DETAILED ACTION
Notice of Pre-AIA or AIA Status
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Claim Status
The amendment of 10/20/2025 has been entered. Claims 1, 5-20, and 30-31 are pending (claim set as filed on 10/20/2025). Claims 11-20 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim.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 03/21/2025. Applicant previously elected invention Group I, drawn to a culture, and the species glycerol as a nutrient source species under disrupted phosphofructokinase conditions, and 3-phosphoglycerate as a nutrient source species under disrupted 3-phospoglycerate kinase conditions in the communications filed on 03/21/2025 and 03/03/2025.
Claims 1, 5-10, and 30-31 are currently under examination and were examined on their merits.
Claim Rejections - 35 USC § 112 (b)
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claims 30-31 are rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
Claims 30 and 31 recite “the method of claim 1” which renders the claim
indefinite for lacking antecedent basis because ‘method' is not recited in claim 1 from which claims 30 and 31 depends. One of ordinary skill in the art would not be able to determine the metes and bounds of the claim, and thus, could not clearly determine how to avoid infringement of claims 30-31.
In the interest of compact prosecution, claims 30-31 are interpreted to the broadest embodiment claimed.
Claim Rejections - 35 USC § 103
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
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 factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
Determining the scope and contents of the prior art.
Ascertaining the differences between the prior art and the claims at issue.
Resolving the level of ordinary skill in the pertinent art.
Considering objective evidence present in the application indicating obviousness or nonobviousness.
Claims 1 and 6-8 are newly rejected under 35 U.S.C. 103 as necessitated by amendment as being unpatentable over Borodina et al. (“Antibiotic Overproduction in Streptomyces coelicolor A3(2) Mediated by Phosphofructokinase Deletion”, published on 09/12/2008, Journal of Biological Chemistry, Vol. 283, No. 37, pages 5186–25199), hereinafter ‘Borodina’, in view of Zeng et al. (“Insights into the simultaneous utilization of glucose and glycerol by Streptomyces albulus M-Z18 for high e-poly-L-lysine productivity”, published on 09/13/2017, Bioprocess Biosyst Eng (2017), Vol. 40, pages 1775-1785), hereinafter ‘Zeng’.
Borodina’s general disclosure relates to the deletion of pfkA2 (SCO5426), one of three annotated pfkA homologues in Streptomyces coelicolor A3(2), which resulted in a higher production of the pigmented antibiotics actinorhodin and undecylprodigiosin (see entire document, including abstract).
Regarding claims 1 and 6, pertaining to a culture, Borodina teaches a culture comprising a culture medium used in culturing under an aerobic condition, wherein the culture medium contains an aerobe that is a bacterium of the genus Streptomyces (“S. coelicolor”; page 25187, right column, paragraph 3; see abstract), a carbon source (“glucose”; page 25188, left column, paragraph 2), and a nutrient other than carbon sources (“NH4Cl“; page 25188, left column, paragraph 2), in an amount sufficient for the aerobe to proliferate under the aerobic environment (page 25188, left column, paragraph 2; page 25192, left column, paragraph 2).
Pertaining to phosphofructokinase, Borodina teaches wherein the Streptomyces has a disrupted gene encoding a metabolic enzyme of glycolysis, wherein the gene is encoding phosphofructokinase (page 25187, left column, paragraph 2; page 25187, right column, paragraphs 2-3), thereby suppressing metabolism from a carbon source to the TCA cycle in the aerobe (page 25192, right column, paragraph 2). It is noted that an increase of glucose-6-phosphate indicates suppressed glycolysis. Borodina teaches the carbon source glucose (page 25188, left column, paragraph 2), and glucose is a substrate and a metabolite for
glycolysis in the upstream of the disrupted phosphofructokinase (see Fig. 4).
Regarding claim 7, Borodina teaches wherein the disrupted gene encoding an enzyme of glycolysis is a gene encoding phosphofructosekinase A2 (page 25187, left column, paragraph 4; page 25187, right column, paragraph 3).
