DETAILED CORRESPONDENCE
Status of the Application
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Claims 23, 25-26, 30-34, 36-37, 40-42, 44-45 and 48 are pending in this application.
Applicant’s amendment to the claims filed 04/29/2026 is acknowledged. This listing of the claims replaces all prior versions and listings of the claims.
Applicant’s remarks filed on 04/29/2026 in response to the non-final rejection mailed on 12/29/2025 are acknowledged and have been fully considered.
Regarding claim 43, applicant is reminded that claim amendments must adhere to 37 CFR 1.121(c)(4) which states that no claim text shall be presented for any claim in the claim listing with the status of “canceled”.
The rejections to claims 46-47 are withdrawn in view of the cancelation of claims 46-47.
The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action.
Election
The elected subject matter is
Group I, corresponding to claims 23, 25-26, 30-34, 36-37, 40 and 48, drawn to the technical feature of a method for conjugating a free thiol group of a moiety of a biomolecule, comprising contacting the biomolecule with a gold-donor agent to form a S-Au-S bond, characterized in that the gold-donor agent is halogen(triarylphosphine)gold (I),
elected with traverse in the reply filed 06/14/2024.
Claims 41-42 and 44-45 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim.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 with traverse in the reply filed on 06/14/2024.
Claims 23, 25-26, 30-34, 36-37, 40 and 48 are being examined on the merits.
Claim Rejections - 35 USC § 112(b)
Claims 23, 25-26, 30-34, 36-37, 40 and 48 are rejected under 35 U.S.C. 112(b) as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor regards as the invention.
The instant rejection is maintained from the previous office action, and any newly recited portion is necessitated by claim amendment.
Claim 23 (claims 25-26, 30-34, 36-37, 40 and 48 dependent therefrom) is indefinite for the phrase “providing a Geobacillus stearothermophilus TRAP protein containing a mutation of the lysine residue at amino acid position 35 to cysteine”. As there is no recited reference sequence for comparison, the location of the recited mutation in the claimed TRAP protein is unknown.
Claim 23 (claims 25-26, 30-34, 36-37, 40 and 48 dependent therefrom) recites “the lysine residue at amino acid position 35” in line 3. There is insufficient antecedent basis for this limitation in the claim.
Claim 48 is indefinite for the recitation of “wherein the Geobacillus stearothermophilus TRAP protein further contains a mutation of the arginine residue at amino acid position 64 to serine”. As there is no recited reference sequence for comparison, the location of the recited mutation in the claimed TRAP protein is unknown.
Claim 48 recites “the arginine residue at amino acid position 64” in line 2. There is insufficient antecedent basis for this limitation in the claim.
Response to Remarks: beginning on page 6 of Applicant’s response to rejections under 35 USC 112(b);
Applicant in summary contends the limitation of claim 46 have been amended into claim 23 to overcome the rejection of claim 23, as claim 46 was not subject to the same rejection;
Applicant further contends one of skill in the art would understand precisely what TRAP protein is being claimed as amended as the TRAP protein is a well-known and well-characterized protein, and the sequence is referenced in the specification on page 12 as PDB accession 4V4F9.
Applicant’s remarks are considered and found not convincing. Claim 46 was previously rejected under 112b citing the indefiniteness of the location of the cysteine as a result of the lack of recited reference sequence for comparison with which to determine the location of the mutation in the claimed protein. Therefore, incorporating the limitations of claim 46 into claim 23 do not overcome the 112b rejections of record.
Regarding the assertion that one of skill in the art would understand precisely what TRAP protein is being claimed:
the claims recite a protein with no defined structure aside from a single mutation at a specified amino acid position, and without a reference sequence for comparison to determine the location of that amino acid position. While Applicant references to the sequence of the TRAP protein as PDB accession 4V4F9 in the instant specification at page 12 in reference to initial atomic coordinate modeling, the referenced PDB Accession No. is not defined as the Geobacillus stearothermophilus TRAP protein recited in the claims. It is also noted that PDB has no apparent record of 4V4F9 [see Appendix A]. Additionally, page 12 of the specification recites “the mutated … residue was assigned to residue 37 in the original PDB file 4V4F but corresponds to position 35 in our analyses” [page 12, bottom], indicating that the positions of amino acids were changed from a sequence in the art. Therefore, the reference to a PDB Accession No. does not clearly and definitely define the structure of the TRAP protein recited in the claims. Furthermore, such reference of amino acid sequence is not adherent to 37 CFR 1.821(c) which dictates disclosures of amino acid sequences of 4 or more amino acids must contain a “Sequence Listing”, which is a separate part of the specification containing each of those amino acid sequences and associated information using the symbols and format in accordance with the requirements of 37 CFR 1.822 and 1.823. Additionally, 37 CFR 1.821(d) establishes where the description or claims of a patent application discuss a sequence that is set forth in the "Sequence Listing," in accordance with paragraph (c) of this section, reference must be made to the sequence by use of the sequence identifier, preceded by "SEQ ID NO:" or the like, in the text of the description or claims, even if the sequence is also embedded in the text of the description or claims of the patent application. Therefore, as the claims do not recite any reference sequence for comparison, the location of the recited amino acid positions of the protein in the claim is considered indefinite.
Claim Rejections - 35 USC § 112(a)
Claim Interpretation: The claims are drawn to a method of conjugating a free thiol group of a moiety of a TRAP protein, comprising
providing a Geobacillus stearothermophilus TRAP (GS-TRAP) protein containing a mutation of the lysine residue at amino acid position 35 to cysteine,
contacting the TRAP protein with a gold-donor agent to form a S-Au-S bond, characterized in that the gold-donor agent is halogen(triarylphosphine)gold (I),
wherein the moiety with the free thiol group is the cysteine moiety at amino acid position 35, and
generating a protein cage biomolecule complex.
