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 .
Response to Amendment
The Amendment filed 02/27/2026 in which claim 14 was amended, new claim 26 was added, and claim 20 was canceled, has been entered. Claims 1-13, 18, 23 and 24 were previously canceled.
Claims 14-17, 19, 21-22 and 25-26 are under examination on the merits.
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.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
(previous rejection, withdrawn as to claims 14, 19, 22) Claims 14, 19, 22, 26 were rejected under 35 U.S.C. 103 as being unpatentable over He et al. in view of Zhu et al. and Fan et al. as evidenced by Telles and Seto, Thermo Fisher 293A cell line User Guide 2016., and Lv et al., further in view of Zhang et al. (prior art of record).
See claims 14, 19, 22 as submitted on 02/27/2026.
Applicant’s amendments to claim 14, have overcome previous rejection to claims 14, 19, 22.
(previous rejection, withdrawn as to claims 15-17) Claims 15-17 were rejected under 35 U.S.C. 103 as being unpatentable over He et al., Zhu et al., Fan et al., and Zhang et al. as applied to claims 14, 19 and 22 above, and further in view of Khayat, et al. (2011) as evidenced by, Thermo Fisher Tech Tip #40, and Beckman Coulter 2025 (prior art of record).
See claims 15-17 as submitted on 02/27/2026.
Applicant’s amendments to claim 14, have overcome previous rejection to claims 15-17.
(previous rejection, withdrawn as to claims 20-21) Claims 20 and 21 were rejected under 35 U.S.C. 103 as being unpatentable over He et al., Zhu et al., Fan et al., and Zhang et al. as applied to claims 14, 19 and 22 above, and further in view of Invitrogen pcDNA3.4 (prior art of record) and GenBank Accession number: AHX25876.1. (prior art of record).
See claims 20 and 21 as submitted on 02/27/2026.
The previous rejection of claim 20 is moot in view of Applicant’s cancelation of this claim.
Applicant’s amendments to claim 14, have overcome previous rejection to claims 21.
(previous rejection, withdrawn as to claims 25) Claim 25 was rejected under 35 U.S.C. 103 as being unpatentable over He et al., Zhu et al., Fan et al., and Zhang et al. as applied to claims 14, 19 and 22 above, and further in view of GenBank Accession number: AHX25876.1 and Luo (prior art of record).
See claim 25 as submitted on 02/27/2026.
Applicant’s amendments to claim 14, have overcome previous rejection to claim 25.
(new rejection, necessitated by amendment as to claims 14, 19, 21, 22, and addition as to new claim 26) Claims 14, 19, 21, 22, 26 are rejected under 35 U.S.C. 103 as being unpatentable over He et al. in view of Zhu et al. and Fan et al., as evidenced by Telles and Seto, Thermo Fisher 293A cell line User Guide 2016., and Lv et al. (prior art of record), further in view of Zhang et al., Invitrogen pcDNA3.4 (prior art of record) and GenBank Accession number: KR559686 (2015) (cited in Applicant’s IDS submitted on 04/17/2025).
See claims 14, 19, 21, 22, 26 as submitted on 02/27/2026.
As to claim 14, it is noted that the amendment of “wherein the plasmid includes a recognition site for NheI, a Kozak sequence, and a recognition site for NotI and wherein the recognition site for NheI and the Kozak sequence are upstream from a start codon of the PCV2 gene, and the recognition site for NotI is incorporated after a termination codon of the PCV2 gene” is already taught by the cited prior art. Specifically, Invitrogen teaches the commercially available construct pcDNA3 and it further teaches including a Kozak sequence in addition to the gene of interest for optimal expression in mammalian cells (page 4, page 8) and GenBank Accession number: KR559686 teaches a gene sequence for the PCV2 capsid protein. Applicant’s remarks concerning the recognition sites has been acknowledged. Amended claim 14 recites, NheI and NotI. It is herein noted that He et al. (prior art of record) teach the use of KpnI and NotI recognition sites as well as the use of NheI and NotI for cloning of the PCV2 protein (page 5, under “Technical Solution”). Accordingly, the teachings of He et al. demonstrate that these restriction sites are in fact recognized as functional equivalents in the prior art and allow for proper cloning of inserts. Therefore, one of ordinary skill in the art would have been able to select either recognition site pair KpnI - NotI or recognition site pair NheI - NotI to clone the PCV2 capsid gene sequence into the pcDNA3 construct. See MPEP 2144.06. Substituting Equivalents Known For The Same Purpose: In order to rely on equivalence as a rationale supporting an obviousness rejection, the equivalency must be recognized in the prior art, and cannot be based on applicant’s disclosure or the mere fact that the components at issue are functional or mechanical equivalents. In re Ruff, 256 F.2d 590, 118 USPQ 340 (CCPA 1958).
