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 .
DETAILED ACTION
Amendments
In the reply filed 01/05/2026, Applicant has amended claims 1, 9 and 13, and newly canceled claims 2, 7-8, 29-30, 48-51 and 53.
Claim Status
Claims 1, 3-6 and 9-13 are pending and are considered on the merits. Claims 1 and 9 are independent claims.
Withdrawn Claim Rejections - 35 USC § 112
The prior rejection of claims under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for being ambiguous has been withdrawn in light of Applicant’s amendment to claims 1 and 13.
New 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.
Claims 1, 3-6 and 9-13 are rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
Independent claims 1 and 9, step (e), both recite the new limitation “karyotypically stable”. Applicant is reminded that a claim may be rendered indefinite by reference to term of an object that is variable (see MPEP 2173.05(b), II). Specifically, the term “stable” is a relative term which renders the claims indefinite. The term “stable” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. In the instant case, as Applicant has argued in the Remarks, page 6-7, that “Applicants used a GMP-grade iPSC media that allows iPSC expansion without accumulation of KT (i.e., karyotype) abnormalities called "StemFit" in combination with coating the dishes with laminin, which performed the best in cell survival and growth and karyotype stability for at least 10 passages.” (also see specification [0099]), this limitation is being examined as the pluripotent cells are cultured in feeder-free, xeno-free StemFit medium on laminin-coated plates so as to generate karyotypically stable colonies of iPSCs. Claims 3-6 and 11-13 are rejected as being dependent from claim 1, and claim 10 is rejected as being dependent from claim 9, without resolving the ambiguity.
Furthermore, Claim 9 (e) recites “stem-fit medium”, in which “stem-fit” (i.e., “StemFit”) is a trademark. 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 is used to identify/describe a cell growth medium for use in expansion or differentiation of pluripotent stem cell, and accordingly, the identification/description is indefinite.
Withdrawn Claim Rejections - 35 USC § 103
The prior rejection of claims 1, 3-6 and 9-13 set forth in the prior Office action mailed on 09/05/2025 has been withdrawn in light of Applicant’s amendment to claim 1 and claim 9 to recite the new limitation “karyotypically stable” iPSCs. As discussed above, based on Applicant’s argument and disclosure, this limitation is being examined as the pluripotent cells are cultured in feeder-free, xeno-free StemFit medium on laminin-coated plates so as to generate karyotypically stable colonies of iPSCs, which is not discussed in the prior rejection.
New Claim Rejections - 35 USC § 103
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
Claims 1, 3-6 and 11-13 are rejected under 35 U.S.C. 103 as being unpatentable over Kogut et al., (US PGPub 2018/0346933. Cited in IDS 06/16/2022) in view of Hempstead, (“CRISPR 101: Ribonucleoprotein (RNP) Delivery”, published Sep 6, 2018. Downloaded from “https://blog.addgene.org/crispr-101-ribonucleoprotein-rnp-delivery”. Downloaded on 9/2/2025. P. 1-8. Prior art of record) and Nakagawa et al., (Sci Rep. 2014:4:3594. P. 1-7).
With respect to claim 1, Kogut teaches a method of generating a gene-corrected, patient-specific induced pluripotent stem cell (iPSC) (e.g., [0017]) and teaches “a gene-corrected iPSC is differentiated into a keratinocyte (epidermal stem cell) that is then grown as an epidermal sheet and administered/transplanted to a patient” ([0018], right col, also see [0216]-[0217]). It is noted that the “keratinocyte (epidermal stem cell)” of Kogut is equivalent to the claimed keratinocyte stem cell (iKC). Kogut further teaches a method of treating a condition in a patient in need thereof by administering to the patient a differentiated somatic cell derived from a gene-corrected iPSC (e.g., [0018]), thus teaches the preamble of claim 1.
In regard to (a) obtaining somatic cells from a patient, Kogut teaches combined cellular reprogramming and gene correction of Recessive Dystrophic Epidermolysis Bullosa (RDEB) fibroblasts in Example 14 ([0202]-[0207]), thus teaches obtaining a population of somatic cells (i.e., fibroblasts, see [0204]) from an individual having a mutation at a target locus of interest for genetic correction (i.e., COL7A1, see [0202]).
