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 .
After further consideration, the finality of the last Office action was found to be
inappropriate and, therefore, the finality of that action is withdrawn. Prosecution is reopened on
the merits.
Receipt is acknowledged of an amendment, filed 09/04/2026. Claims 1-11 are currently under examination.
Any rejection of record in the previous office actions not addressed herein is withdrawn.
Response to Arguments - Claim Rejections - 35 USC § 102
Applicant’s arguments, see pages 7 to 10 of Applicant’s remarks, filed 09/04/2026, with respect to the rejection(s) of claim 11 under 35 U.S.C. 102(a)(1)/(a)(2) have been fully considered and are persuasive. Therefore, the rejection has been withdrawn. However, upon further consideration, a new ground(s) of rejection is made in view of different interpretation of the previously applied reference.
Claim Rejections - 35 USC § 103
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
Claims 1, 9 and 10 are rejected under 35 U.S.C. 103 as being unpatentable over Popma (WO 2018/156818 A1; Cited in a prior Office Action) as applied in view of Das et al (Biotechnology Journal, 11: 71-79; 2016; Cited in a prior Office Action). This rejection was made in the Office action mailed 05/04/2026.
Regarding Claims 1, 9 and 10, Popma teaches a method comprising (i) introducing into a cell (a) a nuclease system that cleaves a GEMS sequence in the genome present in the nucleus of a host cell (e.g., paragraphs [0294]-[0296]; Fig. 10B), Popma teaches a cell comprising three components where the first component is a donor nucleic acid comprising a nucleotide sequence encoding a green fluorescent protein (selector of the claim) operably linked to a tetracycline (Tet)-inducible promoter and a nucleic acid sequence encoding an exogenous gene to be expressed (e.g., paragraphs [0066], [0294]-[0295], [0318]-[0320]; Figs. 7-8 and 10B) and the third component (a nuclease system that targets a genome) which is a Cas9 complexed with a guide RNA [0283] wherein culturing the cell under conditions to induce the expression of the proteins encoded by the donor (e.g., paragraphs [0241] and [0325]; and (iii) using expression of the reporter gene to facilitate identification and selection of expressing cells from a population of cells (e.g., paragraph [0242]); and (iii) using expression of the reporter gene to facilitate identification and selection of expressing cells from a population of cells (e.g., paragraph [0242]). Popma teaches “flanking insertion sequence” as the homology arms wherein this sequence is homologous to a safe harbor site (e.g., AAVs1, Rosa26, CCR5) of said genome [0063], [0140] and [0278].
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Popma does not teach the specific use of the ETT polypeptide and transient expression of the transactivator by the inducible promoter.
Das teaches the rtTA (comprising the TetR DNA binding domain fused to the VP16 transcriptional activator domain; Page 71, Column 2) and Ptet encoding DNA constructs (variants V10, V16 and M2) were transiently transfected into cells to determine which variant performed best (Page 72, Column 2). Das teaches that improved activity and dox-sensitivity are intrinsic properties of the new rtTA proteins and did not result from increased protein expression or stability (Page 72, Column 2). Das teaches the activity of rtTA variants in transiently transfected cells (Page 73, Figure 1). Das teaches the rtTA V16 and M2 variants demonstrated low but detectable background activity in the absence of dox when the DNA constructs were transiently transfected into cells, while the V10 variant did not (Page 78, Column 2). Das teaches that the V10 variant will be the optimal system for most applications in which transiently transfected cells are used, whereas the V16 variant is optimal for applications in which lentivirally transduced cells are used (Page 78, Column 2).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to modify the teachings of Popma to include the transient transfection of the engineered transcriptional transactivator which would allow for transient expression as taught by Das because Popma teaches it is within the ordinary skill in the art to use a genome editing construct to contact a host cell where the editing system comprises an tetracycline (Tet)-inducible promoter operably linked to a regulatory element comprising a target sequence for integration in the genome and Das teaches that the rtTA V10 variant was successful and optimal in transient expression within transfected cells with no background activity in the absence of dox.
One would have been motivated to make such a modification in order to receive the expected benefit of modulation or control of transcription within the host cell with no background activity in the absence of dox as taught by Das.
