DETAILED CORRESPONDENCE
Status of the Application
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Claims 1-14 are pending in this application.
Applicant’s amendment to the claims filed 06/23/2026 is acknowledged. This listing of the claims replaces all prior versions and listings of the claims.
Applicant’s remarks filed on 06/23/2026 in response to the non-final rejection mailed on 03/27/2026 are acknowledged and have been fully considered.
The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action.
Election
The elected invention is
Group I, corresponding to claims 1-8 and new claim 14, drawn to the technical feature of a genetically modified organism in which a nucleic acid encoding any one of the polypeptides described in (a) to (c) below is introduced or the expression of the polypeptide is enhanced and the function of pyruvate kinase is impaired: (a) a polypeptide composed of an amino acid sequence represented by any one of SEQ ID NOs: 1 to 7; (b) a polypeptide composed of the same amino acid sequence as that represented by any one of SEQ ID NOs: 1 to 7, except that one or several amino acids are substituted, deleted, inserted, and/or added, and having an enzymatic activity that catalyzes a reaction to reduce 3- oxoadipyl-CoA to 3-hydroxyadipyl-CoA; (c) a polypeptide composed of an amino acid sequence with a sequence identity of not less than 70% to the sequence represented by any one of SEQ ID NOs: 1 to 7 and having an enzymatic activity that catalyzes a reaction to reduce 3-oxoadipyl-CoA to 3-hydroxyadipyl-CoA, and
Species A2) the polypeptide represented by SEQ ID NO: 2, and
relative to SEQ ID NO: 173 that the 13th amino acid is phenylalanine, the 15th amino acid is glutamine, the 16th amino acid is lysine, the 17th amino acid is glycine, the 19th amino acid is proline, and the 21st amino acid is methionine,
elected in the reply filed 05/08/2024.
New claim 14 is drawn to a genetically modified microorganism wherein a nucleic acid encoding any one of the polypeptides described in (a) to (b) below is introduced by a genetic modification or the expression of the polypeptide is enhanced by a genetic modification: (a) a polypeptide composed of the amino acid sequence of any one of SEQ ID NOs: 1 to 7; (b) a polypeptide composed of the amino acid sequence with a sequence identity of not less than 90% to the sequence of any one of SEQ ID NOs: 1 to 7 and having an enzymatic activity that catalyzes a reaction to reduce 3-oxoadipyl-CoA to 3-hydroxyadipyl-CoA; wherein a function of PykF and/or PykA of the microorganism is impaired by a genetic modification, wherein a production of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid is increased by said genetic modifications. Therefore new claim 14 is drawn to the invention of Group I above.
Claims 9-13 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim. Election was made with traverse in the reply filed on 05/08/2024.
Claims 1-8 and 14 are being examined on the merits only to the extent they read on the elected subject matter.
In view of the elected species
“relative to SEQ ID NO: 173 that the 13th amino acid is phenylalanine, the 15th amino acid is glutamine, the 16th amino acid is lysine, the 17th amino acid is glycine, the 19th amino acid is proline, and the 21st amino acid is methionine”
as recited by claim 3 being free of the prior art of record, the search and examination has been extended to the following species
“relative to SEQ ID NO: 173 that the 13th amino acid is phenylalanine, the 15th amino acid is glutamine, the 16th amino acid is lysine, the 17th amino acid is glycine, the 19th amino acid is arginine, and the 21st amino acid is valine”.
Claim Rejections - 35 USC § 112(b)
The rejection of claim 8 under 35 U.S.C. 112(b) as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor regards as the invention is withdrawn in view of the amendment to limit the PTS system as impaired compared to an otherwise identical microorganism without any of the genetic mutations.
Claims 1-8 and 14 are rejected under 35 U.S.C. 112(b) as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor regards as the invention.
The instant rejection was previously applied to claims 1-8, any newly recited portion is necessitated by claim amendment.
Claim 1 (claims 2-8 dependent therefrom) and 14 are indefinite for the recitation of “the expression of the polypeptide is enhanced by a genetic modification” and “a function of PykF and/or PykA … is impaired by a genetic modification” without a recitation of a standard or reference with which to compare said expression or function.
Claim 14 is indefinite for the recitation of “the expression of the polypeptide is enhanced by a genetic modification”, “a function of PykF and/or PykA … is impaired by a genetic modification”, “wherein a production … is increased by said genetic modifications” without a recitation of a standard or reference with which to compare said expression or function.
Response to remarks: beginning on page 5 of Applicant’s response to rejections under 35 USC 112(b); Applicant in summary contends the amendments to the claims have obviated the rejection.
Applicant’s remarks are considered and found not convincing, as the recited enhancements caused by genetic modification in claim 1 are still recited without a reference for comparison, as well as those recited by new claim 14.
Claim Rejections - 35 USC § 112(a)
Claim Interpretation: Claim 1 as amended is drawn to a genetically modified microorganism wherein a nucleic acid encoding any one of the polypeptides described in (a) to (b) below is introduced by a genetic modification or the expression of the polypeptide is enhanced by a genetic modification: (a) a polypeptide composed of the amino acid sequence of any one of SEQ ID NOs: 1-7; (b) a polypeptide composed of the amino acid sequence with a sequence identity of not less than 90% to the sequence of any one of SEQ ID NOs: 1-7 and having enzymatic activity that catalyzes a reaction to reduce 3-oxoadipyl-CoA to 3-hydroxyadipyl-CoA; wherein a function of PykA and/or PykF of the microorganism is impaired by a genetic modification.
The claims are therefore drawn to a genus of genetic modifications to a cell that is considered to be widely variant that result in the impairment of PykA and/or PykF, and the enhancement of expression of the polypeptides of (a) or (b). Given a broadest reasonable interpretation, the genetic cell modifications are considered to be unlimited, as the modifications are not limited to be carried out on the enzymes recited in the claim.
Claim 14 is drawn to a genetically modified microorganism wherein a nucleic acid encoding any one of the polypeptides described in (a) to (b) below is introduced by a genetic modification or the expression of the polypeptide is enhanced by a genetic modification: (a) a polypeptide composed of the amino acid sequence of any one of SEQ ID NOs: 1 to 7; (b) a polypeptide composed of the amino acid sequence with a sequence identity of not less than 90% to the sequence of any one of SEQ ID NOs: 1 to 7 and having an enzymatic activity that catalyzes a reaction to reduce 3-oxoadipyl-CoA to 3-hydroxyadipyl-CoA; wherein a function of PykF and/or PykA of the microorganism is impaired by a genetic modification, wherein a production of 3-hydroxyadipic acid, a-hydromuconic acid or adipic acid is increased by said genetic modifications.
The claims are therefore drawn to a genus of genetic modifications to a cell that is considered to be widely variant that result in the impairment of PykA and/or PykF, and the enhancement of expression of the polypeptides of (a) or (b), and the enhancement of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid production. Given a broadest reasonable interpretation, the genetic cell modifications are considered to be unlimited, as the modifications are not limited to be carried out on the enzymes recited in the claim.
A. Claims 1-8 and 14 are rejected under 35 U.S.C. 112(a) 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 at the time the application was filed, had possession of the claimed invention.
The instant rejection is maintained from a previous Office action, and any newly recited portion is necessitated by claim amendment.
MPEP 2163.II.A.2.(a).i) states, “Whether the specification shows that applicant was in possession of the claimed invention is not a single, simple determination, but rather is a factual determination reached by considering a number of factors. Factors to be considered in determining whether there is sufficient evidence of possession include the level of skill and knowledge in the art, partial structure, physical and/or chemical properties, functional characteristics alone or coupled with a known or disclosed correlation between structure and function, and the method of making the claimed invention”.
For claims drawn to a genus, MPEP § 2163 states the written description requirement for a claimed genus may be satisfied through sufficient description of a representative number of species by actual reduction to practice, reduction to drawings, or by disclosure of relevant, identifying characteristics, i.e., structure or other physical and/or chemical properties, by functional characteristics coupled with a known or disclosed correlation between function and structure, or by a combination of such identifying characteristics, sufficient to show the applicant was in possession of the claimed genus. See Eli Lilly, 119 F.3d at 1568, 43 USPQ2d at 1406.
