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 .
Election/Restrictions
Applicant’s election of Claims 1-18 and 20 in the reply filed on 07/10/2026 is acknowledged. Because applicant did not distinctly and specifically point out the supposed errors in the restriction requirement, the election has been treated as an election without traverse (MPEP § 818.01(a)).
have been withdrawn.
Claims 19 and 21-27 withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected Inventions, there being no allowable generic or linking claim. Election was made without traverse in the reply filed on 07/10/2026.
Claim Rejections - 35 USC § 102
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
The following is a quotation of the appropriate paragraphs of 35 U.S.C. 102 that form the basis for the rejections under this section made in this Office action:
A person shall be entitled to a patent unless –
(a)(1) the claimed invention was patented, described in a printed publication, or in public use, on sale, or otherwise available to the public before the effective filing date of the claimed invention.
(a)(2) the claimed invention was described in a patent issued under section 151, or in an application for patent published or deemed published under section 122(b), in which the patent or application, as the case may be, names another inventor and was effectively filed before the effective filing date of the claimed invention.
Claims 1, 6 , 9, 12-17 and 20 are rejected under 35 USC § 102(a)(1) and/or 102(a)(2) as being anticipated by Qu at al. (WO 2004113494 A2) herein known as Qu.
Regarding Claim 1, Qu is directed to methods for purifying adeno-associated virus (AAV) virions. More particularly, the invention relates to methods for purifying recombinant AAV (rAAV) virions containing packaged genomes from mixtures of AAV virions containing both packaged rAAV virions and AAV empty capsids lacking said genomes (Page 1, Lines 5-8).
Qu discloses a method for separating adeno-associated virus capsids fully packaged with genetic material from adeno-associated virus capsids not fully packaged with genetic material (Page 1, Lines 5-8).
The method comprising the following steps:
adding a liquid sample comprising adeno-associated virus capsids to a chromatography material (Page2, Lines 20-24, [The invention is based on the discovery of efficient and commercially viable methods for preparing stocks of rAAV virions with reduced amounts of empty capsids. The inventors herein have found that empty capsids can be separated from rAAV virions containing genetic material ("AAV vector particles") using column chromatography techniques]).
wherein the liquid sample comprises adeno-associated virus capsids of a purity of at least 90% and of a concentration of at least 1012 adeno-associated virus capsids/ml, of which at least 10% of the adeno-associated virus capsids are adeno-associated virus capsids fully packaged with genetic material, (Example 5, Page 32, Lines 1-9, [Using Anion Exchange Chromatography To demonstrate that empty capsids and AAV vector particles could be separated using column chromatography techniques, purified empty capsids and AAV vector particles were mixed together to generate samples containing both empty capsids and AAV vector particles. 3 ml of empty capsids with a concentration of 9E+13 particle/ml were mixed with 2 ml of full particles with the concentration of 7E+12 vector genomes (vg) /ml.])
wherein the chromatography material comprises a strong, or partially strong, anion exchange chromatography material (Page 24, Lines 15-24, [A number of suitable anion exchangers for use with the present invention are known and include without limitation, MACRO PREP Q (strong anion-exchanger available from BioRad, Hercules, CA); UNOSPHERE Q (strong anion-exchanger available from BioRad, Hercules, CA); POROS 50HQ (strong anion-exchanger available from Applied Biosystems, Foster City, CA); POROS 50D (weak anion-exchanger available from Applied Biosystems, Foster City, CA)]),
comprising a support and a ligand for binding to the adeno-associated virus capsids, wherein the chromatography material comprises a surface extender connecting the ligand to the support, wherein the surface extender is a polymer, wherein the polymer is selected from:
(i) a polymer having a naturally occurring skeleton, such as a polysaccharide, such as starch, cellulose, dextran, or agarose, and
(ii) a polymer having a synthetic skeleton, such as a polyvinyl alcohol, a polyacrylamide, a polymethacrylamide, or a polyvinyl ether; (Page 22, Lines 28-33, Page 23, Lines 1-4, [Suitable cation exchangers for both the first cation exchange column and the second cation exchange column, if used, include a wide variety of materials, known in the art, useful matrix materials include but are not limited to, cellulose matrices, such as fibrous, microgranular and beaded matrices; agarose, dextran, polyacrylate, polyvinyl, polystyrene, silica and polyether matrices; and composites. Particularly preferred herein are matrices containing the functional ligand R- SO.sub.3.sup.", preferably sulfopropyl or sulfoethyl resins.]).