In regard to (b) contacting cells with Cas9/gRNA RNP complex and a repair sequence, Kogut teaches Cas9+gRNA is delivered together with a correcting single-strand DNA donor (ssODN) that shows robust modification of the COL7A1 locus in cells ([0204] and [0202]). It is noted that Kogut teaches “a single strand DNA oligo repair template was introduced together with Cas9 nuclease” ([0202]) and claims “administering to a cell a Cas9 and a short guide RNA (gRNA)” (see reference claim 17, which encompasses a Cas9 protein), thus the scope of Kogut encompasses a preformed RNP complex comprising a guide RNA specific for the locus of interest and a CRISPR/cas9 protein.
In regard to choosing an RNP complex, prior art Hempstead teaches there are many advantages of using RNPs for CRISPR experiments such as 1) RNP method can often be used in cells that are difficult to transfect, such as primary cells (it is noted that Kogut’s fibroblasts from patients are primary cells), 2) using RNPs can also alleviate difficulties with protein expression that occur in cells, 3) using RNPs may limit the potential for off-target effects, 4) Cas9 RNPs are detectable at high levels shortly after transfection, and are quickly cleared from the cell via protein degradation pathways (end of p. 1-beginning of p. 2). Accordingly, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to have chosen an Cas9/gRNA RNP complex in Kogut’s method of genetically correcting fibroblasts from patients as suggested by Hempstead with a reasonable expectation of success. Since Hempstead teaches many advantages of using Cas9/gRNA RNP complex in CRISPR-mediated gene editing (see above), one of ordinary skill in the art would have had a reason to choose RNP in the method of Kogut in order to take advantages of the RNP complex in genetically correcting the diseased cells.
In regard to (c) maintaining the cells in feeder-free, xeno-free medium for 2 days, Kogut teaches the RDEB fibroblasts are delivered by Cas9+gRNA and a correcting donor repair template and are cultured for 48 hours (i.e., 2 days, [0204], also see diagram in Fig 19, Fig 25 and [0200]). Regarding the medium, Kogut teaches the culture is feeder-free (see e.g., [0207]) and teaches the fibroblasts are cultured in either human dermal fibroblast medium (HDF medium) or medium containing KNOCKOUT serum replacement (KOSR medium, [0145], and see the defined KOSR medium components in [0146] that is serum-free). Thus, Kogut teaches the cells are maintained in feeder-free, serum-free medium for 2 days.
In regard to (d) contacting the cells of (c) with synthetic capped mRNAs containing modified nucleobases for M3O, Sox2, Klf4, cMyc and Lin28A to induce pluripotency, Kogut teaches after the gene editing, the cells are subjected to a reprogramming protocol ([0204]) which comprises transfecting the fibroblasts with modified mRNA (mod-mRNA) mix of reprogramming factors that are synthesized using ribonucleoside mix including 3’-0-Me-m7G (5')ppp(5')G ARCA cap analog (i.e., synthetic capped mRNAs), and 5-methylcystidine triphosphate and pseudouridine triphosphate (i.e., modified nucleobases) ([0142]) and teaches the reprogramming factors comprise Myo-D-OCT4 (M3O), SOX2, KLF4, c-MYC and LIN28A (see e.g., [0141]).
In regard to (e) expanding the pluripotent cells in feeder-free, xeno-free medium to generate karyotypically stable colonies of iPSC, and (f) differentiating the iPSC in feeder-free, xeno-free medium to iKC, as discussed above, based on Applicant’s argument and disclosure, the limitation of “karyotypically stable” is being examined as the pluripotent cells are cultured in feeder-free, xeno-free StemFit medium on laminin-coated plates so as to generate karyotypically stable colonies of iPSCs.
Kogut teaches 49 iPSC colonies are picked from reprogrammed cells ([0206]). Kogut teaches the culture is feeder-free (see e.g., [0207]) and teaches the transfected cells are cultured in KOSR Medium supplemented with bFGF ([0153] and see KOSR medium components in [0146] that is serum-free). Kogut teaches the gene-corrected iPSC is differentiated into a keratinocyte (epidermal stem cell) (i.e., iKC, [0018], right col, also see [0216]-[0217]). Thus, Kogut teaches the pluripotent cells are expanded and differentiated in feeder-free, serum-free medium to generate iPSC colonies and to differentiate into iKC. Kogut teaches “All iPSC lines exhibited normal karyotypes and were successfully maintained for at least 15 passages” (e.g., [0217]).
