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 of claims 1-3, 8, 10-12, 16, 17, 25, 30, 33, 35-37, 40, 42, and 44 (Group I) in the reply filed on 04/01/2026 is acknowledged. Because applicant did not distinctly and specifically point out the supposed errors in the restriction requirement, the election has been treated as an election without traverse (MPEP § 818.01(a)).
Upon further consideration, the restriction requirement between Groups I and II is hereby withdrawn.
Claim 54 is withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim. Election was made without traverse in the reply filed on 04/01/2026.
Accordingly, claims 1-3, 8, 10-12, 16, 17, 25, 30, 33, 35-37, 40, 42, 44, and 52 are pending and under consideration.
Furthermore, because a claimed invention previously withdrawn from consideration under 37 CFR 1.142 has been rejoined, the restriction requirement between Groups I and II as set forth in the Office action mailed on 02/13/2026 is hereby withdrawn. In view of the withdrawal of the restriction requirement as to the rejoined inventions, applicant(s) are advised that if any claim presented in a divisional application is anticipated by, or includes all the limitations of, a claim that is allowable in the present application, such claim may be subject to provisional statutory and/or nonstatutory double patenting rejections over the claims of the instant application.
Once the restriction requirement is withdrawn, the provisions of 35 U.S.C. 121 are no longer applicable. See In re Ziegler, 443 F.2d 1211, 1215, 170 USPQ 129, 131-32 (CCPA 1971). See also MPEP § 804.01.
Regarding the species election in the response filed 04/01/2026, the election of species is hereby withdrawn in view of a search of the prior art.
Priority
Applicant’s claim for the benefit of a prior-filed application under 35 U.S.C. 119(e) or under 35 U.S.C. 120, 121, 365(c), or 386(c) is acknowledged. The earliest effective filing date to which the instant application is entitled is 04/30/2021.
Information Disclosure Statement
Receipt of information disclosure statements on 10/19/2023, 03/27/2025, and 04/24/2026 is acknowledged. The signed and initialed PTO-1449’s have been mailed with this action.
Drawings
The drawings are objected to because:
With regard to Figure 4, while the legend lists six imaging conditions, only two images are actually shown. Neither the drawings themselves nor the associated brief description of the drawings clarify why only two of six images are actually shown. It would be remedial to amend the instant drawings or the associated brief description thereof such that all six images are actually shown or otherwise addressed.
Corrected drawing sheets in compliance with 37 CFR 1.121(d) are required in reply to the Office action to avoid abandonment of the application. Any amended replacement drawing sheet should include all of the figures appearing on the immediate prior version of the sheet, even if only one figure is being amended. The figure or figure number of an amended drawing should not be labeled as “amended.” If a drawing figure is to be canceled, the appropriate figure must be removed from the replacement sheet, and where necessary, the remaining figures must be renumbered and appropriate changes made to the brief description of the several views of the drawings for consistency. Additional replacement sheets may be necessary to show the renumbering of the remaining figures. Each drawing sheet submitted after the filing date of an application must be labeled in the top margin as either “Replacement Sheet” or “New Sheet” pursuant to 37 CFR 1.121(d). If the changes are not accepted by the examiner, the applicant will be notified and informed of any required corrective action in the next Office action. The objection to the drawings will not be held in abeyance.
Claim Objections
Claims 1, 3, 8, 12, 25, 33, 35-37, 40, and 42 are objected to because of the following informalities:
Claim 1 is objected to for reciting “a.”, “i.”, “ii.”, and “b.” to separate system components by including a period following the initial listing of each component. According to MPEP 608.01(m), “Each claim begins with a capital letter and ends with a period. Periods may not be used elsewhere in the claims except for abbreviations. See Fressola v. Manbeck, 36 USPQ2d 1211 (D.D.C. 1995)”. It would be remedial to replace “a.”, “i.”, “ii.”, and “b.” with parentheses, for example “(a)”, “(i)”, “(ii)”, and “(b).”
Claim 1 is further objected to for reciting “an oligonucleotide, optionally the oligonucleotide is double-stranded and comprises a double-stranded region of at least 17 base-pairs,” which does not comport with standard grammatical and/or linguistic conventions. For purposes of comporting with said conventions, it would be remedial to amend the instant claim to recite, for example, “an oligonucleotide, optionally wherein the oligonucleotide is double-stranded and comprises a double-stranded region of at least 17 base-pairs” (bolded and underlined emphasis added). This is merely an example set forth by the Examiner and is not intended to be limiting.
Similarly, claim 3 is objected to for reciting “…a PIWI domain of an Ago, optionally the PIWI domain lacks nuclease activity,” which does not comport with standard grammatical and/or linguistic conventions. For purposes of comporting with said conventions, it would be remedial to amend the instant claim to recites, for example, “…a PIWI domain of an Ago, optionally wherein the PIWI domain lacks nuclease activity” (bolded and underlined emphasis added). This is merely an example set forth by the Examiner and is not intended to be limiting.