According to MPEP 2163.II.A.3.(a).ii), [s]atisfactory disclosure of a ‘representative number’ depends on whether one of skill in the art would recognize that the applicant was in possession of the necessary common attributes or features possessed by the members of the genus in view of the species disclosed. For inventions in an unpredictable art, adequate written description of a genus which embraces widely variant species cannot be achieved by disclosing only one species within the genus…Instead, the disclosure must adequately reflect the structural diversity of the claimed genus, either through the disclosure of sufficient species that are ‘representative of the full variety or scope of the genus,’ or by the establishment of ‘a reasonable structure-function correlation.’"
The factors considered in the Written Description requirement are (1) level of skill and knowledge in the art, (2) partial structure, (3) physical and/or chemical properties, (4) functional characteristics alone or coupled with a known or disclosed correlation between structure and function, and the (5) method of making the claimed invention. Disclosure of any combination of such identifying characteristics that distinguish the claimed invention from other materials and would lead one of skill in the art to the conclusion that the applicant was in possession of the claimed species is sufficient." MPEP § 2163.
The claims recite (in relevant part) a genus of genetic modifications that result in the impairment of PykA and/or PykF functions, the enhancement of expression of one of two polypeptides, and the enhancement of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid production. As stated above, the genetic modifications are considered to be unlimited as the genetic modifications are not limited to be carried out on the enzymes or polypeptides recited in the claim. In this case, the genus of genetic modifications encompasses species that are considered to be widely variant.
The specification discloses the following representative species of modifications: inactivating genes encoding pyruvate kinase (pykF and pykA) and overexpressing genes encoding the polypeptides of any one of SEQ ID NOs: 1-7. Aside from these representative species of genetic modifications, there are no other examples of genetic modifications or conditional modifications in the specification disclosed to result in the claimed activity.
Regarding the level of skill and knowledge in the art of amino acid modification, the reference of Singh et al. (Curr. Protein Pept. Sci. 18:1-11, 2017; cited on the Form PTO-892 mailed 06/11/2024) reviews various protein engineering methods and discloses that despite the availability of an ever-growing database of protein structures and highly sophisticated computational algorithms, protein engineering is still limited by the incomplete understanding of protein functions, folding, flexibility, and conformational changes [see p. 7, column 1, top]. Also, the unpredictability associated with residue substitution is exemplified by the reference of Zhang et al. (Structure 26:1474-1485, 2018; cited on the Form PTO-892 mailed 06/11/2024), which discloses that even a substitution of a surface residue that was predicted to be benign caused significant structural changes and unexpected effects on the function of a polypeptide [p. 1475, column 1].
Further regarding the genus of genetic modifications, it is noted that the field of metabolic engineering can be inherently unpredictable and advances in metabolic pathway engineering are often achieved only by empirical experimentation. According to Guo et al. (Comp Struct Biotechnol J, 2017, 15:161; cited on the attached Form PTO-892) “First, a lot of organisms are difficult to be engineered because of unknown regulation patterns and the lack of engineering tools for non-model organisms [16]. Even for model microorganisms like Escherichia coli and Saccharomyces cerevisiae, which are well studied and equipped with a broad spectrum of biomolecular tools to allow metabolic engineering easily, the effects of heterologous expression of pathways are often unpredictable to guarantee a high productivity…Second, a key challenge in metabolic engineering is balancing the tug-of-war that exists between the cell's physiological and evolutionary objectives on one side and the engineer's process objectives on the other [20]. Such conflict of resource allocation sometimes cannot be well addressed and toxic intermediates could be built up in the unbalanced pathway” [p. 162, column 1, middle].
In view of the high level of unpredictability in the art regarding structure and function of polypeptides, and the high level of unpredictability in the art regarding metabolic engineering, because the genus of genetic modifications is widely variant, and the specification discloses the actual reduction to practice of only 7 representative species among a widely variant genus, one of skill in the art would reasonably conclude that the disclosure fails to provide a representative number of species to describe the genus, and thus, that the applicant was not in possession of the recited genus of genetic modifications. The claimed subject matter is not supported by an adequate written description because a representative number of species has not been described.
B. Claims 1-8 and 14 are rejected under 35 U.S.C. 112(a) because the specification, while being enabling for a genetically modified microorganism modified to express the polypeptide of any of SEQ ID NOs: 1-7 and modified to disrupt the pykA and pykF genes to impair the function of PykA and/or PykF, does not reasonably provide enablement for all genetically modified microorganisms as encompassed by the claims. The specification does not enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and/or use the invention commensurate in scope with these claims.
The instant rejection is maintained from a previous Office action, and any newly recited portion is necessitated by claim amendment.
“The test of enablement is not whether any experimentation is necessary, but whether, if experimentation is necessary, it is undue.” In re Angstadt, 537 F.2d 498, 504, 190 USPQ 214, 219 (CCPA 1976). Factors to be considered in determining whether undue experimentation is required are summarized in In re Wands (858 F.2d 731, 737, 8 USPQ2d 1400, 1404 (Fed. Cir. 1988)) as follows: (A) The breadth of the claims; (B) The nature of the invention; (C) The state of the prior art; (D) The level of one of ordinary skill; (E) The level of predictability in the art; (F) The amount of direction provided by the inventor; (G) The existence of working examples; and (H) The quantity of experimentation needed to make or use the invention based on the content of the disclosure. See MPEP § 2164.01(a). The Factors considered to be most relevant to the instant rejection are addressed in detail below.
The nature of the invention: According to the specification at para 0012, “an object of the present invention is to provide a genetically modified microorganism for producing 3-hydroxyadipic acid, alpha-hydromuconic acid, and/or adipic acid in high yield.” The object of the invention is therefore to provide a mechanism for high yield production of the disclosed metabolites through genetic modification of the organism.
The breadth of the claims: The claims recite (in relevant part) genetic modification(s) that result in the impairment of pyruvate kinase, enhancement of expression of the polypeptides (a) or (b), and enhancement of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid production. As stated above, with the exception of inactivating genes encoding pyruvate kinase and expressing any of the polypeptides of SEQ ID NOs: 1-7, the remaining genetic modifications are considered to be unlimited and are not limited to be carried out on the enzymes or polypeptides in the claim.
The state of the prior art; The level of one of ordinary skill; and The level of predictability in the art: According to MPEP 2164.03, “…what is known in the art provides evidence as to the question of predictability” and “[I]f one skilled in the art cannot readily anticipate the effect of a change within the subject matter to which that claimed invention pertains, then there is lack of predictability in the art.”
As noted above, other than the recited cell comprising the amino acid sequences set forth by SEQ ID NOs: 1-7, the modifications are unlimited. The reference of Singh (supra) reviews various protein engineering methods and discloses that despite the availability of an ever-growing database of protein structures and highly sophisticated computational algorithms, protein engineering is still limited by the incomplete understanding of protein functions, folding, flexibility, and conformational changes [see p. 7, column 1, top].
The unpredictability associated with amino acid modification is exemplified by the reference of Zhang (supra) which discloses that even a mutation that was predicted to be benign caused significant structural changes and unexpected effects on the function of a polypeptide [p. 1475, column 1].
The unpredictability associated with metabolic engineering is exemplified by the reference of Guo (supra) which discloses “First, a lot of organisms are difficult to be engineered because of unknown regulation patterns and the lack of engineering tools for non-model organisms [16]. Even for model microorganisms like Escherichia coli and Saccharomyces cerevisiae, which are well studied and equipped with a broad spectrum of biomolecular tools to allow metabolic engineering easily, the effects of heterologous expression of pathways are often unpredictable to guarantee a high productivity… Second, a key challenge in metabolic engineering is balancing the tug-of-war that exists between the cell's physiological and evolutionary objectives on one side and the engineer's process objectives on the other [20]. Such conflict of resource allocation sometimes cannot be well addressed and toxic intermediates could be built up in the unbalanced pathway” [p. 162, column 1, middle].
As such, one of skill in the art would recognize a high level of unpredictability that all genetic modifications as encompassed by the claims would maintain the desired activity of impairing pyruvate kinase, enhancing the expression of the polypeptides (a) and (b), and enhancing 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid production.
The amount of direction provided by the inventor and The existence of working examples: The specification discloses the following working examples of the recited modification: inactivating genes encoding pyruvate kinase (pykF and pykA) and overexpressing genes encoding the polypeptides of any one of SEQ ID NOs: 1-7. Other than these working examples, the specification fails to disclose any other modifications that would result in the desired impairment of the functions of pyruvate kinase, enhancement of expression of the polypeptides (a) or (b), or enhancement of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid production.
