ETAILED 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 Status
Claims 18 and 20 are amended.
Claim 58 is new.
Claims 1, 6-7, 11 and 16-17 are withdrawn.
Claims 18-20, 22-24, 29, 34, 38, 41 and 57-58 are under examination.
Withdrawn Rejections
Rejections - 35 USC § 102
The rejection of claims 18,20, 23, 29,34, and 38 under 35 U.S.C. 102 (a)(1) as being anticipated by Levenberg et al (WO 2020/261257 A1) is withdrawn considering claim amendment. Applicant amended claim 18 to recite the limitation “(i) culturing the iPSCs or ESCs by dynamic culture on microcarrier beads or as 3D spheroids”. Levenberg et al do not expressly teach culturing iPSCs or ESCs on microcarriers or as 3D spheroids. Accordingly, the rejection is withdrawn.
Edited Rejections Necessitated by Claims Amendment
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 text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action.
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.
Claims 18-20, 22, and 23-24 are rejected under 35 U.S.C. 103 as being unpatentable over Levenberg et al (WO 2020/261257 A1), in view of Rodrigues et al (Society of Chemical Industry, 2018).
Regarding claims 18, 20, and 22, Levenberg et al teach a method for producing advanced extracellular vesicles (EVs) from stem cells. The method involves culturing stem cells on a three-dimensional (3D) porous scaffold in a bioreactor system and collecting EVs from the culture medium. Levenberg discloses that the bioreactor may be operated as a continuous stirred-tank reactor and that culture medium may be circulated through the three-dimensional scaffold at selected flow rates, thereby providing dynamic culture conditions and shear stress to the cells. (See page 4, lines 2-5). (See claim 1). Levenberg et al also identify that embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs), as suitable stem-cell sources for production of EVs. (See claim 21). The method of Levenberg et al also includes collecting the medium; and isolating the secreted EV s dispersed therein, this reads on step (ii) of instant claim 18. (See claim 1). Thus, Levenberg et al teach substantially all aspects of the presently claimed method except, at least expressly, the particular recitation of culturing the iPSCS or ESCs on “microcarrier beads”.
Rodrigues et al supplement Levenberg et al by expressly teaching culturing of human iPSCs on microcarriers in dynamic bioreactors. Rodrigues et al demonstrate culturing of hiPSCs on microcarriers in 100 ml to 500 ml vertical-wheel bioreactors and conclude that the system provides scalable production of hiPSCs in a microcarrier-based system. Rodrigues et al also describe use of commercially available microcarriers for hiPSC culture and operation of the bioreactor in a manner that maintains the microcarriers in suspension. (See abstract). Accordingly, Rodrigues et al established, before the effective filing date of the claimed invention, that microcarrier-based dynamic culture of iPSCs in bioreactors was a known technique. Rodrigues et al further state that culturing of adherent cells in bioreactors using microcarriers, which are small particles that are suspended in vessel, provide higher culture surface area per reactor volume compared to the planar platform, thereby offering a platform for large-scale culture of stem cells. (See the 2nd column- 1st paragraph of the Introduction on page 3597). As such, providing an ordinary skill in the art with the motivation to substitute the microcarriers-based culturing system of Rodrigues for the three-dimensional scaffold of Levenberg to increase the number of cells that could be cultured, and consequently enhancing EV production.
Therefore, it would have been a prima facie obvious to one with ordinary skill in art at the time the invention was filed to employ the microcarrier-based dynamic culture of Rodrigues in the dynamic bioreactor/EV production method of Levenberg. Because Levenberg et al seek enhanced production of EV from stem cells through dynamic three-dimensional culture, while Rodrigues et al seek scalable culture of iPSCs in dynamic microcarrier-based bioreactors. An ordinary skill in the art would have had a reasonable expectation that incorporating the microcarriers as the three-dimensional cell supporting environment into Levenberg’s system would improve the scale of stem-cell culture and further increase EV production. In other words, claim 18 is combining prior art elements according to known methods to yield predictable results. See MPEP 2143 (I)(A).
