Compositions comprising soluble HLA/M. tuberculosis-specific ligand complexes and methods of production and use thereof
Inventors
Hildebrand, William H. • McMurtrey, Curtis • Lewinsohn, David • Lewinsohn, Deborah • Harriff, Melanie
Assignees
Oregon Health and Science University • University of Oklahoma • US Department of Veterans Affairs • Office of General Counsel of VA
Publication Number
US-10857219-B2
Publication Date
2020-12-08
Expiration Date
2035-03-27
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Abstract
Compositions that include one isolated, class I HLA-E trimolecular complex that includes a peptide ligand unique to M. tuberculosis-infected cells are disclosed. Isolated compositions that include the three components of the trimolecular complex and/or a polynucleotide encoding one or more of the three components are also disclosed.
Core Innovation
The invention relates to compositions comprising one isolated, class I HLA-E trimolecular complex that includes a peptide ligand unique to Mycobacterium tuberculosis (Mtb)-infected cells, as well as isolated compositions with the components of the trimolecular complex or polynucleotides encoding them. The methods include producing soluble, truncated HLA-E molecules that bind Mtb-specific peptide ligands and forming trimolecular complexes for identification and utilization.
The problem being addressed is the difficulty in identifying Mtb-derived peptide ligands presented by HLA molecules of infected cells, especially by the non-classical, monomorphic HLA-E. Existing methods yield low peptide quantities and impure samples, causing ambiguous determination of which HLA allele presents peptides and limiting diagnostic and therapeutic targeting to subsets of the population. There is a need for effective identification of Mtb peptide ligands presented by HLA-E of Mtb-infected cells for improved diagnostic and treatment methods.
The invention solves these problems by producing soluble HLA-E molecules in vitro that capture endogenous, Mtb-specific peptide ligands from infected cell lines, allowing purification of high quantities of peptide-bound soluble HLA-E complexes. Comparative ligand mapping and deep ligand sequencing methods are applied to distinguish Mtb-specific peptides that are unique or substantially increased in infected cells compared to uninfected cells. The identified HLA-E/Mtb peptide complexes do not occur naturally and provide a basis for population-wide diagnostics, vaccines, and therapies due to the monomorphic nature of HLA-E.
Claims Coverage
The patent contains one independent claim directed to compositions comprising isolated class I HLA-E trimolecular complexes formed in vitro, each comprising a soluble, truncated HLA-E heavy chain, beta-2-microglobulin, and a unique Mtb-specific peptide ligand from a defined group.
Isolated class I HLA-E trimolecular complexes with specific Mtb peptide ligands
Compositions comprising soluble, truncated HLA-E heavy chains lacking transmembrane and cytoplasmic domains, beta-2-microglobulin, and peptides consisting of any one of SEQ ID NOS:1-29, where the peptide ligands are unique to Mtb-infected cells.
Use of soluble, truncated HLA-E heavy chains
The HLA-E heavy chain in the trimolecular complex is soluble, truncated, and recombinant, produced without the native transmembrane and cytoplasmic domains.
Production in recombinant host cells
The trimolecular complexes can be produced in host cells transfected with constructs encoding the soluble, truncated HLA-E heavy chain, with endogenous peptides loaded in vivo, or with synthetic peptides introduced in vitro or in vivo.
Peptide length specification
The peptides in the trimolecular complexes have lengths ranging from about 8 to about 20 amino acids.
Allele specificity
The compositions specify the use of HLA-E alleles HLA-E*01:01 or HLA-E*01:03 for the heavy chain in the trimolecular complexes.
The claims cover compositions of isolated, in vitro formed soluble class I HLA-E trimolecular complexes with beta-2-microglobulin and Mtb-specific peptide ligands defined by SEQ ID NOS:1-29, employing soluble, truncated HLA-E heavy chains preferably of alleles HLA-E*01:01 or HLA-E*01:03, produced recombinantly with peptides either endogenous or synthetic and of defined length, enabling targeted use of these complexes in diagnostics, therapeutics, and vaccine development.
Stated Advantages
Provides high purity and high concentration of soluble HLA-E peptide complexes, surpassing sensitivity of prior methods that have low yield and impurity.
Enables direct identification of Mtb-specific peptides presented by HLA-E of infected cells, overcoming ambiguity from previous indirect or computational methods.
The monomorphic nature of HLA-E allows for diagnostic and therapeutic applications that are applicable across the entire human population without restriction to subset groups.
The identified peptide/HLA-E complexes serve as biomarkers for Mtb infection, facilitating improved diagnostics, vaccine targets, and therapeutic strategies.
Documented Applications
Use of Mtb-specific HLA-E peptide ligands and trimolecular complexes as biomarkers for identifying Mtb-infected cells.
Development of diagnostics based on detection of Mtb-specific peptide/HLA-E epitopes.
Use of Mtb-specific peptides as vaccine candidates to elicit protective immune responses.
Therapeutic targeting of the entire HLA-E/peptide trimolecular complex for treatment of tuberculosis.
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