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 provisional U.S. Application No. 63/489,628 (filed 03/10/2023).
Election/Restrictions
Applicant’s election of Group I in the reply filed on 07/13/2026 is acknowledged. Because applicant did not distinctly and specifically point out the supposed errors in the restriction requirement, the election has been treated as an election without traverse (MPEP § 818.01(a)).
Claims 1-20 are pending in the instant application, with claims 15-20 withdrawn as directed to an invention nonelected.
Claims 1-14 read on the elected invention and are examined on the merits herein.
Claim Objections
Claim 1 is objected to because of the following informalities: Line 1 of claim 1 recites the term hPSCs; while the use of this term is well known to those in the art as an abbreviation for human pluripotent stem cells, it is proper for the initial introduction of a term to be spelled out in full, with the abbreviation subsequently denoted in enclosed parentheses.
Appropriate correction is required.
Claim Interpretation
The following comments are made to establish broadest reasonable interpretation for the record.
Regarding claim 1: This claim recites, “A method for generating blastoids from naïve hPSCs, the method comprising: exposing said naïve hPSCs to 5i/L/A; aggregating the exposed hPSCs in N2B27; and treating the aggregated hPSCs with a simplified blastocyst induction medium (“BIM”).”
The term blastoid is interpreted as a cell aggregate resembling the pre-implantation blastocyst; this interpretation is supported by the instant specification (pg. 40; lines 22-25).
As evidenced by Dodsworth, et al. (Stem Cell. 2015), naïve or ground state pluripotency is a cellular state in vitro which resembles cells of the pre-implantation epiblast in vivo, while hPSCs correspond to a later or “primed” stage of embryonic development (Abstract); for instance, mouse epiblast stem cells (mEpiSCs) are sourced from post-implantation epiblast cells and are termed primed due to their inability to integrate into a preimplantation blastocyst, although they can still be differentiated into all three germ layers in vitro (pg. 3181; col. 1, par. 1). Thus, the term naïve hPSCs is interpreted to mean PSCs which can be characterized by their resemblance to cells of the pre-implantation epiblast.
As supported by the “Experimental Model and Subject Details” section of the specification (beginning on pg. 55), the terms 5i/L/A and N2B27 are interpreted as referring to culture media; however, there is no clear definition set forth in the instant disclosure for any of the media recited in claim 1. The 5i/L/A medium as it pertains to 35 U.S.C. 112(b) is discussed below; however, as evidenced by Theunissen, et al. (Cell Stem Cell. 2014), this limitation appears to refer to a culture medium comprising N2B27 medium, a mitogen-activated protein kinase (MEK) inhibitor, a glycogen synthase kinase 3 (GSK3) inhibitor, a Rho-associated protein kinase (ROCK) inhibitor, a BRAF (or B-Raf) inhibitor, an Src inhibitor, Activin, and human leukemia inhibitory factor (hLIF) (Fig. 5A). Under broadest reasonable interpretation, the N2B27 medium limitation is interpreted as referring to a basal medium supplement with N2 and B27®, and the blastocyst induction medium limitation is interpreted as referring to any culture medium used for or capable of inducing blastocyst-like characteristics in a cell. The simplified portion of the simplified blastocyst induction medium is not given any patentable weight.
Regarding claims 4-12: Many of these claims are directed to so-called negative limitations, or limitations which are directed to components excluded from the method of the instant invention. This interpretation section will use the limitations recited in claims 4-5 for illustrative purposes in order to establish broadest reasonable interpretation for the negative limitations recited in claims 4-12.
