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
A request for continued examination under 37 CFR 1.114, including the fee set forth in 37 CFR 1.17(e), was filed in this application after final rejection. Since this application is eligible for continued examination under 37 CFR 1.114, and the fee set forth in 37 CFR 1.17(e) has been timely paid, the finality of the previous Office Action has been withdrawn pursuant to 37 CFR 1.114. Applicant’s submission filed on November 4, 2025 has been entered.
The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action.
This action is in response to the papers filed on November 4, 2025. Claims 1 - 16 are currently pending. Claim 1 has been amended in the Applicant’s amendment filed November 4, 2025. No claims have been added or canceled in the Applicant’s amendment filed November 4, 2025.
Applicant's election without traverse of Group I, claims 1 – 6, drawn to a method of preparing human cell-derived microcells; and the election of Species without traverse as follows:
Species (A): normal human chromosome (instant claim 2);
Species (B): a microtubule depolymerization inhibitor as the micronucleus inducer (instant claim 5) in the reply filed September 30, 2024 is acknowledged.
Claims 7 – 16 were previously withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a non-elected invention, there being no allowable generic or linking claim.
Claim 3 – 4, and 6 were previously withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a non-elected species, there being no allowable generic or linking claim.
The restriction requirement was deemed proper and was therefore made FINAL.
The claims will be examined insofar as they read on the elected species.
The examiner acknowledges receiving the Declaration under 37 C.F.R. § 1.132 filed on November 4, 2025 and executed by Dr. Narumi Uno.
Therefore, claims 1, 2, and 5 are under consideration to which the following grounds of rejection are applicable.
Information Disclosure Statement
The information disclosure statements (IDS) submitted on November 4, 2025 has been considered. An initialed copy of the IDSs accompanies this Office Action.
Priority
The present application filed December 30, 2020, is a 35 U.S.C. 371 national stage filing of International Application No. PCT/JP2019/040091, filed October 10, 2019, which claims the benefit of Japanese Patent Application JP2018-191994, filed October 10, 2018.
Withdrawn Objections/Rejections
Claim Rejection - 35 USC § 112(b)
The rejection of claims 1, 2, and 5 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 is withdrawn.
The Applicant has convinced the Examiner that one of ordinary skill in the art would know the metes and bounds of the term “human cell-derived microcells.”
In view of the withdrawn rejection, Applicant’s arguments are moot.
Claim Rejections - 35 USC § 102
The rejection of claims 1, 2, and 5 under 35 U.S.C. 102 (a)(1)/(a)(2) as being anticipated by Samwer et al. (hereinafter referred to as “Samwer”) (Samwer M. et al. DNA Cross-Bridging Shapes a Single Nucleus from a Set of Mitotic Chromosomes. Cell. 2017 Aug 24;170(5):956-972.e23. doi: 10.1016/j.cell.2017.07.038. PMID: 28841419; PMCID: PMC5638020.) is withdrawn.
Samwer does not teach collecting the human cell-derived microcells by centrifugation in a medium supplement with an action polymerization inhibitor, and then selecting the human cell-derived microcells comprising the DNA of interest by drug selection (amended claim 1).
In view of the withdrawn rejection, Applicant’s arguments are moot.
New Objections/Rejections
Claim Rejection - 35 USC § 112(a) Scope of Enablement
The following is a quotation of the first paragraph of 35 U.S.C. 112(a):
(a) IN GENERAL.—The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor or joint inventor of carrying out the invention.
The following is a quotation of the first paragraph of pre-AIA 35 U.S.C. 112:
The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor of carrying out his invention.