Claims 2-5 and 7 are rejected under 35 U.S.C. 103 as being unpatentable over Popma (WO 2018/156818 A1; Cited in a prior Office Action) in view of Das et al (Biotechnology Journal, 11: 71-79; 2016; Cited in a prior Office Action), as applied to claims 1, 9 and 10 above, and in further view of Dechiara et al et al (WO 03020743 A1; Cited in a prior Office Action) as evidenced by Gossen (Science, vol. 268, no. 5218, 23 June 1995; Cited in a prior Office Action). This rejection was made in the Office action mailed 05/04/2026.
The combined teachings of Popma and Das are referenced above as applied to claims 1, 9 and 10.
Regarding claim 2, Pompa teaches sequentially a donor nucleic acid comprising a 5’ homology arm, a nucleotide sequence encoding a green fluorescent protein (selector of the claim) operably linked to a tetracycline (Tet)-inducible promoter and a nucleic acid sequence encoding an exogenous gene to be expressed operably linked to a promoter and a 3’ homology arm (e.g., paragraphs [0066], [0294]-[0295], [0318]-[0320]; Figs. 7-8, 9 and 10B), (ii) culturing the cell under conditions to induce the expression of the proteins encoded by the donor (e.g., paragraphs [0241] and [0325]; and (iii) using expression of the reporter gene to facilitate identification and selection of expressing cells from a population of cells (e.g., paragraph [0242]) but does not teach that the Tet-inducible promoter comprises a minimal promoter. Pompa teaches the insertion of the donor nucleic acid sequence proceeds via homologous or NHEJ in the cell [0306 and 0308].
Pompa and Das do not teach specifically the nucleotide sequence encoding the selector operably linked to a minimal promoter. Pompa and Das do not specifically teach the ETT polypeptide expressed by the second component activates the minimal promoter.
Dechiara teaches constructing a DNA targeting vector, containing a nucleotide sequence comprising: a 5’ homology arm, a protected transgene cassette, and a 3’ homology arm, wherein the protected transgene cassette comprised an exogenous promoter and gene of interest (Page 28; Lines 28-35). Dechiara teaches the additional elements of a transactivator such as tetracycline transactivator, regulatory elements, accessory elements such internal ribosome entry sites (IRES), and recombinases (Page 29; Lines 13-21). Dechiara teaches the promoter is regulated by the regulatory protein, stating for example that if the regulatory protein is reverse tetracycline transactivator (rtTA) then the promoter can be the CMV minimal promoter with Tet operator site (Page 47; Lines 27-35).
As evidenced by Gossen et al (1995), the rtTA protein of Dechiara contains the reverse TetR DBD and VP16 TA of claim 7 (Page 1766, Column 3 and Page 1767, Fig. 1).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to modify the teachings of Popma and Das to include the CMV promoter as a minimal promoter and the promoter is activated via the ETT polypeptide as taught by Dechiara because Popma teaches it is within the ordinary skill in the art to use a genome editing construct to contact a host cell where the editing system comprises a tetracycline (Tet)-inducible promoter operably linked to a regulatory element, where the nucleotide shows transient expression comprising a target sequence for integration in the genome, Das teaches that the rtTA V10 variant was successful and optimal in transient expression within transfected cells with no background activity in the absence of dox and Dechiara teaches a construct comprising the homology arms, gene of interest as well as the transactivator regulatory element such as the reverse tetracycline transactivator that controls the minimal promoter, CMV.
One would have been motivated to make such a modification in order to receive the expected benefit of modulation or control of transcription within the host cell with no background activity in the absence of dox as taught by Dechiara and Das.
Regarding claims 3 and 4, Pompa teaches sequentially a donor nucleic acid comprising a 5’ homology arm, a nucleotide sequence encoding a green fluorescent protein (selector of the claim) operably linked to a tetracycline (Tet)-inducible promoter and a nucleic acid sequence encoding an exogenous gene to be expressed and a 3’ homology arm (e.g., paragraphs [0066], [0294]-[0295], [0318]-[0320]; Figs. 7-8, 9 and 10B), (ii) culturing the cell under conditions to induce the expression of the proteins encoded by the donor (e.g., paragraphs [0241] and [0325]; and (iii) using expression of the reporter gene to facilitate identification and selection of expressing cells from a population of cells (e.g., paragraph [0242]) but does not teach that the Tet-inducible promoter comprises a minimal promoter. Pompa teaches the insertion of the donor nucleic acid sequence proceeds via homologous or NHEJ in the cell [0306 and 0308].