Claim 4 recites the limitation wherein a GSK3α/β inhibitor is omitted; this is interpreted as referring to at least one of the many GSK3α and/or β inhibitors available being omitted from the claimed method. This interpretation is supported by the use of the indefinite article “a” preceding recitation of the singular “GSK3α/β inhibitor”, as opposed to the use of a determiner or quantifier such as “any”, or as opposed to use of the plural form “GSK3α/β inhibitors” (e.g., “…wherein GSK3α/β inhibitors are omitted”); either of these would clearly set forth that the method omits use of any and all GSK3α/β inhibitors. As such, under broadest reasonable, it would be remiss to restrict the scope of this limitation to only methods which do not use any GSK3α/β inhibitor at all. Further, claim 5, which depends from claim 4, specifies the omitted GSK3α/β inhibitor is CHIR99021. In the examples disclosed in the specification, the media recipes disclosed for 5i/L/A (pg. 55; line 15) and BIM (pg. 57; line 1) both include IM-12, a GSK3α/β inhibitor. Indeed, Fig. 2A discloses eight different conditions tested for the claimed method, wherein all eight conditions include IM-12.
Therefore, the negative limitations recited in the instant claims are interpreted as omitting at least one of the respective inhibitors recited therein.
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 1-14 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: This claim suffers from several points of indefiniteness. As a first matter, it is not clear what is meant by 5i/L/A medium. The specification discloses a recipe (pg. 55; line 15), but it is not clear if this recipe is a limiting definition. Likewise, it’s unclear if the recipes for N2B27 medium (pg. 56; line 29) and blastocyst induction medium (pg. 57; line 1) are meant to be limiting definitions.
Further, the recipes include trademarked components. Per MPEP 2173.05(u), 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.
Thus, as the recipes disclosed for the media recite trademarked components, it’s ultimately unclear what is in the recipe set forth in the specification for the media limitations recited in claim 1.
As a second matter, and following the interpretation for N2B27 medium set forth above, the trade name “N2B27” is used to identify/describe a culture medium; specifically, “N2B27” is used in the art to describe a culture medium which comprises a 1:1 mixture of DMEM/F12 media: Neurobasal® media, supplemented with N2 and B27®. However, as B27® supplement is a registered trademark, this limitation is necessarily directed to a particular trademarked product, rendering the claim indefinite.
Lastly, it’s not clear what is meant by the term “simplified” in the simplified blastocyst induction medium limitation recited in the instant claim, as “simplified” is a subjective term. Simplified in comparison to what? The BIM recipe provided on pg. 57 discloses some – but not all – of the media components for BIM are at a lower concentration than in the recipes provided for 5i/L/A (pg. 55) and N2B27 (pg. 56); does “simplified” refer to a reduced concentration of said components? Further, the BIM recipe discloses fewer media components overall in comparison to 5i/L/A, but not in comparison to N2B27; does “simplified” refer to a blastocyst induction media which is “simpler” than the 5i/L/A medium?
Overall, the metes and bounds of claim 1 are not clearly or precisely defined, rendering the claim indefinite.
Claims 2-14 depend from claim 1, inherit its deficiencies, and are likewise rejected as indefinite.
Regarding claim 9: It is not clear what is meant by “Activin/Nodal/TGFβ inhibitor”. Does this refer to a pan inhibitor of the Activin/Nodal/TGFβ pathway? Is this a Markush group, wherein the limitation is considered satisfied if the method excludes use of an Activin inhibitor but includes use of a TGFβ inhibitor? Does the “inhibitor” portion refer solely to the TGFβ portion, and Activin and Nodal refer to the respective proteins? As currently written, the metes and bounds are not clearly or precisely defined.
Further, there is insufficient antecedent basis for the “A83-01” limitation in the instant claim; thus, it is unclear what is meant by “…an amount of A83-01 is increased.” For purposes of examination, and under broadest reasonable interpretation, an amount of A83-01 is increased is interpreted as a medium comprising A83-01 used in the method of claim 1, wherein the medium comprises A83-01 in an amount which is greater than the amount contained in a preceding media used in the method; e.g., blastocyst induction medium comprising A83-01 while 5i/L/A and/or N2B27 do not.
Claim 10 depends from claim 9 inherits its deficiencies, and is likewise rejected as indefinite. It is noted for the record that claim 10 only inherits the deficiencies related to the an amount of A83-01 is increased limitation, as SB431542 is a pan inhibitor of the Activin/Nodal/TGFβ pathway.
