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 .
Claims 25, 30, 51, 53, 55, 58-61, 66, and 69-76, of record 7/27/2026, are pending and subject to prosecution. Claim 73 is amended.
Status of Prior Rejections/Response to Arguments
RE: Rejection of claim 73 under 35 U.S.C. 112(b):
The amendment to claim 73 is effective to obviate the rejection. The rejection is withdrawn.
RE: Rejection of claims 25, 30, 51, 53, 55, 58-61, and 69-75 under 35 U.S.C. 103 over Steiner et al. (Nature Biotechnology, 2010) in view of Gao et al. (Stem Cell Research, 2019), Naujok et al. (BMC Research Notes, 2014), and Laco et al. (Stem Cell Reports, 2018), and evidenced by Watanabe et al. (Nature Biotechnology, 2007), Garcia-Gonzalo et al., (PLoS One, 2008), and Life Technologies (Product data sheet, 2014):
RE: Rejection of claims 25, 30, 51, 53, 55, 58-61, 66, and 69-76 under 35 U.S.C. 103 over Steiner et al. (Nature Biotechnology, 2010) in view of Gao et al. (Stem Cell Research, 2019), Naujok et al. (BMC Research Notes, 2014), and Laco et al. (Stem Cell Reports, 2018), and evidenced by Watanabe et al. (Nature Biotechnology, 2007), Garcia-Gonzalo et al., (PLoS One, 2008), and Life Technologies (Product data sheet, 2014), further in view of Ito et al. (WO 2020022511 A1, machine translation):
The applicant asserts that:
Gao et al. and Naujok et al. do not teach coordinated transient administration of a ROCK inhibitor with a GSK3 inhibitor, as required by independent claim 25, and the cited references do not provide articulated reasons for modifying the method of Steiner et al. (Applicant Remarks, page 6-7).
Laco et al. teach transient treatment with a GSK inhibitor to promote exit from, rather than maintenance of, pluripotency, and address a different problem than the applicant (Applicant Remarks, page 7-8).
The cited references do not teach or suggest the balance among expansion, spheroidal morphology, and maintenance of pluripotency demonstrated by the claimed invention (Applicant Remarks, page 8-9).
The rejections of record are the product of reconstruction based upon the instant disclosure (Applicant Remarks, page 9).
The applicant’s arguments have been fully considered but are not found persuasive.
Regarding the asserted deficiencies of the cited references, prior art is not required to teach or suggest all claim limitations when a person of ordinary skill in the art would have found them obvious. See MPEP 2141(III). The cited references all involve the culture of pluripotent stem cells; that they have different experimental endpoints does not negate any teachings that directly pertain to the culture of pluripotent stem cells, and a person of ordinary skill in the art would be free to apply those teachings to other situations wherein those cells are cultured.
Regarding the argument that Laco et al. teach CHIR99021 as promoting differentiation rather than pluripotency, cardiac differentiation was achieved using embryoid bodies and stem cells at day 11 or 14 following treatment with CHIR99021 and only within a narrow concentration range and at various cell-cycle profiles (See fig. 3). Some pluripotent cell lines showed no or little expression of the cardiac marker troponin after treatment with CHIR99021 (See fig. S2E, S2G, S33A, and S3D), implying that cardiac differentiation is not a global phenomenon. Further, Laco et al. found that CHIR99021 decreases hPSC monolayer protein mass after 24 h exposure (See page 1855, col. 1, ¶1-2 and fig. 1); Naujok et al. likewise noted cytotoxicity associated with CHIR09921 but found that it was best able to balance Wnt activation/pluripotency marker expression and cell viability in ESCs among small molecule GSK inhibitors (See fig. 1-3). The combined teachings of Laco et al. and Naujok et al. would suggest to one of ordinary skill a benefit to short-term exposure to CHIR99021 for pluripotent cells.
In response to applicant's argument that the examiner's conclusion of obviousness is based upon improper hindsight reasoning, it must be recognized that any judgment on obviousness is in a sense necessarily a reconstruction based upon hindsight reasoning. But so long as it takes into account only knowledge which was within the level of ordinary skill at the time the claimed invention was made and does not include knowledge gleaned only from the applicant's disclosure, such a reconstruction is proper. See In re McLaughlin, 443 F.2d 1392, 170 USPQ 209 (CCPA 1971). The teachings of Gao et al., Naujok et al., and Laco et al. were all publicly available prior to the effective filing date of the claimed invention and would have provided a basis for modifying methods of pluripotent stem cell culture, such as that of Steiner et al.
