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 .
Claim Status
As of the Non-Final Office Action mailed 8/26/2025, claims 17, 20-21, 23-24, 26-34, 54-56, 58-59, 62, 64, and 66-70 were pending.
In Applicant's Response filed on 11/25/2025, claims 17, 32, 62, 64, 67-68, and 70 were amended and claims 66 and 69 were canceled.
As such, claims 17, 20-21, 23-24, 26-34, 54-56, 58-59, 62, 64, and 67-68, and 70 are pending and have been examined herein.
Withdrawn Objections and Rejections
The objection of record to claims 62, 64, and 70 for minor informalities has been withdrawn.
The rejection of record of claim 17, 20-21, 23-24, 26-34, 54-56, 58-59, 62, 64, and 66-70 under 35 USC § 112(b) have been withdrawn in view of Applicant’s amendment to the claims. The rejections of canceled claims are moot.
Maintained Rejections
The rejection of record of claims 17, 20-21, 23-24, 26, 28-29, 31, 54, 58-59, 62, 64, 67-68, and 70 under 35 USC § 103 as being unpatentable over Nicholas (WO 2014153230, published 9/25/2014) as evidenced by Nicholas et al (Cell Stem Cell. 2013 May 2; 12(5):573-86; Ref. 73 of NPL in IDS filed 8/18/2020; referenced herein as “Nicholas 2” for purposes of differentiation; previously cited), Miyoshi et al (J Neurosci. 2015 Sep 16;35(37):12869-89; previously cited), and Mei et al (Neuron. 2014 Jul 2;83(1):27-49; previously cited) is maintained. Rejection has been recast below and response to arguments will follow the rejection. Rejections of canceled claims are moot.
The rejection of claims 27 and 32-33 under 35 U.S.C. 103 as being unpatentable over Nicholas et al as evidenced by Miyoshi et al, Mei et al, and Nicholas et al as applied to claims 17, 20-21, 23-24, 26, 28-29, 31, 54, 58-59, 62, 64, 66-68, and 70 above, and further in view of Cameron et al (BMC Neuroscience, 2012; 13:90, 1-23; previously cited) is maintained. Rejection has been recast below and response to arguments will follow the rejection.
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.
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.
Claim(s) 17, 20-21, 23-24, 26, 28-29, 31, 54, 58-59, 62, 64, 67-68, and 70 remain rejected under 35 U.S.C. 103 as being unpatentable over Nicholas et al (WO2014153230 A1, 3/14/2014; published 9/25/2014; previously cited) as evidenced by Nicholas et al (Cell Stem Cell. 2013 May 2; 12(5):573-86; Ref. 73 of NPL in IDS filed 8/18/2020; referenced herein as “Nicholas 2” for purposes of differentiation; previously cited), Miyoshi et al (J Neurosci. 2015 Sep 16;35(37):12869-89; previously cited), and Mei et al (Neuron. 2014 Jul 2;83(1):27-49; previously cited).
Nicholas teaches a method of producing medial ganglionic eminence precursor cells from primate pluripotent stem cells comprising culturing the cells in a serum free medium comprising an activator of sonic hedgehog pathway and a neural inducing supplement to generate the MGE precursor cells (see claim 1 of Nicholas) where the PS cells are human pluripotent stem cells (see claim 5 of Nicholas) (“providing human pluripotent stem cells in culture, differentiating the hPSCs in vitro into medial ganglionic eminence-type precursor cells” as in instant claim 17 in-part and 70 in-part). The cells can also be human embryonic stem cells or induced pluripotent stem cells (see claim 6 of Nicholas) (“wherein the human pluripotent stem cells are human embryonic stem cells” as in instant claim 54; “wherein the human pluripotent stem cells are induced pluripotent stem cells” as in instant claim 59). The pluripotent stem cells can be derived from somatic cells after culturing in reprogramming factors, where the somatic cells can be neurons or neural progenitors (para 54) (“further comprising reprogramming neural or non-neural cells to provide the human pluripotent stem cells” as in instant claim 58). MGE precursor cells express the markers expressed by cells in the MGE region of the developing brain (para 50). In general MGE precursor cells express markers such as, homeobox gene Nkx2.1, LIM- homeobox genes Lhx6, Lhx7, or Lhx8 (same para) (“wherein the cortical interneuron markers further comprises . . . NKX2-1” as in instant claim 23). The reference teaches that the functional MGE precursors are able to be differentiated into functional GABAergic interneurons (para 7) (“wherein the neural precursor cells are capable of differentiating into neural cells that produce GABA in vitro” as in instant claim 20). MGE precursor cells may be transplanted into a target site in the subject that provides appropriate differentiation conditions for the MGE precursor cells to differentiate into interneurons, such as, GABAergic inhibitory interneurons (para 216) (“wherein the neural precursor cells are capable of differentiating into