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 .
Election/Restrictions
Applicant’s election without traverse of Group I (claims 1-5, 7-14, 16-20, and 22) in the reply filed on August 22, 2025 is acknowledged. Applicants further elect Type II CRISPR systems, the same Type II CRISPR system (claim 11), and different promoter (claim 12).
Claims 10, 13, 20, and 23 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention and/or species, there being no allowable generic or linking claim. Election was made without traverse in the reply filed on August 22, 2025.
Claims 1-5, 7-9, 11-12, 14, 16-19, and 22 are under examination.
Information Disclosure Statement
The Information Disclosure Statement filed September 9, 2022 has been considered.
Claim Objections
Claims 4 and 18 are objected to because of the following informalities:
At claim 4, line 5, “corresponding” should be deleted.
At claim 18, line 2, “and” should be changed to “or.”
Appropriate correction is required.
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.
Claims 2, 14, 16, and 19 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.
At claim 2, line 4, it is not clear how the target nucleic acid is located on a fourth promoter. Is the target nucleic acid the promoter itself or is the target nucleic acid under the control of the fourth promoter. Further, what is the relationship between the target nucleic acid and the output nucleic acid?
Claims 14 and 16 depend from claim 2, and are therefore included in this rejection.
A broad range or limitation together with a narrow range or limitation that falls within the broad range or limitation (in the same claim) may be considered indefinite if the resulting claim does not clearly set forth the metes and bounds of the patent protection desired. See MPEP § 2173.05(c). In the present instance, claim 19 recites the broad recitation “a wild type Type II CRISPR nucleic acid and a wild type Type II tracer nucleic acid from a Lactobacillus spp. Type II system. . . and/or an Olsenella spp. Type II system,” and the claim also recites “optionally, wherein the wild type Type II CRISPR nucleic acid-tracer nucleic acid is a wild type Type II CRISPR nucleic acid and a wild type Type II tracr nucleic acid from Streptococcus pyogenes or Streptococcus thermophilus” which is the narrower statement of the range/limitation. The claim is considered indefinite because there is a question or doubt as to whether the feature introduced by such narrower language is (a) merely exemplary of the remainder of the claim, and therefore not required, or (b) a required feature of the claims.
Claim Rejections - 35 USC § 101
35 U.S.C. 101 reads as follows:
Whoever invents or discovers any new and useful process, machine, manufacture, or composition of matter, or any new and useful improvement thereof, may obtain a patent therefor, subject to the conditions and requirements of this title.
Section 33(a) of the America Invents Act reads as follows:
Notwithstanding any other provision of law, no patent may issue on a claim directed to or encompassing a human organism.
Claim 22 is rejected under 35 U.S.C. 101 and section 33(a) of the America Invents Act as being directed to or encompassing a human organism. See also Animals - Patentability, 1077 Off. Gaz. Pat. Office 24 (April 21, 1987) (indicating that human organisms are excluded from the scope of patentable subject matter under 35 U.S.C. 101).
The claim recites a cell comprising a logic-gate-based system for modifying gene expression or a genome comprising A) a least one orthogonal pair of hybridizing synthetic nucleic acid constructs, each comprising i) a CRISPR nucleic acid . . . operably linked to a first promoter and ii) a CRISPR tracer nucleic acid. . . operably linked to a second promoter. . . and B) a least one nucleic acid operably linked to a third promoter and encoding a Type II. . . CRISPR-Cas effector protein. . . wherein the CRISPR nucleic acid of each orthogonal pair . . comprise at least one non-natural mismatch between the repeat sequence of
The instant specification discloses that polynucleotides can be introduced into a host cell in a single transformation event, in separate transformation events, or, for example they can be incorporated into an organism by conventional breeding protocols (page 21, lines 6-8). The specification further discloses that stable introduction of a polynucleotide is intended that the introduced polynucleotide is stably incorporated into the genome of the cell (page 21, lines 17-18). The specification also discloses the provision of organisms and cells comprising the claimed logic-gate-based system, with the organism being a human primate (page 52, lines 1-8.).
