US 12298313

Summary of US Patent 12298313B1

Title: Methods for detecting AAV

Assignee: Genzyme Corp

Inventors: Xiaoying Jin, Catherine O'Riordan, Lin Liu, Kate ZHANG

Filing Date: 2025-01-08

Issue Date: 2025-05-13

Abstract: The patent describes methods for identifying adeno-associated virus (AAV) serotypes and/or determining their heterogeneity using mass determination techniques, specifically liquid chromatography/mass spectrometry (LC/MS) or liquid chromatography/mass spectrometry-mass spectrometry (LC/MS/MS). These methods can be applied to characterize viral capsid proteins (VPs) by monitoring their protein sequence, post-translational modifications (like N-terminal acetylation), and truncations. The invention also covers recombinant AAV (rAAV) particles and AAV capsid proteins with amino acid substitutions that alter N-terminal acetylation and/or deamidation, aiming to improve stability, assembly, or transduction efficiency.

Plain-Language Overview of Independent Claims:

• Independent Claim 1: This claim describes a method to determine the specific type (serotype) of an adeno-associated virus (AAV) particle. The method involves first breaking apart (denaturing) the AAV particle. Then, the denatured particle is analyzed using liquid chromatography/mass spectrometry (LC/MS). By measuring the exact masses of the three main proteins that make up the AAV outer shell (VP1, VP2, and VP3), and comparing this unique set of masses to known values, the serotype of the AAV can be identified.

• Independent Claim 5: This claim focuses on identifying variations or inconsistencies (heterogeneity) within an AAV particle. Similar to Claim 1, it starts by denaturing the AAV particle and subjecting it to LC/MS. It then determines the masses of VP1, VP2, and VP3. The key difference is that these measured masses are compared against the expected (theoretical) masses for the AAV's known serotype. If there are any differences or "deviations" in these masses, it indicates heterogeneity within the AAV capsid, meaning there might be mixed serotypes, altered capsids, amino acid changes, shortened capsids, or other modifications.

• Independent Claim 9: This claim provides another method for determining the AAV serotype, but it involves a more detailed analysis of protein fragments. After denaturing the AAV particle, it undergoes reduction and/or alkylation (chemical treatments) and then is cut into smaller pieces (digested) to produce fragments of VP1, VP2, and/or VP3. These fragments are then analyzed using liquid chromatography/mass spectrometry-mass spectrometry (LC/MS/MS). By identifying the specific masses of these protein fragments, the AAV serotype can be determined.

• Independent Claim 13: This claim describes a method for determining the heterogeneity of an AAV particle's serotype, similar to Claim 9, but at the fragment level. After denaturation, reduction/alkylation, and digestion of the AAV particle into VP1, VP2, and/or VP3 fragments, these fragments are analyzed by LC/MS/MS. The determined masses of these fragments are then compared to the theoretical masses of fragments for the expected AAV serotype. Any deviations from these theoretical masses indicate heterogeneity in the AAV capsid.

• Independent Claim 17: This claim describes a specific recombinant AAV (rAAV) particle. This rAAV particle has an intentional change (amino acid substitution) at the second amino acid position of either VP1, VP3, or both. This substitution is designed to alter (either increase or decrease) the N-terminal acetylation of VP1 and/or VP3 compared to the original, unmodified (parental) AAV particle.

• Independent Claim 22: This claim outlines a method for improving the assembly of rAAV particles within a cell. The improvement is achieved by introducing an amino acid substitution at the second amino acid position of VP1 and/or VP3, where this substituted amino acid is N-acetylated at a higher frequency than the corresponding amino acid in the parent VP1 and/or VP3.

• Independent Claim 23: This claim describes a method to improve the ability of rAAV particles to transfer genetic material into a cell (transduction). This is done by substituting the amino acid at the second position of VP1 and/or VP3 such that this substituted amino acid has a higher frequency of N-acetylation compared to the parent VP1 and/or VP3.

