US 2013 0330349A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2013/0330349 A1 Neufeld et al. (43) Pub. Date: Dec. 12, 2013 (54) HIGHAFFINITY MOLECULES CAPABLE OF Publication Classification BNDING ATYPE A PLEXN RECEPTOR AND USES OF SAME (51) Int. Cl. (75) Inventors: SNEutels,1ryat-11Von as; Noa rRabinoWIcz, E. Kigel, C07K 6/28 (2006.01) Haifa (IL); Asya Varshavsky, Haifa A 6LX39/395 (2006.01) (IL); Ofra Kessler, Haifa (IL) (52) U.S. Cl. (73) Assignee: Rappaport Family Institute for CPC ....... C07K 16/2863 (2013.01); A61K.39/39558 Research in the Medical Sciences, (2013.01) Haifa (IL) USPC ... 424/139.1; 530/387.3: 530/387.9; 435/375 (21) Appl. No.: 14/000,914 (22) PCT Filed: Feb. 23, 2012 (57) ABSTRACT (86). PCT No.: PCT/IL12/SOO57 A high affinity molecule is provided. The high affinity mol S371 (c)(1), ecule comprises a binding domain which binds a type-A (2), (4) Date: Aug. 22, 2013 plexin receptor, wherein said binding domain inhibits prolif Related U.S. Application Data erative signals through said type-A plexin receptor but does (60) Provisional application No. 61/445,567, filed on Feb. not interfere with binding of a neuropilin or semaphorin 6A to 23, 2011. said type-A plexin receptor. Patent Application Publication Dec. 12, 2013 Sheet 1 of 9 US 2013/0330349 A1 à 5 e S s o n ionowolysive,0Povny ked Patent Application Publication Dec. 12, 2013 Sheet 2 of 9 US 2013/0330349 A1 ced S. S. 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Ss s Ss S&ss s sessass Xss s s s 3. cd co go go c d to go go r cyd N was I (u010npu%) IOleIeygoid 80 as “ara”ugaya rea o Co. o. ab Cox |OBU00 WOI-85ue,000 - Sele WNWWeweS Patent Application Publication Dec. 12, 2013 Sheet 8 of 9 US 2013/0330349 A1 9A-YOVH-70- :e?esÁIIIeo ASA-YO:{ENA 9A-70VH-70- Patent Application Publication Dec. 12, 2013 Sheet 9 of 9 US 2013/0330349 A1 US 2013/0330349 A1 Dec. 12, 2013 HGHAFFINITY MOLECULES CAPABLE OF selected from the group consisting of a type A plexin receptor, BNDING ATYPE A PLEXN RECEPTOR AND a semaphorin a co-receptor of the type A plexin receptor and USES OF SAME a ligand of the co-receptor. 0012. According to some embodiments of the invention, FIELD AND BACKGROUND OF THE wherein the co-receptor is an FGFR or a VEGFR-2. INVENTION 0013. According to some embodiments of the invention, the high affinity molecule is selected from the group consist 0001. The present invention, in some embodiments ing of an antibody, a peptide, an aptamer and a small mol thereof, relates to high affinity molecules capable of binding ecule. a type A plexin receptor and uses of same. 0014. According to some embodiments of the invention, 0002 The human plexin gene family comprises at least the type-A plexin receptor comprises Plexin-A4. nine members in four subfamilies. 0015. According to some embodiments of the invention, 0003. The extracellular domains of plexins encompasses the binding of the binding domainto the type-A plexin recep about 500 amino acid semaphorin domains. The highly con tor comprises an affinity of at least 10 M. served cytoplasmic moieties of plexins (about 600 amino 0016. According to some embodiments of the invention, acids), however share no homology with any other known the antibody comprises a monoclonal antibody. protein. 0017. According to some embodiments of the invention, 0004 Plexin-B1 is a receptor for the transmembrane the antibody comprises a bispecific antibody. semaphorin Sema4D (CD 100), and plexin-C1 is a receptor 0018. According to some embodiments of the invention, for the GPI-anchored semaphorin Sema7A (Sema-K1). Type the bispecificantibody binds the type-A plexin receptor and at A plexins tranduce class-6 semaphorin signaling and also least one of an FGFR and semaphorin 6B. interact with neuropilins as co-receptors and tranduce the 0019. According to some embodiments of the invention, signal of class 3 semaphorins. the bispecific antibody binds a type-A1 plexin receptor and at 0005. The human gene related to the class 6 semaphorin least one of VEGFR-2 and semaphorin 6D. family termed semaphorin 6B or SEMA6B was cloned in 0020. According to some embodiments of the invention, 2001 by Correa wt al. (Genomics, 2001, 1:73(3):343-8. Two the bispecific antibody binds to distinct epitopes on the splice variants of this gene were identified. This protein sig type-A plexin receptor. nals by interacting with Plexin A4. The gene was found to be expressed in neural tissues. 