Regarding the argument that the cited references do not teach or suggest the benefits provided by the claimed invention, it is noted that the features upon which applicant relies (i.e., balanced expansion, spheroidal morphology, and maintenance of pluripotency) are not recited in the rejected claims. Although the claims are interpreted in light of the specification, limitations from the specification are not read into the claims. See In re Van Geuns, 988 F.2d 1181, 26 USPQ2d 1057 (Fed. Cir. 1993). Additionally, motivation to modify the teachings of Steiner et al. need not be for the same purpose or to achieve the same results as the applicant. See MPEP 2144(IV).
The rejections are maintained.
Maintained Rejections
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.
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.
Claims 25, 30, 51, 53, 55, 58-61, and 69-75 are rejected under 35 U.S.C. 103 as being unpatentable over Steiner et al. (Nature Biotechnology, 2010), of record, in view of Gao et al. (Stem Cell Research, 2019), of record, Naujok et al. (BMC Research Notes, 2014), of record, and Laco et al. (Stem Cell Reports, 2018), of record, and evidenced by Watanabe et al. (Nature Biotechnology, 2007), of record, Garcia-Gonzalo et al., (PLoS One, 2008), of record, and Life Technologies (Product data sheet, 2014), of record.
Regarding claims 25, 30, 51, 55, 58-59, 61, 69-70, and 73-75: Steiner et al. teach the propagation of hESCs (which read on “pluripotent stem cells”) as aggregates in suspension culture in 12-well tissue culture dishes (which read on “culture vessel” and “multi-well plate”) (See Online Methods, page 1, col. 1, ¶2). The aggregates were cultured in Neurobasal medium (which reads on “cell culture basal medium”) with Knockout serum replacement (which reads on “serum-free”), FGF2 (which reads on “one or more mitogenic growth factors” and “bFGF”), BDNF (which reads on “at least two mitogenic growth factors”) and fibronectin and laminin (which read on “one or more extracellular matrix proteins”) (See Online Methods, page 1, col. 1, ¶2). A ROCK inhibitor (which reads on “inhibits ROCK activity and/or ROCK2 activity”)was present in the culture medium for 48 h following passaging (which reads on “contacting with the first medium is at a seeding step or a passaging step”) to reduce cell death (See page 363, col. 2, full ¶1). Steiner et al. do not teach removal of the ROCK inhibitor 24 h after seeding or the use of a GSK inhibitor.
Gao et al. teach that treatment of dissociated hESCs with the ROCK inhibitor Y-27632 is usually limited to 24 h in order to optimize its pro-survival effects (See page 7, col. 2, ¶1).
Naujok et al. teach that small molecule GSK3 inhibitors can be used to maintain pluripotency in ESCs (See Page 1, col. 1, ¶1). GSK3 inhibitors induced dose-dependent cytotoxicity in mouse ESC lines, with CHIR99021 able to maintain the best balance between Wnt activation/pluripotency marker expression and cell viability (See fig. 1-3).
Laco et al. teach that CHIR99021 has a cytotoxic effect on hPSC monolayer cultures and embryoid bodies, with a gradual loss of cell number in the embryoid bodies and in monolayer cell protein mass after 24 h exposure (See page 1855, col. 1, ¶1-2 and fig. 1).
It would have been obvious to one having ordinary skill in the art prior to the effective filing date of the claimed invention to modify the method of Steiner et al. to comprise limiting ROCK inhibitor treatment of the hESCs to 24 h. One would be motivated to make this modification because Gao et al. teach that 24 h exposure to a ROCK inhibitor is commonly performed for optimizing its effects (See page 7, col. 2, ¶1). There would be a reasonable expectation of success in making this modification because the post-passaging culture medium in the method of Steiner et al. could be readily replaced after 24 h with a medium lacking a ROCK inhibitor, which would read on “contacting the PSCs in the suspension culture with a second composition the day after the contacting with the first composition”.
It also would have been obvious to modify the method of Steiner et al. to comprise the short-term addition of a GSK3 inhibitor such as CHIR99021 to the culture medium. One would be motivated to make this modification because Naujok et al. teach that GSK3 inhibitors promote pluripotency (See page 1, col. 1, ¶1), though the results of Naujok et al. and Laco et al. suggest that exposure time be limited in order to reduce cytotoxicity. There would be a reasonable expectation of success in making this modification because a GSK3 inhibitor such as CHIR99021 could be readily added to the culture medium upon passaging and omitted upon changing of the medium at 24 h.
Regarding claim 53: Following the discussion of claims 25, 30, 51, 55, 58-59, 61, 69-70, and 73-75, Steiner et al. do not expressly teach the ROCK inhibitor used. However, Steiner et al. cite (See Online Methods, page 1, col. 1, ¶2) the work of Watanabe et al., who focus on the effects of the ROCK inhibitor Y-27632 on hESCs (See Abstract), which indicates Y-27632 was the inhibitor used by Steiner et al.