With regard to claim 8, the recitation of “…the second domain comprises an amino acid sequence having a mutation at one or more positions selected from the group consisting of D597 and D699 of human Ago amino acid sequence” (bolded emphasis added), does not comport with standard grammatical and/or linguistic conventions, as there is no article such as “a” preceding the instantly claimed “human Ago amino acid sequence.” It would be remedial to amend the instant claim language to include an article preceding the instantly claimed “human Ago amino acid sequence,” thereby comporting with standard grammatical and/or linguistic conventions.
Claim 8 is further objected to for reciting “…or a corresponding position in a homologous or orthologous Ago protein, optionally the second domain comprises an amino acid sequence having a mutation at one or more positions…,” which does not comport with standard grammatical and/or linguistic conventions. For purposes of comporting with said conventions, it would be remedial to amend the instant claim to recite, for example, “…or a corresponding position in a homologous or orthologous Ago protein, optionally wherein the second domain comprises an amino acid sequence having a mutation at one or more positions…” (bolded and underlined emphasis added). This is merely an example set forth by the Examiner and is not intended to be limiting.
Claim 12 is objected to for reciting “…a cytidine deaminase, optionally the cytidine deaminase is an apolipoprotein B mRNA editing enzyme catalytic polypeptide-like (APOBEC),” which does not comport with standard grammatical and/or linguistic conventions. For purposes of comporting with said conventions, it would be remedial to amend the instant claim to recite, for example, “…a cytidine deaminase, optionally wherein the cytidine deaminase is an apolipoprotein B mRNA editing enzyme catalytic polypeptide- like (APOBEC)” (bolded and underlined emphasis added). This is merely an example set forth by the Examiner and is not intended to be limiting.
Claim 25 is objected to for reciting a number of clauses that do not comport with standard grammatical and/or linguistic conventions, as set forth below. The recitation of “…the polypeptide further comprises a nuclear export signal (NES) sequence, optionally the NES sequence is located between the first and the second domain; the polypeptide further comprises a FLAG octapeptide; or the polypeptide lacks nuclease activity” lacks several requisite “wherein” recitations. For purposes of comporting with standard grammatical and/or linguistic conventions, it would be remedial to amend the instant claim to recite, for example, “…wherein the polypeptide further comprises a nuclear export signal (NES) sequence, optionally wherein the NES sequence is located between the first and the second domain; wherein the polypeptide further comprises a FLAG octapeptide; or wherein the polypeptide lacks nuclease activity” (bolded and underlined emphasis added). This is merely an example set forth by the Examiner and is not intended to be limiting.
Claim 33 is objected to for reciting “…the oligonucleotide comprises a double-stranded region of at least 19 base-pairs, optionally the oligonucleotide comprises a double-stranded region of at least 25 base-pairs,” which does not comport with standard grammatical and/or linguistic conventions. For purposes of comporting with said conventions, it would be remedial to amend the instant claim to recite, for example, “…the oligonucleotide comprises a double-stranded region of at least 19 base-pairs, optionally wherein the oligonucleotide comprises a double-stranded region of at least 25 base-pairs” (bolded and underlined emphasis added). This is merely an example set forth by the Examiner and is not intended to be limiting.
Claim 35 is objected to for reciting “…wherein said strand comprises a mismatch with the target RNA at position 10, counting from 5’-end of said strand,” which does not comport with standard grammatical and/or linguistic conventions, as there is no article such as “the” preceding the recited “5’-end of said strand.” It would be remedial to amend the instant claim language to include an article preceding the instantly claimed “5’-end of said strand,” thereby comporting with standard grammatical and/or linguistic conventions.
Similarly, claim 36 is objected to for reciting “…said strand comprises a C at position…28 counting from 5’-end of said strand, and the strand comprises an A:C mismatch with the target RNA at position…28, counting from 5’-end of said strand; or (ii) said strand comprises a C at position…10, counting from 3’-end of said strand, and the strand comprises an A:C mismatch with the target RNA at position…10, counting from 3’-end of said strand,” which does not comport with standard grammatical and/or linguistic conventions, as there is no article such as “the” preceding the recited “5’-end of said strand” or “3’-end of said strand.” It would be remedial to amend the instant claim language to include an article preceding the instantly claimed “5’-end of said strand” and “3’-end of said strand,” thereby comporting with standard grammatical and/or linguistic conventions.
Similarly, claim 37 is objected to for reciting “…said strand comprises a C at position 25, counting from 5’-end of said strand, and the strand comprises an A:C mismatch with the target RNA at position 25, counting from 5’-end of said strand; or (ii) said strand comprises a C at position 7, counting from 3’-end of said strand, and the strand comprises an A:C mismatch with the target RNA at position 7, counting from 3’-end of said strand,” which does not comport with standard grammatical and/or linguistic conventions, as there is no article such as “the” preceding the recited “5’-end of said strand” or “3’-end of said strand.” It would be remedial to amend the instant claim language to include an article preceding the instantly claimed “5’-end of said strand” and “3’-end of said strand,” thereby comporting with standard grammatical and/or linguistic conventions.
Claim 40 is objected to for reciting “…said loop structure comprises a single-stranded C nucleotide, optionally said loop structure is 5 to 20 nucleotides in length,” which does not comport with standard grammatical and/or linguistic conventions. For purposes of comporting with said conventions, it would be remedial to amend the instant claim to recite, for example, “…said loop structure comprises a single-stranded C nucleotide, optionally wherein said loop structure is 5 to 20 nucleotides in length” (bolded and underlined emphasis added). This is merely an example set forth by the Examiner and is not intended to be limiting.
