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 .
Priority
Acknowledgement is made of Applicants’ claim for benefit of prior-filed US Provisional application no. 63/268344 (filed 2/22/2022).
Election/Restrictions
Applicant's election with traverse of Group I, a decellularized stem cell sheet and method of making such, in the reply filed on 3/23/2026 is acknowledged. The traversal is on the ground(s) that Groups II and III are related, as they both require application of the decellularized stem cell sheet to a tissue site within a subject. This is not found persuasive because the method of Group II does not require application of the decellularized stem cell sheet to a tissue site within a subject. Such a limitation is not required by independent claim 29, and is thus not required by the method of Group II. The restriction set forth why the methods of Group II and III were related, but distinct methods, that conclusion stands. The requirement is still deemed proper and is therefore made FINAL.
Claims 1-38 remain pending. Claims 1-12 read on the elected invention and have been considered on the merits. Claims 13-38 are withdrawn as being directed to non-elected inventions.
Information Disclosure Statement
The listing of references in the specification is not a proper information disclosure statement. 37 CFR 1.98(b) requires a list of all patents, publications, or other information submitted for consideration by the Office, and MPEP § 609.04(a) states, "the list may not be incorporated into the specification but must be submitted in a separate paper." Therefore, unless the references have been cited on the IDS of 3/29/2023, or on a form PTO-892, they have not been considered.
Specification
The use of the term Triton [X-100], which is 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.
The drawings submitted 2/16/2023 are objected to because they contain color images and there is no granted petition to accept color drawings.
Color photographs and color drawings are not accepted in utility applications unless a petition filed under 37 CFR 1.84(a)(2) is granted. Any such petition must be accompanied by the appropriate fee set forth in 37 CFR 1.17(h), one set of color drawings or color photographs, as appropriate, if submitted via the USPTO patent electronic filing system or three sets of color drawings or color photographs, as appropriate, if not submitted via the via USPTO patent electronic filing system, and, unless already present, an amendment to include the following language as the first paragraph of the brief description of the drawings section of the specification:
The patent or application file contains at least one drawing executed in color. Copies of this patent or patent application publication with color drawing(s) will be provided by the Office upon request and payment of the necessary fee.
Color photographs will be accepted if the conditions for accepting color drawings and black and white photographs have been satisfied. See 37 CFR 1.84(b)(2).
Claim Interpretation
The term “decellularization” is defined in the as-filed specification at pages 9-10 as ‘substantial (i.e. at least 75%...) removal of cellular components by use of chemical and biological means.”
Thus, the claims directed to a decellularized stem cell sheet will be interpreted as referring to a sheet-shaped product that has been generated in some capacity from stem cells, yet at least 75% of cellular components have been removed by use of chemical and biological means.
Claim 5 is interpreted as requiring zero cells or dsDNA (yet, by dependency on claim 1, the sheet must still comprise small RNA molecules).
Claim Objections
Claims 5 and 9 are objected to for minor informalities:
In claim 5 the full term “double stranded DNA” should precede the first use of the abbreviation (dsDNA).
In claim 9, at the end of line 4, the words “to the stem cell sheet” appear to be a typographical/editing error. They should be deleted.
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 1-5 and 7-12 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.
Regarding claim 1: The term “small [RNA molecules]” in claim 1 is a relative term which renders the claim indefinite. The term “small” 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.
Claims 2-5 and 7-8 depend from claim 1, inherit the deficiency and thus are rejected on the same basis. It is noted claim 6 is not included in the rejection because claim 6 provides a size limitation for ‘small’.
Regarding claim 9: Claims 9 and 10 contain the trademark/trade name Triton [X-100]. 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 non-ionic surfactant 2-[4-(2,4,4-trimethylpentan-2-yl)phenoxy]ethanol and, accordingly, the identification/description is indefinite. Please note: if the claim is amended to insert the generic chemical name, the specification must, too, be amended to provide antecedent basis.
Claims 11 and 12 depend from claim 9, inherit the deficiency, and are thus rejected on the same basis.