Also, the specification fails to provide guidance for using any cells bearing genetic modifications that are non-functional.
The quantity of experimentation needed to make or use the invention based on the content of the disclosure: While methods of modifying the amino acid sequence of a polypeptide and genetically modifying cells were known at the time of the invention, it was not routine in the art to make and determine a use for genetic modifications recited by the claims.
In view of the overly broad scope of the claims, the lack of guidance and working examples provided in the specification, the high level of unpredictability, and the state of the prior art, undue experimentation would be necessary for a skilled artisan to make and use the entire scope of the claimed invention. Applicants have not provided sufficient guidance to enable one of ordinary skill in the art to make and use the claimed invention in a manner reasonably correlated with the scope of the claims. The scope of the claims must bear a reasonable correlation with the scope of enablement (In re Fisher, 166 USPQ 19 24 (CCPA 1970)). Without sufficient guidance, determination of having the desired biological characteristics is unpredictable and the experimentation left to those skilled in the art is unnecessarily, and improperly, extensive and undue. See In re Wands 858 F.2d 731, 8 USPQ2nd 1400 (Fed. Cir, 1988).
Response to Remarks: beginning on page 5 of Applicant’s response to 112(a) rejections; Applicant in summary contends the amendment to recite “a function of PykF and/or PykA of the microorganism is impaired by a genetic modification” in claims 1 and 14 limit the genus of genetic modifications by identifying the pyruvate kinase function being impaired; Applicant further contends the amended claims are enabled as they recite the impairment of PykA and/or PykF, and the scope of genetic modifications is not unlimited.
Applicant’s remarks are considered and found not convincing. The amendments to recite “a function of PykF and/or PykA of the microorganism is impaired by a genetic modification” do not alter the unlimited genus of genetic modifications. Not only are the genetic modifications recited in the rejection above unlimited in terms of how a gene is being modified, but the recited genetic modifications are not required to be carried out on the genes recited in the claim. As such the recited genus of genetic modifications is considered to be widely variant, and one of skill in the art would reasonably conclude that the applicant was not in possession of the recited genus of genetic modifications.
Regarding the enablement rejection, as stated above: the specification, while being enabling for a genetically modified microorganism modified to express the polypeptide of any of SEQ ID NOs: 1-7 and modified to disrupt the pykA and pykF genes to impair the function of PykF and/or PykA, does not reasonably provide enablement for all genetically modified microorganisms as encompassed by the claims. As amended, the claims do not require any genetic modification be carried out on any particular gene (e.g., the pykA and pykF genes), and as such the genetic modifications named in the rejection are unlimited. Therefore Applicants have not provided sufficient guidance to enable one of ordinary skill in the art to make and use the claimed invention in a manner reasonably correlated with the scope of the claims.
Claim Rejections - 35 USC § 103
Claims 1-5, 8 and 14 are rejected under 35 U.S.C. 103 as being unpatentable over Japanese Application No. JP 2011-515111A (cited on the Form PTO-892 mailed 03/08/2024; reference is made to the machine translation cited on the Form PTO-892 mailed 03/08/2024; herein referred to as 111A) in view of UniProt Accession No. A0A2V4GCF0_SERMA (09/12/2018, 1 page; cited on the Form PTO-892 mailed 08/04/2025; herein referred to as UNI1), and evidentiary references Vuoristo et al. (Trend Biotechnol, 2016, 34:191; cited on the Form PTO-892 mailed 01/17/2025; herein referred to as Vuoristo) and Bueno et al. (Antioxidant Redox Signal, 2012, 16:819; cited on the Form PTO-892 mailed 01/17/2025; herein referred to as Bueno).
The instant rejection is maintained from the previous Office Action and any newly recited portions are necessitated by claim amendment.
Claim 1 (claims 2-5 and 8 dependent therefrom) are drawn to a genetically modified microorganism in which a nucleic acid encoding any one of the polypeptides described in (a) to (b) below is introduced by a genetic modification or the expression of the polypeptide is enhanced by a genetic modification:
(a) a polypeptide composed of the amino acid sequence of SEQ ID NO: 2;
(b) a polypeptide composed of the amino acid sequence with a sequence identity of not less than 90% to the sequence of SEQ ID NO: 2 and having an enzymatic activity that catalyzes a reaction to reduce 3-OA-CoA to 3-HA-CoA;
wherein a function of PykA and/or PykF of the microorganism is impaired by a genetic modification.
Claim 14 is drawn to a genetically modified microorganism wherein a nucleic acid encoding any one of the polypeptides described in (a) to (b) below is introduced by a genetic modification or the expression of the polypeptide is enhanced by a genetic modification:
(a) a polypeptide composed of the amino acid sequence of SEQ ID NO: 2;
(b) a polypeptide composed of the amino acid sequence with a sequence identity of not less than 90% to the sequence of SEQ ID NO: 2 and having an enzymatic activity that catalyzes a reaction to reduce 3-oxoadipyl-CoA to 3-hydroxyadipyl-CoA;
wherein a function of PykF and/or PykA of the microorganism is impaired by a genetic modification,
wherein a production of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid is increased by said genetic modifications.
The limitations in claims 1 and 14 of “a nucleic acid … is introduced” and “the expression of the polypeptide is enhanced” are being interpreted as alternatives in consideration that they are linked with the conjunction “or”.
The limitation in claims 1 and 14 of “wherein a function of PykA and/or PykF is impaired by a genetic modification” is not directed to any specific genetic modification, therefore the limitation encompasses, for example, the genetic modification of introducing a nucleic acid encoding polypeptide (a) or (b) into an organism. Additionally, as impairment of the PykA and/or PykF function resulting from a genetic modification is recited without a reference for comparison, the claim encompasses any level of PykA and/or PykF function.
111A discusses microorganisms to produce adipic acid and other compounds [title], and discloses non-naturally occurring microorganisms with an adipate pathway and methods to produce adipate [abstract].
Regarding claim 1, 111A discloses a genetically modified E. coli strain for the production of adipate wherein the strain has been engineered by introducing nucleic acids encoding enzymes that include PaaH, a 3-hydroxyacyl-CoA dehydrogenase [p 20, section “Example II”, para 2-3] which is used for the reduction of 3-OA-CoA to 3-HA-CoA [p 15, para 5]. In view of the interpretation of the limitations of “a nucleic acid … is introduced or the expression of the polypeptide is enhanced by genetic modification” (emphasis added) as alternatives in consideration that they are linked with the conjunction “or”, the teachings of 111A are considered to correspond to introducing into an organism a nucleic acid encoding a polypeptide for the reduction of 3-OA-CoA to 3-HA-CoA, and wherein introduction of a nucleic acid to an organism is a genetic modification.
Regarding the limitation of a function of PykA and/or PykF, 111A describes the engineering of an organism for the reverse adipate degradation pathway [p 20, section “Example II”, para 2, pathway shown in Figure 2], wherein the initial reaction involves the consumption of acetyl-CoA and ultimately produces adipic acid in addition to oxidizing NADH to NAD+ along the way. One of skill in the art would be expected to reason that acetyl-CoA is produced from pyruvate, and pyruvate is produced from PEP via pyruvate kinase activity as evidenced by Vuoristo [Figure 1A depicting conventional glucose metabolism], wherein pyruvate kinase corresponds to the protein encoded by the genes pykA and/or pykF [instant specification, para 0009]. Therefore in view of the interpretation set forth above regarding the limitation of “a function of PykA and/or PykF is impaired by a genetic modification”, the teachings of 111A are considered to correspond to a cell comprising a genetic modification and having a function of PykA and/or PykF encompassed by the claim.
111A does not teach the sequence limitations of the polypeptide recited in parts (a) and (b) of claim 1.
UNI1 discloses a 3-hydroxybutyryl-CoA dehydrogenase enzyme from Serratia marcescens that shares 94.3% sequence identity with SEQ ID NO: 2 [see Appendix A], and as the polypeptide of UNI1 is encompassed by the sequence requirements of the claim, it is considered to have the enzymatic activity of catalyzing the reduction of 3-OA-CoA to 3-HA-CoA, as the function of the polypeptide is presumed to be inherent to its structure (see MPEP 2112.01(I)).
