EP4580665A2 - Utilisation d'épitopes de lymphocytes t immunogènes pour la vaccination et le diagnostic de la maladie de lyme - Google Patents

Utilisation d'épitopes de lymphocytes t immunogènes pour la vaccination et le diagnostic de la maladie de lyme

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Publication number
EP4580665A2
EP4580665A2 EP23861597.5A EP23861597A EP4580665A2 EP 4580665 A2 EP4580665 A2 EP 4580665A2 EP 23861597 A EP23861597 A EP 23861597A EP 4580665 A2 EP4580665 A2 EP 4580665A2
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EP
European Patent Office
Prior art keywords
seq
polypeptide
lyme disease
subject
composition
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
EP23861597.5A
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German (de)
English (en)
Inventor
Robert B. LOCHHEAD
Rebecca L. DANNER
Joseph R. ROUSE
Lauren PROCHNIAK
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Medical College of Wisconsin
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Medical College of Wisconsin
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Publication date
Application filed by Medical College of Wisconsin filed Critical Medical College of Wisconsin
Publication of EP4580665A2 publication Critical patent/EP4580665A2/fr
Pending legal-status Critical Current

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    • 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
    • A61K39/02—Bacterial antigens
    • A61K39/0225—Spirochetes, e.g. Treponema, Leptospira, Borrelia
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/04—Antibacterial agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/195—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from bacteria
    • C07K14/20—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from bacteria from Spirochaetales (O), e.g. Treponema, Leptospira
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
    • Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
    • Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Definitions

  • compositions and methods for the treatment and/or preventative treatment of Lyme disease in a subject are also disclosed. Also disclosed are methods for detecting Lyme disease in a subject in need thereof, or detecting exposure to Borrelia spp.
  • polypeptides comprising or consisting of fragments of BB0680 (MCP4) or variants thereof.
  • the polypeptides comprise or consist of one or more of the polypeptides presented in Tables 1 and 2. In some QB ⁇ 650053.01002 ⁇ 84442353.1 Page 1 of 153 Atty. Dkt.
  • the polypeptide comprises or consists of SEQ ID NO: 1, or a polypeptide at least about 90% identical to SEQ ID NO: 1.
  • compositions comprising the polypeptides of the present disclosure.
  • the composition comprises a polypeptide comprising or consisting of SEQ ID NO: 1, and optionally an adjuvant.
  • the composition comprises one or more additional polypeptides, such as OspA, OspC, or one or more polypeptides listed in Tables 1 and 2.
  • additional polypeptides such as OspA, OspC, or one or more polypeptides listed in Tables 1 and 2.
  • polypeptides comprising or consisting of fragments of BB0838 (LptD) or variants thereof.
  • the polypeptides comprise or consist of one or more of the polypeptides presented in Tables 3 and 4.
  • the polypeptide comprises or consists of SEQ ID NO: 2, or a polypeptide at least about 90% identical to SEQ ID NO: 2.
  • compositions comprising the polypeptides of the present disclosure.
  • the composition comprises a polypeptide comprising or consisting of SEQ ID NO: 2, and optionally an adjuvant.
  • the composition comprises one or more additional polypeptides, such as OspA, OspC, or one or more polypeptides in listed in Tables 3 and 4.
  • compositions disclosed herein can include one or more polypeptides disclosed herein, such as a polypeptide comprising or consisting of SEQ ID NO:1, a polypeptide at least about 90% identical to SEQ ID NO: 1, a polypeptide comprising or consisting of SEQ ID NO: 2, a polypeptide at least about 90% identical to SEQ ID NO: 2, one or more polypeptides listed in Tables 1, 2, 3, 4, and 5, or a combination thereof.
  • polypeptides disclosed herein such as a polypeptide comprising or consisting of SEQ ID NO:1, a polypeptide at least about 90% identical to SEQ ID NO: 1, a polypeptide comprising or consisting of SEQ ID NO: 2, a polypeptide at least about 90% identical to SEQ ID NO: 2, one or more polypeptides listed in Tables 1, 2, 3, 4, and 5, or a combination thereof.
  • the composition disclosed herein can include one or more of: a polypeptide at least 90% or 95% identical to SEQ ID NO: 1, a polypeptide at least 90% or 95% identical to SEQ ID NO: 2, a polypeptide at least 90% or 95% identical to SEQ ID NO: 3, or a polypeptide at least 90% or 95% identical to SEQ ID NO: 4.
  • a polypeptide at least 90% or 95% identical to SEQ ID NO: 1 a polypeptide at least 90% or 95% identical to SEQ ID NO: 2
  • a polypeptide at least 90% or 95% identical to SEQ ID NO: 3 or a polypeptide at least 90% or 95% identical to SEQ ID NO: 4.
  • methods of treating Lyme disease or a method for the preventative treatment of Lyme disease, the method comprising: administering to a subject the polypeptide or composition as disclosed herein.
  • the subject is asymptomatic for Lyme disease, but may have come in contact with a blacklegged tick selected from Ixodes scapularis or Ixodes pacificus.
  • the subject is asymptomatic.
  • the subject is symptomatic, and exhibits one or more of the following symptoms: erythema migrans, facial palsy, and arthritis; and/or the subject tests positive for Lyme disease in standard two-tier test (STT).
  • STT standard two-tier test
  • kits comprising one or more polypeptides as disclosed herein, and optionally instructions for use of the kit.
  • the one or more polypeptides is linked to a solid support, and comprise or consist of SEQ ID NO:1, a polypeptide at least about 90% identical to SEQ ID NO: 1, a polypeptide comprising or consisting of SEQ ID NO: 2, a polypeptide at least about 90% identical to SEQ ID NO: 2, or a combination thereof.
  • FIG.1 Overview of methodology used to identify immunogenic MHC II peptides in mice infected with Borrelia burgdorferi.
