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 .
Application Status
Applicant’s remarks, sequence listing, and amendments to the claims, drawings, and specification filed December 2, 2025 are acknowledged. Claims 1-4, 7-9, 11, 13-16, and 18-20 were amended, and claims 5-6 were cancelled. Claims 1-4, and 7-20 are pending and under consideration herein.
Examiner’s Comment on Amendments to the Sequence Listing and Specification
Based on Applicant’s remarks, the sequence listing has been amended to “change the sequences of SEQ ID NOs. 3 and 4, which are obvious typing errors, to the correct ones.” Applicant indicates that the correct nucleic acid and amino acid sequences are those corresponding to “Acidaminococcus sp. BV3L6,” which were known in the prior art (see pg. 13 of remarks). These amendments do not introduce new matter because the skilled artisan would have (I) recognized the obvious error (i.e., that original SEQ ID NOs: 3-4 correspond to LbCas12a rather than AsCas12a, see paragraph 23 of the prior action), and (II) recognized that the “appropriate correction” was to substitute original SEQ ID NOs: 3-4 for the prior art sequences corresponding to AsCas12a (i.e., the sequences set forth in instant SEQ ID NOs: 3-4) based on the original disclosure (“AsCas12 [is] derived from… Acidaminococcus BV3L6,” [0004]; AsCas12 derived from Acidaminococcus sp. BV3L6,” [0031]).
Applicant’s remarks also indicate that the specification has been amended to “adjust the expressions” in certain paragraphs “for clarity.” The amended expressions relate to the “expression plasmid” and “helper”/ “tool” plasmid. These amendments do not introduce new matter because the skilled artisan would understand based on the disclosure that the terms “helper” and “tool” refer to the same type of plasmid ([0055]; [0096]), and that the “expression” plasmid is different from the “helper”/ “tool” plasmid ([0081]; [0107]). Thus, the amendments to paragraph [0096] amount to a mere rephrasing where the same meaning remains intact, and the amendments to paragraph [0075] correct an error which would have been recognized by the skilled artisan, and for which the skilled artisan would have recognized the appropriate correction made.
Withdrawn Rejections
The amendments to claim 14 are sufficient to overcome the § 101 rejection raised in the prior action. Cancellation of claims 5-6 is sufficient to overcome the § 112(b) rejections and § 102 and § 103 rejections of claims 5-6. Finally, the amendments to claim 15 are sufficient to overcome the § 102 rejection of claim 15 over Ling. The aforementioned rejections are withdrawn, accordingly.
Applicant’s remarks and amendments to the claims have been thoroughly reviewed, but are not found persuasive to place the claims in condition for allowance for the reasons that follow. Any rejection or objection not reiterated herein has been overcome by amendment.
Priority
Acknowledgment is made of Applicant's claim for priority under 35 U.S.C. 119(a)-(d) or (f), 365(a) or (b), or 386(a), and claim for benefit under 35 U.S.C. 119(e) or under 35 U.S.C. 120, 121, 365(c), or 386(c). The certified copy of Application No. CN201911092554.8, filed November 11, 2019, has been received. However, Applicant has not complied with one or more conditions for receiving the benefit of an earlier filing date as follows:
The disclosure of the prior-filed foreign application, Application No. CN201911092554.8, fails to provide adequate support or enablement in the manner provided by 35 U.S.C. 112(a) or pre-AIA 35 U.S.C. 112, first paragraph for one or more claims of this application. Specifically, the aforementioned application does not set forth the unnatural amino acid pAcF either by name or structure as recited in instant claim 7. The first disclosure of pAcF is in Application No. PCT/CN2020/127992, filed November 11, 2020 (see pg. 6). The aforementioned application also does not appear to set forth the CRISPR family proteins Cas φ, Cas12g, dCas9, dCas12a, or ddCas12a recited in instant claims 2, 4, and 15-16. The first disclosure of the aforementioned CRISPR family proteins is in Application No. PCT/CN2020/127992 (see pg. 6). Accordingly, claims 2, 4, and 7-20 are not entitled to the filing date of the foreign priority document, and the effective filing date of claims 2, 4, and 7-20 is November 11, 2020. Claims 1 and 3 under examination find support in the prior-filed foreign application. The effective filing date of claims 1 and 3 is November 11, 2019.
Claim Objections
Claims 8 and 15 are objected to because of the following informalities:
Claim 8, step (4) should preferably be amended to recite “inducing the expression of the mutated CRISPR protein in the host cell” so that the terminology is consistent throughout the claim.
Claim 15 recites “wherein the CRISPR family protein is an inactivated counterpart is selected from,” which should be amended to recite “wherein the CRISPR family protein is an inactivated counterpart [[is]] selected from,” to improve the grammar of the claim.
Appropriate correction is required.
Claim Rejections - 35 USC § 112(b)
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 1-4, 8-16, and 18-20 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. The rejections that follow are maintained and modified or new, as necessitated by Applicant’s amendments to the claims.
Claim 1 recites that “the unnatural amino acid has orthogonal chemical reactivity.” The specification states that “orthogonal chemical reactivity means, without limitation, that the unnatural amino acid contains a group such as an azide, an alkynyl, an aldehyde, a ketone, a tetrazine or a cyclopropene group” ([0030]). While setting forth examples of structures an unnatural amino acid may comprise to be considered to have “orthogonal chemical reactivity,” the specification uses terms like “without limitation,” and “such as,” such that it is not clear what other structures may/may not meet the scope of the claims, and fails to provide guidance to determine what the chemical reactivity must be “orthogonal” to. For example, while an unnatural amino acid containing an aldehyde may be “orthogonal” in one system or cell, it may be native in another system or cell. The structures of unnatural amino acids having orthogonal chemical reactivity is unclear, which renders the claims indefinite.
Claims 2-3, and 15 are rejected for depending from claim 1 and failing to remedy the indefiniteness.
