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
Election/Restrictions
Applicant's election without traverse the following species in the reply filed on 3/04/2026 is acknowledged.
MSC source: Umbilical and other perinatal tissues.
Dedifferentiation proteins: OCT4, NANOG, KLF-1, and SOX-2.
Injury signal: Interferon gamma.
Priority
Applicant' s claim for the benefit of a prior-filed parent provisional application 63351332, filed on 06/10/2022 under 35 U.S.C. 119(e) or under 35 U.S.C. 120, 121, or 365(c) is acknowledged.
The effective priority for the instant application is 06/10/2022.
Claims Status
Claims 2 and 5-9 are canceled, no claims are newly added, claims 13-17 and 19 have been withdrawn from consideration as being drawn to non-elected subject matter, and claims 1, 3-4, 10-12, 18 and 20 have been considered on the merits.
Claim Objections
Claim 1 is objected to because of the following informalities: the claim recites options a)-d) for cellular proteins. If the cellular proteins were designated differently (for example i)-iv)) this would improve clarity of the claim because the method steps are also designated a)-d).
Appropriate correction is required.
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, 3-4, 10, 12, 18 and 20 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 claim is indefinite because it recites the method step obtained a MSC cellular population” which implies the step has already been achieved and is inconsistent with the tense of the claim. Amending the claim to recite “obtaining a MSC cellular population” would overcome the rejection.
Note claims 3-4, 10-12, 18 and 20 depend from claim 1 and fail to cure the deficiencies.
Regarding claim 20: The term “stable” in claim 20 is a relative term which renders the claim indefinite. The term “stable” 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.
Claim Rejections - 35 USC § 103
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
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, 4, 11 and 12 are rejected under 35 U.S.C. 103 as being unpatentable over Jakob et al (Cells (2021) 10:33;1-18) in view of Okita et al (Nature Methods (2011) 8(5) 409-414), Kim et al (Cell (2008) 132;1049-1061), Nakagawa et al (Nature Biotechnology (2008) 26:1;101-106), Choi et al (eLife (2021) 10;1-27) and Alhashem et al (Virginia Commonwealth theses and Dissertations (2009) ROLES OF KRÜPPEL LIKE FACTORS KLF1, KLF2, AND KLF4 IN EMBRYONIC BETA-GLOBIN GENE EXPRESSION).
Regarding claim 1: The claim is drawn to a method of making conditioned media.
Claim 1 recites the active steps “obtained” (interpreted as obtaining), “introducing” and “inducing differentiation” and “culturing”.
The claim recites “dedifferentiating” in step b). Dedifferentiating is considered to be the result of introducing the claimed proteins into the cell.
The claim recites “to obtain a conditioned media”. This phrase is considered a result of the claimed method does not impart or imply an active step or structure to the claimed method.
Jakob teach reprogramming (dedifferentiating) MSCs into iPSCs and then differentiating the MSC derived iPSCs to MSCs (iP-MSCs) (abstract).
Jakob teach an MSC cell population is obtained; isolation of human tissue-resident MSC from nasal mucosa was performed (p3 ¶1).
Jakob further teach introducing cellular proteins for dedifferentiation; MSCs were electroporated with the plasmids pCXLE-hSK, pCXLE-hUL and pCXLEhOct3/4-shp53-Fas (Jakob p3 ¶3). Jakob teach after the reprogramming procedure (electroporation with the plasmids), colonies with iPS-like morphology gave rise to different step cell lines (p4 ¶1).
Jakob teach the plasmids used for reprogramming are described by Okita. As evidenced by Okita; pCXLE-hSK is an expression plasmid which encodes Sox2 and KLF4 proteins and pCXLE-hOct3/4-shp53 is an expression vector which encodes Oct 3/4 protein (Okita Figure 1 b). This reads on introducing cellular proteins OCT4 and SOX-2 as required by the instant claim 1.
Jakob teach inducing differentiation of the dedifferentiated MSC cells into iP-MSCs (IPSC derived MSCs); Jakob teach MSC-derived iPSCs were cultured using gelatin-coated flat culture and mesenchymal stem cell growth medium 2 (p8 ¶1). Jakob further teach that after almost 4 weeks of culturing the morphology was similar to regular MSCs (p8 ¶1). Jakob also teach the iP-MSCs have MSC properties and show the functional and phenotypic characteristics of the original MSCs, demonstrating the differentiation of the MSC derived iPSCs to (p9 ¶2).
Jakob teach culturing the cells in mesenchymal stem cell growth medium 2 and that after almost 4 weeks the cells appear to be MSCs (p8 ¶1). This reads on culturing said MSCs in a liquid media as required by the claim, as required by the claim.
Because the method of Jakob obtains MSCs in a liquid media, a conditioned media is obtained by the method of Jakob.
While Jakob teach introducing SOX2, KLF4 and OCT4, for dedifferentiation of MSCs, Jakob do not teach introducing KLF1 and NANOG.