  • FIG. 2A-2B Predicted structure of BB0680 (MCP4), with MCP442-462 epitope (Peptide 1) highlighted within the methyl-accepting transducer domain.
  • FIG.3 Percent identify of MCP4 protein in various Borrelia strains, including Bb sensu stricto, Bb sensu lato, and non-Lyme disease spirochete Borrelia hermsii, a relapsing fever spirochete.
  • FIG.4A-4B CD4+ T cell epitope MCP442-462 (Peptide 1) is immunogenic in mice infected with B. burgdorferi. Lymphocytes from mice infected with B. Burgdorferi for 4 weeks were stained with a proliferation dye (Cell-Trace Violet) and stimulated with Peptide 1 or OVA peptide as a negative control and incubated for 5 days.
  • a proliferation dye Cell-Trace Violet
  • FIG. 5 CLUSTAL O(1.2.4) multiple sequence alignment of BB0680 (MCP4).
  • B.hermsii is a relapsing fever Borrelia.
  • B. afzelii, B. garinii, B. garinii-barariensis, and B. mayonii are Bb sensu lato strains.
  • FIG. 5 includes SEQ ID NOS 1309-1317, respectively, in order of appearance from top to bottom.
  • FIG. 6 Table showing MHC class II peptides derived from Borrelia burgdorferi identified by LC/MS/MS.
  • FIG.6 discloses SEQ ID NOS 1324-1325, and 3-4, respectively, in order of appearance.
  • FIG.7 Structure of LptD (BB0838) based on Phyre2 modeling.
  • the C-terminal domain which includes LptD 923-934 , consists of a ⁇ -barrel and an unresolved C-terminal sequence, possibly a plug (yellow).
  • FIG.8 Sequence alignment and phylogenic analysis of LptD across Lyme disease spirochetes. ClustalW as used to align LptD protein sequences from Borrelia burgdorferi, Borrelia califorensis, Borrelia mayonii, Borrelia garinii, Borrelia afzelii, Borrelia baveriensis. The alignment and amino acid conservation of the 50 amino acids downstream from the LptD923-934 epitope (underlined) is shown.
  • Brackets indicate promiscuous core epitopes that are predicted to bind to a large number of HLA-DRB1 alleles (see FIGS.11 and 12 for complete list).
  • LptD sequence from relapsing fever spirochete Borrelia hermsii was used as an outgroup in generation of a phylogenetic relationship tree (below the alignment), with numbers indicating the Baysian distance from the consensus sequence.
  • FIG. 8 discloses SEQ ID NOS 1318-1323, respectively, in order of appearance from top to bottom.
  • FIG.9. Is a schematic depiction of a proposed use of T cell vaccine components for use in a novel Lyme disease vaccine.
  • Lyme disease affects nearly 500,000 individuals in the U.S. annually and is caused by infection with the tick-borne spirochete Borrelia burgdorferi. Although most patients are effectively treated with antibiotic therapy, ⁇ 10% of patients with Lyme disease have persistent symptoms that continue for months to years following antibiotic therapy and apparent spirochetal killing, called post-treatment Lyme disease syndrome (PTLDS), sometimes referred to as “chronic Lyme disease.”
  • PTLDS post-treatment Lyme disease syndrome
  • T helper cells also called CD4+ T cells
  • CD4+ T cells are the “generals” of the adaptive immune system. They regulate all aspects of immune responses to pathogens by directing other components of the immune system to find, neutralize, and kill pathogens during infection, and promote immune resolution and tissue repair post-infection. Dysregulated CD4+ T cell responses can also drive numerous chronic inflammatory, arthritic, neurodegenerative, and fibrotic diseases. B. burgdorferi has evolved mechanisms to evade CD4+ T cell responses during infection.
  • BB0680 is a methyl-accepting chemotaxis protein that is highly conserved amongst both sensu lato and sensu stricto strains of Lyme disease spirochetes (see FIG.3 and FIG.5).
  • Peptide 1 is highly immunogenic in mice infected with B. burgdorferi (see FIGS. 4A and 4B), validating Peptide 1 as target of CD4+ T cell responses to B. burgdorferi infection.
  • Peptide 1 is one of a number of peptides derived from MCP4 predicted to bind to MHC class II alleles in mice and/or humans (Table 1 below (mouse), Table 2 below (human)).
  • Peptide 1 has the amino acid sequence of SEQ ID NO: 1 and is provided below: [0028] KASLEVASSSQNLSSSALQQA [0029] Another peptide identified in the screen was derived from BB0838 (LptD923-934), hereinafter Peptide 2 (SEQ ID NO: 2).
  • LptD is an outer membrane protein (FIG. 7) involved in translocation of lipoproteins such as OspA and OspC from the periplasm to the outer membrane and is essential for cell growth.
  • the LptD 923-934 epitope is located at the interface between the central ⁇ -barrel domain and the C-terminal ⁇ -barrel/plug domain (FIG.7). Based on structure modeling, this epitope is localized at or near the cell surface, within the outer membrane-spanning ⁇ -barrel (FIG.7).
  • LptD is highly conserved amongst both sensu lato and QB ⁇ 650053.01002 ⁇ 84442353.1 Page 5 of 153 Atty. Dkt.
  • Peptide 2 is one of a number of peptides derived from LptD predicted to bind to MHC class II alleles in humans (Table 3 and Table 4 below). It is anticipated that immunization with Peptide 2 will lead to protective immunity in humans and mice. It also anticipated that other epitopes derived from LptD are immunogenic in humans and mice infected with B. burgdorferi.
  • T and B cell reactivity to LptD in humans with early and late Lyme disease, and in patients with PTLDS will be measured.