Borodina teaches a Streptomyces having a disrupted gene wherein the gene is encoding phosphofructokinase A3 (page 25187, right column, paragraph 3). Borodina further discloses wherein “Glycolysis is not completely blocked in the pfkA2 mutant, as demonstrated by the metabolic flux analyses, despite our inability to find remaining Pfk activity in ΔpfkA2 cell extracts. The remaining Pfk activity is most likely provided by PfkA3,” (page 25197, right column, paragraph 3).
Additionally, Borodina discloses that “[t]he pfkA2 deletion strain had an increased carbon flux through the pentose phosphate pathway”, that deletion of pfkA2 “resulted in a higher production of the pigmented antibiotics actinorhodin and undecylprodigiosin”, that “Streptomycetes are exploited for production of a wide range of secondary metabolites, and there is much interest in enhancing the level of production of these metabolites”, that “[s]econdary metabolites are synthesized in dedicated biosynthetic routes, but precursors and co-factors are derived from the primary metabolism” (see abstract).
Borodina does not teach
wherein the culture medium contains an energy source in addition to the carbon source, wherein the energy source contains the energy source in an amount sufficient for the aerobe to proliferate under the aerobic environment for 2 times a level of proliferation compared to in the absence of the energy source, wherein the aerobe allows for energy production by metabolism from the energy source to the TCA cycle in the presence of the energy source for the TCA cycle, and the energy source contains a nutrient driving energy production in the electron transport system of the aerobe selected from the group consisting of a metabolite of glycolysis in the downstream of the metabolic enzyme disrupted, a substrate and a metabolite for the metabolic pathway flowing into the downstream of the metabolic enzyme disrupted in glycolysis, a metabolite of the TCA cycle, and a substrate and a metabolite for a metabolic pathway flowing into the TCA cycle (instant claim 1),
wherein the disrupted gene encoding an enzyme of glycolysis is genes encoding phosphofructokinase A2 and A3 (instant claim 7),
wherein the energy source is the elected glycerol (instant claim 8; see above under Claim Status).
Zeng’s general disclosure relates to the simultaneous utilization of glucose
and glycerol by Streptomyces albulus M-Z18 in ɛ-PL batch fermentation (see entire document, including abstract; note, ɛ -Poly-L-lysine (ɛ -PL)).
Regarding claim 1, pertaining to a culture, Zeng teaches a culture comprising a culture medium used in culturing under an aerobic condition, wherein the culture medium contains an aerobe that is a bacterium of the genus Streptomyces (“Streptomyces albulus M-Z18”; page 1776, right column, paragraph 2; see abstract), an energy source (“glycerol”; page 1776, right column, paragraph 2), a carbon source (“glucose”; page 1776, right column, paragraph 2), and a nutrient other than carbon sources (“(NH4)2SO4“; page 1776, right column, paragraph 2), in an amount sufficient for the aerobe to proliferate under the aerobic environment (see increase of dry cell weight (DCW) in the presence of glycerol and glucose (mixed carbon source) in Fig. 1a; page 1781, left column, paragraph 1). It is noted that proliferation of the aerobe is increased in a culture comprising the energy source glycerol and the carbon source glucose, versus a culture comprising only the carbon source glucose (page, 1776, left column, paragraph 1; see Fig. 1a, note the DCW of about 16 g/L (glycerol and glucose) versus a DCW of about 11 g/L (glucose) at about 30h of culturing in Fig. 1a, resulting in an increased proliferation level of about 1.45 in the presence of glucose and glycerol).
Zeng teaches wherein the aerobe allows for energy production by metabolism from the energy source to the TCA cycle in the presence of the energy source for the TCA cycle, and the energy source contains a nutrient driving energy production in the electron transport system of the aerobe consisting of a substrate and a metabolite for a metabolic pathway flowing into the TCA cycle (see Fig. 2; note the pathway of glycerol into the TCA cycle which leads to ATP (energy) production). It is noted that it is within the skill of an ordinary artisan to recognize that ATP is produced via the electron transport system (see Zeng, Fig. 4).
Additionally, Zeng teaches wherein Streptomyces is cultured to produce a secondary metabolite (“utilization of glucose and glycerol by Streptomyces albulus M-Z18 for high e-poly-L-lysine productivity”; see title and abstract).