Claims 1, 3-4, 8-12, 14, 16, and 20 recite an “unnatural amino acid.” The specification and claims, while providing examples of structures an unnatural amino acid may comprise (e.g., “an azido, an alkynyl”) or consist of (e.g., “pAcF,” “AeF”), do not define an “unnatural amino acid” such that the scope of the term is clear. Specifically, it is not clear whether the term “unnatural” intends to encompass I) any amino acid which is non-proteinogenic (i.e., excluding only proteinogenic amino acids like arginine, lysine, etc.), or II) any amino acid which does not occur “naturally.” In the latter case, it is also not clear what the amino acid must be “unnatural” to, e.g., the CRISPR family protein, some unrecited cell or system, etc. For example, while pyrrolysine would be “unnatural” in a eukaryotic cell, it would be “natural” in some prokaryotes. The skilled artisan would also consider amino acid substitutions within the sequence of a CRISPR family protein to be “unnatural.” Because the scope of amino acids which would be considered unnatural is unclear, the claims are indefinite.
Claims 2, 13, 15, and 18-19 are also rejected for depending from claims 1 or 4 and failing to remedy the indefiniteness. Claims 7 and 17 are not included in this rejection, because the claims clearly set forth the structure(s) required of the unnatural amino acid.
Claim 8 recites a “tool plasmid.” While the tool plasmid must be “for unnatural amino acid insertion,” this functional limitation fails to clarify what would constitute a “tool plasmid.” The specification also does not define the term. The specification provides an exemplary tool plasmid, “pUltra-MjPolyRS” ([0055]), but does not make clear what structural features are required of a plasmid to be considered a “tool” plasmid, which renders the claim indefinite.
Claims 18-19 are rejected for depending from claim 8 and failing to remedy the indefiniteness. Claim 9 is not included in this rejection, because the claim clearly sets forth the structural features of the plasmid.
Claim 14 recites “a site-specific nucleic acid conjugate of the gRNA and the CRISPR family protein.” The specification describes site-specific nucleic acid conjugates of the CRISPR family protein, e.g., “oligo DNA, a donor DNA, or a crRNA” ([0050]). However, the specification does not appear to describe any site-specific nucleic acid conjugates “of the gRNA and the CRISPR family protein.” The structural implications of this limitation are not clear. For example, it is not clear whether the limitation requires a site-specific nucleic acid conjugate “of the gRNA,” i.e., the conjugate is a gRNA, or alternatively, whether the conjugate must be able to conjugate with the gRNA and CRISPR family protein, but itself, is not a gRNA. The claim is indefinite because the site-specific nucleic acid conjugates encompassed by the method are unclear.
Response to Remarks - 35 USC § 112(b)
Applicant’s remarks regarding the § 112(b) rejections raised in the prior action have been reviewed.
Regarding the rejections over the phrases “orthogonal chemical reactivity” and “unnatural amino acid,” Applicant submits that these terms “are well known and established concepts in the art.” Applicant argues that the skilled artisan would “clearly understand the meaning of the terms.” Examiner respectfully disagrees that the skilled artisan would “clearly understand the meaning of the terms” for the reasons described in the preceding paragraphs. It is clear that the exemplary structures in the specification are within the scope of “unnatural amino acids,” e.g., pAcF, NAEK, etc. However, it is not clear what other structures are included in the scope of “unnatural amino acids” or unnatural amino acids comprising “orthogonal chemical reactivity,” because I) based on the specification, unnatural amino acids need only comprise a highly generic structure, e.g., a “ketone” group, “azide” group, or essentially any other group (“such as,” [0030]), such that the scope of “unnatural amino acids” encompasses virtually any amino acid, and II) neither the claim nor specification make clear what the amino acid must be “unnatural” or “orthogonal” to, e.g., the CRISPR family protein, some unrecited cell or system, etc. Applicant’s remarks do specifically address any of the reasons described in the rejection, and the claims have not been amended so as to overcome the lack of clarity. The rejections are maintained, accordingly. Applicant may overcome this rejection by reciting, for example, the specific compounds recited in claims 7 or 17.
Regarding the rejections over the phrase “tool plasmid,” Applicant submits that their amendments to the specification overcome the rejection. Examiner respectfully disagrees. While the amendments to the specification correct an error which inadvertently drew a distinction between a “helper” and “tool” plasmid, neither Applicant’s remarks, nor the amendments to the specification make clear the structural features which are required of a plasmid to be considered a “tool” plasmid. The claim has not been amended so as to overcome the lack of clarity. The rejection is maintained, accordingly. Applicant may overcome this rejection by reciting, for example, the structural elements recited in claim 9.
Claim Rejections - 35 USC § 112(d)
The following is a quotation of 35 U.S.C. 112(d):
(d) REFERENCE IN DEPENDENT FORMS.—Subject to subsection (e), a claim in dependent form shall contain a reference to a claim previously set forth and then specify a further limitation of the subject matter claimed. A claim in dependent form shall be construed to incorporate by reference all the limitations of the claim to which it refers.
The following is a quotation of pre-AIA 35 U.S.C. 112, fourth paragraph:
Subject to the following paragraph [i.e., the fifth paragraph of pre-AIA 35 U.S.C. 112], a claim in dependent form shall contain a reference to a claim previously set forth and then specify a further limitation of the subject matter claimed. A claim in dependent form shall be construed to incorporate by reference all the limitations of the claim to which it refers.
Claims 2 and 15 are rejected under 35 U.S.C. 112(d) or pre-AIA 35 U.S.C. 112, 4th paragraph, as being of improper dependent form for failing to further limit the subject matter of the claim upon which it depends, or for failing to include all the limitations of the claim upon which it depends. The rejections that follow are new and necessitated by Applicant’s amendments to the claims.
Claim 2 recites “wherein the CRISPR family protein comprises Cas9 protein, Cas12a protein, CasX protein, Cas φ protein, Cas12g protein from different species or genera, or an inactivated counterpart thereof having no cleavage activity but retaining binding activity.” Claim 15 recites “wherein the CRISPR family protein is an inactivated counterpart [] selected from dCas9, dCas12a protein, or ddCas12a protein.” The terms “Cas9,” “Cas12a,” “dCas9,” dCas12a,” and “ddCas12a” are interpreted as encompassing any Cas9, Cas12a, dCas9, dCas12a, and ddCas12a from any species. Claim 1 requires that the CRISPR family protein “is SpCas9…” or “is AsCas12a” (emphasis added). Thus, claims 2 and 15 encompass CRISPR family proteins which are either I) broader than the specific Cas9 and Cas12 proteins required of claim 1 (i.e., “comprises Cas9 protein, Cas12a protein… from different species or genera…,” and “dCas9, dCas12a protein, ddCas12a protein”), or II) completely different than the specific Cas9 and Cas12 proteins required of claim 1 (i.e., “CasX protein, Cas φ protein, Cas12g protein… from different species or genera…”). These claims fail to include all the limitations of the claim upon which they depend.