In view of 111A and UNI1, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date to modify the cell of 111A by using the polypeptide of UNI1. One of ordinary skill in the art would have recognized that the PaaH of 111A and the enzyme of UNI1 are both 3-hydroxyacyl-CoA dehydrogenases, and as such both are capable of being incorporated into the cell described by 111A. Thus it would have been obvious to one of ordinary skill in the art to replace the PaaH of 111A with the enzyme of UNI1, as one of ordinary skill in the art would have been able to carry out such a substitution with a reasonable expectation of success because both the 111A and UNI1 disclose enzymes with 3-hydroxyacyl-CoA dehydrogenase activity.
In view of the elected species
“relative to SEQ ID NO: 173 that the 13th amino acid is phenylalanine, the 15th amino acid is glutamine, the 16th amino acid is lysine, the 17th amino acid is glycine, the 19th amino acid is proline, and the 21st amino acid is methionine”
as recited in claim 3 being free of the prior art of record, the search and examination has been extended to the following species:
“relative to SEQ ID NO: 173 that the 13th amino acid is phenylalanine, the 15th amino acid is glutamine, the 16th amino acid is lysine, the 17th amino acid is glycine, the 19th amino acid is arginine, and the 21st amino acid is valine”.
Regarding claims 2-3, UNI1 discloses a 3-hydroxybutyryl-CoA dehydrogenase enzyme from Serratia marcescens that contains the amino acid sequence of SEQ ID NO: 173 and relative to SEQ ID NO: 173 has a 13th amino acid of phenylalanine (F), 15th amino acid of glutamine (Q), a 16th amino acid of lysine (K), a 17th amino acid of glycine (G), a 19th amino acid of arginine (R), and a 21st amino acid of valine (V) [see Appendix B].
Regarding claim 4, 111A discloses the modified organism is E. coli.
Regarding claim 5, the combined microorganism of 111A and UNI1 has the ability to generate 3-oxoadipyl-CoA and CoA from acetyl-CoA and succinyl-CoA and 3-hydroxyadipic acid from 3-hydroxyadipyl-CoA [p 7, para 3 of 111A].
Regarding claim 8, the combined microorganism of 111A and UNI1 as described above is engineered for the reverse adipate degradation pathway [p 20, section “Example II”, para 2, pathway shown in Figure 2 in 111A], wherein the initial reaction involves the consumption of acetyl-CoA and ultimately produces adipic acid in addition to oxidizing NADH to NAD+ along the way. It is understood in the art that acetyl-CoA plays a role in central metabolism for feeding into the TCA cycle to ultimately reduce NAD+ to NADH as part of the generation of energy [as evidenced by Vuoristo, p 191, para 2 and Figure 1A], wherein these reduced NADH molecules are oxidized again to NAD+ during respiration as part of the activity of the phosphotransfer system enzyme ATPase to produce ATP [as evidenced by Bueno, Fig 1A]. Therefore one of skill in the art would recognize a reverse adipate degradation pathway which consumes acetyl-CoA would thereby reduce the activity of the TCA cycle, further reducing the activity of the phosphotransfer system enzyme ATPase used to generate ATP via respiration. Therefore, the combined microorganism of 111A and UNI1 satisfies the limitations recited in the claim.
Regarding claim 14, the cell of claim 14 is substantially similar to the cell of claim 1, except the cell of claim 14 is limited to additionally have increased production of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid as a result of said genetic modifications. The limitation “wherein a production of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid is increased by said genetic modifications” is not directed to any specific genetic modification as it references genetic modifications previously recited in the claim, which are themselves not directed to any specific genetic modification. Therefore the limitation encompasses, for example, the genetic modification of introducing a nucleic acid encoding polypeptide (a) or (b) into an organism. Additionally, as the increased production of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid resulting from said genetic modification is recited without a reference for comparison, the claim encompasses any level of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid production.
Regarding claim 14, as stated in the rejection of claim 1 above regarding the limitations in common with claim 14, 111A discloses a genetically modified E. coli strain for the production of adipate wherein the strain has been engineered by introducing nucleic acids encoding enzymes that include PaaH, a 3-hydroxyacyl-CoA dehydrogenase [p 20, section “Example II”, para 2-3] which is used for the reduction of 3-OA-CoA to 3-HA-CoA [p 15, para 5]. In view of the interpretation of the limitations of “a nucleic acid … is introduced or the expression of the polypeptide is enhanced by genetic modification” (emphasis added) as alternatives in consideration that they are linked with the conjunction “or”, the teachings of 111A are considered to correspond to introducing into an organism a nucleic acid encoding a polypeptide for the reduction of 3-OA-CoA to 3-HA-CoA, and wherein introduction of a nucleic acid to an organism is a genetic modification.
Regarding the limitation of a function of PykA and/or PykF, 111A describes the engineering of an organism for the reverse adipate degradation pathway [p 20, section “Example II”, para 2, pathway shown in Figure 2], wherein the initial reaction involves the consumption of acetyl-CoA and ultimately produces adipic acid in addition to oxidizing NADH to NAD+ along the way. One of skill in the art would be expected to reason that acetyl-CoA is produced from pyruvate, and pyruvate is produced from PEP via pyruvate kinase activity as evidenced by Vuoristo [Figure 1A depicting conventional glucose metabolism], wherein pyruvate kinase corresponds to the protein encoded by the genes pykA and/or pykF [instant specification, para 0009]. Therefore in view of the interpretation set forth above regarding the limitation of “a function of PykA and/or PykF is impaired by a genetic modification”, the teachings of 111A are considered to correspond to a cell comprising a genetic modification and having a function of PykA and/or PykF encompassed by the claim.
Regarding claim 14 and the limitation of production of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid is increased by said genetic modification, 111A describes in Example V [p 25, para 3] a modified E. coli cell with three introduced heterologous genes, wherein the cell is capable of forming adipic acid, which is considered to correspond to a genetic mutation resulting in increased production of adipic acid.
Regarding claim 14 and the limitations of polypeptides (a) and (b), UNI1 discloses a 3-hydroxybutyryl-CoA dehydrogenase enzyme from Serratia marcescens that shares 94.3% sequence identity with SEQ ID NO: 2 [see Appendix A], and as the polypeptide of UNI1 is encompassed by the sequence requirements of the claim, it is considered to have the enzymatic activity of catalyzing the reduction of 3-OA-CoA to 3-HA-CoA, as the function of the polypeptide is presumed to be inherent to its structure (see MPEP 2112.01(I)).
In view of 111A and UNI1, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date to modify the cell of 111A by using the polypeptide of UNI1 to arrive at the claimed invention. One of ordinary skill in the art would have recognized that the PaaH of 111A and the enzyme of UNI1 are both 3-hydroxyacyl-CoA dehydrogenases, and as such both are capable of being incorporated into the cell described by 111A. Thus it would have been obvious to one of ordinary skill in the art to replace the PaaH of 111A with the enzyme of UNI1, as one of ordinary skill in the art would have been able to carry out such a substitution with a reasonable expectation of success because both the 111A and UNI1 disclose enzymes with 3-hydroxyacyl-CoA dehydrogenase activity.
Therefore, the invention of claims 1-5, 8 and 14 would have been obvious to one of ordinary skill in the art before the effective filing date.
Claims 6-7 are rejected under 35 U.S.C. 103 as being unpatentable over 111A and UNI1 as applied to claims 1-5, 8 and 14 above, and further in view of Parthasarathy et al. (Biochem, 2011, 50:3540; cited on the Form PTO-892 mailed 01/17/2025; herein referred to as Parthasarathy).
The instant rejection is maintained from the previous Office Action and any newly recited portions are necessitated by claim amendment.
Claim 6 is drawn to the genetically modified microorganism of claim 1, which has an ability to generate 3-oxoadipyl-CoA and coenzyme A from acetyl-CoA and succinyl-CoA, an ability to generate 2,3-dehydroadipyl-CoA from 3-hydroxyadipyl-CoA, and an ability to generate alpha-hydromuconic acid from 2,3-dehydroadipyl-CoA.
Claim 7 is drawn to the genetically modified microorganism of claim 1, which has an ability to generate 3-oxoadipyl-CoA and coenzyme A from acetyl-CoA and succinyl-CoA, an ability to generate 2,3-dehydroadipyl-CoA from 3-hydroxyadipyl-CoA, an ability to generate adipyl-CoA from 2,3-dehydroadipyl-CoA, and an ability to generate adipic acid from adipyl-CoA.
