Anti PD-l2 antibodies for use in treating fibrotic diseases

Abstract

The present invention relates to isolated monoclonal antibodies that specifically bind to programmed cell death ligand 2 (PD-L2) and block activity against PD-L2 / PD1 interaction for use in the treatment of a fibrotic disease in a mammal.

Description

[0001] Anti PD-L2 antibodies for use in treating fibrotic diseases

[0002] Fibrosis is a pathological condition referring to the excessive accumulation of fibrous connective tissue in an organ or tissue, which can disrupt the normal structure and function of the affected organ or tissue. Typically, fibrosis occurs as a response to chronic inflammation, injury or damage.

[0003] There are several types of fibrosis, each affecting different organs and tissues in the body. For example, pulmonary fibrosis (or "lung f i brosis") involves the excessive buildup of scar tissue in the lung tissue, which can make it difficult for the lungs to function properly, leading to symptoms such as shortness of breath, coughing, and reduced lung capacity. Another type of fibrosis is liver fibrosis, which occurs when there is an excessive accumulation of fibrous connective tissue caused by chronic inflammation, often as a result of conditions like chronic viral hepatitis, alcohol abuse, or non-alcoholic fatty liver disease. If left untreated, liver fibrosis can progress to cirrhosis, which is a severe and irreversible condition. Other types of fibrosis include cardiac (or heart fibrosis), renal (or kidney) fibrosis and gastrointestinal fibrosis, which can affect various parts of the digestive tract, such as the stomach and intestines.

[0004] The aetiology and pathogenesis of fibrosis are very complex and still not completely understood. In the past, the development of an effective treatment has been very challenging due to the complex mechanisms underlying the onset and the development of fibrosis.

[0005] Depending on the type of fibrosis, its severity and cause, different treatments are available. For example, anti-fibrotic treatments under investigation involve medications that specifically target the biochemical pathways involved in fibrosis, such as drugs inhibiting TGF-0 (Transforming Growth Factor-beta) signalling, which plays a key role in fibrosis. Other treatments involve the administration of inflammation-reducing drugs, such as corticosteroids, nonsteroidal anti-inflammatory drugs (NSAIDs), and immunosuppressants, to decelerate fibrosis.

[0006] In the specific case of Idiopathic Pulmonary Fibrosis (IPF), examples of standard therapies include medication with pirfenidone (Esbriet™), which functions by downregulating the productions of growth factors and procollagens I and II and which is effective at slowing down, but not halting, disease progression. Adverse side effects frequently associated with pirfenidone include gastrointestinal effects, such as dyspepsia, nausia, gastritis, gastroesophageal reflux disease and vomiting. Additionally, pirfenidone has been known to elevate hepatic enzyme levels. Some patients have also reported skin-related side effects, including reactions like photosensitivity, rashes, pruritus, and dry skin.

[0007] Other therapies for the treatment of IPF includes medication using nintedanib (sold under the brand names Ofev™ and Vargatef™), which is also used for treating systemic sclerosis associated interstitial lung disease (SSc-ILD, also called scleroderma-associated interstitial lung disease), and chronic interstitial lung diseases (ILDs) in which fibrosis continues to progress. The use of nintedanib in IPF is predicated on its inhibition of PDGFR, FGFR, and VEGFR, which increase fibroblast proliferation, migration, and transformation.

[0008] Apart from the standard therapies using pirfenidone and nintedanib, anti body- based therapies targeting specific molecules or pathways involved in fibrosis are being investigated. For example, antibodies that block the activity of TGF-p have been suggested as potential antifibrotic agents (see J. Varga et a!., "Anti-TGF-0 Therapy in Fibrosis: Recent Progress and Implications for Systemic Sclerosis", Curr. Opin. Rheumatol., 2008, 20(6), 720-728). The anti-PD-Ll antibody Atezolizumab is currently being tested for preliminary efficacy in IPF in a Phase I clinical trial (NCT05515627).

[0009] An alternative approach is suggested in W02022 / 200463A1, which relates to an antibody that binds Axl and anatagonizes Axl activity for use in a method of treating a fibrotic disease, in particular to a fibrotic disorder selected from the group consisting of: arthrofibrosis, Crohn's disease, Dupuytren's contracture, peyronie's disease, strabmisus, scleroderma, keloid, Nephrogenic systemic fibrosis, pulmonary fibrosis, idiopathic pulmonary fibrosis (IPF), cystic fibrosis (CF), progressive massive fibrosis, fibrothorax, systemic sclerosis, cardiac fibrosis, non-alcoholic steatohepatitis (NASH), other types of liver fibrosis, primary biliary cirrhosis, renal fibrosis, reteroperitoneal fibrosis, mediastinal fibrosis, myelofibrosis, and atherosclerosis.

[0010] There is an ongoing need for an efficient treatment of fibrosis in general, and specifically, in IPF, in particular in view of the adverse side effects of a therapy using pirfenidone and its inability to halt disease progression.

[0011] Summary

[0012] In consideration of the above, the present disclosure deals with the object of providing an agent for efficiently treating, ameliorating or preventing (or reducing the risk of) a fibrotic disease.

[0013] The inventors have shown in the Examples that antibodies which specifically bind to programmed cell death ligand 2 (PD-L2) and block activity against PD-L2 / PD-1 (e.g. human PD-L2 and / or PD-1) interaction can be used in in vivo models to slow and / or reverse fibrosis and / or markers of fibrotic diseases.

[0014] Brief Description of the Figures

[0015] Figure 1 is a schematic representation showing the intervention points of the anti-PD-L2 antibodies described herein in the mechanisms underlying the pathogenesis of fibrosis. As shown, anti-PD-L2 antibodies mediate pleiotropic effects by mediating anti-fibrotic effects including reducing senescent cell burden as a first intervention point and, as a second intervention point, reprogramming pro-fibrotic M2 macrophages that promote the fibroblast to myofibroblast transition, which is a direct cause of fibrosis. As also shown, anti-PD-L2 antibody directly inhibits the transition of fibroblasts to myofibroblasts, which relates to a third intervention point.

[0016] Figure 2 shows VH6 / VK3 on a hlgGl wild-type (WT) constant region and effector-reduced hlgGl LALA constant region (in the following referred to as VH6 / VK3 WT and VH6 / VK3 LALA) as well as parental VH0 / VK0 on the hlgGl WT and hlgGl LALA constant regions have similar blocking activity against the cellular hPD-L2 / hPD-l interaction in a bioassay that reports blocking activity in relative light units (RLUs) from a fluorescent reporter in hPD-1 expressing Jurkat cells co-cultured with hPD- L2 expressing CHOK-1 cells.

[0017] Figure 3 shows that the half-life of VHO / VKO-hlgGl WT has an average half-life of 11.88 days, while VHO / VKO-hlgGl LALA has significantly longer average half-life of 13.86 days in the humanized FcRN mouse model. N=6 for each group. An unpaired t-test was applied to determine significance.

[0018] Figure 4. Analysis of PD-L2 expression cell types that drive fibrosis

[0019] Figure 4A: Percentage (left) or mean expression (right) of cells expressing PD-L2 according to bioinformatic analysis of publicly available data set on IPF patients and healthy controls: This plot displays the percentage of cells expressing PD-L2 for five distinct cell types: macrophage, fibroblast, myofibroblast and aberrant basaloid, for each cell type across two conditions: control and IPF (Idiopathic Pulmonary Fibrosis). Each point represents an individual subject (Macrophage: 21 control, 28 IPF; Fibroblast: 12 control, 15 IPF; Myofibroblast: 6 control, 8 IPF; Aberrant basaloid: 10 control, 11 IPF; Senescent: 11 control, 15 IPF). Statistical Significance using wilcoxon test represented as follow: "ns": Not significant (p > 0.1); "*": Significant (p < 0.1); "**": Very significant (p < 0.05); "***": Highly significant (p < 0.01).

[0020] Figure 4B: Analysis by flow cytometry to quantify the MFI (mean fluorescence intensity) respect to the isotype control signal. Lung fibroblasts (NHLF) and IPFs from 2 donors were treated with 5 ng / mL TGF-p for 72h for their conversion into myofibroblasts, and the levels of PD-L2 were analysed in both populations by FACS. PD-L2 is higher in IPF fibroblasts and myofibroblasts compared to normal fibroblasts and TGF-0 also increases PD-L2 expression in both normal and IPF fibroblasts.

[0021] Figure 4C: Flow cytometry data showing % positive PD-L2 expressing M0, Ml or M2 macrophages that were generated as described in Example 5.

[0022] Figure 5A. Flow Cytometry Analysis of Macrophage Reprogramming from M2 to Ml Phenotype upon treatment with VH6 / VK3 WT and LALA. Left panel: Flow cytometry analysis of CD86 expression in macrophages. Untreated M2 macrophages (NT) exhibit low CD86 expression. Treatment with VH6 / VK3 WT and LALA both result in a significant increase in CD86 expression, indicating reprogramming towards an Ml-like phenotype, which is not seen with the equivalent isotype controls. The differences between VH6 / VK3 WT and LALA treatments are not statistically significant. Right Panel: Flow cytometry analysis of CD206 expression in macrophages. Untreated M2 macrophages (NT) show high CD206 expression, consistent with their pro-fibrotic phenotype. Treatment with VH6 / VK3 WT and LALA leads to a statistically significant decrease in CD206 expression, reflecting the shift towards an antifibrotic Ml-like state. This decrease is not seen with the equivalent isotype controls. Treatment of M2 macrophages with the CSF1R inhibitor PLX3997, which is known to induce reprogramming of M2 macrophages to an Ml-like phenotype was used as a positive control and shows an increase in CD86 expression that is lesser than VH6 / VK3 WT and LALA and a decrease in CD206 that is less than VH6 / VK3 WT but greater than VH6 / VK3 LALA.

[0023] Figure 5B. ELISA analysis of cytokine levels in culture supernatants of M2 macrophages treated with VH6 / VK3 WT or LALA. Left panel: A statistically significant reduction in TGF-0 (a cytokine heavily involved in fibrotic processes and tissue scarring) is observed after treatment with both VH6 / VK3 WT and LALA, indicating a decrease in pro-fibrotic activity. Such a reduction is not seen with the equivalent isotype controls. There is no significant difference in TGF- P reduction between the two antibodies. Right panel: A marked increase in TNF-a (a cytokine associated with the resolution of fibrosis) is detected following treatment with both antibodies, suggesting a shift toward an anti-fibrotic, Ml macrophage -like state. This increase is not seen with the equivalent isotype controls. There is no significant difference in TNF-a induction are between the two anti-PD-L2 antibodies.

[0024] Figure 5C. Microscopic Analysis of Macrophage Reprogramming from M2 to Ml Phenotype upon treatment with VH6 / VK3 WT or LALA. Left panel: Anti-PD-L2 antibodies on either wild-type or LALA hlgGl formats stimulated the expansion of reprogrammed macrophages relative to untreated or isotype controls. The increased stimulation of proliferation by anti-PD-L2 antibodies was greater than that of PLX3397 (CSF1R inhibitor, positive control).

[0025] Right panel: Untreated M2 macrophages exhibit an elongated, spindle-shaped morphology typical of the M2 phenotype (top left panel) Microscopy images suggest that both VH6 / VK3 WT and LALA reprogram M2 macrophages into an Ml-like phenotype, characterized by a shift to a more rounded morphology and an increase in proliferation (top right, middle left panels). No significant changes in morphology or proliferation are observed with the isotype controls (middle right, bottom left panels), indicating that the observed effects are specific to the antibodies.

[0026] Figure 6. Anti-fibrotic effect of anti-PD-L2 antibody VH6 / VK3 WT and LALA in myofibroblasts.

[0027] Figure 6A: Representative immunofluorescent images of a-smooth muscle actin 1 (smal) positive staining in IPF donor #27 cells. Smal positive fibers are shown along with cell nuclei (stained with DAPI). Scale bar is shown (50 mm).

[0028] Figure 6B-E: Quantification (intensity of signal) of smal positive fibers (by immunofluorescence) in: (B) normal human lung fibroblasts, (C) fibroblasts from IPF donor #29, (D) fibroblasts from IPF donor #37, and (E) liver stellate cells. Cells were untreated or treated with TGF- P in the presence or absence of all antibodies or isotype controls for three days, and the total area of smal positive signal was stained and quantified. Atezolizumab was used as a positive anti-fibrotic control. Results are representative of two independent experiments. Statistical analyses were performed using one-way ANOVA with post-hoc Sidak's test (B, D): **** p< 0.0001; *** <0.001; ** <0.01.; * p<0.05. Figure 7A shows VH0 / VK0-mIgG2a LALA and VH0 / VK0-mIgG2a WT reduces hydroxyproline content, a marker of fibrosis, compared to vehicle and isotype controls in mice treated with bleomycin. The reduction for VH0 / VK0-mIgG2a LALA is equivalent to that seen with pirfenidone and is greater than the reduction seen with VH0 / VK0-mIgG2a WT. Data indicates Mean ± SEM. ###p <0.01 Vs Normal control + PBS. f**p<0.01 ***p<0.001, Vs Bleomycin + PBS. One-way ANOVA followed by Sidak's multiple comparisons test, n = 6-12.

[0029] Figure 7B shows that VH0 / VK0-mIgG2a LALA and VH0 / VK0-mIgG2a WT reduces the Ashcroft score, compared to vehicle and isotype controls, in mice treated with bleomycin. The reduction for VH0 / VK0-mIgG2a LALA is equivalent to that seen with pirfenidone and greater than that seen with VH0 / VK0-mIgG2a WT. Data indicates Mean ± SEM. ###p <0.01 Vs Normal control + PBS. *p<0.05, ***p<0.001, Vs Bleomycin + PBS. One-way ANOVA followed by Bonferroni multiple comparisons test, n = 6-12.

[0030] Figure 7C shows representative H&E images of lungs analyzed to generate Ashcroft scores illustrating improvements in lung architecture and fibrosis most significantly in bleomycin-treated mice that are also treated with VH0 / VK0-mIgG2a LALA and pirfenidone and to a lesser extent in mice treated with VH0 / VK0-mIgG2a WT.

[0031] Figure 7D shows that VH0 / VK0-mIgG2a LALA reduces the picrosirius red staining, a marker of fibrosis, compared to vehicle and isotype controls, in mice treated with bleomycin. This reduction is equivalent to that seen with pirfenidone and greater than the reduction seen with VH0 / VK0-mIgG2a WT. Data indicates Mean ± SEM. ###p <0.05 Vs Normal control + PBS. ***p<0.05, Vs Bleomycin + PBS. One-way ANOVA followed by Sidak's multiple comparisons test, n = 6-12.

[0032] Figure 7E represents images of lungs stained with picrosirius red illustrating reductions in fibrosis in bleomycin-treated mice that are also treated with VH0 / VK0-mIgG2a LALA and pirfenidone and to a lesser extent in mice treated with VH0 / VK0-mIgG2a WT.

[0033] Figure 7F shows VH0 / VK0-mIgG2a LALA reduces the inflammation score as assessed by a pathologist, compared to vehicle and isotype controls, in mice treated with bleomycin. This reduction is equivalent to that seen with pirfenidone and not as pronounced with VH0 / VK0-mIgG2a WT. Data indicates Mean ± SEM. ###p <0.01 Vs Normal control + PBS. ***p<0.001, Vs Bleomycin + PBS. One-way ANOVA followed by Bonferroni multiple comparisons test, n = 6 -12.

[0034] Figure 8A shows that VH0 / VK0-mIgG2a LALA and VH0 / VK0-mIgG2a WT reduces pl6 staining, a marker of senescence, compared to vehicle and isotype controls, in mice treated with bleomycin. The reduction for VH0 / VK0-mIgG2a LALA is equivalent to that seen with pirfenidone and greater than that seen with VH0 / VK0-mIgG2a WT. Data indicates Mean ± SEM. ###p <0.001 Vs Normal control + PBS. *p<0.05, ***p<0.001, Vs Bleomycin + PBS. One-way ANOVA followed by Bonferroni multiple comparisons test, n = 6-12. Figure 8B shows representative images of lung sections stained with pl6 illustrating reductions in senescent cells burden in bleomycin-treated mice that are also treated with VHO / VKO- m!gG2a LALA and pirfenidone and to a lesser extent in mice treated with VH0 / VK0-mIgG2a WT.

[0035] Figure 8C shows that VH0 / VK0-mIgG2a LALA and VH0 / VK0-mIgG2a WT reduces p21 staining, a marker of senescence, compared to vehicle and isotype controls, in mice treated with bleomycin. The reduction for VH0 / VK0-mIgG2a LALA is equivalent to that seen with pirfenidone and greater than that seen with VH0 / VK0-mIgG2a WT. Data indicates Mean ± SEM. ###p <0.001 Vs Normal control + PBS. ***p<0.001, Vs Bleomycin + PBS. One-way ANOVA followed by Bonferroni multiple comparisons test, n = 6-12.

[0036] Figure 8D shows representative images of lung sections stained with p21, illustrating reductions in senescent cells burden in bleomycin-treated mice that are also treated with VH0 / VK0- m!gG2a LALA and pirfenidone and less significantly in mice treated with VH0 / VK0-mIgG2a WT.

[0037] Figure 9A shows that treatment of the Zmpste24 knockout mouse progeria model with an anti-PD-L2 antibody significantly extends median lifespan by 13% compared to treatment with isotype control. p=0.041 according to log rank Mantel-Cox test. n= 17 for IgG control group and n=15 for anti-PD-L2 group with equivalent sex distributions in each group.

[0038] Figure 9B shows that treatment of females in the Zmpste24 knockout mouse progeria model with an anti-PD-L2 antibody significantly extends median lifespan by 30% compared to treatment with isotype control. p=0.028 according to Gehan-Breslow Wilcoxon test. n= 9 for IgG control group and n=8 for anti-PD-L2 group.

[0039] Figure 10A shows that all Zmpste24 knockout mice treated with anti-PD-L2 antibody have serum creatinine levels, a measure of kidney function, below 0.2 mg / dL, while those treated with IgG control antibody have creatinine levels above detection. N=5 for IgG-treated group and n=5 for anti- PD-L2-treated group.

[0040] Figure 10B shows that Zmpste24 knockout mice treated with anti-PD-L2 antibody have increased glomerular size, an indication of improved renal function, compared to IgG-treated animals. N=3 for both treatment groups

[0041] Figure IOC shows that Zmpste24 knockout mice treated with anti-PD-L2 antibody have decreased expression of various collagen genes colla2, co / 2al, co / 6al and co / 6a2), which are markers of fibrosis, compared to IgG-treated animals. n=9 for the IgG treatment group and n=7 for the anti-PD-L2 treatment group.

[0042] Figure 11A shows that Zmpste24 knockout mice treated with anti-PD-L2 antibody do not have decreased absolute thickness of the inner layers of the inner aorta wall (tunica intima and media) but have decreased absolute thickness of the outer collagen-rich tunica adventitious, which also is reflected as decreased percent thickness and surface area of the 3 layers together. An unpaired Student's t-test was applied to n=4 for each treatment group. Figure 11B is a representative image of Trichome-staining of aortas used to quantify the data in 11A showing a reduction in the collagen-rich tunica adventitious layer.

[0043] Definitions

[0044] The term "aberrant basaloid" refers to a rare type of epithelial cell which can be found in fibrotic lung tissue. They are the predominant senescent cell type in fibrotic lung tissue that drives fibrosis, as evidenced by the fact that deletion of their murine counterparts called "transitional aveolar epithelial cells" reduces fibrosis in the bleomycin mouse model of IPF (see Wang eta / ., "Regulation of epithelial transitional states in murine and human pulmonary fibrosis", J. Clin. Invest., 2023, 133(22), el65612, doi: 10.1172 / JCI165612; which is incorporated by reference herein in its entirety).

[0045] Within the context of the present disclosure, an "antibody" typically comprises at least two heavy (H) chains and two light (L) chains interconnected by disulphide bonds. Each heavy chain is comprised of a heavy chain variable region (abbreviated herein as VH) and a heavy chain constant region. The heavy chain constant region is comprised of three regions, CHI, CH2 and CH3. Each light chain is comprised of a light chain variable region (abbreviated as VL) and a light chain constant region. The light chain constant region is comprised of one region, CL. The VH and VL regions can be further subdivided into regions of hypervariability, termed Complementary Determining Regions (CDR), interspersed with regions that are more conserved, termed framework regions (FW). Each VH and VL is composed of three CDRs and four FWs, arranged from amino-terminus to carboxy-terminus in the following order: FWI, CDRI, FW2, CDR2, FW3, CDR3, FW4. The variable regions of the heavy and light chains contain a binding region that interacts with an antigen. The constant regions of the antibodies can mediate the binding of the immunoglobulin to host tissues and factors, including various cells of the immune system (e.g. effector cells) and the first component (Clq) of the classical complement system. The CDR regions of an Ig-derived region may be determined as described in Ka bat "Sequences of Proteins of Immunological Interest", 5th edit. NIH Publication no. 91-3242 U.S. Department of Health and Human Services (1991); Chothia, J. Mol. Biol., 196 (1987), 901-917 or Chothia, Nature, 1989, 342, 877-883.

[0046] The term "antigen-binding fragment" (of an antibody) refers to that portion of an antibody which comprises the amino acid residues that interact with an antigen and confer on the binding agent its specificity and affinity for the antigen (e.g. the CDRs). The antigen binding region can be derived from any animal species, such as rodents (e.g. rabbit, rat or hamster) and humans. Preferably, the antigen binding region will be of human origin. An antigen-binding fragment of an antibody encompasses antigen-binding fragments of antibodies (e.g. single chain antibodies, including VHHS, Fab and sFab fragments, F(ab')2, Fd fragments, Fv fragments, scFv, and domain antibodies (dAb) fragments (see, e.g. de Wildt eta / ., Eur J. Immunol., 1996; 26(3):629-39; which is incorporated by reference herein in its entirety)).

[0047] The term "chimeric antibody" means an antibody having light and heavy chain genes which have been constructed, typically by genetic engineering, from immunoglobulin variable and / or constant region genes belonging to more than one different species. For example, the variable regions from, e.g. a mouse antibody (e.g. a monoclonal or polyclonal antibody), may be used in conjunction with human constant regions to produce the chimeric antibody. A therapeutic chimeric antibody as described herein is thus a hybrid protein composed of the variable or antigen-binding region from a mouse antibody (e.g. one or more of the CDRs of a mouse antibody) and the constant or effector region from a human antibody, although other mammalian species may be used. The chimeric antibody can also include amino acid sequence obtained from a protein source other than an antibody.

[0048] The expression "competes for binding" or similar terminology as used herein refers to the competition for binding with both linear and conformational epitopes and steric binding competition, which can be measured by various techniques known to the skilled person. In an example, competition is determined by surface plasmon resonance (SPR), such techniques being readily apparent to the skilled person. SPR can be carried out using Biacore™, Proteon™ or another standard SPR technique. Such competition may be due, for example, to the antibodies binding to identical or overlapping epitopes of PD-L2. In a further example, competition is determined by enzyme-linked immunosorbent assays (ELISA), which is also well known and readily apparent to the skilled person. In further example, competition is determined by homogenous time resolved fluorescence (HTRF). In still a further example, competition is determined by fluorescence activated cell sorting (FACS) or by other cell-based assays, such techniques being readily apparent to the skilled person as well.

[0049] As used herein, "conservative" relates to an amino acid substitution. A substitution may be classified as "conservative" when an amino acid residue contained in a polypeptide is replaced with another naturally occurring amino acid of similar character either in relation to polarity, side chain functionality or size. Such conservative substitutions are well known in the art.

[0050] As used herein, a "decrease" or a "reduction" (and other similar terms) may mean a reduction (or decrease) of at least (about) 10%, at least (about) 20%, at least (about) 30%, at least (about) 40%, at least (about) 50%, at least (about) 60%, at least (about) 70%, at least (about) 80%, or at least (about) 90%. A reduction (or decrease) may be a statistically significant reduction (or decrease).

[0051] The term "epitope," as used herein, refers the portion or region of an antigenic molecule (e.g. a peptide), that is specifically bound by the antibody combining site of an antibody. An epitope typically includes at least 3, and more usually, at least 5 or 8 or 10 residues (e.g. amino acids or nucleotides). Typically, an epitope also is less than 20 residues (e.g. amino acids or nucleotides) in length, such as less than 15 residues or less than 12 residues. The term "epitope" encompasses both a linear epitope for which the consecutive amino acids are recognized by the antibody as well as a conformational epitope for which the antibodies recognize amino acids to the extent, they adopt a proper configuration or conformation. Consequently, in some epitopes, the conformation (three-dimensional structure) is as important as the amino acid sequence (primary structure).

