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 included in the prosecution are claims 1-5, 9-11 and 14-16.
Applicants' arguments, filed 03/12/2026, have been fully considered. Rejections and/or objections not reiterated from previous office actions are hereby withdrawn. The following rejections and/or objections are either reiterated or newly applied. They constitute the complete set presently being applied to the instant application.
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.
1. Claims 1-5 and 9-11 are rejected under 35 U.S.C. 103 as being unpatentable over Hwang (KR 10-2019-0080723 A. Jul. 8, 2019) (IDS reference) in view of Cosenza et al. (Mesenchymal stem cells derived exosomes and microparticles protect cartilage and bone from degradation in osteoarthritis, Nov. 24, 2017) (IDS reference) (hereinafter Cosenza), Niibe et al. (US 2021/0139855, Filing date: Apr. 20, 2018) (hereinafter Niibe), and Baraniak et al. (Scaffold-free culture of mesenchymal stem cell spheroids in suspension preserves multilineage potential, Aug. 29, 2014) (hereinafter Baraniak).
Hwang discloses a method for extracting exosomes (i.e., extracellular vesicles) from stem cells (abstract). The method comprises (a) culturing stem cells in a three-dimensional rotating ellipsoid. The stem cells may be induced pluripotent stem cells or mesenchymal stem cells. The term “three-dimensional culture” mean that the cells are cultured in such a state that a cell aggregate of a three-dimensional structure is generated by self-aggregation. The cells are suspended in a culture medium and are plated on a round bottom plate. Centrifugal force is applied thereto to produce a three-dimensional cell aggregate (page 3 of translation). The plate may be a 96-well round bottom plate (i.e., multi-well culture plate) (page 7 of translation). The cultivation of the cell aggregate can be performed by shaking culture. The shaking culture may be performed by orbital shaking (page 4 of translation). At various times of cell culture, exosomes were isolated from each culture. First, the cells and the culture were collected in a tube and centrifuged. The resulting supernatant was transferred to a centrifuge tube and centrifuge for 10-20 minutes. Separated supernatant was obtained and transferred to a polyallomer tube or polycarbonate bottle suitable for the ultracentrifuge rotor and centrifuged for 30 minutes to obtain pellets (page 7 of translation). Exosomes are generally known to have a diameter of 30-100 nm (page 3 of translation).
Hwang differs from the instant claims insofar as not disclosing wherein the culture medium comprises TGF-β.
However, Cosenza discloses wherein mesenchymal stem cells exert chondroprotective effects in preclinical models of osteoarthritis. Most of their therapeutic effects are mediated via soluble mediators, which can be conveyed within extracellular vesicles (EVs). Exosomes were isolated from BM-MSCs. It was found that the exosomes exerted chondroprotective an anti-inflammatory function in vitro and protected mice from developing OA in vivo (abstract). For evaluating the chondroprotective function of EVs, TGF-β3 was added in a proliferative medium (page 2). Exosomes isolated from TGF-β3-pretreated BM-MSCs exerted high chondroprotective effect on OA-like chondrocytes (Figure 2). TGF-β3 pre-activation of BM-MSCs stimulates their pro-chondrogenic and anti-fibroblastic function through the release of factors that might be conveyed by exosomes (page 11).
Accordingly, it would have been prima facie obvious to one of ordinary skill in the art to have incorporated TGF-β3 into the culture medium of Hwang motivated by the desire to formulate exosomes with chondroprotective function as taught by Consenza.
The combined teachings of Hwang and Cosenza do not teach wherein the orbital shaking culture is performed at 50-70 rpm.
However, Niibe discloses culturing mesenchymal stem cells (¶ [0001]). The culturing is performed by shaking culture at a rotational speed of from 20 rpm to 200 rpm (¶ [0025]).
Hwang discloses wherein the cultivation of the cell aggregate can be performed by shaking culture. Accordingly, it would have been prima facie obvious to one of ordinary skill in the art to have performed the shaking culture at a rotational speed of from 20 rpm to 200 rpm since this is a known and effective shaking culture rotational speed for culturing stem cells as taught by Niibe.