It is noted that the specification defines alpha-hydromuconic acid as (E)-hex-2-enedioic acid [para 0002] and 2,3-dehydroadipate [para 0005], which is also known in the art as 2-hexenedioic acid, beta-dihydromuconic acid. It is noted that the instant application refers to 2,3-dehydroadipyl-CoA which is known in the art as 5-carboxy-2-pentenoyl-CoA.
The teachings of 111A and UNI1 as applied to claims 1-5, 8 and 14 are discussed above, and include the microorganism with the ability to generate 3-oxoadipyl-CoA and CoA from acetyl-CoA and succinyl-CoA [p 7, para 3]
111A and UNI1 do not teach the limitation of generating 3-oxoadipyl-CoA and coenzyme A from acetyl-CoA and succinyl-CoA, generating 2,3-dehydroadipyl-CoA from 3-hydroxyadipyl-CoA, and generating alpha-hydromuconic acid from 2,3-dehydroadipyl-CoA.
Regarding claim 6, 111A describes in Example IX the generation of a genetically modified microorganism capable of carrying out the 3-oxoadipate pathway [p 30, paras 1-2 under heading Example IX] characterized by Figure 2, which show the reactions of converting 3-hydroxyadipyl-CoA to 2,3-dehydroadipyl-CoA [reaction 3 of Figure 2].
Parthasarathy discusses substrate specificity of 2-hydroxyglutaryl-CoA dehydratase for the production of adipic acid [title], wherein it is described that expression of six genes from two glutamate fermenting clostridia can also be used to dehydrate the 2-hydroxyapidic acid CoA thioester to 2-hexendioic acid, an unsaturated precursor of the biotechnologically valuable adipic acid [abstract].
Regarding claim 6 and the limitation of generating alpha-hydromuconic acid from 2,3-dehydroadipyl-CoA, Parthasarathy discloses the production of alpha-hydromuconic acid from 2,3-dehydroadipyl-CoA via the activity of the enzymes glutaconate CoA-transferase (GctAB) from Acidaminococcus fermentans [p 3540, col 1, para 2], the scheme of which is shown in the [Figure within the abstract, and Figure 1].
In view of Parthasarathy, it would have been prima facie obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the combined cell of 111A and UNI1 with the genes GctAB, as taught by Parthasarathy, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the combined cell of 111A and UNI1, because Parthasarathy teaches that expression of six genes from two glutamate fermenting clostridia can also be used to dehydrate the 2-hydroxyapidic acid CoA thioester to 2-hexendioic acid, an unsaturated precursor of the biotechnologically valuable adipic acid. One of ordinary skill in the art would have had a reasonable expectation of success because both 111A and Parthasarathy discuss engineering microorganisms to produce adipate.
Regarding claim 7, 111A describes in Example IX the generation of a genetically modified microorganism capable of carrying out the 3-oxoadipate pathway [p 30, paras 1-2 under heading Example IX] characterized by Figure 2, which show the reactions of converting 3-hydroxyadipyl-CoA to 2,3-dehydroadipyl-CoA and further converting 2,3-dehydroadipyl-CoA to adipyl-CoA [reactions 3 and 4 of Figure 2].
Therefore, the invention of claims 6-7 would have been obvious to one of ordinary skill in the art before the effective filing date.
Response to Remarks: beginning on page 6 of Applicant’s response to rejections under 35 USC 103; Applicant in summary contends that the amended claims recite features not taught or suggested by 111A or the other prior art of record, particularly the limitation of 2 distinct genetic modifications related to (1) introduction or expression of a polypeptide and (2) impairment of PykA and/or PykA function, and not a single genetic modification that has both outcomes; Applicant further contends the prior art does not teach or suggest a genetically modified microorganism that produces 3-hydroxyadipic acid, α-hydromuconic acid and/or adipic acid; Applicant further contends that the genetic modifications in claim 1 inherently result in increased production of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid, and therefore the claims do not require the explicit recitation of this feature in the claim.
Applicant’s remarks are considered and found not convincing.
Regarding the assertion that two distinct genetic modifications are required by claim 1, the genetic modifications in the claim are not directed to any particular gene. Therefore there is no limitation or language to exclude the multiple effects recited by the claim to result from a single genetic modification to the same gene, given a broadest reasonable interpretation.
Regarding the assertion that the prior art doesn’t teach or suggest a genetically modified microorganism that produces 3-hydroxyadipic acid, α-hydromuconic acid and/or adipic acid, the rejection of claim 14 above describes the combination of prior art 111A and UNI1, comprising a cell genetically modified to produce a protein encompassed by the claim limitations, and a genetic modification to produce adipic acid. It is noted that the three products of 3-hydroxyadipic acid, α-hydromuconic acid and adipic acid are recited in the claim in the alternative, and therefore the claims do not require a cell genetically modified to produce 3-hydroxyadipic acid, α-hydromuconic acid and/or adipic acid.
Regarding the assertion that the genetic modifications in claim 1 inherently result in increased production of 3-hydroxyadipic acid, α-hydromuconic acid and/or adipic acid, and therefore the claims do not require the explicit recitation of this feature in the claim: the combined cell of 111A and UNI1 as set forth above is considered to correspond to a cell comprising genetic modifications encompassed by the claims, and in consideration of Applicant’s remarks, the combined cell would be considered to inherently result in increased production of 3-hydroxyadipic acid, α-hydromuconic acid and/or adipic acid.
Double Patenting
The provisional rejection of claims 1-4 on the ground of nonstatutory double patenting as being unpatentable over claims 1, 4, 7 and 19 of co-pending Application 18/848396 (herein “reference application”) in view of UNI1 is withdrawn in view of the instant claim amendments.
A. Claims 1-5, 7-8 and 14 are provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claims 1, 4, 7 and 19 of co-pending Application 18/848396 in view of UNI1, 111A and evidentiary references Vuoristo and Beuno.
The instant rejection is maintained from the previous Office Action and any newly recited portions are necessitated by claim amendment.
Regarding instant claim 1, claim 1 of the reference application recites a genetically modified organism, claim 4 of the reference application recites the genetically modified organism wherein a gene is introduced, and claim 19 of the reference application recites the organism contains a genetically modified enzyme having enhanced activities of enzymes that catalyze the reaction to reduce 3-OA-CoA to produce 3-HA-CoA. In view of the interpretation of the limitations of “a nucleic acid … is introduced” and “the expression of the polypeptide is enhanced and the function of pyruvate kinase is impaired…” as alternatives in consideration that they are linked with the conjunction “or”, the genetically modified organism of the reference application is considered to satisfy the limitation of a genetically modified cell with a nucleic acid introduced of instant claim 1.
The claims of the reference application do not recite a polypeptide with the sequence limitations of the instant claims and the impairment of PykA and/or PykF by a genetic modification.
111A discusses microorganisms to produce adipic acid and other compounds [title], and discloses non-naturally occurring microorganisms with an adipate pathway and methods to produce adipate [abstract].
Regarding instant claim 1, 111A discloses a genetically modified E. coli strain for the production of adipate wherein the strain has been engineered by introducing nucleic acids encoding enzymes that include PaaH, a 3-hydroxyacyl-CoA dehydrogenase [p 20, section “Example II”, para 2-3] which is used for the reduction of 3-OA-CoA to 3-HA-CoA [p 15, para 5]. The disclosure of 111A is considered to correspond to introducing into an organism a nucleic acid encoding a polypeptide for the reduction of 3-OA-CoA to 3-HA-CoA.
Regarding the limitation of a function of PykA and/or PykF, 111A describes the engineering of an organism for the reverse adipate degradation pathway [p 20, section “Example II”, para 2, pathway shown in Figure 2], wherein the initial reaction involves the consumption of acetyl-CoA and ultimately produces adipic acid in addition to oxidizing NADH to NAD+ along the way. One of skill in the art would be expected to reason that acetyl-CoA is produced from pyruvate, and pyruvate is produced from PEP via pyruvate kinase activity as evidenced by Vuoristo [Figure 1A depicting conventional glucose metabolism], wherein pyruvate kinase corresponds to the protein encoded by the genes pykA and/or pykF [instant specification, para 0009]. Therefore in view of the interpretation set forth above regarding the limitation of “a function of PykA and/or PykF is impaired by a genetic modification”, the disclosure of 111A is considered to correspond to a cell comprising a genetic modification and having a function of PykA and/or PykF encompassed by the instant claim.
