DETAILED ACTION
Applicant’s response of 06/27/2025 has been received and entered into the application file.
Notice of Pre-AIA or AIA Status
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Election/Restrictions
Claims 1-8 and 16-27 are pending in the instant application.
Applicants have previously elected with traverse of species (ii) contacting the biological cells with a cell cycle inhibiting agent from species Group A, species (iv) kip pathway from species Group B, and species (xvi) cellular marker for proliferation from species Group C, in the reply filed on 01/24/2025.
Claims 3-5, 17-20, and 22-27 are still withdrawn from further consideration pursuant to 37 CFR 1.142(b), as being drawn to a nonelected species, there being no allowable generic or linking claim. Applicant timely traversed the restriction (election) requirement in the reply filed on 01/24/2025. Therefore, claims 1-2, 6-8, 16, and 21 are under examination in the instant application.
Status of Prior Rejections/Response to Arguments
RE: Rejection of claim 7 under 35 U.S.C. 112(d) or pre-AIA 35 U.S.C. 112, 4th paragraph:
Applicants’ amendments filed on 06/27/2025 overcome the rejection of claim 7. Applicants have
specifically amended claim 7 to remove p14ARF, P16INK4a, p18, p19, and p53, which are modulators of different pathways other than the elected species kinase inhibitory protein (kip) pathway and as recited in claim 6, from which claim 7 depends. The rejection is therefore withdrawn.
RE: Rejection of claims 1, 6-8, and 16 under 35 U.S.C. 103 as being unpatentable over Prieur et al., in view of Donovan et al.:
Applicants have traversed the rejection asserting that Donovan does not teach or suggest "contacting the biological cells with a reversible cell cycle inhibiting agent to arrest the cells in a first cell-cycle phase selected from Gl/S or G2, and thereafter removing the agent; (b) following step (a), selecting, from the biological cells, a subset of cells determined to be in mitotic (M) phase, thereby obtaining a subset of cells that are enriched in cells at the same mitotic phase," as recited by Applicant's Amended Independent Claim 1. Applicants specifically argues that Donovan's approach of arresting cells in G1 by continuous TGF-(3 exposure is merely a single-step, same-phase growth-inhibition experiment, rather than a two-step synchronization strategy that (i) uses a reversible block at Gl/S or G2, (ii) removes the blocker, and (iii) harvests the cells only after they have progressed into and accumulated in M phase.
Applicants’ arguments and amendments filed on 06/27/2025 have been fully considered and are persuasive. Applicants have specifically amended claim 1 to recite contacting the biological cells with a reversible cell cycle inhibiting agent to arrest the cells in a first cell-cycle phase selected from Gl/S or G2, and thereafter removing the agent; (b) following step (a), selecting, from the biological cells, a subset of cells determined to be in mitotic (M) phase, thereby obtaining a subset of cells that are enriched in cells at the same mitotic phase.
Therefore, the rejection has been withdrawn. However, upon further consideration, a new ground(s) of rejection is made in view of Silberstein et al.
RE: Rejection of claims 1-2, 6-8, and 16 under 35 U.S.C. 103 as being unpatentable over Prieur et al., in view of Donovan et al. as applied to claims 1, 6-8, and 16, and further in view of Silberstein et al.:
Applicants have traversed the rejection asserting that Silberstein does not teach or suggest "contacting the target cell population with the cell cycle inhibiting agent (TGF beta) to arrest the cell cycle at a first cell cycle phase and then removing the inhibiting agent until the specific desired cell cycle phase is determined to have been reached such that the selected subset of cells is in a second cell cycle phase.
Applicant's arguments have been fully considered but they are not persuasive. The rejection has been maintained.
RE: Rejection of claims 1, 6-8, 16, and 21 under 35 U.S.C. 103 as being unpatentable over Prieur et al., in view of Donovan et al. as applied to claims 1, 6-8, and 16, and further in view of Whitfield et al.:
Applicants have traversed the rejection asserting that Whitfield does not teach or suggest "contacting the biological cells with a reversible cell cycle inhibiting agent to arrest the cells in a first cell-cycle phase selected from Gl/S or G2, and thereafter removing the agent; (b) following step (a), selecting, from the biological cells, a subset of cells determined to be in mitotic (M) phase, thereby obtaining a subset of cells that are enriched in cells at the same mitotic phase,".