Regarding claim 19, following the discussion of claim 18 above, Levenberg in view of Rodrigues et al render obvious claim 18. Rodrigues et al further teach iPSCs derived from fibroblast. Therefore, for the reason discussed above with respect to claim 18, and further in view of Rodrigues teaching of iPSCs derived from fibroblast, instant claim would have been obvious to one with ordinary skill in the art. (See Material and Methods section “ Human induced pluripotent stem cell culture”).
Regarding claim 23-24, following the discussion of claim 18 above, Levenberg in view of Rodrigues renders obvious the method of culturing iPSCs in a dynamic culture on microcarriers and collecting EV from the culture medium. Levenberg et al also disclose a composition comprising extracellular vesicles (EVs) derived from induced pluripotent stem cells (iPSCs) or embryonic stem cells (ESCs). (See claim 28). Levenberg, however, does not expressly disclose that the iPSCs are derived from fibroblasts. However, Rodrigues teaches iPSCs derived from fibroblasts. ( See Material and Methods section “ Human induced pluripotent stem cell culture”). Thus, Rodrigues et al teach the additional limitation of claim 24.
Claim(s) 29,34, and 38 are rejected under 35 U.S.C. 103 as being unpatentable over Levenberg et al, in view of Rodrigues et al as applied to claims 18-20, 22, and 23-24 above, and further in view of Singla et al ( Cells, 2019), and Sun et al (Materials Science & Engineering, 2018) .
Regarding claims 29, and 34, following the discussion of claim 23 above, neither Levenberg nor Rodrigues expressly teach the immunomodulatory properties of the EVs. Specifically, neither Levenberg et al nor Rodrigues teach that the collected EVs can be used to promote secretion of anti-inflammatory cytokines by macrophages, wherein the anti-inflammatory cytokine is IL-10.
Singla, however, supplements the cited prior arts by teaching the immunomodulatory effect of ESCs-derived exosomes (i.e. EVs) on macrophages. Specifically, Singla teaches that treatment with EV increases M2 macrophages and increases the anti-inflammatory cytokine IL-10, while inhibiting pro-inflammatory M1 macrophages and inflammatory signaling. (See abstract, and Fig.7). Singla et al further teach that ESCs-derived exosomes inhibit inflammatory responses associated with proinflammatory macrophages. Accordingly, Singla et al teach the claimed promotion of M2 macrophage differentiation and promotion of secretion of an anti-inflammatory cytokine such as IL10 by macrophages. Therefore, it would have been prima facie obvious to one with ordinary skill in the art to apply the immunomodulatory teachings of Singla to the EV compositions of Levenberg, as modified by Rodrigues, because Singla demonstrates that stem-cell derived exosomes (i.e. EVs) can be used to modulate macrophage phenotype and inflammatory responses. Thus, the proposed combination would have involved applying a known biological function of stem cell-derived EVs to a composition of stem cell-derived EVs for the same purpose of modulating macrophage-mediated inflammation. One with ordinary skill in the art would have had a reasonable expectation of success in applying the teachings of Singla to the EVs composition of Levenberg because Singla demonstrates that such EVs can modulate macrophage phenotype and inflammatory responses. In other words, instant claims are combining prior art elements according to known methods to yield predictable results. See MPEP 2143 (I)(A).
Regarding claim 38, as discussed above with respect to claim 29, Levenberg et al in view of Rodrigues renders obvious the claimed EV’s composition. Neither Levenberg et al nor Rodrigues teach that the additional limitation concerning the IFN gamma.
Sun et al, however, supplement the cited prior arts by teaching the use of human umbilical cord mesenchymal stem cells-derived exosomes (huc-MSCs) for attenuating inflammation and report that exosomes promote polarization of bone-marrow-derived macrophages from M1 phenotype to M2 phenotype. Sun et al further teach that treatment with hucMSCs-derived EV reduces inflammatory cytokines including IFN-gamma, while increasing anti-inflammatory cytokines including IL-10. (See abstract, and Fig.4). Thus, Sun et al teach that stem cells-derived EV can regulate macrophage phenotype and inflammatory cytokine production, including IFN-gamma. Therefore, it would have been obvious to one with ordinary skill in the art at the time the invention was filed to apply the immunomodulatory teachings of Sun to the EV’s composition of Levenberg, as modified by Rodrigues, because Sun et al demonstrate that stem cells-derived EVs can be used to modulate macrophage polarization and attenuate inflammatory cytokines responses including IFN-gamma. In other words, claim 38 is combining prior art elements according to known methods to yield predictable results. See MPEP 2143 (I)(A). It is submitted that Sun teaches EV derived mesenchymal cells derived from umbilical cords rather than iPSCs- or ESCs- derived EVs. However, Sun et al is cited to provide evidence that stem-cell derived EVs were known to exert immunomodulatory effects on macrophages, including the reduction of IFN-gamma, while Levenberg in view of Rodrigues provide iPSCs- or ESCs-derived EV. One with ordinary skill in the art therefore would have been motivated to investigate and apply the known anti-inflammatory effect of stem-cell derived EVs taught by Sun to the iPSC-derived EVs of Levenberg.
