DETAILED ACTION
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
Receipt is acknowledged of the preliminary amendment filed 05/01/2023 in which claims 9, 10, 16, 19, and 20 were amended to remove multiple dependencies.
Claims 1-20 are pending and under consideration.
Priority
This application is a 371 of PCT/US2021/058315 with a filing date of 11/04/2021, which claims benefit of 63/109,909 with a filing date of 11/05/2020.
Claims 1-20 have the effective filing date of 11/05/2020.
Information Disclosure Statement
The information disclosure statement(s) filed on 05/01/2023 and 11/12/2024 have been acknowledged and all references are considered.
Specification
The use of the term(s) “TAKARA BIO ®” and “CELL BIOLABS, INC.®”, which are a trade name or a mark used in commerce, have been noted in this application. The term(s) should be accompanied by the generic terminology; furthermore the term should be capitalized wherever it appears or, where appropriate, include a proper symbol indicating use in commerce such as ™, SM , or ® following the term.
Although the use of trade names and marks used in commerce (i.e., trademarks, service marks, certification marks, and collective marks) are permissible in patent applications, the proprietary nature of the marks should be respected and every effort made to prevent their use in any manner which might adversely affect their validity as commercial marks.
Claim Objections
Claims 8-10, 16, 19-20 are objected to because of the following informalities:
Claim 8 recites the limitation "a complex of claim 5." The claim would be clearer if it recited “the complex of claim 5.” The It would be remedial to amend these claim(s) to recite, “the complex of claim 5.”
Claim(s) 9, 10, 16 19, and 20 recite the limitation "a fusion protein of claim 1." The claims would be clearer if they recited “the fusion protein of claim 1.” It would be remedial to amend these claim(s) to recite, “the fusion protein of claim 1.”
Appropriate correction or clarification 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 8 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).
Claim 8 does not explicitly recite “human organism”, however the instant specification discloses several different cell types that the cell can be as well as “modifying the cells ex vivo using a system as described herein, and reintroducing the cells or their progeny into the individual or an immunologically matched individual for prophylaxis and/or therapy of a condition, disease or disorder, as described above... The disclosure is thus suitable for a wide range of human, veterinary, experimental animal, and cell culture uses.” (paragraph [0049]). Notwithstanding, the instant application discloses treating muscular dystrophy (MD) (paragraph [0006]), and correcting the human dystrophin gene (paragraph [0018]). Therefore, claim 8 encompasses a human cell that can be modified ex vivo and reintroduced into a human, which encompasses cells in vivo in a human (i.e., a human organism), which is excluded from the scope of patentable subject matter.
Limiting claim 8 to “an isolated cell” would obviate the rejection.
Claim Rejections - 35 USC § 112
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claim 4 recites the limitation "further comprising the first linker amino acid sequence that links the MS2 segment to a first nuclear localization signal" in the first and second line of the claim. There is insufficient antecedent basis for this limitation (first linker amino acid sequence) in the claim. This claim depends upon claim 3 which recites “…wherein the T4 DNA polymerase segment and the segment of the MS2 protein are separated by a first linker sequence.” (Emphasis add).
Claim Rejections - 35 USC § 112
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(s) 6 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 6 further comprises, “a guide RNA comprising MS2 protein binding sequences”, however, claim 5 from which it depends on recites, “…a guide RNA comprising MS2 bacteriophage coat protein binding sites.” MS2 protein binding sequences is broader than MS2 bacteriophage coat protein binding sites, and therefore does not further limit claim 5.
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)(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.
Claim(s) 1-3, 9, 16-17, 19, and 20 are rejected under 35 U.S.C. 102(a)(2) as being anticipated by Maresca (US -20230340538 A1, effective filing date of 04/08/2020).
Regarding instant Claim 1, Maresca discloses a fusion protein comprising the NHEJ promoting domain (T4 DNA polymerase in [0008] and claim 9) and an MCP2 (an MS2 Coat protein in [0010], more specifically a MS2 bacteriophage coat protein is “described in references including [0157] Peabody et al, which teaches MS2 bacteriophage coat protein”) see paragraph [0174] and Figure 1B.
Regarding instant Claim 2, Maresca discloses that in some embodiments the fusion
protein comprises a nuclear localization signal (NLS) (Paragraph [0134]).