According to the instant specification, TRAP proteins are trp RNA binding attenuation proteins from Geobacillus- stearothermophilus [p 1, final paragraph], and according to Bayfield et al. (Plos One, 2012, 7:e44309; cited on the Form PTO-892 mailed 09/04/2024; herein referred to as Bayfield), TRAP proteins are responsible for regulating L-tryptophan biosynthesis in many Bacilli and can be composed of either 11 or 12 subunits depending on the sequence [p 1, col 2, para 2].
The limitation “a Geobacillus stearothermophilus TRAP protein containing a mutation of the lysine residue at amino acid position 35 to cysteine” in claim 23 is considered a product-by-process limitation (MPEP 2113.I). The phrase “Geobacillus stearothermophilus TRAP protein” is considered the source of the TRAP protein recited in the claim, and therefore phrase is drawn to the process of making the TRAP protein recited in the claims, as the recited TRAP protein being used in the claimed method contains a mutation. As the transitional phrase “containing” is synonymous with “comprising” which is an inclusive and open-ended term that does not exclude additional unrecited elements (MPEP 2111.03.I), the TRAP protein being used in the claimed method can contain additional mutations. In view of these interpretations, the phrase “Geobacillus stearothermophilus TRAP protein” is not considered structurally limiting on the TRAP protein being used in the claimed method, and therefore the only structural requirement of this TRAP protein is the amino acid cysteine. As there is no sequence for comparison to determine the location of the cysteine residue of the claimed TRAP protein, the TRAP protein of the independent claim 23 is only limited to have a cysteine residue.
Claim 48 limits the claimed TRAP protein further contain mutation of arginine at position 64 serine, however as there is no sequence for comparison to determine the location of the serine residue of the claimed TRAP protein, the TRAP protein of the independent claim 48 is only limited to have a cysteine and a serine residue, and is not limited to have any additional amino acids aside from the residues stated in the claims.
In this case, the genus of recited TRAP proteins encompasses species that are considered to be widely variant with respect to sequence.
A. Claims 23, 25-26, 30-34, 36-37, 40 and 48 are rejected under 35 U.S.C. 112(a) as failing to comply with the written description requirement. The claim(s) contains subject matter which was not described in the specification in such a way as to reasonably convey to one skilled in the relevant art that the inventor or a joint inventor at the time the application was filed, had possession of the claimed invention.
The instant rejection is maintained from the previous Office Action and any newly recited portions are necessitated by claim amendment.
MPEP 2163.II.A.2.(a).i) states, “Whether the specification shows that applicant was in possession of the claimed invention is not a single, simple determination, but rather is a factual determination reached by considering a number of factors. Factors to be considered in determining whether there is sufficient evidence of possession include the level of skill and knowledge in the art, partial structure, physical and/or chemical properties, functional characteristics alone or coupled with a known or disclosed correlation between structure and function, and the method of making the claimed invention”.
For claims drawn to a genus, MPEP § 2163 states the written description requirement for a claimed genus may be satisfied through sufficient description of a representative number of species by actual reduction to practice, reduction to drawings, or by disclosure of relevant, identifying characteristics, i.e., structure or other physical and/or chemical properties, by functional characteristics coupled with a known or disclosed correlation between function and structure, or by a combination of such identifying characteristics, sufficient to show the applicant was in possession of the claimed genus. See Eli Lilly, 119 F.3d at 1568, 43 USPQ2d at 1406.
According to MPEP 2163.II.A.3.(a).ii), [s]atisfactory disclosure of a ‘representative number’ depends on whether one of skill in the art would recognize that the applicant was in possession of the necessary common attributes or features possessed by the members of the genus in view of the species disclosed. For inventions in an unpredictable art, adequate written description of a genus which embraces widely variant species cannot be achieved by disclosing only one species within the genus…Instead, the disclosure must adequately reflect the structural diversity of the claimed genus, either through the disclosure of sufficient species that are ‘representative of the full variety or scope of the genus,’ or by the establishment of ‘a reasonable structure-function correlation.’"
The factors considered in the Written Description requirement are (1) level of skill and knowledge in the art, (2) partial structure, (3) physical and/or chemical properties, (4) functional characteristics alone or coupled with a known or disclosed correlation between structure and function, and the (5) method of making the claimed invention. Disclosure of any combination of such identifying characteristics that distinguish the claimed invention from other materials and would lead one of skill in the art to the conclusion that the applicant was in possession of the claimed species is sufficient." MPEP § 2163.
The claims recite (in relevant part) a genus of TRAP proteins. As stated above, with the exception of a cysteine residue, the amino acid sequences of the genus of TRAP proteins are unlimited. In this case, the genus of recited TRAP proteins encompasses species that are considered to be widely variant with respect to sequence.
The specification discloses the following representative species of the genus of recited variant polypeptides: A TRAP-CS mutant, containing K35C and R64S mutations. The specification does not disclose any additional mutation(s) of the TRAP-CS sequence aside from the K35C and R64S mutations.
Regarding the level of skill and knowledge in the art of amino acid modification, the reference of Singh et al. (Curr. Protein Pept. Sci. 18:1-11, 2017; cited on the Form PTO-892 mailed 09/04/2024) reviews various protein engineering methods and discloses that despite the availability of an ever-growing database of protein structures and highly sophisticated computational algorithms, protein engineering is still limited by the incomplete understanding of protein functions, folding, flexibility, and conformational changes (see p. 7, column 1, top). Also, the unpredictability associated with residue substitution is exemplified by the reference of Zhang et al. (Structure 26:1474-1485, 2018; cited on the Form PTO-892 mailed 09/04/2024), which discloses that even a substitution of a surface residue that was predicted to be benign caused significant structural changes and unexpected effects on the function of a polypeptide (p. 1475, column 1).
In view of the high level of unpredictability in the art of amino acid modification, because the genus of TRAP proteins is widely variant with respect to structure, and the specification discloses the actual reduction to practice of only one representative species among a widely variant genus, but without any accompanying amino acid sequence, one of skill in the art would reasonably conclude that the disclosure fails to provide a representative number of species to describe the genus, and thus, that the applicant was not in possession of the recited genus of TRAP proteins. The claimed subject matter is not supported by an adequate written description because a representative number of species has not been described.