Claims 41 and 57 are rejected under 35 U.S.C. 103 as being unpatentable over Levenberg et al in view of Rodrigues et al as applied to claims 18-20, 22, and 23-24 above, and further in view of Mardpour et al (Applied Materials, 2019), and Brennan et al ( Advanced Functional Material, 2020).
Regarding claims 41 and 57, the teachings of Levenberg et al and Rodrigues are set forth above. Levenberg et al teach a composition comprising extracellular vesicles derived from induced pluripotent stem cells (iPSCs) or embryonic stem cells (ESCs). However, Levenberg et al do not teach a composition that further comprises hydrogel.
Mardpour et al supplement Levenberg et al by demonstrating that embedding stem cell-derived EVs into hydrogel enables controlled and sustained release of the EVs, significantly improving their therapeutic effectiveness in vivo compared to free EVs injection which is characterized by rapid clearance rate. In other words, Mardpour et al teach that biocompatible hydrogel can serve as a sustained release carrier for exosomes to maintain their bioactivity at the injured tissue accelerating the healing process. Therefore, providing an ordinary skill in the art with the motivation to encapsulate stem cells-derived EVs with hydrogel prior to administration. (See abstract). Mardpour et al teach PEG-based hydrogel for the encapsulation of EVs and do not teach hyaluronic acid-based hydrogel. However, Brennan et al supplement Levenberg and Mardpour by stating that “free EVs administered via bolus injections are rapidly sequestered and cleared. However, encapsulating EVs with biomaterials (i.e. hydrogels) offers delivery platforms to enhance EV retention rates and healing efficacy”. (See abstract). Brennan et al also provide examples form preclinical models showing a composition comprising of mesenchymal stem cells conditioned medium (MSC-CM ) distributed within a hyaluronic acid (HA) hydrogel enhanced tissue regeneration after endometrial injury, while a hydrogel composed of HA and chondroitin sulfate incorporating MSC-CM achieved corneal wound healing. (See 2nd column, 2nd pargraph on page 7). It should be noted that MSC-CM also contains EVs. In other words, Brennan et al demonstrate the utility of using hyaluronic acid (HA) hydrogel to encapsulate EVs/CM prior injections to offer a controlled release platform, enhancing EVs/CM retention rates and healing efficacy.
Taken together, instant claims would have been obvious to one of ordinary skill in the art, as there was some teaching, suggestion, or motivation in the prior art that would have led one of ordinary skill to modify the prior art reference or to combine prior art reference teachings to arrive at the claimed invention. Levenberg et al teach a method for producing EVs from stem cells and a composition comprising the same, but do not teach a composition that further comprises hydrogel. Mardpour et al supplement Levenberg et al by demonstrating that embedding stem cell-derived extracellular vesicles into hydrogel enables controlled and sustained release, significantly improving their therapeutic effectiveness in vivo compared to free EVs injection. Brennan et al further provide scientific evidence from preclinical models wherein the use of hyaluronic acid-based hydrogel was successfully used to deliver EVs/CM to injured tissues, enhancing retention rates and healing efficacy. Therefore, an ordinary skill in the art who had reviewed Levenberg could have come across Mardpour and Brennan et al and immediately recognized the strong possibility of encapsulating the EVs of Levenberg with Hyaluronic acid-based hydrogel, as taught by Mardpour and Brennan, to overcome the rapid clearance rates, providing a controlled release platform for healing tissues. There is a reasonable expectation of success, when making a composition comprising EVs derived from stem cells, to encapsulate the EVs with hydrogel comprising hyaluronic acid, in that the encapsulated EVs would be efficiently delivered to the injured tissues.