With respect to all of the other limitations of claim 14, as previously explained, He et al. teach a method for producing porcine circovirus type 2 (PCV2) virus-like particles (VLPs) (Abstract; page 3, ¶ 1) wherein the produced particles are present in the cytoplasm or the nucleus of mammalian cells as verified by immunofluorescence (page 5, ¶ 9). He et al.’s method comprises the following steps:
Providing HEK-293A cells (cultured mammalian cells, as recited in claim 14) (page 4, ¶ 7, 8)
Transfecting HEK-293A cells with a recombinant plasmid pAD-ΔCap-ΔVp2 comprising the PCV2 gene encoding the capsid protein of PVC2 (page 4, ¶ 4, 5, 7, 8)
Washing and centrifuging the HEK-293A cells after they have been transfected (page 6, ¶ 2)
Resuspending the HEK-293A cells in a phosphate buffered saline (PBS) solution after centrifuging (page 6, ¶ 2)
Subjecting the HEK-293A cells to ultrasonic lysis (performing sonication, as recited in claim 14) (page 6, ¶ 2)
Performing two more centrifugation cycles after the HEK-293A cells were sonicated, one cycle for 24000 r/min for 15 min followed by a second cycle 20000 r/min for 10 min (page 6, ¶ 2)
He et al. do not teach the steps of (a) adding valproic acid (VPA) sodium salt to the transfected mammalian cells, wherein the addition of the VPA sodium salt inhibits cell proliferation, (b) performing multiple freeze and thaw cycles on the mammalian cells, and wherein the capsid protein is modified with a secretion signal sequence introduced at an NH2 terminal of the capsid protein.
However, Fan et al. teach the step (a) of adding valproic acid (VPA) sodium salt to mammalian cells for the benefit of enhancing viral gene expression, wherein the addition of the VPA sodium salt inhibits histone deacetylases (HDACs) thereby modifying gene transcription (Abstract; page 1, ¶ 2). As evidenced by Telles and Seto, HDACs act to promote cell cycle progression and proliferation, therefore HDAC inhibitors potently inhibit cell proliferation (Abstract; page 2, ¶ 1).
Zhu et al. teach a method for enhanced replication of porcine circovirus type 2 (PCV2) in mammalian cells (a porcine kidney cell line termed PK15) (Abstract, page 1), comprising the step (b) of performing three freeze-thawed cycles for retrieval of produced viral particles (page 7, ¶ 1).
Zhang et al. teach a method of secreted expression of CPV2 particles comprising a vector comprising the PCV2 capsid protein and the N-terminal secretion signal peptide for increased extracellular expression of a PCV2 capsid protein, wherein the vector induces an antibody response (Abstract, page 2, ¶ 5; page 9, ¶ 2).
It would have been prima facie obvious to a person of ordinary skill in the art, before the effective filing date to have included the teachings of Fan et al., Zhu et al., and Zhang et al. in a method for producing PCV2 viral particles for the benefit of enhancing intracellular and extracellular viral gene expression and later for effective retrieval of produced viral particles, as taught by Fan et al., Zhu et al., and Zhang et al.
