Methods for treating progressive chronic interstitial lung disease

Abstract

The present invention relates to anti-transglutaminase type 2 antibodies that inhibit the transaminase activity of the enzyme for use in the treatment of progressive chronic interstitial lung diseases such as idiopathic pulmonary fibrosis (IPF).

Description

[Technical field]

[0001] Field of the invention: The present invention relates to anti-transglutaminase type 2 antibodies, which inhibit the transaminase activity of the enzyme, for use in the treatment of progressive chronic interstitial lung disease, such as idiopathic pulmonary fibrosis (IPF). [Background technology]

[0002] Background of the invention: Tissue transglutaminase or transglutaminase type 2 (TG2) is an enzyme that forms crosslinks between proteins via epsilon (gamma-glutamyl) lysine dipeptide bonds. High expression of TG2 leads to abnormal protein crosslinks and is associated with several pathologies, such as various types of tissue scarring, fibrosis, formation of neurofibrillary tangles in some brain diseases, and resistance to chemotherapy in some cancers. Various TG2 inhibitors, such as small molecules, silencing RNAs, or antibodies (e.g., Non-Patent Document 1, Non-Patent Document 2, Patent Document 1, Patent Document 2, or Patent Document 3), have been disclosed for the possible treatment of TG2-mediated disorders.

[0003] Progressive chronic interstitial lung diseases include idiopathic pulmonary fibrosis (IPF), which is characterized by the deposition of scar tissue in the pulmonary interstitium and thickening of the alveolar membranes, leading to a progressive decline in lung function and ultimately death. The overall prognosis after a diagnosis of IPF is generally poor, as the disease progresses steadily and ultimately to death.

[0004] Increases in TG2 and LOXL2 are associated with progressive chronic interstitial lung diseases such as idiopathic pulmonary fibrosis (Non-Patent Document 3, Non-Patent Document 4, Non-Patent Document 5). Overexpression or overactivation of TG2 in IPF-derived fibroblasts increases epsilon (gamma-glutamyl) lysine crosslinks between proteins of the extracellular matrix (ECM). Manipulation of TG2 and LOXL2 by the use of small inhibitors or by the use of null mice during fibrosis induction reduces lung inflammation and fibrosis in bleomycin-treated animals (see Non-Patent Document 3, Non-Patent Document 6, Non-Patent Document 7, Non-Patent Document 8, Non-Patent Document 5). For example, in fibroblast adhesion and proliferation assays, it has been shown that the simultaneous addition of TG2 and cystamine (a pan-TG inhibitor) inhibits the increase in adhesion associated with TG2 (see non-patent document 5). Although the primary role of TG2 cross-linking activity in fibrotic remodeling appears to be to slow extracellular matrix (ECM) turnover by incorporating protease-resistant intramolecular cross-links into ECM proteins and to locally activate TGFβ1, there is some emerging evidence that in lung cells, the nonenzymatic role of TG2 in cell adhesion may be modulated by inhibiting its transamidation activity, as exemplified by the use of cystamine to reduce pulmonary fibroblast adhesion in vitro.

[0005] Two approved therapeutic agents, pirfenidone (which appears to function in part through modulation of TGFβ1-related pathways, including collagen synthesis) and nintedanib (which targets multiple tyrosine kinases), are effective in slowing disease progression in some patients, and additional treatment options are urgently needed (Non-Patent Document 9). However, there remains a need to identify additional effective therapies for use in the treatment and prevention of progressive chronic interstitial lung diseases such as IPF, as well as for use in treating subjects with pulmonary fibrosis associated with COVID infection or for use in preventing the development of pulmonary fibrosis in patients with COVID infection. [Prior art documents] [Patent documents]

[0006] [Patent Document 1] International Publication No. 2006 / 100679 [Patent Document 2] International Publication No. 2012 / 146901 [Patent Document 3] International Publication No. 2013 / 175229 [Non-patent literature]

[0007] [Non-Patent Document 1] Siegel et al., 2007 [Non-Patent Document 2] Wang et al., 2020 [Non-Patent Document 3] Olsen et al., 2011 [Non-Patent Document 4] Olsen et al., 2020 [Non-Patent Document 5] Philp et al., 2018 [Non-Patent Document 6] Olsen et al., 2014 [Non-Patent Document 7] Raghav 2017 [Non-Patent Document 8] Vaidya et al., 2017 [Non-Patent Document 9] Margaritopoulos et al., 2016 Summary of the Invention

[0008] Summary of the invention: The object of the present invention is to provide a specific anti-transglutaminase 2 (anti-TG2) antibody for use in the treatment of progressive chronic interstitial lung diseases such as idiopathic pulmonary fibrosis (IPF) or for use in preventing the onset of progressive chronic interstitial lung diseases such as idiopathic pulmonary fibrosis (IPF). Alternatively, the present invention provides a specific anti-transglutaminase 2 (anti-TG2) antibody for use in the treatment of a subject having lung fibrosis associated with COVID infection or for use in preventing the onset of lung fibrosis in a patient suffering from COVID infection.

[0009] In a second aspect, the present invention provides a method for treating or preventing the onset of progressive chronic interstitial lung disease, comprising administering a therapeutically effective amount of an anti-TG2 antibody. Alternatively, the present invention provides a method for treating a subject having pulmonary fibrosis associated with COVID infection or for preventing the development of pulmonary fibrosis in a patient suffering from COVID infection, comprising administering a therapeutically effective amount of an anti-TG2 antibody.

[0010] In a third aspect, the present invention relates to the use of an anti-TG2 antibody for the manufacture of a medicament for the treatment of a progressive chronic interstitial lung disease or for the prevention of the onset of a progressive chronic interstitial lung disease, such as idiopathic pulmonary fibrosis (IPF). Alternatively, the present invention relates to the use of an anti-TG2 antibody for the manufacture of a medicament in the treatment of a subject with pulmonary fibrosis associated with COVID infection or in the prevention of the development of pulmonary fibrosis in patients suffering from COVID infection.

[0011] Definition: The entire specification is intended to be linked as a unified disclosure, and it is to be understood that all combinations of features described herein are contemplated, even if the combinations are not described together in the same sentence or paragraph or section of this specification. With respect to aspects of the invention that may be described or claimed using "a" or "an," these terms should be understood to mean "one or more," unless the context clearly requires a more limited meaning. The term "or" should be understood to encompass alternative or together items, unless the context clearly requires otherwise. When aspects of the invention are described as "comprising" a certain feature, the embodiment is also considered to "consist" of or "consisting essentially of" that feature.

[0012] - The term "tissue transglutaminase", "transglutaminase type 2" or "TG2" refers to an enzyme that forms crosslinks between proteins via epsilon (gamma-glutamyl) lysine dipeptide bonds. TG2 typically refers to the protein having the amino acid sequence set forth in UniProt entry P21980 (SEQ ID NO:41), i.e., human TG2. The term "TG2" may also refer to a protein that is (a) a derivative having one or more amino acid substitutions, modifications, deletions or insertions relative to the amino acid sequence of SEQ ID NO:41 that retains the activity of TG2, or (b) a variant thereof, e.g., the variant typically retains at least about 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94% or 95% identity to SEQ ID NO:41 (or even about 96%, 97%, 98% or 99% identity to SEQ ID NO:41). Protein TG2 is encoded by the gene Tgm2.

[0013] - The term "anti-TG2 antibody" as used herein intends an antibody molecule that binds to TG2 and inhibits its transamidase activity, preventing cross-linking. Examples of such antibodies are described in WO 2013 / 175229. Without being limited thereto, an anti-TG2 antibody that can be used according to the present invention includes, for example, a light chain variable region defined in SEQ ID NO. 24 and a heavy chain variable region defined in SEQ ID NO. 37.

[0014] - The term "antibody" as used herein includes, but is not limited to, monoclonal antibodies, polyclonal antibodies, and recombinant antibodies produced by recombinant techniques as known in the art. "Antibody" includes antibodies of any species, such as human antibodies of any isotype, such as IgG1, IgG2a, IgG2b, IgG3, IgG4, IgE, IgD, and antibodies produced as dimers of these basic structures, including pentamers such as IgGA1, IgGA2, or IgM, and modified variants thereof; non-human primate antibodies, such as antibodies from chimpanzees, baboons, rhesus monkeys, or cynomolgus monkeys; rodent antibodies, such as antibodies from mice or rats; rabbit, goat, or horse antibodies; camelid antibodies (e.g., antibodies from camels or llamas, such as Nanobodies®) and derivatives thereof; antibodies of avian species, such as chicken antibodies; or antibodies of fish species, such as shark antibodies. The term "antibody" also refers to a "chimeric" antibody in which a first portion of at least one heavy and / or light chain antibody sequence is from a first species and a second portion of said heavy and / or light chain antibody sequence is from a second species. Chimeric antibodies of interest herein include "primatized" antibodies that contain variable domain antigen-binding sequences from a non-human primate (e.g., an Old World monkey such as a baboon, rhesus monkey, or cynomolgus monkey) and human constant region sequences. A "humanized" antibody is a chimeric antibody that contains sequences from a non-human antibody. In most cases, a humanized antibody is a human antibody (recipient antibody) in which residues from the recipient's hypervariable regions are replaced by residues from the hypervariable regions [or complementarity determining regions (CDRs)] of a non-human species (donor antibody) such as mouse, rat, rabbit, chicken, non-human primate, etc., to provide the desired specificity, affinity, and activity. In most cases, residues of the human (recipient) antibody outside the CDRs, i.e., in the framework regions (FR), are further replaced by corresponding non-human residues. Furthermore, humanized antibodies may comprise residues that are not found in the recipient antibody or in the donor antibody.These modifications are made to further refine the properties of the antibody. Humanization reduces the immunogenicity of non-human antibodies in humans, facilitating the application of antibodies in the treatment of human diseases. Humanized antibodies and techniques for their production are well known in the art. The term "antibody" also refers to human antibodies, which can be generated as an alternative to humanization. For example, it is possible to generate transgenic animals (e.g. mice) that are capable of producing a full repertoire of human antibodies upon immunization in the absence of endogenous mouse antibody production. Other methods for obtaining human antibodies / antibody fragments in vitro are based on display technologies, such as phage display and ribosome display technology, using recombinant DNA libraries that are at least partially artificially generated or generated from a donor immunoglobulin variable (V) domain gene repertoire. Phage and ribosome display technologies for generating human antibodies are well known in the art. Human antibodies can also be generated from isolated human B cells ex vivo immunized with an antigen of interest, which can then be fused to generate hybridomas and screened for optimal human antibodies. The term "antibody" refers to both glycosylated and aglycosylated antibodies. Furthermore, as used herein, the term "antibody" refers not only to full-length antibodies, but also to antibody fragments, more particularly antigen-binding fragments thereof. Antibody fragments contain at least one heavy or light chain immunoglobulin domain and bind to one or more antigens, as known in the art. Examples of antibody fragments according to the present invention include Fab, modified Fab, Fab', modified Fab', F(ab')2, Fv, Fab-Fv, Fab-dsFv, Fab-Fv-Fv, scFv and Bis-scFv fragments. The fragments may also be single domain antibodies (dAbs) such as diabodies, tribodies, triabodies, tetrabodies, minibodies, sdAbs, VL, VH, VHH or camelid antibodies (e.g. from camel or llama such as Nanobodies®) and VNAR fragments.An antigen-binding fragment according to the invention may also comprise a Fab linked to one or two scFvs or dsscFvs, each scFv or dsscFv binding the same or different targets (e.g. one scFv or dsscFv that binds a therapeutic target and one scFv or dsscFv that increases half-life, e.g. by binding albumin). Examples of such antibody fragments include FabdsscFv (also called BYbe®) or Fab-(dsscFv)2 (also called TrYbe®, see e.g. WO 2015 / 197772). The antibody molecules as defined above, including antigen-binding fragments thereof, are known in the art.