Regarding instant Claim 3, Maresca discloses that the fusion protein further comprises a linker that links that Cas nuclease domain and the reverse transcriptase, DNA polymerase, or DNA ligase (Paragraph [0135]).
Regarding claim 9, Maresca discloses that the fusion protein is transfected into cells indicating that the fusion protein in a composition appropriate for delivery into living cells, thus meeting the structural limitations of the claimed pharmaceutical formulation (Paragraph [0215]),
Regarding instant Claim 16 and Claim 17, Maresca discloses, “In some embodiments, the disclosure provides a kit comprising the fusion protein provided herein. In some embodiments, the fusion protein in the kit is provided as a polynucleotide encoding the fusion protein. In some embodiments, the polynucleotide encoding the fusion protein is provided on a vector, e.g., a vector described herein (Paragraph [0187] and [0190]).
Regarding instant Claim 19, Maresca discloses, “In some embodiments, the present disclosure provides a host cell including any of the expression vectors described herein, e.g., an expression vector including a polynucleotide encoding a nuclease, a fusion protein, or a variant thereof.”, (Paragraph [0078] and [0162]).
Regarding instant Claim 20, Maresca discloses, “In some embodiments, the sequence of interest is reverse transcribed by the reverse transcriptase to generate a first cDNA.”, (Paragraph 0142]).
Claim Rejections - 35 USC § 103
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
Claim(s) 1-8, and 19 is/are rejected under 35 U.S.C. 103 as being unpatentable over Xu et al (Cited on IDS from 05/01/2023, non-patent literature #1) in view of Liu et al (Cited on IDS from 05/01/2023, non-patent literature #2).
Regarding instant Claim 1, which recites:
A fusion protein comprising a T4 DNA polymerase segment and a segment of an MS2 bacteriophage coat protein.
Xu et al teaches a CRISPR-based approach for targeted demethylation of specific genomic loci. Pertaining to instant claim 1, Xu et al teaches a fusion protein of a Tet1-Catalytic Domain to an MS2 RNA element-containing sgRNA2.0 system-guided dCas9 and MS2 bacteriophage coat protein (Page 2, Col 1, Paragraph 3; and Figure 1a)
Xu et al does not teach a T4 DNA polymerase fused to an MS2 bacteriophage coat protein.
Liu et al teaches the use of T4 DNA polymerase for DNA end repairing of the breaks induced by a CRISPR/Cas9 system.
Thus, taking into consideration the success of Xu et al with targeted demethylation of specific genomic loci from the fusion protein of Tet1-CD and MS2 bacteriophage coat protein, and the success of Liu et al with the addition of T4 DNA polymerase for end repairing, it would have been obvious to one of ordinary skill in the art to substitute the Tet1-CD domain of Xu et al with the T4 DNA polymerase of Liu et al to yield the predictable invention of instant claim 1 of a fusion protein comprising a T4 DNA polymerase segment and a segment of an MS2 bacteriophage coat protein.
Regarding instant Claim 2, which recites:
The fusion protein of claim 1, further comprising at least one nuclear localization signal.
In combination, Xu et al and Liu et al teach the disclosed fusion protein of instant claim 1. Xu et al further teaches the full amino-acid sequence of MS2-NLS-Tet1 CD (Materials and methods section under plasmid construction, Page 10, Column 2, Paragraph 2). More specifically, the NLS separating MS2 and Tet1-CD is SPKKKRKVEAS (Supplementary methods, see supplementary materials II).
Regarding instant Claim 3, which recites:
The fusion protein of claim 2, wherein the T4 DNA polymerase segment and the segment of the MS2 protein are separated by a first linker sequence.
In combination, Xu et al and Liu et al teach the disclosed fusion protein of instant claim 2, which is dependent on instant claim 1. Xu et al further teaches that the MS2 protein and Tet1 CD are separated by a linker (Page 2, Column 1, Results section, Lines 9-11).
Regarding instant Claim 4, which recites:
The fusion protein of claim 3, further comprising the first linker amino acid sequence that links the MS2 segment to a first nuclear localization signal, and a second linker sequence that links the T4 DNA polymerase segment to a second nuclear localization signal.
