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 .
Continued Examination Under 37 CFR 1.114
A request for continued examination under 37 CFR 1.114, including the fee set forth in 37 CFR 1.17(e), was filed in this application after final rejection. Since this application is eligible for continued examination under 37 CFR 1.114, and the fee set forth in 37 CFR 1.17(e) has been timely paid, the finality of the previous Office action has been withdrawn pursuant to 37 CFR 1.114. Applicant's submission filed on Sep. 11, 2025 has been entered.
DETAILED ACTION
Acknowledgement is hereby made of receipt and entry of the communication filed on Sept. 11, 2025. Claims 29-36 and 38-46 are pending and currently examined.
Claim Rejections - 35 USC § 103
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
(New Rejection-necessitated by amendment) Claims 29-36 and 38-46 are rejected under 35 U.S.C. 103 as being unpatentable over Jin et al. (US 11698377 B2, patented on Jul. 11, 2023) in view of Xiao et al. (J Virol. 1999 May;73(5):3994-4003) as evidenced by Krokhin et al. (Anal Chem. 2006 Sep 15;78(18):6645-50), Martinez-Navio et al. (Mol Ther. 2016 Feb;24(1):76-86. ), and Cancer Research UK (https://www.cancerresearchuk.org/about-cancer/treatment/targeted-cancer-drugs/types/anti-angiogenics#:~:text=Types%20of%20anti%20angiogenesis%20treatment&text=Some%20drugs%20block%20vascular%20endothelial,ramucirumab).
The amended base claim 29 is directed to a composition comprising a mixed population of recombinant adeno- associated virus (rAAV), each of said rAAV comprising:
(a) an AAV1 capsid comprising a heterogeneous population of AAV1 vp1 proteins, a heterogeneous population of AAV 1 vp2 proteins, and a heterogeneous population of AAV1 vp3 proteins which contain amino acid modifications comprising 50% to 100% asparagines (N) deamidated at each of positions: N57, N383, N512, and N718, based on the numbering of SEQ ID NO: 1, as determined using mass spectrometry, wherein vp1 comprises positions N57, N383, N512 and N718, vp2 comprises positions N383, N512 and N718 and vp3 comprises positions N383, N512 and N718,
wherein the deamidated asparagines are deamidated to an aspartic acid, an isoaspartic acid, an interconverting aspartic acid/isoaspartic acid pair, or combinations thereof; and
(b) a vector genome in the AAV capsid, wherein the vector genome comprises a nucleic acid molecule comprising AAV inverted terminal repeat sequences and a non-AAV nucleic acid sequence encoding a product operably linked to sequences which direct expression of the product in a target cell.
Jin et al. teaches a method for serotyping and/or determining the heterogeneity of a viral particle ( e.g., an adeno-associated virus (AAV) particle) using mass determination, e.g., by employing liquid chromatography/mass spectrometry (LC/MS) or liquid chromatography/mass spectrometry-mass spectrometry (LC/MS/MS) (See column 1, lines 21-30). Jin et al. teaches that the AAV heterogeneity in the composition is due to post-translational modifications such as deamidation. Substitution mutations may be introduced into vp1 and the amino acid substitution results in less deamidation of the AAV capsid (See e.g., column 8, lines 27-42). Jin et al. indicates that the amino acids considered for high rate of deamidation are A35, N57, G58, N382, G383, N511, G512, N715, or G716 of VP1 (numbering in AAV2) and the one or more amino acid substitutions such as N57K or a N57Q substitution can result in a lower frequency of deamidation as compared to deamidation of VP1 and/or VP3 of the parent AAV particle including AAV1 particles (See e.g., column 10, lines 19-60), which can improve the stability, assembly and/or transduction efficiency (See e.g., column 11, lines 14-65). The percentage of the deamidation in AAV1 and AAV2 particles are also taught in Jin’s invention. Fig.6A & 6B show the results of LC/MS/MS analysis comparing the percentage of deamidation in AAV1 and AAV2 