[0052] As used herein, a "fibroblast" refers to a cell which creates and maintains a diverse array of extracellular matrix (ECM)-rich connective tissues to support a broad range of essential, normal organ functions, such as elastic recoiling during normal breathing in the lung. Fibroblasts have a characteristic flat, spindle shape, and are often characterised by the expression of vimentin and PDGFRa (see for example Lendahl et al., Nature Comms, 13(3409), 2022, doi: https: / / doi.org / 10.1038 / s41467-022-30633-9, which is incorporated herein by reference in its entirety).

[0053] As used herein, the term "fibrotic disease" relates to any kind of fibrosis, including pulmonary (or lung) fibrosis, liver fibrosis, in particular selected from non-alcoholic steatohepatitis (NASH) and non-alcoholic fatty liver disease, heart fibrosis, mediastinal fibrosis, in particular lymph node fibrosis, retroperitoneal cavity fibrosis, in particular kidney fibrosis and aorta fibrosis, bone marrow fibrosis, skin fibrosis, as well as scleroderma or systemic sclerosis. In some embodiments a fibrotic disease includes a condition associated therewith. In another embodiment, the fibrotic disease is a fibrotic disease of the eye. As would be generally understood by the skilled person, fibrosis generally refers to tissue which is scarred, or to the accumulation of excess extracellular matrix components in a tissue. The fibrosis may be sufficiently established to impair the normal function of the tissue.

[0054] The term "human antibody" means that the amino acid sequence of the antibody or antigenbinding fragment thereof is fully human, e.g. human heavy and light chain variable and optionally human constant regions.

[0055] The term "humanized antibody" means an antibody or antigen-binding fragment thereof comprising a human framework region and one or more CDRs from a non-human (usually a mouse or rat) immunoglobulin. The non-human immunoglobulin providing the CDRs is called the "donor" and the human immunoglobulin providing the framework is called the "acceptor". Constant regions need not be present, but if they are, they should be substantially identical to human immunoglobulin constant regions, i.e., at least about 85-90%, preferably about 95% or more identical. Hence, all parts of a humanized immunoglobulin, except possibly the CDRs, are substantially identical to corresponding parts of natural human immunoglobulin sequences. A humanized antibody (or fragment) is useful as an effective component in a therapeutic agent described herein since antigenicity of the humanized antibody (or fragment) in the human body is lowered as compared to the parental mouse antibody.

[0056] As used herein, an "increase" (and other similar terms) may mean an increase of at least (about) 10%, at least (about) 20%, at least (about) 30%, at least (about) 40%, at least (about) 50%, at least (about) 60%, at least (about) 70%, at least (about) 80%, or at least (about) 90%. An increase may be a statistically significant increase.

[0057] As used herein, the term "isolated antibody" is intended to refer to an antibody, or antigenbinding fragment thereof, which is substantially free of other antibodies having different antigenic specificities (e.g. an isolated antibody that specifically binds to PD-L2 and is substantially free of antibodies that do not bind to PD-L2). An isolated antibody that specifically binds to an epitope of human PD-L2 may, however, have cross-reactivity to other PD-L2 proteins from different species. However, in particular embodiments, the antibody maintains higher affinity and selectivity for human PD-L2. In addition, an isolated antibody is typically substantially free of other cellular material and / or chemicals.

[0058] As used herein, the term "isolated host cell" (or simply "host cell"), is intended to refer to a cell into which a recombinant expression vector has been introduced. It should be understood that such terms are intended to refer not only to the particular subject cell but to the progeny of such a cell. Because certain modifications may occur in succeeding generations due to either mutation or environmental influences, such progeny may not, in fact, be identical to the parent cell, but are still included within the scope of the term "host cell" as used herein.

[0059] As used herein, the term "KD" is intended to refer to the dissociation equilibrium constant of a particular antibody-antigen interaction, ka is the binding rate constant and kd is the dissociation rate constant. The binding affinity of antibodies disclosed herein may be measured or determined by standard antibody-antigen assays. In certain embodiments herein, the relative KD was calculated by dividing the KD of the humanized variant by that of the VHO / VKO assayed in the same experiment.

[0060] As used herein, a "macrophage" refers to a type of white blood cell which is part of the innate immune system, and which is involved in the detection, phagocytosis and destruction of bacteria and other harmful organisms. Macrophages may be separated from whole blood by the methods described herein (see Example 5B). Various sub-types of macrophages exist.

[0061] The least differentiated macrophage, in its unpolarised and resting state, is referred to as an "MO macrophage", which is characterised by the expression of CD14 and CD68. MO macrophages do not express either CD206 or CD86. MO macrophages differentiate into either Ml or M2 macrophages, depending on the signals they receive from their environment, and therefore are crucial as a precursor population. For a review of macrophages, see Lendeckel et a!., ChemTexts. 8(2), 12, 2022, doi: 10.1007 / s40828-022-00163-4, which is incorporated herein by reference in its entirety.

[0062] "Pro-inflammatory macrophages" and / or "anti-fibrotic macrophages" are often referred to as "Ml macrophages", and the terms are used interchangeably herein. Ml macrophages are are also known as classically activated macrophages, and are further characterised by the expression of CD86. Whilst generally associated with defense against pathogens, tumor resistance, and tissue damage, these Ml macrophages, in certain contexts, can exhibit anti-fibrotic properties. They may limit fibrosis by producing factors (such as cytokines like IFN-y and TNF-a), which can help suppress fibrotic processes by preventing the activation of fibroblasts, preventing collagen deposition and degrading extracellular matrix components. These mechanisms can resolve inflammation and prevent excessive tissue scarring.

[0063] "Anti-inflammatory macrophages" and / or "pro-fibrotic macrophages" are often referred to as "M2 macrophages", and the terms are used interchangeably herein. M2 macrophages are are also known as alternatively activated macrophages, and are characterised by the expression of CD206 (mannose receptor). Whilst generally associated with anti-inflammatory responses, tissue repair, and wound healing by resolving inflammation and restoring homeostasis after tissue injury, these M2 macrophages can promote fibrosis by producing various fibrotic mediators, such as TGF-0, which can stimulate the fibroblast-to-myofibroblast transition. They may contribute to fibrosis when their tissue repair function leads to excessive collagen deposition and scar formation in chronic conditions.

[0064] As used herein, the term "monoclonal antibody" or "mAb", refers to an antibody which displays a single binding specificity and affinity for a particular epitope.

[0065] As used herein, a "myofibroblast" refers to a cell which arises from a fibroblast during tissue injury and repair to support dynamic structural changes within the tissue. Myofibroblasts express a- smooth muscle actin (aSMA) that differentiates them from fibroblasts. aSMA contributes to contractile force in the tissue for biomechanical remodeling. Myofibroblasts secrete a greater volume of ECM components that have a unique composition compared to fibroblasts. Myofibroblasts also modulate the function of resident immune populations and can phagocytose dead cells. Acute myofibroblast activity supports healthy tissue repair, but in contrast chronic myofibroblast activity leads to fibrosis and organ stiffening, which impedes organ function. Myofibroblast abundance is associated with severity of fibrosis.

[0066] As used herein, the term "prevention" refers to the prevention or reduction of signs and symptoms associated with the respective disease in subjects who are at risk for developing the disease, in the specific case a fibrotic disease. In these subjects, a predisposing factor may be retained, but the signs and / or symptoms of the disease do not occur, or take significantly longer to develop. Further, the term also includes the prevention of a further deterioration of the symptoms once the disease has occurred. In that respect, prevention denotes a reduction in the likelihood (chance) of the development and / or progression of the signs and / or symptoms of the disease. Thus, prevention relates to prophylactically treating the disease in that manner (such that, in contrast, therapeutically treating the disease mainly applies after disease development).

[0067] As used herein, the term "senescent cells" refers to cells which are characterized by i) being damaged (e.g. exhibiting alterations in size, shape and / or cellular structure) and being resistant to apoptosis, ii) being durably arrested (meaning that they have stopped dividing a replicating), and iii) having a highly secretory phenotype (in that they secrete various bioactive molecules, including pro- fibrotic and proinflammatory cytokines, chemokines, growth factors, and matrix remodeling enzymes; also referred to as Senescence-Associated Secretory Phenotype or SASP). Markers for senescence in non-tumor cells include pl6 and p21 expression, as will be discussed in the context of Example 7 and Figures 8A and 8B and Figures 8C and 8D. Thus, in one embodiment, the senescent cells described herein express PD-L2 and pl6. In another embodiment, the senescent cells described herein express PD-L2 and p21. In another embodiment, the senescent cells described herein express PD-L2, pl6 and p21. In another embodiment, the senescent cells described herein may additionally express (or have up-regulation of expression of) p-galactosidase, e.g. lysosomal-associated p-galactosidase. In the context of fibrosis, senescent cells are thought to contribute to fibrosis by producing various fibrotic mediators, such as TGF-0, which can stimulate the fibroblast-to-myofibroblast transition. As used herein, "stellate cells" refer to resident liver mesenchymal cells that become activated in response to injury via factors such as TGF-0. Once activated, they secrete collagen and become a major contributor to fibrosis in the liver.

[0068] As used herein, the terms "treat," "treatment," "treating," or "amelioration" refer to therapeutic treatments, wherein the object is to reverse, alleviate, ameliorate, inhibit, slow down or stop the progression or severity of a condition associated with a disease or disorder. The term "treating" includes reducing or alleviating at least one adverse effect or symptom of a condition, disease or disorder. Treatment is generally "effective" if one or more symptoms or clinical markers are reduced. Alternatively, treatment is "effective" if the progression of a disease is reduced or halted. That is, "treatment" includes not just the improvement of symptoms or markers, but also a cessation of, or at least slowing of, progress or worsening of symptoms compared to what would be expected in the absence of treatment. Beneficial or desired clinical results include, but are not limited to, alleviation of one or more symptom(s), diminishment of extent of disease, stabilized (not worsening) state of disease, delay or slowing of disease progression, amelioration or palliation of the disease state, remission (whether partial or total), and / or decreased mortality, whether detectable or undetectable. The term "treatment" of a disease also includes providing relief from the symptoms or side-effects of the disease (including palliative treatment). For treatment to be effective, a complete cure is not contemplated. The method can, in certain aspects, include cure as well.

[0069] As used herein, the term "vector" is intended to refer to a nucleic acid molecule capable of transporting another nucleic acid to which it has been linked. One type of vector is a "plasmid", which refers to a circular double stranded DNA loop into which additional DNA segments may be ligated. Another type of vector is a viral vector, wherein additional DNA segments may be ligated into the viral genome. Certain vectors are capable of autonomous replication in a host cell into which they are introduced (e.g. bacterial vectors having a bacterial origin of replication and episomal mammalian vectors). Other vectors (e.g. non-episomal mammalian vectors) can be integrated into the genome of a host cell upon introduction into the host cell, and thereby are replicated along with the host genome. Moreover, certain vectors are capable of directing the expression of genes to which they are operatively linked. Such vectors are referred to herein as "recombinant expression vectors". In general, expression vectors of utility in recombinant DNA techniques are often in the form of plasmids. In the present specification, "plasmid" and "vector" may be used interchangeably as the plasmid is the most commonly used form of a vector. However, other forms of expression vectors, such as viral vectors (e.g., replication defective retroviruses, adenoviruses and adeno-associated viruses), my be used as described herein, which serve equivalent functions.

[0070] Detailed Description

[0071] Human PD-1 (hPD-1) has been identified as a member of the CD28 / CTLA-4 family of molecules based on its ability to bind to PD-L1. Like CTLA4, hPD-1 is rapidly induced on the surface of T-cells in response to anti-CD3. In contrast to CTLA4, however, hPD-1 is also induced on the surface of B-cells (in response to anti-IgM). hPD-1 is also expressed on a subset of thymocytes and myeloid cells. hPD- 1 polypeptides are inhibitory receptors capable of transmitting an inhibitory signal to an immune cell to thereby inhibit immune cell effector function, or are capable of promoting co-stimulation (e.g. by competitive inhibition) of immune cells, e.g., when present in soluble, monomeric form. An exemplary human PD-1 is shown in UniProtKB / Swiss-Prot Accession No. Q15116.

[0072] PD-1 has 2 ligands, programmed death ligand 1 (PD-L1) and programmed death ligand 2 (PD- L2). An exemplary human PD-L2 is shown in UniProtKB / Swiss-Prot Accession No. Q9BQ51.

[0073] Whilst PD-L1 therapies are being investigated for the treatment of fibrosis (see, for example, the discussion on atezolizumab above, and Zhao et a!., "PD-1 / PD-L1 axis in organ fibrosis", Front. Immunol., 2023, 14: 1145682, doi:10.3389 / fimmu.2023.1145682, and Guo eta / ., "PD-L1 as a Novel Mediator of Lung Fibroblast to Myofibroblast Transition", J. Cell Immunol., 2022, 4(4), 141-144, doi: 10.33696 / Immunology.4.142, both of which are incorporated herein by reference in their entireties), little is known about the potential function and / or implication of PD-L2 in the progress of fibrotic diseases.

[0074] The finding that anti-PD-L2 antibodies can be used in the treatment of a wide range of fibrotic diseases, such as lung, kidney, and heart fibrosis, has been surprising since PD-L2 has not previously been considered as an anti-fibrotic target, and in view of the complex mechanisms involved in the pathogenesis of fibrosis and the difficulties in identifying potential targets for developing a successful treatment strategy.

[0075] Fibrosis is a complex disease, which is still not well understood. However, as explained in Figure 1, there are several general mechanisms which contribute to the formation of fibrotic tissues. It is thought that myofibroblasts are generally responsible for the formation of too much extracellular matrix (ECM)-rich connective tissues which leads to fibrosis and organ stiffening in all fibrotic diseases. Therefore, reduction of the formation of myofibroblasts is key to the treatment and prevention of fibrosis.

[0076] As shown in Figure 1, senescent cells are found in fibrotic tissues and are thought to promote formation of myofibroblasts through the secretion of various pro-fibrotic cytokines, such as TGF-0. TGF-p is known to act on fibroblasts to stimulate the fibroblast to myofibroblast transition.

[0077] M2 Macrophages also contribute to fibrosis by the secretion of various pro-fibrotic cytokines, such as TGF-p, which promotes myofibroblast formation. Thus, as shown in Figure 1, preventing the formation of M2 macrophages (from their Ml macrophage precursor) would be beneficial in the treatment of fibrosis.

[0078] Finally, prevention of the fibroblast to myofibroblast transition through other mechanisms is an important step in the prevention of fibrosis.

[0079] The inventors have found anti-PD-L2 antibodies to mediate pleiotropic effects by mediating anti-fibrotic effects through these three separate mechanisms shown in Figure 1: 1) reducing senescent cell burden, 2) reprogramming pro-fibrotic M2 macrophages that promote the fibroblast to myofibroblast transition, and 3) preventing and / or reversing the formation of myofibroblasts from fibroblasts. As mentioned above, each of these mechanisms is a direct, and general, cause of fibrosis. It is envisaged that improved therapeutic effects may be seen by the targeting of these three separate mechanisms, which are expected to be applicable in all fibrotic diseases.

[0080] In particular, because the mechanisms involved in the pathogenesis of fibrotic diseases are very complex, and not completely understood, a mere assumption that PD-L2 / PD-1 interaction might be involved in the disease would be insufficient to speculate that targeting PD-L2 might give rise to a reasonable expectation of success for a treatment, in the absence of data showing the ability to intervene in the specific mechanisms. The Examples below show three separate and general fibrotic mechanisms are targeted by the anti-PD-L2 antibodies described herein. Hence, the data now presented show that anti-PD-L2 antibodies (such as those defined in claim 1) are suitable for therapy in the treatment of a fibrotic disease.

[0081] As will also be shown in the Examples, anti-PD-L2 antibodies have in particular been found to exhibit a significant effect in the treatment of pulmonary fibrosis, and more particularly of Idiopathic Pulmonary Fibrosis (IPF).

[0082] In one aspect, there is provided an isolated monoclonal antibody (or antigen-binding fragment thereof) that specifically binds to programmed cell death ligand 2 (PD-L2) and blocks activity against PD-L2 / PD1 interaction for use in the treatment of a fibrotic disease in a mammal, wherein the antibody

[0083] (a) binds to human PD-L2 with a dissociation constant (KD) equal from (about) 10'8M to (about) 10'12M; and / or

[0084] (b) blocks in vitro the interaction between human programmed cell death protein 1 (hPD-1) and human PD-L2 with an ECso of from (about) 0.1 to (about) 1.5 nM, optionally in a bioluminescent cell-based assay optionally using Jurkat T cells expressing human PD-1 and a luciferase reporter driven by an NFAT response element (NFAT-RE) and CHO-K1 cells expressing human PD-L2 and an engineered cell surface protein designed to activate cognate TCRs in an antigen-independent manner; and / or

[0085] (c) specifically binds to both human and murine PD-L2; and / or

[0086] (d) competes for binding to PD-L2 with an antibody having a heavy chain variable region having an amino acid sequence of SEQ ID No:l and a light chain variable region having an amino acid sequence of SEQ ID No:9, optionally in an ELISA, SPR or cell-based assay, and optionally wherein the PD-L2 is murine PD-L2.

[0087] According to a particular embodiment, the antibody is defined by a combination of features a) and b), more particularly a combination of features a) and b) only.

[0088] Optionally, in addition to one or more of the features a) to d) defined above, the antibody may further: e) comprise a heavy chain constant region which has reduced or no Fc receptor (FcyR) interactions, or reduced or no immune effector functions. Further optionally, the antibody promotes immune-mediated clearance of senescent cells in fibrotic tissue.

[0089] The immune-mediated clearance may be performed by the innate immune system. The immune-mediated clearance may be performed by the adaptive immune system.

[0090] Without being bound by theory, by removing senescent cells from a fibrotic tissue, the level of fibrosis will be reduced allowing the remaining healthy cells to function more effectively and allowing a recovery or at least stabilization of the tissue. In particular, the accumulation of senescent cells can be avoided.

[0091] Furthermore, given the fact that senescent cells not only lose their functional abilities, but also contribute to inflammation and fibrosis, the inflammatory milieu is dampened by clearing senescent cells, which slows down the initial stages of fibrosis or even helps reverse some of its effects.

[0092] As will be shown in detail in the context of Figure 1 discussed below, this reduction in senescent cell burden relates to a first intervention point of the anti-PD-L2 antibody to mediate anti- fibrotic effects.

[0093] According to a particular embodiment, the antibody comprises each of features a) and b).

[0094] In other embodiments, the antibody comprises each of features c) and d). In another embodiment, the antibody comprises each of features a) and c). In another embodiment, the antibody comprises each of features a) and d). In another embodiment, the antibody comprises each of features b) and c). In another embodiment, the antibody comprises each of features b) and d).

[0095] In other embodiments, the antibody comprises each of features a), b) and c). In another embodiment, the antibody comprises each of features a), b) and d). In another embodiment, the antibody comprises each of features a), c) and d). In another embodiment, the antibody comprises each of features b), c) and d).

[0096] In other embodiments, the antibody comprises each of features a), b), c) and d).

[0097] In another embodiment, the antibody comprises each of features a) to c).

[0098] In an alternative to feature d), the antibody may compete for binding to human PD-L2 with an antibody having any of the combinations of heavy and light variable region sequences described herein, in particular any of humanised VH regions VH1 (SEQ ID NO:2), VH2 (SEQ ID NO:3), VH3 (SEQ ID NO:4), VH4, (SEQ ID NO:5), VH6 (SEQ ID NO:6), VH7 (SEQ ID NO:7) or VH8 (SEQ ID NO:8) when combined with any of the humanised VL regions VK1 (SEQ ID N:10), VK2 (SEQ ID NO: 11) or VK3 (SEQ ID NO: 12). In particular, the antibody may compete for binding to hPD-L2 with an antibody comprising the VH and VL sequences of VH6 / VK3.

[0099] In a further aspect, there is provided an isolated monoclonal antibody that specifically binds to programmed cell death ligand 2 (PD-L2) and blocks activity against hPD-L2 / PD-l interaction, wherein said antibody comprises a heavy chain variable region and a light chain variable region, and comprises a) as heavy chain variable region CDRs: a CDRH1 comprising the amino acid sequence of SEQ ID NO:33 (NYFIE, according to Kabat numbering) or of SEQ ID NO:34 (GYAFSNY, according to Chothia numbering) or alternatively SEQ ID NO:36 (GYSFSNY, according to Chothia numbering), in particular comprising the amino acid sequence of SEQ ID NO: 13 (GYAFSNYFIE) or SEQ ID NO: 14 (GYSFSNYFIE); a CDRH2 region of SEQ ID NO: 15 (LNIPGSGGSNYAEKFKG, according to Kabat numbering) or of SEQ ID NO: 35 (IPGSGG, according to Chothia numbering); and a CDRH3 region of SEQ ID NO: 16 (RRLPPDWYFDV, according to both Kabat and Chothia numbering); and b) as light chain region CDRs: a CDRL1 region of SEQ ID NO: 17 (RSSQSLVHSGGNTYLH, according to both Kabat and Chothia numbering) or SEQ ID NO: 20 (RSSQSLVHSDGNTYLH, according to both Kabat and Chothia numbering); a CDRL2 region of SEQ ID NO: 18 (KVSNRFS, according to both Kabat and Chothia numbering); and a CDRL3 region of SEQ ID NO: 19 (SQSTHVPWT, according to both Kabat and Chothia numbering), for use in treating (or ameliorating or preventing) a fibrotic disease, in particular in a mammal and more particularly in a human.

[0100] According to an alternative embodiment, the antibody comprises the CDR sequences of a) and b) comprising one or two, in particular one, amino acid substitutions, deletions or additions in each of the CDR sequences.

[0101] As reported in international patent application No. WO2023 / 166210A1, antibodies described herein have a high affinity for human PD-L2 while also providing blocking activity against hPD-1. In particular, antibodies described herein have a superior binding activity against human PD-L2 compared to commercially available monoclonal PD-L2 antibodies such as MIH-18, 24F.10C12, D7U86 and TY25.

[0102] In addition to this finding, it has now further been found that anti-PD-L2 antibodies (for example as defined herein) do not only effectively block the hPD-l / hPD-L2 interaction, can be used in the treatment and / or prevention of fibrosis. As described in the Examples below, anti-PD-L2 antibodies have been found to effect a macrophage reprogramming from M2 to Ml phenotype, as well as to directly inhibit the fibroblast to myofibroblast transition, giving direct evidence for their efficacy in the treatment of fibrotic diseases in general. As regards macrophage reprogramming, it has in particular been found that pro-fibrotic M2 macrophages treated with anti PD-L2 antibodies described herein (VH6 / VK3) show a significant increase in CD86 expression and a notable decrease in CD206 expression, reflecting the shift towards an anti-fibrotic Ml-like state. Another anti-PD-L2 antibody (VH0 / VK0), when administered to subjects suffering from fibrosis, showed a significant decrease in the %-CPA (collagen proportionate area) and in the hydroxyproline levels (which as in index of a collagen proliferation) compared to the respective disease control group. In the specific case of pulmonary fibrosis, a decrease in the Ashcroft score (used to assign the extent and severity of fibrosis), of the lung weight and of the respective inflammation score was determined in comparison to the disease control group. Further, a reduction in the expression of pl6 and p21, which are markers for senescence, was seen in subjects suffering from pulmonary fibrosis treated with an anti-PD-L2 antibody as defined herein in comparison to vehicle and isotype controls. This also shows another intervention point in the general treatment of fibrosis through the removal of senescent cells and therefore their secretion of cytokines which induce myofibroblast formation and the major cause of fibrotic diseases.

[0103] Thus, in one embodiment, administration of an anti-PD-L2 antibody (such as the anti-PD-L2 antibodies described herein) results in a decrease (such as a statistically significant decrease) in the %-CPA (collagen proportionate area) in a mouse model of IPF compared to the respective disease control group. In a specific embodiment, the decrease in the %-CPA is at least (about) 5%, at least (about) 10%, at least (about) 15%, at least (about) 20%, at least (about) 25%, at least (about) 30%, at least (about) 35%, or at least (about) 40%, as compared to the disease control group.

[0104] In another embodiment, administration of an anti-PD-L2 antibody (such as an anti-PD-L2 antibody described herein) results in a decrease (such as a statistically significant decrease) in the hydroxyproline levels (which as in index of the quantity of collagen in a tissue OR simply of fibrosis) in a mouse model of IPF compared to the respective disease control group. In a specific embodiment, the decrease in the hydroxyproline level is at least (about) 5%, at least (about) 10%, at least (about) 15%, at least (about) 20%, at least (about) 25%, at least (about) 30%, at least (about) 35%, or at least (about) 38%, as compared to the disease control group.

