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 Arguments
The Amendment filed 03JUN2026 has been entered. No new matter has been entered. Applicant’s amendments have overcome each and every 112(b) rejections and claim objections previously set forth in the Non-Final Office Action mailed 04MAR2026.
Applicant's arguments filed 03JUN2026 have been fully considered.
Regarding claim 1, in response to applicant's arguments against the references individually, one cannot show no obviousness by attacking references individually where the rejections are based on combinations of references. See In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981); In re Merck & Co., 800 F.2d 1091, 231 USPQ 375 (Fed. Cir. 1986).
The previously applied rejection points out that DHELLIN teaches using a nuclease treatment. The question was the timing of when the nuclease may be applied. LIU is cited for teaching a nuclease treatment on/during the chromatography step as an alternative to applying the nuclease treatment separately for the purpose of reducing processing steps. Thus it is obvious to one having ordinary skill in the art to modify the method of DHELLIN to wash the chromatography resin with a nuclease wash buffer comprising a nuclease as taught by LIU in order to reduce steps as is known in the art of biological separations. While LIU may be purifying a different biological material, both DHELLIN and LIU are purifying biological materials and thus separation techniques for one application may be applicable to another application. As stated by the Supreme Court in KSR International Co. v. Teleflex Inc. (KSR), 550 U.S. 398, 82 USPQ2d 1385 (2007):
When a work is available in one field of endeavor, design incentives and other market forces can prompt variations of it, either in the same field or a different one. If a person of ordinary skill can implement a predictable variation, § 103 likely bars its patentability. Id. at 417, 82 USPQ2d at 1396. MPEP 2141.I.
The Applicant argues that separation of one biological molecule would not necessarily represent the optimal purification method for another biological molecule. An obviousness analysis does not need to consider the best or optimal result, only that one having ordinary skill in the art would have considered it obvious to combine or modify the primary reference with features of the secondary reference. See Intel Corp. v. Qualcomm Inc., 21 F.4th 784, 800 (Fed. Cir. 2021) (“It’s not necessary to show that a combination is the best option, only that it be a suitable option.”)
The alleged unexpected results are a result of a salt-activated nuclease (SAN), which are to be expected, because a nuclease is used for DNA digestion (removal) and the salt-activated nuclease allows for high salt tolerance in separations. As far as the results, use of a cation alone does not contain a SAN and thus a residual DNA is structurally intact and may be easier to remove in some cases. Use of a SAN and a minimal amount of a cation would allow for a minimal activity of the SAN and thus a residual DNA may have a combination of undigested and digested DNA and may complicate removal. Use of a SAN with a sufficient amount of a cation would allow for a sufficient amount of activity of the SAN to digest the DNA allowing for a sufficient amount of removal. See e.g. Salt Active Nucleases by ArcticZymes for a further explanation.
LANES is cited for teaching a SAN, which is an improved nuclease known in the art. All refences are analogous arts in the technological area of biological separations and thus may be properly combined in determination of obviousness. Furthermore, LANES is broadly directed towards methods of removing contaminating polynucleotides from a sample using [endonuclease] enzyme[s]. Abstract. Patents are relevant as prior art for all they contain (MPEP 2123).
The Supreme Court has made clear that an obviousness analysis “need not seek out precise teachings directed to the specific subject matter of the challenged claim, for a court can take account of the inferences and creative steps that a person of ordinary skill in the art would employ.” KSR Int’l Co. v. Teleflex Inc., 550 U.S. 398, 418 (2007). That is because “[a] person of ordinary skill is also a person of ordinary creativity, not an automaton.”
SMITH and JONES are cited for teaching various chromatography resins, which are known in the art. SMITH is cited for teaching of positively charged vesicles or negatively charged vesicles and that capture of both types of vesicles may require both an anionic exchange resin and a cationic exchange resin.
CLAIMS ENTERED
Claim 54 appears to be (previously presented) despite being labeled as (Currently Amended).
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 no obviousness.
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(s) 1,3,7,11,17,22,40,41,46,51,53-54,60,132 are rejected under 35 U.S.C. 103 as being unpatentable over DHELLIN (US 6899863) in view of LIU (US 20080286850).
Regarding claims 1,11, DHELLIN teaches a method for preparing membrane vesicles (title) including a method of purifying extracellular vesicles (EVs) (via e.g. chromatography; abstract) comprising:
Treating the sample with a nuclease (e.g. a DNase (Sigma) and an RNase (Sigma); C17/L34), which enable the fine separation of membrane vesicles (e.g. exosomes) from cell proteins or certain macromolecular components (DNA, RNA) or macromolecular complexes (C2/L8-15);
(i) contacting the sample with a chromatography resin (e.g. SOURCE 15Q; C3/L49-58; C17/L37-38); and,
(ii) washing the chromatography resin with a wash buffer (e.g. BTP buffer including NaCl; C17/L41-44);
wherein the wash buffer comprises a cation (e.g. Na+ dissolved in BTP buffer); and
wherein the (ii) washing follows the (i) contacting,
wherein the method results in a reduction of a concentration of residual nucleic acid molecules (via the nuclease) in the sample to purify the EVs.