Regarding claim 11: There is insufficient antecedent basis for the “Activin” and “FGF2” limitations recited in this claim. Claim 12 depends from claim 11 and is likewise rejected as indefinite.
Regarding claim 13: There is insufficient antecedent basis for the “sodium pyruvate” limitation recited in this claim.
Regarding claim 14: It is not clear what the “a seeding density is increased” limitation is referring to. The method of claim 1 does not recite any limitations directed to seeding or plating the hPSCs. While it would be understood by a person having ordinary skill in the art a method comprising use of hPSCs necessarily includes seeding these cells, it would not be understood how or at which step the limitation recited in claim 14 is integrated into said method. Are the hPSCs replated in one of the steps and the seeding density is increased at that step? Is it simply increased compared to a standard or recommended seeding density known to those in the art? The metes and bounds are not clearly or precisely defined, rendering the claim indefinite.
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.
Claim 1 is rejected under 35 U.S.C. 102(a)(1) as being anticipated by Yu, et al. (Nature. 2021).
Yu, et al. (hereinafter Yu) teaches the generation of hPSC-derived blastocyst-like structures, termed blastoids, in vitro (Abstract).
Regarding claim 1: Yu teaches a method for the generation of human blastoids wherein naïve WIBR3 human embryonic stem cells (hESCs) are cultured in 6-well plates on mitotically inactivated mouse embryonic fibroblast (MEF) feeders in 5i/L/A medium until reaching 60-70% confluency (pg. 1 of “Methods”: col. 1, par. 6 to col. 2, par. 1; and col. 2, par. 3). This anticipates:
the method for generating blastoids from naive hPSCs, the method comprising exposing the naive hPSCs to 5i/L/A limitations recited in claim 1.
Yu teaches the 5i/L/A-cultured WIBR3 hESCs are then seeded in 5i/L/A medium supplemented with 5 µM Y-27632 in AggreWell 24-well plates for around 12h of culture until aggregates are formed (pg. 2 of “Methods”; col. 1, par. 1). As the 5i/L/A medium comprises 1:1 DMEM/F12: Neurobasal® medium supplemented with N2 supplement and B27® supplement (pg. 1 of “Methods”; col. 2, par. 1), under broadest reasonable interpretation, this reads on the N2B27 limitation recited in line 4 of claim 1. Thus, this anticipates:
the aggregating the exposed hPSCs in N2B27 limitation recited in claim 1.
Yu teaches the medium is then changed to trophoblast differentiation medium (TDM) and cells are cultured until cavity structures appear (approximately 6-7 days), with media exchanges every 2 days; after cavity structures appear, the medium is replaced with hypoblast differentiation medium (HDM) and cultured for an additional 1-2 days for blastoid formation (pg. 2 of “Methods”; col. 1, pars. 1-3). As the TDM reads on blastocyst induction medium, this anticipates:
the remaining treating the aggregated hPSCs with a simplified blastocyst induction medium (“BIM”) limitation recited in claim 1.
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.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
Claims 1-2 are rejected under 35 U.S.C. 103 as being unpatentable over Yu, et al. (Nature. 2021).
The teachings of Yu are set forth above; claim 1 is anticipated by the same.
Regarding claim 2: Following the above discussion, under broadest reasonable interpretation, the step taught by Yu wherein the cells are cultured in TDM for approximately 6-7 days until cavity structures appear reads on the treating is carried out for about 5 days limitation recited in claim 2.
Yu does not teach the remaining time period limitations recited in the instant claim.
However, Yu explicitly teaches for the generation of blastoids, the initial culturing step is performed until the naïve PSCs are 60-70% confluent (pg. 1 of “Methods”; col. 2; par. 3); additionally, Yu clearly teaches the subsequent culturing step is performed for a time period effective to form aggregates (pg. 2 of “Methods”; col. 1, par. 1). That means the conditions necessarily to achieve 60-70% confluency and aggregating of the cells were result effective variables. Result effective variables would be optimized by routine experimentation by one having ordinary skill in the art; see MPEP 2144.05(II)(A).
