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 .
Receipt is acknowledged of an amendment, filed 02/17/2026. Claims 1-11 are pending.
Any rejection of record in the previous office actions not addressed herein is withdrawn.
This action is FINAL.
Claim Rejections - 35 USC § 102
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 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.
Claim 11 is rejected under 35 U.S.C. 102(a)(1)/(a)(2) as being anticipated by Popma (WO 2018/156818 A1). This rejection was made in the Office action mailed 02/17/2026.
Regarding claim 11, the claim is interpreted that because Popma teaches all of the listed items, then it teaches the kit.
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 TALEN (an ETT) fused to the third component (a nuclease system that targets a genome) which is a Fok1 endonuclease [0142] 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].
Response to Arguments - Claim Rejections - 35 USC § 102
The previous rejection of claim 11 under 35 U.S.C. 102(a)(1)/(a)(2) has been maintained in view of Applicant’s arguments filed on 02/17/2026. Applicant’s arguments have been fully considered but are not found to be persuasive.
Applicant argues Pompa does not teach all the elements of the kit specified in claim 11.
However, the claim recites “A kit comprising a first component, a second component and a third component and, optionally, a cell population; wherein the first component is a donor reporter cassette comprising a nucleotide sequence encoding a selector and a further nucleotide sequence and, optionally, a minimal promoter operably linked to a regulatory element; wherein the second component is an engineered transcriptional transactivator (ETT) polypeptide or a nucleotide sequence encoding an ETT polypeptide; wherein the ETT polypeptide comprises a DNA binding domain (DBD) and at least one transcription activator (TA) domain; and wherein the third component is a nuclease system comprising a genome targeted nuclease and, optionally, a guide RNA (gRNA) comprising at least one targeted genomic sequence.”. Claim 11 does not teach the any specifics of the limitation regarding the engineered transcriptional transactivator.
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 TALEN (an ETT) fused to the third component (a nuclease system that targets a genome) which is a Fok1 endonuclease [0142] wherein culturing the cell under conditions to induce the expression of the proteins encoded by the donor (e.g., paragraphs [0241] and [0325]; and 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].
Pompa properly teaches all the elements within the kit and the kit itself due to listing all the elements that would be required for the kit in one location. Therefore, Applicant’s arguments in regards to the rejection of claim 11 were not found to be persuasive.
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) as applied in view of Das et al (Biotechnology Journal, 11: 71-79; 2016). This rejection was made in the Office action mailed 02/17/2026.
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), 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), (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]). 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 and Ptet encoding DNA constructs (variants V10, V16 and M2) were transiently transfected into cells to determine which variant preformed 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 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) in view of Das et al (Biotechnology Journal, 11: 71-79; 2016), as applied to claims 1, 9 and 10 above, and in further view of Dechiara et al et al (WO 03020743 A1) as evidenced by Gossen (Science, vol. 268, no. 5218, 23 June 1995). This rejection was made in the Office action mailed 02/17/2026.
The 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 does 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), Dechiara teaches the rtTA protein contains the reverse TetR DBD and VP16 TA of claim 7. Gossen also teaches the example of transient transfection expressed by functional transactivator (Page 1768; 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 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 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 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), Dechiara teaches the rtTA protein contains the reverse TetR DBD and VP16 TA of claim 7. Gossen also teaches the example of transient transfection expressed by functional transactivator (Page 1768; 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 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 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), Dechiara teaches the rtTA protein contains the reverse TetR DBD and VP16 TA of claim 7. Gossen also teaches the example of transient transfection expressed by functional transactivator (Page 1768; 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 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.
Pompa and Das do not teach the TA domain as selected from the group consisting of VP16, VP64, VP128, VP160, VPR, p65, Rta, HSF1 and SAM.
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), Dechiara teaches the rtTA protein contains the reverse TetR DBD and VP16 TA of claim 7. Gossen also teaches the example of transient transfection expressed by functional transactivator (Page 1768; 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 CMV promoter as a minimal promoter 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, such as the rtTA protein contains the reverse TetR DBD and VP16 TA.
