Methods and materials for treating HIV

Abstract

This document provides methods and materials for treating a mammal (e.g., a human) having, or at risk of developing, a human immunodeficiency virus (HIV; e.g., an HIV-1) infection. For example, one or more polypeptides including a D1 domain of a cluster determinant 4 (CD4) polypeptide (and / or nucleic acids designed to express a polypeptide including a D1 domain of a CD4 polypeptide) can be administered to a mammal (e.g., a human) having, or at risk of developing, an HIV (e.g., an HIV-1) infection to treat the mammal.

Description

[0001] METHODS AND MATERIALS FOR TREATING HIV

[0002] CROSS-REFERENCE TO RELATED APPLICATIONS

[0003] This application claims the benefit of U.S. Patent Application Serial No. 63 / 557,249, filed on February 23, 2024, and claims the benefit of U.S. Patent Application Serial No. 63 / 717,457, filed on November 7, 2024. The disclosures of the prior applications are considered part of, and are incorporated by reference in, the disclosure of this application.

[0004] STATEMENT REGARDI G FEDERAL FUNDING

[0005] This invention was made with government support under AI036219 awarded by the National Institutes of Health. The government has certain rights in the invention.

[0006] SEQUENCE LISTING

[0007] This application contains a Sequence Listing that has been submitted electronically as an XML file named “48881-0060W01 _SL.xml.” The XML file, created on February 13, 2025, is 56,170 bytes in size. The material in the XML file is hereby incorporated by reference in its entirety.

[0008] TECHNICAL FIELD

[0009] This document relates to methods and materials for treating a mammal (e.g., a human) having, or at risk of developing, a human immunodeficiency virus (HIV; e.g., an HIV-1) infection. For example, one or more polypeptides including a DI domain of a cluster determinant 4 (CD4) polypeptide (and / or nucleic acids designed to express a polypeptide including a DI domain of a CD4 polypeptide) can be administered to a mammal (e.g., a human) having, or at risk of developing, an HIV (e.g., an HIV-1) infection to treat the mammal.

[0010] BACKGROUND

[0011] Human CD4 is the primary HIV receptor. Pre-treatment with soluble CD4 (sCD4) decoys has been explored as a potential strategy to prevent HIV infection. Crystal structures show that the DI domain of CD4 interacts with gpl20, an envelope glycopeptide present on HIV-1 particles that is required for the HIV-1 particles to enter CD4+cells (Bour et al., Microbiol. Rev., 59(l):63-93 (1995)), and engineered sCD4 polypeptides including DI, Dl- D2, and D1-D4 domains have been used to attempt to constrain HIV infection (Haim et al., PLoS Pathog., 5(4):el000360 (2009)). However, previous versions of sCD4 polypeptides exhibited limited and transient efficacy in both preclinical and clinical trials, partially due to the short in vivo half-life of sCD4 (e.g., Zeira et al., Proc. Natl. Acad. Sci. USA, 88(10):4409- 13 (1991)).

[0012] SUMMARY

[0013] This document provides methods and materials for treating a mammal (e g., a human) having, or at risk of developing, an HIV (e.g., an HIV-1) infection. For example, one or more polypeptides comprising a DI domain of a CD4 polypeptide (and / or nucleic acids designed to express a polypeptide comprising a DI domain of a CD4 polypeptide) can be administered to a mammal (e.g., a human) having, or at risk of developing, an HIV (e.g., an HIV-1) infection to treat the mammal. Polypeptides comprising a D I domain of a CD4 polypeptide can also be referred to as CD4-D1 polypeptides.

[0014] As described herein, polypeptides (e.g., recombinant polypeptides) comprising, consisting essentially of, or consisting of SEQ ID NO:4 can bind HIV particles, thus preventing the HIV particles from binding to CD4+cells and entering and infecting the CD4+cells. Having the ability to reduce entry of HIV particles into cells within a mammal (e.g., a human) as described herein (e.g., by administering one or more CD4-D1 polypeptides provided herein and / or nucleic acids designed to express a CD4-D1 polypeptide provided herein) provides a unique and unrealized opportunity to treat mammals having, or at risk of developing, an HIV (e.g., an HIV-1) infection.

[0015] Also as described herein, CD4-D1 polypeptides (e.g., recombinant CD4-D1 polypeptides) comprising, consisting essentially of, or consisting of SEQ ID NO:4 demonstrate favorable pharmacokinetic features for treating mammals having, or at risk of developing, an HIV (e.g., an HIV-1) infection. For example, CD4-D1 polypeptides provided herein demonstrate an extended half-life, enhanced thermostability, and high affinity binding to gpl20 glycopeptides (e.g., as compared to CD4-D1 polypeptides lacking the amino acid sequence set forth in any one of SEQ ID NOs:4-39). For example, CD4-D1 polypeptides provided herein can avoid immune-suppression (e.g., due to reduced or eliminated affinity for a major histocompatibility complex (MHC) such as MHC II).

[0016] In general, one aspect of this document features methods for treating a mammal having an HIV infection where the methods can include, or consist essentially of, administering a CD4-D1 polypeptide to the mammal, where the CD4-D1 polypeptide includes the amino acid sequence set forth in any one of SEQ ID NOs:4-39 with zero, one, two, three, four, or five amino acid residues preceding the amino acid sequence, with zero, one, two, three, four, or five amino acid residues following the amino acid sequence, and / or with zero, one, two, three, four, or five amino acid substitutions within the amino acid sequence. The CD4-D1 polypeptide can include the amino acid sequence set forth in any one of SEQ ID NOs:4-8. The mammal can be a human. The HIV can be HIV-1. The CD4-D1 polypeptide can be effective to reduce a symptom of the HIV infection. The symptom can be fever, headache, muscle aches, joint pain, rash, sore throat, painful mouth sores, swollen lymph glands, swollen lymph nodes on the neck, diarrhea, weight loss, cough, night sweats, fatigue, vomiting, vision loss, or memory loss. The CD4-D1 polypeptide can be effective to reduce a number of HIV particles present in one or more cells within the mammal. The CD4- D1 polypeptide can be effective to reduce a viral load of the mammal. The method can be effective to delay or prevent progression of the HIV infection to acquired immunodeficiency syndrome (AIDS) within the mammal. The mammal can have AIDS. The method can include identifying the mammal as being in need of the CD4-D1 polypeptide prior to the administering step.

[0017] In another aspect, this document features methods for treating a mammal having an HIV infection where the methods can include, or consist essentially of, administering nucleic acid encoding a CD4-D1 polypeptide to the mammal, where the CD4-D1 polypeptide includes the amino acid sequence set forth in any one of SEQ ID NOs:4-39 with zero, one, two, three, four, or five amino acid residues preceding the amino acid sequence, with zero, one, two, three, four, or five amino acid residues following the amino acid sequence, and / or with zero, one, two, three, four, or five amino acid substitutions within the amino acid sequence. The CD4-D1 polypeptide can include the amino acid sequence set forth in any one of SEQ ID NOs:4-8. The mammal can be a human. The HIV can be HIV-1. The CD4-D1 polypeptide can be effective to reduce a symptom of the HIV infection. The symptom can be fever, headache, muscle aches, joint pain, rash, sore throat, painful mouth sores, swollen lymph glands, swollen lymph nodes on the neck, diarrhea, weight loss, cough, night sweats, fatigue, vomiting, vision loss, or memory loss. The CD4-D1 polypeptide can be effective to reduce a number of HIV particles present in one or more cells within the mammal. The CD4- D1 polypeptide can be effective to reduce a viral load of the mammal. The method can be effective to delay or prevent progression of the HIV infection to AIDS within the mammal. The mammal can have AIDS. The method can include identifying the mammal as being in need of the CD4-D1 polypeptide prior to the administering step. The nucleic acid can be in the form of a vector. The vector can be an adeno-associated virus (AAV) vector. The vector can be an expression plasmid.

[0018] In another aspect, this document features methods for treating a mammal at risk of developing an HIV infection where the methods can include, or consist essentially of, administering a CD4-D1 polypeptide to the mammal, where the CD4-D1 polypeptide includes the amino acid sequence set forth in any one of SEQ ID NOs:4-39 with zero, one, two, three, four, or five amino acid residues preceding the amino acid sequence, with zero, one, two, three, four, or five amino acid residues following the amino acid sequence, and / or with zero, one, two, three, four, or five amino acid substitutions within the amino acid sequence. The CD4-D1 polypeptide can include the amino acid sequence set forth in any one of SEQ ID NOs:4-8. The mammal can be a human. The HIV can be HIV-1. The method can be effective to delay or prevent development of an HIV infection within the mammal. The method can include identifying the mammal as being in need of the CD4-D1 polypeptide prior to the administering step.

[0019] In another aspect, this document features methods for treating a mammal at risk of developing an HIV infection where the methods can include, or consist essentially of, administering nucleic acid encoding a CD4-D1 polypeptide to the mammal, where the CD4- D1 polypeptide includes the amino acid sequence set forth in any one of SEQ ID NOs:4-39 with zero, one, two, three, four, or five amino acid residues preceding the amino acid sequence, with zero, one, two, three, four, or five amino acid residues following the amino acid sequence, and / or with zero, one, two, three, four, or five amino acid substitutions within the amino acid sequence. The CD4-D1 polypeptide can include the amino acid sequence set forth in any one of SEQ ID NOs:4-8. The mammal can be a human. The HIV can be HIV-1. The method can be effective to delay or prevent development of an HIV infection within the mammal. The method can include identifying the mammal as being in need of the CD4-D1 polypeptide prior to the administering step. The nucleic acid can be in the form of a vector. The vector can be an AAV vector. The vector can be an expression plasmid.

[0020] In another aspect, this document features methods for treating a mammal having an HIV infection where the methods can include, or consist essentially of, administering a CD4- D1 polypeptide to the mammal, where the CD4-D1 polypeptide includes the amino acid sequence set forth in SEQ ID NO:40, where each X is independently selected from the group consisting of A, S, T, E, D, Q, N, and H. The CD4-D1 polypeptide can include the amino acid sequence set forth in any one of SEQ ID NOs:9-39. The mammal can be a human. The HIV can be HIV-1. The CD4-D1 polypeptide can be effective to reduce a symptom of the HIV infection. The symptom can be fever, headache, muscle aches, joint pain, rash, sore throat, painful mouth sores, swollen lymph glands, swollen lymph nodes on the neck, diarrhea, weight loss, cough, night sweats, fatigue, vomiting, vision loss, or memory loss. The CD4- D1 polypeptide can be effective to reduce a number of HIV particles present in one or more cells within the mammal. The CD4-D1 polypeptide can be effective to reduce a viral load of the mammal. The method can be effective to delay or prevent progression of the HIV infection to AIDS within the mammal. The mammal can have AIDS. The method can include identifying the mammal as being in need of the CD4-D1 polypeptide prior to the administering step.

[0021] In another aspect, this document features methods for treating a mammal having an HIV infection where the methods can include, or consist essentially of, administering nucleic acid encoding a CD4-D1 polypeptide to the mammal, where the CD4-D1 polypeptide includes the amino acid sequence set forth in SEQ ID NO:40, where each X is independently selected from the group consisting of A, S, T, E, D, Q, N, and H. The CD4-D1 polypeptide can include the amino acid sequence set forth in any one of SEQ ID NOs:9-39. The mammal can be a human. The HIV can be HIV-1. The CD4-D1 polypeptide can be effective to reduce a symptom of the HIV infection. The symptom can be fever, headache, muscle aches, joint pain, rash, sore throat, painful mouth sores, swollen lymph glands, swollen lymph nodes on the neck, diarrhea, weight loss, cough, night sweats, fatigue, vomiting, vision loss, or memory loss. The CD4-D1 polypeptide can be effective to reduce a number of HIV particles present in one or more cells within the mammal. The CD4-D1 polypeptide can be effective to reduce a viral load of the mammal. The method can be effective to delay or prevent progression of the HIV infection to AIDS within the mammal. The mammal can have AIDS. The method can include identifying the mammal as being in need of the CD4-D1 polypeptide prior to the administering step. The nucleic acid can be in the form of a vector. The vector can be an AAV vector. The vector can be an expression plasmid.

[0022] In another aspect, this document features methods for treating a mammal at risk of developing an HIV infection where the methods can include, or consist essentially of, administering a CD4-D1 polypeptide to the mammal, where the CD4-D1 polypeptide includes the amino acid sequence set forth in SEQ ID NO:40, where each X is independently selected from the group consisting of A, S, T, E, D, Q, N, and H. The CD4-D1 polypeptide can include the amino acid sequence set forth in any one of SEQ ID NOs:9-39. The mammal can be a human. The HIV can be HIV-1. The method can be effective to delay or prevent development of an HIV infection within the mammal. The method can include identifying the mammal as being in need of the CD4-D1 polypeptide prior to the administering step.

[0023] In another aspect, this document features methods for treating a mammal at risk of developing an HIV infection where the methods can include, or consist essentially of, administering nucleic acid encoding a CD4-D1 polypeptide to the mammal, where the CD4- D1 polypeptide includes the amino acid sequence set forth in SEQ ID NO:40, where each X is independently selected from the group consisting of A, S, T, E, D, Q, N, and H. The CD4- D1 polypeptide can include the amino acid sequence set forth in any one of SEQ ID N0s:9- 39. The mammal can be a human. The HIV can be HIV-1. The method can be effective to delay or prevent development of an HIV infection within the mammal. The method can include identifying the mammal as being in need of the CD4-D1 polypeptide prior to the administering step. The nucleic acid can be in the form of a vector. The vector can be an AAV vector. The vector can be an expression plasmid.

[0024] In another aspect, this document features uses of a composition including a CD4-D1 polypeptide having the amino acid sequence set forth in any one of SEQ ID NOs:4-39 to treat a mammal having an HIV infection. In another aspect, this document features CD4-D1 polypeptides having the amino acid sequence set forth in any one of SEQ ID NOs:4-39 for use in the preparation of a medicament to treat a mammal having an HIV infection.

[0025] In another aspect, this document features CD4-D1 polypeptides having the amino acid sequence set forth in any one of SEQ ID NOs:4-39 for use in the treatment of an HIV infection.

[0026] In another aspect, this document features uses of a composition including nucleic acid encoding a CD4-D1 polypeptide having the amino acid sequence set forth in any one of SEQ ID NOs:4-39 to treat a mammal having an HIV infection.

[0027] In another aspect, this document features nucleic acid encoding a CD4-D1 polypeptide having the amino acid sequence set forth in any one of SEQ ID NOs:4-39 for use in the preparation of a medicament to treat a mammal having an HIV infection.

[0028] In another aspect, this document features nucleic acid encoding a CD4-D1 polypeptide having the amino acid sequence set forth in any one of SEQ ID NOs:4-39 for use in the treatment of an HIV infection.

[0029] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. Although methods and materials similar or equivalent to those described herein can be used to practice the invention, suitable methods and materials are described below. All publications, patent applications, patents, and other references mentioned herein are incorporated by reference in their entirety. In case of conflict, the present specification, including definitions, will control. In addition, the materials, methods, and examples are illustrative only and not intended to be limiting.

[0030] The details of one or more embodiments of the invention are set forth in the accompanying drawings and the description below. Other features, objects, and advantages of the invention will be apparent from the description and drawings, and from the claims.

[0031] BRIEF DESCRIPTION OF THE DRAWINGS

[0032] Figures 1A and IB. Abrogation of MHC II binding by structure-based rational design. Figure 1A) gpl20 / mD1.22 and MHC IVmD1.22 docking structure models. Figure IB) Method for identifying mutations that increased binding free energy between DI and MHC II but decreased binding free energy between DI and gpl20.

[0033] Figures 2A and 2B. Identification of a DI mutation that eliminated MHC II binding. Figure 2A) Subcloning and expression of the recombinant MHC II (HLA-DR). Figure 2B) Biolayer interferometry to test binding to the MHC II recombinant protein.

