WO2026024596A1 - Methods of stimulating an immune response - Google Patents

Methods of stimulating an immune response

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Publication number
WO2026024596A1
WO2026024596A1 PCT/US2025/038411 US2025038411W WO2026024596A1 WO 2026024596 A1 WO2026024596 A1 WO 2026024596A1 US 2025038411 W US2025038411 W US 2025038411W WO 2026024596 A1 WO2026024596 A1 WO 2026024596A1
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WIPO (PCT)
Prior art keywords
hla
molecule
tapbpr
seq
peptide
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PCT/US2025/038411
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French (fr)
Inventor
Nikolaos SGOURAKIS
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Childrens Hospital of Philadelphia CHOP
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Childrens Hospital of Philadelphia CHOP
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Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/46—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
    • C07K14/47—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/385—Haptens or antigens, bound to carriers
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/705—Receptors; Cell surface antigens; Cell surface determinants
    • C07K14/70503—Immunoglobulin superfamily
    • C07K14/70539—MHC-molecules, e.g. HLA-molecules
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2319/00—Fusion polypeptide
    • C07K2319/01—Fusion polypeptide containing a localisation/targetting motif
    • C07K2319/02—Fusion polypeptide containing a localisation/targetting motif containing a signal sequence
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2319/00—Fusion polypeptide
    • C07K2319/01—Fusion polypeptide containing a localisation/targetting motif
    • C07K2319/03—Fusion polypeptide containing a localisation/targetting motif containing a transmembrane segment
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2319/00—Fusion polypeptide
    • C07K2319/40—Fusion polypeptide containing a tag for immunodetection, or an epitope for immunisation
    • C07K2319/43—Fusion polypeptide containing a tag for immunodetection, or an epitope for immunisation containing a FLAG-tag

Definitions

  • the methods comprise contacting a population of mammalian cells or the subject in need thereof with an immunogenic agent and a TAP-binding protein-related (TAPBPR) polypeptide.
  • TAPBPR TAP-binding protein-related
  • MHC-I major histocompatibility complex
  • TCR T cell receptors
  • NK natural killer
  • HLA Human Leukocyte Antigen
  • human MHC-I Class I Human Leukocyte Antigen
  • Antigen processing and presentation (APP) chaperones have more recently emerged as key components for the presentation of metabolite ligands on MHC- related 1 (MR1) molecules (22-24), and have provided powerful tools for in vitro applications, including the generation of pMHC-I libraries encompassing different peptide specificities or the loading of antigens on cells independently of the endogenous processing pathway (18, 25, 26).
  • MR1 MHC- related 1
  • the methods comprise providing a population of mammalian cells having surface MHC class I molecule with an immunogenic agent and a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecules thereby increasing the immunogenicity of the mammalian cells.
  • TAPBPR TAP-binding protein-related
  • the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11.
  • the MHC class I molecule is selected from the group consisting of an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, an HLA-E molecule, an HLA-G molecule, an HLA-F molecule, a MR1 molecule, a CD1 molecule, and a combination thereof.
  • the MHC class I molecule can, for example, be an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, a MR1 molecule, an HLA-E molecule, and/or an HLA-G molecule.
  • the HLA-A molecule, HLA-B molecule, and/or HLA-C molecule can, for example, be selected from an HLA-A01 supertype molecule, an HLA-A02 supertype molecule, an HLA-A03 supertype molecule, an HLA-A24 supertype molecule, an HLA- A01/A24 supertype molecule, an HLA-A01/A03 supertype molecule, an HLA-B08 supertype molecule, an HLA-B44 supertype molecule, an HLA-B58 supertype molecule, and/or an HLA-C01 supertype molecule.
  • the MHC class I molecule can, for example, be selected from HLA-A*01:01, HLA-A*11:01, HLA-A*29:01, HLA-A*30:01, HLA-A*32:01, HLA-A*74:01, HLA-A*24:03, HLA-A*02:06, HLA-A*02:01, HLA- A*02:03, HLA-A*23:01, HLA-A*24:02, HLA-A*68:01, HLA-A*69:01, HLA-A*68:02, HLA-A*29:02, HLA-A*03:01, HLA-B*08:01, HLA-B*57:03, HLA-B*59:01, HLA-B*37:01, HLA-C*01:02, HLA-C*03:04, MR1, HLA-E*01:03, and/or HLA-G*01:01.
  • the mammalian cell is a human cell.
  • the immunogenic agent is selected from an immunogenic peptide, a metabolite, a lipid, a phospholipid, a fatty acid ligand, and/or combinations thereof.
  • the immunogenic peptides are low-to-moderate affinity peptides.
  • the method is performed in vivo.
  • the methods comprise (a) obtaining antigen presenting cells from the subject in need thereof; and (b) providing antigen presenting cells comprising surface MHC class I molecules in vitro with a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7 and an immunogenic agent, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecules thereby creating antigen presenting cells for stimulating an immune response.
  • the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11.
  • the MHC class I molecule is selected from the group consisting of an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, an HLA-E molecule, an HLA-G molecule, an HLA-F molecule, a MR1 molecule, a CD1 molecule, and a combination thereof.
  • the MHC class I molecule can, for example, be an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, a MR1 molecule, an HLA-E molecule, and/or an HLA-G molecule.
  • the HLA-A molecule, HLA-B molecule, and/or HLA-C molecule can, for example, be selected from an HLA-A01 supertype molecule, an HLA-A02 supertype molecule, an HLA-A03 supertype molecule, an HLA-A24 supertype molecule, an HLA- A01/A24 supertype molecule, an HLA-A01/A03 supertype molecule, an HLA-B08 supertype molecule, an HLA-B44 supertype molecule, an HLA-B58 supertype molecule, and/or an HLA-C01 supertype molecule.
  • the MHC class I molecule can, for example, be selected from HLA-A*01:01, HLA-A*11:01, HLA-A*29:01, HLA-A*30:01, HLA-A*32:01, HLA-A*74:01, HLA-A*24:03, HLA-A*02:06, HLA-A*02:01, HLA- A*02:03, HLA-A*23:01, HLA-A*24:02, HLA-A*68:01, HLA-A*69:01, HLA-A*68:02, HLA-A*29:02, HLA-A*03:01, HLA-B*08:01, HLA-B*57:03, HLA-B*59:01, HLA-B*37:01, HLA-C*01:02, HLA-C*03:04, MR1, HLA-E*01:03, and/or HLA-G*01:01.
  • the mammalian cell is a human cell.
  • the immunogenic agent is selected from an immunogenic peptide, a metabolite, a lipid, a phospholipid, a fatty acid ligand, and/or combinations thereof.
  • the immunogenic peptides are low-to-moderate affinity peptides.
  • the antigen presenting cells are dendritic cells.
  • the method further comprises administering the antigen presenting cells to the subject in need thereof.
  • Also provided are methods of stimulating an immune response in a subject in need thereof comprising (a) administering to the subject in need thereof a TAP- binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7, wherein the TAPBPR polypeptide binds to an antigen presenting cell comprising a surface MHC class I molecule in the subject in need thereof; and (b) administering to the subject in thereof an immunogenic agent, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecule of the antigen presenting cell such that the antigen presenting cell stimulates an immune response in the subject in need thereof.
  • TAPBPR TAP- binding protein-related
  • TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7 and an immunogenic agent for use in stimulating an immune response in a subject in need thereof, 4 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 wherein the TAPBPR polypeptide binds to an antigen presenting cell comprising a surface MHC class I molecule; and wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecule of the antigen presenting cell such that the antigen presenting cell stimulates an immune response.
  • TAPBPR TAP-binding protein-related
  • the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10.
  • the MHC class I molecule is selected from the group consisting of an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, an HLA-E molecule, an HLA-G molecule, an HLA-F molecule, a MR1 molecule, a CD1 molecule, and a combination thereof.
  • the MHC class I molecule can, for example, be an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, a MR1 molecule, an HLA-E molecule, and/or an HLA-G molecule.
  • the HLA-A molecule, HLA-B molecule, and/or HLA-C molecule can, for example, be selected from an HLA-A01 supertype molecule, an HLA-A02 supertype molecule, an HLA-A03 supertype molecule, an HLA-A24 supertype molecule, an HLA- A01/A24 supertype molecule, an HLA-A01/A03 supertype molecule, an HLA-B08 supertype molecule, an HLA-B44 supertype molecule, an HLA-B58 supertype molecule, and/or an HLA-C01 supertype molecule.
  • the MHC class I molecule can, for example, be selected from HLA-A*01:01, HLA-A*11:01, HLA-A*29:01, HLA-A*30:01, HLA-A*32:01, HLA-A*74:01, HLA-A*24:03, HLA-A*02:06, HLA-A*02:01, HLA- A*02:03, HLA-A*23:01, HLA-A*24:02, HLA-A*68:01, HLA-A*69:01, HLA-A*68:02, HLA-A*29:02, HLA-A*03:01, HLA-B*08:01, HLA-B*57:03, HLA-B*59:01, HLA-B*37:01, HLA-C*01:02, HLA-C*03:04, MR1, HLA-E*01:03, and/or HLA-G*01:01.
  • the subject is a human.
  • the immunogenic agent is selected from an immunogenic peptide, a metabolite, a lipid, a phospholipid, a fatty acid ligand, and/or combinations thereof.
  • the immunogenic peptides are low-to-moderate affinity peptides.
  • the antigen presenting cells are dendritic cells.
  • the TAPBPR polypeptide further comprises a signal peptide, a flag tag, a linker, and/or a transmembrane domain.
  • the signal peptide can, for example, comprise SEQ ID NO:47.
  • the flag tag can, for example, comprise SEQ ID NO:48.
  • the linker can, for example comprise SEQ ID NO:49.
  • the transmembrane domain can, for example, comprise SEQ ID NO:50.
  • TAPBPR TAP-binding protein-related polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7.
  • the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11.
  • the TAPBPR polypeptide further comprises a signal peptide, a flag tag, a linker, and/or a transmembrane domain.
  • the signal peptide can, for example, comprise SEQ ID NO:47.
  • the flag tag can, for example, comprise SEQ ID NO:48.
  • the linker can, for example comprise SEQ ID NO:49.
  • the transmembrane domain can, for example, comprise SEQ ID NO:50.
  • the host cell is a mammalian cell.
  • FIG. 1A shows a structural model of TAPBPR FLQ in complex with peptide-free HLA-A*02:01/ ⁇ 2m generated using RosettaCM.
  • the G24-R36 loop is colored in orange, and the A29-S32 loop segment is shown as spheres.
  • the side chains of mutations S104F, K211L, and R270Q are shown as magenta sticks.
  • FIG.1B shows a representative SPR sensorgram of graded concentrations of TAPBPR FLQ flown over a streptavidin chip coupled with HLA- A*02:01/TAX9 in excess TAX9 peptide.
  • FIG.1C shows a log-scale comparison of K D values between HLA-A*02:01/TAX9 and TAPBPR.
  • FIG. 1E shows relative peptide dissociation of TAMRA-TAX9 from HLA-A*02:01 by TAPBPR relative to no TAPBPR.
  • FIG. 1F shows correlation of Log10 (MFI ratio) between TAPBPR WT and TAPBPR FLQ plotted in FIG. 6.
  • the dashed line represents a conceptual 1:1 correlation.
  • Two-sample unequal variance student’s t-test was performed, P > 0.12 (not significant, ns), P ⁇ 0.033(*), P ⁇ 0.002(**), and P ⁇ 0.001(***).
  • FIG.2 shows a comparison of the sequences for TAPBPR, and its variants used in this study.
  • FIGs. 3A-3B show the purification of recombinant TAPBPR FLQ ⁇ G24-R36 protein.
  • FIG. 3A shows a SEC trace of TAPBPR FLQ ⁇ G24-R36 mutant. The arrow indicates the protein peak relative to TAPBPR WT and TAPBPR FLQ and is further confirmed by SDS/PAGE analysis.
  • FIGs. 4A-4F show direct interactions between HLA-A allotypes and TAPBPR variants.
  • FIGs. 4A-4F show representative SPR sensorgrams of various concentrations of TAPBPR WT (FIG. 4A), TAPBPR ⁇ ALAS (FIG.
  • FIG. 4B TAPBPR ⁇ G24-R36
  • FIG. 4C TAPBPR FLQ
  • FIG. 4E TAPBPR FLQ ⁇ G24-R36
  • FIG. 4F TAPBPR TN6
  • FIG. 5 shows a graph of a peptide dissociation assay. Comparison of peptide dissociation kinetics of TAMRA fluorophore-labeled TAMRATAX9 peptide (TAMRAKLFGYPVYV (SEQ ID NO:2))-loaded HLA-A*02:01 (40nM) with 1 ⁇ M unlabeled TAX9 peptide in the presence of buffer or 1 ⁇ M TAPBPR TN6 , TAPBPR WT , TAPBPR ⁇ G24-R36 , TAPBPR FLQ , and TAPBPR FLQ ⁇ G24-R36 .
  • TAMRA fluorophore-labeled TAMRATAX9 peptide TAMRAKLFGYPVYV (SEQ ID NO:2)
  • FIG.6 shows binding levels of TAPBPR tetramers on HLA single antigen beads.
  • FIG. 6 middle and bottom, shows bar graphs showing the Log10 Mean Fluorescence Intensity (MFI) levels of tetramerized TAPBPR FLQ , TAPBPR WT , and TAPBPR TN6 binding to HLA molecules on SABs. Tetramer staining upon the W6/32 antibody incubation was used to control for background staining levels.
  • MFI Log10 Mean Fluorescence Intensity
  • FIGs. 7A-7C shows log 10 (MFI ratios) of TAPBPR tetramer binding to 97 common HLA allotypes.
  • FIG. 7A shows a bar graph showing the Log10 (MFI ratios) of TAPBPR FLQ , TAPBPR WT , and TAPBPR TN6 (negative control) binding to the SABs. Plotted data are means ⁇ SD from 2 to 3 independent experiments.
  • FIGs. 7B-7C show graphs of the correlation of Log10 (MFI ratio) for TAPBPR FLQ (FIG. 7B) and TAPBPR WT (FIG.
  • FIGs. 8A-8D demonstrated that TAPBPR FLQ enhanced peptide loading across HLA allotypes expressed on a cellular membrane.
  • FIG. 8A shows a schematic of peptide exchange for MHC-I expressed on the cell surface by soluble TAPBPR.
  • FIG. 8B shows a bar graph summarizing the MFI of fluorescent peptide binding for monoallelic HLA-A*02:01, A*24:02, and A23:01 cell lines with 10 nM fluorescent peptide and 1 ⁇ M TAPBPR and HLA-A*03:01 cell line in the presence of 10 ⁇ M fluorescent peptide and 10 ⁇ M TAPBPR, as indicated, from three independent experiments.
  • Two-way ANOVA was performed relative to TAPBPR TN6 , P > 0.0.1234 (ns), P ⁇ 0.0332(*), P ⁇ 0.0021(**), P ⁇ 0.0002(***), and P ⁇ 0.0001(****).
  • FIG. 8C shows a schematic of peptide exchange for MHC-I expressed on the cell surface by membrane-bound TAPBPR.
  • FIG. 8D shows a bar graph summarizing the MFI of different concentrations of fluorescent peptide binding to HLA-A*02:01 expressed T2 cell line with expression of TAPBPR-TM, as indicated, from two independent experiments. Two- way ANOVA was performed relative to the parental (no transduction), P > 0.0.1234 (ns), P ⁇ 0.0332(*), P ⁇ 0.0021(**), P ⁇ 0.0002(***), and P ⁇ 0.0001(****).
  • FIGs. 9A-9D show flow cytometry gating strategy of monoallelic 722.211 cell lines.
  • FIGs. 9A-9D show flow cytometry gating strategy of monoallelic 722.211 cell lines.
  • FIG. 9A-9D show the results of flow cytometry for monoallelic HLA-A*02:01 (FIG. 9A), 8 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 24:02 (FIG. 9B), 23:01 (FIG. 9C), or 03:01 (FIG. 9D) 722.211 cell lines were thawed and recovered before performing peptide exchange. Cells were sorted by side and forward scatter (SSC-A and FSC-A) followed by single cell isolation (SSC-A and SSC-H). Gating for live cells was determined by LIVE/DEADTM Fixable Violet Dead Cell Stain Kit.
  • FIGs.10A-10D show soluble TAPBPR mediated peptide exchange on surface MHC-I.
  • FIGs. 10A-10D show bar graph summarizing the MFI of fluorescent peptide binding to monoallelic HLA-A*02:01 (FIG. 10A), A*24:02 (FIG. 10B), A23:01 (FIG. 10C), and A*03:01 (FIG.
  • HLA-A*02:01, A*24:02 (FIG. 10B), and A23:01 (FIG. 10C) 721.211 cell lines were incubated in the presence of soluble TAPBPR TN6 , TAPBPR WT , and TAPBPR FLQ at different concentrations, followed by incubation with 10 nM fluorescent peptide for 60 minutes.
  • HLA-A*03:01 monoallelic cell line was incubated in the presence of soluble TAPBPR TN6 , TAPBPR WT , and TAPBPR FLQ , followed by incubation with 10 ⁇ M (FIG.
  • FIG. 11 shows fluorescent peptide binding on monoallelic HLA-A*02:01 cell line. Titration of soluble TAPBPR TN6 , TAPBPR WT , and TAPBPR FLQ at different concentrations to the monoallelic HLA-A*02:01 721.211 cell line with the incubation of 10 nM fluorescent peptide for 60 minutes.
  • FIG. 12 shows fluorescent peptide binding on monoallelic HLA-A*24:02 cell line. Titration of soluble TAPBPR TN6 , TAPBPR WT , and TAPBPR FLQ at different concentrations to the monoallelic HLA-A*24:02 721.211 cell line with the incubation of 10 nM fluorescent peptide for 60 minutes. Gates are shown in black box, and the percentages of events are gated in parentheses.
  • FIG. 13 shows fluorescent peptide binding on monoallelic HLA-A*23:01 cell line. Titration of soluble TAPBPR TN6 , TAPBPR WT , and TAPBPR FLQ at different concentrations to the monoallelic HLA-A*23:01 721.211 cell line with the incubation of 10 nM fluorescent 9 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 peptide for 60 minutes.
  • FIG. 14 shows fluorescent peptide binding on monoallelic HLA-A*03:01 cell line. Titration of soluble TAPBPR TN6 , TAPBPR WT , and TAPBPR FLQ at different concentrations to the monoallelic HLA-A*24:02 721.211 cell line with the incubation of 10 ⁇ M fluorescent peptide for 60 minutes. Gates are shown in black box, and the percentages of events are gated in parentheses.
  • FIG. 15 shows representative flow cytometry gating strategy of T2 cell lines in peptide exchange experiments.
  • Cells were incubated with varying concentrations of fluorescent TAMRA-TAX9 peptide for 60 minutes before washing with FACS buffer, staining with LIVE/DEADTM Fixable Near IR (876) Viability dye, and fixation with 4% PFA in PBS. Analyzed cells were gated on live singlets of appropriate size. Cells incubated with 10 -5 M of TAX9-TAMARA are shown.
  • FIGs. 16A-16C show flow cytometry analysis of T2 cell lines.
  • FIG. 16A shows a graph demonstrating surface TAPBPR expression of TAPBPR FLQ , TAPBPR WT , and TAPBPR TN6 -TM transduced T2 cell lines, as measured by anti-FLAG MFI.
  • FIG. 16B shows a graph demonstrating TAPBPR transduction efficiency of TAPBPR FLQ , TAPBPR WT , and TAPBPR TN6 -TM transduced T2 cell lines relative to the non-transduced parental cell line.
  • FIG. 16C shows a graph demonstrating surface HLA- A*02:01 expression of TAPBPR-FLQ, WT, and TN6 TAPBPR FLQ , TAPBPR WT , and TAPBPR TN6 -TM transduced and parental T2 cell lines from cells selected for peptide exchange assays.
  • FIG. 17 shows fluorescent peptide binding of HLA-A*02:01 mediated by surface TAPBPR. Titration of peptide at different concentrations to TAPBPR TN6 , TAPBPR WT , and TAPBPR FLQ -TM transduced T2 cell lines.
  • FIGs. 18A-18E demonstrated the isolation and cryoEM structure of an MHC-I intermediate bound to a peptide decoy by TAPBPR.
  • 18A shows a size exclusion chromatography purification of a recombinant open HLA-A*02:01/TAPBPR FLQ complex 10 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 prepared by UV-irradiation of KILGFVFJV/HLA-A*02:01 pre-incubated with TAPBPR FLQ at 1:1.3 molar ratio.
  • J 3-amino-3-(2-nitrophenyl)-propionic acid.
  • SDS/PAGE analysis confirms the identity of the TAPBPR FLQ /open HLA-A2 complex peak under nonreducing (NR) or reducing (R) conditions.
  • FIG.18B shows a LC/MS analysis of the purified complex in FIG.18A showing the presence of captured peptide decoy KILGFVF (SEQ ID NO:5) (observed and expected mass-to-charge ratios are 822.52 and 822.50 m/z, respectively).
  • SEQ ID NO:5 captured peptide decoy KILGFVF
  • FIG. 18C shows a reconstructed cryoEM density and refined structural model of the tertiary complex, with TAPBPR FLQ shown in light pink, open HLA-A*02:01 heavy chain in light green, ⁇ 2 m in light blue, and peptide in wheat, as indicated.
  • FIG. 18D show a zoom in on the interactions between the TAPBPR FLQ hairpin and the floor of the MHC-I groove.
  • FIG. 18E shows a zoom in on the interactions between TAPBPR FLQ IgC domain residues and ⁇ 2m, highlighting the sidechain interaction.
  • FIGs. 19A-19B show LC-MS validation of UV-irradiated placeholder ligand.
  • FIGs. 19A-19B show LC-MS validation of UV-irradiated placeholder ligand.
  • FIG. 19A-19B show LC-MS analysis of peptide KILGFVFJV (SEQ ID NO:46) (FIG. 19A) and UV-irradiated peptide KILGFVFJV (SEQ ID NO:46) (FIG. 19B).
  • J 3-amino-3-(2- nitrophenyl)-propionic acid. UV irradiation was performed at 365 nm for 40 minutes at 4 ⁇ C.
  • Left panel the LC chromatogram trace of each sample.
  • Right panel relative abundance for the selected time interval (red box).
  • FIGs.20A-20E show NMR characterization of TAPBPR FLQ interactions with peptide- loaded HLA-A*02:01.
  • FIG. 20A shows a representative region of the 2D 1 H- 13 C HMQC spectral overlay of 13 C AILV methyl-labeled TAX9/HLA-A*02:01 and with a 3-fold molar excess of TAPBPR FLQ .
  • FIG. 20B shows selected NMR resonances (corresponding to I23 and A125) undergoing conformational motion in the slow-exchange regime upon titration of TAPBPR FLQ .
  • the HLA-A*02:01: TAPBPR FLQ ratios for the different titration points were 1: 0, 1: 0.55, 1: 1.10, 1: 1.65, 1: 2.75 and 1: 3.31.
  • FIG. 20C shows that methyl groups of residues undergoing significant chemical shift deviation (CSD) upon binding TAPBPR FLQ are mapped onto the structure of the HLA-A*02:01/TAPBPR FLQ complex.
  • the structure is a Roseta-CM homology model obtained using the H2-D d /TAPBPR crystal structure as a template (PDB ID: 5WER).
  • CSDs are plotted using a heat-map scale 11 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 shown on the right.
  • FIG. 20D shows a correlation plot of CSDs observed for the titration of HLA-A*02:01 with TAPBPR WT versus TAPBPR FLQ .
  • the Pearson correlation coefficient (r) is shown on the plot. The correlations are statistically significant with a P value of ⁇ 0.0001.
  • FIG. 20E shows a close-up of the peptide-binding groove with the TAX9 peptide (shown as green sticks). Select residues distributed throughout the MHC-I groove and their CSDs are labeled. The mutation sites on TAPBPR FLQ are denoted as pink spheres. [0051] FIGs.
  • FIG. 21A-21C show NMR spectra overlay and line shape analysis of peptide-loaded HLA-A*02:01 upon TAPBPR FLQ titration.
  • FIG. 21A shows a 2D 1 H- 13 C HMQC spectral overlay of 13 C AILV methyl-labeled HLA-A*02:01 titrated with TAPBPR FLQ in the molar ratios: 1: 0, 1: 0.55, 1: 1.10, 1: 1.65, 1: 2.75, 1: 3.31.
  • FIG. 21B resonances for selected methyl groups (I23, I52, A125, L126, V247) undergoing slow exchange upon adding TAPBPR FLQ .
  • FIG. 21C shows a representative NMR line shape fitting of TAPBPR FLQ titration onto TAX9/HLA-A*02:01/ ⁇ 2 m complex for heavy chain methyl residues A125 and L126 using TITAN (5).
  • FIG. 22 shows a schematic of a cryo-EM data processing workflow. Cryo-EM data processing workflow including a representative micrograph (top left).
  • FIG. 23 shows cryoEM data quality. Local resolution calculated using Relion’s local resolution tool (top panels). Unmasked (grey dashed), masked (black) and movel versus map (green) fourier shell correlation (FSC) curves (bottom left panel).
  • FSC Fourier shell correlation
  • FIGs. 24A-24C show structural adaptations of the MHC-I groove induced by TAPBPR.
  • FIG. 24A shows an overview of the molecular complex between TAPBPR FLQ /HLA-A*02:01/ ⁇ 2 m with bound peptide decoy. The MHC-I ⁇ 2 helix has been partially removed for simplicity.
  • FIG.24B shows a structural superposition of peptide-loaded HLA-A*02:01 (PDB ID 2VLL) onto the cryoEM structure of KILGFVF (SEQ ID NO:5)/HLA-A*02:01 (PDB ID 9C96, EMDB ID EMD-45360) showing well-resolved density for peptide residues 1-6, and a lack of cryoEM density for residues G71-G91 of HLA- A*02:01. MHC-I A-, B-, D-, E-, F-pockets are noted, and the spheres are showing A150 C ⁇ .
  • FIG. 27 shows a sequence logo of conserved peptide-coordinating residues across 215 common HLA allotypes. Seq2logo visualization (20) depicting the conserved peptide- coordinating residues’ sequence alignment across 215 distinct HLA-A*, B*, and C* allotypes.
  • FIG. 28 shows a structural mechanism of peptide antigen proofreading by TAPBPR.
  • TAPBPR Empty MHC-I is preferentially recognized by TAPBPR in a peptide-receptive, “open” conformation.
  • Chaperoned MHC-I screens a large peptide pool in the endoplasmic reticulum, forming transient peptide/MHC-I/TAPBPR complexes.