In combination, Xu et al and Liu et al teach the disclosed fusion protein of instant claim 3, which is dependent on instant claim 2, which depends upon instant claim 1. Xu et al discloses a linker sequence connecting the MS2 protein to the NLS sequence.
Xu et al does not teach a second linker sequence that links the Tet1-CD to a second nuclear localization signal.
It would have been obvious to try to one of ordinary skill in the art at before the effective filing date of the claimed invention to add an additional linker and nuclear localization signal to the fusion protein, as disclosed by Xu, in order to improve nuclear localization. This is a routine and convention addition as well as in another plasmid disclosed by Xu et al, that contains the dCas9 and Tet1-CD fusion protein, there are two nuclear localization signals (Supplementary methods, Supplementary Materials I).
Regarding instant Claim 5, which recites:
A complex comprising a double stranded DNA template, a Cas enzyme, a guide RNA comprising MS2 bacteriophage coat protein binding sites, a protein comprising a T4 DNA polymerase, and an MS2 binding protein.
Xu et al discloses a complex comprising a double stranded DNA template, a dCas enzyme, a guide RNA comprising MS2 bacteriophage coat protein binding sites, and a protein comprising an MS2 binding protein (see figure 1a for all components listed above).
Xu et al does not disclose a Cas enzyme, or a protein comprising T4 DNA polymerase.
Liu et al discloses a Cas enzyme (Abstract) and a T4 DNA polymerase (Used for end repair of breaks generated by the CRISPR/Cas9 system; abstract).
With the success of the complex disclosed in Xu et al, and of the system disclosed in Liu et al, it would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to substitute the dCas9 enzyme from Xu et al with the Cas9 enzyme of Liu et al, and to substitute the Tet1-CD segment of the MS2-Tet1 CD fusion protein of Xu et al with the T4 DNA polymerase of Liu et al to provide a complex comprising a double stranded DNA template, a Cas enzyme, a guide RNA comprising MS2 bacteriophage coat protein binding sites, a protein comprising a T4 DNA polymerase, and an MS2 binding protein, for efficient repairing of Cas9 generated sticky ends, as disclosed by Liu et al.
Regarding instant Claim 6, which recites:
The complex of claim 5, further comprising a guide RNA comprising MS2 protein binding sequences.
In combination, Xu et al and Liu et al teach the disclosed complex of claim 5, from which instant claim 6 depends. Xu et al further discloses the guide RNA comprising MS2 protein binding sequences. More specifically, Xu et al teaches a strategy for targeted demethylation of specific genomic loci by tethering Tet1-CD both to MS2 RNA element-containing sgRNA2.0 system-guided dCas9 and MS2 bacteriophage coat protein (Page 2, Column 1, Paragraph 3; Abstract).
Regarding instant Claim 7, which recites:
The complex of claim 5, wherein the Cas enzyme is Cas9.
In combination, Xu et al and Liu et al teach the disclosed complex of claim 5, from which instant claim 7 depends.
Xu et al teaches dCas9.
Xu et al does not teach Cas9.
Liu et al discloses Cas9 (Abstract).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to substitute the dCas9 enzyme of Xu et al with the Cas9 enzyme of Liu et al in order to provide the endonuclease activity required for the complex claimed in instant claim 5.
Regarding instant Claim 8, which recites:
A cell comprising a complex of claim 5.
In combination, Xu et al and Liu et al teach the disclosed complex of claim 5, from which instant claim 8 depends. Xu et al further discloses a cell comprising a complex. More specifically, Xu et al discloses that the MS2 coat protein gene fused with Tet1-CD was inserted into pcDNA3.1/hygro(+) vector (Page 10, Materials and Methods, Plasmid construction), Designed sgRNAs were synthesized as oligos, annealed and cloned into pdCas9-Tet1-CD vector at BbsI digestion sites (Page 10, Materials and Methods, Plasmid construction). Xu et al further teaches transfecting three different cell lines with components of the complex to yield stabling expressing cells (Hela Cells - Page 2, Column 2, Paragraph 1 and Figure 1d; HEK-293FT cells – Page 2, Column 2, Paragraph 2 and Figure 1c; SH-SY5Y cells – Page 5, Column 2, Paragraph 2 and Figure 2e).
Regarding instant Claim 19, which recites:
An expression vector encoding a fusion protein of claim 1.