Pompa and Das does not teach specifically the nucleotide sequence encoding the selector operably linked to a minimal promoter and does not sequentially teach the nucleotide sequence before the nucleotide sequence encoding the selector.
Dechiara teaches constructing a DNA targeting vector, containing a nucleotide sequence comprising: a 5’ homology arm, a protected transgene cassette, and a 3’ homology arm, wherein the protected transgene cassette comprised an exogenous promoter and gene of interest (Page 28; Lines 28-35). Dechiara teaches sequentially 5’ homology arm, a splicing acceptor site (SA), a PGK gene followed by a lacZ selector gene operably linked to a promoter and a 3’ homology arm (Page 71, Figures 6-8). Dechiara teaches the additional elements of a transactivator such as tetracycline transactivator, regulatory elements, accessory elements such internal ribosome entry sites (IRES), and recombinases (Page 29; Lines 13-21). Dechiara teaches the promoter is regulated by the regulatory protein, stating for example that if the regulatory protein is reverse tetracycline transactivator (rtTA) then the promoter can be the CMV minimal promoter with Tet operator site (Page 47; Lines 27-35).
As evidenced by Gossen et al (1995), the rtTA protein of Dechiara contains the reverse TetR DBD and VP16 TA of claim 7 (Page 1766, Column 3 and Page 1767, Fig. 1).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to modify the teachings of Popma and Das to include a minimal promoter and the promoter is activated via the ETT polypeptide and the sequential arrangement of the nucleotide of interest before the selectable gene as taught by Dechiara because Popma teaches it is within the ordinary skill in the art to use a genome editing construct to contact a host cell where the editing system comprises an inducible promoter operably linked to a regulatory element, where the nucleotide shows transient expression comprising a target sequence for integration in the genome, Das teaches that the rtTA V10 variant was successful and optimal in transient expression within transfected cells with no background activity in the absence of dox and Dechiara teaches a construct comprising the homology arms, gene of interest as well as the transactivator regulatory element such as the reverse tetracycline transactivator that controls the minimal promoter.
One would have been motivated to make such a modification in order to receive the expected benefit of modulation or control of transcription within the host cell with no background activity in the absence of dox as taught by Dechiara and Das.
Regarding claim 5, Pompa teaches a donor nucleic acid comprising a 5’ homology arm, a nucleotide sequence encoding a green fluorescent protein (selector of the claim) operably linked to a tetracycline (Tet)-inducible promoter and a nucleic acid sequence encoding an exogenous gene to be expressed and a 3’ homology arm (e.g., paragraphs [0066], [0294]-[0295], [0318]-[0320]; Figs. 7-8, 9 and 10B), (ii) culturing the cell under conditions to induce the expression of the proteins encoded by the donor (e.g., paragraphs [0241] and [0325]; and (iii) using expression of the reporter gene to facilitate identification and selection of expressing cells from a population of cells (e.g., paragraph [0242]) but does not teach that the Tet-inducible promoter comprises a minimal promoter.
Pompa and Das do not teach specifically the nucleotide sequence encoding the selector operably linked to a minimal promoter and does not sequentially teach the nucleotide sequence before the nucleotide sequence encoding the selector.
Dechiara teaches constructing a DNA targeting vector, containing a nucleotide sequence comprising: a 5’ homology arm, a protected transgene cassette, and a 3’ homology arm, wherein the protected transgene cassette comprised an exogenous promoter and gene of interest (Page 28; Lines 28-35). Dechiara teaches sequentially 5’ homology arm, a splicing acceptor site (SA), a PGK gene followed by a lacZ selector gene operably linked to a promoter and a 3’ homology arm (Page 71, Figures 6-8). Dechiara teaches the additional elements of a transactivator such as tetracycline transactivator, regulatory elements, accessory elements such internal ribosome entry sites (IRES), and recombinases (Page 29; Lines 13-21). Dechiara teaches the promoter is regulated by the regulatory protein, stating for example that if the regulatory protein is reverse tetracycline transactivator (rtTA) then the promoter can be the CMV minimal promoter with Tet operator site (Page 47; Lines 27-35).