Regarding claim 60: Following the discussion of claims 25, 30, 51, 55, 58-59, 61, 69-70, and 73-75, Steiner et al. do not expressly teach the inclusion of albumin in the medium. Garcia-Gonzalo et al. teach that Knockout serum replacement comprises the lipid-rich albumin AlbuMAX (See table 1), which is bovine serum albumin (See Life Technologies, product data sheet).
Regarding claims 71-72: Following the discussion of claims 25, 30, 51, 55, 58-59, 61, 69-70, and 73-75, Steiner et al. teach that medium changes were performed by replacing 80% of the medium (which reads on “exchanging a portion of the first composition with the second composition” and “exchanging at least 25% of the suspension culture medium with an equivalent amount of fresh second composition”) (See Online Methods, page 1, col. 1, ¶2).
Claims 25, 30, 51, 53, 55, 58-61, 66, and 69-76 are rejected under 35 U.S.C. 103 as being unpatentable over Steiner et al. (Nature Biotechnology, 2010), of record, in view of Gao et al. (Stem Cell Research, 2019), of record, Naujok et al. (BMC Research Notes, 2014), of record, and Laco et al. (Stem Cell Reports, 2018), of record, and evidenced by Watanabe et al. (Nature Biotechnology, 2007), of record, Garcia-Gonzalo et al., (PLoS One, 2008), of record, and Life Technologies (Product data sheet, 2014), of record, further in view of Ito et al. (WO 2020022511 A1, machine translation), of record.
The teachings of Steiner et al., Gao et al., Naujok et al., Laco et al., Watanabe et al., Garcia-Gonzalo et al., and Life Technologies are set forth in the rejection above and are incorporated herein in their entirety.
Regarding claim 66: Following the discussion of claims 25, 30, 51, 53, 55, 58-61, and 69-75, Steiner et al., modified by Gao et al., Naujok et al., and Laco et al., and evidenced by Watanabe et al., Garcia-Gonzalo et al., and Life Technologies, render obvious a method for growing PSCs in suspension but do not teach the agitation of the suspension culture.
Ito et al. teach that stirring or shaking (which read on “agitating”) can be used to efficiently obtain iPSC aggregates having a controlled size (See ¶0034, 0038, 0041, and 0047).
It would have been obvious to one having ordinary skill in the art prior to the effective filing date of the claimed invention to modify the method of Steiner et al., modified by Gao et al., Naujok et al., and Laco et al., and evidenced by Watanabe et al., Garcia-Gonzalo et al., and Life Technologies, to comprise stirring or shaking of the suspension culture, as taught by Ito et al. One would be motivated to make this modification because Ito et al. teach that the size of the stem cell aggregates can be controlled by agitation (See ¶0034 and 0041). There would be a reasonable expectation of success in doing so because the cultures of Steiner et al., modified by Gao et al., Naujok et al., and Laco et al., and evidenced by Watanabe et al., Garcia-Gonzalo et al., and Life Technologies, could be readily shaken or stirred.
Regarding claim 76: Following the discussion of claims 25, 30, 51, 53, 55, 58-61, and 69-75, Steiner et al., modified by Gao et al., Naujok et al., and Laco et al., and evidenced by Watanabe et al., Garcia-Gonzalo et al., and Life Technologies, render obvious a method for growing PSCs in suspension but do not teach the inclusion of a surfactant in the culture media.
Ito et al. teach the use of water-soluble polymers having surface activity (which read on “surfactants”) for preventing precipitation of serum-free medium components in the suspension culture of cells such as pluripotent stem cells (See ¶0004, 0006, 0022, 0034, and 0041). The one or more polymers can be nonionic surfactants (See ¶0011-0013).
It would have been obvious to one having ordinary skill in the art prior to the effective filing date of the claimed invention to modify the culture medium in the method of Steiner et al., modified by Gao et al., Naujok et al., and Laco et al., and evidenced by Watanabe et al., Garcia-Gonzalo et al., and Life Technologies, to comprise at least one surfactant, such as those taught by Ito et al. One would be motivated to make this modification because Ito et al. teach that a surfactant can prevent precipitation of serum-free media components, such as insulin, thereby increasing the efficiency of suspension culture (See ¶0034 and 0041). There would be a reasonable expectation of success in doing so because a surfactant could be readily added to the serum-free medium of Steiner et al., modified by Gao et al., Naujok et al., and Laco et al., and evidenced by Watanabe et al., Garcia-Gonzalo et al., and Life Technologies.
Conclusion
THIS ACTION IS MADE FINAL. 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 JENNIFER S SPENCE, whose telephone number is 571-272-8590. The examiner can normally be reached M-F 8:30-5:30.
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If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Christopher M Babic, can be reached at 571-272-8507. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
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/J.S.S./Examiner, Art Unit 1633
/CHRISTOPHER M BABIC/Supervisory Patent Examiner, Art Unit 1633