[0034] Figure 3. Candidate cavity filling mutations were combined with candidate MHC II- binding abrogation mutations. A wild type (WT) CD4-D1 amino acid sequence (SEQ ID NO: 1) is aligned with the mDl .22 polypeptide with the relative locations of modifications labeled in the sub-libraries 1 and 3 (Libi and Lib3). Libraries 2 and 4 are constructed based on libraries 1 and 3 using error-prone PCR.

[0035] Figure 4. A depletion phage panning strategy to engineer out MHC II binders but retain gpl20 binders.

[0036] Figure 5. CD4-D1 polypeptides with higher thermostability were identified.

[0037] Figure 6. CD4-D1 polypeptides did not bind to MHC II, and MHC II binding negatively correlated with in vivo half-life.

[0038] Figure 7. CD4-D1 polypeptide pharmacokinetics studies in Balb / c mice.

[0039] Figure 8. Neutralization of HIV transmitted / founder (T / F) virus infection by CD4-D1 polypeptides with comparison to 3BNC117.

[0040] Figure 9. Neutralization of HIV- 1 pseudoviruses representing global circulating strains by CD4-D1 polypeptides with comparison to 3BNC117.

[0041] Figure 10. Neutralization of HIV-1 T / F and pseudoviruses by CD4-D1 S60E.

[0042] Figure 11. Sequence alignment of exemplary CD4-D1 polypeptides DI (SEQ ID NO: 1), mDl.22 (SEQ ID NO:3), S60E (SEQ ID NO:4), and 1A2-4 (SEQ ID N0:6).

[0043] Figure 12. Surface plasmon resonance (SPR) binding test with gp!40 demonstrated that 1A2-4 had the highest affinity for gpl40, followed by mDl.22 and S60E.

[0044] Figure 13. The BLItz binding assay showed that both S60E and 1A2-4 variants lost the ability to bind to recombinant MHC II.

[0045] Figure 14. Size exclusion chromatography (SEC) showed that mDl.22, S60E, and 1A2-4 had high purity and were monomeric, without high molecular weight species.

[0046] Figure 15. A thermostability test was performed for mDl .22, S60E, and 1A2-4. S60E showed the highest Tm value, followed by 1A2-4 and mDl.22. Figure 16. A dynamic light scattering (DLS) assay showed that S60E-Fc had high stability and was aggregation resistant when incubated at 37°C for 14 days, whereas both mD1.22-Fc and lA2-4-Fc showed aggregation.

[0047] Figures 17A-17D. Cell binding test with MHC II positive and negative cell lines. The binding effects of CD4-D1 variants were tested on both human and mouse MHC II and MHCIF cell lines. Figures 17A-17B) S60E showed reduced non-specific binding effects compared to 1A2-4 and mD1.22. Figures 17C-17D) 1A2-4, S60E, and mD1.22 did not show non-specific binding effects with human or mouse PBMCs. These results indicate that the binding effects were not dependent on MHC II expression.

[0048] Figures 18. A pharmacokinetic assay showed that S60E had a longer half-life than 1A2-4 and mD1.22-Fc, and even better than the clinical antibody 10-1074.

[0049] Figure 19 Neutralization effects of CD4-D1 variants on different HIV virus types. S60E had similar IC50 value to mD1.22 and 1A2-4 and retained broad neutralizing effects.

[0050] Figure 20. Schematics of exemplary bispecific antibodies designed based on 1A2-4 and S60E.

[0051] Figure 21. A pharmacokinetic assay showed that the bispecific antibodies based on 1 A2-4 and S60E did not have a longer half-life than S60E-Fc alone, which showed the longest half-life.

[0052] Figure 22. A neutralization assay showed that the bispecific antibodies S60E-1074 and 1A2-4-1074 had higher neutralization potential.

[0053] Figure 23. Neutralization of subtype C (CHO5O5) and B (CH077) transmitted / founder viruses by CD4-D1 variants.

[0054] Figure 24. Neutralization of subtype B (2626 and 2864) transmitted / founder viruses by CD4-D1 variants.

[0055] Figure 25. Neutralization of subtype B (pTROl 1 Env) and CRFO (pBJOX2000 env) HIV pseudoviruses by CD4-D1 variants.

[0056] Figure 26. Neutralization of subtype C (pCEl 176 Env) and subtype AC (p246F3 env) HIV pseudoviruses by CD4-D1 variants.

[0057] Figure 27. Neutralization of subtype CRF07 BC (pCHl 19 Env) and subtype G (pX1632 env) HIV pseudoviruses by CD4-D1 variants. DETAILED DESCRIPTION

[0058] This document provides methods and materials for treating a mammal (e.g., a human) having, or at risk of developing, an HIV (e.g., an HIV-1) infection. For example, one or more CD4-D1 polypeptides (e.g., recombinant CD4-D1 polypeptides) provided herein (and / or nucleic acids designed to express a CD4-D1 polypeptide provided herein) can be administered to a mammal (e.g., a human) having, or at risk of developing, an HIV (e.g., an HIV-1) infection to treat the mammal.

[0059] In some cases, the methods and materials described herein can be used to treat an HIV (e.g., an HIV-1) infection. For example, one or more CD4-D1 polypeptides provided herein (and / or nucleic acids designed to express a CD4-D1 polypeptide provided herein) can be administered to a mammal (e.g., a human) in need thereof (e.g., a human having, or at risk of developing, an HIV (e.g., an HIV-1) infection) to slow, delay, or prevent infection of one or more cells within the mammal by one or more HIV particles. In some cases, one or more CD4-D1 polypeptides provided herein (and / or nucleic acids designed to express a CD4-D1 polypeptide provided herein) can be administered to a mammal (e.g., a human) in need thereof (e.g., a human having, or at risk of developing, an HIV (e.g., an HIV-1) infection) to slow, delay, or prevent the development of an HIV (e.g., an HIV-1) infection.

[0060] In some cases, the methods and materials described herein can be used to reduce or eliminate the number of cells within a mammal (e.g., a human) that are infected by one or more HIV (e g., HIV-1) particles. For example, the materials and methods described herein can be used to reduce the number of HIV-infected cells present within a mammal by, for example, 10, 20, 30, 40, 50, 60, 70, 80, 90, 95, or more percent.

[0061] In some cases, the methods and materials described herein can be used to reduce or eliminate the viral load of a mammal (e.g., a human) that is infected by one or more HIV (e.g., HIV-1) particles. For example, the materials and methods described herein can be used to reduce the number of HIV particles present in the blood and / or gastrointestinal (GI) tract of a mammal by, for example, 10, 20, 30, 40, 50, 60, 70, 80, 90, 95, or more percent.

[0062] In some cases, the methods and materials described herein can be used to reduce or eliminate the number of viral particles present in one or more cells within a mammal (e.g., a human) that is infected by one or more HIV (e.g., HIV-1) particles. For example, the materials and methods described herein can be used to reduce the number of viral particles present in one or more cells within a mammal by, for example, 10, 20, 30, 40, 50, 60, 70, 80, 90, 95, or more percent.

[0063] In some cases, the methods and materials described herein can be used to reduce or eliminate one or more symptoms of an HIV (e.g., an HIV-1) infection. For example, one or more CD4-D1 polypeptides provided herein (and / or nucleic acids designed to express a CD4- D1 polypeptide provided herein) can be administered to a mammal (e.g., a human) in need thereof (e.g., a human having an HIV (e.g., an HIV-1) infection) to reduce or eliminate one or more symptoms of an HIV (e.g., an HIV-1) infection. Examples of symptoms of an HIV (e.g., an HIV-1) infection include, without limitation, fever, headache, muscle aches and joint pain, rash, sore throat and painful mouth sores, swollen lymph glands (e.g., swollen lymph nodes on the neck), diarrhea, weight loss, cough, night sweats, fatigue, vomiting, vision loss, and memory loss. In some cases, the materials and methods described herein can be used to reduce the severity of one or more symptoms of an HIV (e.g., an HIV-1) infection in a mammal (e.g., a human) by, for example, 10, 20, 30, 40, 50, 60, 70, 80, 90, 95, or more percent.

[0064] Any appropriate mammal having, or at risk of developing, an HIV (e.g., an HIV-1) infection can be treated as described herein (e.g., by administering one or more CD4-D1 polypeptides provided herein (and / or nucleic acids designed to express a CD4-D1 polypeptide provided herein)). Examples of mammals that can be treated as described herein include, without limitation, humans and non-human primates (e.g., monkeys).

[0065] When treating a mammal (e.g., a human) having, or at risk of developing, an HIV infection as described herein (e.g., by administering one or more CD4-D1 polypeptides provided herein (and / or nucleic acids designed to express a CD4-D1 polypeptide provided herein)), the HIV infection can be any type of HIV infection. Examples of types of HIV infections that can be treated as described herein include, without limitation, HIV-1 infections and HIV-2 infections. In some cases, instead of or in addition to begin used to treat a mammal having, or at risk of developing, an HIV infection, the methods and materials provided herein can be used to treat a mammal having, or at risk of developing, a simian immunodeficiency virus (SIV) infection or a simian-human immunodeficiency virus (SHIV) infection. In some cases, the methods and materials provided herein can be used to slow or prevent the progression of an HIV (e.g., an HIV-1) infection to acquired immunodeficiency syndrome (AIDS). For example, one or more CD4-D1 polypeptides provided herein (and / or nucleic acids designed to express a CD4-D1 polypeptide provided herein) can be administered to a mammal (e.g., a human) having an HIV infection to slow or prevent the progression of the HIV infection to AIDS within the mammal.

[0066] In some cases, the methods and materials provided herein can be used to treat a mammal having an HIV (e.g., an HIV-1) infection that has progressed to acquired immunodeficiency syndrome (AIDS). For example, one or more CD4-D1 polypeptides provided herein (and / or nucleic acids designed to express a CD4-D1 polypeptide provided herein) can be administered to a mammal (e.g., a human) having AIDS to treat the mammal.

[0067] In some cases, a mammal (e.g., a human) can be identified as having an HIV (e.g., an HIV-1) infection. For example, antigen tests (e.g., to identify antigens present on an HIV virus), antibody tests (e.g., to identify the presence of anti-HIV antibodies in blood or saliva), nucleic acid tests (e.g., to identify the presence of nucleic acid encoding an HIV virus in the blood, GI tract, lymphoid organs, and / or central nervous system (CNS) can be used to identify a mammal as having an HIV (e.g., an HIV-1) infection.

[0068] In some cases, a mammal (e.g., a human) can be identified as being at risk of developing an HIV (e.g., an HIV-1) infection. For example, sexual contact with an HIV+partner, sharing needles with an FHV+person, receiving a blood transfusion of blood from an HIV+person, being born to an HIV+mother, breastfeeding from an HIV+mother, and / or having one or more sexually transmitted diseases (e.g., herpes, gonorrhea, and / or syphilis) can be used to identify a mammal as being at risk of developing an HIV (e.g., an HIV-1) infection.

[0069] Once identified as having, or as being at risk of developing, an HIV (e.g., an HIV-1) infection, the mammal (e.g., the human) can be administered, or instructed to self-administer, one or more CD4-D1 polypeptides provided herein (and / or nucleic acids designed to express a CD4-D1 polypeptide provided herein) as described herein.

[0070] Any appropriate CD4-D1 polypeptide provided herein (and / or nucleic acid designed to express a CD4-D1 polypeptide provided herein) can be administered to a mammal (e.g., a human) having, or at risk of developing, an HIV (e.g., an HIV-1) infection as described herein.

[0071] In some cases, a CD4-D1 polypeptide that can be used as described herein (e.g., to treat a mammal having, or at risk of developing, an HIV (e.g., an HIV-1) infection) can have one or more (e.g., one, two, three, four, five, or more) modifications as compared to a wildtype human CD4-D1 polypeptide. For example, a CD4-D1 polypeptide that can be used as described herein can have one or more modifications as compared to an amino acid sequence set forth in SEQ ID NO: 1 (see, e.g., Example 1). In some cases, a nucleic acid encoding a CD4-D1 polypeptide that can be used as described herein can have one or more modifications as compared to a nucleotide sequence set forth in SEQ ID NO:2 (see, e.g., Example 1).

[0072] In some cases, a CD4-D1 polypeptide that can be used as described herein (e.g., to treat a mammal having, or at risk of developing, an HIV (e.g., an HIV-1) infection) can comprise, consist essentially of, or consist of the amino acid sequence set forth in any one of SEQ ID NOs:4-39 (e.g., any one of SEQ ID NOs:4-8). For example, a CD4-D1 polypeptide that can be used as described herein can have an amino acid sequence set forth in any one of SEQ ID NOs:4-39 (e.g., any one of SEQ ID NOs:4-8) (see, e.g., Example 2). In some cases, a nucleic acid encoding a CD4-D1 polypeptide that can be used as described herein can have a nucleotide sequence that encodes the amino acid sequence set forth in any one of SEQ ID NOs:4-39 (e.g., any one of SEQ ID NOs:4-8) (see, e.g., Example 2). In some cases, a nucleic acid encoding a CD4-D1 polypeptide that can be used as described herein can have a nucleotide sequence set forth in any one of SEQ ID NOs:41-46 (see, e.g., Example 2).

[0073] In some cases, a CD4-D1 polypeptide provided herein can comprise, consist essentially of, or consist of the amino acid sequence set forth in any one of SEQ ID NOs:4-39 (e.g., any one of SEQ ID NOs:4-8). In some cases, a CD4-D1 polypeptide provided herein can comprise, consist essentially of, or consist of the amino acid sequence set forth in any one of SEQ ID NOs:4-39 (e.g., any one of SEQ ID NOs:4-8) with zero, one, two, three, four, or five amino acid residues preceding the articulated sequence, with zero, one, two, three, four, or five amino acid residues following the articulated sequence, or with zero, one, two, three, four, or five amino acid substitutions within the articulated sequence, provided that the CD4-D1 polypeptide retains at least some activity exhibited by a CD4-D1 polypeptide that consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-8 (e.g., the ability to reduce or eliminate entry of HIV particles into cells within a mammal (e.g., a human)).

[0074] In some cases, a CD4-D1 polypeptide provided herein (e.g., a CD4-D1 polypeptide that can be used to treat a mammal having, or at risk of developing, an HIV (e.g., an HIV-1) infection) can comprise, consist essentially of, or consist of the amino acid sequence set forth in SEQ ID NO:40 set forth below, where each X is independently a neutral or charged amino acid with a small side chain. Examples of amino acids that are neutral or charged and have a small side chain include, without limitation, A, S, T, E, D, Q, N, and H. For example, a CD4-D1 polypeptide provided herein can comprise, consist essentially of, or consist of the following amino acid sequence:

[0075] KKWYAKKGDTVELTCTASQKKNI QFXWKNSNQIKI LGNQGS FLTKGPSKLNDRVDSRR ELWDQGNFPLI IKNLKIEDSDXYI CXMEDXKEEVQLLVSG (SEQ ID NO:40), where X = A, S, T, E, D, Q, N, or H.

[0076] Examples of CD4-D1 polypeptides that can be used as described herein (e.g., to treat a mammal having, or at risk of developing, an HIV (e.g., an HIV-1) infection) are set forth in Table 1.

[0077] Table 1. Exemplary CD4-D1 polypeptides.

[0078]

[0079] A CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID N0s:4- 39) can be any appropriate length (e.g., can include any number of amino acids). In some cases, a CD4-D1 polypeptide that can be used as described herein can be from about 98 amino acids in length to about 102 amino acids in length (e.g., from about 98 to about 101, from about 98 to about 100, from about 98 to about 99, from about 99 to about 102, from about 100 to about 102, from about 101 to about 102, from about 99 to about 101, from about 99 to about 100, or from about 100 to about 101 amino acids in length), provided that it retains at least some activity exhibited by a CD4-D1 polypeptide that consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39 (e.g., the ability to reduce or eliminate entry of HIV particles into cells within a mammal (e.g., a human)). For example, a CD4-D1 polypeptide that can be used as described herein can be about 99 amino acids in length. A CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID N0s:4- 39) can have any appropriate half-life. In some cases, a CD4-D1 polypeptide that can be used as described herein can have any appropriate serum half-life when administered to a mammal (e.g., a human). For example, a CD4-D1 polypeptide that can be used as described herein can have a half-life of from about 21.9 hours to about 98.9 hours (e.g., from about 22 to about 95, from about 22 to about 85, from about 22 to about 75, from about 22 to about 65, from about 22 to about 50, from about 22 to about 30, from about 25 to about 98, from about 35 to about 98, from about 45 to about 98, from about 55 to about 98, from about 65 to about 98, from about 75 to about 98, from about 25 to about 90, from about 35 to about 80, from about 45 to about 70, from about 55 to about 60, from about 25 to about 45, from about 35 to about 55, from about 45 to about 65, from about 55 to about 75, from about 65 to about 85, or from about 75 to about 95 hours).