[0015] - The term "epitope" refers to the region of an antigen to which an antibody binds. Epitopes can be structurally or functionally defined. Functional epitopes are generally a subset of structural epitopes and contain residues that directly contribute to the affinity of the interaction. Epitopes can also be conformational, i.e., composed of non-linear amino acids. In certain embodiments, epitopes can include determinants that are chemically active surface groups of molecules such as amino acids, sugar side chains, phosphoryl or sulfonyl groups, and, in certain embodiments, can have specific three-dimensional structural characteristics and / or specific charge characteristics.

[0016] - The term "treating" or "treatment" of a disease state includes (i) inhibiting the disease state, i.e., preventing the onset of the disease state or its clinical symptoms, or (ii) alleviating the disease state, i.e., causing the temporary or permanent regression of the disease state or its clinical symptoms.

[0017] - The term "preventing" or "prevention" of a disease state includes preventing the development of clinical symptoms of the disease state in a subject who may be exposed to or susceptible to the disease state, but who has not yet experienced or exhibited symptoms of the disease state.

[0018] Detailed description of the invention: The present invention is based on the inventor's discovery that total TG2 mRNA and total TG2 protein are increased in lung tissues of IPF patients. TG2 is associated with elevated collagen levels that correlate with reduced lung function. It was also discovered that mice lacking TG2 (called knockout or KO mice) are protected from the development of interstitial pulmonary fibrosis after provocation with bleomycin (a drug that causes pulmonary fibrosis and is widely used in animal models of pulmonary fibrosis) and subsequently have reduced lung function. The inventors were then surprisingly able to demonstrate that anti-TG2 antibodies not only attenuate ECM deposition by primary human IPF cells in in vitro assays, but also that prophylactic administration of anti-TG2 antibodies (which inhibit the extracellular protein cross-linking activity of TG2) significantly attenuates pulmonary fibrosis in vivo (in a rabbit pulmonary silicosis model) and prevents further progression of fibrosis when administered starting 28 days after the onset of fibrosis. Furthermore, the inventors surprisingly showed that TG2 was upregulated in postmortem lung samples from patients who died from COVID-19 infection (caused by the SARS-Cov2 virus) with evidence of extensive matrix deposition.

[0019] The main object of the present invention is an anti-transglutaminase 2 (anti-TG2) antibody for use in the treatment of progressive chronic interstitial lung disease or for preventing the onset of progressive chronic interstitial lung disease. For example, an anti-TG2 antibody for use according to the present invention may comprise the following sequence: (i) KASQDINSYLT (LCDR1; SEQ ID NO. 1); LVNRLVD (LCDR2; SEQ ID NO. 2); LQYDDFPYT (LCDR3; SEQ ID NO. 3); THAMS (HCDR1; SEQ ID NO. 4); TISSGGRSTYYPDSVKG (HCDR2; SEQ ID NO. 5); and LISTY (HCDR3; SEQ ID NO. 6); or (ii) KASQDINSYLT (LCDR1; SEQ ID NO. 1); LTNRLMD (LCDR2; SEQ ID NO. 7); LQYVDFPYT (LCDR3; SEQ ID NO. 8); SSAMS (HCDR1; SEQ ID NO. 9); TISSGGRSTYYPDSVKG (HCDR2; SEQ ID NO. 5); and LISPY (HCDR3; SEQ ID NO. 10); or (iii) KASQDINSYLT (LCDR1; SEQ ID NO. 1); RTNRLFD (LCDR2; SEQ ID NO. 11); LQYDDFPYT (LCDR3; SEQ ID NO. 3); SSAMS (HCDR1); TISVGGGKTYYPDSVKG (HCDR2; SEQ ID NO. 9); and LISLY (HCDR3; SEQ ID NO. 12), or (iv) It competes with any one of the antibodies (i) to (iii) above.

[0020] The present invention also provides a method for treating or preventing the onset of progressive chronic interstitial lung disease, comprising administering a therapeutically effective amount of an anti-transglutaminase 2 (anti-TG2) antibody. For example, the anti-TG2 that can be administered in such a method comprises the following sequence: (i) KASQDINSYLT (LCDR1; SEQ ID NO. 1); LVNRLVD (LCDR2; SEQ ID NO. 2); LQYDDFPYT (LCDR3; SEQ ID NO. 3); THAMS (HCDR1; SEQ ID NO. 4); TISSGGRSTYYPDSVKG (HCDR2; SEQ ID NO. 5); and LISTY (HCDR3; SEQ ID NO. 6); or (ii) KASQDINSYLT (LCDR1; SEQ ID NO. 1); LTNRLMD (LCDR2; SEQ ID NO. 7); LQYVDFPYT (LCDR3; SEQ ID NO. 8); SSAMS (HCDR1; SEQ ID NO. 9); TISSGGRSTYYPDSVKG (HCDR2; SEQ ID NO. 5); and LISPY (HCDR3; SEQ ID NO. 10); or (iii) KASQDINSYLT (LCDR1; SEQ ID NO. 1); RTNRLFD (LCDR2; SEQ ID NO. 11); LQYDDFPYT (LCDR3; SEQ ID NO. 3); SSAMS (HCDR1); TISVGGGKTYYPDSVKG (HCDR2; SEQ ID NO. 9); and LISLY (HCDR3; SEQ ID NO. 12), or (iv) It competes with any one of the antibodies (i) to (iii) above.

[0021] The use of an anti-transglutaminase 2 (anti-TG2) antibody for the manufacture of a medicament for the treatment of progressive chronic interstitial lung disease or for the prevention of the onset of progressive chronic interstitial lung disease has also been described. For example, an anti-TG2 for use according to the invention may comprise the following sequence: (i) KASQDINSYLT (LCDR1; SEQ ID NO. 1); LVNRLVD (LCDR2; SEQ ID NO. 2); LQYDDFPYT (LCDR3; SEQ ID NO. 3); THAMS (HCDR1; SEQ ID NO. 4); TISSGGRSTYYPDSVKG (HCDR2; SEQ ID NO. 5); and LISTY (HCDR3; SEQ ID NO. 6); or (ii) KASQDINSYLT (LCDR1; SEQ ID NO. 1); LTNRLMD (LCDR2; SEQ ID NO. 7); LQYVDFPYT (LCDR3; SEQ ID NO. 8); SSAMS (HCDR1; SEQ ID NO. 9); TISSGGRSTYYPDSVKG (HCDR2; SEQ ID NO. 5); and LISPY (HCDR3; SEQ ID NO. 10); or (iii) KASQDINSYLT (LCDR1; SEQ ID NO. 1); RTNRLFD (LCDR2; SEQ ID NO. 11); LQYDDFPYT (LCDR3; SEQ ID NO. 3); SSAMS (HCDR1); TISVGGGKTYYPDSVKG (HCDR2; SEQ ID NO. 9); and LISLY (HCDR3; SEQ ID NO. 12), or (iv) It competes with any one of the antibodies (i) to (iii) above.

[0022] Another object of the present invention is an anti-transglutaminase 2 (anti-TG2) antibody for use in treating a subject with pulmonary fibrosis associated with COVID infection or for preventing the development of pulmonary fibrosis in a subject suffering from COVID infection. For example, an anti-TG2 antibody for use according to the present invention may comprise the following sequence: (i) KASQDINSYLT (LCDR1; SEQ ID NO. 1); LVNRLVD (LCDR2; SEQ ID NO. 2); LQYDDFPYT (LCDR3; SEQ ID NO. 3); THAMS (HCDR1; SEQ ID NO. 4); TISSGGRSTYYPDSVKG (HCDR2; SEQ ID NO. 5); and LISTY (HCDR3; SEQ ID NO. 6); or (ii) KASQDINSYLT (LCDR1; SEQ ID NO. 1); LTNRLMD (LCDR2; SEQ ID NO. 7); LQYVDFPYT (LCDR3; SEQ ID NO. 8); SSAMS (HCDR1; SEQ ID NO. 9); TISSGGRSTYYPDSVKG (HCDR2; SEQ ID NO. 5); and LISPY (HCDR3; SEQ ID NO. 10); or (iii) KASQDINSYLT (LCDR1; SEQ ID NO. 1); RTNRLFD (LCDR2; SEQ ID NO. 11); LQYDDFPYT (LCDR3; SEQ ID NO. 3); SSAMS (HCDR1); TISVGGGKTYYPDSVKG (HCDR2; SEQ ID NO. 9); and LISLY (HCDR3; SEQ ID NO. 12), or (iv) It competes with any one of the antibodies (i) to (iii) above.