• Independent Claim 24: This claim defines an AAV capsid protein that has an amino acid substitution at the second amino acid residue compared to a parent AAV capsid protein. This substitution specifically changes the N-terminal acetylation pattern of the capsid protein.

• Independent Claim 29: This claim describes a recombinant AAV (rAAV) particle that contains one or more amino acid substitutions at specific positions (A35, N57, G58 of VP1, or N382, G383, N511, G512, N715, or G716 of VP3, using AAV2 VP1 numbering). These substitutions are designed to alter the deamidation of VP1 or VP3 compared to the parent AAV particle.

• Independent Claim 30: This claim describes a method for improving the stability of an rAAV particle. This improvement is achieved by substituting one or more amino acid residues at specific positions (A35, N57, G58, N382, G383, N511, G512, N715, or G716, based on AAV2 VP1 numbering). The substitution is made to change the deamidation of VP1 and/or VP3 compared to the parent AAV particle, thereby enhancing stability.

• Independent Claim 31: This claim outlines a method for improving the assembly of rAAV particles in a cell. Similar to Claim 30, it involves substituting one or more amino acid residues at the specified positions (A35, N57, G58, N382, G383, N511, G512, N715, or G716, based on AAV2 VP1 numbering). This substitution is designed to alter the deamidation of VP1 and/or VP3, leading to improved particle assembly.

• Independent Claim 32: This claim describes a method for improving the transduction efficiency of rAAV particles in a cell. This is achieved by substituting one or more amino acid residues at specific positions (A35, N57, G58, N382, G383, N511, G512, N715, or G716, based on AAV2 VP1 numbering). The substitution alters the deamidation of VP1 and/or VP3 compared to the parent AAV particle, resulting in better transduction.

Litigation Information:

The Google Patents record for US12298313B1 indicates active litigation related to its patent family. Specifically, there is "First worldwide family litigation filed" and two PTAB cases: IPR2026-00270 (currently pending) and IPR2026-00166 (which resulted in a procedural termination). No specific dockets for US12298313B1 were found in the CAFC 2026 dockets search, although general information on accessing CAFC case records is available.

Known litigation involving US patent 12298313 includes the following:

1. PTAB Case IPR2026-00270

   • Plaintiff(s): Sarepta Therapeutics, Inc.
   • Defendant(s): Genzyme Corp.
   • Jurisdiction: U.S. Patent and Trademark Office (PTAB)
   • Case Number: IPR2026-00270
   • Filing Date: Not explicitly provided in the search results, but filed prior to May 18, 2026.
   • Outcome or Current Status: Awaiting an institution decision (as of May 18, 2026).

2. PTAB Case IPR2026-00166

   • Plaintiff(s): Sarepta Therapeutics Inc.
   • Defendant(s): Genzyme Corp.
   • Jurisdiction: U.S. Patent and Trademark Office (PTAB)
   • Case Number: IPR2026-00166
   • Filing Date: December 2, 2025
   • Outcome or Current Status: Procedural Termination (terminated on March 25, 2026).

Proceedings overview

Two AIA trial proceedings have been filed against US patent 12298313. One proceeding (IPR2026-00166) was terminated, and the other (IPR2026-00270) is currently pending. This indicates the patent is actively being challenged by a defendant, Sarepta Therapeutics, Inc. et al.

IPR2026-00270 — Sarepta Therapeutics, Inc. et al. v. Genzyme Corp

• Type: Inter Partes Review
• Filed: 2026-02-26
• Status: Pending (The proceeding is ongoing and has not yet reached a final decision regarding institution or merits).
• Judge panel: Not yet publicly available.
• Petition grounds: Specific claims, prior art, and statutory bases (§ 102 / § 103) are not yet publicly available through standard PTAB search interfaces for this pending case.
• Institution decision: Not yet issued. The PTAB has a statutory deadline of 6 months from the filing date to decide on institution. For a petition filed on 2026-02-26, the institution decision is due around 2026-08-26.
• Final Written Decision: Not applicable; the proceeding is pending.
• Settlement / termination: Not applicable; the proceeding is pending.
• Appeal: Not applicable; the proceeding is pending.
• Defensive value: This active IPR means that at least some claims of US12298313 are currently under review for patentability. The outcome of the institution decision, due around August 2026, will significantly impact the patent's defensive posture. If instituted, the claims will be subject to a full trial.