As evidenced by Gossen et al (1995), the rtTA protein of Dechiara contains the reverse TetR DBD and VP16 TA of claim 7 (Page 1766, Column 3 and Page 1767, Fig. 1).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to modify the teachings of Popma and Das to include a minimal promoter and the promoter is activated via the ETT polypeptide and the sequential arrangement of the nucleotide of interest before the selectable gene as taught by Dechiara because Popma teaches it is within the ordinary skill in the art to use a genome editing construct to contact a host cell where the editing system comprises an inducible promoter operably linked to a regulatory element, where the nucleotide shows transient expression comprising a target sequence for integration in the genome, Das teaches that the rtTA V10 variant was successful and optimal in transient expression within transfected cells with no background activity in the absence of dox and Dechiara teaches a construct comprising the homology arms, gene of interest as well as the transactivator regulatory element such as the reverse tetracycline transactivator that controls the minimal promoter, CMV.
One would have been motivated to make such a modification in order to receive the expected benefit of modulation or control of transcription within the host cell with no background activity in the absence of dox as taught by Dechiara and Das.
Regarding claim 7, Pompa teaches a donor nucleic acid comprising a 5’ homology arm, a nucleotide sequence encoding a green fluorescent protein (selector of the claim) operably linked to a tetracycline (Tet)-inducible promoter and a nucleic acid sequence encoding an exogenous gene to be expressed and a 3’ homology arm (e.g., paragraphs [0066], [0294]-[0295], [0318]-[0320]; Figs. 7-8, 9 and 10B), (ii) culturing the cell under conditions to induce the expression of the proteins encoded by the donor (e.g., paragraphs [0241] and [0325]; and (iii) using expression of the reporter gene to facilitate identification and selection of expressing cells from a population of cells (e.g., paragraph [0242]) but does not teach that the Tet-inducible promoter comprises a minimal promoter.
Popma does not teach the specific use of the ETT polypeptide and transient expression of the transactivator by the inducible promoter.
Das teaches the rtTA (comprising the TetR DNA binding domain fused to the VP16 transcriptional activator domain; Page 71, Column 2) and Ptet encoding DNA constructs (variants V10, V16 and M2) were transiently transfected into cells to determine which variant performed best (Page 72, Column 2). Das teaches that improved activity and dox-sensitivity are intrinsic properties of the new rtTA proteins and did not result from increased protein expression or stability (Page 72, Column 2). Das teaches the activity of rtTA variants in transiently transfected cells (Page 73, Figure 1). Das teaches the rtTA V16 and M2 variants demonstrated low but detectable background activity in the absence of dox when the DNA constructs were transiently transfected into cells, while the V10 variant did not (Page 78, Column 2). Das teaches that the V10 variant will be the optimal system for most applications in which transiently transfected cells are used, whereas the V16 variant is optimal for applications in which lentivirally transduced cells are used (Page 78, Column 2).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to modify the teachings of Popma to include the transient transfection of the engineered transcriptional transactivator which would allow for transient expression as taught by Das because Popma teaches it is within the ordinary skill in the art to use a genome editing construct to contact a host cell where the editing system comprises an tetracycline (Tet)-inducible promoter operably linked to a regulatory element comprising a target sequence for integration in the genome and Das teaches that the rtTA V10 variant was successful and optimal in transient expression within transfected cells with no background activity in the absence of dox.
One would have been motivated to make such a modification in order to receive the expected benefit of modulation or control of transcription within the host cell with no background activity in the absence of dox as taught by Das.
Claim 6 is rejected under 35 U.S.C. 103 as being unpatentable over Popma (WO 2018/156818 A1; Cited in a prior Office Action) in view of Das et al (Biotechnology Journal, 11: 71-79; 2016; Cited in a prior Office Action) as applied to claims 1, 9 and 10 above, and in further view of Farzadfard et al (CS Synth Biol. 2013 Oct 18;2(10):604-13). This is a NEW rejection.