Applicant may cancel the claim(s), amend the claim(s) to place the claim(s) in proper dependent form, rewrite the claim(s) in independent form, or present a sufficient showing that the dependent claim(s) complies with the statutory requirements.
Claim Rejections - 35 USC § 102 – Ling
The following is a quotation of the appropriate paragraphs of 35 U.S.C. 102 that form the basis for the rejections under this section made in this Office action:
A person shall be entitled to a patent unless –
(a)(1) the claimed invention was patented, described in a printed publication, or in public use, on sale, or otherwise available to the public before the effective filing date of the claimed invention.
Claims 4, 7-14, and 16-20 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Ling (Ling et al., 8 April 2020, Science Advances, 6: eaaz0051, pg. 1-8 and Supplemental Information; of record). The rejections that follow are maintained and modified as necessitated by Applicant’s amendments to the claims.
As stated above, the effective filing date of claims 2, 4, and 7-20 is November 11, 2020.
Regarding claim 4, Ling teaches a Cas9 protein site-specifically mutated by an unnatural amino acid (“AeF”), wherein the unnatural amino acid bears an azido group (“On the basis of the structure of Streptococcus pyogenes Cas8 (SpyCas9), a total of 11 amino acids were selected as AeF mutation sites (Fig. 1A): K3, D39, H41… K1151, G1367… The codons of these residues were mutated to the TAG codon, and the mutant proteins were expressed…,” pg. 2, left col.; “K1151-AeF and G1367-AeF showed the highest expression yields and quality,” pg. 2, right col.; “we prepared and assessed two AeF-modified Cas9 D576-AeF-G1367-AeF in cell with double-adaptor conjugates,” pg. 4, right col.).
Regarding claims 7 and 17, Ling teaches the unnatural amino acid is “AeF” (Fig. 1A).
Regarding claims 8 and 18-19, Ling teaches a method for preparing an SpCas9 site-specifically mutated by the unnatural amino acid AeF (pgs. 2-3; Fig. 1-2). Ling teaches the method comprises:
(1) selecting one or more specific amino acid sites in the amino acid sequence based on structural information of the CRISPR family protein (“On the basis of the structure of Streptococcus pyogenes Cas9 (SpyCas9), a total of 11 amino acids were selected as AeF mutation sites…,” pg. 2, left col.);
(2) mutating the codon encoding the amino acid at the site selected in step (1) to codon TAG by a genetic engineering method, to obtain a mutated CRISPR protein gene (“The codons of these residues were mutated to the TAG codon,” pg. 2, left col.; “Mutagenesis PCR was conducted using KOD-One PCR Master Mix,” pg. 6, left col.);
(3) ligating the mutated CRISPR protein gene obtained by in step (2) to an expression plasmid by means of molecular cloning, to obtain an expression vector plasmid comprising a mutant sequence (“All plasmids were constructed using Gibson assembly… Mutagenesis PCR was conducted using KOD-One PCR Master Mix,” pg. 6, left col.); and
(4) co-transfecting the expression vector plasmid obtained in (3) and a tool plasmid for unnatural amino acid insertion into a host cell, culturing the host cell in a culture solution supplemented with the unnatural amino acid, and inducing the expression of the mutated protein in the host cell (“E. coli BL21 cells transformed with pET28a-Cas9-mutants and pUltra-MjPolyRS were cultivated in 2YT medium… AeF was added,” pg. 6, left col.; “the mutant proteins were expressed in Escherichia coli BL21 cells with a previously developed polyspecific Methanococcus jannacschii tyrosyl-tRNA synthetase (MjPolyRS)/tRNACUA pair,” pg. 2, left col.).
Regarding claim 9, Ling teaches the tool plasmid encodes tRNA and an aminoacyl-tRNA synthetase that recognizes TAG codon and inserts the unnatural amino acid, AeF (“Methanococcus jannacschii tyrosyl-tRNA synthetase (MjPolyRS)/tRNACUA pair,” pg. 2, left col.).
Regarding claims 10-11, and 20, Ling teaches a vector comprising a nucleic acid sequence encoding the CRISPR family protein site-specifically mutated with the unnatural amino acid (“All plasmids were constructed using Gibson assembly… Mutagenesis PCR was conducted using KOD-One PCR Master Mix,” pg. 6, left col.; “the mutant proteins were expressed in Escherichia coli BL21 cells…,” pg. 2, left col.; “E. coli BL21 cells transformed with pET28a-Cas9-mutants,” pg. 6, left col.). As stated above, the codon corresponding to the unnatural amino acid is TAG (“The codons of these residues were mutated to the TAG codon,” pg. 2, left col.).
Regarding claims 12-13, Ling teaches a site-specific nucleic acid conjugate of the protein, which is a donor DNA (“ssODN conjugate,” at least pgs. 2-3; Fig. 2A).
Regarding claim 14, Ling teaches a method for improving gene editing efficiency and/or reducing off-target effects comprising transfecting a host cell with a gRNA and the CRISPR family protein site-specifically mutated by the unnatural amino acid, and a site-specific nucleic acid conjugate of the CRISPR family protein (“As shown in Fig. 2C, cells cotransfected with the G1367-AeF RNP and DBCO-modified ssODN had a 1.6 times HDR increase as compared to other groups, indicating that covalent conjugation of ssODN donor DNA to Cas9 can enhance HDR efficiency,” pg. 3).
Regarding claim 16, while the claim further limits the related inactivated counterpart of claim 4, the CRISPR family protein may still be any of those encompassed by claim 4, e.g., Cas9. As stated above, Ling teaches the Cas9 protein is an SpCas9 (“SpyCas9”) mutated with the unnatural amino acid bearing an azido group (“On the basis of the structure of Streptococcus pyogenes Cas8 (SpyCas9), a total of 11 amino acids were selected as AeF mutation sites (Fig. 1A): K3, D39, H41… K1151, G1367… The codons of these residues were mutated to the TAG codon, and the mutant proteins were expressed…,” pg. 2, left col.; “K1151-AeF and G1367-AeF showed the highest expression yields and quality,” pg. 2, right col.; “we prepared and assessed two AeF-modified Cas9 D576-AeF-G1367-AeF in cell with double-adaptor conjugates,” pg. 4, right col.).
