(19) TZZ ¥_T (11) EP 2 744 823 B1 (12) EUROPEAN PATENT SPECIFICATION (45) Date of publication and mention (51) Int Cl.: of the grant of the patent: A61K 39/00 (2006.01) C07K 14/705 (2006.01) 02.08.2017 Bulletin 2017/31 C07K 16/28 (2006.01) (21) Application number: 12823966.2 (86) International application number: PCT/US2012/051373 (22) Date of filing: 17.08.2012 (87) International publication number: WO 2013/026004 (21.02.2013 Gazette 2013/08) (54) ANTIBODIES THAT BIND INTEGRIN ALPHA-V BETA-8 INTEGRIN-ALPHA-V BETA-8 BINDENDE ANTIKÖRPER ANTICORPS QUI SE LIENT À L’INTÉGRINE ALPHA-V BÊTA-8 (84) Designated Contracting States: US-A1- 2009 324 604 US-A1- 2011 071 278 AL AT BE BG CH CY CZ DE DK EE ES FI FR GB US-B2- 7 087 405 GR HR HU IE IS IT LI LT LU LV MC MK MT NL NO PL PT RO RS SE SI SK SM TR • CLAUS NEUROHR ET AL: "Activation of Transforming Growth Factor-[beta] by the (30) Priority: 17.08.2011 US 201161524708 P Integrin [alpha]v[beta]8 Delays Epithelial Wound 11.05.2012 US 201261646111 P Closure", AMERICAN JOURNAL OF RESPIRATORY CELL AND MOLECULAR (43) Date of publication of application: BIOLOGY, vol. 35, no. 2, 1 August 2006 25.06.2014 Bulletin 2014/26 (2006-08-01) , pages 252-259, XP055088389, ISSN: 1044-1549, DOI: (73) Proprietor: The Regents of the University of 10.1165/rcmb.2006-0013OC California • Stephanie Cambier ET AL: "Vascular Biology, Oakland, CA 94607-5200 (US) Atherosclerosis and Endothelium Biology Integrin alpha v beta 8 Mediated Activation of (72) Inventors: Transforming Growth Factor-beta by • NISHIMURA, Stephen Perivascular Astrocytes An Angiogenic Control Mill Valley, California 94941 (US) Switch", , 1 June 2005 (2005-06-01), • LOU, Jianlong XP055088666, Retrieved from the Internet: San Bruno, California 94066 (US) URL:http://download.journals.elsevierhealt • BARON, Jody Lynn h.com/pdfs/journals/0002-9440/PIIS00029440 Mill Valley, California 94941 (US) 10624972.pdf [retrieved on 2013-11-15] • MARKS, James • LARS FJELLBIRKELAND ET AL: "Integrin Kensington, California 94707 (US) [alpha]v[beta]8-Mediated Activation of Transforming Growth Factor-[beta] Inhibits (74) Representative: J A Kemp Human Airway Epithelial Proliferation in Intact 14 South Square BronchialTissue", THE AMERICAN JOURNAL OF Gray’s Inn PATHOLOGY, vol. 163, no. 2, 1 August 2003 London WC1R 5JJ (GB) (2003-08-01), pages533-542, XP055088657, ISSN: 0002-9440, DOI: 10.1016/S0002-9440(10)63681-4 (56) References cited: • MU ET AL.: ’The integrin alphavbeta8 mediates WO-A1-2010/022737 WO-A2-2011/020529 epithelial homeostasis through MT1- WO-A2-2011/103490 US-A- 5 635 601 MMP?dependent activation of TGF-betal’ J US-A1- 2004 170 630 US-A1- 2005 002 934 CELLULAR BIOLOGY vol. 157, 29 April 2002, pages 493 - 507, XP055071726 Note: Within nine months of the publication of the mention of the grant of the European patent in the European Patent Bulletin, any person may give notice to the European Patent Office of opposition to that patent, in accordance with the Implementing Regulations. Notice of opposition shall not be deemed to have been filed until the opposition fee has been paid. (Art. 99(1) European Patent Convention). EP 2 744 823 B1 Printed by Jouve, 75001 PARIS (FR) (Cont. next page) EP 2 744 823 B1 • ARAYA ET AL.: ’Integrin-Mediated Transforming Growth Factor-beta Activation Regulates Homeostasis of the Pulmonary Epithelial-Mesenchymal Trophic Unit’ AM J PATHOLOGY vol. 169, August 2008, pages 405 - 415, XP002501190 2 EP 2 744 823 B1 Description BACKGROUND OF THE INVENTION 5 [0001] The multifunctional cytokine transforming growth factor-β (TGF-β) affects immune, endothelial, epithelial, and mesenchymal cells during development and adult life in invertebrate and vertebrate species. In mammals, these functions are mediated by three widely expressed isoforms, TGF- β1, 2, and 3. All three isoforms interact with the same cell surface receptors (TGFBR2 and ALK5) and signal through the same intracellular signaling pathways, which involve either ca- nonical (i.e., SMADs) or noncanonical (i.e., MAPK, JUN, PI3K, PP2A, Rho, PAR6) signaling effectors. In the canonical 10 TGF-β signaling pathway, the signal is propagated from the TGF- β receptor via phosphorylation of cytoplasmic SMAD- 2/3, complex formation with SMAD-4, translocation of the SMAD-2/3/4 complex to the nucleus, and binding to SMAD response elements located in the promoter regions of many genes involved in the fibrogenic response. While the TGF- β isoforms have similar signaling partners, each serves distinct biological functions. The TGF- β isoforms have differences in binding affinities to TGF- β receptors, activation mechanism, signaling intensity or duration, or spatial and/or temporal 15 distribution. [0002] Knockout and conditional deletion models of TGF-β isoforms, receptors, and signaling mediators, as well as function-blocking reagents targeting all TGF- β isoforms, have revealed essential roles for TGF- β in T-cell, cardiac, lung, vascular,and palate development. Mice