The combined teachings of Popma and Das are referenced above as applied to claims 1, 9 and 10.
Regarding Claim 6, Popma does not teach the second component such as the ETT comprising a DNA binding domain fused to a transcriptional activator domain.
Das teaches the rtTA (comprising the TetR DNA binding domain fused to the VP16 transcriptional activator domain; Page 71, Column 2) and Ptet encoding DNA constructs (variants V10, V16 and M2) were transiently transfected into cells to determine which variant performed best (Page 72, Column 2).
Das does not teach the DNA binding domain is a TALE, zinc finger and/or catalytically inactive Cas9 domain.
Farzadfard teaches a functional CRISPR-based transcription factor (crisprTF) which comprises four tandem copies of the Herpes Simplex Viral Protein 16 activation domain (VP64) fused to a nuclease-null (dead) Cas9 (dCas9) under an inducible promoter responsive to conditions such as galactose and anhydrotetracycline (aTc) media to produce activation on synthetic promoters containing a Tet operator (tetO) site (Page 605, Fig. 1). Farzadfard teaches that crisprTFs can be used to build synthetic promoters with tunable strengths by the straightforward engineering of gRNA-binding sites (Page 608, Column 1 bridging Column 2).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to modify the teachings of Popma and Das to include the dCas9-VP64 ETT polypeptide as taught by Farzadfard because Popma and Das teach it is within the ordinary skill in the art to use a DNA binding domain and an effector domain and Farzadfard teaches a functional CRISPR-based transcription factor (crisprTF) which comprises four tandem copies of the Herpes Simplex Viral Protein 16 activation domain (VP64) fused to a nuclease-null (dead) Cas9 (dCas9) under an inducible promoter responsive to conditions such as galactose and anhydrotetracycline (aTc) media to produce activation on synthetic promoters containing a Tet operator (tetO) site.
One would have been motivated to make such a modification in order to receive the expected benefit of crisprTFs that can be used to build synthetic promoters with tunable strengths as taught by Farzadfard.
Claim 8 is rejected under 35 U.S.C. 103 as being unpatentable over Popma (WO 2018/156818 A1; Cited in a prior Office Action) in view of Das et al (Biotechnology Journal, 11: 71-79; 2016; Cited in a prior Office Action) as applied to claims 1, 9 and 10 above, and in further view of Baek et al (US 2019/0062373 A1; Cited in a prior Office Action). This rejection was made in the Office action mailed 05/04/2026.
The combined teachings of Popma and Das are referenced above as applied to claims 1, 9 and 10.
Regarding claim 8, Popma teaches that the construct must include a safe harbor site that is homologous to the safe harbor site of the target sequence, thus implying that the target sequence must have a safe harbor site [0010]. Popma also teaches that the safe harbor site is an adeno-associated virus site 1 (AAVs1) site [0010]. Popma teaches that the safe harbor site is a portion of the genome where one or more donor genes, including transgenes, can integrate with substantially predictable expression and function, but without inducing adverse effects on the host cells or organism [0070].
Popma and Das do not teach the guide RNA is capable of binding to a nucleotide sequence selected from the group consisting of SEQ ID Nos 1-31 of the present disclosure with at least 75% identity thereto.
Baek teaches the use of a guide RNA target sequence that is identical to SEQ ID NO: 26 of the present disclosure and denoted as SEQ ID NO: 36 in Baek [0088]. Baek teaches that the sgRNA targeting human AAVS1 region was synthesized by in vitro transcription using a 118 bp PCR-assembled DNA fragment AAVS1-T23826 as template [0088].
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to modify the teachings of Popma and Das to include the specific sequence for the guide RNA provided as taught by Baek because Popma and Das teach it is within the ordinary skill in the art to use a guide RNA target sequence for the purpose of directing the guide RNA and that safe harbor sites are useful when introducing genes or transgenes to target genomes for substantially predictable expression and function and Baek teaches that the guide RNA target sequence is identical to the sequence of a fragment AAVS1_T23826 template.
One would have been motivated to make such a modification in order to receive the expected benefit of the guide RNA having an identical sequence of the target sequence and expression of the transgene or gene without adverse or deleterious effects taught by Baek and Pompa.