Claim 42 is objected to for several reasons. The recitation of “…said single-stranded C nucleotide is at position 6, 7, 8, 9, 10, 11, 12, counting from 5’-end of the loop structure, or said loop structure is in form of hairpin…” does not comport with standard grammatical and/or linguistic conventions for several reasons. First, there is no conjunction such as “or” separating the recited possible positions of the single-stranded C nucleotide of the instant claim. Furthermore, there are numerous articles missing preceding recited components, including “5’-end of the loop structure” and “hairpin.” For purposes of comporting with said conventions, it would be remedial to amend the instant claim to recite, for example, “…said single-stranded C nucleotide is at position 6, 7, 8, 9, 10, 11, or 12, counting from the 5’-end of the loop structure, or said loop structure is in the form of a hairpin…” (bolded and underlined emphasis added). This is merely an example set forth by the Examiner and is not intended to be limiting.
Appropriate correction is required.
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 52 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).
With regard to claim 52, which recites “a cell comprising a polypeptide or a nucleic acid encoding a polypeptide of claim 1,” the broadest reasonable interpretation of the term “cell” embraces a human having the cell. The instant specification envisions both single cells and populations of cells, wherein said population of cells may be human cells (paragraphs [00270] and [00271]). Therefore, under broadest reasonable interpretation of the claim language and in view of the disclosure of the instant specification, the term “cell” in claim 52 embraces a human having the cell. It would be remedial to amend the instant claim to recite “an isolated cell” to avoid the claim embracing a human organism.
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 35-37, 40, 42, and 44 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 term “substantially” recited in claims 35-37, 40, and 44 (and inherited by instant claim 42) is a relative term which renders the claim indefinite. The term “substantially” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. It would be remedial to clearly delineate the metes and bounds of protection sought by the instant claim set by setting forth a clear threshold for the level of complementarity claimed.
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.
Claim 17 is 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.
Claim 17 depends from claim 16, which recites four possible configurations of the first domain of the system of claim 1. While claim 16 recites positions that may be mutated (i.e. T375), claim 17 generally specifies the exact mutations (i.e. T375Q). However, possible configuration (ii) of claim 17 is identical to that of configuration (ii) of claim 16 in that both recite positions that may be mutated (“T339, R348, A353, V351, V355, T375, K376, E396, S397, E 438, F442, H443, L444, Y445, T448, C451, R455, S486, Q488, R510, I520, V525, P539, G593, K594, and E1008”) rather than the exact mutations claimed. Given that claim 16 may be interpreted such that only configuration (ii) is selected and that claim 17 may also be interpreted such that only configuration (ii) is selected and further that configuration (ii) is identical in both claims 16 and 17, it cannot be considered that claim 17 further limits instant claim 16. It would be remedial to amend the instant claim language such that claim 16 further limits all possible configurations recited at instant claim 16.
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
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 1-3, 10-12, 25, and 52 are rejected under 35 U.S.C. 102(a)(1) and 35 U.S.C. 102(a)(2) as being anticipated by US 2017/0175104 A1 (hereinafter Doudna; as cited in Applicant IDS; of record).
With regard to claim 1, which recites “a system for modifying a target RNA, the system comprising:
a polypeptide or a nucleic acid encoding a polypeptide, wherein the polypeptide comprises:
a first domain comprising a catalytic domain of an RNA modifying enzyme, wherein the RNA modifying enzyme is not a nuclease;
a second domain comprising a MID domain of an Argonaute (Ago) protein; and
an oligonucleotide, optionally [wherein] the oligonucleotide is double-stranded and comprises a double-stranded region of at least 17 base-pairs,”
as previously set forth, Doudna discloses compositions and methods for modifying a single stranded target nucleic acid such as mRNA or other ssRNA (paragraphs [0006], [0010], and [0019]). Specifically, Doudna discloses that the compositions taught therein facilitate the precise and controlled targeting of Ago binding to single stranded target nucleic acids such as ssRNA, mRNA, rRNA, tRNA, microRNA, etc. (paragraph [0005]). The compositions of Doudna are disclosed to comprise an Ago polypeptide (comprising a MID domain (paragraph [0104])) fused to an amino acid sequence that provides for a modification of the target nucleic acid other than cleavage (i.e. lacks nuclease activity) (paragraph [0010]). Furthermore, Doudna discloses that the compositions taught therein are guided to a targeted locus by introducing a guide RNA (i.e. an oligonucleotide) (paragraph [0008]). Thus, it is considered that Doudna discloses each and every limitation of instant claim 1.
With regard to claim 2, which recites “the second domain [of the system of claim 1] further comprises a PAZ domain of an Ago,” the compositions of Doudna are disclosed to comprise an Ago polypeptide, wherein said Ago polypeptide comprises a MID domain, a PAZ domain, and a PIWI domain (paragraphs [0010] and [0104]). Thus, it is considered that Doudna discloses each and every additional limitation of instant claim 2.