[0105] In another embodiment, administration of an anti-PD-L2 antibody (such as the ati-PD-L2 antibodies described herein) results in a decrease in the Ashcroft score, a numerical histopathological scoring system to grade the degree of fibrosis and its impact on lung architecture, in a mouse model of IPF compared to the respective disease control group. In one embodiment, the Ashcroft score is decreased by at least (about) 0.5 points. In another embodiment, the Ashcroft score is decreased by at least (about) 0.6 points, or at least (about) 0.7 points, or at least (about) 0.8 points, or at least (about) 0.9 points, or at least (about) 1 point. In another embodiment, the Ashcroft score is decreased by at least (about) 1.1 points, at least (about) 1.2 points, at least (about) 1.3 points, at least (about) 1.4 points.

[0106] In another embodiment, the Ashcroft score is decreased by at least (about) 10%. In another embodiment, the Ashcroft score is decreased by at least (about) 15%, or at least (about) 20%, or at least (about) 25%, or at least (about) 30%, or at least (about) 35%. In a particular embodiment, the Ashcroft score is reduced by at least (about) 38%.

[0107] In another embodiment, administration of an anti-PD-L2 antibody (such as an anti-PD-L2 antibody described herein) results in a decrease of the respective inflammation score, a numerical histopathological scoring system which reflects the amount of pulmonary immune infiltrate, in a mouse model of IPF compared to the respective disease control group. In one embodiment, the inflammation score is decreased by at least (about) 0.5 points. In another embodiment, the inflammation score is decreased by at least (about) 0.6 points, or at least (about) 0.7 points, or at least (about) 0.8 points, or at least (about) 0.9 points, or at least (about) 1 point. In another embodiment, the inflammation score is decreased by at least (about) 1.1 points, at least (about) 1.2 points, or at least (about) 1.3 points.

[0108] In another embodiment, the inflammation score is decreased by at least (about) 10%. In another embodiment, the inflammation score is decreased by at least (about) 15%, or at least (about) 20%, or at least (about) 25%, or at least (about) 30%, or at least (about) 35%. In another embodiment, the inflammation score is decreased by at least (about) 40%, or at least (about) 45%, or at least (about) 50%. In a particular embodiment, the inflammation score is reduced by at least (about) 53%.

[0109] A detailed definition of the Ashcroft score, i.e. of fibrosis grading by the Ashcroft scale, and inflammation score is given below in the experimental section (Example 7).

[0110] In another embodiment, administration of an anti-PD-L2 antibody (such as an anti-PD-L2 antibody described herein) results in a decrease (such as a statistically significant decrease) of pl6 expression by at least (about) 10%, at least (about) 15%, at least (about) 20%, at least (about) 25%, at least (about) 30%, or at least (about) 35%, as compared to the disease control group. In a particular embodiment, the pl6 expression is reduced by at least (about) 40%. In yet another embodiment, administration of an anti-PD-L2 antibody (such as an anti-PD-L2 antibody described herein) results in a decrease (such as a statistically significant decrease) of p21 expression by at least (about) 10%, at least (about 15%, at least (about) 20%, at least (about) 25%, at least (about) 30%, at least (about) 35%, or at least (about) 40%, as compared to the disease control group.

[0111] These measurements may be taken in a mouse model of fibrosis, for example in a bleomycin- induced IPF model as described in Example 7.

[0112] In another aspect, there is provided an isolated monoclonal antibody (or antigen-binding fragment thereof) that specifically binds to programmed cell death ligand 2 (PD-L2) and blocks activity against PD-L2 / PD1 interaction for use in the treatment of a fibrotic disease in a mammal, or a method of treating or reducing the risk of a fibrotic disease, said method comprising administering an effective amount of an anti-PD-L2 antibody or antigen-binding fragment thereof to a subject in need thereof, and whereby the fibrotic disease is treated, or the risk of the fibrotic disease is reduced, wherein: the antibody is capable of preventing the conversion of fibroblasts to myofibroblasts in an in vitrolPV- model as measured by a reduction in a-smooth muscle actin 1 (smal) positive fibres. There is also provided a method of preventing the formation of myofibroblasts, said method comprising contacting an anti-PD-L2 antibody or antigen-binding fragment thereof with a population of cells comprising fibroblasts, wherein the anti-PD-L2 antibody or fragment is capable of preventing the conversion of fibroblasts to myofibroblasts, in particular as measured by a reduction in a-smooth muscle actin 1 (smal) positive fibres in an in vitro W model.

[0113] In one embodiment, the reduction in smal positive fibres may be compared to a TGF-0 treated control sample. The reduction (such as a statistically significant reduction) may be at least (about) 35%. The reduction may be at least (about) 40%. The reduction may be at least (about) 45%. The reduction may be at least (about) 50%. The reduction may be at least (about) 55%. The reduction may be at least (about) 65%. The reduction may be at least (about) 75%. In particular, the reduction may be at least (about) 60%, or at least (about) 70%, compared to a TGF-p treated control sample.

[0114] In another embodiment, the reduction is comparable (e.g. statistically similar) to a positive anti-fibrotic control. The positive anti-fibrotic control may be an anti-PD-Ll antibody, for example atezolizumab.

[0115] The antibody or fragment may be any of the antibodies or fragments as described elsewhere herein.

[0116] In another embodiment, the reduction in smal positive fibres is measured by a method comprising the steps of:

[0117] - providing fibroblasts from a human donor;

[0118] - treating the fibroblasts with TGF-0;

[0119] - contacting the antibody or fragment with the treated fibroblasts;

[0120] - incubating the fibroblasts in the presence of the TGF-0 and the antibody or fragment;

[0121] - conducting immunofluorescence staining against smooth muscle actin 1 (smal); and

[0122] - quantifying smal positive fibers, in particular by fluorescence-activated cell sorting (FACS).

[0123] The addition of TGF-p induces the transition from a fibroblast to a myofibroblast (see, for example, Desmouliere etai., "Transforming growth factor-beta 1 induces alpha-smooth muscle actin expression in granulation tissue myofibroblasts and in quiescent and growing cultured fibroblasts", J. Cell Biol., 1993, 122(1), 103-111, doi: 10.1083 / jcb.122.1.103 and Sime et al, "Adenovector-mediated gene transfer of active transforming growth factor-betal induces prolonged severe fibrosis in rat lung", J. Clin. Invest., 1997, 100(4), 768-776, doi: 10.1172 / JCI119590, both of which are incorporated herein by reference in their entirety).

[0124] The fibroblasts may be cultured, for example by growing in a fibroblast specific media (e.g. FBM fibroblast Basal Medium) from an IPF sample obtained from a diseased human (or other nonhuman mammalian) subject. In one embodiment, incubation is carried out for 3 days. The TGF-0 may be added at a concentration of (about) 2.5 ng / mL. The antibody may be added at a concentration of (about) 60 pg / mL. In one embodiment, the method is carried out as described in Example 6. There is also provided an isolated monoclonal antibody or antigen-binding fragment thereof that specifically binds to programmed cell death ligand 2 (PD-L2) and blocks activity against PD- L2 / PD1 interaction for use in the treatment of a fibrotic disease in a mammal, or a method of treating or reducing the risk of a fibrotic disease, said method comprising administering an effective amount of an anti-PD-L2 antibody or antigen-binding fragment thereof to a subject in need thereof, and whereby the fibrotic disease is treated or the risk of the fibrotic disease is reduced, and wherein

[0125] (i) the antibody or fragment is capable of reprogramming M2 pro-fibrotic macrophages expressing CD206 on the surface of said macrophage into Ml phenotypic macrophages expressing CD86 on the surface of said macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages.

[0126] There is also provided an isolated monoclonal antibody or antigen-binding fragment thereof that specifically binds to programmed cell death ligand 2 (PD-L2) and blocks activity against PD- L2 / PD1 interaction for use in the treatment of a fibrotic disease in a mammal, or a method of treating or reducing the risk of a fibrotic disease, said method comprising administering an effective amount of an anti-PD-L2 antibody or antigen-binding fragment thereof to a subject in need thereof, and whereby the fibrotic disease is treated or the risk of the fibrotic disease is reduced, and wherein

[0127] (ii) the antibody or fragment is capable of increasing CD86 expression on a macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages.

[0128] There is also provided a method of reprogramming M2 pro-fibrotic macrophages to Ml phenotypic macrophages, said method comprising contacting an anti-PD-L2 antibody or antigenbinding fragment thereof with said M2 macrophages, wherein the anti-PD-L2 antibody or fragment decreases the level (or amount) of the M2 pro-fibrotic macrophages and increases the level (or amount) of the Ml phenotypic macrophages, in particular as measured by (i) observing an increase in CD86 expression on the macrophages.

[0129] The increase in CD86 expression may be compared to an untreated M2 macrophage. The increase (such as a statistically significant increase) may be an increase of at least (about) 40%. The increase may be at least (about) 45%. The increase may be at least (about) 50%. In particular, the increase may be at least (about) 55%, compared to an untreated M2 macrophage. In another embodiment, the increase may be at least (about) 60%.

[0130] In another embodiment, the increase in CD86 expression is measured as compared to an isotype control antibody, and the increase (such as a statistically significant increase) may be at least (about) 40%, or at least (about) 45%. In particular, the increase may be at least (about) 50% or, in particular at least (about) 55%.

[0131] In another embodiment, the increase in CD86 expression is greater than that seen for a positive control. The positive control may be a CSFR inhibitor, for example PLX3397.

[0132] There is also provided an isolated monoclonal antibody or antigen-binding fragment thereof that specifically binds to programmed cell death ligand 2 (PD-L2) and blocks activity against PD- L2 / PD1 interaction for use in the treatment of a fibrotic disease in a mammal, or a method of treating or reducing the risk of a fibrotic disease, said method comprising administering an effective amount of an anti-PD-L2 antibody or antigen-binding fragment thereof to a subject in need thereof, and whereby the fibrotic disease is treated or the risk of the fibrotic disease is reduced, and wherein

[0133] (iii) the antibody or fragment is capable of decreasing CD206 expression on a macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages.

[0134] There is also provided a method of reprogramming M2 pro-fibrotic macrophages to Ml phenotypic macrophages, said method comprising contacting an anti-PD-L2 antibody or antigenbinding fragment thereof with said M2 macrophages, wherein the anti-PD-L2 antibody or fragment decreases the level (or amount) of the M2 pro-fibrotic macrophages and increases the level (or amount) of the Ml phenotypic macrophages, in particular as measured by (ii) observing a decrease in CD206 expression on the macrophages.

[0135] The decrease in CD206 expression may be compared to an untreated M2 macrophage. The decrease (such as a statistically significant decrease) may be a decrease of at least (about) 50%. The decrease may be at least (about) 60%. In particular, the decrease may be at least (about) 65%, or may be at least (about) 70%, compared to an untreated M2 macrophage.

[0136] In another embodiment, the decrease in CD206 expression is measured as compared to an isotype control antibody, and the decrease (such as a statistically significant decrease) may be at least (about) 50%, or at least (about) 55%, or at least (about) 60%, or in particular at least (about) 70%.

[0137] In another embodiment, the decrease in CD206 expression is comparable (e.g. statistically similar) to that seen for a positive control. The positive control may be a CSFR inhibitor, for example PLX3397.

[0138] There is also provided an isolated monoclonal antibody or antigen-binding fragment thereof that specifically binds to programmed cell death ligand 2 (PD-L2) and blocks activity against PD- L2 / PD1 interaction for use in the treatment of a fibrotic disease in a mammal, or a method of treating or reducing the risk of a fibrotic disease, said method comprising administering an effective amount of an anti-PD-L2 antibody or antigen-binding fragment thereof to a subject in need thereof, and whereby the fibrotic disease is treated or the risk of the fibrotic disease is reduced, and wherein

[0139] (iv) the antibody or fragment is capable of reducing TGF-0 production in a macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages.

[0140] There is also provided a method of reprogramming M2 pro-fibrotic macrophages to Ml phenotypic macrophages, said method comprising contacting an anti-PD-L2 antibody or antigenbinding fragment thereof with said M2 macrophages, wherein the anti-PD-L2 antibody or fragment decreases the level (or amount) of the M2 pro-fibrotic macrophages and increases the level (or amount) of the Ml phenotypic macrophages, in particular as measured by (iii) observing a reduction in TGF-p production on the macrophages.

[0141] The reduction in TGF-0 production may be compared to an untreated M2 macrophage. The reduction (such as a statistically significant reduction) may be a reduction of at least (about) 30%. The reduction may be at least (about) 40%. In particular, the reduction may be at least (about) 50%, or may be at least (about) 55%, compared to an untreated M2 macrophage.

[0142] In another embodiment, the reduction in TGF-0 production is measured as compared to an isotype control antibody, and the reduction (such as a statistically significant reduction) may be at least (about) 40%, or at least (about) 50%, or at least (about) 60%, or in particular at least (about) 70%.

[0143] There is also provided an isolated monoclonal antibody or antigen-binding fragment thereof that specifically binds to programmed cell death ligand 2 (PD-L2) and blocks activity against PD- L2 / PD1 interaction for use in the treatment of a fibrotic disease in a mammal, or a method of treating or reducing the risk of a fibrotic disease, said method comprising administering an effective amount of an anti-PD-L2 antibody or antigen-binding fragment thereof to a subject in need thereof, and whereby the fibrotic disease is treated or the risk of the fibrotic disease is reduced, and wherein

[0144] (v) the antibody or fragment is capable of increasing TNF-a production in a macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages.

[0145] There is also provided a method of reprogramming M2 pro-fibrotic macrophages to Ml phenotypic macrophages, said method comprising contacting an anti-PD-L2 antibody or antigenbinding fragment thereof with said M2 macrophages, wherein the anti-PD-L2 antibody or fragment decreases the level (or amount) of the M2 pro-fibrotic macrophages and increases the level (or amount) of the Ml phenotypic macrophages, in particular as measured by (iv) observing an increase in TNF-a production on the macrophages. The increase in TNF-a production may be compared to an untreated M2 macrophage. The increase (such as a statistically significant increase) may be an increase of at least (about) 5-fold. The increase may be at least (about) 5.5-fold. The increase may be at least (about) 6-fold. In particular, the increase may be at least (about) 6.5-fold, or may be at least (about) 7-fold, compared to an untreated M2 macrophage.

[0146] In another embodiment, the increase in TNF-a production is measured as compared to an isotype control antibody, and the increase (such as a statistically significant increase) may be at least (about) 2-fold, or at least (about) 3-fold, or at least (about) 3.5-fold, or in particular at least (about) 4-fold.

[0147] There is also provided an isolated monoclonal antibody or antigen-binding fragment thereof that specifically binds to programmed cell death ligand 2 (PD-L2) and blocks activity against PD- L2 / PD1 interaction for use in the treatment of a fibrotic disease in a mammal, or a method of treating or reducing the risk of a fibrotic disease, said method comprising administering an effective amount of an anti-PD-L2 antibody or antigen-binding fragment thereof to a subject in need thereof, and whereby the fibrotic disease is treated or the risk of the fibrotic disease is reduced, and wherein

[0148] (vi) the antibody or fragment is capable of changing the morphology of the M2 macrophages from an elongated morphology to a rounded morphology, in particular as observed by microscopic analysis of cells in an in vitro model of M2 pro-fibrotic macrophages.

[0149] There is also provided a method of reprogramming M2 pro-fibrotic macrophages to Ml phenotypic macrophages, said method comprising contacting an anti-PD-L2 antibody or antigenbinding fragment thereof with said M2 macrophages, wherein the anti-PD-L2 antibody or fragment decreases the level (or amount) of the M2 pro-fibrotic macrophages and increases the level (or amount) of the Ml phenotypic macrophages, in particular as measured by (v) observing a change in morphology of the M2 macrophages from an elongated morphology to a rounded morphology.

[0150] As shown in Figure 5C and Example 5F, M2 and Ml macrophages each have a specific and characteristic morphology which is easily observable by the skilled person.

[0151] There is also provided an isolated monoclonal antibody or antigen-binding fragment thereof that specifically binds to programmed cell death ligand 2 (PD-L2) and blocks activity against PD- L2 / PD1 interaction for use in the treatment of a fibrotic disease in a mammal, or a method of treating or reducing the risk of a fibrotic disease, said method comprising administering an effective amount of an anti-PD-L2 antibody or antigen-binding fragment thereof to a subject in need thereof, and whereby the fibrotic disease is treated or the risk of the fibrotic disease is reduced, and wherein (vii) the antibody or fragment is capable of increasing the proliferation of macrophages, in particular as measured by number of cells in an in vitro model of M2 pro-fibrotic macrophages.

[0152] There is also provided a method of reprogramming M2 pro-fibrotic macrophages to Ml phenotypic macrophages, said method comprising contacting an anti-PD-L2 antibody or antigenbinding fragment thereof with said M2 macrophages, wherein the anti-PD-L2 antibody or fragment decreases the level (or amount) of the M2 pro-fibrotic macrophages and increases the level (or amount) of the Ml phenotypic macrophages, in particular as measured by (vi) observing an increase in the proliferation of macrophages.

[0153] The increase in the proliferation of macrophages may be compared to an untreated M2 macrophage. The increase (such as a statistically significant increase) may be an increase of at least (about) 1.4-fold. The increase may be at least (about) 1.5-fold. The increase may be at least (about) 1.6-fold. In particular, the increase may be at least (about) 1.7-fold, or may be at least (about) 1.8- fold, compared to an untreated M2 macrophage.

[0154] In another embodiment, the increase in the proliferation of macrophages is measured as compared to an isotype control antibody, and the increase (such as a statistically significant increase) may be at least (about) 1.4-fold, or at least (about) 1.5-fold, or at least (about) 1.6-fold, or in particular at least (about) 1.7-fold.

[0155] In one embodiment, the in vitro model of M2 pro-fibrotic macrophages comprises the steps of providing monocytes from a human donor; treating the monocytes with the cytokines M-CSF1 and IL-4 to polarise the monocytes to M2 macrophages; contacting the antibody with the M2 macrophages; incubating the M2 macrophages in the presence of the cytokines and the antibody or fragment; and for observing an increase in CD86 and / or a decrease in CD206, conducting immunofluorescence staining against CD86 and / or CD206; and quantifying expression of CD86 and / or CD206, in particular by flow cytometry; and / or for observing a reduction in TGF-0 and / or increase in TNF-a, conducting an ELIZA assay to quantify the amount of TGF-0 and / or TNF-a; and / or for observing an increase in the proliferation of macrophages counting the number of macrophages on a cell counter.

[0156] The method may be the methodology as described in Example 5. The antibody or fragment, of the fibrotic disease may be any of the antibodies, fragments or fibrotic diseases described elsewhere herein. Optionally, the antibody or fragment is administered (or contacted) in combination with a therapeutically active agent as described elsewhere herein.

[0157] The M2 macrophages and / or the Ml macrophages may be as described elsewhere herein.

[0158] In the use of the antibody or fragment, or the method of treatment described above, the antibody or fragment may be capable of two of the effects described in parts (i) to (vii). For example, the antibody or fragment may be capable of the effects described in (ii) and (iii). The antibody or fragment may be capable of the effects described in (i) and a further effect selected from (ii), (iii), (iv), (v), (vi) and (vii). The antibody or fragment may be capable of the effects described in (ii) and a further effect selected from (iii), (iv), (v), (vi) and (vii). The antibody or fragment may be capable of the effects described in (iii) and a further effect selected from (iv), (v), (vi) and (vii). The antibody or fragment may be capable of the effects described in (iv) and a further effect selected from (v), (vi) and (vii). The antibody or fragment may be capable of the effects described in (v) and a further effect selected from (vi) and (vii). The antibody or fragment may be capable of the effects described in (vi) and (vii).

[0159] In the use of the antibody or fragment, or the method of treatment described above, the antibody or fragment may be capable of three of the effects described in parts (i) to (vii). Alternatively, the antibody or fragment may be capable of four of the effects described in parts (i) to (vii). Alternatively, the antibody or fragment may be capable of five of the effects described in parts (i) to (vii). Alternatively, the antibody or fragment may be capable of six of the effects described in parts (i) to (vii). Alternatively, the antibody or fragment may be capable of all of the effects described in parts

[0160] (i) to (vii).

[0161] In the method of reprogramming M2 pro-fibrotic macrophages to Ml phenotypic macrophages described above, the antibody or fragment may be capable of two of the effects described in parts (i) to (vi). For example, the antibody or fragment may be capable of the effects described in (ii) and (iii). The antibody or fragment may be capable of the effects described in (i) and a further effect selected from (ii), (iii), (iv), (v) and (vi). The antibody or fragment may be capable of the effects described in

[0162] (ii) and a further effect selected from (iii), (iv), (v) and (vi). The antibody or fragment may be capable of the effects described in (iii) and a further effect selected from (iv), (v) and (vi). The antibody or fragment may be capable of the effects described in (iv) and a further effect selected from (v) and (vi). The antibody or fragment may be capable of the effects described in (v) and (vi).

[0163] In the method of reprogramming M2 pro-fibrotic macrophages to Ml phenotypic macrophages described above, the antibody or fragment may be capable of three of the effects described in parts (i) to (vi). Alternatively, the antibody or fragment may be capable of four of the effects described in parts (i) to (vi). Alternatively, the antibody or fragment may be capable of five of the effects described in parts (i) to (vi). Alternatively, the antibody or fragment may be capable of all of the effects described in parts (i) to (vi). The Examples further show that the efficacy of anti-PD-L2 antibodies not only relates to pulmonary fibrosis but is also applicable to fibrotic diseases in general, by acting to prevent or reduce general fibrotic mechanisms (as discussed herein in relation to Figure 1). Further, markers of fibrosis indicate that anti-PD-L2 antibodies may be useful in the specific treatment of kidney fibrosis (see in particular Figures 10A and 10B) as well as aorta or heart fibrosis (see the data in Figure IOC discussed in Example 8, which is indicative of therapeutic efficacy). As shown in Figure 6E, the effect of anti- PD-L2 antibodies on stellate cells suggests useful application in the treatment of liver fibrosis.

[0164] In a general aspect, there is provided an isolated monoclonal antibody that specifically binds to programmed cell death ligand 2 (PD-L2) and blocks activity against PD-L2 / PD1 interaction for use in the treatment of a fibrotic disease in a mammal (in particular a human subject).

[0165] In another general aspect, there is provided a method of treating or reducing the risk of a fibrotic disease, said method comprising administering an effective amount of an anti-PD-L2 antibody or antigen-binding fragment thereof to a subject in need thereof, and whereby the fibrotic disease is treated or the risk of the fibrotic disease is reduced.

[0166] In another general aspect, there is provided a method of treating or reducing the risk of a fibrotic disease, said method comprising administering an effective amount of an anti-PD-L2 antibody or antigen-binding fragment thereof to a subject in need thereof, wherein the anti-PD-L2 antibody or fragment comprises means for binding to programmed cell death ligand 2 (PD-L2) and means for blocking activity against PD-L2 / PD1 interaction, and whereby the fibrotic disease is treated or the risk of the fibrotic disease is reduced.

[0167] In another general aspect, there is provided a pharmaceutical composition for treating or reducing the risk of a fibrotic disease, said composition comprising an anti-PD-L2 antibody or antigenbinding fragment.

[0168] The antibody or fragment, or the fibrotic disease may be any of the antibodies, fragments or fibrotic diseases described elsewhere herein. Optionally, the antibody or fragment is administered (or contacted) in combination with a therapeutically active agent as described elsewhere herein.

[0169] In any use or method described herein, the method or use may be, in particular, carried out in vivo in a human subject. Alternatively, the method or use may be carried out in vivo in a a nonhuman mammal.

[0170] In any use or method described herein, the subject has or is at risk of a fibrotic disease characterised by the presence of:

[0171] (i) senescent cells; and / or

[0172] (ii) M2 pro-fibrotic macrophages; and / or

[0173] (iii) fibroblasts.

[0174] In any use or method described herein, any contacting of the macrophages and the antibody or fragment may be by administering said antibody or fragment in an effective amount to said or a subject, and optionally whereby said disease is treated, or the risk of said disease is reduced. In embodiments related to the use of antibodies in the treatment of fibrotic diseases, or any other use described herein, the antibody may comprise a mutagenized Fc variant sequence of murine IgG2a subclass exhibiting reduced Fc receptor (FcyR) interaction compared to its parent, wild type Fc region. In any of these embodiments, the mutagenized Fc variant sequence may comprise a mutation of leucine 234 to alanine and leucine 235 to alanine (LALA mutation) according to EU numbering, in particular wherein the heavy chain constant region comprises the amino acid sequence of SEQ ID NO:25.

[0175] In a further aspect, there is provided an isolated variant monoclonal antibody comprising a heavy chain constant region which has reduced or no Fc receptor (FcyR) interactions, or reduced or no immune effector functions, for example a mutagenized Fc variant sequence having reduced Fc receptor (FcyR) interaction or reduced immune effector function compared to the parent (wild-type) Fc region.

