DETAILED ACTION
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
This is the second Office action on the merits of the claims.
All citations to the Manual of Patent Examining Procedure (MPEP) refer to Revision 01.2024, which was released in November 2024.
Status of the Claims
In the Reply filed 11 December 2025, Applicant amended claims 1, 6-11, 14-15, 18-21, 24, 26-27, 30, 32-34, 38, 44 and 51; cancelled claims 2, 25, 29, 31 and 39; and added one new claim, i.e., claim 55. Claims 35-37, 40-43, 45-50, and 52-54 were cancelled previously by Applicant. Claims 1, 3-24, 26-28, 30, 32-34, 38, 44, 51, and 55 are pending.
Status of the Rejections and Objections
The objections to the claims are new and have been necessitated by Applicant’s recent amendments.
The rejection under 35 U.S.C. 112(b) has been modified in view of Applicant’s claim amendments.
The rejection of claims 1, 6, 10-11, 13-14, 18-28, and 38-39 under 35 U.S.C. 102(a)(1) as being anticipated by Liu (CN 110092841 A), as evidenced by Zhang (Reprint of “Mixed-mode chromatography in pharmaceutical and biopharmaceutical applications.” Journal of pharmaceutical and biomedical analysis 130 (2016): 19-34) and Borgohain (“Maximizing expression and yield of human recombinant proteins from bacterial cell factories for biomedical applications.” Advances in microbial biotechnology (2018): 431-468) is withdrawn in view of Applicant’s narrowing amendments to claim 1.
The rejection of claims 1-6, 11-34, 38-39, 44, and 51 under 35 U.S.C. 103 as being unpatentable over Bale in view of Borgohain, Massiah, Menart and, optionally, Baker has been modified in view of Applicant’s combination of narrowing amendments to claim 1. The examiner notes that no claim previously required a particular urea concentration or concentration range. Claim 1, as recently amended, now requires a urea concentration range of 0.05 to 1.0 M. Applicant is alerted that (i) the rationale supporting the §103 rejection has been updated (see paragraph 48 infra) and (ii) different or additional sections of certain references are cited as support, most notably pages 5-7 and 13-15 of Menart.
The two remaining §103 rejections set forth in the previous Office action (23 September 2025) were predicated on the foregoing §103 rejection and have been modified to maintain consistency.
Claim Objections
Claims 1-5, 7-13, 18-19, 28 and 38-42 are objected to because of the following minor informality: In claim 1, the semicolon immediately following “0.5 M” should be replaced with a comma. Appropriate corrected is required.
Claims 4 and 5 are objected to because of the following informality: The word “further” is missing between the word “solution” and the transitional phrase “comprises.” Appropriate corrected is required.
Claim Rejections - 35 U.S.C. 112(b)
The following is a quotation of 35 U.S.C. 112(b):
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.
Claims 6, 18-19, 21, 24, and 34 are rejected under 35 U.S.C. 112(b) as being indefinite for failing to particularly point out and distinctly claim the subject matter that the inventors regard as the invention.
Regarding claim 6, it is unclear what (if anything) differentiates the step of “washing the inclusion bodies with a wash solution comprising urea” (claim 6) from the step of “contacting the inclusion bodies with a solubilization solution comprising urea at a concentration of from 0.05 M to 1.0 M” (claim 1). How does claim 6 further limit claim 1?
Regarding claims 18 and 19, the phrase “such as” leads to confusion over the intended scope of the claim. Is the claim language merely exemplary and, therefore, not further limiting? MPEP § 2173.05(d) (“If stated in the claims, examples and preferences may lead to confusion over the intended scope of a claim. In those instances where it is not clear whether the claimed narrower range is a limitation, a rejection under 35 U.S.C. 112(b) or pre-AIA 35 U.S.C. 112, second paragraph should be made.”).
In further regard to claim 19, the adjective “strong” is a relative term that renders the claim indefinite. MPEP § 2173.05(b)(1) (“Even if the specification uses the same term of degree as in the claim, a rejection is proper if the scope of the term is not understood when read in light of the specification.”).