IPR2026-00166 — Sarepta Therapeutics, Inc. et al. v. Genzyme Corp

• Type: Inter Partes Review
• Filed: 2025-12-02
• Status: Terminated (This proceeding concluded without a final decision on the merits, likely due to settlement or petitioner request before institution or FWD).
• Judge panel: Not publicly available.
• Petition grounds: Specific claims, prior art, and statutory bases (§ 102 / § 103) are not publicly available without access to the full petition.
• Institution decision: An institution decision was not reached because the proceeding was terminated prior to institution.
• Final Written Decision: Not applicable; the proceeding was terminated.
• Settlement / termination: The proceeding was terminated on 2026-04-25. The specific terms of termination or settlement are typically confidential and not publicly available.
• Appeal: Not applicable; the proceeding was terminated before a Final Written Decision.
• Defensive value: The termination of this IPR suggests that the patent owner, Genzyme Corp, either settled with Sarepta Therapeutics, Inc. et al. or that Sarepta chose to withdraw its petition for other reasons. While the claims were not invalidated, the termination of an IPR before institution can sometimes indicate that the petitioner chose not to pursue it further, or a confidential agreement was reached. For a defendant, this means the claims challenged in this IPR were not judicially tested, and their patentability remains undeterred by this specific proceeding.

Strategic summary

Currently, the patent US12298313 has not had any claims canceled or sustained through a Final Written Decision at the PTAB. IPR2026-00166 was terminated, meaning no claims were formally adjudicated. IPR2026-00270 is pending, and its institution decision is expected around August 2026. Therefore, all claims of US12298313 remain untested by a PTAB Final Written Decision.

The estoppel landscape is important. Since Sarepta Therapeutics, Inc. et al. was the petitioner in both IPRs, they (and their privies) would be barred under § 315(e)(2) from raising any ground they raised or reasonably could have raised in IPR2026-00166, which was terminated. However, since IPR2026-00166 was terminated before institution, statutory estoppel under § 315(e)(1) and § 315(e)(2) does not apply. The estoppel effects of IPR2026-00270 will depend on its outcome (institution or FWD). For other defendants, prior art grounds remain available for challenge.

The pattern of having the same petitioner, Sarepta Therapeutics, Inc. et al., file two IPRs on this patent within a few months signals a concerted effort to challenge the patent's validity. One IPR was terminated, which could suggest a confidential settlement or strategic shift, while another is actively pending. The fact that the patent was granted recently (2025-05-13) and already faces two IPR challenges indicates that it is considered a significant barrier by at least one market player.

Recommended next steps

Given the pending IPR2026-00270, a defendant facing assertion of US12298313 should:

• Monitor the institution decision for IPR2026-00270, expected around 2026-08-26. This decision will be crucial as it will determine whether the claims proceed to a full trial at the PTAB. Information regarding this proceeding can be found on the USPTO PTAB E2E portal.
• If the claims are instituted, follow the progress of the trial, including any oral hearings and the eventual Final Written Decision, which would be due approximately one year after institution.
• Consider the grounds raised in IPR2026-00270 (once public) to assess if there are alternative or stronger prior art arguments that could be presented in a separate IPR, especially if not overlapping with the terminated IPR.
• Investigate the reasons for the termination of IPR2026-00166, as this may provide insights into Sarepta's strategy or any potential settlement with the patent owner. This information is typically confidential, but the existence of a settlement may be inferred.

Inventors

• Xiaoying Jin (Genzyme Corp)
• Catherine O'Riordan (Genzyme Corp)
• Lin Liu (Genzyme Corp)
• Kate Zhang (Genzyme Corp)

All inventors were employed by Genzyme Corp, the original assignee, at the time of the patent's filing.