Okita teach introducing OCT4, NANOG, KLF4 and SOX2 into MSCs for dedifferentiation; expression plasmids are introduced (electroporated) into the cells (p413 col 1 ¶4). Okita further teach that the expression plasmids in mixture Y1 encode OCT4, NANOG, KLF4 and SOX2 (Figure 1a and Supplementary Table 2).
It would have been obvious to one of ordinary skill in the art to modify the method of Jakob, drawn to generating conditioned media from culturing MSCs that were dedifferentiated by introducing OCT4, SOX-2, and KLF4 and then differentiated into MSCs with the teachings of Okita, to introduce the NANOG protein for dedifferentiation.
One would have been motivated to modify the method of Jakob with the teaching of Okita, to also introduce the NANOG protein for dedifferentiation because Kim teach Nanog is considered a core pluripotency factor and promotes ES cell self-renewal (p1049 col1 ¶2). Kim also teach that iPS cells that are reprogramed by forced expression of transcriptional factors (Oct4, Sox2, Klf4 and c-My) have enhanced quality when selected for expression of Nanog (p1049 col1 ¶1).
One would have had a reasonable expectation of success because the transcription factors Oct4, Sox2 and Nanog are well-known pluripotency factors.
While Jakob and Okita teach dedifferentiation of MSCs by introducing the KLF-4 protein, Jakob and Okita are silent on introducing the KLF-1 protein.
Nakagawa teach generating iPSCs from somatic cells using family members of the four transcription factors Oct3/4, Sox2, KLF4 and c-Myc (p101 col1 ¶1). Nakagawa teach KLF1 can also generate iPS cells (p101 col2 ¶1).
It would have been obvious to one of ordinary skill in the art to modify the method of Jakob and Okita, drawn to generating conditioned media from culturing MSCs that were dedifferentiated by introducing OCT4, SOX-2, KLF4 and NANOG with the teachings of Nakagawa, that introducing KLF1 is effective for dedifferentiation.
One would have been motivated to modify the method of Jakob and Okita with the teachings of Nakagawa, to include the KLF1 protein in the dedifferentiation step, because Nakagawa teach KLF1 can generate iPSCs. Furthermore, Alhashem teach KLF1 and KLF3 have high homology and overlapping function (p15 ¶1). Alhashem further teach KLF1 and 4 belong to the same subgroup and have structural similarity which supports that the proteins have functional redundancy (p20 ¶3).
One would have had a reasonable expectation of success because methods are drawn to dedifferentiation factors that are well known and well characterized in the art.
Regarding claim 4: The claim recites “wherein said MSC cellular population expresses interleukin-3 receptor”. The phrase is considered an intended result of the method and does not confer a structure or active step to the claimed method.
Regarding claim 11: The teachings of Jakob are discussed above. Jakob teach that the cellular proteins are introduced using a plasmid-based integration-free protocol (p3 ¶3). Jakob introduces the cellular proteins to dedifferentiate the cells. Proteins which dedifferentiate cells are considered pluripotency inducing genes.
Introduction of plasmids is considered to read on “mRNA is introduced” because in order for a protein to be expressed from a plasmid-based system the protein coding sequence must be transcribed to mRNA before the mRNA is translated to protein by cellular machinery.
Regarding claim 20: The claim recites the phrase “where the dedifferentiated MSC cellular population has stable karyotype”. This is considered an intended result and does not confer or imply active steps or structure to the claimed method.
Claim 3 is rejected under 35 U.S.C. 103 as being unpatentable over being unpatentable over Jakob et al, Okita et al, Kim et al, Nakagawa et al, and Choi et al as applied to claims 1, 4, 11 and 12 above, and further in view of Nagamura-Inoue et al (WJSC (2014) 6:2;195-202).
Regarding claim 3: The teachings of Jakob Okita are discussed supra. While Jakob teach MSCs are sourced from nasal mucosa, Jakob do not teach the MSCs are sourced from umbilical and other perinatal tissues. Jakob further teach the MSCs that were differentiated from mucosal MSC derived iPSCs reacquired the major functional properties of the mucosal MSCs (p14 ¶4).
Nagamura-Inoue(2014) teach that mesenchymal stem cells can be isolated from the bone marrow, mobilized peripheral blood, cord blood, umbilical cord (UC), placenta, adipose tissue, dental pulp, fetal liver and lungs (p195 col1 ¶1). Nagamura-Inoue teach that collection of UC-MSCs (umbilical cord MSCs) is noninvasive, and have immunomodulatory properties (p195/196 col2/1 ¶1/1). Nagamura-Inoue teach that UC-MSCs have immunosuppressive properties both in vitro and in vivo, and that the immunosuppressive effects of MSCs is a popular property of MSCs for potential clinical use (p198/199 col2/1 ¶4/2).
It would have been obvious to one of ordinary skill in the art to modify the method of Jakob, drawn to generating conditioned media from culturing MSCs that were generated from iPSCs from dedifferentiated mucosal MSCs by using umbilical MSCs as a source of MSCs, as taught by Nagamura-Inoue.