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 1-4 and 6 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Zhou et al (Stem Cells Int’l, 2020).
Zhou et al teach preparation of decellularized adipose-derived stem cell (ADSC) sheets (See abstract). Specifically, Zhou et al prepare ADSC sheets from canine ADSCs (See Pg 2, “2.3 Formation of ADSC Sheets”), and then subject the sheets to three different decellularization procedures (See Pg 3, “2.4. Decellularization of ADSC Sheets”). For purposes of this rejection, the “Triton X-100 method” and resulting decellularized ADSC sheet is being relied upon.
The ADSC sheet decellularized by Triton X-100 method is described as having cells removed with no cell remnants or cellular debris visible by H&E staining (See Pg 4-5 “3.2. Evaluation of Decellularized Efficiency with Different Methods”), as retaining arrangement of collagen (See id), as having a thickness of 67 ±6.4 µm (See Pg 5 “3.3. Analysis of Retained Cytokines…”), as containing cytokines VEGF, bFGF and TGFß (See id).
Regarding claims 1-3 and 6: The ADSC sheet decellularized by Triton X-100 is considered to read on a decellularized stem cell sheet. The sheet is derived from canine (animal) adipose derived stem cells. The decellularized ADSC sheet contains collagenous ECM. The decellularized ADSC contains growth factors, which reads on bioactive factors secreted by the stem cells.
Zhou et al does not specifically comment on the presence of small RNA molecules, including RNA molecules having a size of less than 200 bp, however, given that the Triton X-100 method does not involve use of an RNase, there is a reasonable basis to conclude that the decellularized ADSC sheet of Zhou et al retained some small RNA molecules present within the ECM, including RNA molecules of less than 200 bp. These are part of normal cellular debris within cells. Zhou et al does note that the decellularized ADSC sheets obtained by the Triton X-100 method retained approximately 19 ± 6 ng/mg (dry weight) DNA (See Pg 5-6 “3.2. Evaluation…”). This is considered to evidence that a small amount of nucleic acid residue remained on/in the decellularized sheets. Without specific steps to eliminate 100% of residual nucleic acids, including RNA, including RNA less than 200 bp, some will inevitably remain. Therefore, the decellularized sheet is considered to contain small RNA molecules, including RNA particles having a size of less than 200 bp.
Regarding claim 4: Zhou et al note the collagen structure of the ECM is substantially maintained in the Triton X-100 decellularized sheet. The growth factors present read on non-collagenous proteins.
Claims 1-7 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Qin et al (US 2020/0121828).
Qin et al discloses methods of preparing a bioactive scaffold for tendon repair. The method involves: (1) decellularizing a fresh tendon tissue to obtain a decellularized tendon sheet scaffold; and
(2) adding ECM materials to the decellularized tendon sheet scaffold (See ¶0011-0013).
More specifically, in Example 1 Qin et al disclose:
For step (1):
providing a fresh tendon tissue and washing it;
freezing and thawing the tendon tissue;
slicing the tendon tissue to form slices;
subjecting the slices to nuclease treatment to obtain decellularized tendon slice scaffolds (DTSs) (See ¶0067).
Then for step (2):
culturing rat tendon-derived stem cells on the DTSs under conditions to form dense cell sheet;
decellularizing the cultured composite by first treating with 0.5% Triton X-100 containing ammonia water for 15 min, then treating with 100 IU DNase for 2 hrs (See ¶0069). Qin et al refer to the decellularized composites as ECM-DTSs (See ¶0102)
Qin et al report cells are completely removed (See ¶0070, 0047, 0099, Fig. 3).
Regarding claims 1-3, 5, 6: The ECM-DTSs read on the decellularized stem cell sheet of the claims. The ECM-DTSs comprises ECM deposited by the rat (animal) tendon-derived stem cells, as well as that of the DTS scaffold. The ECM-DTSs retains growth factors (bioactive factors) from the tendon derived stem cells (See ¶0049, 0101, Fig. 5).