neural cells that produce GABA following transplantation into a mammalian nervous system” as in instant claim 21; “Further comprising differentiating said population of neural precursor cells into GABA-producing neurons” as in instant claim 24). In support of a cortical-like interneuron lineage, robust CXCR7 expression was detected in GFP+ cells (para 261). Although increased ZEB2, ARX, and CXCR4 transcript signals were found at later stages, overall levels were modest, and NKX2.1 and LHX8 continued to be expressed, suggesting a striatal interneuron-like lineage and/or a cortical-like lineage at an immature stage (same para) (“wherein the cortical interneuron markers further comprises one or more of: . . . ARX” as in instant claim 23; “wherein the neural precursor cells express the cell surface markers . . . CXCR4, CXCR7” as in instant claim 26; “wherein the neural precursor cells express CXCR4” as in instant claim 28; “wherein the neural precursor cells express CXCR7” as in instant claim 29; “wherein more immature cells in the population of neural precursor cells have higher LHX8 expression than less immature cells in the population of neural precursor cells” as in instant claim 66; “cells of the population of neural precursor cells express a marker signature for MGE-type cortical interneurons, the marker signature comprising (i) LHX6” as in instant claim 68 in-part). The reference also teaches that MGE precursor cells can be enriched after PS cell cultures are induced to differentiate into MGE precursors (para 190) and that cells enriched for GABAergic neuronal precursors were injected (working example 7 of Nicholas) ) (“isolating cells expressing a cell marker selected from CXCR4, CXCR7 . . thereby generating an in vitro population of neural precursor cells expressing cortical interneuron markers comprising: (i) LHX6, and (ii) MAF and/or MAFB, and having an established MGE-type cortical interneuron fate” as in instant claim 17 in-part and 70 in-part). The reference further teaches that MGE precursors can be isolated and enriched using an affinity tag that is specific for the cell, where the affinity tags can be binding agents that are specific to a marker molecule (para 187) (“wherein isolating comprises using a binding agent to the cell surface marker” as in instant claim 31; “wherein the method does not comprise use of an enhancer-promoter reporter gene expression” as in instant claim 62). The cells can be cryopreserved for a period of 1 day to 10 years (para 205) (“further comprising the step of cryopreserving the neural precursor cells” as in instant claim 34). Please note that while Nicholas does not explicitly teach that greater than 50% of the cells in the population of neural precursor cells express the cortical interneuron markers, the instantly claimed method steps (culturing, differentiating, and isolating) are taught by the reference, such that the result of “at least 50% of cells expressing cortical interneuron markers” would be achieved.
Evidentiary reference Nicholas 2 discloses that most cortical interneurons are born in the MGE of the developing ventral telencephalon midgestation and, subsequently, undergo tangential migration into the neocortex (Introduction, para 1). The reference evidences that interneuron maturation occurs with the progressive acquisition of features that include increasing cell size and branching of processes, expression of subtype markers, formation of GABAergic synapses, ability to fire high-frequency trains of action potentials (APs), and development of more mature electrophysiological properties (same para). Fig. 1A shows schematic of hPSC-derived MGE-like progenitors and the pathways of differentiation into different neuronal subtypes. Of importance, Fig. 1A describes that cortical GABAergic interneurons develop from MGE and initially express NKX2.1, LHX6/8, and DLX, for example, and gradually express only LHX6, CXCR4/7, ARX, etc., while losing NKX2.1 expression (“wherein LHX8 expression was at least reduced in the population of neural precursor cells compared to a population not enriched for cells expressing the cortical interneuron markers” as in instant claim 17 in-part, 68 in-part, and 70 in-part; “wherein LHX8 expression is at least reduced in the population of neural precursor cells after enriching compared to LHX8 expression in a population of MGE-type precursor cells that is depleted for cells expressing the one or more NPCSMs” as in instant claim 64; “wherein NKX2.1 expression is at least reduced in the population of neural precursor cells compared to NKX2.1 expression in a population of MGE-type precursor cells that is not enriched for cells expressing the cortical interneuron markers” as in instant claim 67).