Applicant's arguments have been fully considered but they are not persuasive. Whitfield reference was actually only used to teach the limitation of instant dependent claim 21 of contacting the cells with a labeled agent that binds to or comprises a cellular marker for proliferation. However, in view of the amendments of instant claim 1, the rejection has been withdrawn and a new ground(s) of rejection is made in view of Silberstein et al.
New/Maintained Grounds of Rejection
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.
Claims 1-2, 6-8, and 16 are rejected under 35 U.S.C. 103 as being unpatentable over Prieur et al. (US20140056860A1, filed on 04/10/2012, and published on 02/27/2014), in view of Donovan et al. (Donovan et al., “Transforming growth factor-β and breast cancer: Cell cycle arrest by transforming growth factor-β and its disruption in cancer”. Breast Cancer Res. 2000;2(2):116-24) and Silberstein et al. (Silberstein et al., “Reversible Inhibition of Mammary Gland Growth by Transforming Growth Factor-β”. Science. 1987; 237(4812): 291-293).
Regarding claim 1, Prieur et al. teaches a method for preparing induced pluripotent stem cells (iPSCs) from a target cell population comprising senescent cells (Abstract). Target cell population reads on a plurality of biological cells. Prieur et al. further teaches that “senescent cells” refers to cells that exhibit cell cycle arrest, generally during the G1 transition of the cell cycle (paragraph 0055). Prieur et al. teaches that serial passaging was used in order to induce replicative senescence and make the senescent cells, wherein these senescent cells were maintained more than 2 months in culture without any detectable increase in the cell number, confirming the robustness of cell cycle arrest (paragraph 0191). Prieur et al. also teaches a subset of cells exhibiting up-regulation of the tumor suppressors p16INK4a and p21CIP1 arrested in G1 phase (paragraph 0055). This reads on step (b) of the instant method of selecting, from the biological cells, a subset of cells determined to be at a specific cell cycle phase, thereby obtaining a subset of cells that are enriched in cells at the same specific desired cell cycle phase. Additionally, Prieur et al. teaches the step of culturing said target cell population under appropriate conditions for reprogramming said cells into iPSCs, wherein said appropriate conditions comprises increasing expression in said target cells, of at least the following reprogramming factors: Oct4, Klf4, Sox2, c-Myc, Lin28 (Abstract). The reprogramming factors reads on the transformation agents delivered to the subset of cells to obtain a plurality of iPSCs.
However, Prieur et al. fails to teach contacting the target cell population with a reversible cell cycle inhibiting agent in order to make the senescent cells.
However, Donovan et al. complements Prieur et al. by teaching that cell are sensitive to TGF-β during a discrete period in early G1 phase, until they reach a 'restriction point' 6-10 h after G0 release. When TGF-β is added after this critical time point, cells complete the cell cycle but arrest during the subsequent G1 phase (page 117, column 2, paragraph 4, lines 1-5). Donovan et al. further teaches that "TGF-β can either lengthen G1 transit time or cause arrest in the G1 phase. This cell cycle arrest is usually reversible (Donovan at p. 116, column 2, paragraph 2, lines 1-3). Adding TGF-β to the cells after which cells complete the cell cycle but arrest during the subsequent G1 phase reads on contacting the target cell population with a reversible cell cycle inhibiting agent. Additionally, Donovan et al. teaches that TGF-β can either lengthen G1 transit time or cause arrest in late G1 phase. This cell cycle arrest is usually reversible (page 116, column 2, paragraph 2, lines 1-3).