  • TAPBPR widens the ⁇ 2-1 helix of the peptide binding groove, enhances the dynamics of the ⁇ 1 helix, and induces unstructured F- pockets.
  • chaperone MHC-I exhibits a lower affinity towards incoming peptides and promotes the dissociation of suboptimal peptide decoys.
  • the N terminal peptide forms native-like contacts with its A- and B-pockets.
  • the C-terminus of the peptide interacts with peptide-coordinating residues in the F-pocket to stabilize and close the peptide binding groove, which allosterically triggers the release of TAPBPR.
  • This disclosure is based on the discovery that it is possible to increase the immunogenicity of mammalian cells, to produce antigen presenting cells to stimulate an immune response in a subject, and to stimulate an immune response in a subject in need thereof use a combination of an immunogenic agent and a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecules thereby increasing the immunogenicity of the mammalian cells, producing antigen presenting cells to stimulate an immune response, and stimulating an immune response in a subject.
  • TAPBPR TAP-binding protein-related
  • a concentration range of 1% to 10% (w/v) includes 0.9% (w/v) to 11% (w/v).
  • the use of a numerical range expressly includes all possible subranges, all individual numerical values within that range, including integers within such ranges and fractions of the values unless the context clearly indicates otherwise. [0065] Unless otherwise indicated, the term “at least” preceding a series of elements is to be understood to refer to every element in the series. Those skilled in the art will recognize or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments of the invention described herein. Such equivalents are intended to be encompassed by the invention.
  • the terms “comprises,” “comprising,” “includes,” “including,” “has,” “having,” “contains” or “containing,” or any other variation thereof, will be understood to imply the inclusion of a stated integer or group of integers but not the exclusion of any other integer or group of integers and are intended to be non-exclusive or open-ended.
  • a composition, a mixture, a process, a method, an article, or an apparatus that comprises a list of elements is not necessarily limited to only those elements but can include other elements not expressly listed or inherent to such composition, mixture, process, method, article, or apparatus.
  • a first option refers to the applicability of the first element without the second.
  • a second option refers to the applicability of the second element without the first.
  • a third option refers to the applicability of the first and second elements together. Any one of these options is understood to fall within the meaning, and therefore satisfy the requirement of the term “and/or” as used herein.
  • “subject” means any animal, preferably a mammal, most preferably a human.
  • mammals include, but are not limited to, cows, horses, sheep, pigs, cats, dogs, mice, rats, rabbits, guinea pigs, monkeys, humans, etc., more preferably a human.
  • the words “right,” “left,” “lower,” “upper,” “top,” “bottom,” and “middle” designate directions in the drawings to which reference is made.
  • references that include a numerical parameter would include variations that, using mathematical and industrial principles accepted in the art (e.g., rounding, 15 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 measurement or other systematic errors, manufacturing tolerances, etc.), would not vary the least significant digit.
  • nucleic acids or polypeptide sequences e.g., TAP-binding protein-related (TAPBPR) polypeptides and nucleotide sequences encoding the same
  • TAPBPR TAP-binding protein-related polypeptides and nucleotide sequences encoding the same
  • sequences or subsequences that are the same or have a specified percentage of amino acid residues or nucleotides that are the same, when compared and aligned for maximum correspondence, as measured using one of the following sequence comparison algorithms or by visual inspection.
  • sequence comparison typically one sequence acts as a reference sequence, to which test sequences are compared.
  • test and reference sequences are input into a computer, subsequence coordinates are designated, if necessary, and sequence algorithm program parameters are designated.
  • sequence comparison algorithm calculates the percent sequence identity for the test sequence(s) relative to the reference sequence, based on the designated program parameters.
  • Optimal alignment of sequences for comparison can be conducted, e.g., by the local homology algorithm of Smith & Waterman, Adv. Appl. Math. 1981; 2:482, by the homology alignment algorithm of Needleman & Wunsch, J. Mol. Biol. 1970; 48:443, by the search for similarity method of Pearson & Lipman, Proc. Nat’l. Acad. Sci.
  • HSPs high scoring sequence pairs
  • Extension of the word hits in each direction are halted when: the cumulative alignment score falls off by the quantity X from its maximum achieved value; the cumulative score goes to zero or below, due to the accumulation of one or more negative- scoring residue alignments; or the end of either sequence is reached.
  • the BLAST algorithm parameters W, T, and X determine the sensitivity and speed of the alignment.
  • the BLASTP program uses as defaults a wordlength (W) of 3, an expectation (E) of 10, and the BLOSUM62 scoring matrix (see Henikoff & Henikoff, Proc.
  • the BLAST algorithm In addition to calculating percent sequence identity, the BLAST algorithm also performs a statistical analysis of the similarity between two sequences (see, e.g., Karlin & Altschul, Proc. Nat’l. Acad. Sci. USA 1993; 90:5873-5787).
  • One measure of similarity provided by the BLAST algorithm is the smallest sum probability (P(N)), which provides an indication of the probability by which a match between two nucleotide or amino acid sequences would occur by chance.
  • a nucleic acid is considered similar to a reference sequence if the smallest sum probability in a comparison of the test nucleic acid to the reference nucleic acid is less than about 0.1, more preferably less than about 0.01, and most preferably less than about 0.001.
  • a further indication that two nucleic acid sequences or polypeptides are substantially identical is that the polypeptide encoded by the first nucleic acid is immunologically cross reactive with the polypeptide encoded by the second nucleic acid, as described below.
  • a polypeptide is typically substantially identical to a second polypeptide, for example, where the two peptides differ only by conservative substitutions.
  • isolated means a biological component (such as a nucleic acid, peptide or protein) has been substantially separated, produced apart from, or purified away from other biological components of the organism in which the component naturally 17 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 occurs, i.e., other chromosomal and extrachromosomal DNA and RNA, and proteins.
  • Nucleic acids, peptides and proteins that have been “isolated” thus include nucleic acids and proteins purified by standard purification methods. “Isolated” nucleic acids, peptides and proteins can be part of a composition and still be isolated if the composition is not part of the native environment of the nucleic acid, peptide, or protein. The term also embraces nucleic acids, peptides and proteins prepared by recombinant expression in a host cell as well as chemically synthesized nucleic acids.
  • nucleic acid molecule refers to any polyribonucleotide or polydeoxyribonucleotide, which can be unmodified RNA or DNA or modified RNA or DNA.
  • Polynucleotides include, without limitation single- and double-stranded DNA, DNA that is a mixture of single- and double-stranded regions, single- and double-stranded RNA, and RNA that is a mixture of single- and double-stranded regions, hybrid molecules comprising DNA and RNA that can be single-stranded or, more typically, double-stranded or a mixture of single- and double-stranded regions.
  • polynucleotide refers to triple-stranded regions comprising RNA or DNA or both RNA and DNA.
  • the term polynucleotide also includes DNAs or RNAs containing one or more modified bases and DNAs or RNAs with backbones modified for stability or for other reasons.
  • Modified bases include, for example, tritylated bases and unusual bases such as inosine.
  • polynucleotide embraces chemically, enzymatically or metabolically modified forms of polynucleotides as typically found in nature, as well as the chemical forms of DNA and RNA characteristic of viruses and cells.
  • Polynucleotide also embraces relatively short nucleic acid chains, often referred to as oligonucleotides.
  • the term “vector” is a replicon in which another nucleic acid segment can be operably inserted so as to bring about the replication or expression of the segment.
  • the term “host cell” refers to a cell comprising a nucleic acid molecule of the invention.
  • the “host cell” can be any type of cell, e.g., a primary cell, a cell in culture, or a cell from a cell line.
  • a “host cell” is a cell transfected with a nucleic acid molecule of the invention.
  • a “host cell” is a progeny or potential progeny of such a transfected cell.
  • a progeny of a cell may or may not be identical to the parent cell, e.g., due to mutations or environmental influences that can occur in succeeding generations or integration of the nucleic acid molecule into the host cell genome.
  • the term “expression” as used herein, refers to the biosynthesis of a gene product.
  • the term encompasses the transcription of a gene into RNA.
  • the term also encompasses 18 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 translation of RNA into one or more polypeptides, and further encompasses all naturally occurring post-transcriptional and post-translational modifications.
  • the expressed gene product can be within the cytoplasm of a host cell, into the extracellular milieu such as the growth medium of a cell culture or anchored to the cell membrane.
  • peptide can refer to a molecule comprised of amino acids and can be recognized as a protein by those of skill in the art.
  • the conventional one-letter or three-letter code for amino acid residues is used herein.
  • peptide can be used interchangeably herein to refer to polymers of amino acids of any length.
  • the polymer can be linear or branched, it can comprise modified amino acids, and it can be interrupted by non-amino acids.
  • the terms also encompass an amino acid polymer that has been modified naturally or by intervention; for example, disulfide bond formation, glycosylation, lipidation, acetylation, phosphorylation, or any other manipulation or modification, such as conjugation with a labeling component. Also included within the definition are, for example, polypeptides containing one or more analogs of an amino acid (including, for example, unnatural amino acids, etc.), as well as other modifications known in the art. [0086] The peptide sequences described herein are written according to the usual convention whereby the N-terminal region of the peptide is on the left and the C-terminal region is on the right.
  • the term “low-to-moderate affinity peptide” can refer to any peptide that does not have a high affinity for an MHC-1 complex.
  • a low affinity peptide can, for example, have an IC 50 value of >500 nM for MHC I.
  • a moderate affinity peptide can, for example, have an IC50 value of ⁇ 500 nM and >50 nM for MHC I.
  • a high affinity peptide generally has an IC 50 value of ⁇ 50 nM.
  • Low-to-moderate affinity peptides can be excluded from consideration as candidates for vaccines and can be referred to as a “non-binder.”
  • Low affinity peptides are known in the art, see, e.g., Ebrahimi-Nik et al., “Reversion Analysis Reveals the in vivo immunogenicity of a poorly MHC-I binding cancer neoepitope,” Nature Communications 12:6423 (2021).
  • administering means a method for simulating an immune response as described herein by using a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 95% identity to SEQ ID NO:10 and an immunogenic agent of the invention or a form, composition or medicament thereof.
  • TAPBPR TAP-binding protein-related
  • Such methods include administering an effective amount of the TAP- 19 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 95% identity to SEQ ID NO:10 and the immunogenic agent at different times during the course of a simulating the immune response or concurrently in a combination form.
  • TAPBPR binding protein-related
  • TAPBPR TAP-binding protein-related polypeptide comprising an amino sequence having at least 95% identity to SEQ ID NO:10 and the immunogenic agent that elicits the desired immune response in the subject, that is being sought by a researcher, veterinarian, medical doctor, or other clinician.
  • a high-fidelity chaperone, TAPBPR FLQ , containing three mutations (S104F, K211L, and R270Q) at MHC-I interaction sites was characterized and shown to significantly enhance peptide exchange for multiple HLA allotypes expressed on a cellular membrane.
  • the cryoEM structure of open HLA- A*02:01/ ⁇ 2 m loaded with a peptide decoy captured by TAPBPR FLQ was determined, revealing a key intermediate of the peptide exchange process.
  • the transient proofreading complex utilizes (1) conserved groove residues to capture the backbone of incoming peptides in a native-like conformation, (2) properly conformed hydrophobic A-, B-, and D-pockets to dock the peptide P2 and P3 anchors, and (3) a conformationally heterogenous F-pocket that folds upon binding of high-affinity peptides, allosterically triggering the dissociation of TAPBPR.
  • the combined structural and biochemical analysis provides a complete mechanism of MHC-I antigen proofreading and peptide repertoire selection.
  • the methods comprise providing a population of mammalian cells having a surface MHC class I molecule with an immunogenic agent and a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:7, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecules thereby increasing the 20 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 immunogenicity of the mammalian cells.
  • TAPBPR TAP-binding protein-related
  • the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11.
  • the method is performed in vivo. [0092] Also provided are methods of producing antigen presenting cells to stimulate an immune response in a subject in need thereof.
  • the methods comprise (a) obtaining antigen presenting cells from the subject in need thereof; and (b) providing antigen presenting cells comprising surface MHC class I molecules in vitro with a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:7 and an immunogenic agent, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecules thereby creating antigen presenting cells for stimulating an immune response.
  • TAPBPR TAP-binding protein-related
  • the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11.
  • the TAPBPR polypeptide further comprises a signal peptide, a flag tag, a linker, and/or a transmembrane domain.
  • the signal peptide can, for example, comprise SEQ ID NO:47.
  • the flag tag can, for example, comprise SEQ ID NO:48.
  • the linker can, for example, comprise SEQ ID NO:49.
  • the transmembrane domain can, for example, comprise SEQ ID NO:50.
  • the mammalian cell is a human cell.
  • the mammalian cell is a human cell.
  • a cell displaying MHC class I molecules can be exposed to (i) a soluble extracellular TAPBPR polypeptide as described herein; (ii) a cell having a surface bound TAPBPR polypeptide as described herein; or (iii) a chimeric TAPBPR as described in WO2023/163980, which is herein incorporated by reference in its entirety.
  • the loading of cell-surface MHC class I molecules as described herein can increase the number of MHC class I molecules on the surface of a cell which present the immunogenic agent relative to cells not treated with the TAPBPR.
  • the number of MHC class I molecules on the surface of a cell which present the immunogenic agent may be increased by 30 fold or more, 40 fold or more, 50 fold or more, 60 fold or more, 70 fold or more, 80 fold or more, 90 fold or more, 100 fold or more, 150 fold or more or 200 fold or more of the immunogenic agent in the presence relative to the absence of TAPBPR.
  • Cells may present none or substantially none of the immunogenic agent in the absence of treatment with the TAPBPR. 21 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 [0097]
  • Mammalian cells or cells of the subject may comprise MHC class I molecules on the cell surface.
  • MHC class I molecules are heterodimers comprising an ⁇ chain and ⁇ 2- microglobulin. MHC class I molecules are expressed on all nucleated human cells. An individual inherits a set of HLA-A, -B and -C genes from each parent. These genes are co- dominantly expressed and nucleated cells in mammals express up to 6 different classical MHC class I molecules. MHC class I molecules are highly polymorphic within the ⁇ chain and there is huge variation within the population. MHC class I molecules may include HLA- A molecules, HLA-B molecules, such as HLA-B51, HLA-Bl5, HLA-B38, and HLA-B57 and HLA-C molecules, such as HLA-Cwl. Preferred MHC class I molecules include HLA-A.
  • the MHC class I molecule is selected from the group consisting of an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, an HLA-E molecule, an HLA-G molecule, an HLA-F molecule, a MR1 molecule, a CD1 molecule, and a combination thereof.
  • the MHC class I molecule can, for example, be an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, a MR1 molecule, an HLA-E molecule, and/or an HLA-G molecule.
  • the HLA-A molecule, HLA-B molecule, and/or HLA-C molecule can, for example, be selected from an HLA-A01 supertype molecule, an HLA-A02 supertype molecule, an HLA-A03 supertype molecule, an HLA-A24 supertype molecule, an HLA- A01/A24 supertype molecule, an HLA-A01/A03 supertype molecule, an HLA-B08 supertype molecule, an HLA-B44 supertype molecule, an HLA-B58 supertype molecule, and/or an HLA-C01 supertype molecule.
  • the MHC class I molecule can, for example, be selected from HLA-A*01:01, HLA-A*11:01, HLA-A*29:01, HLA-A*30:01, HLA-A*32:01, HLA-A*74:01, HLA-A*24:03, HLA-A*02:06, HLA-A*02:01, HLA- A*02:03, HLA-A*23:01, HLA-A*24:02, HLA-A*68:01, HLA-A*69:01, HLA-A*68:02, HLA-A*29:02, HLA-A*03:01, HLA-B*08:01, HLA-B*57:03, HLA-B*59:01, HLA-B*37:01, HLA-C*01:02, HLA-C*03:04, MR1, HLA-E*01:03, and/or HLA-G*01:01.
  • the cells may be disease cells, such as cancer cells, cells infected with a pathogen, or other cells that cause disease. Increasing the immunogenicity of disease cells in a subject using a TAPBPR may generate or increase the strength of immune responses against the disease cells in the subject. This may lead to a reduction or eradication of disease cells in the individual and may exert a therapeutic effect.
  • the cells may be antigen presenting cells.
  • the antigen presenting cells are dendritic cells.
  • the loading of the surface MHC class I molecules with immunogenic peptides can increase the ability of the antigen presenting cells to induce immune responses, for example immune responses against the antigenic epitopes contained in the immunogenic peptide.
  • Antigen presenting cells loaded with immunogenic peptides as described above may be used to stimulate T cells in vitro or ex vivo or administered to an individual to stimulate T cells in vivo.
  • the immunogenic peptide can comprise one or more antigenic epitopes.
  • the antigen presenting cells can activate T cells against the antigenic epitopes of the immunogenic peptide.
  • the antigenic epitopes of the immunogenic peptide can be present on disease cells in the subject.
  • the antigen presenting cells can activate T cells capable of generating an immune response against the disease cells in the subject.
  • the method further comprises administering the antigen presenting cells to the subject in need thereof.
  • the immunogenic agent is selected from an immunogenic peptide, a metabolite, a lipid, a phospholipid, a fatty acid ligand, and/or combinations thereof.
  • the immunogenic peptides are low-to-moderate affinity peptides.
  • the immunogenic peptide may comprise an antigen or an epitope that is characteristic of a disease cell.
  • the immunogenic peptide may comprise an antigen or an epitope that is characteristic of a cancer cell or a pathogen-infected cell.
  • Epitopes that are characteristic of cancer cells are well known in the art and include epitopes from tumor antigens.
  • tumor antigens from which immunogenic peptides may be derived include neoantigens, tumor-specific, differentiation and overexpressed proteins, such as ErbB2/Her2 (e.g., RLLQETELV (SEQ ID NO:12)), gpl00 (e.g., IMDQVPFSV (SEQ ID NO:13) and YLEPGPVTA (SEQ ID NO:14)), NY-Eso-1 (e.g., SLLMWITQC (SEQ ID NO:15)), p53 (e.g., LLGRNSFEV (SEQ ID NO:16)), MARTI (e.g., ELAGIGILTV (SEQ ID NO:17)), MAGE-10 (e.g., GLYDGMEHL (SEQ ID NO:18)), human AFP (e.g., FMNKFIYEI (SEQ ID NO:19)), Mesothelin (e.g.
  • ErbB2/Her2 e.g., RLLQ
  • Suitable epitopes for immunogenic peptides can include influenza epitopes (e.g., GILGFVFTL (SEQ ID NO:26), AIMDKNIIL (SEQ ID NO:27)), HIV epitopes (e.g., ILKEPVHGV (SEQ ID NO:28), SLYNTVATL (SEQ ID NO:29), KLTPLCVTL (SEQ ID NO:30)), hepatitis B epitopes (e.g., FLPSDFFPSV (SEQ ID NO:31), WLSLLVPFV (SEQ ID NO:32)), Human cytomegalovirus (CMV) epitopes (e.g., NLVPMVATV (SEQ ID NO:33), VLEETSVML (SEQ ID NO:34)), Epstein Barr virus (EBV) epitopes (e.g., YLLEMLWRL (SEQ ID NO:35), CLGGLLTMV (SEQ ID NO:36)), Varicella-zoster virus epitop
  • TAPBPR TAP- binding protein-related polypeptide comprising an amino sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:7, wherein the TAPBPR polypeptide binds to an antigen presenting cell comprising a surface MHC class I molecule in the subject in need thereof; and (b) administering to the subject in thereof an immunogenic agent, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecule of the antigen presenting cell such that
  • the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11. [00110] In certain embodiments, the TAPBPR polypeptide further comprises a signal peptide, a flag tag, a linker, and/or a transmembrane domain.
  • the signal peptide can, for example, comprise SEQ ID NO:47.
  • the flag tag can, for example, comprise SEQ ID NO:48.
  • the linker can, for example, comprise SEQ ID NO:49.
  • the transmembrane domain can, for example, comprise SEQ ID NO:50.
  • the MHC class I molecule is selected from the group consisting of an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, an HLA-E molecule, an HLA-G molecule, an HLA-F molecule, a MR1 molecule, a CD1 molecule, and a combination thereof.
  • the MHC class I molecule can, for example, be an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, a MR1 molecule, an HLA-E molecule, and/or an HLA-G molecule.
  • the HLA-A molecule, HLA-B molecule, and/or HLA-C molecule can, for example, be selected from an HLA-A01 supertype molecule, an HLA-A02 supertype molecule, an HLA-A03 supertype molecule, an HLA-A24 supertype molecule, an HLA- A01/A24 supertype molecule, an HLA-A01/A03 supertype molecule, an HLA-B08 supertype molecule, an HLA-B44 supertype molecule, an HLA-B58 supertype molecule, and/or an HLA-C01 supertype molecule.
  • the MHC class I molecule can, for example, be selected from HLA-A*01:01, HLA-A*11:01, HLA-A*29:01, HLA-A*30:01, HLA-A*32:01, HLA-A*74:01, HLA-A*24:03, HLA-A*02:06, HLA-A*02:01, HLA- A*02:03, HLA-A*23:01, HLA-A*24:02, HLA-A*68:01, HLA-A*69:01, HLA-A*68:02, HLA-A*29:02, HLA-A*03:01, HLA-B*08:01, HLA-B*57:03, HLA-B*59:01, HLA-B*37:01, HLA-C*01:02, HLA-C*03:04, MR1, HLA-E*01:03, and/or HLA-G*01:01.
  • the subject is a human.
  • the immunogenic agent is selected from an immunogenic peptide, a metabolite, a lipid, a phospholipid, a fatty acid ligand, and/or combinations thereof.
  • the immunogenic peptides are low-to-moderate affinity peptides.
  • the antigen presenting cells are dendritic cells.
  • the subject in need thereof can have a disease or condition, such as a viral infection or other pathogenic infection, or cancer.
  • the subject has cancer.
  • a method of treatment of cancer in a subject can comprise administering the TAPBPR polypeptide to the subject, wherein TAPBPR polypeptide comprises a targeting domain which binds to cancer cells in the subject, and administering an immunogenic peptide to the subject, such that the TAPBPR polypeptide loads the immunogenic peptide onto surface MHC class I molecules of the cancer cells of the subject, thereby eliciting or increasing an immune response in the subject against the cancer cells.
  • Cancer can be characterized by the abnormal proliferation of malignant cancer cells and can include leukemias, such as AML, CML, ALL and CLL, lymphomas, such as Hodgkin lymphoma, non-Hodgkin lymphoma and multiple myeloma, and solid cancers such as sarcomas, skin cancer, melanoma, bladder cancer, brain cancer, breast cancer, uterus 25 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 cancer, ovary cancer, prostate cancer, lung cancer, colorectal cancer, cervical cancer, liver cancer, head and neck cancer, esophageal cancer, pancreas cancer, renal cancer, adrenal cancer, stomach cancer, testicular cancer, cancer of the gall bladder and biliary tracts, thyroid cancer, thymus cancer, cancer of bone, and cerebral cancer, as well as cancer of unknown primary (CUP).
  • leukemias such as AML, C
  • cancer cells within a subject can be immunologically distinct from normal somatic cells in the subject (i.e., the cancerous tumor may be immunogenic).
  • the cancer cells can be capable of eliciting a systemic immune response in the subject against one or more antigens expressed by the cancer cells.
  • the tumor antigens that elicit the immune response can be specific to cancer cells or can be shared by one or more normal cells in the individual.
  • cancer cells within a subject cannot be immunologically distinct from normal somatic cells in the subject until MHC class I molecules on the surface of the cancer cells are loaded with exogenous immunogenic peptide using a TAPBPR polypeptide as described herein.
  • Cancer growth generally refers to any one of a number of indices that indicate change within the cancer to a more developed form.
  • indices for measuring an inhibition of cancer growth include a decrease in cancer cell survival, a decrease in tumor volume or morphology (for example, as determined using computed tomographic (CT), sonography, or other imaging method), a delayed tumor growth, a destruction of tumor vasculature, improved performance in delayed hypersensitivity skin test, an increase in the activity of T cells, and a decrease in levels of tumor-specific antigens.
  • Administration of modified T cells can improve the capacity of the subject to resist cancer growth, in particular growth of a cancer already present in the subject and/or decrease the propensity for cancer growth in the subject.
  • a method of treatment of a viral or pathogenic infection in a subject can comprise administering a TAPBPR polypeptide described above to the subject, wherein the TAPBPR polypeptide comprises a targeting domain which binds to the viral or pathogen-infected cells in the subject, and administering an immunogenic peptide to the subject, such that the TAPBPR polypeptide loads the immunogenic peptide onto surface MHC class I molecules of the viral or pathogen-infected cells of the subject, thereby eliciting or increasing an immune response in the subject against the viral or pathogen-infected cells.
  • Viral infection can, for example, be HIV, EBV, CMV, hepatitis, influenza, polio, human papilloma virus, measles, mumps, rubella, chicken pox, ebola, or zika infection.
  • TAPBPR polypeptides [00124] Also provided is an isolated TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:7.
  • the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11. [00125] In certain embodiments, the TAPBPR polypeptide further comprises a signal peptide, a flag tag, a linker, and/or a transmembrane domain.
  • the signal peptide can, for example, comprise SEQ ID NO:47.
  • the flag tag can, for example, comprise SEQ ID NO:48.
  • the linker can, for example, comprise SEQ ID NO:49.
  • the transmembrane domain can, for example, comprise SEQ ID NO:50.
  • TAPBPR polypeptides load MHC class I molecules on the surface of the cells with an immunogenic agent.
  • the immunogenic agent can, for example, be an immunogenic peptide.
  • the immunogenic peptide can, for example, be an exogenous peptide.
  • An exogenous peptide is a peptide that is not generated naturally by the cells with the MHC class I molecules. For example, it may have been administered to the subject. Exogenous peptide may have the same amino acid sequence as an endogenous peptide that is generated naturally by the cells or a different amino acid sequence.
  • the immunogenicity of the exogenous peptide may be different to the immunogenicity of endogenous peptides displayed in the MHC class I molecules (i.e., it may be higher or lower). In other embodiments, the immunogenicity of the exogenous peptide may be the same as the immunogenicity of one or more endogenous peptides displayed in the MHC class I molecules. For example, the exogenous peptide may have the same amino acid sequence as one or more endogenous peptides.
  • An immunogenic peptide is an exogenous peptide that is capable of generating an immune response in a subject when loaded onto an MHC class I molecule.
  • the immunogenic peptide/MHC class I complex may be recognized by T cells.
  • the presence of MHC class I molecules loaded with an immunogenic peptide on the surface of target cells may induce or increase immune responses against the target cells.
