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( 12 ) United States Patent US010106817B2 (12 ) United States Patent ( 10 ) Patent No. : US 10 , 106 ,817 B2 Weinberger ( 45 ) Date of Patent: Oct . 23 , 2018 ( 54 ) COMPOSITIONS AND METHODS OF USE 2009/ 0210952 A1* 8/ 2009 Wu .. .. .. C12N 15 /63 THEREOF FOR IDENTIFYING ANTI -VIRAL 800 / 13 2011/ 0136896 A1 * 6 /2011 Fu .. .. .. CO7K 14 /005 AGENTS 514 /44 R 2012/ 0156238 AL 6 / 2012 Silverstein et al. ( 71 ) Applicant : The J . David Gladstone Institutes , 2013 / 0345294 A1 * 12/ 2013 Yang .. .. .. .. C12N 15 /63 San Francisco , CA (US ) 514 / 44 R ( 72 ) Inventor: Leor S . Weinberger , San Francisco , 2015 /0218584 A1 * 8 /2015 Payne .. .. .. .. .. .. C12N 15 /85 CA (US ) 435 / 455 ( 73 ) Assignee : The J . David Gladstone Institutes, FOREIGN PATENT DOCUMENTS San Francisco , CA (US ) WO WO 199303769 3 / 1993 Wo WO 199309239 5 / 1993 ( * ) Notice : Subject to any disclaimer , the term of this WO WO 199319191 9 / 1993 patent is extended or adjusted under 35 WO WO 199412649 6 / 1994 U . S . C . 154 ( b ) by 0 days . WO WO 199428938 12 / 1994 WO WO 199500655 1 / 1995 WO WO 199511984 5 / 1995 (21 ) Appl. No. : 14 / 767, 541 WO WO 2009155535 A2 * 12 / 2009 . .. A61K 31/ 7105 ( 22 ) PCT Filed : Feb . 13 , 2014 OTHER PUBLICATIONS ( 86 ) PCT No .: PCT/ US2014 /016296 Zheng C , Brownlie R , Babiuk LA , van Drunen Littel- van den Hurk $ 371 ( c ) ( 1 ) , S . Characterization of the nuclear localization and nuclear export ( 2 ) Date : Aug . 12 , 2015 signals ofbovine herpesvirus 1 VP22 . J Virol. Sep . 2005 ;79 ( 18 ) : 11864 72 . * ( 87) PCT Pub . No. : WO2014 /127148 Bieleski L , Talbot SL . Kaposi' s sarcoma- associated herpesvirus PCT Pub . Date : Aug . 21, 2014 VCyclin open reading frame contains an internal ribosome entry site . J Virol . Feb . 2001; 75 ( 4 ) :1864 - 9 . * (65 ) Prior Publication Data Souza AP, Haut L , Reyes- Sandoval A , Pinto AR . Recombinant viruses as vaccines against viral diseases . Braz J Med Biol Res . Apr. US 2016 /0002668 A1 Jan . 7 , 2016 2005 ; 38 ( 4 ) :509 - 22. Epub Apr. 13 , 2005 . * Teng MW , Bolovan - Fritts C , Dar RD , Womack A , Simpson ML , Related U . S . Application Data Shenk T , Weinberger LS . An endogenous accelerator for viral gene expression confers a fitness advantage . Cell. Dec. 21, 2012 ; 151 (7 ) :1569 ( 60 ) Provisional application No . 61/ 764 , 854, filed on Feb . 80. * 14 , 2013 Asmar J , Wiebusch L , Truss M , Hagemeier C . The putative zinc finger of the human cytomegalovirus IE2 86 - kilodalton protein is (51 ) Int . Ci. dispensable for DNA binding and autorepression , thereby demar GOIN 33 /50 ( 2006 .01 ) cating a concise core domain in the C terminus of the protein . J C12N 15 / 86 ( 2006 .01 ) ( 52 ) U . S . CI. Virol. Nov. 2004 ;78 ( 21 ) : 11853 -64 . * CPC .. .. C12N 15 /86 ( 2013 .01 ) ; GOIN 33 /5023 ( Continued ) ( 2013 . 01 ) ; C12N 2710 / 16122 ( 2013 .01 ) ; CI2N 2710 / 16143 ( 2013 .01 ) ; C12N 2740 / 15043 Primary Examiner — Catherine S Hibbert ( 2013 .01 ) ; C12N 2830 /00 (2013 .01 ) ; C12N 2830 /48 (2013 .01 ) ; C12N 2830 /60 (2013 .01 ) ; ( 74 ) Attorney , Agent, or Firm — Michael B . Rubin ; C12N 2840 / 203 (2013 . 01 ) Bozicevic , Field & Francis LLP (58 ) Field of Classification Search None See application file for complete search history. (57 ) ABSTRACT A recombinant expression vector comprising a nucleotide ( 56 ) References Cited sequence encoding a herpesvirus transactivator , where the nucleotide sequence is operably linked to a herpesvirus U . S . PATENT DOCUMENTS control element is provided as are cell lines genetically 5 , 674 ,730 A * 10 / 1997 Baim .. .. .. .. CO7K 14 / 005 modified to express a herpesvirus transactivator under the 435 /320 . 1 control of a herpesvirus control element. Also provided are 5 , 945 ,276 A * 8 / 1999 Wu .. .. .. .. C12N 15 /85 435 / 235 . 1 methods of identifying agents that disrupt feedback regula 2002/ 0065243 A1 * 5 / 2002 Fung .. A61K 38 / 1709 tion of a herpesvirus transcriptional control element by a 514 /44 A herpesvirus transactivator . 2002/ 0151049 A1 * 10 / 2002 Mueller .. .. .. A61K 38 /47 435 /320 . 1 2005/ 0003343 A1 * 1/ 2005 Palese .. C12N 15 / 1086 435 / 5 24 Claims, 62 Drawing Sheets 2005/ 0266564 Al 12/ 2005 Yao et al . Specification includes a Sequence Listing. US 10 ,106 ,817 B2 Page 2 (56 ) References Cited Choi , K . 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" ; J Virol. 