Claim Rejections - 35 USC § 102 – Ling as evidenced by GenBank
Claim 2 is rejected under 35 U.S.C. 102(a)(1) as being anticipated by Ling (Ling et al., 8 April 2020, Science Advances, 6: eaaz0051, pg. 1-8 and Supplemental Information; of record) as evidenced by GenBank (Chain D, CRISPR-associated endonuclease Cas9/Csn1, PDB: 4OO8_D, available 26 February 2014). The rejection that follows is new and necessitated by Applicant’s amendments to the claims.
As stated above, the effective filing date of claims 2, 4, and 7-20 is November 11, 2020.
Regarding claim 2, Ling teaches a method for site-specific conjugation of a nucleic acid to a CRISPR family protein (pgs. 2-3; Fig. 1-2). Ling teaches the method comprises:
(a) mutating a SpCas9 site-specifically with an unnatural amino acid (“AeF”) at one or more of positions K3, D39, H41, H116, D576, E945, K1151, and G1367 (“On the basis of the structure of Streptococcus pyogenes Cas8 (SpyCas9), a total of 11 amino acids were selected as AeF mutation sites (Fig. 1A): K3, D39, H41… K1151, G1367… The codons of these residues were mutated to the TAG codon, and the mutant proteins were expressed…,” pg. 2, left col.; “K1151-AeF and G1367-AeF showed the highest expression yields and quality,” pg. 2, right col.; “we prepared and assessed two AeF-modified Cas9 D576-AeF-G1367-AeF in cell with double-adaptor conjugates,” pg. 4, right col.); and
(b) conjugating the nucleic acid to the CRISPR family protein site-specifically mutated by the unnatural amino acid obtained in step (a) (pg. 2, right col. to pg. 3; Fig. 2).
Ling teaches the mutations were made relative to the SpCas9 sequence disclosed in “Protein Data Bank code: 4OO8” (Fig. 1, description). As shown in the attached alignment, the amino acid sequence corresponding to 4OO8 disclosed by GenBank comprises the same residues at the recited positions of SEQ ID NO: 1. Thus, as evidenced by GenBank, the mutations disclosed by Ling correspond to the residues at the recited positions in SEQ ID NO: 1.
Claim Rejections - 35 USC § 102 - Choudhary
The following is a quotation of the appropriate paragraphs of 35 U.S.C. 102 that form the basis for the rejections under this section made in this Office action:
A person shall be entitled to a patent unless –
(a)(1) the claimed invention was patented, described in a printed publication, or in public use, on sale, or otherwise available to the public before the effective filing date of the claimed invention.
(a)(2) the claimed invention was described in a patent issued under section 151, or in an application for patent published or deemed published under section 122(b), in which the patent or application, as the case may be, names another inventor and was effectively filed before the effective filing date of the claimed invention.
Claims 4, 7, 10-14, 16-17, and 20 are rejected under 35 U.S.C. 102(a)(1) and 102(a)(2) as being anticipated by Choudhary (Choudhary et al., WO 2019/135816 A9, published 11 July 2019; of record). The rejections that follow are maintained and modified as necessitated by Applicant’s amendments to the claims.
Regarding claims 4, 7 and 17, Choudhary teaches a CRISPR family protein site specifically mutated with an unnatural amino acid, wherein the CRISPR family protein is Cas9, and wherein the unnatural amino acid is pAcF, or bears a tetrazine group (“The CRISPR/Cas protein can be selected from… Cas9… In certain embodiments, the CRISPR/Cas protein comprises one or more unnatural amino acid p-Acetyl Phenylalanine (pAcF), or one or more unnatural amino acid comprising tetrazine,” [0010]-[0016]; [0843]; “wherein the CRISPR/Cas protein comprises one or more unnatural amino acid p-Acetyl Phenylalanine (pAcF), or one or more unnatural amino acid comprising tetrazine,” claim 81, pg. 319).
Regarding claims 10-11, and 20, Choudhary teaches a nucleic acid sequence encoding the CRISPR family protein, and a vector comprising the nucleic acid sequence (“the present invention provides a means for delivering the nucleic acid modifying protein… e.g. particle(s) delivering component(s) of the complex, vector(s) comprising the polynucleotide(s) discussed herein (e.g., encoding the nucleic acid modifying protein, providing the nucleotides encoding the nucleic acid modifying complex,” [0357]). Choudhary teaches the codon corresponding to the unnatural amino acid in the base sequence of the nucleic acid sequence encoding the CRISPR family protein is TAG (“amber (TAG)… the amber codon is the least used in E. coli (~7%) and rarely terminates essential genes. Amber suppression codons will be placed at the optimal sites identified above,” [0843]).
Regarding claims 12-13, Choudhary teaches a site-specific nucleic acid conjugate of a CRISPR family protein, wherein the nucleic acid conjugate is a donor DNA (“the one or more effector components comprise one or more single-stranded oligo donors (ssODNs),” [0012]; “the one or more effector components are linked to the CRISPR/Cas protein… via unnatural amino acids,” [0016]; “engineered Cas proteins can be mono-conjugated with ssODN… engineered CRISPR/Cas proteins can be conjugated with ssODN… using orthogonal conjugation chemistries,” [0092]). The limitation “wherein the CRISPR family protein is the CRISPR family protein-site specifically mutated by an unnatural amino acid of claim 4,” is interpreted as requiring that the nucleic acid conjugate be capable of conjugation to the protein of claim 4. Choudhary teaches that “engineered CRISPR/Cas proteins can be conjugated with ssODN… using orthogonal conjugation chemistries,” ([0092]), wherein unnatural amino acid mutagenesis, e.g., with pAcF or tetrazine-comprising amino acids, is such an orthogonal conjugation chemistry ([0843]). Thus, Choudhary also teaches that the nucleic acid conjugate is capable of conjugation to the protein of claim 4.
