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 .
Continued Examination Under 37 CFR 1.114
A request for continued examination under 37 CFR 1.114, including the fee set forth in 37 CFR 1.17(e), was filed in this application after final rejection. Since this application is eligible for continued examination under 37 CFR 1.114, and the fee set forth in 37 CFR 1.17(e) has been timely paid, the finality of the previous Office action has been withdrawn pursuant to 37 CFR 1.114. Applicant's submission filed on 22 January 2026 has been entered.
DETAILED ACTION
The claim amendment on 01/22/2026 does not comply with 37 CFR 1.121(c). Claim 35 has the status identifier of (Previously presented), but the claim text has been amended.
Amendments to the claims filed on or after July 30, 2003 must comply with 37 CFR 1.121(c) which states:
(c) Claims. Amendments to a claim must be made by rewriting the entire claim with all changes (e.g., additions and deletions) as indicated in this subsection, except when the claim is being canceled. Each amendment document that includes a change to an existing claim, cancellation of an existing claim or addition of a new claim, must include a complete listing of all claims ever presented, including the text of all pending and withdrawn claims, in the application. The claim listing, including the text of the claims, in the amendment document will serve to replace all prior versions of the claims, in the application. In the claim listing, the status of every claim must be indicated after its claim number by using one of the following identifiers in a parenthetical expression: (Original), (Currently amended), (Canceled), (Withdrawn), (Previously presented), (New), and (Not entered).
(2) When claim text with markings is required. All claims being currently amended in an amendment paper shall be presented in the claim listing, indicate a status of “currently amended,” and be submitted with markings to indicate the changes that have been made relative to the immediate prior version of the claims. The text of any added subject matter must be shown by underlining the added text. The text of any deleted matter must be shown by strike-through except that double brackets placed before and after the deleted characters may be used to show deletion of five or fewer consecutive characters. The text of any deleted subject matter must be shown by being placed within double brackets if strike-through cannot be easily perceived. Only claims having the status of “currently amended,” or “withdrawn” if also being amended, shall include markings. If a withdrawn claim is currently amended, its status in the claim listing may be identified as “withdrawn—currently amended.”
Claims 1, 4, 6-7, 9, 19, 22, 26, 28, and 35 are amended. Claims 1-2, 4-9, 16, 19-20, 22-28, 35, 38-41, and 48-51 are pending. Claims 38-41 and 48-51 remain withdrawn. Claims 1-2, 4-9, 16, 19-20, 22-28, and 35 are under examination.
Priority
Receipt is acknowledged of the English translation of the certified copies of CN201910411123.7 as required by 37 CFR 1.55.
The effective filing date is now 17 May 2019.
Claim Rejections - 35 USC § 112
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 1-2, 4-9, 16, 19-20, 22-28, and 35 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.
Regarding claims 1 and 22, it is unclear what the limitation “for performing a triacylglycerol decomposition pathway in the Streptomyces during a stationary growth phase” in the last method step is limiting or requiring. It is unclear if the limitation is merely describing that the amplified gene of the Streptomyces (i.e. the amplified gene is used for performing a triacylglycerol decomposition pathway in the Streptomyces), or if a step subsequent to adding a cumate to induce the expression of the amplified gene of the Streptomyces (i.e. performing a triacylglycerol decomposition pathway in the Streptomyces) is required.
Claim 26 recites that the triacylglycerol decomposition pathway is “strengthened”. It is unclear if the term “strengthened” means the triacylglycerol decomposition is increased, enhanced, somehow reinforced, somehow protected from regulation, or if genes associated with triacylglycerol decomposition are overexpressed. There is no special definition provided within the instant specification, and the plain meaning of the term, make or become stronger, does not elucidate the required method steps.
Regarding claims 9 and 28, it is unclear what is the structural limitation corresponding to each of the acyl coenzyme A synthetases, SCO6196, SCO6968, SCO7244, and SCO4383. It is noted that the sequences for these enzymes was added to the sequence disclosure on 22 January 2026. However, the claims lack the appropriate sequence identifiers. The claims are therefore still indefinite because the structures of the claimed SCO6196, SCO6968, SCO7244, and SCO4383 are not provided or limited in the claims.
Claims 2, 4-9, 16, and 19-20 are dependent on claim 1, and claims 23-28 and 35 are dependent on claim 22, and so those dependent claims are indefinite for the same reasons.
