long cytoplasmic tail 2 (KIR2DL2); killer cell immunoglobulin-like receptor, 2 Ig domains and long cytoplasmic tail 3 (KIR2DL3); killer cell immunoglobulin-like receptor, 3 Ig domains and long cytoplasmic tail 1 (KIR3DL1); killer cell lectin-like receptor D1 (KLRD1); and SLAM family member 7 (SLAMF7).

[0255] In various embodiments, the antibodies or antigen-binding fragments described herein are combined with one or more blockers or inhibitors of one or more T cell inhibitory immune checkpoint proteins or receptors. Exemplary T cell inhibitory immune checkpoint proteins or receptors include CD274 (CD274, PDL1, PD-L1); programmed cell death 1 ligand 2 (PDCD1LG2, PD-L2, CD273); programmed cell death 1 (PDCD1, PD1, PD-1); cytotoxic T lymphocyte-associated protein 4 (CTLA4, CD152); CD276 (B7H3); V-set domains including T cell activation inhibitor 1 (VTCN1, B7H4); V-set immunoregulatory receptor (VSIR, B7H5, VISTA); immunoglobulin superfamily member 11 (IGSF11, VSIG3); TNFRSF14 (HVEM, CD270), TNFSF14 (HVEML); CD272 (B and T lymphocyte associated (BTLA)); PVR-related immunoglobulin domains including (PVRIG, CD112R); Ig domains including These include, but are not limited to, T cell immunoreceptor with in and ITIM domains (TIGIT); lymphocyte activation 3 (LAG3, CD223); hepatitis A virus cell receptor 2 (HAVCR2, TIMD3, TIM3); galectin 9 (LGALS9); killer cell immunoglobulin-like receptor, 3 Ig domains and long cytoplasmic tail 1 (KIR, CD158E1); killer cell immunoglobulin-like receptor, 2 Ig domains and long cytoplasmic tail 1 (KIR2DL1); killer cell immunoglobulin-like receptor, 2 Ig domains and long cytoplasmic tail 2 (KIR2DL2); killer cell immunoglobulin-like receptor, 2 Ig domains and long cytoplasmic tail 3 (KIR2DL3); and killer cell immunoglobulin-like receptor, 3 Ig domains and long cytoplasmic tail 1 (KIR3DL1). In various embodiments, the antibodies or antigen-binding fragments described herein are combined with one or more agonists or activators of one or more T cell stimulatory immune checkpoint proteins or receptors.Exemplary T cell stimulatory immune checkpoint proteins or receptors include, but are not limited to, CD27, CD70; CD40, CD40LG; inducible T cell costimulator (ICOS, CD278); inducible T cell costimulatory ligand (ICOSLG, B7H2); TNF receptor superfamily member 4 (TNFRSF4, OX40); TNF superfamily member 4 (TNFSF4, OX40L); TNFRSF9 (CD137), TNFSF9 (CD137L); TNFRSF18 (GITR), TNFSF18 (GITRL); CD80 (B7-1), CD28; nectin cell adhesion molecule 2 (NECTIN2, CD112); CD226 (DNAM-1); CD244 (2B4, SLAMF4), poliovirus receptor (PVR) cell adhesion molecule (PVR, CD155). See, e.g., Xu, et al., J Exp. See Clin Cancer Res. (2018) 37:110.

[0256] In various embodiments, the antibodies or antigen-binding fragments described herein are combined with one or more blockers or inhibitors of one or more NK cell inhibitory immune checkpoint proteins or receptors. Exemplary NK cell inhibitory immune checkpoint proteins or receptors include killer cell immunoglobulin-like receptor, three Ig domains and a long cytoplasmic tail 1 (KIR, CD158E1); killer cell immunoglobulin-like receptor, two Ig domains and a long cytoplasmic tail 1 (KIR2DL1); killer cell immunoglobulin-like receptor, two Ig domains and a long cytoplasmic tail 2 (KIR2D L2); killer cell immunoglobulin-like receptor, two Ig domains and long cytoplasmic tail 3 (KIR2DL3); killer cell immunoglobulin-like receptor, three Ig domains and long cytoplasmic tail 1 (KIR3DL1); killer cell lectin-like receptor C1 (KLRC1, NKG2A, CD159A); and killer cell lectin-like receptor D1 (KLRD1, CD94). In various embodiments, the antibodies or antigen-binding fragments described herein are combined with one or more agonists or activators of one or more NK cell stimulatory immune checkpoint proteins or receptors. Exemplary NK cell stimulatory immune checkpoint proteins or receptors include, but are not limited to, CD16, CD226 (DNAM-1); CD244 (2B4, SLAMF4); killer cell lectin-like receptor K1 (KLRK1, NKG2D, CD314); SLAM family member 7 (SLAMF7). For example, Davis, et al., Semin Immunol. (2017) 31:64-75; Fang, et al., Semin Immunol. (2 017) 31:37-54; and Chiossone, et al., Nat Rev Immunol. (2018) 18(11):671-688.

[0257] [In some embodiments, the one or more immune checkpoint inhibitors comprise a proteinaceous (e.g., an antibody or fragment thereof, or an antibody mimetic) inhibitor of PD-L1 (CD274), PD-1 (PDCD1), or CTLA4. In some embodiments, the one or more immune checkpoint inhibitors comprise a small organic molecule inhibitor of PD-L1 (CD274), PD-1 (PDCD1), or CTLA4.

[0258] Examples of CTLA4 inhibitors that may be co-administered include ipilimumab, tremelimumab, BMS-986218, AGEN1181, AGEN1884, BMS-986249, MK-1308, REGN-4659, ADU-1604, CS-1002, BCD-145, APL-509, JS-007, BA-3071, ONC-392, AGEN-2041, JHL-1155, KN-044, CG-0161, ATOR-1144, PBI- These include, but are not limited to, 5D3H5, BPI-002, and the multispecific inhibitors FPT-155 (CTLA4 / PD-L1 / CD28), PF-06936308 (PD-1 / CTLA4), MGD-019 (PD-1 / CTLA4), KN-046 (PD-1 / CTLA4), MEDI-5752 (CTLA4 / PD-1), XmAb-20717 (PD-1 / CTLA4), and AK-104 (CTLA4 / PD-1).

[0259] Examples of PD-L1 (CD274) or PD-1 (PDCD1) inhibitors that may be co-administered include pembrolizumab, nivolumab, cemiplimab, pidilizumab, AMP-224, MEDI0680 (AMP-514), spartalizumab, atezolizumab, avelumab, durvalumab, BMS-936559, CK-301, PF-06801591, BGB-A317 (tislelizumab), GLS-010 (WBP-3055), AK-103 (HX-008), AK-105, CS-1003, HLX-10, MGA-012, BI-754091, AGEN-2034, JS-001 (toriparimab), JNJ-63723283, genolimuzumab (CBT-501), LZM-009, BCD-100, LY-3300054, SHR-1201, SHR-1210 (camrelizumab), Sym-021, ABBV-181 (budigalimab), PD1-PIK, BAT-1306, (MSB0010718C), CX-072, CBT-502, TSR-042 (dostallimab ), MSB-2311, JTX-4014, BGB-A333, SHR-1316, CS-1001, WBP-3155, KN-035, IBI-308 (sintilimab), HLX-20, KL-A167, STI-A1014, STI-A1015 (IMC-001), BCD-135, FAZ-053, TQB-2450, MDX 1105-01, GS-4224, GS-4416, INCB086550, MAX10181, and the multispecific inhibitor FPT-155 (CTLA4 / PD-L1 / CD28), PF-06936308(PD-1 / CTLA4), MGD-013(PD-1 / LAG-3), FS-118(LAG-3 / P D-L1)MGD-019(PD-1 / CTLA4), KN-046(PD-1 / CTLA4), MEDI-5752(CTLA4 / PD-1), RO-7121661(PD-1 / TIM-3), XmAb-20717(PD-1 / CTLA4), AK-104(CTLA4 / PD-1), M7824(PD- L1 / TGFβ-EC domain), CA-170 (PD-L1 / VISTA), CDX-527 (CD27 / PD-L1), LY-3415244 (TIM3 / PDL1), and INBRX-105 (4-1BB / PDL1).

[0260] In some embodiments, the small molecule inhibitor of CD274 or PDCD1 is selected from the group consisting of GS-4224, GS-4416, INCB086550, and MAX 10181. In some embodiments, the small molecule inhibitor of CTLA4 comprises BPI-002.

[0261] In various embodiments, the antibodies or antigen-binding fragments described herein are combined with anti-TIGIT antibodies such as etigilimab, BMS-986207, tiragolumab (also known as MTIG-7192A; RG-6058; RO 7092284), AGEN1307, AGEN1327, AGEN1777, COM-902, IBI-939, AB154, MG1131, and EOS884448 (EOS-448).

[0262] TNF receptor superfamily (TNFRSF) member agonists or activators In various embodiments, the antibodies or antigen-binding fragments described herein are agonists of one or more TNF receptor superfamily (TNFRSF) members, such as agonists of one or more of TNFRSF1A (NCBI Gene ID: 7132), TNFRSF1B (NCBI Gene ID: 7133), TNFRSF4 (OX40, CD134; NCBI Gene ID: 7293), TNFRSF5 (CD40; NCBI Gene ID: 958), TNFRSF6 ( FAS, NCBI gene ID: 355), TNFRSF7 (CD27, NCBI gene ID: 939), TNFRSF8 (CD30, NCBI gene ID: 943), TNFRSF9 (4-1BB, CD137, NCBI gene ID: 3604), TNFRSF10A (CD261, DR4, TRAILR1, NCBI gene ID: 8797), TNFRSF10B (CD262, DR5, TRAILR2, NCBI gene ID: 8795), TNFRSF10C (CD263, TR AILR3, NCBI gene ID: 8794), TNFRSF10D (CD264, TRAILR4, NCBI gene ID: 8793), TNFRSF11A (CD265, RANK, NCBI gene ID: 8792), TNFRSF11B (NCBI gene ID: 4982), TNFRSF12A (CD266, NCBI gene ID: 51330), TNFRSF13B (CD267, NCBI gene ID: 23495), TNFRSF13C (CD268, NCBI gene ID: 115650), TNFRSF16 (NGFR, CD271, NCBI Gene ID: 4804), TNFRSF17 (BCMA, CD269, NCBI Gene ID: 608), TNFRSF18 (GITR, CD357, NCBI Gene ID: 8784), TNFRSF19 (NCBI Gene ID: 55504), TNFRSF21 (CD358, DR6, NCBI Gene ID: 27242), and TNFRSF25 (DR3, NCBI Gene ID: 8718).