78 ; pp . 5491 -5499 . content/ dam / sigma- aldrich /docs / Sigma - Aldrich /Brochure / 1 / epitope Deshaies , R . J ., and Ferrell, J. E . , Jr. ; ( 2001) “ Multisite phosphorylation tags - in - protein - research .pdf . * and the countdown to S phase ” ; Cell107 ; pp . 819 - 822 . Dull , T . et al ; ( 1998 ) “ A third - generation lentivirus vector with a Luke G , Escuin H , De Felipe P , Ryan M . 2A to the fore — research , conditional packaging system ” ; J Virol 72 ; pp . 8463 - 8471 . technology and applications. Biotechnol Genet Eng Rev. 2010 ; 26 : 223 Dwarakanath , R . S . et al ; (2001 ) “ The use of recombinant baculoviruses 60 . * for sustained expression of human cytomegalovirus immediate Fukui et al ( " Establishment of a Cell - Based Assay for Screening of early proteins in fibroblasts ” ; ' Virology 284 ; pp . 297 - 307 . Compounds Inhibiting Very Early Events in the Cytomegalovirus Everett, R . D . , and Chelbi- Alix , M . K ., ( 2007 ) “ PML and PML Replication Cycle and Characterization of a Compound Identified nuclear bodies: implications in antiviral defence " ; Biochimie 89 ; pp . Using the Assay " Antimicrobiol ( Year : 2008 ) . * 819 -830 . Lu , Rui et al ( 1998 ) “ The herpesvirus transactivator VP16 mimics Fan , S . et al; (2005 ) “ Valproic acid enhances gene expression from a human basic domain leucine zipper protein , luman , in its inter viral gene transfer vectors ” ; J Virol Methods 125 , pp . 23 -33 . action with HCF ” ; Journal of Virology ; pp . 6291 -6297 . Flannery et al. , ( 1997 ) ; “ Efficient photoreceptor- targeted gene expres Luganini, A . et al ; (2008 ) “ New cell -based indicator assays for the sion in vivo by recombinant adeno -associated virus” ; PNAS 94 : pp . detection of human cytomegalovirus infection and screening of 6916 -6921 . inhibitors of viral immediate -early 2 protein activity ” ; Journal of Flemington ; (2001 ) “ Herpesvirus lytic replication and the cell cycle : Applied Microbiology 105 ( 6 ) ; pp . 1791 - 1801 . arresting new developments ” ; J Virol. 75 ( 10 ) :4475 -4481 . Papworth , Monika et al ( 2003 ) “ Inhibition of herpes simplex virus Flotte et al ., ( 1993 ) " Stable in vivo expression of the cystic fibrosis 1 gene expression by designer zinc - finger transcription factors” transmembrane conductance regulator with an adeno -associated Proc Natl Acad Sci US A . 100 ( 4 ) ; pp . 1621 - 1626 . virus vector” ; PNAS 90 ( 22 ) : 10613 - 10617 . Ali et al ; ( 1996 ) “Gene transfer into the mouse retina mediated by Garcia - Pino , A . et al ; ( 2010 ) “ Allostery and intrinsic disorder an adeno -associated viral vector ” Hum Mol Genet 5 : pp . 591 594 . mediate transcription regulation by conditional cooperativity ” ; Cell Ali et al ; ( 1998 ) " Adeno - Associated Virus Gene Transfer to Mouse 142 , pp . 101 - 111. Retina ” ; Human Gene Therapy 9 ; pp . 81 86 . Gardner, T . S . et al; ( 2000 ) “ Construction of a genetic toggle switch Barrasa , M . I. et al; (2005 ) “ The phosphorylation status of the in Escherichia coli” ; Nature 403 ; pp . 339 - 342 . serine - rich region of the human cytomegalovirus 86 -kilodalton Gautier , I. , et al ; ( 2001) “ Homo - FRET microscopy in living cells to major immediate - early protein IE2 / IEP86 affects temporal viral measure monomer -dimer transition of GFP - tagged proteins" ; Biophys gene expression " ; Journal of Virology 79 , pp . 1428 - 1437 . J 80 ; pp . 3000 - 3008 . Bennett et al; “ Real - Time, Noninvasive in Vivo Assessment of Gebert , S . et al; ( 1997 ) “ The UL84 protein of human cytomegalovirus Adeno - Associated Virus- Mediated Retinal Transduction " ; Investi acts as a transdominant inhibitor of immediate -early -mediated gative Ophthalmology & Visual Science, vol . 38 , No . 13; (Dec . transactivation that is able to prevent viral replication ” ; J Virol 71; 1997 ) pp . 2857 - 2863 pp . 7048 -7060 . Black , H . S . ( 1999 ) “ Stabilized feed -back amplifiers ” ; (Reprinted Haase , A . T ., (2010 ) “ Targeting early infection to prevent HIV - 1 from Electrical Engineering , vol. 53 , p . 114 - 120 , 1934 ) . Proceed mucosal transmission ” ; Nature 464 ; pp . 217 - 223 . ings of the IEEE 87 , pp . 379 - 385 . Hofmann , H ., et al ; (2000 ) “ Covalent modification of the transactiva Bolovan - Fritts, C . and Wiedeman , J . A . (2001 ). “ Human cytomegalovirus tor protein IE2 - p86 of human
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