Claim Rejections - 35 USC § 103
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
Claims 1-2, 4-8, 16, 19, 22-27, and 35 are rejected under 35 U.S.C. 103 as being unpatentable over Hu et al. (CN 104513840 A published 15 April 2015), Horbal et al. (Dual control system – A novel scaffolding architecture of an inducible regulatory device for the precise regulation of gene expression, Metabolic Engineering 37 (2016) 11–23), and Banchio et al. (A Stationary-Phase Acyl-Coenzyme A Synthetase of Streptomyces coelicolor A3(2) Is Necessary for the Normal Onset of Antibiotic Production, APPLIED AND ENVIRONMENTAL MICROBIOLOGY, Sept. 2002, p. 4240–4246, Vol. 68, No. 9), and as evidenced by LibreTexts (Oxidation of Fatty Acids, https://chem.libretexts.org/@go/page/234043, accessed on 01 April 2025), Yang et al. (Enhancement of acyl-CoA precursor supply for increased avermectin B1a production by engineering meilingmycin polyketide synthase and key primary metabolic pathway genes, Microbial Biotechnology. 2024;17:e14470.), and Gibson et al. (Enzymatic Assembly of Overlapping DNA Fragments, Methods in Enzymology, Volume 498, 2011).
The limitation “…for performing a triacylglycerol decomposition pathway in a Streptomyces during a stationary growth phase” recited in claim 1 is an intended result or effect of practicing the method, but does not recite an active step to execute the method for improving the production of a polyketide compound in a Streptomyces. As such, where the prior art teaches all of the steps of claim 1, the limitation is considered to be necessarily met.
The limitation “…to obtain a plasmid triacylglycerol degradation regulation module having a sequence as set forth in SEQ ID NO: 35…” recited in claim 1 is an intended result of practicing the method steps prior to the limitation, but does not recite an active step to execute the claimed method. Practicing the amplifying steps for the cumate inducible promoter and Streptomyces gene, and the Gibson assembly step results in the plasmid triacylglycerol degradation regulation module having a sequence as set forth in SEQ ID NO: 35. Thus, where the art teaches the amplifying steps for the cumate inducible promoter and Streptomyces gene, and the Gibson assembly step of claim 1, the limitation is considered to be necessarily met.
Regarding claim 1, Hu teaches a method for improving yield of polyketide compounds in engineered Streptomyces hygroscopius (Hu Abstract and pg. 3 para. 6) comprising the steps of amplifying a promoter ERM*E using primers P3 and P4 to obtain a fragment containing the ERM*E promoter and amplifying acyl coenzyme A DPTE using primers P1 and P2 to obtain a fragment containing the complete DPTE gene (Hu pg. 4 Embodiment 2 steps 1 and 2). The ERM*E promoter and DPTE fragments were assembled into plasmid pSET152 using the Gibson assembly method of digesting the ERM*E fragment with restriction enzymes BamHI and NdeI, digesting the DPTE fragment with NdeI and XbaI, digesting the pSET152 plasmid with BamHI and XbaI, and the ligating all of the fragments together using T4 DNA ligase to obtain a plasmid pSET152erm*E-DPTEF (Hu fig. 1 and pg. 4 last para. through pg. 5 para. 1). The pSET152erm*E-DPTEF plasmid was then integrated into the genome of actinomycete Streptomyces hygroscopicus by intergenic conjugation through E. coli (Hu pg. 3 para. 6 and pg. 5 Example 3).
Hu does not teach amplifying a cumate inducible promoter using primers CuF and CuR or amplifying a gene of Streptomyces using primers 6196F and 6196R.
Horbal teaches a library of cumate inducible promoters which can be used to modulate gene expression of secondary metabolite gene clusters (Horbal pg. 14 Results section 3.1). Horbal also teaches inducing the promoters with different concentrations of cumate (denoted Cu) in order to induce the expression of the genes following the promoters (Horbal Fig. 1).
Banchio teaches Streptomyces coelicolor acyl coenzyme A synthetase encoded by the fadD1 gene, also called the sco6196 gene, as evidenced by Yang (Yang pg. 8 right col. para. 1), modulates the activation of polyketide actinorhodin synthesis (Banchio discussion).
Hu, Horbal, and Banchio do not teach the primers CuF, CuR, 6196F, and 6196R.
However, it would have been a matter of routine optimization by one of ordinary skill in the art to determine adequate primer sequences in order to amplify Horbal’s cumate inducible promoter and Banchio’s fadD1 (sco6196) acyl coenzyme A synthetase gene. In depth instructions on how to design effective primers for amplifying DNA fragments for use in Gibson assembly are readily available and well known in the art, as evidenced by Gibson (Gibson pgs. 352-353 sec. 2 and Fig. 15.1). By referencing the DNA sequences of the cumate inducible promoter as disclosed by Horbal and fadD1 (sco6196) acyl coenzyme A synthetase gene as disclosed by Bachio, one of ordinary skill in the art would use the readily available guidance of Gibson Assembly method as disclosed by Hu to construct PCR primers that would predictably amplify the cumate inducible promoter and fadD1 (sco6196) acyl coenzyme A synthetase gene fragments, and prepare those fragments for Gibson assembly.