The combined teachings of Hwang, Cosenza, and Niibe do not teach wherein the round bottom plate has wells with a size of 300-500 µm and wherein 300-500 cells are seeded per each well.
However, Baraniak discloses wherein MSCs, corresponding to approximately 300, 600, or 1,000 cells/microwell were added to AggrewellTM 6-well inserts containing an array of 400x400 µm-sized microwells to form MSC spheroids (page 3).
Generally, it is prima facie obvious to select a known material for incorporation into a composition, based on its recognized suitability for its intended use. See MPEP 2144.07. Hwang discloses culturing stem cells in a well plate. Accordingly, it would have been obvious to one of ordinary skill in the art to have cultured the stem cells in AggrewellTM 6-well inserts containing an array of 400x400 µm-sized microwells since this is a known and effective well plate for culturing stem cells as taught by Baraniak.
It would have been prima facie obvious to one of ordinary skill in the art to have seeded approximately 300, 600, or 1,000 cells/microwell since Hwang does not disclose how many cells are to be seeded in each well and this is a known and effective amount for culturing stem cells as taught by Baraniak.
In regards to instant claim 1 reciting wherein cell aggregates formed in step (a) are spheroids having a diameter of 120-200 µm, as noted in the instant specification in Example 1, uniform spheroids having a diameter of 120-200 µm were produced by seeding about 400 umbilical cord-derived mesenchymal stem cells per well, wherein the well plate is AggreWellTM400 with a size of 400 µm. Thus, cell aggregate size depends on the particular well culture plate and number of cells seeded per each well. Instant claim 4 recites a well plate with a size of 300-500 µm and instant claim 5 recites wherein 300-500 cells are seeded per each well. As discussed above, it would have been obvious to one of ordinary skill in the art to have cultured the stem cells in AggrewellTM 6-well inserts containing an array of 400x400 µm-sized microwells and to have seeded approximately 300, 600, or 1,000 cells/microwell. Accordingly, since cell aggregate size depends on the particular well culture plate and number of cells seeded per each well and the method of the prior art uses substantially the same well culture plate and seeds substantially the same number of cells/microwell as claimed, the cell aggregates of the prior art would necessarily have substantially the same diameter as claimed.
2. Claims 14-16 are rejected under 35 U.S.C. 103 as being unpatentable over Hwang (KR 10-2019-0080723 A. Jul. 8, 2019) (IDS reference) in view of Cosenza et al. (Mesenchymal stem cells derived exosomes and microparticles protect cartilage and bone from degradation in osteoarthritis, Nov. 24, 2017) (IDS reference) (hereinafter Cosenza), Niibe et al. (US 2021/0139855, Filing date: Apr. 20, 2018) (hereinafter Niibe), Baraniak et al. (Scaffold-free culture of mesenchymal stem cell spheroids in suspension preserves multilineage potential, Aug. 29, 2014) (hereinafter Baraniak), and further in view of Elliman et al. (US 2019/0015331, Jan. 17, 2019) (hereinafter Elliman).
The teachings of Hwang, Cosenza, Niibe, and Baraniak are discussed above. Hwang, Cosenza, Niibe, and Baraniak do not teach wherein peroxiredoxin-4, thioredoxin reductase 1, prostaglandin G/H synthase 2, RUVBL2, and CORO1A are highly expressed in the exosomes.
However, Elliman discloses a mesenchymal stem cell (MSC)-derived exosome composition (¶ [0003]). The exosome composition comprises proteins that contribute to therapeutic efficacy or that mediate paracrine signaling to effect therapeutic efficacy. Such proteins include (¶ [0062]) PRDX4 (peroxiredoxin-4) (page 21), TXNRD1 (thioredoxin reductase 1) (page 20), PTGS2 (prostaglandin G/H synthase 2) (page 22), RUVBL2 (RuvB-like 2) (page 20), and CORO1A (page 22).
Accordingly, it would have been prima facie obvious to one ordinary skill in the art to have incorporated PRDX4 (peroxiredoxin-4), TXNRD1 (thioredoxin reductase 1), PTGS2 (prostaglandin G/H synthase 2), RUVBL2 (RuvB-like 2), and CORO1A into the exosomes of Hwang motivated by the desire to have the exosomes provide a therapeutic effect as taught by Elliman.