This renders obvious the remaining the exposing is carried out for about 1 day and the aggregating is carried out for about 2 days limitations recited in claim 2.
Claims 1, 3, 6-7, and 14 are rejected under 35 U.S.C. 103 as being unpatentable over Yu, et al. (Nature. 2021) in view of Theunissen, et al. (US 2021/0230538).
The teachings of Yu are set forth above; claim 1 is anticipated by the same.
Theunissen, et al. (hereinafter Theunissen) teaches compounds and methods useful in changing the pluripotency state of a vertebrate cell to a more naïve state (Abstract).
Regarding claim 3: Following the above discussion, Yu does not teach the cell line of the instant claim.
However, Theunissen teaches naïve WIBR3 OCT4-ΔPE-GFP hESCs (par. 0101); these read on the WIBR3 OCT4-GFP naive hPSCs of the instant claim.
It would have been prima facie obvious for a person having ordinary skill in the art to have modified the method of Yu by substituting the naïve WIBR3 hESCs with the naïve WIBR3 OCT4-ΔPE-GFP hESCs of Theunissen. This conclusion of obviousness is based on the ‘substitution rationale’. The use of the naïve WIBR3 OCT4-ΔPE-GFP hESCs in place of the naïve WIBR3 hESCs is a predictable use of prior art elements according to their established functions as embryonic stem cells derived from the WIBR3 cell line in a naïve state of pluripotency, leading to the predictable result of the generation of blastoids. This rationale aligns with the principle of a simple substitution of one known element for another to obtain predictable results; see MPEP 2143(I)(B).
Thus, the modified method of Yu renders obvious the wherein the naïve hPSCs are WIBR3 OCT4-GFP naïve hPSCs limitation recited in claim 3.
Regarding claims 6-7: Following the above discussion, Yu does teach the inclusion of BRAF inhibitor SB590885 in the 5i/L/A medium (pg. 1 of “Methods”; col. 2, par. 1) and the trophoblast differentiation medium (pg. 2 of “Methods”; col. 1, par. 1).
However, Theunissen teaches BRAF inhibitors SB590885 and GDC-0879 are both appropriate BRAF inhibitors for use with naïve hESCs (pars. 0005, 0117, 0122); thus, Theunissen teaches SB590885 and GDC-0879 as functional equivalents.
Therefore, it would have been prima facie obvious to a person having ordinary skill in the art to have further modified the method of Yu by substituting SB590885 with GDC-0879 in the media. This conclusion of obviousness is based on the ‘substitution rationale’. The use of the GDC-0879 in place of the SB590885 is a predictable use of prior art elements according to their established functions as BRAF inhibitors, leading to the predictable result of the inhibition of BRAF. This rationale aligns with the principle of a simple substitution of one known element for another to obtain predictable results; see MPEP 2143(I)(B). Thus, this modified method of Yu renders obvious:
the wherein a BRAF inhibitor is omitted limitation recited in claim 6; and
the wherein the omitted BRAF inhibitor is SB590885 limitation recited in claim 7.
Regarding claim 14: The indefiniteness issues of the instant claim are set forth above. However, the instant specification discloses working examples; see “A protocol for efficient blastoid formation from naïve PSCs derived in 5i/L/A” (starting at pg. 41; line 24), and “Generation of blastoids” (starting at pg. 56; line 16). In these examples, after the initial seeding, the hPSCs do not appear to be re-plated in any subsequent steps; further, the former explicitly states blastoids were originally generated at low efficiency (~5-10%) consistent with the original report (“Condition 1”), and thus different conditions were optimized to increase efficiency, including a higher seeding density for “Condition 8” as seen in Figs. 2A-C (pg. 41, line 25 to pg. 42, line 11). Therefore, it is reasonable to conclude the limitation of claim 14 is referring to a higher seeding density than the seeding density used for “Condition 1”; however, said seeding density is not readily found in the disclosure.