[0023] The present invention also provides a method for treating a patient with pulmonary fibrosis associated with COVID infection or for preventing the development of pulmonary fibrosis in a patient suffering from COVID infection, comprising administering a therapeutically effective amount of an anti-transglutaminase 2 (anti-TG2) antibody. For example, an anti-TG2 that can be administered in such a method can comprise the following sequence: (i) KASQDINSYLT (LCDR1; SEQ ID NO. 1); LVNRLVD (LCDR2; SEQ ID NO. 2); LQYDDFPYT (LCDR3; SEQ ID NO. 3); THAMS (HCDR1; SEQ ID NO. 4); TISSGGRSTYYPDSVKG (HCDR2; SEQ ID NO. 5); and LISTY (HCDR3; SEQ ID NO. 6); or (ii) KASQDINSYLT (LCDR1; SEQ ID NO. 1); LTNRLMD (LCDR2; SEQ ID NO. 7); LQYVDFPYT (LCDR3; SEQ ID NO. 8); SSAMS (HCDR1; SEQ ID NO. 9); TISSGGRSTYYPDSVKG (HCDR2; SEQ ID NO. 5); and LISPY (HCDR3; SEQ ID NO. 10); or (iii) KASQDINSYLT (LCDR1; SEQ ID NO. 1); RTNRLFD (LCDR2; SEQ ID NO. 11); LQYDDFPYT (LCDR3; SEQ ID NO. 3); SSAMS (HCDR1); TISVGGGKTYYPDSVKG (HCDR2; SEQ ID NO. 9); and LISLY (HCDR3; SEQ ID NO. 12), or (iv) It competes with any one of the antibodies (i) to (iii) above.

[0024] Also described is the use of an anti-transglutaminase 2 (anti-TG2) antibody for the manufacture of a medicament in the treatment of a subject with pulmonary fibrosis associated with COVID infection or in the prevention of the development of pulmonary fibrosis in a subject suffering from COVID infection. For example, an anti-TG2 for use according to the invention may comprise the following sequence: (i) KASQDINSYLT (LCDR1; SEQ ID NO. 1); LVNRLVD (LCDR2; SEQ ID NO. 2); LQYDDFPYT (LCDR3; SEQ ID NO. 3); THAMS (HCDR1; SEQ ID NO. 4); TISSGGRSTYYPDSVKG (HCDR2; SEQ ID NO. 5); and LISTY (HCDR3; SEQ ID NO. 6); or (ii) KASQDINSYLT (LCDR1; SEQ ID NO. 1); LTNRLMD (LCDR2; SEQ ID NO. 7); LQYVDFPYT (LCDR3; SEQ ID NO. 8); SSAMS (HCDR1; SEQ ID NO. 9); TISSGGRSTYYPDSVKG (HCDR2; SEQ ID NO. 5); and LISPY (HCDR3; SEQ ID NO. 10); or (iii) KASQDINSYLT (LCDR1; SEQ ID NO. 1); RTNRLFD (LCDR2; SEQ ID NO. 11); LQYDDFPYT (LCDR3; SEQ ID NO. 3); SSAMS (HCDR1); TISVGGGKTYYPDSVKG (HCDR2; SEQ ID NO. 9); and LISLY (HCDR3; SEQ ID NO. 12), or (iv) It competes with any one of the antibodies (i) to (iii) above.

[0025] In the context of the present invention as a whole, a chronic progressive lung fibrotic disease is characterized by an increase in a marker in a subject sample, for example, TG2 activity, TG2 expression (e.g., an increase in mRNA encoding TG2 or TG2 antigen), TG2 export, or any combination thereof, and the subject sample is a cell or tissue (e.g., lung cell or lung tissue) associated with the disease. The increase in the marker can be determined by any means in the cell / tissue associated with the disease. The increase in a marker in a subject sample is typically determined by comparing the level of the marker in the subject sample with the level of the same marker in normal cells of the same tissue type (i.e., basal level; e.g., basal TG2 activity, basal expression level (mRNA level and / or protein level) and / or basal level of TG2 export). A subject sample in which the level of at least one marker is 10% or more, 15% or more, 20% or more, 25% or more, or even 30% or more compared to the basal level of the marker is considered to show an increase in the marker. For example, an increase in TG2 expression (alternatively referred to as TG2 overexpression) can be determined by determining the amount of TG2 mRNA in the patient's lung cells. Chronic progressive pulmonary fibrosis cells / tissues are characterized, for example, by an increase in the amount of TG2 mRNA (representing overexpression) in the subject's lung cells compared to normal cells of the same tissue type. The expression of TG2 mRNA can be increased by any amount, such as 10% or more, 15% or more, 20% or more, 25% or more, or even 30% or more compared to the basal level.The amount of mRNA can be measured using known methods, such as quantitative reverse transcription polymerase chain reaction (qRT-PCR), real-time qRT-PCR, quantigene assay (Affymetrix / Thermo Fisher), RNA sequencing by Northern blotting or using microarrays, various in situ hybridization methods (e.g., RNAscope). Alternatively, overexpression can be determined by determining the amount of TG2 antigen in the patient's lung cells. Thus, chronic progressive pulmonary fibrosis cells are characterized by an increased amount (overexpression) of TG2 protein (or TG2 antigen) in the subject's lung cells, such as compared to normal cells of the same tissue type. The expression of TG2 protein can be increased by any amount, such as 10% or more, 15% or more, 20% or more, 25% or more, or even 30% or more, compared to basal levels. The amount of protein can be measured using known methods such as immunohistochemistry, Western blotting, mass spectrometry or fluorescence activated cell sorting (FACS), including the use of the anti-TG2 antibody of the present invention. The threshold for determining expression may vary depending on the technique used and may be verified against the immunohistochemistry score. Alternatively, chronic progressive pulmonary fibrosis cells are characterized by elevated TG2 activity in the subject's lung cells compared to normal cells of the same tissue type. TG2 activity can be increased by any amount, such as 10% or more, 15% or more, 20% or more, 25% or more, or even 30% or more, compared to basal levels. TG2 activity can be measured using known methods, such as via frozen biopsy (TG-ISA), exhaled breath condensate (EBC) or bronchoalveolar lavage fluid (BALF).

[0026] The anti-TG2 antibody for use, the method for treatment or prevention, or the use of an anti-TG2 according to the invention, for example for the treatment or prevention of a progressive chronic interstitial lung disease or for the treatment / prevention of pulmonary fibrosis associated with COVID infection in a subject, may comprise: Thus, the method may include the steps of: (a) measuring TG2 expression, TG2 activity or TG2 transport in a sample (e.g., lung cells) from a subject; (b) comparing the measurement results obtained from (a) with corresponding measurements in normal cells / tissues (e.g., lung cells); and (c) if increased expression (i.e., overexpression of TG2), increased activity or increased transport is observed, administering an anti-TG2 antibody to the patient, thereby treating or preventing pulmonary fibrosis associated with progressive chronic interstitial lung disease or COVID infection. The TG2 expression level measured in step (a) may be either mRNA or protein level, and the increase may be any increase in expression as described above. It is not necessary to obtain a corresponding measurement in normal cells / tissues each time a comparison is made. The corresponding measurement may be obtained at any time before the comparison is made, and may be the average TG2 expression level or TG2 activity in normal cells / tissues.

[0027] In the context of the present invention as a whole, the progressive chronic interstitial lung disease is selected from the group consisting of idiopathic pulmonary fibrosis (IPF), desquamative interstitial pneumonia (DIP), acute interstitial pneumonia (AIP; otherwise known as Hamman-Rich syndrome), hypersensitivity pneumonitis (HSP), non-specific interstitial pneumonia (NSIP), respiratory bronchiolitis associated interstitial lung disease (RB-ILD), cryptopathic organizing pneumonia (COP; otherwise known as Bronchiolitis Obliterans Organizing Pneumonia or BOOP), sarcoidosis, asbestosis and lymphocytic interstitial pneumonia (LIP). Preferably, the progressive chronic interstitial lung disease is selected from the group consisting of idiopathic pulmonary fibrosis (IPF).

[0028] In the context of the present invention as a whole, the anti-TG2 antibody preferably binds to an epitope within the core region of transglutaminase type 2 (TG2) and inhibits TG2 activity, wherein said core region consists of amino acids 143 to 473 of TG2 (e.g. SEQ ID NO: 41) and the TG2 activity inhibited is TG2 cross-linking of lysine and glutamine via an N-ε (γ-glutamyl) lysine isopeptide bond. More preferably, the antibody binds to a region comprising or consisting of amino acids 304 to 326 of TG2 (e.g. SEQ ID NO: 41) or a part of this region. The antibody comprises or consists of an intact antibody. Alternatively, it may comprise or consist of antigen-binding fragments such as (but are not limited to) Fv fragments (e.g., single chain Fv fragments or disulfide-linked Fv fragments); Fab fragments; and Fab-like fragments (e.g., Fab' fragments or F(ab)2 fragments), single domain antibodies (or any other fragments defined herein or known by the skilled artisan). Preferably, the anti-TG2 antibody used in accordance with the present invention is a) comprises six CDRs selected from the group consisting of: (i) KASQDINSYLT (LCDR1; SEQ ID NO. 1); LVNRLVD (LCDR2; SEQ ID NO. 2); LQYDDFPYT (LCDR3; SEQ ID NO. 3); THAMS (HCDR1; SEQ ID NO. 4); TISSGGRSTYYPDSVKG (HCDR2; SEQ ID NO. 5); and LISTY (HCDR3; SEQ ID NO. 6); or (ii) KASQDINSYLT (LCDR1; SEQ ID NO. 1); LTNRLMD (LCDR2; SEQ ID NO. 7); LQYVDFPYT (LCDR3; SEQ ID NO. 8); SSAMS (HCDR1; SEQ ID NO. 9); TISSGGRSTYYPDSVKG (HCDR2; SEQ ID NO. 5); and LISPY (HCDR3; SEQ ID NO. 10); or (iii) KASQDINSYLT (LCDR1; SEQ ID NO. 1); RTNRLFD (LCDR2; SEQ ID NO. 11); LQYDDFPYT (LCDR3; SEQ ID NO. 3); SSAMS (HCDR1); TISVGGGKTYYPDSVKG (HCDR2; SEQ ID NO. 9); and LISLY (HCDR3; SEQ ID NO. 12), b) a light chain variable domain having a sequence as defined in any one of SEQ ID NO:13 to SEQ ID NO:27 and a heavy chain variable domain having a sequence as defined in any one of SEQ ID NO:28 to SEQ ID NO:40; c) a light chain variable domain having at least 80% identity or similarity, preferably at least 90% identity or similarity or preferably at least 95% identity or similarity, to a sequence defined in any one of SEQ ID NOs: 13 to 27, and a heavy chain variable domain having at least 80% identity or similarity, preferably at least 90% identity or similarity or preferably at least 95% identity or similarity, to a sequence defined in any one of SEQ ID NOs: 28 to 40, d) Competing the binding of an epitope comprising or consisting of amino acids 304 to 326 of TG2 (eg, SEQ ID NO. 41) or a part of this region with an antibody defined in a), b) or c) above.