b. eluting the adeno-associated virus capsids fully packaged with genetic material from the chromatography material; wherein the adeno-associated virus capsids eluted in step (b) are eluted into eluate fractions (Example2, Page 30, Lines 11-21 [The empty capsids and AAV vector particles were prepared as described above and loaded on columns separately. Three resins, Q SEPHAROSE, POROS 50HQ and UNOSPHERE Q, were used for column chromatography runs which were performed using the same programs to compare the elution profiles between empty and full particles. AAV empty capsids eluted at a lower salt concentration (eluted in earlier fractions) as compared with the AAV vector particles (eluted in later fractions); this phenomenon was observed for all three anionic resins tested. In the case of the UNOSPHERE Q resin, the empty capsids were eluted in fraction 10 in a salt gradient from 0 to 500 mM NaCl, while the AAV vector particles eluted in fraction 11; in the case of the Q SEPHAROSE column, the empty capsids were eluted in fractions 11, 12 and 13 while the AAV vector particles were eluted in fractions of 13, 14 and 15.]),
which eluate fractions combined comprise at least 50% of the adeno-associated virus capsids of the liquid sample added in step (a), of which at least 60% of the adeno-associated virus capsids are fully packaged with genetic material. (Page 25, Lines 28-32, Lines Using the techniques described above, more than 90% of the AAV empty capsids can be separated away from the AAV vector particles. Moreover, high recovery of AAV vector particles is readily achieved, i.e., more preferably more than 60% of the AAV vector particles can be recovered.]; Example 7, Page 33, Lines 30-33, [Using these techniques, approximately 60% of the vector was recovered. The vector recovered was essentially free of empty capsids (<10 of the particles were empty capsids) based on the SDS-PAGE silver staining analysis (Figure 3)).
Regarding Claim 6, Qu discloses the method according to claim 1, wherein the surface extender is dextran. (Page 22, Lines 28-33, Page 23, Lines 1-4, [Suitable cation exchangers for both the first cation exchange column and the second cation exchange column, if used, include a wide variety of materials, known in the art, useful matrix materials include but are not limited to, cellulose matrices, such as fibrous, microgranular and beaded matrices; agarose, dextran, polyacrylate, polyvinyl, polystyrene, silica and polyether matrices; and composites. Particularly preferred herein are matrices containing the functional ligand R- SO.sub.3.sup.", preferably sulfopropyl or sulfoethyl resins.]).
Regarding Claim 9, Qu discloses the method according to claim 1, wherein steps (a) and (b) comprise applying a buffer having a pH of from about 6.0 to about 10.5, such as from about 7.5 to about 9.5, optionally wherein said buffer comprises tris(hydroxymethyl)amino-methane (i.e., Tris), 1,3-bis(tris(hydroxymethyl)methylamino) propane (i.e., bis-Tris propane), triethanolamine, N-methyldiethanolamine, Diethanolamine, 1,3-diaminopropane, or ethanolamine ( Page 25, Lines 3- 11, [In particular, on the anion exchange column, empty capsids can separated from AAV vector particles using an appropriate buffer at a pH of from about pH 5 to pH 12, preferably pH 6 to pH 10, and even more preferably pH 7 to pH 9.5, such as pH 7.1, 7.2, 7.3, 7.4...8.0, 8.1, 8.2, 8.3, 8.4, 8.5...9.0, 9.1, 9.2, 9.3, 9.4, 9.5, or any pH between the stated ranges. Appropriate buffers for use with the anion exchange columns are well known in the art and are generally cationic or zwitterionic in nature. Such buffers include, without limitation, buffers with the following buffer ions: N-methylpiperazine; piperazine; Bis-Tris; Bis-Tris propane; Triethanolamine; Tris; N-methyldiethanolamine; 1,3-diaminopropane; ethanolamine; acetic acid, and the like]).
Regarding Claim12, Qu discloses the method according to claim 1, wherein the chromatography material is a polishing chromatography material (Page 22, Lines 24-27, [Fractions collected from the first cation exchange column are then applied to a second ion exchanger, i.e., a second cation exchange column and/or an anion exchange column, using different elution conditions that separate AAV empty capsids from AAV vector particles]).
Regarding Claim 13, Qu discloses The method according to claim 1, wherein the liquid sample added in step (a), is a pre-purified liquid sample (Page27, Lines 24-28, [The AAV preparation containing both AAV vector particles and empty capsids was diluted 2-fold to reduce the salt concentration and further purified through a Q SEPHAROSE column, the flow-through AAV vector particles and empty capsids were collected, and concentration and buffer exchange was performed using ultra-filtration and dialfilfration techniques. The final POROS 50HS column-purified product contained both AAV empty capsids as well as AAV vector particles containing recombinant vector genomes]).