Given the teaching of Prieur et al. of using senescent cells that were induced by serial culturing and the teaching of Donovan et al. that senescence can be induced by contacting cells with the reversible inhibitor TGF- β and since both references teach creating senescent cells, it would have been prima facie obvious to one of the ordinary skills in the art before the effective filing date of the claimed invention to have modified the cells of Prieur et al. to involve the step of contacting the cells with TGF-beta for the purpose of generating senescent cells and substituting the technique of Donovan et al. to induce senescence in cells in the method of Prieur et al. with predictable results.
However, Donovan et al. fails to teach that contacting the target cell population with the reversible cell cycle inhibiting agent (TGF- β) to arrest the cell cycle at in a first cell cycle phase selected from Gl/S or G2, and then removing the agent;(b) following step (a), selecting, from the biological cells, a subset of cells determined to be in mitotic (M) phase, thereby obtaining a subset of cells that are enriched in cells at the same mitotic phase.
However, Silberstein et al. teaches to investigate the reversibility of TGF- β induced inhibition, glands were treated for 4 days to induce inhibition, after which the implant was surgically removed. Eleven days
after TGF- β removal, end bud growth in the treated and contralateral control glands was determined (Table 2 and page 291, column 3, paragraph 4, lines 1-6). Silberstein et al. further teaches that the end buds are bulbous, highly mitotic (page 291, column 2, paragraph 2, lines 10-12).
Therefore, it would have been prima facie obvious to one of the ordinary skills in the art before the effective filing date of the claimed invention to have contacted the target cell population of Prieur et al. with TGF- β for 4 days to arrest the cell cycle at a first cell cycle phase and the removed the TGF- β until the specific desired second cell cycle phase is determined to have been reached with a reasonable expectation of success. One would have been motivated to have done so in order to facilitate later estimation of growth and to reach the specific desired M cell cycle phase.
Regarding claim 2: Following discussion of claim 1 above, Prieur et al. in view of Donovan et al. fails to teach contacting the target cell population with the cell cycle inhibiting agent (TGF- β) to arrest the cell cycle at a first cell cycle phase and then removing the inhibiting agent until the specific desired cell cycle phase is determined to have been reached such that the selected subset of cells is in a second cell cycle phase.
However, Silberstein et al. teaches to investigate the reversibility of TGF- β induced inhibition, glands were treated for 4 days to induce inhibition, after which the implant was surgically removed. Eleven days
after TGF- β removal, end bud growth in the treated and contralateral control glands was determined (Table 2 and page 291, column 3, paragraph 4, lines 1-6). This reads on contacting the target cell population with the cell cycle inhibiting agent (TGF- β) to arrest the cell cycle at a first cell cycle phase and then removing the inhibiting agent until the specific desired cell cycle phase is determined to have been reached such that the selected subset of cells is in a second cell cycle phase.
Therefore, it would have been prima facie obvious to one of the ordinary skills in the art before the effective filing date of the claimed invention to have contacted the target cell population of Prieur et al. with TGF- β for 4 days to arrest the cell cycle at a first cell cycle phase and the removed the TGF- β until the specific desired second cell cycle phase is determined to have been reached with a reasonable expectation of success. One would have been motivated to have done so in order to facilitate later estimation of growth and to reach the specific desired second cell cycle phase.
Regarding claims 6 and 7: Following discussion of claim 1 above, Donovan et al. further teaches that TGF-β induces p15INK4B and stabilizes the p15 protein, leading to p15 binding and inhibition of cdk4 and cdk6. Cyclin D1 and KIP molecules dissociate from cdk4 and cdk6, and p27 accumulates in cyclin E-cdk2 complexes, inhibiting the latter (page 118, column 1, paragraph 4, lines 6-10). This reads on that the inhibiting agent (TGF-β) modulates the kinase inhibitory protein (kip) pathway as recited in claim 6 and p27 reads on the protein modulated by the inhibiting agent as recited in claim 7.
Regarding claim 8: Following discussion of claim 6 above, Donovan et al. specifically teaches that TGF-β is added to the cells, such that the cells complete the cell cycle but arrest during the subsequent G1 phase (page 117, column 2, paragraph 4, lines 1-5).