  • Peptide 2 has the amino acid sequence of SEQ ID NO: 2 and is provided below: [0031] NVFDFQFLFAMK [0032] A peptide from BB0559 (Crr) was also identified in the screen (Crr 123-138 ), hereinafter Peptide 3 (SEQ ID NO: 3). Crr is a PTS system glucose-specific component.
  • Peptide 3 has the amino acid sequence of SEQ ID NO: 3 and is provided below: [0034] HSESVITPVVIANSDE [0035] A peptide from oppAIV was also identified in the screen (oppAIV138-152), herein after Peptide 4 (SEQ ID NO: 4). oppAIV is an oligopeptide ABC transporter. [0036] Peptide 4, has the amino acid sequence of SEQ ID NO: 4 and is provided below: [0037] NAEEYFDGKANESE [0038] Additional immunogenic T cell epitopes are identified and described with respect to Table 5 below in the Examples.
  • compositions and methods for treating, detecting, and diagnosing infection by a Borrelia sp. may be further described using definitions and terminology as follows.
  • the definitions and terminology used herein are for the purpose of describing particular embodiments only, and are not intended to be limiting.
  • QB ⁇ 650053.01002 ⁇ 84442353.1 Page 6 of 153 Atty. Dkt. No.650053.01002 [0041]
  • the singular forms “a,” “an,” and “the” include plural forms unless the context clearly dictates otherwise.
  • the terms “include” and “including” have the same meaning as the terms “comprise” and “comprising.”
  • the terms “comprise” and “comprising” should be interpreted as being “open” transitional terms that permit the inclusion of additional components further to those components recited in the claims.
  • the terms “consist” and “consisting of” should be interpreted as being “closed” transitional terms that do not permit the inclusion of additional components other than the components recited in the claims.
  • the term “consisting essentially of” should be interpreted to be partially closed and allowing the inclusion only of additional components that do not fundamentally alter the nature of the claimed subject matter.
  • No.650053.01002 having 1-3 members refers to groups having 1, 2, or 3 members.
  • a group having 6 members refers to groups having 1, 2, 3, 4, or 6 members, and so forth.
  • the modal verb “may” refers to the preferred use or selection of one or more options or choices among the several described embodiments or features contained within the same. Where no options or choices are disclosed regarding a particular embodiment or feature contained in the same, the modal verb “may” refers to an affirmative act regarding how to make or use and aspect of a described embodiment or feature contained in the same, or a definitive decision to use a specific skill regarding a described embodiment or feature contained in the same.
  • the modal verb “may” has the same meaning and connotation as the auxiliary verb “can.”
  • sequences are then said to be “substantially identical.”
  • This definition also refers to the complement of a test sequence, in the context of nucleic acids.
  • the identify exists over a region that is about or at least about 5, 10, 15, 20, 50, 100, or 1000, amino acids in length, to about, less than about, or at least about 220, 100 or 1000 amino acids or nucleotides in length.
  • the identity exists over a region that is at least about 5, 10, 15, 20, 21 amino acids in length (e.g., with reference to SEQ ID NO: 1) to about 100, about 20 to about 75, about 30 to about 50 amino acids or nucleotides in length.
  • Non-limiting examples of polypeptide sequences provided herein comprise sequences that are substantially identical to SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, or SEQ ID NO: 4.
  • polypeptides that are at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% identical to SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, or SEQ ID NO: 4 is provided herein.
  • Polypeptides comprising a difference of 1, 2, 3, 4, 5, 6 or 7, 8, 9 or 0 amino acids as compared to SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, or SEQ ID NO: 4 are also contemplated herein.
  • sequence comparison typically one sequence acts as a reference sequence, to which test sequences are compared.
  • test and QB ⁇ 650053.01002 ⁇ 84442353.1 Page 8 of 153 Atty. Dkt. No.650053.01002 reference sequences are entered into a computer, subsequence coordinates are designated, if necessary, and sequence algorithm program parameters are designated. Preferably, default program parameters can be used, or alternative parameters can be designated.
  • sequence comparison algorithm then calculates the percent sequence identities for the test sequences relative to the reference sequence, based on the program parameters.
  • BLAST and BLAST 2.0 are described in Altschul et al., Nuc. Acids Res.25:3389-3402 (1977) and Altschul et al., J. Mol. Biol. 215:403-410 (1990), respectively.
  • the software for performing BLAST analyses is publicly available through the website of the National Center for Biotechnology Information (NCBI).
  • NCBI National Center for Biotechnology Information
  • BLAST and BLAST 2.0 are used, with the parameters described herein, to determine percent sequence identity for the nucleic acids and proteins.
  • a BLAST 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 are extended in both directions along each sequence for as far as the cumulative alignment score can be increased.
  • 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).
  • 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 NCBI BLASTN or BLASTP program is used to align sequences.
  • the BLASTN or BLASTP program uses the defaults used by the NCBI.
  • the BLASTN program (for nucleotide sequences) uses as defaults: a word size (W) of 28; an expectation threshold (E) of 10; max matches in a query range set to 0; match/mismatch scores of 1, ⁇ 2; linear gap costs; the filter for low complexity regions used; and mask for lookup table only used.
  • the BLASTP program (for amino acid sequences) uses as defaults: a word size QB ⁇ 650053.01002 ⁇ 84442353.1 Page 9 of 153 Atty. Dkt.
  • polypeptide polypeptide
  • peptide protein
  • W amino acid residues
  • E expectation threshold
  • max matches in a query range set to 0
  • BLOSUM62 matrix see Henikoff & Henikoff, Proc. Natl. Acad. Sci. USA 89:10915 (1992)
  • gap costs of existence: 11 and extension: 1 and conditional compositional score matrix adjustment.
  • polypeptide polypeptide
  • protein protein
  • amino acid polymers in which one or more amino acid residue is an artificial chemical mimetic of a corresponding naturally occurring amino acid, as well as to naturally occurring amino acid polymers and non-naturally occurring amino acid polymer.