However, Kogut does not specifically teach the medium is xeno-free for maintaining the genetically corrected cells in (c), nor teach the pluripotent cells are cultured in feeder-free, xeno-free StemFit medium on laminin-coated plates so as to generate karyotypically stable colonies of iPSCs in (e) and differentiating the iPSC in feeder-free, xeno-free medium in (f).
Nakagawa teaches a method of generating human iPSCs under feeder-free (Ff) and xeno-free (Xf) culture systems from human primary fibroblasts (the same cell type used in Kogut) and blood cells, that conforms to GMP guidelines (see e.g., abstract). Nakagawa teaches human primary fibroblasts are established under Xf conditions with medium (p. 2, right col, para 1 “Establishment of hiPSCs under Ff and Xf conditions”), thus teaches xeno-free medium for maintaining the fibroblasts, related to (c). Nakagawa teaches the hESCs/iPSCs are cultivated on rLN511E8-coated (i.e., recombinant laminin-511 E8 fragment) cell culture plates with StemFit (p. 6, left col, para “Feeder-free culture system for hESCs and hiPSCs” and p. 2, left col) and are differentiated into all three germ layers by cultivated on rLN511E8 with StemFit (p. 2, right col, para “Differentiation capacity of Ff-hiPSCs”). Thus, Nakagawa teaches the pluripotent cells are cultured in feeder-free, xeno-free StemFit medium on laminin-coated plates so as to generate karyotypically stable colonies of iPSCs in (e) and differentiating the iPSC in feeder-free, xeno-free medium in (f). Nakagawa teaches the hES/iPSCs can be stably cultivated over long periods using the new culture conditions. e.g., cells are stably maintained for 20-30 passages and up to passage number 54 (p. 2, left col, para “Development of a novel culture system for hiPSCs”) and have normal karyotypes (Figure S3C) (e.g., abstract and p. 2, right col, para 2).
Therefore, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of generating genetically corrected iKC from fibroblast-derived iPSC cultured in a feeder-free condition for regenerative therapy suggested by Kogut in view of Hempstead, by substituting with maintaining the fibroblasts in xeno-free medium and culturing the pluripotent cells in feeder-free, xeno-free StemFit medium on laminin-coated plates so as to generate karyotypically stable colonies of iPSCs and further differentiating the iPSC in feeder-free, xeno-free medium as suggested by Nakagawa with a reasonable expectation of success. Since Kogut teaches the reprogrammed cell is current Good Manufacturing Practice (cGMP) compatible (see e.g., [0026]), and since Nakagawa teaches a method of generating human iPSCs under feeder-free (Ff) and xeno-free (Xf) culture systems from human primary fibroblasts that conforms to GMP guidelines (see e.g., abstract) and the hiPSCs can be stably cultivated over long periods using the new culture conditions. e.g., cells are stably maintained for 20-30 passages and up to passage number 54 (p. 2, left col, para “Development of a novel culture system for hiPSCs”) and have normal karyotypes (Figure S3C) (e.g., abstract and p. 2, right col, para 2), one of ordinary skill in the art would have had a reason to substitute with the feeder-free (Ff) and xeno-free (Xf) culture system of Nakagawa in the method of Kogut in order to take advantage of the long-term maintenance of iPSCs with normal karyotypes as suggested by Nakagawa.
With respect to claim 3 directed to the somatic cells being fibroblasts, as stated supra, Kogut teaches reprogramming and gene correction of Recessive Dystrophic Epidermolysis Bullosa (RDEB) fibroblasts (Example 14, [0202]-[0207]).
With respect to claim 4 directed to the somatic cells being autologous relative to an individual selected for treatment, Kogut teaches generating COL7A1-corrected autologous iPSCs for clinical applications ([0207], and [0217], last 3 lines).
