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 .
Election/Restrictions
Applicant’s election without traverse of Group I (claims 1-17) drawn to recombinant miropin polypeptides/compositions; and Applicants’ election of species without traverse: SEQ ID NO: 4 (i.e., RVK miropin) and Claims 1-3 read on the elected species. (It is noted that the longer sequence of SEQ ID NO. 4 includes the shorter RVK sequence included in SEQ ID NO: 3 of claim 2.), in the reply filed on June 16, 2026 is acknowledged. Claims 4-17 are withdrawn.
The species of group I, therefore claims 1-3 which read on the elected species has been considered.
Claims 1-3 are hereby examined on the merits.
Claims 4-19 and 21 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim. Election was made without traverse in the reply filed on June 16, 2026.
Priority
This application is a 371 of PCT/US2022/071178, filed March 16, 2022. This application claims benefit of U.S. Provisional Application No. 63/176,945, filed April 20, 2021.
Status of Claims
Claims 1-19, and 21 are pending. Claims 4-19 and 21 are withdrawn for further consideration.
Claims 1-3 are currently examined on the merits herein.
Information Disclosure Statement
The IDS submitted on 10/18/2023 and 11/10/2025 are under consideration.
Claim Objections
Claim 1 is objected to because of the following informalities: a period (.) is missing at the end of the claim. Appropriate correction is required.
Claim 2 is objected to because of the following informalities: has a comma (,) instead of a period (.) at the end of the claim. Appropriate correction is required.
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.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
Claims 1-3 are rejected under 35 U.S.C. 103 as being unpatentable over Goulas et. al. (“A structure-derived snap-trap mechanism of a multispecific serpin from the dysbiotic human oral microbiome”, Theodoros Goulas, Miroslaw Ksiazek, Irene Garcia-Ferrer, Alicja M. Sochaj-Gregorczyk, Irena Waligorska, Marcin Wasylewski, Jan Potempa and F. Xavier Gomis-Rüth, J. Biol. Chem. 292(26) 10883–10898, published May 16, 2017), in view of Betts et. al. (“Amino Acid Properties and Consequences of Substitutions”, Matthew J. Betts, Robert B. Russell, Bioinformatics for Geneticists, Chapter 14, published February 21, 2003).
Goulas et. al. teaches polypeptides including a fragment of T. forsythia miropin (Gen-BankTM code WP_041590947; UniProt code G8UQY8) spanning residues Glu39–Glu408 (see Materials and Method, 1st paragraph under section “Protein production and purification”, page 10892) including amino acid sequence 362TAVEMVKTSS371 (see Figure 1B (ii) and (iii), page 10885) where TAVEMVKTSS is the sequence specific to wild type Reactive-Center Loop (RCL)
region of miropin. Goulas et. al. teaches miropin uses the serpin-type suicidal mechanism similar to a snap trap, the protein transits from a metastable native form to a relaxed triggered or induced form after cleavage of a reactive-site target bond in an exposed reactive-center loop (see Abstract). Goulas et. al. further teaches that miropin uniquely block many serine and cysteine endopeptidases of disparate architecture and substrate specificity owing to several potential target bonds within the reactive-center loop and to plasticity in accommodating extra beta-strands of variable length (see Abstract).
Goulas et. al. elaborates T. forsythia inhabits dental plaques in periodontal pockets that
contain an inflammatory exudate: gingival crevicular fluid. This fluid is rich in defensive cysteine endopeptidases (CEPs) and serine endopeptidases (SEPs) such as neutrophil elastase and cathepsin G, from the host immune system (see page 10884, Endopeptidase inhibition mechanism of T. forsythia miropin, paragraph 2). P. gingivalis secretes the CEPs calpain-like peptidase Tpr, periodontain, and gingipains K and R, as well as SEP PepK; and T. denticola contributes with CEP dentipain and SEP dentilysin. To keep these peptidases in check, T. forsythia possesses miropin, a serine proteinase inhibitor from the serpin family (see page 10884, Endopeptidase inhibition mechanism of T. forsythia miropin, paragraph 2) [Note: This is in direct correlation with the instant specification which exemplifies - recombinant miropin with remarkable broaden inhibitory spectrum, including apart from neutrophil elastase and Lys-gingipain (Kgp), also Arg-gingipain (Rgp) (see instant Example 1)].