[0029] [Table A-1]

[0030] [Table A-2]

[0031] [Table A-3]

[0032] Whether an antibody binds to the same epitope as another antibody or competes for binding with another antibody can be easily determined using routine methods known in the art. For example, to determine whether a test antibody binds to the same epitope as a reference antibody of the present invention, the reference antibody is bound to a protein or peptide under saturating conditions. The ability of the test antibody to bind to the protein or peptide is then evaluated. If the test antibody can bind to the protein or peptide after saturation binding with the reference antibody, it can be concluded that the test antibody binds to a different epitope than the reference antibody. On the other hand, if the test antibody cannot bind to the protein or peptide after saturation binding with the reference antibody, the test antibody may bind to the same epitope as the epitope bound by the reference antibody of the present invention. To determine whether an antibody competes for binding with a reference antibody, the binding methodology described above is performed in two orientations. In the first orientation, the reference antibody is allowed to bind to the protein / peptide under saturating conditions and then binding of the test antibody to the protein / peptide molecule is assessed. In the second orientation, the test antibody is allowed to bind to the protein / peptide under saturating conditions and then binding of the reference antibody to the protein / peptide is assessed. In either orientation, if only the first antibody (saturating antibody) is able to bind to the protein / peptide, it is concluded that the test antibody and the reference antibody compete for binding to the protein / peptide. As will be appreciated by those skilled in the art, an antibody that competes with a reference antibody for binding will not necessarily bind to the same epitope as the reference antibody, but may sterically inhibit binding of the reference antibody by binding to an overlapping or adjacent epitope.

[0033] Two antibodies bind to the same or overlapping epitope if each competitively inhibits (blocks) the binding of the other antibody to the antigen, i.e., a 1-, 5-, 10-, 20- or 100-fold excess of one antibody inhibits binding of the other antibody by at least 50%, 75%, 90% or even 99% when measured in a competitive binding assay. Alternatively, two antibodies have the same epitope if essentially all amino acid mutations in the antigen reduce or eliminate binding of one antibody and reduce or eliminate binding of the other antibody. If some amino acid mutations reduce or eliminate binding of one antibody and also reduce or eliminate binding of the other antibody, then the two antibodies have overlapping epitopes. Additional routine experiments (e.g. peptide mutations and binding analysis) can then be performed to confirm whether the observed lack of binding of the test antibody is indeed due to binding to the same epitope as the reference antibody or whether steric hindrance (or another phenomenon) is responsible for the observed lack of binding. This type of experiment can be performed using ELISA, RIA, surface plasmon resonance, flow cytometry or other quantitative or qualitative antibody binding assays available in the art.

[0034] Any subject can be treated according to the present invention. Preferably, the subject is a human. However, the subject may also be other mammals, such as non-human primates, horses, cows, sheep, pigs, dogs, cats, rabbits, rats, mice, guinea pigs, or hamsters. Alternatively, the term patient may be used interchangeably in place of subject.

[0035] Any of the anti-TG2 antibodies of the present invention can be incorporated into a pharmaceutical composition suitable for administration to a subject in any manner, including, but not limited to, topical, intranasal, intradermal, intravenous, subcutaneous, or intramuscular. Typically, the pharmaceutical composition comprises an anti-TG2 antibody and one or more pharma- ceutically acceptable adjuvants and / or carriers. Accordingly, also described herein is a pharmaceutical composition for use in the treatment of progressive chronic interstitial lung diseases, such as idiopathic pulmonary fibrosis (IPF), comprising an anti-TG2 antibody and one or more pharma- ceutical acceptable adjuvants and / or carriers.The pharmaceutical composition of the present invention may be part of a kit with instructions for use, including instructions for use and, optionally, devices for intravenous, subcutaneous or intramuscular administration to an individual in need thereof.

[0036] As used herein, a "pharmaceutically acceptable carrier" includes any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, etc. that are physiologically compatible and suitable for administration to a subject for the methods and uses described herein. Examples of pharmaceutically acceptable carriers include one or more of water, saline, phosphate buffered saline, dextrose, glycerol, ethanol, and the like, and combinations thereof. Depending on the route of administration or the type of formulation (such as liquid, freeze-dried, or spray-dried formulation), isotonicity agents, e.g., sugars, polyalcohols such as mannitol, sorbitol, or sodium chloride, may be included in the composition. Pharmaceutically acceptable carriers may contain minor amounts of auxiliary substances, such as wetting or emulsifying agents, preservatives or buffers, which enhance the shelf life or effectiveness of the antibody or antibody portion.

[0037] The pharmaceutical compositions of the present invention may be in a variety of forms. These include, for example, liquid solutions (e.g., injection or infusion solutions), dispersions or suspensions, powders, and liposomes. The preferred form will depend on the intended mode of administration or therapeutic application. Exemplary preferred compositions are in the form of injectable or infusible solutions, such as compositions similar to those used for passive immunization of humans with other antibodies.

[0038] The appropriate dosage of the anti-TG2 antibody of the present invention can be determined by a skilled physician. The actual dosage of the active ingredient in the pharmaceutical composition of the present invention can be varied to obtain an amount of the active ingredient that is effective to achieve the desired therapeutic response for a particular patient, composition and mode of administration and is not toxic to the patient. The selected dosage will depend on various pharmacokinetic factors, such as the route of administration, the time of administration, the rate of excretion of the antibody, the duration of treatment, other drugs, compounds and / or materials used in combination with the particular antibody, the age, sex, weight, condition, general health and medical history of the patient being treated.

[0039] Suitable doses can be, for example, in the range of about 0.01 pg / kg to about 1000 mg / kg body weight of the patient to be treated, typically about 0.1 pg / kg to about 100 mg / kg body weight. The dosage regimen can be adjusted to provide the optimum desired response (e.g., therapeutic response). For example, a single dose can be administered, or several doses can be administered over time. As used herein, a dosage unit form refers to a physically discrete unit suitable as a unitary dosage for the subject to be treated, each unit containing a predetermined amount of active compound calculated to produce the desired therapeutic effect in association with the required pharmaceutical arena. Administration can be a single dose or multiple doses. Multiple doses can be administered by the same or different routes and to the same or different sites. In the context of the present invention as a whole, the anti-TG2 antibody can be co-administered with one or more other therapeutic agents. The co-administration of two or more agents can be accomplished in a variety of ways. They can be administered together in a single composition, or in separate compositions as part of a combination therapy. For example, one can be administered before the other, separately, or after the other, sequentially, simultaneously with the other, or simultaneously. [Brief description of the drawings]

[0040] [Figure 1]Collagen staining is elevated in lung biopsies from IPF patients. (A) Percentage of tissue section area positive for picosirius red staining for dense collagen fibers associated with mature scar tissue. (B) Percentage of non-airway collagen in total tissue section. Non-fibrotic (patients requiring lung biopsy without ILD) served as control samples (n=8). IPF / UIP samples were pooled and averaged (n=9). *p<0.05 by unpaired, 2-sided t-tests.

[0041] [Diagram 2] TG2 mRNA is elevated in biopsies from patients with IPF (RNAscope). Formalin-fixed paraffin-embedded lung tissue samples were probed for expression of Tgm2 using the in-situ hybridization technique RNAscope. Red / pink staining represents Tgm2 mRNA transcripts. (A): Total number of Tgm2 positive cells within the biopsy. (B): Tgm2 positive cells by staining intensity. Staining intensity is represented as 1+ (1-6 probes per cell), 2+ (7-13 probes per cell), 3+ (≥14 probes per cell) (p<0.015). Only 2 of 10 "non-fibrotic" biopsies analyzed are shown, the rest failed the RNA quality QC check.

[0042] [Diagram 3](A) TG2 antigen is elevated in biopsies from patients with IPF. TG2 immunohistochemical staining of paraffin-fixed sections from nonfibrotic and IPF samples was analyzed, and the percentage of stained area was calculated by high content image analysis. Nonfibrotic samples (n=7) and IPF / UIP samples (n=10) were averaged. p=0.03 by unpaired, 2-sided t-tests. (B) Lung function measurements in patients, using forced vital capacity (FVC) as percent predicted (n=10). Linear regression was performed between FVC and collagen, as determined by PSR staining (see circles; P=0.0128, R2=0.5597) and SHG staining (see triangles; P=0.0983, R2=0.3043).

[0043] [Figure 4] Knockout (KO) mice are protected from increased collagen as measured by second harmonic generation microscopy, or SHG, in the bleomycin model of interstitial lung disease. (A): Percentage of interstitial collagen area in wild-type and TG2 KO mice. (B): Fold increase in pulmonary interstitial collagen in response to bleomycin in wild-type and TG2 KO mice (n=9-12).

[0044] [Diagram 5] TG2 knockout mice do not have elevated mRNA (RT QPCR) for major interstitial collagens in the bleomycin model of interstitial lung disease. N=5 mice / group. ANOVA with Tukey's post-hoc test.

[0045] [Figure 6]TG2 knockout mice preserved lung function in a bleomycin interstitial lung disease model. (A): Resistance (Rrs=respiratory system resistance (total); Rn=Newtonian (airway) resistance). (B): Compliance (Crs=dynamic compliance (respiratory system compliance), Cst=quasi-static compliance). (C): Pressure volume loop (K=curvature of the contractile edge of the PV loop, PV=pressure volume). (D): Elastance (Ers=respiratory system elastance (total), H=tissue elastance); **P<0.01, ***P<0.001 by ANOVA with Tukey's post-hoc test.