One of ordinary skill in the art would have had a reasonable expectation of success for introducing a step of adding VPA sodium salt to mammalian cells and another step for performing three freeze-thaw cycles to retrieve intracellular and extracellular virus particles given that the methods of virus production, viral gene expression, and viral particle retrieval are known, successfully demonstrated, and commonly used as evidenced by the applied prior art.
Regarding claim 19, it is noted that no amendments were introduced to claim 19 in the amendment filed on 02/27/2026. As previously explained, He et al. further teach HEK-293 A cells (page 6, ¶ 2) which is a subcloned cell line of HEK-293 cells, as evidenced by Thermo Fisher 293A cell line User Guide (page 4, ¶ 1). Hence, HEK-293 A cells constitute a species within the HEK-293 genus. Accordingly, the teachings of He et al. meet the limitations of claim 19.
Regarding claims 21 and 26, it is noted that no amendments were introduced to claim 21 in the amendment filed on 02/27/2026. As previously explained, Invitrogen teaches the commercially available construct pcDNA3.4 and GenBank Accession number: KR559686 teaches a gene sequence for the PCV2d capsid protein.
Regarding claim 22, it is noted that no amendments were introduced to claim 22. As previously explained, Zhu et al. further teach the VLPs from the PCV2 strain BJW which correspond to the group PCV2b VLPs as evidenced by Lv et al. (Table 2, 3rd row) and GenBank Accession number: KR559686 teaches a gene sequence for the PCV2d capsid protein.
Accordingly, claims 14, 19, 21, 22 and 26 would have been prima facie obvious to one of ordinary skill in the art before the effective filing date, especially in the absence of evidence to the contrary.
(new rejection, necessitated by amendment as to claims 15-17) Claims 15-17 are rejected under 35 U.S.C. 103 as being unpatentable over He et al., Zhu et al., Fan et al., Zhang et al., Invitrogen pcDNA3.4 and GenBank Accession number: KR559686 as applied to claims 14, 19, 21, 22, 26 above, and further in view of Khayat, et al. (2011) as evidenced by, Thermo Fisher Tech Tip #40, and Beckman Coulter 2025 (prior art of record).
See claims 15-17 as submitted on 02/27/2026.
Regarding claim 15, it is noted that no amendments were introduced to claim 15 in the amendment filed on 02/27/2026. As previously explained, He et al. further teach washing mammalian cells previously transfected with PBS and centrifuging at 1000 r/min for 15 min and at 24000 r/min for 10 minutes. (page 6, ¶ 2). The centrifuging speed taught by He et al. can be expressed in units of gravity (g) depending on the size of the rotor. As evidenced by Thermo Fisher Tech Tip # 40, the centrifuging speeds taught by He et al. can range between 45 x g and 168 x g for the first centrifuging cycle and between 26,000 x g and 96,600 x g for the second centrifuging cycle, depending on the size of the rotor used.
He et al. does not teach the size of the rotor used, therefore the precise centrifuging speeds are not indicated.
However, Khayat et al. 2011 teach a method for producing purified PCV2 VLPs for the purpose of purifying and further solving the crystal structure of such particles (Abstract). Khayat et al. 2011 further teach PCV2 VLP purification comprising centrifuging the cells at 15,300 x g for 15 min, 184,048 x g for 2.5 hours, and 16,100 x g for 10 min (page 3, column 1, ¶ 6).
In view of the teachings of He et al. and Khayat et al 2011, the claimed centrifuging speeds and times as recited in claim 15 are considered to be those determined by routine optimization according to one of ordinary skill in the art.
It would have been prima facie obvious to a person of ordinary skill in the art, before the effective filing date, to have optimized the centrifuging speeds and times for the benefit of maximizing PCV2 viral particle retrieval and achieving a high degree of purification.
One of ordinary skill in the art would have had a reasonable expectation of success for optimizing the centrifuging speeds and times to maximize PCV2 viral particle retrieval and later purification given that the methods of virus production, viral particle retrieval, and viral particle purification are known, successfully demonstrated, and commonly used as evidenced by the applied prior art.