Regarding claim 8, pertaining to the energy source, Zeng teaches wherein the energy source is glycerol (page 1776, right column, paragraph 2) (instant claim 8).
While Borodina does not teach wherein the culture medium contains an energy source in addition to the carbon source, wherein the energy source contains the energy source in an amount sufficient for the aerobe to proliferate under the aerobic environment for 2 times a level of proliferation compared to in the absence of the energy source, wherein the aerobe allows for energy production by metabolism from the energy source to the TCA cycle in the presence of the energy source for the TCA cycle, and the energy source contains a nutrient driving energy production in the electron transport system of the aerobe selected from the group consisting of a metabolite of glycolysis in the downstream of the metabolic enzyme disrupted, a substrate and a metabolite for the metabolic pathway flowing into the downstream of the metabolic enzyme disrupted in glycolysis, a metabolite of the TCA cycle, and a substrate and a metabolite for a metabolic pathway flowing into the TCA cycle. (instant claim 1), wherein the energy source is the elected glycerol (instant claim 8), it would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have combined Borodina’s culture with Zeng’s teachings on a culture comprising a culture medium containing the energy source glycerol and the carbon source glucose, in order to have created a culture, wherein the culture medium contains the energy source glycerol in addition to the carbon source glucose, wherein the energy source contains the energy source in an amount sufficient for the aerobe to proliferate under the aerobic environment for at a higher level of proliferation compared to in the absence of the energy source, wherein the aerobe allows for energy production by metabolism from the energy source to the TCA cycle in the presence of the energy source for the TCA cycle, and the energy source contains a nutrient driving energy production in the electron transport system of the aerobe selected from the group consisting of a metabolite of glycolysis in the downstream of the metabolic enzyme disrupted, a substrate and a metabolite for the metabolic pathway flowing into the downstream of the metabolic enzyme disrupted in glycolysis, a metabolite of the TCA cycle, and a substrate and a metabolite for a metabolic pathway flowing into the TCA cycle. One would have been motivated to do so to increase cell growth and production of secondary metabolites.
A skilled artisan would have reasonably expected success in combining Borodina’s with Zeng’s teachings since both references are directed to Streptomyces and production of metabolites of interest (see Borodina, abstract, and Zeng, abstract).
While modified Borodina does not teach a proliferation level of 2 times a level of proliferation compared to in the absence of the energy source (instant claim 1), the instantly recited proliferation level would be within the realm of routine experimentation since Zeng teaches an increased proliferation level of about 1.45 in the presence of combined glucose and glycerol, versus only glucose (see Fig.1a; note the DCW (dry cell weight) of about 16 g/L (glycerol and glucose) versus a DCW of about 11 g/L (glucose) at about 30h of culturing in Fig. 1a, resulting in an increased proliferation level of about 1.45 in the presence of glucose and glycerol) . It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have optimized the proliferation level of Streptomyces in order to maximize both cell growth and production of secondary metabolites.
While modified Borodina does not teach wherein the instantly recited proliferation level is achieved by providing the energy source in an amount sufficient to obtain said proliferation level, the energy source amount would have been within the realm of routine experimentation since Zeng teaches providing glycerol in an amount of 30 g/L (page 1776, right column, paragraph 2). It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have determined the optimal amount of energy and carbon source in order to achieve the instantly recited proliferation level for achieving maximal cell growth and production of secondary metabolites.
While modified Borodina does not teach wherein the disrupted gene encoding an enzyme of glycolysis is genes encoding phosphofructokinase A2 and A3 (instant claim 7), It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have combined modified Borodina’s Streptomyces comprising a disrupted gene encoding phosphofructokinase A2 with Borodina’s teachings on the phosphofructokinase activity of phosphofructokinase A3, in order to have created a Streptomyces wherein the disrupted gene encoding an enzyme of glycolysis is genes encoding phosphofructokinase A2 and A3. One would have been motivated to do so, in order to engineer a Streptomyces strain with strongly inhibited glycolysis in order to increase production of secondary metabolites of interest (see Borodina above).