particles produced by the TTx and PCL methods. The T9 peptide YLGPFNGLDK (SEQ ID NO: 9) was used to monitor potential deamidation site N57 in both AAV1 and AAV2. FIGS. 8A & 8B show the results of LC/MS/MS analysis comparing the percentage of deamidation in AAV1 and AAV2 particles produced by the TTx and PCL methods. The T67 peptides SANVDFTVDNNGLYTEPR (SEQ ID NO: 13) and SVNVDFTVDTNGVYSEPR (SEQ ID NO: 14) were used to monitor potential deamidation site N715 in AAV1 and AAV2, respectively ((See e.g., Column 13). Here the description above indicates the NG pairs as the deamidation sites. Jin et al. also discloses that the "deamidation" refers to a chemical reaction in which an amide functional group in the side chain of asparagine or glutamine is removed or converted to another functional group. For example, asparagine may be converted to aspartic acid or isoaspartic acid (See column 27, lines 54-60). Jin et al. also discloses that the AAV particle is deamidated to a higher extent compared to a parental AAV particle. In some embodiments, the AAV particle is more than about any of 45 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100% more deamidated compared to a parent AAV particle. In some embodiments, the AAV particle is deamidated between about any of 5%-10%, 10%-15%, 15%-20%, 20%-25%, 25%-30%, 30%-35%, 35%-40%, 40%-55%, 45%-50%, 50%-55%, 55%-60%, 60%-65%, 65%-70%, 70%-75%, 75%-80%, 80%-85%, 85%-90%, 90%-95%, 95%-100%, 5-25%, 25-50%, 50-75%, 75%-100%, 5-50% or 50%-100% more than a parent AAV particle (See e.g., column 28, lines 42-54), which teaches a 50% to 100% deamidation amino acid is present in the heterogeneous population of AAV particles.heterogeneous, Jin et al. discloses that the "Heterogeneity" when used in reference to an AAV capsid refers to an AAV capsid characterized by one or more capsid polypeptides observed to deviate from a reference mass of a VP1, VP2, and/or VP3 polypeptide, or fragment thereof. A reference mass may include, without limitation, a theoretical, predicted, or expected mass of a VP1, VP2, and/or VP3 polypeptide, e.g., of a known AAV serotype. For example, an AAV capsid may be said to display heterogeneity if it demonstrates one or more of the following properties (without limitation): a mixed serotype, a variant capsid, a capsid amino acid substitution, a truncated capsid, or a modified capsid (See column 19, lines 41-60), where the substitution comprises a substitution with Asp at N57 of VP1, N382 of VP3, N511 ofVP3, or N715 of VP3, and results in a higher frequency of deamidation as compared to deamidation of VP1 and/or VP3 of the parent AAV particle (See e.g., column 10, lines 32-36). As for the vector genome in the AAV capsid, Jin et al. teaches that the viral particle comprises an AAV1 ITR, an AAV2 ITR, an AAV3 ITR, an AAV4 ITR, an AAV5 ITR, an AAV6 ITR… an AAV11 ITR, or an AAV12 ITR. In some embodiments, the AAV particle comprises an AAV vector encoding a heterologous transgene (See Column 6, lines 57-64). Jin et al. also discloses that a "recombinant viral vector" refers to a recombinant polynucleotide vector comprising one or more heterologous sequences (i.e., nucleic acid sequence not of viral origin). In the case of recombinant AAV vectors, the recombinant nucleic acid is flanked by at least one, e.g., two, inverted terminal repeat sequences (ITRs) (See Column 16, lines 36-41). Jin et al. also discloses that in some embodiments, the heterologous nucleic acid is operably linked to a promoter [See column 33, lines 26-50). Jin et al. also teaches that different AAV serotypes are used to optimize transduction of particular target cells or to target specific cell types within a particular target tissue (See column 43, lines 11-13). Jin et al. also teaches that in some embodiments, the heterogeneity comprises one or more of mixed serotypes, variant capsids, capsid amino acid substitutions, truncated capsids, or modified capsids (See e.g., column 4, lines 8-11).