Regarding Claim 14, Qu discloses the method according to claim 1, further comprising (al) pre- purifying adeno-associated virus capsids by separating adeno-associated virus capsids from an adeno-associated virus capsid-containing cell culture harvest, thereby obtaining a pre-purified liquid sample comprising adeno-associated virus capsids, before adding said pre-purified liquid sample comprising adeno-associated virus capsids to the chromatography material according to step (a), optionally wherein said pre-purifying comprises subjecting the adeno-associated virus capsid-containing cell culture harvest to chromatography, or to clarification followed by chromatography (Page22, Lines 4-13, [After harvesting the transfected host cell, a lysate is formed by disrupting the transfected host cells using techniques suitable for large-scale production, such as microfluidization. The lysate is then filtered (for example, through a 0.45 μm filter) and purified using column chromatographic methods as described herein. Other techniques have also been reported to determine the infectious titer of any AAV vector. See, e.g., Zhen et al., "An Infectious Titer Assay for Adeno-associated Virus (AAV) Vectors with Sensitivity Sufficient to Detect Single Infectious Events." Hum. Gene Ther. (2004) In Press. The purified AAV stock is then treated to remove empty capsids using column chromatography techniques, in a particularly preferred method of the invention).
Regarding Claim 15, Qu discloses the method according to claim 1, further comprising subjecting the eluate fractions comprising adeno-associated virus capsids fully packaged with genetic material, eluted in step (b), to one or more of the following steps:c1) concentrating the adeno-associated virus capsids to a pharmaceutically relevant dose,c2) replacing a buffer applied in step (b), with a pharmaceutically acceptable buffer, and/or c3) sterilizing the eluate fractions comprising adeno-associated virus capsids, thereby obtaining a pharmaceutical composition comprising adeno-associated virus capsids. (Page27, Lines 26-28, [The flow-through AAV vector particles and empty capsids were collected, and concentration and buffer exchange was performed using ultra-filtration and diafiltration techniques]; Page28, Lines 1-4, [The purified empty capsids and AAV vector particles were separately dialyzed against buffer composed of 10 mM sodium phosphate pH 7.4, 140 mM NaCl and 5% sorbital using dialysis cassettes (Pierce) at 4°C with three buffer exchanges]).
Regarding Claim 16, Qu discloses the method according to claim 1, wherein the adeno-associated virus capsids are capsids of adeno-associated virus serotype 8 (AAV8), adeno-associated virus serotype 5 (AAV5), adeno-associated virus serotype 1 (AAV1), adeno-associated virus serotype 2 (AAV2), adeno-associated virus serotype 4 (AAV4), adeno-associated virus serotype 6 (AAV6), adeno-associated virus serotype 7 (AAV7), adeno-associated virus serotype 9 (AAV9), or adeno-associated virus serotype 10 (AAV 10), or a variant thereof. (Page8, Lines 7-10, [By an "AAV vector" is meant a vector derived from an adeno-associated virus serotype, including without limitation, AAV-1, AAV-2, AAV-3, AAV-4, AAV-5, AAV- 6, AAV-7 and AAV-8. AAV vectors can have one or more of the AAV wild-type genes deleted in whole or part]).
Regarding Claim 17, Qu discloses the method according to claim 1, wherein the elution buffer of step (b) comprises a kosmotropic salt, wherein the salt comprises (i) an anion selected from a group consisting of CO32-, SO42-, S2O32-, H2PO4, HPO42-, acetate, citrate, and Cl-, and (ii) a cation selected from a group consisting of NH4+, K+, Na+, and Li+;optionally wherein the salt is sodium acetate (Example 4, Page31, Lines 26-29, [the column was washed with either 150 mM sodium acetate, 160 mM sodium acetate or 170 mM sodium acetate. The empty capsids were eluted most efficiently when 170 mM NaAc was used]; Example 5, Page32, Lines 11-14 [The column chromatography elution profile described above was used, and the data indicated that the empty capsids were eluted in the salt condition of 170 mM sodium acetate]).