With regard to claim 3, which recites “the second domain [of the system of claim 1] further comprises a PIWI domain of an Ago, optionally [wherein] the PIWI domain lacks nuclease activity,” as set forth above, the compositions of Doudna are disclosed to comprise an Ago polypeptide, wherein said Ago polypeptide comprises a MID domain, a PAZ domain, and a PIWI domain (paragraphs [0010] and [0104]). Thus, it is considered that Doudna discloses each and every additional limitation of instant claim 3.
With regard to claim 10, which recites “the RNA modifying enzyme [of the system of claim 1] is an RNA deaminase, an RNA methylase, or an RNA demethylase,” the fusion partner of the system of Doudna is further disclosed to be an RNA deaminase (paragraphs [0010] and Claim
[0121]), as instantly claimed. Thus, it is considered that Doudna discloses each and every additional limitation of instant claim 10.
With regard to claim 11, which recites “the catalytic domain of the RNA modifying enzyme [of the system of claim 10] is a deaminase domain of an RNA deaminase,” as set forth above, the fusion partner of the system of Doudna is disclosed to be an RNA deaminase (paragraphs [0010] and [0121]), which must comprise a deaminase domain by definition. Thus, it is considered that Doudna discloses each and every additional limitation of instant claim 11.
With regard to claim 12, which recites “the RNA deaminase [of the system of claim 11] is Adenosine Deaminase Acting on RNA (ADAR) or a cytidine deaminase, optionally [wherein] the cytidine deaminase is an apolipoprotein B mRNA editing enzyme catalytic polypeptide-like (APOBEC),” as set forth above, the fusion partner of the system of Doudna is disclosed to be an RNA deaminase (paragraphs [0010] and [0121]). Doudna further discloses that the RNA deaminase may be an ADAR enzyme or a C to U editing enzyme (i.e. a cytidine deaminase), as instantly claimed (paragraph [0121]). Thus, it is considered that Doudna discloses each and every additional limitation of instant claim 12.
With regard to claim 25, which recites “the polypeptide [of the system of claim 1] further comprises a linker between the first domain and the second domain; the polypeptide further comprises a nuclear export signal (NES) sequence…; the polypeptide further comprises a FLAG octapeptide; or the polypeptide lacks nuclease activity,” Doudna further discloses that the fusion protein of the system taught therein may comprise a FLAG tag (paragraph [0112]), which is considered to read on the instantly claimed “FLAG octapeptide” per paragraphs [00109] and [00110] of the instant specification. Thus, it is considered that Doudna discloses each and every additional limitation of instant claim 25.
With regard to claim 52, which recites “a cell comprising a polypeptide or a nucleic acid encoding a polypeptide of claim 1,” as set forth above, Doudna anticipates the system of instant claim 1. Doudna further discloses genetically modified host cells comprising said system (paragraphs [0227] and [0228]). Thus, it is considered that Doudna discloses each and every limitation of instant claim 52.
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.
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 8 is rejected under 35 U.S.C. 103 as being unpatentable over US 2017/0175104 A1 (hereinafter Doudna; as cited in Applicant IDS; of record) as applied to claim 1 above, and further in view of Diedrichs and Haber, 2007 (hereinafter Diedrichs).
The disclosure of Doudna is described above and applied as before (see section Claim Rejections - 35 USC § 102). However, this disclosure does not teach the Ago2 mutations of instant claim 8.
With regard to claim 8, which recites “the second domain [of the system of claim 1] comprises an amino acid sequence having a mutation at one or more positions selected from the group consisting of D597 and D699 of human Ago2 amino acid sequence, or a corresponding position in a homologous or orthologous Ago protein, optionally [wherein] the second domain comprises an amino acid sequence having a mutation at one or more positions selected from the group consisting of D597A and D699A of human Ago2 amino acid sequence, or a corresponding position in a homologous or orthologous Ago protein,” as set forth above, Doudna discloses compositions and methods for modifying a single stranded target nucleic acid such as mRNA or other ssRNA (paragraphs [0006], [0010], and [0019]). Specifically, Doudna discloses that the compositions taught therein facilitate the precise and controlled targeting of Ago binding to single stranded target nucleic acids such as ssRNA, mRNA, rRNA, tRNA, microRNA, etc. (paragraph [0005]). Doudna further discloses that the Ago of the systems taught therein comprises an amino acid sequence having one or more mutations that exhibit reduced nuclease activity (paragraph [0011]). However, Doudna discloses that said mutation may be D516A, not the instantly claimed mutations.
This deficiency is cured by Diedrichs. Diedrichs discloses that residue D597 is essential for the catalytic endonuclease activity of the Ago2 PIWI domain (page 1101, column 1, paragraph 1). Diedrichs further discloses that the D597A mutation abrogates Ago2 RNase (i.e. nuclease) activity (Figure 5). Thus, it is considered that Diedrichs discloses each and every additional limitation of instant claim 8.