B. Claims 23, 25-26, 30-34, 36-37, 40 and 48 are rejected under 35 U.S.C. 112(a) because the specification, while being enabling for the TRAP-CS protein containing K25C and R64S mutations, does not reasonably provide enablement for all TRAP proteins as encompassed by the claims. The specification does not enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and/or use the invention commensurate in scope with these claims.
The instant rejection is maintained from the previous Office Action and any newly recited portions are necessitated by claim amendment.
“The test of enablement is not whether any experimentation is necessary, but whether, if experimentation is necessary, it is undue.” In re Angstadt, 537 F.2d 498, 504, 190 USPQ 214, 219 (CCPA 1976). Factors to be considered in determining whether undue experimentation is required are summarized in In re Wands (858 F.2d 731, 737, 8 USPQ2d 1400, 1404 (Fed. Cir. 1988)) as follows: (A) The breadth of the claims; (B) The nature of the invention; (C) The state of the prior art; (D) The level of one of ordinary skill; (E) The level of predictability in the art; (F) The amount of direction provided by the inventor; (G) The existence of working examples; and (H) The quantity of experimentation needed to make or use the invention based on the content of the disclosure. See MPEP § 2164.01(a). The Factors considered to be most relevant to the instant rejection are addressed in detail below.
The nature of the invention: According to the specification at p 3, “The subject matter of the invention is a method for conjugation of free thiol group(s) moiety(s) of biomolecules, leading to the biomolecular complex formation, comprising a reaction between biomolecules and gold-donor agent in which -S-Au-S- bond is formed, wherein a gold- donor agent is halogen(triarylphosphine)gold (I). Preferably the biomolecules used in the method are selected from the group comprising peptides, polypeptides, proteins.” The object of the invention is therefore to provide a method for bioconjugation of biomolecules, utilizing thioaurate formation chemistry.
The breadth of the claims: The claims recite (in relevant part) a TRAP protein used in the method for bioconjugation and formation of the -S-Au-S- bond. As stated above, with the exception of a cysteine, the amino acid sequence of the TRAP polypeptide is unlimited.
The state of the prior art; The level of one of ordinary skill; and The level of predictability in the art: According to MPEP 2164.03, “…what is known in the art provides evidence as to the question of predictability” and “[I]f one skilled in the art cannot readily anticipate the effect of a change within the subject matter to which that claimed invention pertains, then there is lack of predictability in the art.”
As noted above, the amino acid sequence of the recited polypeptides is unlimited. The reference of Singh (supra) reviews various protein engineering methods and discloses that despite the availability of an ever-growing database of protein structures and highly sophisticated computational algorithms, protein engineering is still limited by the incomplete understanding of protein functions, folding, flexibility, and conformational changes (see p. 7, column 1, top).
The unpredictability associated with amino acid modification is exemplified by the reference of Zhang (supra) which discloses that even a mutation that was predicted to be benign caused significant structural changes and unexpected effects on the function of a polypeptide (p. 1475, column 1).
While the instant specification discloses a TRAP protein wherein the only identified sequence features are mutations to introduce cysteine [p 9, Example 1], both the instant specification at [p 5, final paragraph] and Heddle et al. (Small, 2007, 11:1950; cited on the Form PTO-892 mailed 09/04/2024) at [p 1951, col 2, para 1] disclose that TRAP proteins lack naturally-occurring cysteine residues that are understood to be required to use the claimed method, indicating that not all TRAP proteins would be able for use in the claimed method without the modification to add a cysteine in the correct location.
As such, one of skill in the art would recognize a high level of unpredictability that all TRAP proteins as encompassed by the claims would be contain the required amino acids to maintain the desired activity/utility.
The amount of direction provided by the inventor and The existence of working examples: The specification discloses the following representative species of the recited TRAP proteins: A TRAP-CS protein with K25C and R64S mutations.
The specification does not disclose any additional mutation(s) or amino acids of the TRAP-CS sequence aside from the K35C and R64S mutations and other than this working example, the specification fails to disclose any other TRAP protein.
The quantity of experimentation needed to make or use the invention based on the content of the disclosure: While methods of modifying the amino acid sequence of a polypeptide were known at the time of the invention, it was not routine in the art to make and determine a use for all TRAP polypeptides as recited by the claims.
In view of the overly broad scope of the claims, the lack of guidance and working examples provided in the specification, the high level of unpredictability, and the state of the prior art, undue experimentation would be necessary for a skilled artisan to make and use the entire scope of the claimed invention. Applicants have not provided sufficient guidance to enable one of ordinary skill in the art to make and use the claimed invention in a manner reasonably correlated with the scope of the claims. The scope of the claims must bear a reasonable correlation with the scope of enablement (In re Fisher, 166 USPQ 19 24 (CCPA 1970)). Without sufficient guidance, determination of having the desired biological characteristics is unpredictable and the experimentation left to those skilled in the art is unnecessarily, and improperly, extensive and undue. See In re Wands 858 F.2d 731, 8 USPQ2nd 1400 (Fed. Cir, 1988).
C. Claims 23, 25-26, 30-34, 36-37, 40 and 48 are newly rejected under 35 U.S.C. 112(a) as failing to comply with the written description requirement. The claim(s) contains subject matter which was not described in the specification in such a way as to reasonably convey to one skilled in the relevant art that the inventor or a joint inventor, at the time the application was filed, had possession of the claimed invention.
This is a new matter rejection and is necessitated by amendment.
MPEP § 2163.II.A.3.(b) states, “when filing an amendment an applicant should show support in the original disclosure for new or amended claims.” See also MPEP 714.02. MPEP § 2163.II.A.3.(b) further states, “[i]f the originally filed disclosure does not provide support for each claim limitation, or if an element which applicant describes as essential or critical is not claimed, a new or amended claim must be rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112, para. 1, as lacking adequate written description.” According to MPEP § 2163.I.B, “While there is no in haec verba requirement, newly added claim limitations must be supported in the specification through express, implicit, or inherent disclosure” and “The fundamental factual inquiry is whether the specification conveys with reasonable clarity to those skilled in the art that, as of the filing date sought, applicant was in possession of the invention as now claimed. See, e.g., Vas-Cath, Inc., 935 F.2d at 1563-64, 19 USPQ2d at 1117.”
