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
Receipt is acknowledged of certified copies of papers required by 37 CFR 1.55.
Claims 1-15, 17, 18 are pending and have been considered on the merits herein.
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 4 and 12 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.
Claim 4 recites the limitation "wherein in purification steps b) to e)…". There is insufficient antecedent basis for this limitation in the claim. The steps ((b) to e)) are claimed to be production process steps in claim 4.
Additionally, the claim refers to “wherein said production process comprises the steps of…a)-e)”. It is suggested applicants amend the claim to read “wherein purification further comprises the steps of…” to more distinctly claim (and further add to) the steps of the purification process as seen in claim 1.
Claim 12 recites the limitation "purified O-EPA conjugate drug substances" in claim 4. There is insufficient antecedent basis for this limitation in the claim.
The claim should depend from claim 11 which recites obtaining a purified O-EPA conjugate as a pharmaceutical drug substance.
Claim Rejections - 35 USC § 103
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
Claim(s) 1, 2, 4-13, 15, 18 is/are rejected under 35 U.S.C. 103 as being unpatentable over van den Dobbelsteen et al. (Vaccine, 2016, IDS) in view of WO2014110246 and WO2020191082 (IDS) supported by Sarawat et al. (Biomed. Res. Int., 2013, IDS) and Hong et al. (J. Liq. Chrom.& related tech., vol. 35, p. 2923-2950, 2012).
Regarding claim 1, Van den Dobbelsteen teach a process for production of a purified O-polysaccharide -ExoProtein A carrier (O-EPA) conjugate comprising obtaining the O-EPA conjugate as a bioconjugate obtained from prokaryotic host cells incubated in a bioreactor, i.e. ExPEC-4V which is composed of the O1, O2, O6 and O25 E. coli antigens coupled to a detoxified variant of Exotoxin A from Pseudomonas aeruginosa (EPA) (p. 4153, 1st whole col. Including section 2.1 and 2.2),
and purification of the conjugate comprising
providing a periplasmic fraction of prokaryotic host cells that express the bioconjugate (p. 4153, section 2.1. and 2.2),
subjecting the periplasmic fraction to a first anion exchange chromatography (AEX1), i.e. enriched and purified over two anion exchange chromatography (AEC) steps using Q Ceramic HyperD® and Source15Q resins where the product is captured on the first Q Ceramic HyperD® column, buffer adjusted and pooled product fractions were loaded onto the hydrophobic interaction chromatography (HIC) matrix (p. 4153, section 2.2),
followed by a hydrophobic interaction chromatography (HIC), i.e. buffer adjusted and pooled product fractions were loaded onto the hydrophobic interaction chromatography (HIC) matrix (p. 4153, section 2.2),
and followed by a second anion exchange chromatography (AEX2), i.e. products from the HIC are purified and polished with a second AEC step (Source 15Q) and concentrated by tangential flow filtration (TFF) (section 2.2).
Regarding claims 4 and 7, van denDobbelsteen teach the production process to comprise the steps of
a) providing a filtered periplasmic fraction of prokaryotic host cells comprising the O-EPA conjugate, i.e. the bioconjugate product is pre-purified using negative mode AEC…filter cartridge (section 2.2).
b) subjecting an adjusted load of the filtered periplasmic fraction to a first anion exchange chromatography (AEX 1) step to obtain a first AEX eluate (AEX1) , i.e. the bioconjugate product is pre-purified using negative mode AEC…filter cartridge and subsequently captured on the first AEC column (section 2.2) and thus is taken to be an adjusted load suitable for binding to the AEX medium according to applicants’ definition on p. 8 of the specification “adjusting a load of a process intermediate to conditions suitable for applying said process intermediate on a chromatography resin”, and
c) subjecting an adjusted load eluate to a hydrophobic interaction chromatography (HIC) step to obtain a HIC eluate i.e. buffer adjusted and pooled product fractions (from the first AEC column) were loaded onto the hydrophobic interaction chromatography (HIC) matrix (p. 4153, section 2.2), and
d) subjecting the HIC eluate obtained in step (c) to a second anion exchange chromatography (AEX2) step to obtain a second AEX eluate as product, i.e. products from the HIC are purified and polished with a second AEC step (Source 15Q) and concentrated by tangential flow filtration (TFF) (section 2.2),
and adjusting conditions to allow binding of the O-EPA conjugate to a chromatography medium and subsequently adjusted to allow elution of the O-EPA conjugate from said medium (section 2.2).