Claims 1, 5, and 30 are newly rejected under 35 U.S.C. 103 as necessitated by amendment as being unpatentable over Borodina et al. (“Antibiotic Overproduction in Streptomyces coelicolor A3(2) Mediated by Phosphofructokinase Deletion”, published on 09/12/2008, Journal of Biological Chemistry, Vol. 283, No. 37, pages 5186–25199), hereinafter ‘Borodina’, in view of Zeng et al. (“Insights into the simultaneous utilization of glucose and glycerol by Streptomyces albulus M-Z18 for high e-poly-L-lysine productivity”, published on 09/13/2017, Bioprocess Biosyst Eng (2017), Vol. 40, pages 1775-1785), hereinafter ‘Zeng’, and in view of Miasnikov et al. (WO 01/53306 A2, published on 07/26/2001), hereinafter ‘Miasnikov’.
Modified Borodina’s teachings have been set forth above.
Modified Borodina does not teach wherein
wherein the aerobe further has a disrupted gene encoding transaldolase (instant claim 5),
wherein the aerobe further has a disrupted gene encoding glucose 6-phosphate isomerase to inhibit production of fructose 6-phosphate (F6P) (instant claim 30).
Miasnikov’s general disclosure relates to methods of manufacturing five-carbon sugars and sugar alcohols as well as other compounds derived from pentose-phosphate pathway from readily available substrates such a hexoses using metabolically engineered microbial hosts. (see entire document, including abstract).
Regarding claim 5, pertaining to a gene encoding a transaldolase, Miasnikov teaches wherein transaldolase results in the production of fructose-6-phosphate (page 13, lines 18-20), and further teaches wherein inactivation of the transaldolase gene leads to decreased carbon flow out of the PPP in the direction of the glycolytic pathway intermediates” (page 15, lines 3-5; note PPP, pentose phosphate pathway).
Regarding claim 30, pertaining to a glucose 6-phosphate isomerase, Miasnikov teaches disrupting glucose 6-phosphate isomerase (page 19, lines 15-18).
Additionally, Miasnikov teaches “disruption in the gene that encodes an enzyme that
regulates the distribution of carbon flow” (page 19, line 6-8).
While modified Borodina does not teach wherein the aerobe further has a disrupted gene encoding transaldolase (instant claim 5), and wherein the aerobe further has a disrupted gene encoding glucose 6-phosphate isomerase to inhibit production of fructose 6-phosphate (F6P) (instant claim 30), it would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have combined modified Borodina’s teachings with Miasnikov’s teachings on transaldolase and glucose 6-phosphate isomerase, to have created a culture wherein the aerobe further has a disrupted gene encoding transaldolase, and wherein the aerobe further has a disrupted gene encoding glucose 6-phosphate isomerase to inhibit production of fructose 6-phosphate (F6P). One would have been motivated to do so in order to further limit glycolysis and thus to maximize the production of secondary metabolites (see Borodina, abstract). A skilled artisan would have reasonably expected success in the combination of modified Borodina’s and Miasnikov’s teachings, since both are directed to increasing metabolites in the pentose phosphate pathway (see Borodina, abstract, and Miasnikov, abstract).
Claims 1, 9-10, and 31 are newly rejected under 35 U.S.C. 103 as necessitated by amendment as being unpatentable over Borodina et al. (“Antibiotic Overproduction in Streptomyces coelicolor A3(2) Mediated by Phosphofructokinase Deletion”, published on 09/12/2008, Journal of Biological Chemistry, Vol. 283, No. 37, pages 5186–25199), hereinafter ‘Borodina’, in view of Zeng et al. (“Insights into the simultaneous utilization of glucose and glycerol by Streptomyces albulus M-Z18 for high e-poly-L-lysine productivity”, published on 09/13/2017, Bioprocess Biosyst Eng (2017), Vol. 40, pages 1775-1785), hereinafter ‘Zeng’, in view of Boisart et al. (WO 2018/138289 A1, published on 08/02/2018), hereinafter ‘Boisart’, and in view of Saier et al. (“A Transport System for Phosphoenolpyruvate, 2-Phosphoglycerate, and 3-Phosphoglycerate in Salmonella typhimurium, published on 07/10/1975, Journal of Biological Chemistry, Vol. 250, No. 13, pages 5089-5096), hereinafter ‘Saier’.
