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 third 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.
Continued Examination
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and the fee set forth in 37 CFR 1.17(e) has been timely paid, the finality of the previous Office action has been withdrawn pursuant to 37 CFR 1.114.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 26 May 2026 has been entered.
Status of the Claims
Applicant amended claims 1, 4-5, 14, 18-19, and 24; cancelled claims 6, 15, 21, and 34; and added one new claim, i.e., claim 56. Claims 2, 25, 29, 31, 35-37, 39-43, 45-50, and 52-54 were cancelled previously by Applicant. Claims 1, 3-5, 7-14, 16-20, 22-24, 26-28, 30, 32-33, 38, 44, 51, and 55-56 are pending.
Status of the Rejections and Objections
The objections to the claims set forth in the previous Office action (23 March 2026) are withdrawn in view of Applicant’s amendments.
The objection to claim 14 set forth in this Office action is new.
The rejection of claims 1, 3-5, 7-14, 16-20, 22-24, 26-28, 30, 32-33, 38, 44, 51, and 55-56 under 35 U.S.C. 112(a) is new.
The rejection of claims 6, 18-19, 21, 24, and 34 under 35 U.S.C. 112(b) set forth in the previous Office action is withdrawn in view of Applicant’s amendments.
The rejection of claims 1, 3-5, 7-14, 16-20, 22-24, 26-28, 30, 32-33, 38, 44, 51, and 55-56 under 35 U.S.C. 112(b) set forth in this Office action is new.
The rejection of claims 1, 3-6, 11-24, 26-28, 30, 32-33, 38, 44, 51, and 55 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 recent amendments. Applicant’s amendment to claim 1 is considered in paragraph 57 of this Office action. Also, this rejection has been expanded to apply to (new) claim 56, which is considered in paragraph 69 of this Office action. Applicant’s arguments are considered in paragraphs 70-75 of this Office action.
The two remaining §103 rejections set forth in the previous Office action (23 March 2026) were predicated on the foregoing §103 rejection and have been modified to maintain consistency.
Claim Objections
Claim 14 is objected to because of the following minor informality: The strain identifiers (B-strain, K12-strain) should not be italicized because they are not part of the binomial name for the corresponding bacteria. Appropriate corrected is required.
Claim Rejections – 35 U.S.C. 112(a) – Written Description
The following is a quotation of 35 U.S.C. 112(a):
The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor or joint inventor of carrying out the invention.
Claims 1, 3-5, 7-14, 16-20, 22-24, 26-28, 30, 32-33, 38, 44, 51, and 55-56 are rejected under 35 U.S.C. 112(a) for failing to comply with the written description requirement.
Claim 1, as recently amended, now contains subject matter that was not described in the specification in such a way as to reasonably convey to one skilled in the relevant art that the inventors, at the time the application was filed, had possession of the invention, as now claimed.
Referring to the specification of the present application as originally filed 02 December 2022 (WO 2021/252687), paragraph [0147] — which is located on page 31 and is reproduced in relevant part below — supports a urea concentration range of 0.05 M to 3.0 M, as well as various narrower ranges encompassed therein, such as 0.05 M to 0.9 M. MPEP § 2163.05(III).
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However, paragraph [0147] provides no support for the broadening adverb <about> recited in the italicized segment of the following limitation from claim 1: “urea at a concentration of about 0.05 M, 0.1 M, 0.2 M, 0.3 M, 0.4 M, 0.5 M, 0.6 M, 0.7 M, 0.8 M, or 0.9 M.” Emphasis added.
No other relevant disclosure concerning the claimed urea concentration range is included in the specification.
Given that none of Applicant’s claims, as originally filed, compensates for the deficiency in the specification identified above, the present application does not reasonably convey to persons skilled in the art that the inventor, at the time the application was filed, had possession of the urea concentration a range of “about 0.05 M, 0.1 M, 0.2 M…” now recited in claim 1. Thus, claim 1 recites new matter. 35 U.S.C. 132(a) (“No amendment shall introduce new matter into the disclosure of the invention.”); see also MPEP § 608.04. Accordingly, it is appropriate to reject claim 1 and all claims depending thereon under 35 U.S.C. 112(a).