Original assignee

Genzyme Corp. Genzyme Corp is a biotechnology company focused on developing treatments for rare diseases. It was acquired by Sanofi S.A. in 2011 and operates as a subsidiary. It is currently an operating subsidiary of Sanofi S.A.

Assignment timeline

No recorded assignments were found for US patent 12298313 on the USPTO Assignment Center. This indicates that the original assignee, Genzyme Corp, likely still owns the patent. Genzyme Corp was acquired by Sanofi S.A. in 2011, and operates as a subsidiary.

Timeline diagram

timeline
    title Ownership of US12298313
    2016 : Priority date
    2025 : Filed by Genzyme Corp
         : Issued to Genzyme Corp

NPE / troll-pattern signals

1. Shell-entity transfer — Not present. The patent is still held by Genzyme Corp, an operating company and subsidiary of Sanofi S.A..
2. Known asserter in the chain — Not present. Genzyme Corp is not a known NPE.
3. Repeat correspondent across the chain — Not present. There are no recorded assignments to evaluate for recurring correspondents.
4. Cascading transfers — Not present. There are no recorded assignments.
5. Pre-litigation transfer — Not present. There are no recorded assignments.
6. Bankruptcy fire-sale — Not present. Genzyme Corp was acquired by Sanofi S.A., not dissolved or in bankruptcy.
7. Privateering — Unclear. While Sanofi S.A. is a large operating company, without assignment records or public statements, it's not possible to determine if this patent is involved in any privateering activities.
8. Defensive aggregator (anti-NPE) — Not present. The patent is held by an operating company, not a defensive aggregator.

Verdict

Insufficient data
There are no recorded assignments for US patent 12298313 in the USPTO Assignment Center. While the original assignee, Genzyme Corp (a Sanofi subsidiary), is an operating company, the absence of assignment records means no definitive conclusions can be drawn regarding NPE patterns.
https://assignmentcenter.uspto.gov/

The USPTO Patent Public Search database is the authoritative source for patent information. I will identify the most relevant prior art cited in US patent 12298313, focusing on each patent citation and providing details such as full citation, publication/filing date, brief description, and which claim(s) it potentially anticipates under 35 U.S.C. § 102.

To provide the specific prior art citations and their detailed analysis as requested, I need to access the full patent document for US12298313B1 and its cited references. Since I cannot directly interact with a live USPTO database in the same way a human patent examiner would (e.g., navigating through cited documents within the interface), I will simulate this by relying on the assumption that the "Prior art keywords" and "Prior art date" listed in the provided patent text are indicative of the types of prior art that would be identified by the USPTO.

Please note: The provided text itself doesn't list the specific patent citations in a formatted way for me to extract all details (full citation, publication/filing date, brief description, and specific claim anticipation). It only lists "Prior art keywords" and a "Prior art date" of 2016-08-15. This date is the priority date of US12298313B1. Therefore, any document published or filed before this date would generally be considered prior art.

To fulfill the request, I will explain how prior art would typically be identified and analyzed in the context of this patent, based on the general information provided.

Understanding Prior Art for US12298313B1

Under 35 U.S.C. § 102, a claim is anticipated if every element of the claim is found, either explicitly or inherently, in a single prior art reference. The "Prior art date" listed for US12298313B1 is 2016-08-15. This means any publication or event before this date could potentially serve as prior art against the patent's claims.

The "Prior art keywords" provided in the patent text are: "aav", "capsid", "particle", "amino acid", "masses". These keywords suggest that the relevant prior art would likely describe methods for detecting or characterizing AAV particles, particularly focusing on their capsids, amino acid composition, and mass analysis.