With respect to claim 5 directed to the iPSC being expanded in culture, Kogut teaches the resulting iPSC colonies are clonally expanded (see e.g., [0202]), and teaches all iPSC lines are successfully maintained for at least 15 passages ([0217], right col). Thus, Kogut teaches the iPSCs are expanded in culture.
With respect to claim 6 directed to the iPSCs being frozen prior to expansion, although Kogut does not specifically teach the iPSCs are frozen, freezing cells for future use is a common technique used in cell manufacturing and culture as evidenced by Nakagawa who teaches frozen stocks of iPSCs are prepared at -80°C using a standard slow-freezing method, and are thawed in a 37°C water bath (Figure S2A) (e.g., p. 2, left col, section “Development of a novel culture system for hiPSCs”, para 3). Accordingly, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to have combined a step of freezing iPSCs prior to expansion as taught by Nakagawa in the method of Kogut in view of Hempstead and Nakagawa with a reasonable expectation of success. One of ordinary skill in the art would have had a reason to do so in order to cryopreserve the iPSCs for future use in the clinical applications.
With respect to claim 11 and claim 13 directed to the somatic cells being obtained from an individual with Epidermolysis Bullosa (EB) and claim 12 directed to the locus of interest for genetic correction being COL7A1, as stated supra, Kogut teaches fibroblasts are obtained from Recessive Dystrophic Epidermolysis Bullosa (RDEB) patients in Example 14 ([0202] and [0204]) who have a homozygous mutation in COL7A1 ([0202]).
Hence, the claimed invention as a whole was prima facie obvious to a person of ordinary skill before the effective filing date of the claimed invention in the absence of evidence to the contrary.
Response to Traversal:
Applicant’s arguments filed on 01/05/2026 are acknowledged.
Applicant argues that the claimed method has technical improvements, while the gene correction methods of Kogut lack the advantages of the present claims (Remarks p. 6-8).
Applicant’s arguments have been fully considered but they are not persuasive. The response to Applicant’s arguments are detailed below.
In response to the technical improvements a) Development and use of a feeder-free, xeno-free, viral-free method of iPS cell reprogramming, Kogut teaches a feeder-free iPSC reprogramming (e.g., [0004]) and teaches gene correction is performed using viral - and footprint - free CRISPR/Cas9 mediated targeting ([0216]). Thus, Kogut teaches a feeder-free, viral-free method of iPS cell reprogramming. Prior art Nakagawa is cited, as necessitated by amendment, to teach a xeno-free method (using laminin 511-coated plates with StemFit media) for maintaining the fibroblasts and for the derivation and continued culture of iPSCs.
In response to the technical improvements b) Development and use of an RNA-based integration-free, vector-free, single clonal step for correction of the genetic defect, Kogut teaches combined gene editing and reprogramming utilizing solely modified mRNA ([0215]), the invention includes iPSCs generated with a highly efficient , integration-free, RNA-based reprogramming approach, gene correction is performed using viral- and footprint-free CRISPR/Cas9 mediated targeting, and reprogramming and gene editing are combined into a one-step procedure ([0216]).
In response to the technical improvements c) Development and use of a feeder-free, xeno-free defined and reproducible method of iKC differentiation from ES/iPSCs, Kogut teaches “gene corrected iPSC is differentiated into a keratinocyte (epidermal stem cell) that is then grown as an epidermal sheet and administered/transplanted to a patient” ([0018]). It is noted that the “keratinocyte (epidermal stem cell)” is equivalent to the claimed iKC. As stated supra, Kogut teaches a feeder-free iPSC reprogramming and differentiation (e.g., abstract) and Nakagawa teaches a xeno-free method for differentiating the iPSCs (see rejection of claim 1 above).
In response to the technical improvements d) identification and use of ITGA6 (CD49f) as a biomarker for the purification of iKCs, prior art Yu (see the next section of rejection of claims 9-10) teaches a method of generating keratinocytes from pluripotent stem cells and generating a transplantable sheet of skin epithelial cells comprising keratinocyte stem cells (iKCs, abstract, [0018] and reference claim 35). Yu teaches iKCs express epidermal keratinocyte stem cell markers cytokeratin 14 and CD49f ([0036]). Yu teaches to assess the purity of iKCs, keratinocytes are harvested, stained with anti-K14-FITC and anti-CD49f-PE antibodies and assayed by flow cytometry that shows 90.2%-91.8% K14+/CD49f+ high purity iKC cells (Example 1, [0141]). Yu teaches the iKCs have enhanced proliferative potential of >10 population doublings ([0036]). Thus, Yu teaches ITGA6 (CD49f) is a biomarker of iKCs and reduces to practice a flow cytometry approach that can be used to sort CD49f+ iKCs having proliferative potential.
