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 .
DETAILED ACTION
This action is in response to the papers filed on October 20, 2025. Pursuant to amendment filed November 08, 2022, claims 1-16 are amended. Claims 17-18 are newly added. Claims 1, 7, and 13 are independent claims.
Election/Restrictions
Applicant's election with traverse of Group II, drawn to a eukaryotic host cell, method of making the cell, and a vector system, in the reply filed on October 20, 2025 is acknowledged. The traversal is on the grounds that the cited references do not disclose a PAH lacking a functional N-terminal domain. This is not found persuasive because the ordinary artisan would have recognized the prior art teaches PAH variants lacking the N-terminal regulatory domain and their established use in engineered expression systems.
Specifically, Luo teaches that amino acid hydroxylases, including PheH/PAH enzymes, may be employed as functionally active variants or fragments and that N-terminal truncations removing residues not necessary for catalytic activity can be used to improve expression and stability (Example 5, pg. 50, lines 10-27). Likewise, Daubner teaches a phenylalanine hydroxylase lacking the N-terminal regulatory domain, specifically a catalytic-domain PAH construct comprising PAH Δ 116, human PAH lacking the N-terminal 116 amino acid residues, Δ1-116, demonstrating that removal of the N-terminal regulatory region yields an enzymatically active PAH lacking regulatory the known regulatory constraints, explicitly describing a truncated PAH lacking the regulatory domain (first 116 amino acids) (pg. 296, column 2, para. 1-5; Section: Results, pg. 298-299). This explicit disclosure of the lack of a functional N-terminal regulatory domain in the PAH due to a deletion to form a truncated PAH is consistent with the instant specification’s definition of the PAH lacking a functional N-terminal domain (See instant specification pg. 3, lines 7-10, pg. 19, para. 1 through pg. 20, para. 1).
Additionally, as already communicated, Ding teaches implementing a complete phenylalanine hydroxylating system by co-expressing phenylalanine hydroxylase (PAH or PheH) together with the BH4-biosyntnetic enzymes guanosine triphosphate cyclohydrase I (GTPCH), or GTP cyclohydrase (GCH1), and PTPS in a multicistronic construct (Abstract, Fig. 1 and 4, pg. 674, column 1, para. 1 through column 2, para. 2.; pg. 677, column 2.). Hiller teaches selecting eukaryotic host cells in tyrosine-free medium based on outgrowth of cells expressing exogenous PAH- pathway enzymes, or tyrosine prototrophy selection pressure (Abstract; [0054-0062]; [0076-0-83]; Examples 1-3).
Therefore, the ordinary artisan would have found it obvious, in view of the teachings of Ding and Hiller, and further in view of Luo and Daubner, to employ a PAH lacking its N-terminal regulatory domain in combination with GCH1 for use in a eukaryotic tyrosine-selection system, with a reasonable expectation of success. The ordinary artisan would have been motivated to do so because the prior art teaches that BH4 availability limits PAH activity and that removal or modification of the N-terminal regulatory domain is a known approach to obtaining catalytically competent PAH variants suitable for engineered expression and selection under tyrosine-limiting conditions. Hence, before the effective filing date, the ordinary artisan would have recognized the N-terminal regulatory domain of PAH limits enzyme activity and introduces activation lag, and removing it yields a constitutively active catalytic enzyme where expression under a control sequence provides predictable, robust, phenylalanine to tyrosine conversion in the host cell, making it better suited for reliable metabolic selection and engineered expression systems.
The requirement is still deemed proper and is therefore made FINAL.
Claims 1-6 and 17 are withdrawn from further consideration pursuant to 37 CFR 1.142(b), as being drawn to a nonelected groups of inventions, there being no allowable generic or linking claim. Applicant timely traversed the restriction (election) requirement in the reply filed on October 20, 2025. The restriction requirement between Groups I-II is still deemed proper and made FINAL.
It is noted that when a final requirement for restriction is made by the examiner, applicant may file a petition under 37 CFR 1.144 for review of the restriction requirement. The propriety of a requirement to restrict, if traversed, is reviewable by petition under 37 CFR 1.144. In re Hengehold, 440 F.2d 1395, 169 USPQ 473 (CCPA 1971). Petition may be deferred until after final action on or allowance of claims to the invention elected, but must be filed not later than appeal. A petition will not be considered if reconsideration of the requirement was not requested (See § 1.181.).
Therefore, claims 7-16 and 18 are currently under examination to which the following grounds of rejection are applicable.
Priority
The present application, filed on May 13, 2022, is a 35 U.S.C. 371 national stage filing of the International Application No. PCT/EP2020/081923, filed November 12, 2020. Acknowledgment is made of applicant’s claim for foreign priority under 35 U.S.C. 119 (a)-(d). The certified copy has been filed in parent European Patent Application EP19209232.8, filed November 14, 2019, and EP20185798.4, filed July 14, 2020. Receipt is acknowledged of priority documents EP19209232.8 and EP20185798.4 required by 37 CFR 1.55.