Accordingly, Jin et al. teaches a composition comprising a mixed population of recombinant adeno- associated virus (rAAV) comprising AAV1 VP1, VP2 and VP3 that the heterogeneous population is caused by the high ratio of the deamidation, where can be determined by mass spectrometry. At the same time, Jin et al. also teaches a vector genome in the AAV capsid comprising ITRs and a transgene encoded by a non-AAV nucleic acid sequence.
Although Jin does not teach the specific positions of the amino acids of deamidation of SEQ ID NO: 1, it is a reference sequence so one of ordinary skill in the art can locate the specific amino acid positions in different capsid proteins. Nevertheless, Xiao et al. teaches the SEQ ID NO: 1.
Xiao et al. studies the Gene Therapy Vectors Based on Adeno-Associated Virus Type 1. It teaches a complete sequence of adeno-associated virus type 1 (AAV-1) and discloses the AAV-1 sequence with the GenBank accession # AF063497, where the CDS of” capsid protein” is identical to the claimed SEQ ID NO: 1 with an initial amino acid M (See below the aligned sequences in grey shadow). The aligned sequences are the amino acids sequences compared between the AAV1 VP1 amino acids sequences and the claimed SEQ ID NO: 1. It shows as: Query-SEQ ID NO: 1; Sbjct- 1“capsid protein”; N57-green, N383-blue, N512-pink and N-718-yellow), where the NG pairs (Asn-Gly) have been highlighted and considered as a “hot spot” associated with deamidation. This can be evidenced by Krokhin’s study. Krokhin et al. describes the deamidation of -Asn-Gly- Sequences during Sample Preparation for Proteomics: Consequences for MALDI and HPLC-MALDI Analysis. It teaches that the peptides containing -Asn-Gly- sequences typically show ~70-80% degree of deamidation after standard overnight (~12 h) tryptic digestion at 37 °C (See Abstract), and the deamidation of Asn and Gln residues is one of the best known and studied posttranslational modifications in proteins. Since the degree of deamidation often controls the biological activity and functions of proteins, the reaction is particularly important in large-molecule therapeutics (See page 6645, left column, paragraph 2) and the highest deamidation rate was found where -Asn- is followed by -Gly- (See page 6647, left column, paragraph 1).
Query 1 MAADGYLPDWLEDNLSEGIREWWDLKPGAPKPKANQQKQDDGRGLVLPGYKYLGPFNGLD 60
MAADGYLPDWLEDNLSEGIREWWDLKPGAPKPKANQQKQDDGRGLVLPGYKYLGPFNGLD
Sbjct 1 MAADGYLPDWLEDNLSEGIREWWDLKPGAPKPKANQQKQDDGRGLVLPGYKYLGPFNGLD 60
Query 61 KGEPVNAADAAALEHDKAYDQQLKAGDNPYLRYNHADAEFQERLQEDTSFGGNLGRAVFQ 120
KGEPVNAADAAALEHDKAYDQQLKAGDNPYLRYNHADAEFQERLQEDTSFGGNLGRAVFQ