Claim 11 is rejected under 35 U.S.C. 103 as being unpatentable over Popma (WO 2018/156818 A1; Cited in a prior Office Action) as applied in view of Das et al (Biotechnology Journal, 11: 71-79; 2016; Cited in a prior Office Action) and Ahern et al (The Scientist 1995, 9(15):20; Pgs. 1-7). This is a NEW REJECTION.
Regarding claim 11, Popma teaches a cell comprising three components where the first component is a donor nucleic acid comprising a nucleotide sequence encoding a green fluorescent protein (selector of the claim) operably linked to a tetracycline (Tet)-inducible promoter and a nucleic acid sequence encoding an exogenous gene to be expressed (e.g., paragraphs [0066], [0294]-[0295], [0318]-[0320]; Figs. 7-8 and 10B), the second component is a Cas9 (a DNA binding domain) and the third component (a nuclease system that targets a genome) which is a guide RNA [0283] wherein culturing the cell under conditions to induce the expression of the proteins encoded by the donor (e.g., paragraphs [0241] and [0325]; and (iii) using expression of the reporter gene to facilitate identification and selection of expressing cells from a population of cells (e.g., paragraph [0242]). Popma teaches “flanking insertion sequence” as the homology arms wherein this sequence is homologous to a safe harbor site (e.g., AAVs1, Rosa26, CCR5) of said genome [0063], [0140] and [0278].
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Popma does not teach the specific use of transient expression of the transactivator by the inducible promoter.
Das teaches the rtTA (comprising the TetR DNA binding domain fused to the VP16 transcriptional activator domain; Page 71, Column 2) and Ptet encoding DNA constructs (variants V10, V16 and M2) were transiently transfected into cells to determine which variant performed best (Page 72, Column 2). Das teaches that improved activity and dox-sensitivity are intrinsic properties of the new rtTA proteins and did not result from increased protein expression or stability (Page 72, Column 2). Das teaches the activity of rtTA variants in transiently transfected cells (Page 73, Figure 1). Das teaches the rtTA V16 and M2 variants demonstrated low but detectable background activity in the absence of dox when the DNA constructs were transiently transfected into cells, while the V10 variant did not (Page 78, Column 2). Das teaches that the V10 variant will be the optimal system for most applications in which transiently transfected cells are used, whereas the V16 variant is optimal for applications in which lentivirally transduced cells are used (Page 78, Column 2).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to modify the teachings of Popma to include the transient transfection of the engineered transcriptional transactivator which would allow for transient expression as taught by Das because Popma teaches it is within the ordinary skill in the art to use a genome editing construct to contact a host cell where the editing system comprises a tetracycline (Tet)-inducible promoter operably linked to a regulatory element comprising a target sequence for integration in the genome and Das teaches that the rtTA V10 variant was successful and optimal in transient expression within transfected cells with no background activity in the absence of dox.
One would have been motivated to make such a modification in order to receive the expected benefit of modulation or control of transcription within the host cell with no background activity in the absence of dox as taught by Das.
Popma and Das do not teach the collective items of the invention as a kit.
Ahern teaches the major advantages of purchasing products that come already prepared -- such as restriction enzymes, gels, and buffers --are that it enables investigators to save time and money (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 modify the teachings of Popma and Das to include the elements described above for the method of selecting genome edited cells within a kit as taught by Ahern because Popma teaches it is within the ordinary skill in the art to use a genome editing construct to contact a host cell where the editing system comprises a tetracycline (Tet)-inducible promoter operably linked to a regulatory element comprising a target sequence for integration in the genome, Das teaches that the rtTA V10 variant was successful and optimal in transient expression within transfected cells with no background activity in the absence of dox and Ahern teaches the major advantages of purchasing products that come already prepared -- such as restriction enzymes, gels, and buffers --are that it enables investigators to save time and money.
One would have been motivated to make such a modification in order to receive the expected benefit of purchasing all the elements for the method of selecting genome edited cells within a kit as taught by Popma, Das and Ahern.