  • amino acid refers to naturally occurring and synthetic amino acids, as well as amino acid analogs and amino acid mimetics that function in a manner similar to the naturally occurring amino acids.
  • Naturally occurring amino acids are those encoded by the genetic code, as well as those amino acids that are later modified, e.g., hydroxyproline, ⁇ - carboxyglutamate, and O-phosphoserine.
  • Amino acid analogs refers to compounds that have the same basic chemical structure as a naturally occurring amino acid, i.e., an a carbon that is bound to a hydrogen, a carboxyl group, an amino group, and an R group, e.g., homoserine, norleucine, methionine sulfoxide, methionine methyl sulfonium. Such analogs have modified R groups (e.g., norleucine) or modified peptide backbones, but retain the same basic chemical structure as a naturally occurring amino acid.
  • Amino acid mimetics refers to chemical compounds that have a structure that is different from the general chemical structure of an amino acid, but that functions in a manner similar to a naturally occurring amino acid.
  • Amino acids may be referred to herein by either their commonly known three letter symbols or by the one-letter symbols recommended by the IUPAC-IUB Biochemical Nomenclature Commission. Nucleotides, likewise, may be referred to by their commonly accepted single-letter codes.
  • “Conservatively modified variants” applies to both amino acid and nucleic acid sequences. With respect to particular nucleic acid sequences, conservatively modified variants refers to those nucleic acids which encode identical or essentially identical amino acid sequences, or where the nucleic acid does not encode an amino acid sequence, to essentially identical sequences. Because of the degeneracy of the genetic code, a large number of functionally identical nucleic acids encode any given protein.
  • the codons GCA, GCC, GCG and GCU all encode the amino acid alanine.
  • the codon can be altered to any of the corresponding codons described without altering the encoded polypeptide.
  • Such nucleic acid variations are “silent QB ⁇ 650053.01002 ⁇ 84442353.1 Page 10 of 153 Atty. Dkt. No.650053.01002 variations,” which are one species of conservatively modified variations. Every nucleic acid sequence herein which encodes a polypeptide also describes every possible silent variation of the nucleic acid.
  • Lyme disease refers to an infection by a Borrelia bacterium, which is also known as Borreliosis. At least nine species of Borrelia are known to cause Lyme disease, including Borrelia afzelii, Borrelia bissettiae, Borrelia burgdorferi, Borrelia garinii, B. hermsii, Borrelia lusitaniae, Borrelia mayonii, Borrelia spielmanii, and Borrelia valaisiana.
  • Additional Borrelia strains include, without limitation, B.burgdorferi-JD1, B.burgdorferi-N40, B.burgdorferi-B31, B.burgdorferi-ZS7, and B.garinii-bavariensis.
  • Lyme disease is commonly caused by the transfer of Borrelia bacteria to mammalian hosts by certain hard ticks of the Ixodidae family. In the U.S., Lyme disease is most commonly the result of the transfer of Borrelia burgdorferi bacteria by the blacklegged tick, also known as a deer tick (Ixodes scapularis or Ixodes pacificus).
  • Ixodes ricinus and Ixodes persulcatus act as Borreliosis vectors. Without early detection and treatment, the bacteria travel through the bloodstream and affect various tissues in the host. The infection may develop into an inflammatory condition that affects multiple systems, starting with skin, joints, and nervous system and moving to organs.
  • subject may be used interchangeably with the terms “individual” and “patient” and includes human and non-human subjects.
  • subjects may be any animal that can be infected with a bacterium, such as Borrelia burgdorferi.
  • the subject is a mammal, such as a human, dog, cat, or livestock, such as cattle, pigs, or sheep.
  • a subject may include a wild, domesticated, or captive population of animals such as deer, elk, bison, etc.
  • a subject may include one or more wild or domesticated bird species, such as chickens, ducks, or turkeys.
  • the methods include detecting the presence of Borrelia burgdorferi in a subject. In some embodiments, the methods include detecting the presence of Borrelia afzelii, Borrelia burgdorferi, Borrelia garinii, or Borrelia mayonii in a subject.
  • the terms “treat” or “treatment” encompass both “preventative” and “curative” treatment. “Preventative” treatment is meant to indicate a postponement of development of a disease, a symptom of a disease, or medical condition, suppressing symptoms that may appear, or reducing the risk of developing or recurrence of a disease or symptom.
  • Treatment includes reducing the severity of or suppressing the worsening of an existing disease, symptom, or condition.
  • treatment includes ameliorating or preventing the worsening of existing disease symptoms, preventing additional symptoms from occurring, ameliorating or preventing the underlying systemic causes of symptoms, inhibiting the disorder or disease, e.g., arresting the development of the disorder or disease, relieving the disorder or disease, causing regression of the disorder or disease, relieving a condition caused by the disease or disorder, or stopping the symptoms of the disease or disorder.
  • antibodies are gamma globulin proteins that can be found in blood or other bodily fluids of vertebrates and are used by the immune system to identify foreign materials, such as bacteria, viruses, and toxins.
  • Antibodies bind, by non-covalent interactions, with high affinity to other molecules or structures known as antigens. This binding is specific in the sense that an antibody molecule will only bind to a specific structure with high affinity.
  • the unique part of the antigen recognized by an antibody molecule is called an epitope, or antigenic determinant.
  • the part of the antibody molecule binding to the epitope is sometimes called paratope and resides in the so-called variable domain, or variable region (Fv) of the antibody.
  • an epitope comprises, or consists of SEQ ID NO: 2, or a polypeptide at least 90% identical to SEQ ID NO: 2, or an epitope provided in Tables 3 and 4.