deficientin TGF- β1 either diein utero, owing to defects inyolk sac vasculogenesis, or survive to adulthood with severe multiorgan autoimmunity. Genetic deletion of TGF-β signaling mediator Smad2 20 reveals that it is essential in early patterning and mesodermal formation. Mice lacking Smad3 are viable and fertile, but exhibit limb malformations, immune dysregulation, colitis, colon carcinomas, and alveolar enlargement. In adult tissues, the TGF-β pathway is involved in the immune, mesenchymal, and epithelial cell interactions to maintain homeostasis in response to environmental stress. [0003] The homeostatic pathways mediated by TGF-β are perturbed in response to chronic repetitive injury. TGF-β 25 is a major profibrogenic cytokine in response to injury, delaying epithelial wound healing. TGF-β inhibits epithelial pro- liferation and migration, promotes apoptosis, and expands the mesenchymal compartment by inducing fibroblast recruit- ment, fibroblast contractility, and extracellular matrix deposition. Intratracheal transfer of adenoviral recombinant TGF- β 1 to the rodent lung dramatically increases fibroblast accumulation and expression of type I and type III collagen around airways and in the pulmonary interstitium. Neutralizing anti-TGF-β antibodies can block bleomycin or radiation-induced 30 pulmonary fibrosis. [0004] Increased TGF-β activity can play a role in fibrotic lung disease, glomerulosclerosis, and restenosis of cardiac vessels, primarily mediated by TGF-β. TGF-β1 function in humans is complex, as indicated by hereditary disorders involving either TGF-β1 itself or its signaling effectors. Mutations that increase the activity of the TGF-β pathway lead to defects in bone metabolism (ie, Camurati-Engelmann disease), in connective tissue (ie, Marfan syndrome), and in 35 aortic aneurysms (ie, Loeys-Dietz syndrome). Mutations that lead to decreased activity of the TGF- β pathway correlate with cancer. The role of TGF-β as a tumor suppressor in cancer is not straightforward, however, because TGF-β can also enhance tumor growth and metastasis. [0005] Despite the multiple essential functions of TGF-β, a single dose or short-term administration of a pan-TGF-β neutralizing antibody is well tolerated. No side effects are observed in rodents at doses that inhibit organ fibrosis or 40 carcinoma cell growth and metastasis. This treatment also effectively inhibits experimental fibrosis. Single-dose phase I/II clinical trials using neutralizing pan-TGF-β antibodies are ongoing for metastatic renal cell carcinoma, melanoma, focal segmental glomerulosclerosis, and idiopathic pulmonary fibrosis (Genzyme Corporation, available at genzyme- clinicalresearch.com). Neurohr et al discloses a murine monoclonal αvβ8-blocking antibody that significantly increases the degree of wound closure by inhibiting TGF- β activation (Am J Respir Cell Mol Biol, Vol 35, pp 252-259, 2006). 45 BRIEF SUMMARY [0006] Provided herein are antibody compositions that can be used for diagnosis and treatment of disorders associated with elevated TGF- β activity mediated by αvβ8. In some embodiments, provided is a humanized antibody that specifically 50 binds αvβ8 integrin, wherein the antibody inhibits release of active, mature TGFβ peptide, but does not significantly inhibit adhesion of latent TGF β to αvβ8 integrin on a αvβ8-expressing cell, and wherein the antibody has a heavy chain variable region of SEQ ID NO:8 and a light chain variable region of SEQ ID NO:9. [0007] The disclosure also provides an isolated antibody that specifically binds αvβ8, wherein the isolated antibody inhibits release of active, mature TGFβ peptide, but does not significantly inhibit adhesion of latent TGFβ to αvβ8 on a 55 αvβ8-expressing cell, and wherein the isolated antibody binds fixedα vβ8-expressing cells (e.g., formalin fixed). The antibody with these activities is referred to as 11E8, which term includes affinity-matured, humanized, chimeric, and labeled versions of the 11E8 antibodies, as well as αvβ8-binding fragments thereof. In some aspects of the disclosure, the isolated antibody specifically binds to an epitope on β8 that is within SEQ ID NO:1. In some aspects of the disclosure, 3 EP 2 744 823 B1 the antibody comprises the heavy chain CDRs shown in SEQ ID NO:10 (SEQ ID NOs:48, 49, and 50). In some aspects of the disclosure, the antibody comprises the light chain CDRs shown in SEQ ID NO:11 (SEQ ID NOs:51, 52, and 53). In some aspects of the disclosure, the antibody comprises the heavy chain CDRs shown in SEQ ID NO:10 and the light chain CDRs shown in SEQ ID NO:11. In some aspects of the disclosure, the antibody comprises the heavy chain variable 5 region shown in SEQ ID NO:10. In some aspects of the disclosure, the antibody comprises the light chain variable region shown in SEQ ID NO:11. In some aspects of the disclosure, the antibody comprises the heavy chain variable region shown in SEQ ID NO:10 and the light chain variable region shown in SEQ ID NO:11.