Given that Doudna discloses systems for facilitating the precise and controlled targeting of Ago binding to single stranded target nucleic acids such as ssRNA, wherein said Ago comprises an amino acid sequence having one or more mutations that exhibit reduced nuclease activity, and that Diedrichs discloses that the D597A mutation abrogates Ago2 RNase activity, it would have been obvious to someone of ordinary skill in the art before the effective filing date of the claimed invention to mutate the Ago of Doudna such that is comprises the D597A mutation disclosed in Diedrichs to predictably abrogate Ago nuclease activity, thereby facilitating its precise and controlled binding to single-stranded target nucleic acids such as ssRNA. One would have been motivated to make such a modification in order to receive the expected benefit of generating a system capable of precisely controlling Ago binding to single-stranded target nucleic acids such as ssRNA without cleaving said target nucleic acids.
Claims 16 and 17 are rejected under 35 U.S.C. 103 as being unpatentable over US 2017/0175104 A1 (hereinafter Doudna; as cited in Applicant IDS; of record), as applied to claim 1 above, and further in view of US 2021/0093667 A1 (hereinafter Zhang).
The disclosure of Doudna is described above and applied as before (see section Claim Rejections - 35 USC § 102). However, this disclosure does not teach the hADAR2 mutations of instant claims 16 and 17.
With regard to claims 16 and 17, which respectively recite “the system of claim 1, wherein: (i) the first domain comprises an amino acid sequence having a mutation at one or more positions selected from the group consisting of T375…of human ADAR2 (hADAR2) amino acid sequence, or a corresponding position in a homologous or orthologous ADAR protein,” wherein the mutation is specifically “T375Q,” as set forth above, Doudna anticipates the system of instant claim 1. Furthermore, as set forth above, Doudna discloses that the fusion partner of the system taught therein may be an RNA deaminase such as an ADAR enzyme (paragraphs [0010] and [0121]). However, Doudna does not disclose the instantly claimed ADAR mutations.
This deficiency is cured by Zhang. Zhang discloses systems, methods, and compositions for targeting and editing nucleic acids such as RNA by targeting said RNA with a Cas13 fused to ADAR2 (abstract; Figure 1). Zhang further discloses that the ADAR2 enzymes taught therein may comprise a T375Q mutation, which is associated with increased specificity and increased efficiency (paragraph [1477]); Figure 84). Thus, it is considered that Zhang discloses each and every additional limitation of instant claims 16 and 17.
Given that Doudna discloses systems for facilitating the precise and controlled targeting of Ago binding to single-stranded target nucleic acids such as ssRNA, wherein said Ago is fused to an ADAR enzyme that edits said target nucleic acid, and that Zhang discloses ADAR2 mutations including T375Q, which is associated with increased specificity and increased efficiency, it would have been obvious to someone of ordinary skill in the art before the effective filing date of the claimed invention to modify the ADAR enzyme disclosed in Doudna to comprise the T375Q mutation disclosed in Zhang to predictably target ssRNA with greater efficiency and specificity. One would have been motivated to make such a modification in order to receive the expected benefit of generating a system capable of targeting ssRNA with greater efficiency and specificity.
Claims 30 and 33 are rejected under 35 U.S.C. 103 as being unpatentable over US 2017/0175104 A1 (hereinafter Doudna; as cited in Applicant IDS; of record), as applied to claim 1 above, and further in view of Meister and Tuschl, 2004 (hereinafter Meister).
The disclosure of Doudna is described above and applied as before (see section Claim Rejections - 35 USC § 102). However, this disclosure does not teach the oligonucleotide limitations of instant claims 30 and 33.
With regard to claims 30 and 33, which respectively recite “the system of claim 1, wherein the oligonucleotide is double-stranded and comprises at least one 3’-single stranded overhang or a blunt end,” and “wherein the oligonucleotide comprises a double-stranded region of at least 19 base-pairs,” as set forth above, Doudna anticipates the system of instant claim 1. However, the guide of Doudna is not specifically disclosed to be double-stranded, as instantly claimed (paragraph [0061]). Doudna does disclose that the guide RNA associates with and directs the activities of the Ago polypeptides (and fusions thereof) disclosed therein and further that the guide RNA may be derived from a precursor RNA via modification or cleavage (paragraph [0080]).
As is known to those of ordinary skill in the art and as set forth above, Argonaute (as part of the RISC complex) binds to single stranded RNA following cleavage of dsRNA by Dicer, which subsequently targets specified mRNA for cleavage (reviewed in Meister: see Figure 2). Furthermore, Dicer is known to process long dsRNA into 21-23 nucleotide dsRNA intermediates with 2-nucleotide 3’ overhangs (reviewed in Meister: see page 343, column 2, paragraph 3) that are suitable for interaction with the RISC complex, which comprises Argonaute (reviewed in Meister: see Figure 2 and associated caption). Therefore, one of ordinary skill in the art would have been aware, prior to the effective filing date of the instant application, that dsRNA is processed by Dicer into 21-23 nucleotide dsRNA intermediates that are then processed by the RISC complex, which comprises Argonaute. This processing by Dicer and subsequent processing by the RISC complex is considered to read on the derivation of guide RNAs disclosed in Doudna and set forth above. Thus, it is considered that Doudna and Meister collectively disclose each and every additional limitation of instant claims 30 and 33.
