THE NATIONALUS009816080B2 TITOLARE DI TUTE (12 ) United States Patent ( 10 ) Patent No. : US 9 ,816 , 080 B2 Lu et al. ( 45 ) Date of Patent: Nov . 14 , 2017 ( 54 ) DELIVERY OF CASO VIA ( 52 ) U . S . CI. CPC . .. C12N 9 / 16 ( 2013 .01 ) ; CO7K 7 / 06 ARRDC1 -MEDIATED MICROVESICLES (2013 . 01) ; C07K 14 / 4718 (2013 .01 ) ; C12N (ARMMS ) 9 /22 ( 2013 .01 ) ; @( 71) Applicants : President and Fellows of Harvard ( Continued ) College, Cambridge , MA (US ) ; The (58 ) Field of Classification Search Board of Trustees of the Leland CPC . .. C12N 9 / 16 ; C12N 2810 /85 ; CO7K 14 /315 ; Stanford Junior University , Stanford , CO7K 14 /47 ; CO7K 2319/ 01 ; CA (US ) ( Continued ) (56 ) References Cited @( 72 ) Inventors : Quan Lu , Newton , MA (US ) ; Qiyu Wang , Boston , MA (US ) ; Stanley N . U . S . PATENT DOCUMENTS Cohen , Palo Alto , CA (US ) 7 ,479 ,573 B2 1 / 2009 Chu et al. (73 ) Assignees : President and Fellows of Harvard 8 ,680 ,069 B2 3 / 2014 de Fougerolles et al. College, Cambridge , MA (US ) ; The (Continued ) Board of Trustees of the Leland Stanford Junior University , Stanford , FOREIGN PATENT DOCUMENTS CA (US ) EP 2 604 255 Al 6 / 2013 WOWO 2011/ 127219 A1 10 / 2011 @( * ) Notice : Subject to any disclaimer , the term of this (Continued ) patent is extended or adjusted under 35 U . S . C . 154 ( b ) by 0 days. OTHER PUBLICATIONS (21 ) Appl. No. : 14 / 929 ,177 International Search Report and Written Opinion for PCT/ US2013 / 024839 , dated May 28 , 2013 . (22 ) Filed : Oct . 30 , 2015 (Continued ) (65 ) Prior Publication Data Primary Examiner — Jeffrey E Russel (74 ) Attorney, Agent, or Firm — Wolf, Greenfield & US 2016 /0206566 A1 Jul . 21, 2016 Sacks , P . C . ( 57 ) ABSTRACT Related U . S . Application Data Methods, systems, compositions and strategies for the deliv (60 ) Provisional application No .62 / 073 , 241, filed on Oct . ery of WW domain -containing fusion proteins into cells in vivo , ex vivo , or in vitro via ARMMsare provided .Methods , 31 , 2014 . systems, compositions and strategies for the delivery of (51 ) Int. Ci. Cas9 proteins and / or Cas9 variants into cells in vivo , ex CO7K 14 /47 ( 2006 . 01) vivo , or in vitro via fusion to ARMM associated proteins COZK 19 /00 (2006 .01 ) ( e . g . , ARRDC1 or TSG101 ) are also provided . ( Continued ) 17 Claims, 9 Drawing Sheets Interaction between the PPXY motif of [ 21 ??d th = } } } ??? ?f the PPXY } } } {{ Cas9 fu patel ? WW US 9 ,816 ,080 B2 Page 2 (51 ) Int. Cl. Genbank Submission ; NIH /NCBI , Accession No . NP _ 848495 . C12N 9 / 16 ( 2006 .01 ) Skarnes et al . , Feb . 26 , 2014 . C12N 15 / 87 Genbank Submission ; NIH /NCBI , Accession No . NP _ 006283 . ( 2006 .01 ) Rush et al ., May 4 , 2014 . 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