[0263] Examples of anti-TNFRSF4 (OX40) antibodies that may be co-administered include, but are not limited to, MEDI6469, MEDI6383, MEDI0562 (tabolixizumab), MOX R0916, PF-04518600, RG-7888, GSK-3174998, INCAGN1949, BMS-986178, GBR-8383, ABBV-368, and those described in WO2016179517, WO2017096179, WO2017096182, WO2017096281, and WO2018089628.

[0264] Examples of anti-TNFRSF5 (CD40) antibodies that may be co-administered include, but are not limited to, RG7876, SEA-CD40, APX-005M, and ABBV-428.

[0265] In some embodiments, the anti-TNFRSF7 (CD27) antibody varlilumab (CDX-1127) is co-administered.

[0266] Examples of anti-TNFRSF9 (4-1BB, CD137) antibodies that may be co-administered include, but are not limited to, urelumab, utomilumab (PF-05082566), AGEN2373, and ADG-106.

[0267] Examples of anti-TNFRSF18 (GITR) antibodies that can be co-administered include, but are not limited to, MEDI1873, FPA-154, INCAGN-1876, TRX-518, BMS-986156, MK-1248, GWN-323, and those described in WO2017096179, WO2017096276, WO2017096189, and WO2018089628. In some embodiments, an antibody or fragment thereof that co-targets TNFRSF4 (OX40) and TNFRSF18 (GITR) is co-administered. Such antibodies are described, for example, in WO2017096179 and WO2018089628.

[0268] Interleukin receptor agonists In some embodiments, the antibodies or antigen-binding fragments described herein are directed to interleukin receptor agonists, such as IL-2, IL-7, IL-15, IL-10, IL-12 agonists; examples of IL-2 receptor agonists, such as Proleukin (aldesleukin, IL-2); pegylated IL-2 (e.g., NKTR-214); modified variants of IL-2 (e.g., THOR-707), bempegaldesleukin, AIC-284, ALKS, -4230, CUI-101, Neo-2 / 15; IL-15 receptor agonists such as ALT-803, NKTR-255, hetIL-15, interleukin-15 / Fc fusion proteins, AM-0015, NIZ-985, SO-C101, IL-15 scintrin (pegylated IL-15), P-22339, and IL-15-PD-1 fusion protein N-809; examples of IL-7 include CYT-107.

[0269] Examples of additional interleukin receptor agonists that may be combined with the antibodies or antigen-binding fragments described herein include interferon alpha; interferon alpha-2b; interferon alpha-n3; pegylated interferon alpha; interferon gamma; Flt3 agonists such as CDX-301; Gepone; Normferon, pegylated interferon alpha-2a, pegylated interferon alpha-2b, RPI-MN.

[0270] Bispecific and trispecific natural killer (NK)-cell engagement In various embodiments, the antibodies or antigen-binding fragments described herein are directed to bispecific NK cell engaging (BiKE) or trispecific NK cell engaging (TriKE) (e.g., Fc-less) or NK cell activating receptors, such as CD16A, C-type lectin receptors (CD94 / NKG2C, NKG2D, NKG2E / H, and and NKG2F), natural killer cytotoxicity receptors (NKp30, NKp44, and NKp46), killer cell C-type lectin-like receptors (NKp65, NKp80), Fc receptors FcγR (mediating antibody-dependent cellular cytotoxicity), SLAM family receptors (2B4, SLAM6, SLAM7, etc.), killer cell immunoglobulin-like receptors (KIR) (KIR-2DS and KIR-3DS), DNAM-1, and CD137 (4-1BB), are combined with bispecific antibodies (e.g., with Fc) against. Exemplary anti-CD16 bispecific antibodies, BiKE or TriKE that may be co-administered include AFM26 (BCMA / CD16A) and AFM-13 (CD16 / CD30). Optionally, the anti-CD16 binding bispecific molecule may or may not have an Fc. BiKE and TriKE are described, for example, in Felices, et al., Methods Mol Biol. (2016) 1441:333-346; Fang, et al. I., Semin Immunol. (2017) 31:37-54. Examples of trispecific NK cell engagement (TRiKE) include OXS-3550, and CD16-IL-15-B7H3TriKe.

[0271] Phosphatidylinositol 3-kinase (PI3K) inhibitors In some embodiments, the antibody or antigen-binding fragment described herein is combined with a PI3K inhibitor. Examples of PI3K inhibitors include idelalisib, alpelisib, bupallisib, CAI orotate, copanlisib, duvelisib, gedatricisib, neratinib, panulisib, perifosine, pictilisib, piralalisib, pukitinib mesylate, rigosertib, rigosertib sodium, sonolicisib, taselisib, AMG-319, AZD-8186, BAY-1082439, CLR-1401, CLR-457, CUDC-907, DS-7423, EN-3342, GSK-2126458, GSK-2269577, GSK -2636771, INCB-040093, LY-3023414, MLN-1117, PQR-309, RG-7666, RP-6530, RV-1729, SAR-245409, SAR-260301, SF-1126, TGR-1202, UCB-5857, VS-5584, XL-765, and ZSTK-474.

[0272] Alpha-4 / beta-7 antagonists In some embodiments, the antibodies or antigen-binding fragments described herein are combined with an alpha-4 / beta-7 antagonist. Examples of integrin alpha-4 / beta-7 antagonists include PTG-100, TRK-170, abrilumab, etrolizumab, carotegrass methyl, and vedolizumab.

[0273] Pharmacokinetic Enhancers In some embodiments, the antibodies or antigen-binding fragments described herein are combined with a pharmacokinetic enhancer. Examples of pharmacokinetic enhancers include cobicistat and ritonavir.

[0274] Additional Therapeutic Agents Examples of additional therapeutic agents include those described in WO 2004 / 096286 (Gilead Sciences); WO 2006 / 015261 (Gilead Sciences); WO 2006 / 110157(Gilead Sciences);WO 2012 / 003497(Gilead Sciences);WO 2012 / 003498(Gilead Sciences);WO 2012 / 145728(Gilead Sciences);WO 2013 / 006738(Gilead Sciences);WO 2013 / 159064(Gilead WO 2014 / 100323 (Gilead Sciences), US 2013 / 0165489 (University of Pennsylvania), US 2014 / 02213 78 (Japan Tobacco), US 2014 / 0221380 (Japan Tobacco); WO 2009 / 062285 (Boehringer Ingelheim); WO 2010 / 130034 (Boehringer Ingelheim); WO 2013 / 006792 (Pharma Resources), US 20140221356 (Gilead Sciences), US 20100143301 (Gilead Sciences), and WO 2013 / 091096 (Boehringer Ingelheim).

[0275] HIV combination therapy In certain embodiments, the antibodies or antigen-binding fragments described herein are selected from the group consisting of ATRIPLA® (efavirenz, tenofovir disoproxil fumarate, and emtricitabine); COMPLERA® (EVIPLERA®; rilpivirine, tenofovir disoproxil fumarate, and emtricitabine); STRIBILD® (elvitegravir, cobicistat, tenofovir disoproxil fumarate, and emtricitabine); TRUVADA® (tenofovir disoproxil fumarate and emtricitabine; TDF+FTC); DESCOVY® (tenofovir arafenib, ribavirin ... tenofovir alafenamide and emtricitabine; ODEFSEY® (tenofovir alafenamide, emtricitabine, and rilpivirine); GENVOYA® (tenofovir alafenamide, emtricitabine, cobicistat, and elvitegravir); adefovir; adefovir dipivoxil; cobicistat; emtricitabine; tenofovir; tenofovir disoproxil; tenofovir disoproxil fumarate; tenofovir alafenamide; tenofovir alafenamide hemifumarate; TRIUMEQ® (dolutegravir, abacavir, and lamivudine); dolutegravir, abacavir sulfate, and lamivudine.Raltegravir; Raltegravir and lamivudine; Maraviroc; Enfuvirtide; ALUVIA® (KALETRA®; Lopinavir and ritonavir); COMBIVIR® (Zidovudine and lamivudine; AZT+3TC); EPZICOM® (LIVEXA®; Abacavir sulfate and lamivudine; ABC+3TC); TRIZIVIR® (Abacavir sulfate, Zidovudine, and Lamivudine; ABC+AZT+3TC); Rilpivirine; Rilpivirine hydrochloride; Atazanavir sulfate and cobicistat; Atazanavir and cobicistat; Darunavir and cobicistat; Atazanavir; Atazanavir sulfate; Dolutegravir; Elvitegravir; Ritonavir; Atazanavir sulfate and ritonavir; Darunavir; Lamivudine; Prola and abacavir sulfate; aldesleukin; zalcitabine; tipranavir; amprenavir; delavirdine; delavirdine mesylate; Radha-108 (Receptor); lamivudine and tenofovir disoproxil fumarate; efavirenz, lamivudine, and tenofovir disoproxil fumarate; phosphazides; lamivudine, nevirapine, and zidovudine; abacavir; and abacavir sulfate.

[0276] It will be understood by one of skill in the art that the additional therapeutic agent may fall into more than one class described above.

[0277] In certain embodiments, the antibodies or antigen-binding fragments described herein are combined with an HIV nucleoside or nucleotide inhibitor of reverse transcriptase and an HIV non-nucleoside inhibitor of reverse transcriptase. In other specific embodiments, the antibodies or antigen-binding fragments described herein are combined with an HIV nucleoside or nucleotide inhibitor of reverse transcriptase and an HIV non-nucleoside inhibitor of reverse transcriptase. and an HIV protease inhibitor compound. In further embodiments, the antibodies or antigen-binding fragments described herein are combined with an HIV nucleoside or nucleotide inhibitor of reverse transcriptase, an HIV non-nucleoside inhibitor of reverse transcriptase, and a pharmacokinetic enhancer. In some embodiments, the antibodies or antigen-binding fragments described herein are combined with at least one HIV nucleoside inhibitor of reverse transcriptase, an integrase inhibitor, and a pharmacokinetic enhancer. In other embodiments, the antibodies or antigen-binding fragments described herein are combined with two HIV nucleoside or nucleotide inhibitors of reverse transcriptase.