Thus, claim 22 is deemed to encompass human cells and tissues, which are present or intended to be present in a human organism, and which is non-statutory subject matter. As such the recitation of the limitation “isolated” cell would be remedial. See 1077 Off. Gaz. Pat. Office 24 (April 21, 1987).
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 1, 3-5, 8-9, 11, 17-19, and 22 are rejected under 35 U.S.C. 103 as being unpatentable over Garreau de Loubresse et al. (U.S. Patent Application Publication No. 2019/0233806, published August 1, 2019, and cited in the Information Disclosure Statement filed September 9, 2022) in view of Lee et al. (24(3) Molecular Therapy 645-654 (2016), and cited in the Information Disclosure Statement filed September 9, 2022) and Briner et al. (56 Molecular Cell 333-339 (2014), and cited in the Information Disclosure Statement filed September 9, 2022).
Regarding claims 1, 8-9, 11-12, and 17, Garreau de Loubresse a logic-gate-based system for modifying gene expression or genome modification (paragraph [0091]). Garreau de Loubresse discloses a pair of hybridizing nucleic acid constructs (paragraph [0094]). Garreau de Loubresse discloses at least two different guide RNA systems to be used with Cas9 (a Type II CRISPR effector protein), with the first system comprising a CRISPR nucleic acid construct comprising a repeat sequence and at least one spacer sequence having substantial complementarity to a target nucleic acid (paragraph [0095] and Figure 1). Garreau de Loubresse discloses engineering sgRNAs to implement conditional logic, which allows for programmable sensing of endogenous or synthetic RNA components using logic such as "see sequence A, cut/bind sequence B", as well as more complex logic operations such c "see sequence A AND B, cut/bind sequence C" (2-lnput AND gate), the system comprising: at least one orthogonal (or independent) pair of hybridizing synthetic nucleic acid constructs, each of the at least one orthogonal pair (paragraph [0094]). Garreau de Loubresse discloses that there are at least two different guide RNA systems that can be used with Cas9 is a native system originally discovered in Streptococcus pyogenes (paragraph [0094 and Figure 1, top). Garreau de Loubresse discloses that the system includes two distinct RNA molecules referred to as tracrRNA and crRNA, with both RNA molecules are used to activate Cas9. Comprising a CRISPR nucleic acid (e.g., crRNA, crDNA) construct comprising a repeat sequence and at least one spacer sequence having substantial complementarity to a target nucleic acid (para [0095], a two-component guide system is used, the approximate length of domain "a" (also referred to as protospacer) is 20 nucleotides (+/-10 nt), the approximate length of domain "b" is 10 nucleotides (nt) (+/.1 nt) (paragraph [0095 and Figure 1). Garreau de Loubresse further discloses a trans-activating CRISPR (tracr) nucleic acid construct comprising a sequence that is complementary to the repeat sequence of the CRISPR nucleic acid of the same orthogonal pair of hybridizing synthetic nucleic acid constructs the approximate length of domain 'c' is 10 nucleotides (+/-10 nt), and the approximate length of domain 'd' is 20 nucleotides (+/-10 nt).; (paragraph [0095] and Figure 1).
Regarding claims 3, Garreau de Loubresse discloses active CRISPR-Cas nucleases, CRISPR-Cas nickases, and inactive CRISPR-Cas proteins (paragraph [0242]).
Regarding claim 18, Garreau de Loubresse discloses multiple guide RNAs and outputs, using AND and OR Boolean logic functions (paragraphs [0067]-[0070] and Figures 16-19).
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Regarding claim 19, Garreau de Loubresse discloses that the wild type CRISPR system can be a Streptococcus Cas9 (Type II) CRISPR system (Figure 1).
Regarding claim 22, Garreau de Loubresse discloses cells comprising the CRISPR-Cas9 logic-gate-based system and DNA complexes (paragraphs [0138] and [0154]-[0159]).