0021. According to some embodiments of the invention, 0006 WO 2001/14420 teaches compositions and methods the high affinity binding molecule binds an epitope on an related to newly isolated plexins. Plexin specific binding extracellular domain of the Type A plexin receptor, the extra agents are disclosed and their use in the treatment of onco cellular domain being selected from the group consisting of a logical diseases is envisaged. Specifically disclosed is the sema domain (pfam number PFO1403) and an IgG domain. nucleic acid sequence and amino acid sequence of plexin A4. 0022. According to some embodiments of the invention, WO 2001/14420 also contemplates suppressing or altering the high affinity molecule induces internalization of the aberrant cell growth involving a signaling between plexin and plexin receptor. neuropilin using an agent (e.g., an antibody) which interferes 0023. According to an aspect of some embodiments of the with the binding between a plexin and a neuropilin. present invention there is provided an isolated antibody com 0007 U.S. Patent Application 20060228710 provides a prising an antigen recognition domain which binds a type A comprehensive list of molecular targets, such as Semaphorin plexin receptor, wherein the antibody induces internalization 6B, which can be used in the diagnosis and treatment of of the type A plexin receptor upon binding thereto. CaCC. 0024. According to some embodiments of the invention, 0008 U.S. Patent Application 20060127902 discloses a the type-A plexin receptor is selected from the group consist method of treating glioma using an antisemaphorin 6B anti ing of Plxn-A1, Plxn-A2, Plxn-A3 and Plxin-A4. body. 0025. According to some embodiments of the invention, the isolated antibody binds an epitope on an extracellular 0009 WO 2007000672 discloses peptidic antagonists of domain of the Type A plexin receptor, the domain being class III Sempahorins/neuropilins complexes comprising an selected from the group consisting of a sema domain (pfam amino acid sequence which is derived from the transmem number PFO1403) and an IgG domain. brane domain of plexin-A4 and uses thereof in the treatment 0026. According to an aspect of some embodiments of the of diseases associated with abnormal angiogenesis. present invention there is provided a method of reducing angiogenesis in a tissue, the method comprising contacting SUMMARY OF THE INVENTION the tissue with the high affinity molecule or composition or 0010. According to an aspect of some embodiments of the the antibody, thereby reducing angiogenesis in the tissue. present invention there is provided a high affinity molecule 0027. According to some embodiments of the invention, comprising a binding domain which binds a type-A plexin the contacting is effected ex-vivo. receptor, wherein the binding domain inhibits proliferative 0028. According to some embodiments of the invention, signals through said type-A plexin receptor but does not the tissue comprises a cancer tissue. interfere with binding of a neuropilin or semaphorin 6A to the 0029. According to an aspect of some embodiments of the type-A plexin receptor. present invention there is provided a method of treating an 0011. According to an aspect of some embodiments of the angiogenesis-related disorderina Subject in need thereof, the present invention there is provided a composition of matter method comprising administering to the Subject a therapeu comprising at least two distinct high affinity molecules the at tically effective amount of the high affinity biding molecule least two distinct high affinity molecules capable of binding or composition or the isolated antibody, thereby treating the and inhibiting signaling from a plexin signaling molecule angiogenesis-related disorder. US 2013/0330349 A1 Dec. 12, 2013 0030. According to an aspect of some embodiments of the lar network formed were assessed at various time points. present invention there is provided a method of treating can Represented photos of tube formation assay with HUVEC cer in a Subject in need thereof, the method comprising that were knocked down with shRNA for plexin-A1, plexin administering to the subject a therapeutically effective A3 or plexin-A4. FIG. 3B Spheroids (500 cells/spheroid) amount of the high affinity binding molecule or composition containing HUVEC expressing a non-targeted shRNA (sh or the isolated antibody of claim, thereby treating cancer. control) or HUVEC expressing a plexin-A4 targeting shRNA 0031. According to an aspect of some embodiments of the (sh-plexA4) were seeded on collagen and stimulated to sprout present invention there is provided a pharmaceutical compo with 5 ng/ml bFGF.