DETAILED ACTION
Status of the Application
Claims 1-20 are pending.
The amendment filed on 08/01/2024 amending the specification is acknowledged.
The amendment filed on 03/12/2025 amending claims 4-6, 8, 13-15, 17, 19 and 20 is acknowledged.
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Priority
This application is a CON of PCT/US2023/061733 and claims priority to 63/305,657 filed on 02/01/2022.
Specification
The disclosure is objected to because of the following informalities: On page 4, the disclosure recites "The DAHP synthase may AroG*". Appropriate correction is required.
The description of FIG. 2 on page 5 of the specification is objected to in the recitation of “FIG.2 illustrates are steps”. Appropriate correction is required.
Claim Rejections - 35 USC § 112(a) Written Description
The following is a quotation of the first paragraph of 35 U.S.C. 112(a):
(a) IN GENERAL.—The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor or joint inventor of carrying out the invention.
The following is a quotation of the first paragraph of pre-AIA 35 U.S.C. 112:
The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor of carrying out his invention.
Claims 1-20 are rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, as failing to comply with the written description requirement. The claim(s) contains subject matter which was not described in the specification in such a way as to reasonably convey to one skilled in the relevant art that the inventor or a joint inventor, or for applications subject to pre-AIA 35 U.S.C. 112, the inventor(s), at the time the application was filed, had possession of the claimed invention.
As stated in MPEP 2111.01, during examination, the claims must be interpreted as
broadly as their terms reasonably allow. Claim 1 is directed in part to a method of producing hydroxytyrosol using a recombinant host microorganism that has been transformed with a decarboxylase-encoding polynucleotide and a second polynucleotide encoding a hydroxylase having at least 90 % identity to SEQ ID NO: 2, where the host is cultured in a medium comprising glucose and hydroxytyrosol is collected. Claims 2-9 depend directly or indirectly therefrom claim 1.
Claim 1 recites a method requiring a hydroxylase having at least 90 % identity to SEQ ID NO: 2, but the specification does not reasonably convey possession of the full breadth of hydroxylase variants encompassed by the claim. The disclosure appears to support, at most a specific exemplified sequence (SEQ ID NO: 2) and does not show possession of the claimed genus as a whole. The claim encompasses a large genus of proteins that can have up to 53 amino acid modifications within SEQ ID NO: 2 ((
53
=
0.1
×
527
); SEQ ID NO: 2 has 527 amino acids).
Claim 4 is directed in part to the method of claim 1, wherein the decarboxylase has a sequence with at least 95 % identity to SEQ ID NO: 4. However, the specification does not reasonably convey possession of the full breadth of decarboxylase variants encompassed by the claim. The disclosure appears to support, at most a specific exemplified sequence (SEQ ID NO: 4) and does not show possession of the claimed genus as a whole. The claim encompasses a large genus of proteins that can have up to 31 amino acid modifications within SEQ ID NO: 4 ((
31
=
0.05
×
604
); SEQ ID NO: 4 has 604 amino acids).
Claim 10 is directed in part to a recombinant host microorganism comprising a metabolic pathway for producing hydroxytyrosol, where the host has been transformed with a decarboxylase-encoding polynucleotide and a hydroxylase-encoding polynucleotide, and the hydroxylase has at least 90 % identity to SEQ ID NO:2. Claims 11-20 depend, directly or indirectly, therefrom claim 10.
Claim 10 recites a method requiring a hydroxylase having at least 90 % identity to SEQ ID NO: 2, but the specification does not reasonably convey possession of the full breadth of hydroxylase variants encompassed by the claim. The disclosure appears to support, at most a specific exemplified sequence (SEQ ID NO: 2) and does not show possession of the claimed genus as a whole.
The specification discloses the host organisms as non-limiting examples including bacteria of the genera Escherichia, Bacillus, Corynebacterium, and fungi of the genera Saccharomyces and Yarrowia. This by itself does not reasonably convey possession of the full scope of recombinant host microorganisms encompassed by the claim.
Claim 19 is directed in part to the recombinant host microorganism of claim 10, wherein the recombinant host microorganism is of a genus selected from Escherichia, Bacillus, Corynebacterium, Saccharomyces and Yarrowia.
The specification’s identification of these genera as non-limiting examples does not reasonably convey possession of recombinant host microorganisms across the full breadth of the claimed genus set. The specification appears to provide only E. coli as an actual example of a host microorganism (Example’s 2-3).
