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 .
Status of the Claims
Claims 1-18 are pending in the application.
Claim for Foreign Priority
This application is a PCT national Stage application of PCT/KR2022/001307, filed Jan. 25, 2022, is acknowledged and claims foreign priority to KR10-2021-0026617.
Information Disclosure Statement
The information disclosure statement (IDS) submitted on 8/25/2023 was filed prior to the mailing of the instant first Office action on the merits. The submission is in compliance with the provisions of 37 CFR 1.97. Accordingly, the information disclosure statements are being considered by the examiner. A copy of Form PTO/SB/08 is attached to the instant Office action.
Objections to the Specification
The use of the term TritonX-100 and Zwiiergent 3-16, which are a trade name or a mark used in commerce, has been noted in this application. The term should be accompanied by the generic terminology; furthermore the term should be capitalized wherever it appears or, where appropriate, include a proper symbol indicating use in commerce such as ™, SM , or ® following the term.
Although the use of trade names and marks used in commerce (i.e., trademarks, service marks, certification marks, and collective marks) are permissible in patent applications, the proprietary nature of the marks should be respected and every effort made to prevent their use in any manner which might adversely affect their validity as commercial marks.
Drawings
The presented drawings (Figure 1 through Figure 5) have been considered by the examiner.
Claim Rejections - 35 USC § 112
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claim 4 is rejected under 35 USC 112(b) because it recites the limitation "performed twice" in the claim language. It is not clear if the applicants mean that the primary centrifugation and secondary centrifugation is each performed twice or if they mean “twice” by employing primary centrifugation once and the second time is by utilizing secondary centrifugation.
Claim 9 is rejected under 35 USC 112(b) for recitation of trademarks/tradenames in the claims without providing the generic names. Claim 9 contains the trademark/trade name TritonX-100 and Zwittergent 3-16 in lines two and three of the claim. Where a trademark or trade name is used in a claim as a limitation to identify or describe a particular material or product, the claim does not comply with the requirements of 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph. See Ex parte Simpson, 218 USPQ 1020 (Bd. App. 1982). The claim scope is uncertain since the trademark or trade name cannot be used properly to identify any particular material or product. A trademark or trade name is used to identify a source of goods, and not the goods themselves. Thus, a trademark or trade name does not identify or describe the goods associated with the trademark or trade name. In the present case, the trademark/trade name is used to identify/describe surfactants and, accordingly, the identification/description is indefinite.
Claim 18 is rejected because it recites the limitation "a surfactant is additionally treated" in the claim. This language is not clear because there is no treatment step involving a surfactant listed in the claims and one of ordinary skill in the art would not know where and how it relates to the preamble of claim 17 or that of claim 1.
Claim Rejections - 35 USC § 102
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.
Claims 1, 3-4, 8, 10-15 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Tschantz, et. al., Expression, purification and characterization of the human membrane transporter protein OATP2B1 from Sf9 insect cells, Protein Expression and Purification, 57, (2008) 163–171).
Claim 1 is drawn to a method of producing a purified target protein by disrupting cells with a sonicator, a homogenizer, or a microfluidizer (instant claim 1 i)) to obtain a cell membrane fraction(s) (instant claim 1 ii)). A sonicator, a homogenizer, or a microfluidizer is then utilized to disrupt the cell membrane fractions (instant claim 1 iii) to separate target proteins (instant claim iv)). Regarding claim 1, Tschantz anticipates the applicant’s method for purifying target proteins by disrupting cells with a sonicator (page 165, first complete paragraph, line 5) (instant claim i)) to obtain cell membrane fraction(s) (page 166, line 2) (instant claim ii)). Tschantz then employs a homogenizer to disrupt the cell membrane fraction (page 166, lines 4-5) (Instant claim 1 iii)) to separate target proteins (page 166, lines 8-10) (instant claim iv)).
Claims 3-4 are drawn to a method wherein the cell membrane fractions are obtained by differential centrifugation; wherein the differential centrifugation is performed twice by primary and secondary centrifugation. It should be noted that the claims do not limit the claims to any specific xg of centrifugation force and is open ended. Tschantz anticipates (differential centrifugation) be employing primary centrifugation at 1000g (page 165, first full paragraph, line 6) and secondary centrifugation at 100,000g (page 166, line 1).
Claim 8 which depends from claim 1 is drawn to a method wherein an extraction buffer containing a surfactant is added to the cell membrane fractions. Tschantz anticipates this with the utilization of surfactant ASB-14-4 (page 166, line 6) to extract protein from the membrane fraction.
Claims 10-11 is drawn to a method where the separating uses ultracentrifugation. Tschantz anticipates these steps by employing ultracentrifugation at 100,000g after the membrane protein was extracted from the membrane protein fractions (page 166, line 10).