In combination, Xu et al and Liu et al teach the disclosed fusion protein of claim 1. Xu et al further teaches that the MS2 coat protein gene fused with Tet1-CD was inserted into pcDNA3.1/hygro(+) vector (Page 10, Materials and Methods, Plasmid construction).
Claim 10 is rejected under 35 U.S.C. 103 as being unpatentable over Liu et al (Cited on IDS from 05/01/2023, non-patent literature #2) in view of Xu et al (Cited on IDS from 05/01/2023, non-patent literature #1).
Regarding instant Claim 10, which recites:
A method for producing an indel at a selected chromosome locus in a cell, the method comprising introducing into the cell a fusion protein of claim 1, a Cas enzyme, and a guide RNA comprising MS2 protein binding sites, such that the T4 DNA polymerase and the MS2 binding protein, the Cas enzyme, and the guide RNA produce the indel at the selected chromosome locus.
Liu et al discloses a method for producing an indel in a selected DNA molecule, the method comprising a Cas enzyme (Cas9 enzyme – Page 2, Column 1, Paragraph 2), a guide RNA (sgRNA3 – Page 2, Column 1, Paragraph 2), and a T4 DNA polymerase (Page 2, Column 1, Paragraph 2), such that the T4 DNA polymerase, the case enzyme, and the guide RNA produce the indel in a selected DNA molecule (Page 2, Column 2, Paragraph 3, and FIG 1F,G,H, and I).
Liu et al does not disclose a method for producing an indel at a selected chromosome locus in a cell, the method comprising introducing into the cell a fusion protein of claim 1, a Cas enzyme, and a guide RNA comprising MS2 protein binding sites, such that the T4 DNA polymerase and the MS2 binding protein, the Cas enzyme, and the guide RNA produce the indel at the selected chromosome locus.
Xu et al discloses producing a DNA modification at a selected chromosome locus in a cell (Page 2, Column 1, Paragraph 3; Page 2, Column 2, Paragraph 2; and Page 5, Column 1, Paragraph 1), the method comprising introducing into a cell a fusion protein comprising MS2 bacteriophage coat protein, and a guide RNA comprising MS2 protein binding sites. More specifically, Xu et al discloses that the MS2 coat protein gene fused with Tet1-CD was inserted into pcDNA3.1/hygro(+) vector and guide RNAs were cloned into pdCas9-Tet1-CD vector, and consequently transfected into a HEK-293FT cell-line (Page 2, Column 1, Paragraph 3; Page 2, Column 2, Paragraph 2; and Page 10, Materials and Methods, Plasmid construction, and cell culture and transfection).
Given the success of Liu et al with producing indels in a selected DNA molecule and the success of Xu et al for introducing a CRISPR/Cas system into a cell comprising an MS2 fusion protein for targeted demethylation of specific genomic loci, it would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to apply the known technique of Xu et al (a method of producing a DNA modification at a selected chromosome locus in a cell through introducing into a cell a fusion protein and a guide RNA with MS2 protein binding sites) to the known technique of Liu et al (a method for producing indels in a selected DNA molecule) to yield the predictable method for producing an indel at a selected chromosome locus in a cell comprising introducing into the cell a fusion protein of instant claim 1 as disclosed by the combination of Xu et al and Lui et al (see above rejection regarding claim 1), the guide RNA comprising MS2 protein binding sites, and the Cas Enzyme, such that the T4 DNA polymerase and the MS2 binding protein, the Cas enzyme and the guide RNA produce the indel at the selected chromosome locus.
Claim 11 is rejected under 35 U.S.C. 103 as being unpatentable over Liu et al (Cited on IDS from 05/01/2023, non-patent literature #2) in view of Xu et al (Cited on IDS from 05/01/2023, non-patent literature #1) in further view of Shen et al (Cited on IDS from 05/01/2023, non-patent literature #4).
Regarding instant Claim 11, which recites:
The method of claim 10, wherein the indel corrects a mutation in an open reading frame encoded by the selected chromosome locus.
In combination, Liu et al in view of Xu et al disclose the method of instant claim 10, for which instant claim 11 depends upon.
Liu et al does not disclose wherein the indel corrects a mutation in an open reading frame encoded by the selected chromosome locus.
