COMPOSITIONS AND METHODS FOR BASE SPECIFIC MITOCHONDRIAL GENE EDITING

DETAILED ACTION Notice of Pre-AIA or AIA Status The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA . Applicant cancels claims 1-83 and added new claims 84-106 in the amendment filed on 7/29/26. Election/Restrictions Applicant’s election without traverse of group I (new claims 84-104) in the reply filed on 7/29/26 is acknowledged. New claims 105-106 (directed to group II) are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim. Election was made without traverse in the reply filed on 7/29/26. Claim Objections Claims 86-88 are objected to because of the following informalities: because SEQ ID NO: 2 and 7 are nucleotide sequences and are not amino acid sequences. There appears to be a typographical error. Appropriate correction is required. The limitation in these claims will be considered to read on a nucleic acid comprising SEQ ID NO: 2(7). Claim Interpretation Claims 97 and 101-104 are considered independent claims because they are directed to a nucleic acid encoding the protein of claim 84 because reciting “of claim 84” is a reference to a limitation in another claim (See MPEP 2173.05(f)) or another way to recite the protein of claim 84 without including the entire limitation of claim 84 in the claim. Claim Rejections - 35 USC § 103 In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status. The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action: A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made. The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows: 1. Determining the scope and contents of the prior art. 2. Ascertaining the differences between the prior art and the claims at issue. 3. Resolving the level of ordinary skill in the pertinent art. 4. Considering objective evidence present in the application indicating obviousness or nonobviousness. This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention. Claims 84, 89-97, and 101-104 are rejected under 35 U.S.C. 103 as being unpatentable over The Regents of the University of California (WO 2019051097, published on 3/14/19 and EFD 9/8/17). ‘097 teaches a fusion polypeptide comprising DNA modifying enzyme (cytidine deaminase or adenosine deaminase) and RNA-guided endonuclease. See Pages 129-137. A polynucleotide sequence can encode the fusion polypeptide (pages 75-81). The fusion polypeptide can have a nuclear localization signal(s) (NLS) at the N-terminal or C-terminal (page 9). The fusion polypeptide can have one or more localization signal peptides, including NLS and mitochondrial localization signals for targeting the mitochondria (pages 72-74). In some cases, the RNA-guided endonuclease is inactive Cas9, including nickases and dCas9 (pages 18-30). The inactive Cas9 can have a mutation at an amino acid position corresponding to residue D10 (aspartate to alanine) and can cleave the complementary strand of a double stranded target nucleic acid but has reduced ability to cleave the non-complementary strand of the double stranded nucleic acid target (thus resulting in a single strand break (SSB) instead of a double stranded break (DSB) (page 25). However, ‘097 does not specifically teach a covalently attaching a mitochondrial localization amino acid sequence to the base editor fusion protein. It would have been prima facie obvious to a person of ordinary skill in the art before the time of the effective filing date to a covalently attaching a mitochondrial localization amino acid sequence to the base editor fusion protein to assist in delivery of the protein to the mitochondrial of a cell to be edited, namely to arrive at the claimed invention. See MPEP 2413(I)A and B. ‘097 teaches that the fusion protein comprises either cytidine deaminase or adenosine deaminase. ‘097 further teaches that the fusion polypeptide can have one or more localization signal peptides, mitochondrial localization signals for targeting the mitochondria (pages 72-74). One of ordinary skill in the art would have