One would be motivated to modify the method of Jakob with the teaching of Nagamura-Inoue, to source MSCs from umbilical tissues because Nagamura-Inoue teach UC-MSCs have desirable immunosuppressive properties and Jakob teach MSCs that are generated from iPSCs which have been derived from dedifferentiated MSCs show functional and phenotypic characteristics of the original MSCs (Jakob p9 ¶1).
One would have had a reasonable expectation of success because the inventions are both drawn to MSCs.
Claim 10 is rejected under 35 U.S.C. 103 as being unpatentable over being unpatentable over Jakob et al, Okita et al, Kim et al, Nakagawa et al, and Choi et al as applied to claims 1, 4, 11 and 12 above, and further in view of Kubara et al (Stem Cell Reports (2018) 11;380-394).
Regarding claim 10: The teachings of Jakob, Okita and Nakagawa are discussed supra. While Jakob, Okita and Nakagawa teach dedifferentiation by introducing the proteins OCT4, NANOG, KLF-1 and SOX-2, they do not teach administering k-RAS to dedifferentiate the MSC population.
Kubara teach oncogenic KRAS is associated with enhanced stemness in some cancers such as colon cancer and pancreatic cancers (p380 col1 ¶1). Kubara teach that the oncogenic KRAS mutant G13C enhances retention of self-renewal capacity in iPSCs (p385 col2 ¶3). Kubara further teach that KRAS G13C is an activated version of the protein which confers retained self-renewal of undifferentiated cells (p390 col1 ¶2). Kubara teach that KRAS has a significant role in the maintenance of iPSC stemness which affects aspects such as self-renewal and differentiation (p391 col1 ¶2).
Sasine teach overexpression of wild-type KRAS promotes hematopoietic stem cell expansion and self-renewal without inducing malignancy (title).
It would have been obvious to one of ordinary skill in the art to modify the method of Jakob, Okita, and Nakagawa drawn to generating conditioned media from culturing MSCs that were generated from iPSCs from dedifferentiated MSCs by dedifferentiating the MSCs using OCT4, NANOG, KLF-1 and SOX2 with the teachings of Kubara and Sasine, to over express (administer) KRAS for dedifferentiation because Kubara teach that KRAS activity enhances retention of self-renewal capacity in iPSCs and Sasine teach overexpression of wild type KRAS promotes stem cell expansion and self-renewal.
One would have been motivated to include KRAS in the dedifferentiation step taught by Jakob, Okita and XXX because Sasine teach wild-type KRAS can expand normal stem cells and progenitors without causing malignancy (pS424 col2 ¶2).
One would have had a reasonable expectation of success in including KRAS in the dedifferentiation step because Kubara teach KRAS enhances iPSC self-renewal and Sasine teach overexpression of wild KRAS doesn’t cause malignant disease.
Claims 12 and 18 are rejected under 35 U.S.C. 103 as being unpatentable over being unpatentable over Jakob et al, Okita et al, Kim et al, Nakagawa et al, and Choi et al as applied to claims 1, 4, 11 and 12 above, and further in view of Yang et al (Cell Death and Disease (2020) 11:1-12).
Regarding claims 12 and 18: The teachings of Jakob, Okita, and Nakagawa are discussed supra. Jakob, Okita and Nakagawa do not teach the MSCs are activated with interferon gamma.
Yang teach mesenchymal stem cells have tremendous therapeutic effects for a variety of diseases due to strong immunomodulatory ability (p1 col1 ¶1). Yang further teach the immunomodulatory property of MSCs is largely attributed to secreted paracrine effectors (p1 col1 ¶1). One of ordinary skill in the art would understand that secreted effectors are secreted into the media in which cells are cultured. Yang teach that administration of exosomes derived from MSCs is reported to ameliorate disease phenotypes in animal models (p1 col2 ¶1). Yang teach that INF-gamma treatment increases the number of exosomes derived from MSCs and that exosomes derived from INF-gamma-activated MSCs show higher efficacy to treat colitis (p2 col1 ¶3, p7 col1 ¶1).
It would have been obvious to one of ordinary skill in the art to modify the method of Jakob, Okita, and Nakagawa drawn to generating conditioned media from culturing MSCs that were generated from iPSCs from dedifferentiated MSCs by activating the MSCs with INF-gamma as taught by Yang.
One would have been motivated to include activate the MSCs with INF-gamma, as taught by Yang, because Yang teach that exosomes, which are secreted into MSC media, have greater numbers and improved therapeutic properties compared to exosomes from MSCs that aren’t activated with INF-gamma.
One would have had a reasonable expectation of success in using INF-gamma to activate MSCs in the method of Jakob, Okita and XXX because both inventions are drawn to MSCs and Jakob teach MSCs derived from IPSCs that are from dedifferentiated MSCs retain functional and phenotypic characteristics of the original MSCs (Jakob p9 ¶1).
Conclusion
No claims are allowed.
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/ANDREA LYNNE MORRIS SPENCER/Examiner, Art Unit 1631
/TAEYOON KIM/Primary Examiner, Art Unit 1631
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