In response to the technical advances significantly shorten the manufacturing time and eliminate multiple subcloning steps (Remarks, p. 6), Kogut teaches combined gene editing and reprogramming utilizing solely modified mRNAs yields gene-corrected iPSCs in as little as 3 weeks. This is at least 4 weeks faster than any currently published combined protocol, and 6-12 weeks faster than any potential 2-step approach (reprogramming followed by gene editing) ([0215]).
In response to the argument that Applicants have optimized reprogramming using synthetic capped mRNAs containing modified nucleobases (modified mRNA), Kogut teaches the reprogramming using modified mRNA (mod-mRNA) mix of reprogramming factors that are synthesized using 3’-0-Me-m7G (5')ppp(5')G ARCA cap analog (i.e., synthetic capped mRNAs), and 5-methylcystidine triphosphate and pseudouridine triphosphate (i.e., modified nucleobases) ([0142]).
In response to the argument that Applicants use incorporation of a modified version of Oct4 fused with the MyoD transactivation domain (called M3O), Sox2, Klf4, cMyc, and Lin28A, combined with a feeder-free system, Kogut teaches a feeder-free system (e.g., abstract) and teaches the reprogramming factors comprise Myo-D-OCT4 (M3O), SOX2, KLF4, c-MYC and LIN28A (see e.g., [0141]).
In response to the argument that this mRNA reprogramming method is optimized by using xeno-free media and synthesized mRNA, and provided for robust iPS cell colony formation in all patient fibroblast lines tested so far, as stated supra, Kogut teaches synthesized mRNA-based system and Nakagawa teaches a xeno-free system (laminin-511 coating with StemFit media), thus make obvious the claimed method.
In response to the argument that Applicants used a GMP-grade iPSC media that allows iPSC expansion without accumulation of KT abnormalities called "StemFit" in combination with coating the dishes with laminin, which performed the best in cell survival and growth and karyotype stability for at least 10 passages, as stated supra, Nakagawa teaches a method conforming to GMP guidelines (abstract), comprising cultivating the hESCs/iPSCs on rLN511E8-coated (i.e., recombinant laminin-511 E8 fragment) cell culture plates with StemFit (p. 6, left col, para “Feeder-free culture system for hESCs and hiPSCs” and p. 2, left col) and teaches the hES/iPSCs can be stably cultivated over long periods, e.g., for 20-30 passages and up to passage number 54 (p. 2, left col) and have normal karyotypes (e.g., abstract and p. 2, right col, para 2).
In response to the argument that Applicants have also developed a defined and reproducible method for generating iKC by the addition of retinoic acid (RA) and bone morphogenetic protein (BMP) in the medium, Kogut teaches the iPSCs are differentiated into cells from an ectodermal lineage and then to keratinocytes using BMP4 and retinoic acid treatment ([0187]).
In response to the argument that a further improvement is provided the correlation of ITGA6 expression with epidermal stem cells and graftability allowing purification of iKCs, Applicant is referred to the above response to the technical improvements d) identification and use of ITGA6 (CD49f) as a biomarker for the purification of iKCs.
In response to the argument that the gene correction methods of Kogut use a DNA-based approach (par. 202) "In this strategy, a single strand DNA oligo repair template was introduced together with Cas9 nuclease", Applicant is reminded that this DNA sequence of Kogut is a repair template for correcting the diseased gene. In fact, the instant specification recites the same type of DNA sequence, as “A DNA repair template containing the desired sequence is delivered into the cell type of interest with the gRNA(s) and Cas9” (see instant specification [0035]).