[0080] A CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4- 39) can be a stable polypeptide. In some cases, a CD4-D1 polypeptide that can be used as described herein can be stable under particular conditions. For example, a CD4-D1 polypeptide that can be used as described herein can be thermostable. For example, a CD4- D1 polypeptide that can be used as described herein can have a melting temperature of from about 72.8°C to about 79.8°C (e.g., from about 73CC to about 78°C, from about 73°C to about 77°C, from about 73°C to about 76°C, from about 73°C to about 75°C, from about 73°C to about 74°C, from about 74°C to about 79°C, from about 75°C to about 79°C, from about 76°C to about 79°C, from about 77°C to about 79°C, from about 78°C to about 79°C, from about 74 C to about 78°C, from about 75°C to about 77°C, from about 74°C to about 76°C, from about 75°C to about 77°C, or from about 76°C to about 78CC).

[0081] A CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4- 39) can have any appropriate binding affinity for a gpl20 glycopeptide. A CD4-D1 polypeptide that can be used as described herein can have a binding affinity for any appropriate gpl20 glycopeptide. For example, a CD4-D1 polypeptide that can be used as described herein can have binding affinity for a gpl20 glycopeptide as described in, for example, Kwong et al., Nature, 393(6686):648-59 (1998). In some cases, a CD4-D1 polypeptide that can be used as described herein can have a high affinity for a gpl20 glycopeptide. For example, a CD4-D1 polypeptide that can be used as described herein can have an EC50 for a gpl20 glycopeptide of from about 3.4 nM to about 1.4 nM (e.g., from about 3 nM to about 1.4 nM, from about 2.7 nM to about 1.4 nM, from about 2.5 nM to about 1.4 nM, from about 2.2 nM to about 1.4 nM, from about 2.0 nM to about 1.4 nM, from about 1.7 nM to about 1.4 nM, from about 3.4 nM to about 1.6 nM, from about 3.4 nM to about 1.8 nM, from about 3.4 nM to about 2.0 nM, from about 3.4 nM to about 2.3 nM, from about 3.4 nM to about 2.5 nM, from about 3.4 nM to about 2.8 nM, from about 3.4 nM to about 3.0 nM, from about 3.0 nM to about 1.8 nM, from about 2.8 nM to about 2.0 nM, from about 3.0 nM to about 2.5 nM, or from about 2.5 nM to about 2.0 nM).

[0082] A CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4- 39) can have little or no affinity for a MHC complex. A CD4-D1 polypeptide that can be used as described herein can have little or no binding affinity for any appropriate MHC complex. For example, a CD4-D1 polypeptide that can be used as described herein can have little or no binding affinity for a MHC II complex. In some cases, a CD4-D1 polypeptide that can be used as described herein can have an equilibrium dissociation constant (KD) for a MHC complex that is effectively infinity. In some cases, a CD4-D1 polypeptide that can be used as described herein can have a KD for a MHC complex of greater than about 9.5 nm (e.g., greater than about 10 nm, greater than about 12 nm, greater than about 15 nm, greater than about 18 nm, greater than about 20 nm, greater than about 22 nm, or greater than about 25 nm).

[0083] In some cases, a CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39) can avoid immune-suppression. For example, when a CD4-D1 polypeptide that can be used as described herein is administered to a mammal (e.g., a human), the CD4-D1 polypeptide is not recognized (e.g., recognized and destroyed) by that mammal’s immune system. For example, when a CD4-D1 polypeptide that can be used as described herein is administered to a mammal (e.g., a human), the CD4-D1 polypeptide is not recognized (e.g., recognized and destroyed) by antigen presenting cells (APCs) present within that mammal.

[0084] In some cases, two or more (e.g., two, three, four, or more) CD4-D1 polypeptides provided herein (e.g., two or more polypeptides that each comprise, consist essentially of, or consist of the amino acid sequence set forth in any one of SEQ ID NOs:4-39) can be conjugated together (e.g., to form a multivalent molecule such as a bivalent molecule). For example, a nucleic acid can be designed to encode a first CD4-D1 polypeptide provided herein followed by a second CD4-D1 polypeptide provided herein such that the encoded polypeptide is a fusion polypeptide including both the first CD4-D1 polypeptide and the second CD4-D1 polypeptide. In some cases, two or more CD4-D1 polypeptides provided herein can be conjugated together to form a bivalent molecule.

[0085] In some cases, a CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39) can be conjugated to one or more additional molecules. For example, a nucleic acid can be designed to encode a CD4-D1 polypeptide provided herein followed by (or preceded by) one or more additional polypeptides such that the encoded polypeptide is a fusion polypeptide including both the CD4-D1 polypeptide and the one or more additional polypeptides. In some cases, an additional molecule that can be conjugated to a CD4-D1 polypeptide provided herein can stabilize the CD4-D1 polypeptide. In some cases, an additional molecule that can be conjugated to a CD4-D1 polypeptide provided herein can be a targeting moiety. For example, an additional molecule that can be conjugated to a CD4-D1 polypeptide provided herein can be an antibody or antibody fragment (e.g., such that the CD4-D1 polypeptide provided herein can be used as a bispecific molecule). In some cases, an additional molecule that can be conjugated to a CD4-D1 polypeptide provided herein can induce an immune response. For example, an additional molecule that can be conjugated to a CD4-D1 polypeptide provided herein can be an immunogen (e.g., such that the CD4-D1 polypeptide provided herein can be used as a vaccine composition). Examples of additional molecules that can be conjugated to a CD4-D1 polypeptide provided herein can include, without limitation, those described elsewhere (see, e.g., Saha et al., Biochemistry, 50(37):7891-900 (2011); Chen et al., J. Virol., 85(18):9395-405 (2011); Chen et al., J. Virol., 88(2): 1125-39 (2014); Caskey et al., Nat. Med., 23(2): 185-191 (2017); Suphaphiphat et al., Nat. Commim., 14(1):6224 (2023); Gaebler et al., Nature, 606(7913):368-374 (2022); Chen et al., Antiviral. Res., 88(1): 107-15 (2010); Chen et al., Antiviral. Res., 116:62-6 (2015); Chen et al., J. Virol., 88(2): 1125-39 (2014); Chen et al., Proc. Natl. Acad. Sci. USA, 105(44): 17121-6 (2008); International Patent Application Publication No. WO 2011 / 146891; U.S. Patent Application Publication No. 2018 / 0086812; and U.S. Patent Application Publication No. 2018 / 0118815).

[0086] In some cases, one or more (e.g., one or two) CD4-D1 polypeptides where each CD4- D1 polypeptide comprises, consists essentially of, or consists of the amino acid sequence set forth in SEQ ID NO:4 can be conjugated to a Fc polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in SEQ ID NO:47.

[0087] In some cases, one or more (e g., one or two) CD4-D1 polypeptides where each CD4- DI polypeptide comprises, consists essentially of, or consists of the amino acid sequence set forth in SEQ ID NO:5 can be conjugated to a Fc polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in SEQ ID NO:47.

[0088] In some cases, one or more (e.g., one or two) CD4-D1 polypeptides where each CD4- D1 polypeptide comprises, consists essentially of, or consists of the amino acid sequence set forth in SEQ ID NO:4 can be conjugated to at least a portion of an m36.4 antibody that comprises, consists essentially of, or consists of the amino acid sequence set forth in SEQ ID NO:48.

[0089] In some cases, one or more (e.g., one or two) CD4-D1 polypeptides where each CD4- DI polypeptide comprises, consists essentially of, or consists of the amino acid sequence set forth in SEQ ID NO:5 can be conjugated to at least a portion of an m36.4 antibody that comprises, consists essentially of, or consists of the amino acid sequence set forth in SEQ ID NO:48.

[0090] In some cases, one or more (e.g., one or two) CD4-D1 polypeptides where each CD4- DI polypeptide comprises, consists essentially of, or consists of the amino acid sequence set forth in SEQ ID NO:4 can be conjugated to at least a portion of an 10-1074 antibody that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:49-50.

[0091] In some cases, one or more (e g., one or two) CD4-D1 polypeptides where each CD4-

[0092] DI polypeptide comprises, consists essentially of, or consists of the amino acid sequence set forth in SEQ ID N0:5 can be conjugated to at least a portion of an 10-1074 antibody that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:49-50.

[0093] Any appropriate method can be used to deliver one or more CD4-D1 polypeptides provided herein and / or nucleic acid designed to express a CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39 and / or nucleic acid designed to express a CD4-D1 polypeptide that comprises, consists essentially of, or consist of the amino acid sequence set forth in any one of SEQ ID NOs:4-39) to a mammal (e.g., a human). In some cases, one or more CD4-D1 polypeptides provided herein (and / or nucleic acid designed to express a CD4-D1 polypeptide provided herein) can be administered systemically to a mammal (e.g., a human). For example, one or more CD4-D1 polypeptides provided herein (and / or nucleic acid designed to express a CD4-D1 polypeptide provided herein) can be administered systemically to a mammal (e.g., a human) by intravenous injection, intramuscular injection, or subcutaneous injection. In some cases, one or more CD4-D1 polypeptides provided herein (and / or nucleic acid designed to express a CD4-D1 polypeptide provided herein) can be administered locally to any particular location within a mammal (e.g., a human). For example, one or more CD4-D1 polypeptides provided herein (and / or nucleic acid designed to express a CD4-D1 polypeptide provided herein) can be administered locally within a mammal (e g., a human) using intravaginal rings, rectal gels, subdermal implants, or intravitreal injections.

[0094] Any appropriate method can be used to obtain a CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39). For example, a CD4-D1 polypeptide that can be used as described herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39) can be obtained by synthesizing the polypeptide of interest using appropriate polypeptide synthesizing techniques.

[0095] When one or more nucleic acids designed to express a CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39) are administered to a mammal (e.g., a human), the nucleic acid can be in the form of a vector (e.g., a viral vector or a non-viral vector).

[0096] When a vector used to deliver nucleic acid encoding a CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39) to a mammal (e.g., a human) is a viral vector, any appropriate viral vector can be used. A viral vector can be derived from a positive-strand virus or a negative-strand virus. A viral vector can be derived from a virus with a DNA genome or an RNA genome. In some cases, a viral vector can be a chimeric viral vector. In some cases, a viral vector can infect dividing cells. In some cases, a viral vector designed to deliver nucleic acid encoding a nucleoporin polypeptide (or fragment thereof) can infect non-dividing cells. Examples of virus-based vectors that can be used to deliver nucleic acid encoding a CD4-D1 polypeptide that can be used as described herein to a mammal (e.g., a human) include, without limitation, virus-based vectors based on, adeno- associated viruses (AAV), retroviruses, lentiviruses, baculoviruses, and vaccinia viruses.

[0097] When a vector used to deliver nucleic acid encoding a CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39) to a mammal (e.g., a human) is a non-viral vector, any appropriate non-viral vector can be used. In some cases, a non-viral vector can be an expression plasmid (e.g., a cDNA expression vector).

[0098] When nucleic acid encoding a CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39) is administered to a mammal, the nucleic acid can be used for transient expression of a CD4-D1 polypeptide that can be used as described herein or for stable expression of a CD4-D1 polypeptide that can be used as described herein. In cases where a nucleic acid encoding a CD4-D1 polypeptide that can be used as described herein is used for stable expression of the CD4-D1 polypeptide, the nucleic acid encoding a CD4-D1 polypeptide that can be used as described herein can be engineered to integrate into the genome of a cell. Nucleic acid can be engineered to integrate into the genome of a cell using any appropriate method. For example, gene editing techniques (e g., CRISPR or TALEN gene editing) can be used to integrate nucleic acid designed to express a CD4-D1 polypeptide that can be used as described herein into the genome of a cell. In addition to nucleic acid encoding a CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39), a vector (e.g., a viral vector or a non-viral vector) can contain one or more regulatory elements operably linked to the nucleic acid encoding a CD4- D1 polypeptide that can be used as described herein. Such regulatory elements can include promoter sequences, enhancer sequences, response elements, signal peptides, internal ribosome entry sequences, polyadenylation signals, terminators, and inducible elements that modulate expression (e.g., transcription or translation) of a nucleic acid. The choice of regulatory element(s) that can be included in a vector depends on several factors, including, without limitation, inducibility, targeting, and the level of expression desired. For example, a promoter can be included in a vector to facilitate transcription of a nucleic acid encoding a CD4-D1 polypeptide that can be used as described herein. A promoter can be a naturally occurring promoter or a recombinant promoter. A promoter can be ubiquitous or inducible (e.g., in the presence of tetracycline), and can affect the expression of a nucleic acid encoding a polypeptide in a general or tissue-specific manner. Examples of promoters that can be used to drive expression of a CD4-D1 polypeptide that can be used as described herein in cells include, without limitation, CMV promoters, SV40 promoters, and EFla promoters. As used herein, “operably linked” refers to positioning of a regulatory element relative to a nucleic acid encoding a polypeptide in such a way as to permit or facilitate expression of the encoded polypeptide. For example, a vector can contain a promoter and nucleic acid encoding a CD4- D1 polypeptide that can be used as described herein. In this case, the promoter is operably linked to a nucleic acid encoding a CD4-D1 polypeptide that can be used as described herein such that it drives expression of the CD4-D1 polypeptide in cells.

[0099] In some cases, nucleic acid encoding a CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39) can contain nucleic acid encoding a detectable label. For example, a vector can include nucleic acid encoding a CD4-D1 polypeptide that can be used as described herein and nucleic acid encoding a detectable label positioned such that the encoded polypeptide is a fusion polypeptide that includes a CD4-D1 polypeptide fused to a detectable polypeptide. In some cases, a detectable label can be a peptide tag. In some cases, a detectable label can be a fluorescent molecule (e.g., fluorescent polypeptides). Examples of detectable labels that can be used as described herein include, without limitation, Fc tags, His tags, HA tags, Myc-tags, FLAG-tags, GST tags, MBP tags, GFP tags, and S tags.

[0100] Nucleic acid encoding a CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39) can be produced using any appropriate techniques including, without limitation, common molecular cloning, polymerase chain reaction (PCR), chemical nucleic acid synthesis techniques, and combinations of such techniques. For example, PCR or RT-PCR can be used with oligonucleotide primers designed to amplify nucleic acid (e.g., genomic DNA or RNA) encoding a CD4-D1 polypeptide that can be used as described herein.