[0278] In certain embodiments, the antibodies or antigen-binding fragments described herein are combined with abacavir sulfate, tenofovir, tenofovir disoproxil, tenofovir disoproxil fumarate, tenofovir disoproxil hemifumarate, tenofovir alafenamide, or tenofovir alafenamide hemifumarate.

[0279] In certain embodiments, the antibodies or antigen-binding fragments described herein are combined with tenofovir, tenofovir disoproxil, tenofovir disoproxil fumarate, tenofovir alafenamide, or tenofovir alafenamide hemifumarate.

[0280] In certain embodiments, the antibodies or antigen-binding fragments described herein are combined with a first additional therapeutic agent selected from the group consisting of abacavir sulfate, tenofovir, tenofovir disoproxil, tenofovir disoproxil fumarate, tenofovir alafenamide, and tenofovir alafenamide hemifumarate, and a second additional therapeutic agent selected from the group consisting of emtricitabine and lamivudine.

[0281] In certain embodiments, the antibodies or antigen-binding fragments described herein are combined with a first additional therapeutic agent selected from the group consisting of tenofovir, tenofovir disoproxil, tenofovir disoproxil fumarate, tenofovir alafenamide, and tenofovir alafenamide hemifumarate, and a second additional therapeutic agent, wherein the second additional therapeutic agent is emtricitabine.

[0282] In some embodiments, the antibodies or antigen-binding fragments described herein are combined with one or more additional therapeutic agents at a therapeutically effective dosage ranging, e.g., from 1 mg to 50 mg, 75 mg, 100 mg, 150 mg, 200 mg, 250 mg, 300 mg, 400 mg, 500 mg, 1000 mg, or 1500 mg of the antibody or antigen-binding fragment. In some embodiments, the antibodies or antigen-binding fragments described herein are combined with one or more additional therapeutic agents at a therapeutically effective dosage ranging, e.g., from about 0.1 mg / kg to about 0.5 mg / kg, 1 mg / kg, 2 mg / kg, 3 mg / kg, 4 mg / kg, 5 mg / kg, 8 mg / kg, 10 mg / kg, 15 mg / kg, 20 mg / kg, 25 mg / kg, 30 mg / kg, 35 mg / kg, 40 mg / kg, 45 mg / kg, or 50 mg / kg of the antibody or antigen-binding fragment. In some embodiments, the antibodies or antigen-binding fragments described herein are combined with one or more additional therapeutic agents at a therapeutically effective dose ranging, for example, from about 5 mg to about 10 mg, 20 mg, 25 mg, 50 mg, 100 mg, 125 mg, 150 mg, 250 mg, 300 mg, 500 mg, 1000 mg, or 1500 mg of the antibody or antigen-binding fragment.

[0283] In some embodiments, an antibody or antigen-binding fragment described herein is combined with 5-30 mg of tenofovir alafenamide fumarate, tenofovir alafenamide hemifumarate, or tenofovir alafenamide, and 200 mg of emtricitabine. is combined with 5-10, 5-15, 5-20, 5-25, 25-30, 20-30, 15-30, or 10-30 mg of tenofovir alafenamide fumarate, tenofovir alafenamide hemifumarate, or tenofovir alafenamide, and 200 mg of emtricitabine. In some embodiments, an antibody or antigen-binding fragment described herein is combined with 10 mg of tenofovir alafenamide fumarate, tenofovir alafenamide hemifumarate, or tenofovir alafenamide, and 200 mg of emtricitabine. In some embodiments, an antibody or antigen-binding fragment described herein is combined with 25 mg of tenofovir alafenamide fumarate, tenofovir alafenamide hemifumarate, or tenofovir alafenamide, and 200 mg of emtricitabine. In some embodiments, an antibody or antigen-binding fragment described herein is combined with an agent provided herein at a dose of either the antibody or antigen-binding fragment (e.g., 1 mg to 500 mg of antibody or antigen-binding fragment as described herein) as each combination of doses is specifically and individually listed.

[0284] In some embodiments, the antibodies or antigen-binding fragments described herein are combined with 200-400 mg of tenofovir disoproxil fumarate, tenofovir disoproxil hemifumarate, or tenofovir disoproxil, and 200 mg of emtricitabine. In some embodiments, the antibodies or antigen-binding fragments described herein are combined with 200-250, 200-300, 200-350, 250-350, 250-400, 350-400, 300-400, or 250-400 mg of tenofovir disoproxil fumarate, tenofovir disoproxil hemifumarate, or tenofovir disoproxil, and 200 mg of emtricitabine. In some embodiments, an antibody or antigen-binding fragment described herein is combined with 300 mg of tenofovir disoproxil fumarate, tenofovir disoproxil hemifumarate, or tenofovir disoproxil, and 200 mg of emtricitabine. The antibody or antigen-binding fragment may be combined with an agent provided herein in any dosage amount (e.g., 1 mg to 500 mg of antibody or antigen-binding fragment), as each combination of dosage amounts is specifically and individually described.

[0285] Long-acting HIV inhibitors In some embodiments, the antibody or antigen-binding fragment described herein can be co-administered with a long-acting HIV inhibitor. Examples of drugs being developed as long-acting HIV inhibitors include, but are not limited to, cabotegravir LA, rilpivirine LA, any integrase LA, VM-1500 LAI, maraviroc (LAI), tenofovir implant, MK-8591 implant, long-acting dolutegravir.

[0286] In one embodiment, the kit comprises an antibody or antigen-binding fragment described herein in combination with one or more (e.g., 1, 2, 3, 1 or 2, or 1-3) additional therapeutic agents.

[0287] HIV vaccine In some embodiments, the antibodies or antigen-binding fragments described herein are combined with an HIV vaccine. Examples of HIV vaccines include peptide vaccines, recombinant subunit protein vaccines, live vector vaccines, DNA vaccines, HIV MAG DNA vaccines, CD4-derived peptide vaccines, combination vaccines, adenovirus vector vaccines (e.g., Ad5, Ad26, or Ad35), simian adenovirus (chimpanzee, gorilla, resus, i.e., rhAd), adeno-associated virus vector vaccines, chimpanzee adenovirus vaccines (e.g., ChAdOX1, ChA, d68, ChAd3, ChAd63, ChAd83, ChAd155, ChAd157, Pan5, Pan6, Pan7, Pan9), coxsackievirus-based vaccines, enterovirus-based vaccines, gorilla adenovirus vaccines, lentivirus vector-based vaccines, two-segment or three-segment arenavirus-based vaccines (e.g., LCMV, Pichinde), trimer-based HIV-1 vaccines, measles virus-based vaccines, flavivirus vector-based vaccines, tobacco mosaic virus vector-based vaccines, varicella-zoster virus-based vaccines, and avian serovar IgE vaccines. poxvirus-based vaccines (such as modified vaccine virus Ankara (MVA), NYVAC derived from an orthopoxvirus, and ALVAC (canarypox virus) strains derived from an avipoxvirus); fowlpoxvirus-based vaccines; rhabdovirus-based vaccines such as vesicular stomatitis virus (VSV) and Maraba virus; recombinant human CMV (rhCMV)-based vaccines; alphavirus-based vaccines such as Semliki Forest virus, Venezuelan equine encephalitis virus, and Sindbis virus (e.g., Lauer, et al., Clin Vaccine Immunol. (2017) 24(1): e00298-16); LNP-formulated mRNA-based therapeutic vaccines; and LNP-formulated self-replicating RNA / self-amplifying RNA vaccines.

[0288] Examples of HIV vaccines include anti-CD40.Env-gp140 vaccine, Ad4-EnvC150, BG505 SOSIP.664 gp140 adjuvant vaccine, BG505 SOSIP.GT1.1 gp140 adjuvant vaccine, Chimigen HIV vaccine, ConM SOSIP.v7 gp140, rgp120(AIDSVAX), ALVAC HIV(vCP1521 / AIDSVAX B / E(gp120)(RV144), monomeric gp120 HIV-1 subtype C vaccine, MPER-656 liposomal subunit vaccine, Remune, ITV-1, Contre Vir, Ad5-ENVA-48, DCVax-001 (CDX-2401), Vacc-4x, Vacc-C5, VAC-3S, multiclade DNA recombinant adenovirus-5 (rAd5), rAd5 gag-pol env A / B / C vaccine, Pennvax-G, Pennvax-GP, Pennvax-G / MVA-CMDR, HIV-TriMix-mRNA vaccine, HIV-LAMP-vax, Ad35, Ad35-GRIN, NAcGM3 / VSSP ISA-51, poly-ICLC adjuvanted vaccine, TatImmune, GTU-multiHIV (FIT-06), ChAdV63.HIVconsv, gp140δV2.TV1+ MF-59, rVSVIN HIV-1 gag vaccine, SeV-EnvF, SeV-Gag vaccine, AT-20, DNK-4, ad35-Grin / ENV, TBC-M4, HIVAX, HIVAX-2, N123-VRC-34.01-derived epitope-based HIV vaccine, NYVAC-HIV-PT1, NYVAC-HIV-PT4, DNA-HIV-PT123, rAAV1-PG9DP, GOVX-B11, GOVX-B21, GOVX-C55, TVI-HIV-1, Ad-4 (Ad4-env Clade C + Ad4-mGag), Paxvax, EN41-UGR7C, EN41-FPA2, ENOB-HV-11, PreVaxTat, AE-H, MYM-V101, CombiHIVvac, ADVAX, MYM-V201, MVA-CMDR, MagaVax, DNA-Ad5 gag / pol / nef / nev (HVTN505), MVATG-17401, ETV-01, CDX-1401, DNA and Sev vector vaccines expressing SCaVII, rcAD26.MOS1.HIV-Env, Ad26.Mod.HIV vaccine, Ad26.Mod.HIV+MVA mosaic vaccine+gp140, AGS-004, AVX-101, AVX-201, PEP-6409, SAV-001, ThV-01, TL-01, TUTI-16, VGX-3300, VIR-1111, IHV-001, and virus-like particle vaccines such as pseudovirion vaccines, CombiVICHvac, LFn-p24 B / C fusion vaccine, GTU-based DNA vaccine, HIV gag / pol / nef / env D NA vaccine, anti-TAT HIV vaccine, conjugate polypeptide vaccine, dendritic cell vaccine, gag-based DNA vaccine, GI-2010, gp41 HIV-1 vaccine, HIV vaccine (PIKA adjuvant), I i-key / MHC class II epitope hybrid peptide vaccine, ITV-2, ITV-3, ITV-4, LIPO-5, multi-clade Env vaccine, MVA vaccine, Pennvax-GP, pp71-deficient HCMV vector HIV gag vaccine, recombinant peptide vaccine (HIV infection), NCI, rgp160 HIV vaccine, RNActive HIV vaccine, SCB-703, Tat Oyi vaccine, TBC-M4, therapeutic HIV vaccine, UBI HIV gp120, Vacc-4x + romidepsin, mutant gp120 polypeptide vaccine, rAd5 gag-pol env A / B / C vaccine, DNA.HTI and MVA.HTI, VRC-HIVDNA016-00-VP+VRC-HIVADV014-00-VP, INO-6145, JNJ-9220, gp145 C.6980; eOD-GT8 60mer-based vaccine, PD-201401, env(A,B,C,A / E) / gag(C) DNA vaccine, gp120(A,B,C,A / E) protein vaccine, PDPHV-201401, Ad4-EnvCN54, EnvSeq-1 Envs HIV-1 vaccine (GLA-SE adjuvanted), HIV p24gag prime boost plasmid DNA vaccine, HIV-1 iglb12 neutralizing VRC-01 antibody-stimulating anti-CD4 vaccine, MVA-BN HIV-1 vaccine regimen, UBI These include, but are not limited to, HIV gp120, an mRNA-based prophylactic vaccine, VPI-211, and TBL-1203HI.