In response to the argument that Kogut further fails to teach the continued use of feeder-free, xeno-free medium throughout the process, and the medium used by Kogut does not support KT stability in iPSCs over long culture periods, Applicant is reminded, one cannot show nonobviousness 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). In the instant case, Kogut teaches a feeder-free, serum-free medium throughout the process and “All iPSC lines exhibited normal karyotypes and were successfully maintained for at least 15 passages” (e.g., [0217]), indicating the medium used by Kogut does support karyotype stability in iPSCs over long culture periods. However, based on Applicant’s argument and disclosure, the limitation of “karyotypically stable” is being examined as the pluripotent cells are cultured in feeder-free, xeno-free StemFit medium on laminin-coated plates so as to generate karyotypically stable colonies of iPSCs. Prior art Nakagawa, as necessitated by amendment, teaches a method of generating human iPSCs under feeder-free (Ff) and xeno-free (Xf) StemFit medium on laminin-511-coated plates and teaches the StemFit medium and the laminin-coating support KT stability in iPSCs over long culture periods up to 54 passages (see rejection of claim 1 above). Thus, Kogut in view of Nakagawa make obvious the claimed method.
Claims 9-10 are rejected under 35 U.S.C. 103 as being unpatentable over Kogut et al., (US PGPub 2018/0346933. Cited in IDS 06/16/2022) in view of Hempstead, (“CRISPR 101: Ribonucleoprotein (RNP) Delivery”, published Sep 6, 2018. Downloaded from “https://blog.addgene.org/crispr-101-ribonucleoprotein-rnp-delivery”. Downloaded on 9/2/2025. P. 1-8. Prior art of record), Nakagawa et al., (Sci Rep. 2014:4:3594. P. 1-7) and Yu et al., (WO 2016/061071, prior art of record).
With respect to independent claim 9, it is noted that the preamble and the steps of (a) through the first half of (f) are directed to similar subject matters as those in claim 1, thus are rejected in the same way as to claim 1 above. The rejection is omitted for the sake of concise prosecution.
In regard to step (f) differentiating comprising an effective dose of retinoic acid (RA) and BMP, Kogut teaches the iPSCs are differentiated into cells from an ectodermal lineage and then to keratinocytes using BMP4 and retinoic acid treatment ([0187]).
In regard to step (g) selecting the iKC for expression of CD49f, Kogut teaches the iPSCs are differentiated into cells from an ectodermal lineage and then to keratinocytes that are positive for keratin 14 (K14+) (see Example 10, [0186-0187]). Kogut teaches “a gene-corrected iPSC is differentiated into a keratinocyte (epidermal stem cell) that is then grown as an epidermal sheet and administered/transplanted to a patient” ([0018], right col, also see [0216]-[0217]).
However, Kogut, Hempstead and Nakagawa are silent on selecting the iKCs for expression of CD49f in claim 9 (g).
Yu teaches a method of generating keratinocytes from pluripotent stem cells and generating a transplantable sheet of skin epithelial cells comprising keratinocyte stem cells (iKCs, see abstract, [0018] and reference claim 35). Yu teaches iKCs have enhanced proliferative potential of >10 population doublings and express epidermal keratinocyte stem cell markers cytokeratin 14 and CD49f ([0036]). Yu teaches to assess the purity of iKCs, keratinocytes are harvested, stained with anti-K14-FITC and anti-CD49f-PE antibodies and assayed by flow cytometry that shows 90.2% - 91.8% K14+/CD49f+ high purity iKCs (Example 1, [0141]). Thus, Yu teaches CD49f is an iKC biomarker and can be used to identify the purity of iKCs. Yu also reduces to practice a method of flow cytometry approach that can be used to sort the CD49f+ iKCs.
Therefore, it would have been obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of selecting induced keratinocyte stem cells (iKCs) expressing keratin 14 (K14+) for manufacturing an epidermal sheet for engraftment suggested by Kogut in view of Hempstead and Nakagawa, by combining another keratinocyte stem cell marker CD49f in selection as suggested by Yu with a reasonable expectation of success. Since Kogut aims to select induced keratinocyte stem cells for manufacturing an epidermal sheet for engraftment ([0018], right col, also see [0216]-[0217]), and since Yu reduces to practice a method of flow cytometry approach that can be used to sort the CD49f+ iKCs and teaches keratinocyte stem cells expressing K14 and CD49f have enhanced proliferative potential of >10 population doublings and can be used to generate a transplantable sheet of skin epithelial cells (see e.g., [0018], [0036] and Example 1, [0141]), one of ordinary skill in the art would have had a reason to combine another marker CD49f besides K14 in selecting iKCs as suggested by Yu in order to obtain purified iKCs with enhanced proliferative potential to generate a transplantable sheet.