Response to Arguments - Claim Rejections - 35 USC § 103
The previous rejection of claims 1, 9 and 10 under 35 U.S.C. 103 has been maintained in view of Applicant’s arguments filed on 09/04/2026. Applicant’s arguments have been fully considered but are not found to be persuasive.
Applicant argues Pompa fails to teach the second component in its entirety therefore it is not possible to modify the engineered transcriptional transactivator of Pompa to include transient transfection is flawed due to Pompa does not teach it in any form.
However, Pompa teaches the use of two of the three components where the first component is a donor nucleic acid comprising a nucleotide sequence encoding a green fluorescent protein (selector of the claim) operably linked to a tetracycline (Tet)-inducible promoter and a nucleic acid sequence encoding an exogenous gene to be expressed (e.g., paragraphs [0066], [0294]-[0295], [0318]-[0320]; Figs. 7-8 and 10B) and the third component (a nuclease system that targets a genome) which is a Cas9 complexed with a guide RNA [0283] wherein culturing the cell under conditions to induce the expression of the proteins encoded by the donor (e.g., paragraphs [0241] and [0325]. Popma does not teach the EET secondary component (DNA binding domain and transcriptional activator domain to activate expression of GFP operably linked to the tetracycline (Tet)-inducible promoter which is necessary for activation of GFP expression in the presence of tetracycline [0065 and 0224]), however, Das teaches the rtTA (comprising the TetR DNA binding domain fused to the VP16 transcriptional activator domain; Page 71, Column 2) and Ptet encoding DNA constructs (variants V10, V16 and M2) were transiently transfected into cells to determine which variant performed best (Page 72, Column 2). Specifically, Das teaches that the rtTA V10 variant was successful and optimal in transient expression within transfected cells with no background activity in the absence of dox. Therefore, one would have been motivated to make such a modification in order to receive the expected benefit of modulation or control of transcription within the host cell with no background activity in the absence of dox as taught by Das.
Applicant continues to argue that the Das reference does not envisage a scenario in which the rtTA variant is transiently present in the target cell and the selector is stably inserted into a target locus of the genome of the target cell; specifically, Das only teaches both in the chromosome, or both transiently present as plasmids.
Although Das teaches both in the chromosome or both transiently present as plasmids; the current claim limitations require the selector to be chromosomally inserted, the current limitations of the claims read on that the other can be transiently transfected in an episome. Therefore, Das teaches the rtTA and Ptet-luciferase DNA constructs are episomally present in the cells (Page 76, Column 2) due to the rtTA proteins binding more efficiently to episomal than integrated Ptet sequence elements due to differences in the plasmid DNA versus chromatin DNA structure (Page 78, Column 2). In addition, Das teaches the GFP selector is chromosomally inserted along with the Ptet promoter (Page 77, Column 2).
Applicant argues that the instant invention’s design provides advantages that were not appreciated in the prior art, as explained in detail in Applicant's remarks from February 2026, such as avoiding problems associated with constitutive expression of a selector component, specifically, ensuring insertion of the selector at a target locus without risking constitutive expression of the selector.
However, as recited above, Popma teaches inducible expression of the reporter. Specifically, Pompa teaches the use of two of the three components where the first component is a donor nucleic acid comprising a nucleotide sequence encoding a green fluorescent protein (selector of the claim) operably linked to a tetracycline (Tet)-inducible promoter and a nucleic acid sequence encoding an exogenous gene to be expressed (e.g., paragraphs [0066], [0294]-[0295], [0318]-[0320]; Figs. 7-8 and 10B) and the third component (a nuclease system that targets a genome) which is a Cas9 complexed with a guide RNA [0283] wherein culturing the cell under conditions to induce the expression of the proteins encoded by the donor (e.g., paragraphs [0241] and [0325]. Therefore, nothing unexpected is shown.
Conclusion
Any inquiry concerning this communication or earlier communications from the examiner should be directed to ALEXANDRA ROSE LIPPOLIS whose telephone number is (703)756-5450. The examiner can normally be reached Monday-Friday, 8:00am to 5:00pm EST.
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/ALEXANDRA ROSE LIPPOLIS/Examiner, Art Unit 1637
/Jennifer Dunston/Supervisory Patent Examiner, Art Unit 1637