Claim 23 (claims 25-26, 30-34, 36-37, 40 and 48 dependent therefrom) has been amended to recite “providing a Geobacillus stearothermophilus TRAP protein containing a mutation of the lysine residue at amino acid position 35 to cysteine”, and
Claim 48 has been amended to recite “wherein the Geobacillus stearothermophilus TRAP protein further contains a mutation of the arginine residue at position 64 to serine”.
Applicant has not acknowledged any support for the instant amendments to the claims, and there is no apparent descriptive support for the limitations “providing a Geobacillus stearothermophilus TRAP protein containing a mutation of the lysine residue at amino acid position 35 to cysteine” and “wherein the Geobacillus stearothermophilus TRAP protein further contains a mutation of the arginine residue at position 64 to serine” in the original application as filed. Absent descriptive support, the noted limitation is considered to introduce new matter into the claims. Applicant is invited to show support for the limitations at issue.
Response to Remarks: beginning on page 8 of Applicant’s response to 35 U.S.C. 112(a) rejections; Applicant in summary contends a skilled artisan would understand that Applicant was in possession of the GS-TRAP protein containing a lysine residue at position 35 since the amino acid sequence of GS-TRAP is well-known, as is the location of position 35; Applicant further contends a skilled artisan would understand that a Geobacillus stearothermophilus TRAP protein containing a mutation of the lysine residue at position 35 could be generated without undue experimentation since the amino acid sequence of the GS-TRAP is well-known, as is the location of the lysine residue at position 35; while Applicant has not responded to the 112(a) rejection of new matter set forth in the previous office action, the instant remarks are considered a bona fide response to the rejections of record.
Applicant’s remarks are considered and found not convincing. As stated in the rejection above:
the limitation “a Geobacillus stearothermophilus TRAP protein containing a mutation of the lysine residue at amino acid position 35 to cysteine” in claim 23 is considered a product-by-process limitation (MPEP 2113.I). The phrase “Geobacillus stearothermophilus TRAP protein” is considered the source of the TRAP protein recited in the claim, and therefore phrase is drawn to the process of making the TRAP protein recited in the claims, as the recited TRAP protein being used in the claimed method contains a mutation. As the transitional phrase “containing” is synonymous with “comprising” which is an inclusive and open-ended term that does not exclude additional unrecited elements (MPEP 2111.03.I), the TRAP protein being used in the claimed method can contain additional mutations. In view of these interpretations, the phrase “Geobacillus stearothermophilus TRAP protein” is not considered structurally limiting on the TRAP protein being used in the claimed method, and therefore the only structural of this TRAP protein is the amino acid cysteine. As there is no sequence for comparison to determine the location of the cysteine residue of the claimed TRAP protein, the TRAP protein of the independent claim 23 is only limited to have a cysteine residue.
For these reasons, the claims are drawn to a genus of protein that are widely variant with respect to structure, and as the specification discloses the actual reduction to practice of only one representative species among a widely variant genus, but without any accompanying amino acid sequence, one of skill in the art would reasonably conclude that the disclosure fails to provide a representative number of species to describe the genus, and thus, that the applicant was not in possession of the recited genus of TRAP proteins.
As the TRAP protein recited in the claims is only limited to having a cysteine, the scope of the claimed method includes all TRAP proteins of unlimited structure with the exception of the recited cysteine, wherein the location of the recited cysteine is unlimited. In view of the overly broad scope of the claims, the lack of guidance and working examples provided in the specification, the high level of unpredictability, and the state of the prior art, undue experimentation would be necessary for a skilled artisan to make and use the entire scope of the claimed invention.
Claim Rejections - 35 USC § 103
Claims 23, 25-26, 30, 32-34, 36-37, 40 and 48 are rejected under 35 U.S.C. 103 as being unpatentable over Igashiri-Kamiyama et al. (Dalton Trans, 2011, 40:7249; cited on the IDS filed 05/25/2021; herein referred to as IK), Caddy et al. (Zeitshrift fur Naturforschung B, 2007, 62:460; cited on the IDS submitted 05/25/2021; herein referred to as Caddy), and Heddle et al. (Small, 2007, 11:1950; cited on the Form PTO-892 mailed 09/04/2024; herein referred to as Heddle) and evidentiary reference UniProt Accession No. MTRB_GEOSE (2 pages 11/02/2016; cited on the Form PTO-892 mailed 03/31/2025; herein referred to as UNI).
The instant rejection is maintained from the previous Office Action and any newly recited portions are necessitated by claim amendment.
Claim 23 is drawn to a method of conjugating a free thiol group of a moiety of a TRAP protein, comprising
(i) providing a Geobacillus stearothermophilus TRAP (GS-TRAP) protein containing a mutation of the lysine residue at amino acid position 35 to cysteine,
(ii) contacting the TRAP protein with a gold-donor agent to form a S-Au-S bond, characterized in that the gold-donor agent is halogen(triarylphosphine)gold(I),
wherein the moiety with the free thiol group is the cysteine moiety at amino acid position 35, and
(iii) generating a protein cage biomolecule complex.
As stated in the rejection of claims under 35 USC 112(a) above, the limitation “a Geobacillus stearothermophilus TRAP protein containing a mutation of the lysine residue at amino acid position 35 to cysteine” in claim 23 is considered a product-by-process limitation (MPEP 2113.I). The phrase “Geobacillus stearothermophilus TRAP protein” is considered the source of the TRAP protein used in the claimed method, and therefore phrase is drawn to the process of making the TRAP protein used in the claimed method, as the TRAP protein used in the claimed method contains a mutation. As the transitional phrase “containing” is synonymous with “comprising” which is an inclusive and open-ended term that does not exclude additional unrecited elements (MPEP 2111.03.I), the TRAP protein being used in the claimed method can contain additional mutations. In view of these interpretations, the phrase “Geobacillus stearothermophilus TRAP protein” is not considered structurally limiting on the TRAP protein used in the claimed method, and therefore the only structural requirement of this TRAP protein is the amino acid cysteine. As there is no sequence for comparison to determine the location of the cysteine residue of the claimed TRAP protein, the TRAP protein of the independent claim 23 is only limited to have a cysteine residue, and therefore encompasses a protein with a cysteine residue at any location.
