# US 12,181,476 B2 · Use of amino acid sequences from Mycobacterium tuberculosis or corresponding nucleic acids for diagnosis and prevention of tubercular infection, diagnostic kit and vaccine therefrom > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 12,181,476 B2 - Document ID: US12181476B2 - Status: Granted - Kind: B2 - Application number: 17/576,308 - Filed: 2022-01-14 - Published: 2022-04-28 - Granted: 2024-12-31 - Earliest claimed: 2010-07-23 - Assignees: Cellestis Limited - Applicants: Cellestis Limited - Inventors: Francesca Mariani; Massimo Amicosante; Vittorio Colizzi; Cesare Saltini - Web page: https://patentgrep.com/patent/US12181476B2 ## CPC - G01N 33/56972: {White blood cells} - A61K 38/10: Peptides having 12 to 20 amino acids {(A61K38/043 - A61K38/046 take precedence)} - A61K 38/164: {from bacteria} - A61P 31/04: Antibacterial agents - A61P 31/06: for tuberculosis - C07K 7/08: having 12 to 20 amino acids (gastrins C07K14/595; somatostatins C07K14/655; melanotropins C07K14/68) - C07K 14/35: from Mycobacteriaceae (F) - G01N 33/5695: {Mycobacteria} - G01N 33/6866: {Interferon} - G01N 2333/35: from Mycobacteriaceae (F) - G01N 2333/57: IFN-gamma - G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q) - A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D) - A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS - C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00) ## Abstract The present invention refers to the use of gene sequences or portions thereof characterized in that the same belong to the classes of in vitro and ex vivo induced, repressed or conserved genes in Mycobacterium tuberculosis currently infected human macrophages and to corresponding peptides or consensus peptides or proteins for the preparation of specific bio-markers for the diagnosis and prevention of active or latent disease. ## Claims ### Claim 1 A method for detecting a Mycobacterium species infection in a test subject, comprising: (a) incubating in vitro a blood sample or a lavage sample that comprises lymphocytes from the test subject with at least one isolated protein or peptide that comprises SEQ ID NO: 43, 44, 45, 46, or 47, wherein said step of incubating is for a time and under conditions sufficient to stimulate the lymphocytes to produce an effector molecule; and (b) detecting a test level of the effector molecule, wherein detection of a test level that is higher than a control level that is detected following incubation of control lymphocytes from a healthy control subject, indicates that the test subject is infected with, or has been previously exposed to, the Mycobacterium species. ### Claim 2 (depends on 1) The method of claim 1 wherein the Mycobacterium species is selected from M. tuberculosis, M. bovis, M. bovis BCG, M. africanum, M. canetti, M. caprae, M. microti, M. pinnipedii, M. avium, M. avium paratuberculosis, M. avium silvaticum, M. avium “hominissuis”, M. colombiense, M. asiaticum, M. gordonae, M. gastri, M. kansasii, M. hiberniae, M. nonchromogenicum, M. terrae, M. triviale, M. ulcerans, M. pseudoshottsii, M. shottsii, M. triplex, M. genavense, M. florentinum, M. lentiflavum, M. palustre, M. kubicae, M. parascrofulaceum, M. heidelbergense, M. interjectum, M. simiae, M. branderi, M. cookii, M. celatum, M. bohemicum, M. haemophilus, M. malmoense, M. szulgai, M. leprae, M. lepraemurium, M. lepromatosis, M. africanum, M. botniense, M. chimaera, M. conspicuum, M. doricum, M. farcinogenes, M. heckeshornense, M. intracellulare, M. lacus, M. marinum, M. monacense, M. montefiorense, M. murale, M. nebraskense, M. saskatchewanense, M. scrofulaceum, M. shimoidei, M. tusciae, M. xenopi, M. intermedium, M. abscessus, M. chelonae, M. bolletii, M. fortuitum, M. fortuitum subsp. acetamidolyticum, M. boenickei, M. peregrinum, M. porcinum, M. senegalense, M. septicum, M. neworleansense, M. houstonense, M. mucogenicum, M. mageritense, M. brisbanense, M. cosmeticum, M. parafortuitum, M. austroafricanum, M. diernhoferi, M. hodleri, M. neoaurum, M. frederiksbergense, M. aurum, M. vaccae, M. chitae, M. fallax, M. confluentis, M. flavescens, M. madagascariense, M. phlei, M. smegmatis, M. goodii, M. wolinskyi, M. thermoresistibile, M. gadium, M. komossense, M. obuense, M. sphagni, M. agri, M. aichiense, M. alvei, M. arupense, M. brumae, M. canariasense, M. chubuense, M. conceptionense, M. duvalii, M. elephantis, M. gilvum, M. hassiacum, M. holsaticum, M. immunogenum, M. massiliense, M. moriokaense, M. psychrotolerans, M. pyrenivorans, M. vanbaalenii, M. pulveris, M. arosiense, M. aubagnense, M. caprae, M. chlorophenolicum, M. fluoroanthenivorans, M. kumamotonense, M. novocastrense, M. parmense, M. phocaicum, M. poriferae, M. rhodesiae, M. seoulense and M. tokaiense. ### Claim 3 (depends on 1) The method of claim 1 wherein the Mycobacterium species is Mycobacterium tuberculosis. ### Claim 4 (depends on 1) The method of claim 1 wherein at least one of: (a) heparin is present in the step of incubating, (b) heparin and an added carbohydrate are present in the step of incubating, and (c) at least one Mycobacterium protein, or a peptide fragment thereof or a chemical analog thereof, is present in the step of incubating, said Mycobacterium protein being selected from ESAT6, CFP10, TB7.7 and PPD. ### Claim 5 (depends on 1) The method of claim 1 wherein the effector molecule is selected from interferon-γ, TNF-α, and an interleukin. ### Claim 6 (depends on 1) The method of claim 1 wherein the effector molecule is interferon-γ. ### Claim 7 (depends on 1) The method of claim 1, wherein the subject is a human subject. ### Claim 8 (depends on 1) The method of claim 1, wherein the subject is a non-human animal. ### Claim 9 (depends on 1) The method of claim 1, wherein in the step of incubating there is present one or more Mycobacterium proteins, wherein the one or more Mycobacterium proteins are selected from the group consisting of ESAT6, CFP10, TB7.7 and PPD, and wherein the detected test level of the effector molecule is greater than a detectable effector molecule level that can be produced by said lymphocytes incubated only with said one or more Mycobacterium proteins selected from the group consisting of ESAT6, CFP10, TB7.7 and PPD. ### Claim 10 (depends on 1) The method of claim 1, wherein in the step of incubating there is further present one or more isolated proteins or peptides comprising at least one of the amino acid sequences selected from the group consisting of SEQ ID NOS: 1-42 and 48-74. ### Claim 11 (depends on 1) The method of claim 1, wherein in the step of incubating there is further present an isolated protein or peptide comprising SEQ ID NO: 48, 49, or 50. ### Claim 12 (depends on 1) The method of claim 1, wherein in the step of incubating there is further present an isolated protein or peptide comprising SEQ ID NO: 64, 65, or 66. ### Claim 13 (depends on 1) The method of claim 1, wherein in the step of incubating there is further present at least 5 isolated proteins or peptides comprising SEQ ID NOS: 43-47, respectively. ### Claim 14 (depends on 1) The method of claim 1, wherein in the step of incubating there is further present at least 3 isolated proteins or peptides comprising SEQ ID NOS: 48-50, respectively. ### Claim 15 (depends on 1) The method of claim 1, wherein in the step of incubating there is further present at least 3 isolated proteins or peptides comprising SEQ ID NOS: 64-66, respectively.