Modified Borodina’s teachings have been set forth above.
Modified Borodina does not teach
wherein the disrupted gene encoding an enzyme of glycolysis is a phosphoglycerate kinase gene (instant claim 9), wherein the energy source is the elected molecule 3-phosphoglycerate (instant claim 10; see above under Claim Status),
wherein the aerobe further has a disrupted gene encoding a phosphoglycerate kinase (instant claim 31).
Boisart’s general disclosure relates to genetically modified microorganism, capable of using carbon dioxide as at least a partial carbon source for the production of molecules of interest (paragraph [0003]).
Regarding claims 9 and 31, pertaining to the disrupted gene, Boisart teaches wherein a disrupted gene for encoding an enzyme for glycolysis is a phosphoglycerate kinase (paragraphs [0021], [0053]). Additionally, Boisart teaches wherein the genetically modified microorganism according to the invention is a Streptomyces (paragraph [0032]).
Saier’s general disclosure relates to “a permease system which allows S. typhimurium to utilize several phosphoglycerates for growth” (see entire document, including page 5094, right column, paragraph 3).
Regarding to claim 10, pertaining to 3-phosphoglycerate as a energy source, Saier teaches bacterial strains with the ability to use 3-phosphoglycerate as an energy source for growth (“3-phosphoglycerate, … as sole source of carbon and energy”; page 5089, left column, paragraph 2 - page 5089, right column, paragraph 1; page 5090, left column, paragraph 2; page 5095, right column, paragraph 1).
While modified Borodina does not teach wherein the disrupted gene encoding an enzyme of glycolysis is a phosphoglycerate kinase gene (instant claim 9), or wherein the aerobe has an additional disrupted gene encoding a phosphoglycerate kinase (instant claim 31), and wherein the energy source is 3-phosphoglycerate (instant claim 10), it would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention, to have modified Borodina’s culture with Boisart’s teachings on a disrupted 3-phosphoglycerate kinase gene and with Saier’s teachings on bacterial strains having the ability to use 3-phosphoglycerate as an energy source, to have created a culture wherein the disrupted gene encoding a glycolysis enzyme, or an additional disrupted gene, is a gene encoding phosphoglycerate kinase, and wherein the energy source is 3-phosphoglycerate. One would have been motivated to do so in order to inhibit, or to further inhibit glycolysis for increased production of secondary metabolites while maintaining energy production by using 3-phosphoglycerate as an energy source (Zeng, Fig. 2, see 3-phosphoglycerate in glycolysis feeding into the TCA; see Borodina, abstract). A skilled artisan would have reasonably expected success in combining modified Borodina’s, Boisart’s, and Saier’s teachings, since all teachings are directed to microbial metabolism.
Response to Arguments
Applicant has traversed the previous rejections of claims under 35 U.S.C. 103. Applicant’s arguments have been considered but are moot in light of Applicant’s amendment filed on 10/20/2025, which introduced the new limitation “Streptomyces” in base claim 1 and thus narrowed the scope of the claim. Borodina is still relied upon in the above rejections, but is not relied upon for teaching ‘culture medium’ recited in claim 1.
In Applicant’s reply, Applicant describes “unexpected results of a culture medium
comprising anaerobe that is a bacterium of the genus Streptomyces” (remarks, page 10).
The Examiner responds that the significance of the impact of culture medium and gene disruptions in the aerobe on cell growth and obtained metabolites is unclear since no statistical evaluation of the presented data in the instant application is provided (see paragraphs [0056] -[0057] and Figures 1-5). Therefore, proper interpretation of the presented results is not possible (see MPEP 716.02(b) I).
Conclusion
No claims are allowed.
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). 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.
Correspondence Information
Any inquiry concerning this communication or earlier communications from the examiner should be directed to SANDRA ZINGARELLI whose telephone number is (703)756-1799. The examiner can normally be reached M-F 9-5.
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/SANDRA ZINGARELLI/ Examiner, Art Unit 1653
/SHARMILA G LANDAU/Supervisory Patent Examiner, Art Unit 1653