IK discusses the creation of multinuclear and metallosupramolecular compounds from thiol-containing amino acids [title].
Regarding claim 23 and the limitation of conjugating a free thiol group of a moiety of a biomolecule, comprising contacting the biomolecule with a gold-donor agent to form a S-Au-S bond, IK teaches a mononuclear AuI complex [Au(D-pen)2]3- comprising two D-pen ligands bound to an AuI center through thiolato S atoms [abstract, also shown as a reactant in Scheme 1], wherein the D-pen ligands are amino acids bearing free-thiols. IK further discloses the use of such thiolato metal complexes as a metalloligand is of interest in the rational construction of well-organized S-bridged multinuclear structures.
IK does not teach the use of halogen(triarylphosphine)gold (I) donor agents in step (ii), TRAP proteins containing cysteine in step (i), or generating a protein cage in step (iii).
Caddy describes the introduction of phosphine-gold(I) precursors into Cysteine-modified neuropeptides [title], and discusses that gold(I) has a known preference for thiol ligands.
Regarding claim 23 and the limitations in step (ii) of the use of halogen(triarylphosphine)gold (I) donor agents, Caddy teaches chlorine(triphenylphosphine)gold(I) [(Ph3P)AuCl, shown as molecule 1 in Scheme 2] is used as gold donor to form an S-Au linked biomolecule comprising the Ph3P group and a Boc-protected cysteine [shown as molecule 3 in Scheme 2].
Heddle describes using ring-shaped protein TRAP to capture and confine gold nanodots [title], and discusses ring proteins, rather than commonly used spherical protein cages, can be used to constrain gold nanodots via formation of gold-protein complexes [abstract].
Regarding claim 23 and the limitation of a of generating a protein cage biomolecule complex comprising the TRAP proteins that contain cysteine in steps (i) and (iii), Heddle teaches cysteine-bearing mutants of the B. stearothermophilus TRAP protein can be used to provide sulfur atoms for the formation of thioaurate bonds that fix gold nanodots into the central cavity of the TRAP ring [p 1951, col 2, para 1]. The TRAP protein of Heddle is based off of the B. stearothermophilus TRAP [p 1951, col 2, para 1] with the PDB Accession No. 1QAW, which is considered to correspond to UniProt Accession No. MTRB_GEOSE (2 pages 11/02/2016; cited on the Form PTO-892 mailed 03/31/2025; herein referred to as UNI). In view of the interpretation set forth above, and the lack of a reference sequence for comparison as described in the rejection of the claims under 35 USC 112(b) above, it is unclear where in the claimed TRAP protein the mutations are to occur, and therefore the TRAP protein of Heddle that comprises a cysteine is considered to satisfy the structural limitations of “a Geobacillus stearothermophilus TRAP protein containing a mutation of the lysine residue at amino acid position 35 to cysteine” set forth in the claim.
In view of IK, Heddle and Caddy, it would have been prima facie obvious to one of ordinary skill in the art to modify the method of Caddy by using free thiol-containing amino acids as disclosed by IK that occur in the mutant TRAP protein of Heddle, because Caddy discloses the use of a halogen(triarylphosphine)gold (I) donor agent to form S-Au linked biomolecules, and IK discloses the formation of S-Au-S linked biomolecules, and Heddle discloses the use of cysteine in mutant TRAP proteins used for the formation of thioaurate bonds. One of ordinary skill in the art would have been motivated to modify the method of Caddy because IK teaches the production of thiolato metal complexes is of interest in the rational construction of well-organized S-bridged multinuclear structures, and Heddle teaches cysteine-bearing TRAP protein mutants can function as a protein cage wherein the sulfur atoms of cysteine form a thioaurate bond with gold. One of ordinary skill in the art would have had a reasonable expectation of success because Caddy, IK and Heddle all discuss methods of generating S-Au linked biomolecules.
Regarding claim 25, Heddle teaches the complex of gold nanodot and TRAP protein [p 1951, col 2, para 1], wherein TRAP is comprised of multiple units of the same biomolecule [p 1951, col 1, para 2].
Regarding claim 26, Heddle teaches the mutated TRAP contains symmetry [p 1951, col 1, para 5], wherein one of skill in the art would conclude the complex comprising gold nanodot within the TRAP ring would be symmetrical.
Regarding claims 30 and 32, Caddy teaches the use of the gold donor chlorine(triphenylphosphine)gold(I) [(Ph3P)AuCl, shown as molecule 1 in Scheme 2], which corresponds to the halogen chlorine and a polysubstituted phenyl.
Regarding claim 33, Heddle teaches the immobilization of TRAP-gold complexes on SiO2 followed by the thorough washing with pure water [p 1955, col 2, para 5], corresponding to the purifying of the conjugation product.
Regarding claim 34, Heddle teaches the sequence optimization of the genes encoding TRAP from Bacillus stearothermophilus and subsequent expression in E. coli BL21 (DE3) cells followed by purification via Q-sepharose and HiLoad superdex 200 gel filtration columns [p 1955, col 1, para 2].
Regarding claim 36, Heddle teaches the TRAP adsorption and Au-nanoparticle immobilization on an Si substrate were carried out by a method comprising incubating TRAP in a solution 10 mM Tris-HCl and 50 mM NaCl containing Au-nanoparticles at room temperature for 3 hours [p 1955, col 2, para 2], and Caddy teaches the combination of 0.167 mmol Boc-cysteine (corresponding to the biomolecule) with 0.167 mmol Ph3PAuCl (corresponding to the gold donor) [p 463, col 1, final paragraph], which falls within the claimed range of biomolecule : gold donor ratios of (3 to 1) : (1 to 4).