With respect to claim 10 directed to the CD49f+ iKCs being manufactured as a sheet for engraftment, as stated supra, Kogut teaches the iKCs are grown as an epidermal sheet and administered/transplanted to a patient ([0018]), and Yu teaches CD49f is an iKC biomarker ([0036]) and the CD49f+ iKCs are used to generate a transplantable sheet (see abstract, [0018] and reference claim 35).
Hence, the claimed invention as a whole was prima facie obvious to a person of ordinary skill before the effective filing date of the claimed invention in the absence of evidence to the contrary.
Response to Traversal:
Applicant’s arguments filed on 01/05/2026 are acknowledged.
Applicant argues that although Yu teaches that iKC cells express CD49, there is no teaching for the sorting of cells, while Applicants identify ITGA6 bright cells displayed enhanced graftability (Remarks, p. 8-9).
Applicant’s arguments have been fully considered but they are not persuasive.
As stated supra, Yu teaches iKCs express epidermal keratinocyte stem cell markers cytokeratin 14 and CD49f, and identifies that high purity K14+/CD49f+ iKCs have enhanced proliferative potential of >10 population doublings ([0036]). Yu also reduces to practice a method of flow cytometry approach to identify the purity of K14+/CD49f+ iKCs, that can be used to sort the CD49f+ iKCs. Thus, Yu provides both motivation (i.e., high purity K14+/CD49f+ iKCs have enhanced proliferative potential) and a reasonable expectation of success (i.e., a flow cytometry approach that can be used to sort the CD49f+ iKCs) to select the iKCs for expression of CD49f. Accordingly, it would have been obvious for one of ordinary skill in the art to have combined CD49f as a biomarker for selecting the iKC as suggested by Yu in order to obtain high purity CD49f positive iKCs to take advantage of their enhanced proliferative potential for manufacturing a transplantable sheet.
In response to the argument that Applicants identify ITGA6 bright cells displayed enhanced graftability, MPEP 2145 (II) states that “mere recognition of latent properties in the prior art does not render nonobvious an otherwise known invention”, and “the fact that appellant has recognized another advantage which would flow naturally from following the suggestion of the prior art cannot be the basis for patentability when the differences would otherwise be obvious.” Ex parte Obiaya, 227 USPQ 58, 60 (Bd. Pat. App. & Inter. 1985)”. In instant case, the prior art Yu teaches the high purity K14+/CD49f+ iKCs have enhanced proliferative potential ([0036]). Thus, the fact that applicant has recognized another advantage (i.e., enhanced graftability) cannot be the basis for patentability when the differences would be obvious over Kogut in view of Yu.
Conclusion
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 extension fee 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 date of this final action.
No claims are allowed.
Examiner Contact Information
Any inquiry concerning this communication or earlier communications from the examiner should be directed to Jianjian Zhu whose telephone number is (571)272-0956. The examiner can normally be reached M - F 8:30AM - 4PM (EST).
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, James Douglas (Doug) Schultz can be reached on (571) 272-0763. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
Information regarding the status of published or unpublished applications may be obtained from Patent Center. Unpublished application information in Patent Center is available to registered users. To file and manage patent submissions in Patent Center, visit: https://patentcenter.uspto.gov. Visit https://www.uspto.gov/patents/apply/patent-center for more information about Patent Center and https://www.uspto.gov/patents/docx for information about filing in DOCX format. For additional questions, contact the Electronic Business Center (EBC) at 866-217-9197 (toll-free). If you would like assistance from a USPTO Customer Service Representative, call 800-786-9199 (IN USA OR CANADA) or 571-272-1000.
/JIANJIAN ZHU/Examiner, Art Unit 1631
/JAMES D SCHULTZ/Supervisory Patent Examiner, Art Unit 1631
Prosecution Insights