Regarding Claim 20, Qu discloses the method according to claim 1, wherein the not fully packaged adeno-associated virus capsids are empty adeno-associated virus capsids and/or partially packaged adeno-associated virus capsids. (Abstract, [Methods for separating AAV empty capsids from mixtures of AAV vector particles and AAV empty capsids are described. The methods use column chromatography techniques and provide for commercially viable levels of recombinant AAV virions]; Page 1, Lines 5-8, [The invention relates to methods for purifying adeno-associated virus (AAV) virions. More particularly, the invention relates to methods for purifying recombinant AAV (rAAV) virions containing packaged genomes from mixtures of AAV virions containing both packaged rAAV virions and AAV empty capsids lacking said genomes]).
Claim Rejections - 35 USC § 103
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
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.
Claim 2-3 and 18 are rejected under 35 U.S.C. 103 as being unpatentable over Qu at al. (WO 2004113494 A2) herein known as Qu, as applied to the claims above, and in view of Spitali (WO 2012013682 A2) herein known as Spitali, and further in view of Kumpalume (US-8580931-B2), herein known as Kumpalume.
Regarding claim 2-3 and 18, Qu teaches all the limitations in the claims as set forth above.
Qu discloses wherein the elution buffer of step (b) comprises sodium acetate (Claim18) (Example 4, Page31, Lines 26-29, [the column was washed with either 150 mM sodium acetate, 160 mM sodium acetate or 170 mM sodium acetate. The empty capsids were eluted most efficiently when 170 mM NaAc was used]; Example 5, Page 32, Lines 11-14 [The column chromatography elution profile described above was used, and the data indicated that the empty capsids were eluted in the salt condition of 170 mM sodium acetate]).
However, Qu is silent to the method according to claim 1, wherein the ligand of the strong anion exchange chromatography material is defined by the following Formula I:
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wherein R1 is selected from Cl-C3 alkyl, and R2 and R3 are independently selected from Cl-C3 alkyl, CH2OH, and CH2CHOHCH3 (claim 2). The method according to claim 2, wherein each of R1, R2, and R3 is CH3 (claim 3), The method according to claim 1, wherein the chromatography material is defined by Formula IV:
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and wherein the elution buffer of step (b) comprises sodium acetate, optionally wherein the adeno-associated virus capsids are capsids of adeno-associated virus serotype 9 (AAV9) or a variant thereof (Claim 18).
Spitali is directed to the field of protein purification. More specifically, it relates to a process for the purification of antibody fragments (Page 1, Lines 2-4).
Spitali discloses wherein the ligand of the strong anion exchange chromatography material is defined by the following Formula I: (Example1, Page 21 , Lines 31-34, [The CDP870 Fab' pool was loaded onto a Capto Q™ anion exchange chromatography column from GE Healthcare [highly cross-linked rigid agarose bead (mean particle size 90 μηη) with a quaternary amine anion exchange ligand attached via a dextran linker]).
Kumpalume is directed to methods for the purification of alpha-1-antitrypsin (Col.1, Lines 10-11).
Kumpalume discloses wherein R1 is selected from Cl-C3 alkyl, and R2 and R3 are independently selected from Cl-C3 alkyl, CH2OH, and CH2CHOHCH3. (claim 2). The method according to claim 2, wherein each of R1, R2, and R3 is CH3 (claim 3), wherein the chromatography material is defined by Formula IV (Claim 18) (Col. 7, Lines 26-37, [Of particular utility as the anion exchange substrate in step (a1) or step (a2) is quaternary amino linked agarose, for example quaternary amino linked Sepharose.RTM. (Amersham Biosciences), in particular the substrate marketed under the name Capto Q.RTM. (Amersham Biosciences). Capto Q.RTM. is a high-capacity strong quaternary ammonium (O) anion exchanger coupled to a chemically modified (dextran coated), high-flow agarose matrix. The quaternary amino group in Capto Q is --N.sup.+(CH.sub.3).sub.3. Sepharose. RTM. is the commonly used trade name for agarose beads. Other suitable anion exchange substrates include cellulose, dextran and polymer-based beads.])
It would have been obvious to one of ordinary skill in the art before the effective filing date of the invention to modify Qu ‘s method for separating adeno-associated virus capsids, as taught by
Spitali and Kumpalume wherein the ligand of the strong anion exchange chromatography material is defined by the Formula I, wherein R1 is selected from Cl-C3 alkyl, and R2 and R3 are independently selected from Cl-C3 alkyl, CH2OH, and CH2CHOHCH3, (claim 2), wherein each of R1, R2, and R3 is CH3 (claim 3), wherein the chromatography material is defined by Formula IV, optionally wherein the adeno-associated virus capsids are capsids of adeno-associated virus serotype 9 (AAV9) or a variant thereof (Claim 18), because Capto Q.RTM. is a high-capacity strong quaternary ammonium (O) anion exchanger coupled to a chemically modified (dextran coated), high-flow agarose matrix (See Kumpalume, [Col. 7, Lines 26-37]), yielding nothing more than predictable results.