Given that Doudna discloses that the guide RNAs taught therein may be derived from a precursor RNA via modification or cleavage, and that Meister discloses that the natural substrate for Argonaute, as part of the RISC complex, is dsRNA that has been processed by Dicer into 21-23 nucleotide dsRNA intermediates, it would have been obvious to someone of ordinary skill in the art before the effective filing date of the claimed invention to guide Argonaute fused to ADAR (as disclosed in Doudna) via providing dsRNA that has been processed into 21-23 nucleotide dsRNA intermediates (as disclosed in Meister) to predictably target a dsRNA intermediate sequence-specified mRNA sequence for modification with said Argonaute-ADAR fusion (as disclosed in Doudna). One would have been motivated to make such a modification in order to receive the expected benefit of generating a system capable of targeting a user-specified mRNA sequence for modification with an Argonaute-ADAR fusion, as is known in the art.
Claim 35 is rejected under 35 U.S.C. 103 as being unpatentable over US 2017/0175104 A1 (hereinafter Doudna; as cited in Applicant IDS; of record), as applied to claim 1 above, and further in view of Theotokis et al., 2017 (hereinafter Theotokis).
The disclosure of Doudna is described above and applied as before (see section Claim Rejections - 35 USC § 102). However, this disclosure does not teach the oligonucleotide limitations of instant claim 35.
With regard to claim 35, which recites “the oligonucleotide [of the system of claim 1] is double-stranded and comprises a strand having a nucleotide sequence substantially complementary to a target RNA and wherein said strand comprises a mismatch with the target RNA at position 10, counting from [the] 5’-end of said strand,” as set forth above regarding instant claims 30 and 33, it is considered that Doudna and Meister collectively disclose utilizing a double-stranded oligonucleotide to guide an Argonaute-ADAR fusion polypeptide to a targeted RNA site. Furthermore, Doudna discloses that the guide RNA taught therein hybridizes to a target site of the target nucleic acid (paragraph [0080]), meaning said oligonucleotide comprises a strand having a nucleotide sequence complementary to a target RNA, as instantly claimed. While Doudna discloses that hybridization may involve base mismatches, the positions of which are important (paragraph [0035]), they are silent as to the instantly claimed mismatch position.
This deficiency is cured by Theotokis. Theotokis discloses that Argonaute slices between positions 10-11 and requires full Watson-Crick base pairing for said slicing (page 22, column 2, paragraph 1). Therefore, one of ordinary skill in the art would be aware that in the absence of full Watson-Crick base pairing at positions 10-11, Argonaute will not slice the target RNA. As set forth above, Doudna discloses that the Ago of the systems taught therein comprises an amino acid sequence having one or more mutations that exhibit reduced nuclease activity (paragraph [0011]) to facilitate binding of a single stranded target nucleic acid (i.e. RNA) (abstract; paragraph [0005]). Therefore, it is considered that one of ordinary skill in the art would be motivated to reduce Argonaute nuclease activity to facilitate binding of a single stranded target nucleic acid, be it via disrupting full Watson-Crick base pairing at positions 10-11 (as disclosed in Theotokis) or via mutating Argonaute itself (as disclosed in Doudna). Thus, it is considered that Doudna and Theotokis collectively disclose each and every additional limitation of instant claim 35.
Given that Doudna discloses that the Argonaute taught therein may be altered to reduce its nuclease activity for purposes of binding a single stranded target nucleic acid (i.e. RNA), that Meister and Doudna collectively disclose that dsRNA oligonucleotides target Argonaute to a user-specified RNA site, and that Theotokis discloses that full Watson-Crick base pairing is required at positions 10-11 for Argonaute to slice the targeted RNA duplex, it would have been obvious to someone of ordinary skill in the art before the effective filing date of the claimed invention to introduce a mismatch at position 10 (as disclosed in Theotokis) to predictably prevent Argonaute from cleaving the targeted RNA and instead to bind it (as disclosed in Doudna). One would have been motivated to make such a modification in order to receive the expected benefit of generating a system capable of targeting a user-specified RNA sequence for editing with an Argonaute-ADAR fusion by binding said fusion protein to the specified target site with said Argonaute.
Claims 36, 37, 40, 42, and 44 are rejected under 35 U.S.C. 103 as being unpatentable over US 2017/0175104 A1 (hereinafter Doudna; as cited in Applicant IDS; of record), as applied to claim 1 above, and further in view of Meister and Tuschl, 2004 (hereinafter Meister), Aquino-Jarquin, 2020 (hereinafter Aquino-Jarquin), and Bass, 2002 (hereinafter Bass).
The disclosure of Doudna is described above and applied as before (see section Claim Rejections - 35 USC § 102). However, this disclosure does not teach the oligonucleotide limitations of instant claims 36, 37, 40, 42, and 44.
