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(12) Patent Application Publication (10) Pub. No.: US 2015/0335707 A1 Swamynathan Et Al

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(12) Patent Application Publication (10) Pub. No.: US 2015/0335707 A1 Swamynathan Et Al US 20150335707A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2015/0335707 A1 Swamynathan et al. (43) Pub. Date: Nov. 26, 2015 (54) USE OF SLURP1ASAN (22) Filed: Aug. 7, 2015 MUNOMODULATORY MOLECULE IN THE Related U.S. Application Data OCULAR SURFACE (63) Continuation of application No. 14/071,559, filed on (71) Applicant: University of Pittsburgh - Of the Nov. 4, 2013, now Pat. No. 9,132,193. Commonwealth System of Higher (60) Provisional application No. 61/722,712, filed on Nov. Education, Pittsburgh, PA (US) 5, 2012. Publication Classification (72) Inventors: Shivalingappa Kottur Swamynathan, Glenshaw, PA (US); Sudha (51) Int. Cl. Swamynathan, Glenshaw, PA (US); A638/17 (2006.01) Kristine-Ann Gallegos Buela, A6II 45/06 (2006.01) (52) U.S. Cl. Pittsburgh, PA (US); Robert Lee CPC ............... A61K 38/177 (2013.01); A61K 45/06 Hendricks, Pittsburgh, PA (US) (2013.01) (57) ABSTRACT (73) Assignee: University of Pittsburgh - Of the Methods for treating inflammation are disclosed. Such as for treating ocular inflammation. In some embodiments, the ocu Commonwealth System of Higher lar inflammation is inflammation of an ocular Surface, such as Education, Pittsburgh, PA (US) keratitis. The methods include administering to a subject with inflammation a therapeutically effective amount of SLURP1, or a nucleic acid encoding SLURP1, thereby treating the (21) Appl. No.: 14/821,567 inflammation. Patent Application Publication Nov. 26, 2015 Sheet 1 of 30 US 2015/0335707 A1 in mouse Corneas FIG. 1A Patent Application Publication Nov. 26, 2015 Sheet 2 of 30 US 2015/0335707 A1 i. PN21 No Primary Ab iv., PN21 Anti-Slurpi. A urp Ab FIG. B. Patent Application Publication Nov. 26, 2015 Sheet 3 of 30 US 2015/0335707 A1 FIG. C Patent Application Publication Nov. 26, 2015 Sheet 4 of 30 US 2015/0335707 A1 Developmental Expression of Kifa Patent Application Publication Nov. 26, 2015 Sheet 5 of 30 US 2015/0335707 A1 Patent Application Publication Nov. 26, 2015 Sheet 6 of 30 US 2015/0335707 A1 Patent Application Publication Nov. 26, 2015 Sheet 7 of 30 US 2015/0335707 A1 (IZ ?INH Patent Application Publication Nov. 26, 2015 Sheet 8 of 30 US 2015/0335707 A1 exiter weeters ested ... xxiii-x ig scies: 88:38-3: 3. 8-8, 8.3 : 8xxx FIG. 3A Promoter Activity in HCE cells ng pc-Kifa o 100 500 0 100 Reporter 500 bp surp1-Luc 150 bp Slurp1-Luc FIG. 3B Patent Application Publication Nov. 26, 2015 Sheet 9 of 30 US 2015/0335707 A1 Relative Promoter A o : so - is si s: . E. Patent Application Publication Nov. 26, 2015 Sheet 10 of 30 US 2015/0335707 A1 ChiP of KLF4 bound to SLURP1 promoter input FIG 3E Niciecticia Sequence of Siiii pi *soxia 8 for soter ra:ss.xistic}t S:t Site *:::::si:ic: :::::::ii C&xict FG, 3F Patent Application Publication Nov. 26, 2015 Sheet 11 of 30 US 2015/0335707 A1 2 o e s Patent Application Publication Nov. 26, 2015 Sheet 12 of 30 US 2015/0335707 A1 Relative levels in HSV-infected Corneas Patent Application Publication Nov. 26, 2015 Sheet 13 of 30 US 2015/0335707 A1 Slurp 1 expression in HSV-1 infected Corneas i. 1 DPS, No Primary Ab ii. 2 DPS, No Primary Ab i. 1 DPS, Anti-S FIG.SB Patent Application Publication Nov. 26, 2015 Sheet 14 of 30 US 2015/0335707 A1 C. Reiative levels in LPS-injected Corneas : s. sas-------------------------------------------------------------------------------------------------------------------------------------------. FIG.SC Patent Application Publication Nov. 26, 2015 Sheet 15 of 30 US 2015/0335707 A1 nopidae Patent Application Publication Nov. 26, 2015 Sheet 16 of 30 US 2015/0335707 A1 & HSV-1 infected corneas Io Wr Abraded a KCN Araded WT infected Kifa CN infected F.G. 6B Patent Application Publication Nov. 26, 2015 Sheet 17 of 30 US 2015/0335707 A1 si?ep+qrtoo+t-upuogºugeâffugensninsuo?daopaagenuasaudax) {{{{{} Patent Application Publication Nov. 26, 2015 Sheet 18 of 30 US 2015/0335707 A1 in m ock-& HSV-1 infected corneas Gated on CDS. Cells WTAbraded a Kf4CN Abraded WT infected y KidCN infected F.G. 6D Patent Application Publication Nov. 26, 2015 Sheet 19 of 30 US 2015/0335707 A1 Effect of Cytokines on Kift and Surp FG, 7A Kf4 and Surp in HSV-infected GKO corneas & FIG 7B Patent Application Publication Nov. 26, 2015 Sheet 20 of 30 US 2015/0335707 A1 CD45+ ceils in SV-infected GKO corneas O 40 W Abraded GKO Abraded WT infected : GKO infected FG. 7C Patent Application Publication Nov. 26, 2015 Sheet 21 of 30 US 2015/0335707 A1 Gr-1+ CD11b-- cells in HSV-infected GKO Corneas e Gated on CD45+ Cells w 8. 