Claims 12-15 are drawn to the method of claim 1 wherein the target proteins are membrane proteins, wherein the cells are animal cells, and the animal cells are insect cells. The insect cells are any one selected from the group consisting of BTl-Tn-5B1-4 cells of Trichoplusiani, LD652Y cells of Lymantria dispar, Sf9 cells of Spodoptera frugiperda, Sf21 cells of Spodoptera frugiperda, Kcl cells of Drosophila, SL2 cells of Drosophila, and mosquito cell lines. Tschantz anticipates these claims wherein membrane proteins are purified from the insect cell line Sf9 of Spodoptera frugiperda (page 165, section titled “Purification of OATP2B1-FLAG”).
For the reasons outlined above, claims 1, 3-4, 8, 10-15 are rejected.
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.
Claims 1-18 are rejected under 35 U.S.C. 103 as being unpatentable over Tschantz, et. al., Expression, purification and characterization of the human membrane transporter protein OATP2B1 from Sf9 insect cells, Protein Expression and Purification, 57, (2008) 163–171 (examiner cited) in view of Microfluidics Processor User Guide, 2014) (examiner cited), Hofmann, et. al, (Wilson and Walker's Principles and Techniques of Biochemistry and Molecular Biology, Centrifugation and Ultracentrifugation Chapter 12, 424-453, 2017) (examiner cited) and Burgess, et. al., Methods of Enzymology, Chapter 35, 2009 (examiner cited).
Claim 1 is drawn to a method of producing a purified target protein by disrupting cells with a sonicator, a homogenizer, or a microfluidizer (instant claim 1 i)) to obtain a cell membrane fraction(s) (instant claim 1 ii)). A sonicator, a homogenizer, or a microfluidizer is then utilized to disrupt the cell membrane fractions (instant claim 1 iii) to separate target proteins (instant claim iv)).
Regarding claim 1, Tschantz teaches the applicant’s method for purifying target proteins by disrupting cells with a sonicator (page 165, first complete paragraph, line 5) (instant claim i)) to obtain cell membrane fraction(s) (page 166, line 2) (instant claim ii)). Tschantz then employs a homogenizer to disrupt the cell membrane fraction (page 166, lines 4-5) (Instant claim 1 iii)) to separate target proteins (page 166, lines 8-10) (instant claim iv)). Tschantz teaches the above cellular lysis methods utilizing a sonicator and a homogenizer in the development of their strategy to express, purify and characterize the human membrane transporter protein OATP2B1 from Sf9 insect cells.
Claims 3-4 are drawn to a method wherein the cell membrane fractions are obtained by differential centrifugation; wherein the differential centrifugation is performed twice by primary and secondary centrifugation. Tschantz teaches (differential centrifugation) be employing primary centrifugation at 1000g (page 165, first full paragraph, line 6) and secondary centrifugation at 100,000g (page 166, line 1).
Claim 8 is drawn to a method wherein an extraction buffer containing a surfactant is added to the cell membrane fractions. Tschantz teaches this with the utilization of surfactant ASB-14-4 (page 166, line 6) to extract protein from the membrane fraction.
Claims 10-11 is drawn to a method where the separating uses ultracentrifugation. Tschantz teaches these steps by employing ultracentrifugation at 100,000g after the membrane protein was extracted from the membrane protein fractions (page 166, line 10).
Claims 12-15 are drawn to the method of claim 1 wherein the target proteins are membrane proteins, wherein the cells are animal cells, and the animal cells are insect cells. The insect cells are any one selected from the group consisting of BTl-Tn-5B1-4 cells of Trichoplusiani, LD652Y cells of Lymantria dispar, Sf9 cells of Spodoptera frugiperda, Sf21 cells of Spodoptera frugiperda, Kcl cells of Drosophila, SL2 cells of Drosophila, and mosquito cell lines. Tschantz teaches these claims wherein membrane proteins are purified from the insect cell line Sf9 of Spodoptera frugiperda (page 165, section titled “Purification of OATP2B1-FLAG”).
Tschantz does not teach the use of a microfluidizer to perform a differential cellular lysis strategy in order to obtain target proteins from cell membrane protein fractions. However, the reference of Microfluidics does teach how to utilize a microfluidizer to develop such a strategy.
Microfluidics (page 4, last paragraph) outlines the operating capacity of a microfluidizer for cellular lysis as being between 500 psi and 40,000 psi (instant claims 1-2, 7) and Microfluidics (page 3, Figure 5) also teaches and compares the shear rates available to a Microfluidizer (and other common cellular lysis laboratory instruments). Furthermore, utilizing a Microfluidizer to correspond to the shear rates for cellular lysis of similar technologies, including that of a homogenizer and a sonicator, is taught by Microfluidics (page 4, Figure 6). Specifically, Microfluidics teaches one how to attune a microfluidizer to give the cellular lysis disruptive power of a sonicator up to and including to the power of mimicking a homogenizer, and then goes further beyond the homogenizer’s capabilities to that of the cell lysis power of a microfluidizer processor.