  • an epitope comprises or consists of a polypeptide provided in Table 5, or a polypeptide at least 90% identical to one or more of the polypeptides provided in Table 5.
  • the solid support may formed from glass, ceramic, polymers (e.g., plastic, latex, polystyrene, polyacrylamide, polyvinylchloride, polypropylene, polyethylene, polylactic acid), cellulose (e.g., paper).
  • a polypeptide is immobilized on a solid support.
  • the polypeptide is SEQ ID NO: 1.
  • the polypeptide is SEQ ID NO: 2.
  • the immobilized polypeptide is bound to the solid support, either reversibly or irreversibly, to form a platform for an antibody binding reaction.
  • the term "bead” refers to a microbead or relatively small bead having a diameter less than about 500 ⁇ m, typically with a diameter of less than about 100 ⁇ m, 50 ⁇ m, or 10 ⁇ m. While a bead may vary in shape, typically a bead is substantially spherical, QB ⁇ 650053.01002 ⁇ 84442353.1 Page 15 of 153 Atty. Dkt. No.650053.01002 e.g. a microsphere.
  • Non-limiting examples of surfactants include polysorbate 20, polysorbate 80, Triton X-100, and pluronic F68.
  • the term "cut-off value" refers to a dividing point for a quantitative result of a diagnostic test or screen to be indicative of a positive or negative result based on measurements of the level of a molecular marker(s). This disclosure provides methods of detecting Lyme disease and/or Borrelia infection in a subject comprising the step of comparing a measured level of molecular marker to a predetermined value.
  • comparing a measured value of an antibody against an epitope e.g., SEQ ID NO: 1 and/or SEQ ID NO: 2 to a predetermined cut-off value of the antibody may be used to decide whether the method indicates a positive or negative result of Lyme disease and/or Borrelia infection in the subject.
  • a "cut-off value” refers to a value above which a measured epitope antibody level in a sample is indicative of the positive presence of a Borrelia infection and/or Lyme disease in the subject from which the sample was derived.
  • SST standard two-tier test
  • the first required test is the Enzyme Immunoassay (EIA) or Immunofluorescence Assay (IFA). If this test yields negative results, the provider should consider an alternative diagnosis; or in cases where the patient has had symptoms for less than or equal to 30 days, the provider may treat the patient and follow up with a convalescent serum. If the first test yields positive or equivocal results, two options are available: 1) If the patient has had symptoms for less than or equal to 30 days, an IgM Western Blot is performed; 2) if the patient has had symptoms for more than 30 days, the IgG Western Blot is performed. The IgM should not be used if the patient has been ill for more than 30 days.
  • EIA Enzyme Immunoassay
  • IFA Immunofluorescence Assay
  • the STT has several well-known shortcomings. Two of the most notable include low sensitivity and specificity in early infection, and the inability to determine infection status after antibody therapy. Diagnostic compositions and methods disclosed herein e.g., that incorporate Peptide 1 (SEQ ID NO: 1) and/or Peptide 2 (SEQ ID NO: 2), such as ELISpot assays, MCH tetramer assays, and proliferation assays, overcome these limitations.
  • compositions comprising an epitope of the BB0680/MCP4 peptide, wherein the epitope comprises or consists of SEQ ID NO: 1.
  • the composition comprises one or more additional antigens, such as outer surface protein A, and outer surface protein C (OspA and OspC, respectively).
  • the additional antigen is a naturally occurring antigen; in some embodiments, the antigen is recombinant.
  • OspA sequence is shown below (NCBI: Ref. Seq: WP_075552755.1).
  • the compositions disclosed herein can include an epitope of the BB0838/LptD peptide and an epitope of the BB0680/MCP4 peptide.
  • the compositions disclosed herein can include an epitope of the BB0838/LptD peptide, wherein the epitope comprises or consists of SEQ ID NO: 2, and an epitope of the BB0680/MCP4 peptide, wherein the epitope comprises or consists of SEQ ID NO: 1.
  • the compositions may also include one or more additional antigens, such as outer surface protein A, and outer surface protein C (OspA and OspC, respectively).
  • the additional antigen is a naturally occurring antigen; in some embodiments, the antigen is recombinant.
  • the compositions disclosed herein can include an epitope of a BB0559/Crr peptide, e.g., Peptide 3, SEQ ID NO: 3, and/or an epitope of oppAIV, e.g., Peptide 4, SEQ ID NO: 4.
  • the compositions disclosed herein comprise an adjuvant.
  • compositions disclosed herein are provided in lyophilized form.
  • Methods for the treating and diagnosis/detection of infection with Borrelia sp.
  • the methods comprise contacting an antibody-containing sample from a subject with a polypeptide of SEQ ID NO: 1 and/or with a polypeptide of SEQ ID NO: 2.
  • the method includes allowing the binding of antibodies in the sample to the polypeptide; contacting the bound antibodies with a detectable binding agent; detecting the detectable binding agent; measuring the level of antibodies bound to the polypeptide; and comparing the measured levels of bound antibody to a value, such as a predetermined cutoff value or a value calculated based on the level of a maker in the sample measured in parallel or a control sample (e.g. a sample from a Borrelia sp. exposure na ⁇ ve subject).
  • the level of antibodies bound to the polypeptide can then be compared to a value to inform a diagnosis or treatment decision, such as, a diagnosis of Lyme disease or a Borrelia infection.
  • the level of antibodies bound to the polypeptide can be used to monitor the progression of a treatment for Lyme disease or a Borrelia infection.
  • the level of antibodies bound to the polypeptide, or after a series of measurements over a duration of weeks or months, can be used to estimate when the subject was exposed to Borrelia sp. QB ⁇ 650053.01002 ⁇ 84442353.1 Page 19 of 153 Atty. Dkt. No.650053.01002 [00100]
  • the measured level is greater than a predetermined antibody value for the polypeptide, and the subject is diagnosed as positive for Lyme disease.