[0289] D.Kit In other aspects, the present disclosure further provides kits useful for performing diagnostic and prognostic assays using the antibodies, polypeptides, and nucleic acids of the present invention. The kits of the present invention include suitable containers containing HIV antibodies, polypeptides, or nucleic acids of the present invention in either labeled or unlabeled form. Furthermore, when the antibodies, polypeptides, or nucleic acids are provided in a labeled form suitable for indirect binding assays, the kits further include reagents for performing the appropriate indirect assays. For example, the kits may include one or more suitable containers containing an enzyme substrate or a derivatizing agent, depending on the nature of the label. Control samples and / or instructions for use may also be included. The present invention also provides kits for detecting the presence of HIV antibodies or nucleotide sequences of HIV antibodies of the present invention in biological samples by PCR or mass spectrometry.

[0290] In some embodiments, the kit further comprises a pharma- ceutically effective amount of a pharma- ceutically acceptable dosage unit of an anti-HIV agent. The two pharma-ceutically acceptable dosage units may be in the form of a single pharma-ceutically acceptable dosage unit. In some embodiments, the anti-HIV agent is selected from the group consisting of a non-nucleoside reverse transcriptase inhibitor, a protease inhibitor, an entry or fusion inhibitor, and an integrase inhibitor.

[0291] In some embodiments, the kit also includes a container that contains the composition and may also contain informational material. The informational material may be explanatory, educational, transactional, or other material related to the methods described herein and / or the use of the agent for therapeutic benefit. In one embodiment, the kit also includes an additional therapeutic agent as described above. For example, the kit includes a first container that contains the composition and a second container for the additional therapeutic agent.

[0292] The informational material of the kit is not limited in format. In some embodiments, the informational material may include information regarding the composition's manufacture, concentration, expiration date, batch or manufacturing site information, etc. In one embodiment, the informational material may include information regarding the composition's manufacture, concentration, expiration date, batch or manufacturing site information, etc. In one embodiment, the informational material may include information regarding the composition's manufacture, e.g., in an appropriate dose, dosage form, or mode of administration (e.g., a dose, dosage form, or mode of administration described herein). In one embodiment, the instructions provide a dosing regimen, dosing schedule, and / or route of administration of the composition or additional therapeutic agent. The information can be provided in a variety of formats, including printed text, computer readable material, video recording, audio recording, or information containing links or addresses to substantive material.

[0293] The kit may include one or more containers for the composition. In some embodiments, the kit includes separate containers, dividers, or compartments for the composition and informational material. For example, the composition may be in a bottle or vial and the informational material may be in a plastic sleeve or packet. In other embodiments, the separate elements of the kit are included in a single, undivided container. For example, the composition is included in a bottle or vial having informational material attached thereto in the form of a label. In some embodiments, the kit includes a plurality (e.g., a pack) of individual containers, each of which includes one or more unit dosage forms (e.g., dosage forms described herein) of the agent.

[0294] The kit may include a device suitable for administration of the composition or other suitable delivery device. The device may be provided pre-loaded with one or both of the agents or may be empty but suitable for loading. Such a kit may include a syringe capable of injecting the antibody contained within the kit into an animal, such as a human.

[0295] E. Definition To aid in understanding the detailed description of the compositions and methods according to the present disclosure, some explicit definitions are provided to facilitate a clear disclosure of the various aspects of the disclosure. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which this disclosure belongs.

[0296] The term "antibody" (Ab) as used herein includes monoclonal antibodies, polyclonal antibodies, multispecific antibodies (e.g., bispecific and polyreactive antibodies), and antibody fragments. Thus, the term "antibody" as used in any context herein is meant to include, but is not limited to, any specific binding member, immunoglobulin class and / or isotype (e.g., IgG1, IgG2, IgG3, IgG4, IgM, IgA, IgD, IgE, and IgM); as well as biologically relevant fragments or specific binding members thereof, including but not limited to Fab, F(ab')2, Fv, and scFv (single chain or related entities). In the art, an antibody is understood to be a glycoprotein comprising at least two heavy (H) chains and two light (L) chains interconnected by disulfide bonds or antigen-binding portions thereof. The heavy chain is composed of a heavy chain variable region (VH) and a heavy chain constant region (CH1, CH2, and CH3). The light chain is composed of a light chain variable region (VL) and a light chain constant region (CL). The variable regions of both heavy and light chains are composed of framework regions (FWR) and complementarity determining regions (CDR). The four FWR regions are relatively conserved, while the CDR regions (CDR1, CDR2, and CDR3) represent hypervariable regions and are arranged from the NH2-terminus to the COOH-terminus as follows: FWR1, CDR1, FWR2, CDR2, FWR3, CDR3, FWR4. The variable regions of the heavy and light chains contain binding domains that interact with antigens, while the constant regions, depending on the isotype, may mediate the binding of immunoglobulins to host tissues or factors.

[0297] The definition of "antibody" as used herein also includes chimeric antibodies, humanized antibodies, and recombinant antibodies, transgenic non-human animals, and human antibodies produced from antibodies selected from libraries using enrichment techniques available to those of skill in the art.

[0298] A "neutralizing antibody" is one that neutralizes the ability of HIV to initiate and / or perpetuate infection in host and / or target cells in vitro. The present disclosure provides neutralizing human monoclonal antibodies, which recognize antigens from HIV, e.g., gp120 polypeptide. In some embodiments, a "neutralizing antibody" can inhibit entry of HIV-1 virus, e.g., SF162 and / or JR-CSF, with a neutralization index >1.5 or >2.0 (Kostrikis LG et al., J. Virol., 70(1): 445-458 (1996)).

[0299] In some embodiments, the neutralizing antibody is a broadly neutralizing antibody (such as a monoclonal antibody) that targets HIV-L. The term "broadly neutralizing antibody" refers to an antibody that neutralizes multiple HIV-1 virus species (from various clades and different strains within a clade) in a neutralization assay. A broadly neutralizing antibody can neutralize at least 2, 3, 4, 5, 6, 7, 8, 9 or more different strains of HIV-1, strains belonging to the same or different clades. In some embodiments, a broadly neutralizing antibody can neutralize multiple HIV-1 species belonging to at least 2, 3, 4, 5, or 6 different clades. In some embodiments, the inhibitory concentration of the antibody can be less than about 0.0001 pg / mL, less than about 0.001 pg / mL, less than about 0.01 pg / mL, less than about 0.1 pg / mL, less than about 0.5 pg / mL, less than about 1.0 pg / mL, less than about 5 pg / mL, less than about 10 pg / mL, less than about 25 mg / mL, less than about 50 mg / mL, or less than about 100 mg / mL to neutralize about 50% of input virus in a neutralization assay.

[0300] "Antibody fragment" includes a portion of an intact antibody, such as the antigen-binding or variable region of the intact antibody. Examples of antibody fragments include, but are not limited to, Fab, Fab', F(ab')2, and Fv fragments; diabodies; linear antibodies (see, e.g., U.S. Pat. No. 5,641,870; Zapata et al., Protein Eng. 8(10): 1057-1062

[1995] ); single-chain antibody molecules; single-chain variable fragments; and multispecific antibodies formed from antibody fragments.

[0301] The term "variable" refers to the fact that certain segments of the variable (V) domains vary widely in sequence among antibodies. The V domains mediate antigen binding and define the specificity of a particular antibody for a particular antigen. However, the variability is evenly distributed throughout the 110 amino acid span of the variable region. Instead, the V regions are composed of relatively invariant stretches called framework regions (FRs) of 15-30 amino acids, separated by short regions of extreme variability called "hypervariable regions", each 9-12 amino acids in length. The unique heavy and light chain variable regions are each composed of four FRs, adopting a primarily beta-sheet configuration, connected by three hypervariable regions, which form loops and in some cases form part of the beta-sheet structure. The hypervariable regions of each chain are held in close proximity by the FRs and, together with the hypervariable regions of the other chain, contribute to the formation of the antigen-binding site of antibodies (see, e.g., Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md. (1991)).

[0302] The term "hypervariable region" as used herein refers to the amino acid residues of an antibody which are responsible for antigen-binding. Said hypervariable region typically comprises amino acid residues from the "complementarity determining regions" ("CDRs").