Regarding claim 5, the O-polysaccharide is selected from Escherichia (E. coli), (section 2.1, Discussion section 4, 1st 2 parags.).
Regarding claim 6, the prokaryotic host cells from which the O-EPA conjugate is obtained comprise genetic information encoding an O-polysaccharide and a recombinant ExoProtein A (EPA) and a metabolic apparatus that carries out N- glycosylation of the EPA with the O-polysaccharide thereby producing the O-EPA conjugate in vivo in the periplasm of the prokaryotic host cells, i.e. “E. coli produces two specific surface polysaccharides: the lipopolysaccharide (LPS) O antigen (p. 4152, last parag.) “A recent technology called in vivo bioconjugation allows the biosynthesis of polysaccharide and carrier protein within E. coli cells, and their subsequent in vivo coupling using a specific oligosaccharyl transferase from the N-linked protein glycosylation system. This process allows in vivo conjugation of O-polysaccharides to specific consensus sites for N-glycosylation of any carrier protein and removes the requirement for chemical detoxification of LPS and further conjugation processes, in a cost-effective manner. We investigated the immunogenicity and safety of a 4-valent ExPEC vaccine (ExPEC-4V) produced for the first time by the process of bioconjugation. ExPEC-4V is composed of the 01, 02, 06 and 025 antigens coupled to a detoxified variant of Exotoxin A from Pseudomonas aeruginosa (EPA)” (p. 4153, parag. 1-3, and section 2.1 teaching production of the ExPEC-4V bioconjugate using engineered E. coli).
Regarding claim 11, the AEX eluate is formulated into Tris-buffered saline at pH 7.4, thus a pharmaceutically acceptable buffer, to obtain the ExPEC-4V pharmaceutical drug substance (section 2.2, 2nd to last parag.).
Regarding claims 12 and 18, the reference teaches combining several purified O-polysaccharides to produce a multivalent drug product, i.e. ExPEC-4V (4-valent) containing O25, O1, O2 and O6 E. coli O-serotypes (p. 4153, sections 2.3.3, section 3.2, 3.4, Fig. 1, p. 4158, 2nd col, 2-4th parag.).
Regarding claim 15, van denDobbelsteen teach releasing the O-EPA conjugate from the periplasm by osmotic shock treatment (section 2.2).
Van denDobbelsteen does not teach a hydroxyapatite chromatography step according to claim 1.
WO2014110246 teaches recombinantly produced protein purification (from cell culture) methods comprising a combination of anionic exchange chromatography, followed by mixed-mode hydroxyapatite HA chromatography, followed by hydrophobic interaction chromatography (HIC) and tangential flow ultrafiltration with an intervening viral filtration step resulting in a pharmaceutically acceptable drug substance into a final formulation buffer (0006, 0007, 0009, 0012-0018, 0022, 0032, 0108, 0111, 0257). The method involves pooling eluate from the chromatography steps and adjusting the pH of the pooled eluate (0006). The reference teaches that the method allows for simple and effective purification methods to facilitate large scale production of proteins (0006).
Regarding claim 13, WO246 teaches using a very high salt concentration for eluting contaminants and polypeptides from the exchange resin (0138) and teach that the protein is loaded onto the anionic exchange resin using a loading buffer at a salt concentration and pH so that the polypeptide and contaminant bind to the anion exchange resin, which is further washed with column volumes of loading buffer followed by one or more column volumes of buffer wherein the salt concentration is increased and finally eluted with an elution buffer of increasing salt concentration (0142). The reference teaches that the salt concentration is the mobile phase is a linear gradient in which the salt concentration is increased stepwise (0143).