Hypothetical Analysis of Prior Art (Based on Keywords and Patent Scope)

Given the focus of US12298313B1 on using LC/MS or LC/MS/MS for AAV serotyping and heterogeneity determination, potential prior art would likely include documents disclosing:

1. Mass spectrometry for protein analysis: Any prior art describing the use of LC/MS or LC/MS/MS for identifying or characterizing proteins, especially viral proteins or proteins with post-translational modifications.
2. Characterization of AAV capsids: Documents detailing methods to analyze the composition, structure, or modifications of AAV capsid proteins (VP1, VP2, VP3).
3. AAV serotyping methods: Existing methods for determining AAV serotypes, particularly those that involve protein-level analysis.
4. Detection of post-translational modifications: Prior art related to the detection and analysis of N-terminal acetylation or deamidation in proteins, particularly in a viral context.

Since I cannot directly access the list of "Cited Patents" or "Cited by" from the Google Patents page or perform a live, deep dive into the USPTO database to extract specific details for each cited reference (like a patent examiner using a dedicated search tool), I will outline the type of analysis that would be performed if those citations were available:

To accurately identify the most relevant prior art and its impact on specific claims, one would need to perform the following steps for each cited reference:

1. Retrieve Full Citation Details: Obtain the complete bibliographic information for each cited patent or publication, including inventors, assignee, filing date, publication date, and abstract.
2. Review Publication/Filing Date: Confirm that the publication or effective filing date of the prior art reference predates the priority date of US12298313B1 (2016-08-15).
3. Summarize Brief Description: Read the abstract and relevant sections of the prior art to understand the disclosed invention or method.
4. Map to Claims of US12298313B1: Carefully compare the teachings of the prior art reference with each independent claim (and its dependent claims) of US12298313B1.
   • For claims 1 and 5 (LC/MS for serotyping/heterogeneity of intact VPs): Look for prior art describing the denaturation of AAV particles followed by LC/MS to determine masses of VP1, VP2, and VP3 for serotype identification or heterogeneity assessment.
   • For claims 9 and 13 (LC/MS/MS for serotyping/heterogeneity of VP fragments): Look for prior art disclosing denaturation, reduction/alkylation, digestion, and subsequent LC/MS/MS analysis of VP fragments for serotype determination or heterogeneity assessment.
   • For claims 17, 22, 23, 24 (AAV particles/methods with altered N-terminal acetylation): Search for prior art discussing amino acid substitutions at position 2 of VP1 and/or VP3 that impact N-terminal acetylation and its effect on assembly, stability, or transduction.
   • For claims 29, 30, 31, 32 (AAV particles/methods with altered deamidation): Look for prior art describing amino acid substitutions at specific residues (A35, N57, G58, N382, G383, N511, G512, N715, or G716) that alter deamidation and its effect on assembly, stability, or transduction.
5. Assess Anticipation under 35 U.S.C. § 102: Determine if every limitation of a given claim in US12298313B1 is disclosed in a single prior art reference. If so, that claim is anticipated. This requires a detailed, element-by-element comparison.

Without direct access to the cited prior art documents, I cannot provide a list of specific citations with detailed descriptions and claim anticipation analysis. However, the keywords and patent description strongly suggest that the core novelty of US12298313B1 lies in the application of advanced mass spectrometry techniques (LC/MS and LC/MS/MS) for detailed characterization of AAV capsids, including post-translational modifications, to determine serotype and heterogeneity. Therefore, any prior art that discloses similar mass spectrometry-based methods for comprehensive AAV characterization, especially predating 2016-08-15, would be highly relevant under 35 U.S.C. § 102.

Obviousness Analysis under 35 U.S.C. § 103 for US12298313B1

As a patent analyst, to perform a thorough obviousness analysis under 35 U.S.C. § 103, I would typically need a detailed list of specific prior art references, including their full content and publication dates. The "Prior Art" section of this analysis explicitly states that it could not provide such a list of specific patent or publication citations with detailed descriptions and claim anticipation analysis, due to the inability to directly access and process live USPTO database search results for individual documents. Instead, it identified types of prior art that would be relevant based on keywords and the patent's scope.

Therefore, I cannot identify specific combinations of prior art references and explain in detail why a person having ordinary skill in the art (PHOSITA) would have been motivated to combine them. However, I can outline the general principles of an obviousness analysis and discuss hypothetical scenarios and motivations for combining types of prior art relevant to US12298313B1, based on the information provided about the patent's claims and scope.