Regarding claim 21, the following trademark or tradename is recited therein: “Superose 6.” A trademark or trade name is used to identify a source of goods, and not the goods themselves. Thus, a trademark or trade name does not identify or describe the goods associated with the trademark or trade name. The trademark/tradename recited in claim 21 is used to identify or describe a size-exclusion chromatography column and, consequently, causes confusion as to the scope of the claim. Therefore, claim 21 does not comply with the requirements of 35 U.S.C. 112(b). MPEP § 2173.05(u). Applicant is required to delete the trademark/tradename and, if desired, replace it with generic terminology describing the corresponding column.
Regarding claim 24, it is unclear how the functional limitations now recited in this claim further limit the production process of claim 1. How must the process of claim 1 be further defined to yield a “compB” protein (product) that satisfies the “assembly competent” functional limitation of claim 24? A person having ordinary skill in the art, even after reviewing the specification of the present application, would not be able to discern the scope of the foregoing functional limitation with reasonable certainty. MPEP § 2173.05(g) (“the use of functional language in a claim may fail ‘to provide a clear-cut indication of the scope of the subject matter embraced by the claim’ and thus be indefinite”).
Regarding claim 34, this application discloses a total of 62 sequences. Accordingly, the highest sequence ID number is 62. Because there is no sequence corresponding to SEQ ID NO: 169, this claim is indefinite and cannot be examined further on the merits.
Claim Rejections - 35 U.S.C. 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 set forth in Graham v. John Deere Co., 383 U.S. 1, 148 USPQ 459 (1966), that are applied for establishing a background for determining obviousness under 35 U.S.C. 103(a) 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, 3-6, 11-24, 26-28, 30, 32-33, 38, 44, 51, and 55 are rejected under 35 U.S.C. 103 as being unpatentable over Bale (“Accurate design of megadalton-scale two-component icosahedral protein complexes.” Science 353, 389-394 & SM1-SM40 (2016)) in view of Borgohain (“Maximizing expression and yield of human recombinant proteins from bacterial cell factories for biomedical applications.” Advances in microbial biotechnology (2018): 431-468), Massiah (“Obtaining soluble folded proteins from inclusion bodies using Sarkosyl, Triton X‐100, and CHAPS: application to LB and M9 minimal media.” Current protocols in protein science 84.1 (2016): 6-13), Menart (WO 2004/015124 A1) and, optionally, Baker (US 2016/0122392 A1).
Bale is directed to the accurate design of megadalton-scale two-component icosahedral protein complexes. This rejection contains citations to the primary article (pages 389-394), as well as citations to the Supplementary Materials, which are in the following format: SM[page number].
Bale discloses: “Nature provides many examples of self- and co-assembling protein-based molecular machines, including icosahedral protein cages that serve as scaffolds, enzymes, and compartments for essential biochemical reactions and icosahedral virus capsids, which encapsidate and protect viral genomes and mediate entry into host cells. Inspired by these natural materials, we report the computational design and experimental characterization of co-assembling, two-component, 120-subunit icosahedral protein nanostructures with molecular weights (1.8 to 2.8 megadaltons) and dimensions (24 to 40 nanometers in diameter) comparable to those of small viral capsids.” (Emphasis added) Abstract.
Bale discloses: “We set out to design two-component icosahedral protein complexes capable of packaging macromolecular cargo through controlled in vitro assembly. The twofold, threefold, and fivefold rotational axes present within icosahedral symmetry provide three possible ways to construct such complexes from pairwise combinations of oligomeric building blocks; we refer to these architectural types as I53, I52, and I32 (fig. S1). The I53 architecture is formed from a combination of 12 pentameric building blocks and 20 trimeric building blocks aligned along the fivefold and threefold icosahedral symmetry axes, respectively (Fig. 1, A to E; I53 stands for icosahedral assembly constructed from pentamers and trimers).” (Emphasis added) Pages 390-391.
Bale discloses that genes encoding the 71 pairs of I53 sequences were synthesized and cloned into a variant of the pET29b expression vector (an expression plasmid). Page SM9.