In further regard to claim 56, as established above, the present application (as originally filed) does not support the adverb <about> as recited in the following limitation: “urea at a concentration of about 0.5 M.” Consequently, claim 56 recites new matter. 35 U.S.C. 132(a) (quoted above); see also MPEP § 608.04. Accordingly, it is appropriate to reject claim 56 under 35 U.S.C. 112(a).
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 1, 3-5, 7-14, 16-20, 22-24, 26-28, 30, 32-33, 38, 44, 51, and 55-56 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 1, it is unclear whether the adverb <about> applies to “0.05 M” only or — instead — applies to the entire set of recited urea concentration values. For example, is the maximum urea concentration 0.9 M? Alternatively, is it about 0.9 M?
Regarding claims 1, 12, 28 and 56, the adverb <about> is a relative term that generates significant uncertainty. MPEP § 2173.05(b)(III)(A). The definition of “about” set forth on page 13 of the specification, as originally filed (WO 2021/252687), is reproduced below:
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Although Applicant’s definition of “about” is informative, it fails to clarify the claims because it includes a series of exemplary ranges that progressively narrow in scope. MPEP § 2173.05(c)(I) (“Use of a narrow numerical range that falls within a broader range in the same claim may render the claim indefinite when the boundaries of the claim are not discernible.”) and MPEP § 2173.05(d) (“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 sum, claims 1, 12, 28 and 56 and all claims depending thereon are indefinite.
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, 3-5, 11-14, 16-20, 22-24, 26-28, 30, 32-33, 38, 44, 51, and 55-56 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.
The foregoing urea range of 1-2 M (Menart at p. 15) is overlapped by the corresponding range now recited in claim 1, which has a maximum of about 0.9 M. 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.”). In the alternative, the urea range of 1-2 M (Menart) is close enough to the corresponding range recited in claim 1, which has a maximum of 0.9 M, to support a prima facie case of obviousness. MPEP § 2144.05(I) (“Similarly, a prima facie case of obviousness exists where the claimed ranges or amounts do not overlap with the prior art but are merely close.”). The examiner acknowledges that Example 8 of Menart (page 32) employs urea at a concentration of 2 M, not 1 M. However, it is important to recognize that “[a] reference may be relied upon for all that it would have reasonably suggested to one having ordinary skill in the art, including nonpreferred embodiments.” MPEP § 2123(I); see also MPEP § 2123(II) (“Disclosed examples and preferred embodiments do not constitute a teaching away from a broader disclosure or nonpreferred embodiments.”).
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 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. Additionally, 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 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 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 claim 20, 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 claim 20 does not require 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 “compB” protein of the present application, which is assembly competent: SEQ ID NO: 34.
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).
Regarding claim 56, the urea range of 1-2 M (Menart at p. 15) is close enough to the claimed range of “about 0.5 M” to support a finding of prima facie obviousness. MPEP § 2144.05(I) (“Similarly, a prima facie case of obviousness exists where the claimed ranges or amounts do not overlap with the prior art but are merely close.”). This conclusion is based, at least in part, on Applicant’s expansive definition of the adverb <about>, which is set forth on page 13 of the specification, as originally filed (WO 2021/252687). MPEP § 2111 (“During patent examination, the pending claims must be ‘given their broadest reasonable interpretation consistent with the specification.’”), quoting Phillips v. AWH Corp., 415 F.3d 1303, 1316 (Fed. Cir. 2005) (en banc).