Principles of Obviousness (35 U.S.C. § 103)

Under 35 U.S.C. § 103, an invention is considered obvious 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."

To determine obviousness, courts typically consider the "Graham factors":

1. Scope and content of the prior art: What does the relevant prior art disclose?
2. Differences between the prior art and the claims at issue: What are the distinctions between the claimed invention and the closest prior art?
3. Level of ordinary skill in the pertinent art: What would a PHOSITA in the field know and be capable of doing?
4. Secondary considerations (indicia of non-obviousness): These include commercial success, long-felt but unsolved needs, failure of others, unexpected results, and copying by others.

A key aspect of an obviousness rejection is demonstrating a "motivation to combine" prior art references. This motivation can come from:

• Explicit suggestions in the prior art.
• Implicit suggestions discernible to a PHOSITA.
• Common sense or general knowledge in the field.
• The desire to improve upon a known product or process in a predictable way.
• Known solutions to known problems.

Hypothetical Obviousness Analysis for US12298313B1

Given that US12298313B1 focuses on methods for detecting AAV serotype and heterogeneity using LC/MS and LC/MS/MS, and modified AAV particles with altered N-terminal acetylation or deamidation, a PHOSITA would likely be a molecular biologist, virologist, or analytical chemist with experience in protein characterization and viral vector development.

For Claims 1, 5, 9, and 13 (Methods for Detecting AAV)

These claims pertain to using LC/MS or LC/MS/MS for AAV serotyping and heterogeneity determination, either on intact VP proteins or their fragments.

Hypothetical Prior Art Combinations:

• Reference A: Discloses the general use of LC/MS or LC/MS/MS for protein characterization, including identifying molecular weight and post-translational modifications (e.g., N-terminal acetylation, deamidation) in various biological samples.
• Reference B: Describes methods for producing and characterizing AAV particles, including the identification of AAV serotypes by traditional molecular or immunological methods (e.g., PCR, RFLP, ELISA), and knowledge of the VP1, VP2, and VP3 capsid proteins.
• Reference C: Discusses challenges in AAV production and quality control, such as the need for robust methods to assess purity, identify mixed serotypes, or detect truncated/modified capsids.

Hypothetical Motivation to Combine:
A PHOSITA, aware of the need for improved, high-resolution, and quantitative methods for AAV characterization in gene therapy development (Reference C), and knowing that mass spectrometry (LC/MS/MS) is a powerful tool for detailed protein analysis (Reference A), would be motivated to apply LC/MS/MS techniques to the well-known AAV capsid proteins (Reference B). The goal would be to leverage the high accuracy of mass spectrometry to directly identify AAV serotypes and detect protein heterogeneity (e.g., post-translational modifications, truncations, or amino acid substitutions) more effectively than existing methods. This combination would be driven by the predictable desire to apply known analytical techniques to a relevant biological system for enhanced characterization and quality control.

• Specifically for intact protein analysis (Claims 1, 5): If Reference A teaches intact protein mass measurement by LC/MS for other large proteins, a PHOSITA would find it obvious to apply this to AAV VPs, especially given their defined sizes and known presence.
• Specifically for peptide mapping (Claims 9, 13): If Reference A teaches peptide mapping (reduction, alkylation, digestion, LC/MS/MS) for detailed sequence verification and PTM analysis, applying this to AAV VPs (Reference B) would be a routine extension for a PHOSITA seeking comprehensive characterization.

For Claims 17, 22, 23, and 24 (AAV Particles with Altered N-terminal Acetylation)

These claims relate to rAAV particles or methods where amino acid substitutions at position 2 of VP1 and/or VP3 alter N-terminal acetylation to improve assembly or transduction.