UNI1 discloses a 3-hydroxybutyryl-CoA dehydrogenase enzyme from Serratia marcescens that shares 94.3% sequence identity with SEQ ID NO: 2 [see Appendix A], and as the polypeptide of UNI1 is encompassed by the sequence requirements of the claim, it is considered to have the enzymatic activity of catalyzing the reduction of 3-OA-CoA to 3-HA-CoA, as the function of the polypeptide is presumed to be inherent to its structure (see MPEP 2112.01(I)).
In view of 111A and UNI1, it would have been obvious to one of ordinary skill in the art before the effective filing date to modify the cell of the claims of the reference application by using the microorganism of 111A and the enzyme of UNI1.
One of ordinary skill in the art would have recognized that the microorganism of the reference application and the microorganism of 111A are both microorganisms genetically modified for the production of adipic acid, and as such both are capable of being incorporated into the claim of the reference application. Thus it would have been obvious to one of ordinary skill in the art to replace the microorganism of the claim of the reference application with the microorganism of 111A, as one of ordinary skill in the art would have been able to carry out such a substitution with a reasonable expectation of success because the claim of the reference application and 111A disclose microorganisms genetically modified to produce adipic acid.
One of ordinary skill in the art would have recognized that the enzyme of the 111A and the enzyme of UNI1 both have 3-hydroxyacyl-CoA dehydrogenase activity, and as such both are capable of being incorporated into the combined microorganism of the reference application and 111A. Thus it would have been obvious to one of ordinary skill in the art to replace the enzyme of 111A with the enzyme of UNI1, as one of ordinary skill in the art would have been able to carry out such a substitution with a reasonable expectation of success because 111A and UNI1 disclose enzymes with 3-hydroxyacyl-CoA dehydrogenase activity.
Regarding instant claims 2-3, UNI1 discloses a 3-hydroxybutyryl-CoA dehydrogenase enzyme from Serratia marcescens that contains the amino acid sequence of SEQ ID NO: 173 and relative to SEQ ID NO: 173 has a 13th amino acid of phenylalanine (F), 15th amino acid of glutamine (Q), a 16th amino acid of lysine (K), a 17th amino acid of glycine (G), a 19th amino acid of arginine (R), and a 21st amino acid of valine (V) [see Appendix A].
Regarding instant claim 4, claim 7 of the reference application recites the organism is an Escherichia or Serratia.
Regarding instant claim 5, 111A discloses a microorganism that contains the ability to generate 3-oxoadipyl-CoA and CoA from acetyl-CoA and succinyl-CoA and 3-hydroxyadipic acid from 3-hydroxyadipyl-CoA [p 7, para 3].
Regarding instant claim 7, 111A describes in Example IX the generation of a genetically modified microorganism capable of carrying out the 3-oxoadipate pathway [p 30, paras 1-2 under heading Example IX] characterized by Figure 2, which show the reactions of converting 3-hydroxyadipyl-CoA to 2,3-dehydroadipyl-CoA and further converting 2,3-dehydroadipyl-CoA to adipyl-CoA [reactions 3 and 4 of Figure 2].
Regarding instant claim 8, the microorganism of 111A as described above is engineered for the reverse adipate degradation pathway [p 20, section “Example II”, para 2, pathway shown in Figure 2], wherein the initial reaction involves the consumption of acetyl-CoA and ultimately produces adipic acid in addition to oxidizing NADH to NAD+ along the way. It is understood in the art that acetyl-CoA plays a role in central metabolism for feeding into the TCA cycle to ultimately reduce NAD+ to NADH as part of the generation of energy [as evidenced by Vuoristo, p 191, para 2 and Figure 1A], wherein these reduced NADH molecules are oxidized again to NAD+ during respiration as part of the phosphotransfer system to produce ATP via ATPase activity [as evidenced by Bueno, Fig 1A]. Therefore one of skill in the art would recognize a reverse adipate degradation pathway which consumes acetyl-CoA would thereby reduce the activity of the TCA cycle, further reducing the activity of the phosphotransfer system used to generate ATP via respiration. Therefore, the microorganism of 111A satisfies the limitations recited in the claim.
Regarding instant claim 14, the cell of the instant claim is substantially similar to the cell of instant claim 1, except the cell of the instant claim is limited to additionally have increased production of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid as a result of said genetic modifications. The limitation “wherein a production of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid is increased by said genetic modifications” is not directed to any specific genetic modification as it references genetic modifications previously recited in the claim, which are themselves not directed to any specific genetic modification. Therefore the limitation encompasses, for example, the genetic modification of introducing a nucleic acid encoding polypeptide (a) or (b) into an organism. Additionally, as the increased production of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid resulting from said genetic modification is recited without a reference for comparison, the instant claim encompasses any level of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid production.
Regarding instant claim 14, as stated in the rejection of instant claim 1 above regarding the limitations in common with instant claim 14, 111A discloses a genetically modified E. coli strain for the production of adipate wherein the strain has been engineered by introducing nucleic acids encoding enzymes that include PaaH, a 3-hydroxyacyl-CoA dehydrogenase [p 20, section “Example II”, para 2-3] which is used for the reduction of 3-OA-CoA to 3-HA-CoA [p 15, para 5]. In view of the interpretation of the limitations of “a nucleic acid … is introduced or the expression of the polypeptide is enhanced by genetic modification” (emphasis added) as alternatives in consideration that they are linked with the conjunction “or”, the disclosure of 111A is considered to correspond to introducing into an organism a nucleic acid encoding a polypeptide for the reduction of 3-OA-CoA to 3-HA-CoA, and wherein introduction of a nucleic acid to an organism is a genetic modification.
Regarding the limitation of a function of PykA and/or PykF, 111A describes the engineering of an organism for the reverse adipate degradation pathway [p 20, section “Example II”, para 2, pathway shown in Figure 2], wherein the initial reaction involves the consumption of acetyl-CoA and ultimately produces adipic acid in addition to oxidizing NADH to NAD+ along the way. One of skill in the art would be expected to reason that acetyl-CoA is produced from pyruvate, and pyruvate is produced from PEP via pyruvate kinase activity as evidenced by Vuoristo [Figure 1A depicting conventional glucose metabolism], wherein pyruvate kinase corresponds to the protein encoded by the genes pykA and/or pykF [instant specification, para 0009]. Therefore in view of the interpretation set forth above regarding the limitation of “a function of PykA and/or PykF is impaired by a genetic modification”, the disclosure of 111A is considered to correspond to a cell comprising a genetic modification and having a function of PykA and/or PykF encompassed by the claim.
Regarding instant claim 14 and the limitation of production of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid is increased by said genetic modification, 111A describes in Example V [p 25, para 3] a modified E. coli cell with three introduced heterologous genes, wherein the cell is capable of forming adipic acid, which is considered to correspond to a genetic mutation resulting in increased production of adipic acid.
Regarding instant claim 14 and the limitations of polypeptides (a) and (b), UNI1 discloses a 3-hydroxybutyryl-CoA dehydrogenase enzyme from Serratia marcescens that shares 94.3% sequence identity with SEQ ID NO: 2 [see Appendix A], and as the polypeptide of UNI1 is encompassed by the sequence requirements of the claim, it is considered to have the enzymatic activity of catalyzing the reduction of 3-OA-CoA to 3-HA-CoA, as the function of the polypeptide is presumed to be inherent to its structure (see MPEP 2112.01(I)).
In view of 111A and UNI1, it would have been obvious to one of ordinary skill in the art before the effective filing date to modify the cell of the claims of the reference application by using the microorganism of 111A and the enzyme of UNI1.
One of ordinary skill in the art would have recognized that the microorganism of the reference application and the microorganism of 111A are both microorganisms genetically modified for the production of adipic acid, and as such both are capable of being incorporated into the claim of the reference application. Thus it would have been obvious to one of ordinary skill in the art to replace the microorganism of the claim of the reference application with the microorganism of 111A, as one of ordinary skill in the art would have been able to carry out such a substitution with a reasonable expectation of success because the claim of the reference application and 111A disclose microorganisms genetically modified to produce adipic acid.