Bale discloses: “Expression plasmids were transformed into BL21(DE3) E. coli cells. Cells were grown in LB medium supplemented with 50 mg L-1 of kanamycin (Sigma) at 37° C until an OD600 of 0.8 was reached. Protein expression was induced by addition of 0.5 mM isopropyl-thio-β-D-galactopyranoside (Sigma) and allowed to proceed for either 5 h at 22 °C or 3 h at 37 °C before cells were harvested by centrifugation.” Page SM10.
Bale discloses: “The designed proteins were screened for soluble expression and co-purification as follows. Cells collected from 2 to 4 mL expression cultures were lysed by sonication in 25 mM TRIS pH 8.0, 250 mM NaCl, 1 mM DTT, 20 mM imidazole supplemented with 1 mM phenylmethanesulfonyl floride and the lysates cleared by centrifugation. A portion of each soluble fraction was saved for analysis by SDS-PAGE. The remaining portion of each soluble fraction was applied to His MultiTrap FF nickel-coated filter plates preequilibrated with 25 mM TRIS pH 8.0, 250 mM NaCl, 1 mM DTT, 20 mM imidazole running buffer (GE Healthcare).” Page SM10.
In Figures 4D and 4E (page 392), Bale discloses that two-component icosahedral protein complexes are formed using I53-50A.1PT1 and I53-50B.4PT1 as variants of the trimeric and pentameric components of I53-50. In Table S4, Bale discloses the amino acid sequence for I53-50B.4PT1. Page SM38. That sequence satisfies the new SEQ ID NO: 40 limitation now recited in claim 1 (as recently amended) of the present application.
Bale is silent regarding inclusion bodies and the processing thereof. It follows that Bale also does not disclose the new urea limitation — i.e., “urea at a concentration of from 0.05 M to 1.0 M, optionally 0.5 M” — now recited in claim 1, as recently amended. Moreover, Bale is silent regarding the negative limitation concerning denaturing or refolding now recited in claim 1.
As explained below, the following three references compensate for this deficiency: Borgohain, Massiah, and Menart.
Borgohain is directed to maximizing yield of recombinant proteins from bacterial cells.
Borgohain teaches: “Inclusion bodies are densely packed intracellular insoluble protein aggregates which are formed when a gene of interest is overexpressed in the cytoplasm of E. coli. Inclusion body formation is advantageous: (i) it helps in higher yield of protein in a pure form, (ii) protects the protein from intracellular proteases, (iii) homogeneity of protein reduces the purification step, and (iv) entrapped protein can be easily isolated based on its size and density. Since, these inclusion bodies are resistant to proteolysis, they contain a large amount of relatively pure protein of interest. However, these bodies are sites of misfolded proteins. Formation of these inclusion bodies is mainly due to the usage of strong promoters, high inducer concentrations, inability to form correct, or any, intra- or intermolecular disulphide bonds in the reducing intracellular environment, failure of bacteria to provide all post-translational modifications that a protein requires to fold, imbalance between in vitro protein solubilization and aggregation, and so forth.” (Emphasis added) Pages 452-453.
Borgohain teaches: “BL21 (DE) mutants [of E. coli] carry λDE3 lysogen with gene T7 RNA polymerase under the control of lac UV5 promoter. Presence of T7 RNA polymerase in these cells induces specific expression of gene(s) cloned downstream to a T7 promoter at a higher rate.” (Emphasis added) Page 439; see also Table 18.3 on page 446 (T7 promoter has a higher rate of transcription initiation) and page 447 (E. coli BL21(DE3) has a T7 promoter).
Borgohain teaches: “Isolation of inclusion bodies can be achieved by a treatment with lysozyme before cell homogenization to enable cell disruption. Inclusion bodies are isolated by low speed centrifugation of bacterial cells that have been mechanically ruptured either by high pressure homogenization or by sonication.” Page 453.
Massiah is directed to obtaining soluble folded proteins from E. coli inclusion bodies.