Even so, Yu discloses that while approximately 30,000 cells are seeded into the wells of the AggreWell plate, this density is for the WIBR3 cells; Yu explicitly states the starting cell number needs to be optimized for other cell lines (pg. 2 of “Methods”; col. 1, par. 1).
Therefore, following the above discussion, it would have been prima facie obvious to a person having ordinary skill in the art to have further modified the method of Yu by increasing the cell seeding density for the WIBR3 OCT4-GFP hESCs of Theunissen. This conclusion of obviousness is based on the ‘obvious to try rationale’. Yu clearly teaches the seeding density needs to be optimized for different cell lines, and there are only three possible outcomes for a known seeding density which is being optimized by a skilled artisan: a decreased seeding density, an equivalent seeding density, or an increased seeding density. Therefore, it would be obvious for the skilled artisan to try all three of these possible conditions, which is well within the purview of those in the art. This rationale aligns with choosing from a finite number of identified, predictable solutions with a reasonable expectation of success; see MPEP 2143(I)(E).
This renders obvious the wherein a seeding density is increased limitation recited in claim 14.
Claims 1 and 4-5 are rejected under 35 U.S.C. 103 as being unpatentable over Yu, et al. (Nature. 2021) in view of Zhang (US 2020/0277568), as evidenced by Force and Woodgett (J Biol Chem. 2009).
The teachings of Yu are set forth above; claim 1 is anticipated by the same.
Zhang teaches a small molecule composition for reprogramming cells (Abstract).
Regarding claims 4-5: Following the above discussion, Yu does teach the inclusion of CHIR99021 in the tropohoblast differentiation medium (pg. 2 of “Methods”; col. 1, par. 1).
However, Zhang teaches GSK3β inhibitors CHIR99021, TWS119, and BIO for inhibiting GSK3β signaling in the transdifferentiation of fibroblasts (pars. 0005, 0026). As evidenced by Force and Woodgett, small molecule inhibitor BIO inhibits both -α and -β isoforms of GSK3 (pg. 9643; col. 2, par. 2). Thus, CHIR99021 and BIO are functional equivalents.
Therefore, it would have been prima facie obvious to a person having ordinary skill in the art to have further modified the method of Yu by substituting CHIR99021 with BIO in the trophoblast differentiation medium. This conclusion of obviousness is based on the ‘substitution rationale’. The use of BIO in place of CHIR99021 is a predictable use of prior art elements according to their established functions as GSK3α/β inhibitors, leading to the predictable result of the inhibition of GSK3 signaling. This rationale aligns with the principle of a simple substitution of one known element for another to obtain predictable results; see MPEP 2143(I)(B). Thus, this modified method of Yu renders obvious:
the wherein a GSK3α/β inhibitor is omitted limitation recited in claim 4; and
the wherein the omitted GSK3α/β inhibitor is CHIR99021 limitation recited in claim 5.
Claims 1 and 8 are rejected under 35 U.S.C. 103 as being unpatentable over Yu, et al. (Nature. 2021) in view of Blanchard, et al. (J Biol Chem. 2000).
The teachings of Yu are set forth above; claim 1 is anticipated by the same.
Blanchard, et al. (hereinafter Blanchard) teaches leukemia inhibitory factor (LIF) signaling via the heterodimeric receptor complex comprising the LIF receptor α subunit (LIFRα) and the common signal transducing subunit for interleukin (IL)-6 cytokine receptors, gp130 (Abstract).
Regarding claim 8: Following the above discussion, Yu does teach the inclusion of rhLIF in the 5i/L/A medium (pg. 1 of “Methods”; col. 2, par. 1) and the trophoblast differentiation medium (pg. 2 of “Methods”; col. 1, par. 1).