  • the measured level is less than a predetermined antibody value for the polypeptide, the subject is diagnosed as negative for Lyme disease.
  • the method further comprises treating the subject for Lyme disease.
  • predetermined value and “control value” are used interchangeably.
  • the predetermined value or control value is a cutoff value based on a na ⁇ ve control antibody level, either measured independently or measured using the same sample.
  • a suitable cutoff value can be determined using routine techniques known to the skilled worker.
  • the cutoff value is calculated from a measured na ⁇ ve control level adjusted by a standard deviation or parameter based on the standard deviation.
  • a cutoff value can be a mean na ⁇ ve control value + 2 standard deviations.
  • a subject is tested multiple times, and the control value (or predetermined value) comprises the test value of a previous sample from the same subject (e.g., to determine whether a subject is responding favorably to a treatment).
  • the control value (the predetermined value) is greater than the test value if the subject is responsive to treatment, or the control value (the predetermined value) is equal to or less than the test value if the subject is non-responsive to treatment. While it is understood that in some situations a control value is not a predetermined value per-se, the terms "predetermined value” and "control value" are intended to encompass such situations.
  • control sample e.g., a known na ⁇ ve control sample, or a known positive sample
  • control value is not necessarily “predetermined.”
  • the control sample will have a value indicative of a predetermined infection status (e.g., na ⁇ ve or positive).
  • predetermined infection status e.g., na ⁇ ve or positive.
  • multiple different polypeptides are employed in testing.
  • the one or more polypeptides can be configured on a solid support.
  • the detecting comprises detecting the bound antibodies or antibody-epitope complexes using a detectable binding agent.
  • the sample comprises or consists of a blood sample (e.g. whole blood), plasma sample, and/or serum sample.
  • the detecting step comprises immobilizing antibody-epitope complexes to a solid support.
  • the solid support comprises or consists of a plate, bead, flow cell, flow chamber, microfluidic chamber, and/or microchip comprising a plurality of microfluidic chambers.
  • the detecting step comprises flowing the reagent- composition through a flow-chamber.
  • the detection comprises a microfluidic chamber, microchip comprising a plurality of microfluidic chambers, or lateral flow assay.
  • the subject is human and the detectable binding agent comprises an anti-human IgG comprising a detectable label or conjugated to a detectable agent.
  • the detectable agent comprises an enzyme, such as a horseradish peroxidase.
  • a T cell assay is provided.
  • the T cell assay is configured as an ELISpot assay, an MHC tetramer assay, or a cell proliferation assay.
  • the term "ELISpot” assay refers to the "enzyme-linked immune absorbent spot” assay. The ELISpot assay is often used to quantitatively measure the frequency of cytokine secretion for a single cell.
  • the ELISpot Assay is also a form of immunostaining since it is classified as a technique that uses antibodies to detect a protein analyte, with the word analyte referring to any biological or chemical substance being identified or measured.
  • an "MHC tetramer assay” refers to an assay method that uses tetrameric proteins to detect and quantify T cells that are specific for a given antigen within a blood sample.
  • the tetramers used in the assay are made up of four major histocompatibility complex (MHC) molecules, which are found on the surface of most cells in the body.
  • MHC major histocompatibility complex
  • MHC molecules present peptides to T-cells as a way to communicate the presence of viruses, bacteria, cancerous mutations, or other antigens in a cell. If a T-cell's receptor matches the peptide being presented by an MHC molecule, an immune response is triggered.
  • MHC tetramers that are bioengineered to present a specific peptide can be used to find T-cells with receptors that match that peptide.
  • the tetramers are labeled with a detectable label, such as a fluorophore, allowing tetramer-bound T-cells to be analyzed with flow cytometry.
  • the subject has not previously been treated for Lyme disease.
  • the subject is asymptomatic for Lyme disease, but may have come in contact with a blacklegged tick selected from Ixodes scapularis, Ixodes pacificus, Ixodes ricinus, and Ixodes persulcatus.
  • the possible contact between the subject and the tick was within 3 months, within 2 months, within 1 month, within 2 weeks, within 1 week, within 3 days, or within 1 day.
  • the subject is asymptomatic for Lyme disease and has previously been treated for Lyme disease.
  • kits include a polypeptide comprising SEQ ID NO: 2, or a polypeptide at least 90% identical to SEQ ID NO: 2, configured on a solid support or in a reagent composition.
  • the kits include a polypeptide comprising SEQ ID NO: 1 and/or SEQ ID NO: 2.
  • the kits include a polypeptide at least 90% identical to SEQ ID NO: 1 and/or a polypeptide at least 90% identical to SEQ ID NO: 2.
  • the additional reagent or device is a flow chamber, bead composition, ELISpot assay reagent and/or platform, ELISA reagent, anti-human IgG antibody, and/or a detectable reagent such as a HRP substrate.
  • a kit includes reagents and components for an MHC tetramer assay.
  • the kit comprises instructions for using the components of the kit in the performance of a method described herein.
  • the kits comprise instructions, and are configured for in-home use.
  • kits and components there may be used to evaluate T and/or B cell responses to the epitope(s), which provide novel diagnostic biomarkers for active infection of B. burgdorferi or post-treatment Lyme disease syndrome (PTLDS).
  • the kits are configured for ELISpot assays, MCH tetramer assays, and/or T cell proliferation assays, and include SEQ ID NO: 1 and/or SEQ ID NO: 2.
  • Table 1 below shows the predicted mouse MHCII binding of peptides derived from BB0680 determined using NetMHCIIpan (V 4.0).