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.
Claim 6 is rejected under 35 U.S.C. 103 as being unpatentable over Popma (WO 2018/156818 A1) in view of Das et al (Biotechnology Journal, 11: 71-79; 2016) as applied to claims 1, 9 and 10 above, and in further view of Zhang et al (US 2015/0291966 A1). This rejection was made in the Office action mailed 02/17/2026.
The teachings of Popma and Das are referenced above as applied to claims 1, 9 and 10.
Regarding Claim 6, Popma and Das teaches the DNA binding domain where the transcriptional activator-like effector nuclease such as a TALE based DNA binding domain fused to an effector domain [Popma; 0142] and the ETT polypeptide comprising a DNA binding domain with a transcription activator (Das; Page 72, Column 2 and Page 73, Figure 1).
Popma and Das do not teach the TA domain being VP64.
Zhang teaches the use of a DNA binding domain such as a TALE DNA-binding domain fused to the synthetic VP64 transcriptional activator [0056]. Zhang teaches the system comprise a TALE fused to CRY2 and the cryptochrome binding partner CIBl fused to VP64, a transcription activor where in the inactive state, the TALE localizes its fused CRY2 domain to the promoter region of the gene of interest and at this point, CIBl is unable to bind CRY2, leaving the CIB1-VP64 unbound in the nuclear space; however, upon stimulation with 488 nm (blue) light, CRY2 undergoes a conformational change, revealing its CIB 1 binding site [0398]. Zhang teaches rapid binding of CIBl results in recruitment of the fused VP64 domain, which induces transcription of the target gene [0398].
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 DNA binding domain and TA domain taught by Zhang 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 Zhang teaches that the use of the TetR inducible system, comprising the TetR fused to VP64 transcriptional activator, is useful for being able to control gene expression with the presence or absence of tetracycline [0234].
One would have been motivated to make such a modification in order to receive the expected benefit of controlling the gene expression of the cassette taught by Zhang.
Claim 8 is rejected under 35 U.S.C. 103 as being unpatentable over Popma (WO 2018/156818 A1) in view of Das et al (Biotechnology Journal, 11: 71-79; 2016) as applied to claims 1, 9 and 10 above, and in further view of Baek et al (US 2019/0062373 A1). This rejection was made in the Office action mailed 02/17/2026.
The 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.
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 02/17/2026. 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 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 TALEN (an ETT) fused to the third component (a nuclease system that targets a genome) which is a Fok1 endonuclease [0142] wherein culturing the cell under conditions to induce the expression of the proteins encoded by the donor (e.g., paragraphs [0241] and [0325].
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.
However, the Applicant is only considering the use of the Das system on its own. When taken in combination, 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 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. 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.
The previous rejection of claims 2-5 and 7 under 35 U.S.C. 103 has been maintained in view of Applicant’s arguments filed on 02/17/2026. Applicant’s arguments have been fully considered but are not found to be persuasive.
Applicant argues Pompa and Das fails to teach a method in which a selector is stably inserted into a target locus of the genome of the target cell and the ETT is present in transient form, and the advantage associated therewith. Applicant continues to argue that Pompa and Das fails to teach i. the nucleotide sequence encoding the selector operably linked to a minimal promoter; ii. the ETT polypeptide expressed by the second component activates the minimal promoter; iii. 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; and iv. the TA domain as selected from the group consisting of VP16, VP64, VP128, VP160, VPR, p65, Rta, HSFI and SAM. Applicant argues that the disclosure in Dechiara does not remedy the deficiencies in Popma and Das. Dechiara does not teach a method in which the selector is stably inserted into a target locus of the genome of the target cell and the ETT is present in transient form, or the advantages associated therewith but rather, Deschiara reinforces the teachings of Das, which fails to suggest the invention presently claimed, specifically applicant argues that Dechiara does not teach that the integration of a rtTA variant into a target locus together with the selector can be immunogenic or detrimental to long term cell viability.