Claims 1, 2, and 5 are newly rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, because the Specification, while being enabling for
A method for preparing human cell-derived microcells from human cells comprising a DNA of interest, the method comprising culturing the cells in a medium supplemented with microtubule polymerization inhibitor, wherein the microtubule polymerization inhibitor is a mixed reagent of colchicine and vincristine (MAS) at a concentration of 0.2 μg/mL, or a microtubule depolymerization inhibitor and a spindle checkpoint inhibitor, wherein the microtubule depolymerization inhibitor is paclitaxel, and the spindle checkpoint inhibitor is reversine, and wherein the DNA of interest is a human chromosome,
does not reasonably provide enablement for culturing the human cells in a medium comprising a genus of micronucleus polymerization inhibitors, except for 1,2,3, l 0-Tetramethoxy-7-(methylamino)-6, 7-dihydrobenzo[a]heptalen-9(5H)-one, a genus of micronucleus depolymerization inhibitors, and a genus of spindle checkpoint inhibitors, or any combination of these inhibitors, or wherein the DNA of interest is any DNA sequence.
The Specification does not enable any person skill in the art to which it pertains, or with which it is most nearly connected, to use the invention commensurate in scope with these claims. The claims, when given the broadest possible interpretation, encompass a method for preparing human cell-derived microcells comprising a DNA of interest, wherein the human cells are cultured in a medium comprising at least one micronucleus least one micronucleus inducer, selected from the group consisting of a microtubule polymerization inhibitor, except for 1,2,3, l 0-Tetramethoxy-7-(methylamino)-6, 7-dihydrobenzo[a]heptalen-9(5H)-one, a microtubule depolymerization inhibitor, and a spindle checkpoint inhibitor to produce human cell-derived microcells, and collecting the human cell-derived microcells comprising the DNA of interest by centrifugation in a medium supplemented with a genus of actin polymerization inhibitors, and then selecting the human cell-derived microcells comprising the DNA of interest by drug selection. The Specification does not enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to use the invention commensurate in scope with these claims.
The test of enablement is whether one skilled in the art could make and use the claimed invention from the disclosures in the patent coupled with information known in the art without undue experimentation (United States v. Telectronics, Inc., 8 USPQ2d 1217 (Fed. Cir. 1988)). Whether undue experimentation is required is not based on a single factor but is rather a conclusion reached by weighing many factors (See Ex parte Forman, 230 USPQ 546 (Bd. Pat. App. & Inter, 1986) and In re Wands, 8USPQ2d 1400 (Fed. Cir. 1988); these factors include the following:
Nature of invention. The invention encompasses a method for preparing human cell-derived microcells comprising a DNA of interest, wherein the human cells are cultured in a medium comprising at least one micronucleus least one micronucleus inducer, selected from the group consisting of a microtubule polymerization inhibitor, except for 1,2,3, l 0-Tetramethoxy-7-(methylamino)-6, 7-dihydrobenzo[a]heptalen-9(5H)-one, a microtubule depolymerization inhibitor, and a spindle checkpoint inhibitor to produce human cell-derived microcells, and collecting the human cell-derived microcells comprising the DNA of interest by centrifugation in a medium supplemented with an actin polymerization inhibitor, and then selecting the human cell-derived microcells comprising the DNA of interest by drug selection.
Scope of the invention. The invention encompasses a method of preparing human cell-derived microcells that comprises DNA of interest.
Number of working examples and guidance. In the instant case, Applicant provides 7 relevant working examples. Example 1 teaches the search of micronucleus inducer for human iPS cells, wherein 22 conditions in total were designed based on treatments with colcemid 0.25 μg/mL, MAS 0.2 μg/mL, paclitaxel (at a range of concentrations, and reversine (Paragraph [0117]). Examples 2, 6, and 7 teach the transfer of a chromosome, wherein the micronucleation induction treatment using paclitaxel (1 nM to 1 μM) and reversine (1 nM to 1 μM) (Paragraph [0134]). Examples 3 and 4 teaches the transfer of a chromosome, wherein the micronucleation induction treatment using MAS (0.2 μg/mL) (Paragraph [0140]). These examples only teach the transfer of chromosomes. Further, the micronucleus induction treatment comprises either of MAS at 0.2 μg/mL or the combination of paclitaxel and reversine.