Regarding claims 1, 4, 7-10, anion exchange chromatography is followed by mixed-mode HA chromatography comprising washing steps to eluate the protein, i.e. adjust load of the eluate (0169, 0174, 0175, 0179, 0180, 0264). Regarding step c) of claim 4, the AEX1 eluate is adjusted and subsequently purified using the HA chromatography step (0264, 0265). The reference teaches that purity following mixed mode HA chromatography greatly improves (0182). Regarding applicants’ limitations drawn to steps b-1) of claim 7, c-3) of claim 8, d-6) of claim 9, e-10) of claim 10, the reference teaches at each step of AEX1 (0143, for example), HA (0171, for example), HIC (0185, 0186, for example) using loading buffers suitable for binding to the medium, which according to applicants definition of adjusting conductivity (see p. 28, lines 14-20) is how conductivity is adjusted.
HA chromatography is followed by hydrophobic interaction chromatography (HIC) utilizing the attraction of a molecule for a polar or non-polar environment, i.e. the hydrophobicity or hydrophilicity of residues on the exposed, outer surface of a protein. Thus, proteins are fractionated based upon their varying degrees of attraction to a hydrophobic matrix (0184). Regarding step d) of claim 4, the HA eluate is subsequently adjusted and loaded onto the HIC column, washed two or more times and eluted from the column (0185, 0188, 0191, 0267, 0268). Purity following HIC is improved (0194).
Regarding claim 17, the HIC eluate is subjected to an additional chromatography step, which is then followed by one or more downstream ultrafiltration steps, including tangential flow ultrafiltration using the eluate from the chromatography steps as the starting material (0196, 0202) followed by a viral inactivation filtration step (0275, 0276). The reference additionally teaches that the filtration and viral inactivation may be implemented during the purification process (Ex. 2, 0246-0249, 0257, 0273, 0275).
Thus, before the effective filing date of the claimed invention, processes for producing a purified O-EPA conjugate comprising providing a periplasmic fraction of a prokaryotic host cell expressing a bioconjugate to two anion exchange chromatography (AEX) steps and HIC was known. The art teaches that the bioconjugates can be purified using any protein purification methods known in the art to include chromatography steps and TFF. Secondary reference, WO246 teaches a recombinant protein purification process which includes a first anion exchange chromatography (AEX), followed by HA chromatography, followed by HIC, a second chromatography step, followed by TFF and viral inactivation filtration, in which with each additional step resulting in increased purity of the drug product. Therefore, all the claimed elements were known in the prior art and one skilled in the art could have combined the elements as claimed by known methods with no change in their respective functions and the combination would have yielded predictable results to one of ordinary skill in the art at the time of the invention. The combination of chromatography steps and filtration steps results in a product of higher purity and thus, these purification steps were recognized as part of the ordinary capabilities of a posita and thus a posita would have been capable of applying these techniques with a reasonable expectation of successfully purifying the O-EPA conjugate.
The references do not teach cells incubated in a bioreactor with a volume between 100-20000L according to claim 1, and 150L and 5000L according to claim 3.
WO2020191082 teaches a process for producing a purified O-EPA bioconjugate obtained from a prokaryotic cell, i.e. E. coli polysaccharide conjugated to carrier protein exotoxin A of P. aeruginosa (EPA) (0010-0015, 00153-00156, 00178), wherein the O-EPA is a conjugate drug product composition which is a multivalent vaccine comprising at least E. coli antigen polysaccharides O25B, O1A, O2 and O6A (0017-0019, 00119). The bioconjugates can be purified by any known method in the art for purification of a protein including anion exchange chromatography or other standard techniques for purification of proteins (00196). WO082 teaches bioconjugate production by culturing cells in bioreactors of 200L volume (00315).
Therefore, large scale production and purification methods were known in the art before the effective filing date of the claimed invention and it would have been obvious to one of ordinary skill in the art to apply known production and purification methods to large scale production with a reasonable expectation of successfully producing and purifying the O-EPA conjugate.