[0101] In some cases, one or more CD4-D1 polypeptides provided herein and / or nucleic acid designed to express a CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39 and / or nucleic acid designed to express a CD4-D1 polypeptide that comprises, consists essentially of, or consist of the amino acid sequence set forth in any one of SEQ ID NOs:4-39) can be formulated into a composition (e.g., a pharmaceutical composition) for administration to a mammal (e.g., a human). For example, one or more CD4-D1 polypeptides that can be used as described herein can be formulated into a pharmaceutically acceptable composition for administration to a mammal (e.g., a human) having, or at risk of developing, an HIV (e.g., an HIV-1) infection. In some cases, one or more CD4-D1 polypeptides provided herein (and / or nucleic acid designed to express a CD4- D1 polypeptide provided herein) can be formulated together with one or more pharmaceutically acceptable carriers (additives), excipients, and / or diluents. Examples of pharmaceutically acceptable carriers, excipients, and diluents that can be used in a composition described herein include, without limitation, sucrose, lactose, starch (e.g., starch glycolate), cellulose, cellulose derivatives (e.g., modified celluloses such as microcrystalline cellulose and cellulose ethers like hydroxypropyl cellulose (HPC) and cellulose ether hydroxypropyl methylcellulose (HPMC)), xylitol, sorbitol, mannitol, gelatin, polymers (e.g., polyvinylpyrrolidone (PVP), polyethylene glycol (PEG), crosslinked polyvinylpyrrolidone (crospovidone), carboxymethyl cellulose, polyethylene-polyoxypropylene-block polymers, and crosslinked sodium carboxymethyl cellulose (croscarmellose sodium)), titanium oxide, azo dyes, silica gel, fumed silica, talc, magnesium carbonate, vegetable stearin, magnesium stearate, aluminum stearate, stearic acid, antioxidants (e.g., vitamin A, vitamin E, vitamin C, retinyl palmitate, and selenium), citric acid, sodium citrate, parabens (e.g., methyl paraben and propyl paraben), petrolatum, dimethyl sulfoxide, mineral oil, serum proteins (e.g., human serum albumin), glycine, sorbic acid, potassium sorbate, water, salts or electrolytes (e.g., saline, protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, and zinc salts), colloidal silica, magnesium tri silicate, polyacrylates, waxes, wool fat, and lecithin.

[0102] A composition (e.g., a pharmaceutical composition) containing one or more CD4-D1 polypeptides provided herein and / or nucleic acid designed to express a CD4-D1 polypeptide provided herein (e g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39 and / or nucleic acid designed to express a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39) can be formulated into any appropriate dosage form. Examples of dosage forms include solid or liquid forms including, without limitation, gels, liquids, suspensions, and solutions (e.g., sterile solutions).

[0103] In some cases, a composition (e.g., a pharmaceutical composition) containing one or more CD4-D1 polypeptides provided herein and / or nucleic acid designed to express a CD4- D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39 and / or nucleic acid designed to express a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39) can be designed for parenteral (e.g., intravenous, intramuscular, subcutaneous, intravaginal, rectal, subdermal, and intravitreal) administration. Compositions suitable for parenteral administration include aqueous and non-aqueous sterile injection solutions that can contain anti-oxidants, buffers, bacteriostats, and solutes which render the formulation isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions which may include suspending agents and thickening agents.

[0104] Compositions (e.g., pharmaceutical compositions) containing one or more CD4-D1 polypeptides provided herein and / or nucleic acid designed to express a CD4-D1 polypeptide provided herein (e g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39 and / or nucleic acid designed to express a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39) can be presented in unit-dose or multidose containers, for example, sealed ampules and vials, and may be stored in a freeze dried (lyophilized) condition requiring only the addition of the sterile liquid carrier, for example water for injections, immediately prior to use. Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules, and tablets.

[0105] An effective amount of a composition (e.g., a pharmaceutical composition) containing one or more CD4-D1 polypeptides provided herein and / or nucleic acid designed to express a CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39 and / or nucleic acid designed to express a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39) can be any amount that can treat a mammal having, or at risk of developing, an HIV (e.g., an HIV-1) infection without producing significant toxicity to the mammal. For example, an effective amount of one or more CD4-D1 polypeptides that can be used as described herein can be from about 10 milligrams of polypeptide(s) per kilogram body weight of the mammal (mg / kg) per day to about 100 mg / kg per day (e.g., from about 10 mg / kg to about 90 mg / kg, from about 10 mg / kg to about 80 mg / kg, from about 10 mg / kg to about 70 mg / kg, from about 10 mg / kg to about 60 mg / kg, from about 10 mg / kg to about 50 mg / kg, from about 10 mg / kg to about 40 mg / kg, from about 10 mg / kg to about 30 mg / kg, from about 10 mg / kg to about 20 mg / kg, from about 20 mg / kg to about 100 mg / kg, from about 30 mg / kg to about 100 mg / kg, from about 40 mg / kg to about 100 mg / kg, from about 50 mg / kg to about 100 mg / kg, from about 60 mg / kg to about 100 mg / kg, from about 70 mg / kg to about 100 mg / kg, from about 80 mg / kg to about 100 mg / kg, from about 90 mg / kg to about 100 mg / kg, from about 20 mg / kg to about 90 mg / kg, from about 30 mg / kg to about 80 mg / kg, from about 40 mg / kg to about 70 mg / kg, from about 50 mg / kg to about 60 mg / kg, from about 20 mg / kg to about 40 mg / kg, from about 30 mg / kg to about 50 mg / kg, from about 40 mg / kg to about 60 mg / kg, from about 50 mg / kg to about 70 mg / kg, from about 60 mg / kg to about 80 mg / kg, or from about 70 mg / kg to about 90 mg / kg). The effective amount can remain constant or can be adjusted as a sliding scale or variable dose depending on the mammal’s response to treatment. Various factors can influence the actual effective amount used for a particular application. For example, the frequency of administration, duration of treatment, use of multiple treatment agents, route of administration, and severity of the condition may require an increase or decrease in the actual effective amount administered.

[0106] The frequency of administration of a composition (e.g., a pharmaceutical composition) containing one or more CD4-D1 polypeptides provided herein and / or nucleic acid designed to express a CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39 and / or nucleic acid designed to express a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39) can be any frequency that can treat a mammal (e.g., a human) having, or at risk of developing, an HIV (e.g., an HIV-1) infection without producing significant toxicity to the mammal. For example, the frequency of administration can be from about two times a day to about once a month, from about once a day to about twice a month, or from about once a week to about every two weeks. The frequency of administration can remain constant or can be variable during the duration of treatment. A course of treatment with a composition containing one or more CD4-D1 polypeptides that can be used as described herein can include rest periods. As with the effective amount, various factors can influence the actual frequency of administration used for a particular application. For example, the effective amount, duration of treatment, use of multiple treatment agents, route of administration, and severity of the condition may require an increase or decrease in administration frequency.

[0107] An effective duration for administering a composition (e.g., a pharmaceutical composition) containing one or more CD4-D1 polypeptides provided herein and / or nucleic acid designed to express a CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39 and / or nucleic acid designed to express a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39) can be any duration that treat a mammal (e.g., a human) having, or at risk of developing, an HIV (e.g., an HIV-1) infection without producing significant toxicity to the mammal. For example, the effective duration can vary from several days to several weeks, months, or years. In some cases, the effective duration for the treatment of a mammal (e.g., a human) having, or at risk of developing, an HIV (e.g., an HIV-1) infection can be the duration of the life of the mammal. Multiple factors can influence the actual effective duration used for a particular treatment. For example, an effective duration can vary with the frequency of administration, effective amount, use of multiple treatment agents, route of administration, and severity of the condition being treated.

[0108] In some cases, the one or more CD4-D1 polypeptides provided herein and / or nucleic acid designed to express a CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39 and / or nucleic acid designed to express a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39) can be used as the sole active agent(s) used to treat a mammal (e.g., a human) having, or at risk of developing, an HIV (e.g., an HIV-1) infection.

[0109] In some cases, the methods and materials described herein can include one or more (e.g., one, two, three, four, five or more) additional agents used to treat a mammal (e.g., a human) having, or at risk of developing, an HIV (e.g., an HIV-1) infection. For example, an agent that can be used to treat a mammal having, or at risk of developing an HIV infection can be an antiretroviral therapy (ART). In some cases, an ART can include administration of one or more of a non-nucleoside reverse transcriptase inhibitor (NNRTI), a nucleoside or nucleotide reverse transcriptase inhibitor (NRTI), a protease inhibitor, an integrase inhibitor, and an entry or fusion inhibitor. For example, an agent that can be used to a treat a mammal having, or at risk of developing, an HIV infection can be an antibody-drug conjugate (ADC). Examples of agents used to treat an HIV (e.g., an HIV-1) infection that can be administered to a mammal (e.g., a human) having, or at risk of developing, an HIV (e.g., an HIV-1) infection together with one or more CD4-D1 polypeptides provided herein and / or nucleic acid designed to express a CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39 and / or nucleic acid designed to express a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39) include, without limitation, efavirenz (e.g., SUSTIVA®), rilpivirine (e g., EDURANT®), doravirine (e.g., PIFELTRO®), abacavir (e.g., ZIAGEN®), tenofovir disoproxil fumarate (e.g., VIREAD®), emtricitabine (e.g., EMTRIVA®), lamivudine (e.g., EPIVIR®), zidovudine (e.g., RETROVIR®), emtricitabine / tenofovir disoproxil fumarate (e.g., TRUVADA®), emtricitabine / tenofovir alafenamide fumarate (e.g., DESCOVY®), atazanavir (e.g., REYATAZ®), darunavir (e.g., PREZISTA®), lopinavir / ritonavir (e.g., KALETRA®), bictegravir sodium / emtricitabine / tenofovir alafenamide fumarate (e.g., BIKTARVY®), raltegravir (e.g., ISENTRESS®), dolutegravir (e.g., TIVICAY™), cabotegravir (e.g., VOCABRIA), enfuvirtide (e.g., FUZEON®), maraviroc (e.g., SELZENTRY®), cobi cistat (e g., TYBOST™), and ibalizumab (e.g., TROGARZO®). In some cases, the one or more additional agents can be administered together with one or more CD4-D1 polypeptides provided herein and / or nucleic acid designed to express a CD4-D1 polypeptide provided herein. In some cases, the one or more additional agents can be administered independent of the one or more CD4-D1 polypeptides provided herein (and / or nucleic acid designed to express a CD4-D1 polypeptide provided herein). When the one or more additional agents are administered independent of the one or more CD4-D1 polypeptides that can be used as described herein, the one or more CD4-D1 polypeptides that can be used as described herein can be administered first, and the one or more additional agents administered second, or vice versa.

[0110] In some cases, the methods and materials described herein can include subjecting a mammal having, or at risk of developing, an HIV (e.g., an HIV-1) infection to one or more (e.g., one, two, three, four, five or more) additional therapies (e g., therapeutic interventions) that are effective to treat an HIV (e.g., an HIV-1) infection. Examples of therapies that can be used to treat an HIV (e.g., an HIV-1) infection include, without limitation, CAR-T cell therapies. In some cases, the one or more additional treatments that are effective to treat an HIV (e.g., an HIV-1) infection can be performed at the same time as the administration of the one or more CD4-D1 polypeptides provided herein and / or nucleic acid designed to express a CD4-D1 polypeptide provided herein (e.g., a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39 and / or nucleic acid designed to express a polypeptide that comprises, consists essentially of, or consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-39). In some cases, the one or more additional treatments that are effective to treat an HIV (e.g., an HIV-1) infection can be performed before and / or after the administration of the one or more CD4-D1 polypeptides provided herein and / or nucleic acid designed to express a CD4-D1 polypeptide provided herein.

[0111] The invention will be further described in the following examples, which do not limit the scope of the invention described in the claims.

[0112] EXAMPLES

[0113] Example 1: Exemplary Human CD4-D1 Sequences

[0114] Exemplary wild type human CD4-D1 polypeptide:

[0115] KKWLGKKGDTVELTCTASQKKS IQFHWKNSNQIKILGNQGS FLTKGPSKLNDRADSRRSLW

[0116] DQGNFPLI IKNLKIEDSDTYICEVEDQKEEVQLLVFG

[0117] (SEQ ID NO:!)

[0118] Nucleic Acid encoding an exemplary wild-type human CD4-D1 polypeptide:

[0119] AAGAAGGTGGTGCTGGGCAAGAAGGGCGACACCGTGGAGCTGACCTGCACCGCCAGCCAGAA

[0120] GAAGAGCATCCAGTTCCACTGGAAGAACAGCAACCAGATCAAGATCCTGGGCAACCAGGGCA

[0121] GCTTCCTGACCAAGGGACCTAGCAAGCTGAACGACAGGGCAGACAGCCGGCGGAGCCTGTGG

[0122] GACCAGGGAAACTTCCCACTGATCATCAAGAACCTGAAGATCGAGGACAGCGACACCTACAT

[0123] CTGCGAGGTGGAGGACCAGAAGGAGGAGGTGCAGCTGCTGGTGTTCGGC

[0124] (SEQ ID NO:2)

[0125] Example 2: Exemplary Recombinant CD4-D1 Sequences

[0126] Exemplary recombinant CD4-D1 polypeptides:

[0127] Nucleic acid encoding an exemplary recombinant CD4-D1 polypeptide:

[0128] Example 3: Engineering next generation of long-lasting soluble CD4-D1 domains as HIV entry inhibitors

[0129] This Example describes the design and engineering of polypeptides that include a DI domain of a CD4 polypeptide (referred to herein as a CD4-D1 polypeptide) that can be used to prevent HIV infection. For example, this Example describes the design and engineering of small CD4-D1 polypeptides having increased stability and prolonged half-life. Materials and Methods

[0130] Phage Display, Biopanning, and Clone Characterization

[0131] Cavity filling hotspots and MHC Il-binding abrogation sites were identified by in silico structural modeling combined with folding and binding free energy calculations. Then cavity -filling hotspots were combined with MHC Il-binding abrogation sites along with folding residues in the D1-D2 interface to identify optimal clones using phage display technology. Specifically, in the phage libraries, these three groups of hotspots (i.e., residues 5 and 98 for modifying the D1 / D2 interface, residues 6, 55, and 86 for filling cavities, and residue 60 for abrogating MHC II binding) are randomized by using degenerated codons introduced into overlapping primers

[0132] The entire CD4-D1 gene variants were assembled by overlapping PCR and then subcloned into phagemid pComb3x, which was further packaged into phage particles under aid of helper phage M13KO7.

[0133] The new CD4-D1 variants were identified by a competitive phage panning strategy, in which the CD4-D1 phage particles were first depleted against MHC II-peptide complex coated beads. The beads display multimers of MHC II due to biotin-streptavidin coupling. The depleted CD4-D1 phage was then pull-down gpl20 from different HIV isolates. The panning process was done for 3 rounds, followed by ELISA screening of monoclonal phage clones. Those clones exhibiting high binding signals were sanger sequenced and extensively characterized in terms of gpl20 binding, thermostability, MHC II binding, HIV-1 neutralization, and in vivo half-life in mice.

[0134] Results

[0135] It was examined whether enhancing molecular folding compactness can increase molecular themostability and decrease propensity to protease degradation. Molecular interior cavities in CD4-D1 were identified through in silico structural computations, and those cavities were filled by mutagenesis into residues with bulkier side chains (Figure 3). To decease the “sink” effects of MHC II, the interaction interface differences between CD4 / MHC II and CD4 / gpl20 were scrutinized and compared, and sCD4 variants with abolished MHC II binding but preserved binding affinity to gp!20 were identified (Figure 1 and Figure 2B). In addition to extending the half- / / / c in vivo, elimination of MHC II binding from sCD4 decreased potential negative effects on the immune response resulting from CD4- MHC binding.

[0136] The cavity filling mutations and the MHC II binding elimination mutations were combined into one molecule through phage display method (Figure 3). A comprehensive phage display library was generated that contains all possible combinations of mutations. To facilitate binder identification, MHC II-peptide complex (HLA-DR) was recombinantly expressed using mammalian cell expression system (expi293 cells). The recombinant MHC II alpha and beta chains contain a Fos / Jun motif to promote its heterodimerization as well as a HIS tag for purification and AVI tag for biotin labeling. By using Ni-NTA chromatography, pure protein was obtained, and its folding intactness was validated by using anti-human HLA-DR antibody (Figure 2A). To prepare phage panning baits, MHC II was biotinylated followed by ligation onto streptavidin modified dyna beads. Then a competitive phage panning method was performed to obtain DI variants with abrogated MHC II binding but with retained gp!20 binding (Figure 4). Several CD4-D1 variants were obtained from phage panning and screening.

[0137] The thermostability of the CD4-D1 variants was compared using temperature ramping CD spectrum, in which proteins unfolding was monitored by CD spectrum along with increased temperature. Results showed that clones 1A2-4 and 1D9 exhibited significantly increased melting temperatures than the prototypic CD4-D1 mD1.22 (Figure 5).