With regard to claims 36 and 37, which respectively recite “the oligonucleotide [of the system of claim 1] is double-stranded and comprises a strand having a nucleotide sequence substantially complementary to a target RNA and wherein: (i) said strand comprises a C at position 21, 22, 23, 24, 25, 26, 27 or 28, counting from [the] 5’-end of said strand, and the strand comprises an A:C mismatch with the target RNA at position 21, 22, 23, 24, 25, 26, 27 or 28, counting from [the] 5’ end of said strand; or (ii) said strand comprises a C at position 4, 5, 6, 7, 8, 9 or 10, counting from [the] 3’-end of said strand, and the strand comprises an A:C mismatch with the target RNA at position 4, 5, 6, 7, 8, 9 or 10, counting from [the] 3’-end of said strand,” and further that “...(i) said strand comprises a C at position 25, counting from [the] 5’-end of said strand, and the strand comprises an A:C mismatch with the target RNA at position 25, counting from [the] 5’-end of said strand; or (ii) said strand comprises a C at position 7, counting from [the] 3’-end of said strand, and the strand comprises an A:C mismatch with the target RNA at position 7, counting from [the] 3’-end of said strand,” as set forth above regarding instant claims 30 and 33, it is considered that Doudna and Meister collectively disclose utilizing a double-stranded oligonucleotide to guide an Argonaute-ADAR fusion polypeptide to a targeted RNA site. Furthermore, Doudna discloses that the guide RNA taught therein hybridizes to a target site of the target nucleic acid (paragraph [0080]), meaning said oligonucleotide comprises a strand having a nucleotide sequence complementary to a target RNA, as instantly claimed. While Doudna discloses that hybridization may involve base mismatches, the positions of which are important (paragraph [0035]), they are silent as to the instantly claimed mismatch position.
This deficiency is cured by Aquino-Jarquin and Bass. Aquino-Jarquin discloses a number of novel engineered programmable systems for ADAR-mediated RNA editing (abstract; Figure 1), which comprise guides with mismatched A:C residues to promote site-specific deamination (page 1066, column 1, paragraph 2; Figure 1C). However, Aquino-Jarquin is silent as to the specific positions at which said mismatch occurs. This deficiency is cured by Bass, which discloses that ADAR specificity displays a wide range in that depending on the substrate, the enzyme can deaminate half of the adenosines before the reaction stops, precisely target one adenosine in the midst of hundreds, or do anything in between these two extremes (page 834, paragraph 3). Bass further discloses that molecules that are less stable because they contain mismatches, bulges, or loops, are deaminated at many fewer sites at reaction completion (i.e. are deaminated more selectively so that deamination occurs at only a few sites) (page 834, paragraph 3). Finally, Bass also discloses that the exact position of loops and other unpaired sequences found in ADAR substrates is critical in maintaining enzyme specificity (page 834, paragraph 2).
Per MPEP 2144.05(II)(A), optimization within prior art conditions through routine experimentation is not inventive. Given that Bass discloses that mismatches facilitate greater ADAR specificity and/or selectivity and further that the exact positions of these mismatches are critical to maintaining enzyme specificity, one of ordinary skill in the art would have been motivated to practice routine experimentation to identify those mismatch positions that most effectively increase ADAR specificity and/or selectivity.
Thus, it is considered that Doudna, Meister, Aquino-Jarquin, and Bass collectively disclose each and every additional limitation of instant claims 36 and 37.
Given that Doudna discloses that the guide RNAs taught therein may be derived from a precursor RNA via modification or cleavage, that Meister discloses that the natural substrate for Argonaute, as part of the RISC complex, is dsRNA that has been processed by Dicer into 21-23 nucleotide dsRNA intermediates, that Aquino-Jarquin discloses that ADAR-mediated RNA editing is facilitated by providing guides with mismatched A:C residues to promote site-specific deamination, and that Bass discloses that mismatches facilitate greater ADAR specificity and/or selectivity and further that the exact positions of these mismatches are critical to maintaining enzyme specificity, one of ordinary skill in the art would have been motivated to practice routine experimentation to identify those mismatch positions that most effectively increase ADAR specificity and/or selectivity to predictably generate an effective guide oligonucleotide capable of specifically targeting a user-specified RNA site for ADAR-mediated editing. One would have been motivated to make such a modification in order to receive the expected benefit of generating an effective guide oligonucleotide capable of specifically targeting a user-specified RNA site for ADAR-mediated editing.
With regard to claim 40, which recites “the oligonucleotide [of the system of claim 1] is double-stranded and comprises a strand having a nucleotide sequence substantially complementary to a target RNA, wherein the target RNA forms a loop structure when hybridized to said strand, and wherein said loop structure comprises a single-stranded C nucleotide,” as set forth above, as set forth above regarding instant claims 30 and 33, it is considered that Doudna and Meister collectively disclose utilizing a double-stranded oligonucleotide to guide an Argonaute-ADAR fusion polypeptide to a targeted RNA site. Furthermore, Doudna discloses that the guide RNA taught therein hybridizes to a target site of the target nucleic acid (paragraph [0080]), meaning said oligonucleotide comprises a strand having a nucleotide sequence complementary to a target RNA, as instantly claimed. While Doudna discloses that hybridization may involve base mismatches, the positions of which are important (paragraph [0035]), they are silent as to the instantly claimed loop structure.