Regarding the presence of small RNA molecules and the absence of dsDNA: Qin et al is silent about these two parameters. However, Qin et al subject the cultured composite to 100 IU DNase for 2 hrs, which is substantially the same nuclease treatment carried out in the instant specification (Instant specification subjects TDSC sheets to 150 U/mL DNase for 2 hrs (See Pg 22)). The instant specification teaches this treatment was effective to remove dsDNA content (See Pg 22, and Fig. 3B) and appears to be basis for support for the claimed product which comprises small RNA molecules of less than 200 bp in length. Because the prior art subjects the tendon stem cell-derived cell composite to the same nuclease treatment as the instant application, the resulting nucleic acid content (both dsDNA, small RNA molecules, and RNA molecules less than 200 bp) must be the same in the prior art as reported in the instant application. Where the claimed and prior art products… are produced by identical or substantially identical processes, a prima facie case of anticipation is established. "When the PTO shows a sound basis for believing that the products of the applicant and the prior art are the same, the applicant has the burden of showing that they are not." See MPEP 2112.01.
Regarding claim 4: Qin et al reports the DTSs contains collagen (See ¶0006). Qin et al also report the ECM-DTSs contains growth factors, such as VEGF. Thus the ECM-DTSs contains both collagenous and non-collagenous proteins.
Regarding claim 7: Qin et al reports the ECM-DTSs contains growth factors, including VEGF. Qin et al is silent about the presence of BMP2. However, given that Qin et al culture tendon-derived stem cells, the same type of cells as the instant application teaches to secrete VEGF and BMP2, then there is reasonable basis to conclude that the tendon derived stem cells in Qin et al will also secrete VEGF and BMP2, and these will be present in the ECM-DTSs. Burden of proof is shifted to Applicants. See MPEP 2112.01.
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 1-7 are rejected under 35 U.S.C. 103 as being unpatentable over Lui et al (US 2012/0189587), in view of Zhou et al (Stem Cells Int’l, 2020).
Lui et al teach cell sheets formed from a stem cell for the promotion of tissue repair and bioartificial tissue engineering (See ¶0070). The cell sheets are formed by treating stem cells so that the treated stem cells become embedded in their self-secreted ECM, thereby forming a cell sheet comprising the cells and the ECM (See ¶0071). The stem cell used can be tendon-derived stem cells (TDSCs) isolated from animals (See ¶0076). Lui et al teach an embodiment wherein the above described cell sheet is rendered acellular through decellularization (See ¶0088).
Regarding claims 1-3, 6: The decellularized TDSC sheet is comparable to the decellularized stem cell sheet of the claims.
Lui et al does not provide details on the decellularization process, nor characterize the resulting decellularized TDSC. When a prior patent does not teach details of a protocol, it is prima facie obvious to look to other prior art in the same field to determine appropriate steps. In this case it would have been obvious to utilize the decellularization process of Zhou et al, as they teach decellularization of similar stem cell sheets. The teachings of Zhou et al are set forth above. The decellularization method of Zhou et al successfully removes cells and substantially all nucleic acid material, but retains the ECM structure, proteins and growth factors secreted by the stem cells. As such, it is reasonable to conclude that applying the decellularization method of Zhou et al to the TDSC sheet of Lui et al would result in a decellularized TDSC sheet that also retains the ECM structure, proteins and growth factors secreted by the TDSCs. There is a reasonable expectation of success based on the teachings of Zhao et al that their method (specifically the Triton X-100 method) removed cells and preserved the ECM and growth factors within the ECM. As such, there is a reasonable expectation that applying the Triton X-100 method to the TDSC of Lui et al would result in the decellularized TDSC sheet which retains ECM, small RNA molecules, including those sized under 200 bp, and bioactive factors secreted by the cells.
Regarding claim 4: Lui et al teaches the ECM contains collagen and the growth factor BMP2 (See ¶0034, Fig. 3A-D). Therefore the TDSC sheet would contain collagen and BMP2, which reads on non-collagenous protein.