Claim 4 is rejected under 35 U.S.C. 103 as being unpatentable over Qu at al. (WO 2004113494 A2) herein known as Qu, as applied to the claims above, and in view of GE Healthcare Life Sciences (Capto™ S, Capto Q, and Capto DEAE, 2012), herein known as GE.
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(Table1, GE)
Regarding claim 4, Qu teaches all the limitations in the claims as set forth above.
However, Qu is silent to the method according to claim 1, wherein the ligand of the strong, or partially strong, anion exchange chromatography material is defined by the following Formula II:
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(II) wherein: m is an integer of from 1 to 3; R1 and R2 are independently selected from a Cl-C3 alkyl; R3, and R4 are independently selected from C1-C3 alkyl and CH2CHOHCH3; and Rs is selected from hydrogen, a Cl- C3 alkyl and CH2CHOHCH3; provided that if m is 1, the ligand of the strong, or partially strong, anion exchange chromatography material is defined by the following Formula III:
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(III) wherein n is an integer of from 0 to 3; provided that if n is 0, R3 and R4 are independently selected from Cl-C3 alkyl, and Rs is hydrogen or CH2CHOHCH3.
GE is directed to Capto S, Capto Q and Capto DEAE. Capto S, Capto Q and Capto DEAE are, respectively, strong anion and weak anion exchange media for packed bed chromatography that increase speed and throughput in capture and intermediate purification (Page 1).
Ge discloses the method according to claim 1, wherein the ligand of the strong, or partially strong, anion exchange chromatography material is defined by the following Formula II, wherein: m is an integer of from 1 to 3; R1 and R2 are independently selected from a Cl-C3 alkyl; R3, and R4 are independently selected from C1-C3 alkyl and CH2CHOHCH3; and Rs is selected from hydrogen, a Cl- C3 alkyl and CH2CHOHCH3; provided that if m is 1, the ligand of the strong, or partially strong, anion exchange chromatography material is defined by the following Formula III, wherein n is an integer of from 0 to 3; provided that if n is 0, R3 and R4 are independently selected from Cl-C3 alkyl, and Rs is hydrogen or CH2CHOHCH3. (Table1; [Page1, Section Anion and cation exchangers with fast mass transfer and high dynamic binding capacities, [For ion exchange, Capto S uses a sulfonate group, Capto Q uses a quartenary amine group and Capto DEAE uses a diethylaminoethyl group. The groups are linked to a high flow agarose base matrix modified with a dextran surface extender which further increases capacities and mass transfer properties. Fast mass transfer ensures high dynamic binding capacity over a wide range of residence times. High binding capacity also contributes to shortening the overall processing time as the total number of cycles may be reduced])
It would have been obvious to one of ordinary skill in the art before the effective filing date of the invention to modify Qu ‘s method for separating adeno-associated virus capsids, as taught by GE, wherein the ligand of the strong, or partially strong, anion exchange chromatography material is defined by the following Formula II, wherein: m is an integer of from 1 to 3; R1 and R2 are independently selected from a Cl-C3 alkyl; R3, and R4 are independently selected from C1-C3 alkyl and CH2CHOHCH3; and Rs is selected from hydrogen, a Cl- C3 alkyl and CH2CHOHCH3; provided that if m is 1, the ligand of the strong, or partially strong, anion exchange chromatography material is defined by the following Formula III, wherein n is an integer of from 0 to 3; provided that if n is 0, R3 and R4 are independently selected from Cl-C3 alkyl, and Rs is hydrogen or CH2CHOHCH3, because the groups are linked to a high flow agarose base matrix modified with a dextran surface extender which further increases capacities and mass transfer properties. Fast mass transfer ensures high dynamic binding capacity over a wide range of residence times. High binding capacity also contributes to shortening the overall processing time as the total number of cycles may be reduced (See GE, [Page1, Section Anion and cation exchangers with fast mass transfer and high dynamic binding capacities]), yielding nothing more than predictable results.
Claim 5 is rejected under 35 U.S.C. 103 as being unpatentable over Qu at al. (WO 2004113494 A2) herein known as Qu, as applied to the claims above, and in view of GE Healthcare Life Sciences (Capto™ S, Capto Q, and Capto DEAE, 2012), herein known as GE, as applied to the claims above, and further in view of Russo et al. (US 20140378532 A1) herein known as Russo.