[0176] The variant antibody may comprise a heavy chain constant region which has (i) a wild-type constant region sequence which has reduced or no Fc receptor (FcyR) interactions, (ii) a wild-type constant region sequence which has reduced or no immune effector functions, or (iii) a mutagenized Fc variant sequence of the IgG isotype exhibiting reduced Fc receptor (FcyR) interaction or reduced immune effector function compared to the parent Fc region, and for part (iii), wherein the isolated monoclonal antibody is a chimeric, humanized or human antibody. Optionally, the antibody is a chimeric, humanized or human antibody.

[0177] These variant antibodies are also referred to as "effector-reduced" anti-PD-L2 antibodies. It is understood that the term "effector-reduced" also covers "effector-disabled", e.g. having essentially no effector function.

[0178] As will be shown in the context of the in vivo experimental data discussed below, efficacy in the treatment of subjects suffering from fibrosis, in particular pulmonary fibrosis, in a dose-dependent manner has also been shown for an effector-reduced anti-PD-L2 antibody variant. Specifically, a pronounced reduction in hydroxyproline content, in the %-CPA, in the Ashcroft score, as well as in the inflammation score was seen in mice suffering from bleomycin-induced IPF when treated with an effector-reduced anti-PD-L2 antibody (specifically VH0 / VK0-mIgG2a LALA), the effect being equivalent to the one seen for treatment with pirfenidone. In addition to that, pharmacokinetic studies revealed a relatively long half-life for an effector-reduced anti-PD-L2 antibody.

[0179] In addition, data obtained by studies in a mouse model indicate that an effector-reduced anti- PD-L2 antibody described below also exhibits a clear beneficial effect in the treatment of a disease that is driven by the accumulation of senescent cells (such as pathologically accelerated senescence), or premature aging, in particular progeria (Hutchinson-Gilford progeria syndrome).

[0180] Thus, in one embodiment, there is provided an anti-PD-L2 antibody or antigen-binding fragment thereof (such as an anti-PD-L2 antibody as described elsewhere herein) for use in the treatment of progeria syndrome (such as Hutchinson-Gilford progeria syndrome). In another embodiment, there is provided an anti-PD-L2 antibody or antigen-binding fragment thereof (such as an anti-PD-L2 antibody as described elsewhere herein) for use in reducing the effects of premature ageing.

[0181] In one embodiment, administration of an anti-PD-L2 antibody or antigen-binding fragment thereof (such as an anti-PD-L2 antibody described herein) results in an extended median lifespan by at least (about) 5% in a subject suffering from a disease that is driven by the accumulation of senescent cells (e.g. pathologically accelerated senescence) or premature aging, and specifically progeria (e.g. Hutchinson-Gilford progeria syndrome). In this regard, in a particular embodiment, by the administration of the antibody or fragment, median lifespan is extended by at least (about) 6%, at least (about) 8%, at least (about) 10%, at least (about) 12%, or at least (about) 13%, as compared to an untreated control group. In one embodiment, the subject is a female mouse.

[0182] In another embodiment, the administration of an anti-PD-L2 antibody or antigen-binding fragment thereof (such as an anti-PD-L2 antibody described herein) results in an extended median lifespan by at least (about) 10 days in a subject (e.g. a mouse) suffering from a disease selected from a disease that is driven by the accumulation of senescent cells (e.g. pathologically accelerated aging), premature aging and progeria (e.g. Hutchinson-Gilford progeria syndrome). In another embodiment, the median extended lifespan is at least (about) 15 days in a subject (e.g. a mouse).

[0183] In another embodiment, the administration of an anti-PD-L2 antibody or antigen-binding fragment thereof (such as an anti-PD-L2 antibody described herein) results in an extended median lifespan by at least (about) 25 days in a subject (e.g. a female mouse) suffering from a disease selected from a disease that is driven by the accumulation of senescent cells (e.g. pathologically accelerated aging), premature aging and progeria (e.g. Hutchinson-Gilford progeria syndrome). In another embodiment, the median extended lifespan is at least (about) 30 days, or at least (about) 35 days in a subject (e.g. a female mouse).

[0184] In one embodiment, the reduction of the effects of premature aging and / or the extension of lifespan is seen in a mouse model of Hutchinson-Gilford progeria syndrome (e.g. as further described in Example 8), and optionally the subject is a female mouse.

[0185] In a particular embodiment, the anti-PD-L2 antibody or antigen-binding fragment thereof (for example for any of the uses or methods described herein) comprises a) as heavy chain variable region CDRs: a CDRH1 comprising the amino acid sequence of SEQ ID NO:33 (NYFIE, according to Kabat numbering) or of SEQ ID NO:36 (GYSFSNY, according to Chothia numbering), in particular comprising the amino acid sequence of SEQ ID NO: 14 (GYSFSNYFIE); a CDRH2 comprising the amino acid sequence of SEQ ID NO: 15 (LNIPGSGGSNYAEKFKG, according to Kabat numbering) or of SEQ ID NO:35 (IPGSGG, according to Chothia numbering); and a CDRH3 comprising the amino acid sequence of SEQ ID NO: 16 (RRLPPDWYFDV, according to both Kabat and Chothia numbering); and b) as light chain variable region CDRs a CDRL1 comprising the amino acid sequence of SEQ ID NO: 17 (RSSQSLVHSGGNTYLH, according to both Kabat and Chothia numbering); a CDRL2 comprising the amino acid sequence of SEQ ID NO: 18 (KVSNRFS, according to both Kabat and Chothia numbering); and a CDRL3 comprising the amino acid sequence of SEQ ID NO: 19 (SQSTHVPWT, according to both Kabat and Chothia numbering), or which comprises the CDR sequences of a) and b) comprising one or two, in particular one, amino acid substitutions, deletions or additions in one or more (e.g. all) of the CDR sequences.

[0186] These antibodies have been found to have a particularly high affinity for human PD-L2 while also providing blocking activity against hPD-1. Furthermore, they have a low isomerization risk, which is a significant advantage.

[0187] In another particular embodiment, the isolated monoclonal antibody or antigen-binding fragment thereof (for example for any of the uses or methods described herein) comprises: a) as heavy chain variable region CDRs a CDRH1 comprising the amino acid sequence of SEQ ID NO:33 (NYFIE, according to Kabat numbering) or of SEQ ID NO: 34 (GYAFSNY, according to Chothia numbering), in particular comprising the amino acid sequence of SEQ ID NO: 13 (GYAFSNYFIE); a CDRH2 comprising the amino acid sequence of SEQ ID NO: 15 (LNIPGSGGSNYAEKFKG, according to Kabat numbering) or of SEQ ID NO:35 (IPGSGG, according to Chothia numbering); and a CDRH3 comprising the amino acid sequence of SEQ ID NO: 16 (RRLPPDWYFDV, according to both Kabat and Chothia numbering); and b) as light chain variable region CDRs a CDRL1 comprising the amino acid sequence of SEQ ID NO: 20 (RSSQSLVHSDGNTYLH, according to both Kabat and Chothia numbering); a CDRL2 comprising the amino acid sequence of SEQ ID NO: 18 (KVSNRFS, according to both Kabat and Chothia numbering); and a CDRL3 comprising the amino acid sequence of SEQ ID NO: 19 (SQSTHVPWT, according to both Kabat and Chothia numbering), or which comprises the CDR sequences of a) and b) comprising one or two, in particular one, amino acid substitutions, deletions or additions in one or more (e.g. all) of the CDR sequences. These antibodies have also been found to have a particularly high affinity for human and / or mouse PD-L2, while also providing blocking activity against hPD-1.

[0188] According to a further embodiment, the antibody or antigen-binding fragment thereof is a humanised or chimeric antibody whose three heavy chain CDRs are derived from a murine IGHV gene segment which is IGHV1-54 (SEQ ID No:37), a murine IGHD gene segment, and murine IGHJ gene segment which is IGHJ1 (SEQ ID No:38), and whose three light chain CDRs derived from a murine IGKV gene segment which is IGKV1-110 (SEQ ID No:39) and a murine IGKJ gene segment which is IGKJ1 (SEQ ID No:40).

[0189] SEQ ID No:37 (Murine IGHV1-54, IMGT Accession ID:AC074329) cag gtc cag ctg cag cag tct gga get gag etg gta agg cct ggg act tea gtg aag gtg tcc tgc aag get tct gga tac gee ttc act aat tac ttg ata gag tgg gta aag cag agg cct gga cag ggc ett gag tgg att gga gtg att aat cct gga agt ggt ggt act aac tac aat gag aag ttc aag ggc aag gca aca ctg act gca gac aaa tcc tcc age act gcc tac atg cag etc age age ctg aca tct gag gac tct geg gtc tat ttc tgt gca aga

[0190] SEQ ID No:38 (Murine IGHJ 1-03, IMGT Accession ID:X63164)

[0191] C TAC TGG TAC TTC GAT GTC TGG GGC ACA GGG ACC ACG GTC ACC GTC TCC TCA G

[0192] SEQ ID No:39 (Murine IGKV1-110, IMGT Accession ID:D00080) gat gtt gtg atg acc caa act cca etc tcc ctg cct gtc agt ett gga gat caa gcc tcc ate tct tgc aga tct agt cag age ett gta cac agt aat gga aac acc tat tta cat tgg tac ctg cag aag cca ggc cag tct cca aag etc ctg ate tac aaa gtt tcc aac ega ttt tct ggg gtc cca gac agg ttc agt ggc agt gga tea ggg aca gat ttc aca etc aag ate age aga gtg gag get gag gat ctg gga tt tat ttc tgc tct caa agt aca cat gtt cct cc

[0193] SEQ ID No:40 (Murine IGK 1, IMGT Accession ID:IMGT000068)

[0194] G TGG ACG TTC GGT GGA GGC ACC AAG CTG GAA ATC AAA C

[0195] Within the context of the present disclosure, VH0 / VK0 is a chimeric antibody (mouse VHO on a hlgGl backbone, and mouse VKO on human light chain constant region). In particular, the V region of the antibodies described herein have sequences that are devoid of or reduced in significant T cell epitopes. Eight heavy chain variable regions (VHO (mouse), VH1 to VH4 and VH6 to VH8 (all humanized)) and four light chain variable regions (VKO (mouse) and VK1 to VK3 (humanized)) sequences were found to provide particularly effective antibodies.

[0196] VHO, VH1, VH2, VH3, VH4, VH6, VH7 and VH8 all comprise the amino acid sequence of SEQ ID NO:33 (NYFIE, according to Kabat numbering) or of SEQ ID NO:34 (GYAFSNY, according to Chothia numbering) or alternatively SEQ ID NO:36 (GYSFSNY, according to Chothia numbering), in particular of SEQ ID NO: 13 (GYAFSNYFIE) or SEQ ID NO: 14 (GYSFSNYFIE), as CDRH1, SEQ ID NO: 15 (LNIPGSGGSNYAEKFKG, according to Kabat numbering) or of SEQ ID NO:35 (IPGSGG, according to Chothia numbering) as CDRH2; and SEQ ID NO: 16 (RRLPPDWYFDV, according to both Kabat and Chothia numbering) as CDRH3. In one embodiment, the antibody or antigen-binding fragment thereof may comprise any of the CDR sequences described herein with one or two amino acid substitutions, deletions or additions (in particular one or two substitutions, e.g. one substitution) to the given SEQ ID No. For example, the antibody or fragment may comprise one or two (e.g. one) amino acid substitutions, deletions or additions (e.g. substitution(s)) in CDRH1. For example, the antibody or fragment may comprise one or two (e.g. one) amino acid substitutions, deletions or additions (e.g. substitution(s)) in CDRH2. For example, the antibody or fragment may comprise one or two (e.g. one) amino acid substitutions, deletions or additions (e.g. substitution(s)) in CDRH3. For example, the antibody or fragment may comprise one or two (e.g. one) amino acid substitutions, deletions or additions (e.g. substitution(s)) in CDRL1. For example, the antibody or fragment may comprise one or two (e.g. one) amino acid substitutions, deletions or additions (e.g. substitution(s)) in CDRL2. For example, the antibody or fragment may comprise one or two (e.g. one) amino acid substitution, deletion or addition (e.g. substitution) in CDRL3. The substitution may be a conservative substitution.

[0197] VKO, VK1, VK2 and VK3 all comprise the amino acid sequences of SEQ ID NO: 17 (RSSQSLVHSGGNTYLH, according to both Kabat and Chothia numbering) or SEQ ID NO:20 (RSSQSLVHSDGNTYLH, according to both Kabat and Chothia numbering) as CDRL1; SEQ ID NO: 18 (KVSNRFS, according to both Kabat and Chothia numbering) as CDRL2; and SEQ ID NO: 19 (SQSTHVPWT, according to both Kabat and Chothia numbering) as CDRL3.

[0198] The specific amino acid sequences of VHO, VH1, VH2, VH3, VH4, VH6, VH7 and VH8 and of VKO, VK1, VK2 and VK3 are listed in Table 1 below.

[0199] Table 1: VH and VK variable region sequences

[0200] In particular, the isolated monoclonal antibody or antigen-binding fragment thereof comprises: (a) a heavy chain variable region having a complementarity determining region 1 (CDRH1) comprising the amino acid sequence of SEQ ID NO:33 (NYFIE, according to Kabat numbering) or of SEQ ID NO:34 (GYAFSNY, according to Chothia numbering), in particular the amino acid sequence or alternatively SEQ ID NO:36 (GYSFSNY, according to Chothia numbering), in particular comprising the amino acid sequence of SEQ ID NO: 13 (GYAFSNYFIE) or SEQ ID NO: 14 (GYSFSNYFIE); a CDRH2 comprising the amino acid sequence of SEQ ID NO: 15 (LNIPGSGGSNYAEKFKG, according to Kabat numbering) or of SEQ ID NO:35 (IPGSGG, according to Chothia numbering); and a CDRH3 comprising the amino acid sequence of SEQ ID NO: 16 (RRLPPDWYFDV, according to both Kabat and Chothia numbering), wherein the remaining part of the heavy chain variable region sequence has at least 95% identity to a heavy chain variable region sequence selected from the group consisting of the sequences listed in Table 1; and

[0201] (b) a light chain variable region having a CDRL1 comprising the amino acid sequence of SEQ ID NO: 17 (RSSQSLVHSGGNTYLH, according to both Kabat and Chothia numbering) or SEQ ID NO:20 (RSSQSLVHSDGNTYLH, according to both Kabat and Chothia numbering); a CDRL2 comprising the amino acid sequence of SEQ ID NO: 18 (KVSNRFS, according to both Kabat and Chothia numbering); and a CDRL3 comprising the amino acid sequence of SEQ ID NO: 19 (SQSTHVPWT, according to both Kabat and Chothia numbering), wherein the remaining part of the heavy chain variable region sequence has at least 95% identity to a heavy chain variable region sequence selected from the group consisting of the sequences listed in Table 1.

[0202] All humanized variant antibodies containing VH1, VH2, VH3, VH4, VH6, VH7 and VH8 as heavy chains and VK1, VK2 and VK3 as light chains bind human PD-L2 antigen with high affinity, as shown in Table 2. Table 2 relates to single cycle kinetic parameters of chimeric (VHO / VKO) and humanized variants (tested as cell culture supernatants) binding to human PD-L2 antigen as determined using the Biacore 8K.

[0203] Table 2: KD (Biacore) binding affinities of selected antibodies to hPD-L2

[0204] In a particular embodiment, the monoclonal antibody or antigen-binding fragment thereof (for example for any of the uses or methods described herein) comprises a heavy chain variable region and a light chain variable region, wherein:

[0205] (a) the heavy chain variable region comprises the amino acid sequence of SEQ ID NO: 1 (VHO) and the light chain variable region comprises the amino acid sequence of SEQ ID NO:9 (VKO); or

[0206] (b) the heavy chain variable region comprises the amino acid sequence of SEQ ID NO:6 (VH6) and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 10 (VK1), of SEQ ID NO: 11 (VK2) or of SEQ ID NO: 12 (VK3); or

[0207] (c) the heavy chain variable region comprises the amino acid sequence of SEQ ID NO:7 (VH7) and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 11 (VK2); or

[0208] (d) the heavy chain variable region comprises the amino acid sequence of SEQ ID NO:8 (VH8) and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 11 (VK2).

[0209] In another embodiment, the antibodies or antigen-binding fragments described herein (for example for any of the uses or methods described herein) may comprise a VH region which is at least (about) 80%, such as at least (about) 85% identical to any of the SEQ ID No:s of any of the heavy chain variable regions described herein. In another embodiment, the VH region is at least (about) 90%, such as at least (about) 95% identical (or (about) 96%, (about) 97%, or (about) 98% identical) to any of the SEQ ID No:s of any of the heavy chain variable regions described herein.

[0210] In another embodiment, the antibodies or antigen-binding fragments described herein (for example for any of the uses or methods described herein) may comprise a VL region which is at least (about) 80%, such as at least (about) 85% identical to any of the SEQ ID No:s of any of the light chain variable regions described herein. In another embodiment, the VL region is at least (about) 90%, such as at least (about) 95% identical (or (about) 96%, (about) 97%, or (about) 98%) identical to any of the SEQ ID No:s of any of the light chain variable regions described herein.

[0211] Based upon expression, iTopeTM-AI scores and TCEM™ (Jn sih'co tools provided by Abzena that rapidly screen antibodies and other proteins for potential immunogenicity), the above mentioned chimeric VH0 / VK0 antibody (SEQ ID NO: 1 / SEQ ID NO:9) and six humanized antibodies VH4 / VK2 (SEQ ID NO:5 / SEQ ID NO: 11), VH6 / VK3 (SEQ ID NO:6 / SEQ ID NO: 12), VH6 / VK1 (SEQ ID NO:6 / SEQ ID NO: 10), VH6 / VK2 (SEQ ID NO:6 / SEQ ID NO: 11), VH7 / VK2 (SEQ ID NO:7 / SEQ ID NO: 11) and VH8 / VK2 (SEQ ID NO:7 / EQ ID NO: 11) are especially particularised. The variant VH6 / VK3 (SEQ ID NO:6 / SEQ ID NO: 12) has the advantage that there is no isomerisation risk which might occur with the other 5 humanized variants VH4 / VK2 (SEQ ID NO:5 / SEQ ID NO: 11), VH6 / VK1 (SEQ ID NO:6 / SEQ ID NO: 10), VH6 / VK2 (SEQ ID NO:6 / SEQ ID NO: 11), VH7 / VK2 (SEQ ID NO:7 / SEQ ID NO: 11) and VH8 / VK2 (SEQ ID NO:7 / SEQ ID NO: 11). In one aspect, the isolated antibody or antigen-binding fragment thereof binds to human recombinant PD-L2 with a dissociation constant (KD) equal to or less than (about) 10'8M. In another embodiment, the KD may be equal to or less than (about) 10'9M. In another embodiment, the KD may be equal to or less than (about) 10'10M. The KD may be equal to or less than (about) 10'11M. In another embodiment, the KD may be equal to or higher than (about) 10'12M. In another embodiment, the KD may be equal to or higher than (about) 10'13M.

[0212] In another aspect, the isolated antibody or antigen-binding fragment thereof blocks in vitro the interaction between human programmed cell death protein 1 (PD-1) and human PD-L2 with an ECso (half maximal effective concentration) equal to or less than (about) 1.3 nM (such as less than 1 nM, or less than 0.65 nM) optionally in a bioluminescent cell-based assay, optionally using Jurkat T cells expressing human PD-1 and a luciferase reporter driven by an NFAT response element (NFAT- RE) and CHO-K1 cells expressing human PD-L2 and an engineered cell surface protein designed to activate cognate TCRs in an antigen-independent manner. Such antibodies have a high clinical efficacy.

[0213] It was found that the antibodies described herein (ECso = 0.219 nM) bind cellular PD-L2 better than commercial antibodies MIH-18 (ECso = 0.618 nM), 24F.10C12 (ECso = 0.70 nM), and TY25 (0.64 nM). In addition, it was found that VH6 / VK6 has superior blocking activities against cellular hPD- L2 / hPD-l interaction compared to commercial antibodies MIH-18, 24F.10C12, and D7U86.

[0214] In one embodiment, the ECso is equal to or less than (about) 1.5 nM. In one embodiment, the ECso is equal to or less than (about) 1 nM. In one embodiment, the ECso is equal to or less than (about) 0.65 nM. In one embodiment, the ECso is equal to or higher than (about) 0.1 nM. In one embodiment, the ECso is equal to or higher than (about) 0.05 nM.

[0215] Experimental data clearly indicate that the chimeric antibody VH0 / VK0 (SEQ ID NO: 1 / SEQ ID NO:9) and the five humanized antibodies VH6 / VK3 (SEQ ID NO:6 / SEQ ID NO: 12), VH6 / VK1 (SEQ ID NO:6 / SEQ ID NO: 10), VH6 / VK2 (SEQ ID NO:6 / SEQ ID NO: 11), VH7 / VK2 (SEQ ID NO:7 / SEQ ID NO: 11) and VH8 / VK2 (SEQ ID NO:7 / SEQ ID NO: 11) have similar physiological properties regarding the interaction with hPD-L2. In particular, it is shown that the difference of one amino acid at position 3 of CDRH1 (A vs S) and the difference of one amino acid at position 10 (G vs D) of CDRL1 does not negatively impact the physiological properties. Especially good results could be obtained with VH0 / VK0 (SEQ ID NO: 1 / SEQ ID NO:9), VH6 / VK3 (SEQ ID NO:6 / SEQ ID NO: 12), VH6 / VK2 (SEQ ID NO:6 / SEQ ID NO: 11) and VH7 / VK2 (SEQ ID NO:7 / SEQ ID NO: 11). In another aspect, the isolated antibody specifically binds both to human and murine PD-L2.

[0216] Further, it has been shown that parental and humanized variant antibodies have a superior binding activity for recombinant and cellular mPD-L2 than the commercially available anti-mPD-L2 TY- 25. The antibody VH0 / VK0 was found to show cross-reactivity to recombinant mPD-L2 by ELISA, whereas commercially available MIH-18 shows no cross-reactivity to recombinant mPD-L2. The humanised variant VH8 / VK2 also shows cross-reactivity to recombinant mPD-L2. In another aspect, the isolated antibody or antigen-binding fragment thereof binds to human PD-L2 with a dissociation constant (KD) equal to or less than 10'8M (or 10'10M) and at the same time blocks in vitro the interaction between human programmed cell death protein 1 (PD-1) and human PD-L2 with an ECso equal to or less than 1.3 nM, or equal to or less than 1 nM, equal to or less than 0.65 nM, optionally in a bioluminescent cell-based assay and optionally using Jurkat T cells expressing human PD-1 and a luciferase reporter driven by an NFAT response element (NFAT-RE) and CHO-K1 cells expressing human PD-L2 and an engineered cell surface protein designed to activate cognate TCRs in an antigen-independent manner.

[0217] In yet another aspect, the isolated antibody or antigen-binding fragment thereof binds to human PD-L2 with a dissociation constant (KD) equal to or less than 10'8M (or 10'10M) and specifically binds both to human and murine PD-L2 which eases pre-clinical testing.

[0218] In yet another aspect, the isolated antibody or antigen-binding fragment thereof binds to human PD-L2 with a dissociation constant (KD) equal to or less than 10'8M (or 10'10M) and at the same time blocks in vitro the interaction between human Programmed cell death protein 1 (PD-1) and human PD-L2 with an ECso equal to or less than 1.3 nM (such as less than 1 nM, or less than 0.65 nM), optionally in a bioluminescent cell-based assay and optionally using Jurkat T cells expressing human PD-1 and a luciferase reporter driven by an NFAT response element (NFAT-RE) and CHO-K1 cells expressing human PD-L2 and an engineered cell surface protein designed to activate cognate TCRs in an antigen-independent manner and specifically binds both to human and murine PD-L2.

[0219] The antibodies described herein have a superior binding activity against cellular hPD-L2 and blocking activity against the cellular hPD-l / hPD-L2 interaction compared to a panel of commercially available anti-PD-L2 antibodies. Furthermore, the antibodies described herein showed a higher blocking activity than MIH-18, 24F.10C12, D7U86 and associated isotype negative controls against cellular hPD-L2 / hPD-l interaction as measured by fluorescent reporter in hPD-1 expressing Jurkat cells co-cultured with hPD-L2 expressing CHOK-1 cells. In still another embodiment, the isolated monoclonal antibody or antigen-binding fragment thereof is chimeric, humanized or human.