Goulas et. al. assessed the inhibitory capacity of miropin against physiologically
relevant SEPs and CEPs and found that it uniquely blocks many serine and cysteine endopeptidases of disparate architecture and substrate specificity owing to several potential
target bonds within the RCL (see page 10884, Endopeptidase inhibition mechanism of T. forsythia miropin, paragraph 6). Goulas et. al. analyzed its native and peptidase-induced wild
type forms, as well as two mutants affecting the RCL and a mutant ablating a disulfide bond between vicinal residues (see page 10884, Endopeptidase inhibition mechanism of T. forsythia miropin, paragraph 6). Goulas et. al. produced two mutants : mutant K368A/T369K, mutant V367K/K368A, whereas the pI of the molecule was kept intact (see page 85, right col, paragraph 2).
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Goulas et. al. illustrates the RCL of the wild type (panel (ii)) and double-point
mutants V367K/K368A (panel (iii)) and K368A/T369K (panel (iv)) (see Figure 1. Page 85) [marked in green box above]. Goulas et. al. also disclose wild-type miropin and mutant K368A/T369K inhibited trypsin with stoichiometry of inhibition ~1.5, whereas V367K/K368A did so with stoichiometry of inhibition ~5. The kass values, in contrast, were very similar ~105 M-1s-1 thus indicating that the reactive-site bonds are equivalent with respect to inhibitory power in miropin (see Figure 2, page 10886, left col, paragraph 1).
Goulas et. al. is silent about the exact same mutations in the reactive center loop (RCL) region; i.e., where 362TAVEMV367K368T369SS371 is mutated to amino acid sequences 362TAVEMR367V368K369SS371 as in the instant claim.
Betts et. al. elaborates the nature of mutations and the properties of amino acids in a variety of different protein contexts to interpreting the effect that a particular amino acid change will have on protein structure and function (see 14.1. INTRODUCTION, paragraph 3, page 292). Betts et. al discuss general preferences for amino acid substitutions and important specific details regarding their possible structure and functional roles (see 14.5 AMINO ACID QUICK REFERENCE, paragraph 1, page 299-300). Betts et. al explains with examples like, (i) Valine - Being hydrophobic, valine prefers to be buried in protein hydrophobic cores. Valine can be substituted by other hydrophobic, particularly aliphatic, amino acids. (see 14.5.4 Valine (Val, V), 14.5.4.1 Substitutions); (ii) Lysine - Lysine can be substituted by arginine or other polar amino acids (see 14.5.11 Lysine (Lys, K), 14.5.11.1 Substitutions); and (iii) Threonine - Threonine can be substituted with other polar amino acids (see 14.5.17 Threonine (Thr, T), 14.5.17.1 Substitutions). Betts et. al. explains the rationale for the interpretation of effects seen by an amino acid substitution and select logical amino acids for mutagenesis experiments (see 14.7 A SUMMARY OF THE THOUGHT PROCESS, last paragraph, page 314).
Betts et. al. teaches site-directed mutagenesis is a powerful tool for discovering the importance of an amino acid in the function of the protein. Gross changes in amino acid type can reveal sites that are important in maintaining the structure of the protein. Conversely, when investigating functionally interesting sites it is important to choose replacement residues that are unlikely to affect structure dramatically, for example by choosing ones of a similar size
to the original (see 14.6.2 Site-directed Mutagenesis, page 312).