Thus, the earliest possible priority for the instant application is November 14, 2019.
Information Disclosure Statement
The information disclosure statements (IDS) submitted on 05/13/2022, 11/08/2022, 03/04/2024 are acknowledged. The submission is in compliance with the provisions of 37 CFR 1.97. Accordingly, the information disclosure statements are being considered by the examiner.
Claim Interpretation
The claims are directed to a eukaryotic cell comprising “exogenous” nucleic acids. Under the broadest reasonable interpretation, consistent with the specification, the term “exogenous” refers to any nucleic acid or material that is introduced into the cell from outside the cell or produced inside the cell prior to introduction, regardless of whether the sequence itself is naturally occurring or identical to an endogenous sequence. As expressly defined in the Specification, an “exogenous nucleic acid” includes nucleic acids that are not naturally produced in the cell, as well as nucleic acids whose sequences may also be found endogenously in the same organism or cell, provided they are introduced into the cell or are under non-native regulatory control (see instant specification, pg. 8, para. 1). For instance, under the broadest reasonable interpretation, claim 11 recites “exogenous nucleic acids” and encompasses introduced copies of PAH, GCH1, and the product-encoding sequence regardless of whether identical or homologous sequences are endogenously present in the host cell, including where such nucleic acids are integrated into the genome or operably linked to non-native regulatory elements. Therefore, it is not limited to non-naturally occurring or non-endogenous gene sequences.
Under this definition, a nucleic acid encoding an enzyme which is also endogenously encoded in the host cell encompasses an “exogenous” nucleic acid when introduced by techniques such as transformation, transfection, or integration, such as where the introduced nucleic acid is operably linked to a heterologous or constitutively active promoter. Thus, the ordinary artisan would reasonably understand “exogenous” to encompass introduced copies of native genes, modified variants thereof, truncated forms, or identical coding sequences placed under different regulatory elements, and would not interpret the term as limited to non-naturally occurring sequences or sequences absent from the host organism.
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.
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 7-16 and 18 are rejected under 35 U.S.C. 103 as being unpatentable over Ding et al. (Mol Ther. 2008 Apr;l6(4):673-81. Epub 2008 Mar 11., see IDS), in view of Hiller et al. (US 2020/0056190 A1, filed March 16, 2017; prior published September 20, 2018, WO 2018/167621, see IDS), and further in view of Daubner et al. (Arch Biochem Biophys. 1997 Dec 15;348(2):295-302.) and Luo et al. (WO 2017/167866 A1, see IDS).
Regarding claims 7-9 and 18, Ding teaches a eukaryotic host cell comprising exogenous nucleic acids encoding phenylalanine hydroxylase (PAH), GTP cyclohydrolase I (GCH1), along with third and 6-pyruvoyltetrahydrobiopterin synthase (PTPS) introduced via vector systems to enable phenylalanine hydroxylation through coordinated expression of PAH and BH4-biosyntnetic enzymes, under the independent control sequences, the CMV promoter (Abstract; pg. 674, column 2, para. 1; Table 1-2; Fig. 3-4).
Ding does not explicitly teach the lack of a functional N-terminal regulatory domain or specifically teach where the eukaryotic cell is a Chinese hamster ovary (CHO) cell.
However, before the effective filing date, Hiller teaches the use of PAH-based systems in eukaryotic host cells, including CHO cells, as a tyrosine prototrophy selection marker system (Abstract; [0019]). Hiller teaches the PAH-based systems can be used in eukaryotic host cells, including CHO cells, to confer tyrosine prototrophy and enable growth in tyrosine-deficient media. Hiller explicitly states:
“provided herein is a host cell containing one or more nucleic acid constructs provided herein containing the PAH and PCBD1 genes, wherein the host cell further comprises an exogenous copy of one or more genes selected from the group consisting of… GCH1 ([0163]).
Hiller further teaches that host cells may be genetically modified to alter expression of endogenous PAH, including by mutation or deletion, to enhance reliance on introduced PAH pathway components, and the selection or evaluation based on their ability to proliferate under tyrosine-limiting conditions and control elements, such as enhancers or promoters ([0031-0033]; [0107-0108]). Hence, the teachings of Hiller provide the selection context and host-cell application for PAH/GCH1 constructs, as taught by Ding. The combination of Ding and Hiller establish the host-cell context, selection pressure, and practical application of PAH-centered systems in CHO cells, but do not specifically teach the deletion of the PAH N-terminal domain.