[0220] In one embodiment, the antibody comprises a heavy chain constant region in the wild-type format of the IgG isotype, specifically of the human IgGl or the murine IgG2a subclass. According to these embodiments, the heavy chain constant region of the antibody may comprise the amino acid sequence of SEQ ID NO:21 (mIgG2a) or of SEQ ID NO:22 (hlgGl) and according to a particular embodiment, the heavy chain thus comprises the amino acid sequence of SEQ ID NO:23 (VH0 / VK0- mIgG2a) or of SEQ ID NO:24 (VH6 / VK3-hIgGl). Both the wild-type and LALA heavy chains in the hlgGl formats may optionally comprise a terminal lysine (not shown in SEQ ID NO:22, SEQ ID NO:24, SEQ ID NO:26 and SEQ ID NO:28).

[0221] Table 3: HC constant region sequences (effector enabled)

[0222] According to a particular embodiment, the antibody comprises a constant region sequence which has reduced or no Fc receptor (FcyR) interactions, or reduced or no immune effector functions. In some embodiments, the constant region comprises no naturally occurring FcyR interactions. In some embodiments, the constant region comprises no naturally occurring immune effector functions. In some embodiments, the antibody comprises a mutagenized Fc variant sequence of the IgG isotype exhibiting reduced Fc receptor (FcyR) interaction (or reduced immune effector function) compared to its parent, wild type Fc region.

[0223] The variant antibody may comprise a heavy chain constant region which has (i) a wild-type constant region sequence which has reduced or no Fc receptor (FcyR) interactions, (ii) a wild-type constant region sequence which has reduced or no immune effector functions, or (iii) a mutagenized Fc variant sequence of the IgG isotype exhibiting reduced Fc receptor (FcyR) interaction or reduced immune effector function compared to the parent Fc region, and for part (iii), wherein the isolated monoclonal antibody is a chimeric, humanized or human antibody. Optionally, the antibody is a chimeric, humanized or human antibody.

[0224] As mentioned above, a specific variant antibody in the context of this application is referred to as an effector-reduced anti-PD-L2 antibody. Surprisingly, these constant regions have a relatively long half-life, which may lead to improvements in therapy, for example by reduced frequency of dosing and / or fewer side effects, as will be shown in the context of the Examples below.

[0225] Specifically, a variant antibody according to this embodiment may comprise a mutagenized Fc variant sequence comprising a mutation of leucine 234 to alanine and leucine 235 to alanine (LALA mutation) according to EU numbering, which has been found to be particularly useful owed to the variant's high therapeutic efficacy in the mouse fibrosis model of Example 7 below. According to a particular embodiment, the heavy chain constant region thus comprises the amino acid sequence of SEQ ID NO:25 (mIgG2a-LALA) or of SEQ ID NO:26 (hlgGl-LALA) and according to a particular embodiment, the heavy chain thus comprises the amino acid sequence of SEQ ID NO:27 (VHO / VKO- mIgG2a-LALA format) or of SEQ ID NO:28 (VH6 / VK3-hIgGl-LALA format).

[0226] Other constant regions which have reduced effector functions include mutagenized constant regions of the human IgG4 class, and e.g. comprises a Leu235Glu mutation (i.e. where the wild type leucine residue is mutated to a glutamic acid residue). In another embodiment, the heavy chain constant region comprises a Ser228Pro mutation to increase stability. In another embodiment, the heavy chain constant region is a human IgG4 comprising both the Leu235Glu mutation and the Ser228Pro mutation (commonly referred to as "IgG4-PE"). Other such mutagenised constant regions are described in Wilkinson et al. "Fc-engineered antibodies with immune effector functions completely abolished", PLOS One, 16(2), e0260954, 2021, doi:https: / / doi.org / 10.1371 / journal.pone.0260954, which is incorporated herein by reference in its entirety. Any of these constant regions may be used with the variable regions described herein, and it is expected that they will have similarly high efficacy. Table 4: HC constant regions (effector disabled)

[0227] However, there may be no difference in the light chain constant region of the antibody. In one embodiment, the light chain constant region thus comprises the amino acid sequence of SEQ ID NO:29 or SEQ ID NO:30. In a particular embodiment, the light chain thus comprises the amino acid sequence of SEQ ID NO:31 (VHO / VKO LALA and wild-type) or of SEQ ID NO:32 (VH6 / VK3 LALA and wild -type).

[0228] Table 5: LC constant region sequences

[0229] As mentioned above, the disclosure relates to an antibody with a wild-type heavy chain constant region and an antibody with a variant heavy chain constant region for use in treating (or for ameliorating or preventing) a fibrotic disease, in particular in a mammal.

[0230] As will be shown in the context of the Examples (in particular, Figures 7A to 7F and 8A to 8D), an anti-PD-L2 antibody having a wild-type heavy chain constant region as well as having a variant heavy chain constant region have in particular been found to be effective in the treatment of a pulmonary fibrosis. Specifically, an antibody having a wild-type heavy chain constant region as well as an antibody having a variant heavy chain constant region displayed a significant decrease in the %- CPA and in hydroxyproline levels (which as in index of a collagen proliferation) compared to the respective disease control group. In the specific case of pulmonary fibrosis, a decrease in the Ashcroft score, of the lung weight and of the respective inflammation score was determined in comparison to the disease control group.

[0231] According to a specific embodiment, there is provided the antibody or antigen-binding fragment thereof defined elsewhere herein (e.g. above) for use in treating (or for ameliorating or preventing) pulmonary fibrosis, in particular in Idiopathic Pulmonary Fibrosis (IPF). With regard to the treatment described herein, the antibody or fragment may be administered in combination with a further therapeutically active agent, in particular a therapeutically active agent selected from the group consisting of pirfenidone, nintedanib, hydronidone, luspatercept, pamrevlumab, parsaclisib, fedratinib, momelotinib, pacritinib, jaktinib, saroglitazar, lanifibranor, obeticholic acid, semaglutide, elexacaftor, ivacaftor, resmetirom, belapectin, azemiglitazone potassium, aramchol, imetelstat, KRT-232, ruxolitinib and panobinostat and combinations thereof.

[0232] Additionally or alternatively, the antibody or fragment may be administered in combination with a therapeutically active agent selected from the group consisting of HEC-585, PLN-74809, BG00011, Sotatercept, BI089-100, Efruxifermin, Pegbelfermin, Aldafermin, ZSP1603, MK-3655, Itacitinib, Ilginatinib, SM04646, PRI-724, CC-90001, Epeleuton, Tipelukast, PAT-1251, Pemafibrate, Cilofexor, Nidufexor, TERN-101, Vonafexor, EDP-305, Tropifexor, JKB-121, JKB-122, Tirzepatide, Cotadutide, IDL-2965, TRK-250, AVID200, Omipalisib, HEC-68498, Buparlisib, Umbralisib, MG-S-2525, PXS-5382A, ZSP0678, FDL176, Posenacaftor, GLPG2451, PU-H71, IQNIS-ENaCRx, AZD5634, BI 443651, Idelalisib, CHF 6333, Ribociclib, ARO-HSD, CB4211, Interferon gamma, BBT-877, ZL-2102, CB-280, N-6022, PRT-543 and combinations thereof.

[0233] Additionally or alternatively, the antibody or fragment may be administered in combination with a therapeutically active agent selected from the group consisting of RXC007, Tezacaftor, VX-121, LYT-100, CSL312, BI 1015550, Rencofilstat, ZED1227, LASN01, Metformin, Treprostinil, HCC 45, HTD1801, Duplimab, BAX602, Allopurinol, ASC42, Sacroglitizar magnesium, CNP-104, Setanaxib, EP547, Apibaxan, OCE-205 and Efzofitimod.

[0234] According to further aspect, there is provided an isolated monoclonal antibody having a variant heavy chain constant region (i.e. effector reduced or effector-disabled), as mentioned above. In particular, this isolated monoclonal antibody having a variant heavy chain constant region is chimeric, humanized or human.

[0235] There is provided an isolated variant monoclonal antibody that specifically binds to programmed cell death ligand 2 (PD-L2) and blocks activity against PD-L2 / hPD-l interaction, wherein said variant antibody comprises six CDR sequences in the heavy chain variable region and in the light chain variable region, and comprises a) as heavy chain variable region CDRs a CDRH1 comprising the amino acid sequence of SEQ ID NO:33 (NYFIE, according to Kabat numbering) or of SEQ ID NO:34 (GYAFSNY, according to Chothia numbering) or alternatively SEQ ID NO:36 (GYSFSNY, according to Chothia numbering), in particular comprising the amino acid sequence of SEQ ID NO: 13 (GYAFSNYFIE) or SEQ ID NO: 14 (GYSFSNYFIE); a CDRH2 having the amino acid sequence of SEQ ID NO: 15 (LNIPGSGGSNYAEKFKG, according to Kabat numbering) or of SEQ ID NO:35 (IPGSGG, according to Chothia numbering); and a CDRH3 having the amino acid sequence of SEQ ID NO: 16 (RRLPPDWYFDV according to both Kabat and Chothia numbering); and b) as light chain variable region CDRs a CDRL1 having the amino acid sequence of SEQ ID NO: 17 (RSSQSLVHSGGNTYLH, according to Kabat and Chothia numbering) or SEQ ID NO:20 (RSSQSLVHSDGNTYLH, according to both Kabat and Chothia numbering); a CDRL2 having the amino acid sequence of SEQ ID NO: 18 (KVSNRFS, according to both Kabat and Chothia numbering); and a CDRL3 having the amino acid sequence of SEQ ID NO: 19 (SQSTHVPWT, according to both Kabat and Chothia numbering), and wherein the variant antibody comprises (i) a wild-type constant region sequence which has reduced or no Fc receptor (FcyR) interactions, (ii) a wild-type constant region sequence which has reduced or no immune effector functions, or (iii) a mutagenized Fc variant sequence of the IgG isotype exhibiting reduced Fc receptor (FcyR) interaction (or reduced immune effector function) compared to its parent, wild type Fc region, and for part (iii), wherein the isolated monoclonal antibody is chimeric, humanized or human. Specifically, the parent Fc region thereby relates to the wild-type sequence of the respective IgG format. Optionally, the isolated monoclonal antibody is chimeric, humanized or human.

[0236] In embodiments relating to variant antibodies perse, in one embodiment, the constant region is not an effector-reduced or an effector disabled mouse constant region. In another embodiment, the constant region is not a mutagenized Fc variant sequence of murine IgG2a subclass exhibiting reduced Fc receptor (FcyR) interaction compared to its parent, wild type Fc region. In a further embodiment, the constant region is not a murine IgG2a constant region comprising a mutation of leucine 234 to alanine and leucine 235 to alanine (LALA mutation) according to EU numbering. Additionally or alternatively, the constant region is not a murine IgG2a constant region comprising a mutation of leucine 234 to alanine, leucine 235 to alanine, and proline 329 to glycine (LALAPG mutation) according to EU numbering. When the constant region is not a murine IgG2a constant region comprising a mutation of leucine 234 to alanine and leucine 235 to alanine (LALA mutation) according to EU numbering, the constant region may not be a constant region comprising the amino acid sequence of SEQ ID No:25.

[0237] In embodiments relating to variant antibodies per se, in one embodiment the antibody may comprise (i) a mutagenized Fc variant sequence of human IgGl subclass subclass exhibiting reduced Fc receptor (FcyR) interaction compared to its parent, wild type Fc region, or (ii) a mutagenized Fc variant sequence of murine IgG2a subclass exhibiting reduced Fc receptor (FcyR) interaction compared to its parent, wild type Fc region wherein the mutagenized Fc variant sequence of murine IgG2a does not comprise a mutation of leucine 234 to alanine and leucine 235 to alanine (LALA mutation) according to EU numbering, or a mutation of leucine 234 to alanine, leucine 235 to alanine, and proline 329 to glycine (LALAPG mutation) according to EU numbering. Surprisingly, also an effector-reduced anti-PD-L2 antibody has been found to exhibit a high efficacy in the treatment of subjects suffering fibrosis in a dose-dependent manner, as mentioned above. In addition, data obtained by studies in a mouse model support the finding that an effector- reduced anti-PD-L2 antibody also exhibits a clear effect in the treatment of other disorders, such as a disease that is driven by the accumulation of senescent cells (e.g. pathologically accelerated senescence) or premature aging, in particular progeria.

[0238] Specifically, a significant increase of the median lifespan (i.e. the timespan relating to a 50% survival rate) was observed for mice treated with an anti-PD-L2 antibody as defined above compared to a control IgG antibody. Anti-PD-L2 and IgG control -treated mice of both sexes combined showed a median lifespan of 137 days and 121 days respectively. Anti-PD-L2 and IgG control-treated females mice showed a median lifespan of 159.5 days versus 121 days, respectively. Moreover, pharmacokinetic studies revealed a further increased half-life and AUC as well as a further decreased clearance of effector-reduced anti-PD-L2 antibody in comparison to wild-type anti-PD-L2, as shown in Figure 3. Hence, the therapeutic potential of effector-reduced anti-PD-L2 antibodies in vivo is even higher than for wild-type anti-PD-L2. Without being bound by theory, it is expected that any constant region having reduced or no effector functions may possess similar advantages.

[0239] There is provided a pharmaceutical composition comprising an isolated monoclonal antibody or antigen-binding fragment thereof described herein and a pharmaceutically acceptable excipient.

[0240] There is also provided an isolated nucleic acid sequence that encodes the amino acid sequence of the light chain variable region and / or the heavy chain variable region of an anti-PD-L2 antibody or antigen-binding fragment thereof described herein. A nucleic acid is "isolated" or "rendered substantially pure" when purified away from other cellular components or other contaminants, e.g., other cellular nucleic acids or proteins, by standard techniques, including alkaline / SDS treatment, CsCI banding, column chromatography, agarose gel electrophoresis and others well known in the art.

[0241] There is also provided a recombinant expression vector comprising such a nucleic acid sequence.

[0242] There is also provided an isolated host cell which comprises a recombinant expression vector described herein and / or expresses an antibody or antigen-binding fragment described herein.

[0243] The present application additionally relates to the following items:

[0244] 1. An isolated monoclonal antibody, or antigen binding fragment thereof, that specifically binds to programmed cell death ligand 2 (PD-L2) and blocks activity against PD-L2 / PD1 interaction for use in the treatment of a fibrotic disease in a mammal, wherein the antibody

[0245] (a) binds to human PD-L2 with a dissociation constant (KD) equal from (about) 10'8M to (about) 10'12M; and / or

[0246] (b) blocks in vitro the interaction between human programmed cell death protein 1 (hPD-1) and human PD-L2 with an ECso of from (about) 0.1 to (about) 1.5 nM, optionally in a bioluminescent cell-based assay, and optionally using Jurkat T cells expressing human PD-1 and a luciferase reporter driven by an NFAT response element (NFAT-RE) and CHO-K1 cells expressing human PD-L2 and an engineered cell surface protein designed to activate cognate TCRs in an antigen-independent manner; and / or

[0247] (c) specifically binds to both human and murine PD-L2; and / or

[0248] (d) competes for binding to PD-L2 with an antibody having a heavy chain variable region having an amino acid sequence of SEQ ID No:l and a light chain variable region having an amino acid sequence of SEQ ID No:9, optionally in an ELISA, SPR or cell-based assay.

[0249] 2. The antibody or fragment according to item 1, wherein the antibody further

[0250] (e) comprises a heavy chain constant region which has reduced or no Fc receptor (FcyR) interactions, or reduced or no immune effector functions.

[0251] 3. The antibody or fragment according to item 1 or item 2, which comprises each of features a) and b).

[0252] 4. The antibody or fragment according any one of the preceding items, wherein the antibody promotes immune-mediated clearance of senescent cells in fibrotic tissue.

[0253] 5. The antibody or fragment according to any one of the preceding items, whose administration to a subject suffering from fibrosis, in particular from IPF, results as compared to the respective disease control group

[0254] A. in a decrease in the %-CPA (collagen proportionate area), in particular a decrease by at least (about) 5%, at least (about) 10%, at least (about) 15%, at least (about) 20%, or at least (about) 25%, at least (about) 30%, at least (about) 35%, or at least (about) 40%; and / or

[0255] B. in a decrease in the hydroxyproline levels, in particular a decrease by at least (about) 5%, at least (about) 10%, at least (about) 15%, at least (about) 20%, at least (about) 25%, at least (about) 30%, at least (about) 35%, or at least (about) 38%; and / or

[0256] C. in a decrease in the Ashcroft score as compared to the disease control group, in particular a decrease by at least 0.5 points, at least 0.6 points, at least (about) 0.7 points, at least (about) 0.8 points, at least (about) 0.9 points, at least (about) 1 point, at least (about) 1.1 points, at least (about) 1.2 points, at least (about) 1.3 points, or at least (about) 1.4 points; and / or

[0257] D. in a decrease of the inflammation score, in particular a decrease by at least (about) 0.5 points, at least (about) 0.6 points, at least (about) 0.7 points, at least (about) 0.8 points, at least (about) 0.9 points, at least (about) 1 point, at least (about) 1.1 points, at least (about) 1.2 points, or at least (about) 1.3 points; and / or E. in a decrease in pl6 expression by at least (about) 10%, at least (about) 15%, at least (about) 20%, at least (about) 25%, at least (about) 30%, at least (about) 35%, or at least (about) 40%; and / or

[0258] F. in a decrease in p21 expression by at least (about) 10%, at least (about) 15%, at least (about) 20%, at least (about) 25%, at least (about) 30%, at least (about) 35%, or at least (about) 40%.

[0259] 6. The antibody or fragment according to item 5, wherein the subject is a mouse and the fibrosis is a bleomycin-induced model of IPF.

[0260] 7. The antibody or fragment according to any one of the preceding items, wherein the antibody is a humanised or chimeric antibody whose three heavy chain CDRs are derived from a murine IGHV gene segment which is IGHV1-54 (SEQ ID No:37), a murine IGHD gene segment, and murine IGHJ gene segment which is IGHJ1 (SEQ ID No: 38), and whose three light chain CDRs are derived from a murine IGKV gene segment which is IGKV1-110 (SEQ ID No:39) and a murine IGKJ gene segment which is IGKJ1 (SEQ ID No:40).

[0261] 8. The antibody or fragment according to any one of the preceding items, wherein said antibody comprises three CDR sequences in the heavy chain variable region, and three CDR sequences in the light chain variable region, and comprises i. as heavy chain variable region CDRs a CDRH1 comprising the amino acid sequence of SEQ ID NO:33 (NYFIE, according to Kabat numbering) or of SEQ ID NO:34 (GYAFSNY, according to Chothia numbering) or alternatively SEQ ID NO:36 (GYSFSNY, according to Chothia numbering), in particular comprising the amino acid sequence of SEQ ID NO: 13 (GYAFSNYFIE) or SEQ ID NO: 14 (GYSFSNYFIE); a CDRH2 comprising the amino acid sequence of SEQ ID NO: 15 (LNIPGSGGSNYAEKFKG, according to Kabat numbering) or of SEQ ID NO:35 (IPGSGG, according to Chothia numbering); and a CDRH3 comprising the amino acid sequence of SEQ ID NO: 16 (RRLPPDWYFDV, according to both Kabat and Chothia numbering); and ii. as light chain variable region CDRs a CDRL1 comprising the amino acid sequence of SEQ ID NO: 17 (RSSQSLVHSGGNTYLH, according to both Kabat and Chothia numbering) or SEQ ID NO:20 (RSSQSLVHSDGNTYLH, according to both Kabat and Chothia numbering); a CDRL2 comprising the amino acid sequence of SEQ ID NO: 18 (KVSNRFS, according to both Kabat and Chothia numbering); and a CDRL3 comprising the amino acid sequence of SEQ ID NO: 19 (SQSTHVPWT, according to both Kabat and Chothia numbering), or which comprises the CDR sequences of a) and b) comprising one or two, in particular one, amino acid substitutions, deletions or additions in one or more of the CDR sequences.

[0262] 9. The antibody or fragment according to item 8, which comprises: i. as heavy chain variable region CDRs a CDRH1 comprising the amino acid sequence of SEQ ID NO:33 (NYFIE, according to Kabat numbering) or of SEQ ID NO:36 (GYSFSNY, according to Chothia numbering), in particular comprising the amino acid sequence of SEQ ID NO: 14 (GYSFSNYFIE); a CDRH2 comprising the amino acid sequence of SEQ ID NO: 15 (LNIPGSGGSNYAEKFKG, according to Kabat numbering) or of SEQ ID NO:35 (IPGSGG, according to Chothia numbering); and a CDRH3 comprising the amino acid sequence of SEQ ID NO: 16 (RRLPPDWYFDV, according to both Kabat and Chothia numbering); and ii. as light chain variable region CDRs a CDRL1 comprising the amino acid sequence of SEQ ID NO: 17 (RSSQSLVHSGGNTYLH, according to both Kabat and Chothia numbering); a CDRL2 comprising the amino acid sequence of SEQ ID NO: 18 (KVSNRFS, according to both Kabat and Chothia numbering); and a CDRL3 comprising the amino acid sequence of SEQ ID NO: 19 (SQSTHVPWT, according to both Kabat and Chothia numbering), or which comprises the CDR sequences of a) and b) comprising one or two, in particular one, amino acid substitutions, deletions or additions in one or more of the CDR sequences.

[0263] 10. The antibody or fragment according to item 8, which comprises: i. as heavy chain variable region CDRs a CDRH1 comprising the amino acid sequence of SEQ ID NO:33 (NYFIE, according to Kabat numbering) or of SEQ ID NO:34 (GYAFSNY, according to Chothia numbering), in particular comprising the amino acid sequence of SEQ ID NO: 13 (GYAFSNYFIE); a CDRH2 comprising the amino acid sequence of SEQ ID NO: 15 (LNIPGSGGSNYAEKFKG, according to Kabat numbering) or of SEQ ID NO:35 (IPGSGG, according to Chothia numbering); and a CDRH3 comprising the amino acid sequence of SEQ ID NO: 16 (RRLPPDWYFDV, according both to Kabat and Chothia numbering); and ii. as light chain variable region CDRs a CDRL1 comprising the amino acid sequence of SEQ ID NO:20 (RSSQSLVHSDGNTYLH, according to both Kabat and Chothia numbering); a CDRL2 comprising the amino acid sequence of SEQ ID NO: 18 (KVSNRFS, according to both Kabat and Chothia numbering); and a CDRL3 comprising the amino acid sequence of SEQ ID NO: 19 (SQSTHVPWT, according to both Kabat and Chothia numbering), or which comprises the CDR sequences of a) and b) comprising one or two, in particular one, amino acid substitutions, deletions or additions in one or more of the CDR sequences. The antibody or fragment according to any one of the preceding items, wherein

[0264] I. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO:1 (VHO) and the light chain variable region comprises the amino acid sequence of SEQ ID NO:9 (VKO); or

[0265] II. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO:6 (VH6) and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 12 (VK3), preferably, the heavy chain variable region comprises the amino acid sequence of SEQ ID NO:6 (VH6) and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 12 (VK3). The antibody according to any one of item 1 or items 3 to 11, wherein the antibody comprises a heavy chain constant region which comprises the amino acid sequence of SEQ ID NO:21 or of SEQ ID NO:22. The antibody according to any one of items 1 to 11, wherein the antibody comprises a heavy chain constant region which has reduced or no Fc receptor (FcyR) interactions, or reduced or no immune effector functions, and in particular comprises a mutagenized Fc variant sequence of an IgG isotype exhibiting reduced Fc receptor (FcyR) interaction or reduced immune effector function compared to its parent, wild type Fc region. The antibody according to item 13, wherein the antibody comprises a mutagenized Fc variant sequence of murine IgG2a subclass exhibiting reduced Fc receptor (FcyR) interaction compared to its parent, wild type Fc region, optionally wherein the mutagenized Fc variant sequence comprises a mutation of leucine 234 to alanine and leucine 235 to alanine (LALA mutation) according to EU numbering, and further optionally wherein the heavy chain constant region comprises the amino acid sequence of SEQ ID NO:25. An isolated variant monoclonal antibody which specifically binds to programmed cell death ligand 2 (PD-L2) and blocks activity against PD-L2 / PD-1 interaction, wherein said variant antibody comprises three CDR sequences in the heavy chain variable region and three CDRs in the light chain variable region, and comprises i. as heavy chain variable region CDRs a CDRH1 comprising the amino acid sequence of SEQ ID NO:33 (NYFIE, according to Kabat numbering) or of SEQ ID NO:34 (GYAFSNY, according to Chothia numbering) or alternatively SEQ ID NO:36 (GYSFSNY, according to Chothia numbering), in particular comprising the amino acid sequence of SEQ ID NO: 13 (GYAFSNYFIE) or SEQ ID NO: 14 (GYSFSNYFIE); a CDRH2 comprising the amino acid sequence of SEQ ID NO: 15 (LNIPGSGGSNYAEKFKG, according to Kabat numbering) or of SEQ ID NO:35 (IPGSGG, according to Chothia numbering); and a CDRH3 comprising the amino acid sequence of SEQ ID NO: 16 (RRLPPDWYFDV, according to both Kabat and Chothia numbering); and ii. as light chain variable region CDRs a CDRL1 comprising the amino acid sequence of SEQ ID NO: 17 (RSSQSLVHSGGNTYLH, according to both Kabat and Chothia numbering) or SEQ ID NO:20 (RSSQSLVHSDGNTYLH, according to both Kabat and Chothia numbering); a CDRL2 comprising the amino acid sequence of SEQ ID NO: 18 (KVSNRFS, according to both Kabat and Chothia numbering); and a CDRL3 comprising the amino acid sequence of SEQ ID NO: 19 (SQSTHVPWT, according to both Kabat and Chothia numbering), or which variant antibody comprises the CDR sequences of a) and b) comprising one or two, in particular one, amino acid substitutions, deletions or additions in one or more of the CDR sequences, and wherein the variant antibody comprises a heavy chain constant region which has (i) a wildtype constant region sequence which has reduced or no Fc receptor (FcyR) interactions, (ii) a wild-type constant region sequence which has reduced or no immune effector functions, or (iii) a mutagenized Fc variant sequence of the IgG isotype exhibiting reduced Fc receptor (FcyR) interaction or reduced immune effector function compared to the parent Fc region, and for part (iii), wherein the isolated monoclonal antibody is a chimeric, humanized or human antibody.