[0046] [Figure 7] (A): Representative images of TGFβ1 stimulated cells showing uptake of 5-BP (i.e. transglutaminase activity) immediately (left panel), 1 hour (middle panel) and 3 hours (right panel) after scratching, showing loss of activity within the first hour after scratching. (B): Fibronectin and TG2 activity were quantified in the cell layer (away from the scratch) and at the scratch border for non-fibrotic (NF) and fibrotic (F) fibroblasts with and without TGFβ1 treatment. 5-BP was added immediately after introducing the scratch injury and cells were fixed 1 hour later. Each point represents staining in one well. P values ​​are as indicated. (C): Monolayers of TGFβ1 treated lung fibroblasts were scratched and 5-BP was added immediately as above and treated with vehicle (left panel) or the TG2 blocking antibody 300 uM rbBB7 (right panel). rbBB7 completely inhibited 5-BP uptake as shown by the absence of staining.

[0047] [Figure 8]Rabbit Pulmonary Silicosis Study Design. New Zealand White rabbits were administered saline or silica after bronchoscopy incision in the right lower lobe of the lung. Six experimental groups were established: Five animals (group 1) were treated with saline and left for 56 days as the "healthy" group; 40 were treated with silica; 10 animals (group 2) were withdrawn after 28 days and used as indicators of disease at the start of treatment administration; 10 animals were treated with vehicle (group 3, n=5) or control IgG rb922 (group 4, n=5) and carried out as "untreated" controls for 56 days (because the results for the vehicle and control IgG groups were essentially similar, this group was combined into a single group of 10 untreated controls). Ten animals were treated with the TG2 blocking rabbit antibody rbBB7 from days 28 to 56 as a therapeutic group (group 5), and 10 animals were treated with rbBB7 from days 1 to 56 as a "prophylactic" group. rbBB7 was administered every 5 days. Terminal analyses consisted of TG2 activity (TG ISA), TG2 antigen (TG2 Ant), Picosirius red staining (PSR), Masson's Trichrome staining (MT), second harmonic generation quantification of lung collagen (HG), and various messenger RNAs (mRNAs).

[0048] [Figure 9] rbBB7 serum exposure in a rabbit silicosis model. Prophylactic (from day 1, plots A and B) and therapeutic (from day 26 to day 56, plots C and D) treatments were administered on two separate occasions and therefore plotted separately.

[0049] [Figure 10]In a rabbit silicosis model of ILD, rbBB7 reduced total lung collagen as measured by Picrosirius red staining. Collagen was measured in "parenchymal cells" (excluding large airways containing pre-existing collagen unrelated to the disease process) (A) and within active fibrotic lesions (B). Silica d28 and silica d56 groups were discontinued on days 28 or 56, respectively, and either received nothing (d28) or vehicle / control IgG (d56) and were not treated with any drug. Silica+rbBB7 (d28-d56) is the treatment group receiving rbBB7 from days 28 to 56. Silica+rbBB7 (days 1-56) is the prophylactic group receiving rbBB7 starting 1 day before silica injection. Each point represents one rabbit. Statistics between groups are 1-way Students t-tests, showing suppression of fibrosis in both the prophylactic and treatment groups. When analyzed by ANOVA, the overall treatment effect was significant (lesional collagen, p = 0.002; parenchymal collagen, p = 0.0037).

[0050] [Figure 11] rbBB7 reduced total lung collagen as measured by second harmonic generation microscopy (SHG) in a rabbit silicosis model of ILD. Silica d28 and silica d56 groups were discontinued on days 28 and 56, respectively, and either received nothing (d28) or vehicle / control IgG (d56) and were not treated with any drug. Silica+rbBB7 (d28-d56) is the treatment group receiving rbBB7 from days 28 to 56. Silica+rbBB7 (days 1-56) is the prophylactic group receiving rbBB7 starting 1 day before silica injection. Each point represents one rabbit. Statistics between groups are one-way Student's t-tests, showing that fibrosis is suppressed in both the prophylactic and treatment groups.

[0051] [Figure 12]rbBB7 inhibits TG2 activity in a rabbit silicosis model. TG2 antigen (A) and in situ extracellular TG (ISA) activity (B) were measured at the time of animal termination using high-content image analysis of lung sections stained for TG2 antigen using immunofluorescence or TG2 activity by uptake of fluorochrome-labeled cadaverine, respectively, in the study described in Figure 11. Staining was quantified as the area percentage of tissue defined by nuclear DAPI staining. *=P<0.05 by ANOVA. Data show that TG2 antigen levels were unchanged, but TG activity was clearly increased in the silica-treated group, decreased in the therapeutically treated group, and significantly decreased in the prophylactically treated group.

[0052] [Figure 13] TG2 mRNA (RNAscope) was upregulated in postmortem lung samples from patients who died from COVID-19 infection (caused by the SARS-Cov2 virus). Formalin-fixed, paraffin-embedded lung tissue samples were probed for Tgm2 expression using the in-situ hybridization technique RNAscope. Black dots / stains represent Tgm2 mRNA transcripts. TG2 mRNA expression was compared between normal and IPF lung tissue.

[0053] [Figure 14] Expression of Tgm2 in postmortem bovine (Covid) lungs. Expression of Tgm2 was assessed in lung tissue from normal, IPF, and subjects who died as a result of COVID infection. Expression was calculated as the percentage area of ​​tissue sections stained for Tgm2. Tgm2 was significantly elevated in postmortem COVID tissue compared to healthy control tissue (*=P<0.05 by ANOVA). EXAMPLES

[0054] material: Anti-TG2 antibody: The anti-TG2 mAb used in the following examples contained a light chain variable region defined in SEQ ID NO:25 and a heavy chain variable region defined in SEQ ID NO:38, which is a rabbitised version of the original BB7 and is referred to in the following examples as rbBB7. Zampilimab (also known as UCB7858, derived from antibody DC1) is an anti-TG2 antibody with a variable light chain according to SEQ ID NO.24 and a variable heavy chain according to SEQ ID NO.37, a humanized antibody that specifically binds to human TG2. rbBB7 was developed to mimic its effects in animal models such as rabbits. Zampilimab / DC1 and rbBB7 / BB7 have been shown to behave similarly. They bind to the same epitope in the TG2 core (aa 313-325 of SEQ ID NO.41), have nearly identical IC50 (0.25 vs. 0.3 nM) and Kd (<50 vs. <60 pm) against human TG2, and inhibit ECM accumulation comparably in in vitro cell-based evaluations. The only notable difference is the inferior IC50 of Zampilimab against rabbit TG2 (103 vs. 8 nM). Thus, the findings from the following examples using rbBB7 are fully applicable to Zampilimab and other anti-TG2 antibodies as described herein.

[0055] Example 1 - Collagen and TG2 expression in IPF: The aim of this study was to determine whether the presence of collagen and expression of TG2 are associated with pulmonary fibrosis in human patients.

[0056] method: Human tissue: Lung biopsy samples were obtained from the NIH and the University of Rochester. Ten non-fibrotic and ten IPF patients were analyzed.

[0057] Tissue sample preparation: Formalin-fixed, paraffin-embedded human samples were serially sectioned and stained to correlate the degree of fibrosis with TG2. Staining was performed to assess fibrosis as measured by collagen content and transglutaminase type 2 (TG2) mRNA and protein expression.

[0058] Picrosirius Red Staining (PSR): Sample sections were stained with picrosirius red, which stains collagen fibers bright red and non-collagenous regions pink or tan. Whole slides were scanned with a Zeiss Axio Z.1 scanner using Zeiss Zen 2.6 (Blue Edition) software. Images were processed with multiphase analysis using Definiens Tissue Studio software. For image processing, tissue masks were created to identify areas of lesion (active fibrosis), parenchyma (prefibrosis), white space, or airway collagen. The percentage of PSR-positive staining within each mask was calculated as follows: ([% area of ​​mask] × [% positive staining in mask]) / 100 = % collagen in area. The sum of the collagen content within the lesion and parenchymal regions represents the total non-airway collagen content within the biopsy.

[0059] TG2 Protein Expression: Immunohistochemistry for TG2 was performed using the automated staining platform Leica Bond RX with the following staining protocol: Dewax (Bond Dewax Solution, 72°C, 30 min), H1(20) antigen retrieval (Bond ER solution 1, 100°C), and DAB 30 Min Marker detection (using mouse anti-TG2 antibody DH2 (UCB, internal antibody) at 83 ng / ml). Staining was quantified using multiphase field analysis in Definiens tissue studio software.

[0060] TG2 mRNA: TG2 mRNA was assessed in formalin-fixed, paraffin-embedded (FFPE) tissues using the RNAscope In Situ Hybridization (ISH) assay using the RNAscope 2.5 LS Reagent Kit Red (Advanced Cell Diagnostics) and the Leica Bond Polymer Refine Red Detection Kit on a Leica Bond RX processor according to the manufacturer's instructions. Tissue quality was assessed by performing RNAscope analysis for mRNA of the housekeeping gene Homo sapiens ubiquitin C mRNA. Sections were taken at 5 μm thickness onto Superfrost Plus Gold slides and dried overnight at 37°C following the 'Bake and Dewax' protocol in the Leica Bond RX factory. Slides were placed in the staining rack of the Leica BOND RX without pretreatment, baked in place at 60°C, and then dewaxed using ethanol before rehydration. Heat-induced RNA recovery was performed by incubation in recovery buffer ER2 (pH 9, AR9640 Leica) at 95°C for 15 min, followed by protease treatment (Advanced Cell Diagnostics) for 15 min, and peroxidase blocking with two rinses in distilled water between pretreatments. Briefly, 20 ZZ probe pairs targeting relevant genomic nuclear protein genes were designed (target nucleotides Target 160-2563) and synthesized by Advanced Cell Diagnostics. Sections were exposed to ISH target probes and incubated at 42°C for 2 h. Hs-TGM2 (Advanced Cell Diagnostics). A probe against the bacterial gene DapB mRNA was used as a negative control for each run. After washing with water, ISH signals were amplified using a company-provided pre-amplifier and an amplifier coupled to alkaline phosphatase (AP) and incubated with red substrate-developing solution for 10 min at room temperature.Sections were then counterstained with hematoxylin, air-dried, and mounted with Ecomount permanent mounting medium (Biocare Medical). Images were acquired with an Olympus slide scanner and quantified based on the number of cells stained positive for TG2 mRNA and the staining intensity (probe number) within each cell using Halo image analysis software from imaging specialist Oracle Bio.

