US010703789B2 ( 12 ) United States Patent ( 10 ) Patent No.: US 10,703,789 B2 De Fougerolles et al. (45 ) Date of Patent: * Jul. 7 , 2020 (54 ) MODIFIED POLYNUCLEOTIDES FOR THE (2013.01 ) ; A61K 38/36 ( 2013.01 ) ; A61K PRODUCTION OF SECRETED PROTEINS 38/363 ( 2013.01 ) ; A61K 38/44 ( 2013.01) ; A61K 38/4833 (2013.01 ) ; A61K 38/4846 ( 71 ) Applicant : Moderna TX , Inc., Cambridge, MA (2013.01 ) ; A61K 39/3955 ( 2013.01) ; A61K (US ) 47/10 (2013.01 ) ; A61K 47/54 (2017.08 ) ; A61K 47/542 (2017.08 ) ; A61K 48/0033 ( 2013.01 ) ; ( 72 ) Inventors: Antonin De Fougerolles, Waterloo A61K 48/0066 (2013.01 ) ; A61K 48/0075 ( BE ) ; Justin Guild , Framingham , MA (2013.01 ) ; CO7K 14/47 ( 2013.01 ) ; CO7K (US ) 14/475 ( 2013.01) ; CO7K 14/505 (2013.01 ) ; ( 73 ) Assignee : Moderna TX , Inc., Cambridge , MA CO7K 14/525 (2013.01 ) ; C07K 14/56 (US ) (2013.01 ) ; CO7K 14/565 ( 2013.01 ) ; CO7K 14/745 (2013.01 ) ; C07K 14/75 ( 2013.01) ; ( * ) Notice: Subject to any disclaimer , the term of this CO7K 16/2887 ( 2013.01 ) ; CO7K 16/32 patent is extended or adjusted under 35 ( 2013.01) ; CO7K 19/00 ( 2013.01) ; C12N U.S.C. 154 (b ) by 0 days . 9/0069 ( 2013.01) ; C12N 9/644 ( 2013.01 ) ; C12N 15/85 (2013.01 ) ; C12N 15/88 This patent is subject to a terminal dis ( 2013.01 ) ; C12Y 113/12007 (2013.01 ) ; C12Y claimer . 304/21005 (2013.01 ) ; C12Y 304/21022 (2013.01 ) ; A61K 9/0019 (2013.01 ) ; A61K (21 ) Appl. No.: 16 /438,978 48/00 (2013.01 ) ; C12N 2840/00 (2013.01 ) ( 22 ) Filed : Jun . 12 , 2019 (58 ) Field of Classification Search CPC CO7H 21/02 , C12N 15/67; C12N 15/11 (65 ) Prior Publication Data See application file for complete search history . US 2020/0017565 A1 Jan. 16 , 2020 (56 ) References Cited Related U.S. Application Data U.S. PATENT DOCUMENTS (63 ) Continuation of application No. 14 /987,328 , filed on Jan. 4 , 2016 , now Pat . No. 10,385,106 , which is a 5,489,677 A 2/1996 Sanghvi et al . (Continued ) 5,591,722 A 1/1997 Montgomery et al. (Continued ) ( 51 ) Int. Cl. A61K 48/00 ( 2006.01 ) FOREIGN PATENT DOCUMENTS A61K 38/17 ( 2006.01) CA 2028849 A1 9/1991 A61K 47/54 ( 2017.01 ) CA 2473135 A1 6/2003 A61K 9/127 (2006.01 ) COZK 14/535 ( 2006.01 ) ( Continued ) C12N 15/88 (2006.01 ) A61K 9/50 ( 2006.01 ) OTHER PUBLICATIONS COZK 14/47 ( 2006.01 ) Anderson et al ., “ Incorporation of pseudouridine into mRNA enhances A61K 31/7088 ( 2006.01) translation by diminishing PKR activation , ” Nucleic Acids Res . CO7K 19/00 (2006.01 ) 38( 17 ) : 5884-92 ( 2010 ) . CI2N 15/85 ( 2006.01 ) A61K 38/18 ( 2006.01 ) ( Continued ) A61K 38/19 (2006.01 ) A61K 38/48 (2006.01 ) Primary Examiner — Antonio Galisteo Gonzalez A61K 9/14 ( 2006.01 ) A61K 47/10 ( 2017.01 ) (74 ) Attorney, Agent, or Firm -Clark & Elbing LLP A61K 38/21 (2006.01 ) A61K 38/36 ( 2006.01 ) (57 ) ABSTRACT A61K 38/44 ( 2006.01 ) A61K 39/395 ( 2006.01 ) A pharmaceutical composition which has a plurality of lipid (Continued ) nanoparticles that has a mean particle size ofbetween 80 nm (52 ) U.S. CI. and 160 nm and contains a modified mRNA encoding a CPC CO7K 14/535 (2013.01 ) ; A61K 9/1271 polypeptide. The lipid nanoparticles include a cationic lipid , (2013.01 ) ; AKIK 9/1272 ( 2013.01) ; A6IK a neutral lipid , a cholesterol, and a PEG lipid . The mRNA 9/1277 ( 2013.01) ; A61K 9/14 ( 2013.01) ; A61K contains a 5 '- cap , 5 '- UTR , N1- methyl - pseudouridine , a 9/5031 ( 2013.01) ; A61K 31/7088 (2013.01 ) ; 3 '- UTR , and a poly - A region with at least 100 nucleotides . A61K 38/1767 ( 2013.01) ; A61K 38/1816 ( 2013.01 ) ; A61K 38/1866 ( 2013.01) ; A61K 38/191 (2013.01 ) ; A61K 38/193 ( 2013.01) ; 14 Claims, 14 Drawing Sheets A61K 38/212 ( 2013.01 ) ; A61K 38/215 Specification includes a Sequence Listing . US 10,703,789 B2 Page 2 Related U.S. Application Data CO7K 14/56 (2006.01 ) continuation of application No. 14 / 750,004 , filed on C07K 14/565 ( 2006.01) Jun . 25 , 2015 , now Pat . No. 9,828,416 , which is a CO7K 14/745 (2006.01 ) continuation of application No. 13 / 791,921 , filed on COZK 14/75 ( 2006.01) Mar. 9 , 2013 , now Pat. No. 9,192,651 . CO7K 16/28 ( 2006.01) CO7K 16/32 ( 2006.01 ) (60 ) Provisional application No.61 / 737,174 , filed on Dec. 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