[0266] 16. The antibody according to item 15, which is a chimeric, humanized or human antibody.

[0267] 17. The antibody according to item 15 or item 16, which comprises the CDR combinations as defined in item 9 or item 10; or which comprises the VH and VL sequences as defined in item 11.

[0268] 18. The antibody as defined in any one of items 15 to 17 for use in treating a fibrotic disease. 19. The antibody according to any one of the preceding items, wherein the antibody comprises (i) a mutagenized Fc variant sequence of human IgGl subclass subclass exhibiting reduced Fc receptor (FcyR) interaction compared to its parent, wild type Fc region, or (ii) a mutagenized Fc variant sequence of murine IgG2a subclass exhibiting reduced Fc receptor (FcyR) interaction compared to its parent, wild type Fc region wherein the mutagenized Fc variant sequence of murine IgG2a does not comprise a mutation of leucine 234 to alanine and leucine 235 to alanine (LALA mutation) according to EU numbering, or a mutation of leucine 234 to alanine, leucine 235 to alanine, and proline 329 to glycine (LALAPG mutation) according to EU numbering.

[0269] 20. The antibody according to item 19(i), wherein the mutagenized Fc variant sequence comprises a mutation of leucine 234 to alanine and leucine 235 to alanine (LALA mutation) according to EU numbering.

[0270] 21. The antibody according to item 20, wherein the heavy chain constant region comprises the amino acid sequence of SEQ ID NO: 26.

[0271] 22. The antibody or fragment according to any one of the preceding items, wherein the light chain constant region comprises the amino acid sequence of SEQ ID NO: 29 or of SEQ ID NO: 30.

[0272] 23. The antibody or fragment according to any one of the preceding items for use in treating, ameliorating or preventing pulmonary fibrosis, in particular in Idiopathic Pulmonary Fibrosis (IPF).

[0273] 24. The antibody or fragment according to any one of the preceding items for use in treating a fibrotic disease selected from lung fibrosis, liver fibrosis, in particular selected from non-alcoholic steatohepatitis (NASH) and non-alcoholic fatty liver disease, kidney fibrosis and heart fibrosis.

[0274] 25. The antibody or fragment according to any one of the preceding items, wherein the antibody is administered in combination with a further therapeutically active agent, in particular a therapeutically active agent selected from the group consisting of pirfenidone, nintedanib hydronidone, luspatercept, pamrevlumab, parsaclisib, fedratinib, momelotinib, pacritinib, jaktinib, saroglitazar, lanifibranor, obeticholic acid, semaglutide, elexacaftor, ivacaftor, resmetirom, belapectin, azemiglitazone potassium, aramchol, imetelstat, KRT-232, ruxolitinib and panobinostat and combinations thereof.

[0275] 26. A composition comprising the antibody or fragment as defined in any one of the preceding items in combination with a pharmaceutically acceptable excipient.

[0276] 27. An isolated nucleic acid sequence that encodes the amino acid sequence of the antibody or fragment as defined in any one of items 1 to 22.

[0277] 28. A recombinant expression vector comprising the nucleic acid of item 27. An isolated host cell comprising the recombinant expression vector of item 28. An isolated monoclonal antibody or antigen-binding fragment thereof that specifically binds to programmed cell death ligand 2 (PD-L2) and blocks activity against PD-L2 / PD1 interaction for use in the treatment of a fibrotic disease in a mammal, wherein the antibody is capable of preventing the conversion of fibroblasts to myofibroblasts in an in vitrolPV- model as measured by a reduction in a-smooth muscle actin 1 (smal) positive fibres. A method of treating or reducing the risk of a fibrotic disease, said method comprising administering an effective amount of an anti-PD-L2 antibody or antigen-binding fragment thereof to a subject in need thereof, wherein the anti-PD-L2 antibody or fragment is capable of preventing the conversion of fibroblasts to myofibroblasts in an in vitro W model as measured by a reduction in a-smooth muscle actin 1 (smal) positive fibres, and whereby the fibrotic disease is treated or the risk of the fibrotic disease is reduced. A method of preventing the formation of myofibroblasts, said method comprising contacting an anti-PD-L2 antibody or antigen-binding fragment thereof with a population of cells comprising fibroblasts, wherein the anti-PD-L2 antibody or fragment is capable of preventing the conversion of fibroblasts to myofibroblasts, in particular as measured by a reduction in a-smooth muscle actin 1 (smal) positive fibres in an in vitro W model. The antibody or fragment according to item 30, or the method according to item 31 or item 32, wherein the reduction in smal positive fibres is at least (about) 40%, or at least (about) 50%, in particular at least (about) 60%, or at least (about) 70%, compared to a TGF-0 treated control sample. The antibody or fragment according to item 30 or item 33, or the method according to any one of items 31 to 33, wherein the reduction in smal positive fibres is measured by a method comprising the steps of:

[0278] - providing fibroblasts from a human donor;

[0279] - treating the fibroblasts with TGF-0;

[0280] - contacting the antibody or fragment with the treated fibroblasts;

[0281] - incubating the fibroblasts in the presence of the TGF-0 and the antibody or fragment;

[0282] - conducting immunofluorescence staining against smooth muscle actin 1 (smal); and

[0283] - quantifying smal positive fibers, in particular by fluorescence-activated cell sorting (FACS). An isolated monoclonal antibody or antigen-binding fragment thereof that specifically binds to programmed cell death ligand 2 (PD-L2) and blocks activity against PD-L2 / PD1 interaction for use in the treatment of a fibrotic disease in a mammal, wherein (i) the antibody or fragment is capable of reprogramming M2 pro-fibrotic macrophages expressing CD206 on the surface of said macrophage into Ml phenotypic macrophages expressing CD86 on the surface of said macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages; and / or

[0284] (ii) the antibody or fragment is capable of increasing CD86 expression on a macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages; and / or

[0285] (iii) the antibody or fragment is capable of decreasing CD206 expression on a macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages; and / or

[0286] (iv) the antibody or fragment is capable of reducing TGF-0 production in a macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages; and / or

[0287] (v) the antibody or fragment is capable of increasing TNF-a production in a macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages; and / or

[0288] (vi) the antibody or fragment is capable of changing the morphology of the M2 macrophages from an elongated morphology to a rounded morphology, in particular as observed by microscopic analysis of cells in an in vitro model of M2 pro-fibrotic macrophages; and / or

[0289] (vii) the antibody or fragment is capable of increasing the proliferation of macrophages, in particular as measured by number of cells in an in vitro model of M2 pro-fibrotic macrophages. A method of treating or reducing the risk of a fibrotic disease, said method comprising administering an effective amount of an anti-PD-L2 antibody or antigen-binding fragment thereof to a subject in need thereof, wherein

[0290] (i) the antibody or fragment is capable of reprogramming M2 pro-fibrotic macrophages expressing CD206 on the surface of said macrophage into Ml phenotypic macrophages expressing CD86 on the surface of said macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages; and / or

[0291] (ii) the antibody or fragment is capable of increasing CD86 expression on a macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages; and / or

[0292] (iii) the antibody or fragment is capable of decreasing CD206 expression on a macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages; and / or

[0293] (iv) the antibody or fragment is capable of reducing TGF-0 production in a macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages; and / or

[0294] (v) the antibody or fragment is capable of increasing TNF-a production in a macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages; and / or

[0295] (vi) the antibody or fragment is capable of changing the morphology of the M2 macrophages from an elongated morphology to a rounded morphology, in particular as observed by microscopic analysis of cells in an in vitro model of M2 pro-fibrotic macrophages; and / or (vii) the antibody or fragment is capable of increasing the proliferation of macrophages, in particular as measured by number of cells in an in vitro model of M2 pro-fibrotic macrophages, and whereby the fibrotic disease is treated or the risk of the fibrotic disease is reduced. A method of reprogramming M2 pro-fibrotic macrophages to Ml phenotypic macrophages, said method comprising contacting an anti-PD-L2 antibody or antigen-binding fragment thereof with said M2 macrophages, wherein the anti-PD-L2 antibody or fragment decreases the level (or amount) of the M2 pro-fibrotic macrophages and increases the level (or amount) of the Ml phenotypic macrophages, in particular as measured by:

[0296] (i) observing an increase in CD86 expression on the macrophages; and / or

[0297] (ii) observing a decrease in CD206 expression on the macrophages; and / or

[0298] (iii) observing a reduction in TGF-0 production on the macrophages; and / or

[0299] (iv) observing an increase in TNF-a production on the macrophages; and / or

[0300] (v) observing a change in morphology of the M2 macrophages from an elongated morphology to a rounded morphology; and / or

[0301] (vi) observing an increase in the proliferation of macrophages. The antibody or fragment according to item 35, or the method according to item 36 or item 37, wherein, independently from each other,

[0302] - the increase in CD86 expression according to item 35(H), item 36(H) or item 37(i) is an increase of at least (about) 40%, or at least (about) 45%, or at least (about) 50% or, in particular at least (about) 55%, compared to an untreated M2 macrophage; and / or

[0303] - the decrease in CD206 expression according to item 35(iii), item 36(iii) or item 37(H) is a decrease of at least (about) 50%, or at least (about) 60%, or in particular at least (about) 70%, compared to an untreated M2 macrophage; and / or

[0304] - the reduction in TGF-0 production according to item 35(iv), item 36(iv) or item 37(iii) is a reduction of at least (about) 30%, or at least (about) 40%, in particular at least (about) 50%, compared to an untreated M2 macrophage; and / or

[0305] - the increase in TNF-a production according to item 35(v), or item 36(v) or item 37(iv) is an increase of at least (about) 5-fold, or at least (about) 5.5-fold, or at least (about) 6-fold, and in particular at least (about) 6.5-fold, compared to an untreated M2 macrophage; and / or.

[0306] - the increase in the proliferation of macrophages according to item 35(vii), or item 36(vii) or item 37(v) is an increase of at least (about) 1.4-fold, or at least (about) 1.5-fold, or at least (about) 1.6-fold, and in particular at least (about) 1.7-fold, compared to untreated M2 macrophages. The antibody or fragment according to item 35 or item 38, or the method according to any one of items 36 to 38, wherein the in vitro model of M2 pro-fibrotic macrophages comprises the steps of

[0307] - providing monocytes from a human donor;

[0308] - treating the monocytes with the cytokines M-CSF1 and IL-4 to polarise the monocytes to M2 macrophages;

[0309] - contacting the antibody with the M2 macrophages;

[0310] - incubating the M2 macrophages in the presence of the cytokines and the antibody or fragment; and

[0311] - for observing an increase in CD86 and / or a decrease in CD206, conducting immunofluorescence staining against CD86 and / or CD206; and quantifying expression of CD86 and / or CD206, in particular by flow cytometry;

[0312] - and / or for observing a reduction in TGF-0 and / or increase in TNF-a, conducting an ELIZA assay to quantify the amount of TGF-0 and / or TNF-a;

[0313] - and / or for observing an increase in the proliferation of macrophages counting the number of macrophages on a cell counter. The antibody or fragment according to any one of items 30, 33 to 35, or 38 or 39, or the method according to any one of items 31 to 34, 36 or 37 to 39, wherein the anti-PD-L2 antibody or fragment is as defined in any one of items 1 to 22, and optionally wherein the fibrotic disease is as defined in item 23 or item 24 and further optionally wherein the antibody or fragment is administered in combination with a therapeutically active agent as defined in item 25. A method of treating or reducing the risk of a fibrotic disease, said method comprising administering an effective amount of an anti-PD-L2 antibody or antigen-binding fragment thereof to a subject in need thereof, wherein the anti-PD-L2 antibody or fragment is as defined in any one of items 1 to 22, or 30 to 39, and whereby the fibrotic disease is treated or the risk of the fibrotic disease is reduced, and optionally wherein the fibrotic disease is as defined in item 23 or item 24 and further optionally wherein the antibody or fragment is administered in combination with a therapeutically active agent as defined in item 25. A method according to any one of items 31 to 34, 36 or 37 to 39, or 41, wherein the anti-PD-L2 antibody or fragment thereof comprises means for binding to programmed cell death ligand 2 (PD-L2) and means for blocking activity against PD-L2 / PD1 interaction. A method of treating or reducing the risk of a fibrotic disease, said method comprising administering an effective amount of an anti-PD-L2 antibody or antigen-binding fragment thereof to a subject in need thereof, wherein the anti-PD-L2 antibody or fragment comprises means for binding to programmed cell death ligand 2 (PD-L2) and means for blocking activity against PD- L2 / PD1 interaction, and whereby the fibrotic disease is treated or the risk of the fibrotic disease is reduced, and optionally wherein the fibrotic disease is as defined in item 23 or item 24 and further optionally wherein the antibody or fragment is administered in combination with a therapeutically active agent as defined in item 25.

[0314] 44. The antibody or fragment according to any one of items 1 to 14, or 30, or 33 to 35, or 38 or 39, or the method according to any one of items 31 to 34, 36 or 37 to 43, wherein the method or use is carried out in vivo in a human subject.

[0315] 45. The antibody or fragment according to any one of items 1 to 14, or 30, or 33 to 35, or 38, or 39 or 44, or the method according to any one of items 31 to 34, 36 or 37 to 44, wherein the subject has or is at risk of a fibrotic disease characterised by the presence of:

[0316] (i) senescent cells; and / or

[0317] (ii) M2 pro-fibrotic macrophages; and / or

[0318] (iii) fibroblasts.

[0319] 46. The method or use according to item 45, wherein the contacting of the macrophages and the antibody or fragment is by administering said antibody or fragment in an effective amount to said or a subject, and optionally whereby said disease is treated, or the risk of said disease is reduced.

[0320] 47. A pharmaceutical composition for treating or reducing the risk of a fibrotic disease, said composition comprising an anti-PD-L2 antibody or antigen-binding fragment as defined in any one of items 1 to 22 or 30 to 39, and optionally wherein the fibrotic disease is as defined in item 23 or item 24 and further optionally wherein the antibody or fragment is administered in combination with a therapeutically active agent as defined in item 25.

[0321] Examples

[0322] The present invention is further illustrated by the following examples. Insofar as a use of the antibody other than in oncology is concerned, VHO / VKO-mlgGl is in the context of this application explicitly excluded from its scope.

[0323] Example 1: Generation and Characterisation of humanised anti-PD-L2 antibodies

[0324] The generation and initial characterisation of a parental murine antibody and it's humanised variants VH0 / VK1 (SEQ ID NO: 1 / SEQ ID NO: 10); VH1 / VK0 (SEQ ID NO:2 / SEQ ID NO:9); VH1 / VK1 (SEQ ID NO:2 / SEQ ID NO: 10); VH1 / VK2 (SEQ ID NO:2 / SEQ ID NO: 11); VH1 / VK3 (SEQ ID NO:2 / SEQ ID NO: 12); VH2 / VK1 (SEQ ID NO: 3 / SEQ ID NO: 10); VH2 / VK2(SEQ ID NO: 3 / SEQ ID NO: 11); VH2 / VK3 (SEQ ID NO:3 / SEQ ID NO: 12); VH3 / VK1(SEQ ID NO:4 / SEQ ID NO: 10); VH3 / VK2 (SEQ ID NO:4 / SEQ ID NO: 11); VH3 / VK3 (SEQ ID NO:4 / SEQ ID NO: 12); VH4 / VK1 (SEQ ID NO:5 / SEQ ID NO: 10); VH4 / VK2 (SEQ ID NO:5 / SEQ ID NO: 11); VH4 / VK3(SEQ ID NO:5 / SEQ ID NO: 12); VH6 / VK1 (SEQ ID NO:6 / SEQ ID NO: 10); VH6 / VK2(SEQ ID NO:6 / SEQ ID NO: 11); VH6 / VK3 (SEQ ID NO:6 / SEQ ID NO: 12); VH7 / VK1(SEQ ID NO:7 / SEQ ID NO: 10); VH7 / VK2 (SEQ ID NO:7 / SEQ ID NO: 11); VH7 / VK3 SEQ ID NO:7 / SEQ ID NO: 12); VH8 / VK1 (SEQ ID NO:8 / SEQ ID NO: 10); VH8 / VK2 (SEQ ID NO:8 / SEQ ID NO: 11); VH8 / VK3 (SEQ ID NO:8 / SEQ ID NO: 12) are described in detail in PCT / EP2023 / 055506, published as WO2023 / 166210A1, see page 28, line 24 to page 35, line 2, which Examples are specifically incorporated herein by reference.

[0325] Example 2: Reporter assay blocking activity on cellular hPD-L2 / hPD-l interaction

[0326] Cellular blocking activity of the antibodies of Example 1 was assessed by with the Promega (Southampton, Hampshire, UK) PD-1 / PD-L2 blockade bioassay (Promega, Cat No. CS187131-1), which is a bioluminescent cell-based assay that is used to measure the potency and stability of antibodies and other biologies designed to block the hPD-l / PD-L2 interaction. In brief, CHO-K1 cells overexpressing hPD-L2 were thawed and plated in a 96-well plate. The following day, appropriate concentrations of antibodies were added to the wells. Jurkat PD-1 effector cells, expressing a luminescent NFAT reporter for T cell activation, were subsequently thawed and added to the wells immediately. The co-culture was incubated for 6 hours and read using the SpectraMax i3x reader (Molecular Devices, Wokingham, UK). GraphPad Prism 9.0 (GraphPad Software, La Jolla, Ca) was used for the data analysis and data was plotted using a four-parameter non-linear regression model. Results are shown in Figure 2.

[0327] Example 3: Pharmacokinetic study of IgGl WT anti-PD-L2 and IgGl LALA anti-PD-L2 mAb

[0328] Twelve 15 week old B6.Cg-FcgrttmlDcrTg(FCGRT)32Dcr / DcrJ mice (Tg32, JAX stock# 014565) that were homozygous for the Tg(FCGRT)32Dcr transgene were distributed into 3 groups with 6 mice per group. Body weights were measured just prior to IV injecting the mice. At 0 hours on Day 0, test antibodies were administered as IV injections at 5 mg / kg and 5 mL / kg. 25 pL blood samples were collected from each mouse according to the bleeding schedule at 5 minutes, 1 day, 3 days, 7 days, 10 days, 14 days, 17 days, 21 days, 24 days, and 28 days. The blood samples were collected into 1 pL K3EDTA, processed to plasma, diluted 1 / 10 in 50% glycerol in PBS, transferred into specialized 96- well storage plates, and stored at -20°C. The Mabtech (3850-1AD-6) hlgG Fc ELISA was validated for test antibody sensitivity and dynamic range for all test antibodies. All plasma samples were assessed via a validated Mabtech (3850-1AD-6) hlgG Fc ELISA to quantify test antibody concentrations. Data points consistent with antidrug antibody formation were eliminated from the half-life calculation.

[0329] Surprisingly, a significantly longer half-life was measured for VHO / VKO-hlgGl LALA (13.86 days) as compared VHO / VKO-hlgGl WT (11.88 days). Thus, variant VHO / VKO-hlgGl LALA has the potential of reduced frequency of dosing and / or fewer side effects, which emphasizes that it is of particular interest for therapy, and that other antibodies on this type of variant effector reduced constant regin may equally show such benefits. Example 4: Expression of PD-L2 on various cell types

[0330] A. Bioinformatic analysis of PD-L2 expression in cell types in IPF versus normal controls:

[0331] The single-cell dataset from the Adams et a!., "Single-cell RNA-seq reveals ectopic and aberrant lung-resident cell populations in idiopathic pulmonary fibrosis", Science Advances, 2020, 6(28), eabal983, doi:10.1126 / sciadv.abal983 was downloaded from GEO ncbi (Gene Ontology Expression database accession number GSE136831). The dataset comprises samples from the following patients: 32 IPF, 18 COPD, and 28 control donor lungs. The matrix extracted were imported and normalized using seurat package in R.

[0332] Results are shown in Figure 4A, which shows that PD-L2 is significantly more highly expressed in aberrant baseloid cells, compared to healthy individuals. Aberrant baseloid cells, a prominent fibrosis-driving senescent cell population which is highly over-represented in IPF patients but not healthy individuals, express a subset of basal markers and epithelial-mesenchymal markers. They also express senescent markers and have been proposed to be a senescent population in IPF that drives fibrosis, as deletion of their murine counterparts called "transitional aveolar epithelial cells" reduces fibrosis in the bleomycin mouse model of IPF (see Wang et al. supra).

[0333] B. Flow cytometry to assess surface levels of PD-L2 on fibroblasts, myofibroblasts or macrophage populations:

[0334] Normal human lung fibroblasts (NHLF) or Idiopathic Pulmonary Fibrosis (IPF) fibroblasts or myofibroblasts, generated as described in Example 6, or MO, Ml or M2 macrophages generated as described in Example 5, were collected and stained with antibodies against PD-L2 conjugated to Allophycocyanin (APC) (Miltenyi #130-116-564). A population of 10,000 live cells was captured in all fluorescence-activated cell sorting (FACS) experiments. FACS analysis was performed using FACS Gallios Flow Cytometry system and FlowJo software package.

[0335] Figure 4B shows that PD-L2 is more highly expressed in IPF fibroblasts compared to NHLF controls and that PD-L2 is higher in myofibroblasts compared to fibroblasts in both NHLFs and IPF fibroblasts.

[0336] Figure 4C shows that PD-L2 is more highly expressed in pro-fibrotic M2 macrophages compared to M0 or Ml macrophages which are not pro-fibrotic, suggesting that targeting PD-L2 on pro-fibrotic macrophages might be an anti-fibrotic therapeutic strategy.

[0337] Example 5: Anti-PD-L2 antibodies reprogram pro-fibrotic macrophages into non-fibrotic Ml macrophages

[0338] A. PD-L2 and isotvoe antibody preparation

[0339] All anti-PD-L2 antibodies (VH6 / VK3, HC V region SEQ ID No:6, LC V region SEQ ID No: 12) and isotype controls (directed against hen egg lysozyme) on either wild-type hlgGl (SEQ ID No:22) or hlgGl-LALA (SEQ ID No:26) formats that were used in Examples 5 and 6 were manufactured by Biointron Biological (Shanghai, China). B. Macrophage polarization and treatment with antibodies to assess effect on reprogramming

[0340] Monocytes were isolated from whole blood from healthy donors obtained at Banc de Sang I Teixits (BST) by RosetteSep™ Human Monocyte Enrichment Cocktail incubation prior to Ficoll density gradient centrifugation. Monocytes were seeded at 700,000 cells / well with Roswell Park Insitute Medium (RPMI) supplemented with glutamine (GLUTAMAX, Gibco #13462629), 10% fetal bovine serum (Gibco #10500064) and 1% penicillin streptomycin (Merck #P0781-100ML) in a 6-well plate with the following cytokines:

[0341] • Ml Polarization: 10 ng / mL GM-CSF (Peprotech #00-03-50UG)

[0342] • M2 Polarization: 10 ng / mL M-CSF1 (Peprotech #300-25-500UG) + 10 ng / mL IL-4 (Peprotech #00-04-50 ug)

[0343] Monocytes were left to polarize for 7 days in an incubator (37 °C, 5% CO2). After polarization, macrophages were treated for 72 hours as follows:

[0344] • 20 pg / mL VH6 / VK3 WT or isotype control

[0345] • 20 pg / mL VH6 / VK3 LALA or isotype control

[0346] • 10 ng / mL of PLX3397 (Selleckchem Ref: S7818) which is a CSF1R inhibitor molecule, as positive control of reprogramming

[0347] C. Flow cytometry to assess reprogramming markers on macrophages

[0348] Repolarization of macrophages was evaluated by expression of surface markers by flow cytometry after the different treatments using the following antibodies:

[0349] • CD206 (MRC1) as marker for M2 polarized macrophages APC anti-human CD206 (MMR) (Biolegend #321110). Macrophages were detached using Trypsin-EDTA (0.05%), phenol red (Gibco #25300054), washed twice with PBS, resuspended in 200 pL of PBS + 1% BSA and incubated protected from light for 1 hour at 4 °C with the antibody at a 1: 150 dilution. After incubation, cells are washed twice with PBS, resuspended in 200 pL PBS and analyzed by flow cytometry using a Beckman Coulter Gallios flow cytometer.