Regarding claim 16: Following discussion of claim 1 above, Prieur et al. further teaches contacting the target cell population with propidium iodide that binds with DNA, using flow cytometry to analyze cells stained with Propidium iodide and obtaining senescent cells by cell culture assessed by FACS (paragraphs 0065 and 0187). propidium iodide reads on the detectable dye that the target cell population is contacted with.
However, Prieur et al. fails to teach detecting a signal indicative of the presence and/or quantity of the labeled agent within and/or on the biological cells based on the detectable dye and selecting, as the subset of cells, biological cells that are determined to comprise a substantially similar quantity of the labeled agent based on the detected signal.
However, it would have been prima facie obvious to one of the ordinary skills in the art before the effective filing date of the claimed invention to have understood that a subset of cells would be selected that comprise a similar amount of propidium iodide, provided the teaching of cells obtained from donors being cultured and assessed by FACS for cellular senescence (paragraphs 0187 and 0191), as cells that incorporate a similar amount of the dye can be considered senescent, and used for further experiments for reprogramming into IPSCs.
Claims 1-2, 6-8, 16, and 21 are rejected under 35 U.S.C. 103 as being unpatentable over Prieur et al. (US20140056860A1, filed on 04/10/2012, and published on 02/27/2014), in view of Donovan et al. (Donovan et al., “Transforming growth factor-β and breast cancer: Cell cycle arrest by transforming growth factor-β and its disruption in cancer”. Breast Cancer Res. 2000;2(2):116-24) and Silberstein et al. (Silberstein et al., “Reversible Inhibition of Mammary Gland Growth by Transforming Growth Factor-β”. Science. 1987; 237(4812): 291-293), as applied to claims 1-2, 6-8, and 16 above, and further in view of Whitfield et al. (Whitfield et al., “Common markers of proliferation”. Nat Rev Cancer 6, 99–106 (2006)).
Regarding claims 1-2, 6-8, and 16, the discussions of Prieur et al., Donovan et al., and Silberstein et al. are set forth in detail above.
Regarding claim 21: Following discussion of claim 16 above, Prieur et al. in view of Donovan et al. and Silberstein et al. fails to teach contacting the cells with a labeled agent that binds to or comprises a cellular marker for proliferation.
However, Whitfield et al. teaches that proliferation signature was first identified through both the gene expression profiling of human tumour samples and through a comparison of normal cell lines that had undergone cell-cycle arrest with those that were proliferating. This signature was found to be characterized by genes whose increased expression was correlated with increased proliferation rates in the cells under analysis. This analysis revealed a set of genes (which included the cell cycle transcriptional regulator E2F1 as well as the replication-initiation complex proteins minichromosome maintenance 2–6 (MCM2–MCM6)), whose increased expression patterns were correlated with tumour cell proliferation rates, as assessed by the Ki67-labelling index (page 99, column 1, paragraph 2, lines 1-5). Whitfield et al. further teaches that the Ki67-based labelling index is commonly used in clinical assessment, and the expression levels of proliferating cell nuclear antigen (PCNA) can also be used (page 103, column 2, paragraph 1, lines 13-19). Ki67, PCNA, MCMs are cellular markers for proliferation that are used to assess the increased expression patterns that were correlated with tumour cell proliferation rates.
Therefore, it would have been prima facie obvious to one of the ordinary skills in the art before the effective filing date of the claimed invention to have modified the labeled agent that the target cell population of Prieur et al. is contacted with such that it bonds to a cellular marker for proliferation, such as Ki67 or MCM with a reasonable expectation of success. One would have been motivated to have done so because cellular markers for proliferation are vital to assess and quantify cell growth and division, crucial for understanding normal cell health, tumour progression, and clinical assessments.
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 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 HANAN ISAM ABUZEINEH whose telephone number is (571)272-9596. The examiner can normally be reached Mon- Fri 8:30-5:00.
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Hanan Isam Abuzeineh
/H.I.A./Examiner, Art Unit 1633
/CHRISTOPHER M BABIC/Supervisory Patent Examiner, Art Unit 1633