This deficiency is cured by Aquino-Jarquin and Bass. Aquino-Jarquin discloses a number of novel engineered programmable systems for ADAR-mediated RNA editing (abstract; Figure 1), which comprise guides with mismatched A:C residues to promote site-specific deamination (page 1066, column 1, paragraph 2). As depicted in Figure 1B, said mismatched A:C may comprise an A residue in the guide strand and a C residue in the target sequence to facilitate cytidine deamination to uridine. Per Aquino-Arquin, the RESCUE approach (depicted in Figure 1B) produces this conversion via a rationally mutated ADAR2DD (see section RESCUE). However, Aquino-Jarquin is silent as to the instantly claimed loop structure. This deficiency is cured by Bass. As set forth above, Bass discloses that ADAR specificity displays a wide range in that depending on the substrate, the enzyme can deaminate half of the adenosines before the reaction stops, precisely target one adenosine in the midst of hundreds, or do anything in between these two extremes (page 834, paragraph 3). Bass further discloses that molecules that are less stable because they contain mismatches, bulges, or loops, are deaminated at many fewer sites at reaction completion (i.e. are deaminated more selectively so that deamination occurs at only a few sites) (page 834, paragraph 3). Finally, Bass also discloses that the exact position of loops and other unpaired sequences found in ADAR substrates is critical in maintaining enzyme specificity (page 834, paragraph 2). Thus, it is considered that Doudna, Meister, Aquino-Jarquin, and Bass collectively disclose and/or motivate each and every additional limitation of instant claim 40.
With regard to claim 42, which recites “said single-stranded C nucleotide [of the system of claim 40] is at position 6, 7, 8, 9, 10, 11, [or] 12 counting from [the] 5’-end of the loop structure, or said loop structure is in [the] form of [a] hairpin and said C nucleotide is present in a single stranded region of the hairpin,” as set forth above, Bass discloses that molecules comprising loops are deaminated at many fewer sites at reaction completion (i.e. are deaminated more selectively so that deamination occurs at only a few sites) and further that the exact position of said loops is critical in maintaining enzyme specificity (page 834, paragraphs 2 and 3). While Bass does not disclose the instantly claimed position of the single-stranded C (disclosed in Aquino-Jarquin as set forth above: see Figure 1B) within such a loop structure, per MPEP 2144.05(II)(A), optimization within prior art conditions through routine experimentation is not inventive. Given that Aquino-Jarquin discloses that an unpaired C in the target sequence is required for effective deamination thereof via an engineered ADAR, and that Bass discloses that loops facilitate greater ADAR specificity and/or selectivity and further that the exact positions of these loops are critical to maintaining enzyme specificity, one of ordinary skill in the art would have been motivated to practice routine experimentation to identify those loop structures that most effectively increase ADAR selectivity to specifically deaminate a targeted C residue, as disclosed in Aquino-Jarquin. Thus, it is considered that Aquino-Jarquin and Bass collectively disclose and/or motivate each and every additional limitation of instant claim 42.
With regard to claim 44, which recites “(i) said loop structure [of the system of claim 40] is at a position opposite of position 8, 9, 10, 11, 12 or 13, counting from the 3’-end or 5’-end of said strand having a nucleotide sequence substantially complementary to the target RNA; or (ii) said loop structure is at a position opposite of position 8, 9, 10, 11, 12 or 13 counting from the 5’-end of said strand having a nucleotide sequence substantially complementary to the target RNA,” as set forth above, Bass discloses that molecules comprising loops are deaminated at many fewer sites at reaction completion (i.e. are deaminated more selectively so that deamination occurs at only a few sites) and further that the exact position of said loops is critical in maintaining enzyme specificity (page 834, paragraphs 2 and 3). While Bass does not disclose the instantly claimed positioning of said loop, per MPEP 2144.05(II)(A), optimization within prior art conditions through routine experimentation is not inventive. Given that Bass discloses that loops facilitate greater ADAR specificity and/or selectivity and further that the exact positions of these loops are critical to maintaining enzyme specificity, one of ordinary skill in the art would have been motivated to practice routine experimentation to identify those loop structures that most effectively increase ADAR selectivity to specifically deaminate a targeted C residue (as disclosed in Aquino-Jarquin and set forth above). Thus, it is considered that Bass discloses and/or motivate each and every additional limitation of instant claim 44.
Given that Doudna discloses that the guide RNAs taught therein may be derived from a precursor RNA via modification or cleavage, that Meister discloses that the natural substrate for Argonaute, as part of the RISC complex, is dsRNA that has been processed by Dicer into 21-23 nucleotide dsRNA intermediates, that Aquino-Jarquin discloses that ADAR-mediated RNA editing is facilitated by providing guides with mismatched A:C residues to promote site-specific deamination (i.e. either A to I or C to U), and that Bass discloses that internal loops and mismatches facilitate greater ADAR specificity and/or selectivity and further that the exact positions of these loops are critical to maintaining enzyme specificity, one of ordinary skill in the art would have been motivated to practice routine experimentation to identify those mismatch and loop positions that most effectively increase ADAR specificity and/or selectivity to predictably generate an effective guide oligonucleotide capable of specifically targeting a user-specified RNA site for ADAR-mediated editing. One would have been motivated to make such a modification in order to receive the expected benefit of generating an effective system for specifically targeting a user-specified RNA site for ADAR-mediated editing.
Conclusion
No claims are allowed.
Claims 1, 3, 8, 12, 25, 33, 35-37, 40, and 42 are objected to.
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/SARAH E ALLEN/ Examiner, Art Unit 1637
/J. E. ANGELL/ Primary Examiner, Art Unit 1637
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