Tschantz and Microfluidics does not teach the rotor speeds utilized by the applicant for primary and secondary centrifugation but Hofmann does
Claim 5 is drawn to a method wherein primary centrifugation is performed at a speed of 8,000g to 11,000g; Hofmann teaches a primary centrifugation at 10,000g (page 441, figure 12.5(b)). Claim 6 is drawn to a method wherein the secondary centrifugation is performed at a speed of 140,000g to 160,000g; Hofmann teaches a secondary centrifugation at 150,000g (page 441, figure 12.5(b)).
Tschantz, Microfluidics, and Hofmann do not teach the use of a surfactant cited in claim 9, however this is remedied by the teachings of Burgess.
Claim 9 is drawn to a method wherein the surfactant is any one selected from the group consisting of SDS, TritonX-100, Nonidet P-40, octyl-b-D-glucopyranoside, n-dodecyl-b-D- maltoside, and Zwittergent 3-16.
Claim 18 is drawn to a method wherein a surfactant is additionally treated (Examiner is interpreting this claim limitation that it is drawn to a method wherein a surfactant is additionally added. The phrase “additionally treated” does not make any scientific sense). The teachings of Burgess (page 623, 3rd paragraph) teaches addition of TritonX-100 (instant claims 9, 18) as being utilized to solubilize membrane proteins. Claim 16 is drawn to a method wherein the target protein purification method comprises: in step i), performing homogenization 10 times to 20 times; in step ii), performing primary centrifugation at a speed of 2,500xg to 3,500xg for 10 minutes to 30 minutes and then performing secondary centrifugation at a speed of 4,000xg to 6,000xg for 20 minutes to 40 minutes; in step iii), performing homogenization three times to seven times; and in step iv), performing centrifugation at a speed of 4,000xg to 6,000xg for 20 minutes to 40 minutes. Claim 17 is drawn to a method wherein the target protein purification method comprises: in step i), performing sonication for 5 minutes to 30 minutes; in step ii), performing primary centrifugation at a speed of 100xg to 1,000xg for 1 minute to 3 minutes and then performing secondary centrifugation at a speed of 12,000xg to 14,000xg for 10 minutes to 20 minutes; in step iii), performing sonication for 10 minutes to 30 minutes; and in step iv), performing centrifugation at a speed of 10,000xg to 15,000xg for 20 minutes to 40 minutes.
Claims 16-17 are rejected as utilizing routine optimization. "[W]here the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation." In re Aller, 220 F.2d 454, 456, 105 USPQ 233, 235 (CCPA 1955) (MPEP 2144.05). The applicant applies and modifies workable ranges of commercially available instrumentation in both their cellular perturbation and centrifugation steps. See also KSR Int’l Co. v. Teleflex Inc., 550 U.S. 398, 416, 82 USPQ2d 1385, 1395 (2007) (identifying "the need for caution in granting a patent based on the combination of elements found in the prior art.").
At the time of the applicant’s invention, it would have been obvious to one of ordinary skill in the art to replace the cell lysis method using the sonicator taught by Tschantz with the use of a Microfluidizer using the teachings of Microfluidics User guide by modifying the settings on a Microfluidizer to obtain the varying shear forces necessary to perform a differential cellular lysis strategy of insect cells in order to obtain cellular membrane protein fractions in high yield. It would have also been obvious to applying the teachings of Burgess and Hofmann to utilize a surfactant in cell membrane fractions to solubilize the membrane proteins which are generally insoluble and employ the centrifugation techniques as outlined by the applicant. One would also have been motivated to include the use of a Microfluidizer and varying the pressure (psi) to achieve the smooth cellular lysis with a higher yield of the membrane fractions. One would have had a reasonable expectation of success because Tschantz readily provides all the steps to isolate the cell membrane fractions and the Microfluidizer publication provides the advantages of using a Microfluidizer to obtain the membrane fragments in higher yield. Therefore, the method of claims 1-18 would have been obvious to one of ordinary skill in the art at the time of invention.
For the reasons outlined above, claims 1-18 are rejected.
Status of the claims:
• Claims 1-18 are pending.
• Claims 1-18 are rejected.
• No claim is in condition for allowance.
Conclusion
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/A.S./Examiner, Art Unit 1656
/MANJUNATH N RAO/Supervisory Patent Examiner, Art Unit 1656