Massiah teaches: “Recombinant proteins, especially those of eukaryotic origin, can have a tendency to aggregate or become packaged into inclusion bodies (IB) in E. coli because of a number of reasons: too high a concentration in the cell, the protein may be too large or partially toxic to the cell. Furthermore, the type of vector employed, the codon type, and the bacterial cell type can also contribute to IB formation. Inclusion bodies are protein aggregates and in E. coli will appear as a white speck under high magnification light microscopy. While it is still not clear, proteins in IB may not be completely unfolded but rather natively folded or close to it, based on the observation that protocols using sarkosyl [(sodium lauroyl sarcosinate)] can rescue functional proteins without refolding steps.” (Emphasis added) Pages 1-2, bridging paragraph.
Menart is directed to a process for the production of a biologically active heterologous protein.
Menart teaches: “For intracellular production of heterologous G-CSF in the bacterium E. coli, the protein is accumulated in the form of inclusion bodies (classical inclusion bodies). In experiments of secretion into E. coli periplasm, G-CSF is accumulated either in the form of classical inclusion bodies or there has been no report on biological activity of G-CSF produced in this way. Page 1.
Menart teaches: “From the aforementioned, it is clear that in almost all described experiments of isolation of G-CSF from the bacterium E. coli in the prior art, G-CSF is found in classical inclusion bodies.” Page 1. “Similar observations on the formation of classical inclusion bodies also apply to the production process of heterologous protein other than G-CSF.” (Emphasis added) Page 2.
Menart teaches: “Processes for the production of recombinant proteins from classical inclusion bodies comprise lysis and disruption of the cells followed by centrifuging. The pellet comprising a large proportion of classical inclusion bodies is usually washed with detergents. … A further step in obtaining recombinant proteins is the solubilisation of classical inclusion bodies requiring generally the use of rather strong denaturants.” (Emphasis added) Page 2; see also page 37 at claim 27.
Menart teaches: “It is an object of the invention to provide an improved process for the production of a heterologous protein, which involves obtaining the heterologous protein from non-classical inclusion bodies which are formed in the organism in which the heterologous protein is expressed.” (Emphasis added) Page 4. Menart defines non-classical inclusion bodies as “inclusion bodies which are more soluble (in media under non-denaturating conditions such as non-denaturating aqueous solutions) than the classical inclusion bodies and which comprise a certain amount of correctly folded precursor of a heterologous protein.” (Emphasis added) Page 7.
Menart teaches: “In another particular aspect of the invention, the precursors of the heterologous protein found in the inclusion bodies are kept, during the process of isolation and purification of the heterologous protein from the inclusion bodies, under conditions which are non-denaturating for the heterologous protein. A particularly preferred embodiment of the process for the production of biologically active heterologous protein of the present invention accordingly further comprises the solubilisation of inclusion bodies, which is preferably preceded by a washing step, under non-denaturating and preferably native conditions and enables the direct isolation of biologically active proteins, without the need of using denaturants or applying a denaturation/renaturation process. Since the present invention enables the production of inclusion bodies having a substantial proportion of correctly folded precursor of the heterologous protein (non-classical inclusion bodies), this particular aspect of the invention provides a very efficient way to the production of the protein, without a denaturation/renaturation step being required.” (Emphasis added) Pages 5-6 at bridging paragraph.
Menart teaches that the preferred cultivation temperature range for the accumulation of the correctly folded precursor in non-classical inclusion bodies is significantly lower than 37°C, namely, between about 20°C and about 30°C, with the most preferred temperature being about 25°C. Page 13.
Menart teaches that the proportion of the correctly folded precursor in non-classical inclusion bodies also depends on the induction mode and, thereafter, identifies IPTG (isopropyl-thio-β-D-galactopyranoside) as the most-preferred mode of induction. Page 13.
Menart provides additional guidance on increasing the proportion of the correctly folded precursor in non-classical inclusion bodies by optimizing (i) the mode of fermentation and (ii) the composition of the cultivation medium. Pages 13-14.
Menart teaches: “The higher solubility of non-classical inclusion bodies which occurs due to a higher solubility of a correctly folded precursor of heterologous protein in the inclusion bodies indicates that the solubilisation can be advantageously performed under mild conditions, without the addition of strong denaturants, strongly alkaline solutions or denaturating concentrations of detergents.” (Emphasis added) Page 15. “For the solubilisation of the inclusion bodies,” Menart continues, “the solvents to be used can be selected from the group consisting of: urea in non-denaturing concentrations (1-2 M, preferably in a buffer at a pH of below 10 and more preferably at a pH of about 8.0) ….” (Emphasis added) Id.; see also page 32 (Example 8) and page 37 (claim 29).