  • the prediction outcome for each molecule includes the following columns in the table: Pos Residue number (starting from 0); MHC MHC molecule name; Peptide Amino acid sequence; Of Starting position offset of the optimal binding core (starting from 0); Core Binding core register; Core_Rel Reliability of the binding core, expressed as the fraction of networks in the ensemble selecting the optimal core; Score_EL Eluted ligand prediction score; %Rank_EL Percentile rank of eluted ligand prediction score; BindLevel (SB: strong binder, WB: weak binder). The peptide will be identified as a strong binder if the % Rank is below the 1% threshold for the strong binders.
  • the prediction outcome for each molecule includes the following columns in the table: Pos Residue number (starting from 0); MHC MHC molecule name; Peptide Amino acid sequence; Of Starting position offset of the optimal binding core (starting from 0); Core Binding core register; Core_Rel Reliability of the binding core, expressed as the fraction of networks in the ensemble selecting the optimal core; Score_EL Eluted ligand prediction score; %Rank_EL Percentile rank of eluted ligand prediction score; BindLevel (SB: strong binder, WB: weak binder). The peptide will be identified as a strong binder if the % Rank is below the 1% threshold for the strong binders.
  • the peptide will be identified as a weak binder if the % Rank is above the 1% threshold of the strong binders but below the 5% threshold for the weak binders.
  • Table 3 Predicted human CD4+ T cell epitopes from B. burgdorferi LptD Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO W B W W B W W W W W B W W W W W W W W W W W W B QB ⁇ 650053.01002 ⁇ 84442353.1 Page 54 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W W W W W W W W B W W W W W W W W W W B B W W W W W W B W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 57 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W W W W W W W W W W W W B B W W W W W W W W B B W W B B QB ⁇ 650053.01002 ⁇ 84442353.1 Page 58 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W B B W W W B B W W B W W B W B W B W B W W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 59 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: B W W W W W W B W B W W W W W W W W W W W W W W W W W B W B QB ⁇ 650053.01002 ⁇ 84442353.1 Page 61 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W W W W W W W W B W W W W W W B B W W W W W W B B QB ⁇ 650053.01002 ⁇ 84442353.1 Page 62 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: B W W W B B W W W W W W W W W B B W B B W W W B B B W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 63 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W B W W W W W B B B W W W W W W W B B QB ⁇ 650053.01002 ⁇ 84442353.1 Page 64 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W W W W W W W W W W W W W B W W W W W W W W B W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 66 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W B W W W W W B W B W W W W W W W B W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 67 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W B W W W W W W W W W W W W W W W W W W W W W W W B W B W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 70 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W W W W W W B W W W W W W W W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 72 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W B W W W B B B W W B W B W W W W W W B W B B QB ⁇ 650053.01002 ⁇ 84442353.1 Page 75 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: B W B B W W B W W W W B W B B B W W W W W W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 76 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W B B B W W W W W W W W W W W B B W W W W W B B W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 77 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W B B W W B B W W W W W W W W W W W B W W W W W W W W B QB ⁇ 650053.01002 ⁇ 84442353.1 Page 78 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: B W W B W W W W W W W W W W W W B W W W W W W W W W W B B W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 79 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W W W W B W W W W W B W W B W B B W W W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 80 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 81 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W B W B W W W W W W W W B W W W W B W W W B W W W B W W B W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 84 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W W W W W W W W W W W W W B W B W B W W B W W W W B QB ⁇ 650053.01002 ⁇ 84442353.1 Page 85 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W B B W W B W W W W B B B W W B W W W W B B QB ⁇ 650053.01002 ⁇ 84442353.1 Page 86 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W B W W W W W W W W W W W W W W W W W W W W W W B W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 87 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W B W W W W W W W W W B W W W W W W W W W W W W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 88 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W B W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 90 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: B W W W W W W W W B W B W W W W W W W W W W W W W W W W W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 95 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W W W W W W W W W W B W W W W W W W W W W W W W W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 105 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W B W W W W W B W W W W W B W B W W W W B W W W B B W W B B QB ⁇ 650053.01002 ⁇ 84442353.1 Page 108 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 113 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W W W W W W W W W W W W W W W W W W W B W W W W W B B QB ⁇ 650053.01002 ⁇ 84442353.1 Page 114 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W B W B W W W W B W W B B W B W B W B W W W W B W W B B W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 115 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W B W W W W W W W W W W W W W W W W W W B W B W W W W B W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 116 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W B B B W W W W W W W W W B B W B W W W W W B W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 117 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W B W W W W W W W W W B W W W W B W B W W W W W B W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 118 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W B W W W W B W W B W W W W W W W W W B W B W W B QB ⁇ 650053.01002 ⁇ 84442353.1 Page 119 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W B W W W W W W B W B W W W W B B W W W W B B W B W B W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 120 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: B B W B W W W W W W W W W W W W W W W W W W W W W W W W W W W B W B W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 121 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 122 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 123 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W W W W W W W W W W W W W W W B W W W W W W W W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 124 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W B W W W B B W W B W B W W B W B W B B B W W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 125 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 126 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W B W B W W W W W W W W W B B B W B W W B W B W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 127 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W B B B W B W W W W W W W W W W W W W W W B W W W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 128 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W B W W W W W W W B W W W W W W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 129 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W B W W W W W W W W W B W W W W W B B W W W W W W W W B B QB ⁇ 650053.01002 ⁇ 84442353.1 Page 130 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W W W B W W W W W W W W W W W W W W W W W W W W W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 131 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W W B W W W W W W W W W W W W W W W W W W W W W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 132 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 133 of 153 Atty. Dkt.
  • No.650053.01002 Pos MHC SEQ ID Peptide Of SEQ Core Core_R Identity Score_EL %Rank Bind NO: ID el _EL Level NO: W W W W W W W W W W W B W W W W W W W W W W W W W W W W W W W W W W QB ⁇ 650053.01002 ⁇ 84442353.1 Page 134 of 153 Atty. Dkt.
