Patent SG-11201810772X-A (SG)
Title: Diet controlled expression of a nucleic acid encoding cas9 nuclease and uses thereof
Inventor: RAVASSARD PHILIPPE
IPC: C12N15/85
Filed: 2017-06-02
Abstract: INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCT) (19) World Intellectual Property 11111 11111 11 1101 11111 111111110111111101 OE 1110111011111 11101111111110111111111 Organization International Bureau (10) International Publication Number (43) International Publication Date ......0\"\" WO 2017/207797 Al 07 December 2017 (07.12.2017) W I P0 I P C T (51) International Patent Classification: AO, AT, AU, AZ, BA, BB, BG, BH, BN, BR, BW, BY, BZ, C12N 9/22 (2006.01) Cl 2N 15/85 (2006.01) CA, CH, CL, CN, CO, CR, CU, CZ, DE, DJ, DK, DM, DO, C12N 15/867 (2006.01) DZ, EC, EE, EG, ES, FI, GB, GD, GE, GH, GM, GT, HN, (21) International Application Number: HR, HU, ID, IL, IN, IR, IS, JP, KE, KG, KH, KN, KP, KR, PCT/EP2017/063549 KW, KZ, LA, LC, LK, LR, LS, LU, LY, MA, MD, ME, MG, MK, MN, MW, MX, MY, MZ, NA, NG, NI, NO, NZ, OM, (22) International Filing Date: PA, PE, PG, PH, PL, PT, QA, RO, RS, RU, RW, SA, SC, 02 June 2017 (02.06.2017) SD, SE, SG, SK, SL, SM, ST, SV, SY, TH, TJ, TM, TN, TR, TT, TZ, UA, UG, US, UZ, VC, VN, ZA, ZM, ZW. (25) Filing Language: English (26) Publication Language: English (84) Designated States (unless otherwise indicated, for every kind of regional protection available): ARIPO (BW, GH, (30) Priority Data: GM, KE, LR, LS, MW, MZ, NA, RW, SD, SL, ST, SZ, TZ, EP16172964 03 June 2016 (03.06.2016) EP UG, ZM, ZW), Eurasian (AM, AZ, BY, KG, KZ, RU, TJ, (71) Applicants: INSERM (INSTITUT NATIONAL DE TM), European (AL, AT, BE, BG, CH, CY, CZ, DE, DK, EE, ES, FI, FR, GB, GR, HR, HU, IE, IS, IT, LT, LU, LV, LA SANTE ET DE LA RECHERCHE MEDICALE) MC, MK, MT, NL, NO, PL, PT, RO, RS, SE, SI, SK, SM, [FR/FR]; 101, rue de Tolbiac, 75013 PARIS (FR). ICM TR), OAPI (BF, BJ, CF, CG, CI, CM, GA, GN, GQ, GW, (INSTITUT DU CERVEAU ET DE LA MOELLE KM, ML, MR, NE, SN, TD, TG). EPINIERE) [FR/FR]; Hopital Pitie Salpetriere, 47/83, boulevard de l'Hopital, 75013 PARIS (FR). CENTRE Published: NATIONAL DE LA RECHERCHE SCIENTIFIQUE — with international search report (Art. 21(3)) (CNRS) [FR/FR]; 3, me Michel Ange, 75016 PARIS (FR). _ with sequence listing part of description (Rule 5.2(a)) UNIVERSITE PIERRE ET MARIE CURIE (PARIS 6) [FR/FR]; 4, place Jussieu, 75005 PARIS (FR). COM- MISSARIAT A L'ENERGIE ATOMIQUE ET AUX ENERGIES ALTERNATIVES (CEA) [FR/FR]; 25, me Leblanc, 75015 PARIS (FR). ASSISTANCE PUBLIQUE - HOPITAUX DE PARIS [FR/FR]; 3 Avenue Victoria, 75004 Paris (FR). Inventors: RAVASSARD, Philippe; Institut du Cerveau et de la Moelle Epiniere (ICM), CNRS UMR7225; INSERM (72) U975, Hopital Pitie Salpetriere Bat. ICM, 47, boulevard de l'Hopital, 75013 PARIS (FR). MALLET, Jacques; In- stitut du Cerveau et de la Moelle Epiniere (ICM), CNRS UMR7225; INSERM U975, Hopital Pitie Salpetriere Bat. ICM, 47, boulevard de l'Hopital, 75013 PARIS (FR). SER- GUERA, Ch6; INSERM US27, CEA MIRCen, 18, route du Panorama, 92265 FONTENAY-AUX-ROSES (FR). Agent: CABINET NONY; 11, me Saint-Georges, 75009 = (74) PARIS (FR). Designated States (unless otherwise indicated, for every kind of national protection available): AE, AG, AL, AM, (81) ,_ 1 (54) Title: DIET CONTROLLED EXPRESSION OF A NUCLEIC ACID ENCODING CAS9 NUCLEASE AND USES THEREOF (57) : The present invention relates to genome editing by the mean of Cas nucleases. The inventors found that the expression IN of Cas nucleases may be finely controlled by the use of regulatory elements comprising a minimal promoter and at least one amino acid cT r , response element (AARE) nucleic acid, which are responsive to a diet deficient in at least one essential amino acid, or tunicamycin. N For example, a FLAG-Cas9-GFP fusion and a Cas9-FLAG-RFP fusion could be expressed in 293 T cells. In addition, in the presence 0 of a donor plasmid bearing a puromycin resistant gene, integration of the said puromycin resistant gene may be performed at the site --.... \" of the safe harbour locus AASV1 on the genome of 293 T cells. Therefore, the invention relates to a nucleic acid for the controlled IN expression of a nucleic acid encoding a Cas nuclease in an individual, comprising (i) a regulatory polynucleotide comprising a minimal 11 © promoter and from one to twenty AARE nucleic acids, and (ii) a nucleic acid encoding a Cas nuclease, which is placed under the control ei of the said regulatory polynucleotide. C