[0061] result: Presence of collagen (Figure 1): Collagen was visualized using picrosirius red (PSR) staining. Whole biopsy scans showed denser tissue and less airspace in IPF biopsies compared with nonfibrotic samples, although some of these "nonfibrotic" controls had obvious remodelling occurring (photos not shown). Four regions were defined within the tissue: (1) active fibrotic lesion area was assigned based on the high density of red collagen fibers, (2) total parenchyma was defined by low density of red collagen fibers in areas dominated by pink or yellow staining of collagen-free tissue, (3) air spaces were defined by white spaces or unstained areas of the slide within the tissue periphery, and (4) airway collagen was defined as dense areas of collagen surrounding the airways (this is a normal feature of lung histopathology and is not expected to change significantly in fibrotic disease). Comparing IPF and non-IPF sections, lesion size and dense collagen areas increased, whereas air space and parenchymal tissue areas decreased. The proportion of non-airway collagen in the tissue also increased significantly (Figure 1A / B).

[0062] Expression of TG2 (Figures 2 and 3): Tissue transglutaminase 2 is encoded by the Tgm2 gene. Quantification of the corresponding mRNA revealed an overall significant increase in the number of TG2 mRNA positive cells per area in IPF biopsies (Figure 2A). The number of probes per cell was also quantified by subdividing cells expressing low, medium, and high TG2 RNA per cell (Figure 2B). The number of cells expressing high TG2 mRNA was significantly increased threefold in the IPF group compared to the nonfibrotic group. Immunohistochemical staining for TG2 confirmed the localization of protein expression in tissues. Observations showed that TG2 positive staining was localized to the epithelium in nonfibrotic tissues and was strongly localized to fibrotic areas in IPF samples. IPF samples showed a 42% increase in the area of ​​TG2 positive staining in whole biopsies compared to nonfibrotic samples (Figure 3A). As shown in Figure 3B, the reduction in forced vital capacity (FVC) was significantly associated with the percentage of non-airway collagen determined by PSR staining (circles; P = 0.0128, R2 = 0.5597). The decline in FVC was also associated with collagen detected by SHG, but not significantly (triangles, P = 0.0983, R2 = 0.3043).

[0063] Conclusions of Example 1: Total lung collagen was significantly increased in IPF tissue and correlated with decreased lung function (R2 0.56, p=0.01), consistent with the expected disease phenotype and previous reports. IPF lung tissue showed increased expression of TG2 mRNA and detectable TG2 protein in fixed samples.

[0064] Example 2 - Knockout Animal Model TG2 knockout mice have previously been shown to be protected from fibrotic remodeling in a pulmonary bleomycin model of interstitial lung disease (Olsen et al., 2011), but there remains a critical need to correlate histological protection with functional benefits in the lung when TG2 is reduced.

[0065] method: All animal procedures were performed under approved accreditation. Wild-type and TG2 knockout mice: Mice used in these experiments were either commercially available male or female C57BL / 6J (The Jackson Laboratories, n=20) or in-house bred TG2 knockout mice (n=21). The TG2 knockout mice were the Tgm2tm1.1Rmgr (Victor Chang Institute) strain, in which exons 6-8 of the Tgm2 gene were deleted by Cre-mediated recombination. This strain was then backcrossed to C56BL / 6J for at least 10 generations and confirmed by genome scanning.

[0066] Bleomycin model: Mice were oropharyngeal aspirated with 2 U / kg bleomycin (Fresenius Kabi) in 40 μl saline or 40 μl saline alone under isoflurane anesthesia. After 21 days, mice were euthanized according to the current guidelines of the American Veterinary Medical Association Panel on Euthanasia. The heart and lungs were removed en bloc. The right bronchus was severed and the right lung lobe was frozen in liquid nitrogen. The left lung was inflated with 2% low melting point agarose and placed in 10% neutral buffered formalin or inflated with neutral buffered formalin and placed in formalin overnight.

[0067] Histology: Mouse lung pieces were fixed overnight in 10% neutral buffered formalin, then transferred to 70% ethanol and embedded in paraffin. Five micron sections were stained with H&E, Masson's Trichrome or Picrosirius Red using standard methods. In some experiments, collagen staining was quantified with Oracle Definiens Tissue studio software.

[0068] Pulmonary function tests: For pulmonary function tests, mice were anesthetized, tracheally cannulated, and connected to a ventilator connected to a FlexiVent device (SciReq). Mice were anesthetized with 2 mg / kg vecuronium bromide intraperitoneally (ip) to prevent spontaneous respiratory efforts, and a series of forced ventilation maneuvers, usually lasting approximately 10 min, were performed with the FlexiVent, which were used to derive pulmonary function parameters. Mice were removed from the ventilator, euthanized, and tissues were harvested.

[0069] RNA quantification: The right middle lobe was snap frozen in liquid nitrogen at the time of euthanasia and stored at -80°C until RNA purification. RNA purification was performed using Qiazol extraction, followed by disruption of the tissue in a bullet blender and purification using the RNAeasy kit (Qiagen) according to the manufacturer's protocol. Reverse transcription to cDNA was completed using iScript Supermix (Bio-Rad). Real-time PCR was performed using 1 ng of sample cDNA and pre-validated rabbit primers targeting COL1A1, COL3A1, fibronectin, Tgm2 and GADPH (housekeeping genes).

[0070] result: Lung collagen expression (Figures 4 and 5): Second harmonic generation assessment of lung collagen (Figure 4) confirmed previous cytochemical staining assessments that TG2 KO mice showed a reduced increase in collagen levels compared to wild-type C56BL / 6 mice after bleomycin treatment (Olson et al., 2011). However, TG2 deficiency resulted in a clear and significant reduction in interstitial collagen gene expression that was greater than would be realistically expected given our current understanding of the mechanism of action of TG2 in fibrosis (Figure 5).

[0071] Pulmonary function after bleomycin treatment (Figure 6): Prior to tissue collection, lung function of mice was measured using a FlexiVent device. Resistance of the respiratory system to forced ventilation was measured in two stages. The first, Rrs or total resistance, is the resistance of the entire respiratory system to forced ventilation. Rrs increased significantly in C57BL / 6 mice in response to bleomycin, whereas the slight increase in TG2 KO mice was not significant and was significantly lower than Rrs in C57BL / 6 mice (Figure 6A). The second, Rn or Newtonian resistance, estimates how much of the total system resistance is due to changes in airway resistance (e.g., airway narrowing in asthma). In both wild-type and TG2 KO mice, Newtonian resistance was unaffected by bleomycin (Figure 6A). Assuming that the chest wall was also unaffected by bleomycin, the apparent difference in Rrs between wild-type and KO mice must be due to changes in the resistance of the lung parenchyma (i.e., interstitial lung disease).

[0072] Compliance represents the ease with which the respiratory system expands. Fibrosis generally reduces compliance because the lungs become stiffer, scarred, and less able to expand. Dynamic compliance is measured during tidal breathing. Static compliance is measured in human patients during a deep breath-hold maneuver. The Flexivent device measures quasi-static compliance during a single deep inflation by the ventilator (because mice cannot hold their breath on command). Bleomycin caused a significant loss of dynamic and quasi-static compliance in C57BL / 6 mice (Figure 6B). TG2 KO mice also had reduced compliance, but this was preserved compared to C57BL / 6, losing only about half the compliance of C57BL / 6 mice.

[0073] PV loop area reflects the ability to recruit alveoli to breath by a forced ventilation maneuver, with smaller PV loop areas indicating less recruitable alveolar volume. K measures the curvature of the contractile edge of the PV loop. A decrease in K suggests a faster initial contraction after breathing, which is consistent with increased elastic stiffness (the lungs do not stretch as much on inflation, so they collapse faster on contraction). In C57BL / 6 mice, both the K PV curve and PV loop area were significantly reduced in response to bleomycin (Figure 6C). In contrast, TG2 KO mice were protected from this decline, with neither showing a significant decrease.

[0074] Elastance measures the elastic stiffness of lung tissue and is the inverse of compliance. Ers is the elastance of the entire respiratory system, including contributions from tissue, chest wall, and airways, and H is the elastance of tissue only, calculated from two different mathematical models of lung tissue. Compared to saline-treated mice, bleomycin significantly increased the elastic stiffness of lung tissue in C57BL / 6 mice, but not in TG2 KO mice (Figure 6E). This change in total system elastance (Ers), as well as findings related to tissue elastance (H), are attributable to alterations in lung tissue. Taken together, TG2 KO mice were functionally protected in the bleomycin interstitial lung disease model in all parameters used to assess respiratory function. Compared to wild-type mice, there was no increase in total resistance, no increase in elastic stiffness, no loss of pressure-volume loops, and the loss of compliance was significantly reduced.

[0075] Conclusions of Example 2: The finding that TG2 KO mice were protected from loss of lung function following bleomycin-induced interstitial pulmonary fibrosis was particularly surprising. C57BL / 6 mice showed decreased compliance, increased elastic stiffness, increased tissue resistance, and decreased airspace recruitment upon bleomycin administration, all consistent with human IPF and other fibrotic ILDs. On the contrary, TG2 KO mice were protected from these effects, with approximately 50% reduced compliance loss and increased elastance, while airway recruitment and resistance were unchanged. These surprising and exciting results indicate that TG2 deficiency not only affects tissue histology, but also, most importantly, lung function.