Regarding claim 5, modified Qu teaches all the limitations in the claims as set forth above.
Qu discloses wherein the ligand is defined by Formula III and comprises
n is 0; R3 and R4 are ethyl; and R5 is hydrogen or CH2CHOHCH3 (Table1; [Page1, Section Anion and cation exchangers with fast mass transfer and high dynamic binding capacities, [For ion exchange, Capto S uses a sulfonate group, Capto Q uses a quartenary amine group and Capto DEAE uses a diethylaminoethyl group.])
However, modified Qu is silent to comprises a combination of two or more of the following structures (i)-(iv):
n is 0; R3 and R4 are ethyl; and R5 is hydrogen or CH2CHOHCH3;
n is 1; R1, R2, R3, R4 are ethyl; and R5 is hydrogen or CH2CHOHCH3;
n is 2; each R1 and R2 is ethyl; R3 and R4 is ethyl; and R5 is hydrogen or CH2CHOHCH3;
n is 3; each R1 and R2 is ethyl; R3 and R4 is ethyl; and Rs is hydrogen o CH2CHOHCH3.
Russo is directed to glycogen-based cationic polymers, to complexes comprising the said polymers and at least one anionic compound, and to the use of the said complexes for delivering anionic compounds [0001].
Russo discloses structures (ii) n is 1; R1, R2, R3, R4 are ethyl; and R5 is hydrogen or CH2CHOHCH3;
([0015], [DEAE-dextran is a polycationic polymer, obtained by reacting N,N-diethylaminoethyl chloride and dextran]; [0016], [DEAE-Dextran, represented by the following structural formula, has two substituents comprising nitrogenous residues, in which the nitrogen atoms have different physicochemical characteristics from each other];
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(DEAE-Dextran, Russo)
[0017], The first substituent comprises a tertiary amine function (indicated as N') with a pK.sub.a of about 9.5, which, at physiological pH, is in ionized form. The second substituent, known as the "tandem", comprises a quaternary ammonium group (N.sup.+), which has a permanent positive charge and influences the acidity of the second tertiary amine function (indicated as N'') which has a pK.sub.a of about 5.7 and then, at physiological pH, is in non-ionized form]).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the invention to modify modified Qu ‘s method for separating adeno-associated virus capsids, as taught by Russo, wherein the ligand is defined by Formula III and comprises a combination of two or more of the following structures (i)-(iv):
n is 0; R3 and R4 are ethyl; and R5 is hydrogen or CH2CHOHCH3;
n is 1; R1, R2, R3, R4 are ethyl; and R5 is hydrogen or CH2CHOHCH3;
n is 2; each R1 and R2 is ethyl; R3 and R4 is ethyl; and R5 is hydrogen or CH2CHOHCH3;
n is 3; each R1 and R2 is ethyl; R3 and R4 is ethyl; and Rs is hydrogen o CH2CHOHCH3.
Because one of ordinary skill in the art could have combined the elements as claimed by known methods to yield predictable result (See MPEP 2143(I)(A, Examples 1-8)
Claim 7 is rejected under 35 U.S.C. 103 as being unpatentable over Qu at al. (WO 2004113494 A2) herein known as Qu, as applied to the claims above, and in view of Blanche et al. (US 6537793 B2) herein known as Blanche.
Regarding claim 7, Qu teaches all the limitations in the claims as set forth above.
However, Qu is silent to the method according to claim 6, wherein the dextran has a molecular weight of from about 10 to about 2000 kDa, such as about 40 kDa.
Blanche is directed to a new method for the purification and quantification of viral particles. More particularly, the invention relates to a method of purifying and quantifying adenoviruses by ion-exchange chromatography. The invention also relates to a method of identifying various adenovirus serotypes.
Blanche discloses the method according to claim 6, wherein the dextran has a molecular weight of from about 10 to about 2000 kDa, such as about 40 kDa. (Col. 3, Lines 48-54, [By way of polymer of natural origin, there may be mentioned in particular polymers of a polysaccharide nature chosen from starch, cellulose, dextran and agarose. Preferably, the degree of polymerization of the flexible arm is about 30 monomeric units, more preferably, the flexible arm is a dextran having a mean molecular weight of about 5000 Da]).