60. wr Abraded A GKO Abraded y GKo infected FIG. 7D Patent Application Publication Nov. 26, 2015 Sheet 22 of 30 US 2015/0335707 A1 V8 Patent Application Publication Nov. 26, 2015 Sheet 23 of 30 US 2015/0335707 A1 Patent Application Publication Nov. 26, 2015 Sheet 24 of 30 US 2015/0335707 A1 Patent Application Publication Nov. 26, 2015 Sheet 25 of 30 US 2015/0335707 A1 Number of CD11b+ Gr-1+ Cells 600. 400. 200 Patent Application Publication Nov. 26, 2015 Sheet 26 of 30 US 2015/0335707 A1 FIG. 9B Patent Application Publication Nov. 26, 2015 Sheet 27 of 30 US 2015/0335707 A1 x Controi 8.iisgfni 23.8.8 xxiiigfri , , O. O, ,5 ,4 Surg. , , , . gfrn Coliagen-i Coliagen-W Fibronectin Patent Application Publication Nov. 26, 2015 Sheet 28 of 30 US 2015/0335707 A1 ionuoo Sista 83:38 At Sirg An 3. F.G. 13 Patent Application Publication Nov. 26, 2015 Sheet 29 of 30 US 2015/0335707 A1 R 38: }{3 383 888;&BP-Siugigim} MBRi888R-Siip agai FIG. 14A F.G. 14B 38:33 is ks : 3:38 F.G. 5 Patent Application Publication Nov. 26, 2015 Sheet 30 of 30 US 2015/0335707 A1 F.G. 6A F.G. (B FIG.16C " ' ". "Mr. P. FIG. 16D 3. gif Contro: Protein 4. F.G. 17 Sip F.G. 18 US 2015/0335707 A1 Nov. 26, 2015 USE OF SLURP1ASAN In particular embodiments, methods are disclosed for treating MUNOMODULATORY MOLECULE IN THE corneal inflammation Such as keratitis, intraocular inflamma OCULAR SURFACE tion Such as uveitis or conjunctival inflammation Such as conjunctivitis in a Subject. CROSS REFERENCE TO RELATED 0009 Infurther embodiments, the methods include select APPLICATIONS ing a Subject with ocular inflammation, and administering to the subject a therapeutically effective amount of Slurp1 0001. This is a continuation of U.S. patent application Ser. polypeptide, or a polynucleotide encoding the Slurp1 No. 14/071,559, filed Nov. 4, 2013, which in turn claims the polypeptide, thereby treating the Subject. In some specific benefit of U.S. Provisional Application No. 61/722,712, filed non-limiting examples, the Subject is human. In additional Nov. 5, 2012. Both of the prior applications are incorporated non-limiting examples, the ocular inflammation is keratitis, herein by reference in their entirety. Such as keratitis caused by non-traumatic etiologies such as ACKNOWLEDGMENT OF GOVERNMENT an infection, for example viral or bacterial infection, or SUPPORT inflammation caused by non-infectious means, such as trauma. 0002 This invention was made with government support 0010. The foregoing and other objects, features, and under Grant Nos. NIH EYO16875, NIHEY 10359, and NIH advantages of the invention will become more apparent from P30EY08098 awarded by the National Eye Institute of the the following detailed description, which proceeds with ref National Institutes of Health. The government has certain erence to the accompanying figures. rights in the invention. BRIEF DESCRIPTION OF THE DRAWINGS FIELD (0011 FIGS. 1A-1C. Corneal expression of Slurp1. A. 0003. This disclosure relates to the field of inflammatory QPCR demonstrating post-eyelid opening increase in Slurp1 disorders, specifically to the use of Slurp1 and agonists expression Slurp 1 expression increases more than 15-fold thereof for the treatment of ocular inflammation. between post-natal day 11 (PN11) and PN21. B. Immunof luorescent staining of PN11, PN21 and PN56 mouse corneas BACKGROUND showing elevated expression of Slurp 1 (red) in corneal epi 0004 Injury, infection and allergy of the eye stimulate thelium in post-eyelid opening stages. C. Immunofluorescent inflammatory reactions. Humoral and cellular immunity are staining demonstrating expression of SLURP1 (green) in involved in many external eye diseases. Inflammatory corneal human corneas. Post-mortem corneal sections from a healthy disease can be infectious or non-infections, and can be vision 52 year-old male organ donor were used. Nuclei are stained threatening. The infectious causes of ocular inflammation are with DAPI (blue) and corresponding no antibody controls numerous, and include bacteria, viruses, fungi and parasites. are shown in B and C. Signals emanating from the Descem The initial events of exogenous ocular infections involve et's membrane (in panel C-iv) appear to be due to autofluo microbial adherence, invasion and multiplication. Recurrent rescence, as they were detected in no primary antibody con viral infections can also cause ocular inflammation. trols (panel C-iii) as well. Scale bars: 25 um in B and 50 mm 0005. The initial stages of ocular inflammation, such as in C. inflammation of a Surface of the eye, are often non-specific. 0012 FIGS. 2A-2D. Downregulation of Slurp 1 expres These early manifestations include pain, warmth, redness and sion in the Klf4CN cornea. A. Changes in Klf4 expression Swelling. Common symptoms of inflammatory disease of the during mouse corneal development. Absolute numbers of outer eye are itching, discomfort, dryness, redness, tearing, Klf4 transcripts per ng total RNA were calculated using the standard curve method of QPCR with total RNA from mouse discharge, and blurred vision.
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