  • immunogenic peptides are known in the art and may for example be candidates in vaccines for cancer or infection.
  • immunogenic peptides for loading onto MHC class I may be antigens naturally expressed on a subject’s own tumor; neoantigens or other peptides derived from tumors; or peptides derived from pathogens, such as viruses.
  • the invention relates to isolated nucleic acids encoding the TAPBPR polypeptides disclosed herein. It will be appreciated by those skilled in the art that the coding sequence of a protein can be changed (e.g., replaced, deleted, inserted, etc.) without changing the amino acid sequence of the protein.
  • nucleic acid sequences encoding TAPBPR polypeptides of the invention can be altered without changing the amino acid sequences of the proteins.
  • the invention relates to vectors comprising the isolated nucleic acids disclosed herein. Any vector known to those skilled in the art in view of the present disclosure can be used, such as a plasmid, a cosmid, a phage vector or a viral vector.
  • the vector is a recombinant expression vector such as a plasmid.
  • the vector can include any element to establish a conventional function of an expression vector, for example, a promoter, ribosome binding element, terminator, enhancer, selection marker, and origin of replication.
  • the promoter can be a constitutive, inducible, or repressible promoter.
  • a number of expression vectors capable of delivering nucleic acids to a cell are known in the art and can be used herein for production of a TAPBPR polypeptide in the cell. Conventional cloning techniques or artificial gene synthesis can be used to generate a recombinant expression vector according to embodiments of the invention.
  • the invention relates to a host cell comprising an isolated nucleic acid or a vector comprising a nucleic acid as disclosed herein.
  • any host cell known to those skilled in the art in view of the present disclosure can be used for recombinant expression of TAPBPR polypeptides of the invention.
  • the host cells 28 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 are E. coli TG1 or BL21 cells (for expression of, e.g., an scFv or Fab antibody), CHO-DG44 or CHO-K1 cells or HEK293 cells (for expression of, e.g., a full-length IgG antibody).
  • the recombinant expression vector is transformed into host cells by conventional methods such as chemical transfection, heat shock, or electroporation, where it is stably integrated into the host cell genome such that the recombinant nucleic acid is effectively expressed.
  • the host cell is a mammalian cell.
  • Pharmaceutical Compositions [00133]
  • the TAPBPR polypeptide and/or the immunogenic agent can be administered alone, or usually the TAPBPR polypeptide and/or the immunogenic agent can be administered in the form of a pharmaceutical composition, which may comprise at least one component in addition to the TAPBPR polypeptide and/or the immunogenic agent.
  • compositions can comprise, in addition to the TAPBPR polypeptide and/or the immunogenic agent, a pharmaceutically acceptable excipient, carrier, buffer, stabilizer or other materials well known to those skilled in the art.
  • pharmaceutically acceptable as used herein pertains to compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgement, suitable for use in contact with the tissues of a subject (e.g., human) without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio.
  • Each carrier, excipient, etc. must also be “acceptable” in the sense of being compatible with the other ingredients of the formulation.
  • the precise nature of the carrier or other material will depend on the route of administration, which may be by bolus, infusion, injection or any other suitable route.
  • the TAPBPR polypeptide and/or the immunogenic agent can be administered in combination.
  • the TAPBPR polypeptide and/or the immunogenic agent can be formulated in the same pharmaceutical composition.
  • the TAPBPR polypeptide and/or the immunogenic agent can be formulated in separate pharmaceutical compositions.
  • the TAPBPR polypeptide and/or the immunogenic agent can be provided in a lyophilized form for reconstitution prior to administration.
  • a lyophilized TAPBPR polypeptide and/or immunogenic agent can be re-constituted in sterile water and mixed with saline prior to administration to an individual.
  • parenteral for example sub-cutaneous, intra-tumoral, intra-muscular or intra-venous administration, e.g., by injection, the pharmaceutical composition comprising 29 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 the TAPBPR polypeptide and/or the immunogenic agent described herein, nucleic acid or cell can be in the form of a parenterally acceptable aqueous solution which is pyrogen-free and has suitable pH, isotonicity and stability.
  • isotonic vehicles such as Sodium Chloride Injection, Ringers Injection, and Lactated Ringers Injection.
  • Preservatives, stabilizers, buffers, antioxidants and/or other additives may be employed as required including buffers such as phosphate, citrate and other organic acids; antioxidants, such as ascorbic acid and methionine; preservatives (such as octadecyldimethylbenzyl ammonium chloride; hexamethonium chloride; benzalkonium chloride; benzethonium chloride; phenol, butyl or benzyl alcohol; alkyl parabens, such as methyl or propyl paraben; catechol; resorcinol; cyclohexanol; 3'- pentanol; and m-cresol); low molecular weight polypeptides; proteins, such as serum albumin, gelatin or immunoglobulfate, and others.
  • compositions and formulations can conveniently be presented in unit dosage form and can be prepared by any methods well known in the art of pharmacy. Such methods include the step of bringing into association the chimeric protein described herein with the carrier which constitutes one or more accessory ingredients.
  • compositions are prepared by uniformly and intimately bringing into association the active compound with liquid carriers or finely divided solid carriers or both, and then, if necessary, shaping the product.
  • a pharmaceutical composition comprising the TAPBPR polypeptide and/or the immunogenic agent, as described herein, can be administered alone or in combination with other treatments, either simultaneously or sequentially dependent upon the condition to be treated.
  • EMBODIMENTS [00139] The invention provides also the following non-limiting embodiments.
  • Embodiment 1 is a method of increasing the immunogenicity of mammalian cells, the method comprising providing a population of mammalian cells having surface MHC class I molecule with an immunogenic agent and a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecules thereby increasing the immunogenicity of the mammalian cells.
  • TAPBPR TAP-binding protein-related
  • Embodiment 2 is the method of embodiment 1, wherein the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11.
  • Embodiment 3 is the method of embodiment 1 or 2, wherein the MHC class I molecule is selected from the group consisting of an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, an HLA-E molecule, an HLA-G molecule, an HLA-F molecule, a MR1 molecule, a CD1 molecule, and a combination thereof.
  • Embodiment 4 is the method of embodiment 3, wherein the MHC class I molecule is an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, a MR1 molecule, an HLA-E molecule, and/or an HLA-G molecule.
  • Embodiment 5 is the method of embodiment 4, wherein the HLA-A molecule, HLA- B molecule, and/or HLA-C molecule is selected from an HLA-A01 supertype molecule, an HLA-A02 supertype molecule, an HLA-A03 supertype molecule, an HLA-A24 supertype molecule, an HLA-A01/A24 supertype molecule, an HLA-A01/A03 supertype molecule, an HLA-B08 supertype molecule, an HLA-B44 supertype molecule, an HLA-B58 supertype molecule, and/or an HLA-C01 supertype molecule.
  • Embodiment 6 is the method of embodiment 4, wherein the MHC class I molecule is selected from HLA-A*01:01, HLA-A*11:01, HLA-A*29:01, HLA-A*30:01, HLA-A*32:01, HLA-A*74:01, HLA-A*24:03, HLA-A*02:06, HLA-A*02:01, HLA-A*02:03, HLA- A*23:01, HLA-A*24:02, HLA-A*68:01, HLA-A*69:01, HLA-A*68:02, HLA-A*29:02, HLA-A*03:01, HLA-B*08:01, HLA-B*57:03, HLA-B*59:01, HLA-B*37:01, HLA-C*01:02, HLA-C*03:04, MR1, HLA-E*01:03, and/or HLA-G*01
  • Embodiment 7 is the method of any one of embodiments 1-6, wherein the mammalian cell is a human cell.
  • Embodiment 8 is the method of any one of embodiments 1-7, wherein the immunogenic agent is selected from an immunogenic peptide, a metabolite, a lipid, a phospholipid, a fatty acid ligand, and/or combinations thereof. 31 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1
  • Embodiment 9 is the method of any one of embodiments 1-8, wherein the method is performed in vivo.
  • Embodiment 10 is a method of producing antigen presenting cells to stimulate an immune response in a subject, the method comprising: (a) obtaining antigen presenting cells from the subject; and (b) providing antigen presenting cells comprising surface MHC class I molecules in vitro with a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7 and an immunogenic agent, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecules thereby creating antigen presenting cells for stimulating an immune response.
  • TAPBPR TAP-binding protein-related
  • Embodiment 11 is the method of embodiment 10, wherein the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11.
  • Embodiment 12 is the method of embodiment 10 or 11, wherein the MHC class I molecule is selected from the group consisting of an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, an HLA-E molecule, an HLA-G molecule, an HLA-F molecule, a MR1 molecule, a CD1 molecule, and a combination thereof.
  • Embodiment 13 is the method of embodiment 12, wherein the MHC class I molecule is an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, a MR1 molecule, an HLA-E molecule, and/or an HLA-G molecule.
  • Embodiment 14 is the method of embodiment 13, wherein the HLA-A molecule, HLA-B molecule, and/or HLA-C molecule is selected from an HLA-A01 supertype molecule, an HLA-A02 supertype molecule, an HLA-A03 supertype molecule, an HLA-A24 supertype molecule, an HLA-A01/A24 supertype molecule, an HLA-A01/A03 supertype molecule, an HLA-B08 supertype molecule, an HLA-B44 supertype molecule, an HLA-B58 supertype molecule, and/or an HLA-C01 supertype molecule.
  • Embodiment 15 is the method of embodiment 13, wherein the MHC class I molecule is selected from HLA-A*01:01, HLA-A*11:01, HLA-A*29:01, HLA-A*30:01, HLA- A*32:01, HLA-A*74:01, HLA-A*24:03, HLA-A*02:06, HLA-A*02:01, HLA-A*02:03, HLA-A*23:01, HLA-A*24:02, HLA-A*68:01, HLA-A*69:01, HLA-A*68:02, HLA- A*29:02, HLA-A*03:01, HLA-B*08:01, HLA-B*57:03, HLA-B*59:01, HLA-B*37:01, HLA-C*01:02, HLA-C*03:04, MR1, HLA-E*01:03, and/or HLA-G*01
  • Embodiment 16 is the method of any one of embodiments 10-15, wherein the mammalian cell is a human cell.
  • Embodiment 17 is the method of any one of embodiments 10-16, wherein the immunogenic agent is selected from an immunogenic peptide, a metabolite, a lipid, a phospholipid, a fatty acid ligand, and/or combinations thereof.
  • Embodiment 18 is the method of any one of embodiments 10-17, wherein the antigen presenting cells are dendritic cells.
  • Embodiment 19 is the method of any one of embodiments 10-18, wherein the method further comprises administering the antigen presenting cells to the subject.
  • Embodiment 20 is a method of stimulating an immune response in a subject in need thereof, the method comprising: (a) administering to the subject in need thereof a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7, wherein the TAPBPR polypeptide binds to an antigen presenting cell comprising a surface MHC class I molecule in the subject; and (b) administering to the subject an immunogenic agent, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecule of the antigen presenting cell such that the antigen presenting cell stimulates an immune response in the subject.
  • TAPBPR TAP-binding protein-related
  • Embodiment 21 is the method of embodiment 20, wherein the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11.
  • Embodiment 22 is the method of embodiment 20 or 21, wherein the MHC class I molecule is selected from the group consisting of an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, an HLA-E molecule, an HLA-G molecule, an HLA-F molecule, a MR1 molecule, a CD1 molecule, and a combination thereof.
  • Embodiment 23 is the method of embodiment 22, wherein the MHC class I molecule is an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, a MR1 molecule, an HLA-E molecule, and/or an HLA-G molecule.
  • Embodiment 24 is the method of embodiment 23, wherein HLA-A molecule, HLA- B molecule, and/or HLA-C molecule is selected from an HLA-A01 supertype molecule, an HLA-A02 supertype molecule, an HLA-A03 supertype molecule, an HLA-A24 supertype molecule, an HLA-A01/A24 supertype molecule, an HLA-A01/A03 supertype molecule, an 33 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 HLA-B08 supertype molecule, an HLA-B44 supertype molecule, an HLA-B58 supertype molecule, and/or an HLA-C01 supertype molecule.
  • Embodiment 25 is the method of embodiment 23, wherein the MHC class I molecule is selected from HLA-A*01:01, HLA-A*11:01, HLA-A*29:01, HLA-A*30:01, HLA- A*32:01, HLA-A*74:01, HLA-A*24:03, HLA-A*02:06, HLA-A*02:01, HLA-A*02:03, HLA-A*23:01, HLA-A*24:02, HLA-A*68:01, HLA-A*69:01, HLA-A*68:02, HLA- A*29:02, HLA-A*03:01, HLA-B*08:01, HLA-B*57:03, HLA-B*59:01, HLA-B*37:01, HLA-C*01:02, HLA-C*03:04, MR1, HLA-E*01:03, and/or HLA-G*01
  • Embodiment 26 is the method of any one of embodiments 20-25, wherein the subject is a human.
  • Embodiment 27 is the method of any one of embodiments 20-26, wherein the immunogenic agent is selected from an immunogenic peptide, a metabolite, a lipid, a phospholipid, a fatty acid ligand, and/or combinations thereof.
  • Embodiment 28 is the method of any one of embodiments 20-27, wherein the antigen presenting cells are dendritic cells.
  • Embodiment 29 is the method of any one of embodiments 1-28, wherein the TAPBPR polypeptide further comprises a signal peptide, a flag tag, a linker, and/or a transmembrane domain.
  • Embodiment 30 is the method of embodiment 29, wherein: (a) the signal peptide comprises SEQ ID NO:47; (b) the flag tag comprises SEQ ID NO:48; (c) the linker comprises SEQ ID NO:49; and (d) the transmembrane domain comprises SEQ ID NO:50.
  • Embodiment 31 is an isolated TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 95% identity to SEQ ID NO:10 or at least 90% identity to SEQ ID NO:7.
  • Embodiment 32 is the isolated TAPBPR polypeptide of embodiment 31, wherein the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO: 9, SEQ ID NO:10, or SEQ ID NO:11.
  • Embodiment 33 is the isolated TAPBPR polypeptide of embodiment 31 or 32, wherein the TAPBPR polypeptide further comprises a signal peptide, a flag tag, a linker, and/or a transmembrane domain.
  • Embodiment 34 is the isolated TAPBPR polypeptide of embodiment 33, wherein: (a) the signal peptide comprises SEQ ID NO:47; 34 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 (b) the flag tag comprises SEQ ID NO:48; (c) the linker comprises SEQ ID NO:49; and (d) the transmembrane domain comprises SEQ ID NO:50. [00174] Embodiment 35 is an isolated nucleic acid encoding the TAPBPR polypeptide of any one of embodiments 30-34. [00175] Embodiment 36 is an isolated vector comprising the isolated nucleic acid of embodiment 35.
  • Embodiment 37 is a host cell comprising the vector of embodiment 36.
  • Embodiment 34 is the host cell of embodiment 37, wherein the cell is a mammalian cell.
  • EXAMPLES [00179] The following Materials and Methods are used in Examples discussed below. Materials and Methods A. Peptides [00180] All peptide sequences are given as standard single-letter codes.
  • High-affinity peptide TAX9 (LLFGYPVYV) (SEQ ID NO:1) was purchased from Genscript, USA; Piscataway, NJ, at >90% purity.
  • the fluorophore-labeled and photolabile peptides using TAMRA as 5- Carboxytetramethylrhodamine and J as Fmoc-3-amino-3-(2-nitrophenyl)-propionic acid, were purchased from Biopeptek Inc, Malvern, USA. Peptides were solubilized in distilled water and centrifuged at 14000 rpm for 15 minutes. The concentration of each peptide solution was measured and calculated using the absorbance and extinction coefficient at 205 nm wavelength.
  • TAMRA 5- Carboxytetramethylrhodamine
  • J Fmoc-3-amino-3-(2-nitrophenyl)-propionic acid
  • Plasmids encoding the luminal domain of MHC-I heavy chain HLA-A*02:01 and light chain human ⁇ 2 m were provided by the NIH tetramer facility (Emory University; Atlanta, GA). Plasmids encoding open HLA-A*02:01 (G120C) and open ⁇ 2m (G120C) were site mutated in-house.
  • HLA-A*02:01 For the generation of peptide-loaded HLA-A*02:01 (pHLA-A*02:01) molecules, in vitro refolding was performed by slowly diluting a 200 mg mixture of MHC-I heavy chain and ⁇ 2m at a 1:3 molar ratio over 24 hours in refolding buffer (0.4 M L-Arginine HCl, 100 mM Tris pH 8, 2 mM EDTA, 4.9 mM reduced L-glutathione, 0.57 mM oxidized L- glutathione) containing 10 mg of the desired peptide. The mixture was protected from light when refolded with photolabile peptides.
  • the secreted TAPBPR molecules were purified using a high-density metal affinity agarose resin (ABT, Madrid).
  • the eluted proteins were further purified by SEC using a HiLoad 16/600 Superdex 200 pg column at a 1 mL/min flow rate in 150 mM NaCl, 20 mM sodium phosphate buffer, pH 7.4.
  • TAPBPR protein or pMHC-I molecules (7 ⁇ M) were mixed with 10X SYPRO Orange dye in a buffer of 150 mM NaCl and 20 mM sodium phosphate (pH 7.2) to a final volume of 20 ⁇ l.
  • Samples were loaded into a MicroAmp Optical 384-well plate and run in triplicates. The experiment was performed on a QuantStudio 5 real-time polymerase chain reaction (PCR) machine with excitation and emission wavelengths set to 470 and 569nm.
  • the thermal stabilities of TAPBPR or pMHC-I were generated by plotting the first derivative of each melting curve and extracting the peak as the melting temperature (Tm).
  • the chip was primed by flowing over 1 ⁇ M TAX9 peptide solution before each binding assay to prevent peptide 36 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 release from the pMHC-I molecules.
  • Various concentrations from 0.1 up to 300 ⁇ M of TAPBPR and its variants were injected onto the chip at 25 ⁇ C at a flow rate of 30 ⁇ L/min for 60s followed by 180s dissociation time.
  • the SPR sensorgrams and equilibrium dissociation constants (K D ) were analyzed using the surface-bound analysis settings in Biacore X100 evaluation software (Cytiva).
  • TAMRA-TAX9/HLA-A*02:01 protein at a concentration of 40 nM in the presence of 1 ⁇ M non-labeled TAX9 peptide was incubated without chaperone or with 1 ⁇ M TAPBPR TN6 , TAPBPR WT , TAPBPR ⁇ G24-R36 , TAPBPR FLQ , and TAPBPR FLQ ⁇ G24-R36 .
  • the kinetic dissociations were monitored for 10 hours, and fluorescence polarization were recorded approximately every 90 seconds. Excitation and emission values used to monitor the fluorescence of TAMRA-labeled peptides were 531 and 595 nm. All experiments were performed at RT in triplicates.
  • beads were incubated with the anti– HLA class I antibody W6/32 (Abcam, ab22432) at 550 rpm for 30 minutes at RT and washed three times before tetramer addition.
  • W6/32 anti– HLA class I antibody
  • Abcam, ab22432 anti– HLA class I antibody
  • the levels of TAPBPR bound to the beads were 37 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 measured using the Luminex 100 Liquid Array Analyzer System, and the results were analyzed in GraphPad Prism v10. G.
  • target cell lines were seeded at 200,000 cells per well for peptide exchange. These cells were then treated with recombinant TAPBPR FLQ , TAPBPR WT , and TAPBPR TN6 at different concentrations, as indicated, for 15 minutes at 37 °C.
  • Cells were split every 1-2 days for up to 4 days before analysis by flow cytometry for TAPBPR expression for TAPBPR FLQ , TAPBPR WT , and TAPBPR TN6 -TM transduced cell lines.
  • Cells with similar levels of transduction efficiency and TAPBPR expression were selected for downstream assays, as measured by staining for FLAG (Clone: L5 in APC).
  • Expression levels of cell surface A02 were quantified by staining with an anti- HLA-A2 antibody (Clone: BB7.2 in Brilliant Violet 785).
  • T2 cells transduced with TAPBPR variants were incubated with indicated fluorescent TAX9-TAMRA peptide conjugate concentrations for 60 minutes in serum-free DMEM.
  • the mixture was UV-irradiated at a high intensity in 150 mM NaCl, 20 mM sodium phosphate, pH 7.4 at 4 ⁇ C for 40 minutes with a wavelength of 365 nm.
  • the complex was then purified by SEC using a Superdex 200 pg Increase 10/300 GL, and the eluted peaks were further analyzed by SDS–polyacrylamide gel electrophoresis (PAGE) to identify all components and confirm the complex.
  • the open HLA-A*02:01/TAPBPR FLQ complex was then prepared at a 0.2 mg/mL concentration with 1XPBS, pH 7.4 for cryoEM. J.
  • the HLA-A*02:01: TAPBPR FLQ ratios were: 1: 0, 1: 0.55, 1: 1.10, 1: 1.65, 1: 2.75, and 1: 3.31.
  • Two-dimensional methyl 1 H- 13 C methyl SOFAST HMQC experiments were recorded at 298 K at a field strength of 600 MHz with a total number of 136 scans, a recycle delay (d1) of 0.2 s and an acquisition time of 30 milliseconds in the indirect dimension.
  • LC-MS Liquid chromatography-mass spectroscopy
  • CryoEM sample preparation and data collection [00193] The CryoEM sample was prepared as previously described. Briefly, the open peptide-loaded HLA-A*02:01/TAPBPR FLQ complex was obtained by mixing open HLA- A*02:02/TAX9 and TAPBPR FLQ at a 1:1.3 molar ratio. The mixture was incubated at 4 ⁇ C for 1 hour and followed by 40-minute UV irradiation, and the complex was purified by size- exclusion chromatography. The peak fraction at 0.2 mg/mL concentration was used for grid preparation.
  • the sample was applied to freshly plasma cleaned Quantifoil Cu 300 2/2 (Quantifoil; Germany) grids and was plunge frozen in liquid ethane using the Vitroblot Mark IV (Thermo Fisher; Waltham, MA) operated at 4°C and 100% humidity.
  • the dataset was collected on a Titan Krios G3i 300 kV electron microscope (Thermo Fisher Scientific) with a 20 eV energy filter and equipped with a K3 Summit camera (Gatan).
  • Super-resolution images were collected over thirty-five frames with a dose of 40.5 e-/A 2 at a nominal magnification of x105,000, resulting in a pixel size of 0.418 ⁇ /pixel.
  • CryoEM data processing was performed using Relion 5.0 (6) and cryoSPARC 4.5 (7). 5,381 collected movies were motion corrected using the Relion implementation of MotionCor2 (8), binning to a pixel size of 0.836 ⁇ /pixel. The defocus values were determined using CTFFIND-4 (9) and then suboptimal micrographs were removed, resulting in a set of 4,791 micrographs. From this point, processing proceeded in cryoSPARC.
  • Templates for particle picking were generated using blob picker and 2D classification on a subset of the dataset, which was also used to generate a good ab initio model. Template picking resulted in 4,707,523 particles, which were extracted and binned to a pixel size of 3.344 ⁇ /pix. These particles were split evenly into five subsets and each subset was subjected to hetero refinement, using a single copy of the good ab initio model and five copies of a noise volume. After this first round of hetero refinement, a collective 1,119,002 particles were sorted into the good class.
  • 340,226 particles were re-extracted to their final box size of 0.836 A/pixel and subjected to non-uniform refinement (10), resulting in a 3.3 ⁇ structure. These particles were then transferred to Relion and reconnected to the original motion corrected micrographs. The 340,226 particles were refined in Relion using Blush regularization (11), resulting in a 3.3 ⁇ structure. The use of Blush regularization significantly improved the map quality compared to cryoSPARC refinement or Relion refinement without Blush regularization. The particles were then subjected to two rounds of CTF refinement (12) and Bayesian polishing, resulting in a 3.0 ⁇ structure.
  • TAPBPR FLQ shows enhanced editing of peptide-loaded MHC-I
  • contradictory peptide unloading/levering (38, 39) or trapping (40) mechanisms have been proposed involving the TAPBPR G24-R36 loop, whether the loop actively participates in cargo editing of peptide-loaded MHC-I molecules remains unclear (35). This controversy was sought to be resolved and TAPBPR regions that promote interactions with peptide-loaded molecules for antigen editing were sought to be identified.
  • nanomolar soluble TAPBPR FLQ can promote loading of exogenous antigens, albeit the use of a higher peptide concentration relative to the other HLA allotypes (FIG.8B, FIG.10D, and FIG. 14).
  • TAPBPR FLQ -TM (18) expressed on the cell surface of TAP transporter deficient T2 cells that lack the endogenous antigen processing machinery can directly enhance the loading of exogenous, fluorescently labeled peptides (FIG. 8C) was evaluated.
  • TAPBPR FLQ can promote exchange at significantly lower peptide concentrations relative to T2 cells expressing TAPBPR WT -TM (FIG. 8D, FIG. 43 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 15). It was further demonstrated that surface-expressed TAPBPR FLQ -TM increased the expression level of HLA-A*02:01 on the T2 cell surface, as indicated by staining with the allele-specific BB7.2 anti-human A2 antibody PE (FIG. 16).
  • Example 3 CryoEM structure of a TAPBPR/MHC-I complex bound to a peptide decoy [00201]
  • the enhanced affinity of the TAPBPR FLQ mutant for HLA- I may enable the capture of transient low-affinity peptide-editing complexes that would not be resolvable with TAPBPR WT .
  • the enhanced peptide editor, TAPBPR FLQ was leveraged.
  • a tertiary complex prepared using open HLA-A*02:01 (G120C) refolded with ⁇ 2m (H31C) and a photocleavable peptide (KILGFVFJV (SEQ ID NO:46), J 3-amino-3-(2-nitrophenyl)- propionic acid), used here as a conditional ligand (43, 44) was isolated.
  • KILGFVFJV photocleavable peptide
  • J 3-amino-3-(2-nitrophenyl)- propionic acid
  • the ⁇ 2-1 helix comprising residues E148-H151, adopted a conformation that was widened by approximately 3 ⁇ in the TAPBPR-bound complex relative to the fully loaded pMHC-I structure (measured at the C ⁇ atom of A150, FIG. 24B), in line with observations made on the basis of empty MHC-I/TAPBPR complexes (27, 28).
  • the cryoEM structure revealed a significant alteration of the ⁇ 1 helix and adjacent loop (residues G71-G91) (FIG.
  • cryo- EM structure demonstrated that the initial capture of peptides for proofreading by TAPBPR occurred through native-like interactions with conserved MHC-I residues within the A-, B-, and D-pockets that were properly conformed to receive peptide.