One of ordinary skill in the art would have recognized that the enzyme of the 111A and the enzyme of UNI1 both have 3-hydroxyacyl-CoA dehydrogenase activity, and as such both are capable of being incorporated into the combined microorganism of the reference application and 111A. Thus it would have been obvious to one of ordinary skill in the art to replace the enzyme of 111A with the enzyme of UNI1, as one of ordinary skill in the art would have been able to carry out such a substitution with a reasonable expectation of success because 111A and UNI1 disclose enzymes with 3-hydroxyacyl-CoA dehydrogenase activity.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claim 6 is provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claims 1, 4, 7 and 19 of co-pending Application 18/848396 in view of UNI1and 111A and as evidenced by Vuoristo and Beuno as applied to claims 1-5, 7-8 and 14 above, and further in view of Parthasarathy.
The instant rejection is maintained from the previous Office action, and any newly recited portion is necessitated by claim amendment.
The claims of the reference application and the disclosures of UNI1 and 111A as applied to claims 1-5, 7-8 and 14 are discussed above. The claims of the reference application do not recite a microorganism with an ability to generate a-hydromuconic acid from 2,3-dehydroadipyl-CoA.
Parthasarathy discusses substrate specificity of 2-hydroxyglutaryl-CoA dehydratase for the production of adipic acid [title], wherein it is described that expression of six genes from two glutamate fermenting clostridia can also be used to dehydrate the 2-hydroxyapidic acid CoA thioester to 2-hexendioic acid, an unsaturated precursor of the biotechnologically valuable adipic acid [abstract].
Regarding instant claim 6 and the limitation of generating alpha-hydromuconic acid from 2,3-dehydroadipyl-CoA, Parthasarathy discloses the production of alpha-hydromuconic acid from 2,3-dehydroadipyl-CoA via the activity of the enzymes glutaconate CoA-transferase (GctAB) from Acidaminococcus fermentans [p 3540, col 1, para 2], the scheme of which is shown in the [Figure within the abstract, and Figure 1].
In view of Parthasarathy, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the claims of the reference application with the genes GctAB, as taught by Parthasarathy, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the claims of reference application, because Parthasarathy discloses that expression of six genes from two glutamate fermenting clostridia can also be used to dehydrate the 2-hydroxyapidic acid CoA thioester to 2-hexendioic acid, an unsaturated precursor of the biotechnologically valuable adipic acid. One of ordinary skill in the art would have had a reasonable expectation of success because the claims of the reference application, 111A and Parthasarathy are related to engineering microorganisms to produce adipate.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
B. Claims 1-5, 7-8 and 14 are provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claim 1 of co-pending Application 17/924145 (herein “reference application”) in view of 111A and UNI1 and evidentiary references Vuoristo and Beuno.
The instant rejection was set forth in the previous Office action with a typo in the co-pending Application No. of 18/924145, which has been corrected here as co-pending Application No. 17/924125. The instant rejection therefore is maintained from the previous Office Action and any newly recited portions are necessitated by claim amendment.
Regarding instant claim 1, claim 1 of the reference application recites a genetically modified microorganism comprising a gene with a function to increase productivity of 3-hydroxyadipic acid, alpha-hydroxymuconic acid and/or adipic acid. In view of the interpretation of the limitations of “a nucleic acid … is introduced” and “the expression of the polypeptide is enhanced” as alternatives in consideration that they are linked with the conjunction “or”, the genetically modified organism of the reference application is considered to satisfy the limitation of a genetically modified cell with a nucleic acid introduced of instant claim 1.
The claims of the reference application do not recite a polypeptide with the sequence limitations recited in the instant claim or the impairment of PykA and/or PykF by a genetic modification.
111A discusses microorganisms to produce adipic acid and other compounds [title], and discloses non-naturally occurring microorganisms with an adipate pathway and methods to produce adipate [abstract].
Regarding instant claim 1, 111A discloses a genetically modified E. coli strain for the production of adipate wherein the strain has been engineered by introducing nucleic acids encoding enzymes that include PaaH, a 3-hydroxyacyl-CoA dehydrogenase [p 20, section “Example II”, para 2-3] which is used for the reduction of 3-OA-CoA to 3-HA-CoA [p 15, para 5]. The disclosure of 111A is considered to correspond to introducing into an organism a nucleic acid encoding a polypeptide for the reduction of 3-OA-CoA to 3-HA-CoA.
Regarding the limitation of a function of PykA and/or PykF, 111A describes the engineering of an organism for the reverse adipate degradation pathway [p 20, section “Example II”, para 2, pathway shown in Figure 2], wherein the initial reaction involves the consumption of acetyl-CoA and ultimately produces adipic acid in addition to oxidizing NADH to NAD+ along the way. One of skill in the art would be expected to reason that acetyl-CoA is produced from pyruvate, and pyruvate is produced from PEP via pyruvate kinase activity as evidenced by Vuoristo [Figure 1A depicting conventional glucose metabolism], wherein pyruvate kinase corresponds to the protein encoded by the genes pykA and/or pykF [instant specification, para 0009]. Therefore in view of the interpretation set forth above regarding the limitation of “a function of PykA and/or PykF is impaired by a genetic modification”, the disclosure of 111A is considered to correspond to a cell comprising a genetic modification and having a function of PykA and/or PykF encompassed by the instant claim.
UNI1 discloses a 3-hydroxybutyryl-CoA dehydrogenase enzyme from Serratia marcescens that shares 94.3% sequence identity with SEQ ID NO: 2 [see Appendix A], and as the polypeptide of UNI1 is encompassed by the sequence requirements of the claim, it is considered to have the enzymatic activity of catalyzing the reduction of 3-OA-CoA to 3-HA-CoA, as the function of the polypeptide is presumed to be inherent to its structure (see MPEP 2112.01(I)).
In view of 111A and UNI1, it would have been obvious to one of ordinary skill in the art before the effective filing date to modify the cell of the claims of the reference application by using the microorganism of 111A and the enzyme of UNI1.
One of ordinary skill in the art would have recognized that the microorganism of the reference application and the microorganism of 111A are both microorganisms genetically modified for the production of adipic acid, and as such both are capable of being incorporated into the claim of the reference application. Thus it would have been obvious to one of ordinary skill in the art to replace the microorganism of the claim of the reference application with the microorganism of 111A, as one of ordinary skill in the art would have been able to carry out such a substitution with a reasonable expectation of success because the claim of the reference application and 111A disclose microorganisms genetically modified to produce adipic acid.
One of ordinary skill in the art would have recognized that the enzyme of the 111A and the enzyme of UNI1 both have 3-hydroxyacyl-CoA dehydrogenase activity, and as such both are capable of being incorporated into the combined microorganism of the reference application and 111A. Thus it would have been obvious to one of ordinary skill in the art to replace the enzyme of 111A with the enzyme of UNI1, as one of ordinary skill in the art would have been able to carry out such a substitution with a reasonable expectation of success because 111A and UNI1 disclose enzymes with 3-hydroxyacyl-CoA dehydrogenase activity.
Regarding instant claims 2-3, UNI1 discloses a 3-hydroxybutyryl-CoA dehydrogenase enzyme from Serratia marcescens that contains the amino acid sequence of SEQ ID NO: 173 and relative to SEQ ID NO: 173 has a 13th amino acid of phenylalanine (F), 15th amino acid of glutamine (Q), a 16th amino acid of lysine (K), a 17th amino acid of glycine (G), a 19th amino acid of arginine (R), and a 21st amino acid of valine (V) [see Appendix A].
Regarding instant claim 4, claim 1 of the reference application recites the organism is an Escherichia or Serratia.
Regarding instant claim 5, 111A discloses a microorganism that contains the ability to generate 3-oxoadipyl-CoA and CoA from acetyl-CoA and succinyl-CoA and 3-hydroxyadipic acid from 3-hydroxyadipyl-CoA [p 7, para 3].
Regarding instant claim 7, 111A describes in Example IX the generation of a genetically modified microorganism capable of carrying out the 3-oxoadipate pathway [p 30, paras 1-2 under heading Example IX] characterized by Figure 2, which show the reactions of converting 3-hydroxyadipyl-CoA to 2,3-dehydroadipyl-CoA and further converting 2,3-dehydroadipyl-CoA to adipyl-CoA [reactions 3 and 4 of Figure 2].