Before the effective filing date of the claimed invention, the foregoing teachings of Borgohain, Massiah, and Menart would have motivated a person having ordinary skill in the art to infer that inclusion bodies composed of I53 pentameric and trimeric protein building blocks were likely present in the BL21(DE3) E. coli cells of Bale following inducement of protein expression and incubation (see page SM10). MPEP § 2144.01 (“[I]n considering the disclosure of a reference, it is proper to take into account not only specific teachings of the reference but also the inferences which one skilled in the art would reasonably be expected to draw therefrom.”). The teachings of Massiah and (especially) Menart would have motivated the person having ordinary skill in the art to modify Bale’s process for the production of the I53 pentameric and trimeric protein building blocks (the heterologous proteins) by selecting a cultivation temperature, an inducer, a fermentation mode, and/or cultivation medium suitable for encouraging the formation of non-classical inclusion bodies, which (i) are more soluble than classical inclusion bodies and (ii) in contrast to classical inclusion bodies, contain heterologous proteins in their natural conformation, thereby avoiding the need to denature and/or renature (refold) them. Given that the production process of Bale already can employ a relatively low cultivation temperature (22°C) and an inducer (IPTG) recommended by Menart, the examiner finds that further modification of that process by incorporating one or more of the remaining teachings of Menart would have been undertaken with a reasonable expectation of success, for the purpose of increasing the yield of the I53 pentameric and trimeric protein building blocks. MPEP § 2143.02(I) (“Where there is a reason to modify or combine the prior art to achieve the claimed invention, the claims may be rejected as prima facie obvious provided there is also a reasonable expectation of success.”). Therefore, in accordance with Menart, the person having ordinary skill in the art would have been motivated to solubilize the non-classical inclusion bodies in urea (1-2 M) under non-denaturing concentrations. MPEP § 2144.05(I) (overlapping, approaching, and similar ranges, amounts, and proportions). In sum, claims 1, 27, and 38 are prima facie obvious.
Regarding claim 3, Menart teaches that the urea solution is buffered and, most preferably, has a pH of about 8.0. Page 15.
Regarding claims 4 and 5, Menart teaches that the solubilization solution can comprise “low concentrations of Zwittergents” (page 15), which is tradename for a class of commercially-available zwitterionic surfactants. Alternatively, Massiah teaches that “3-[(3-cholamidopropyl) dimethylammonio]-1-propanesulfonate (aka CHAPS or CHAPSO) is a zwitterionic detergent with nondenaturing properties used for enhancing protein solubility.” Page 4. Furthermore, the optional reference (Baker) teaches that CHAPS increases the solubility of I53 building block proteins. Para. [0127]. Additionally, Applicant is referred to MPEP § 2144.06(I) (combining equivalents known for the same purpose).
Regarding claim 6, Applicant is referred to claim 27 on page 37 of Menart. See also Menart at pages 14-15 (bridging paragraph). The selection of any order of performing process steps is prima facie obvious in the absence of new or unexpected results. MPEP § 2144.04(IV)(C), citing In re Burhans, 154 F.2d 690 (CCPA 1946).
Regarding claims 11 and 13-14, Applicant is referred to Figure 18.3 of Borgohain. Page 444. Bale (the primary reference) utilizes BL21(DE3) E. coli cells (page SM10), which are a B-strain, as evidenced by page 439 of Borgohain.
Regarding claim 12, as discussed above, Menart teaches that the preferred cultivation temperature range for the accumulation of the correctly folded precursor in non-classical inclusion bodies is significantly lower than 37°C, namely, between about 20°C and about 30°C, with the most preferred temperature being about 25°C. Page 13. Applicant is referred to MPEP § 2144.05(I) (“In the case where the claimed ranges ‘overlap or lie inside ranges disclosed by the prior art’ a prima facie case of obviousness exists.”).