  • the F-pocket was partially formed in this intermediate state of the complex and folded upon annealing of the C-terminus of high-affinity peptides with robust P9 anchors to allosterically promote the dissociation of TAPBPR from the pMHC-I complex (31).
  • TABBPR widened the peptide binding groove via the ⁇ 2-1 helix and enhanced the dynamic movements alongside the ⁇ 1 helix and F pocket.
  • the key peptide-coordinating residues within the E- and F-pockets lose their interactions with the peptide C-terminus. Therefore, TAPBPR-bound MHC-I exhibits an overall lower affinity towards incoming peptides and promotes the dissociation of transiently bound peptide decoys.

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Abstract

Provided herein are methods of increasing the immunogenicity of mammalian cells, methods of producing antigen presenting cells to stimulate an immune response in a subject, and methods of stimulating an immune response in a subject in need thereof. The methods comprise providing mammalian cells, antigen presenting cells, or the subject with an immunogenic agent and a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7. wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecules thereby increasing the immunogenicity of the mammalian cells, producing antigen presenting cells to stimulate an immune response, and stimulating an immune response in a subject.

Description

CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 METHODS OF STIMULATING AN IMMUNE RESPONSE CROSS-REFERENCE TO RELATED APPLICATIONS [0001] This application claims priority to U.S. Provisional Application 63/674,123 (filed on July 22, 2024) which is incorporated by reference in its entirety. FIELD OF THE INVENTION [0002] This invention relates to the methods of increasing the immunogenicity of mammalian cells and methods of stimulating an immune response in a subject in need thereof. The methods comprise contacting a population of mammalian cells or the subject in need thereof with an immunogenic agent and a TAP-binding protein-related (TAPBPR) polypeptide. FEDERAL FUNDING LEGEND [0003] This invention was made with government support under Grant Nos. AI143997 and GM125034 awarded by the National Institutes of Health. The government has certain rights in the invention. REFERENCE TO SEQUENCE LISTING SUBMITTED ELECTRONICALLY [0004] This application contains a sequence listing, which is submitted electronically. The contents of the electronic sequence listing (074313.9WO1 Sequence Listing.xml; Size: 47,710 bytes; and Date of Creation: July 9, 2025) is herein incorporated by reference in its entirety. BACKGROUND OF THE INVENTION [0005] Class I major histocompatibility complex (MHC-I) proteins display the intracellular proteome onto the cell surface for immunosurveillance by sampling a large pool of peptide fragments (1-3). Optimal high-affinity-peptide-loaded MHC-I (pMHC-I) molecules are recognized by T cell receptors (TCR) and natural killer (NK) receptors, triggering downstream cellular activation and clonal expansion and resulting in the clearance of infected, or aberrant cells (4-6). While the immunogenic peptide repertoires of the Class I Human Leukocyte Antigen (HLA, human MHC-I) proteins are defined by their extremely polymorphic grooves (7-9), selection and optimization of the repertoires in the cellular pathway are mediated by two dedicated molecular chaperones, tapasin (10-12) and TAP binding protein-related (TAPBPR) (13, 14). TAPBPR is a homolog of tapasin that is not part of the peptide loading complex (PLC). These chaperones serve as an essential quality control checkpoint for pMHC-I molecules, further optimizing the peptide cargo of the MHC-I molecules en route to the cell surface (15, 16). Polymorphic residues at the MHC-I groove and chaperone interaction 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 surfaces confer a wide range of dependence on chaperones for peptide loading and cell surface expression across various HLA allotypes, with important ramifications for viral control among different individuals (17-21). Antigen processing and presentation (APP) chaperones have more recently emerged as key components for the presentation of metabolite ligands on MHC- related 1 (MR1) molecules (22-24), and have provided powerful tools for in vitro applications, including the generation of pMHC-I libraries encompassing different peptide specificities or the loading of antigens on cells independently of the endogenous processing pathway (18, 25, 26). [0006] Two independently solved crystal structures of mouse MHC-I bound to human TAPBPR (27, 28), together with studies of TAPBPR and Tapasin in complex with human MHC-I molecules by X-ray crystallography, nuclear magnetic resonance (NMR) spectroscopy, and cryogenic electron microscopy (cryo-EM) (29-31, 21, 32-34) have provided key insights into the chaperoning mechanism. Empty MHC-I adopts an open, peptide-receptive conformation where the short α2-1 helix shifted outwards by 3 Å to induce a widened peptide binding groove (27, 28). Based on these structures obtained for the empty, resting state of the complex, it has been proposed that the binding of high-affinity peptides triggers the closure of the MHC-I groove, which allosterically promotes the release of pMHC-I from the chaperone (27, 28, 31). However, the precise molecular mechanism of antigen exchange by TAPBPR remains enigmatic (35, 36). A range of biophysical studies have demonstrated that the affinity of TAPBPR for MHC-I is negatively correlated with peptide occupancy (18, 21, 30). TAPBPR preferentially interacts with nascent or suboptimal peptide-loaded MHC-I molecules, exchanging low- for high-affinity peptides. Due to the transient nature of these complexes, there is currently no structural visualization of the intermediate peptide/MHC-I/TAPBPR complex, precluding a detailed understanding of how TAPBPR edits the peptide repertoire. Thus, there is an unmet need to understand the complete mechanism of MHC-1 antigen proofreading and the peptide repertoire selection. BRIEF SUMMARY OF THE INVENTION [0007] Provided herein are methods of increasing the immunogenicity of mammalian cells. The methods comprise providing a population of mammalian cells having surface MHC class I molecule with an immunogenic agent and a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecules thereby increasing the immunogenicity of the mammalian cells. In certain 2 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 embodiments, the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11. [0008] In certain embodiments, the MHC class I molecule is selected from the group consisting of an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, an HLA-E molecule, an HLA-G molecule, an HLA-F molecule, a MR1 molecule, a CD1 molecule, and a combination thereof. The MHC class I molecule can, for example, be an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, a MR1 molecule, an HLA-E molecule, and/or an HLA-G molecule. The HLA-A molecule, HLA-B molecule, and/or HLA-C molecule can, for example, be selected from an HLA-A01 supertype molecule, an HLA-A02 supertype molecule, an HLA-A03 supertype molecule, an HLA-A24 supertype molecule, an HLA- A01/A24 supertype molecule, an HLA-A01/A03 supertype molecule, an HLA-B08 supertype molecule, an HLA-B44 supertype molecule, an HLA-B58 supertype molecule, and/or an HLA-C01 supertype molecule. In certain embodiments, the MHC class I molecule can, for example, be selected from HLA-A*01:01, HLA-A*11:01, HLA-A*29:01, HLA-A*30:01, HLA-A*32:01, HLA-A*74:01, HLA-A*24:03, HLA-A*02:06, HLA-A*02:01, HLA- A*02:03, HLA-A*23:01, HLA-A*24:02, HLA-A*68:01, HLA-A*69:01, HLA-A*68:02, HLA-A*29:02, HLA-A*03:01, HLA-B*08:01, HLA-B*57:03, HLA-B*59:01, HLA-B*37:01, HLA-C*01:02, HLA-C*03:04, MR1, HLA-E*01:03, and/or HLA-G*01:01. [0009] In certain embodiments, the mammalian cell is a human cell. [0010] In certain embodiments, the immunogenic agent is selected from an immunogenic peptide, a metabolite, a lipid, a phospholipid, a fatty acid ligand, and/or combinations thereof. In certain embodiments, the immunogenic peptides are low-to-moderate affinity peptides. [0011] In certain embodiments, the method is performed in vivo. [0012] Also provided are methods of producing antigen presenting cells to stimulate an immune response in a subject in need thereof. The methods comprise (a) obtaining antigen presenting cells from the subject in need thereof; and (b) providing antigen presenting cells comprising surface MHC class I molecules in vitro with a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7 and an immunogenic agent, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecules thereby creating antigen presenting cells for stimulating an immune response. In certain embodiments, the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11. 3 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 [0013] In certain embodiments, the MHC class I molecule is selected from the group consisting of an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, an HLA-E molecule, an HLA-G molecule, an HLA-F molecule, a MR1 molecule, a CD1 molecule, and a combination thereof. The MHC class I molecule can, for example, be an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, a MR1 molecule, an HLA-E molecule, and/or an HLA-G molecule. The HLA-A molecule, HLA-B molecule, and/or HLA-C molecule can, for example, be selected from an HLA-A01 supertype molecule, an HLA-A02 supertype molecule, an HLA-A03 supertype molecule, an HLA-A24 supertype molecule, an HLA- A01/A24 supertype molecule, an HLA-A01/A03 supertype molecule, an HLA-B08 supertype molecule, an HLA-B44 supertype molecule, an HLA-B58 supertype molecule, and/or an HLA-C01 supertype molecule. In certain embodiments, the MHC class I molecule can, for example, be selected from HLA-A*01:01, HLA-A*11:01, HLA-A*29:01, HLA-A*30:01, HLA-A*32:01, HLA-A*74:01, HLA-A*24:03, HLA-A*02:06, HLA-A*02:01, HLA- A*02:03, HLA-A*23:01, HLA-A*24:02, HLA-A*68:01, HLA-A*69:01, HLA-A*68:02, HLA-A*29:02, HLA-A*03:01, HLA-B*08:01, HLA-B*57:03, HLA-B*59:01, HLA-B*37:01, HLA-C*01:02, HLA-C*03:04, MR1, HLA-E*01:03, and/or HLA-G*01:01. [0014] In certain embodiments, the mammalian cell is a human cell. [0015] In certain embodiments, the immunogenic agent is selected from an immunogenic peptide, a metabolite, a lipid, a phospholipid, a fatty acid ligand, and/or combinations thereof. In certain embodiments, the immunogenic peptides are low-to-moderate affinity peptides. [0016] In certain embodiments, the antigen presenting cells are dendritic cells. [0017] In certain embodiments, the method further comprises administering the antigen presenting cells to the subject in need thereof. [0018] Also provided are methods of stimulating an immune response in a subject in need thereof, the method comprising (a) administering to the subject in need thereof a TAP- binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7, wherein the TAPBPR polypeptide binds to an antigen presenting cell comprising a surface MHC class I molecule in the subject in need thereof; and (b) administering to the subject in thereof an immunogenic agent, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecule of the antigen presenting cell such that the antigen presenting cell stimulates an immune response in the subject in need thereof. Also provided are TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7 and an immunogenic agent for use in stimulating an immune response in a subject in need thereof, 4 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 wherein the TAPBPR polypeptide binds to an antigen presenting cell comprising a surface MHC class I molecule; and wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecule of the antigen presenting cell such that the antigen presenting cell stimulates an immune response. In certain embodiments, the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10. [0019] In certain embodiments, the MHC class I molecule is selected from the group consisting of an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, an HLA-E molecule, an HLA-G molecule, an HLA-F molecule, a MR1 molecule, a CD1 molecule, and a combination thereof. The MHC class I molecule can, for example, be an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, a MR1 molecule, an HLA-E molecule, and/or an HLA-G molecule. The HLA-A molecule, HLA-B molecule, and/or HLA-C molecule can, for example, be selected from an HLA-A01 supertype molecule, an HLA-A02 supertype molecule, an HLA-A03 supertype molecule, an HLA-A24 supertype molecule, an HLA- A01/A24 supertype molecule, an HLA-A01/A03 supertype molecule, an HLA-B08 supertype molecule, an HLA-B44 supertype molecule, an HLA-B58 supertype molecule, and/or an HLA-C01 supertype molecule. In certain embodiments, the MHC class I molecule can, for example, be selected from HLA-A*01:01, HLA-A*11:01, HLA-A*29:01, HLA-A*30:01, HLA-A*32:01, HLA-A*74:01, HLA-A*24:03, HLA-A*02:06, HLA-A*02:01, HLA- A*02:03, HLA-A*23:01, HLA-A*24:02, HLA-A*68:01, HLA-A*69:01, HLA-A*68:02, HLA-A*29:02, HLA-A*03:01, HLA-B*08:01, HLA-B*57:03, HLA-B*59:01, HLA-B*37:01, HLA-C*01:02, HLA-C*03:04, MR1, HLA-E*01:03, and/or HLA-G*01:01. [0020] In certain embodiments, the subject is a human. [0021] In certain embodiments, the immunogenic agent is selected from an immunogenic peptide, a metabolite, a lipid, a phospholipid, a fatty acid ligand, and/or combinations thereof. In certain embodiments, the immunogenic peptides are low-to-moderate affinity peptides. [0022] In certain embodiments, the antigen presenting cells are dendritic cells. [0023] In certain embodiments, the TAPBPR polypeptide further comprises a signal peptide, a flag tag, a linker, and/or a transmembrane domain. The signal peptide can, for example, comprise SEQ ID NO:47. The flag tag can, for example, comprise SEQ ID NO:48. The linker can, for example comprise SEQ ID NO:49. The transmembrane domain can, for example, comprise SEQ ID NO:50. [0024] Also provided is an isolated TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7. In certain 5 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 embodiments, the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11. [0025] In certain embodiments, the TAPBPR polypeptide further comprises a signal peptide, a flag tag, a linker, and/or a transmembrane domain. The signal peptide can, for example, comprise SEQ ID NO:47. The flag tag can, for example, comprise SEQ ID NO:48. The linker can, for example comprise SEQ ID NO:49. The transmembrane domain can, for example, comprise SEQ ID NO:50. [0026] Also provided are isolated nucleic acids encoding the TAPBPR polypeptides disclosed herein. [0027] Also provided are isolated vectors comprising the isolated nucleic acids disclosed herein. [0028] Also provided are host cells comprising the vectors disclosed herein. In certain embodiments, the host cell is a mammalian cell. [0029] Further aspects, features and advantages of the present invention will be better appreciated upon a reading of the following detailed description of the invention and claims. BRIEF DESCRIPTION OF THE DRAWINGS [0030] The foregoing summary, as well as the following detailed description of preferred embodiments of the present application, will be better understood when read in conjunction with the appended drawings. It should be understood, however, that the application is not limited to the precise embodiments shown in the drawings. [0031] FIGs.1A-1F demonstrated that a high-fidelity TAPBPRFLQ variant promotes peptide editing. FIG. 1A shows a structural model of TAPBPRFLQ in complex with peptide-free HLA-A*02:01/β2m generated using RosettaCM. The G24-R36 loop is colored in orange, and the A29-S32 loop segment is shown as spheres. The side chains of mutations S104F, K211L, and R270Q are shown as magenta sticks. FIG.1B shows a representative SPR sensorgram of graded concentrations of TAPBPRFLQ flown over a streptavidin chip coupled with HLA- A*02:01/TAX9 in excess TAX9 peptide. FIG.1C shows a log-scale comparison of KD values between HLA-A*02:01/TAX9 and TAPBPR. Results of three independent experiments (mean ± SD) are shown as scatter plots. FIG. 1D shows peptide dissociation curves of fluorophore-labeled TAMRA-TAX9 peptide-loaded HLA-A*02:01 in the presence of excess unlabeled TAX9 peptide with buffer, TAPBPRWT, TAPBPRTN6, or TAPBPRFLQ. Data are means for n = 3 independent experiments. FIG. 1E shows relative peptide dissociation of TAMRA-TAX9 from HLA-A*02:01 by TAPBPR relative to no TAPBPR. Error bars (SD) 6 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 were propagated from three independent experiments. FIG. 1F shows correlation of Log10 (MFI ratio) between TAPBPRWT and TAPBPRFLQ plotted in FIG. 6. The dashed line represents a conceptual 1:1 correlation. Two-sample unequal variance student’s t-test was performed, P > 0.12 (not significant, ns), P < 0.033(*), P < 0.002(**), and P < 0.001(***). [0032] FIG.2 shows a comparison of the sequences for TAPBPR, and its variants used in this study. Sequence alignment of the human wild-type TAPBPRWT (SEQ ID NO:7), the loop- shorten mutant TAPBPR∆ALAS (SEQ ID NO:8), the loop-deleted mutant TAPBPR∆G24-R36 (SEQ ID NO:9), the engineered variant TAPBPRFLQ (SEQ ID NO:10), the loop-deleted variant TAPBPRFLQ∆G24-R36 (SEQ ID NO:11). The loop region and amino acid mutations are indicated. Alignments were performed using ClustalOmega (18) and processed with ESPript 3 (19). [0033] FIGs. 3A-3B show the purification of recombinant TAPBPRFLQ∆G24-R36 protein. FIG. 3A shows a SEC trace of TAPBPRFLQ∆G24-R36 mutant. The arrow indicates the protein peak relative to TAPBPRWT and TAPBPRFLQ and is further confirmed by SDS/PAGE analysis. FIG. 3B shows DSF of TAPBPRFLQ∆G24-R36 (Tm = 49.2 ˚C, green) relative to TAPBPRWT (Tm = 51.5 ˚C, red). [0034] FIGs. 4A-4F show direct interactions between HLA-A allotypes and TAPBPR variants. FIGs. 4A-4F show representative SPR sensorgrams of various concentrations of TAPBPRWT (FIG. 4A), TAPBPR∆ALAS (FIG. 4B), TAPBPR∆G24-R36 (FIG. 4C), TAPBPRFLQ (FIG. 4D), TAPBPRFLQ∆G24-R36 (FIG. 4E), and TAPBPRTN6 (FIG. 4F) flowed over a streptavidin chip coupled with HLA-A*02:01/TAX9 in molar excess TAX9 peptide. The concentrations of analyte are noted. The equilibrium constant, KD, is the mean ± SD for n = 3 independent experiments. The KDs of TAPBPRWT, TAPBPR∆ALAS, and TAPBPR∆G24-R36 are estimated values since saturation of binding cannot be reached. [0035] FIG. 5 shows a graph of a peptide dissociation assay. Comparison of peptide dissociation kinetics of TAMRA fluorophore-labeled TAMRATAX9 peptide (TAMRAKLFGYPVYV (SEQ ID NO:2))-loaded HLA-A*02:01 (40nM) with 1 μM unlabeled TAX9 peptide in the presence of buffer or 1 μM TAPBPRTN6, TAPBPRWT, TAPBPR∆G24-R36, TAPBPRFLQ, and TAPBPR FLQ∆G24-R36. The plateau for each dissociation was individually extracted by fitting one phase decay, and the relative dissociation was then calculated using quation % ^^^^^^^^^^^^^^^^ ^^^^^^^^^^^^^^^^^^^^^^^^ = ^ ^౫^^^౨ ^^^౯ ^^^౦^౨^^^ the e ୪ୟ^^ୟ^ ି^୪ୟ^^ୟ^ ^୪ୟ^^ୟ^^౫^^^౨ ^^^౯ . Error bar (SD) was propagated from three independent experiments. 7 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 [0036] FIG.6 shows binding levels of TAPBPR tetramers on HLA single antigen beads. FIG. 6, top, shows a graph demonstrating levels of folded MHC-I molecules captured on single antigen beads (SABs), related to FIG. 1F, which were detected using the primary anti-HLA Class I antibody W6/32 (Abcam, ab22432) and the secondary anti-mouse PE-conjugated antibody (Abcam, ab97024). Similar levels of peptide-loaded MHC-I molecules were observed across different HLA allotypes. FIG. 6, middle and bottom, shows bar graphs showing the Log10 Mean Fluorescence Intensity (MFI) levels of tetramerized TAPBPRFLQ, TAPBPRWT, and TAPBPRTN6 binding to HLA molecules on SABs. Tetramer staining upon the W6/32 antibody incubation was used to control for background staining levels. The plotted data were generated based on n=2 or 3 independent experiments, and the standard deviation is depicted as error bars. [0037] FIGs. 7A-7C shows log10 (MFI ratios) of TAPBPR tetramer binding to 97 common HLA allotypes. FIG. 7A shows a bar graph showing the Log10 (MFI ratios) of TAPBPRFLQ, TAPBPRWT, and TAPBPRTN6 (negative control) binding to the SABs. Plotted data are means ± SD from 2 to 3 independent experiments. FIGs. 7B-7C show graphs of the correlation of Log10 (MFI ratio) for TAPBPRFLQ (FIG. 7B) and TAPBPRWT (FIG. 7C) relative to TAPBPRTN6 plotted in FIG.6. The dashed line represents a conceptual 1:1 correlation. [0038] FIGs. 8A-8D demonstrated that TAPBPRFLQ enhanced peptide loading across HLA allotypes expressed on a cellular membrane. FIG. 8A shows a schematic of peptide exchange for MHC-I expressed on the cell surface by soluble TAPBPR. FIG. 8B shows a bar graph summarizing the MFI of fluorescent peptide binding for monoallelic HLA-A*02:01, A*24:02, and A23:01 cell lines with 10 nM fluorescent peptide and 1 μM TAPBPR and HLA-A*03:01 cell line in the presence of 10 μM fluorescent peptide and 10 μM TAPBPR, as indicated, from three independent experiments. Two-way ANOVA was performed relative to TAPBPRTN6, P > 0.0.1234 (ns), P < 0.0332(*), P < 0.0021(**), P < 0.0002(***), and P < 0.0001(****). FIG. 8C shows a schematic of peptide exchange for MHC-I expressed on the cell surface by membrane-bound TAPBPR. FIG. 8D shows a bar graph summarizing the MFI of different concentrations of fluorescent peptide binding to HLA-A*02:01 expressed T2 cell line with expression of TAPBPR-TM, as indicated, from two independent experiments. Two- way ANOVA was performed relative to the parental (no transduction), P > 0.0.1234 (ns), P < 0.0332(*), P < 0.0021(**), P < 0.0002(***), and P < 0.0001(****). [0039] FIGs. 9A-9D show flow cytometry gating strategy of monoallelic 722.211 cell lines. FIGs. 9A-9D show the results of flow cytometry for monoallelic HLA-A*02:01 (FIG. 9A), 8 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 24:02 (FIG. 9B), 23:01 (FIG. 9C), or 03:01 (FIG. 9D) 722.211 cell lines were thawed and recovered before performing peptide exchange. Cells were sorted by side and forward scatter (SSC-A and FSC-A) followed by single cell isolation (SSC-A and SSC-H). Gating for live cells was determined by LIVE/DEAD™ Fixable Violet Dead Cell Stain Kit. Gates are shown in the black box, and the percentages of events are gated in parentheses. The acquisition was performed on CytoFLEX LX (Beckman Coulter), and the data were analyzed by FlowJo v10.10.0. [0040] FIGs.10A-10D show soluble TAPBPR mediated peptide exchange on surface MHC-I. FIGs. 10A-10D show bar graph summarizing the MFI of fluorescent peptide binding to monoallelic HLA-A*02:01 (FIG. 10A), A*24:02 (FIG. 10B), A23:01 (FIG. 10C), and A*03:01 (FIG. 10D) 721.211 cell lines in the presence of soluble TAPBPR at different concentrations, as indicated, from 3 independent experiments. HLA-A*02:01, A*24:02 (FIG. 10B), and A23:01 (FIG. 10C) 721.211 cell lines were incubated in the presence of soluble TAPBPRTN6, TAPBPRWT, and TAPBPRFLQ at different concentrations, followed by incubation with 10 nM fluorescent peptide for 60 minutes. HLA-A*03:01 monoallelic cell line was incubated in the presence of soluble TAPBPRTN6, TAPBPRWT, and TAPBPRFLQ, followed by incubation with 10 μM (FIG. 10D) fluorescent peptide for 60 minutes. Two-way ANOVA was performed relative to the TAPBPRTN6, P > 0.0.1234 (ns), P < 0.0332(*), P < 0.0021(**), P < 0.0002(***), and P < 0.0001(****). [0041] FIG. 11 shows fluorescent peptide binding on monoallelic HLA-A*02:01 cell line. Titration of soluble TAPBPRTN6, TAPBPRWT, and TAPBPRFLQ at different concentrations to the monoallelic HLA-A*02:01 721.211 cell line with the incubation of 10 nM fluorescent peptide for 60 minutes. Gates are shown in black box, and the percentages of events are gated in parentheses. The acquisition was performed on CytoFLEX LX (Beckman Coulter), and the data were analyzed by FlowJo v10.10.0. [0042] FIG. 12 shows fluorescent peptide binding on monoallelic HLA-A*24:02 cell line. Titration of soluble TAPBPRTN6, TAPBPRWT, and TAPBPRFLQ at different concentrations to the monoallelic HLA-A*24:02 721.211 cell line with the incubation of 10 nM fluorescent peptide for 60 minutes. Gates are shown in black box, and the percentages of events are gated in parentheses. The acquisition was performed on CytoFLEX LX (Beckman Coulter), and the data were analyzed by FlowJo v10.10.0. [0043] FIG. 13 shows fluorescent peptide binding on monoallelic HLA-A*23:01 cell line. Titration of soluble TAPBPRTN6, TAPBPRWT, and TAPBPRFLQ at different concentrations to the monoallelic HLA-A*23:01 721.211 cell line with the incubation of 10 nM fluorescent 9 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 peptide for 60 minutes. Gates are shown in black box, and the percentages of events are gated in parentheses. The acquisition was performed on CytoFLEX LX (Beckman Coulter), and the data were analyzed by FlowJo v10.10.0. [0044] FIG. 14 shows fluorescent peptide binding on monoallelic HLA-A*03:01 cell line. Titration of soluble TAPBPRTN6, TAPBPRWT, and TAPBPRFLQ at different concentrations to the monoallelic HLA-A*24:02 721.211 cell line with the incubation of 10 μM fluorescent peptide for 60 minutes. Gates are shown in black box, and the percentages of events are gated in parentheses. The acquisition was performed on CytoFLEX LX (Beckman Coulter), and the data were analyzed by FlowJo v10.10.0. [0045] FIG. 15 shows representative flow cytometry gating strategy of T2 cell lines in peptide exchange experiments. Cells were incubated with varying concentrations of fluorescent TAMRA-TAX9 peptide for 60 minutes before washing with FACS buffer, staining with LIVE/DEAD™ Fixable Near IR (876) Viability dye, and fixation with 4% PFA in PBS. Analyzed cells were gated on live singlets of appropriate size. Cells incubated with 10-5M of TAX9-TAMARA are shown. The acquisition was performed on CytoFLEX LX (Beckman Coulter), and the data were analyzed by FlowJo v10.10.0. [0046] FIGs. 16A-16C show flow cytometry analysis of T2 cell lines. FIG. 16A shows a graph demonstrating surface TAPBPR expression of TAPBPRFLQ, TAPBPRWT, and TAPBPRTN6-TM transduced T2 cell lines, as measured by anti-FLAG MFI. FIG. 16B shows a graph demonstrating TAPBPR transduction efficiency of TAPBPRFLQ, TAPBPRWT, and TAPBPRTN6-TM transduced T2 cell lines relative to the non-transduced parental cell line. Cells with similar transduction efficiency and levels of FLAG expression were selected for downstream peptide exchange assays. FIG. 16C shows a graph demonstrating surface HLA- A*02:01 expression of TAPBPR-FLQ, WT, and TN6 TAPBPRFLQ, TAPBPRWT, and TAPBPRTN6-TM transduced and parental T2 cell lines from cells selected for peptide exchange assays. [0047] FIG. 17 shows fluorescent peptide binding of HLA-A*02:01 mediated by surface TAPBPR. Titration of peptide at different concentrations to TAPBPRTN6, TAPBPRWT, and TAPBPRFLQ-TM transduced T2 cell lines. The acquisition was performed on CytoFLEX LX (Beckman Coulter), and the data were analyzed by FlowJo v10.10.0. [0048] FIGs. 18A-18E demonstrated the isolation and cryoEM structure of an MHC-I intermediate bound to a peptide decoy by TAPBPR. FIG. 18A shows a size exclusion chromatography purification of a recombinant open HLA-A*02:01/TAPBPRFLQ complex 10 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 prepared by UV-irradiation of KILGFVFJV/HLA-A*02:01 pre-incubated with TAPBPRFLQ at 1:1.3 molar ratio. J = 3-amino-3-(2-nitrophenyl)-propionic acid. SDS/PAGE analysis confirms the identity of the TAPBPRFLQ/open HLA-A2 complex peak under nonreducing (NR) or reducing (R) conditions. The bands of TAPBPRFLQ, open HLA-A2 heavy chain (HC), and β2m, as well as the positions of comigrated molecular-weight standards, are indicated. FIG.18B shows