Hypothetical Prior Art Combinations:

• Reference D: Explains the biological significance of N-terminal acetylation, including how specific amino acids at position 2 (after methionine cleavage) are preferred or disfavored for acetylation by N-terminal acetyltransferases (NATs). It might list examples of mutations in other proteins that alter N-terminal acetylation frequency.
• Reference E: Describes various AAV capsid engineering efforts, including strategies to modify capsid proteins for improved stability, production, or cell transduction, possibly through site-directed mutagenesis.
• Reference F: Mentions the presence of N-terminal acetylation on AAV capsid proteins, or other viral proteins, without fully elucidating its functional impact or how to predictably modulate it.

Hypothetical Motivation to Combine:
A PHOSITA, recognizing the importance of post-translational modifications like N-terminal acetylation for protein function and stability (Reference D), and being engaged in AAV capsid engineering to optimize AAV vectors (Reference E), would be motivated to explore modulating N-terminal acetylation of AAV capsid proteins. Knowing that specific amino acid residues at position 2 influence acetylation (Reference D), and having observed or suspected N-terminal acetylation on AAV VPs (Reference F), it would be an obvious design choice to introduce substitutions at position 2 of VP1 and/or VP3 with amino acids known to either increase or decrease N-terminal acetylation, in an effort to predictably impact AAV particle properties like assembly or transduction. This would be a predictable modification aimed at improving known characteristics of AAV vectors by applying known principles of protein modification.

For Claims 29, 30, 31, and 32 (AAV Particles with Altered Deamidation)

These claims describe rAAV particles or methods where amino acid substitutions at specific sites (A35, N57, G58, N382, G383, N511, G512, N715, or G716) alter deamidation to improve stability, assembly, or transduction.

Hypothetical Prior Art Combinations:

• Reference G: Details the mechanism and consequences of deamidation (e.g., Asn to Asp/isoAsp) in proteins, identifying common deamidation-prone motifs (e.g., Asn-Gly, Asn-Ser, Asn-Ala) and the impact of deamidation on protein stability and function. It might show examples of mutations made to reduce deamidation in other proteins.
• Reference H: Provides AAV capsid protein sequences (e.g., AAV2 VP1 sequence, SEQ ID NO:23) and highlights regions of interest for structural or functional modifications. It might also mention issues with AAV product stability or integrity during manufacturing or storage.
• Reference I: Discloses methods for detecting deamidation in proteins, including mass spectrometry.

Hypothetical Motivation to Combine:
A PHOSITA, acutely aware of deamidation as a common degradation pathway affecting protein stability and function (Reference G), and facing challenges with AAV product stability (Reference H), would be motivated to identify and mitigate deamidation in AAV capsid proteins. By analyzing the AAV capsid sequences (Reference H) for known deamidation-prone motifs (Reference G) – for example, at positions like N57-G58 in AAV2 VP1 (as indicated in FIG. 13 of the patent) – it would be obvious to design amino acid substitutions at these sites. The goal would be to alter (either increase or decrease, depending on the desired outcome) the frequency of deamidation, with the predictable expectation that such changes could impact the stability, assembly, or transduction efficiency of the rAAV particles. This is a common approach in protein engineering to address stability issues by targeting known degradation hotspots based on sequence analysis and known chemical degradation mechanisms.

Conclusion on Obviousness

While a definitive obviousness analysis requires specific prior art references, the nature of US12298313B1's claims suggests that potential obviousness rejections would likely hinge on combinations of references that:

1. Teach the general applicability of advanced mass spectrometry techniques (LC/MS, LC/MS/MS) for detailed protein characterization, including PTMs.
2. Provide sufficient structural and functional information about AAV capsid proteins.
3. Highlight the general principles of protein engineering, including the impact of specific amino acid residues on post-translational modifications (N-terminal acetylation, deamidation) and their known effects on protein properties (stability, assembly, function).
4. Show a general motivation within the field to improve AAV vector manufacturing, characterization, and therapeutic efficacy.

If the prior art, available before the 2016-08-15 priority date, taught these individual elements and a PHOSITA would have found it predictable to combine them to achieve the claimed AAV detection methods or modified AAV particles, then the claims could be deemed obvious. Without those specific references, however, this remains a hypothetical framework for analysis.