[0350] • CD86 as marker for Ml polarized macrophages Anti-CD86 Antibody [BU63] (PE) (Abeam Ab77226). Macrophages were detached using Trypsin-EDTA (0.05%), phenol red (Gibco #25300054), were washed twice with PBS, resuspended in 200 pL PBS + 1% BSA and incubated protected from light for 1 hour at 4 °C with the antibody at a 1: 150 dilution. After incubation, cells are washed twice with PBS, resuspended in 200 pL PBS and analyzed by flow cytometry using a Beckman Coulter Gallios flow cytometer.

[0351] Results are shown in Figure 5A.

[0352] Treatment with VH6 / VK3 WT and LALA both result in a significant increase in CD86 expression, indicating reprogramming towards an Ml-like phenotype, which is not seen with the equivalent isotype controls. Untreated M2 macrophages (NT) exhibit low CD86 expression (Figure 5A, left). Treatment with VH6 / VK3 WT and LALA leads to a notable decrease in CD206 expression, reflecting the shift towards an antifibrotic Ml-like state. This decrease is not seen with the equivalent isotype controls. Untreated M2 macrophages (NT) show high CD206 expression, consistent with their pro-fibrotic phenotype (Figure 5A, right). This suggests that anti-PD-L2 antibodies, and specifically VH6 / VK3, on either a WT or LALA format can have therapeutic anti-fibrotic effects by changing the pro-fibrotic phenotype of M2 macrophages to a non-fibrosis-promoting phenotype.

[0353] D. ELISA to measure cytokines

[0354] After polarization and treatment of macrophages as described in Example 5B, consumed media was aspirated and substituted by fresh Roswell Park Institute Medium (RPMI) supplemented with glutamine (GLUTAMAX, Gibco #13462629), 10% fetal bovine serum (Gibco #10500064) and 1% penicillin streptomycin (Merck #P0781-100ML) and left for 5 days to accumulate cytokines to evaluate by ELISA using either Human TGF-pi ELISA Kit (Invitrogen #BMS249-4) and Human TNF-a ELISA Kit (Invitrogen #KHC3011). After 5 days of culture, supernatants were obtained. For each of TGF-p and TNF-a, 100 pL of standards, controls, and diluted samples, were added to the appropriate wells of the 96-well plate and incubated at room temperature for 2 hours on a shaker. After incubation, wells were washed four times with wash buffer to remove unbound components. 100 pL biotinylated anti-TNF-a or TGF-pi detection antibody was added to each well, followed by a 1-hour incubation at room temperature on a shaker. The washing step was repeated as described above. 100 pL Streptavidin-HRP conjugate was added to each well, and the plate was incubated for 30 minutes at room temperature on a shaker followed by four washes with wash buffer. 100 pL TMB substrate solution was added to each well, and the plate was incubated in the dark at room temperature for 10- 20 minutes until sufficient colour development occurred. The reaction was stopped by adding 100 pL stop solution to each well, resulting in a colour change from blue to yellow. Absorbance was measured at 450 nm using a microplate reader, with a background correction at 540 nm. A standard curve was generated from the standards, and concentrations in the samples were calculated based on this curve. All samples were analyzed in duplicate, and data were expressed as mean concentrations.

[0355] Results are shown in Figure 5B.

[0356] TGF-p is a cytokine heavily involved in fibrotic processes and tissue scarring. A significant reduction in TGF-0 production is observed after treatment with both VH6 / VK3 WT and LALA, indicating a decrease in pro-fibrotic activity. The decrease in TGF-0 levels is similar to that observed in Ml macrophages, suggesting a shift to an non-pro-fibrotic, Ml-like state. Such a reduction is not seen with the equivalent isotype controls. There is no significant difference in TGF-0 reduction between the two antibodies.

[0357] TNF-a is a cytokine associated with inflammation and the resolution of fibrosis. A significant increase in TNF-a is observed following treatment with both antibodies. The increase in TNF-a levels is similar to that observed in Ml macrophages, suggesting a shift to a non-pro-fibrotic, Ml-like state. This increase is not seen with the equivalent isotype controls. There is no significant difference in TNF-a induction between the two antibodies.

[0358] E. Quantification of proliferation / cell number for macrophages in response to antibody treatment:

[0359] 500,000 M2 macrophages were seeded into 6-well plates and left to attach overnight in an incubator (37 °C, 5% CO2). Macrophages were then treated as in Example 5B and after 72h of treatment, cells were rinsed with PBS and detached by adding 0.25% trypsin-EDTA solution (Gibco #25300054) and incubating at 37 °C for 15 minutes until cells detach. Trypsin was then neutralized by adding an equal volume of complete medium to each well and cell suspensions were collected from each well and transferred into a centrifuge tube. Cell suspension was gently mixed to ensure even distribution of cells, and 10 pL of suspension was mixed with 10 pL of 0.4% trypan blue solution (Invitrogen #T10282) and allowed to sit for 2-3 minutes at room temperature. Finally, 10 pL of mixture were loaded into a Countess™ cell counting chamber slide (Invitrogen #C10228) and cells were counted with a Countess™ 3 automated cell counter.

[0360] Results are shown in Figure 5C, left panel. Anti-PD-L2 antibodies on either wild-type or hlgGl- LALA formats stimulated the expansion of reprogrammed macrophages relative to untreated or isotype controls. The increased stimulation of proliferation by anti-PD-L2 antibodies was greater than that of PLX3397 (CSF1R inhibitor, positive control).

[0361] F. Microscopy

[0362] Images were captured at 20x magnification in brighfield on a ZEISS™ model AxioVert.Al FL- LED microscope and camera AXIOCAM 305.

[0363] Results are shown in Figure 5C, right panels. Untreated or isotypte-treated M2 macrophages exhibit an elongated, spindle-shaped morphology typical of the M2 phenotype. These microscopy images suggest that both VH6 / VK3 WT and LALA reprogram M2 macrophages into an Ml-like phenotype, characterized by a shift to a more rounded morphology and an increase in cell density. Treatment of M2 macrophages with the CSF1R inhibitor PLX3997, which is known to induce reprogramming of M2 macrophages to an Ml-like phenotype, showed an effect on cell morphology and cell density that was similar to that seen with anti-PD-L2 treatment.

[0364] Example 6: Assessment of fibroblast to myofibroblast transition

[0365] A. Assessment of fibroblast to myofibroblast transitionbv immunofluorescence for smal

[0366] NHLFs (Lonza #CC-2512) and fibroblasts from IPF donors (Lonza #CC-7231 (donor #29) and #CC-7231 (donor #37)) were grown in Fibroblast Basal Medium (Lonza #CC-3131), supplemented with FGM2 singleQuots (Lonza #CC-4126 that that includes insulin (#CC-4021J), rhFGF-B (#CC- 4065J), GA-1000 (#40813) and fetal bovine serum (#CC-4101J) at a dilution of 1: 1000. Stellate cells (Innoprot #P10653) were grown in SteCM medium supplemented with SteCGS IX (Innoprot #P60126), 2% FBS and 1% P / S solution.

[0367] The transition from fibroblasts to myofibroblast was induced using TGF-0 as previously described (see Desmouliere eta / ., "Transforming growth factor-beta 1 induces alpha-smooth muscle actin expression in granulation tissue myofibroblasts and in quiescent and growing cultured fibroblasts", J. Cell Biol., 1993, 122(1), 103-111, doi: 10.1083 / jcb.122.1.103 and Sime et al, "Adenovector-mediated gene transfer of active transforming growth factor-betal induces prolonged severe fibrosis in rat lung", J. Clin. Invest., 1997, 100(4), 768-776, doi: 10.1172 / JCI 119590).

[0368] Briefly, 5,000-10,000 NHLF, stellate and IPF cells (from the two donors) were plated in a 96- well plate. The following day, cell media was changed to DMEM (Gibco #41965-047)) supplemented with 0.5% FBS (Gibco #10500064), 1% P / S, and 2.5 ng / mL TGFp (Thermofisher, #PHG9214) for 72 hours. The following antibodies were added to cells 6 hours after addition of TGF-0:

[0369] • 60 pg / mL VH6 / VK3 WT or Isotype

[0370] • 60 pg / mL VH6 / VK3 LALA or Isotype

[0371] • 60 pg / mL Atezolizumab as positive anti-fibrotic control (MedChemExpress #HY-P9904)

[0372] After 3 days of incubation with TGF-0 and anti-PD-L2 antibodies, immunofluorescence staining was conducted against a-smooth muscle actin 1 (smal). Briefly, cells were washed and fixed in 4% PFA for 20 min at room temperature, then permeabilized with 0,1% Triton (Merck #X-100) in PBS, 20 min at room temperature, followed by a blocking in blocking solution containing 10% Goat Serum (Sigma Aldrich #G9023) in PBS, 1 hour at room temperature. Then cells were incubated with anti-PD- L2 antibodies or isotype control in blocking solution containing 10% goat serum for 1 hour at room temperature. Cells were then washed in PBS and incubated with goat anti-rabbit IgG Alexafluor 488 (Thermofisher #A11008), 1:300 in PBS, 1 hour at room temperature. Cells then were washed and mounted in Ibidi Mounting medium with DAPI (Ibidi #50011). To perform the quantification of smal positive fibers in all samples, 5 to 6 representative fields per sample were analyzed using Fiji Image] software, measuring the intensity of smal fluorescent expression. DAPI was used to stain cell nuclei. Results are representative of two independent experiments, and one-way ANOVA with Tukey's multiple comparison test was used to compare statistically significant differences between all conditions.

[0373] Results are shown in Figure 6B-E which show a decrease in smal expression in TGF-0-treated IPF fibroblasts and stellate cells upon treatment with VH6 / VK3 hlgGl WT or VH6 / VK3 hlgGl-LALA compared to untreated or isotype-treated cells. This effect was not seen in TGF-0-treated NHLF, suggesting that only TGF-0-treated pulmonary fibroblasts from patients with disease respond to anti- PD-L2 monoclonal antibodies. TGF-0-treated IPF fibroblasts and stellate cells also showed a decrease in smal when treated with the anti-PD-Ll mAb Atezolizumab which was equal or less than the effect seen with anti-PD-L2 mAbs.

[0374] B. Microscopy

[0375] Images were captured using the Leica AF7000 widefield fluorescence microscope and the 20x Dry N.A. 0.40 magnification objective.

[0376] Results are shown in Figure 6A. Smal staining of fibroblasts after TGF-0 exposure shows that treatment with anti-PD-L2 antibodies or anti-PD-Ll antibody Atezolizumab reduces the appearance of smal fibers compared to untreated or isotype-treated fibroblasts. This indicates that anti-PD-L2 therapy can block the fibroblast to myofibroblast transition and have therapeutic anti-fibrotic effects.

[0377] Example 7: Evaluation of anti-PD-L2 mAb in the Bleomvcin-induced Idiopathic Pulmonary Fibrosis in C57BL / 6 mice

[0378] Male C57BL / 6 Ntac mice with 23±2g were provided. A day prior to Bleomycin instillation, i.e., day (-1), animals were randomized to the respective groups based on the body weight and Bleomycin instillation was done on day 0. On day 0, animals were anesthetized using a combination of ketamine + xylazine (70 + 5 mg / kg, i.p.) and were mounted on a vertical board, suspended by its upper incisors. Using the fiber optic cable light source, the intra-tracheal catheter was introduced into the trachea through mouth. The fiber optic cable was retracted slowly by leaving the intra-tracheal catheter in place, without disturbing. The Bleomycin (1 mg equivalent to 1.5 - 2 U / kg in 50 ■ L / mouse dose volume) solution was placed with micropipette into the hub of the intra-tracheal catheter and the solution was sucked into the lung on its own and additionally 300 pL of air was pushed gently to ensure the uniform distribution of the bleomycin solution. Post-instillation mice were returned to their respective cages and observed for recovery. Similarly, non-fibrotic mice without Bleomycin received phosphate buffered saline (PBS) via intra-tracheal route. Mice receiving antibodies were injected three times weekly intraperitoneally with 10 mg / kg of respective antibody formulated in PBS. Mice were treated with Pirfenidone at 100 mg / kg formulated in 0.5% w / v Methyl Cellulose by oral gavage twice daily. All treatments were administered between day 7 and day 21 post-bleomycin.

[0379] A. Hydroxyproline Assay

[0380] On day 21, lung tissue was collected and snap frozen immediately in liquid nitrogen, stored at -80°C, the right lobes of individual lungs were used for hydroxyproline assay. Hydroxyproline level was estimated using following procedure. Hydrolysis of the lung tissue from each mouse with 12 N HCI (1:20) in an oven at 110°C was carried out for overnight. The hydroxyproline content was determined by oxidizing the samples with 0.2 M Chloramine-T and reacting with Ehrlich's reagent. The color change was measured in spectrophotometry at 550 nm.

[0381] B. Histological Analysis

[0382] For histological examination, on day 21, mice were euthanized; lung tissue (left lobe) was collected and fixed with 10% NBF. The tissues were embedded in paraffin wax and sectioned. The lung tissue sections were stained with H&E and Picrosirius Red for light microscope analysis. Ashcroft's score scale was used for grading fibrosis and %CPA was calculated with Leica Application Suite Image Analysis software. Remaining tissue blocks were stored for immunohistochemistry.

[0383] C. Fibrosis Grading bv Ashcroft Scale

[0384] All the H&E stained tissue sections were examined by light microscopy. The whole section was scored for fibrosis independently by Ashcroft scale in 10X objective lens as per the grading system below according to Ralf-Harto Hubner et al., "Standardized quantification of pulmonary fibrosis in histological samples", Biotechniques, 44(4), 507-11, 514-7, 2008, doi: 10.2144 / 000112729, incorporated herein by reference in its entirety.

[0385] Table 6: Ashcroft score Further, the tissue sections were evaluated for inflammation score, as described in Szapiel et a / ., "Bleomycin-induced interstitial pulmonary disease in the nude, athymic mouse", Am. Rev. Respir. Dis., 120(4), 893-9, 1979, doi:10.1164 / arrd.l979.120.4.89, incorporated herein by reference in its entirety.

[0386] Table 7: Inflammation Scale D. Collagen Proportion Area (% CPA) Measurement

[0387] All the Sirius red stained tissue sections were examined by light microscopy in 10X objective lens. Randomly selected five fields (~1243.31 pm X 932.24 pm per field) from each lung was subjected to collagen deposition area measurement by using Leica Application Suite - X (LAS-X) software. The percentage collagen proportion area was calculated by dividing the collagen tissue area by the total tissue area. E. Immunohistochemical analysis

[0388] Immunohistochemical analyses were performed using commercially available p21 (Anti-p21 antibody [HUGO291] (abl07099)), pl6 (Recombinant Anti-ARPC5 / pl6 ARC antibody [EP1551Y] (ab51243)) and secondary analysis with Goat Anti-Rabbit IgG H&L (HRP). The subsequent left lung sections were deparaffinized and rehydrated subsequently and the endogenous peroxidase was blocked with peroxide block. Antigen retrieval was performed as per the manufacturer's instructions.

[0389] Following the antigen retrieval, sections were washed with three changes of Tris-buffered saline (TBS, ab64248) for 3 minutes and the endogenous peroxidase was blocked with hydrogen peroxide block followed by protein block (provided in secondary antibody kit) to prevent non-specific binding of primary antibody and incubated with primary antibodies at 1: 100 dilution overnight at 4° to 8°C. Following adequate washing with TBS, serially the sections previously incubated with p21 primary antibody were incubated with secondary antibody -Goat Anti-Rat IgG H&L (HRP) (ab97057) (1:200 dilution) for 60 mins, and sections previously incubated with pl6 primary antibody were incubated with Rabbit specific horseradish peroxidase / 3,3' Diaminobenzidine (HRP / DAB) (ABC) IHC Detection Kit (ab64261) secondary antibody (1:200 dilution) for 60 mins, washed with three changes of TBS for 3 minutes each, followed by incubation with Streptavidin HRP (ab64269) for 30 mins, washed with three changes of TBS for 3 minutes, and then incubated with DAB solution (1 drop) at room temperature for specific time (1-3 minutes), the DAB solution was discarded, and the slide was washed with distilled water. Next the sections were counterstained with hematoxylin Gill no 3 (Himedia (S076)) (30-60 Sec) and rinsed thrice in distilled water followed by dehydration, clearing with xylene and mounted with D.P.X mountant (Spectrochem (010469)).

[0390] F. Statistical Analysis on bleomycin mouse model samples

[0391] Raw data were analyzed across all groups, using one-way analysis of variance (One-way ANOVA) or two-way analysis of variance (Two-way ANOVA) along with the appropriate multiple comparison post-test (Dunnett's, Bonferroni and Sidak multiple comparison test). Significance for all tests was set at p<0.05.

[0392] G. Results

[0393] As shown in Figures 7A to 7B, therapeutic treatment using VH0 / VK0-mIgG2a LALA and VH0 / VK0-mIgG2a WT resulted in a reduction of hydroxyproline content and Ashcroft score compared to vehicle and isotype controls, the effect seen with VH0 / VK0-mIgG2a LALA being equivalent to that seen with pirfenidone and greater than that seen with VH0 / VK0-mIgG2a WT.

[0394] In addition, a significant reduction in picrosirius red staining was seen in mice subject to a treatment with VH0 / VK0-mIgG2a LALA or VH0 / VK0-mIgG2a WT, the effect seen with VH0 / VK0- mIgG2a LALA being equivalent to that seen with pirfenidone. The effect was confirmed by a histological analysis represented in Figure 7E showing significant improvements in lung architecture and fibrosis for treatment with VH0 / VK0-mIgG2a WT, VH0 / VK0-mIgG2a LALA and pirfenidone, in particular with VH0 / VK0-mIgG2a LALA and pirfenidone. Further, the reduction in the inflammation score was pronounced in mice subject to treatment with VH0 / VK0-mIgG2a WT and VH0 / VK0-mIgG2a LALA, the effect seen with VH0 / VK0-mIgG2a LALA being equivalent to that seen with pirfenidone (Figure 7F).

[0395] As shown in Figure 8A and Figure 8C, treatment with VH0 / VK0-mIgG2a LALA and VHO / VKO- m!gG2a WT resulted in reduction of pl6 and p21 staining compared to vehicle and isotype controls, the reduction being particularly pronounced for VH0 / VK0-mIgG2a LALA (equivalent to that seen with pirfenidone). In addition, images of lung sections stained with p21 illustrated reductions in senescent cells burden in mice that are treated with VH0 / VK0-mIgG2a LALA, VH0 / VK0-mIgG2a WT and pirfenidone, the effect being more significant in mice treated with VH0 / VK0-mIgG2a LALA and pirfenidone.

[0396] The therapeutic and anti-fibrotic effect, and high activity to clear senescent cells with the antibody on the LALA format is unexpected since effector proficient antibodies that depend on depleting cell populations for their activity typically show higher potency when enabled with greater effector function.

[0397] Example 8: Assessing role of anti-PD-L2 mAb in premature aging using Zmpste24-KO mouse model

[0398] A. Mouse husbandry and longevity study

[0399] Zmpste24-KO mice (C57BL / 6N) were kept under a 12 h-12 h light-dark cycle, between 20 and 22° C and with unrestricted access to food and water. The procedure was approved by the Ethical Committee for Animal Experimentation (CEEA) at the Parc Cientific de Barcelona (Spain) and the Generalitat de Catalunya.

[0400] At 40 days of age, Zmpste24-KO animals were randomly assigned to either treated or control groups. Treated animals were administered 100 pg anti-PD-L2 antibody (TY25, BioXCell) twice per week via intraperitoneal injection. Control animals were administered an equivalent volume of PBS. Body weight was monitored weekly. Starting at day 100 of life, and upon increased frailty (determined by a decrease of more than a 20% of body weight compared to the maximum body weight for a given mouse). Euthanasia was performed when the animals were considered unable to reach food or water or when continued monitoring by veterinaries detected signs of excessive pain and weight loss. Penis prolapse, when present, was monitored and was not considered a cause for euthanasia.

[0401] B. Creatinine in serum

[0402] Blood was extracted by cardiac puncture at the time of euthanasia and serum was obtained by centrifugation (1500 x g, 10 minutes, 4° C). Creatinine determinations in mouse sera were performed using VetScan Comprehensive Diagnostic Profile reagent rotors and a VetScan VS2 Chemistry Analyzer.

[0403] C. Gene expression

[0404] For gene expression determinations, kidneys are cut in half and snap frozen in liquid nitrogen upon euthanasia. RNA was extracted from 10 mg of frozen kidney cortex using a Fastprep24 homogenizer (MP Biomedical) and the RNeasy extraction kit (Qiagen). cDNA was obtained using iScript cDNA synthesis kit (BioRad) and gene expression was analyzed in

[0405] The following primer combinations were used:

[0406] Table 8: Primer combinations

[0407] D. Aorta histology

[0408] For histological analysis in aorta, the descending aorta was excised and fixed overnight with 10% neutral buffered formalin, followed by a wash with PBS, dehydration and paraffin embedding. Sections were stained with a Massson's Trichrome Stain Kit to identify collagen fibers (AR17311-2, Artisan, Agilent). Relative thickness of the tunica adventitia was calculated using Qpath 0.4.3.

[0409] E. Statistical analysis

[0410] For longevity studies, log-rank test was applied to determine statistical significance. For the rest of data, two-tailed T-tests were applied for isolated pairwise comparisons and, for comparisons between three groups, one way ANOVA followed by Tukey post-test (for pairwise comparisons) was applied. GraphPad Prism 8.0.1 was used for statistical analysis.

[0411] As shown in Figure 9A, Zmpste24-KO mice treated with anti-PD-L2 antibody showed a median lifespan significantly extended by 13% as compared to treatment with isotype control, the effect being more pronounced in females (30% extension of median lifespan as compared to treatment with isotype control, see Figure 9B).

[0412] In addition, Zmpste24 2) mice treated with anti-PD-L2 antibody had reduced serum creatinine levels (below 0.2 mg / dL) and thus exhibited improved kidney function as compared to those treated with IgG control antibody (Figure 10A). Further, increased glomerular size and hence improved renal function was measured for Zmpste24 KO mice treated with anti-PD-L2 antibody as compared to the subjects treated with IgG control antibody (Figure 10B).

[0413] As evidenced by the data shown in Figure 10C, also expression of various collagen genes colla2, co / 2al, co / 6al and col6a2), which are markers of fibrosis, was found to be significantly reduced for in Zmpste24 knockout mice treated with anti-PD-L2 antibody as compared to those treated with the IgG-control antibody.

[0414] No decrease in the absolute thickness of the inner layers of the inner aorta wall (tunica intima and media) was seen in Zmpste24 2) mice treated with anti-PD-L2 antibody. However, a decreased absolute thickness of the outer collagen-rich tunica adventitious was seen, as shown in Figures 11A and 11B.

Claims

Claims1. An isolated monoclonal antibody, or antigen binding fragment thereof, that specifically binds to programmed cell death ligand 2 (PD-L2) and blocks activity against PD-L2 / PD1 interaction for use in the treatment of a fibrotic disease in a mammal, wherein the antibody(a) binds to human PD-L2 with a dissociation constant (KD) equal from (about) 10'8M to (about) 10'12M; and / or(b) blocks in vitro the interaction between human programmed cell death protein 1 (hPD-1) and human PD-L2 with an ECso of from (about) 0.1 to (about) 1.5 nM, optionally in a bioluminescent cell-based assay, and optionally using Jurkat T cells expressing human PD-1 and a luciferase reporter driven by an NFAT response element (NFAT-RE) and CHO-K1 cells expressing human PD-L2 and an engineered cell surface protein designed to activate cognate TCRs in an antigen-independent manner; and / or(c) specifically binds to both human and murine PD-L2; and / or(d) competes for binding to PD-L2 with an antibody having a heavy chain variable region having an amino acid sequence of SEQ ID No:1 and a light chain variable region having an amino acid sequence of SEQ ID No:9, optionally in an ELISA, SPR or cell-based assay.