Regarding claim 15, Borgohain teaches: “The production efficiency and bioactivity of the produced recombinant proteins are greatly influenced by the strain of E. coli used. In routine practice, BL21 and K12 and their mutant stains are most widely used (Table 18.1).” Page 439; see also Table 18.1 on pages 437-438.
Regarding claims 16 and 17, Applicant is referred to Table 18.3 of Borgohain (page 446), which lists eight commonly used promoter systems, for example, T7 and PhoA. Table 18.3 teaches that PhoA has the advantage of tightly controlled and selective induction, whereas T7 has the disadvantages of lower cell densities and leaky expression.
Regarding claims 18 and 19, Bale discloses: “Cells collected from 2 to 4 mL expression cultures were lysed by sonication in 25 mM TRIS pH 8.0, 250 mM NaCl, 1 mM DTT, 20 mM imidazole supplemented with 1 mM phenylmethanesulfonyl floride and the lysates cleared by centrifugation.” Page SM10.
Regarding claims 20 and 21, Bale discloses the amino acid sequence for I53-50B.4PT1, which is a “compB” protein. Page SM38. That sequence matches SEQ ID NO: 34 of the present application, which is also referred to in claims 30 and 32-33 of the present application as “I53-50B.4PosT1.” This provides a sound basis for the examiner’s position that the claimed solubility range is satisfied, regardless of whether solubility is measured by gel filtration chromatography or another method. MPEP § 2112(V) (once a reference teaching product appearing to be substantially identical is made the basis of a rejection, and the examiner presents evidence or reasoning to show inherency, the burden of production shifts to the applicant). Even though neither claim 20 nor claim 21 requires an active (manipulative) step of purifying or otherwise filtering via gel chromatography, the examiner notes — in the interest of compact prosecution — that Bale discloses purification on a Superose 6 Increase 10/300 gel filtration column. Page SM16.
Regarding claims 22-24, 26 and 28, Bale discloses gel filtration chromatography on page SM11 and polyacrylamide gel electrophoresis (SDS-PAGE) on pages 393 (center column) and SM10. The remaining claimed features are expressions of various intended results of engaging in the active (manipulative) steps of the method of production recited in claim 1 and, therefore, are not afforded patentable weight. MPEP § 2111.04(I) (a “‘whereby clause in a method claim is not given weight when it simply expresses the intended result of a process step positively recited’”), quoting Hoffer v. Microsoft Corp., 405 F.3d 1326, 1329 (Fed. Cir. 2005) (quoting Minton v. Nat’l Ass’n of Securities Dealers, Inc., 336 F.3d 1373, 1381 (Fed. Cir. 2003)). Also, where the claimed and prior art products are produced by identical or substantially identical processes, a prima facie case of obviousness has been established. MPEP § 2112.01(I). In further regard to claim 24 (as recently amended), Applicant is referred below to the rejection of claims 44, 51, and 55, which includes a discussion about the following “compA” protein of the present application: SEQ ID NO: 29.
Regarding claims 30 and 32-33, Applicant is referred to the I53 pentameric and trimeric proteins disclosed in Table S4 of Bale (pages SM36-SM38). For example, Table S4 at page SM38 discloses the amino acid sequence for I53-50B.4PT1, which is the same as Applicant’s I53-50B.4PosT1 (SEQ ID NO: 34). Alternatively, the optional reference (Baker) teaches I53-50B.1 (SEQ ID NO: 32), I53-50B.1NegT2 (SEQ ID NO:33), or I53-50B.4PosT1 (SEQ ID NO: 34), and the I53-50B genus (SEQ ID NO: 40). Page 5 at Table 1.