[0303] As used herein, the term "monoclonal antibody" refers to a substantially homogeneous population of antibodies. The term "polyclonal antibody" refers to an antibody obtained from a population, i.e., the individual antibodies making up the population are identical except for possible naturally occurring mutations that may be present in minor amounts. The term "polyclonal antibody" refers to a preparation containing different antibodies directed against different determinants ("epitopes").

[0304] As used herein, "subject" refers to humans and non-human animals. Examples of non-human animals include all vertebrates, such as mammals, such as non-human mammals, non-human primates (especially higher primates), dogs, rodents (e.g., mice or rats), guinea pigs, cats, and rabbits, and non-mammals, such as birds, amphibians, and reptiles. In one embodiment, the subject is a human. In another embodiment, the subject is an animal suitable as an experimental animal or disease model.

[0305] The term "polypeptide" is used in its conventional sense, i.e., as a sequence of amino acids. Said polypeptide is not limited to a particular length of the product. Peptides, oligopeptides, and proteins are included in the definition of polypeptide, and such terms may be used interchangeably herein, unless otherwise indicated. The term also includes post-expression modifications of the polypeptide, such as glycosylation, acetylation, phosphorylation, and the like, as well as other modifications known in the art, both naturally occurring and non-naturally occurring. A polypeptide may be an entire protein or a subsequence thereof. A particular polypeptide of interest in the context of the present invention is an amino acid subsequence that includes a CDR, a VH, and a VL that can bind to an antigen or an HIV-infected cell.

[0306] A polypeptide "variant" as the term is used herein is a polypeptide that typically differs from the polypeptide specifically disclosed herein in one or more substitutions, deletions, additions and / or insertions. Such variants may be naturally occurring or may be synthetically produced, for example, by modifying one or more of the above polypeptide sequences of the invention as described herein and / or using any of a number of techniques known in the art, and evaluating one or more biological activities of the polypeptide.

[0307] "Homology" or "sequence identity" refers to the percentage of residues of a polynucleotide or polypeptide sequence variant that are identical to the non-mutated sequence after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent homology. In certain embodiments, polynucleotide and polypeptide variants have at least about 70%, at least about 75%, at least about 80%, at least about 90%, at least about 95%, at least about 98%, or at least about 99% polynucleotide or polypeptide homology with the polynucleotides or polypeptides described herein.

[0308] Such variant polypeptide sequences share 70% or more (i.e., 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or more) sequence identity with the sequences described in the present application. In further embodiments, the described invention provides polypeptide fragments comprising contiguous stretches of various lengths of the amino acid sequences disclosed herein. For example, the invention provides peptide sequences comprising at least about 5, 10, 15, 20, 30, 40, 50, 75, 100, 150 or more contiguous peptides of one or more sequences disclosed herein, and all intermediate lengths therebetween.

[0309] The terms "nucleic acid" and "polynucleotide" are used interchangeably herein and refer to single- or double-stranded RNA, DNA, or mixed polymers. Polynucleotides include genomic sequences, extragenomic and plasmid sequences, and sequences that express a polypeptide or It may comprise smaller engineered gene segments that can be adapted to express a polypeptide.

[0310] "Isolated nucleic acid" is a nucleic acid that is substantially separated from other genomic DNA sequences, and proteins or complexes such as ribosomes and polymerases that naturally accompany the natural sequence. The term includes nucleic acid sequences that are removed from their naturally occurring environment, including recombinant or cloned DNA isolates, and chemically synthesized analogs or biologically synthesized analogs by heterologous systems. Substantially pure nucleic acid includes nucleic acid in isolated form. Thus, it refers to the nucleic acid that is originally isolated, and does not exclude genes or sequences that are later added to the isolated nucleic acid by the hand of man.

[0311] A polynucleotide "variant" as the term is used herein is a polynucleotide that typically differs from a polynucleotide specifically disclosed herein in one or more substitutions, deletions, additions, and / or insertions. Such variants may be naturally occurring or may be synthetically produced, for example, by modifying one or more of the polynucleotide sequences of the invention as described herein and / or using any of a number of techniques known in the art, and evaluating one or more biological activities of the encoded polypeptide.

[0312] As used herein, the term "cell" can be any cell, including, but not limited to, cells of eukaryotic multicellular species, such as mammalian or human cells (as opposed to, e.g., unicellular yeast cells). A cell can exist as a single entity or be part of a larger population of cells. Such a "larger population of cells" can include, for example, a cell culture (either mixed or pure), a tissue (e.g., endothelial, epithelial, mucosal or other tissue), an organ (e.g., lung, liver, muscle and other organs), an organ system (e.g., circulatory system, respiratory system, gastrointestinal system, urinary system, nervous system, integumentary system or other organ system), or an organism (e.g., a bird, a mammal, etc.).

[0313] As used herein, a "label" refers to a detectable compound or composition that is directly or indirectly attached to an antibody to produce a "labeled" antibody. Labels can also be conjugated to the polypeptides and / or nucleic acid sequences disclosed herein. The label may be detectable by itself (e.g., radioisotope labels or fluorescent labels) or, in the case of enzymatic labels, may catalyze a chemical alteration of a substrate compound or composition that is detectable. The described antibodies and polypeptides of the invention can also be modified to include epitope tags or labels, for example, for use in purification or diagnostic applications. Suitable detection means include, but are not limited to, the use of labels such as radionucleotides, enzymes, coenzymes, fluorophores, chemiluminescent materials, chromogens, enzyme substrates or cofactors, enzyme inhibitors, prosthetic group complexes, free radicals, particles, dyes, and the like.

[0314] The term "assessing" includes any form of measurement, including determining whether an element is present. The terms "determining," "measuring," "assessing," "calculating," and "analyzing" are used interchangeably and include quantitative and qualitative determinations. Assessment can be relative or absolute. "Assessing the presence" includes determining the amount of something present and / or determining whether it is present or absent. As used herein, the terms "determining," "measuring," and "assessing" and "analyzing" are used interchangeably and include both quantitative and qualitative determinations.

[0315] The terms "treating" or "treatment" or "palliative" are used interchangeably and refer to both curative treatment and preventative or prophylactic measures; the purpose is to alleviate the targeted pathological condition. The objective of the present invention is to prevent or slow (alleviate) the condition or disorder. Those in need of treatment include those already with the disorder, as well as those who are predisposed to the disorder or those for whom the disorder is to be prevented. A subject or mammal has been successfully "treated" for an infection if, after administration of a therapeutic amount of an antibody according to the methods of the present invention, the patient exhibits an observable and / or measurable reduction, with or without one or more of the following: a reduction in the number of infected cells or the absence of infected cells; a reduction in the percentage of total infected cells; and / or some alleviation of one or more of the symptoms associated with the particular infection; a reduction in morbidity and mortality, as well as an improvement in quality of life issues. The above parameters for assessing the success of treatment and improvement of a disease are readily measurable by routine procedures familiar to physicians.

[0316] "Effective amount" or "therapeutically effective amount" refers to an amount of a compound or agent that can produce a medically desirable result in a treated subject. The therapy can be performed in vivo or ex vivo, alone or in combination with other drugs or therapies. A therapeutically effective amount can be administered in one or more administrations, applications, or dosages, and is not intended to be limited to a particular formulation or route of administration.

[0317] The term "disease" as used herein is generally intended to be synonymous with the terms "disorder" and "condition" (as in medical condition), and are used interchangeably, all reflecting an abnormal condition of the human or animal body, or one of its parts, that impairs normal functioning, and is usually manifested by distinguishing signs and symptoms, causing a decrease in the duration or quality of the human or animal's life.

[0318] The terms "reduce", "reduce", "reduction", "reduction" or "suppress" are all used herein to generally mean a statistically significant amount of reduction. However, and for the avoidance of doubt, "reduced", "reduction" or "reduction" or "suppress" means a reduction of at least 10% compared to a reference level, e.g., a reduction of at least about 20%, or at least about 30%, or at least about 40%, or at least about 50%, or at least about 60%, or at least about 70%, or at least about 80%, or at least about 90%, or a reduction of up to 100% (e.g., no value compared to a reference sample), or a reduction of 10-100%.

[0319] As used herein, the term "modulate" is meant to refer to any change in a biological state, ie, an increase, a decrease, and the like.

[0320] The terms "increased", "increase" or "enhance" or "activate" are all used herein to mean an increase, typically a statically significant amount; for the avoidance of doubt, the terms "increased", "increase", "enhance", or "activate" mean an increase of at least 10% compared to a reference level, for example an increase of at least about 20%, or at least about 30%, or at least about 40%, or at least about 50%, or at least about 60%, or at least about 70%, or at least about 80%, or at least about 90%, or up to 100%, or an increase of between 10-100%, or an increase of at least about 2-fold, or at least about 3-fold, or at least about 4-fold, or at least about 5-fold, or at least about 10-fold, or any increase between 2-10-fold compared to a reference level.

[0321] The terms "effective amount," "effective dose," or "effective dosage" are defined as an amount sufficient to achieve, or at least partially achieve, a desired effect. A "therapeutically effective amount" or "therapeutically effective dosage" of a drug or therapeutic agent is defined as an amount sufficient to achieve, or at least partially achieve, a desired effect. A "prophylactically effective amount" or "prophylactically effective dose" of a drug is an amount of a drug that promotes regression of a disease as evidenced by a decrease in the severity of disease symptoms, an increase in the frequency and duration of disease symptom-free periods, or prevention of disability or damage due to the affliction of the disease, when used in combination with other therapeutic agents. A "prophylactically effective amount" or "prophylactically effective dose" of a drug is an amount of a drug that, when administered alone or in combination with other therapeutic agents to a subject at risk of developing a disease or suffering a recurrence of the disease, inhibits the onset or recurrence of the disease. The ability of a therapeutic or prophylactic agent to promote regression of a disease or inhibit the onset or recurrence of a disease can be evaluated using a variety of methods known to those skilled in the art, such as human subjects in clinical trials, animal model systems predictive of efficacy in humans, or assaying the activity of the agent in in vitro assays.

[0322] Thus, a dose expressed as [g, mg, or other unit] / kg (or g, mg, etc.) usually refers to [g, mg, or other unit] "per kg (or g, mg, etc.) of body weight," even if the term "body weight" is not explicitly mentioned.