Claim Rejections - 35 USC § 112(a) Enablement
Claims 1-20 are rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, as failing to comply with the enablement requirement. The claim(s) contains subject matter which was not described in the specification in such a way as to enable one skilled in the art to which it pertains, or with which it is most nearly connected, to make and/or use the invention.
Factors to be considered in determining whether undue experimentation is required are
summarized in In re Wands (858 F.2d 731, 737, 8 USPQ2nd 1400 (Fed. Cir. 1988)) as follows:
1) quantity of experimentation necessary, 2) the amount of direction or guidance presented, 3)
the presence and absence of working examples, 4) the nature of the invention, 5) the state of
prior art, 6) the relative skill of those in the art, 7) the predictability or unpredictability of the art,
and 8) the breadth of the claims. The factors which have led the Examiner to conclude that the
specification fails to teach how to make and/or use the claimed invention without undue
experimentation, are addressed in detail below.
The breadth of the claims. Claim 1 broadly encompass a method of producing hydroxytyrosol by culturing a recombinant host microorganism in a glucose-containing medium and collecting hydroxytyrosol. Claims 2-9 further narrow the method by reciting sequence-identity and overexpression limitations. The specification does not enable the full scope of this method claim set without undue experimentation.
Claim 10 is directed to a broad class of recombinant hydroxytyrosol-producing microorganisms. Claims 11-20 further narrow the microorganism with related pathway features, including hosts of multiple genera, enzyme variants defined by sequence identity thresholds, and additional overexpression limitations. The claim group is broad because it reaches not only the exemplified E. coli embodiment, but also other microbial hosts and variant enzymes that may require additional optimization and testing to practice.
The amount of direction or guidance presented and the existence of working
examples. The specification provides limited direction or guidance for practicing the full scope of claims 1-9 and appears to include only a limited number of working examples. Although claim 1 is directed to a method of producing hydroxytyrosol using a recombinant host microorganism in a glucose-containing medium, the disclosure appears to exemplify only a narrow subset of enzyme constructs and host conditions. Claims 2-9 further narrow claim 1 by adding sequence-identity and overexpression limitations, however, the specification still does not enable the full scope of the claimed method set without undue experimentation Accordingly, the amount of direction provided and the limited working examples weigh against enablement because a person of ordinary skill in the art would need to engage in more than routine experimentation to practice the full scope of the claimed method set.
The specification provides limited direction and guidance for practicing the full scope of claims 10-20 and appears to include only limited examples. Although the disclosure may teach recombinant hydroxytyrosol production in E. coli using a glucose-containing medium, the claims extend to recombinant microorganisms across multiple genera and to enzyme variants defined by sequence identity thresholds and additional overexpression limitations. The specification does not appear to provide comparable guidance or working examples for practicing the invention across that broader scope, and a person of ordinary skill in the art would therefore need to engage in substantial additional experimentation to identify suitable hosts, sequence variants, and expression conditions. Accordingly, the amount of direction or guidance and the existence of working examples weigh against enablement for the full scope of claims 10-20.
The quantity of experimentation required to practice the claimed invention based
on the teachings of the specification. A skilled artisan would have to determine which decarboxylase and hydroxylase sequences retain activity across the claimed sequence identity ranges, select and optimize a suitable host microorganism and adjust expression and culture conditions to obtain hydroxytyrosol. The full scope of the claims would likely require substantial trial-and-error and iterative optimization rather than merely routine optimization.
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.
Claims 1-20 are rejected under 35 U.S.C. 103 as being unpatentable over Bisquert et al. (Microbial Biotechnology, published 10/12/2021), UniProt database entry D0LBD6_GORB4 (4-hydroxyphenylacetate 3-hydroxylase, Gordonia bronchialis, 2010), in view of UniProt database entry C4R7I0_KOMPG (Phenylpyruvate decarboxylase, catalyzes decarboxylation of phenylpyruvate to phenylacetaldehyde, Komagataella phaffi, 2016).