Regarding claims 44, 51 and 55, Bale discloses: “The ability of I53-50A.1 and I53-50B.4PT1 to assemble to the designed icosahedral architecture upon mixing in vitro was analyzed by mixing purified components in a 1:1 molar ratio with each component present at a subunit concentration of 50 μM or 100 μM.” Page SM16; see also page 392 at Figure 4E. On page SM38, Bale discloses the amino acid sequences for I53-50A.1 and I53-50B.4PT1. Those sequences respectively match SEQ ID NO: 29 and SEQ ID NO: 34 of the present application, which is combination (xxiii) of claim 55. In further regard to claim 51, Bale teaches that the two-component icosahedral protein complexes disclosed therein mimic viral capsids (Abstract) and are an attractive starting point for vaccine design (page 393, right column). Baker teaches: “For vaccine design, antigenic epitopes from pathogens could be fused or conjugated to the nanostructure exterior to stimulate development of adaptive immune responses to the displayed epitopes, with adjuvants and other immunomodulatory compounds attached to the exterior and/or encapsulated in the cage interior to help tailor the type of immune response generated for each pathogen.” Para. [0102]; see also paras. [0027] and [0092]. A person having ordinary skill in the art would have determined immunostimulatory amounts (effective amounts) through routine experimentation. MPEP § 2144.05(II)(A) (“‘[W]here the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation.’”), quoting In re Aller, 220 F.2d 454, 456 (CCPA 1955).
Claims 7-9 are rejected under 35 U.S.C. 103 as being unpatentable over Bale in view of Borgohain, Massiah, Menart and, optionally, Baker, as applied above to claims 1, 3-6, 11-24, 26-28, 30, 32-33, 38, 44, 51 and 55, and further in view of Creighton (US 4,977,248) and Wingfield (Overview of the Purification of Recombinant Proteins. Curr. Protoc. Protein Sci. (2015) 80:6.1.1-6.1.35).
Bale discloses that the two-component icosahedral protein complexes resembling viral capsids were purified on a Superose 6 Increase 10/300 gel filtration column. Page SM16. However, Bale is silent as to whether anion-exchange chromatography could be used instead to purify them. As explained below, Creighton and Wingfield compensate for this deficiency.
Creighton is directed to “a process for the production of a soluble native protein, in which an insoluble form of the protein is produced by host cells transformed with a vector including a gene coding for the protein, and as such relates to the field of protein production using recombinant DNA biotechnology.” Column 1, lines 10-16.
Creighton teaches: “A method for the renaturation of unfolded proteins comprises reversibly immobilizing the denatured protein on a solid phase and inducing folding of the immobilized protein by progressively reducing with time the concentration of a denaturing agent in the solvent in contact with the solid phase. The refolded protein is recovered from the solid phase in native form. The proteins can be folded and recovered in high yield in a small volume of buffer.” Abstract.
Creighton teaches: “For instance the solid phase may be an ion-exchange resin such as an agarose or similar material e.g. Q-sepharose or S-sepharose, Pharmacia Mono Q FPLC, Pharmacia Mono S FPLC, or cellulose, e.g. CM-cellulose, DEAE-cellulose, phospho-cellulose, or Amberlite of which CM-cellulose and Pharmacia Mono Q FPLC are preferred.” Column 3, lines 13-20. Diethylaminoethyl-(DEAE) cellulose resins, which are anion exchange resins, are utilized in Example 1 of Creighton. Column 4, lines 9-11; column 5, lines 30-35.
Creighton teaches: “The process may be applied advantageously to proteins produced by recombinant DNA biotechnology which are produced within host cells in the form of insoluble protein aggregates.” Column 2, lines 15-20. E. coli bacteria are identified as exemplary host cells. Column 2, lines 66-68.
Wingfield is directed to the purification of recombinant proteins from E. coli. Title/Abstract.
Wingfield teaches: “Recombinant proteins expressed in E. coli that are located in the low-speed pellet fraction (see Fig. 6.1.2) following cell lysis are highly aggregated (i.e., inclusion bodies).” Page 6.1.14, left column.
Wingfield teaches it is well established to purify proteins extracted from inclusion bodies with ion-exchange chromatography; see Figure 6.1.5 on page 6.1.15; and exemplifies DEAE-Sepharose, which is an anion exchange resin (see page 6.1.17, left column).