Regarding instant claim 8, the microorganism of 111A as described above is engineered for the reverse adipate degradation pathway [p 20, section “Example II”, para 2, pathway shown in Figure 2], wherein the initial reaction involves the consumption of acetyl-CoA and ultimately produces adipic acid in addition to oxidizing NADH to NAD+ along the way. It is understood in the art that acetyl-CoA plays a role in central metabolism for feeding into the TCA cycle to ultimately reduce NAD+ to NADH as part of the generation of energy [as evidenced by Vuoristo, p 191, para 2 and Figure 1A], wherein these reduced NADH molecules are oxidized again to NAD+ during respiration as part of the phosphotransfer system to produce ATP via ATPase activity [as evidenced by Bueno, Fig 1A]. Therefore one of skill in the art would recognize a reverse adipate degradation pathway which consumes acetyl-CoA would thereby reduce the activity of the TCA cycle, further reducing the activity of the phosphotransfer system used to generate ATP via respiration. Therefore, the microorganism of 111A satisfies the limitations recited in the claim.
Regarding instant claim 14, the cell of the instant claim is substantially similar to the cell of instant claim 1, except the cell of the instant claim is limited to additionally have increased production of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid as a result of said genetic modifications. The limitation “wherein a production of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid is increased by said genetic modifications” is not directed to any specific genetic modification as it references genetic modifications previously recited in the claim, which are themselves not directed to any specific genetic modification. Therefore the limitation encompasses, for example, the genetic modification of introducing a nucleic acid encoding polypeptide (a) or (b) into an organism. Additionally, as the increased production of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid resulting from said genetic modification is recited without a reference for comparison, the instant claim encompasses any level of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid production.
Regarding instant claim 14, as stated in the rejection of instant claim 1 above regarding the limitations in common with instant claim 14, 111A discloses a genetically modified E. coli strain for the production of adipate wherein the strain has been engineered by introducing nucleic acids encoding enzymes that include PaaH, a 3-hydroxyacyl-CoA dehydrogenase [p 20, section “Example II”, para 2-3] which is used for the reduction of 3-OA-CoA to 3-HA-CoA [p 15, para 5]. In view of the interpretation of the limitations of “a nucleic acid … is introduced or the expression of the polypeptide is enhanced by genetic modification” (emphasis added) as alternatives in consideration that they are linked with the conjunction “or”, the disclosure of 111A is considered to correspond to introducing into an organism a nucleic acid encoding a polypeptide for the reduction of 3-OA-CoA to 3-HA-CoA, and wherein introduction of a nucleic acid to an organism is a genetic modification.
Regarding the limitation of a function of PykA and/or PykF, 111A describes the engineering of an organism for the reverse adipate degradation pathway [p 20, section “Example II”, para 2, pathway shown in Figure 2], wherein the initial reaction involves the consumption of acetyl-CoA and ultimately produces adipic acid in addition to oxidizing NADH to NAD+ along the way. One of skill in the art would be expected to reason that acetyl-CoA is produced from pyruvate, and pyruvate is produced from PEP via pyruvate kinase activity as evidenced by Vuoristo [Figure 1A depicting conventional glucose metabolism], wherein pyruvate kinase corresponds to the protein encoded by the genes pykA and/or pykF [instant specification, para 0009]. Therefore in view of the interpretation set forth above regarding the limitation of “a function of PykA and/or PykF is impaired by a genetic modification”, the disclosure of 111A is considered to correspond to a cell comprising a genetic modification and having a function of PykA and/or PykF encompassed by the claim.
Regarding instant claim 14 and the limitation of production of 3-hydroxyadipic acid, α-hydromuconic acid or adipic acid is increased by said genetic modification, 111A describes in Example V [p 25, para 3] a modified E. coli cell with three introduced heterologous genes, wherein the cell is capable of forming adipic acid, which is considered to correspond to a genetic mutation resulting in increased production of adipic acid.
Regarding instant claim 14 and the limitations of polypeptides (a) and (b), UNI1 discloses a 3-hydroxybutyryl-CoA dehydrogenase enzyme from Serratia marcescens that shares 94.3% sequence identity with SEQ ID NO: 2 [see Appendix A], and as the polypeptide of UNI1 is encompassed by the sequence requirements of the claim, it is considered to have the enzymatic activity of catalyzing the reduction of 3-OA-CoA to 3-HA-CoA, as the function of the polypeptide is presumed to be inherent to its structure (see MPEP 2112.01(I)).
In view of 111A and UNI1, it would have been obvious to one of ordinary skill in the art before the effective filing date to modify the cell of the claims of the reference application by using the microorganism of 111A and the enzyme of UNI1.
One of ordinary skill in the art would have recognized that the microorganism of the reference application and the microorganism of 111A are both microorganisms genetically modified for the production of adipic acid, and as such both are capable of being incorporated into the claim of the reference application. Thus it would have been obvious to one of ordinary skill in the art to replace the microorganism of the claim of the reference application with the microorganism of 111A, as one of ordinary skill in the art would have been able to carry out such a substitution with a reasonable expectation of success because the claim of the reference application and 111A disclose microorganisms genetically modified to produce adipic acid.
One of ordinary skill in the art would have recognized that the enzyme of the 111A and the enzyme of UNI1 both have 3-hydroxyacyl-CoA dehydrogenase activity, and as such both are capable of being incorporated into the combined microorganism of the reference application and 111A. Thus it would have been obvious to one of ordinary skill in the art to replace the enzyme of 111A with the enzyme of UNI1, as one of ordinary skill in the art would have been able to carry out such a substitution with a reasonable expectation of success because 111A and UNI1 disclose enzymes with 3-hydroxyacyl-CoA dehydrogenase activity.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claim 6 is provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over Application 17/924145 (herein “reference application”) in view of 111A and UNI1 as applied to claims 1-5, 7-8 and 14 above, and further in view of Parthasarathy.
The instant claim is maintained from the previous Office action, and any newly recited portions are necessitated by claim amendment.
The claims of the reference application and the disclosures of UNI1 and 111A as applied to claims 1-5, 7-8 and 14 are discussed above. The claims of the reference application do not recite a microorganism with an ability to generate a-hydromuconic acid from 2,3-dehydroadipyl-CoA.
Parthasarathy discusses substrate specificity of 2-hydroxyglutaryl-CoA dehydratase for the production of adipic acid [title], wherein it is described that expression of six genes from two glutamate fermenting clostridia can also be used to dehydrate the 2-hydroxyapidic acid CoA thioester to 2-hexendioic acid, an unsaturated precursor of the biotechnologically valuable adipic acid [abstract].
Regarding instant claim 6 and the limitation of generating alpha-hydromuconic acid from 2,3-dehydroadipyl-CoA, Parthasarathy discloses the production of alpha-hydromuconic acid from 2,3-dehydroadipyl-CoA via the activity of the enzymes glutaconate CoA-transferase (GctAB) from Acidaminococcus fermentans [p 3540, col 1, para 2], the scheme of which is shown in the [Figure within the abstract, and Figure 1].
In view of Parthasarathy, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to modify the claims of the reference application with the genes GctAB, as taught by Parthasarathy, to arrive at the claimed invention. One of ordinary skill in the art would have been motivated to modify the claims of reference application, because Parthasarathy discloses that expression of six genes from two glutamate fermenting clostridia can also be used to dehydrate the 2-hydroxyapidic acid CoA thioester to 2-hexendioic acid, an unsaturated precursor of the biotechnologically valuable adipic acid. One of ordinary skill in the art would have had a reasonable expectation of success because the claims of the reference application, 111A and Parthasarathy are related to engineering microorganisms to produce adipate.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Response to Remarks: beginning on page 10 of Applicant’s response to double patenting rejections; Applicant in summary request the rejections be held in abeyance until the allowable subject matter is found.
Applicant’s request is acknowledge, and the rejections above have been updated in view of the current amendments to the claims.
Conclusion
Status of the Application:
Claims 1-15 are pending.
Claims 9-13 are withdrawn.
Claims 1-8 and 14 are rejected.
No claim is in condition for allowance.
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.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to JOSEPH SPANGLER whose telephone number is (571)270-0314. The examiner can normally be reached M-F 7:30 am - 4:30 pm.
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/JOSEPH R SPANGLER/
Examiner
Art Unit 1656
/David Steadman/Primary Examiner, Art Unit 1656
APPENDIX A
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SEQUENCE ALIGNMENT OF SEQ ID NO: 2 WITH UNIPROT ACCESSION NO. A0A2V4GCF0_SERMA (REFERENCE UNI1)
APPENDIX B
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SEQUENCE ALIGNMENT OF SEQ ID NO: 173 WITH UNIPROT ACCESSION NO. A0A2V4GCF0_SERMA (REFERENCE UNI1)
Prosecution Insights