2. The antibody or fragment according to claim 1, wherein the antibody further(e) comprises a heavy chain constant region which has reduced or no Fc receptor (FcyR) interactions, or reduced or no immune effector functions.

3. The antibody or fragment according to claim 1 or claim 2, which comprises each of features a) and b).

4. The antibody or fragment according any one of the preceding claims, wherein the antibody promotes immune-mediated clearance of senescent cells in fibrotic tissue.

5. The antibody or fragment according to any one of the preceding claims, whose administration to a subject suffering from fibrosis, in particular from IPF, results as compared to the respective disease control groupA. in a decrease in the %-CPA (collagen proportionate area), in particular a decrease by at least (about) 5%, at least (about) 10%, at least (about) 15%, at least (about) 20%, or at least (about) 25%, at least (about) 30%, at least (about) 35%, or at least (about) 40%; and / orB. in a decrease in the hydroxyproline levels, in particular a decrease by at least (about) 5%, at least (about) 10%, at least (about) 15%, at least (about) 20%, at least (about) 25%, at least (about) 30%, at least (about) 35%, or at least (about) 38%; and / orC. in a decrease in the Ashcroft score as compared to the disease control group, in particular a decrease by at least 0.5 points, at least 0.6 points, at least (about) 0.7 points, at least (about) 0.8 points, at least (about) 0.9 points, at least (about) 1 point, at least (about) 1.1 points, at least (about) 1.2 points, at least (about) 1.3 points, or at least (about) 1.4 points; and / orD. in a decrease of the inflammation score, in particular a decrease by at least (about) 0.5 points, at least (about) 0.6 points, at least (about) 0.7 points, at least (about) 0.8 points, at least (about) 0.9 points, at least (about) 1 point, at least (about) 1.1 points, at least (about) 1.2 points, or at least (about) 1.3 points; and / orE. in a decrease in pl6 expression by at least (about) 10%, at least (about) 15%, at least (about) 20%, at least (about) 25%, at least (about) 30%, at least (about) 35%, or at least (about) 40%; and / orF. in a decrease in p21 expression by at least (about) 10%, at least (about) 15%, at least (about) 20%, at least (about) 25%, at least (about) 30%, at least (about) 35%, or at least (about) 40%.

6. The antibody or fragment according to claim 5, wherein the subject is a mouse and the fibrosis is a bleomycin-induced model of IPF.

7. The antibody or fragment according to any one of the preceding claims, wherein the antibody is a humanised or chimeric antibody whose three heavy chain CDRs are derived from a murine IGHV gene segment which is IGHV1-54 (SEQ ID No:37), a murine IGHD gene segment, and murine IGHJ gene segment which is IGHJ1 (SEQ ID No: 38), and whose three light chain CDRs are derived from a murine IGKV gene segment which is IGKV1- 110 (SEQ ID No:39) and a murine IGKJ gene segment which is IGKJ1 (SEQ ID No:40).

8. The antibody or fragment according to any one of the preceding claims, wherein said antibody comprises three CDR sequences in the heavy chain variable region, and three CDR sequences in the light chain variable region, and comprises: i. as heavy chain variable region CDRs a CDRH1 comprising the amino acid sequence of SEQ ID NO:33 (NYFIE, according to Kabat numbering) or of SEQ ID NO: 34 (GYAFSNY, according to Chothia numbering) or alternatively SEQ ID NO:36 (GYSFSNY, according to Chothia numbering), in particular comprising the amino acid sequence of SEQ ID NO: 13 (GYAFSNYFIE) or SEQ ID NO: 14 (GYSFSNYFIE);a CDRH2 comprising the amino acid sequence of SEQ ID NO: 15 (LNIPGSGGSNYAEKFKG, according to Kabat numbering) or of SEQ ID NO:35 (IPGSGG, according to Chothia numbering); and a CDRH3 comprising the amino acid sequence of SEQ ID NO: 16 (RRLPPDWYFDV, according to both Kabat and Chothia numbering); and ii. as light chain variable region CDRs a CDRL1 comprising the amino acid sequence of SEQ ID NO: 17 (RSSQSLVHSGGNTYLH, according to both Kabat and Chothia numbering) or SEQ ID NO:20 (RSSQSLVHSDGNTYLH, according to both Kabat and Chothia numbering); a CDRL2 comprising the amino acid sequence of SEQ ID NO: 18 (KVSNRFS, according to both Kabat and Chothia numbering); and a CDRL3 comprising the amino acid sequence of SEQ ID NO: 19 (SQSTHVPWT, according to both Kabat and Chothia numbering), or which comprises the CDR sequences of a) and b) comprising one or two, in particular one, amino acid substitutions, deletions or additions in one or more of the CDR sequences.

9. The antibody or fragment according to claim 8, which comprises: i. as heavy chain variable region CDRs a CDRH1 comprising the amino acid sequence of SEQ ID NO:33 (NYFIE, according to Kabat numbering) or of SEQ ID NO: 36 (GYSFSNY, according to Chothia numbering), in particular comprising the amino acid sequence of SEQ ID NO: 14 (GYSFSNYFIE); a CDRH2 comprising the amino acid sequence of SEQ ID NO: 15 (LNIPGSGGSNYAEKFKG, according to Kabat numbering) or of SEQ ID NO:35 (IPGSGG, according to Chothia numbering); and a CDRH3 comprising the amino acid sequence of SEQ ID NO: 16 (RRLPPDWYFDV, according to both Kabat and Chothia numbering); and ii. as light chain variable region CDRs a CDRL1 comprising the amino acid sequence of SEQ ID NO: 17 (RSSQSLVHSGGNTYLH, according to both Kabat and Chothia numbering); a CDRL2 comprising the amino acid sequence of SEQ ID NO: 18 (KVSNRFS, according to both Kabat and Chothia numbering); and a CDRL3 comprising the amino acid sequence of SEQ ID NO: 19 (SQSTHVPWT, according to both Kabat and Chothia numbering), orwhich comprises the CDR sequences of a) and b) comprising one or two, in particular one, amino acid substitutions, deletions or additions in one or more of the CDR sequences.

10. The antibody or fragment according to claim 8, which comprises: i. as heavy chain variable region CDRs a CDRH1 comprising the amino acid sequence of SEQ ID NO:33 (NYFIE, according to Kabat numbering) or of SEQ ID NO:34 (GYAFSNY, according to Chothia numbering), in particular comprising the amino acid sequence of SEQ ID NO: 13 (GYAFSNYFIE); a CDRH2 comprising the amino acid sequence of SEQ ID NO: 15 (LNIPGSGGSNYAEKFKG, according to Kabat numbering) or of SEQ ID NO:35 (IPGSGG, according to Chothia numbering); and a CDRH3 comprising the amino acid sequence of SEQ ID NO: 16 (RRLPPDWYFDV, according both to Kabat and Chothia numbering); and ii. as light chain variable region CDRs a CDRL1 comprising the amino acid sequence of SEQ ID NO:20 (RSSQSLVHSDGNTYLH, according to both Kabat and Chothia numbering); a CDRL2 comprising the amino acid sequence of SEQ ID NO: 18 (KVSNRFS, according to both Kabat and Chothia numbering); and a CDRL3 comprising the amino acid sequence of SEQ ID NO: 19 (SQSTHVPWT, according to both Kabat and Chothia numbering), or which comprises the CDR sequences of a) and b) comprising one or two, in particular one, amino acid substitutions, deletions or additions in one or more of the CDR sequences.

11. The antibody or fragment according to any one of the preceding claims, whereinI. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO:1 (VHO) and the light chain variable region comprises the amino acid sequence of SEQ ID NO:9 (VKO); orII. the heavy chain variable region comprises the amino acid sequence of SEQ ID NO:6 (VH6) and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 12 (VK3), preferably, the heavy chain variable region comprises the amino acid sequence of SEQ ID NO:6 (VH6) and the light chain variable region comprises the amino acid sequence of SEQ ID NO: 12 (VK3).

12. The antibody according to any one of claim 1 or claims 3 to 11, wherein the antibody comprises a heavy chain constant region which comprises the amino acid sequence of SEQ ID NO:21 or of SEQ ID NO:22.

13. The antibody according to any one of claims 1 to 11, wherein the antibody comprises a heavy chain constant region which has reduced or no Fc receptor (FcyR) interactions, or reduced or no immune effector functions, and in particular comprises a mutagenized Fc variant sequence of an IgG isotype exhibiting reduced Fc receptor (FcyR) interaction or reduced immune effector function compared to its parent, wild type Fc region.

14. The antibody according to claim 13, wherein the antibody comprises a mutagenized Fc variant sequence of murine IgG2a subclass exhibiting reduced Fc receptor (FcyR) interaction compared to its parent, wild type Fc region, optionally wherein the mutagenized Fc variant sequence comprises a mutation of leucine 234 to alanine and leucine 235 to alanine (LALA mutation) according to EU numbering, and further optionally wherein the heavy chain constant region comprises the amino acid sequence of SEQ ID NO:2515. An isolated variant monoclonal antibody which specifically binds to programmed cell death ligand 2 (PD-L2) and blocks activity against PD-L2 / PD-1 interaction, wherein said variant antibody comprises three CDR sequences in the heavy chain variable region and three CDRs in the light chain variable region, and comprises i. as heavy chain variable region CDRs a CDRH1 comprising the amino acid sequence of SEQ ID NO:33 (NYFIE, according to Kabat numbering) or of SEQ ID NO: 34 (GYAFSNY, according to Chothia numbering) or alternatively SEQ ID NO:36 (GYSFSNY, according to Chothia numbering), in particular comprising the amino acid sequence of SEQ ID NO: 13 (GYAFSNYFIE) or SEQ ID NO: 14 (GYSFSNYFIE); a CDRH2 comprising the amino acid sequence of SEQ ID NO: 15 (LNIPGSGGSNYAEKFKG, according to Kabat numbering) or of SEQ ID NO:35 (IPGSGG, according to Chothia numbering); and a CDRH3 comprising the amino acid sequence of SEQ ID NO: 16 (RRLPPDWYFDV, according to both Kabat and Chothia numbering); and ii. as light chain variable region CDRs a CDRL1 comprising the amino acid sequence of SEQ ID NO: 17 (RSSQSLVHSGGNTYLH, according to both Kabat and Chothia numbering) or SEQ ID NO:20 (RSSQSLVHSDGNTYLH, according to both Kabat and Chothia numbering); a CDRL2 comprising the amino acid sequence of SEQ ID NO: 18 (KVSNRFS, according to both Kabat and Chothia numbering); anda CDRL3 comprising the amino acid sequence of SEQ ID NO: 19 (SQSTHVPWT, according to both Kabat and Chothia numbering), or which variant antibody comprises the CDR sequences of a) and b) comprising one or two, in particular one, amino acid substitutions, deletions or additions in one or more of the CDR sequences, and wherein the variant antibody comprises a heavy chain constant region which has (i) a wild-type constant region sequence which has reduced or no Fc receptor (FcyR) interactions, (ii) a wild-type constant region sequence which has reduced or no immune effector functions, or (iii) a mutagenized Fc variant sequence of the IgG isotype exhibiting reduced Fc receptor (FcyR) interaction or reduced immune effector function compared to the parent Fc region, and for part (iii), wherein the isolated monoclonal antibody is a chimeric, humanized or human antibody.

16. The antibody according to claim 15, which is a chimeric, humanized or human antibody.

17. The antibody according to claim 15 or claim 16, which comprises the CDR combinations as defined in claim 9 or claim 10; or which comprises the VH and VL sequences as defined in claim 1118. The antibody as defined in any one of claims 15 to 17 for use in treating a fibrotic disease.

19. The antibody according to any one of the preceding claims, wherein the antibody comprises (i) a mutagenized Fc variant sequence of human IgGl subclass subclass exhibiting reduced Fc receptor (FcyR) interaction compared to its parent, wild type Fc region, or (ii) a mutagenized Fc variant sequence of murine IgG2a subclass exhibiting reduced Fc receptor (FcyR) interaction compared to its parent, wild type Fc region wherein the mutagenized Fc variant sequence of murine IgG2a does not comprise a mutation of leucine 234 to alanine and leucine 235 to alanine (LALA mutation) according to EU numbering, or a mutation of leucine 234 to alanine, leucine 235 to alanine, and proline 329 to glycine (LALAPG mutation) according to EU numbering.

20. The antibody according to claim 19(i), wherein the mutagenized Fc variant sequence comprises a mutation of leucine 234 to alanine and leucine 235 to alanine (LALA mutation) according to EU numbering.

21. The antibody according to claim 20, wherein the heavy chain constant region comprises the amino acid sequence of SEQ ID NO: 26.

22. The antibody or fragment according to any one of the preceding claims, wherein the light chain constant region comprises the amino acid sequence of SEQ ID NO: 29 or of SEQ ID NO: 30.

23. The antibody or fragment according to any one of the preceding claims for use in treating, ameliorating or preventing pulmonary fibrosis, in particular in Idiopathic Pulmonary Fibrosis (IPF).

24. The antibody or fragment according to any one of the preceding claims for use in treating a fibrotic disease selected from lung fibrosis, liver fibrosis, in particular selected from non-alcoholic steatohepatitis (NASH) and non-alcoholic fatty liver disease, kidney fibrosis and heart fibrosis.

25. The antibody or fragment according to any one of the preceding claims, wherein the antibody is administered in combination with a further therapeutically active agent, in particular a therapeutically active agent selected from the group consisting of pirfenidone, nintedanib hydronidone, luspatercept, pamrevlumab, parsaclisib, fedratinib, momelotinib, pacritinib, jaktinib, saroglitazar, lanifibranor, obeticholic acid, semaglutide, elexacaftor, ivacaftor, resmetirom, belapectin, azemiglitazone potassium, aramchol, imetelstat, KRT-232, ruxolitinib and panobinostat and combinations thereof.

26. A composition comprising the antibody or fragment as defined in any one of the preceding claims in combination with a pharmaceutically acceptable excipient.

27. An isolated nucleic acid sequence that encodes the amino acid sequence of the antibody or fragment as defined in any one of claims 1 to 22.

28. A recombinant expression vector comprising the nucleic acid of claim 27.

29. An isolated host cell comprising the recombinant expression vector of claim 28.

30. An isolated monoclonal antibody or antigen-binding fragment thereof that specifically binds to programmed cell death ligand 2 (PD-L2) and blocks activity against PD-L2 / PD1 interaction for use in the treatment of a fibrotic disease in a mammal, wherein the antibody is capable of preventing the conversion of fibroblasts to myofibroblasts in an in vitrolPV- model as measured by a reduction in a-smooth muscle actin 1 (smal) positive fibres.

31. A method of treating or reducing the risk of a fibrotic disease, said method comprising administering an effective amount of an anti-PD-L2 antibody or antigen-binding fragment thereof to a subject in need thereof, wherein the anti-PD-L2 antibody or fragment is capable of preventing the conversion of fibroblasts to myofibroblasts in an in vitro W model as measured by a reduction in a-smooth muscle actin 1 (smal) positive fibres, and whereby the fibrotic disease is treated or the risk of the fibrotic disease is reduced.

32. A method of preventing the formation of myofibroblasts, said method comprising contacting an anti-PD-L2 antibody or antigen-binding fragment thereof with a population of cells comprising fibroblasts, wherein the anti-PD-L2 antibody or fragment is capable of preventing the conversion of fibroblasts to myofibroblasts, in particular as measured by a reduction in a-smooth muscle actin 1 (smal) positive fibres in an in vitro W model.

33. The antibody or fragment according to claim 30, or the method according to claim 31 or claim 32, wherein the reduction in smal positive fibres is at least (about) 40%, or at least (about) 50%, in particular at least (about) 60%, or at least (about) 70%, compared to a TGF-0 treated control sample.

34. The antibody or fragment according to claim 30 or claim 33, or the method according to any one of claims 31 to 33, wherein the reduction in smal positive fibres is measured by a method comprising the steps of:- providing fibroblasts from a human donor;- treating the fibroblasts with TGF-0;- contacting the antibody or fragment with the treated fibroblasts;- incubating the fibroblasts in the presence of the TGF-0 and the antibody or fragment;- conducting immunofluorescence staining against smooth muscle actin 1 (smal); and- quantifying smal positive fibers, in particular by fluorescence-activated cell sorting (FACS).

35. An isolated monoclonal antibody or antigen-binding fragment thereof that specifically binds to programmed cell death ligand 2 (PD-L2) and blocks activity against PD-L2 / PD1 interaction for use in the treatment of a fibrotic disease in a mammal, wherein(i) the antibody or fragment is capable of reprogramming M2 pro-fibrotic macrophages expressing CD206 on the surface of said macrophage into Ml phenotypic macrophages expressing CD86 on the surface of said macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages; and / or(ii) the antibody or fragment is capable of increasing CD86 expression on a macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages; and / or(iii) the antibody or fragment is capable of decreasing CD206 expression on a macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages; and / or(iv) the antibody or fragment is capable of reducing TGF-0 production in a macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages; and / or(v) the antibody or fragment is capable of increasing TNF-a production in a macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages; and / or(vi) the antibody or fragment is capable of changing the morphology of the M2 macrophages from an elongated morphology to a rounded morphology, in particular as observed by microscopic analysis of cells in an in vitro model of M2 pro-fibrotic macrophages; and / or(vii) the antibody or fragment is capable of increasing the proliferation of macrophages, in particular as measured by number of cells in an in vitro model of M2 pro-fibrotic macrophages.

36. A method of treating or reducing the risk of a fibrotic disease, said method comprising administering an effective amount of an anti-PD-L2 antibody or antigen-binding fragment thereof to a subject in need thereof, wherein:(i) the antibody or fragment is capable of reprogramming M2 pro-fibrotic macrophages expressing CD206 on the surface of said macrophage into Ml phenotypic macrophages expressing CD86 on the surface of said macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages; and / or(ii) the antibody or fragment is capable of increasing CD86 expression on a macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages; and / or(iii) the antibody or fragment is capable of decreasing CD206 expression on a macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages; and / or(iv) the antibody or fragment is capable of reducing TGF-0 production in a macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages; and / or(v) the antibody or fragment is capable of increasing TNF-a production in a macrophage, in particular as measured in an in vitro model of M2 pro-fibrotic macrophages; and / or(vi) the antibody or fragment is capable of changing the morphology of the M2 macrophages from an elongated morphology to a rounded morphology, in particular as observed by microscopic analysis of cells in an in vitro model of M2 pro-fibrotic macrophages; and / or(vii) the antibody or fragment is capable of increasing the proliferation of macrophages, in particular as measured by number of cells in an in vitro model of M2 pro-fibrotic macrophages, and whereby the fibrotic disease is treated or the risk of the fibrotic disease is reduced.

37. A method of reprogramming M2 pro-fibrotic macrophages to Ml phenotypic macrophages, said method comprising contacting an anti-PD-L2 antibody or antigen-binding fragment thereof with said M2 macrophages, wherein the anti-PD-L2 antibody or fragment decreases the level (or amount) of the M2 pro-fibrotic macrophages and increases the level (or amount) of the Ml phenotypic macrophages, in particular as measured by:(i) observing an increase in CD86 expression on the macrophages; and / or(ii) observing a decrease in CD206 expression on the macrophages; and / or(iii) observing a reduction in TGF-0 production on the macrophages; and / or(iv) observing an increase in TNF-a production on the macrophages; and / or(v) observing a change in morphology of the M2 macrophages from an elongated morphology to a rounded morphology; and / or(vi) observing an increase in the proliferation of macrophages.

38. The antibody or fragment according to claim 35, or the method according to claim 36 or claim 37, wherein, independently from each other:- the increase in CD86 expression according to claim 35(H), claim 36(H) or claim 37(i) is an increase of at least (about) 40%, or at least (about) 45%, or at least (about) 50% or, in particular at least (about) 55%, compared to an untreated M2 macrophage; and / or- the decrease in CD206 expression according to claim 35(iii), claim 36(iii) or claim 37(H) is a decrease of at least (about) 50%, or at least (about) 60%, or in particular at least (about) 70%, compared to an untreated M2 macrophage; and / or- the reduction in TGF-0 production according to claim 35(iv), claim 36(iv) or claim 37(iii) is a reduction of at least (about) 30%, or at least (about) 40%, in particular at least (about) 50%, compared to an untreated M2 macrophage; and / or- the increase in TNF-a production according to claim 35(v), or claim 36(v) or claim 37(iv) is an increase of at least (about) 5-fold, or at least (about) 5.5-fold, or at least (about) 6- fold, and in particular at least (about) 6.5-fold, compared to an untreated M2 macrophage; and / or.- the increase in the proliferation of macrophages according to claim 35(vii), or claim 36(vii) or claim 37(v) is an increase of at least (about) 1.4-fold, or at least (about) 1.5-fold, or at least (about) 1.6-fold, and in particular at least (about) 1.7-fold, compared to untreated M2 macrophages.

39. The antibody or fragment according to claim 35 or claim 38, or the method according to any one of claims 36 to 38, wherein the in vitro model of M2 pro-fibrotic macrophages comprises the steps of- providing monocytes from a human donor;- treating the monocytes with the cytokines M-CSF1 and IL-4 to polarise the monocytes to M2 macrophages;- contacting the antibody with the M2 macrophages;- incubating the M2 macrophages in the presence of the cytokines and the antibody or fragment; and- for observing an increase in CD86 and / or a decrease in CD206, conducting immunofluorescence staining against CD86 and / or CD206; and quantifying expression of CD86 and / or CD206, in particular by flow cytometry;- and / or for observing a reduction in TGF-0 and / or increase in TNF-a, conducting an ELIZA assay to quantify the amount of TGF-0 and / or TNF-a;- and / or for observing an increase in the proliferation of macrophages counting the number of macrophages on a cell counter.

40. The antibody or fragment according to any one of claims 30, 33 to 35, or 38 or 39, or the method according to any one of claims 31 to 34, 36 or 37 to 39, wherein the anti-PD-L2 antibody or fragment is as defined in any one of claims 1 to 22, and optionally wherein the fibrotic disease is as defined in claim 23 or claim 24 and further optionally wherein the antibody or fragment is administered in combination with a therapeutically active agent as defined in claim 25.

41. A method of treating or reducing the risk of a fibrotic disease, said method comprising administering an effective amount of an anti-PD-L2 antibody or antigen-binding fragment thereof to a subject in need thereof, wherein the anti-PD-L2 antibody or fragment is as defined in any one of claims 1 to 22, or 30 to 39, and whereby the fibrotic disease is treated or the risk of the fibrotic disease is reduced, and optionally wherein the fibrotic disease is as defined in claim 23 or claim 24 and further optionally wherein the antibody or fragment is administered in combination with a therapeutically active agent as defined in claim 25.

42. A method according to any one of claims 31 to 34, 36 or 37 to 39, or 41, wherein the anti-PD-L2 antibody or fragment thereof comprises means for binding to programmed cell death ligand 2 (PD-L2) and means for blocking activity against PD-L2 / PD1 interaction.

43. A method of treating or reducing the risk of a fibrotic disease, said method comprising administering an effective amount of an anti-PD-L2 antibody or antigen-binding fragment thereof to a subject in need thereof, wherein the anti-PD-L2 antibody or fragment comprises means for binding to programmed cell death ligand 2 (PD-L2) and means for blocking activity against PD- L2 / PD1 interaction, and whereby the fibrotic disease is treated or the risk of the fibrotic disease is reduced, and optionally wherein the fibrotic disease is as defined in claim 23 or claim 24 and further optionally wherein the antibody or fragment is administered in combination with a therapeutically active agent as defined in claim 25.

44. The antibody or fragment according to any one of claims 1 to 14, or 30, or 33 to 35, or 38 or 39, or the method according to any one of claims 31 to 34, 36 or 37 to 43, wherein the method or use is carried out in vivo in a human subject.

45. The antibody or fragment according to any one of claims 1 to 14, or 30, or 33 to 35, or 38, or 39 or 44, or the method according to any one of claims 31 to 34, 36 or 37 to 44, wherein the subject has or is at risk of a fibrotic disease characterised by the presence of:(i) senescent cells; and / or(ii) M2 pro-fibrotic macrophages; and / or(iii) fibroblasts.

46. The method or use according to any claim 45, wherein the contacting of the macrophages and the antibody or fragment is by administering said antibody or fragment in an effective amount to said or a subject, and whereby said disease is treated, or the risk of said disease is reduced.

47. A pharmaceutical composition for treating or reducing the risk of a fibrotic disease, said composition comprising an anti-PD-L2 antibody or antigen-binding fragment as defined in any one of claims 1 to 22 or 30 to 39, and optionally wherein the fibrotic disease is as defined in claim 23 or claim 24 and further optionally wherein the antibody or fragment is administered in combination with a therapeutically active agent as defined in claim 25.

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