Regarding claim 14, Choudhary teaches a method for improving gene editing efficiency and/or reducing off-target effects comprising transfecting a host cell with a gRNA and the CRISPR family protein site-specifically mutated by the unnatural amino acid, and a site-specific nucleic acid conjugate of the CRISPR family protein (“Following a double-strand break in a knock-in experiment… ssODN[] is integrated at the break site. This integration can be facilitated if the ssODN is readily available at the break site… Increased efficiency of repair is highly desirable in disease models and therapies,” [0007]; “RNA molecules of the invention are delivered in liposome or lipofectin formulations and the like… Liposomal transfection reagents such as lipofectamine...,” [0640]-[0641]).
Regarding claim 16, while the claim further limits the related inactivated counterpart of claim 4, the CRISPR family protein may still be any of those encompassed by claim 4, e.g., Cas9. As stated above, Choudhary teaches the CRISPR family protein is Cas9 (“The CRISPR/Cas protein can be selected from… Cas9… In certain embodiments, the CRISPR/Cas protein comprises one or more unnatural amino acid p-Acetyl Phenylalanine (pAcF), or one or more unnatural amino acid comprising tetrazine,” [0013]).
Response to Remarks - 35 USC § 102
Applicant’s remarks regarding the § 102 rejections raised in the prior action have been considered.
Regarding the § 102 rejections over Ling, Applicant submits that Ling “is published by the same inventors of the present application on April 8, 2020… [and] is within the grace period for filing a patent application after a disclosure.” Applicant submits that because Ling is a “grace period” disclosure, Ling does not apply to the claims. While Examiner agrees that Ling was published one year or less before the effective filing date of the claimed invention, Examiner respectfully disagrees that Ling is “published by the same inventors”; the authors of Ling include the inventors of the instant application, and seven other authors. Ling, therefore, applies as prior art under 102(a)(1), and Applicant’s remarks are insufficient to disqualify the disclosure of Ling. Should Applicant wish to disqualify Ling on the basis that the grace-period disclosure is made by the inventors, “[t]he Office has provided a mechanism in 37 CFR 1.130 for filing an affidavit or declaration to establish that a disclosure made no earlier than one year before the effective filing date of the claimed invention is not prior art under 35 U.S.C. 102(a) due to an exception in 35 U.S.C. 102(b).” See MPEP 717.01.
Applicant’s remarks do not address the § 102 rejections raised over Choudhary.
Notice to Joint Inventors
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.
Claim Rejections - 35 USC § 103 – Choudhary as evidenced by PDB: 5F9R in view of Chatterjee
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 1-3, and 15 are rejected under 35 U.S.C. 103 as being unpatentable over Choudhary (Choudhary et al., WO 2019/135816 A9, published 11 July 2019; of record) as evidenced by PDB: 5F9R (“pdb_00005f9r,” deposited 27 January 2016, https://www.rcsb.org/structure/5F9R, accessed 11 March 2026), in view of Chatterjee (Chatterjee et al., 2013, Biochemistry, 52, pg. 1828-1837; of record). The rejections that follow are new and necessitated by Applicant’s amendments to the claims.
The teachings of Choudhary are described above, and applied hereinafter.
Regarding claim 1, as stated in the preceding paragraphs, Choudhary teaches a CRISPR family protein mutated site specifically with an unnatural amino acid, to which is conjugated a nucleic acid (“ssODN”). Choudhary also teaches a method for site-specific conjugation of a nucleic acid to a CRISPR family protein (“For unnatural amino acid mutagenesis, genetic code expansion can be utilized by adding an engineered pyrrolysyl tRNA (PylT)/tRNA synthetase pair to the translational machinery of the cells to enable the site-specific incorporation of p-Acetyl Phenylalanine (pAcF) into CRISPR/Cas9,” [0843]). Choudhary teaches the method comprises:
(a) mutating the CRISPR family protein site-specifically with an unnatural amino acid (“Amber suppression codons will be placed at the optimal sites identified above,” “the site-specific incorporation of p-Acetyl Phenylalanine (pAcF) into CRISPR/Cas9,” [0843]); and
(b) reaction of the unnatural amino acid pAcF with aminooxy group (“pAcF is proposed to be used as the unnatural amino acid that can react with aminooxy group,” [0843]).
Choudhary teaches the unnatural amino acid has orthogonal chemical reactivity (“This method relies on a unique codon-tRNA pair and corresponding aminoacyl tRNA synthetase (aaRS) for each unnatural amino acid that does not cross-react with any of the endogenous tRNAs, aaRSs, amino acids or codons in the host organism,” [00843]).
Choudhary teaches the CRISPR family protein is SpCas9 (“Most preferably, the Cas9 enzyme is from, or is derived from spCas9 (S. pyogenes Cas9),” [0097]-[0098]). Choudhary teaches that “Amber suppression codons will be placed at the optimal sites identified above” ([0843]), wherein the above “optimal sites identified above” include E945 of SpCas9 (“Guided by the Cas9 crystal structure (PDB: 5F9R)… E945C,” [0835]). Choudhary teaches the residues are relative to the SpCas9 sequence disclosed in “PDB: 5F9R” ([00835]). As shown in the attached alignment, the amino acid sequence disclosed by PDB_5F9R comprises the same residues at the recited positions of SEQ ID NO: 1. Thus, as evidenced by PDB_5F9R, Choudhary teaches that the CRISPR family protein is SpCas9 mutated with an unnatural amino acid at position E945 relative to SEQ ID NO: 1.
With respect to step (b) of the instantly claimed method, Choudhary does not literally set forth a step of attaching the aminooxy group to the nucleic acid conjugate (“ssODN”), which would thereby enable conjugation of the nucleic acid and CRISPR family protein comprising the unnatural amino acid.