Response to Applicant’s Arguments
The arguments raised by Applicant in the Reply filed 26 March 2026 are considered below:
First, Applicant argues: “Indeed, the Specification demonstrates the criticality of the claimed concentrations. For example, FIG. 5 shows and paragraph [0070] teaches that I53-50B protein ‘begins to extract at 50mM urea.’ The working examples (Examples 2 and 3) demonstrate successful solubilization at 0.5 M urea — which is a concentration well below Menart's teachings — while preserving assembly competence.” Reply, page 11. As explained below, the foregoing argument is not persuasive.
Example 2 utilizes urea at only one concentration, i.e., 0.5 M. Page 53 at para. [0221]. Example 3, referring to Figure 5, provides that “[a]n initial experiment determined at which concentration of urea the compB protein became solubilized, beginning at 50mM and increasing to 8M, using aliquots of lysate and equal volumes of PBS with increasing urea molarity for 2 hours.” Page 56 at para. [0229]. Figure 5 evidences that the compB protein was successfully solubilized at each of the following urea concentrations: 50 mM, 75 mM, 0.25 M, 0.5 M, 1.0 M, 1.5 M, 2.0 M, 3.0 M, 4.0 M, 8.0 M (see dark bands at y-axis = 15). The bands at 2.0 M and 1.5 M, for example, are thicker and more intense than the bands at 1.0 M and 0.5 M, suggesting that the former was somewhat better at solubilizing the compB protein than the latter, which does not seem to support Applicant’s assertion of criticality. MPEP § 2144.05(III)(A) (“‘The law is replete with cases in which the difference between the claimed invention and the prior art is some range or other variable within the claims. In such a situation, the applicant must show that the particular range is critical, generally by showing that the claimed range achieves unexpected results relative to the prior art range.’”), quoting In re Woodruff, 919 F.2d 1575 (Fed. Cir. 1990). More importantly, Figure 5 does not establish there is a critical difference between a urea concentration of 0.9 M (claim 1) and the corresponding urea concentration range of 1-2 M taught in Menart. In regard to claim 1, Applicant has not satisfied its burden of showing there is a critical difference between (i) 0.9 M or perhaps “about” 0.9 M and (ii) the minimum concentration of 1.0 M taught in Menart. MPEP § 2144.05(II)(A) (“Generally, differences in concentration or temperature will not support the patentability of subject matter encompassed by the prior art unless there is evidence indicating such concentration or temperature is critical.”).
Second, Applicant argues: “Finally, Menart’s teachings are directed to G-CSF, which is a small cytokine. The claimed compB proteins are pentameric building blocks for massive icosahedral nanostructures. There are no teachings in Menart that would suggest to a skilled artisan that its methodology, which was optimized for G-CSF, would work for structurally distinct, much larger multimeric protein complexes, particularly at urea concentrations below what Menart teaches.” Reply, page 11. As explained below, the foregoing argument is not persuasive.
Applicant is reminded that urea is not used to solubilize the megadalton-scale two-component icosahedral protein complexes, which are formed from a combination of 12 pentameric building blocks and 20 trimeric building blocks. Instead, it is used to solubilize the building blocks, themselves. Table S4 (Bale) at page SM38 discloses the amino acid sequence for I53-50B.4PT1. That pentameric building block has a length of 167 amino acids, which is close to the length of human granulocyte colony-stimulating factor (G-CSF), which has either 174 or 177 amino acids, as it exists in two forms. Furthermore, Menart teaches: “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.
The §103 rejection is maintained.
* * *
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, 3-5, 11-14, 16-20, 22-24, 26-28, 30, 32-33, 38, 44, 51 and 55-56, 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, 3-5, 11-14, 16-20, 22-24, 26-28, 30, 32-33, 38, 44, 51 and 55-56, 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, 3-5, 7-14, 16-20, 22-24, 26-28, 30, 32-33, 38, 44, 51, and 55-56 are rejected.
Claim 14 is also objected to.
No claim is allowed.
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/P.A./
08 August 2026
/BETHANY P BARHAM/Supervisory Patent Examiner, Art Unit 1611
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