[0323] The term "agent" is used herein to denote a compound, a mixture of compounds, a biological macromolecule (e.g., a nucleic acid, an antibody, a protein or portion thereof, e.g., a peptide), or an extract made from biological material such as a bacterial, plant, fungus, or animal (e.g., mammalian) cell or tissue. The activity of such an agent may qualify it as a "therapeutic agent," which is a biologically, physiologically, or pharmacologically active substance (or substances) that acts locally or systemically in a subject.

[0324] The terms "therapeutic agent," "agent capable of treating," or "medical agent" are used interchangeably and refer to a molecule or compound that provides some beneficial effect upon administration to a subject, including enabling a diagnostic determination; ameliorating a disease, symptom, disorder, or pathological condition; reducing or preventing the onset of a disease, symptom, disorder, or condition; and generally combating a disease, symptom, disorder, or pathological condition.

[0325] As used herein, "combination" therapy is meant to include administration of two or more therapeutic agents in a coordinated manner, unless otherwise clear from the context, including but not limited to simultaneous administration. Specifically, combination therapy includes both simultaneous administration (e.g., simultaneous administration of formulations or simultaneous administration of separate therapeutic compositions) and sequential or sequential administration, provided that administration of one therapeutic agent is in some way conditioned on administration of the other therapeutic agent. For example, one therapeutic agent may be administered only after the other therapeutic agent has been administered and allowed to act for a certain period of time. See, for example, Kohrt et al. (2011) Blood 117:2423. Administration "in combination" with one or more additional therapeutic agents includes simultaneous (concurrent) and sequential administration in any order.

[0326] "Sample", "test sample", and "patient sample" may be used interchangeably herein. The sample may be a sample of serum, urine plasma, amniotic fluid, cerebrospinal fluid, cells (e.g., antibody-producing cells) or tissue. Such samples may be used directly as obtained from a patient, or may be pretreated by filtration, distillation, extraction, concentration, centrifugation, inactivation of interfering components, addition of reagents, etc., to modify the characteristics of the sample in some way, as discussed herein or known in the art. The terms "sample" and "biological sample" as used herein generally refer to biological material that is being tested and / or suspected of containing an analyte of interest, such as an antibody. The sample may be any tissue sample from the subject. The sample may contain proteins from the subject.

[0327] As used herein, the terms "inhibit" and "antagonize" refer to an inhibitor or inhibitory agent that inhibits or antagonizes the expression, stability, function or activity of a molecule, reaction, interaction, gene, mRNA, and / or protein. By inhibitors, we mean to measurably reduce or completely prevent the activation of a stimulus. Inhibitors are compounds that, for example, bind to or partially or completely block a stimulus, or reduce, prevent, delay, inactivate, desensitize, or downregulate the stability, expression, function, and activity of proteins, genes, and mRNAs, e.g., antagonists.

[0328] "Parenteral" administration of the compositions includes, for example, subcutaneous (sc), intravenous (iv), intramuscular (im), or intrasternal injection, or infusion techniques.

[0329] As used herein, the term "pharmaceutical composition" refers to a mixture of at least one compound useful within the present invention with other chemical components, such as carriers, stabilizers, diluents, dispersants, suspending agents, thickeners, and / or excipients, that facilitate administration of the compound to an organism.

[0330] Multiple techniques of administering a compound exist in the art, including, but not limited to, intravenous, oral, aerosol, parenteral, ophthalmic, pulmonary, and topical administration.

[0331] As used herein, the term "pharmacologically acceptable" refers to a material, such as a carrier or diluent, that does not abrogate the biological activity or properties of a composition and is relatively non-toxic, i.e., the material may be administered to an individual without causing undesirable biological effects or interacting in a deleterious manner with any of the components of the composition in which it is contained.

[0332] The term "pharmaceutically acceptable carrier" includes pharmaceutically acceptable salts, pharmaceutically acceptable materials, compositions, or carriers such as liquid or solid fillers, diluents, excipients, solvents, or encapsulating materials that are involved in transporting the compounds of the present invention into or to a subject so that they can perform their intended function. Typically, such compounds are carried or transported from one organ or part of the body to another. Each salt or carrier must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not deleterious to the subject. Some examples of materials that may be useful as pharma- ceutically acceptable carriers include: sugars, such as lactose, glucose, sucrose, and the like; starches, such as corn starch and potato starch; cellulose and cellulose derivatives, such as sodium carboxymethylcellulose, ethylcellulose, cellulose acetate, and the like; powdered tragacanth; malt; gelatin; talc; excipients, such as cocoa butter and suppository wax; oils, such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil, soybean oil, and the like; glycols, such as propylene glycol; glycerin, sorbitol, mannitol, and polyethylene. Polyols such as ethylene glycol; esters such as ethyl oleate and ethyl laurate; agar; buffers such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic saline; Ringer's solution; ethyl alcohol; phosphate buffer; diluent; granulating agent; lubricant; binder; disintegrant; wetting agent; emulsifier; coloring agent; release agent; coating agent; sweetener; flavor; flavoring; preservative; antioxidant; plasticizer; gelling agent; thickener; hardener; setting agent; suspending agent; surfactant; humectant; carrier; stabilizer; and other non-toxic compatible substances used in formulation, or any combination thereof. As used herein, "pharmaceutically acceptable carrier" also includes all coatings, antibacterial and antifungal agents, and absorption retardants, etc., that are compatible with the activity of the compound and are physiologically acceptable to the subject. Supplementary active compounds can also be incorporated into the composition.

[0333] As used herein, the term "pharmaceutically acceptable salts" refers to salts of the administered compound prepared from pharmaceutically acceptable non-toxic acids, such as inorganic acids, organic acids, solvates, hydrates, or clathrates thereof.

[0334] As used herein, the term "in vitro" refers to events that take place not within a multicellular organism, but in an artificial environment, e.g., a test tube or reaction vessel, cell culture, etc.

[0335] As used herein, the term "in vivo" refers to events that occur within a multicellular organism, such as a non-human animal.

[0336] It should be noted that as used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural references unless the context clearly dictates otherwise.

[0337] The terms "including," "including," "including," or "having" and variations thereof are meant to encompass the items listed thereafter and equivalents thereof, as well as additional subject matter, unless expressly stated otherwise.

[0338] The phrases "in one embodiment," "in various embodiments," "in some embodiments," and the like are used repeatedly. Such phrases do not necessarily refer to the same embodiment, although they may, unless the context dictates otherwise.

[0339] The term "and / or" or " / " means any one of the items, any combination of the items, or all of the items with which this term is associated.

[0340] The word "substantially" does not exclude "completely". For example, a composition that is "substantially free" of Y may be completely free of Y. If desired, the word "substantially" may be omitted from the definition of the invention.

[0341] As used herein, the term "approximately" or "about" when applied to one or more values ​​of interest refers to a value similar to the stated reference value. In some embodiments, the term "approximately" or "about" refers to a range of values ​​that fall within 25%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less in either direction (greater or less) of the stated reference value, unless otherwise stated or clear from the context (except where such number exceeds 100% of possible values). Unless otherwise stated herein, the term "about" is intended to include values, e.g., weight percentages, close to the stated range that are equivalent in terms of the functionality of the individual components, the composition, or the embodiment.

[0342] Whenever values ​​and ranges are provided herein, it is to be understood that all values ​​and ranges encompassed by these values ​​and ranges are meant to be included within the scope of the present invention. Moreover, all values ​​that fall within these ranges, as well as the upper or lower limits of a range of values, are also contemplated by the application.

[0343] As used herein, the term "each," when used in reference to a collection of items, is intended to identify an individual item in the collection, but does not necessarily refer to every item in the collection. Exceptions may occur where explicit disclosure or context clearly indicates otherwise.

[0344] Any and all examples provided herein, or the use of exemplary language (e.g., "such as"), are intended merely to better illuminate the invention and, unless otherwise claimed, are not intended to be construed as limiting the scope of the invention. No limitation on the scope of the disclosure is intended. No language in the specification should be construed as indicating any non-claimed element as essential to the practice of the invention.

[0345] All methods described herein may be performed in any suitable order unless otherwise indicated herein or clearly contradicted by context. For any of the methods provided, the steps of the method may occur simultaneously or sequentially. When the steps of the method occur sequentially, the steps may occur in any order unless otherwise specified.

[0346] Where a method includes a combination of steps, unless otherwise stated herein, all combinations or subcombinations of said steps are included within the scope of the disclosure.

[0347] Each publication, patent application, patent, and other reference cited herein is incorporated by reference in its entirety to the extent that it does not conflict with this disclosure. Publications disclosed herein are provided solely for their disclosure prior to the filing date of the present invention. Nothing herein should be construed as an admission that the present invention is not entitled to antedate such publication by virtue of prior invention. Further, publication dates provided may be different from the actual publication dates, which may need to be independently confirmed.

[0348] It is understood that the examples and embodiments described herein are for illustrative purposes only, and that various modifications or changes in light thereof will be suggested to those skilled in the art and are to be included within the spirit and scope of this application and the appended claims.

[0349] F. Working Example Example 1 This example describes the materials and methods used in the subsequent examples.

[0350] Polyreactivity assay: Polyreactivity assays were performed using ELISA detection of nonspecific binding to baculovirus extracts as described (Hotzel et al. 2012, mAbs, 4:6, 753-760). Briefly, a solution of 1% baculovirus particles in 100 mM sodium bicarbonate buffer pH 9.6 was absorbed into the wells of a 384-well ELISA plate (Nunc Maxisorp) using a Tecan Freedom Evo liquid handling robot, and the plate was incubated overnight at 4°C. The plate was blocked with 0.5% BSA in PBS for 1 h at room temperature. Purified IgG (diluted to 1 μg / mL in PBS, 0.5% BSA) was added to the blocked assay plate and incubated for 3 h at room temperature. Bound IgG was detected as a luminescence signal at 425 nm using an HRP-labeled anti-human IgG (H&L) secondary antibody (Genscript) and SuperSignal ELISA Femto Maximum Sensitivity Substrate (Thermo Fisher Scientific).

[0351] Neutralization assay: HIV neutralization potency was determined using a luciferase-based TZM-bl assay as previously described (Montefiori, DC 1019, Methods Mol. Biol. 485, 395-405).