Regarding claim 7: For the reasons discussed above, it is reasonable to conclude that because Lui et al use the same cells to generate the stem cell sheet, the proteins, including growth factors secreted into the ECM would be the same as reported in the instant application. The instant application reports TDSC secrete BMP2 and VEGF (See Example 1 at Pg 23 and Fig. 4M and 4N). Thus there is reasonable basis to conclude that the tendon derived stem cells in Lui et al will also secrete VEGF and BMP2, and these will be present in the acellular TDSC sheet decellularized by the method of Zhao et al. Burden of proof is shifted to Applicants. See MPEP 2112.01.
Claims 9-12 are rejected under 35 U.S.C. 103 as being unpatentable over Zhou et al (Stem Cells Int’l, 2020), in view of Gilpin et al (BioMed Research International, 2017) and Qin et al (US 2020/0121828).
Zhou et al teach preparation of decellularized adipose-derived stem cell (ADSC) sheets (See abstract). Specifically, Zhou et al prepare ADSC sheets from canine ADSCs (See Pg 2, “2.3 Formation of ADSC Sheets”), and then subject the sheets to three different decellularization procedures (See Pg 3, “2.4. Decellularization of ADSC Sheets”). For purposes of this rejection, the “Triton X-100 method” and resulting decellularized ADSC sheet is being relied upon.
Regarding claims 9 and 10: Zhou et al decellularizes an ADSC sheet. The “Triton X-100 method” involves:
isolating ADSCs from canines (See Pg 2, “2.2. Primary Cell Isolation and Culture”,
culturing the ADSCs under conditions to form a cell sheet (See Pg 2, “2.3. Formation of ADSC Sheets”),
placing the cell sheets in a decellularization solution comprising 1% Triton X-100, 0.02% EDTA (equivalent to about 0.5 mM), 10 mM Tris, and 1 µg/mL aprotinin with shaking for 24 hrs at room temperature, then
rinsing with PBS for 24 hrs (See Pg 3 “2.4. Decellularization of ADSC Sheets”).
The step of isolating the ADSCs from canines reads on claimed step (i).
The step of culturing the ADSCs under conditions to form cell sheets reads on claimed step (ii).
The step of placing the cell sheets in the decellularization solution is comparable to claimed step (iii). The decellularization solution of Zhou et al differs from that in the current claim only in the concentrations of the agents present. Zhou et al use Triton X-100, Tris, EDTA and aprotinin. The effect of each of these agents, individually was known (See, e.g. Gilpin et al). Modification of the concentrations, particularly of the Triton X-100 and EDTA would have been a matter of routine optimization. One having ordinary skill in the art would understand that the concentration of decellularization agents can be modified based on the duration of exposure, temperature, and/or presence of other synergistic agents. Gilpin et al discuss various combinations of agents and factors that can be used to work together to advance decellularization. Generally, differences in concentration will not support the patentability of subject matter encompassed by the prior art unless there is evidence indicating such concentration or temperature is critical. See MPEP 2144.05.
Zhou et al does not teach further treating the stem cell sheet with DNase I. Gilpin et al teach that nucleases, such as DNase I were known to break down DNA fragments left following cell removal (See Table 1 & Pg 5, “2.1.3. Enzymatic Assisted Decellularization”). Qin et al also exemplifies use of DNase in decellularization of stem cell sheets (See Qin et al ¶0069). Thus DNase was a known agent with a known benefit in decellularization protocols. Zhou et al wants low levels of residual DNA, therefore it would have bene prima facie obvious to have utilized DNase I, an agent known to reduce DNA content following detergent decellularization protocols, in the Triton X-100 protocol of Zhou et al. One would have been motivated in order to further reduce the DNA content, and one would have had a reasonable expectation of success based on the disclosure of Gilpin et al and successful protocol of Qin et al.
Regarding claims 11-12: The method of Zhou et al concludes with a 24 hr rinse in PBS at room temperature. This reads on the rinsing with a buffer steps of claims 11 and 12.
Conclusion
Any inquiry concerning this communication or earlier communications from the examiner should be directed to ALLISON M FOX whose telephone number is (571)272-2936. The examiner can normally be reached M-F 10-6 EST.
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/ALLISON M FOX/Primary Examiner, Art Unit 1633
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