been motivated to combine the teaching to attach a mitochondrial localization sequence for targeting the mitochondria. ‘097 teaches using a dCas9 or Cas9 nickase in the fusion protein, including a SpCas9D10A. Since there are four locations to insert the localization signal peptide into the fusion polypeptide, it would have been obvious to try connecting the mitochondrial localization amino acid at the N-terminal of the RNA-guided DNA endonuclease enzyme (dCas9). See MPEP 2143(I)E. ‘097 teaches making a vector (viral vector or lipid particle) comprising the fusion polynucleotide or a nucleic acid sequence encoding the fusion polypeptide. When making a lipid particle comprising the nucleic acid, a person of ordinary skill in the art would have been motivated to complex the nucleic acid with the lipid particle since that is how lipid particles comprising a nucleic acid are made. It would have been obvious for one of ordinary skill in the art to make a pharmaceutical composition comprising the fusion polypeptide or vector comprising the fusion polypeptide or complexed with the nucleic acid encoding thereof to store for future usage or to assist in delivery of the fusion polypeptide to a cell. Therefore, the invention as a whole would have been prima facie obvious to one ordinary skill in the art before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Claims 85, 86 and 88 are rejected under 35 U.S.C. 103 as being unpatentable over The Regents of the University of California (WO 2019051097, published on 3/14/19 and EFD 9/8/17) as applied to claims 84, 89-97, and 101-104 above, and further in view of Anderson (US 20140066388). ‘097 does not specifically teach using a cytochrome c oxidase subunit VII sequence having at least 90% sequence identity to SEQ ID NO: 1 or 2 as the mitochondrial localization sequence. However, Anderson teaches making a nanoparticle comprising an inhibitor and a mitochondrial location sequence (MLS) to deliver the inhibitor to the mitochondria, including an MLS peptide comprising SEQ ID NO: 2, which is 100% identical to instant SEQ ID NO: 1. See page 12. Instant SEQ ID NO: 1 (Qy) and SEQ ID NO: 2 of ‘388 (Db) Qy 1 MSVLTPLLLRGLTGSARRLPVPRAKIHSL 29 Db 1 MSVLTPLLLRGLTGSARRLPVPRAKIHSL 29 It would have been prima facie obvious to a person of ordinary skill in the art before the time of the effective filing date to combine the teaching of ‘097 taken with Anderson to use instant SEQ ID NO: 2 as the MLS peptide in the fusion polypeptide taught by ‘097 to deliver the polypeptide to mitochondria Therefore the invention as a whole would have been prima facie obvious to one ordinary skill in the art before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Claims 85, 87 and 88 are rejected under 35 U.S.C. 103 as being unpatentable over The Regents of the University of California (WO 2019051097, published on 3/14/19 and EFD 9/8/17) as applied to claims 84, 89-97, and 101-104 above, and further in view of Liu et al. (US 20250011748, EFD 1/28/20). ‘097 does not specifically teach using a cytochrome c oxidase subunit VII sequence having at least 90% sequence identity to SEQ ID NO: 6 or 7 as the mitochondrial localization sequence. However, Liu claims a non-naturally occurring polypeptide variant comprising a double stranded DNA deaminase activity further comprising a targeting sequence, wherein the targeting sequence is mitochondrial targeting sequence (MTS) comprising SEQ ID NO: 13 (page 441). Instant SEQ ID NO: 6 (Qy) and SEQ ID NO: 13 (Db). Qy 1 MLSRAVCGTSRQLAPVLGYLGSRQKHSLPD 30 Db 1 MLSRAVCGTSRQLAPVLGYLGSRQKHSLPD 30 It would have been prima facie obvious to a person of ordinary skill in the art before the time of the effective filing date to combine the teaching of ‘097 taken with Liu to use instant SEQ ID NO: 6 as the MLS peptide in the fusion polypeptide to deliver the polypeptide to the mitochondria. Therefore the invention as a whole would have been prima facie obvious to one ordinary skill in the art