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 .
Information Disclosure Statement
The information disclosure statement (IDS) submitted on 01/03/2024 has been considered.
Sequence Compliance
Specific deficiency – Nucleotide and/or amino acid sequences appearing in the drawings are not identified by sequence identifiers in accordance with 37 CFR 1.821(d). Sequence identifiers for nucleotide and/or amino acid sequences must appear either in the drawings or in the Brief Description of the Drawings.
Required response – Applicant must provide:
Replacement and annotated drawings in accordance with 37 CFR 1.121(d) inserting the required sequence identifiers;
AND/OR
A substitute specification in compliance with 37 CFR 1.52, 1.121(b)(3) and 1.125 inserting the required sequence identifiers into the Brief Description of the Drawings, consisting of:
A copy of the previously-submitted specification, with deletions shown with strikethrough or brackets and insertions shown with underlining (marked-up version);
A copy of the amended specification without markings (clean version); and
A statement that the substitute specification contains no new matter.
Appropriate action is required so that the sequences disclosed in the specification comply with the sequence rules as discussed in MPEP 2421, MPEP 2422, and 37 CFR 1.821 through 1.825. Figure 8 fails to comply with the sequence rules.
The sequence rules embrace all unbranched nucleotide sequences with ten or more nucleotide bases and all unbranched, non-D amino acid sequences with four or more amino acids, provided that there are at least 10 "specifically defined" nucleotides or 4 "specifically defined" or amino acids. The rules apply to all sequences in a given application, whether claimed or not. All such sequences are relevant for the purposes of building a comprehensive database and properly assessing prior art. It is therefore essential that all sequences, whether only disclosed or also claimed, be included in the database. See MPEP 2421.02.
Applicant should carefully review the entire specification to ensure compliance with the sequence rules. Applicant should provide a corresponding sequence identifier (SEQ ID NO) with every appearance of a sequence embraced by the sequence rules. If a sequence embraced by the sequence rules is lacking a corresponding sequence identifier (SEQ ID NO) in the instant Sequence Listing, Applicant should provide the sequence in a substitute computer readable form (CRF) copy and a substitute paper copy of the Sequence Listing, as an amendment specifically directing its entry into the application. Applicant should also provide a statement that the content of the paper and computer readable copies are the same and, where applicable, include no new matter, as required by 37 C.F.R.1.821(e) or 1.821(f) or 1.821(g) or 1.825(b) or 1.825(d). Applicant should provide a statement indicating any changes made to the Sequence Listing. Appropriate action is required in reply to this Office action. See attached PTO-2301.
Status of Application/Claims
Claims 1-13, 15-20, and 48-50, filed 10/19/2023, are pending. Claims 1-13, 15-20, and 48-50 are the subject of the present Official action.
Claim Rejections - 35 USC § 112
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claim 6 is rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
With regard to Claim 6, residue 713 is listed as part of both the N-dCas9 protein and the C-dCas9 protein. Additionally, the sequence length of SEQ ID NO:1 is 1367 residues, and the claim recites that the C-dCas9 protein consists of the 713 to 1368 segment.
Claims 10-11 are rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
Claim 10 recites the limitation "third polynucleotide sequence". There is insufficient antecedent basis for this limitation in the claim.
Dependent claim 11 is included in the basis of the rejection because it does not correct the deficiencies of the claim upon which it depends.
Claims 13 and 49 are rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
Regarding claim 13, the phrase "preferably" renders the claim indefinite because it is unclear whether the limitation(s) following the phrase are part of the claimed invention. See MPEP § 2173.05(d). Additionally, the MPEP states that use of a narrow numerical range that falls within a broader range in the same claim may render the claim indefinite when the boundaries of the claim are not discernible. Description of examples and preferences is properly set forth in the specification rather than in a single claim. A narrower range or preferred embodiment may also be set forth in another independent claim or in a dependent claim. If stated in a single claim, preferences lead to confusion over the intended scope of the claim. See MPEP § 2173.05(c). It is unclear if two or three NLS are part of the claimed invention.
