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 .
Priority
This application is a 371 of PCT/JP2020/037902 filed 10/06/2020. Applicant’s claim for the benefit of a prior-filed application under 35 U.S.C. 119(e) or under 35 U.S.C. 120, 121, 365(c), or 386(c) is acknowledged.
Acknowledgment is made of applicant’s claim for foreign priority under 35 U.S.C. 119 (a)-(d) based on JP 2019-187501 filed 10/11/2019. Receipt is acknowledged of certified copies of papers required by 37 CFR 1.55.
Status of the Claims
Claim 10 is amended. Claim 11 is cancelled. Claims 10, 12 and 13 are pending (claim set filed 12/15/2025) and are examined on the merits herein.
Withdrawal of Rejections
The response and amendment filed on 12/15/2025 are acknowledged. All of the amendment and arguments have been thoroughly reviewed and considered.
For the purposes of clarity of the record, the reasons for the Examiner's withdrawal and/or maintaining if applicable, of the substantive or essential claim rejections are detailed directly below and/or in the Examiner's response to arguments section.
Maintained/Modified Rejections
The following rejections are maintained and/or modified taking into consideration amendment to claim 10 filed on 12/15/2025.
Claim Rejections - 35 USC § 103
The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action.
Claims 10, 12 and 13 are rejected under 35 U.S.C. 103 as being unpatentable over Ohtsu (Ohtsu et al. BBRC, 1967, 29, 635-641 on record in IDS) in view of Maeno (Maeno et al. Research in Microbiology, 2019, 170, 35-42), Marinescu (Marinescu et al. Scientific Reports, 2018, 8, 12278, 1-11) and Brenner (US 20090202680 A1) and as evidenced by NBRC Culture Catalogue (NBRC Culture Catalogue, [retrieved on 07/25/2024]. Retrieved from the Internet: <https://www.nite.go.jp/nbrc/catalogue/NBRCCatalogueDetailServlet?ID=NBRC&CAT=00003516>) and Kodama (Kodama J. Agr. Chem. Soc. Japan, 1956, 30, 219-224).
Regarding claims 10, 12 and 13, Ohtsu teaches that Lactobacillus fructosus which was renamed to Fructobacillus fructosus, as evidenced by NBRC Culture Catalogue (p.1, Synonymous Name), possesses NAD biosynthesis pathway and contains nicotinamide mononucleotide (NMN) and NAD pyrophosphorylases (Title, p. 637, 2nd paragraph, Table 1). Ohtsu showed the crude extracts of cultured L. fructosus to have NMN and NAD pyrophosphorylase activities (p. 636, Table 1). NMN pyrophosphorylase was purified from L. fructosus by Ohtsu and was shown to catalyze conversion of nicotinamide to NMN in the presence of PP-ribose-P and ATP since the product of reaction was isolated from the incubation mixture and identified as NMN (p. 637, 2nd paragraph). These findings indicate production of NMN by culturing Fructobacillus bacteria. The culturing of L. fructosus was performed for 24 hours at 26°C (p. 636, 1st paragraph) that reads on claim 10 limitation of 8 to 36 hours culturing. Ohtsu teaches L. fructosus strain identified by Kodama (p. 635, p. 641, Reference: Kodama J. Agr. Chem. Soc. Japan, 30, 219 (1956)). Kodama identified that strain as strain 353 as evidenced by Kodama (Kodama J. Agr. Chem. Soc. Japan, 1956, 30, 219-224). That strain is the same as strain NBRC3516 as evidenced by NBRC Culture Catalogue (p. 1, Other No.). Ohtsu teaches the same process step of Fructobacillus culturing, the same time period of culturing and the same strain NBRC3516 as claimed and thus it will inherently produce NMN and nicotinamide riboside (NR).
Ohtsu does not teach culturing Fructobacillus at 28°C to 36°C, culturing in a fructose containing culture medium and isolating NMN and NR from the bacterial cell bodies.
Regarding claim 10, Maeno teaches that Fructobacillus bacteria prefer D-fructose over D-glucose as the main carbon source (Abstract). Maeno mentions that these bacteria are found in the environment rich in D-fructose, such as flowers and fruits (Abstract). Additionally, Maeno describes culturing Fructobacillus in a medium containing fructose at 30°C (p. 36, left column, 4th paragraph).
Regarding claim 10, Marinescu teaches production of NMN in Escherichia coli genetically modified to express nicotinamide phosphoribosyl transferase (Nampt) and phosphoribosyl pyrophosphate (PRPP) synthetase from other bacteria (Abstract). Nampt catalyzes conversion of nicotinamide to NMN in the presence of phosphoribosyl pyrophosphate (p. 1, 2nd paragraph). The enzyme catalyzing the same reaction was identified by Ohtsu in F. fructosus and called NMN pyrophosphorylase (p. 637, last paragraph) as described above. Marinescu discloses production of NMN in bacterial cells for up-to 23.57 mM (or 18.7 mg per gram of protein) (p. 3, last paragraph, Figure 3C,D). Marinescu measured the amount of NMN exported to the medium or leaked from the dead cells and its concentration was very low (below 1 x 10-2 mM (p. 3, last paragraph, Fig. 3A,B). Marinescu mentions developed method of purification of NMN from bacterial cells (p. 2, 2nd paragraph).