Bisquert et al. teaches a method of producing hydroxytyrosol by culturing a recombinant Saccharomyces cerevisiae transformed with HpaBC from Escherichia coli (4-hydroxyphenylacetate hydroxylase) which converts tyrosol to hydroxytyrosol in a medium comprising glucose (YPD or SC medium) through the shikimate and Ehrlich pathways (see Figure 1). Bisquert et al. teaches that phenylpyruvate decarboxylase ARO10 catalyzes the entrance into the Ehrlich pathway by the conversion of 4-hydroxyphenylpyruvate (4HPP) into 4-hydroxyphenyl acetaldehyde (4HPAA) (see Effect of individual ARO genes overexpression on tyrosol and HT production and Figure 1). Bisquert et al. teaches overexpression of ARO4, a DAHP synthase and a ARO7, a chorismite mutase that catalyzes the conversion of chorismite into prephenate (see Figure 1). The entire pathway is shown in Figure 1 below. Bisquert et al. teaches that yeast genes involved in obtaining tyrosol from tyrosine through the Ehrlich pathway have been heterologously expressed in E. coli to overproduce tyrosol and hydroxytyrosol (see Metabolic engineering of the shikimate and Ehrlich pathways to increase tyrosol and HT production).
PNG
media_image1.png
816
1085
media_image1.png
Greyscale
UniProt database entry D0LBD6_GORB4 is a 100% sequence match to SEQ ID NO: 2.
UniProt database entry C4R7I0_KOMPG is a 100% sequence match to SEQ ID NO: 4.
Claims 1-20 are directed in part to a method and recombinant host microorganism comprising a metabolic pathway producing hydroxytyrosol in a medium comprising glucose and a decarboxylase enzyme capable of decarboxylating 4-hydroxyphenylpyruvaic acid to produce 4-hydroxyphenylacetaldehyde and a hydroxylase that converts tryosol to hydroxytyrosol, wherein the host microorganism is transformed to overexpress a DAHP synthase, wherein the host microorganism is further transformed to overexpress an enzyme that has catalytic activity to convert chorismite to prephenate, wherein the recombinant microorganism is selected from Escherichia, Bacillus, Corynebacterium, Saccharomyces, and Yarrowia.
It would have been obvious to one of ordinary skill in the art before the effective filling date of the claimed invention to modify the method/microorganism of Bisquert et al. by substituting the 4-hydroxyphenylacetate hydroxylase and phenylpyruvate decarboxylase with another 4-hydroxyphenylacetate hydroxylase and phenylpyruvate decarboxylase known in the prior art.
A person of ordinary skill in the art is motivated to substitute the 4-hydroxyphenylacetate hydroxylase and phenylpyruvate decarboxylase of the claimed invention for the 4-hydroxyphenylacetate hydroxylase and phenylpyruvate decarboxylase of Bisquert et al. because the art teaches the enzymes have the same desired biochemical function that would predictably achieve the same end product (hydroxytyrosol), making the substitution a routine and predictable design choice.
One of ordinary skill in the art has a reasonable expectation of success because the proposed substitution uses known enzymes with known catalytic roles to perform the same transformation under the same or similar conditions. Where the prior art already demonstrates that the target product can be obtained by related enzymes acting in the same pathway, there is no indication that the substitutions would produce unexpected results or require undue experimentation. Therefore, the invention as a whole would have been prima facie obvious to a
person of ordinary skill in the art before the effective filing date of the claimed invention.
Conclusion
No claims at this time are allowed.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to DALTON KIEFER, PhD whose telephone number is (571)272-1235. The examiner can normally be reached M-F 7:30-5 EST.
Examiner interviews are available via telephone, in-person, and video conferencing using a USPTO supplied web-based collaboration tool. To schedule an interview, applicant is encouraged to use the USPTO Automated Interview Request (AIR) at http://www.uspto.gov/interviewpractice.
If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Robert Mondesi can be reached at 408 918-7584. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
Information regarding the status of published or unpublished applications may be obtained from Patent Center. Unpublished application information in Patent Center is available to registered users. To file and manage patent submissions in Patent Center, visit: https://patentcenter.uspto.gov. Visit https://www.uspto.gov/patents/apply/patent-center for more information about Patent Center and https://www.uspto.gov/patents/docx for information about filing in DOCX format. For additional questions, contact the Electronic Business Center (EBC) at 866-217-9197 (toll-free). If you would like assistance from a USPTO Customer Service Representative, call 800-786-9199 (IN USA OR CANADA) or 571-272-1000.
/DALTON EDWARD KIEFER/Examiner, Art Unit 1652
/ROBERT B MONDESI/Supervisory Patent Examiner, Art Unit 1652
Prosecution Insights