Regarding the limitation of claim 1 drawn to “wherein said purification does not comprise a size-exclusion chromatography (SEC) step”. While van denDobbelsteen teach an SEC step, WO246 does not, and further, when using the large scale claimed volume (100L-20000L), one would not have a reasonable expectation of successfully using SEC with a large volume because using large volumes greater than 5-10% of the total column volume decreases resolution, causing peak distortion (See Hong, Sample load section). Additionally, Sarawat who teach purification methods of recombinant proteins to include Anion exchange, HA chromatography (mixed-mode), SEC, and HIC chromatography, teach that sample volumes needed for SEC are very small (p. 6, 1st col., last parag.) and that TFF appears to be gradually replacing SEC in downstream processing (p. 6, 2nd col, 1st parag.). Thus, not using SEC would have been obvious to a posita before the effective filing date of the claimed invention.
Claim(s) 14 is/are rejected under 35 U.S.C. 103 as being unpatentable over van den Dobbelsteen et al. (Vaccine, 2016, IDS) in view of WO2014110246 and WO2020191082 (IDS) supported by Sarawat et al. (Biomed. Res. Int., 2013, IDS) and Hong et al. (J. Liq. Chrom.& related tech., vol. 35, p. 2923-2950, 2012) as applied to claims 1, 2, 4-13, 15, 18 above, and further in view of WO2015/124769 A1 (IDS).
Regarding claim 14, while van denDobbelsteen teach that the cells are cultured and harvesting by centrifugation (section 2.1, last parag.), however the temperature of culture is not taught.
WO769 teaches a process for producing a purified O-EPA bioconjugate obtained from a prokaryotic cell, i.e. E. coli polysaccharide conjugated to carrier protein exotoxin A of P. aeruginosa (EPA) (0004, 0011-0013, 0098, 00161), wherein the O-EPA is a conjugate drug product composition which is a multivalent vaccine comprising at least E. coli antigen polysaccharides O25B, O1A, O2 and O6A (00207, 00208, 00222). The bioconjugate is produced by growing host cells at a temperature of 35°C (0307) and purifying the bioconjugate from the periplasmic space (00300). The bioconjugates can be purified using two consecutive anionic exchange chromatography (00353). The composition is disclosed to be stored frozen (00230).
Thus, before the effective filing date of the claimed invention, culture temperatures for producing O-EPA conjugates were known and therefore, a posita has good reason to pursue known culture temperatures within his or her technical grasp with a reasonable expectation of successfully culturing and harvesting prokaryotic host cells.
Claim(s) 17 is/are rejected under 35 U.S.C. 103 as being unpatentable over van den Dobbelsteen et al. (Vaccine, 2016, IDS) in view of WO2014110246 and WO2020191082 (IDS) supported by Sarawat et al. (Biomed. Res. Int., 2013, IDS) and Hong et al. (J. Liq. Chrom.& related tech., vol. 35, p. 2923-2950, 2012) as applied to claims 1, 2, 4-13, 15, 18 above, and further in view of WO2018/077853 (IDS).
The references do not teach step f-16) of claim 17 drawn to portioning and freezing the purified O-EPA.
WO853 teaches a conjugate drug product composition comprising at least one E. coli O polysaccharide bound to exotoxin A of P. aeruginosa (O-EPA), wherein the composition is a multivalent composition comprising E. coli antigen polysaccharides O25B, O1A, O2 and O6A (0009, 0019, 0043, 0045, 0046, 0060)(ExPEC multivalent vaccine), and freezing said ExPEC vaccines (0005-0008, 0025, 0085). The product compositions are formulated to be stable when stored in a frozen state (see Ex. 1-4, for example). WO853 teaches that methods to prepare the composition for storage by buffer exchange including TTF, chromatography and purifications steps (0086).
Before the effective filing date of the claimed invention, methods of formulating O-EPA conjugate drug substance/products for freezing were known and practiced in the prior art of record. Thus, it would have been obvious to one of ordinary skill in the art to pursue known options of purifying and formulating O-EPA conjugate drug substance/products within his or her technical grasp with a reasonable expectation of successfully storing/freezing the product.
Conclusion
Any inquiry concerning this communication or earlier communications from the examiner should be directed to TIFFANY MAUREEN GOUGH whose telephone number is (571)272-0697. The examiner can normally be reached M-Thu 8-5.
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/TIFFANY M GOUGH/ Examiner, Art Unit 1651
/MELENIE L GORDON/Supervisory Patent Examiner, Art Unit 1651