Regarding claim 16, it is noted that there was no amendment to claim 16. As previously explained, Zhu et al. further teach incubating mammalian cells at 37 °C, and storing at −80 °C (page 7, column 1, ¶ 2; page 4, column 1, ¶ 1). Therefore, the teachings of Zhu et al. on freezing and thawing temperatures meet the limitations of claim 16.
Regarding claim 17, it is noted that there was no amendment to claim 17. As previously explained, the cited prior art teaches a wide range of centrifuging speeds and times. Khayat et al. 2011 further teach a step of clarifying the pellet by centrifugation at 16,100 x g for 10 min at 4°C (page 3, column 1, ¶ 6), demonstrating that adding a step of centrifuging after the VLPs have been pelleted is well known and commonly practiced in the art. Accordingly, the centrifuging speeds and times recited in claim 17 are considered to be those determined by routine optimization according to one of skill in the art in view of the teachings of the cited prior art.
Accordingly, claims 15-17 would have been prima facie obvious to one of ordinary skill in the art before the effective filing date, especially in the absence of evidence to the contrary.
(new rejection, necessitated by amendment as to claim 25) Claim 25 is rejected under 35 U.S.C. 103 as being unpatentable over He et al., Zhu et al., Fan et al., Zhang et al., Invitrogen pcDNA3.4 and GenBank Accession number: KR559686 as applied to claims 14, 19, 21, 22, 26 above, and further in view of GenBank Accession number: AHX25876.1 and Luo (prior art of record).
See claim 25 as submitted on 02/27/2026.
Regarding claim 25, it is noted that there was no amendment to claim 25. As previously explained, the amino acid sequence of instant SEQ ID NO: 4 is 249 residues long. The sequence consists of the following:
Amino acid residues 1-16 – secretion signal peptide
Amino acid residues 17-249 – PCV2 capsid protein
As explained above, GenBank Accession number: AHX25876.1 teaches a gene sequence for the PCV2 capsid protein which shares 100% sequence identity with amino acid residues 17-249 in instant SEQ ID NO: 4. See alignment below (Qy is instant SEQ ID NO: 4; Db is AHX25876.1).
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401
780
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GenBank Accession number: AHX25876.1 does not teach amino acid residues 1-16 encoding a secretion signal peptide.
However, Luo teaches a method of producing VLPs in mammalian cells wherein the VLPs comprise fusion proteins (¶¶ [0031]-[0039]). Luo further teaches said fusion protein comprises a signal peptide to facilitate secretion of the VLPs. Said signal peptide may comprise a hemagglutinin (HA) signal peptide (¶¶ [0031]-[0039]). Luo further teaches an HA signal peptide SEQ ID NO: 96 which shares 100% sequence identity with amino acid residues 1-16 in instant SEQ ID NO: 4. See alignment below (Qy is instant SEQ ID NO: 4; Db is Luo’s HA secretion signal peptide).
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169
882
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It would have been prima facie obvious to one of ordinary skill in the art before the effective filing date to have incorporated the secretion signal peptide of Luo to the PCV2 capsid protein sequence in GenBank Accession number: AHX25876.1 for the benefit of facilitating secretion of the VLPs thereby maximizing VLP yields. See MPEP 2144.07. The selection of a known material based on its suitability for its intended use supported a prima facie obviousness determination in Sinclair & Carroll Co. v. Interchemical Corp., 325 U.S. 327, 65 USPQ 297 (1945).
One of ordinary skill in the art would have had reasonable expectation of success in incorporating the secretion signal peptide of Luo into the PCV2 capsid protein sequence in GenBank Accession number: AHX25876.1 given that the sequences are well known in the art and the methods of fusing signal peptide sequences to viral proteins are well known, successfully demonstrated, and commonly used as evidenced by the applied prior art.
Therefore, such a sequence comprising the entire length of the amino acid sequence of instant SEQ ID NO: 4 was an obvious embodiment to one of ordinary skill in the art before the effective filing date in view of the teachings of the GenBank Accession number: AHX25876.1 and Luo, especially in the absence of evidence to the contrary.