Regarding claim 37, Heddle teaches the spin drying of the immobilized TRAP-gold complexes on SiO2 [p 1955, col 2, para 5], which encompasses centrifugation.
Regarding claim 40, Heddle teaches the complexation of gold nanodot to mutated TRAP bearing one cysteine [p 1951, col 2, para 1] as discussed in the rejection of claim 23 above, wherein the mutant TRAP contains 12 subunits [p 1951, col 1, para 3], referred to as biomolecule units in the instant application. One of skill in the art would reasonably expect that a gold nanoparticle would react with two of such mutant TRAP proteins as suggested by the IK [as shown by the reactant in Scheme 1], thereby resulting in a protein complex consisting of 24 biomolecule units.
Regarding claim 48, in view of indefiniteness of the claim as set forth above, the phrases “wherein the Geobacillus stearothermophilus TRAP protein further contains a mutation of the arginine residue at position 64 to serine”, as stated in the rejection of claims under 35 USC 112(a) above, the limitation is considered a product-by-process limitation (MPEP 2113.I). The phrase “Geobacillus stearothermophilus TRAP protein” is considered the source of the TRAP protein used in the claimed method, and therefore phrase is drawn to the process of making the TRAP protein used in the claimed method, as the TRAP protein used in the claimed method contains a mutation. As the transitional phrase “containing” is synonymous with “comprising” which is an inclusive and open-ended term that does not exclude additional unrecited elements (MPEP 2111.03.I), the TRAP protein being used in the claimed method can contain additional mutations. In view of these interpretations, the phrase “Geobacillus stearothermophilus TRAP protein” is not considered structurally limiting on the TRAP protein used in the claimed method, and therefore the only structural requirement of this TRAP protein is the amino acid cysteine and the amino acid serine. As there is no sequence for comparison to determine the locations of the cysteine and serine residues of the claimed TRAP protein, the TRAP protein of claim 48 is only required to have a cysteine residue and a serine reside, and therefore encompasses a protein with a cysteine residue and a serine at any location.
Regarding claim 48, as there is no sequence for reference recited in the claims from which to determine the relative position 64 of the claimed TRAP protein, the TRAP protein of Heddle comprising a serine [see UNI] is considered to be encompassed by the structural limitations of claim 48.
Therefore, the invention of claims 23, 25-26, 30, 32-34, 36-37, 40 and 48 would have been obvious to one of ordinary skill in the art before the effective filing date.
Claim 31 is rejected under 35 U.S.C. 103 as being unpatentable over IK, Caddy, and Heddle as applied to claims 23, 25-26, 30, 32-34, 36-37, 40 and 48 above, and further in view of Hamon et al. (ChemComm, 2015, 15:16119; cited on the Form PTO-892 mailed 09/04/2024; herein referred to as Hamon).
The instant rejection is maintained from the previous Office Action and any newly recited portions are necessitated by claim amendment.
Claim 31 is drawn to the method of claim 23, wherein the gold-donor agent is chloro[diphenyl(3-sulfonatophenyl)phosphine]gold (I).
The teachings of IK, Caddy and Heddle as applied to claims 23, 25-26, 30, 32-34, 36-37, 40 and 48 are discussed above. These references do not teach chloro[diphenyl(3-sulfonatophenyl)phosphine]gold (I).
Hamon discusses an aqueous one-pot route to gold/quantum rod heterostructured nanoparticles functionalized with DNA [title], and describes that Au(I) organometallic chemistry provides a versatile approach to controlling surface grafting [abstract].
Regarding claim 31, Hamon teaches the use of the monovalent gold complex chloro-Au(I) known as chloro[diphenyl(3-sulfonatophenyl) phosphine]gold(I) for the conjugation of a heterostructured nanoparticle (HNP) with said gold complex [Scheme 1].
In view of Hamon, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date to modify the combined method of Heddle, Caddy and IK by using chloro[diphenyl(3-sulfonatophenyl) phosphine]gold(I), since the simple substitution of one known element for another results in a predictable result. One of ordinary skill in the art would have recognized that both chloro[diphenyl(3-sulfonatophenyl) phosphine]gold(I) and chlorine(triphenylphosphine)gold(I) are halogen(triarylphosphine)gold(I) molecules used as gold-donor agents for the conjugation of gold-biomolecule complexes, and as such both are capable of being incorporated into methods as described by Heddle, Caddy and IK. Thus it would have been obvious to one of ordinary skill in the art to replace chlorine(triphenylphosphine)gold(I) with chloro[diphenyl(3-sulfonatophenyl) phosphine]gold(I), as one of ordinary skill in the art would have been able to carry out such a substitution with a reasonable expectation of success because both Caddy and Hamon discuss the use of halogen(triarylphosphine)gold(I) molecules as gold-donor agents.
Therefore, the invention of claim 31 would have been obvious to one of ordinary skill in the art before the effective filing date.
Response to remarks: beginning on page 9 of Applicant’s response to rejections under 35 USC 103; Applicant in summary contends the amended claims recite “a Geobacillus stearothermophilus TRAP protein containing a mutation of the lysine residue at amino acid position 35 to cysteine”, which is neither taught nor suggested by the prior art of record, and therefore the claims are not obvious over the prior art of record.
Applicant’s remarks are considered and found not convincing. The claim interpretations and teachings of the art are described in detail above. Briefly, the phrase “a Geobacillus stearothermophilus TRAP protein” does not structurally limit the TRAP protein used in the claimed method as the GS-TRAP protein is considered the source protein used in the method of generating the protein used in the claimed method which has a cysteine mutation. As the claims recite no reference sequence for comparison, the location of the cysteine mutation is indefinite, and therefore the TRAP protein used in the method is only required to have a cysteine amino acid, which can occur at any location in the protein. As Heddle teaches the use of a TRAP protein containing a cysteine from Bacillus stearothermophilus, which is recognized as the same organism Geobacillus stearothermophilus as confirmed by Applicant’s response [page 7, middle], the protein of Heddle is encompassed by the structural limitations of the TRAP protein used in the claimed method.