[0138] Another aim was to extend half-life of CD4-D1. It was examined whether MHC II binding may serve a sink for CD4-D1, such that abrogation of MHC II binding could extend CD4-D1 half-life. MHC II binding and in vivo half-life of the clones was tested in balb / c mice in the context of human IgGl Fc. Fc fusion protein binding was compared the MHC II. Results showed that while mD1.22, and two other positive controls (D63W and LSEVhLS) exhibited substantial binding to MHC II (large AUC in the binding curves), and the new CD4-D1 variants from phage panning, including 1A2-4, 1A2-3 and S60E, 1D9 and 2C6, showed significantly decreased binding to MHC II. Impressively, 1A2-4 and S60E binding was below detection limit (Figure 6). Then the variants in vivo PK in mice was compared. Results showed that 1A2-4 and S60E exhibits prolonged durations than other compounds. A negative correlation was found between MHC II binding and in vivo half-life (Figure 6). Based on these results, 1 A2-4 and S60E were chosen for further characterization. The PK of 1A2-4 and S60E was ascertained in the same mice for monitoring longer time up to two weeks (Figure 7). Results showed that 1A2-4 and S60E had significantly extended half-life compared to mD1.22, and comparable to the clinical benchmark, bnAb 10-1074. It should be noted that 1A2-4 and S60E half-life can be further extended by engineering fc with mutations such as LS and YTE.

[0139] The neutralization of 1A2-4 and S60E against HIV T / F and pseudoviruses was tested with comparison to mD1.22 and clinical benchmark, bnAb 3BNC117 (Figures 8, 9, and 10). Results showed that 1A2-4 and S60E broadly neutralized all HIV-1 variants tested, in contrast to 3BNC117 showing inefficacy to some of HIV- 1 variants. And 1A2-4 was more effective than 3BNC117 in some HIV strains. 1 A2-4 and S60E neutralization potency was similar as mD1.22. A sequence alignment is shown in Figure 11.

[0140] In summary, CD4-D1 variants 1A2-4 and S60E showed enhanced thermostability, abrogated MHC II binding, potent and broad HIV neutralization, and extended in vivo serum half-life as tested in a mouse model.

[0141] Together, these results demonstrate that these CD4-D1 variant polypeptides can be used to prevent HIV-1 infection. For example, the CD4-D1 variant polypeptide can be used as a potent, broad, long-lasting, smaller, and highly stable sCD4-Dl molecule for prevention of HIV infection.

[0142] Example 4: Design and characterization of CD4-D1 polypeptides

[0143] The results in this Example re-present and expand on at least some of the results provided in other Examples.

[0144] This Example also describes the design and characterization of bispecific molecules including two CD4-D1 polypeptides described herein.

[0145] Materials and Methods

[0146] Surface Plasmon Resonance (SPR) Binding Test

[0147] SPR experiments were performed using a Biacore X100 (Cytiva). Gpl40sc in ~0.3 pM solution was immobilized onto a CM5 sensor chip (Cytiva) by amine coupling. The DI and the mutants were diluted in HBS-EP buffer (10 mM HEPES, 150 mM NaCl, 3 mM EDTA, and 0.005% surfactant P20, pH 7.4) and injected over an immobilized surface for 90 seconds at a rate of 50 pL / minute. Samples were allowed to dissociate for 600 seconds. After each sample injection, the surface was regenerated by injecting regeneration solution (10 mM Glycine / HCl buffer, with 10% glycerol at pH 2.0). To determine the kinetic values, ka, kd, and KD, data were analyzed by fitting the results to a 1 : 1 kinetic model using the SPR analysis software Scrubber (BioLogic Software). The experimental sensorgrams were subtracted from both the reference flow cell data and the average of blank buffer injection data.

[0148] BLItz Method

[0149] The binding of the CD4-D1 variant Fc fusion polypeptides to a recombinant human major histocompatibility complex class II (MHC-II) protein was detected using biolayer interferometry BLItz (ForteBio, Menlo Park, CA). The biotinylated MHC-II protein was mounted on streptavidin biosensors (ForteBio: 18-5019) for 2 minutes and equilibrated with Dulbecco’s phosphate-buffered saline (DPBS) (pH = 7.4) to establish the baselines. A 1000 nM Dl-Fc polypeptide (80 pg / mL) was loaded for association and monitored for 2 minutes. Then, the Dl-Fc polypeptides were allowed to dissociate in DPBS for 4 minutes. The sensorgrams were recorded for each CD4-D1 variants for comparison.

[0150] Size Exclusion Chromatography

[0151] The purity and structure of the antibodies were analyzed using Superdex 200 Increase 10 / 300 GL chromatography. Ferritin, aldolase, conalbumin, ovalbumin, carbonic anhydrase, and ribonuclease were used as standard proteins for calibration. A 500 pL sample mixture containing the standard proteins was loaded into a column and separated by AKTA explorer machine. For protein analysis, 100 pL of filtered S60E, 1A2-4, and mD1.22 in 1 * DPBS (Gibco, Waltham, MA, USA) were analyzed. Polypeptides were eluted by DPBS buffer at a flow rate of 0.5 mL / minute.

[0152] Thermostability Test

[0153] Purified polypeptides were diluted to 15 pM in 20 mM zytid buffer at pH 7.4. Circular dichroism (CD) spectra were recorded on a JASCO J810 spectrometer. Thermal stability of the polypeptides was measured by recording the CD signal at 226 nm with a temperature range of 30-90°C and a heating rate of 0.2°C / minute. Thermal melting temperatures were determined by fitting the temperature dependent data using Igor Pro software (WaveMetrics, Portland, OR). To assess the degree of polypeptide refolding after thermal denaturation, the CD spectra, from 200 nm to 280 nm wavelength, were recorded before and after the thermal stability experiments.

[0154] Dynamic Light Scattering (DLS)

[0155] The aggregation resistance of mD1.22-Fc, S60E-Fc, and lA2-4-Fc was measured by dynamic light scattering (DLS). The buffer was changed to DPBS and filtered using a 0.22 pm filter. The protein concentration was adjusted to 1 mg / mL. In total, a 500 pL antibody sample was incubated at 37°C without shaking. On days 0, 1, 2, 8 and 14, samples were taken for DLS measurements on Zetasizer Nano ZS ZEN3600 to determine the protein size distribution.

[0156] Cell Binding Test

[0157] To assess the cell surface binding ability of the CD4-D1 polypeptides with MHC II, cells including Jurkat MHC IT cells, 293 T cells, HuT-R5 MHC II+cells, M0LT-R5 MHC IF cells, J558 mouse MHC II+cells, and CHO-K1 MHC IL cells were incubated with 1A2-4, S60E, and mDl .22 His / Fc polypeptides for 0.5 hour at 4°C. The cells were then stained with PE-conjugated anti-FLAG or PE-conjugated goat anti-human IgG for 30 minutes at 4°C. Secondary antibodies used included anti-flag-PE (Figures 17A and 17C) and anti-Fc-PE (Figures 17 B and 17D), and D1D2-Fc served as a positive control antibody.

[0158] To assess the non-specific binding effects of the CD4-D1 polypeptides with peripheral blood mononuclear cells (PBMCs), human or mouse PBMCs were incubated with 1A2-4, S60E, and mDl.22 His / Fc polypeptides for 0.5 hours at 4°C. The cells were then stained with PE-conjugated anti-FLAG or PE-conjugated goat anti-human IgG for 30 minutes at 4°C. Secondary antibodies used included anti-flag-PE (Figures 17A and 17C) and anti-Fc-PE (Figures 17 B and 17D), and D1D2-Fc served as a positive control antibody.

[0159] Pharmacokinetic study

[0160] Balb / c mice (6-8 weeks old) were randomly divided into groups (n=3) and given intravenous tail vein injection of mD1.22-Fc, S60E-Fc, lA2-4-Fc, and 10-1074, at a dose of 25 mg / kg. From each mouse, 10 pL blood was collected at various time points: 1 hour, 4 hour, 8 hour, 1 day, 3 days, 5 days, 7 days, 9 days, 12 days, and 15 days. The samples were then diluted by 10-fold with PBS. The cells were removed by centrifugation at 1000*g for 10 minutes, and serum samples were stored at -80°C. The total antibody concentration was determined by ELISA. Briefly, a 96-well plate (half area, high-binding) was coated with a gpl40 recombinant protein (4 pg / mL) at 4°C overnight. The plate was then blocked with a blocker (5% milk in PBS (MPBS)) at 37°C for 1 hour. After washing, the diluted serum was added to each well and incubated at 37°C for 1 hour. After washing the plate three times with PBS-Tween (0.05%), an HRP-conjugated anti-human Fc antibody was added and incubated for another 1 hour. The plate was washed, and TMB was added for color development. Concentration was calculated by a standard curve.

[0161] Neutralization Test

[0162] To test if the CD4-D1 variants can inhibit HIV-1 replication, a TZM-bl HIV-1 phenotyping assay was run. The TZM-bl cells were used for quantitative analysis of HIV replication. TZM-bl cells were generated from HeLa cells that stably expressed large amounts of CD4 and CCR5. These cells have separately integrated copies of the luciferase and 13-galactosidase genes under the control of HIV-1 promoter. TZM-bl cells were plated at 10,000 cells per well and cultured overnight. The cells were then treated with serial dilutions of CD4-D1 variants, and infected with a dilution of infectious HIV-1 virus normalized to an output of 140000 relative light units (RLU) as determined by endpoint dilution. After a 48- hour incubation at 37°C, cells were lysed, and luminescence was measured in RLU using a commercially available luciferase detection system. The 50% in vitro concentration (IC50) was determined by calculating the concentration of antibodies needed to inhibit 50% of HIV- 1 replication. A batch control virus was run with each experimental setup.

[0163] Results

[0164] SPR binding kinetic with gpl-10

[0165] The SPR results demonstrated that 1 A2-4 had the highest affinity with gp!40, while the affinity of S60E was slightly lower than mD1.22 (Table 2 and Figure 12). Table 2. Summary of the SPR analysis.

[0166] Binding kinetic with human MHC II

[0167] The BLItz assay showed that both S60E and 1A2-4 mutations lost their ability to bind to MHC II (Figure 13).

[0168] SEC test showed high purity and monomeric without high molecule weight species The SEC assay showed that DI and the mutants are monomeric.

[0169] S60E had the highest thermal melting temperature (Tm) level

[0170] The thermostability test showed that S60E had the highest Tm level, followed by 1A2-4 and mD1.22 (Table 3 and Figure 15).

[0171] Table 3. Summary of thermostability test results

[0172] S60E-Fc was resistant to aggregation

[0173] The DLS assay showed that S60E-Fc was highly stable and aggregation resistant after incubation at 37 °C for 14 days, whereas mD1.22-Fc and lA2-4-Fc showed aggregation (Figure 16).

[0174] Flow cytometry assay of IA2-4, S60E, and mDI.22 Cell binding test with MHCII positive and negative cells (Figures 17A-17B)

[0175] Cell binding test with human and mouse PBMCs (Figures 17C-17D)

[0176] Pharmacokinetic assay of 1A2-4, S60E, and mD1.22 S60E showed longer half-life than 1A2-4 and the clinical antibody 10-1074 (Figure

[0177] 18).

[0178] Neutralization effects of 1A2-4, S60E, and mD1.22 on different HIV virus type

[0179] S60E showed similar IC50 value to mD1.22 and 1A2-4, while retaining broad neutralization effects (Table 4 and Figures 19A-19B).

[0180] Table 4. IC5o levels of 1A2-4, S60E, and mD1.22

[0181] Pharmacokinetic assay of bispecific antibodies

[0182] Bispecific antibodies were designed as shown in Figure 20. The pharmacokinetic (PK) assay showed that S60E-Fc had the longest half-life. The bispecific antibodies showed shorter half-life than S60E-Fc alone (Figure 21 and Table 5).

[0183] Table 5. Summary of PK results

[0184] Neutralization assay of bispecific antibodies

[0185] The results showed that the bispecific antibodies S60E-1074 and 1A2 -4-1074 had higher neutralization potential (Figure 22 and Table 6).

[0186] Table 6. Summary of neutralization test results

[0187] Example 5: Exemplary Fc Polypeptide Sequences DKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNW YVDGVEVHNAI<TI<PREEQYNSTYRVVSVLTVLHQDWLNGI<EYI<CI<VSNI<ALPAPI EKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPEN NYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSP GK (SEQ ID NO:47)

[0188] Example 6: Exemplary Molecules that can Bind a gp!20 Glycopeptide m36.4 single-domain antibody:

[0189] QVQLVQSGGGLVQPGGSLRLSCAASAFDFSDYEMSWVREAPGKGLEWIGEINDSGN TIYNPSLKSRVTISRDNSKNTLYLQMNTLRAEDTAIYYCAIYGGNSGGEYWGQGTLV TVSS (SEQ ID NO:48)

[0190] 1074 antibody heavy chain (HC):

[0191] QVQLQESGPGLVKPSETLSVTCSVSGDSMNNYYWTWIRQSPGKGLEWIGYISDRESA

[0192] TYNPSLNSRVVISRDTSKNQLSLKLNSVTPADTAVYYCATARRGQRIYGVVSFGEFF YYYSMDVWGKGTTVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVT

[0193] VSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVD KRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHED

[0194] PEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVS NKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWE

[0195] SNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQ KSLSLSPGK (SEQ ID NO:49)

[0196] 1074 antibody light chain (LC):

[0197] SYVRPLSVALGETARISCGRQALGSRAVQWYQHRPGQAPILLIYNNQDRPSGIPERFS GTPDINFGTRATLTISGVEAGDEADYYCHMWDSRSGFSWSFGGATRLTVLGQPKAA P S VTLFPP S SEELQ ANKATLVCLISDF YPGAVT VAWKAD S SP VKAGVETTTP SKQ SN

[0198] NKYAASSYLSLTPEQWKSHRSYSCQVTHEGSTVEKTVAPTECS (SEQ ID NO:50)

[0199] LSEV HC:

[0200] KKVVYGKKGDTVELTCTASQKKNIQFHWKNSNQIKILGNQGSFLTKGPSKLNDRVD

[0201] SRRSLWDQGNFPLIIKNLKPEDSDTYICEVEDQKEEVQLVVVGDKTHTASTKGPSVF PLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYELV

[0202] SVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVF LFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYN STYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPS RDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLT VDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPGKDKTHTKKVVYGKKGDTVEL TCTASQKKNIQFHWKNSNQIKILGNQGSFLTKGPSKLNDRVDSRRSLWDQGNFPLIIK NLKPEDSDTYICEVEDQKEEVQLVVVG (SEQ ID N0 51)

[0203] LSEV LC:

[0204] QVQLVQSGGGLVQPGGSLRLSCAASAFDFSDYEMSWVREAPGKGLEWIGEINDSGN TIYNPSLKSRVTISRDNSKNTLYLQMNTLRAEDTAIYYCAIYGGNSGGEYWGQGTLV TVSSDKTHTRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQ SGNSQESVTEQDSKDSTYSLLSSLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRG EC (SEQ ID NO:52)

[0205] LSEVdelta HC:

[0206] KKVVYGKKGDTVELTCTASQKKNIQFHWKNSNQIKILGNQGSFLTKGPSKLNDRVD SRRSLWDQGNFPLIIKNLKPEDSDTYICEVEDQKEEVQLVVVGDKTHTASTKGPSVF

[0207] PLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYELV SVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVF LFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYN STYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPS RDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLT VDKSRWQQGNVFSCSVLHEALHSHYTQKSLSLSPGK (SEQ ID NO:53)

[0208] Example 7: Neutralization effects of CD4-D1 variants

[0209] The results in this Example re-present and expand on at least some of the results provided in other Examples.

[0210] Bispecific antibodies consisting of a CD4-D1 variants (1A2-4 or S60E) and the bnAb 10-1074 exhibited more potent neutralization of HIV subtype C (CH0505) and B (CH077) than the tetravalent bispecific Ig-like compound LSEVh-LS made of CD4-D1 and the domain antibody m36.4 targeting HIV Env coreceptor binding site (Figure 23).