DHELLIN does not teach the nuclease wash buffer comprises a nuclease. However, LIU teaches mdck cell lines supporting viral growth to high titers and bioreactor process using the same (title), wherein the virus is treated with Benzonase or other a non-specific endonuclease either e.g. before the material is clarified, or at the same time as an affinity chromatography process, which may reduce processing steps (par. [0323]).
Therefore, before the effective filing date of the invention, it would have been obvious to one of ordinary skill in the art to modify the method of DHELLIN to wash the chromatography resin with a nuclease wash buffer comprising a nuclease as taught by LIU in order to reduce steps for enabling the fine separation of membrane vesicles from cell DNA and RNA as is known in the art. The references are combinable, because they are in the same technological environment of biological separations. See MPEP 2141 III (A) and (G).
Regarding claim 3, DHELLIN teaches the chromatography resin is e.g. an anion exchange (AEX) resin (C2/L40).
Regarding claim 7, DHELLIN teaches the nuclease is an endonuclease of e.g. benzonase (par. [0114]).
Regarding claim 17, DHELLIN teaches the nuclease wash buffer further comprises an anion of e.g. Cl- dissolved in BTP buffer.
Regarding claim 22, while DHELLIN teaches a RPMI buffer (C17/L30-31), LIU teaches a suitable loading buffer (e.g. a sucrose-phosphate buffer) comprising a salt concentration of 11mM (or 0.011M), which anticipates the claimed range of from about 0.01 M to about 2.0 M, wherein the salt of the loading buffer is KPO4 (potassium phosphate; par. [0329]). It is obvious to one having ordinary skill in the art to select a suitable loading buffer known in the art for the chromatography step.
Regarding claim 40, DHELLIN teaches an example of a chromatography resin that is washed twice after the binding stage: a step to wash out non-binding molecules (i.e. until the OD drops to zero) and a second step to reinforce the interactions of the exosomes with the support (C16/L1-6).
It is obvious to one having ordinary skill in the art that such an additional washing would be “nuclease-free” in order to stop any nuclease enzymatic activity and wash away unwanted material.
Regarding claim 41, DHELLIN teaches the wash buffer comprises a salt at a concentration of 12mM (0.012 M), which anticipates the claimed range of e.g. at least about 0.01 M,
wherein the salt in the wash buffer is e.g. NaCl (BTP buffer; C17/L41-44).
Regarding claim 46, DHELLIN’s modified method teaches (iii) eluting the EVs from the chromatography resin by contacting the chromatography resin with an elution buffer (saline; C17/L44-46),
wherein (iii) occurs after (ii) contacting the chromatography resin with a nuclease wash buffer (nuclease wash buffer to wash the vesicles on the resin, LIU par. [0323], then elute the vesicles from the resin),
wherein the elution buffer comprises a salt concentration of 150mM-1M (DHELLIN C17/L44-46), which overlaps the claimed range of at least about 1.0 M to about 5.0 M and therefore establishes a case of prima facie obviousness. See MPEP 2144.05 I. It would have been obvious to one of ordinary skill in the art to select the instantly claimed range from the prior art range because prior art teaches the same utility over the selected range;
wherein the elution buffer has the result of releasing EVs from the chromatography resin (that’s the purpose of an elution buffer).
Regarding claim 51, DHELLIN teaches collecting an eluent after contacting the chromatography resin with the elution buffer (C14/L22-25).
Regarding claim 53, DHELLIN teaches the sample contacted with the chromatography resin comprises a starting concentration of the nucleic acid molecules (the sample has nucleic acid contaminants; see C10/L21-26; C12/L17-19), and
wherein the eluent comprises an eluted concentration of the nucleic acid molecules (as shown by peak analysis of the eluate),
wherein the eluted concentration of the nucleic acid molecules is less than that of the starting concentration of the nucleic acid molecules (materials are eluted with 25 column volumes and thus very dilute; C17/L44-46).
Differences in concentration […] will not support the patentability of subject matter encompassed by the prior art unless there is evidence indicating such concentration […] is critical (MPEP 2144.05.II.A. The concentration has not been established to provide any criticality or to provide any unexpected result/benefit over the prior art of record. It has been held that where the general conditions of a claim are disclosed in the prior art, discovering the optimum or workable ranges involves only routine skill in the art. In re Aller, 105 USPQ 233. Further, it has been held that discovering an optimum value of a result effective variable involves only routine skill in the art. In re Boesch, 617 F.2d 272, 205 USP 215 (CCPA 1980).+
Regarding claim 54, DHELLIN teaches subjecting the sample to an additional chromatography resin comprising e.g. an anion exchange chromatography (AEX) resin (“at least one anion exchange […] chromatography treatment of the sample”; abstract).