a LC/MS analysis of the purified complex in FIG.18A showing the presence of captured peptide decoy KILGFVF (SEQ ID NO:5) (observed and expected mass-to-charge ratios are 822.52 and 822.50 m/z, respectively). FIG. 18C shows a reconstructed cryoEM density and refined structural model of the tertiary complex, with TAPBPRFLQ shown in light pink, open HLA-A*02:01 heavy chain in light green, β2m in light blue, and peptide in wheat, as indicated. FIG. 18D show a zoom in on the interactions between the TAPBPRFLQ hairpin and the floor of the MHC-I groove. FIG. 18E shows a zoom in on the interactions between TAPBPRFLQ IgC domain residues and β2m, highlighting the sidechain interaction. [0049] FIGs. 19A-19B show LC-MS validation of UV-irradiated placeholder ligand. FIGs. 19A-19B show LC-MS analysis of peptide KILGFVFJV (SEQ ID NO:46) (FIG. 19A) and UV-irradiated peptide KILGFVFJV (SEQ ID NO:46) (FIG. 19B). J = 3-amino-3-(2- nitrophenyl)-propionic acid. UV irradiation was performed at 365 nm for 40 minutes at 4˚C. Left panel: the LC chromatogram trace of each sample. Right panel: relative abundance for the selected time interval (red box). In FIG. 19A, the presence of KILGFVFJV (SEQ ID NO:46) is noted (observed and expected mass-to-charge ratios are 1114.63 and 1114.63 m/z, respectively). In contrast, UV irradiation of KILGFVFJV (SEQ ID NO:46) in (FIG. 19B) results in a peptide fragment KILGFVF (SEQ ID NO:5) (observed and expected mass-to- charge ratios are 822.53 and 822.50 m/z, respectively). [0050] FIGs.20A-20E show NMR characterization of TAPBPRFLQ interactions with peptide- loaded HLA-A*02:01. FIG. 20A shows a representative region of the 2D 1H-13C HMQC spectral overlay of 13C AILV methyl-labeled TAX9/HLA-A*02:01 and with a 3-fold molar excess of TAPBPRFLQ. FIG. 20B shows selected NMR resonances (corresponding to I23 and A125) undergoing conformational motion in the slow-exchange regime upon titration of TAPBPRFLQ. The HLA-A*02:01: TAPBPRFLQ ratios for the different titration points were 1: 0, 1: 0.55, 1: 1.10, 1: 1.65, 1: 2.75 and 1: 3.31. FIG. 20C shows that methyl groups of residues undergoing significant chemical shift deviation (CSD) upon binding TAPBPRFLQ are mapped onto the structure of the HLA-A*02:01/TAPBPRFLQ complex. The structure is a Roseta-CM homology model obtained using the H2-Dd/TAPBPR crystal structure as a template (PDB ID: 5WER). CSDs are plotted using a heat-map scale 11 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 shown on the right. FIG. 20D shows a correlation plot of CSDs observed for the titration of HLA-A*02:01 with TAPBPRWT versus TAPBPRFLQ. The Pearson correlation coefficient (r) is shown on the plot. The correlations are statistically significant with a P value of < 0.0001. FIG. 20E shows a close-up of the peptide-binding groove with the TAX9 peptide (shown as green sticks). Select residues distributed throughout the MHC-I groove and their CSDs are labeled. The mutation sites on TAPBPRFLQ are denoted as pink spheres. [0051] FIGs. 21A-21C show NMR spectra overlay and line shape analysis of peptide-loaded HLA-A*02:01 upon TAPBPRFLQ titration. FIG. 21A shows a 2D 1H-13C HMQC spectral overlay of 13C AILV methyl-labeled HLA-A*02:01 titrated with TAPBPRFLQ in the molar ratios: 1: 0, 1: 0.55, 1: 1.10, 1: 1.65, 1: 2.75, 1: 3.31. FIG. 21B resonances for selected methyl groups (I23, I52, A125, L126, V247) undergoing slow exchange upon adding TAPBPRFLQ. FIG. 21C shows a representative NMR line shape fitting of TAPBPRFLQ titration onto TAX9/HLA-A*02:01/β2m complex for heavy chain methyl residues A125 and L126 using TITAN (5). [0052] FIG. 22 shows a schematic of a cryo-EM data processing workflow. Cryo-EM data processing workflow including a representative micrograph (top left). [0053] FIG. 23 shows cryoEM data quality. Local resolution calculated using Relion’s local resolution tool (top panels). Unmasked (grey dashed), masked (black) and movel versus map (green) fourier shell correlation (FSC) curves (bottom left panel). Representative 2D classes generated in cryoSPARC from final particles (middle right panel) and angular distribution from cryoSPARC reconstruct only based on Relion angular assignment (bottom right panel). [0054] FIGs. 24A-24C show structural adaptations of the MHC-I groove induced by TAPBPR. FIG. 24A shows an overview of the molecular complex between TAPBPRFLQ/HLA-A*02:01/β2m with bound peptide decoy. The MHC-I α2 helix has been partially removed for simplicity. FIG.24B shows a structural superposition of peptide-loaded HLA-A*02:01 (PDB ID 2VLL) onto the cryoEM structure of KILGFVF (SEQ ID NO:5)/HLA-A*02:01 (PDB ID 9C96, EMDB ID EMD-45360) showing well-resolved density for peptide residues 1-6, and a lack of cryoEM density for residues G71-G91 of HLA- A*02:01. MHC-I A-, B-, D-, E-, F-pockets are noted, and the spheres are showing A150 Cα. FIG. 24C shows detailed interactions between conserved MHC-I groove residues with the backbone of peptide residues P1-P3, as indicated (top and side views). [0055] FIG.25 shows fit quality of peptide in the MHC Groove. Map density and full residue Q-scores for the key residues of the MHC groove (top panels) and peptide (bottom panels). 12 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 Map is contoured at σ = 10 (top left, middle panels) or σ = 5 (top right, bottom panels). The overall average Q-score for the full structure is 0.52, and the expected average Q-score in a map at 3.0 Å resolution is 0.49. [0056] FIG. 26 shows a half map density for the peptide. Peptide density for half map 1 (left) and half map 2 (right). Maps are contoured at σ = 5. [0057] FIG. 27 shows a sequence logo of conserved peptide-coordinating residues across 215 common HLA allotypes. Seq2logo visualization (20) depicting the conserved peptide- coordinating residues’ sequence alignment across 215 distinct HLA-A*, B*, and C* allotypes. [0058] FIG. 28 shows a structural mechanism of peptide antigen proofreading by TAPBPR. Empty MHC-I is preferentially recognized by TAPBPR in a peptide-receptive, “open” conformation. Chaperoned MHC-I screens a large peptide pool in the endoplasmic reticulum, forming transient peptide/MHC-I/TAPBPR complexes. TAPBPR widens the α2-1 helix of the peptide binding groove, enhances the dynamics of the α1 helix, and induces unstructured F- pockets. Thus, chaperone MHC-I exhibits a lower affinity towards incoming peptides and promotes the dissociation of suboptimal peptide decoys. When chaperoned MHC-I proofreads a high-affinity peptide, the N terminal peptide forms native-like contacts with its A- and B-pockets. The C-terminus of the peptide interacts with peptide-coordinating residues in the F-pocket to stabilize and close the peptide binding groove, which allosterically triggers the release of TAPBPR. DETAILED DESCRIPTION OF THE INVENTION [0059] This disclosure is based on the discovery that it is possible to increase the immunogenicity of mammalian cells, to produce antigen presenting cells to stimulate an immune response in a subject, and to stimulate an immune response in a subject in need thereof use a combination of an immunogenic agent and a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecules thereby increasing the immunogenicity of the mammalian cells, producing antigen presenting cells to stimulate an immune response, and stimulating an immune response in a subject. [0060] Various publications, articles and patents are cited or described in the background and throughout the specification; each of these references is herein incorporated by reference in its entirety. Discussion of documents, acts, materials, devices, articles or the like which has been included in the present specification is for the purpose of providing context for the 13 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 invention. Such discussion is not an admission that any or all of these matters form part of the prior art with respect to any inventions disclosed or claimed. [0061] For clarity of disclosure, and not by way of limitation, the detailed description of the invention is divided into subsections that describe or illustrate certain features, embodiments, or applications of the present invention. Definitions [0062] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood to one of ordinary skill in the art to which this invention pertains. Otherwise, certain terms used herein have the meanings as set forth in the specification. [0063] It must be noted that as used herein and in the appended claims, the singular forms “a,” “an,” and “the” include plural reference unless the context clearly dictates otherwise. [0064] Unless otherwise stated, any numerical values, such as a concentration or a concentration range described herein, are to be understood as being modified in all instances by the term “about.” Thus, a numerical value typically includes ± 10% of the recited value. For example, a concentration of 1 mg/mL includes 0.9 mg/mL to 1.1 mg/mL. Likewise, a concentration range of 1% to 10% (w/v) includes 0.9% (w/v) to 11% (w/v). As used herein, the use of a numerical range expressly includes all possible subranges, all individual numerical values within that range, including integers within such ranges and fractions of the values unless the context clearly indicates otherwise. [0065] Unless otherwise indicated, the term “at least” preceding a series of elements is to be understood to refer to every element in the series. Those skilled in the art will recognize or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments of the invention described herein. Such equivalents are intended to be encompassed by the invention. [0066] As used herein, the terms “comprises,” “comprising,” “includes,” “including,” “has,” “having,” “contains” or “containing,” or any other variation thereof, will be understood to imply the inclusion of a stated integer or group of integers but not the exclusion of any other integer or group of integers and are intended to be non-exclusive or open-ended. For example, a composition, a mixture, a process, a method, an article, or an apparatus that comprises a list of elements is not necessarily limited to only those elements but can include other elements not expressly listed or inherent to such composition, mixture, process, method, article, or apparatus. Further, unless expressly stated to the contrary, “or” refers to an inclusive or and 14 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 not to an exclusive or. For example, a condition A or B is satisfied by any one of the following: A is true (or present) and B is false (or not present), A is false (or not present) and B is true (or present), and both A and B are true (or present). [0067] As used herein, the conjunctive term “and/or” between multiple recited elements is understood as encompassing both individual and combined options. For instance, where two elements are conjoined by “and/or,” a first option refers to the applicability of the first element without the second. A second option refers to the applicability of the second element without the first. A third option refers to the applicability of the first and second elements together. Any one of these options is understood to fall within the meaning, and therefore satisfy the requirement of the term “and/or” as used herein. Concurrent applicability of more than one of the options is also understood to fall within the meaning, and therefore satisfy the requirement of the term “and/or.” [0068] As used herein, the term “consists of,” or variations such as “consist of” or “consisting of,” as used throughout the specification and claims, indicate the inclusion of any recited integer or group of integers, but that no additional integer or group of integers can be added to the specified method, structure, or composition. [0069] As used herein, the term “consists essentially of,” or variations such as “consist essentially of” or “consisting essentially of,” as used throughout the specification and claims, indicate the inclusion of any recited integer or group of integers, and the optional inclusion of any recited integer or group of integers that do not materially change the basic or novel properties of the specified method, structure or composition. See M.P.E.P. § 2111.03. [0070] As used herein, “subject” means any animal, preferably a mammal, most preferably a human. The term “mammal” as used herein, encompasses any mammal. Examples of mammals include, but are not limited to, cows, horses, sheep, pigs, cats, dogs, mice, rats, rabbits, guinea pigs, monkeys, humans, etc., more preferably a human. [0071] The words “right,” “left,” “lower,” “upper,” “top,” “bottom,” and “middle” designate directions in the drawings to which reference is made. [0072] It should also be understood that the terms “about,” “approximately,” “generally,” “substantially,” and like terms, used herein when referring to a dimension or characteristic of a component of the preferred invention, indicate that the described dimension/characteristic is not a strict boundary or parameter and does not exclude minor variations therefrom that are functionally the same or similar, as would be understood by one having ordinary skill in the art. At a minimum, such references that include a numerical parameter would include variations that, using mathematical and industrial principles accepted in the art (e.g., rounding, 15 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 measurement or other systematic errors, manufacturing tolerances, etc.), would not vary the least significant digit. [0073] The terms “identical” or percent “identity,” in the context of two or more nucleic acids or polypeptide sequences (e.g., TAP-binding protein-related (TAPBPR) polypeptides and nucleotide sequences encoding the same), refer to two or more sequences or subsequences that are the same or have a specified percentage of amino acid residues or nucleotides that are the same, when compared and aligned for maximum correspondence, as measured using one of the following sequence comparison algorithms or by visual inspection. [0074] For sequence comparison, typically one sequence acts as a reference sequence, to which test sequences are compared. When using a sequence comparison algorithm, test and reference sequences are input into a computer, subsequence coordinates are designated, if necessary, and sequence algorithm program parameters are designated. The sequence comparison algorithm then calculates the percent sequence identity for the test sequence(s) relative to the reference sequence, based on the designated program parameters. [0075] Optimal alignment of sequences for comparison can be conducted, e.g., by the local homology algorithm of Smith & Waterman, Adv. Appl. Math. 1981; 2:482, by the homology alignment algorithm of Needleman & Wunsch, J. Mol. Biol. 1970; 48:443, by the search for similarity method of Pearson & Lipman, Proc. Nat’l. Acad. Sci. USA 1988; 85:2444, by computerized implementations of these algorithms (GAP, BESTFIT, FASTA, and TFASTA in the Wisconsin Genetics Software Package, Genetics Computer Group, 575 Science Dr., Madison, WI), or by visual inspection (see generally, Current Protocols in Molecular Biology, F.M. Ausubel et al., eds., Current Protocols, a joint venture between Greene Publishing Associates, Inc. and John Wiley & Sons, Inc., 1995 Supplement (Ausubel)). [0076] Examples of algorithms that are suitable for determining percent sequence identity and sequence similarity are the BLAST and BLAST 2.0 algorithms, which are described in Altschul et al., J. Mol. Biol. 1990; 215: 403-410 and Altschul et al., Nucleic Acids Res. 1997; 25: 3389-3402, respectively. Software for performing BLAST analyses is publicly available through the National Center for Biotechnology Information. This algorithm involves first identifying high scoring sequence pairs (HSPs) by identifying short words of length W in the query sequence, which either match or satisfy some positive-valued threshold score T when aligned with a word of the same length in a database sequence. T is referred to as the neighborhood word score threshold (Altschul et al, supra). These initial neighborhood word hits act as seeds for initiating searches to find longer HSPs containing them. The word hits 16 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 are then extended in both directions along each sequence for as far as the cumulative alignment score can be increased. [0077] Cumulative scores are calculated using, for nucleotide sequences, the parameters M (reward score for a pair of matching residues; always > 0) and N (penalty score for mismatching residues; always < 0). For amino acid sequences, a scoring matrix is used to calculate the cumulative score. Extension of the word hits in each direction are halted when: the cumulative alignment score falls off by the quantity X from its maximum achieved value; the cumulative score goes to zero or below, due to the accumulation of one or more negative- scoring residue alignments; or the end of either sequence is reached. The BLAST algorithm parameters W, T, and X determine the sensitivity and speed of the alignment. The BLASTN program (for nucleotide sequences) uses as defaults a wordlength (W) of 11, an expectation (E) of 10, M=5, N=-4, and a comparison of both strands. For amino acid sequences, the BLASTP program uses as defaults a wordlength (W) of 3, an expectation (E) of 10, and the BLOSUM62 scoring matrix (see Henikoff & Henikoff, Proc. Natl. Acad. Sci. USA 1989; 89:10915). [0078] In addition to calculating percent sequence identity, the BLAST algorithm also performs a statistical analysis of the similarity between two sequences (see, e.g., Karlin & Altschul, Proc. Nat’l. Acad. Sci. USA 1993; 90:5873-5787). One measure of similarity provided by the BLAST algorithm is the smallest sum probability (P(N)), which provides an indication of the probability by which a match between two nucleotide or amino acid sequences would occur by chance. For example, a nucleic acid is considered similar to a reference sequence if the smallest sum probability in a comparison of the test nucleic acid to the reference nucleic acid is less than about 0.1, more preferably less than about 0.01, and most preferably less than about 0.001. [0079] A further indication that two nucleic acid sequences or polypeptides are substantially identical is that the polypeptide encoded by the first nucleic acid is immunologically cross reactive with the polypeptide encoded by the second nucleic acid, as described below. Thus, a polypeptide is typically substantially identical to a second polypeptide, for example, where the two peptides differ only by conservative substitutions. Another indication that two nucleic acid sequences are substantially identical is that the two molecules hybridize to each other under stringent conditions. [0080] As used herein, the term “isolated” means a biological component (such as a nucleic acid, peptide or protein) has been substantially separated, produced apart from, or purified away from other biological components of the organism in which the component naturally 17 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 occurs, i.e., other chromosomal and extrachromosomal DNA and RNA, and proteins. Nucleic acids, peptides and proteins that have been “isolated” thus include nucleic acids and proteins purified by standard purification methods. “Isolated” nucleic acids, peptides and proteins can be part of a composition and still be isolated if the composition is not part of the native environment of the nucleic acid, peptide, or protein. The term also embraces nucleic acids, peptides and proteins prepared by recombinant expression in a host cell as well as chemically synthesized nucleic acids. [0081] As used herein, the term “polynucleotide,” synonymously referred to as “nucleic acid molecule,” “nucleotides” or “nucleic acids,” refers to any polyribonucleotide or polydeoxyribonucleotide, which can be unmodified RNA or DNA or modified RNA or DNA. “Polynucleotides” include, without limitation single- and double-stranded DNA, DNA that is a mixture of single- and double-stranded regions, single- and double-stranded RNA, and RNA that is a mixture of single- and double-stranded regions, hybrid molecules comprising DNA and RNA that can be single-stranded or, more typically, double-stranded or a mixture of single- and double-stranded regions. In addition, “polynucleotide” refers to triple-stranded regions comprising RNA or DNA or both RNA and DNA. The term polynucleotide also includes DNAs or RNAs containing one or more modified bases and DNAs or RNAs with backbones modified for stability or for other reasons. “Modified” bases include, for example, tritylated bases and unusual bases such as inosine. A variety of modifications can be made to DNA and RNA; thus, “polynucleotide” embraces chemically, enzymatically or metabolically modified forms of polynucleotides as typically found in nature, as well as the chemical forms of DNA and RNA characteristic of viruses and cells. “Polynucleotide” also embraces relatively short nucleic acid chains, often referred to as oligonucleotides. [0082] As used herein, the term “vector” is a replicon in which another nucleic acid segment can be operably inserted so as to bring about the replication or expression of the segment. [0083] As used herein, the term “host cell” refers to a cell comprising a nucleic acid molecule of the invention. The “host cell” can be any type of cell, e.g., a primary cell, a cell in culture, or a cell from a cell line. In one embodiment, a “host cell” is a cell transfected with a nucleic acid molecule of the invention. In another embodiment, a “host cell” is a progeny or potential progeny of such a transfected cell. A progeny of a cell may or may not be identical to the parent cell, e.g., due to mutations or environmental influences that can occur in succeeding generations or integration of the nucleic acid molecule into the host cell genome. [0084] The term “expression” as used herein, refers to the biosynthesis of a gene product. The term encompasses the transcription of a gene into RNA. The term also encompasses 18 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 translation of RNA into one or more polypeptides, and further encompasses all naturally occurring post-transcriptional and post-translational modifications. The expressed gene product can be within the cytoplasm of a host cell, into the extracellular milieu such as the growth medium of a cell culture or anchored to the cell membrane. [0085] As used herein, the terms “peptide,” “polypeptide,” or “protein” can refer to a molecule comprised of amino acids and can be recognized as a protein by those of skill in the art. The conventional one-letter or three-letter code for amino acid residues is used herein. The terms “peptide,” “polypeptide,” and “protein” can be used interchangeably herein to refer to polymers of amino acids of any length. The polymer can be linear or branched, it can comprise modified amino acids, and it can be interrupted by non-amino acids. The terms also encompass an amino acid polymer that has been modified naturally or by intervention; for example, disulfide bond formation, glycosylation, lipidation, acetylation, phosphorylation, or any other manipulation or modification, such as conjugation with a labeling component. Also included within the definition are, for example, polypeptides containing one or more analogs of an amino acid (including, for example, unnatural amino acids, etc.), as well as other modifications known in the art. [0086] The peptide sequences described herein are written according to the usual convention whereby the N-terminal region of the peptide is on the left and the C-terminal region is on the right. Although isomeric forms of the amino acids are known, it is the L-form of the amino acid that is represented unless otherwise expressly indicated. [0087] As used herein, the term “low-to-moderate affinity peptide” can refer to any peptide that does not have a high affinity for an MHC-1 complex. A low affinity peptide can, for example, have an IC50 value of >500 nM for MHC I. A moderate affinity peptide can, for example, have an IC50 value of <500 nM and >50 nM for MHC I. A high affinity peptide generally has an IC50 value of <50 nM. Low-to-moderate affinity peptides can be excluded from consideration as candidates for vaccines and can be referred to as a “non-binder.” Low affinity peptides are known in the art, see, e.g., Ebrahimi-Nik et al., “Reversion Analysis Reveals the in vivo immunogenicity of a poorly MHC-I binding cancer neoepitope,” Nature Communications 12:6423 (2021). [0088] The term “administering” with respect to the methods of the invention, means a method for simulating an immune response as described herein by using a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 95% identity to SEQ ID NO:10 and an immunogenic agent of the invention or a form, composition or medicament thereof. Such methods include administering an effective amount of the TAP- 19 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 95% identity to SEQ ID NO:10 and the immunogenic agent at different times during the course of a simulating the immune response or concurrently in a combination form. [0089] The term “effective amount” means that amount of the TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 95% identity to SEQ ID NO:10 and the immunogenic agent that elicits the desired immune response in the subject, that is being sought by a researcher, veterinarian, medical doctor, or other clinician. Methods of increasing the immunogenicity of mammalian cells and methods of stimulating an immune response in a subject in need thereof [0090] To examine the editing function of TAPBPR and capture the biologically relevant peptide-bound state of its MHC-I complex, protein engineering has been leveraged herein to enhance the stability of MHC-I molecules that are loaded with suboptimal low-to-moderate- affinity peptides (37), as well as deep scanning mutagenesis for affinity maturation of TAPBPR towards peptide-loaded molecules (18). A high-fidelity chaperone, TAPBPRFLQ, containing three mutations (S104F, K211L, and R270Q) at MHC-I interaction sites was characterized and shown to significantly enhance peptide exchange for multiple HLA allotypes expressed on a cellular membrane. Finally, the cryoEM structure of open HLA- A*02:01/β2m loaded with a peptide decoy captured by TAPBPRFLQ was determined, revealing a key intermediate of the peptide exchange process. It was found that the transient proofreading complex utilizes (1) conserved groove residues to capture the backbone of incoming peptides in a native-like conformation, (2) properly conformed hydrophobic A-, B-, and D-pockets to dock the peptide P2 and P3 anchors, and (3) a conformationally heterogenous F-pocket that folds upon binding of high-affinity peptides, allosterically triggering the dissociation of TAPBPR. Taken together, the combined structural and biochemical analysis provides a complete mechanism of MHC-I antigen proofreading and peptide repertoire selection. [0091] Thus, provided herein are methods of increasing the immunogenicity of mammalian cells. The methods comprise providing a population of mammalian cells having a surface MHC class I molecule with an immunogenic agent and a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:7, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecules thereby increasing the 20 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 immunogenicity of the mammalian cells. In certain embodiments, the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11. In certain embodiments, the method is performed in vivo. [0092] Also provided are methods of producing antigen presenting cells to stimulate an immune response in a subject in need thereof. The methods comprise (a) obtaining antigen presenting cells from the subject in need thereof; and (b) providing antigen presenting cells comprising surface MHC class I molecules in vitro with a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:7 and an immunogenic agent, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecules thereby creating antigen presenting cells for stimulating an immune response. In certain embodiments, the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11. [0093] In certain embodiments, the TAPBPR polypeptide further comprises a signal peptide, a flag tag, a linker, and/or a transmembrane domain. The signal peptide can, for example, comprise SEQ ID NO:47. The flag tag can, for example, comprise SEQ ID NO:48. The linker can, for example, comprise SEQ ID NO:49. The transmembrane domain can, for example, comprise SEQ ID NO:50. [0094] In certain embodiments, the mammalian cell is a human cell. In certain embodiments, the mammalian cell is a human cell. [0095] A cell displaying MHC class I molecules can be exposed to (i) a soluble extracellular TAPBPR polypeptide as described herein; (ii) a cell having a surface bound TAPBPR polypeptide as described herein; or (iii) a chimeric TAPBPR as described in WO2023/163980, which is herein incorporated by reference in its entirety. [0096] The loading of cell-surface MHC class I molecules as described herein can increase the number of MHC class I molecules on the surface of a cell which present the immunogenic agent relative to cells not treated with the TAPBPR. For example, the number of MHC class I molecules on the surface of a cell which present the immunogenic agent may be increased by 30 fold or more, 40 fold or more, 50 fold or more, 60 fold or more, 70 fold or more, 80 fold or more, 90 fold or more, 100 fold or more, 150 fold or more or 200 fold or more of the immunogenic agent in the presence relative to the absence of TAPBPR. Cells may present none or substantially none of the immunogenic agent in the absence of treatment with the TAPBPR. 