As previously discussed above, Pompa teaches the use of 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 TALEN (an ETT) fused to the third component (a nuclease system that targets a genome) which is a Fok1 endonuclease [0142] wherein culturing the cell under conditions to induce the expression of the proteins encoded by the donor (e.g., paragraphs [0241] and [0325]. When taken in combination, 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 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. 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.
Pompa and Das does 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), Dechiara teaches the rtTA protein contains the reverse TetR DBD and VP16 TA of claim 7. Gossen also teaches the example of transient transfection expressed by functional transactivator (Page 1768; 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 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 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.
The previous rejection of claim 6 under 35 U.S.C. 103 has been maintained in view of Applicant’s arguments filed on 02/17/2026. Applicant’s arguments have been fully considered but are not found to be persuasive.
Applicant argues that the rejection is improper for reasons previously discussed above. Applicant continues to argue that Pompa and Das fails to teach the TA domain being VP64 and Zhang allegedly teaches the TA domain being VP64, however Applicant argues that Zhang does not remedy the deficiencies of Pompa and Das, because Zhang does not teach a method in which the selector is stably inserted into a target locus of the genome of the target cell and the ETT is present in transient form, or the advantages associated therewith.
As previously stated above, Pompa teaches the use of 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 TALEN (an ETT) fused to the third component (a nuclease system that targets a genome) which is a Fok1 endonuclease [0142] wherein culturing the cell under conditions to induce the expression of the proteins encoded by the donor (e.g., paragraphs [0241] and [0325]. When taken in combination, 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 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. 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.
Popma and Das do not teach the TA domain being VP64.
Zhang teaches the use of a DNA binding domain such as a TALE DNA-binding domain fused to the synthetic VP64 transcriptional activator [0056]. Zhang teaches the system comprise a TALE fused to CRY2 and the cryptochrome binding partner CIBl fused to VP64, a transcription activor where in the inactive state, the TALE localizes its fused CRY2 domain to the promoter region of the gene of interest and at this point, CIBl is unable to bind CRY2, leaving the CIB1-VP64 unbound in the nuclear space; however, upon stimulation with 488 nm (blue) light, CRY2 undergoes a conformational change, revealing its CIB 1 binding site [0398]. Zhang teaches rapid binding of CIBl results in recruitment of the fused VP64 domain, which induces transcription of the target gene [0398].
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 DNA binding domain and TA domain taught by Zhang 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 Zhang teaches that the use of the TetR inducible system, comprising the TetR fused to VP64 transcriptional activator, is useful for being able to control gene expression with the presence or absence of tetracycline [0234].
One would have been motivated to make such a modification in order to receive the expected benefit of controlling the gene expression of the cassette taught by Zhang.
The previous rejection of claim 8 under 35 U.S.C. 103 has been maintained in view of Applicant’s arguments filed on 02/17/2026. Applicant’s arguments have been fully considered but are not found to be persuasive.
Applicant argues that the rejection is improper for reasons previously discussed above. Applicant continues to argue that Pompa and Das fails to teach the specific 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 and Baek allegedly teaches a guide RNA target sequence that is identical to SEQ ID NO: 26 of the present disclosure. Applicant continues to argue that Baek does not remedy the deficiencies of Pompa and Das because Baek does not teach a method in which the selector is stably inserted into a target locus of the genome of the target cell and the ETT is present in transient form, or the advantages associated therewith.
As stated previously above, Pompa teaches the use of 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 TALEN (an ETT) fused to the third component (a nuclease system that targets a genome) which is a Fok1 endonuclease [0142] wherein culturing the cell under conditions to induce the expression of the proteins encoded by the donor (e.g., paragraphs [0241] and [0325]. When taken in combination, 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 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. 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.
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.
Conclusion
No claims are allowed.
THIS ACTION IS MADE FINAL. Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
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/ALEXANDRA ROSE LIPPOLIS/Examiner, Art Unit 1637
/CELINE X QIAN/Primary Examiner, Art Unit 1637
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