State of the art. Although the field of microcells is highly developed, the
method of preparing human cell derived microcells from human cells specific micronucleus inducers is not highly developed. The art must therefore be considered to be poorly developed.
Unpredictability of the art. Before the effective filing date of the claimed
invention, it was known in the art that colchicine was in the microcell-mediated chromosome transfer, wherein murine donor cells containing additional human chromosome with a resistance gene were treated for 48h with colchicine, as evidenced by Passerini V. et al. (Passerini et al. The presence of extra chromosomes leads to genomic instability. Nat Commun. 2016 Feb 15;7:10754. doi: 10.1038/ncomms10754. PMID: 26876972; PMCID: PMC4756715.) (pg. 10, left column, third paragraph). This reference teaches a specific concentration of the colchicine that is used in chromosome transfer.
Further, it was known in the art that Colcemid and Cytochalasin B was replaced with TN-16 + Griseofulvin and Latrunculin B in combination with a Collage/Laminin surface coating increases the efficiency of HAC transfer to recipient cells by almost sixfold and is possibly less damaging to HAC than the standard MMCT method, as evidenced by Liskovyh et al. (Liskovykh M. et al. Moving toward a higher efficiency of microcell-mediated chromosome transfer. Mol Ther Methods Clin Dev. 2016 Jun 22;3:16043. doi: 10.1038/mtm.2016.43. PMID: 27382603; PMCID: PMC4916947.). This reference teaches the variability of the MMCT protocol, and the variability of micronucleus inducers.
It was known in the art that vincristine, at low concentrations, announces perturbation/unfolding of DNA and chromatin structure, whereas vinchristine at higher concentration precedes DNA and chromatin into compaction, as evidenced by Mohammadgholi A. et al. (Mohammadgholi A. et al. Mechanism of the interaction of plant alkaloid vincristine with DNA and chromatin: spectroscopic study. DNA Cell Biol. 2013 May;32(5):228-35. doi: 10.1089/dna.2012.1886. Epub 2013 Apr 16. PMID: 23590199; PMCID: PMC3651685.) (pg. 233, right column, third concentration). This reference teaches that vinchristine is concentration dependent.
Additionally, the prior art of Samwer (Samwer M. et al. Cell. 2017 Aug 24;170(5):956-972.e23) discloses that live HeLA cells are cultured and imaged in the presence of taxol, wherein the drug-treated cells entering mitosis disassembled the NE and released individualized mitotic chromosomes into the cytoplasm, however, Samwer does not teach the step of collecting micronuclei by centrifugation and drug selection (page 9 of Applicants’ remarks filed on 11/4/2025) where “microcells are released from outside of the cells by centrifugation, and each of the released microcells have a single micronuclei and cell membrane, and then, microcells comprising the DNA of interest are selected by drug selection . “(page 10 of Applicants’ remarks filed on 11/4/2025). The applicant is one record saying that when paclitaxel and reversine are used, multiple micronuclei are formed, instead of a single nucleus (pg. 11 of Applicants’ remarks filed on 11/4/2025). The applicant is on record saying that, in reference to Figure 3 of Barrientos et al., the cells are treated with colcemid for micronucleation, and then subjected to centrifugation to release microcells (pg. 12 of Applicants’ remarks filed 11/4/2025). Thus, paclitaxel and reversine must be used to generate microcells, followed by centrifugation.
Amount of Experimentation Required. Given the unpredictability of the art, the
variability in the chromosome transfer protocol, the specific micronucleus inducers used, and the particular concentrations of micronucleus inducers, the skilled artisan would have to conduct undue, and unpredictable experimentation to practice the claimed invention generating the human cell derived microcells.
Conclusion
Claims 1, 2, and 5 remain rejected.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to VYOMA SHAILESH THAKKER whose telephone number is (571)272-2954. The examiner can normally be reached M-F 8:30 - 5:30 EST.
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/VYOMA SHUBHAM TIWARI/Examiner, Art Unit 1634
/MARIA G LEAVITT/Supervisory Patent Examiner, Art Unit 1634
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