Dependent claim 49 is included in the basis of the rejection because it does not correct the deficiencies of the claim upon which it depends.
Claim 15 is rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
Claim 15 recites the limitation "third expression cassette". There is insufficient antecedent basis for this limitation in the claim.
Claim Rejections - 35 USC § 103
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.
Claims 1-6, 9-13, 15-18, and 48-50 are rejected under 35 U.S.C. 103 as being unpatentable over Church et al. (WO2016112242A1; hereinafter Church), in further view of Liu et al., 2014 (US10077453B2; hereinafter Liu).
With regard to Claims 1 and 9, Church teaches viral delivery of two separate polynucleotides with the first nucleotide encoding the N-terminal lobe of a nuclease-null Cas9 (dCas9) protein fused on its C-terminus with the Rhodothermus marinus (Rma) N-split-intein and the second nucleotide encoding the C-terminal lobe of a dCas9 protein with the complementary Rma C-split-intein. Church teaches that this reduces the protein coding sequences to 2.5kb and 2.2kb respectively, and provides >2 kb within each AAV for incorporation of a transcriptional activator, gRNAs, and fusion domains. See, for example, the abstract and page 36, lines 29-33 (Example VIII) of Church. Church teaches that the two separate nucleotide sequences are DNA sequences that comprise at least one coding sequence operably linked to a promoter that directs transcription of the coding sequence, which meets the definition of the first and second expression cassette of the instant case. See, for example, Fig.1, page 52 lines 21-34, and pages 53-56 of Church.
Church does not teach a transcriptional activator located at the N-terminus to the N-terminal half of the dCas9 protein. Church also does not teach an epigenetic modifier tethered to the dCas9 protein.
Liu teaches viral delivery of a polynucleotide encoding a dCas9 protein with a transcriptional activator located at the N-terminus to the dCas9 protein and an epigenetic modifier at the C-terminus to the dCas9 protein. See, for example, column 71, paragraph 1 and column 74, paragraph 1 of Liu. Liu also teaches that the epigenetic modifier domain encodes the full protein or the catalytic domain of the epigenetic modifier TET1 hydroxylase. See, for example, column 1, paragraph 4 and column 18, lines 24-32 of Liu. Liu also teaches that the target cell of the viral vector can be a eukaryotic cell in a human subject. See, for example, column 8 lines 45-49 of Liu.
It would have been prima facie obvious to one of ordinary skill in the art to modify the split dCas9 system of Church to further comprise a transcriptional activator at the N-terminus and an epigenetic modifier such as the catalytic domain of TET1 hydroxylase at the C-terminus of the dCas9 protein, in view of Liu, to reversibly enhance protein expression through epigenetic modification rather than cutting the gene through alternate gene editing techniques. One would have been motivated to make such a modification with a reasonable expectation of success to overcome the limited capacity of AAV which requires sequences to be under 4.7 kb. It would have been prima facie obvious to one of ordinary skill in the art to select the catalytic domain of human TET1 hydroxylase because Liu teaches that the target cell of the viral vector can be a eukaryotic cell in a human subject. One of ordinary skill in the art would have been motivated to make such a selection with a reasonable expectation of success for the purpose of administering a gene therapy to a human subject.
With regard to Claims 2-3, Church also teaches the delivery of a polynucleotide sequence encoding a guide RNA comprising a scaffold and a spacer region, wherein the spacer region hybridizes to a sequence complementary to a target sequence adjacent to the 5′ end of a protospacer adjacent motif (PAM), with both the target sequence and the PAM located within 1 kilobase (kb) of a target gene transcription start site. See, for example, page 2 lines 12-13, page 13 lines 6-12, and page 20 lines 20-21 of Church. Church teaches that the nucleotide sequence is a DNA sequence that comprises at least one coding sequence operably linked to a promoter that directs transcription of the coding sequence, which meets the definition of the third expression cassette of the instant case. See, for example, pages 25-26 (Example I) of Church. Church also teaches that the guide RNA sequence can be delivered as part of the expression cassettes encoding the N-terminal half of the dCas9 protein and the C-terminal half of the dCas9 protein, described as first and second expression cassettes in the instant case. See, for example, Fig. 4 and Fig. 18 of Church.