before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Claims 85, 87 and 88 are rejected under 35 U.S.C. 103 as being unpatentable over The Regents of the University of California (WO 2019051097, published on 3/14/19 and EFD 9/8/17) as applied to claims 84, 89-97, and 101-104 above, and further in view of Corral-Debrinski et al. (US 20090306188). ‘097 does not specifically teach using a superoxide dismutase 2 (SOD2) sequence having at least 90% sequence identity to SEQ ID NO: 6 or 7 as the mitochondrial localization sequence. However, Corral-Debrinski teaches a SOD2 mitochondrial targeting sequence (MTS) comprising SEQ ID NO: 31 that is 100% sequence identity to instant SEQ ID NO: 7. See Figure 10. Qy 1ATGTTGAGCCGGGCAGTGTGCGGCACCAGCAGGCAGCTGGCTCCGGTTTTGGGGTATCTG 60 Db 1ATGTTGAGCCGGGCAGTGTGCGGCACCAGCAGGCAGCTGGCTCCGGTTTTGGGGTATCTG 60 Qy 61 GGCTCCAGGCAGAAGCACAGCCTCCCCGAC 90 Db 61 GGCTCCAGGCAGAAGCACAGCCTCCCCGAC 90 It would have been prima facie obvious to a person of ordinary skill in the art before the time of the effective filing date to combine the teaching of ‘097 taken with Corral-Debrinski to use instant SEQ ID NO: 6 as the MLS peptide in the fusion polypeptide to deliver the polypeptide to the mitochondria. Therefore the invention as a whole would have been prima facie obvious to one ordinary skill in the art before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Claims 85 and 98-100 are rejected under 35 U.S.C. 103 as being unpatentable over The Regents of the University of California (WO 2019051097, published on 3/14/19 and EFD 9/8/17) as applied to claims 84, 89-97, and 101-104 above, and further in view of Bin Li (US 20200263172, EFD 7/1/19). ‘097 does not specifically teach using a nucleotide sequence having at least 90% sequence identity to SEQ ID NO: 8 at the 3’ end of the sequence. However, Li teaches a recombinant nucleic acid comprising a mitochondria targeting sequence (MTS), a mitochondria protein coding sequence and a 3’ UTR, wherein the 3’ UTR comprises a sequence selected from SEQ ID NOs: 111-125. SEQ ID NO: 119 (Db) of Li is 100% identical to instant SEQ ID NO: 8 (Qy). Qy 1 ACCACGATCGTTATGCTGAGTATGTTAAGCTCTTTATGACTGTTTTTGTAGTGGTATAGA 60 Db 1 ACCACGATCGTTATGCTGAGTATGTTAAGCTCTTTATGACTGTTTTTGTAGTGGTATAGA 60 Qy 61 GTACTGCAGAATACAGTAAGCTGCTCTATTGTAGCATTTCTTGATGTTGCTTAGTCACTT 120 Db 61 GTACTGCAGAATACAGTAAGCTGCTCTATTGTAGCATTTCTTGATGTTGCTTAGTCACTT 120 Qy 121 ATTTCATAAACAACTTAATGTTCTGAATAATTTCTTACTAAACATTTTGTTATTGGGCAA 180 Db 121 ATTTCATAAACAACTTAATGTTCTGAATAATTTCTTACTAAACATTTTGTTATTGGGCAA 180 Qy 181 GTGATTGAAAATAGTAAATGCTTTGTGTGATTGA 214 Db 181 GTGATTGAAAATAGTAAATGCTTTGTGTGATTGA 214 Li further teaches that MTS can be cytochrome c oxidase subunit VIII (COX8) (page 49). SEQ ID NO: 119 and COX8 can be used to target proteins to the mitochondria (pages 47-49). It would have been prima facie obvious to a person of ordinary skill in the art before the time of the effective filing date to combine the teaching of ‘097 taken with Li to use COX8 as the MTS or instant SEQ ID NO: 8 at the 3’ end of the nucleotide sequence encoding the fusion polypeptide to deliver the fusion polypeptide to the mitochondria. Therefore the invention as a whole would have been prima facie obvious to one ordinary skill in the art before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Conclusion See attached PTO-326 for disposition of claims. Any inquiry concerning this communication or earlier communications from the examiner should be directed to Brian Whiteman whose telephone number is (571)272-0764. The examiner can normally be reached on Monday thru Friday; 6:00 AM to 3:00PM. 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Should you have questions on access to the Private PAIR system, contact the Electronic Business Center (EBC) at 866-217-9197 (toll-free). If you would like assistance from a USPTO Customer Service Representative or access to the automated information system, call 800-786-9199 (IN USA OR CANADA) or 571-272-1000. /BRIAN WHITEMAN/ Primary Examiner, Art Unit 1636