Regarding claim 60, DHELLIN teaches the sample is contacted with e.g. an AEX resin (C2/L40).
DHELLIN does not explicitly teach repeating the steps of contacting the sample with the chromatography resin; however, it has been held that it is common sense to one having ordinary skill in the art to repeat steps unless an unexpected result is established. Perfect Web Tech., Inc. v. InfoUSA, Inc., 587 F.3d 1324, 1328-29, 92 USPQ2d 1849, 1854 (Fed. Cir. 2009). See MPEP 2143.I.E.Example 9. In this case, repeating steps would be obvious to one having ordinary skill in the art in order to further improve purification of the liquid by subjecting the sample to the chromatography resin multiple times.
Regarding claim 132, DHELLIN is silent as to the concentration of EVs in the sample. However, differences in concentration […] will not support the patentability of subject matter encompassed by the prior art unless there is evidence indicating such concentration […] is critical (MPEP 2144.05.II.A. The concentration has not been established to provide any criticality or to provide any unexpected result/benefit over the prior art of record. It has been held that where the general conditions of a claim are disclosed in the prior art, discovering the optimum or workable ranges involves only routine skill in the art. In re Aller, 105 USPQ 233. Further, it has been held that discovering an optimum value of a result effective variable involves only routine skill in the art. In re Boesch, 617 F.2d 272, 205 USP 215 (CCPA 1980).
Claim(s) 30,33,111,133-134 are rejected under 35 U.S.C. 103 as being unpatentable over DHELLIN (US 6899863) in view of LIU (US 20080286850) and LANES (US 20140370514).
Regarding claims 30,133, LIU teaches the nuclease wash buffer comprises 50 units/mL (par. [0329]), which anticipates the claimed range of 1-100 units/mL. DHELLIN does not teach the nuclease wash buffer comprises SAN. However, LANES teaches endonucleases (title) and that the use of a salt active nuclease is convenient as salt can be added to limit DNA-protein interactions and remove DNA contaminations (par. [0141]). Note that such a SAN is coordinated with Mg2+ ions (LANES par. [0034]).
Therefore, before the effective filing date of the invention, it would have been obvious to one of ordinary skill in the art to modify the method of DHELLIN to provide the nuclease wash buffer comprising SAN as taught by LANES in order to conveniently remove DNA contaminations. The references are combinable, because they are in the same technological environment of separations. See MPEP 2141 III (A) and (G).
Regarding claim 134, DHELLIN teaches a method for preparing membrane vesicles (title) including a method of purifying extracellular vesicles (EVs) (via e.g. chromatography; abstract) comprising:
Treating the sample with a nuclease (e.g. a DNase (Sigma) and an RNase (Sigma); C17/L34), which enable the fine separation of membrane vesicles (e.g. exosomes) from cell proteins or certain macromolecular components (DNA, RNA) or macromolecular complexes (C2/L8-15);
(i) contacting the sample with an anion exchange resin (e.g. SOURCE 15Q; C3/L49-58; C17/L37-38); and,
(ii) washing the chromatography resin with a wash buffer (e.g. BTP buffer including NaCl; C17/L41-44);
wherein the wash buffer comprises a cation (e.g. Na+ dissolved in BTP buffer); and
wherein the (ii) washing follows the (i) contacting,
wherein the method results in a reduction of a concentration of residual nucleic acid molecules (via the nuclease) in the sample to purify the EVs.
DHELLIN does not teach the nuclease wash buffer comprises a nuclease. However, LIU teaches mdck cell lines supporting viral growth to high titers and bioreactor process using the same (title), wherein the virus is treated with Benzonase or other a non-specific endonuclease either e.g. before the material is clarified, or at the same time as an affinity chromatography process, which may reduce processing steps (par. [0323]).
Therefore, before the effective filing date of the invention, it would have been obvious to one of ordinary skill in the art to modify the method of DHELLIN to wash the chromatography resin with a nuclease wash buffer comprising a nuclease as taught by LIU in order to reduce steps for enabling the fine separation of membrane vesicles from cell DNA and RNA as is known in the art. The references are combinable, because they are in the same technological environment of biological separations. See MPEP 2141 III (A) and (G).
DHELLIN does not teach the nuclease wash buffer comprises SAN. However, LANES teaches endonucleases (title) and that the use of a salt active nuclease is convenient as salt can be added to limit DNA-protein interactions and remove DNA contaminations (par. [0141]). Note that such a SAN is coordinated with Mg2+ ions (LANES par. [0034]).