21 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 [0097] Mammalian cells or cells of the subject may comprise MHC class I molecules on the cell surface. MHC class I molecules are heterodimers comprising an α chain and δ2- microglobulin. MHC class I molecules are expressed on all nucleated human cells. An individual inherits a set of HLA-A, -B and -C genes from each parent. These genes are co- dominantly expressed and nucleated cells in mammals express up to 6 different classical MHC class I molecules. MHC class I molecules are highly polymorphic within the α chain and there is huge variation within the population. MHC class I molecules may include HLA- A molecules, HLA-B molecules, such as HLA-B51, HLA-Bl5, HLA-B38, and HLA-B57 and HLA-C molecules, such as HLA-Cwl. Preferred MHC class I molecules include HLA-A. [0098] In certain embodiments, the MHC class I molecule is selected from the group consisting of an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, an HLA-E molecule, an HLA-G molecule, an HLA-F molecule, a MR1 molecule, a CD1 molecule, and a combination thereof. The MHC class I molecule can, for example, be an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, a MR1 molecule, an HLA-E molecule, and/or an HLA-G molecule. The HLA-A molecule, HLA-B molecule, and/or HLA-C molecule can, for example, be selected from an HLA-A01 supertype molecule, an HLA-A02 supertype molecule, an HLA-A03 supertype molecule, an HLA-A24 supertype molecule, an HLA- A01/A24 supertype molecule, an HLA-A01/A03 supertype molecule, an HLA-B08 supertype molecule, an HLA-B44 supertype molecule, an HLA-B58 supertype molecule, and/or an HLA-C01 supertype molecule. In certain embodiments, the MHC class I molecule can, for example, be selected from HLA-A*01:01, HLA-A*11:01, HLA-A*29:01, HLA-A*30:01, HLA-A*32:01, HLA-A*74:01, HLA-A*24:03, HLA-A*02:06, HLA-A*02:01, HLA- A*02:03, HLA-A*23:01, HLA-A*24:02, HLA-A*68:01, HLA-A*69:01, HLA-A*68:02, HLA-A*29:02, HLA-A*03:01, HLA-B*08:01, HLA-B*57:03, HLA-B*59:01, HLA-B*37:01, HLA-C*01:02, HLA-C*03:04, MR1, HLA-E*01:03, and/or HLA-G*01:01. [0099] In some embodiments, the cells may be disease cells, such as cancer cells, cells infected with a pathogen, or other cells that cause disease. Increasing the immunogenicity of disease cells in a subject using a TAPBPR may generate or increase the strength of immune responses against the disease cells in the subject. This may lead to a reduction or eradication of disease cells in the individual and may exert a therapeutic effect. [00100] In other embodiments, the cells may be antigen presenting cells. Loading the surface MHC I molecules of antigen presenting cells with an exogenous immunogenic agent useful in 22 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 increasing or eliciting immune responses, for example T cell immune responses, against disease cells in vivo, can thereby also exert a therapeutic effect. [00101] In certain embodiments, the antigen presenting cells are dendritic cells. The loading of the surface MHC class I molecules with immunogenic peptides can increase the ability of the antigen presenting cells to induce immune responses, for example immune responses against the antigenic epitopes contained in the immunogenic peptide. Antigen presenting cells loaded with immunogenic peptides as described above may be used to stimulate T cells in vitro or ex vivo or administered to an individual to stimulate T cells in vivo. [00102] The immunogenic peptide can comprise one or more antigenic epitopes. The antigen presenting cells can activate T cells against the antigenic epitopes of the immunogenic peptide. For example, the antigenic epitopes of the immunogenic peptide can be present on disease cells in the subject. The antigen presenting cells can activate T cells capable of generating an immune response against the disease cells in the subject. [00103] In certain embodiments, the method further comprises administering the antigen presenting cells to the subject in need thereof. [00104] In certain embodiments, the immunogenic agent is selected from an immunogenic peptide, a metabolite, a lipid, a phospholipid, a fatty acid ligand, and/or combinations thereof. In certain embodiments, the immunogenic peptides are low-to-moderate affinity peptides. [00105] In some embodiments, the immunogenic peptide may comprise an antigen or an epitope that is characteristic of a disease cell. For example, the immunogenic peptide may comprise an antigen or an epitope that is characteristic of a cancer cell or a pathogen-infected cell. [00106] Epitopes that are characteristic of cancer cells are well known in the art and include epitopes from tumor antigens. Suitable antigens and epitopes are described elsewhere herein. Preferred tumor antigens from which immunogenic peptides may be derived include neoantigens, tumor-specific, differentiation and overexpressed proteins, such as ErbB2/Her2 (e.g., RLLQETELV (SEQ ID NO:12)), gpl00 (e.g., IMDQVPFSV (SEQ ID NO:13) and YLEPGPVTA (SEQ ID NO:14)), NY-Eso-1 (e.g., SLLMWITQC (SEQ ID NO:15)), p53 (e.g., LLGRNSFEV (SEQ ID NO:16)), MARTI (e.g., ELAGIGILTV (SEQ ID NO:17)), MAGE-10 (e.g., GLYDGMEHL (SEQ ID NO:18)), human AFP (e.g., FMNKFIYEI (SEQ ID NO:19)), Mesothelin (e.g., SLLFLLFSL (SEQ ID NO:20)), MAGE-A4 (e.g., GVYDGREHTV (SEQ ID NO:21)), MART-I (e.g., EAAGIGILTV (SEQ ID NO:22), ELAGIGILTV (SEQ ID NO:23)) and 5T4 (e.g., FLTGNQLAV (SEQ ID NO:24), RLARLALVL (SEQ ID NO:25). 23 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 [00107] Other tumor antigens and epitopes are well known in the art (see for example the Cancer Research Institute NY on-line peptide database; Tumor T cell antigen database, Olsen et al (2017) Cancer Immunol Immunother. doi: 10.1007/s00262-0l 7-1978-y; Immune Epitope and Analysis Resource, Vita et al. Nucleic Acids Res.2014 Oct.9. pii: gku938). [00108] Epitopes that are characteristic of pathogen-infected cells are well known in the art and include epitopes from viral proteins. Suitable epitopes for immunogenic peptides can include influenza epitopes (e.g., GILGFVFTL (SEQ ID NO:26), AIMDKNIIL (SEQ ID NO:27)), HIV epitopes (e.g., ILKEPVHGV (SEQ ID NO:28), SLYNTVATL (SEQ ID NO:29), KLTPLCVTL (SEQ ID NO:30)), hepatitis B epitopes (e.g., FLPSDFFPSV (SEQ ID NO:31), WLSLLVPFV (SEQ ID NO:32)), Human cytomegalovirus (CMV) epitopes (e.g., NLVPMVATV (SEQ ID NO:33), VLEETSVML (SEQ ID NO:34)), Epstein Barr virus (EBV) epitopes (e.g., YLLEMLWRL (SEQ ID NO:35), CLGGLLTMV (SEQ ID NO:36)), Varicella-zoster virus epitopes (e.g., ILIEGIFFV (SEQ ID NO:37)), Measles epitopes (e.g., ILPGQDLQYV (SEQ ID NO:38)), ZIKA (e.g., FLVEDHGFGV (SEQ ID NO:39), KSYFVRAAK (SEQ ID NO:40)), and Ebola virus epitopes. Other viral epitopes are well known in the art (see for example Immune Epitope and Analysis Resource, Vita et al Nucleic Acids Res.2014 Oct.9. pii: gku938). [00109] Also provided are methods of stimulating an immune response in a subject in need thereof, the method comprising (a) administering to the subject in need thereof a TAP- binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:7, wherein the TAPBPR polypeptide binds to an antigen presenting cell comprising a surface MHC class I molecule in the subject in need thereof; and (b) administering to the subject in thereof an immunogenic agent, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecule of the antigen presenting cell such that the antigen presenting cell stimulates an immune response in the subject in need thereof. In certain embodiments, the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11. [00110] In certain embodiments, the TAPBPR polypeptide further comprises a signal peptide, a flag tag, a linker, and/or a transmembrane domain. The signal peptide can, for example, comprise SEQ ID NO:47. The flag tag can, for example, comprise SEQ ID NO:48. The linker can, for example, comprise SEQ ID NO:49. The transmembrane domain can, for example, comprise SEQ ID NO:50. 24 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 [00111] In certain embodiments, the MHC class I molecule is selected from the group consisting of an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, an HLA-E molecule, an HLA-G molecule, an HLA-F molecule, a MR1 molecule, a CD1 molecule, and a combination thereof. The MHC class I molecule can, for example, be an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, a MR1 molecule, an HLA-E molecule, and/or an HLA-G molecule. The HLA-A molecule, HLA-B molecule, and/or HLA-C molecule can, for example, be selected from an HLA-A01 supertype molecule, an HLA-A02 supertype molecule, an HLA-A03 supertype molecule, an HLA-A24 supertype molecule, an HLA- A01/A24 supertype molecule, an HLA-A01/A03 supertype molecule, an HLA-B08 supertype molecule, an HLA-B44 supertype molecule, an HLA-B58 supertype molecule, and/or an HLA-C01 supertype molecule. In certain embodiments, the MHC class I molecule can, for example, be selected from HLA-A*01:01, HLA-A*11:01, HLA-A*29:01, HLA-A*30:01, HLA-A*32:01, HLA-A*74:01, HLA-A*24:03, HLA-A*02:06, HLA-A*02:01, HLA- A*02:03, HLA-A*23:01, HLA-A*24:02, HLA-A*68:01, HLA-A*69:01, HLA-A*68:02, HLA-A*29:02, HLA-A*03:01, HLA-B*08:01, HLA-B*57:03, HLA-B*59:01, HLA-B*37:01, HLA-C*01:02, HLA-C*03:04, MR1, HLA-E*01:03, and/or HLA-G*01:01. [00112] In certain embodiments, the subject is a human. [00113] In certain embodiments, the immunogenic agent is selected from an immunogenic peptide, a metabolite, a lipid, a phospholipid, a fatty acid ligand, and/or combinations thereof. In certain embodiments, the immunogenic peptides are low-to-moderate affinity peptides. [00114] In certain embodiments, the antigen presenting cells are dendritic cells. [00115] In certain embodiments, the subject in need thereof can have a disease or condition, such as a viral infection or other pathogenic infection, or cancer. [00116] In some preferred embodiments, the subject has cancer. A method of treatment of cancer in a subject can comprise administering the TAPBPR polypeptide to the subject, wherein TAPBPR polypeptide comprises a targeting domain which binds to cancer cells in the subject, and administering an immunogenic peptide to the subject, such that the TAPBPR polypeptide loads the immunogenic peptide onto surface MHC class I molecules of the cancer cells of the subject, thereby eliciting or increasing an immune response in the subject against the cancer cells. [00117] Cancer can be characterized by the abnormal proliferation of malignant cancer cells and can include leukemias, such as AML, CML, ALL and CLL, lymphomas, such as Hodgkin lymphoma, non-Hodgkin lymphoma and multiple myeloma, and solid cancers such as sarcomas, skin cancer, melanoma, bladder cancer, brain cancer, breast cancer, uterus 25 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 cancer, ovary cancer, prostate cancer, lung cancer, colorectal cancer, cervical cancer, liver cancer, head and neck cancer, esophageal cancer, pancreas cancer, renal cancer, adrenal cancer, stomach cancer, testicular cancer, cancer of the gall bladder and biliary tracts, thyroid cancer, thymus cancer, cancer of bone, and cerebral cancer, as well as cancer of unknown primary (CUP). [00118] In some embodiments, cancer cells within a subject can be immunologically distinct from normal somatic cells in the subject (i.e., the cancerous tumor may be immunogenic). For example, the cancer cells can be capable of eliciting a systemic immune response in the subject against one or more antigens expressed by the cancer cells. The tumor antigens that elicit the immune response can be specific to cancer cells or can be shared by one or more normal cells in the individual. In other embodiments, cancer cells within a subject cannot be immunologically distinct from normal somatic cells in the subject until MHC class I molecules on the surface of the cancer cells are loaded with exogenous immunogenic peptide using a TAPBPR polypeptide as described herein. [00119] A subject with cancer can display at least one identifiable sign, symptom, or laboratory finding that is sufficient to make a diagnosis of cancer in accordance with clinical standards known in the art. Examples of such clinical standards can be found in textbooks of medicine such as Harrison's Principles of Internal Medicine, 15th Ed., Fauci A S et al., eds., McGraw-Hill, New York, 2001. In some instances, a diagnosis of a cancer in a subject can include identification of a particular cell type (e.g., a cancer cell) in a sample of a body fluid or tissue obtained from the individual. [00120] In particular, treatment can include inhibiting cancer growth, including complete cancer remission, and/or inhibiting cancer metastasis. Cancer growth generally refers to any one of a number of indices that indicate change within the cancer to a more developed form. Thus, indices for measuring an inhibition of cancer growth include a decrease in cancer cell survival, a decrease in tumor volume or morphology (for example, as determined using computed tomographic (CT), sonography, or other imaging method), a delayed tumor growth, a destruction of tumor vasculature, improved performance in delayed hypersensitivity skin test, an increase in the activity of T cells, and a decrease in levels of tumor-specific antigens. Administration of modified T cells can improve the capacity of the subject to resist cancer growth, in particular growth of a cancer already present in the subject and/or decrease the propensity for cancer growth in the subject. [00121] In other preferred embodiments, the subject has a viral or pathogenic infection. 26 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 [00122] A method of treatment of a viral or pathogenic infection in a subject can comprise administering a TAPBPR polypeptide described above to the subject, wherein the TAPBPR polypeptide comprises a targeting domain which binds to the viral or pathogen-infected cells in the subject, and administering an immunogenic peptide to the subject, such that the TAPBPR polypeptide loads the immunogenic peptide onto surface MHC class I molecules of the viral or pathogen-infected cells of the subject, thereby eliciting or increasing an immune response in the subject against the viral or pathogen-infected cells. [00123] Viral infection can, for example, be HIV, EBV, CMV, hepatitis, influenza, polio, human papilloma virus, measles, mumps, rubella, chicken pox, ebola, or zika infection. TAPBPR polypeptides [00124] Also provided is an isolated TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity to SEQ ID NO:7. In certain embodiments, the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11. [00125] In certain embodiments, the TAPBPR polypeptide further comprises a signal peptide, a flag tag, a linker, and/or a transmembrane domain. The signal peptide can, for example, comprise SEQ ID NO:47. The flag tag can, for example, comprise SEQ ID NO:48. The linker can, for example, comprise SEQ ID NO:49. The transmembrane domain can, for example, comprise SEQ ID NO:50. [00126] TAPBPR polypeptides load MHC class I molecules on the surface of the cells with an immunogenic agent. The immunogenic agent can, for example, be an immunogenic peptide. The immunogenic peptide can, for example, be an exogenous peptide. An exogenous peptide is a peptide that is not generated naturally by the cells with the MHC class I molecules. For example, it may have been administered to the subject. Exogenous peptide may have the same amino acid sequence as an endogenous peptide that is generated naturally by the cells or a different amino acid sequence. [00127] In some embodiments, the immunogenicity of the exogenous peptide may be different to the immunogenicity of endogenous peptides displayed in the MHC class I molecules (i.e., it may be higher or lower). In other embodiments, the immunogenicity of the exogenous peptide may be the same as the immunogenicity of one or more endogenous peptides displayed in the MHC class I molecules. For example, the exogenous peptide may have the same amino acid sequence as one or more endogenous peptides. Loading of MHC 27 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 class I molecules with the exogenous peptide as described here may increase the total amount of peptide with the amino acid sequence that is displayed on the cells and may thereby increase the immunogenicity of the cells. [00128] An immunogenic peptide is an exogenous peptide that is capable of generating an immune response in a subject when loaded onto an MHC class I molecule. For example, the immunogenic peptide/MHC class I complex may be recognized by T cells. The presence of MHC class I molecules loaded with an immunogenic peptide on the surface of target cells may induce or increase immune responses against the target cells. [00129] Suitable immunogenic peptides are known in the art and may for example be candidates in vaccines for cancer or infection. In some embodiments, immunogenic peptides for loading onto MHC class I may be antigens naturally expressed on a subject’s own tumor; neoantigens or other peptides derived from tumors; or peptides derived from pathogens, such as viruses. [00130] In another general aspect, the invention relates to isolated nucleic acids encoding the TAPBPR polypeptides disclosed herein. It will be appreciated by those skilled in the art that the coding sequence of a protein can be changed (e.g., replaced, deleted, inserted, etc.) without changing the amino acid sequence of the protein. Accordingly, it will be understood by those skilled in the art that nucleic acid sequences encoding TAPBPR polypeptides of the invention can be altered without changing the amino acid sequences of the proteins. [00131] In another general aspect, the invention relates to vectors comprising the isolated nucleic acids disclosed herein. Any vector known to those skilled in the art in view of the present disclosure can be used, such as a plasmid, a cosmid, a phage vector or a viral vector. In some embodiments, the vector is a recombinant expression vector such as a plasmid. The vector can include any element to establish a conventional function of an expression vector, for example, a promoter, ribosome binding element, terminator, enhancer, selection marker, and origin of replication. The promoter can be a constitutive, inducible, or repressible promoter. A number of expression vectors capable of delivering nucleic acids to a cell are known in the art and can be used herein for production of a TAPBPR polypeptide in the cell. Conventional cloning techniques or artificial gene synthesis can be used to generate a recombinant expression vector according to embodiments of the invention. [00132] In another general aspect, the invention relates to a host cell comprising an isolated nucleic acid or a vector comprising a nucleic acid as disclosed herein. Any host cell known to those skilled in the art in view of the present disclosure can be used for recombinant expression of TAPBPR polypeptides of the invention. In some embodiments, the host cells 28 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 are E. coli TG1 or BL21 cells (for expression of, e.g., an scFv or Fab antibody), CHO-DG44 or CHO-K1 cells or HEK293 cells (for expression of, e.g., a full-length IgG antibody). According to particular embodiments, the recombinant expression vector is transformed into host cells by conventional methods such as chemical transfection, heat shock, or electroporation, where it is stably integrated into the host cell genome such that the recombinant nucleic acid is effectively expressed. In certain embodiments, the host cell is a mammalian cell. Pharmaceutical Compositions [00133] The TAPBPR polypeptide and/or the immunogenic agent can be administered alone, or usually the TAPBPR polypeptide and/or the immunogenic agent can be administered in the form of a pharmaceutical composition, which may comprise at least one component in addition to the TAPBPR polypeptide and/or the immunogenic agent. Thus, pharmaceutical compositions can comprise, in addition to the TAPBPR polypeptide and/or the immunogenic agent, a pharmaceutically acceptable excipient, carrier, buffer, stabilizer or other materials well known to those skilled in the art. The term “pharmaceutically acceptable” as used herein pertains to compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgement, suitable for use in contact with the tissues of a subject (e.g., human) without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio. Each carrier, excipient, etc., must also be “acceptable” in the sense of being compatible with the other ingredients of the formulation. The precise nature of the carrier or other material will depend on the route of administration, which may be by bolus, infusion, injection or any other suitable route. [00134] The TAPBPR polypeptide and/or the immunogenic agent can be administered in combination. In some embodiments, the TAPBPR polypeptide and/or the immunogenic agent can be formulated in the same pharmaceutical composition. In other embodiments, the TAPBPR polypeptide and/or the immunogenic agent can be formulated in separate pharmaceutical compositions. [00135] In some embodiments, the TAPBPR polypeptide and/or the immunogenic agent can be provided in a lyophilized form for reconstitution prior to administration. For example, a lyophilized TAPBPR polypeptide and/or immunogenic agent can be re-constituted in sterile water and mixed with saline prior to administration to an individual. [00136] For parenteral, for example sub-cutaneous, intra-tumoral, intra-muscular or intra-venous administration, e.g., by injection, the pharmaceutical composition comprising 29 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 the TAPBPR polypeptide and/or the immunogenic agent described herein, nucleic acid or cell can be in the form of a parenterally acceptable aqueous solution which is pyrogen-free and has suitable pH, isotonicity and stability. Those of relevant skill in the art are well able to prepare suitable solutions using, for example, isotonic vehicles, such as Sodium Chloride Injection, Ringers Injection, and Lactated Ringers Injection. Preservatives, stabilizers, buffers, antioxidants and/or other additives may be employed as required including buffers such as phosphate, citrate and other organic acids; antioxidants, such as ascorbic acid and methionine; preservatives (such as octadecyldimethylbenzyl ammonium chloride; hexamethonium chloride; benzalkonium chloride; benzethonium chloride; phenol, butyl or benzyl alcohol; alkyl parabens, such as methyl or propyl paraben; catechol; resorcinol; cyclohexanol; 3'- pentanol; and m-cresol); low molecular weight polypeptides; proteins, such as serum albumin, gelatin or immunoglobulins; hydrophilic polymers, such as polyvinylpyrrolidone; amino acids, such as glycine, glutamine, asparagines, histidine, arginine, or lysine; monosaccharides, disaccharides and other carbohydrates including glucose, mannose or dextrins; chelating agents, such as EDTA; sugars, such as sucrose, mannitol, trehalose or sorbitol; salt-forming counter-ions, such as sodium; metal complexes (e.g. Zn-protein complexes); and/or non-ionic surfactants, such as TWEEN™, PLURONICS™ or polyethylene glycol (PEG). Suitable carriers, excipients, etc., can be found in standard pharmaceutical texts, for example, Remington's Pharmaceutical Sciences, 18th edition, Mack Publishing Company, Easton, Pa., 1990. [00137] Pharmaceutical compositions and formulations can conveniently be presented in unit dosage form and can be prepared by any methods well known in the art of pharmacy. Such methods include the step of bringing into association the chimeric protein described herein with the carrier which constitutes one or more accessory ingredients. In general, the compositions are prepared by uniformly and intimately bringing into association the active compound with liquid carriers or finely divided solid carriers or both, and then, if necessary, shaping the product. [00138] A pharmaceutical composition comprising the TAPBPR polypeptide and/or the immunogenic agent, as described herein, can be administered alone or in combination with other treatments, either simultaneously or sequentially dependent upon the condition to be treated. EMBODIMENTS [00139] The invention provides also the following non-limiting embodiments. 30 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 [00140] Embodiment 1 is a method of increasing the immunogenicity of mammalian cells, the method comprising providing a population of mammalian cells having surface MHC class I molecule with an immunogenic agent and a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecules thereby increasing the immunogenicity of the mammalian cells. [00141] Embodiment 2 is the method of embodiment 1, wherein the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11. [00142] Embodiment 3 is the method of embodiment 1 or 2, wherein the MHC class I molecule is selected from the group consisting of an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, an HLA-E molecule, an HLA-G molecule, an HLA-F molecule, a MR1 molecule, a CD1 molecule, and a combination thereof. [00143] Embodiment 4 is the method of embodiment 3, wherein the MHC class I molecule is an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, a MR1 molecule, an HLA-E molecule, and/or an HLA-G molecule. [00144] Embodiment 5 is the method of embodiment 4, wherein the HLA-A molecule, HLA- B molecule, and/or HLA-C molecule is selected from an HLA-A01 supertype molecule, an HLA-A02 supertype molecule, an HLA-A03 supertype molecule, an HLA-A24 supertype molecule, an HLA-A01/A24 supertype molecule, an HLA-A01/A03 supertype molecule, an HLA-B08 supertype molecule, an HLA-B44 supertype molecule, an HLA-B58 supertype molecule, and/or an HLA-C01 supertype molecule. [00145] Embodiment 6 is the method of embodiment 4, wherein the MHC class I molecule is selected from HLA-A*01:01, HLA-A*11:01, HLA-A*29:01, HLA-A*30:01, HLA-A*32:01, HLA-A*74:01, HLA-A*24:03, HLA-A*02:06, HLA-A*02:01, HLA-A*02:03, HLA- A*23:01, HLA-A*24:02, HLA-A*68:01, HLA-A*69:01, HLA-A*68:02, HLA-A*29:02, HLA-A*03:01, HLA-B*08:01, HLA-B*57:03, HLA-B*59:01, HLA-B*37:01, HLA-C*01:02, HLA-C*03:04, MR1, HLA-E*01:03, and/or HLA-G*01:01. [00146] Embodiment 7 is the method of any one of embodiments 1-6, wherein the mammalian cell is a human cell. [00147] Embodiment 8 is the method of any one of embodiments 1-7, wherein the immunogenic agent is selected from an immunogenic peptide, a metabolite, a lipid, a phospholipid, a fatty acid ligand, and/or combinations thereof. 