Sbjct 61 KGEPVNAADAAALEHDKAYDQQLKAGDNPYLRYNHADAEFQERLQEDTSFGGNLGRAVFQ 120
Query 121 AKKRVLEPLGLVEEGAKTAPGKKRPVEQSPQEPDSSSGIGKTGQQPAKKRLNFGQTGDSE 180
AKKRVLEPLGLVEEGAKTAPGKKRPVEQSPQEPDSSSGIGKTGQQPAKKRLNFGQTGDSE
Sbjct 121 AKKRVLEPLGLVEEGAKTAPGKKRPVEQSPQEPDSSSGIGKTGQQPAKKRLNFGQTGDSE 180
Query 181 SVPDPQPLGEPPATPAAVGPTTMASGGGAPMADNNEGADGVGNASGNWHCDSTWLGDRVI 240
SVPDPQPLGEPPATPAAVGPTTMASGGGAPMADNNEGADGVGNASGNWHCDSTWLGDRVI
Sbjct 181 SVPDPQPLGEPPATPAAVGPTTMASGGGAPMADNNEGADGVGNASGNWHCDSTWLGDRVI 240
Query 241 TTSTRTWALPTYNNHLYKQISSASTGASNDNHYFGYSTPWGYFDFNRFHCHFSPRDWQRL 300
TTSTRTWALPTYNNHLYKQISSASTGASNDNHYFGYSTPWGYFDFNRFHCHFSPRDWQRL
Sbjct 241 TTSTRTWALPTYNNHLYKQISSASTGASNDNHYFGYSTPWGYFDFNRFHCHFSPRDWQRL 300
Query 301 INNNWGFRPKRLNFKLFNIQVKEVTTNDGVTTIANNLTSTVQVFSDSEYQLPYVLGSAHQ 360
INNNWGFRPKRLNFKLFNIQVKEVTTNDGVTTIANNLTSTVQVFSDSEYQLPYVLGSAHQ
Sbjct 301 INNNWGFRPKRLNFKLFNIQVKEVTTNDGVTTIANNLTSTVQVFSDSEYQLPYVLGSAHQ 360
Query 361 GCLPPFPADVFMIPQYGYLTLNNGSQAVGRSSFYCLEYFPSQMLRTGNNFTFSYTFEEVP 420
GCLPPFPADVFMIPQYGYLTLNNGSQAVGRSSFYCLEYFPSQMLRTGNNFTFSYTFEEVP
Sbjct 361 GCLPPFPADVFMIPQYGYLTLNNGSQAVGRSSFYCLEYFPSQMLRTGNNFTFSYTFEEVP 420
Query 421 FHSSYAHSQSLDRLMNPLIDQYLYYLNRTQNQSGSAQNKDLLFSRGSPAGMSVQPKNWLP 480
FHSSYAHSQSLDRLMNPLIDQYLYYLNRTQNQSGSAQNKDLLFSRGSPAGMSVQPKNWLP
Sbjct 421 FHSSYAHSQSLDRLMNPLIDQYLYYLNRTQNQSGSAQNKDLLFSRGSPAGMSVQPKNWLP 480
Query 481 GPCYRQQRVSKTKTDNNNSNFTWTGASKYNLNGRESIINPGTAMASHKDDEDKFFPMSGV 540
GPCYRQQRVSKTKTDNNNSNFTWTGASKYNLNGRESIINPGTAMASHKDDEDKFFPMSGV
Sbjct 481 GPCYRQQRVSKTKTDNNNSNFTWTGASKYNLNGRESIINPGTAMASHKDDEDKFFPMSGV 540
Query 541 MIFGKESAGASNTALDNVMITDEEEIKATNPVATERFGTVAVNFQSSSTDPATGDVHAMG 600
MIFGKESAGASNTALDNVMITDEEEIKATNPVATERFGTVAVNFQSSSTDPATGDVHAMG
Sbjct 541 MIFGKESAGASNTALDNVMITDEEEIKATNPVATERFGTVAVNFQSSSTDPATGDVHAMG 600
Query 601 ALPGMVWQDRDVYLQGPIWAKIPHTDGHFHPSPLMGGFGLKNPPPQILIKNTPVPANPPA 660
ALPGMVWQDRDVYLQGPIWAKIPHTDGHFHPSPLMGGFGLKNPPPQILIKNTPVPANPPA
Sbjct 601 ALPGMVWQDRDVYLQGPIWAKIPHTDGHFHPSPLMGGFGLKNPPPQILIKNTPVPANPPA 660
Query 661 EFSATKFASFITQYSTGQVSVEIEWELQKENSKRWNPEVQYTSNYAKSANVDFTVDNNGL 720
EFSATKFASFITQYSTGQVSVEIEWELQKENSKRWNPEVQYTSNYAKSANVDFTVDNNGL
Sbjct 661 EFSATKFASFITQYSTGQVSVEIEWELQKENSKRWNPEVQYTSNYAKSANVDFTVDNNGL 720
Query 721 YTEPRPIGTRYLTRPL 736
YTEPRPIGTRYLTRPL
Sbjct 721 YTEPRPIGTRYLTRPL 736
Accordingly, Xiao teaches a AAV1 gene delivery vector with a GenBank accession number of AF063497 that is identical to the claimed SEQ ID NO. 1. Based on the released sequence of capsid protein and the teaching of Jin and Krokhin, the four high deamidation NG sites in SEQ ID NO: 1, which is identical to the AAV1 VP1 sequences, are at N57, N383, N512, and N718 as claimed.