In regards to the proteins being highly expressed, since these proteins contribute to therapeutic efficacy as taught by Elliman, it would have taken no more than the relative skills of one of ordinary skill in the art through routine experimentation to have expressed the proteins such that it is highly expressed in order for a desired therapeutic efficacy to be achieved. Where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation. MPEP 2144.05(II)(A). One of ordinary skill in the art would have had a reasonable expectation of success since Elliman discloses in paragraph [0106] wherein an exosome composition may be genetically modified to overexpress a protein.
In regards to instant claim 14 reciting wherein the stem cell-derived extracellular vesicles are produced by the method of claim 1, even though product-by-process claims are limited by and defined by the process, determination of patentability is based on the product itself. The patentability of a product does not depend on its method of production. If the product in the product-by-process claim is the same as or obvious from a product of the prior art, the claim is unpatentable even though the prior product was made by a different process. See MPEP 2113. Accordingly, since an exosome comprising the claimed highly expressed proteins would have been obvious as discussed above, the claim is unpatentable even though the prior art does not disclose the same method as Applicant of arriving at an exosome comprising the highly expressed proteins.
Response to Arguments
Applicant argues that it is the specific combination of the claimed steps with the specific combination of culture medium composition and orbital shaking speed that resulted in the unexpected increased yield of homogenous extracellular vesicles and improved inhibitory effect on proliferative T cells. It is the specific combination of utilizing a culture medium comprising TGF-β3 and the specific orbital shaking rate of 50-70 rpm that led to the unexpected results.
The Examiner submits that Applicant’s showing appears to be probative of unexpected results; however, the claims are not commensurate in scope. To obtain an increased yield of homogenous extracellular vesicles and improved inhibitory effect on proliferative T cells, Applicant performed the method described in Examples 1 and 2. The method requires using AggreWellTM 400 treated with an F127 solution as a multi-well culture plate. One of ordinary skill in the art would not reasonably expect the results from the showing with any multi-well culture plate since the bottom shape of a culture plate determines how cells attach, grow, and behave during experiments as evidenced by GMP Plastic (The Science Behind Cell Culture Plate Bottom Shapes, Why Do Cell Culture Plates Have Different Bottom Shapes?). Also, plates from different manufacturers yield different cell culture results as evidenced by Mansoury et al. (The edge effect: A global problem. The trouble with culturing cells in 96-well plates, Highlights).
Also, the method requires seeding about 400 umbilical cord-derived mesenchymal stem cells per well. One of ordinary skill in the art would not reasonably expect the results from the showing with any number of cells seeded since cell seeding determines the initial cell concentration, which ultimately affects cell growth, nutrient availability, and space for proliferation as evidenced by ThermoFisher (Seeding, Subculturing, and Maintaining Cells, Seeding density).
Additionally, the method requires a culture medium containing TGF-β3. The claims do not recite an amount of TGF-β3. One of ordinary skill in the art would not reasonably expect the results from the showing with any amount of TGF-β3 since it would not be reasonable to expect a miniscule amount of TGF-β3 to be any different than having no amount of TGF-β3.
Furthermore, the method requires shake culturing for 3 days at 37°C. One of ordinary skill in the art would not reasonably expect the results from the showing with any length of time and temperature since cells need a specific amount of time and adequate temperature to be cultured.
Moreover, the method requires repeated centrifuging at a specific rate for a specific amount of time. One of ordinary skill in the art would not reasonably expect the results from the showing without the centrifuging steps since centrifugation causes exosomes to be isolated from cell culture supernatants as evidenced by ThermoFisher (Centrifuges for almost any application, Centrifuges for cell culture workflows).
As such, the rejection is maintained since the claims are not commensurate in scope.
Response to Declaration
Declarant’s argument is addressed above.
Conclusion
Claims 1-5, 9-11 and 14-16 are rejected.
No claims are allowed.
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.
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/TRACY LIU/Primary Examiner, Art Unit 1614
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