[0211] Bispecific antibodies consisting of a CD4-D1 variant (1A2-4 or S60E) exhibited neutralization of HIV subtype B (2626 and 2864). lA2-4-LSEVh-LS was more potent for T / F 2626 than a 1074-based bispecific antibody, while neutralization against T / F 2864 was similar (Figure 24). Bispecific antibodies consisting of a CD4-D1 variant 1A2-4 or S60E and a bnAb 10- 1074 were more potent than other CD4-D1 variants for neutralization of pTROl 1 Env pseudotyped HIV virus. On pBJOX2000 env pseudoviruses, however, neutralization was similar across different CD4-D1 variants (Figure 25).

[0212] Bispecific antibodies consisting of a CD4-D1 variant 1 A2-4 or S60E and a bnAb 10- 1074 were more potent than other CD4-D1 variants for neutralization of subtype C (pCEl 176 Env) and subtype AC (p246F3 env) HIV pseudoviruses. The particular CD4-D1 variant used in the bispecific antibody did not affect neutralization of pBJOX2000 env pseudoviruses (Figure 26).

[0213] Bispecific antibodies consisting of a CD4-D1 variant 1A2-4 or S60E and a bnAb 10- 1074 were more potent than other CD4-D1 variants for neutralization of subtype CRF07 BC (pCHl 19 Env) and subtype G (pX1632 env). The particular CD4-D1 variant used in the bispecific antibody did not affect neutralization of pX1632 env pseudoviruses (Figure 27).

[0214] Example 8: Treating HIV infection

[0215] A human identified as having an HIV (e.g., an HIV-1) infection is administered one or more CD4-D1 polypeptides set forth in any one of SEQ ID NOs:4-39. The administered CD4-D1 polypeptides can slow or delay an HIV infection of cells within the human.

[0216] Example 9: Treating HIV infection

[0217] A human identified as having an HIV (e.g., an HIV-1) infection is administered nucleic acid encoding one or more CD4-D1 polypeptides set forth in any one of SEQ ID NOs:4-39. The administered nucleic acid can encode the CD4-D1 polypeptide(s) within the human and slow or delay an HIV infection of cells within of the human.

[0218] Example 10: Prophylactic treatment of an HIV infection

[0219] A human identified as being at risk of developing an HIV (e.g., an HIV-1) infection is administered one or more CD4-D1 polypeptides set forth in any one of SEQ ID NOs:4-39. The administered CD4-D1 polypeptides reduce the human’s level of susceptibility to developing an HIV infection of cells within the human. Example 11: Prophylactic treatment of an HIV infection

[0220] A human identified as being at risk of developing an HIV (e.g., an HIV-1) infection is administered nucleic acid encoding one or more CD4-D1 polypeptides set forth in any one of SEQ ID NOs:4-39. The administered nucleic acid can encode the CD4-D1 polypeptide(s) within the human and reduce the human’s level of susceptibility to developing an HIV infection of cells within the human.

[0221] Example 12: Exemplary Embodiments

[0222] Embodiment 1. A CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

[0223] Embodiment 2. A nucleic acid comprising a nucleic acid sequence encoding a CD4-D1 polypeptide of embodiment 1.

[0224] Embodiment 3. A method for treating a mammal having an HIV infection, wherein said method comprises administering a CD4-D1 polypeptide to said mammal, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

[0225] Embodiment 4. The method of embodiment 3, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

[0226] Embodiment 5. The method of embodiment 3, wherein said CD4-D1 polypeptide consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

[0227] Embodiment 6. The method of any one of embodiments 3-4, wherein said mammal is a human.

[0228] Embodiment 7. The method of any one of embodiments 3-6, wherein said HIV is HIV- 1. Embodiment 8. The method of any one of embodiments 3-7, wherein said CD4-D1 polypeptide is effective to reduce a symptom of said HIV infection.

[0229] Embodiment 9. The method of embodiment 8, wherein said symptom is selected from the group consisting of fever, headache, muscle aches, joint pain, rash, sore throat, painful mouth sores, swollen lymph glands, swollen lymph nodes on the neck, diarrhea, weight loss, cough, night sweats, fatigue, vomiting, vision loss, and memory loss.

[0230] Embodiment 10. The method of any one of embodiments 3-9, wherein said CD4-D1 polypeptide is effective to reduce a number of HIV particles present in one or more cells within said mammal.

[0231] Embodiment 11. The method of any one of embodiments 3-10, wherein said CD4-D1 polypeptide is effective to reduce a viral load of said mammal.

[0232] Embodiment 12. The method of any one of embodiments 3-11, wherein said method is effective to delay or prevent progression of said HIV infection to AIDS within said mammal.

[0233] Embodiment 13. The method of any one of embodiments 3-11, wherein said mammal has AIDS.

[0234] Embodiment 14. The method of any one of embodiments 3-13, wherein said method comprises identifying said mammal as being in need of said CD4-D1 polypeptide prior to said administering step.

[0235] Embodiment 15. A method for treating a mammal having an HIV infection, wherein said method comprises administering nucleic acid encoding a CD4-D1 polypeptide to said mammal, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in any one of SEQ ID NOs:4-39. Embodiment 16. The method of embodiment 15, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

[0236] Embodiment 17. The method of embodiment 15, wherein said CD4-D1 polypeptide consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

[0237] Embodiment 18. The method of any one of embodiments 15-17, wherein said mammal is a human.

[0238] Embodiment 19. The method of any one of embodiments 15-18, wherein said HIV is

[0239] HIV-1.

[0240] Embodiment 20. The method of any one of embodiments 15-19, wherein said CD4-D1 polypeptide is effective to reduce a symptom of said HIV infection.

[0241] Embodiment 21. The method of embodiment 20, wherein said symptom is selected from the group consisting of fever, headache, muscle aches, joint pain, rash, sore throat, painful mouth sores, swollen lymph glands, swollen lymph nodes on the neck, diarrhea, weight loss, cough, night sweats, fatigue, vomiting, vision loss, and memory loss.

[0242] Embodiment 22. The method of any one of embodiments 15-21, wherein said CD4-D1 polypeptide is effective to reduce a number of HIV particles present in one or more cells within said mammal.

[0243] Embodiment 23. The method of any one of embodiments 15-22, wherein said CD4-D1 polypeptide is effective to reduce a viral load of said mammal.

[0244] Embodiment 24. The method of any one of embodiments 15-23, wherein said method is effective to delay or prevent progression of said HIV infection to AIDS within said mammal. Embodiment 25. The method of any one of embodiments 15-23, wherein said mammal has AIDS.

[0245] Embodiment 26. The method of any one of embodiments 15-25, wherein said method comprises identifying said mammal as being in need of said CD4-D1 polypeptide prior to said administering step.

[0246] Embodiment 27. The method of any one of embodiments 15-26, wherein said nucleic acid is in the form of a vector.

[0247] Embodiment 28. The method of embodiment 27, wherein said vector is an AAV vector.

[0248] Embodiment 29. The method of embodiment 27, wherein said vector is an expression plasmid.

[0249] Embodiment 30. A method for treating a mammal at risk of developing an HIV infection, wherein said method comprises administering a CD4-D1 polypeptide to said mammal, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

[0250] Embodiment 31. The method of embodiment 30, wherein said CD4-D 1 polypeptide comprises the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

[0251] Embodiment 32. The method of embodiment 30, wherein said CD4-D1 polypeptide consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

[0252] Embodiment 33. The method of any one of embodiments 30-32, wherein said mammal is a human.

[0253] Embodiment 34. The method of any one of embodiments 30-33, wherein said HIV is

[0254] HIV-1. Embodiment 35. The method of any one of embodiments 30-34, wherein said method is effective to delay or prevent development of an HIV infection within said mammal.

[0255] Embodiment 36. The method of any one of embodiments 30-35, wherein said method comprises identifying said mammal as being in need of said CD4-D1 polypeptide prior to said administering step.

[0256] Embodiment 37. A method for treating a mammal at risk of developing an HIV infection, wherein said method comprises administering nucleic acid encoding a CD4-D1 polypeptide to said mammal, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

[0257] Embodiment 38. The method of embodiment 37, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

[0258] Embodiment 39. The method of embodiment 37, wherein said CD4-D1 polypeptide consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

[0259] Embodiment 40. The method of any one of embodiments 37-39, wherein said mammal is a human.

[0260] Embodiment 41. The method of any one of embodiments 37-40, wherein said HIV is

[0261] HIV-1.

[0262] Embodiment 42. The method of any one of embodiments 37-41, wherein said method is effective to delay or prevent development of an HIV infection within said mammal.

[0263] Embodiment 43. The method of any one of embodiments 37-42, wherein said method comprises identifying said mammal as being in need of said CD4-D1 polypeptide prior to said administering step. Embodiment 44. The method of any one of embodiments 37-43, wherein said nucleic acid is in the form of a vector.

[0264] Embodiment 45. The method of embodiment 44, wherein said vector is an AAV vector.

[0265] Embodiment 46. The method of embodiment 44, wherein said vector is an expression plasmid.

[0266] Embodiment 47. A method for treating a mammal having an HIV infection, wherein said method comprises administering a CD4-D1 polypeptide to said mammal, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in SEQ ID NO:40, wherein each X is independently selected from the group consisting of A, S, T, E, D, Q, N, and H.

[0267] Embodiment 48. The method of embodiment 47, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in any one of SEQ ID NOs:9-39.

[0268] Embodiment 49. The method of embodiment 47, wherein said CD4-D1 polypeptide consists of the amino acid sequence set forth in any one of SEQ ID NOs:9-39.

[0269] Embodiment 50. The method of any one of embodiments 47-49, wherein said mammal is a human.

[0270] Embodiment 51. The method of any one of embodiments 47-50, wherein said HIV is

[0271] HIV-1.

[0272] Embodiment 52. The method of any one of embodiments 47-51, wherein said CD4-D1 polypeptide is effective to reduce a symptom of said HIV infection.

[0273] Embodiment 53. The method of embodiment 52, wherein said symptom is selected from the group consisting of fever, headache, muscle aches, joint pain, rash, sore throat, painful mouth sores, swollen lymph glands, swollen lymph nodes on the neck, diarrhea, weight loss, cough, night sweats, fatigue, vomiting, vision loss, and memory loss.

[0274] Embodiment 54. The method of any one of embodiments 47-53, wherein said CD4-D1 polypeptide is effective to reduce a number of HIV particles present in one or more cells within said mammal.

[0275] Embodiment 55. The method of any one of embodiments 47-54, wherein said CD4-D1 polypeptide is effective to reduce a viral load of said mammal.

[0276] Embodiment 56. The method of any one of embodiments 47-55, wherein said method is effective to delay or prevent progression of said HIV infection to AIDS within said mammal.

[0277] Embodiment 57. The method of any one of embodiments 47-55, wherein said mammal has AIDS.

[0278] Embodiment 58. The method of any one of embodiments 47-57, wherein said method comprises identifying said mammal as being in need of said CD4-D1 polypeptide prior to said administering step.

[0279] Embodiment 59. A method for treating a mammal having an HIV infection, wherein said method comprises administering nucleic acid encoding a CD4-D1 polypeptide to said mammal, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in SEQ ID NO:40, wherein each X is independently selected from the group consisting of A, S, T, E, D, Q, N, and H.

[0280] Embodiment 60. The method of embodiment 59, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in any one of SEQ ID NOs:9-39.

[0281] Embodiment 61. The method of embodiment 59, wherein said CD4-D1 polypeptide consists of the amino acid sequence set forth in any one of SEQ ID NOs:9-39. Embodiment 62. The method of any one of embodiments 59-61, wherein said mammal is a human.

[0282] Embodiment 63. The method of any one of embodiments 59-62, wherein said HIV is

[0283] HIV-1.

[0284] Embodiment 64. The method of any one of embodiments 59-63, wherein said CD4-D1 polypeptide is effective to reduce a symptom of said HIV infection.

[0285] Embodiment 65. The method of embodiment 64, wherein said symptom is selected from the group consisting of fever, headache, muscle aches, joint pain, rash, sore throat, painful mouth sores, swollen lymph glands, swollen lymph nodes on the neck, diarrhea, weight loss, cough, night sweats, fatigue, vomiting, vision loss, and memory loss.

[0286] Embodiment 66. The method of any one of embodiments 59-65, wherein said CD4-D1 polypeptide is effective to reduce a number of HIV particles present in one or more cells within said mammal.

[0287] Embodiment 67. The method of any one of embodiments 59-66, wherein said CD4-D1 polypeptide is effective to reduce a viral load of said mammal.

[0288] Embodiment 68. The method of any one of embodiments 59-67, wherein said method is effective to delay or prevent progression of said HIV infection to AIDS within said mammal.

[0289] Embodiment 69. The method of any one of embodiments 59-67, wherein said mammal has AIDS.

[0290] Embodiment 70. The method of any one of embodiments 59-69, wherein said method comprises identifying said mammal as being in need of said CD4-D1 polypeptide prior to said administering step. Embodiment 71. The method of any one of embodiments 59-70, wherein said nucleic acid is in the form of a vector.

[0291] Embodiment 72. The method of embodiment 71, wherein said vector is an AAV vector.

[0292] Embodiment 73. The method of embodiment 71, wherein said vector is an expression plasmid.

[0293] Embodiment 74. A method for treating a mammal at risk of developing an HIV infection, wherein said method comprises administering a CD4-D1 polypeptide to said mammal, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in SEQ ID NO:40, wherein each X is independently selected from the group consisting of A, S, T, E, D, Q, N, and H.

[0294] Embodiment 75. The method of embodiment 74, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in any one of SEQ ID NOs:9-39.

[0295] Embodiment 76. The method of embodiment 74, wherein said CD4-D1 polypeptide consists of the amino acid sequence set forth in any one of SEQ ID NOs:9-39.

[0296] Embodiment 77. The method of any one of embodiments 74-76, wherein said mammal is a human.

[0297] Embodiment 78. The method of any one of embodiments 74-77, wherein said HIV is

[0298] HIV-1.

[0299] Embodiment 79. The method of any one of embodiments 74-78, wherein said method is effective to delay or prevent development of an HIV infection within said mammal. Embodiment 80. The method of any one of embodiments 74-79, wherein said method comprises identifying said mammal as being in need of said CD4-D1 polypeptide prior to said administering step.

[0300] Embodiment 81. A method for treating a mammal at risk of developing an HIV infection, wherein said method comprises administering nucleic acid encoding a CD4-D1 polypeptide to said mammal, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in SEQ ID NO:40, wherein each X is independently selected from the group consisting of A, S, T, E, D, Q, N, and H.

[0301] Embodiment 82. The method of embodiment 81, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in any one of SEQ ID NOs:9-39.

[0302] Embodiment 83. The method of embodiment 81, wherein said CD4-D1 polypeptide consists of the amino acid sequence set forth in any one of SEQ ID NOs:9-39.

[0303] Embodiment 84. The method of any one of embodiments 81-83, wherein said mammal is a human.

[0304] Embodiment 85. The method of any one of embodiments 81-84, wherein said HIV is

[0305] HIV-1.

[0306] Embodiment 86. The method of any one of embodiments 81-85, wherein said method is effective to delay or prevent development of an HIV infection within said mammal.

[0307] Embodiment 87. The method of any one of embodiments 81-86, wherein said method comprises identifying said mammal as being in need of said CD4-D1 polypeptide prior to said administering step.

[0308] Embodiment 88. The method of any one of embodiments 81-87, wherein said nucleic acid is in the form of a vector. Embodiment 89. The method of embodiment 88, wherein said vector is an AAV vector.

[0309] Embodiment 90. The method of embodiment 88, wherein said vector is an expression plasmid.

[0310] Embodiment 91. The use of a composition comprising a CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-39 to treat a mammal having an HIV infection.

[0311] Embodiment 92. A CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-39 for use in the preparation of a medicament to treat a mammal having an HIV infection.

[0312] Embodiment 93. A CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-39 for use in the treatment of an HIV infection.