Betts et. al. further explains direct involvement in the formation and cleavage of bonds is only one of a combination of methods that an enzyme can use to catalyse a reaction. Transition states can be stabilized by complementary shape and electrostatics of the binding site of the enzyme and substrates can be precisely positioned, lowering the entropy of activation (see 14.6.2 Site-directed Mutagenesis, page 312). Betts et. al. teaches polar amino acids prefer to be surrounded by water. Those that are buried within the protein usually participate in hydrogen bonds with other side chains or the protein mainchain that essentially replace the water. Some of these carry a charge at typical biological pHs: lysine and arginine are positively charged (see 14.4.2 Polar Amino Acids, page 299). Regarding aliphatic side chains, Betts et. al. teaches the amino acids with aliphatic side chains are alanine, isoleucine, leucine, proline and valine; they contain largely non-reactive and flexible side chains that are ideally suited for packing in the protein interior; aliphatic side chains are very non-reactive, and are thus rarely involved directly in protein function, although they can play a role in substrate recognition. In particular, hydrophobic amino acids can be involved in binding/recognition of hydrophobic ligands (see 14.4.1.1 Aliphatic Side Chains, page 298). Betts et. al. teaches there are several other amino acids also contain aliphatic regions; for example, arginine, lysine, glutamate and glutamine are amphipathic, meaning that they contain hydrophobic and polar areas. All contain two or more aliphatic carbons that connect the protein backbone to the non-aliphatic portion of the side chain. In some instances it is possible for such amino acids to play a dual role, with part of the side chain being buried in the protein and another being exposed to water(see 14.4.1.1 Aliphatic Side Chains, page 298). Therefore, Betts et. al elaborates with examples like, (i) Valine - Being hydrophobic, valine prefers to be buried in protein hydrophobic cores. Valine can be substituted by other hydrophobic, particularly aliphatic, amino acids. (see 14.5.4 Valine (Val, V), 14.5.4.1 Substitutions); (ii) Lysine - Lysine can be substituted by arginine or other polar
amino acids (see 14.5.11 Lysine (Lys, K), 14.5.11.1 Substitutions); and (iii) Threonine - Threonine can be substituted with other polar amino acids (see 14.5.17 Threonine (Thr, T), 14.5.17.1 Substitutions).
It would have been obvious to combine the teaching of Goulas et. al. and Betts et. al. before the effective filing date of the claimed invention by utilizing the role for amino
acids of different classes in protein structure and function and site-directed mutations, mutations that allow adaptive evolution and post-translational modification in the potential protease inhibitor, and arrive at the instantly claimed invention. One of ordinary skill in the art would have been motivated to utilize the teachings of Goulas et. al. that to keep these peptidases (i.e., gingipains K and R) in check, T. forsythia possesses miropin, a serine proteinase inhibitor from the serpin family (see page 84, Endopeptidase inhibition mechanism of T. forsythia miropin, paragraph 3); with a reasonable expectation of success because Betts et. al. discuss the nature of mutations and the properties of amino acids in a variety of different protein contexts (see page 292, paragraph 2), the rationale for the interpretation of effects seen by an amino acid substitution and select logical amino acids for mutagenesis experiments (see 14.7 A SUMMARY OF THE THOUGHT PROCESS, last paragraph, page 314) and consideration of the physical and chemical properties of the amino acids again guiding the choice of replacements, along with knowledge of the structure of the protein(see 14.6.2 Site-directed Mutagenesis, page 312). Thus, one skilled in the art can predict peptide interactions that could uncover mechanisms of enzyme function and the use of site-directed mutagenesis to investigate mechanisms of enzyme catalysis, with an expectation to succeed in developing serpin-type proteinase inhibitor called miropin (Goulas et. al., see Abstract) capable of
inhibiting a broad spectrum of proteases (see Goulas et. al., Endopeptidase inhibition mechanism of T. forsythia miropin, page 84, left col, paragraph 3).
Regarding claims 1 and 2: According to Goulas et. al. the following mutations were performed –
[AltContent: rect](i) V367 Lysine (K)
[AltContent: rect](ii) K368 Alanine (A)
(iii) T369 remains same as Lysine (K)
Betts et. al discuss general preferences for amino acid substitutions and important specific details regarding their possible structure and functional roles (see 14.5 AMINO ACID QUICK REFERENCE, paragraph 1, page 299-300). Betts et. al. explains the rationale for the interpretation of effects seen by an amino acid substitution and select logical amino acids for mutagenesis experiments (see 14.7 A SUMMARY OF THE THOUGHT PROCESS, last paragraph, page 314).
Betts et. al. teaches (i) arginine (R) is a positively-charged, polar amino acid; prefers to substitute for the other positively-charged amino acid, lysine (K) (see 14.5.10 Arginine (Arg, R), 14.5.10.1 Substitutions, page 303) and (ii) being hydrophobic, valine (V) prefers to be buried in protein hydrophobic cores; valine is also Cβ branched (see 14.5.4 Valine (Val, V), 14.5.4.1 Substitutions, 14.5.4.2 Structure, page 301) and alanine (A) also contains a normal Cβ carbon, meaning that it is generally as hindered as other amino acids (see14.5.1 Alanine (Ala, A),
14.5.1.1 Substitutions, 14.5.1.2 Structure, page 300). Betts et. al. teaches the amino acids with aliphatic side chains are alanine, isoleucine, leucine, proline and valine; they contain largely non-reactive and flexible side chains that are ideally suited for packing in the protein interior;
aliphatic side chains are very non-reactive, and are thus rarely involved directly in protein function, although they can play a role in substrate recognition. In particular, hydrophobic amino acids can be involved in binding/recognition of hydrophobic ligands (see 14.4.1.1 Aliphatic Side Chains, page 298).