However, the ordinary artisan would have recognized that, while Ding and Hiller teach the use of PAH and GCH1 in eukaryotic host cells and in tyrosine-based selection contexts, the specific form of PAH employed could be modified to optimize function in the engineered systems, further in view of the teachings of Daubner and Luo.
Daubner teaches that phenylamine hydroxylase lacking the N-terminal regulatory domain remains catalytically active and shows no lag (Abstract; pg. 295, column 1, para. 2; pg. 297-298, bridging para.), and Luo teaches that aromatic amino acid hydroxylases may be used as variants or fragments, including N-terminally modified forms, in engineered expression systems (pg. 50, Example 5). Luo also teaches nucleic acids and vectors encoding the corresponding nucleic acids, such as the phenylalanine hydroxylase and GCH1. Luo teaches GCH1 variants of E.coli and provides cells comprising such variants together with a pterin-4a-carbinolamine dehydratase (PCD) and at least one of a tryptophan hydroxylase (TPH), a tyrosine hydroxylase (TH) and a phenylalanine hydroxylase, or PheH. The variant provides for an increased hydroxylation activity of at least one of the TPH, TH and PheH as compared to native E. coli GCH1, used to overcome tyrosine auxotrophy (claim 1; p. 28, lines 21-34).
Before the effective filing date of the instant application, it would have been obvious to the ordinary artisan to employ an N-terminal regulatory domain deficient PAH, as taught by Daubner and Luo, in the PAH/GCH1 host-cell and selection systems taught by Ding and Hiller, with a reasonable expectation of success. The ordinary artisan would have been motivated to do so because Ding and Hiller teach PAH/GCH1 based host-cell and selection systems, while Daubner and Luo teach that N-terminal modification or removal of the PAH regulatory domain is an effective approach to obtaining catalytically competent PAH variants suitable for engineered expression.
Regarding claim 10, the combined teachings of Ding, Hiller, Daubner, and Luo renders claim 7 obvious. Additionally, Hiller teaches the utility of mammalian host cells ([0114]).
Regarding claims 11, the combined teachings of Ding, Hiller, Daubner, and Luo renders claim 7 obvious. Ding additionally teaches the stable introduction of the expression cassettes into the mammalian cells using viral vectors (pg. 678, column 1-2, bridging para.), and Hiller teaches the generation and selection of stable eukaryotic host cells with stable genomic incorporation ([0124]).
Regarding claims 12, the combined teachings of Ding, Hiller, Daubner, and Luo renders claim 7 obvious. Hiller additional explicitly teaches genetically modifying host cells by mutating or deleting endogenous PAH genes or modifying their regulatory sequences to reduce endogenous pathway activity and thereby enhance reliance on introduced PAH-based systems ([0033-0034]; [0107-0108]). As already communicated, Daubner teaches that PAH lacking the N-terminal regulatory domain remains catalytically active, and Luo teaches that a aromatic amino acid hydroxylases may be employed as variants or fragments, including N-terminally modified forms, in engineered systems. Thus, the ordinary artisan would have found it obvious to reduce or abolish endogenous PAH activity in the host cells of Ding and Hiller while employing an N-terminal regulatory domain deficient PAH, as taught by Daubner and Luo, with a reasonable expectation of success of generating the selection marker-based system.
Regarding claims 13-14, the claims further recite a vector system and conventional vector generation elements, such as cloning site/multiple cloning site insertion. The disclosures of vector-based systems by Ding and Hiller have already been communicated, and these are conventionally utilized in the art. For instance, see Hiller’s teachings on the utility of such vectors (claim 9; [0268]; [0272]). Additionally, Hiller plainly states,
“a nucleotide sequence of interest may alternatively, for example, be a sequence which provides a regulatory or structural function (e.g. a promoter or enhancer sequence), or which serves a different purpose, such as a restriction enzyme sequence for cloning purposes (e.g. a nucleotide sequence of interest may be a multiple cloning site” ([0072]).
Hence, before the effective filing date of the instant application, the ordinary artisan would have found it obvious to implement vector based PAH/GCH1 systems of Ding and Hiller comprising conventional vector-generation elements, including cloning sites or multiple cloning sites for insertion of a nucleic acid of interest, as taught by Hiller, with a reasonable expectation of success.
Regarding claims 15-16, the combined teachings of Ding, Hiller, Daubner, and Luo renders claim 7 obvious. Furthermore, Hiller expressly teaches expression, recovery, and purification of the product ([0138-0140]).
Conclusion
Claims 7-16 and 18 are rejected. No claims are allowed.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to JOEL D LEVIN whose telephone number is (571)270-0616. The examiner be reached 8:00 am to 5:00 pm, Monday through Friday.
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If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Christopher Babic can be reached at (571) 272-8507. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
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/J.D.L./Examiner, Art Unit 1633
/CHRISTOPHER M BABIC/Supervisory Patent Examiner, Art Unit 1633