[0076] Example 3 - In vitro inhibitory activity of anti-TG2 antibodies: method: Scratch wound experiment: Primary lung fibroblasts were cultured in calcium-supplemented ATCC RPE medium for 7 days. Some cells were left untreated, while others were treated with 1 ng / ml TGFβ, as indicated. Monolayers were scratched and 250 μM of the TG2 substrate 5-BP was added immediately, 1 h later, or 3 h later. Cells were then harvested, washed, and fixed in methanol 1 h after 5-BP addition (whole cell mounts). Fibronectin was detected by antibody staining, and TG2 activity was detected with HRP-streptavidin to detect the 5-BP label. The intensity of the fluorescent signal was quantified at the scratch border and in the cell layer distant from the scratch. With the short incubation times used, 5-BP only detects extracellular or cell surface TG2 activity, but not intracellular activity.

[0077] result: Quantification and inhibition of TG2 activity in scratch assay (Figure 7): Scratch assessments were performed in both native and TGFβ1-treated conditions using eight cell primary cell lines (three nonfibrotic and five IPF lines) derived from different human donors. TG2 activity was rapidly elevated after scratching, as indicated by incorporation of the transglutaminase substrate 5-biotinylated pentylamine (5BP) into the extracellular matrix visualized by raw fluorescence intensity (Figure 7A, left panel). However, timed substrate administration and cessation of the experiment 1 h after 5BP substrate administration showed that this activation was transient, with all activity disappearing within 1 h of scratching, and no activity was recorded when 5BP was administered 1 h after scratching (Figure 7A, middle, right panel). As expected, IPF cell lines expressed more fibronectin than nonfibrotic lines. Both fibrotic and nonfibrotic lines showed increased TG2 activity after scratching, with IPF lines being significantly more active than nonfibrotic lines. TG2 activation was enhanced by TGFβ and was greater in IPF fibroblasts than in nonfibrotic fibroblasts (Figure 7B), an important finding that has implications for the role of TG2 in IPF. Administration of a TG2-blocking antibody (rbBB7) completely blocked the increase in TG activity along the scratch, confirming that the increase in activity was solely due to TG2 and that TG2 is elevated extracellularly in response to injury.

[0078] Conclusions of Example 3: Primary human lung fibroblasts in culture export TG2 out of the cell but are inactive unless activated by scratch injury. Activation is transient, lasting less than an hour. Fibroblasts from IPF donors export more TG2 activity than nonfibrotic fibroblasts. This activity is completely inhibited by rbBB7. This data provides evidence that activated TG2 in IPF can be inhibited with rbBB7 and anti-TG2 antibodies such as Zampilimab. This data constitutes the antifibrotic response of Zampilimab in primary human lung cells.

[0079] Example 4 - In vivo inhibitory activity of anti-TG2 antibodies: method: Intervention studies with rbBB7, vehicle or control antibody treatment: The overall study design (Figure 8) involved 45 animals (rabbits) receiving 10 mg of silica prophylactically and therapeutically for 56 days. Five rabbits were injected with saline and harvested on day 56 (healthy group). Ten rabbits were treated with silica and harvested on the 28th day (morbidity level at the start of treatment). Ten rabbits were dosed with silica, treatment with rbBB7 (100 mg / kg) was initiated on day 28, and harvested on day 56 (treatment dose). Ten rabbits were treated with rbBB7 (100 mg / kg) starting one day before silica injection (i.e., treated on day -1) and harvested on day 56 (prophylaxis). · Five rabbits were administered silica and treatment with rb922 control antibody (100 mg / kg) was started one day prior to silica injection and harvested on day 56 (control antibody). Five rabbits were administered silica, vehicle treatment was started one day before silica injection, and harvested on day 56 (vehicle control).

[0080] This design allows antibody controls and vehicle controls to be combined to form an untreated group of 10 animals if there are no differences between groups. Rabbits were injected subcutaneously in the neck every 5 days with rbBB7, rb922 control antibody or vehicle. Rabbits were weighed prior to injection and antibody injection volume was adjusted accordingly to obtain a dose of 100 mg / kg. Rabbits receiving vehicle control received the same volume of vehicle as those receiving antibody. Rabbits began with an average weight of 2.5 kg and after 56 days averaged 3.2 kg.

[0081] Pharmacokinetics: Blood was collected from rabbits treated with rbBB7 every 5 or 10 days. The first blood draw was 5 days after the first rbBB7 injection. To minimize risk to the animals, a maximum of four samples were collected from each rabbit. Blood draws were always performed immediately prior to the next scheduled injection of rbBB7 to calculate trough (minimum) exposure values. Blood draws were always performed immediately prior to the next scheduled injection of rbBB7 to calculate trough (minimum) exposure values. Prophylactic dosing (from day 1, plots A and B) and therapeutic dosing (from day 26 to day 56, plots C and D) were performed on two separate occasions and therefore plotted separately. Blood was allowed to clot at room temperature for 45 minutes and then centrifuged at 300 × g for 15 minutes. Serum was decanted and stored frozen at -80 °C. rbBB7 was quantified in serum using mass spectrometry.

[0082] Tissue Processing: Lungs were inflated with 30% sucrose in PBS through the trachea as a cryoprotectant to provide resistance to the cutting blade and prevent easy tearing or collapse. The right lower lobe was placed in a custom block and divided into 5-6 sections, each approximately 4 mm thick. The central piece was placed in 10% formalin for 24 hours, then transferred to 70% ethanol and processed for histology. Left and right (mid and distal) slices were frozen at -80°C for later analysis.

[0083] Histology and analysis: Formalin-fixed lung slices were processed as a single piece and embedded in one large paraffin block. Sections were cut, stained with H&E, Trichrome, and Picrosirius red (PSR), and imaged on an Olympus or Zeiss slide scanner. For collagen quantification, PSR whole-slide images were processed for multiphase analysis using Definiens Tissue studio software. This software implemented an algorithm to distinguish between "lesion" and non-lesion / unaffected parenchyma regions based on cellular density and disappearance of unstained vacuoles. Collagen (bright red stained fibers) in lesional and non-lesional regions was then quantified as a percentage of the total area. Analysis of the same paraffin sections was repeated using multiphoton microscopy / second harmonic generation (SHG) and collagen quantification using a Genesis 200 scanner. SHG microscopy is a type of two-photon (2P) microscope that can detect fibrillar collagen without exogenous labels. Fibrogenic collagens include collagen types 1-3, 5, 11, 24 and 27. Some of these fibrous collagens, such as types I, III and V, are major players in pulmonary fibrosis (Kottmann et al. 2015).

[0084] Immunohistochemistry for TG2 antigen and activity: TG2 antigen was detected by immunostaining of frozen slices of lung tissue (swollen in 30% sucrose at the time of collection and then snap frozen) according to standard protocols. Transaminase (cross-linking) activity was measured in frozen slices of lung tissue using the transglutaminase in situ activity (TG ISA) assay using the incorporation of biotinylated cadaverine as previously described. The use of sucrose to swell the lungs interfered with this assessment, and the extra washing cycles required to remove sucrose considerably reduced the measured TG activity. Although the absolute TG2 activity was lower than in a similar experiment without sucrose, the relative differences between groups should be the same since all tissues were treated identically.

[0085] result: Intervention trial using RbBB7: Forty-three of the 45 rabbits successfully completed the experimental regimen (two rabbits died during the first silica injection, likely due to the effects of anesthesia). No adverse effects were observed in rabbits treated with rbBB7; they gained weight to the same extent as control rabbits (data not shown) and did not experience any of the physical problems associated with repeated injections of therapeutic antibodies.

[0086] Pharmacokinetics (Figure 9): Five days after the first rbBB7 injection (day 4 after silica administration, first sample for trough PK), serum levels averaged 395 μg / ml. By the next sampling point (day 19), rbBB7 levels were in the region of 700 μg / ml and maintained a steady state of 600-700 μg / ml on average (measured 5 days after the previous injection at trough). There was no significant decline in antibody levels over the 56 days, which would suggest the development of an allergic and anti-antibody response. These levels are consistent with the therapeutic efficacy in the rabbit model. Sampling was not performed after all treatments to reduce stress to the animals.

[0087] Histological analysis of fibrosis (Figure 10): Rabbit lung sections stained with H&E or Picrosirius red show the presence of fibrosis in silica-treated rabbits (photos not shown). Definiens image analysis was performed on PSR-stained sections to quantify the extent of fibrosis. This software quantified collagen (red) staining not only in the lesional areas but also in the entire lung lobe (i.e., normal parenchymal tissue or nonlesional areas plus lesional areas) (photos not shown). Looking at staining in the entire lobe, there was no clear change in PSR staining until day 28, but there was an average increase of 35% between days 28 and 56 (Figure 10A). rbBB7 showed a significant therapeutic effect in reducing collagen content. Looking at staining in the entire lobe, the increase in collagen was reduced by 50% in both the protective and therapeutic treatment groups, but this only became significant with the protective regimen. Evaluation of collagen PSR staining in the lesion alone also showed a notable decrease in the 92% increase in collagen between days 28 and 56, with a significant decrease of 58% and 48% in the treatment and protective groups, respectively (Figure 10B).

[0088] Analysis of collagen content by second harmonic generation (SHG) microscopy (Figure 11): SHG was used to detect total fibrillar collagen in silica-treated lobes. Rabbit lung tissue sections were scanned and collagen quantified. SHG did not detect an increase in collagen 28 days after silica injection, which may reflect that collagen was not yet mature enough to be detected by SHG. However, the level of detectable fibrillar collagen more than doubled between days 28 and 56 after silica injection. Treatment with rbBB7 significantly reduced collagen accumulation in both early and late treatment groups. Both treatment groups showed a 52% reduction in the mean SHG-detectable collagen levels from baseline, but only the protective treatment group reached significance.

[0089] Changes in TG2 activity (Figure 12): Frozen sections were prepared from one of the frozen lung sections and TG2 antigen and activity were measured. TG2 antigen levels were not altered by silica infusion and were unaffected by rbBB7 treatment (Figure 12A). Despite the use of sucrose in the lung inflation fluid, which significantly impairs TG ISA assessment, TG activity was significantly increased by 2.3-fold by silica treatment at both 28 and 56 days after silica infusion. RbBB7 reduced TG2 activity in both silica groups (Figure 12B). The 45% reduction in the therapeutic group was not significant (likely due to high variability), but the 85% reduction in the protective group was significant and comparable to normal lung levels.