Therefore,
[AltContent: rect](i) V367 Lysine (K) OR Arginine (R)
[AltContent: rect](ii) K368 Alanine (A) OR Valine (V)
[AltContent: textbox ( )](iii) T369 Lysine (K)
with an expectation to mutate 362TAVEMV367K368T369SS371 to amino acid sequence 362TAVEMR367V368K369SS371.
Goulas et. al. also teaches Miropin is capable of broadly inhibiting SEPs and CEPs of disparate classes, architectures, and substrate specificities; which is achieved by offering several target bonds of the reactive center loop (RCL) for cleavage within a bait region, instead of a single reactive-site bond as found in canonical serpins. In addition, promiscuous inhibition is facilitated by the capacity to insert strands deviating from the canonical length into the central sheets, while keeping the prey peptidase bound and inactivated (see page 10892, left col, paragraph 6).
Thus the prior art renders the claimed sequence obvious, and a person of ordinary skill in the art would reasonably expect the same sequence to have the same properties, i.e. that of inhibiting different proteases than the wildtype.
Regarding claim 3: Goulas et. al. discloses The atomic coordinates and structure factors (codes 5NCS, 5NCT, 5NCU, and 5NCW) have been deposited in the Protein Data Bank (http://wwpdb.org/) (see page 83 footnote, left col) (Note: 5NCS (wild type) is equivalent to instant SEQ ID NO: 1, and the instant specification states that the amino acid sequence SEQ ID NO:4 that comprises the amino acid sequence TAVEMRVKSS (SEQ ID NO:3) corresponds to the recombinant miropin amino acid sequence or a variant thereof having at least 65%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO:4 that comprises the amino acid sequence TAVEMRVKSS (SEQ ID NO:3) (see [0037], TABLE-US-00003). Miropin Wild Type and Miropin SEQ ID NO: 4 (as in instant claim) sequence alignment results show (A) Query Match 89.4% and (B) Best Local Similarity 99.2%.
Therefore, the presently claimed invention was prima facie obvious to one of ordinary skill in the art at the time of the effective filing date.
Claim Rejections - 35 USC § 101
35 U.S.C. 101 reads as follows:
Whoever invents or discovers any new and useful process, machine, manufacture, or composition of matter, or any new and useful improvement thereof, may obtain a patent therefor, subject to the conditions and requirements of this title.
Claims 1-3 are rejected under 35 U.S.C. 101 because the claimed invention is directed to a natural phenomenon/natural product without significantly more. Claims 1-3, recite a naturally occurring product, which is not markedly different from its naturally occurring counterpart. See MPEP §§ 2106 et seq.
Claims 1-3 are directed to a recombinant miropin polypeptide, comprising the amino acid sequence TAVEMRVKSS (SEQ ID NO: 3); wherein the recombinant miropin comprises the
amino acid sequence SEQ ID NO:4, or a variant thereof having at least 65% sequence identity to SEQ ID NO:4.
Note: The Miropin Wild Type – Miropin SEQ ID NO: 4 (as in instant claim) sequence alignment results show (A) Query Match 89.4% and (B) Best Local Similarity 99.2%.
The claims do not include additional elements that are sufficient to amount to significantly more than the judicial exception for the following reasons. This judicial exception is not integrated into a practical application because the claimed composition comprises naturally occurring components. The claim(s) does/do not include additional elements that are sufficient to amount to significantly more than the judicial exception because for the following reasons.
Step 1: Is the claim to a process, machine, manufacture or composition of matter? The instant claims are directed to a statutory patent-eligible subject matter category, a composition of matter.
Step 2a Prong 1: Is the claim directed to a law of nature, a natural phenomenon (Product of nature), or an abstract idea? The claims are directed to a natural phenomenon, specifically a natural-based product limitation. The amino acid sequence SEQ ID NO:4 that comprises the amino acid sequence TAVEMRVKSS (SEQ ID NO:3) corresponds to the recombinant miropin amino acid sequence or a variant thereof having at least 65%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO:4 that comprises the amino acid sequence TAVEMRVKSS (SEQ ID NO:3) (see [0037], TABLE-US-00003).