Response to Arguments
Applicant's arguments filed 02/27/2026 have been fully considered but they are not persuasive.
Applicant contends on page 6 of the Remarks submitted on 02/27/2026:
Applicant respectfully disagrees with the Office's position. Specifically, the Office appears to consider, without evidence, the combination of KpnI and NotI recognition sites to be equivalent to the combination of NheI and NotI recognition sites, as now recited in claim 14. However, MPEP § 2142 indicates that "rejections on obviousness cannot be sustained with mere conclusory statements; instead, there must be some articulated reasoning with some rational underpinning to support the legal conclusion of obviousness" (quoting In re Kahn, 441 F.3d 977, 988, 78 USPQ2d 1329, 1336 (Fed. Cir. 2006).
In response:
As noted above, He et al. already teach the restriction sites NheI, KpnI, and NotI for cloning of the PCV2 protein. The teachings of He et al. demonstrate that these restriction sites are in fact recognized as functional equivalents in the prior art and allow for proper cloning of inserts. Accordingly, one of ordinary skill in the art would have been able to select either recognition site pair KpnI - NotI or recognition site pair NheI - NotI to clone the PCV2 capsid gene sequence into the pcDNA3 construct. The interchangeable use of restriction enzymes to cleave nucleic acids at fixed positions with respect to their recognition sequence is a well-established everyday practice in the art relying on basic concepts in molecular biology, such recognition sequences are routinely included in multiple cloning sites within constructs as illustrated by the pcDNA3 vector. The restriction sites in question NheI, KpnI, and NotI are well recognized in the prior art as functional equivalents, as evidenced by the teachings of He et al.
Applicant contends on page 7 of the Remarks submitted on 02/27/2026:
The signal peptide from Luo referred to in the Office Action (Luo, SEQ ID NO: 96) is one of a laundry list of approximately 109 different signal peptides mentioned in Luo, and there is no teaching or suggestion that this particular signal peptide provides any advantage or benefit in the compositions or methods of Luo, let alone in an unrelated method for producing PCV2 VLPs as recited here. This is further evidenced by the fact that Luo only mentions SEQ ID NO: 96 once in the application, and only as one among the long list of other signal peptides. The only teachings in Luo are related to VLPs and fusion protein related to hepatitis B, and thus there is no teaching or suggestion in Luo that it would have been suitable to incorporate the sequence of SEQ ID NO: 96 in Luo into a PCV2 capsid protein sequence of GenBank, for inclusion in the presently claimed method. Moreover, even if a person of ordinary skill in the art would look to the teachings of Luo (which Applicant does not admit), Luo provides no motivation for specifically selecting SEQ ID NO: 96 among the list of more than 100 other signal peptides.
In response:
Luo was cited for teaching a sequence for a signal peptide well known in the art. As explained before, addition of known signal peptides to constructs comprising viral capsid proteins for the purpose of redirecting localization of particles has been extensively studied in the art and further it is routinely practiced (see Luo, see Zhang et al.) In the instant case, the teachings of Zhang et al. demonstrate that such addition of a signal peptides can indeed boost secretion of the produced PCV2 VLPs and maximize yields (see Zhang et al. Fig. 3). Further is noted that there is no indication that any particular signal peptide from those known in the art and taught by Luo for example, presents advantageous features over any other known signal peptide. The cited prior art teaches the exact elements of the entire amino acid sequence of SEQ ID NO:4 as well as clear motivations for adding a secretion signal peptide to the N-terminus of a PCV2 capsid protein.
Conclusion
No claims are allowed.
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 MARLENE V BUCKMASTER whose telephone number is (703)756-5371. The examiner can normally be reached M-F 8-5.
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/MARLENE V BUCKMASTER/Examiner, Art Unit 1672
/THOMAS J. VISONE/Supervisory Patent Examiner, Art Unit 1672
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