Before the effective filing date of the claimed invention, the teachings of Creighton and Wingfield would have motivated a person having ordinary skill in the art to modify the purification process disclosed in Bale by substituting DEAE media (anion exchange media) for the Superose 6 Increase 10/300 gel filtration column (agarose matrix), in an effort to optimize the purification process through routine experimentation. MPEP § 2144.07 (the selection of a known material based on its suitability for its intended use can support a prima facie obviousness determination). The examiner notes that Creighton teaches recovering the recombinant protein from the anion-exchange resin via elution with a salt (NaCl) gradient. Column 3, lines 56-58; see also column 4, lines 34-36. Therefore, claim 7 is prima facie obvious.
Regarding claim 8, Creighton teaches washing the resin column with a buffer solution containing urea. Column 4, lines 27-29. Wingfield teaches: “Other components often added to buffers to promote protein solubility during purification include nonionic or zwitterionic detergents, low concentrations of urea (1 to 2M), and salt (0.5 to 1 M NaCl).” (Emphasis added) Page 6.1.24, right column; see also Wingfield at page 6.1.14, left column and Bale at page SM16 (“buffers contained 500 mM NaCl and 0.75% 3-[(3-Cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS), which was found to minimize precipitation and aggregation of the individual protein components”).
Regarding claim 9, Creighton teaches a linear elution gradient of 0M to 1.0M NaCl. Column 5, lines 10-11; see also Bale at page SM16 (“buffers contained 500 mM NaCl and 0.75% 3-[(3-Cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS), which was found to minimize precipitation and aggregation of the individual protein components”). Applicant is referred to MPEP § 2144.05(I) (overlapping, approaching, and similar ranges, amounts, and proportions).
Claim 10 is rejected under 35 U.S.C. 103 as being unpatentable over Bale in view of Borgohain, Massiah, Menart and, optionally, Baker, as applied above to claims 1, 3-6, 11-24, 26-28, 30, 32-33, 38, 44, 51 and 55, and further in view of Liu (CN 110092841 A), as evidenced by Zhang (Reprint of “Mixed-mode chromatography in pharmaceutical and biopharmaceutical applications.” Journal of pharmaceutical and biomedical analysis 130 (2016): 19-34).
Bale discloses that the two-component icosahedral protein complexes resembling viral capsids were purified on a Superose 6 Increase 10/300 gel filtration column. Page SM16. However, Bale is silent as to whether mixed-mode chromatography could be used instead to purify them. As explained below, Liu compensates for this deficiency.
Liu, which is directed to “a recombinant virus-like particle expressed based on the inclusion body form, a preparation method and application thereof” (para. [0002]), published in Chinese. The examiner obtained an English machine translation from the European Patent Office. Unless otherwise indicated, all citations refer to that translation, which accompanies this Office action.
Liu teaches: “The recombinant virus-like particles are obtained by purification.” Para. [0013]. The method of purification comprises gel filtration or combined chromatography, preferably combined (mixed-mode) chromatography. Para. [0068]. “Preferably, the medium used in the combined chromatography is Capto core 700.” Para. [0069]; see also para. [0132]. Capto Core 700 is a commercially-available mixed-mode resin, as evidenced by page 22, right column, of Zhang (Reprint of “Mixed-mode chromatography in pharmaceutical and biopharmaceutical applications.” Journal of pharmaceutical and biomedical analysis 130 (2016): 19-34).
Before the effective filing date of the claimed invention, the teachings of Liu would have motivated a person having ordinary skill in the art to modify the purification process disclosed on page SM16 of Bale by substituting Capto Core 700 media (mixed-mode resin) for the Superose 6 Increase 10/300 gel filtration column (agarose matrix), in an effort to enhance the efficiency of the purification process during the course of routine experimentation. Therefore, claim 10 is prima facie obvious. MPEP § 2144.07 (the selection of a known material based on its suitability for its intended use can support a prima facie obviousness determination).
Conclusion
Claims 1, 3-24, 26-28, 30, 32-34, 38, 44, 51, and 55 are rejected.
Claims 1, 3-24, 26-28, 30, 32-34, 38, 44, 51, and 55 are also objected to.
No claim is 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.
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/P.A./
19 March 2026
/BETHANY P BARHAM/Supervisory Patent Examiner, Art Unit 1611