With regard to Claim 4, Church also teaches a dCas9-gRNA system with a transcription activator VP64. See, for example, page 19 lines 22-34 of Church.
With regard to Claim 5, Church also teaches the use of a Streptococcus pyogenes dCas9 protein. See, for example, page 18 lines 21-24 of Church.
Note that the scope of Claim 6 is indefinite as described above in the rejection under 35 U.S.C. 112(b).
With regard to Claim 6, Church teaches the sequence of a dCas9 system with the first portion of the Cas9 protein being the N-terminal lobe of the Cas9 protein up to amino acid V713 and the second portion of the Cas9 protein being the C-terminal lobe of the Cas9 protein beginning at D714. See, for example, page 9 lines 5-9 of Church.
Church does not teach the sequence consisting of SEQ ID NO:1 of the instant case.
Liu teaches the sequence of a dCas9 protein consisting of SEQ ID NO:1 of the instant case. See search results dated 02/20/26, SEQ ID NO:1 (File Names: 20260220_155502_us-18-490-443-1_copy_1_713.align150.rai and 20260220_155502_us-18-490-443-1_copy_714_1367.align150.rai), Result 18 duplicates and Result 9 duplicates.
It would have been prima facie obvious to one of ordinary skill in the art to search for a dCas9 sequence such as the SEQ ID NO:5 taught by Liu to be used as a split N-dCas9 and C-dCas9 in the method of Church. One of ordinary skill in the art would find the Cas9 sequence taught by Liu to be suitable for use in the method of Church. One of ordinary skill in the art would be motivated to apply the sequences of Liu in the method of Church with a reasonable expectation of success for the purpose of epigenetic modification taught by Liu to overcome the limited capacity of AAV which requires sequences to be under 4.7 kb.
With regard to Claim 10, Church also teaches that the first, second, or third polynucleotide sequences are operably linked to a promoter. See, for example, Fig.1, pages 25-26 (Example I), page 52 lines 21-34, and pages 53-56 of Church.
With regard to Claim 11, Church also teaches the use of a CMV promoter. See, for example, Fig. 13 and page 40 lines 24-34 (Example X) of Church.
With regard to Claims 12 and 48, Church also teaches the use of at least one nuclear localization signal (NLS) located at the N-terminus to the transcriptional activator of the N-terminal dCas9 protein. See Fig. 1 of Church. Liu teaches an NLS sequence (MAPKKKRKVGIHRGVP), listed as SEQ ID NO:42 of Liu, comprising the SV40 NLS sequence (PKKKRKV) located at the N-terminus to the transcriptional activator of the N-terminal dCas9 protein. See, for example, column 59, lines 19-28 of Liu.
With regard to Claims 13 and 49, Church also teaches the use of at least one NLS located at the C-terminus to the C-terminal half of the dCas9 protein. See Fig. 1 of Church.
Church does not teach an NLS located between the C-terminal half of the dCas9 and an epigenetic modifier.
Liu teaches that a linker can be inserted between the NLS and the epigenetic modifier and/or dCas9 domain. Liu also teaches that the NLS can be located at the C-terminal of the epigenetic modifier and/or the dCas9 domain. Liu also teaches that the NLS can be located between the epigenetic modifier and the dCas9 domain. See column 74, lines 31-37 of Liu. Liu teaches an NLS sequence (MAPKKKRKVGIHRGVP), listed as SEQ ID NO:42 of Liu, comprising the SV40 NLS sequence (PKKKRKV) located at the C-terminus to the C-terminal half of the dCas9 protein. See, for example, column 59, lines 19-28 of Liu.