However, Blanchard teaches leukemia inhibitory factor (LIF) signals via the heterodimeric receptor complex comprising the LIF receptor α subunit (LIFRα) and the common signal transducing subunit for interleukin (IL)-6 cytokine receptors, gp130 (Abstract). LIFRα is a transmembrane protein with low affinity for LIF until combined with the gp130 subunit to form the high affinity LIF receptor complex, wherein LIF binding to LIFRα induces heterodimerization with gp130 (pg. 28793; col. 2, pars. 2-3). Oncostatin M (OSM) is a cytokine closely related to LIF (Abstract); both are members of the IL-6-type family of cytokines (pg. 28793; col. 1, par. 1). OSM also uses LIFRα and gp130 subunits to form a high affinity OSM receptor complex (pg. 28793; col. 2, par. 2).
Therefore, it would have been prima facie obvious to a person having ordinary skill in the art to have further modified the method of Yu by substituting rhLIF with OSM in the media. This conclusion of obviousness is based on the ‘substitution rationale’. The use of OSM in place of rhLIF is a predictable use of prior art elements according to their established functions as members of the IL-6-type family of cytokines, leading to the predictable result of ligand binding to LIFRα and forming a heterodimer receptor complex with gp130. This rationale aligns with the principle of a simple substitution of one known element for another to obtain predictable results; see MPEP 2143(I)(B).
Thus, this modified method of Yu renders obvious the wherein a leukemia inhibitory factor (LIF) is omitted limitation recited in claim 8.
Claims 1 and 9-10 are rejected under 35 U.S.C. 103 as being unpatentable over Yu, et al. (Nature. 2021) in view of Keller and Protze (US 2020/0353011).
The teachings of Yu are set forth above; claim 1 is anticipated by the same.
Keller and Protze (hereinafter Keller) teaches inhibitors of the activin/nodal/TGFβ pathway (par. 0063).
Regarding claims 9-10: Following the above discussion, Yu teaches the trophoblast differentiation medium comprises A83-01, while the 5i/L/A medium does not (pgs. 1 and 2 of “Methods”); this reads on the an amount of A83-01 is increased limitation recited in claim 9.
Yu does teach the inclusion of SB431542 in the trophoblast differentiation medium (pg. 2 of “Methods”; col. 1, par. 1).
However, Keller teaches SB431542 and SB505124 are both small molecule inhibitors of the Activin/Nodal/TGFβ pathway (par. 0063).
Therefore, it would have been prima facie obvious to a person having ordinary skill in the art to have further modified the method of Yu by substituting SB431542 with SB505124 in the trophoblast differentiation medium. This conclusion of obviousness is based on the ‘substitution rationale’. The use of SB505124 in place of SB431542 is a predictable use of prior art elements according to their established functions as small molecule inhibitors of the Activin/Nodal/TGFβ pathway, leading to the predictable result of inhibition of said pathway. This rationale aligns with the principle of a simple substitution of one known element for another to obtain predictable results; see MPEP 2143(I)(B). Thus, this modified method of Yu renders obvious:
the remaining wherein an Activin/Nodal/TGFβ inhibitor is omitted limitation recited in claim 9; and
the wherein the omitted Activin/Nodal/TGFβ inhibitor is SB431542 limitation recited in claim 10.
Claims 1 and 11-12 are rejected under 35 U.S.C. 103 as being unpatentable over Yu, et al. (Nature. 2021) in view of Parent, et al. (WO 2021/222297), Kim, et al. (Int J Mol Sci. 2021), and Theunissen, et al. (Cell Stem Cell. 2014).
The teachings of Yu are set forth above; claim 1 is anticipated by the same.
Parent, et al. (hereinafter Parent) teaches functional analogs of Activin A (par. 0188).
Kim, et al. (hereinafter Kim) teaches human DJ-1 in comparison to human fibroblast growth factor 2 in embryonic stem cell culture (Abstract).
Theunissen, et al. teaches the derivation of 5i/L/A medium (see e.g. pg. 479).
Regarding claims 11-12: Following the above discussion, Yu does teach the inclusion of bFGF (i.e., FGF2) in the hypoblast differentiation medium (pg. 620; col. 2, par. 1), as well as the inclusion of Activin A in the 5i/L/A medium (pg. 1 of “Methods”; col. 2, par. 1) and the trophoblast differentiation medium (pg. 2 of “Methods”; col. 1, par. 1).