Shen et al teaches wherein an indel corrects a mutation in an open reading frame. More specifically, Shen et al teaches that template-free Cas9 editing is predictable and capable of precise repair to a predicted genotype, enabling corrections of human disease-associated mutations (Page 2, Summary), wherein Shen et al experimentally confirmed insertions and deletions in 195 human disease-relevant alleles (Page 2, Summary) through using inDelphi to estimate the fraction of SpCas9 gRNAs targeting exons* and introns in the human genome to support precise end-joining repair (Page 6, Paragraph 3).
*a part of an open reading frame
It would have been obvious to one of ordinary skill in the art at the time before the effective filing date of the invention to utilize the combined method of claim 10, as taught by Lui et al in view of Xu et al, and modify the disclosed method with the teachings of Shen et al, to arrive at the predictable method wherein the indel corrects a mutation in an exon (a part of an open reading frame) at a selected chromosome locus. This modification could predictably yield the benefits of correcting human disease-associated mutations, as suggested by Shen et al (Introduction, Paragraph 1).
Claim(s) 12-15 is/are rejected under 35 U.S.C. 103 as being unpatentable over Liu et al (Cited on IDS from 05/01/2023, non-patent literature #2) and Xu et al (Cited on IDS from 05/01/2023, non-patent literature #1) in view of Shen et al (Cited on IDS from 05/01/2023, non-patent literature #4) in further view of Min et al (Cited on IDS from 05/01/2023, non-patent literature #5).
Regarding instant Claim 12, which recites:
The method of claim 11, wherein the selected chromosome locus comprises a mutation in a gene that is correlated with a monogenic disease.
In combination, Liu et al, Xu et al, and Shen et al, teach the method of claim 11 for which instant claim 12 is dependent upon.
Liu et al does not disclose wherein the selected chromosome locus comprises a mutation in a gene that is correlated with a monogenic disease.
Min et al suggests a chromosome locus comprising a mutation in a gene that is correlated with a monogenic disease. More specifically, Min et al suggests, “More than 800 monogenic disorders result in degeneration or dysfunction of skeletal muscle… Roughly two-thirds of dystrophin mutations involve exon deletions that disrupt the dystrophin open reading frame (ORF)…”, (Page 1-2, Introduction, Lines 1 and 11-12).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to take the teachings of Liu et al, Xu et al, and Shen et al, and the suggestion of Min et al, to yield the predictable method of producing an indel at a selected chromosome locus in a cell comprising a mutation in a gene that is correlated with a monogenic disease, in order to correct said gene for treating a patient that has said monogenic disease, as suggested by Min et al.
Regarding instant Claim 13, which recites:
The method of claim 12, wherein the monogenic disease is muscular dystrophy, and wherein the gene encodes a mutated dystrophin protein.
In combination, Liu et al, Xu et al, Shen et al, and Min et al teach the method of claim 12 for which instant claim 13 is dependent upon.
Liu et al does not disclose wherein the monogenic disease is muscular dystrophy, and wherein the gene encodes a mutated dystrophin protein.
Min et al discloses the monogenic disease, muscular dystrophy. More specifically, Min et al suggests, “… Among the most severe is Duchenne muscular dystrophy (DMD), which is caused by mutations in the X-linked dystrophin gene… DMD is the most common lethal monogenic disorder… More than 3,000 different dystrophin mutations have been shown to cause DMD…Roughly two-thirds of dystrophin mutations involve exon deletions that disrupt the dystrophin open reading frame (ORF), and point mutations or duplications account for the rest”, (Page 1-2, Introduction, Paragraph 1). Min et al further suggests, “Most recently, gene editing has been explored as a possible means of permanently removing genetic mutations that cause DMD, thereby restoring production of the missing dystrophin protein.”, (Page 2, Introduction, Paragraph 4).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to take the teachings of Liu et al, Xu et al, and Shen et al, and the suggestion of Min et al, to yield the predictable method of producing an indel in a mutated dystrophin gene that encodes a mutated dystrophin protein in order to correct said gene and to produce the corrected dystrophin protein in patients with the monogenic disease of muscular dystrophy, as suggested by Min et al.
Regarding instant Claim 14, which recites:
The method of claim 13, wherein the indel corrects the gene encoding the mutated dystrophin protein.