Actually, besides the deamidation of N57 of AAV1 taught by Jin et al., Jin et al. also teaches a NG-sites deamidation at N382, N511 and N713 (See Table 7, column 63 and below). After comparing Jin’s SEQ ID NOs: 10, 11 and 13 with Xiao’s AF063497 (AAV1) and the instant SEQ ID NO: 1, it discloses that the AAV1 deamidation sites of Jin matches with the claimed N57, N383, N512 and N718 (See the tables 4-6 below prepared by the Examiner).
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Accordingly, Jin et al. also teaches the four sites deamidation of AAV1 as claimed.
It would have been prima facie obvious for one having ordinary skill in the art before the effective filing date of the claimed invention to use known AAV1 capsid sequences, such as the AAV1 sequence of Xiao, in Jin’s invention and use the mass spectrometry method of Jin to determine the deamidation percentages in Xiao’s AAV1 VP1 sequence in the heterogeneous AAV1 composition. Based on the teachings of Krokhin regarding the NG pair sites for being a high frequency of deamidation, one of skill in the art would have been motivated to use the method of Jin to identify and modify the deamidations of AAV1 of Xiao, and there would be a reasonable expectation of success to identify and determine a AAV1 composition comprising the mixed populations of VP1, VP2 and VP3 with deamidation as claimed.
As for the “…comprising 50% to 100% asparagines (N) deamidated at each of positions: N57, N383, N512, and N718, -based on the numbering of SEQ ID NO: 1…’ as claimed in the base claim 1, Jin et al. teaches that in some embodiments, the AAV particle is deamidated at about 50%-100% ranges (See e.g., column 28, lines 42-53). Because Xiao et al. teaches an identical sequence to the claimed SEQ ID NO: 1 that contains only four “NG” pairs sites as claimed, based on the teaching of Jin, it is reasonably considered that the asparagine (N) deamidated at each of positions of N57, N383, N512, and N718 in SEQ ID NO: 1 is at 50%-100% ranges. At the same time, Jin et al. also discloses that “the extracted ion chromatograms (XIC) of peptides containing NG sites (T9, T49, and T67 in AA1 and AAV2 VP) and their corresponding deamidated species were used for calculation of deamidation levels” (See column 62, lines 64-67), where the T9 is SEQ ID NO: 9 (AAV1, see Table 7 above and Fig. 6A), T49 is SEQ ID NO: 11 (AAV1, see Table 7 above and column 13, lines 21-26) and T67 is SEQ ID NO: 13 (AAV1, see Table 7 above and column 13, lines 27-33). Here this description further discloses that the deamidation percentage Jin taught is based on the AAV1 NG pairs at positions: N57, N383, N512, and N718 (See Tables 4-6 above). Also, Krokhin et al. teaches the peptides containing -Asn-Gly- sequences (NG pair) typically show ~70-80% degree of deamidation (See Abstract).
Thus, the invention as a whole was clearly prima facie obvious to one of ordinary skill in the art before the effective filing date of the claimed invention.
Regarding claim 30, it requires that the AAV1 Vp1, VP2 and VP3 are produced from a nucleic acid sequence encoding a selected AAV vp 1 amino acid sequence. Jin et al. teaches that the rAAV particle is produced by an AAV
producer cell comprising nucleic acid encoding the rAAV vector and nucleic acid encoding AAV rep and cap functions, and providing nucleic acid encoding AAV helper functions (See e.g., column 8, lines 1-19). It discloses an amino acid sequence of “major coat protein VP1” containing the VP1, VP2 and VP3 of AAV2. The table 3 of Jin shows the theoretical masses of predicted sequences of 13 AAV serotypes based on sequence alignment and the intact protein analysis of several AAV serotypes (See below Table 3). Nevertheless, Xiao et al. discloses the AAV1 major coat protein VP1 nucleic acid sequence that encoding the VP1-3 (See Fig. 2 and the below).