  • the prediction outcome for each molecule includes the following columns in the table: Pos Residue number (starting from 923); MHC molecule name; Peptide Amino acid sequence; Of Starting position offset of the optimal binding core (starting from 0); Core Binding core register; Core_Rel Reliability of the binding core, expressed as the fraction of networks in the ensemble selecting the optimal core; Score_EL QB ⁇ 650053.01002 ⁇ 84442353.1 Page 135 of 153 Atty. Dkt. No.650053.01002 Eluted ligand prediction score; %Rank_EL Percentile rank of eluted ligand prediction score; BindLevel (SB: strong binder, WB: weak binder).
  • the peptide will be identified as a strong binder if the % Rank is below the 1% threshold for the strong binders.
  • the peptide will be identified as a weak binder if the % Rank is above the 1% threshold of the strong binders but below the 5% threshold for the weak binders.
  • Table 4 Predicted human CD4+ T cell epitopes within B. burgdorferi LptD 922-972 Pos MHC SEQ Peptide Of SEQ Core Core_ Identity Score_EL %Rank Bind ID ID ID Rel _EL Level NO: NO: B B B B B B B B B B B B B B B B B B B B B B B B B B B B B B B B B B B B B B QB ⁇ 650053.01002 ⁇ 84442353.1 Page 136 of 153 Atty.
  • Peptide 1 (SEQ ID NO: 1) is a methyl-accepting chemotaxis protein that is highly conserved amongst both sensu lato and sensu stricto strains of Lyme disease spirochetes (FIG. 3, FIG. 5). Peptide 1 is highly immunogenic in mice infected with B. burgdorferi (FIG.
  • Peptide 1 is one of a number of peptides derived from MCP4 predicted to bind to MHC class II alleles in mice and/or humans (FIG. 5). It is anticipated that immunization with Peptide 1 will lead to protective immunity in mice and humans. It is also anticipated that other epitopes derived from MCP4 are immunogenic in mice and humans infected with B. burgdorferi. [00130] Example 2 - Test the therapeutic potential of generating immune responses to Peptide 1 in mice. QB ⁇ 650053.01002 ⁇ 84442353.1 Page 138 of 153 Atty. Dkt.
  • mice will be immunized with one or more of peptide 1 (SEQ ID NO: 1) and/or Peptide 2 (SEQ ID NO: 2) and tested for protection against B. burgdorferi infection. It is anticipated that immunized mice will exhibit fewer or no symptoms of infection compared to unimmunized controls mice. It is further anticipated that mice immunized with a vaccine composition comprising SEQ ID NO: 1 and/or SEQ ID NO: 2, or a polypeptide at least 90% identical to SEQ ID NO: 1 and/or a polypeptide at least 90% identical to SEQ ID NO: 2, will exhibit fewer or no symptoms of infection compared to mice immunized with prior art vaccines.
  • mice immunized with a vaccine composition comprising SEQ ID NO: 1 and/or SEQ ID NO: 2, or a polypeptide at least 90% identical to SEQ ID NO: 1 and/or a polypeptide at least 90% identical to SEQ ID NO: 2, will exhibit an increase in CD4+ T-cell population as compared to unimmunized control mice, or mice immunized with prior art vaccines.
  • Example 3 Determine immune responses to target antigen in human patients during active infection and in patients with post-treatment Lyme disease syndrome (PTLDS).
  • Clause 26 A method for testing a subject for the presence of Borrelia sp. bacteria, the method comprising: contacting a subject sample to the polypeptide of clause 1.
  • Clause 27 The method of clause 26, wherein the polypeptide comprises SEQ ID NO: 2.
  • Clause 28 The method of clause 27, wherein the polypeptide is SEQ ID NO: 2.
  • Clause 29 The method of clause 26, wherein the polypeptide is linked to a solid support.
  • Clause 30 The method of clause 26, wherein the subject sample comprises a blood or serum sample. [00176] Clause 31.
  • Clause 37 The composition of any one of clauses 33-35, wherein the at least one additional polypeptide comprises OspA or OspC.
  • Clause 37 The composition of any one of clauses 33-36, wherein the composition is in lyophilized form.
  • Clause 38 A method of treating Lyme disease, or a method for the preventative treatment of Lyme disease, the method comprising: administering to a subject the polypeptide of clause 32, or the composition of any of clauses 33-37.
  • QB ⁇ 650053.01002 ⁇ 84442353.1 Page 144 of 153 Atty. Dkt. No.650053.01002 [00184] Clause 39.
  • Clause 40 The method of clause 39, wherein the possible contact was within 3 months.
  • Clause 41 The method of clause 39, wherein the possible contact was within 2 months.
  • Clause 42 The method of clause 39, wherein the possible contact was within 1 month.
  • Clause 43 The method of clause 39, wherein the possible contact was within 2 weeks.
  • Clause 44 The methods of clause 39, wherein the possible contact was within 1 week. [00190] Clause 45.

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Abstract

L'invention concerne des compositions et des méthodes de traitement et de traitement préventif de la maladie de Lyme. L'invention concerne également des compositions et des procédés de détection et de diagnostic d'une infection par un Borrelia sp.
EP23861597.5A 2022-09-02 2023-09-01 Utilisation d'épitopes de lymphocytes t immunogènes pour la vaccination et le diagnostic de la maladie de lyme Pending EP4580665A2 (fr)

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AU8151898A (en) * 1997-06-20 1999-01-04 Human Genome Sciences, Inc. Lyme disease vaccines
WO2008031133A2 (fr) * 2006-09-15 2008-03-20 Intercell Ag Antigènes de borrelia
HRP20250954T1 (hr) * 2014-01-09 2025-10-10 Valneva Austria Gmbh Mutantni fragmenti ospa i postupci i upotrebe koji se na njih odnose

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