31 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 [00148] Embodiment 9 is the method of any one of embodiments 1-8, wherein the method is performed in vivo. [00149] Embodiment 10 is a method of producing antigen presenting cells to stimulate an immune response in a subject, the method comprising: (a) obtaining antigen presenting cells from the subject; and (b) providing antigen presenting cells comprising surface MHC class I molecules in vitro with a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7 and an immunogenic agent, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecules thereby creating antigen presenting cells for stimulating an immune response. [00150] Embodiment 11 is the method of embodiment 10, wherein the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11. [00151] Embodiment 12 is the method of embodiment 10 or 11, wherein the MHC class I molecule is selected from the group consisting of an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, an HLA-E molecule, an HLA-G molecule, an HLA-F molecule, a MR1 molecule, a CD1 molecule, and a combination thereof. [00152] Embodiment 13 is the method of embodiment 12, wherein the MHC class I molecule is an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, a MR1 molecule, an HLA-E molecule, and/or an HLA-G molecule. [00153] Embodiment 14 is the method of embodiment 13, wherein the HLA-A molecule, HLA-B molecule, and/or HLA-C molecule is selected from an HLA-A01 supertype molecule, an HLA-A02 supertype molecule, an HLA-A03 supertype molecule, an HLA-A24 supertype molecule, an HLA-A01/A24 supertype molecule, an HLA-A01/A03 supertype molecule, an HLA-B08 supertype molecule, an HLA-B44 supertype molecule, an HLA-B58 supertype molecule, and/or an HLA-C01 supertype molecule. [00154] Embodiment 15 is the method of embodiment 13, wherein the MHC class I molecule is selected from HLA-A*01:01, HLA-A*11:01, HLA-A*29:01, HLA-A*30:01, HLA- A*32:01, HLA-A*74:01, HLA-A*24:03, HLA-A*02:06, HLA-A*02:01, HLA-A*02:03, HLA-A*23:01, HLA-A*24:02, HLA-A*68:01, HLA-A*69:01, HLA-A*68:02, HLA- A*29:02, HLA-A*03:01, HLA-B*08:01, HLA-B*57:03, HLA-B*59:01, HLA-B*37:01, HLA-C*01:02, HLA-C*03:04, MR1, HLA-E*01:03, and/or HLA-G*01:01. 32 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 [00155] Embodiment 16 is the method of any one of embodiments 10-15, wherein the mammalian cell is a human cell. [00156] Embodiment 17 is the method of any one of embodiments 10-16, wherein the immunogenic agent is selected from an immunogenic peptide, a metabolite, a lipid, a phospholipid, a fatty acid ligand, and/or combinations thereof. [00157] Embodiment 18 is the method of any one of embodiments 10-17, wherein the antigen presenting cells are dendritic cells. [00158] Embodiment 19 is the method of any one of embodiments 10-18, wherein the method further comprises administering the antigen presenting cells to the subject. [00159] Embodiment 20 is a method of stimulating an immune response in a subject in need thereof, the method comprising: (a) administering to the subject in need thereof a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7, wherein the TAPBPR polypeptide binds to an antigen presenting cell comprising a surface MHC class I molecule in the subject; and (b) administering to the subject an immunogenic agent, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecule of the antigen presenting cell such that the antigen presenting cell stimulates an immune response in the subject. [00160] Embodiment 21 is the method of embodiment 20, wherein the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11. [00161] Embodiment 22 is the method of embodiment 20 or 21, wherein the MHC class I molecule is selected from the group consisting of an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, an HLA-E molecule, an HLA-G molecule, an HLA-F molecule, a MR1 molecule, a CD1 molecule, and a combination thereof. [00162] Embodiment 23 is the method of embodiment 22, wherein the MHC class I molecule is an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, a MR1 molecule, an HLA-E molecule, and/or an HLA-G molecule. [00163] Embodiment 24 is the method of embodiment 23, wherein HLA-A molecule, HLA- B molecule, and/or HLA-C molecule is selected from an HLA-A01 supertype molecule, an HLA-A02 supertype molecule, an HLA-A03 supertype molecule, an HLA-A24 supertype molecule, an HLA-A01/A24 supertype molecule, an HLA-A01/A03 supertype molecule, an 33 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 HLA-B08 supertype molecule, an HLA-B44 supertype molecule, an HLA-B58 supertype molecule, and/or an HLA-C01 supertype molecule. [00164] Embodiment 25 is the method of embodiment 23, wherein the MHC class I molecule is selected from HLA-A*01:01, HLA-A*11:01, HLA-A*29:01, HLA-A*30:01, HLA- A*32:01, HLA-A*74:01, HLA-A*24:03, HLA-A*02:06, HLA-A*02:01, HLA-A*02:03, HLA-A*23:01, HLA-A*24:02, HLA-A*68:01, HLA-A*69:01, HLA-A*68:02, HLA- A*29:02, HLA-A*03:01, HLA-B*08:01, HLA-B*57:03, HLA-B*59:01, HLA-B*37:01, HLA-C*01:02, HLA-C*03:04, MR1, HLA-E*01:03, and/or HLA-G*01:01. [00165] Embodiment 26 is the method of any one of embodiments 20-25, wherein the subject is a human. [00166] Embodiment 27 is the method of any one of embodiments 20-26, wherein the immunogenic agent is selected from an immunogenic peptide, a metabolite, a lipid, a phospholipid, a fatty acid ligand, and/or combinations thereof. [00167] Embodiment 28 is the method of any one of embodiments 20-27, wherein the antigen presenting cells are dendritic cells. [00168] Embodiment 29 is the method of any one of embodiments 1-28, wherein the TAPBPR polypeptide further comprises a signal peptide, a flag tag, a linker, and/or a transmembrane domain. [00169] Embodiment 30 is the method of embodiment 29, wherein: (a) the signal peptide comprises SEQ ID NO:47; (b) the flag tag comprises SEQ ID NO:48; (c) the linker comprises SEQ ID NO:49; and (d) the transmembrane domain comprises SEQ ID NO:50. [00170] Embodiment 31 is an isolated TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 95% identity to SEQ ID NO:10 or at least 90% identity to SEQ ID NO:7. [00171] Embodiment 32 is the isolated TAPBPR polypeptide of embodiment 31, wherein the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO: 9, SEQ ID NO:10, or SEQ ID NO:11. [00172] Embodiment 33 is the isolated TAPBPR polypeptide of embodiment 31 or 32, wherein the TAPBPR polypeptide further comprises a signal peptide, a flag tag, a linker, and/or a transmembrane domain. [00173] Embodiment 34 is the isolated TAPBPR polypeptide of embodiment 33, wherein: (a) the signal peptide comprises SEQ ID NO:47; 34 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 (b) the flag tag comprises SEQ ID NO:48; (c) the linker comprises SEQ ID NO:49; and (d) the transmembrane domain comprises SEQ ID NO:50. [00174] Embodiment 35 is an isolated nucleic acid encoding the TAPBPR polypeptide of any one of embodiments 30-34. [00175] Embodiment 36 is an isolated vector comprising the isolated nucleic acid of embodiment 35. [00176] Embodiment 37 is a host cell comprising the vector of embodiment 36. [00177] Embodiment 34 is the host cell of embodiment 37, wherein the cell is a mammalian cell. [00178] Without further description, it is believed that one of ordinary skill in the art can, using the preceding description and the following illustrative examples, make and utilize the present invention and practice the claimed methods. The following working examples, therefore, specifically point out the preferred embodiments of the present invention, and are not to be construed as limiting in any way the remainder of the disclosure. EXAMPLES [00179] The following Materials and Methods are used in Examples discussed below. Materials and Methods A. Peptides [00180] All peptide sequences are given as standard single-letter codes. High-affinity peptide TAX9 (LLFGYPVYV) (SEQ ID NO:1) was purchased from Genscript, USA; Piscataway, NJ, at >90% purity. The fluorophore-labeled and photolabile peptides, using TAMRA as 5- Carboxytetramethylrhodamine and J as Fmoc-3-amino-3-(2-nitrophenyl)-propionic acid, were purchased from Biopeptek Inc, Malvern, USA. Peptides were solubilized in distilled water and centrifuged at 14000 rpm for 15 minutes. The concentration of each peptide solution was measured and calculated using the absorbance and extinction coefficient at 205 nm wavelength. B. Recombinant protein expression, refolding, and purification [00181] DNA Plasmids encoding the luminal domain of MHC-I heavy chain HLA-A*02:01 and light chain human β2m were provided by the NIH tetramer facility (Emory University; Atlanta, GA). Plasmids encoding open HLA-A*02:01 (G120C) and open β2m (G120C) were site mutated in-house. Paired plasmids were individually transformed into Escherichia coli 35 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 BL21 (DE3) cells (New England Biolabs; Ipswich, MA). Proteins were expressed in the Luria-Broth medium, and inclusion bodies were collected and purified as previously described (1). For the generation of peptide-loaded HLA-A*02:01 (pHLA-A*02:01) molecules, in vitro refolding was performed by slowly diluting a 200 mg mixture of MHC-I heavy chain and β2m at a 1:3 molar ratio over 24 hours in refolding buffer (0.4 M L-Arginine HCl, 100 mM Tris pH 8, 2 mM EDTA, 4.9 mM reduced L-glutathione, 0.57 mM oxidized L- glutathione) containing 10 mg of the desired peptide. The mixture was protected from light when refolded with photolabile peptides. Refolding proceeded for four days, and proteins were purified by size exclusion chromatography (SEC) using a HiLoad 16/600 Superdex 75 pg column at 1 mL/min with 150 mM NaCl, 20 mM Tris buffer, pH 8.0. [00182] The luminal domain of TAPBPR and its variant proteins containing the non-disulfide bonded free cysteine mutation (C94S) tagged with BSP and 6-His was stably expressed in the Drosophila melanogaster S2 cell line (2). The cultures were induced with 1 mM CuSO4, and the supernatant was collected after four days. The secreted TAPBPR molecules were purified using a high-density metal affinity agarose resin (ABT, Madrid). The eluted proteins were further purified by SEC using a HiLoad 16/600 Superdex 200 pg column at a 1 mL/min flow rate in 150 mM NaCl, 20 mM sodium phosphate buffer, pH 7.4. C. Differential scanning fluorimetry (DSF) [00183] DSF was used to measure the thermal stability of the TAPBPR proteins and the pMHC-I molecules. TAPBPR protein or pMHC-I molecules (7 μM) were mixed with 10X SYPRO Orange dye in a buffer of 150 mM NaCl and 20 mM sodium phosphate (pH 7.2) to a final volume of 20 μl. Samples were loaded into a MicroAmp Optical 384-well plate and run in triplicates. The experiment was performed on a QuantStudio 5 real-time polymerase chain reaction (PCR) machine with excitation and emission wavelengths set to 470 and 569nm. The thermal stabilities of TAPBPR or pMHC-I were generated by plotting the first derivative of each melting curve and extracting the peak as the melting temperature (Tm). The thermal stability was measured by gradually increasing temperature at a rate of 1°C/min between 25° and 95°C. Data analysis and fitting were performed in GraphPad Prism v10. D. Surface plasmon resonance [00184] SPR experiments were conducted in triplicate using a Biacore X100 instrument (Cytiva) in SPR buffer (150 mM NaCl, 20 mM sodium phosphate pH 7.4, 0.1% Tween-20). Approximately 1000 resonance units (RU) of biotinylated TAX9/HLA-A*02:01 were immobilized at 10 μL/min on a streptavidin-coated chip (Cytiva). The chip was primed by flowing over 1 μM TAX9 peptide solution before each binding assay to prevent peptide 36 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 release from the pMHC-I molecules. Various concentrations from 0.1 up to 300 μM of TAPBPR and its variants were injected onto the chip at 25˚C at a flow rate of 30 μL/min for 60s followed by 180s dissociation time. The SPR sensorgrams and equilibrium dissociation constants (KD) were analyzed using the surface-bound analysis settings in Biacore X100 evaluation software (Cytiva). The representative SPR sensorgrams and fitted saturation curves were prepared in GraphPad Prism v9. The KDs of TAPBPRWT, TAPBPR∆ALAS, and TAPBPR∆G24-R36 are estimated values since saturation of binding cannot be reached. E. Fluorescence polarization [00185] The fluorescent peptide dissociation of the fluorophore-labeled peptide-loaded MHC-I was monitored by fluorescence polarization (FP). TAMRA-TAX9/HLA-A*02:01 protein at a concentration of 40 nM in the presence of 1 μM non-labeled TAX9 peptide was incubated without chaperone or with 1 μM TAPBPRTN6, TAPBPRWT, TAPBPR∆G24-R36, TAPBPRFLQ, and TAPBPR FLQ∆G24-R36. The kinetic dissociations were monitored for 10 hours, and fluorescence polarization were recorded approximately every 90 seconds. Excitation and emission values used to monitor the fluorescence of TAMRA-labeled peptides were 531 and 595 nm. All experiments were performed at RT in triplicates. Raw parallel (III) and perpendicular emission intensities (I⊥) were collected and converted to polarization (mP) values using the equation 1000*[(III-(G*I⊥))/(III+(G*I⊥))]. An optimized G-factor was determined to be 0.33 for TAMRA-labeled peptides in calculating baseline fluorescence and overall fluorescence polarization. The plateau for each dissociation was extracted by fitting one phase decay in GraphPad Prism 10. The relative dissociation was calculated using the ^^^^^^^^^^^^ = ^୪ୟ^^ୟ^^౫^^^౨ ^^^౯ି^୪ୟ^^ୟ ^^^౦^౨^^^ equation % ^^^^^^^^^^^^^^^^ ^^^^^^^^^^^^ ^ ^୪ୟ^^ୟ^^౫^^^౨ ^^^౯ . Error bar (SD) was F. Single antigen bead (SAB) screen [00186] TAPBPRFLQ, TAPBPRWT, and TAPBPRTN6 (7 μM) PE-tetramers were mixed with 4 μl of the LABScreen SAB suspension (OneLambda Inc., CA, USA) in a 96-well plate. The samples were incubated for 1 hour and 550 rpm at RT, washed four times in wash buffer (OneLambda Inc., CA, USA) to remove excess tetramers, and resuspended in phosphate- buffered saline (PBS; pH 7.2). For the negative controls, beads were incubated with the anti– HLA class I antibody W6/32 (Abcam, ab22432) at 550 rpm for 30 minutes at RT and washed three times before tetramer addition. To test the levels of peptide-loaded MHC-I molecules on the beads, we used the same W6/32 antibody and the secondary anti-mouse PE-conjugated antibody (Abcam, ab97024) for detection. The levels of TAPBPR bound to the beads were 37 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 measured using the Luminex 100 Liquid Array Analyzer System, and the results were analyzed in GraphPad Prism v10. G. Fluorescent peptide exchange [00187] 721.221 Human HLA negative B-lymphoblastoid cells and monoallelic HLA- A*02:01, 24:02, 23:01, and 03:01 722.211 cell lines were thawed and expanded in RPMI complete medium (RPMI-1640 with 25 mM HEPES & L-Glutamine, 10% heat-inactivated FBS, 1 mM sodium pyruvate, and 1% pen-strep). These cell lines were maintained at a density of 200,000 – 250,000 cells/mL. Cells were resuspended at a concentration of 2X106 cells/mL and 200,000 per well were plated and then stained with LIVE/DEAD™ Fixable Violet for 10 minutes at RT and for 30 minutes at 4°C in 2% BSA containing one of the following fluorophore-labeled antibodies: anti-HLA A2 antibody PE, anti-HLA A24 antibody FITC, anti-HLA A03 antibody APC. Cells were washed three rounds to remove excess unbound fluorophore-labeled antibodies The acquisition was then performed on CytoFLEX LX (Beckman Coulter), and the data were analyzed by FlowJo v10.10.0. [00188] Upon validating the surface expression of HLA-A*02:01, 24:02, 23:01, and 03:01, target cell lines were seeded at 200,000 cells per well for peptide exchange. These cells were then treated with recombinant TAPBPRFLQ, TAPBPRWT, and TAPBPRTN6 at different concentrations, as indicated, for 15 minutes at 37 °C. After 15 minutes, the corresponding fluorophore-labeled peptide (10nM TAMRA-KLFGYPVYV (SEQ ID NO:2) for HLA- A*02:01, 10nM TAMRA-KYNPIRTTF (SEQ ID NO:3) for HLA-24:02 and 23:01, 10 μM FITC-KLIETYFSK (SEQ ID NO:4) for HLA-A*03:01) was added to the cells and incubated at 37 for 60 minutes. Following the peptide treatment, the cells were washed three times in 1xPBS and harvested. The acquisition was performed on CytoFLEX LX (Beckman Coulter; Brea, CA) using the PE or FITC channel, and the data were analyzed by FlowJo v10.10.0. H. Lentiviral production, transduction, and peptide exchange on TAPBPR-TM transduced cells [00189] Three plasmids were used for lentivirus production: pMD2.G (envelope plasmid encoding VSV-G), psPAX2 (packaging plasmid), and pSFFV (transfer plasmid containing FLAG-tagged TAPBPR variants). Lenti-X 293 cells were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM) supplemented with 10% fetal bovine serum (FBS). For lentivirus production, Lenti-X 293 cells were grown to 70-80% confluence in a T75 flask and were co- transfected with psPAX2, pSFFV, and pMD2.G using Lipofectamine 3000. Lentivirus containing supernatant was collected every 24 hours for 3 total harvests and concentrated 38 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 using Lenti-X concentrator as suggested by the manufacturer (Takara; San Jose, CA). The viral pellet was resuspended in phosphate-buffered saline (PBS), flash-frozen in liquid nitrogen, and stored at -80°C for further use. Transductions were performed using retronectin-coated plates (Takara) and different titers of lentivirus along with 1x105 T2 cells in a 96-well plate. Cells were split every 1-2 days for up to 4 days before analysis by flow cytometry for TAPBPR expression for TAPBPRFLQ, TAPBPRWT, and TAPBPRTN6-TM transduced cell lines. Cells with similar levels of transduction efficiency and TAPBPR expression were selected for downstream assays, as measured by staining for FLAG (Clone: L5 in APC). Expression levels of cell surface A02 were quantified by staining with an anti- HLA-A2 antibody (Clone: BB7.2 in Brilliant Violet 785). T2 cells transduced with TAPBPR variants were incubated with indicated fluorescent TAX9-TAMRA peptide conjugate concentrations for 60 minutes in serum-free DMEM. Cells were then washed with FACS buffer, stained with LIVE/DEAD Fixable Near IR 876 (Invitrogen) for viability, and fixed with 4% PFA in 1x PBS. Cells were subsequently analyzed by flow cytometry using a CytoFlex LX cytometer. I. Peptide-loaded open HLA-A*02:01/TAPBPRFLQ complex purification [00190] Open KILGFVFJV (SEQ ID NO:46)/HLA-A*02:01/β2m developed with enhanced stability in previous work (3) was mixed with TAPBPRFLQ at a 1:1.3 molar ratio. The mixture was UV-irradiated at a high intensity in 150 mM NaCl, 20 mM sodium phosphate, pH 7.4 at 4˚C for 40 minutes with a wavelength of 365 nm. The complex was then purified by SEC using a Superdex 200 pg Increase 10/300 GL, and the eluted peaks were further analyzed by SDS–polyacrylamide gel electrophoresis (PAGE) to identify all components and confirm the complex. The open HLA-A*02:01/TAPBPRFLQ complex was then prepared at a 0.2 mg/mL concentration with 1XPBS, pH 7.4 for cryoEM. J. NMR Titrations and Chemical Shift Mapping [00191] NMR chemical shift mapping for the TAX9/HLA-A*02:01 molecule upon binding to TAPBPRFLQ was performed in an analogous fashion to TAPBPRWT, described previously (4). 13CH3 AILV-labeled HLA-A*02:01 at 103 μM was titrated with increasing concentrations of unlabeled TAPBPRFLQ in matched NMR buffer (20 mM sodium phosphate, 100 mM NaCl, 10% D2O (v/v) at pH 7.2). The HLA-A*02:01: TAPBPRFLQ ratios were: 1: 0, 1: 0.55, 1: 1.10, 1: 1.65, 1: 2.75, and 1: 3.31. Two-dimensional methyl 1H-13C methyl SOFAST HMQC experiments were recorded at 298 K at a field strength of 600 MHz with a total number of 136 scans, a recycle delay (d1) of 0.2 s and an acquisition time of 30 milliseconds in the indirect dimension. Chemical shift deviations (CSD) were computed 39 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 using the equation ΔδCH3 = [ ½ (Δδ1H2 + Δδ13C2/4)] ½ for the AILV methyl groups, where Δδ1H and Δδ13C are the changes in proton and carbon chemical shifts, respectively, upon saturation with TAPBPRFLQ. Using the TITAN software, the dissociation constant was obtained by fitting the slow-exchanging NMR resonances (5). For this analysis, the titration spectra were processed using an exponential window function with 4 Hz and 10 Hz line broadening in the direct and indirect dimensions, respectively, and fit using a two-state binding model in TITAN with bootstrap error analysis of 100 replicas. K. Liquid chromatography-mass spectroscopy (LC-MS) [00192] LC-MS was carried out with the passage of protein complex or peptide solution at a concentration range of 5-10 μM a Waters Acquity C8 column followed by electron ion spray–MS performed on a Waters Acquity UPLC SQD instrument with a mass detection range of 0-2000 m/z. Analysis and deconvolution of LC-MS data were performed with Xcalibur and UniDec. L. CryoEM sample preparation and data collection [00193] The CryoEM sample was prepared as previously described. Briefly, the open peptide-loaded HLA-A*02:01/TAPBPRFLQ complex was obtained by mixing open HLA- A*02:02/TAX9 and TAPBPRFLQ at a 1:1.3 molar ratio. The mixture was incubated at 4 ˚C for 1 hour and followed by 40-minute UV irradiation, and the complex was purified by size- exclusion chromatography. The peak fraction at 0.2 mg/mL concentration was used for grid preparation. The sample was applied to freshly plasma cleaned Quantifoil Cu 300 2/2 (Quantifoil; Germany) grids and was plunge frozen in liquid ethane using the Vitroblot Mark IV (Thermo Fisher; Waltham, MA) operated at 4°C and 100% humidity. [00194] The dataset was collected on a Titan Krios G3i 300 kV electron microscope (Thermo Fisher Scientific) with a 20 eV energy filter and equipped with a K3 Summit camera (Gatan). Super-resolution images were collected over thirty-five frames with a dose of 40.5 e-/A2 at a nominal magnification of x105,000, resulting in a pixel size of 0.418 Å/pixel. The defocus range was set from -0.8 to -3.0 μm. M. CryoEM Data Processing [00195] CryoEM data processing was performed using Relion 5.0 (6) and cryoSPARC 4.5 (7). 5,381 collected movies were motion corrected using the Relion implementation of MotionCor2 (8), binning to a pixel size of 0.836 Å/pixel. The defocus values were determined using CTFFIND-4 (9) and then suboptimal micrographs were removed, resulting in a set of 4,791 micrographs. From this point, processing proceeded in cryoSPARC. 40 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 Templates for particle picking were generated using blob picker and 2D classification on a subset of the dataset, which was also used to generate a good ab initio model. Template picking resulted in 4,707,523 particles, which were extracted and binned to a pixel size of 3.344 Å/pix. These particles were split evenly into five subsets and each subset was subjected to hetero refinement, using a single copy of the good ab initio model and five copies of a noise volume. After this first round of hetero refinement, a collective 1,119,002 particles were sorted into the good class. The 1,119,002 particles were combined and again split into two even subsets and subjected to the same hetero refinement protocol again, resulting in 628,062 particles sorted into the good class. The 628,062 particles were then combined and re-extracted to a pixel size of 1.672 Å/pixels, and any overlapping particles were removed. These particles were then subjected to eight additional rounds of the same hetero refinement protocol, until particle loss to the bad classes became negligible, resulting in a set of 340,226 particles. Throughout this process, particles from both the good class and the bad classes were subjected to 2D classification as a sanity check to ensure good particles were not discarded. These 340,226 particles were re-extracted to their final box size of 0.836 A/pixel and subjected to non-uniform refinement (10), resulting in a 3.3 Å structure. These particles were then transferred to Relion and reconnected to the original motion corrected micrographs. The 340,226 particles were refined in Relion using Blush regularization (11), resulting in a 3.3 Å structure. The use of Blush regularization significantly improved the map quality compared to cryoSPARC refinement or Relion refinement without Blush regularization. The particles were then subjected to two rounds of CTF refinement (12) and Bayesian polishing, resulting in a 3.0 Å structure. As significant heterogeneity for the peptide was still present, the 340,226 particles were then locally refined centered on the MHC peptide groove, followed by classification without angular assignment of the same region. This resulted in a set of 88,714 particles with the best density for the peptide in the MHC groove. These particles were then subjected to an overall refinement with Blush regularization, resulting in a 3.0 Å structure after postprocessing in Relion. N. Model building [00196] The model for the MHC-I/TAPBPR complex was built iteratively using ISOLDE (13), Coot (14), and the PHENIX (15) software package, using PDB 2VLL as the starting model. Images of the models and maps for figures were generated using ChimeraX (16) and Pymol (17). 41 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 Example 1: TAPBPRFLQ shows enhanced editing of peptide-loaded MHC-I [00197] While contradictory peptide unloading/levering (38, 39) or trapping (40) mechanisms have been proposed involving the TAPBPR G24-R36 loop, whether the loop actively participates in cargo editing of peptide-loaded MHC-I molecules remains unclear (35). This controversy was sought to be resolved and TAPBPR regions that promote interactions with peptide-loaded molecules for antigen editing were sought to be identified. Towards this, either wildtype TAPBPRWT or an engineered version TAPBPRFLQ, containing 3 mutations outside the loop region (18), were leveraged to directly measure binding to TAX9-loaded HLA-A*02:01 (A02) molecules by surface plasmon resonance (SPR). Experiments were run in the presence of a large molar excess of high-affinity TAX9 peptide to prevent peptide dissociation upon binding to TAPBPR. Different mutants of TAPBPRFLQ and TAPBPRWT, containing partial (A29-S32 deletion, TAPBPR∆ALAS) and complete loop deletions (G24-R36 deletion, TAPBPR∆G24-R36 and TAPBPRFLQ∆G24-R36) as well as TAPBPRTN6 (E205K, R207E, Q209S, and Q272S), which does not interact with peptide- loaded MHC-I (41, 30, 26) (FIG. 1A, FIG. 2, and FIG. 3) were also compared. It was demonstrated that TAX9-loaded A02 exhibited a significantly enhanced affinity (reduced KD) for TAPBPRFLQ, relative to TAPBPRWT (FIG. 1B and 1C). Meanwhile, loop deletions on both TAPBPRWT and TAPBPRFLQ had a more modest effect on binding to pMHC-I (FIG. 1B and 1C, FIG. 4), likely due to the loss of hydrophobic interactions with the rim of the MHC-I α1 and α2 helices, in agreement with previous mapping studies (38, 40). [00198] It was hypothesized that TAPBPR binding to peptide-loaded MHC-I molecules should correlate with peptide dissociation in vitro, and, thus, a fluorescence polarization (FP) assay employing A02 refolded with a TAMRA-labelled TAX9 peptide was applied to directly assess peptide unloading from the MHC-I (37). It was found that TAPBPRWT and its loop deletion mutant, TAPBPR∆G24-R36, demonstrated a similar low level of peptide unloading, relative to the negative controls (FIG. 1D and 1E, FIG. 5). Notably, both TAPBPRFLQ and TAPBPRFLQ∆G24-R36 significantly enhanced peptide unloading function (FIG.1D and 1E, FIG. 5). Introducing the G24-R36 loop deletion on either TAPBPRWT or TAPBPRFLQ had no significant impact on peptide editing function in vitro (FIG. 1E, FIG. 5). Taken together, the results suggest that, while the G24-R36 loop has an important role in maintaining TAPBPR’s structural integrity for pMHC-I recognition, surfaces outside the loop are essential for promoting interactions with peptide-loaded MHC-I and can be manipulated to further enhance the chaperone’s editing capability (18). 