Garreau de Loubresse fails to disclose or suggest the specific orthogonal pairs of hybridizing synthetic nucleic acid constructs comprising at least one nucleotide mismatch. Garreau de Loubresse fails to disclose or suggest that the CRISPR nucleic acid is operably linked to a first promoter, the tracr nucleic acid operably linked to a second promoter, which, when both are expressed in a cell a synthetic CRISPR nucleic acid-tracr nucleic acid (crRNA-tracrRNA) hybrid is formed having a secondary structure comprising a lower stem, and optionally, an upper stem.
Regarding claim 1, Lee discloses a comparison of the Neisseria meningitidis (Nme) CRISPR- Cas9 system and the Streptococcus pyogenes (Spy) CRISPR-Cas9 systems targeting the same protospacer sequence (abstract). Lee discloses that the native crRNA and tracr RNA as well as a chimeric single guide RNA for the Nme CRIEPR- cas9 system were also compared (abstract, page 647, and Figure 1). Lee discloses that the crRNA is linked to a first U6 promoter and the tracr RNA is linked to a second U6 promoter (Figure 1A). This is interpreted as a CRISPR-Cas9 two-component system where the CRISPR nucleic acid is operably linked to a first promoter and the tracr nucleic acid is operably linked to a second promoter. Lee further discloses the activity of the Nme 2-RNA system and the Nme sgRNA system at the IL2RG locus in HEK293T cells, which is interpreted as both the crRNA and the tracrRNA are expressed in a cell (Figure 1C). Lee discloses a synthetic CRISPR nucleic acid-tracr nucleic acid (crRNA-tracrRNA) hybrid is formed having a secondary structure comprising a lower stem (Figures 1A-1B). Lee discloses that the Nme Cas9 protein is operably linked to the EF1α promoter, which forms a complex with the synthetic CRISPR nucleic acid-tracr nucleic acid hybrid by the orthogonal pair of hybridizing nucleic acid constructs, showing operable linkage with two different promoters (Figures 1a-1b).
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Regarding claims 4-5, Lee discloses that the native crRNA and tracr RNA as well as a chimeric single guide RNA for the Nme CRIEPR- cas9 system were also compared (abstract, page 647, and Figure 1). Lee discloses that the crRNA is linked to a first U6 promoter and the tracr RNA is linked to a second U6 promoter, with the promoters being the same (Figure 1a).
Regarding claim 12, Lee discloses that the Nme Cas9 protein is operably linked to the EF1α promoter, which is a third promoter and which forms a complex with the synthetic CRISPR nucleic acid-tracr nucleic acid hybrid by the orthogonal pair of hybridizing nucleic acid constructs, showing operable linkage with two different promoters (Figures 1a-1b).
Regarding claims 1, 8-9, and 11, Briner discloses guide RNA functional modules that are necessary for the activity of RNA-guided Cas9 endonuclease and identifies and characterizes six conserved modules within native crRNA:tracrRNA duplexes and single guide RNAs (sgRNAs) that direct Cas9 endonuclease activity (abstract). Briner discloses that the bulge and nexus are necessary for DNA cleavage and demonstrate that the nexus and hairpins are instrumental in defining orthogonality between systems (page 334, Figure 1, sgRNA Functional Modules (A)). Briner discloses the six modules include the spacer responsible for DNA targeting: the upper stem, bulge, and lower stem formed by the CRISPR repeat: tracrRNA duplex and the nexus and hairpins from the 3' end of the tracrRNA, wherein variants of these functional module affect Cas9 activity (Figure 1, (B), sgRNA guide variants). Briner discloses only the altered modules are shown, which mutated nucleotides are represented (page 334, column1 1, first paragraph to column 2, first paragraph). Briner discloses focusing on the five sgRNA modules located downstream of the spacer sequence, and tested 77 unique sgRNA variants with modified sequences (Table S1). Briner discloses that mutations in both the lower and upper stems were relatively tolerant to sequence variations (Figure 1, V1-V6 ). Briner discloses that abrogation of cleavage in cells required incorporation of two concurrent mismatches within the lower stem (Figure 1, V4; Figure S1, GV24).
