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 .
Status of the Claims
Claims 1-6, 8, and 10-14 are pending and examined herein.
Priority
The present application, filed 12/29/2023, is a 371 of PCT/EP2022/068207, filed 06/30/2022, which claims foreign priority of EP21185395.7, filed 07/13/2021. The benefit is acknowledged and the claims examined herein are treated as having an effective filing date of 07/13/2021.
Information Disclosure Statement
The Information Disclosure Statement(s) filed 12/29/2023 are acknowledged and have been considered.
Claim Rejections - 35 USC § 112
The following is a quotation of the first paragraph of 35 U.S.C. 112(a):
(a) IN GENERAL.—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.
The following is a quotation of the first paragraph of pre-AIA 35 U.S.C. 112:
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 of carrying out his invention.
Claims 1–6, 8, and 10-14 are rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, 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, or for applications subject to pre-AIA 35 U.S.C. 112, the inventor(s), at the time the application was filed, had possession of the claimed invention.
Independent claim 1 recites an affinity ligand comprising an amino acid sequence with at least 90% sequence identity to at least 56 amino acids of SEQ ID NO: 1, wherein the affinity ligand is stable under alkaline conditions of 0.1 M NaOH incubation for at least 6 h and has a binding affinity of less than 100 nM for a ubiquitin mutein. Here, claim 1 is not limited to SEQ ID NO: 1 itself, but instead encompasses a genus of affinity ligands defined by partial sequence identity and functional properties.
The specification describes the invention as directed to novel affinity ligands for affinity purification and detection of ubiquitin muteins (p. 1). The specification identifies specific affinity ligands, including SEQ ID NOs: 1–5 (p. 4), and discloses that SEQ ID NO: 1 is a 58-amino-acid artificial affinity ligand (p. 9). The specification further recites broad variant language, including affinity ligands having at least 90% sequence identity to at least 56 amino acids of SEQ ID NO: 1 (p. 2).
However, the disclosure does not reasonably convey possession of the full claimed genus. Although the specification recites affinity ligands having at least 90% sequence identity to at least 56 amino acids of SEQ ID NO: 1 and identifies certain amino acid positions that may be substituted or deleted, the specification does not adequately identify the boundaries of the claimed genus. In particular, the specification does not adequately explain whether the identity comparison is based on 56 consecutive amino acids, any 56 amino acids selected from SEQ ID NO: 1, a local alignment of 56 residues, or another comparison methodology. The specification also does not adequately explain why identity over 56 residues, rather than the full 58-residue SEQ ID NO: 1, is sufficient to define affinity ligands possessing the claimed ubiquitin-mutein binding and alkaline stability.
This omission is significant because claim 1 permits numerous sequence variants. For a 58-amino-acid sequence, at least 90% sequence identity permits multiple amino acid differences. When the comparison may be made over at least 56 amino acids, the claim further permits partial-sequence variants while still requiring the same functional profile. Although the specification identifies certain positions that may be substituted or deleted, it does not identify which combinations of those modifications preserve the claimed functional properties throughout the full breadth of the claimed genus.
The specification includes extensive position-by-position substitution language for SEQ ID NO: 1 and related affinity ligands. For example, the disclosure lists numerous possible amino acid alternatives at positions corresponding to SEQ ID NO: 1 (pp. 17-19). But those lists do not establish possession of the full claimed genus because they do not correlate the disclosed substitutions with retention of binding affinity of less than 100 nM for a ubiquitin mutein and stability under alkaline conditions of 0.1 M NaOH incubation for at least 6 h. The specification does not show which combinations of those substitutions work, which combinations fail, or which combinations were actually made and tested.
The working examples are also limited to particular disclosed species. The Examples describe library/selection work and testing of specific affinity ligands. The specification provides SPR binding data and alkaline stability data for selected affinity ligands, including specific ligands tested after NaOH incubation (pp. 29-35, including Tables 1-4). This evidence supports possession of the particular tested affinity ligands, but it does not reasonably convey possession of all affinity ligands within the claimed ≥90% sequence identity to at least 56 amino acids of SEQ ID NO: 1 genus that also satisfy both functional limitations. In particular, the disclosure does not provide a representative number of species commensurate in scope with the full claimed genus. The specification identifies only a limited number of selected affinity ligand sequences and tested embodiments, while claim 1 encompasses numerous non-exemplified variants defined by ≥90% sequence identity to at least 56 amino acids of SEQ ID NO: 1. The sequence listing similarly identifies specific affinity ligand sequences, including SEQ ID NOs: 1–5, fusion proteins, ubiquitin muteins, and multimeric constructs. However, disclosure of specific species and broad substitution lists does not, by itself, demonstrate possession of the entire genus of variants covered by claim 1, especially where the claimed genus is also defined by two functional requirements: high-affinity binding to a ubiquitin mutein and alkaline stability after NaOH exposure.
The specification also does not provide common structural features sufficient to identify the members of the claimed genus. The disclosure does not identify a conserved binding motif, critical contact residues, structural determinants of ubiquitin-mutein recognition, or structural determinants of NaOH stability for SEQ ID NO: 1 variants. Nor does the specification identify structural features common to the members of the claimed genus such that one of ordinary skill in the art would recognize which non-exemplified variants fall within the claimed genus while retaining the claimed binding affinity and alkaline stability. Without such common structural features or structure-function correlation, the specification does not reasonably convey that Applicant was in possession of the full scope of affinity ligands having at least 90% sequence identity to at least 56 amino acids of SEQ ID NO: 1 while retaining less than 100 nM binding affinity and 0.1 M NaOH stability.