Double Patenting
Claims 23, 25-26, 30, 32-34, 36-37, 40 and 48 are provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claim 54 of copending Application No. 18/547,242 (herein “reference application”) in view of IK, Caddy, Heddle and evidentiary reference UNI.
The instant rejection is maintained from the previous Office Action and any newly recited portions are necessitated by claim amendment.
Regarding instant claim 23, claim 54 of the reference application recites a method of making an artificial TRAP-cage comprising conjugation of the TRAP ring units via at least one free thiol linkage with a cross-linker.
The claims of the reference application do not recite a TRAP protein with a cysteine, contacting the TRAP with a gold-donor agent to form a S-Au-S bond, and the gold-donor agent is halogen(triarylphosphine)gold(I).
IK discusses the creation of multinuclear and metallosupramolecular compounds from thiol-containing amino acids [title].
Regarding instant claim 23 and the limitation of conjugating a free thiol group of a moiety of a biomolecule, comprising contacting the biomolecule with a gold-donor agent to form a S-Au-S bond, IK discloses a mononuclear AuI complex [Au(D-pen)2]3- comprising two D-pen ligands bound to an AuI center through thiolato S atoms [abstract, also shown as a reactant in Scheme 1], wherein the D-pen ligands are amino acids bearing free-thiols. IK further discloses the use of such thiolato metal complexes as a metalloligand is of interest in the rational construction of well-organized S-bridged multinuclear structures.
Caddy describes the introduction of phosphine-gold(I) precursors into Cys-modified neuropeptides [title], and discusses that gold(I) has a known preference for thiol ligands.
Regarding instant claim 23 and the limitation of the use of halogen(triarylphosphine)gold (I) donor agents, Caddy discloses chlorine(triphenylphosphine)gold(I) [(Ph3P)AuCl, shown as molecule 1 in Scheme 2] is used as gold donor to form an S-Au linked biomolecule comprising the Ph3P group and a Boc-protected cysteine [shown as molecule 3 in Scheme 2].
Heddle describes using ring-shaped protein TRAP to capture and confine gold nanodots [title], and discusses ring proteins, rather than commonly used spherical protein cages, can be used to constrain gold nanodots via formation of gold-protein complexes [abstract].
Regarding instant claim 23 and the limitation of a of generating a protein cage biomolecule complex comprising the TRAP proteins that contain cysteine in steps (i) and (iii), Heddle discloses cysteine-bearing mutants of the B. stearothermophilus TRAP protein can be used to provide sulfur atoms for the formation of thioaurate bonds that fix gold nanodots into the central cavity of the TRAP ring [p 1951, col 2, para 1]. The TRAP protein of Heddle is based off of the B. stearothermophilus TRAP [p 1951, col 2, para 1] with the PDB Accession No. 1QAW, which is considered to correspond to UniProt Accession No. MTRB_GEOSE (2 pages 11/02/2016; cited on the Form PTO-892 mailed 03/31/2025; herein referred to as UNI). In view of the lack of a reference sequence for comparison as described in the rejection of the claims under 35 USC 112(b) above, it is unclear where in the claimed TRAP protein the mutations are to occur, and therefore the TRAP protein of Heddle that comprises a cysteine is considered to satisfy the limitation of “a TRAP protein with a cysteine at the outer perimeter of the TRAP protein at the amino acid position corresponding to position 35 in Geobacillus stearothermophilus”.
In view of Heddle, IK and Caddy, it would have been obvious to one of ordinary skill in the art to modify the method of the reference application by using halogen(triarylphosphine)gold (I) donor agents disclosed by Caddy with the free thiol-containing amino acids as disclosed by IK that occur in the mutant TRAP protein of Heddle, because Caddy discloses the use of a halogen(triarylphosphine)gold (I) donor agent to form S-Au linked biomolecules, and IK discloses the formation of S-Au-S linked biomolecules, and Heddle discloses the use of cysteine in mutant TRAP proteins used for the formation of thioaurate bonds. One of ordinary skill in the art would have been motivated to modify the method of the reference application because Caddy discloses gold(I) has a known preference for thiol ligands, IK discloses the production of thiolato metal complexes is of interest in the rational construction of well-organized S-bridged multinuclear structures, and Heddle discloses the mutated TRAP protein can function as a protein cage. One of ordinary skill in the art would have had a reasonable expectation of success because the reference application, Caddy, IK and Heddle all discuss methods of generating thiol-linked biomolecules.
Regarding instant claim 25, Heddle discloses the complex of gold nanodot and TRAP protein [p 1951, col 2, para 1], wherein TRAP is comprised of multiple units of the same biomolecule [p 1951, col 1, para 2].
Regarding instant claim 26, Heddle discloses the mutated TRAP contains symmetry [p 1951, col 1, para 5], wherein one of skill in the art would conclude the complex comprising gold nanodot within the TRAP ring would be symmetrical.
Regarding instant claims 30 and 32, Caddy discloses the use of the gold donor chlorine(triphenylphosphine)gold(I) [(Ph3P)AuCl, shown as molecule 1 in Scheme 2], which corresponds to the halogen chlorine and a polysubstituted phenyl.
Regarding instant claim 33, Heddle discloses the immobilization of TRAP-gold complexes on SiO2 followed by the thorough washing with pure water [p 1955, col 2, para 5], corresponding to the purifying of the conjugation product.
Regarding instant claim 34, Heddle discloses the sequence optimization of the genes encoding TRAP from Bacillus stearothermophilus and subsequent expression in E. coli BL21 (DE3) cells followed by purification via Q-sepharose and HiLoad superdex 200 gel filtration columns [p 1955, col 1, para 2].