Regarding claim 10, Brenner teaches production of nicotinamide riboside (NR) by fungal strain deficient in import of NR (Abstract). Since the strain cannot import NR, NR is isolated from the medium by solubilizing NR with methanol and subjecting to column chromatography, i.e. SP-SEPHADEX chromatography, as described in the working example (paragraphs 0007 and 0053). Brenner mentions other alternative methods for NR isolation, such as solid phase extraction and porous graphitic carbon and hydrophilic interaction chromatography (paragraph 0019).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to follow Maeno guidance and use fructose-containing medium and 30°C for culturing Fructobacillus in Ohtsu teaching. One would have been motivated to do that because Maeno teaches that Fructobacillus bacteria prefer D-fructose over D-glucose as the main carbon source and hence it would provide more favorable conditions for Fructobacillus culturing. One would be motivated to try culturing temperature 30°C to achieve more efficient production of nicotinamide mononucleotide and nicotinamide riboside. A skilled artisan would have reasonably expected success in this combination since both Ohtsu and Maeno teach culturing Fructobacillus bacteria and it is within the skill of the artisan in the field to optimize culturing conditions.
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to follow Marinescu teaching and isolate NMN and NR produced based on Ohtsu and Maeno teachings from the bacterial cells. One would be motivated to do so since Marinescu teaches production of NMN in E. coli expressing Nampt and the same enzyme is endogenously present in F. fructosus based on Ohtsu teaching and Marinescu confirmed production of NMN mostly in bacterial cells with very low secretion in the medium. Additionally, Marinescu mentions the method of isolation of NMN from bacterial cells. A skilled artisan would have reasonably expected success in this combination because Ohtsu and Marinescu teach production of NMN in bacteria and Ohtsu and Maeno teach culturing F. fructosus.
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to apply method of isolation of NR from medium of NR producing cells taught by Brenner for isolation of NR from bacterial cells bodies producing NMN and NR based on Ohtsu, Maeno, Marinescu teachings. One would be motivated to do so with reasonably expected success since Brenner suggests different techniques for NR purification that can be applied to cell lysate. A skilled artisan would have reasonably expected success in this combination because Ohtsu provides Fructobacillus fructosus for NMN and NR production, Maeno describes the optimal carbon source for Fructobacillus, Marinescu points to bacterial cell bodies for isolation of NMN and NR and Marinescu and Brenner provide methods of NMN and NR isolation.
Thus, combination of teachings of Ohtsu, Maeno, Marinescu and Brenner as evidenced by Kodama and NBRC Culture Catalogue renders claims 10, 12 and 13 obvious.
Response to Arguments
Applicant's arguments filed 12/15/2025 have been fully considered but they are not persuasive.
Applicant argues (addressing p. 3-4 of the Remarks) that the prior art of Ohtsu and Maeno does not teach selecting bacterial cell bodies for recovery of NMN and NR as recited in amended claim 10. Applicant refers to Table 2 showing that in fructose-containing medium NMN and NR were found specifically in bacterial cell bodies and were not detected in the supernatant while in fructose-free medium (Table 1) NMN and NR were not detected in the cells. Applicant further argues that the unique and surprising result that NMN and NR are obtained within bacterial cells only when fructose-containing medium was used was not predicted by prior art. These arguments are not persuasive because:
Current rejection is based on combination of prior art of Ohtsu, Maeno, Marinescu and Brenner as described above. Although Ohtsu does not explicitly teach production of NMN in cells of F. fructosus, Marinescu teaches E. coli expressing the same enzyme, Nampt, as taught by Ohtsu (though named at that time as NMN pyrophosphorylase), catalyzing the same reaction of conversion of nicotinamide to NMN and producing NMN preferentially in the cell bodies as opposite to secretion to medium (Abstract, p. 3, last paragraph). Maeno provides the optimal medium for Fructobacillus, i.e. containing fructose as carbon source (Abstract) and describes that F. fructosus strain metabolized fructose within 1 day compared to 5 days for D-glucose (p. 39, left column). Therefore, it is expected that F. fructosus with endogenous Nampt will produce NMN in the cell bodies in metabolically favorable for Fructobacillus conditions. Additionally, Ohtsu teaches the same strain of F. fructosus as instant strain as described in the rejection and evidenced by Kodama and NBRC Culture Catalogue. MPEP 2112.01: “Products of identical chemical composition cannot have mutually exclusive properties." In re Spada, 911 F.2d 705, 709, 15 USPQ2d 1655, 1658 (Fed. Cir. 1990). A chemical composition and its properties are inseparable. Therefore, if the prior art teaches the identical chemical structure, the properties applicant discloses and/or claims are necessarily present.” Thus, the F. fructosus taught by Ohtsu will necessarily have the same properties of NMN and NR production in the cell bodies when grown on fructose.
Assuming arguendo applicant has shown surprising and unexpected data, claims are not commensurate in scope with the unexpected results. MPEP 716.02: “Whether the unexpected results are the result of unexpectedly improved results or a property not taught by the prior art, the "objective evidence of nonobviousness must be commensurate in scope with the claims which the evidence is offered to support." In instant case, claim 10 has a limitation for the culture temperature of 28-36°C, culturing period of 8-36 hours and fructose-containing medium. The specification provides Example 2 for production of NR and NMN by culturing NBRC3516 strain and the culturing conditions are very specific, i.e. temperature - 30°C, duration - 12 hours, culturing type – static culture, MRS medium and single and specific fructose concentration in the medium, i.e. 2 w/w% (paragraph 0066, 0073-0075). Thus, it was not shown that the unexpected results occurred over the entire claimed range and hence claim 10 is not commensurate in scope with the unexpected results.
Therefore, the 35 U.S.C. 103 rejection is maintained and modified necessitated by amendment of claim 10.
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.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to LIOUBOV G KOROTCHKINA whose telephone number is (571)270-0911. The examiner can normally be reached Monday-Friday: 8:00-5:30.
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If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Sharmila G Landau can be reached at (571)272-0614. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
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/L.G.K./Examiner, Art Unit 1653
/SHARMILA G LANDAU/Supervisory Patent Examiner, Art Unit 1653