Of note, the modified SEQ ID NO: 3 described above have only taken into consideration 1-3 amino acid substitutions relative to SEQ ID NO: 3. Other alterations, such as additions or deletions, have the potential to also read on natural products.
Step 2a Prong 2: Does the claim recite additional elements that integrate the judicial exception into a practical application? This judicial exception is not integrated into a practical application because the composition being claimed comprises naturally occurring components (MPEP § 2106.04(d)(III)).
Step 2b: Does the claim recite additional elements that amount to significantly more than the judicial exception? Claims 1-3 only recite the amino acid sequence, which is a natural phenomenon exception. None of the claims above integrate the above amino acid sequence into a practical application. At most, the claims recite the sequences in a composition or as a protease inhibitor (claims 1-3, respectively) but without significantly more.
Examiner’s Note
A plausible mechanism of action of Miropin; first pathogen-derived serpin with the unusual ability to efficiently inhibit different proteases at several active sites (see Abstract) as taught by Ksiazek et. al. (“Miropin, a Novel Bacterial Serpin from the Periodontopathogen Tannerella forsythia, Inhibits a Broad Range of Proteases by Using Different Peptide Bonds within the Reactive Center Loop”; Miroslaw Ksiazek, Danuta Mizgalska, Jan J Enghild, Carsten Scavenius, Ida B Thogersen, Jan Potempa; J Biol Chem.;290(1):658–670; published November 11, 2014, cited in IDS filed 11/10/2023). Ksiazek et. al. disclosed serpin from T. forsythia (miropin) inhibits a broad array of proteases with divergent specificities (see Results in the Abstract section). Ksiazek et. al. characterized a novel serpin (miropin) from the human pathogen Tannerella forsythia, a bacterium implicated in initiation and progression of human periodontitis; and also disclosed that in contrast to other serpins, miropin efficiently inhibited a broad range of proteases (see page 658, left col, paragraph 1). Ksiazek et. al. further elaborates that the unusually broad specificity of miropin for target proteases is achieved through different active sites within the reactive center loop upstream of the P1-P1’ site, which was predicted from an alignment of the primary structure of miropin with those of well studied human and prokaryotic serpins; thus, miropin is unique among inhibitory serpins, and it has apparently evolved the ability to inhibit a multitude of proteases at the expense of a high stoichiometry of inhibition and a low association rate constant (see page 658, left col, paragraph 1). Ksiazek et. al. expressed, purified, and characterized miropin (i.e., T. forsythia ATCC 43037 was grown, which is a wild-type T. forsythia is a Gram-negative, asaccharolytic, anaerobic implicated in the development and progression of periodontitis in humans rod-shaped bacteria [page 659, left col, paragraph 3, Figure 1A multisequence alignment with wild-type T. forsythia Miropin]. Ksiazek et. al. explains in periodontal pockets, subgingival dental plaque (a dwelling place of T. forsythia) is bathed in gingival crevicular fluid. This inflammatory exudate contains high levels of neutrophil-derived serine proteases (e.g. neutrophil elastase; and by degrading bacterial proteins, these proteases play an important role in the innate immune system of the host (see page 659, left col, paragraph 4). Ksiazek et. al. teaches miropin efficiently inhibits a broad range of proteases with vastly different specificities; and the ability of miropin to inhibit a wide variety of proteases is apparently due to the use of different peptide bonds within the reactive center loop (RCL) (see page 665, right col, paragraph 1-2). And finally, Ksiazek et. al. teaches miropin has acquired the ability to inhibit efficiently an unusually broad range of proteases by using different sites in the RCL; this evolution was probably driven by adaptation to the biological niche of subgingival bacterial biofilm on the dental surface, which is rich in serine proteases originating from other bacteria and the host (see page 669, right col, paragraph 2). Therefore, this is presumed to be sufficient information for a person of skill in the art to know which residues are important for inhibiting broad array of proteases with divergent specificities.
As such, there are no elements added to the judicial exception sufficient to render the claims significantly more than the exception (MPEP § 2106.05).
Taken together, the claims are drawn to patent ineligible subject matter and are rejected here. Therefore, claims 1-3 are patent ineligible.
Conclusion
No claim is allowed.
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/KOYELI BANERJEE/Examiner, Art Unit 1658
/Melissa L Fisher/Supervisory Patent Examiner, Art Unit 1658
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