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Regarding claims 31 and 32, they require the capsid comprises 65% to 100% deamidated asparagine at each position 57, and 75% to 100% deamidated asparagine at positions N383, N512, and/or N718 relative to the numbering of AAV1, as determined using mass spectrometry.
Jin et al. teaches that the N57 position of the NG-pair site is conserved among AAV1, AAV8 or AAV 9, where the deamidation percentages should be the same as what Jin taught (See Fig. 13, and below). Here Jin et al. teaches that in some embodiments, the AAV particle is deamidated to a higher extent compared to a parental AAV particle. In some embodiments, the AAV particle is more than about any of 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100% more deamidated compared to a parent AAV particle (See e.g., Column 28, lines 42-47), where one of the deamidation can be N57, N382, N511 and or N715 (See e.g., column 26, lines 58-67). Because the amino acids numbers in Jin et al. is based on AAV2 (See e.g., column 10, lines 52-56), the deamidated amino acids of N57, N382, N511 and or N715 in AAV2 is equal to the amino acids numbers N57, N383, N512 (See Table 1 below). Jin et al. does not point out the N717 (AAV2)/N718 9AAv1) deamidation, however, the sequence analysis shows that both AAV2 (Seq ID NO: 3 of Jin) and AAV1 (Seq ID No: 1 as claimed) include the NG pair which has a high deamidation rate as
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disclosed in Krokhin et al.
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Regarding claims 33 and 34, it requires all or a subpopulation of the AAV vp 1 proteins and/or vp3 proteins have a truncation of about 1 to about 5 amino acids at its N-terminus or -C-terminus respectively. Jin et al. teaches that in some embodiments, the heterogeneity comprises one or more of mixed serotypes, variant capsids, capsid amino acid substitutions, truncated capsids, or modified capsids (See column 2, lines 51-54). It teaches that the N-terminus of an AAV capsid protein (e.g., VP1 or VP3) may refer to the first amino acid after the initiating methionine, which in some cases may be removed by, e.g., a Met-aminopeptidase (See Column 26, lines 4-17). In Fig. 3, Jin et al. also teaches that the complete N-terminal and C-terminal peptides were covered by Lys-C digests as underlined in FIG. 3 (See Fig. 3; Column 55, lines 20-31), which indicates the truncated subpopulation of the AAV VP1 or VP3 proteins. Jin et al. discloses that in particular, a potential deamidation site is found at N57/G58 in the phospholipase A2 domain (Ca++ binding site), which is conserved among AAV 1, 2, 8 and others. The following experiments were aimed at exploring whether deamidation at N57 can lead to reduced potency and/or truncation of AAV2 (See Column 62, lines 26-41). It would have been prima facie obvious for one having ordinary skill in the art before the effective filing date of the claimed invention to set up an experimental to determine the truncation peptides and study the relations between the N57 deamidation and the truncation through N-terminus and C-terminus with a serial amino acids truction in order to improve/identify the stability, assembly and transduction of rAAAV particles.
Regarding claims 35, 36 and 38, they require that the AAV1 capsid comprises AAV1 vp l proteins, AAV1 vp2 proteins and/or AAV1 vp2 proteins and AAV1 vp3 proteins which are 70% to 100% N deamidated at each of positions: N57, N383, N512, N718, based on the numbering of SEQ ID NO: 1, where the amino acid sites are numbered in AAV1.