Claims 18 and 58 is rejected under 35 U.S.C. 103 as being unpatentable over Levenberg et al in view of in view of Rigamonti et al (Stem Cell Reports, 2016).
Regarding claim 58, following the discussion of claim 18 above, Levenberg et al teach a method of producing EV by culturing pluripotent stem cells in dynamic culture and collecting EV from the culture medium. Levenberg, however, does not expressly teach culturing iPSCs or ESCs as three-dimensional spheroids.
Rigamonti et al supplement Levenberg et al by teaching culturing human pluripotent stem cells in 3D spheres in suspension culture. Specifically, Rigamonti et al describe 3D spheres formed from human ESCs and iPSCs and maintained in suspension culture. (See “hPSCs Adaption and Maintenance in Spinner Flasks” on page 1005). Therefore, it would have been prima facie obvious to one with ordinary skill in the art to culture the ESCs or iPSCs of Levenberg as three dimensional spheroids as taught by Rigamonti et al, because Rigamonti et al demonstrate that culturing these cells as spheroids was known and successful culture technique. Also, the use of such 3D culture configuration would have provided a known alternative culture format for expanding these cells in dynamic culture. Accordingly, such a modification would have involved using known culture technique for its known purpose, and with a reasonable expectation of success. In other words, claim 58 is also combining prior art elements according to known methods to yield predictable results. See MPEP 2143 (I)(A).
Response to Arguments
Applicants’ arguments filed 07/30/2026 have been fully considered but they are not persuasive.
Applicants argue that Levenberg et al’s method requires growing the cells in a fixed, three-dimensional porous scaffold through which medium is perfused in a flow chamber, whereas the claimed method as amended utilizes cells culturing as 3D spheroid or on microcarriers.
Applicant’s arguments with respect to claim 18 rejection under U.S.C. 35 102 have been considered and are persuasive. Therefore, the rejection is withdrawn. However, upon further consideration a new ground of rejection is made over Levenberg et al in view of Rodrigues et al.
Applicants also argue that the immunodulator properties recited in claims 29, 34, and 38, cannot be considered inherent in the EVs of Levenberg because Levenberg’s EV were generated from MSCs whereas Applicants EV are generated from iPSCs. Applicant further points to comparative experimental results allegedly demonstrating differences in the protein composition and immunomodulator properties of EV derived MSCs and iPSCs including differences in IL-10 and IFN-gamma production.
In view of Applicants’ argument and the evidence relied upon in the remarks, the office agrees that the immunodulator properties recited in claims 29, 34, and 38, cannot be considered inherent properties. Accordingly, the previous rejection of claims 29,34, and 38 based on inherency is withdrawn. However, the current rejection does not rely upon inherency. Applicants’ argument still does not overcome the new rejection based on the express teachings of the additional prior arts discussed above. (see the rejection above).
Applicants also further argue the instantly claimed methods for making EVs are different from Levenberg et al. and produce EVs with distinctly different properties from those of Levenberg et al. Applicants further argue that neither Mardpour nor Brennan remedies these alleged differences because Mardpour and Brennan are directed to hydrogel-based delivery of EV rather than to production of the claimed iPSCs or ESCs.
This is also not found persuasive because the rejection does not rely upon a
determination that the EVs of Levenberg inherently possess the allegedly different properties identified by Applicants. Rather, the rejection is based on the combined teachings of the applied references. As per the arguments against Mardpour and Brennan, Applicants appear to argue reference individually. 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., 800F.2d 1091, 231 USPQ 375 (Fed. Cir. 1986)
Conclusion
No claim is allowed.
Applicants’ amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). Applicants are 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 FATIMAH KHALAF MATALKAH whose telephone number is (703)756-5652. The examiner can normally be reached Monday-Friday,7:30 am-4:30 pm EST.
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/FATIMAH KHALAF MATALKAH/ Examiner, Art Unit 1638
/Tracy Vivlemore/ Supervisory Primary Examiner, Art Unit 1638
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