Blanche also discloses flexible ion-exchange arms have been developed for preparative and industrial chromatography of biomolecules. These supports have been more particularly designed for the step of capturing biomolecules from crude mixtures which have been simply clarified. Their performance was optimized in terms of a very high capacity for attachment of solutes to the support, of a very low counterpressure at a high linear liquid flow rate, of a low cost as well as of a very high chemical resistance to the cleaning agents used for regeneration (Col.3, Lines 29-41), advantageously, the flexible arm is of a hydrophilic nature. (Col.3, Line 24).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the invention to modify Qu ‘s method for separating adeno-associated virus capsids, as taught by Blanche, wherein the dextran has a molecular weight of from about 10 to about 2000 kDa, such as about 40 kDa, because
The performance of flexible ion-exchange arms was optimized in terms of a very high capacity for attachment of solutes to the support, of a very low counterpressure at a high linear liquid flow rate, of a low cost as well as of a very high chemical resistance to the cleaning agents used for regeneration, advantageously, the flexible arm (dextran) is of a hydrophilic nature (See Blanche, [Col.3, Lines 24 and 29-41]), yielding nothing more than predictable results.
Claim 8 and 11 are rejected under 35 U.S.C. 103 as being unpatentable over Qu at al. (WO 2004113494 A2) herein known as Qu, as applied to the claims above, and in view of Berg et al. (US 6428707 B1) herein known as Berg.
Regarding claim 8 and 11, Qu teaches all the limitations in the claims as set forth above.
However, Qu is silent to the method according to claim 6, wherein the density of dextran is from about 5 to about 30 mg dextran per ml of the strong, or partially strong, anion exchange chromatography material (Claim 8), the method according to claim 1, wherein the support of the chromatography material comprises particles, nanofibres, a monolith, or a membranous structure; optionally wherein the particles are particles having a homogeneous porosity and being at least partly permeable to adeno-associated virus capsids; optionally wherein the particles are substantially spherical particles; optionally wherein the nanofibres comprise electrospun polymer nanofibres; optionally wherein the membranous structure comprises a nonwoven web of polymer nanofibers (Claim 11).
Berg is directed to a process for the separation of a substance by adsorption to non-packed beds containing beads which exhibit structures (ligands) with affinity to the substance. Non-packed beds may be generated by expanding/fluidizing sedimented beads by an upward or a downward flow, the direction of flow being depending on the density of the beads used. A liquid sample containing the substance to be adsorbed is introduced into the flow after expansion. A less effective non-packed bed is generated by agitating suspendible beads with the aid of a turbulent flow or by mechanical stirring (Col.1, Lines 8-18).
Berg discloses wherein the density of dextran is 22 or 29 mg dextran per ml of the strong, or partially strong, anion exchange chromatography material which anticipate the claimed range of about 5 to about 30 mg dextran per ml of the strong, or partially strong, anion exchange chromatography material (Claim 8). (Example 1, Col.8, Lines 20-29; Example 2, Col.8, Lines 37-42; Example 3, Lines 44-48, [Introduction of quaternary aminomethyl groups (Q-groups) (anion exchanger). 250 ml of glycidyltrimethyl ammonium chloride (G-MAC, 70%) were added under stirring to a solution containing 250 ml of the matrix from Example 1]).
Berg also discloses wherein the support of the chromatography material comprises particles, nanofibres, a monolith, or a membranous structure; optionally wherein the particles are particles having a homogeneous porosity and being at least partly permeable to adeno-associated virus capsids; optionally wherein the particles are substantially spherical particles; optionally wherein the nanofibres comprise electrospun polymer nanofibres; optionally wherein the membranous structure comprises a nonwoven web of polymer nanofibers (Claim 11). (Col.5, Lines 22-26, [The base matrix of the beads may be of organic or inorganic nature as known for beads used in chromatography on fluidized and packed beads. It may be porous or non-porous. It is often a polymer, such as glass, a synthetic polymer or a biopolymer]).
Berg also discloses We have now realized that the above-mentioned problems can be solved by utilizing beads in which the affinity structure/ligand is linked to the base matrix of the beads via an extender (Col.3, Lines 6-9), The positive effect caused by an extender is believed to reside in the fact that it will provide the inner surfaces (pore surfaces) and/or outer surfaces of the beads with a flexible polymer layer that is permeable to macromolecules and other molecules allowed to pass the bed. This will cause an increase in the effective interacting volume as well as in the steric availability of the affinity structures/ligands for the substance to be adsorbed. This in turn will increase the mass transfer rate as well as the total capacity available (Col3., Lines 11-19).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the invention to modify Qu ‘s method for separating adeno-associated virus capsids, as taught by Berg, wherein the density of dextran is from about 5 to about 30 mg dextran per ml of the strong, or partially strong, anion exchange chromatography material (Claim 8), wherein the support of the chromatography material comprises particles, nanofibres, a monolith, or a membranous structure; optionally wherein the particles are particles having a homogeneous porosity and being at least partly permeable to adeno-associated virus capsids; optionally wherein the particles are substantially spherical particles; optionally wherein the nanofibres comprise electrospun polymer nanofibres; optionally wherein the membranous structure comprises a nonwoven web of polymer nanofibers (Claim 11), because the positive effect caused by an extender is believed to reside in the fact that it will provide the inner surfaces (pore surfaces) and/or outer surfaces of the beads with a flexible polymer layer that is permeable to macromolecules and other molecules allowed to pass the bed. This will cause an increase in the effective interacting volume as well as in the steric availability of the affinity structures/ligands for the substance to be adsorbed. This in turn will increase the mass transfer rate as well as the total capacity available (See Berg, (Col3., Lines 11-19), yielding nothing more than predictable results.