[0313] Embodiment 94. The use of a composition comprising nucleic acid encoding a CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-39 to treat a mammal having an HIV infection.

[0314] Embodiment 95. Nucleic acid encoding a CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-39 for use in the preparation of a medicament to treat a mammal having an HIV infection.

[0315] Embodiment 96. Nucleic acid encoding a CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-39 for use in the treatment of an HIV infection.

[0316] Embodiment 97. A method for treating a mammal having an HIV infection, wherein said method comprises administering, to said mammal, a fusion polypeptide or a nucleic acid construct encoding said fusion polypeptide to said mammal, said fusion polypeptide comprising a first polypeptide and a second polypeptide, wherein said first polypeptide is a first CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

[0317] Embodiment 98. The method of clam 97, wherein said mammal is a human.

[0318] Embodiment 99. The method of any one of embodiments 97-98, wherein said HIV is

[0319] HIV-1.

[0320] Embodiment 100. The method of any one of embodiments 97-99, wherein said second polypeptide is a second CD4-D1 polypeptide different from said first CD4-D1 polypeptide.

[0321] Embodiment 101. The method of embodiment 100, wherein both of said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently comprise the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

[0322] Embodiment 102. The method of embodiment 100, wherein both of said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently comprise the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

[0323] Embodiment 103. The method of embodiment 100, wherein said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently consist of the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

[0324] Embodiment 104. The method of any one of embodiments 97-99, wherein said second polypeptide is an Fc polypeptide.

[0325] Embodiment 105. The method of embodiment 104, wherein said Fc polypeptide comprises the amino acid sequence set forth in SEQ ID NO:47. Embodiment 106. The method of any one of embodiments 97-105, wherein said fusion polypeptide further comprises at least a portion of an m36.4 antibody.

[0326] Embodiment 107. The method of embodiment 106, wherein said portion of said m36.4 antibody comprises the amino acid sequence set forth in SEQ ID NO:48.

[0327] Embodiment 108. The method of any one of embodiments 97-105, wherein said fusion polypeptide further comprises at least a portion of an 10-1074 antibody.

[0328] Embodiment 109. The method of embodiment 108, wherein said portion of said 10-1074 antibody comprises the amino acid sequence set forth in SEQ ID NO:49 and / or SEQ ID NO:50.

[0329] Embodiment 110. The method of any one of embodiments 97-109, wherein said fusion polypeptide is effective to reduce a symptom of said HIV infection.

[0330] Embodiment 111. The method of embodiment 110, wherein said symptom is selected from the group consisting of fever, headache, muscle aches, joint pain, rash, sore throat, painful mouth sores, swollen lymph glands, swollen lymph nodes on the neck, diarrhea, weight loss, cough, night sweats, fatigue, vomiting, vision loss, and memory loss.

[0331] Embodiment 112. The method of any one of embodiments 97-109, wherein said fusion polypeptide is effective to reduce a number of HIV particles present in one or more cells within said mammal.

[0332] Embodiment 113. The method of any one of embodiments 97-109, wherein said fusion polypeptide is effective to reduce a viral load of said mammal.

[0333] Embodiment 114. The method of any one of embodiments 97-113, wherein said method is effective to delay or prevent progression of said HIV infection to AIDS within said mammal. Embodiment 115. The method of any one of embodiments 97-113, wherein said mammal has AIDS.

[0334] Embodiment 116. The method of any one of embodiments 97-115, wherein said method comprises identifying said mammal as being in need of said fusion polypeptide prior to said administering step.

[0335] Embodiment 117. A method for treating a mammal at risk of developing an HIV infection, wherein said method comprises administering, to said mammal, a fusion polypeptide or a nucleic acid construct encoding said fusion polypeptide to said mammal, said fusion polypeptide comprising a first polypeptide and a second polypeptide, wherein said first polypeptide is a first CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

[0336] Embodiment 118. The method of clam 117, wherein said mammal is a human.

[0337] Embodiment 119. The method of any one of embodiments 117-118, wherein said HIV is

[0338] HIV-1.

[0339] Embodiment 120. The method of any one of embodiments 117-119, wherein said second polypeptide is a second CD4-D1 polypeptide different from said first CD4-D1 polypeptide.

[0340] Embodiment 121. The method of embodiment 120, wherein both of said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently comprise the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

[0341] Embodiment 122. The method of embodiment 120, wherein both of said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently comprise the amino acid sequence set forth in any one of SEQ ID NOs:4-8. Embodiment 123. The method of embodiment 120, wherein said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently consist of the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

[0342] Embodiment 124. The method of any one of embodiments 117-119, wherein said second polypeptide is an Fc polypeptide.

[0343] Embodiment 125. The method of embodiment 124, wherein said Fc polypeptide comprises the amino acid sequence set forth in SEQ ID NO:47.

[0344] Embodiment 126. The method of any one of embodiments 117-125, wherein said fusion polypeptide further comprises at least a portion of an m36.4 antibody.

[0345] Embodiment 127. The method of embodiment 126, wherein said portion of said m36.4 antibody comprises the amino acid sequence set forth in SEQ ID NO:48.

[0346] Embodiment 128. The method of any one of embodiments 117-125, wherein said fusion polypeptide further comprises at least a portion of an 10-1074 antibody.

[0347] Embodiment 129. The method of embodiment 128, wherein said portion of said 10-1074 antibody comprises the amino acid sequence set forth in SEQ ID NO:49 and / or SEQ ID NO:50.

[0348] Embodiment 130. The method of any one of embodiments 117-129, wherein said method is effective to delay or prevent development of an HIV infection within said mammal.

[0349] Embodiment 131. The method of any one of embodiments 117-130, wherein said method comprises identifying said mammal as being in need of said fusion polypeptide prior to said administering step. Embodiment 132. A method for treating a mammal having an HIV infection, wherein said method comprises administering, to said mammal, a fusion polypeptide or a nucleic acid construct encoding said fusion polypeptide, said fusion polypeptide comprising a first polypeptide and a second polypeptide, wherein said first polypeptide is a first CD4-D1 polypeptide comprising the amino acid sequence set forth in SEQ ID NO:40, wherein each X is independently selected from the group consisting of A, S, T, E, D, Q, N, and H.

[0350] Embodiment 133. The method of clam 132, wherein said mammal is a human.

[0351] Embodiment 134. The method of any one of embodiments 132-133, wherein said HIV is

[0352] HIV-1.

[0353] Embodiment 135. The method of any one of embodiments 132-134, wherein said second polypeptide is a second CD4-D1 polypeptide different from said first CD4-D1 polypeptide.

[0354] Embodiment 136. The method of embodiment 135, wherein both of said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently comprise the amino acid sequence set forth in any one of SEQ ID NOs:9-39.

[0355] Embodiment 137. The method of embodiment 135, wherein said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently consist of the amino acid sequence set forth in any one of SEQ ID NOs:9-39.

[0356] Embodiment 138. The method of any one of embodiments 132-134, wherein said second polypeptide is an Fc polypeptide.

[0357] Embodiment 139. The method of embodiment 138, wherein said Fc polypeptide comprises the amino acid sequence set forth in SEQ ID NO:47.

[0358] Embodiment 140. The method of any one of embodiments 132-139, wherein said fusion polypeptide further comprises at least a portion of an m36.4 antibody. Embodiment 141. The method of embodiment 140, wherein said portion of said m36.4 antibody comprises the amino acid sequence set forth in SEQ ID NO:48.

[0359] Embodiment 142. The method of any one of embodiments 132-139, wherein said fusion polypeptide further comprises at least a portion of an 10-1074 antibody.

[0360] Embodiment 143. The method of embodiment 142, wherein said portion of said 10-1074 antibody comprises the amino acid sequence set forth in SEQ ID NO:49 and / or SEQ ID NO:50.

[0361] Embodiment 144. The method of any one of embodiments 132-143, wherein said fusion polypeptide is effective to reduce a symptom of said HIV infection.

[0362] Embodiment 145. The method of embodiment 144, wherein said symptom is selected from the group consisting of fever, headache, muscle aches, joint pain, rash, sore throat, painful mouth sores, swollen lymph glands, swollen lymph nodes on the neck, diarrhea, weight loss, cough, night sweats, fatigue, vomiting, vision loss, and memory loss.

[0363] Embodiment 146. The method of any one of embodiments 132-143, wherein said fusion polypeptide is effective to reduce a number of HIV particles present in one or more cells within said mammal.

[0364] Embodiment 147. The method of any one of embodiments 132-143, wherein said fusion polypeptide is effective to reduce a viral load of said mammal.

[0365] Embodiment 148. The method of any one of embodiments 132-147, wherein said method is effective to delay or prevent progression of said HIV infection to AIDS within said mammal. Embodiment 149. The method of any one of embodiments 132-147, wherein said mammal has AIDS.

[0366] Embodiment 150. The method of any one of embodiments 132-149, wherein said method comprises identifying said mammal as being in need of said fusion polypeptide prior to said administering step.

[0367] Embodiment 151. A method for treating a mammal at risk of developing an HIV infection, wherein said method comprises administering, to said mammal, a fusion polypeptide or a nucleic acid construct encoding said fusion polypeptide, said fusion polypeptide comprising a first polypeptide and a second polypeptide, wherein said first polypeptide is a first CD4-D1 polypeptide comprising the amino acid sequence set forth in SEQ ID NO:40, wherein each X is independently selected from the group consisting of A, S, T, E, D, Q, N, and H.

[0368] Embodiment 152. The method of clam 151, wherein said mammal is a human.

[0369] Embodiment 153. The method of any one of embodiments 151-152, wherein said HIV is

[0370] HIV-1.

[0371] Embodiment 154. The method of any one of embodiments 151-153, wherein said second polypeptide is a second CD4-D1 polypeptide different from said first CD4-D1 polypeptide.

[0372] Embodiment 155. The method of embodiment 154, wherein both of said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently comprise the amino acid sequence set forth in any one of SEQ ID NOs:9-39.

[0373] Embodiment 156. The method of embodiment 154, wherein said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently consist of the amino acid sequence set forth in any one of SEQ ID NOs:9-39. Embodiment 157. The method of any one of embodiments 151-153, wherein said second polypeptide is an Fc polypeptide.

[0374] Embodiment 158. The method of embodiment 157, wherein said Fc polypeptide comprises the amino acid sequence set forth in SEQ ID NO:47.

[0375] Embodiment 159. The method of any one of embodiments 151-158, wherein said fusion polypeptide further comprises at least a portion of an m36.4 antibody.

[0376] Embodiment 160. The method of embodiment 106, wherein said portion of said m36.4 antibody comprises the amino acid sequence set forth in SEQ ID NO:48.

[0377] Embodiment 161. The method of any one of embodiments 151-158, wherein said fusion polypeptide further comprises at least a portion of an 10-1074 antibody.

[0378] Embodiment 162. The method of embodiment 161, wherein said portion of said 10-1074 antibody comprises the amino acid sequence set forth in SEQ ID NO:49 and / or SEQ ID NO:50.

[0379] Embodiment 163. The method of any one of embodiments 151-162, wherein said method is effective to delay or prevent development of an HIV infection within said mammal.

[0380] Embodiment 164. The method of any one of embodiments 151-163, wherein said method comprises identifying said mammal as being in need of said fusion polypeptide prior to said administering step.

[0381] Embodiment 165. A fusion polypeptide comprising a first polypeptide and a second polypeptide for use in the preparation of a medicament to treat a mammal having an HIV infection, wherein said first polypeptide is a first CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-39. Embodiment 166. The fusion polypeptide of clam 165, wherein said mammal is a human.

[0382] Embodiment 167. The fusion polypeptide of any one of embodiments 165-166, wherein said HIV is HIV- 1

[0383] Embodiment 168. The fusion polypeptide of any one of embodiments 165-167, wherein said second polypeptide is a second CD4-D1 polypeptide different from said first CD4-D1 polypeptide.

[0384] Embodiment 169. The fusion polypeptide of embodiment 168, wherein both of said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently comprise the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

[0385] Embodiment 170. The fusion polypeptide of embodiment 168, wherein both of said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently comprise the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

[0386] Embodiment 171. The fusion polypeptide of embodiment 168, wherein said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently consist of the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

[0387] Embodiment 172. The fusion polypeptide of any one of embodiments 165-167, wherein said second polypeptide is an Fc polypeptide.

[0388] Embodiment 173. The fusion polypeptide of embodiment 172, wherein said Fc polypeptide comprises the amino acid sequence set forth in SEQ ID NO:47.

[0389] Embodiment 174. The fusion polypeptide of any one of embodiments 165-173, wherein said fusion polypeptide further comprises at least a portion of an m36.4 antibody. Embodiment 175. The fusion polypeptide of embodiment 174, wherein said portion of said m36.4 antibody comprises the amino acid sequence set forth in SEQ ID NO:48.

[0390] Embodiment 176. The fusion polypeptide of any one of embodiments 165-173, wherein said fusion polypeptide further comprises at least a portion of an 10-1074 antibody.

[0391] Embodiment 177. The fusion polypeptide of embodiment 176, wherein said portion of said 10-1074 antibody comprises the amino acid sequence set forth in SEQ ID NO:49 and / or SEQ ID NO:50.

[0392] Embodiment 147. A fusion polypeptide comprising a first polypeptide and a second polypeptide for use in the treatment of an HIV infection, wherein said first polypeptide is a first CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

[0393] Embodiment 148. The fusion polypeptide of embodiment 147, wherein said HIV is HIV- 1.

[0394] Embodiment 149. The fusion polypeptide of any one of embodiments 147-148, wherein said second polypeptide is a second CD4-D1 polypeptide different from said first CD4-D1 polypeptide.

[0395] Embodiment 150. The fusion polypeptide of embodiment 149, wherein both of said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently comprise the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

[0396] Embodiment 151. The fusion polypeptide of embodiment 149, wherein both of said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently comprise the amino acid sequence set forth in any one of SEQ ID NOs:4-8. Embodiment 152. The fusion polypeptide of embodiment 149, wherein said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently consist of the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

[0397] Embodiment 153. The fusion polypeptide of any one of embodiments 147-148, wherein said second polypeptide is an Fc polypeptide.

[0398] Embodiment 154. The fusion polypeptide of embodiment 153, wherein said Fc polypeptide comprises the amino acid sequence set forth in SEQ ID NO:47.

[0399] Embodiment 155. The fusion polypeptide of any one of embodiments 147-154, wherein said fusion polypeptide further comprises at least a portion of an m36.4 antibody.

[0400] Embodiment 156. The fusion polypeptide of embodiment 155, wherein said portion of said m36.4 antibody comprises the amino acid sequence set forth in SEQ ID NO:48.

[0401] Embodiment 157. The fusion polypeptide of any one of embodiments 147-154, wherein said fusion polypeptide further comprises at least a portion of an 10-1074 antibody.

[0402] Embodiment 158. The fusion polypeptide of embodiment 157, wherein said portion of said 10-1074 antibody comprises the amino acid sequence set forth in SEQ ID NO:49 and / or SEQ ID NO:50.

[0403] Embodiment 159. The use of a composition comprising a fusion polypeptide comprising a first polypeptide and a second polypeptide to treat a mammal having an HIV infection, wherein said first polypeptide is a first CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

[0404] Embodiment 160. The use of clam 159, wherein said mammal is a human. Embodiment 161. The use of any one of embodiments 159-160, wherein said HIV is HIV-1.

[0405] Embodiment 162. The use of any one of embodiments 159-161, wherein said second polypeptide is a second CD4-D1 polypeptide different from said first CD4-D1 polypeptide.

[0406] Embodiment 163. The use of embodiment 162, wherein both of said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently comprise the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

[0407] Embodiment 164. The use of embodiment 162, wherein both of said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently comprise the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

[0408] Embodiment 165. The use of embodiment 162, wherein said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently consist of the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

[0409] Embodiment 166. The use of any one of embodiments 159-161, wherein said second polypeptide is an Fc polypeptide.

[0410] Embodiment 167. The use of embodiment 166, wherein said Fc polypeptide comprises the amino acid sequence set forth in SEQ ID NO:47.