Accordingly, the specification reasonably supports, at most, the specifically disclosed and tested affinity ligand embodiments described therein, but does not reasonably convey possession of the broader genus recited in claim 1 - namely all affinity ligands having at least 90% sequence identity to at least 56 amino acids of SEQ ID NO: 1 that bind a ubiquitin mutein with less than 100 nM affinity and remain stable after 0.1 M NaOH incubation for at least 6 h.
Claims 2–6 depend from claim 1 and further limit the ubiquitin mutein, monomer/dimer form, fusion protein, or non-immunoglobulin-binding protein features. Claim 8 recites an affinity separation matrix comprising the affinity ligand of claim 1. Claim 10 recites a method employing an affinity chromatography matrix comprising the affinity ligand of claim 1. Claim 11 recites a method for affinity purifying a ubiquitin mutein using the affinity ligand of claim 1, and claim 12 depends from claim 11. Claim 13 recites a method for analyzing for the presence of a ubiquitin mutein using an affinity ligand according to claim 1. Claim 14 recites a polynucleotide encoding the affinity ligand according to claim 1. Since claims 2–6, 8, and 10-14 all require the insufficiently described affinity ligand genus of claim 1, these claims are rejected for the same reasons.
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claims 4 and 12 are rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
Claim 4 recites “The affinity ligand according to claim 4…”; however, claim 4 improperly refers back to itself. This circular dependency renders the scope of claim 4 unclear because it is uncertain which preceding claim provides the basis for the further limitation recited in claim 4. For purposes of compact prosecution, claim 4 has been interpreted as depending from claim 1. Appropriate correction is required.
Claim 12 recites the limitation “wherein in step (d) the elution pH is 3.5 or higher” in line 2. There is insufficient antecedent basis for this limitation in the claim. Specifically, there is insufficient antecedent basis for “the elution pH” in claim 12. Claim 11 recites “eluting said ubiquitin mutein from said affinity chromatography matrix,” but does not previously introduce “an elution pH.” Accordingly, the scope of claim 12 is unclear. Appropriate correction is required.
Closest Prior Art
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The closest structural prior art identified is E. Fiedler et al. (US 2020/0172580 A1), which teaches artificial scaffold-library proteins and methods for selecting target-binding scaffold proteins. This reference is considered the closest structural prior art because it discloses related engineered scaffold proteins and binder-selection methodologies. However, the reference does not teach or suggest an affinity ligand comprising an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 1. As shown below, the closest disclosed scaffold sequence is only approximately 64.3% query match (approximately 74.1% best local similarity) to instant SEQ ID NO: 1.
Furthermore, the reference does not teach or suggest a protein/polypeptide capable of binding a ubiquitin mutein, nor does it disclose the claimed combination of binding affinity of less than 100 nM together with stability under alkaline conditions of 0.1 M NaOH incubation for at least six hours.
M. Fiedler et al. (Affilin (TM) Molecules: Novel Ligands for Bioseparation. Food and Bioproducts Processing. Vol. 84, No. C1, March 2006) represents the closest prior art directed to engineered affinity ligands for bioseparation. Fiedler teaches Affilin™ scaffold ligands generated by phage display, nanomolar binding affinity, immobilization onto chromatography matrices, affinity purification, and regeneration under harsh cleaning conditions. However, the disclosed Affilin ligand is directed to proNGF, rather than a ubiquitin mutein. Accordingly, Fiedler does not teach or suggest an affinity ligand that binds a ubiquitin mutein as recited in the instant claims.
Dias et al. (The Future of Protein Scaffolds as Affinity Reagents for Purification. Biotechnology and Bioengineering. Vol. 114, No. 3, March 2017) reviews engineered protein scaffolds as affinity purification reagents and teaches desirable properties of affinity ligands, including robustness, immobilization, and cleaning-in-place stability. However, Dias is directed generally to scaffold-based affinity reagents and does not disclose or suggest a protein affinity ligand capable of binding a ubiquitin mutein.
Randles et al. (Ubiquitin and Its Binding Domains. Frontiers in Bioscience. Vol. 17, No. 7, June 2012) teaches ubiquitin-binding domains and ubiquitin receptors that recognize native ubiquitin and ubiquitin chains. Although Walters demonstrates that ubiquitin-binding proteins were known, the reference is directed to native ubiquitin and does not teach or suggest proteins capable of binding ubiquitin muteins as required by the instant claims.
Bosse-Doenecke et al. (US 2019/0292266 A1) teaches ubiquitin muteins (AFFILIN® proteins) as engineered binding proteins directed to unrelated target antigens. Thus, the ubiquitin mutein functions as the binding protein, rather than as the target of another affinity ligand. The reference therefore teaches the reverse binding relationship and does not teach or suggest a protein affinity ligand capable of binding a ubiquitin mutein as required by claim 1.
Accordingly, although the prior art demonstrates that engineered scaffold ligands, affinity purification ligands, ubiquitin-binding proteins, and ubiquitin muteins were individually known, no reference, either alone or in combination, teaches or reasonably suggests the claimed affinity ligand comprising an amino acid sequence with at least 90% sequence identity to at least 56 amino acids of SEQ ID NO: 1 that binds a ubiquitin mutein, with a binding affinity of less than 100 nM and is stable under alkaline conditions of 0.1 M NaOH incubation for at least six hours. Therefore, independent claim 1, as well as claims depending therefrom, are considered free of the prior art presently of record.
Conclusion
No claims are allowable.
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/E.O./Examiner, Art Unit 1677
/BAO-THUY L NGUYEN/Supervisory Patent Examiner, Art Unit 1677 July 10, 2026
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