Based on the description above, Jin et al. teaches a composition comprising AAV particles wherein the AAV particles comprise one or more amino acid substitutions at amino acid residue A35, N57, G58, N382, G383, N511, G512, N715, or G716 of VP1 or VP3, residue numbering based on VP1 of AAV2, wherein the amino acid substitution alters deamidation as compared to deamidation of VP1 and/or VP3 of the parent AAV particle (See column 10, lines 52-67). Among the predicted/potential deamidation sites, the N57G58 is a highly conserved deamidation site among the AAV serotypes (See Fig. 13 and below). Therefore, the AAV1 numbers the N57. Since the SEQ ID NO: 1 is identical to AAV 1 capsid protein VP1 (See table 1 above), the aligned sequences teaches the potential NG deamidation sites as: the N382 in AAV2 is the N383 of AAV1 and the N511in AAV2 is N512 of AAV1 (See Table 1 above, where they are all in a N-G pairs. Jin et al. teaches that the AAV particle is more than about any of 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%,55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100% more deamidated compared to a parent AAV particle (See column 28, lines 42-63), where the percentage ranges of the deamidated asparagine claimed in the instant application is taught in Jin’s deamidation ranges.
Regarding claims 39-41, Jin et al. teaches that in some embodiments, the viral particle comprises an AAV ITR sequence. For example, an expression cassette may be flanked on the 5' and 3' end by at least one functional AAV ITR sequence to contain the heterologous transgene encoding a heterologous polypeptide (See e.g., column 35, lines 42-45; column 17, lines 33-41), where the heterologous transgene (i.e., nucleic acid sequence not of viral origin) that is flanked by at least one, e.g., two, inverted terminal repeat sequences (ITRs) (See e.g., column 16, lines 36-41). Also, Jin et al. teaches that an "AAV inverted terminal repeat (ITR)" sequence is an approximately 145-nucleotide sequence that is present at both termini of the native single-stranded AAV genome (See e.g., column 18, lines 10-13), and the viral particles comprising a recombinant self-complementing genome (See e.g., column 42, lines 41-45).
Regarding claim 42-43, Jin et al. teaches that in some embodiments, the heterologous nucleic acid encodes a therapeutic nucleic acid. In some embodiments, a therapeutic nucleic acid may include without limitation an siRNA, an shRNA, an RNAi, a miRNA, an antisense RNA, a ribozyme or a DNAzyme (See column 31, lines 12-16).
Regarding claims 44, 45 and 46, Jin et al. teaches in some embodiments, an AAV particle of the present disclosure (e.g., a rAAV particle) is in a pharmaceutical composition. The pharmaceutical compositions may be suitable for any mode of administration described herein or known in the art (See e.g., column 49, lines 47-63). As for the immunoglobulin product, Jin et al. teaches that the AAV vector may comprise as a transgene, a nucleic acid encoding a protein or functional RNA. Here one of the transgenes is the anti-angiogenic polypeptide (See e.g., claim 30, lines 31-50). The anti-angiogenic polypeptide can be an antibody such as bevacizumab (Avastin). Bevacizumab is a monoclonal antibody that targets vascular endothelial growth factor (VEGF), effectively blocking angiogenesis by preventing VEGF from binding to its receptors on endothelial cells (See Cancer Research UK, downloaded on 1-30-2025), which also teaches claim 45. Also, claim 45 does not provide any structural differences from the Ab of claim 44.
As for the claim 46, it is a common knowledge and technique that one of skilled in the art to select expressing an anti-viral immunoglobulin in claim 44 based on needs. Many researches can be evidenced for using AAV to deliver antiviral antibodies. For example, Martinez-Navio et al. teaches using AAV vector to deliver antibodies against HIV and SIV in Rhesus Monkeys (See Abstract).
Responses to Applicant’s Remarks
Applicant’s arguments filed on Sept. 11, 2025 has been received and fully considered.
Applicant’s arguments on the rejections under 35 U.S.C. 103 are not persuasive.