However, Chatterjee teaches similar methods in which probes attached to an aminooxy group were conjugated to a protein site-specifically mutated with the unnatural amino acid, pAcF (pg. 1830).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have attached the nucleic acid conjugate of Choudhary to an aminooxy group, in order to conjugate the nucleic acid to the CRISPR family protein comprising pAcF taught by Choudhary. It would have amounted to applying a known step to a similar method, in order to yield a similar protein comprising a conjugated unnatural amino acid. The skilled artisan would have had a reasonable expectation of success in attaching the nucleic acid conjugate to an aminooxy group because such a step is alluded to by Choudhary, and because aminooxy groups were known in the art, and suitable for conjugation with pAcF-mutated proteins as evidenced by Chatterjee. The skilled artisan would have been motivated to attach the nucleic acid conjugate to an aminooxy group because based on Choudhary and Chatterjee, doing so would allow the skilled artisan to prepare the nucleic acid conjugated-CRISPR family protein. The skilled artisan would have been further motivated because Choudhary suggests that a nucleic acid conjugated-CRISPR family protein will improve gene editing efficiency, by localizing the donor DNA to the break site ([0007]). Thus, producing the protein of Choudhary would be likely to improve gene editing efficiency, which Choudhary teaches is “highly desirable in disease models and therapies” ([0007]).
Regarding claim 2, as stated above, Choudhary teaches the CRISPR family protein is Cas9 (“The CRISPR/Cas protein can be selected from… Cas9… In certain embodiments, the CRISPR/Cas protein comprises one or more unnatural amino acid p-Acetyl Phenylalanine (pAcF), or one or more unnatural amino acid comprising tetrazine,” [0013]; [0843]).
Regarding claim 3, Choudhary teaches the unnatural amino acid is pAcF or comprises a tetrazine group ([0843]).
Regarding claim 15, Choudhary teaches the CRISPR family protein is an inactivated counterpart dCas9 ([0098]).
Claim Rejections - 35 USC § 103 – Choudhary and Chatterjee as evidenced by Young
Claims 8-9, and 18-19 are rejected under 35 U.S.C. 103 as being unpatentable over Choudhary (Choudhary et al., WO 2019/135816 A9, published 11 July 2019; of record) and Chatterjee (Chatterjee et al., 2013, Biochemistry, 52, pg. 1828-1837; of record) as evidenced by Young (Young et al., 2010, JMB, 295, pg. 361-374; of record). The rejections that follow are maintained from the prior action.
Regarding claims 8-9, Choudhary teaches a method for preparing a CRISPR family protein site-specifically mutated with an unnatural amino acid ([0843]). Choudhary teaches the method comprises:
(1) selecting one or more specific amino acid sites in the amino acid sequence of the CRISPR family protein based on structural information (“Amber suppression codons will be placed at the optimal sites identified above,” [0843]; “Guided by the Cas9 crystal structure… sites were chosen that sampled multiple Cas9 domains… an eGFP disruption assay was used to determine the [nuclease] activity of the labelled mutants in cells,” [0835]);
(2) mutating the codon encoding the amino acid at the one or more selected sites to codon TAG by a genetic engineering method (“amber (TAG)… Amber suppression codons will be placed at the optimal sites identified above,” [0843]); and
(4) adding to a cell comprising a sequence encoding the CRISPR family protein, a tRNA and aminoacyl-tRNA synthetase, which can insert the unnatural amino acid pAcF into the CRISPR family protein during translation (“adding an engineered pyrrolysyl tRNA (PylT)/tRNA synthetase pair to the translational machinery of the cells to enable the site-specific incorporation of p-Acetyl Phenylalanine (pAcF) into CRISPR/Cas9,” [0843]).
Choudhary does not set forth step (3) of the instantly claimed method. With respect to step (4) of the instantly claimed method, Choudhary does not teach that the step comprises co-transfecting a cell with a vector comprising the sequence encoding the CRISPR family protein and a plasmid encoding the tRNA and aminoacyl-tRNA synthetase, and culturing the transfected cell in a culture solution supplemented with the unnatural amino acid.
However, Chatterjee teaches a method of preparing a protein site-specifically mutated with the unnatural amino acid taught by Choudhary, i.e., pAcF (pg. 1830, left col.; pg. 1831-1832; Fig. 2). With respect to step (3), Chatterjee teaches that the vector comprising the sequence encoding the TAG codon-mutated protein was previously reported in Young et al. (“The previously reported GFP (Tyr151-TAG)11 was used to evaluate the suppression efficiencies of the new suppressor plasmids,” pg. 1831). Young teaches that the sequence encoding the TAG codon-mutated protein was inserted into an expression vector by molecular cloning (“we inserted the gene for GFP with a Tyr151TAG mutation under the control of the T7 promoter in a pET101 vector,” pg. 362, right col.).
Regarding step (4), Chatterjee teaches that the pAcF-modified protein was expressed from a cell co-transfected with the aforementioned vector and a plasmid encoding a tRNA that recognizes the TAG codon and an aminoacyl-tRNA synthetase for inserting pAcF (“pUltra, harboring MjTyrRS (pAcF) and tRNACUATyr, were cotransformed with pET101-GFP (Tyr151TAG) into the BL21 (DE3) strain of E. coli,” pg. 1831, right col.; pg. 1830, left col.; Fig. 2). Chatterjee teaches that the transfected cell was cultured in a solution supplemented with the unnatural amino acid (“The relevant UAAs were also added at this point to a final concentration of 1 mM. Protein expression was continued,” pg. 1830, left col.).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have adapted the method of Choudhary with steps (3) and (4) taught by Chatterjee, in order to arrive at the method of instant claims 8-9. It would have amounted to applying known steps to a similar method, in order to yield a similar protein comprising a conjugated unnatural amino acid. The skilled artisan would have had a reasonable expectation of success in using Chatterjee’s steps because Choudhary and Chatterjee both describe a method in which a host cell is supplemented with a specific tRNA and aminoacyl-tRNA synthetase for site-specifically inserting the unnatural amino acid pAcF into a protein at a TAG codon, and because Chatterjee provides a detailed method, utilizing available plasmids (e.g., pUltra), which achieves the outcome desired by Choudhary on a protein of interest. The skilled artisan would have been motivated to use Chatterjee’s steps because Choudhary’s steps for producing the conjugated CRISPR family protein are generic, and Chatterjee’s steps would, alternatively, provide the skilled artisan with steps and materials ready for use in producing Choudhary’s CRISPR family protein. The skilled artisan would have been further motivated because Choudhary suggests that a nucleic acid conjugated-CRISPR family protein will improve gene editing efficiency, by localizing the donor DNA to the break site ([0007]). Thus, producing the protein of Choudhary would be likely to improve gene editing efficiency, which Choudhary teaches is “highly desirable in disease models and therapies” ([0007]).