Regarding instant claim 36, Heddle discloses the TRAP adsorption and Au-nanoparticle immobilization on an Si substrate were carried out by a method comprising incubating TRAP in a solution 10 mM Tris-HCl and 50 mM NaCl containing Au-nanoparticles at room temperature for 3 hours [p 1955, col 2, para 2], and Caddy discloses the combination of 0.167 mmol Boc-cysteine (corresponding to the biomolecule) with 0.167 mmol Ph3PAuCl (corresponding to the gold donor) [p 463, col 1, final paragraph], which falls within the claimed range of biomolecule : gold donor ratios of (3 to 1) : (1 to 4).
Regarding instant claim 37, for the sake of advancing prosecution, the phrase “comprising at least one of filtration, crystallization, centrifugation, and column chromatography” is being interpreted as “comprising at least one selected from the group consisting of filtration, crystallization, centrifugation, and column chromatography”. Heddle discloses the spin drying of the immobilized TRAP-gold complexes on SiO2 [p 1955, col 2, para 5], which encompasses centrifugation.
Regarding instant claim 40, Heddle discloses the complexation of gold nanodot to mutated TRAP bearing one cysteine [p 1951, col 2, para 1] as discussed in the rejection of claim 23 above, wherein the mutant TRAP contains 12 subunits [p 1951, col 1, para 3], referred to as biomolecule units in the instant application. One of skill in the art would reasonably expect that a gold nanoparticle would react with two of such mutant TRAP proteins as disclosed by the IK [as shown by the reactant in Scheme 1], thereby resulting in a protein complex consisting of 24 biomolecule units.
Regarding instant claim 48, as there is no sequence for reference recited in the claims from which to determine the relative position 64 of the claimed TRAP protein, the TRAP protein of Heddle comprising a serine [see UNI] is considered to be encompassed by the limitations of instant claim 48.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claim 31 is provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claim 54 of copending Application No. 18/547,242 in view of IK, Caddy and Heddle as applied to claims 23, 25-26, 30, 32-34, 36-37, 40 and 48 above, and further in view of Hamon.
The instant rejection is maintained from the previous Office Action and any newly recited portions are necessitated by claim amendment.
Claim 31 of the current application is drawn to the method of instant claim 23, wherein the gold-donor agent is chloro[diphenyl(3-sulfonatophenyl)phosphine]gold (I).
The claims of the reference application and the disclosures of IK, Caddy and Heddle as applied to claims 23, 25-26, 30, 32-34, 36-37, 40 and 48 are discussed above. The claims of the reference application do not recite chloro[diphenyl(3-sulfonatophenyl)phosphine]gold (I).
Hamon discusses an aqueous one-pot route to gold/quantum rod heterostructured nanoparticles functionalized with DNA [title], and describes that Au(I) organometallic chemistry provides a versatile approach to controlling surface grafting [abstract].
Regarding instant claim 31, Hamon discloses the use of the monovalent gold complex chloro-Au(I) known as chloro[diphenyl(3-sulfonatophenyl) phosphine]gold(I) for the conjugation of a heterostructured nanoparticle (HNP) with said gold complex [Scheme 1].
It would have been obvious to one of ordinary skill in the art before the effective filing date to modify the combined method of the reference application, Heddle, Caddy and IK by using chloro[diphenyl(3-sulfonatophenyl) phosphine]gold(I), since the simple substitution of one known element for another results in a predictable result. One of ordinary skill in the art would have recognized that both chloro[diphenyl(3-sulfonatophenyl) phosphine]gold(I) and chlorine(triphenylphosphine)gold(I) are halogen(triarylphosphine)gold(I) molecules used as gold-donor agents for the conjugation of gold-biomolecule complexes, and as such both are capable of being incorporated into methods as described by the reference application, Heddle, Caddy and IK. Thus it would have been obvious to one of ordinary skill in the art to replace chlorine(triphenylphosphine)gold(I) with chloro[diphenyl(3-sulfonatophenyl) phosphine]gold(I), as one of ordinary skill in the art would have been able to carry out such a substitution with a reasonable expectation of success because both Caddy and Hamon discuss the use of halogen(triarylphosphine)gold(I) molecules as gold-donor agents.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Response to Remarks: beginning on page 9 of Applicant’s response to non-statutory double patenting rejections; Applicant in summary contends the claims are not obvious for the reasons discussed in the response to rejections under 35 USC 103.
Applicant’s response is considered and found not convincing. As the response to double patenting rejections is the same as the response to 35 USC 103 rejections, the following section is repeated from above and any changes relate to specifics of co-pending Application 18/547242 (“reference application”).
The claim interpretations and teachings of the art are described in detail above. Briefly, the phrase “a Geobacillus stearothermophilus TRAP protein” does not structurally limit the TRAP protein used in the claimed method as the GS-TRAP protein is considered the source protein used in the method of generating the protein used in the claimed method which has a cysteine mutation. As the claims recite no reference sequence for comparison, the location of the cysteine mutation is indefinite, and therefore the TRAP protein used in the method is only required to have a cysteine amino acid, which can occur at any location in the protein. As Heddle teaches the use of a TRAP protein containing a cysteine from Bacillus stearothermophilus, which is recognized as the same organism Geobacillus stearothermophilus as confirmed by Applicant’s response [page 7, middle], the protein of Heddle is encompassed by the structural limitations of the TRAP protein used in the claimed method.
Conclusion
Status of the Application:
Claims 23, 25-26, 30-34, 36-37, 40-42, 44-45 and 48 are pending.
Claims 41-42 and 44-45 are withdrawn.
Claims 23, 25-26, 30-34, 36-37, 40 and 48 are rejected.
No claim is in condition for allowance.
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to JOSEPH SPANGLER whose telephone number is (571)270-0314. The examiner can normally be reached M-F 7:30 am - 4:30 pm.
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/JOSEPH R SPANGLER/
Examiner
Art Unit 1656
/David Steadman/Primary Examiner, Art Unit 1656
APPENDIX A
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Protein Data Bank search results for 4V4F9, carried out on 07/15/2026, https://www.rscb.org
Prosecution Insights