1). Applicant’s amendment on “…comprising 50% to 100% asparagines (N) deamidated at each of positions: N57, N383, N512, and N718, -based on the numbering of SEQ ID NO: 1…” does not overcome the teaching of Jin in view of Xiao and Krokhin. Xiao teaches an identical sequence as SEQ ID NO: 1 claimed in the instant application, where the SEQ ID NO:1 contains the only four NG sites at positions N57, N383, N512, and N718. Krokhin teaches that the deamidation of -Asn-Gly- Sequences (NG pair) during sample preparation and the peptides containing NG pair typically show about 70-80% degree of deamidation (See Abstract, Krokhin et al.). Because Jin teaches that in some embodiments, the AAV particle is deamidated between about 50% to 100% (See column 28, lines 42-52), the deamidation percentage at N57, N383, N512, and N718 can be in the range of 50% to 100% as claimed. Furthermore, Jin et al. discloses that “the extracted ion chromatograms (XIC) of peptides containing NG sites (T9, T49, and T67 in AA1 and AAV2 VP) and their corresponding deamidated species were used for calculation of deamidation levels” (See column 62, lines 64-67), where the T9 is SEQ ID NO: 9 (AAV1, see Table 7 above and Fig. 6A), T49 is SEQ ID NO: 11 (AAV1, see Table 7 above and column 13, lines 21-26) and T67 is SEQ ID NO: 13 (AAV1, see Table 7 above and column 13, lines 27-33). Here this description further confirms that the deamidation percentage Jin taught is based on the AAV1 NG pairs at positions: N57, N383, N512, and N718 (See Tables 4-6 above).
2). Applicant argued that Jin does not teaches or suggest the deamidated AAV1 amino acid residues that are defined in the instant application, or the recited high level of deamidation for each of these positions in an AAV1 capsid (See instant remarks, page 8).
Applicant’s argument is not persuasive.
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Jin’s invention mainly uses the AAV2 as example to study the deamidations. However, Jin also teaches the deamidation on AAV1 N57 (See Fig. 6A, sheet 11 and below).
In Jin’s table 7 (See Table 7, column 63 and above), Jin’s peptides of AAV1, SEQ ID NO: 10, 11 and 13, shows the identical sequence or over 95% homologues to SEQ ID NO: 1 and AAV1 AF063497 (See Table 4-6), where the deamidation sites taught by Jin here matches the claimed deamidated sites of N57, N383, N512 and N718.
As for the deamidation percentage, based on the description in 1) above, Jin teaches a deamidation percentage include the range of 50% to 100% at each position at N57, N383, N512, and N718. In addition, Jin uses the same methods of TTx (the triple transfection method) to produce the AAV particles for the deamidation as the applicant used (See the instant specification, e.g., column 63; column 46, lines 40-51), therefore, the results of the percentage of the deamidation should be inherently similar to each other by the same method. Here, Jin teaches that FIGS. 6A & 6B show the results of LC/MS/MS analysis comparing the percentage of deamidation in AAV1 and AAV2 particles produced by the TTx and PCL methods (See column 13, lines 15-20), and the instant specification discloses that FIG. SA and FIG. 8B show the results of in vitro analysis of the impact of genetic deamidation on vector performance. (FIG. SA) Titers of wtAAV8 and genetic deamidation mutant vectors produced by small-scale triple transfection in 293 cells, as measured by quantitative PCR (qPCR). Titers are reported relative to the wtAAV8 control (See instant specification, [0024]).
3). Applicant argued the combination references of Xiao, Krokhin, Martinez-Navia et al., and Cancer Research UK not teaching the deamidated capsid as recited in the claims (See instant Remarks, page 8).
Applicant’s argument is not persuasive.
The prior arts of Xiao, Krokhin, Martinez-Navia et al., and Cancer Research UK used here is for addressing a specific limitation as claimed. For example, Xiao teaches that the SEQ ID NO: 1 claimed in the instant application is a AAV1 sequences and prove that the claimed SEQ ID NO:1 is taught by a prior art. Krokhin teaches the Deamidation of -Asn-Gly- Sequences during Sample Preparation for Proteomics and discloses the NG “hot spot” for deamidation. The Caner Research UK teaches the Bevacizumab. Martinez-Navio et al. teaches using AAV vector to deliver antibodies against HIV and SIV in Rhesus Monkeys. Therefore, it is applicable to use these references to combine with Jin’s invention for the current office action.
Conclusion
No claims are allowed.
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/RUIXUE WANG/ Examiner, Art Unit 1671
/NICOLE KINSEY WHITE/ Primary Examiner, Art Unit 1671