In combination, Liu et al, Xu et al, Shen et al, and Min et al teach the method of claim 13 for which instant claim 14 is dependent upon.
Liu et al does not disclose wherein the indel corrects the gene encoding the mutated dystrophin gene.
Min et al suggests, wherein an indel corrects the gene encoding the mutated dystrophin protein. More specifically, Min et al suggests, “In the absence of an exogenous DNA template, a sgRNA can direct Cas9 to introduce a double-stranded break (DSB) in DNA, which is subsequently repaired through an imprecise process known as nonhomologous end-joining (NHEJ), resulting in insertions and deletions (indels). This type of editing has been especially effective in deleting splice donor or acceptor site sequences in out-of-frame exons, thereby allowing restoration of the ORF of the dystrophin gene.” (Page 4, Paragraph 2).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to take the teachings of Liu et al, Xu et al, and Shen et al, and the suggestion of Min et al, to yield the predictable method of producing an indel that corrects the gene encoding the mutated dystrophin protein, as suggested by Min et al, in order to provide a method that corrects dystrophin genes encoding a mutated dystrophin protein, develop a treatment method for patients suffering from muscular dystrophy.
Regarding instant Claim 15, which recites:
The method of claim 14, wherein the indel comprises a one or two base pair insertion.
In combination, Liu et al, Xu et al, Shen et al, and Min et al teach the method of claim 14 for which instant claim 15 is dependent upon.
Liu et al does not teach wherein the indel comprises a one or two base pair insertion.
Shen et al teaches a one base pair insertion. More specifically, Shen et al teaches, “Building on this idea of precision gRNAs, we used inDelphi to design 14 gRNAs for high-precision template-free editing yielding predictable 1-bp insertion genotypes in endogenous human disease-relevant loci and experimentally confirmed highly precise editing (median 61% among edited products) in two human cell lines.”, (Page 3, Paragraph 2).
It would have been obvious to one of ordinary skill in the art before the effective filing date to combine the teachings of Liu et al, Xu et al, and Min et al, with the teachings of Shen et al, to yield the predictable method of producing an indel that corrects the gene encoding the mutated dystrophin protein as disclosed by the combinations of Liu et al, Xu et al, Shen et al, and Min et al, wherein that indel comprises a one or two base pair insertion, in order to provide a method for producing an indel that comprises one base pair insertion, in order to correct human disease associated mutations, which is further taught by Shen et al.
Claim(s) 9, 16-18 is/are rejected under 35 U.S.C. 103 as being unpatentable over Xu et al (Cited on IDS from 05/01/2023, non-patent literature #1) in view of Liu et al (Cited on IDS from 05/01/2023, non-patent literature #2) in further view of Chevessier-Tuennesen et al (Cited on IDS from 05/01/2023, Foreign Patent Document #1).
Regarding instant Claim 9, which recites:
A pharmaceutical formulation comprising a fusion protein of claim 1.
In combination, Xu et al and Liu et al teach the disclosed fusion protein of claim 1 of which this claim depends on.
Xu et al and Liu et al do not disclose a pharmaceutical formulation.
Chevessier-Tuennesen et al discloses a pharmaceutical composition containing RNAs encoding CRISPR-associated proteins for the treatment and/or prophylaxis of diseases amenable to treatement with CRISPR-associated proteins. More specifically, Chevessier-Tuennesen et al discloses, “The present invention relates to artificial nucleic acids, in particular RNAs, encoding CRISPR-associated proteins, and (pharmaceutical) compositions and kit-of-parts comprising the same. Said artificial nucleic acids, in particular RNAs, (pharmaceutical) compositions and kits are inter alia envisaged for use in medicine, for instance in gene therapy, and in particular in the treatment and/or prophylaxis of diseases amenable to treatment with CRISPR-associated proteins, e.g. by gene editing, knock-in, knock-out or modulating the expression of target genes of interest.” (Page 1, Lines 6-11).
Given the success of Xu et al for introducing a CRISPR/Cas system into a cell comprising an MS2 fusion protein for targeted demethylation of specific genomic loci, the success of Liu et al for producing indels in a given DNA molecule utilizing T4 DNA polymerase, and the success of Chevessier-Tuennesen et al for generating pharmaceutical compositions containing RNAs encoding CRISPR-associated proteins, it would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to combine the teachings of Xu et al and Liu et al of the fusion protein with a pharmaceutical composition as taught by Chevessier-Tuennesen et al since the fusion protein disclosed by Xu et al is part of a DNA modification system that could be used as a medical therapeutic (Page 10, Column 2, Paragraph 1 and abstract).