It would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date of the present invention to modify Hu’s method of improving polyketide production in a Streptomyces by substituting the ERM*E promoter used in Hu’s method with Horbal’s cumate inducible promoter, and Hu’s Streptomyces gene DPTE with Banchio’s Streptomyces gene fadD1 (sco6196), thereby obtaining a triacylglycerol degradation regulation module.
One of ordinary skill in the art would have been motivated to substitute Hu’s ERM*E promoter with Horbal’s cumate inducible promoter in order to selectively control the transcription of the acyl coenzyme A synthetase gene using cumate. One of ordinary skill in the art would have a reasonable expectation of success because Horbal taught the effectiveness of a cumate inducible promoter controlling expression of downstream genes upon addition of cumate (Horbal fig. 2b).
One of ordinary skill in the art would have been motivated to substitute Hu’s acyl coenzyme A synthetase gene DPTE with Banchio’s acyl coenzyme A synthetase gene fadD1 (sco6196) in order to overexpress the transcription levels of FadD1 (SCO6196) in the Streptomyces to improve its polyketide production. One of ordinary skill in the art would have a reasonable expectation of success because Hu’s method is taught to improve the production of polyketides in a Streptomyces species by overproducing the expression of an acyl coenzyme A synthetase, and Hu’s DPTE and Banchio’s FadD1 (SCO6196) are both acyl coenzyme A synthetases produced by Streptomyces species.
Regarding claim 22, Hu teaches that polyketides are secondary metabolites (Hu pg. 2 Background), and Banchio teaches that the polyketides are synthesized during the stationary phase of the Streptomyces (Banchio Fig. 6). The obvious method of Hu in view of Horbal and Banchio improves the production of polyketides by inducing selective control over the expression of acyl coenzyme A synthetase SCO6196, which is normally expressed in the stationary phase when the secondary metabolism naturally begins and is correlated with SCO6196 levels (Banchio Figs. 3A and 6). Inducing the expression of acyl coenzyme A synthetase SCO6196 leads to increased secondary metabolites like polyketides, thereby switching the primary metabolism of Streptomyces to its secondary metabolism where the polyketides are synthesized. Therefore, performing the obvious method of Hu in view of Horbal and Banchio switches the primary metabolism of a Streptomyces to its secondary metabolism.
Regarding claims 2 and 23, acyl-coenzyme A synthetase is an enzymatic component that is part of the fatty acid (such as triacylglycerol) catabolism through β-oxidation, as evidenced by LibreTexts (Pg. 1 para. 2 and chemical equation).
Regarding claim 4, Hu teaches their method can improve the yield of polyketide avermectin (Hu pg. 2 Background).
Regarding claims 6 and 25, Hu teaches a method for improving yield of polyketide compounds in engineered Streptomyces hygroscopius (Hu Absract and pg. 3 para. 6).
Regarding claims 5, 7-8, 16, 19, 24, 26-27, and 35, these claims further limit the phrase “for performing a triacylglycerol decomposition pathway in the Streptomyces” recited in claim 1. This phrase is an intended result or effect of practicing the method, and not an active step to be performed in order to execute the method. Since Hu teaches a method for improving yield of polyketide compounds in engineered Streptomyces hygroscopius comprising the steps of claims 1 and 22, the limitations of these claims are considered necessarily met.
Claims 9 and 28 is rejected under 35 U.S.C. 103 as being unpatentable over Hu in view of Horbal and Banchio as applied to claims 1-2, 4-8, 16, 19, 22-27, and 35 above, and further in view of Risdian et al. (Biosynthesis of Polyketides in Streptomyces, Microorganisms 2019, 7, 124; doi:10.3390/microorganisms70501 24).
The obvious method of Hu in view of Horbal and Banchio enhances the expression level of the acyl coenzyme A synthetase enzyme SCO6196. However, Hu, Horbal, and Banchio do not teach the improvement in yield of polyketide compounds in the species Streptomyces coelicolor.
Risdian teaches that Streptomyces coelicolor produces polyketides actinorhodin (Risdian pg. 2-3 bridging para.) and germicidin A (Risdian pg. 11 sec. 4.1 para. 1). Risdian teaches that germicidin A is known to inhibit spore germination and be antibacterial against various Gram-positive bacteria (Risdian pg. 11 sec. 4.1 para. 1).