Claim 10 is rejected under 35 U.S.C. 103 as being unpatentable over Qu at al. (WO 2004113494 A2) herein known as Qu, as applied to the claims above, and in view of Chunlei et al. (Developing an Anion Exchange Chromatography Assay for Determining Empty and Full Capsid Contents in AAV6.2, Year: 2019) herein known as Chunlei.
Regarding claim 10, Qu teaches all the limitations in claim 1 as set forth above.
However, Qu is silent to the method according to claim 1, wherein step (b) comprises applying a buffer comprising a compound which improves separation between capsids fully packaged with genetic material and capsids not fully packaged with genetic material, optionally wherein said compound is selected from a carbohydrate, a divalent metal ion, and a detergent; optionally wherein the carbohydrate is selected from sucrose, sorbitol, and a polysaccharide; optionally wherein the divalent metal ion is selected from Mg2+, and Mn2+,optionally wherein the divalent metal ion is present in the form of a salt, optionally in combination with chloride ions or sulphate ions; optionally wherein the detergent is selected from poloxamer and polysorbate.
Chunlei is directed to Developing an Anion Exchange Chromatography Assay for Determining Empty and Full Capsid Contents in AAV6.2 (Title).
Chunlei discloses wherein step (b) comprises applying a buffer comprising a compound which improves separation between capsids fully packaged with genetic material and capsids not fully packaged with genetic material, optionally wherein said compound is selected from a carbohydrate, a divalent metal ion, and a detergent; optionally wherein the carbohydrate is selected from sucrose, sorbitol, and a polysaccharide; optionally wherein the divalent metal ion is selected from Mg2+, and Mn2+,optionally wherein the divalent metal ion is present in the form of a salt, optionally in combination with chloride ions or sulphate ions; optionally wherein the detergent is selected from poloxamer and polysorbate. (Result section, Page2, Right Col., [The structural integrity of AAV capsids under chromatographic conditions is a prerequisite for a separation method of empty and full capsids. The on-column stability of AAV capsids has not been reported previously as a concern. Divalent cations are known to stabilize non-enveloped viruses under thermal stress, and Urabe et al. included MgCl2 in the sample dilution buffer prior to loading AAV1 on preparative-scale columns. In our case, the presence of 2 mM MgCl2 resulted in the disappearance of the broad peaks and a dramatic increase in the intensity of the first two peaks (Figure 1B)])
It would have been obvious to one of ordinary skill in the art before the effective filing date of the invention to modify Qu ‘s method for separating adeno-associated virus capsids, as taught by Chunlei, wherein the method according to claim 1, wherein step (b) comprises applying a buffer comprising a compound which improves separation between capsids fully packaged with genetic material and capsids not fully packaged with genetic material, optionally wherein said compound is selected from a carbohydrate, a divalent metal ion, and a detergent; optionally wherein the carbohydrate is selected from sucrose, sorbitol, and a polysaccharide; optionally wherein the divalent metal ion is selected from Mg2+, and Mn2+,optionally wherein the divalent metal ion is present in the form of a salt, optionally in combination with chloride ions or sulphate ions; optionally wherein the detergent is selected from poloxamer and polysorbate, because divalent cations are known to stabilize non-enveloped viruses under thermal stress and the presence of 2 mM MgCl2 resulted in the disappearance of the broad peaks and a dramatic increase in the intensity of the first two peaks (See Chunlei, [Result section, Page2, Right Col.]), yielding nothing more than predictable results.
Conclusion
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/MAHMOUD MOTAZ ABDEL LATIF/Examiner, Art Unit 1773 /BENJAMIN L LEBRON/Supervisory Patent Examiner, Art Unit 1773