Regarding claim 18, Choudhary teaches the CRISPR family protein is SpCas9 (“Most preferably, the Cas9 enzyme is from, or is derived from spCas9 (S. pyogenes Cas9),” [0097]-[0098]).
Regarding claim 19, as stated above, Choudhary teaches the unnatural amino acid is pAcF ([0843]).
Claim Rejections - 35 USC § 103 – Choudhary in view of Chatterjee, Reddington, and PDB_5B43
Claims 1-3 are rejected under 35 U.S.C. 103 as being unpatentable over Choudhary (Choudhary et al., WO 2019/135816 A9, published 11 July 2019; of record) in view of Chatterjee (Chatterjee et al., 2013, Biochemistry, 52, pg. 1828-1837; of record), Reddington (Reddington et al., 2012, Chem. Commun., 48, pg. 8419-8421 and Supplemental Information; of record) and PDB_5B43 (5B43, deposited 30 March 2016, accessed 24 July 2025; of record). The rejections that follow are new and necessitated by Applicant’s amendments to the claims.
Cancelled claim 6 was previously rejected over the combination of Choudhary, Reddington, and PDB_5B43. Applicant’s amendments incorporate features of cancelled claim 6 into instant claim 1. Hereinafter, Reddington and PDB_5B43 are applied to instant claim 1 and dependents solely in the interest of compact prosecution, given that the AsCas12a limitations are recited in the alternative to the SpCas9 limitations, which have already been addressed above.
Regarding claim 1, as stated in the preceding paragraphs, Choudhary teaches a CRISPR family protein mutated site specifically with an unnatural amino acid, to which is conjugated a nucleic acid (“ssODN”). Choudhary also teaches a method for site-specific conjugation of a nucleic acid to a CRISPR family protein (“For unnatural amino acid mutagenesis, genetic code expansion can be utilized by adding an engineered pyrrolysyl tRNA (PylT)/tRNA synthetase pair to the translational machinery of the cells to enable the site-specific incorporation of p-Acetyl Phenylalanine (pAcF) into CRISPR/Cas9,” [0843]). Choudhary teaches the method comprises:
(a) mutating the CRISPR family protein site-specifically with an unnatural amino acid (“Amber suppression codons will be placed at the optimal sites identified above,” “the site-specific incorporation of p-Acetyl Phenylalanine (pAcF) into CRISPR/Cas9,” [0843]); and
(b) reaction of the unnatural amino acid pAcF with aminooxy group (“pAcF is proposed to be used as the unnatural amino acid that can react with aminooxy group,” [0843]).
Choudhary also teaches the unnatural amino acid has orthogonal chemical reactivity (“This method relies on a unique codon-tRNA pair and corresponding aminoacyl tRNA synthetase (aaRS) for each unnatural amino acid that does not cross-react with any of the endogenous tRNAs, aaRSs, amino acids or codons in the host organism,” [00843]).
With respect to step (b) of the instantly claimed method, Choudhary does not literally set forth a step of attaching the aminooxy group to the nucleic acid conjugate (“ssODN”), which would thereby enable conjugation of the nucleic acid and CRISPR family protein comprising the unnatural amino acid.
However, Chatterjee teaches similar methods in which probes attached to an aminooxy group were conjugated to a protein site-specifically mutated with the unnatural amino acid, pAcF (pg. 1830).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have attached the nucleic acid conjugate of Choudhary to an aminooxy group, in order to conjugate the nucleic acid to the CRISPR family protein comprising pAcF taught by Choudhary. It would have amounted to applying a known step to a similar method, in order to yield a similar protein comprising a conjugated unnatural amino acid. The skilled artisan would have had a reasonable expectation of success in attaching the nucleic acid conjugate to an aminooxy group because such a step is alluded to by Choudhary, and because aminooxy groups were known in the art, and suitable for conjugation with pAcF-mutated proteins as evidenced by Chatterjee. The skilled artisan would have been motivated to attach the nucleic acid conjugate to an aminooxy group because based on Choudhary and Chatterjee, doing so would allow the skilled artisan to prepare the nucleic acid conjugated-CRISPR family protein. The skilled artisan would have been further motivated because Choudhary suggests that a nucleic acid conjugated-CRISPR family protein will improve gene editing efficiency, by localizing the donor DNA to the break site ([0007]). Thus, producing the protein of Choudhary would be likely to improve gene editing efficiency, which Choudhary teaches is “highly desirable in disease models and therapies” ([0007]).
Neither Choudhary nor Chatterjee teach that the CRISPR family protein is an AsCas12a comprising an unnatural amino acid at one or more of the recited positions.
However, Choudhary teaches that the CRISPR family protein may be Cas12a (“Cpf1”, [0013]). Choudhary teaches the overall organization of and function of specific residues in AsCas12a ([0167]-[0187]). Choudhary teaches that selection of optimal sites for mutation is chosen on the basis of the structure of the CRISPR family protein (“Amber suppression codons will be placed at the optimal sites identified above,” [0843]; “Guided by the Cas9 crystal structure… sites were chosen that sampled multiple Cas9 domains… an eGFP disruption assay was used to determine the [nuclease] activity of the labelled mutants in cells,” [0835]).
Reddington teaches a protein site-specifically mutated by an unnatural amino acid, pAzF, which is prepared through a method similar to that of Choudhary (“The non-canonical amino acid… AzPhe… was site-specifically introduced into sfGFP using a reprogrammed TAG amber stop codon,” pg. 8419, right col.; Fig. 1). Reddington selected four surface sites for incorporation of the unnatural amino acid, and compared the site’s effects on the structure and function of the resulting proteins (pg. 8419, right col. to pg. 8420, left col.). Reddington concludes that “incorporation of [AzPhe] does not significantly perturb structure or function at the targeted sites” (pg. 8420, left col.). Reddington also compared the site’s effects on conjugation of the unnatural amino acid to a DBCO-functionalized probe (pg. 8420, left col.). Reddington concludes that the site’s location influenced the efficiency of conjugation, likely due to “local surface microenvironments of a protein” (pg. 8420). Reddington states that “it is clear that the choice of residue is important in terms of efficiency of modification and the influence on function,” and suggests optimizing the placement of unnatural amino acids through scanning mutagenesis will be advantageous (pg. 8420, right col. to pg. 8421).