It would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date of the present invention to apply the obvious method of Hu in view of Horbal and Banchio to the Risdian’s Streptomycin coelicolor in order to advantageously enhance production of antibacterial and anti-spore polyketide compound germicidin A produced by Risdian’s Streptomyces coelicolor. One of ordinary skill in the art would have a reasonable expectation of success because Hu taught their method of improving polyketide yield to be useful in engineered Streptomyces cells, and so one of ordinary skill in the art would reasonably expect the method of Hu in view of Horbal and Banchio to successfully work in Streptomyces coelicolor.
Claim 20 is rejected under 35 U.S.C. 103 as being unpatentable over Hu in view of Horbal and Banchio as applied to claims 1-2, 4-8, 16, 19, 22-27, and 35 above, and further in view of Zhang et al. (Suitable extracellular oxidoreduction potential inhibit rex regulation and effect central carbon and energy metabolism in Saccharopolyspora spinosa, Microbial Cell Factories 2014, 13:98).
Hu, Horbal, and Banchio do not teach a step of increasing a NADH/NAD+ ratio in the Streptomyces.
Zhang teaches that polyketides are synthesized in the stationary growth phase of fermentation, and that the ratio of NADH/NAD+ has significant influence on polyketide production (Zhang Abstract). Zhang also teaches that the NADH/NAD+ ratio is significantly higher in stationary growth phase than in either lag or exponential growth phases under normal conditions (Zhang Fig. 2 control group). Therefore, Zhang teaches that a high NADH/NAD+ ratio is present in stationary growth phase, which allows for the production of polyketides secondary metabolites.
It would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date of the present invention to increase the ratio of NADH/NAD+ of the Streptomyces of Hu in view of Horbal and Banchio. One of ordinary skill in the art would have been motivated to do so in order to advantageously promote the secondary metabolism conditions which positively impact therapeutic polyketide production under normal growth conditions, such as a high NADH/NAD+ ratio. One of ordinary skill in the art would have had reasonable expectations of success in this endeavor because Zhang teaches that polyketides are synthesized in the stationary growth phase, the ratio of NADH/NAD+ has significant influence on polyketide production, and the NADH/NAD+ ratio is significantly higher in stationary growth phase than in either lag or exponential growth phases under normal conditions. Thus, a high NADH/NAD+ ratio is present in stationary growth phase, which enhances the production of therapeutic polyketides secondary metabolites.
Response to Arguments
Applicant's arguments filed 22 January 2026 have been fully considered but they are not persuasive. Applicant argues that that none of the cited references disclose the use of the specific sequences recited in the claims, nor the step of adding cumate to induce expression of the amplified gene of the Streptomyces (Remarks pg. 36).
In response, it is noted that the specific sequences of the promoters are not sufficient to differentiate the instant invention from the prior art because it would have been a matter of routine optimization by one of ordinary skill in the art to determine adequate primer sequences in order to amplify Horbal’s cumate inducible promoter and Banchio’s fadD1 (sco6196) acyl coenzyme A synthetase gene. In depth instructions on how to design effective primers for amplifying DNA fragments for use in Gibson assembly are readily available and well known in the art, as evidenced by Gibson (Gibson pgs. 352-353 sec. 2 and Fig. 15.1). By referencing the DNA sequences of the cumate inducible promoter as disclosed by Horbal and fadD1 (sco6196) acyl coenzyme A synthetase gene as disclosed by Bachio, one of ordinary skill in the art would use the readily available guidance of Gibson Assembly method as disclosed by Hu to construct PCR primers that would predictably amplify the cumate inducible promoter and fadD1 (sco6196) acyl coenzyme A synthetase gene fragments, and prepare those fragments for Gibson assembly.
The specific sequence of the triacylglycerol degradation regulation module set forth in SEQ ID NO: 35 is the result of practicing the method steps prior to the limitation. Practicing the amplifying steps for the cumate inducible promoter and Streptomyces gene, and practicing the Gibson assembly step results in the triacylglycerol degradation regulation module having a sequence as set forth in SEQ ID NO: 35. Thus, since Hu in view of Horbal and Banchio teach the amplifying and assembly steps that produce the triacylglycerol degradation regulation module, the limitation is considered to be necessarily met.
Horbal teaches the addition of a variety of different concentrations of cumate to a cumate-inducible promoter to induce expression of the gene following the promoter, as shown in Horbal’s Fig. 1. Thus, when considered in combination with Hu and Banchio, it would have been obvious to one of ordinary skill in the art to add cumate to Horbal’s cumate-inducible promoter in order to induce expression of Banchio’s acyl coenzyme A synthetase gene fadD1 (sco6196).
Conclusion
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/LOUISE W HUMPHREY/ Supervisory Patent Examiner, Art Unit 1657
/Alexander M Duryee/Examiner, Art Unit 1657
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