[0352] Non-human primate half-life: Macaque half-life was determined as described (Shingai, M. et al., 2014, J. Exp. Med. 211, 2061-74). Briefly, macaques were dosed with 10 mg / kg purified IgG and antibodies were detected in serum by ELISA and / or neutralizing activity over a 70-day period. Half-life was determined by fitting data from days 8 to 70.

[0353] Pharmacokinetics in mice: The designed reagents were radiolabeled with 125I using the iodine method and purified by SEC (Yazaki PJ et al., 2008, Nucl Med Biol., 35(2), 151-8). The half-life and tissue distribution in mice were determined as previously described (Yazaki PJ et al., 2013, PEDS 26(3), 187-193). Briefly, NSG mice were injected with mouse IVIG (4 h prior) and then injected with approximately 3 μg of labeled Ab by tail vein injection (approximately 3-6 microcuries). Blood samples were taken to count the radiolabeled protein. These data are plotted as a function of time for blood clearance. Mice were sacrificed at 96 h, and organs and carcasses were weighed and counted, and the counts were normalized to body weight and plotted.

[0354] Example 2 The parent antibody NIH45-46G54W is composed of NIH45-46 HC and NIH45-46HC with the G54W mutation paired with NIH45-46LC. The novel antibody variants have mutations in one or both of these chains.

[0355] The single chains from which the novel antibody variants are constructed are as follows:

[0356] [Table 4]

[0357] The novel antibody reagent is composed of the following HC / LC pair (LS indicates the aforementioned mutations in the Fc region, e.g., M428L / N434S): HC1 / SAP10 HC1 / SAP10-LS HC1 / SAP10T HC1 / SAP10T-LS HC1 / SAP8 HC1 / SAP3 HC1 / NIH45-46 LC HC1 / m2 NIH45-46 G54W / SAP10 NIH45-46 G54W / SAP3 NIH45-46 G54W / SAP8 The G54W mutant of NIH45-46 (NIH45-46G54W and NIH45-4 6m2) have a short in vivo half-life (Shingai, M. et al., 2014, J. Exp. Med. 211, 2061-74). A short half-life correlates with a high level of polyreactivity (Hotzel, I., et al., 2012, m Abs 4, 753-760). Our novel engineered mutant HC1 / SAP10 exhibits reduced polyreactivity (Figure 2) and retains potency (Figure 1, Tables 1 and 2). Although the neutralizing potency of HC1 / SAP10 is slightly reduced compared to NIH45-46G54W and NIH45-46m2, it still exhibits higher neutralizing potency than other anti-HIV antibodies such as N6 and 3BNC117. It is comparable to bNAbs (Figure 1).

[0358] The half-life of bNAb NIH45-46 m2 in non-human primates (NHPs) could not be determined due to rapid clearance (Shingai, M. et al., 2014, J. Exp. Med. 211, 2061-74). In contrast, our novel mutant HC1 / SAP10-LS has a half-life and tissue distribution similar to that of 3BNC117 in mice (Figure 3), with a half-life of 8.4 days in NHPs.

[0359] Neutralization data for the new reagents are shown in Table 1 and summarized in Table 2.

[0360] [Table 5-1]

[0361] [Table 5-2]

[0362]

Table 5-3

[0363]

Table 6-1

[0364]

Table 6-2

[0365]

Table 7

Claims

1. Residue numbering is shown according to Kabat numbering. a heavy chain having a heavy chain amino acid sequence that is at least 75% identical to SEQ ID NO:1, comprising one or more heavy chain substitutions at one or more residues selected from the group consisting of S5, S21, G54, G55, V57, T68, T70, V73, S75, F79, S82, D85, V89, Y97, and P112; or a light chain comprising one or more light chain substitutions at one or more residues selected from the group consisting of: T5, S7, T10, S12, S14, T18, I20, S22, R24, Q27, S28, S30, R59, S61, S63, W65, D68, N70, S72, S74, and S78, and having a light chain amino acid sequence that is at least 75% identical to SEQ ID NO:2; 2. An isolated anti-HIV antibody, or an antigen-binding portion thereof, comprising:

2. a heavy chain having a heavy chain polypeptide sequence at least 75% identical to SEQ ID NO:1, said heavy chain amino acid sequence comprising one or more heavy chain substitutions at one or more residues selected from the group consisting of S5, S21, G54, G55, V57, T68, T70, V73, S75, F79, S82, D85, V89, Y97, and P112; and a light chain having a light chain amino acid sequence at least 75% identical to SEQ ID NO:2, said light chain amino acid sequence comprising one or more light chain substitutions at one or more residues selected from the group consisting of T5, S7, T10, S12, S14, T18, 120, S22, R24, Q27, S28, S30, R59, S61, S63, W65, D68, N70, S72, S74, and S78; The isolated anti-HIV antibody or antigen-binding portion thereof of claim 1 .

3. 3. The isolated anti-HIV antibody or antigen-binding portion thereof of claim 1 or 2, wherein the one or more heavy chain substitutions comprise: a substitution of S5D or S5E, or a conservative substitution of Asp or Glu at residue S5; a substitution of S21D or S21E, or a conservative substitution of Asp or Glu at residue S21; Substitution of G54W or conservative substitution of Trp at residue G54; a substitution of G55D or G55E, or a conservative substitution of Asp or Glu at residue G55; a substitution of V57D or V57E, or a conservative substitution of Asp or GIu at residue V57; a substitution of T68D or T68E, or a conservative substitution of Asp or Glu at residue T68; a substitution of T70D or T70E, or a conservative substitution of Asp or Glu at residue T70; a substitution of V73D or V73E, or a conservative substitution of Asp or Glu at residue V73; a substitution of S75D or S75E, or a conservative substitution of Asp or Glu at residue S75; a substitution of F79D, F79E, F79Y or F79H, or a conservative substitution of Asp, Glu, Tyr or His at residue F79; a substitution of S82D or S82E, or a conservative substitution of Asp or Glu at residue S82; a substitution of D85E or a conservative substitution of Asp at residue D85; Substitution of V89D or V89E, or Asp or Glu at residue V89 conservative substitution; a substitution of Y97D or Y97E, or a conservative substitution of Asp or Glu at residue Y97; a substitution of P112D or P112E, or a conservative substitution of Asp or Glu at residue P112; Or a combination thereof.

4. The isolated anti-HIV antibody or antigen-binding portion thereof of any one of claims 1 to 3, wherein the one or more light chain substitutions comprise: a substitution of T5D or T5E, or a conservative substitution of Asp or Glu at residue T5; a substitution of S7D or S7E, or a conservative substitution of Asp or Glu at residue S7; a substitution of T10D or T10E, or a conservative substitution of Asp or Glu at residue T10; a substitution of S12D or S12E, or a conservative substitution of Asp or Glu at residue S12; a substitution of S14D or S14E, or a conservative substitution of Asp or Glu at residue S14; a substitution of T18D or T18E, or a conservative substitution of Asp or Glu at residue T18; a substitution of I20D or I20E, or a conservative substitution of Asp or Glu at residue I20; a substitution of S22D or S22E, or a conservative substitution of Asp or Glu at residue S22; a substitution of R24D or R24E, or a conservative substitution of Asp or Glu at residue R24; a substitution of Q27D or Q27E, or a conservative substitution of Asp or Glu at residue Q27; a substitution of S28Y or S28H, or a conservative substitution of Tyr or His at residue S28; a substitution of S30D or S30E, or a conservative substitution of Asp or Glu at residue S30; a substitution of R59D or R59E, or a conservative substitution of Asp or Glu at residue R59; a substitution of S61D or S61E, or a conservative substitution of Asp or Glu at residue S61; a substitution of S63D or S63E, or a conservative substitution of Asp or Glu at residue S63; a substitution of W65D, W65E, W65Y or W65H, or a conservative substitution of Asp, Glu, Tyr or His at residue W65; a substitution of D68E, or a conservative substitution of Asp at residue D68; a substitution of N70D or N70E, or a conservative substitution of Asp or Glu at residue N70; a substitution of S72D or S72E, or a conservative substitution of Asp or Glu at residue S72; a substitution of S74D or S74E, or a conservative substitution of Asp or Glu at residue S74; a substitution of S78D or S78E, or a conservative substitution of Asp or Glu at residue S78; Or a combination thereof.

5. 5. The isolated anti-HIV antibody or antigen-binding portion thereof of claim 4, wherein the one or more heavy chain substitutions further comprise the following substitutions: M428L and N434S.

6. 6. The isolated anti-HIV antibody or antigen-binding portion thereof of claim 5, wherein the light chain amino acid sequence comprises one or more of the following light chain substitutions: Q27E, S28H, and S30D; Q27E, S28H, S30D, and S74T; Q27E, S28H, S30D, S74T, M428L, and N434S; Q27E, S28Y, and S30D; Q27D and S28H; S28Y; T5D, T10D, S12D, S14D, I20D, and S22D; or S61D, S63D, W65D, N70D, S72D, and S74D.

7. (i) the heavy chain amino acid sequence comprises one or more heavy chain substitutions including G54W and T68D; and the light chain amino acid sequence comprises: (a) Q27E, S28H, S30D, and S74T; (b) Q27E, S28H, S30D, S74T, M428L, and N434S; (c) Q27E, S28Y, and S30D; (d) Q27D and S28H; or (e) S28Y, or (ii) the heavy chain amino acid sequence contains one or more heavy chain substitutions including G54W; and the light chain amino acid sequence contains (f) Q27E, S28H, and S30D; (g) Q27E, S28Y, and S30D; or (h) Q27D and S28H, comprising one or more light chain substitutions of The isolated anti-HIV antibody or antigen-binding portion thereof according to any one of claims 1 to 6.

8. The isolated anti-HIV antibody or antigen-binding portion thereof according to any one of claims 1 to 7, wherein the heavy chain amino acid sequence comprises an amino acid sequence of SEQ ID NO:3 to SEQ ID NO:

35.

9. The isolated anti-HIV antibody or antigen-binding portion thereof according to any one of claims 1 to 8, wherein the light chain amino acid sequence comprises the amino acid sequences of SEQ ID NO:2 and SEQ ID NO:36 to SEQ ID NO:

80.