42 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 Example 2: TAPBPRFLQ promotes exogenous antigen loading on the cellular surface [00199] To evaluate the MHC-I allelic interaction landscape of TAPBPR, a single antigen bead (SAB) assay encompassing 97 common HLA allotypes was applied (18, 42). The phycoerythrin (PE) mean fluorescent intensity (MFI) of SABs upon incubation with TAPBPR PE-tetramers was measured. The levels of nonspecific background binding for TAPBPR PE-tetramers to HLAs were referenced relative to SABs that were pre-incubated with W6/32, a pan-allelic HLA monoclonal antibody that blocks the TAPBPR interaction surface on MHC-I, and further compared to the staining with TAPBPRTN6 PE-tetramer (FIG. 6 and FIG. 7A). Analysis of MFI ratios for TAPBPRFLQ revealed enhanced binding across multiple HLA allotypes, including HLA-A*02:01, HLA-A*02:03, HLA-A*02:06, HLA- A*69:01, HLA-A*68:01, HLA-A*23:01, HLA-A*24:02, and HLA-A*24:03, relative to TAPBPRWT (FIG. 1F, FIG. 7B and 7C). These results demonstrated that recombinant TAPBPRFLQ exhibits enhanced recognition of multiple HLA-A allotypes in vitro, suggesting that it can be used in non-native cellular compartments to promote antigen editing (26). [00200] Subsequently, whether TAPBPRFLQ could enhance the exchange of peptides on different MHC-I molecules expressed on a cellular surface was tested. Using a flow cytometry-based method, the loading of fluorophore-labeled peptides upon incubation with different soluble TAPBPR variants on monoallelic 722.211 cell lines expressing either HLA- A*02:01, 24:02, 23:01, or 03:01 (FIG. 8A, FIG. 9) was assessed. Consistent with the bead- based binding data, it was found that a nanomolar-range concentration of TAPBPRFLQ can readily exchange peptides on HLA-A*02:01, relative to the negative control TAPBPRTN6, whereas TAPBPRWT shows no significant enhancement in loading (FIG. 8B, FIG. 10A, and FIG. 11). Similarly, using monoallelic HLA-A*24:02 and HLA-A*23:01 cell lines, it was observed that TAPBPRFLQ could promote robust peptide exchange at a concentration that is 2 orders of magnitude lower than TAPBPRWT (FIG. 8B, FIG. 10B-10C, and FIGs. 12-13). For HLA-A*03:01, showing the lower levels of binding to TAPBPR PE-tetramers in the SAB assay, nanomolar soluble TAPBPRFLQ can promote loading of exogenous antigens, albeit the use of a higher peptide concentration relative to the other HLA allotypes (FIG.8B, FIG.10D, and FIG. 14). Finally, whether TAPBPRFLQ-TM (18) expressed on the cell surface of TAP transporter deficient T2 cells that lack the endogenous antigen processing machinery can directly enhance the loading of exogenous, fluorescently labeled peptides (FIG. 8C) was evaluated. The results demonstrated that TAPBPRFLQ can promote exchange at significantly lower peptide concentrations relative to T2 cells expressing TAPBPRWT-TM (FIG. 8D, FIG. 43 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 15). It was further demonstrated that surface-expressed TAPBPRFLQ-TM increased the expression level of HLA-A*02:01 on the T2 cell surface, as indicated by staining with the allele-specific BB7.2 anti-human A2 antibody PE (FIG. 16). A02 molecules on T2 cells expressing TAPBPRFLQ-TM were receptive to peptide binding at a picomolar peptide concentration (FIG.8D, FIG.17). In summary, the results demonstrated that TAPBPRFLQ is a high-fidelity TAPBPR variant that can have a wide range of applications in promoting peptide exchange across different HLA allotypes directly on the cell surface and serve as a bait to capture peptide-editing MHC-I/TAPBPR complex. Example 3: CryoEM structure of a TAPBPR/MHC-I complex bound to a peptide decoy [00201] Next, it was reasoned that the enhanced affinity of the TAPBPRFLQ mutant for HLA- I may enable the capture of transient low-affinity peptide-editing complexes that would not be resolvable with TAPBPRWT. Thus, to define the structural basis of peptide antigen proofreading by TAPBPR, the enhanced peptide editor, TAPBPRFLQ, was leveraged. A tertiary complex prepared using open HLA-A*02:01 (G120C) refolded with β2m (H31C) and a photocleavable peptide (KILGFVFJV (SEQ ID NO:46), J = 3-amino-3-(2-nitrophenyl)- propionic acid), used here as a conditional ligand (43, 44) was isolated. Upon UV irradiation, a soluble TAPBPRFLQ/A02 complex was purified by size-exclusion chromatography (FIG. 18A). Further analysis by LC-MS revealed that a 7-mer peptide fragment (KILGFVF) (SEQ ID NO:5) was captured within the TAPBPR/MHC-I complex (FIG. 18B, FIG. 19). The presence of a high-affinity complex was confirmed using NMR experiments performed by titrating unlabeled TAPBPRFLQ on isotopically methyl-labeled (AILV) HLA-A*02:01 using established methods (21, 31, 40). The NMR titrations further confirmed that TAPBPRFLQ binding to peptide-loaded HLA-A*02:01 was in the slow exchange regime, with a low micromolar-range affinity, as determined using a line shape analysis of the 2D methyl HSQC spectra (KD = 13.9 μM) (FIG. 20A-20B, FIG. 21, and Table 1). Analysis of methyl 13C/1H Chemical Shift Deviations (CSDs) further suggests that TAPBPRFLQ docks on MHC-I using an overall similar binding mode as determined in the previous solution mapping of interactions between TAX9/HLA-A*02:01 with TAPBPRWT (FIG.20C-20E) (21, 40). Finally, the structure of the purified complex was solved using cryoEM. Briefly, purified peptide- loaded open HLA-A*02:01/TAPBPRFLQ was used to screen grids, and more than 5,000 micrographs were collected (FIG. 22). This dataset revealed a fully assembled MHC- I/TAPBPR complex with a well-defined peptide-binding groove, though with some heterogeneity in peptide occupancy. The dataset was sorted to obtain a set of particles with 44 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 the best occupancy of the first residues of the peptide decoy, obtaining a structure at an overall resolution of 3.0 Å (FIGs.22-23, Table 2). [00202] Table 1: Dissociation equilibrium constants of TAPBPRFLQ binding to TAX9/HLA- A*02:01/β2m fitted from NMR line shape analysis of different methyl resonances. These residues undergo slow exchange and are used to obtain globally fitted KD (KD = 13.9 ± 0.7 μM) by performing line shape analysis using TITAN with bootstrap error analysis of 100 replicas. Residue KD (μM) I23δ1 11.7 ± 0.3 pMHC-I/TAPBPR complex EMD-45360 45 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 pMHC-I/TAPBPR complex EMD-45360 PDB 9C96 tron density reveals key chaperoning interactions with the nascent MHC-I (FIG. 18C). In agreement with previous studies (27, 28), it was observed that the N-terminal immunoglobulin V (IgV)–like domain of TAPBPRFLQ cradles the MHC-I peptide binding groove, while the C-terminal IgC domain nestles between the MHC-I α3 and β2m domains to create a pseudo 3-fold symmetric arrangement (FIG. 18C). Consistent with the crystal structures of empty mouse MHC-I/TAPBPR complexes (27, 28), as well as extensive mapping of human MHC-I/TAPBPR interactions by NMR, hydrogen-deuterium exchange, and deep scanning mutagenesis (18, 21, 31), the overall binding mode of TAPBPR was polarized towards the α2 helix of the MHC-I peptide binding groove (FIG.18C). Interactions between TAPBPR and the MHC-I groove were mediated through G212 and T259, forming polar contacts with Q115, K121, and D122 on strands β7 and β8, located under the peptide binding groove (FIG. 18D). Interactions between TAPBPR residues Q336 and F331 with L231 from α3 and residues I92-K94 from β2m was also observed, while D309 of TAPBPR formed a hydrogen bond with the T4 sidechain from β2m (FIG. 18E). In summary, a triad of domain interactions between β2m, MHC-I α3, and C terminal TAPBPR mediated stable docking, and allowed TAPBPR to stabilize the floor and α2-1 helix of the peptide binding groove, setting the stage for peptide editing. [00205] To understand the mechanism of peptide editing by TAPBPR and conformational changes induced by peptide binding to the MHC-I/TAPBPR complex, a structural comparison of the partially loaded TAPBPR/HLA-A*02:01/KILGFVF (SEQ ID NO:5) 46 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 cryoEM structure was performed (FIG. 24A) relative to the X-ray structure of fully loaded HLA-A*02:01/GILGFVFTL (SEQ ID NO:6) (45). It was found that the α2-1 helix, comprising residues E148-H151, adopted a conformation that was widened by approximately 3 Å in the TAPBPR-bound complex relative to the fully loaded pMHC-I structure (measured at the Cα atom of A150, FIG. 24B), in line with observations made on the basis of empty MHC-I/TAPBPR complexes (27, 28). Notably, in contrast to the previous empty TAPBPR/MHC-I crystal structures as well as the fully loaded pMHC-I structure, the cryoEM structure revealed a significant alteration of the α1 helix and adjacent loop (residues G71-G91) (FIG. 24B), where a lack of clear electron density indicates the presence of conformational heterogeneity impacting the F-pocket of the peptide-binding groove (FIG.25). This finding is in agreement with the previous NMR studies of the MHC-I/TAPBPR complex and reflects the dynamic transitions of the F-pocket between peptide-deficient and peptide-bound states in a solution environment (31). Compared to the original influenza epitope GILGFVFTL (SEQ ID NO:6), the heptamer peptide decoy within the TAPBPR editing complex exhibits a native- like backbone conformation and side chain rotamer placement (FIG. 24C, FIGs. 25-26). Despite the highly polymorphic character of the MHC-I groove, a relatively conserved cluster of residues (Y7, E63, K66, Y99, Y159, and Y171) located within the A-, B-, and D-pockets mediate stable docking interactions with the peptide backbone and hydrophobic sidechain anchors at P2 and P3 (FIG. 24C, FIG. 27). However, the dynamic character of the G71-G91 region in the editing complex leads to missing interactions between the C-terminus of the peptide with residues D77, T80, Y84, R97, T143, and W147 of the E- and F-pockets (FIG. 24C). Placed in the context of previous NMR and X-ray studies (21, 27, 28, 31, 40), the cryo- EM structure demonstrated that the initial capture of peptides for proofreading by TAPBPR occurred through native-like interactions with conserved MHC-I residues within the A-, B-, and D-pockets that were properly conformed to receive peptide. The F-pocket was partially formed in this intermediate state of the complex and folded upon annealing of the C-terminus of high-affinity peptides with robust P9 anchors to allosterically promote the dissociation of TAPBPR from the pMHC-I complex (31). Discussion [00206] The molecular chaperones tapasin and TAPBPR play important roles in stabilizing nascent MHC-I molecules, optimizing the repertoire of bound peptide cargo, and mediating quality control of peptide-loaded molecules, in tandem with other components of the MHC-I antigen processing pathway (1, 2, 46-48). Advances in sample preparation and the use of complementary structural techniques have provided a range of resting-state (empty) MHC- 47 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 I/chaperone complex structures, which have established the basis of tapasin and TAPBPR recognition of peptide-deficient MHC-I molecules (49, 50). Notwithstanding, the molecular mechanism of peptide editing is enigmatic and highly controversial in the literature (35). In previous work, solution NMR methyl probes have been used to study a peptide-bound MHC- I/TAPBPR intermediate, and it was shown that it involves a transient conformational state (approximately 200-millisecond lifetime) (31). A large body of work has established that dynamics at residues distributed throughout the MHC-I peptide binding groove were dampened upon binding of high-affinity peptides, and allosterically coupled to the TAPBPR binding surfaces on the underside and α2-1 helix of the groove to promote chaperone release from the pMHC-I (21, 31, 40). Recent X-ray structures of HLA-B*08:01 bound to 20mer peptides with protruding N-terminus have provided clues into the peptide-bound intermediate of the loading process (51). Despite these insights, a high-resolution structure of the transient peptide-bound chaperoned MHC-I has been missing due to its transient nature and dynamic complexity, hindering structure determination by X-ray crystallography or cryoEM. [00207] Here, recent advances in protein engineering of the individual components, a high- fidelity TAPBPRFLQ variant (18), and an ultra-stable, open HLA-A*02:01 molecule (37) prepared using a photocleavable conditional peptide ligand (52), were leveraged to isolate a TAPBPR/MHC-I intermediate bound to a heptamer peptide decoy. The insights gained from the peptide-loaded MHC-I/TAPBPR cryoEM structure allowed for the following proposed mechanism for peptide proofreading on MHC-I (FIG. 28). Empty MHC-I were preferentially recognized and stabilized by TAPBPR in a peptide-receptive, “open” state. Chaperoned MHC-I molecules screened the large peptide pool within the endoplasmic reticulum compartment, transiently forming the peptide/MHC-I/TAPBPR complex. Meanwhile, TABBPR widened the peptide binding groove via the α2-1 helix and enhanced the dynamic movements alongside the α1 helix and F pocket. As a result, the key peptide-coordinating residues within the E- and F-pockets lose their interactions with the peptide C-terminus. Therefore, TAPBPR-bound MHC-I exhibits an overall lower affinity towards incoming peptides and promotes the dissociation of transiently bound peptide decoys. When chaperoned MHC-I proofreads a high-affinity peptide, it interacts with the N terminal peptide through native-like contacts formed within the A-, B-, and D-pockets. The C-terminus of the high-affinity peptide interacts with peptide-coordinating residues in the E- and F- pockets to stabilize and close the peptide binding groove, which allosterically triggered the release of TAPBPR (31). The resulting high-affinity peptide-loaded MHC-I that has been proofread by TAPBPR can finally traffic to the cell surface for antigen presentation. 48 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 [00208] The TAPBPR G24-R36 loop has been previously suggested to directly mediate peptide editing by competing with the peptide C-terminus for binding to the MHC-I F-pocket (28, 38, 39). More recent studies using solution NMR have challenged this “scooping/ levering” mechanism, by explicitly showing that the TAPBPR loop adopts a disordered conformation, which forms hydrophobic contacts with the rim of the α2-1 helices, instead of entering the empty MHC-I groove in a solution environment (38-40). The engineered high- affinity TAPBPRFLQ corroborates this model, by showing that the S104F mutation on the edge of the TAPBPR loop contributes to enhanced binding on peptide-loaded MHC-I molecules. 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Tampé, Structure of the TAPBPR–MHC I complex defines the mechanism of peptide loading and editing. Science 358, 1060–1064 (2017). 53 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 29. G. Dong, P. A. Wearsch, D. R. Peaper, P. Cresswell, K. M. Reinisch, Insights into MHC class I peptide loading from the structure of the tapasin/ERp57 heterodimer. Immunity 30, 21–32 (2009). 30. G. I. Morozov, H. Zhao, M. G. Mage, L. F. Boyd, J. Jiang, M. A. Dolan, R. Venna, M. A. Norcross, C. P. McMurtrey, W. Hildebrand, P. Schuck, K. Natarajan, D. H. Margulies, Interaction of TAPBPR, a tapasin homolog, with MHC-I molecules promotes peptide editing. Proc. Natl. Acad. Sci. U. S. A.113, E1006–E1015 (2016). 31. A. C. McShan, K. Natarajan, V. K. Kumirov, D. Flores-Solis, J. Jiang, M. Badstübner, J. S. Toor, C. R. Bagshaw, E. L. Kovrigin, D. H. Margulies, N. G. Sgourakis, Peptide exchange on MHC-I by TAPBPR is driven by a negative allostery release cycle. Nat. Chem. 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Natarajan, Chaperone function in antigen presentation by MHC class I molecules—tapasin in the PLC and TAPBPR beyond. Front. Immunol.14, 1179846 (2023). 37. Y. Sun, M. C. Young, C. H. Woodward, J. N. Danon, H. V. Truong, S. Gupta, T. J. Winters, J. Font-Burgada, G. M. Burslem, N. G. Sgourakis, Universal open MHC-I molecules for rapid peptide loading and enhanced complex stability across HLA allotypes. Proc. Natl. Acad. Sci.120, e2304055120 (2023). 38. L. Sagert, F. Hennig, C. Thomas, R. Tampé, A loop structure allows TAPBPR to exert its dual function as MHC I chaperone and peptide editor. eLife 9, e55326 (2020). 54 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 39. F. T. Ilca, A. Neerincx, C. Hermann, A. Marcu, S. Stevanović, J. E. Deane, L. H. Boyle, TAPBPR mediates peptide dissociation from MHC class I using a leucine lever. eLife 7, e40126. 40. A. C. McShan, C. A. Devlin, G. I. Morozov, S. A. Overall, D. Moschidi, N. Akella, E. Procko, N. G. 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Natarajan, Chaperones and Catalysts: How Antigen Presentation Pathways Cope With Biological Necessity. Front. Immunol.13 (2022). 55 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 50. D. H. Margulies, J. Jiang, K. Natarajan, “Structure and Function of Molecular Chaperones that Govern Immune Peptide Loading” in Macromolecular Protein Complexes II: Structure and Function, J. R. Harris, J. Marles-Wright, Eds. (Springer International Publishing, Cham, 2019; https://doi.org/10.1007/978-3-030-28151-9_10)Subcellular Biochemistry, pp.321–337. 51. L. Li, X. Peng, M. Batliwala, M. Bouvier, Crystal structures of MHC class I complexes reveal the elusive intermediate conformations explored during peptide editing. Nat. Commun.14, 5020 (2023). 52. P. H. N. Celie, M. Toebes, B. Rodenko, H. Ovaa, A. Perrakis, T. N. M. Schumacher, UV-Induced Ligand Exchange in MHC Class I Protein Crystals. J. Am. Chem. Soc. 131, 12298–12304 (2009). [00209] It will be appreciated by those skilled in the art that changes could be made to the embodiments described above without departing from the broad inventive concept thereof. It is understood, therefore, that this invention is not limited to the particular embodiments disclosed, but it is intended to cover modifications within the scope of the present invention as defined by the present description. 56 4934-3954-3379.1

Claims

CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 CLAIMS It is claimed: 1. A method of increasing the immunogenicity of mammalian cells, the method comprising providing a population of mammalian cells having surface MHC class I molecule with an immunogenic agent and a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecules thereby increasing the immunogenicity of the mammalian cells. 2. The method of claim 1, wherein the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO: 8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11. 3. The method of claim 1, wherein the MHC class I molecule is selected from the group consisting of an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, an HLA-E molecule, an HLA-G molecule, an HLA-F molecule, a MR1 molecule, a CD1 molecule, and a combination thereof. 4. The method of claim 3, wherein the MHC class I molecule is an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, a MR1 molecule, an HLA-E molecule, and/or an HLA-G molecule. 5. The method of claim 4, wherein the HLA-A molecule, HLA-B molecule, and/or HLA-C molecule is selected from an HLA-A01 supertype molecule, an HLA-A02 supertype molecule, an HLA-A03 supertype molecule, an HLA-A24 supertype molecule, an HLA- A01/A24 supertype molecule, an HLA-A01/A03 supertype molecule, an HLA-B08 supertype molecule, an HLA-B44 supertype molecule, an HLA-B58 supertype molecule, and/or an HLA-C01 supertype molecule. 6. The method of claim 4, wherein the MHC class I molecule is selected from HLA- A*01:01, HLA-A*11:01, HLA-A*29:01, HLA-A*30:01, HLA-A*32:01, HLA-A*74:01, HLA-A*24:03, HLA-A*02:06, HLA-A*02:01, HLA-A*02:03, HLA-A*23:01, HLA- A*24:02, HLA-A*68:01, HLA-A*69:01, HLA-A*68:02, HLA-A*29:02, HLA-A*03:01, HLA-B*08:01, HLA-B*57:03, HLA-B*59:01, HLA-B*37:01, HLA-C*01:02, HLA- C*03:04, MR1, HLA-E*01:03, and/or HLA-G*01:01. 7. The method of any one of claims 1-6, wherein the mammalian cell is a human cell. 8. The method of any one of claims 1-6, wherein the immunogenic agent is selected from an immunogenic peptide, a metabolite, a lipid, a phospholipid, a fatty acid ligand, and/or combinations thereof. 57 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 9. The method of any one of claims 1-6, wherein the method is performed in vivo. 10. A method of producing antigen presenting cells to stimulate an immune response in a subject, the method comprising: (a) obtaining antigen presenting cells from the subject; and (b) providing antigen presenting cells comprising surface MHC class I molecules in vitro with a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7 and an immunogenic agent, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecules thereby creating antigen presenting cells for stimulating an immune response. 11. The method of claim 10, wherein the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:11. 12. The method of claim 10, wherein the MHC class I molecule is selected from the group consisting of an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, an HLA- E molecule, an HLA-G molecule, an HLA-F molecule, a MR1 molecule, a CD1 molecule, and a combination thereof. 13. The method of claim 12, wherein the MHC class I molecule is an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, a MR1 molecule, an HLA-E molecule, and/or an HLA-G molecule. 14. The method of claim 13, wherein the HLA-A molecule, HLA-B molecule, and/or HLA-C molecule is selected from an HLA-A01 supertype molecule, an HLA-A02 supertype molecule, an HLA-A03 supertype molecule, an HLA-A24 supertype molecule, an HLA- A01/A24 supertype molecule, an HLA-A01/A03 supertype molecule, an HLA-B08 supertype molecule, an HLA-B44 supertype molecule, an HLA-B58 supertype molecule, and/or an HLA-C01 supertype molecule. 15. The method of claim 13, wherein the MHC class I molecule is selected from HLA- A*01:01, HLA-A*11:01, HLA-A*29:01, HLA-A*30:01, HLA-A*32:01, HLA-A*74:01, HLA-A*24:03, HLA-A*02:06, HLA-A*02:01, HLA-A*02:03, HLA-A*23:01, HLA- A*24:02, HLA-A*68:01, HLA-A*69:01, HLA-A*68:02, HLA-A*29:02, HLA-A*03:01, HLA-B*08:01, HLA-B*57:03, HLA-B*59:01, HLA-B*37:01, HLA-C*01:02, HLA- C*03:04, MR1, HLA-E*01:03, and/or HLA-G*01:01. 16. The method of any one of claims 10-15, wherein the mammalian cell is a human cell. 58 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 17. The method of any one of claims 10-15, wherein the immunogenic agent is selected from an immunogenic peptide, a metabolite, a lipid, a phospholipid, a fatty acid ligand, and/or combinations thereof. 18. The method of any one of claims 10-15, wherein the antigen presenting cells are dendritic cells. 19. The method of any one of claims 10-15, wherein the method further comprises administering the antigen presenting cells to the subject. 20. A method of stimulating an immune response in a subject in need thereof, the method comprising: (a) administering to the subject in need thereof a TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7, wherein the TAPBPR polypeptide binds to an antigen presenting cell comprising a surface MHC class I molecule in the subject; and (b) administering to the subject an immunogenic agent, wherein the TAPBPR polypeptide loads the immunogenic agent onto the surface MHC class I molecule of the antigen presenting cell such that the antigen presenting cell stimulates an immune response in the subject. 21. The method of claim 20, wherein the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO: 9, SEQ ID NO:10, or SEQ ID NO:11. 22. The method of claim 20, wherein the MHC class I molecule is selected from the group consisting of an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, an HLA- E molecule, an HLA-G molecule, an HLA-F molecule, a MR1 molecule, a CD1 molecule, and a combination thereof. 23. The method of claim 22, wherein the MHC class I molecule is an HLA-A molecule, an HLA-B molecule, an HLA-C molecule, a MR1 molecule, an HLA-E molecule, and/or an HLA-G molecule. 24. The method of claim 23, wherein the HLA-A molecule, HLA-B molecule, and/or HLA-C molecule is selected from an HLA-A01 supertype molecule, an HLA-A02 supertype molecule, an HLA-A03 supertype molecule, an HLA-A24 supertype molecule, an HLA- A01/A24 supertype molecule, an HLA-A01/A03 supertype molecule, an HLA-B08 supertype molecule, an HLA-B44 supertype molecule, an HLA-B58 supertype molecule, and/or an HLA-C01 supertype molecule. 59 4934-3954-3379.1 CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 25. The method of claim 23, wherein the MHC class I molecule is selected from HLA- A*01:01, HLA-A*11:01, HLA-A*29:01, HLA-A*30:01, HLA-A*32:01, HLA-A*74:01, HLA-A*24:03, HLA-A*02:06, HLA-A*02:01, HLA-A*02:03, HLA-A*23:01, HLA- A*24:02, HLA-A*68:01, HLA-A*69:01, HLA-A*68:02, HLA-A*29:02, HLA-A*03:01, HLA-B*08:01, HLA-B*57:03, HLA-B*59:01, HLA-B*37:01, HLA-C*01:02, HLA- C*03:04, MR1, HLA-E*01:03, and/or HLA-G*01:01. 26. The method of any one of claims 20-25, wherein the subject is a human. 27. The method of any one of claims 20-25, wherein the immunogenic agent is selected from an immunogenic peptide, a metabolite, a lipid, a phospholipid, a fatty acid ligand, and/or combinations thereof. 28. The method of any one of claims 20-25, wherein the antigen presenting cells are dendritic cells. 29. The method of any one of claims 1-6, 10-15, or 20-25, wherein the TAPBPR polypeptide further comprises a signal peptide, a flag tag, a linker, and/or a transmembrane domain. 30. The method of claim 29, wherein: (a) the signal peptide comprises SEQ ID NO:47; (b) the flag tag comprises SEQ ID NO:48; (c) the linker comprises SEQ ID NO:49; and (d) the transmembrane domain comprises SEQ ID NO:50. 31. An isolated TAP-binding protein-related (TAPBPR) polypeptide comprising an amino sequence having at least 90% identity to SEQ ID NO:7. 32. The isolated TAPBPR polypeptide of claim 31, wherein the TAPBPR polypeptide comprises the amino acid sequence of SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO: 9, SEQ ID NO:10, or SEQ ID NO:11. 33. The isolated TAPBPR polypeptide of claim 31, wherein the TAPBPR polypeptide further comprises a signal peptide, a flag tag, a linker, and/or a transmembrane domain. 34. The isolated TAPBPR polypeptide of claim 33, wherein: (a) the signal peptide comprises SEQ ID NO:47; (b) the flag tag comprises SEQ ID NO:48; (c) the linker comprises SEQ ID NO:49; and (d) the transmembrane domain comprises SEQ ID NO:50. 35. An isolated nucleic acid encoding the TAPBPR polypeptide of any one of claims 31- 34. CHOP Ref.: 2025-002PCT Attorney Docket No.: 074313.11027/9WO1 36. An isolated vector comprising the isolated nucleic acid of claim 35. 37. A host cell comprising the vector of claim 36. 38. The host cell of claim 37, wherein the cell is a mammalian cell. 61 4934-3954-3379.1
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