However, Parent teaches small molecule IDE1 is a functional analog of Activin A (par. 0188).
Additionally, Kim discloses human DJ-1 (hDJ-1) as a replacement for human fibroblast growth factor 2 (hFGF2) in hPSC cell culture as both bind to FGFR-1 (pg. 2; pars. 1-2). Kim teaches hDJ-1 can maintain the pluripotency of hPSCs in feeder-free conditions without the need for hFGF2 in the culture media (“Conclusions”; pg. 11).
Therefore, it would have been prima facie obvious to a person having ordinary skill in the art to have further modified the method of Yu by (1) substituting Activin A with IDE1 in the 5i/L/A and trophoblast differentiation media, and (2) substituting FGF2 with hDJ-1 in the hypoblast differentiation medium. This conclusion of obviousness is based on the ‘substitution rationale’. Theunissen, et al. teaches the supplementation of 5i/L medium with FGF and Activin A enhanced the kinetics of OCT4-ΔPE-GFP induction, indicating the additional growth factor support prolongs the time window wherein human ESCs are amenable to a naïve state of pluripotency (pg. 477; col. 1, par. 2). Thus, a skilled artisan would have a reasonable expectation that replacing Activin A with its functional analog IDE1 (as taught by Parent; par. 0188) and replacing FGF2 with another FGFR-1 ligand hDJ-1 (as taught by Kim; pgs. 2 and 11) would maintain the naïve state of pluripotency in the hESCs required for the generation of blastoids. This rationale aligns with the principle of a simple substitution of one known element for another to obtain predictable results; see MPEP 2143(I)(B). Thus, this modified method of Yu renders obvious:
the wherein Activin and FGF2 are omitted limitation recited in claim 11; and
the wherein the Activin and the FGF2 are omitted from the aggregating step limitation recited in claim 12.
For clarity of record, it is noted the unmodified method of Yu excludes FGF2 in the aggregating step, as hypoblast differentiation medium is not used until after aggregation has occurred (Yu, et al.; pgs. 1 and 2 of “Methods”).
Claims 1 and 13 are rejected under 35 U.S.C. 103 as being unpatentable over Yu, et al. (Nature. 2021) in view of Brinster and Troike (J Anim Sci. 1979).
The teachings of Yu are set forth above; claim 1 is anticipated by the same.
Brinster and Troike (hereinafter Brinster) teaches culture conditions for blastocyst development (Summary).
Regarding claim 13: Following the above discussion, Yu does not teach the inclusion of sodium pyruvate in the media.
However, Brinster teaches pyruvate is an important energy source for early embryos, and inclusion of sodium pyruvate in culture media greatly improves the ability of the media to allow in vitro development of the embryos (pg. 30: col. 2, par. 1; and pg. 31: Table 6). Brinster further discloses pyruvate, and not glucose, is the preferred energy source for early pre-implantation embryos (pg. 32; col. 1, par. 3).
Therefore, it would have been prima facie obvious to a person having ordinary skill in the art to have further modified the method of Yu by including sodium pyruvate in the trophoblast and hypoblast differentiation media, as taught by Brinster. This conclusion of obviousness is based on the ‘teaching, suggestion, or motivation rationale’. One would be motivated to do so in order to provide an energy source to support blastoid development, as taught by Brinster (pgs. 30-32); further, as the blastoid is akin to a pre-implantation blastocyst, and as Brinster teaches the use of sodium pyruvate in culture media for early pre-implantation embryos (Table 6; pg. 32), the skilled artisan would have more than a reasonable expectation of success.
Thus, this modified method of Yu renders obvious the wherein an amount of sodium pyruvate is increased limitation recited in claim 13.
Conclusion
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/GINA PRONZATI/Examiner, Art Unit 1633
/ALLISON M FOX/Primary Examiner, Art Unit 1633
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