Therefore, before the effective filing date of the invention, it would have been obvious to one of ordinary skill in the art to modify the method of DHELLIN to provide the nuclease wash buffer comprising SAN as taught by LANES in order to conveniently remove DNA contaminations. The references are combinable, because they are in the same technological environment of separations. See MPEP 2141 III (A) and (G).
Regarding the concentration of SAN and the concentration of Mg2+ ions, differences in concentration […] will not support the patentability of subject matter encompassed by the prior art unless there is evidence indicating such concentration […] is critical (MPEP 2144.05.II.A. The concentration has not been established to provide any criticality or to provide any unexpected result/benefit over the prior art of record. It has been held that where the general conditions of a claim are disclosed in the prior art, discovering the optimum or workable ranges involves only routine skill in the art. In re Aller, 105 USPQ 233. Further, it has been held that discovering an optimum value of a result effective variable involves only routine skill in the art. In re Boesch, 617 F.2d 272, 205 USP 215 (CCPA 1980).
Regarding claim 33, DHELLIN’s modified method teaches the nuclease wash buffer comprises 1-20mM (or 0.001-0.020 M; LANES par. [0035]), which overlaps the claimed range of about 0.01- 1.0 M of the cation and therefore establishes a case of prima facie obviousness. See MPEP 2144.05 I. It would have been obvious to one of ordinary skill in the art to select the instantly claimed range from the prior art range because prior art teaches the same utility over the selected range; and,
wherein the cation comprises Mg2+ (LANES par. [0035]); and,
DHELLIN’s modified method teaches the nuclease wash buffer is contacted with the chromatography resin 3 times (LIU par. [0329]).
Regarding claim 111, LANES teaches the eluent is contacted with the nuclease wash buffer by incubation for e.g. 15min., which anticipates the claimed range of at about 1 minute to about 14 days, and at or around 35-37°C (par. [0041]), which overlaps the claimed range of e.g. about 37ºC and therefore establishes a case of prima facie obviousness. See MPEP 2144.05 I. It would have been obvious to one of ordinary skill in the art to select the instantly claimed range from the prior art range because prior art teaches the same utility over the selected range.
Claim(s) 56 is rejected under 35 U.S.C. 103 as being unpatentable over DHELLIN (US 6899863) in view of LIU (US 20080286850), SMITH (US 20070078374), and JONES (US 12083448).
Regarding claim 56, DHELLIN does not teaches a CEX resin. However, SMITH teaches iontophoretic delivery of vesicle-encapsulated active agents (title) including an inner ion selective membrane, which may be an anion exchange membrane (par. [0048,0051]) and an outermost ion selective membrane, which may be a cation exchange membrane (par. [0048, 0053]). SMITH teaches that vesicles can be positively or negatively charged (par. [0022]) and that an anion exchange membrane and a cation exchange membrane may be used to capture the desired charged vesicles (par. [0073]).
Therefore, before the effective filing date of the invention, it would have been obvious to one of ordinary skill in the art to modify the method of DHELLIN to provide for a CEX resin after an AEX resin as taught by SMITH in order to capture the desired membrane vesicles. The references are combinable, because they are in the same technological environment of separations. See MPEP 2141 III (A) and (G).
DHELLIN does not teaches a MMC resin. However, JONES teaches purification and labeling of extracellular vesicles using a mixed mode resin composition (title) and that a mixed mode resin has several advantages including inter alia it is simple to use, fast, does not dilute the EV concentration and provides EVs with a high purity (C3/L24-32).
Therefore, before the effective filing date of the invention, it would have been obvious to one of ordinary skill in the art to modify the method of DHELLIN to provide for a MMC resin as taught by JONES in order to improve EV purity. The references are combinable, because they are in the same technological environment of separations. See MPEP 2141 III (A) and (G).
Regarding the arrangement of the resins (e.g. AEX, CEX, MMC order or AEX, MMC, CEX order), the arrangement is a simple rearrangement of parts. This design choice does not change the operation of the device and thus one having ordinary skill in the art would have found it obvious absent persuasive evidence that a new and unexpected result is produced (see MPEP 2144.04(VI)(C)).
Telephonic Inquiries
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 LIAM A ROYCE whose telephone number is (571)270-0352. The examiner can normally be reached M-F ~08:00~15:00.
Examiner interviews are available via telephone, in-person, and video conferencing using a USPTO supplied web-based collaboration tool. To schedule an interview, applicant is encouraged to use the USPTO Automated Interview Request (AIR) at http://www.uspto.gov/interviewpractice.
If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Ben Lebron can be reached at (571) 272-0475. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
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LIAM A. ROYCE
Primary Examiner
Art Unit 1773
/Liam Royce/ Primary Examiner, Art Unit 1773
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