Therefore, it would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have applied these nucleotide variants in the functional modules of crRNA: tracrRNA, as taught by Briner to the composition of Garreau de Loubresse and Lee, such that, instead of switch domain a synthetic oligonucleotide comprising variants of said functional modules of Briner, would have provided a genetic logic-gate-based circuits that switch the gene expression on-off conditionally.
Claims 2, 7, 14, and 16 are rejected under 35 U.S.C. 103 as being unpatentable over Garreau de Loubresse in view of Lee and Briner, as applied to claims 1, 3-5, 8-9, 11, 17-19, and 22 above, and further in view of Gander et al. (8 Nature Communications 15459, 1-11 (2017)).
Garreau de Loubresse discloses active CRISPR-Cas nucleases, CRISPR-Cas nickases, and inactive CRISPR-Cas proteins (paragraph [0242]). Garreau de Loubresse discloses an input sequence a-b-c- (trigger) binding to complex 1, displacing the intermediate DNA strand, binding to, complexing to, and displacing an output gDNA (paragraph [0140] and Figures 16-19).
Lee discloses that the native crRNA and tracr RNA as well as a chimeric single guide RNA for the Nme CRIEPR- cas9 system were also compared (abstract, page 647, and Figure 1). Lee discloses that the crRNA is linked to a first U6 promoter and the tracr RNA is linked to a second U6 promoter, with the promoters being the same (Figure 1A). Lee discloses that the Nme Cas9 protein is operably linked to the EF1α promoter, which is a third promoter and which forms a complex with the synthetic CRISPR nucleic acid-tracr nucleic acid hybrid by the orthogonal pair of hybridizing nucleic acid constructs, showing operable linkage with two different promoters (Figures 1A-1B). Therefore, Lee discloses that the promoter operably linked to the output nucleic acid can be synthetic or naturally occurring heterologous promoters, or can be endogenous to the cell or output nucleic acid promoter (Figures 1A-1B).
Briner discloses guide RNA functional modules that are necessary for the activity of RNA-guided Cas9 endonuclease and identifies and characterizes six conserved modules within native crRNA:tracrRNA duplexes and single guide RNAs (sgRNAs) that direct Cas9 endonuclease activity (abstract). Briner discloses that the bulge and nexus are necessary for DNA cleavage and demonstrate that the nexus and hairpins are instrumental in defining orthogonality between systems
Thus, Garreau de Loubresse, Lee, and Briner disclose a logic-gate-based system for modifying gene expression or modifying a genome, as discussed above.
Garreau de Loubresse, Lee, and Briner fail to disclose or suggest a fourth promoter operably linked to an output nucleic acid, or the types of promoters for each operably linked nucleic acid.
Gander discloses NOR logic circuits based on CRISPR-Cas9 with the logic circuits having up to seven guide RNAs, which is interpreted as including at least 4 promoters driving expression of the guide RNAs (abstract). Gander discloses a variety of promoters that can provide expression control of the circuit components (Figure 1).
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It would have been obvious to one with ordinary skill in the art before the effective filing date of the claimed invention to increase the number of logic circuits of the logic-gate-based CRISPR-Cas9 system disclosed and suggested by Garreau de Loubresse, Lee, and Briner according to the multi-logic-circuits of Gander because increasing the number of logic circuits will provide greater control over the expression of genes in a cell/genome modifications required by the instant claims. One of ordinary skill in the art would have been motivated to increase the logic circuit numbers and corresponding CRISPR-Cas9 systems under the control of a variety (same or different) promoters in order to direct gene expression in a controlled and reproducible manner.
Conclusion
Any inquiry concerning this communication or earlier communications from the examiner should be directed to NANCY J LEITH whose telephone number is (313)446-4874. The examiner can normally be reached Monday - Thursday 8:00 AM - 6:30 PM.
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If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, NEIL HAMMELL can be reached at (571) 270-5919. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
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NANCY J. LEITH
Primary Examiner
Art Unit 1636
/NANCY J LEITH/Primary Examiner, Art Unit 1636