Regarding instant Claim 16, which recites:
A kit comprising a fusion protein of claim 1, or an expression vector encoding said fusion protein.
In combination, Xu et al and Liu et al teach the disclosed fusion protein of claim 1 of which this claim depends on.
Xu et al and Liu et al do not disclose kit comprising said fusion proteins.
Chevessier-Tuennesen et al discloses a kit (Page 1, Lines 6-11).
Given the success of Xu et al for introducing a CRISPR/Cas system into a cell comprising an MS2 fusion protein for targeted demethylation of specific genomic loci, the success of Liu et al for producing indels in a given DNA molecule utilizing T4 DNA polymerase, and the success of Chevessier-Tuennesen et al for generating a “kit-of-parts” containing RNAs encoding CRISPR-associated proteins, it would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to combine the teachings of Xu et al and Liu et al of the fusion protein with a “kit-of-parts” as taught by Chevessier-Tuennesen et al since the fusion protein disclosed by Xu et al is part of a DNA modification system that could be used in basic research as well as medical therapeutics (Page 10, Column 2, Paragraph 1 and abstract).
Regarding instant Claim 17, which recites:
The kit of claim 16, further comprising a Cas enzyme or an expression vector encoding a Cas enzyme.
In combination, Xu et al and Liu et al teach the disclosed fusion protein of claim 1 of which claim 16 depends on, and in combination, Xu et al, Liu et al, and Chevessier-Tuennesen et al teach the disclosed kit of claim 16, of which claim 17 depends upon.
Xu et al and Liu et al do not disclose a kit comprising a Cas enzyme or an expression vector encoding a Cas enzyme.
Chevessier-Tuennesen et al discloses a kit comprising an expression vector (Page 6-7, lines 38-13) encoding a Cas enzyme (Page 28, Lines 27-41). It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to combine the fusion protein as disclosed in Xu et al with the kit comprising an expression vector encoding a Cas enzyme as disclosed by Chevessier-Tuennesen et al in order to yield the predictable results of combining the DNA modifying fusion protein with the DNA modifying capabilities in a kit.
Regarding instant Claim 18, which recites:
The kit of claim 17, further comprising a guide RNA or an expression vector encoding said guide RNA, wherein the guide RNA comprises MS2 protein binding sequences, and wherein the guide RNA comprises a sequence targeted to a selected chromosome locus.
In combination, Xu et al and Liu et al teach the disclosed fusion protein of claim 1 of which claim 16 depends on, and in combination, Xu et al, Liu et al, and Chevessier-Tuennesen et al teach the disclosed kit of claim 16 and 17, of which claim 18 depends upon.
Xu et al further discloses a guide RNA comprising MS2 protein binding sequences (Figure 1a; Page 2, Column 1, Paragraph 4; and Abstract), and wherein the guide RNA comprising a sequence targeted to a selected chromosome locus (Page 2, Column 2, Paragraph 1; Supplementary Table 1; and Figure 1a).
Claim 20 is rejected under 35 U.S.C. 103 as being unpatentable over Xu et al (Cited on IDS from 05/01/2023, non-patent literature #1) in view of Liu et al (Cited on IDS from 05/01/2023, non-patent literature #2) in further view of Okayama (Cited on IDS from 05/01/2023, non-patent literature #3).
Regarding instant Claim 20, which recites:
A cDNA encoding a fusion protein of claim 1.
In combination, Xu et al and Liu et al teach the disclosed fusion protein of claim 1 of which claim 20 depends upon.
Xu et al and Liu et al do not disclose a cDNA encoding a fusion protein.
Okayama discloses cDNA.
It would have been obvious to try by one of ordinary skill in the art before the effective filing date of the invention to make cDNA, as disclosed by Okayama encoding the fusion protein, as disclosed by Xu et al, to yield the predictable results of improving expression of the fusion protein.
Conclusion
No claims are allowed.
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/L.M.T./Examiner, Art Unit 1637
/J. E. ANGELL/Primary Examiner, Art Unit 1637