10. The isolated anti-HIV antibody is SEQ ID NOs: 81, 84, 87, 90, 93, and 96; SEQ ID NOs: 81, 84, 87, 99, 93, and 96; SEQ ID NOs: 81, 84, 87, 100, 93, and 96; SEQ ID NOs: 81, 84, 87, 101, 93, and 96; or SEQ ID NOs: 81, 84, 87, 102, 93, and 96; (shown in Kabat numbering); SEQ ID NOs: 82, 85, 88, 91, 94, and 97 (shown in IMGT numbering); or SEQ ID NOs: 83, 86, 89, 92, 95, and 98 (shown in Chothia numbering); The set of CDR sequences includes CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3, each of which comprises a sequence shown in The isolated anti-HIV antibody or antigen-binding portion thereof according to any one of claims 1 to 9.

11. The isolated anti-HIV antibody or antigen-binding portion thereof according to any one of claims 1 to 10, wherein the heavy chain amino acid sequence and the light chain amino acid sequence comprise the respective amino acid sequences set forth in the set of sequences selected from the group consisting of SEQ ID NOs: 3 and 36; SEQ ID NOs: 3 and 38; SEQ ID NOs: 3 and 39; SEQ ID NOs: 4 and 2; SEQ ID NOs: 4 and 38; SEQ ID NOs: 4 and 39; SEQ ID NOs: 4 and 41; and SEQ ID NOs: 34 and 37.

12. the isolated anti-HIV antibody is a bispecific antibody comprising a first antigen-binding arm that binds a first antigen and a second antigen-binding arm that binds a second antigen; The first and second antigens are different, and the first antigen-binding arm comprises a heavy chain amino acid sequence and a light chain amino acid sequence according to any one of claims 2 to 11. The isolated anti-HIV antibody or antigen-binding portion thereof according to any one of claims 1 to 11.

13. the second antigen is selected from the group consisting of CD3, FcγRI (CD64), FcγRII (CD32), FcγRIII (CD16); CD89, CCR5, CD4, CD8, CD28, CD137, CTLA-4, gp41, killer cell immunoglobulin-like receptor, three Ig domains, and long cytoplasmic tail 1 (KIR3DL1), killer cell immunoglobulin-like receptor, three Ig domains, and long cytoplasmic tail 1 (KIR3DL1), killer cell immunoglobulin-like receptor, Killer cell immunoglobulin-like receptor, two Ig domains and a long cytoplasmic tail 1 (KIR2DL1), Killer cell immunoglobulin-like receptor, two Ig domains and a long cytoplasmic tail 3 (KIR2DL3), Killer cell lectin-like receptor Cl (KLRC1), Killer cell lectin-like receptor C2 (KLRC2), Killer cell lectin-like receptor C3 (KLRC3), Killer cell lectin-like receptor C4 (KLRC4), Killer cell lectin-like receptor D1 (KLRD1), Killer cell lectin-like receptor K 1 (KLRK1), natural cytotoxicity-inducing receptor 3 (NCR3 or NKp30), natural cytotoxicity-inducing receptor 2 (NCR2 or NK-p44), natural cytotoxicity-inducing receptor 1 (NCR1 or NK-p46), CD226 (DNAM-1), cytotoxic and regulatory T cell molecule (CRTAM or CD355), signaling lymphocyte activation molecule family member 1 (SLAMF1), CD48 (SLAMF2), lymphocyte antigen 9 (LY9 or SLAMF3), C 13. The isolated anti-HIV antibody or antigen-binding portion thereof of claim 12, wherein the antibody is selected from the group consisting of D244 (2B4 or SLAMF4), CD84 (SLAMF5), SLAM family member 6 (SLAMF6 or NTB-A), SLAM family member 7 (SLAMF7 or CRACC), CD27 (TNFRSF7), semaphorin 4D (SEMA4D or CD100), CD160 (NK1), and the second epitope of gp120.

14. An isolated nucleic acid comprising a sequence encoding the CDR, heavy chain variable region, or light chain variable region, or an antigen-binding portion thereof, of the anti-HIV antibody of any one of claims 1 to 13.

15. A vector comprising the nucleic acid of claim 14.

16. A cultured cell comprising the vector of claim 15.

17. Obtaining the cultured cell of claim 16; culturing the cultured cells in a medium under conditions that permit expression of the polypeptide encoded by the vector and assembly of the antibody or fragment thereof; and purifying said antibody or fragment from said cultured cells or from the medium of said cultured cells; A method for producing an anti-HIV antibody or a fragment thereof, comprising:

18. (i) at least one anti-HIV antibody or antigen-binding portion thereof according to any one of claims 1 to 13, a nucleic acid according to claim 14, or a vector according to claim 15; and (ii) a pharma- ceutically acceptable carrier; 13. A pharmaceutical composition comprising:

19. 20. The pharmaceutical composition of claim 18, further comprising a second therapeutic agent.

20. 20. The pharmaceutical composition of claim 19, wherein the second therapeutic agent comprises an antiviral agent or one or more additional antibodies.

21. 21. The pharmaceutical composition of claim 20, wherein the one or more additional antibodies comprise a second anti-HIV antibody or antigen-binding portion thereof, or a third antibody that binds to a third antigen.

22. The second antigen is selected from the group consisting of CD3, FcγRI (CD64), FcγRII (CD32), FcγRIII (CD16); CD89, CCR5, CD4, CD8, CD28, CD137, CTLA-4, gp41, killer cell immunoglobulin-like receptor, three Ig domains, and long cytoplasmic tail 1 (KIR3DL1), killer cell immunoglobulin-like receptor, three Ig domains, and long cytoplasmic tail 1 (KIR3DL1), killer cell immunoglobulin-like Killer cell immunoglobulin-like receptor, two Ig domains and a long cytoplasmic tail 1 (KIR2DL1), Killer cell immunoglobulin-like receptor, two Ig domains and a long cytoplasmic tail 3 (KIR2DL3), Killer cell lectin-like receptor Cl (KLRC1), Killer cell lectin-like receptor C2 (KLRC2), Killer cell lectin-like receptor C3 (KLRC3), Killer cell lectin-like receptor C4 (KLRC4), Killer cell lectin-like receptor D1 (KLRD1), Killer cell lectin-like receptor Cutin-like receptor K1 (KLRK1), natural cytotoxicity-inducing receptor 3 (NCR3 or NKp30), natural cytotoxicity-inducing receptor 2 (NCR2 or NK-p44), natural cytotoxicity-inducing receptor 1 (NCR1 or NK-p46), CD226 (DNAM-1), cytotoxic and regulatory T cell molecule (CRTAM or CD355), signaling lymphocyte activation molecule family member 1 (SLAMF1), CD48 (SLAMF2), lymphocyte antigen 9 (LYMPH-9), and IL-11 (IL-11) are also involved.

22. The pharmaceutical composition of claim 21, wherein the epitope is selected from the group consisting of: CD244 (2B4 or SLAMF4), CD84 (SLAMF5), SLAM family member 6 (SLAMF6 or NTB-A), SLAM family member 7 (SLAMF7 or CRACC), CD27 (TNFRSF7), semaphorin 4D (SEMA4D or CD100), CD160 (NK1), and the second epitope of gp120.

23. 21. The pharmaceutical composition of claim 20, wherein the antiviral agent is selected from the group consisting of non-nucleoside reverse transcriptase inhibitors, protease inhibitors, entry or fusion inhibitors, and integrase inhibitors.

24. Methods for preventing or treating HIV infection or HIV-related diseases, including: Identifying a patient in need of said prevention or treatment; and Administering to said patient a first therapeutic agent comprising a therapeutically effective amount of at least one anti-HIV antibody or antigen-binding portion thereof according to any one of claims 1-13.

25. 25. The method of claim 24, further comprising administering a second therapeutic agent.

26. 4. The method of claim 3, wherein the second therapeutic agent comprises an antiviral agent or one or more additional antibodies.

25. The method according to claim 25.

27. 27. The method of claim 26, wherein the one or more additional antibodies comprise a second anti-HIV antibody or antigen-binding portion thereof, or a third antibody that binds to a third antigen.

28. The second antigen is selected from the group consisting of CD3, FcγRI (CD64), FcγRII (CD32), FcγRIII (CD16); CD89, CCR5, CD4, CD8, CD28, CD137, CTLA-4, gp41, killer cell immunoglobulin-like receptor, three Ig domains, and long cytoplasmic tail 1 (KIR3DL1), killer cell immunoglobulin-like receptor, three Ig domains, and long cytoplasmic tail 1 (KIR3DL1), killer cell immunoglobulin-like Killer cell immunoglobulin-like receptor, two Ig domains and a long cytoplasmic tail 1 (KIR2DL1), Killer cell immunoglobulin-like receptor, two Ig domains and a long cytoplasmic tail 3 (KIR2DL3), Killer cell lectin-like receptor Cl (KLRC1), Killer cell lectin-like receptor C2 (KLRC2), Killer cell lectin-like receptor C3 (KLRC3), Killer cell lectin-like receptor C4 (KLRC4), Killer cell lectin-like receptor D1 (KLRD1), Killer cell lectin-like receptor Cutin-like receptor K1 (KLRK1), natural cytotoxicity-inducing receptor 3 (NCR3 or NKp30), natural cytotoxicity-inducing receptor 2 (NCR2 or NK-p44), natural cytotoxicity-inducing receptor 1 (NCR1 or NK-p46), CD226 (DNAM-1), cytotoxic and regulatory T cell molecule (CRTAM or CD355), signaling lymphocyte activation molecule family member 1 (SLAMF1), CD48 (SLAMF2), lymphocyte antigen 9 (LYMPH-9), and IL-11 (IL-11) are also involved.

28. The pharmaceutical composition of claim 27, wherein the epitope is selected from the group consisting of: CD244 (2B4 or SLAMF3), CD84 (SLAMF5), SLAM family member 6 (SLAMF6 or NTB-A), SLAM family member 7 (SLAMF7 or CRACC), CD27 (TNFRSF7), semaphorin 4D (SEMA4D or CD100), CD160 (NK1), and the second epitope of gp120.

29. A kit comprising a pharma- ceutically effective amount of a pharma- ceutically acceptable dosage unit of at least one isolated anti-HIV antibody, or antigen-binding portion thereof, according to any one of claims 1-13.

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