With regard to Claim 15, Church teaches the nucleotide sequences comprising a coding sequence encoding two or three guide RNAs. See, for example, page 2 lines 12-13, page 20 lines 20-21, Fig. 4, and Fig. 18 of Church. Church teaches that the nucleotide sequence is a DNA sequence that comprises at least one coding sequence operably linked to a promoter that directs transcription of the coding sequence, which meets the definition of an expression cassette of the instant case. See, for example, pages 25-26 (Example I) of Church.
With regard to Claims 16-18 and 50, Church teaches the delivery of the nucleotide sequences of the N-terminal split protein, C-terminal split protein, and guide RNAs each present in separate viral vectors such as AAV vectors which are considered pharmaceutically acceptable carriers. See page 4 lines 29-31, Fig. 4, page 5 lines 22-25, and Fig. 13 of Church. Church teaches that the three nucleotide sequences are DNA sequences that comprise at least one coding sequence operably linked to a promoter that directs transcription of the coding sequence, which meets the definition of the first, second, and third expression cassettes of the instant case. See, for example, Fig.1, pages 25-26 (Example I), page 52 lines 21-34, and pages 53-56 of Church.
Claims 7-8 are rejected under 35 U.S.C. 103 as being unpatentable over Church (WO2016112242A1) and Liu (US10077453B2; 2014) as applied to Claims 1-6, 9-13, 15-18, and 48-50 above, and in further view of Xiang-Qin Liu et al. (PNAS, 1997; hereinafter XQ Liu) and Huang et al., 2020 (CN111117985A; hereinafter Huang).
With regard to Claim 7, Church teaches the use of an intein of Rhodothermus marinus (Rma). See page 11 lines 1-4 of Church.
Church does not teach that the intein is derived from the DNA helicase DnaB protein of Rma. XQ Liu teaches the discovery of the Rma intein derived from the DNA helicase DnaB protein. Given the lack of comprehensive documentation of all the potential inteins in the Rma genome and the existence of a well-characterized intein derived from the Rma DNA helicase DnaB protein, it would have been prima facie obvious to one of ordinary skill in the art that the intein of Church would be derived from the DNA helicase DnaB protein of Rma as taught by XQ Liu.
With regard to Claim 8, Church teaches sequences comprising the Rma N-split-intein in SEQ ID NO:34 and the Rma C-split-intein in SEQ ID NO:36, which are 100% matches for the 1 to 102 and 103 to 154 segments of SEQ ID NO:2 of the instant case. See the sequence alignments below.
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Church does not teach sequences consisting of the Rma N-split-intein and the Rma C-split-intein.
Huang teaches methods for splitting the Cas9 protein using an Rma intein sequence listed as SEQ ID NO:6 of Huang, which is a 100% match for SEQ ID NO:2 of the instant case. Huang teaches splitting the intein from residues 1 to 102 and 103 to 154 for use as the N-split-intein and the C-split-intein, respectively. See, for example, Claim 4 of Huang, the sequence alignment below, and the search results dated 02/19/2026, SEQ ID NO:2 (File Name: 20260219_151155_us-18-490-443-2.rag), Result 1.
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It would have been prima facie obvious to one of ordinary skill in the art to search for a Rma intein sequence such as the SEQ ID NO:6 taught by Huang to be used as an N-split-intein and C-split-intein in the method of Church. One of ordinary skill in the art would find the Rma intein sequence taught by Huang to be suitable for use in the method of Church. One of ordinary skill in the art would be motivated to apply the sequence of Huang in the method of Church with a reasonable expectation of success to overcome the limited capacity of AAV which requires sequences to be under 4.7 kb.