Neither Choudhary nor Reddington provide the crystal structure of AsCas12a in complex with crRNA and target DNA, such that the skilled artisan could predict which residues would be amenable to incorporation with an unnatural amino acid. However, PDB_5B43 teaches the crystal structure of AsCas12a in complex with crRNA and target DNA. The structure disclosed by PDB_5B43 would allow the skilled artisan to reasonably predict which residues would be amendable to incorporation of an unnatural amino acid, e.g., surface exposed residues and/or residues that are not involved in interactions with crRNA and/or target DNA. As shown in the attached alignment, the amino acid sequence disclosed by PDB_5B43 comprises the same residues at the recited positions of SEQ ID NO: 3.
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have optimized the site for incorporation of the unnatural amino acid in Choudhary’s CRISPR family protein, AsCas12a. It would have amounted to choosing from a finite number of identified, predictable potential solutions, with a reasonable expectation of success.
It was known prior to the effective filing date that the placement of an unnatural amino acid must be optimized. Indeed, Choudhary and Reddington each employ such optimization processes to identify residues which when modified, do not affect the protein function, and/or allow for effective conjugation to a functionalized group. Reddington suggests using scanning mutagenesis to define the optimal sites for modification with an unnatural amino acid.
There are a finite number of amino acids in AsCas12a which may be mutagenized with an unnatural amino acid. Among these amino acids are those at the instantly recited positions. Based on the crystal structure disclosed by PDB, the skilled artisan could have reasonably predicted which residues were amendable to incorporation of an unnatural amino acid, e.g., surface exposed residues and/or residues uninvolved in interactions with crRNA or target DNA. The skilled artisan could have pursued these finite solutions with a reasonable expectation of success because the structure and residues of AsCas12a were known as evidenced by Choudhary and PDB_5B43, and methods to incorporate an unnatural amino acid at a desired location in a protein were known as evidenced by the prior art, including at least Choudhary and Reddington cited herein. The skilled artisan would have been motivated to select and mutagenize amino acids in AsCas12a given the teachings of Choudhary and Reddington regarding the need to optimize the site at which the unnatural amino acid is incorporated in order to preserve the function of the bioconjugated protein, and the conjugate (i.e., in the instant case, the ssODN).
Regarding claim 2, the CRISPR family protein rendered obvious above is a Cas12a protein.
Regarding claim 3, Choudhary teaches the unnatural amino acid is pAcF or comprises a tetrazine group ([0843]).
Response to Remarks - 35 USC § 103
Applicant’s remarks regarding the § 103 rejections raised in the prior action have been considered.
Regarding the § 103 rejections over Choudhary in view of Chatterjee, or Choudhary and Chatterjee as evidenced by Young, Applicant submits that the amendments to claim 1 “to incorporate the features of claims 5-6” overcome the rejections because “the Examiner does not hold claim 6 [as] being anticipated” by the combination of references. Examiner agrees that claim 6 was not rejected over Choudhary and Chatterjee, or the references as evidenced by Young. However, the features of now cancelled claim 6 are not required of amended claim 1, as they are stated in the alternative to features previously recited in cancelled claim 5. The features of cancelled claim 5, which are now incorporated into amended claim 1, were identified in the prior action has being taught by Choudhary as evidenced by PDB_5F9R (see paragraphs 50-51). Furthermore, the features of claim 1 are not required of claims 8-9, or 18-19, because these claims refer to claim 4, not claim 1. Applicant’s remarks are not found convincing to overcome the rejections over Choudhary in view of Chatterjee, or Choudhary and Chatterjee as evidenced by Young.
Regarding the § 103 rejection of now cancelled claim 6 over Choudhary in view of Reddington and PDB_5B43, Examiner notes that neither an AsCas12a, nor a substitution at one of the recited positions therein is required of the amended claims, because these limitations are recited in the alternative with the SpCas9 limitations. The SpCas9 limitations are addressed in the rejection over Choudhary and Chatterjee. The references previously applied to cancelled claim 6 are applied herein to claim 1 and dependents herein, solely in the interest of compact prosecution. Applicant’s remarks regarding the rejection of cancelled claim 6 are addressed as they apply to the rejection of claim 1 and dependents above.
Applicant submits that “it is commonly accepted in the art that the appropriate mutation site cannot be easily predicted simply relying on the data from crystal structure which cannot always provide information reflecting the true nature of a protein.” This argument is not accompanied by evidence to support this position, and Applicant’s position is rebutted by evidence in the prior art cited in the rejection. For example, Choudhary uses the crystal structure of a Cas protein to identify the optimal sites for incorporation of an unnatural amino acid. Reddington also uses the crystal structure of a protein to identify optimal sites for incorporation of an unnatural amino acid. The crystal structure of AsCas12a was known, and means to incorporate an unnatural amino acid at identified sites were known. There is substantial evidence that the skilled artisan would have had a reasonable expectation of success in arriving at the claimed residues, which are among the finite number of identified, predictable potential solutions in the known AsCas12a protein sequence.
Applicant also argues that the prior art “teaches away from making substitution M806” because Yamano (Yamano et al., 2016, Cell, 165(4):949-62 and Supplemental Information) identifies this residue as a “key residue.” Examiner has reviewed Applicant’s evidence that Yamano “teaches away” from a substitution at M806 in AsCas12a. Fig. S4 referenced by Applicant corresponds to a “Multiple Sequence Alignment of Cpf1 Proteins,” in which triangles correspond to “key residues” disclosed in Fig. 3. Fig. 3 identifies that M806 corresponds to a residue in a region which interacts with the “5’-handle” of a crRNA. Yamano does not explicitly teach away from modifying this residue. Examiner also notes that the claims under consideration are directed to a method, not a product, and no activity is required of the CRISPR family protein-nucleic acid conjugate obtained from the method. Even so, Reddington provides evidence that incorporation of an unnatural amino acid does not significantly perturb structure or function at targeted sites (pg. 8420, left col.). The rejection sufficiently describes why a skilled artisan would have had a reasonable expectation of success in arriving at one or more of the claimed residues, and Applicant’s remarks are not sufficient to overcome this rejection.
Conclusion
No claims are allowed.
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
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/JENNA L PERSONS/Examiner, Art Unit 1637
/Soren Harward/Primary Examiner, TC 1600