[0411] Embodiment 168. The use of any one of embodiments 159-167, wherein said fusion polypeptide further comprises at least a portion of an m36.4 antibody.

[0412] Embodiment 169. The use of embodiment 168, wherein said portion of said m36.4 antibody comprises the amino acid sequence set forth in SEQ ID NO:48. Embodiment 170. The use of any one of embodiments 159-167, wherein said fusion polypeptide further comprises at least a portion of an 10-1074 antibody.

[0413] Embodiment 171. The use of embodiment 170, wherein said portion of said 10-1074 antibody comprises the amino acid sequence set forth in SEQ ID NO: 49 and / or SEQ ID NO:50.

[0414] OTHER EMBODIMENTS

[0415] It is to be understood that while the invention has been described in conjunction with the detailed description thereof, the foregoing description is intended to illustrate and not limit the scope of the invention, which is defined by the scope of the appended claims. Other aspects, advantages, and modifications are within the scope of the following claims.

Claims

WHAT IS CLAIMED IS:

1. A CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

2. A nucleic acid comprising a nucleic acid sequence encoding a CD4-D1 polypeptide of claim 1.

3. A method for treating a mammal having an human immunodeficiency virus (HIV) infection, wherein said method comprises administering a CD4-D1 polypeptide or a nucleic acid encoding said CD4-D1 polypeptide to said mammal, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

4. The method of claim 3, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

5. The method of claim 3, wherein said CD4-D1 polypeptide consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

6. The method of any one of claims 3-5, wherein said mammal is a human.

7. The method of claim 6, wherein said HIV is HIV-1.

8. The method of any one of claims 3-6, wherein said CD4-D1 polypeptide is effective to reduce a number of HIV particles present in one or more cells within said mammal.

9. The method of any one of claims 3-6, wherein said CD4-D1 polypeptide is effective to reduce a viral load of said mammal.

10. The method of any one of claims 3-6, wherein said method is effective to delay or prevent progression of said HIV infection to acquired immunodeficiency syndrome (AIDS) within said mammal.

11. The method of any one of claims 3-6, wherein said mammal has AIDS.

12. The method of any one of claims 3-11, wherein said method comprises identifying said mammal as being in need of said CD4-D1 polypeptide prior to said administering step.

13. The method of any one of claims 3-12, wherein said nucleic acid is in the form of a vector.

14. The method of claim 13, wherein said vector is an adeno-associated virus (AAV) vector.

15. The method of claim 13, wherein said vector is an expression plasmid.

16. A method for treating a mammal at risk of developing an HIV infection, wherein said method comprises administering a CD4-D1 polypeptide or a nucleic acid encoding said CD4- D1 polypeptide to said mammal, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

17. The method of claim 16, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

18. The method of claim 16, wherein said CD4-D1 polypeptide consists of the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

19. The method of any one of claims 16-18, wherein said mammal is a human.

20. The method of claim 19, wherein said HIV is HIV-1.

21. The method of any one of claims 16-18, wherein said method is effective to delay or prevent development of an HIV infection within said mammal.

22. The method of any one of claims 16-21, wherein said method comprises identifying said mammal as being in need of said CD4-D1 polypeptide prior to said administering step.

23. The method of any one of claims 16-22, wherein said nucleic acid is in the form of a vector.

24. The method of claim 23, wherein said vector is an adeno-associated virus (AAV) vector.

25. The method of claim 23, wherein said vector is an expression plasmid.

26. A method for treating a mammal having an human immunodeficiency virus (HIV) infection, wherein said method comprises administering a CD4-D1 polypeptide or a nucleic acid encoding a CD4-D1 polypeptide to said mammal, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in SEQ ID NO:40, wherein each X is independently selected from the group consisting of A, S, T, E, D, Q, N, and H.

27. The method of claim 26, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in any one of SEQ ID NOs:9-39.

28. The method of claim 26, wherein said CD4-D1 polypeptide consists of the amino acid sequence set forth in any one of SEQ ID NOs:9-39.

29. The method of any one of claims 26-28, wherein said mammal is a human.

30. The method of claim 29, wherein said HIV is HIV-1.

31. The method of any one of claims 26-30, wherein said CD4-D1 polypeptide is effective to reduce a number of HIV particles present in one or more cells within said mammal.

32. The method of any one of claims 26-30, wherein said CD4-D1 polypeptide is effective to reduce a viral load of said mammal.

33. The method of any one of claims 26-30, wherein said method is effective to delay or prevent progression of said HIV infection to acquired immunodeficiency syndrome (AIDS) within said mammal.

34. The method of any one of claims 26-30, wherein said mammal has AIDS.

35. The method of any one of claims 26-34, wherein said method comprises identifying said mammal as being in need of said CD4-D1 polypeptide prior to said administering step.

36. The method of any one of claims 26-35, wherein said nucleic acid is in the form of a vector.

37. The method of claim 36, wherein said vector is an adeno-associated virus (AAV) vector.

38. The method of claim 36, wherein said vector is an expression plasmid.

39. A method for treating a mammal at risk of developing an HIV infection, wherein said method comprises administering a CD4-D1 polypeptide or a nucleic acid encoding said CD4- D1 polypeptide to said mammal, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in SEQ ID NO:40, wherein each X is independently selected from the group consisting of A, S, T, E, D, Q, N, and H.

40. The method of claim 39, wherein said CD4-D1 polypeptide comprises the amino acid sequence set forth in any one of SEQ ID NOs:9-39.

41. The method of claim 39, wherein said CD4-D1 polypeptide consists of the amino acid sequence set forth in any one of SEQ ID NOs:9-39.

42. The method of any one of claims 39-41, wherein said mammal is a human.

43. The method of claim 42, wherein said HIV is HIV-1.

44. The method of any one of claims 39-41, wherein said method is effective to delay or prevent development of an HIV infection within said mammal.

45. The method of any one of claims 39-41, wherein said method comprises identifying said mammal as being in need of said CD4-D1 polypeptide prior to said administering step.

46. The method of any one of claims 39-45, wherein said nucleic acid is in the form of a vector.

47. The method of claim 46, wherein said vector is an adeno-associated virus (AAV) vector.

48. The method of claim 46, wherein said vector is an expression plasmid.

49. The use of a composition comprising a CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-39 to treat a mammal having an HIV infection.

50. A CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-39 for use in the preparation of a medicament to treat a mammal having an HIV infection.

51. A CD4-D 1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-39 for use in the treatment of an HIV infection.

52. The use of a composition comprising nucleic acid encoding a CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-39 to treat a mammal having an HIV infection.

53. Nucleic acid encoding a CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-39 for use in the preparation of a medicament to treat a mammal having an HIV infection.

54. Nucleic acid encoding a CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-39 for use in the treatment of an HIV infection.

55. A method for treating a mammal having, or at risk of developing, an human immunodeficiency virus (HIV) infection, wherein said method comprises administering, to said mammal, a fusion polypeptide or a nucleic acid construct encoding said fusion polypeptide to said mammal, said fusion polypeptide comprising a first polypeptide and a second polypeptide, wherein said first polypeptide is a first CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

56. The method of clam 55, wherein said mammal is a human.

57. The method of claim 56, wherein said HIV is HIV-1.

58. The method of any one of claims 55-57, wherein said second polypeptide is a second CD4-D1 polypeptide different from said first CD4-D1 polypeptide.

59. The method of claim 58, wherein both of said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently comprise the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

60. The method of claim 58, wherein both of said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently comprise the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

61. The method of claim 58, wherein said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently consist of the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

62. The method of claim 58, wherein said second polypeptide is an Fc polypeptide.

63. The method of claim 62, wherein said Fc polypeptide comprises the amino acid sequence set forth in SEQ ID NO:47.

64. The method of any one of claims 55-63, wherein said fusion polypeptide further comprises at least a portion of an m36.4 antibody.

65. The method of claim 64, wherein said portion of said m36.4 antibody comprises the amino acid sequence set forth in SEQ ID NO:48.

66. The method of any one of claims 55-63, wherein said fusion polypeptide further comprises at least a portion of an 10-1074 antibody.

67. The method of claim 66, wherein said portion of said 10-1074 antibody comprises the amino acid sequence set forth in SEQ ID NO:49 and / or SEQ ID NO:50.

68. The method of any one of claims 55-67, wherein said fusion polypeptide is effective to reduce a number of HIV particles present in one or more cells within said mammal.

69. The method of any one of claims 55-67, wherein said fusion polypeptide is effective to reduce a viral load of said mammal.

70. The method of any one of claims 55-67, wherein said method is effective to delay or prevent progression of said HIV infection to acquired immunodeficiency syndrome (AIDS) within said mammal.

71. The method of any one of claims 55-67, wherein said mammal has AIDS.

72. The method of any one of claims 55-71, wherein said method comprises identifying said mammal as being in need of said fusion polypeptide prior to said administering step.

73. The method of any one of claims 55-67, wherein said method is effective to delay or prevent development of an HIV infection within said mammal.

74. A method for treating a mammal having, or at risk of developing, an HIV infection, wherein said method comprises administering, to said mammal, a fusion polypeptide or a nucleic acid construct encoding said fusion polypeptide, said fusion polypeptide comprising a first polypeptide and a second polypeptide, wherein said first polypeptide is a first CD4-D1 polypeptide comprising the amino acid sequence set forth in SEQ ID NO:40, wherein each X is independently selected from the group consisting of A, S, T, E, D, Q, N, and H.

75. The method of clam 74, wherein said mammal is a human.

76. The method of claim 75, wherein said HIV is HIV-1.

77. The method of any one of claims 74-76, wherein said second polypeptide is a second CD4-D1 polypeptide different from said first CD4-D1 polypeptide.

78. The method of claim 77, wherein both of said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently comprise the amino acid sequence set forth in any one of SEQ ID NOs:9-39.

79. The method of claim 77, wherein said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently consist of the amino acid sequence set forth in any one of SEQ ID NOs:9-39.

80. The method of any one of claims 74-76, wherein said second polypeptide is an Fc polypeptide.

81. The method of claim 80, wherein said Fc polypeptide comprises the amino acid sequence set forth in SEQ ID NO:47.

82. The method of any one of claims 74-76, wherein said fusion polypeptide further comprises at least a portion of an m36.4 antibody.

83. The method of claim 82, wherein said portion of said m36.4 antibody comprises the amino acid sequence set forth in SEQ ID NO:48.

84. The method of any one of claims 74-76, wherein said fusion polypeptide further comprises at least a portion of an 10-1074 antibody.

85. The method of claim 84, wherein said portion of said 10-1074 antibody comprises the amino acid sequence set forth in SEQ ID NO:49 and / or SEQ ID NO:50.

86. The method of any one of claims 74-85, wherein said fusion polypeptide is effective to reduce a number of HIV particles present in one or more cells within said mammal.

87. The method of any one of claims 74-85, wherein said fusion polypeptide is effective to reduce a viral load of said mammal.

88. The method of any one of claims 74-85, wherein said method is effective to delay or prevent progression of said HIV infection to acquired immunodeficiency syndrome (AIDS) within said mammal.

89. The method of any one of claims 74-85, wherein said mammal has AIDS.

90. The method of any one of claims 74-89, wherein said method comprises identifying said mammal as being in need of said fusion polypeptide prior to said administering step.

91. The method of any one of claims 74-85, wherein said method is effective to delay or prevent development of an HIV infection within said mammal.

92. A fusion polypeptide comprising a first polypeptide and a second polypeptide for use in the preparation of a medicament to treat a mammal having an HIV infection, wherein said first polypeptide is a first CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

93. The fusion polypeptide of clam 92, wherein said mammal is a human.

94. The fusion polypeptide of claim 93, wherein said HIV is HIV-1.

95. The fusion polypeptide of any one of claims 92-94, wherein said second polypeptide is a second CD4-D1 polypeptide different from said first CD4-D1 polypeptide.

96. The fusion polypeptide of claim 95, wherein both of said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently comprise the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

97. The fusion polypeptide of claim 95, wherein both of said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently comprise the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

98. The fusion polypeptide of claim 95, wherein said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently consist of the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

99. The fusion polypeptide of claim 95, wherein said second polypeptide is an Fc polypeptide.

100. The fusion polypeptide of claim 99, wherein said Fc polypeptide comprises the amino acid sequence set forth in SEQ ID NO:47.

101. The fusion polypeptide of any one of claims 92-94, wherein said fusion polypeptide further comprises at least a portion of an m36.4 antibody.

102. The fusion polypeptide of claim 101, wherein said portion of said m36.4 antibody comprises the amino acid sequence set forth in SEQ ID NO:48.

103. The fusion polypeptide of any one of claims 92-94, wherein said fusion polypeptide further comprises at least a portion of an 10-1074 antibody.

104. The fusion polypeptide of claim 103, wherein said portion of said 10-1074 antibody comprises the amino acid sequence set forth in SEQ ID NO:49 and / or SEQ ID NO:50.

105. A fusion polypeptide comprising a first polypeptide and a second polypeptide for use in the treatment of an HIV infection, wherein said first polypeptide is a first CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

106. The fusion polypeptide of claim 105, wherein said HIV is HIV-1.

107. The fusion polypeptide of any one of claims 105-106, wherein said second polypeptide is a second CD4-D1 polypeptide different from said first CD4-D1 polypeptide.

108. The fusion polypeptide of claim 107, wherein both of said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently comprise the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

109. The fusion polypeptide of claim 107, wherein both of said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently comprise the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

110. The fusion polypeptide of claim 107, wherein said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently consist of the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

111. The fusion polypeptide of any one of claims 105-106, wherein said second polypeptide is an Fc polypeptide.

112. The fusion polypeptide of claim 111, wherein said Fc polypeptide comprises the amino acid sequence set forth in SEQ ID NO:47.

113. The fusion polypeptide of any one of claims 105-106, wherein said fusion polypeptide further comprises at least a portion of an m36.4 antibody.

114. The fusion polypeptide of claim 113, wherein said portion of said m36.4 antibody comprises the amino acid sequence set forth in SEQ ID NO:48.

115. The fusion polypeptide of any one of claims 105-106, wherein said fusion polypeptide further comprises at least a portion of an 10-1074 antibody.

116. The fusion polypeptide of claim 115, wherein said portion of said 10-1074 antibody comprises the amino acid sequence set forth in SEQ ID NO:49 and / or SEQ ID NO:50.

117. The use of a composition comprising a fusion polypeptide comprising a first polypeptide and a second polypeptide to treat a mammal having an HIV infection, wherein said first polypeptide is a first CD4-D1 polypeptide comprising the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

118. The use of clam 117, wherein said mammal is a human.

119. The use of claim 118, wherein said HIV is HIV- 1.

120. The use of any one of claims 117-118, wherein said second polypeptide is a second CD4-D1 polypeptide different from said first CD4-D1 polypeptide.

121. The use of claim 120, wherein both of said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently comprise the amino acid sequence set forth in any one of SEQ ID NOs:4-39.

122. The use of claim 120, wherein both of said first CD4-D1 polypeptide and said second CD4-D1 polypeptide each independently comprise the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

123. The use of claim 120, wherein said first CD4-D1 polypeptide and said second CD4- D1 polypeptide each independently consist of the amino acid sequence set forth in any one of SEQ ID NOs:4-8.

124. The use of any one of claims 117-118, wherein said second polypeptide is an Fc polypeptide.

125. The use of claim 124, wherein said Fc polypeptide comprises the amino acid sequence set forth in SEQ ID NO:47.

126. The use of any one of claims 117-118, wherein said fusion polypeptide further comprises at least a portion of an m36.4 antibody.

127. The use of claim 126, wherein said portion of said m36.4 antibody comprises the amino acid sequence set forth in SEQ ID NO:48.

128. The use of any one of claims 117-118, wherein said fusion polypeptide further comprises at least a portion of an 10-1074 antibody.

129. The use of claim 128, wherein said portion of said 10-1074 antibody comprises the amino acid sequence set forth in SEQ ID NO: 49 and / or SEQ ID NO:50.

Images