Claim 19 is rejected under 35 U.S.C. 103 as being unpatentable over Church (WO2016112242A1) and Liu (US10077453B2; 2014) as applied to Claims 1-6, 9-13, 15-18, and 48-50 above, and in further view of Halmai et al. (Nucleic Acids Research, 2020; hereinafter Halmai).
With regard to Claim 19, Church does not teach CDKL5 as a target gene.
Halmai teaches the use of a dCas9 system and guide RNAs targeting CDKL5, a gene causative for infantile epilepsy, for the purpose of editing DNA methylation of the gene promoter. Halmai further teaches that a fusion of the catalytic domain of TET1 to dCas9 targeted to the CDKL5 promoter using three guide RNAs causes reactivation of the inactive allele. See page 2372 of Halmai.
It would have been prima facie obvious to one of ordinary skill in the art to modify the dCas9 system of Church and Liu to target the CDKL5 gene target in view of Halmai. One of ordinary skill in the art would have been motivated to make such a combination with a reasonable expectation of success because Halmai teaches that artificial escape from X chromosome inactivation through editing DNA methylation of the CDKL5 promoter has potential as a therapy for individuals suffering from X-linked disorders. See pages 2372 and 2384 of Halmai.
Claim 20 is rejected under 35 U.S.C. 103 as being unpatentable over Church (WO2016112242A1) and Liu (US10077453B2; 2014) as applied to Claims 1-6, 9-13, 15-19, and 48-50 above, and in further view of Halmai Fink et al. (WO2021081135A2).
With regard to Claim 20, Liu does not teach the target sequences listed as SEQ IDs NO:12-14.
Halmai Fink teaches methods of editing the CDKL5 gene and lists SEQ ID NOs:1-3 that are a 100% match for SEQ ID NOs:12-14 of the instant case. See search results dated 02/05/2026, SEQ ID NOs:12-14 (File Names: 20260204_114254_us-18-490-443-12.align450.rnpbm, 20260204_114254_us-18-490-443-13.align450.rnpbm, and 20260204_114254_us-18-490-443-14.align450.rnpbm), Result 1 for each sequence. The sequences listed are consensus sequences for targeting CDKL5, and their use in the context of Church and the secondary references Liu and Halmai would have been obvious to one of ordinary skill in the art.
Claim 20 is rejected under 35 U.S.C. 103 as being unpatentable over Church (WO2016112242A1) and Liu (US10077453B2; 2014) as applied to Claims 1-6, 9-13, 15-19, and 48-50 above, and in further view of Halmai (Nucleic Acids Research, 2020) and Fyffe-Maricich et al., (WO2021163322A1).
With regard to Claim 20, Liu does not teach the target sequences listed as SEQ IDs NOs:12-14.
As stated above, Halmai teaches the use of a dCas9 system and guide RNAs targeting CDKL5. See page 2372 of Halmai. Fyffe-Maricich teaches the CDKL5 gene-specific endogenous promoter as SEQ ID NO:14 of Fyffe-Maricich. The CDKL5 gene comprises the sequence of SEQ ID NOs:12-14 of the instant case. See the sequence alignments with SEQ ID NO:14 of Fyffe-Maricich below and in the sequence search results dated 02/05/2026, SEQ ID NOs:12-14 (File Names: 20260204_114254_us-18-490-443-12.align450.rng (Result 5), 20260204_114254_us-18-490-443-13.align450.rng (Result 3), and 20260204_114254_us-18-490-443-12.align450.rng (Result 4)). Therefore, Halmai teaches targeting the CDKL5 gene which comprises SEQ ID NO:12 as claimed.
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Conclusion
Any inquiry concerning this communication or earlier communications from the examiner should be directed to HANNAH PHILIPOSE whose telephone number is (571)272-9562. The examiner can normally be reached Monday-Friday 7:30am-5pm.
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/H.P./Examiner, Art Unit 1631
/JAMES JOSEPH GRABER/Examiner, Art Unit 1631