[0090] Conclusions of Example 4: The surprising findings of this example are that 1) both early prophylactic treatment (treatment administered every 5 days starting the day before silica injection) and late treatment (treatment administered every 5 days starting 28 days after silica injection) were effective in suppressing histological fibrosis, and 2) as an antibody inhibitor that can only target extracellular TG2, it was able to protect animals from fibrosis as well as total TG2 knockout (in a mouse model), indicating that lung fibrosis is mainly due to extracellular TG2 activity. The reduction in fibrosis was confirmed independently by two methods of analyzing collagen in lung tissue (PSR staining and SHG microscopy). Anti-TG2 antibodies also blocked TG2 enzyme activity detected in frozen lung sections. In other words, anti-TG2 antibodies (which inhibit the extracellular protein cross-linking activity of TG2) significantly suppressed lung fibrosis when administered prophylactically to rabbits in a silicosis model, and could block further progression of fibrosis when administered from 28 days after the onset of lung remodeling.

[0091] Example 5 - TG2 is upregulated in COVID-19 infection method: Human tissue: Lung tissue samples were obtained from Tissue Solutions Ltd. Samples from 22 patients who died from COVID-19 infection were analyzed, as well as six tissue samples from normal lungs and IPF patients. Patients with known lung disease, including chronic obstructive pulmonary disease (COPD), were excluded. Tissue sample preparation: Formalin-fixed, paraffin-embedded human samples were serially sectioned and stained to assess transglutaminase type 2 (TG2) mRNA expression. TG2 mRNA (RNAscope): A similar protocol to that detailed in Example 1 was used.

[0092] result: As shown in Figures 13 and 14, TG2 mRNA expression was significantly elevated in postmortem tissue samples obtained from patients who died of COVID-19 compared to expression in normal lung tissue (P<0.05). IPF samples are included as a reference only. This high expression was accompanied by evidence of extensive matrix deposition (picrosil red staining - data not shown).

[0093] Conclusions of Example 5: During the inflammatory response and remodeling of lung tissue that occurs in severe SARS-cov2 virus infection, the inventors have shown that TG2 is highly expressed in most postmortem lung samples from patients who died of COVID-19 infection. It is expected that anti-TG2 antibodies, such as Zampilimab, can delay remodeling not only in the acute phase but also in the long-term chronic phase of COVID disease.

[0094] References: 1)Siegel et al.,2007,Pharmacol.Ther.,115(2):232-245 2)Wang et al.,2020,3 Biotech.,10:287 3) International Publication No. 2006 / 100679 4) International Publication No. 2012 / 146901 5) International Publication No. 2013 / 175229 6)Olsen et al., 2011, Am.J.Respir.Crit.Care Med.,184:699-707 7)Olsen 2020,Am.J.Respir.Crit.Care Med.,201:A1963 8) Philp et al., 2018, Am.J.Resp.Cell Mol.Biol., 58(5):594-603 9) Olsen et al., 2014, Am.J.Resp.Cell Mol.Biol., 50(4):737-747 10)Raghu,2017,Eur.Respir.Rev.2017;26:170071 11)Vaidya et al.,2017,Curr.Med.Chem.,24:1-20 12)Margaritopoulos et al.,2016,Core Evidence,11:11-22 13) International Publication No. 2015 / 197772 14)Kottmann et al.,2015,Resp.Res.,16:61

Claims

1. An anti-transglutaminase 2 (anti-TG2) antibody for use in treating a subject with progressive chronic interstitial lung disease or preventing the onset of progressive chronic interstitial lung disease.

2. 2. The anti-TG2 antibody for use according to claim 1, wherein the lung disease is selected from the group consisting of idiopathic pulmonary fibrosis (IPF), desquamative interstitial pneumonia (DIP), acute interstitial pneumonia (AIP; otherwise known as Hamman-Rich syndrome), hypersensitivity pneumonitis (HSP), nonspecific interstitial pneumonia (NSIP), respiratory bronchiolitis-associated interstitial lung disease (RBILD), cryptogenic organizing pneumonia (COP; otherwise known as Bronchitis Obliterans Organizing Pneumonia or BOOP), sarcoidosis, asbestosis, and lymphocytic interstitial pneumonia (LIP).

3. An anti-transglutaminase 2 (anti-TG2) antibody for use in treating a subject with pulmonary fibrosis associated with COVID infection or for preventing the development of pulmonary fibrosis in a subject suffering from COVID infection.

4. The anti-TG2 antibody for use according to claim 1 or 3, wherein the lung disease or pulmonary fibrosis is characterized by an increase in a marker in the subject's sample, the marker being selected from the group consisting of TG2 activity, mRNA encoding TG2, TG2 antigen, enhanced transport of TG2, or any combination thereof.

5. 4. The anti-TG2 antibody for use according to claim 1 or 3, wherein the antibody binds to an epitope within the core region of transglutaminase type 2 (TG2) and inhibits TG2 activity, the core region consisting of amino acids 143 to 473 of TG2, and the TG2 activity inhibited is TG2 cross-linking with lysine and glutamine via an N-ε(γ-glutamyl)lysine isopeptide bond.

6. the antibody or antigen-binding fragment thereof a. comprising or consisting of an intact antibody; b. comprising or consisting of an antigen-binding fragment; An anti-TG2 antibody for use according to claim 1 or 3.

7. 4. The anti-TG2 antibody for use according to claim 1 or 3, wherein the antibody comprises the following sequence: (i) KASQDINSYLT (LCDR1; SEQ ID NO. 1); LVNRLVD (LCDR2; SEQ ID NO. 2); LQYDDFPYT (LCDR3; SEQ ID NO. 3); THAMS (HCDR1; SEQ ID NO. 4); TISSGGRSTYYPDSVKG (HCDR2; SEQ ID NO. 5); and LISTY (HCDR3; SEQ ID NO. 6); or (ii) KASQDINSYLT (LCDR1; SEQ ID NO. 1); LTNRLMD (LCDR2; SEQ ID NO. 7); LQYVDFPYT (LCDR3; SEQ ID NO. 8); SSAMS (HCDR1; SEQ ID NO. 9); TISSGGRSTYYPDSVKG (HCDR2; SEQ ID NO. 5); and LISPY (HCDR3; SEQ ID NO. 10); or (iii) KASQDINSYLT (LCDR1; SEQ ID NO. 1); RTNRLFD (LCDR2; SEQ ID NO. 11); LQYDDFPYT (LCDR3; SEQ ID NO. 3); SSAMS (HCDR1); TISVGGGKTYYPDSVKG (HCDR2; SEQ ID NO. 9); and LISLY (HCDR3; SEQ ID NO. 12).

8. An anti-TG2 antibody for use according to claim 1 or 3, the antibody comprising: a) a light chain variable domain having a sequence as defined in any one of SEQ ID NOs: 13 to 27 and a heavy chain variable domain having a sequence as defined in any one of SEQ ID NOs: 28 to 40; b) a light chain variable domain having at least 80% identity or similarity, preferably at least 90% identity or similarity or preferably at least 95% identity or similarity, to a sequence set forth in any one of SEQ ID NOs: NO. 13 to NO. 27, and a heavy chain variable domain having at least 80% identity or similarity, preferably at least 90% identity or similarity or preferably at least 95% identity or similarity, to a sequence set forth in any one of SEQ ID NOs: NO. 28 to NO.

40.

9. 4. An anti-TG2 antibody for use according to claim 1 or 3, wherein the antibody competes with antibody X and / or antibody Y for binding to TG2: wherein said antibody X has the following sequence: (i) KASQDINSYLT (LCDR1; SEQ ID NO. 1); LVNRLVD (LCDR2; SEQ ID NO. 2); LQYDDFPYT (LCDR3; SEQ ID NO. 3); THAMS (HCDR1; SEQ ID NO. 4); TISSGRSTYPDSVKG (HCDR2; SEQ ID NO. 5); and LISTY (HCDR3; SEQ ID NO. 6); or (ii) KASQDINSYLT (LCDR1; SEQ ID NO. 1); LTNRLMD (LCDR2; SEQ ID NO. 7); LQYVDFPYT (LCDR3; SEQ ID NO. 8); SSAMS (HCDR1; SEQ ID NO. 9); TISSGGRSTYYPDSVKG (HCDR2; SEQ ID NO. 5); and LISPY (HCDR3; SEQ ID NO. 10); or (iii) KASQDINSYLT (LCDR1; SEQ ID NO. 1); RTNRLFD (LCDR2; SEQ ID NO. 11); LQYDDFPYT (LCDR3; SEQ ID NO. 3); SSAMS (HCDR1); TISVGGKTYPDSVKG (HCDR2; SEQ ID NO. 9); and LISLY (HCDR3; SEQ ID NO. 12), and wherein antibody Y consists of: a) a light chain variable domain having a sequence as defined in any one of SEQ ID NO: 13 to SEQ ID NO: 27, and a heavy chain variable domain having a sequence as defined in any one of SEQ ID NO: 28 to SEQ ID NO: 40; b) a light chain variable domain having at least 80% identity or similarity, preferably at least 90% identity or similarity, or preferably at least 95% identity or similarity, to a sequence defined in any one of SEQ ID NOs: NO. 13 to NO. 27, and a heavy chain variable domain having at least 80% identity or similarity, preferably at least 90% identity or similarity, or preferably at least 95% identity or similarity, to a sequence defined in any one of SEQ ID NOs: NO. 28 to NO.

40.

10. 1. A method for treating a subject having a progressive chronic interstitial lung disease or for preventing the development of a progressive chronic interstitial lung disease in a subject, the method comprising administering to the subject a therapeutically effective amount of an anti-TG2 antibody.

11. 1. A method for treating a subject having pulmonary fibrosis associated with COVID infection or for preventing the development of pulmonary fibrosis in a subject suffering from COVID infection, said method comprising administering to said subject a therapeutically effective amount of an anti-TG2 antibody.

12. Use of an anti-transglutaminase 2 (TG2) antibody for the manufacture of a medicament for treating a subject with progressive chronic interstitial lung disease or for preventing the onset of progressive chronic interstitial lung disease.

13. Use of an anti-transglutaminase 2 (TG2) antibody for the manufacture of a medicament for treating a subject with pulmonary fibrosis associated with COVID infection or for preventing the development of pulmonary fibrosis in a subject suffering from COVID infection.

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