Miyoshi evidences about GABAergic cortical interneurons (title). The reference evidences the genes enriched in MGE derived interneurons include LHX6, CXCR7, MAFB (Table 2) (“cells expressing cortical interneuron markers comprising (i) LHX6, and (ii) MAFB” as in instant claim 17 in-part; 68 in-part; and 70 in-part; “wherein the cortical interneuron markers comprise LHX6 and MAFB” as in instant claim 55; “wherein the cortical interneuron markers comprise LHX6 . . . and MAFB” as in instant claim 56 in-part).
Finally, evidentiary reference Mei discusses neuregulin-erbb signaling in the nervous system (title). Specifically, the reference evidences that Erbb kinases, in particular Erbb4, are critical for the assembly of the GABAergic circuitry including interneuron migration, axon and dendrite development, and synapse formation (Figure 3). Erbb4 is present on progenitors in the median ganglionic eminence (MGE) where interneurons are born, and later in the migratory streams in cells positive for Dlx, a marker of tangentially migrating neurons (“Assembly of neuronal circuitry” para 2). This shows that neuronal precursor cells, particularly those in the MGE, express ErbB4 (“MGE-type precursor cells . . . expressing a cell surface marker selected from . . . ERBB4” as in instant claim 17 in-part and 70 in-part; “wherein the neural precursor cells express ERBB4” as in instant claim 30).
Therefore, it would have been obvious prior to the effective filing date of the instantly claimed invention to produce MGE precursor cells from human pluripotent stem cells that are isolated for particular markers as taught by Nicholas, to arrive at the instantly claimed invention. As Nicholas shows that neural precursor cells can be created from pluripotent stem cells, one of ordinary skill would have been motivated to use the protocol of Nicholas with a reasonable expectation of advantageously obtaining cells that can become GABAergic interneurons when implanted into the central nervous system as taught by the prior art.
Regarding instant claim 68, the composition of human pluripotent stem cell-derived neural precursor cells is provided by the method of Nicholas and Nicholas further teaches the implantation of said composition. Since Nicholas taught the steps of the method as claimed, it would be expected that transplantation of the composition of neural precursor cells would give rise to MGE-type cortical interneurons as set forth in the whereby clause at the end of claim 68. Accordingly, claim 68 is rendered prima facie obvious.
Response to Arguments
Applicant's arguments and the Declaration filed by Dr. Rubenstein have been fully considered but they are not persuasive.
On p. 12-13 of Remarks, Applicant argues that the cells of Nicholas are different from the instantly claimed population of cells because the previous Office action does not state that the cells of Nicholas express MAF and/or MAFB, that the population of cells in Nicholas continue to express LHX8, and that the Nicholas reference fails to recognize the recited combination of marker expression as being specific to hPSC-derived MGE-like cells that specifically give rise to cortical interneurons. Applicant argues that the cells of Nicholas used MKX2.1 to identify MGE-like cells, which Applicant states is expressed in many lineages emanating from MGE. On p. 14 of Remarks, applicant argues that Fig. 1A of the Nicholas 2 reference relates to endogenous cortical interneuron development from MGE precursors and does not teach or suggest how the markers relevant in the in vivo context relate to the Nicholas MGE-like cells in culture. Applicant argues that the reliance on evidentiary reference Miyoshi is misplaced because it relates to in vivo cortical interneuron development, and even if one of ordinary skill would look to Miyoshi, that it does not show that the markers are specific to cortical interneurons within MGE lineage and rather compares MGE-derived and CGE-derived precursor cells developed in vivo. Applicant makes similar arguments related to evidentiary reference Mei.
In response, the examiner disagrees. First, 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., 800 F.2d 1091, 231 USPQ 375 (Fed. Cir. 1986). It is the teaching, suggestions, and motivations of the combination of prior art and evidentiary references cited that creates a prima facie case of obviousness. The examiner also notes that it is not necessary that the prior art suggest the combination to achieve the same advantage or result discovered by applicant. See, e.g., In re Kahn, 441 F.3d 977, 987, 78 USPQ2d 1329, 1336 (Fed. Cir. 2006) (motivation question arises in the context of the general problem confronting the inventor rather than the specific problem solved by the invention); Cross Med. Prods., Inc. v. Medtronic Sofamor Danek, Inc., 424 F.3d 1293, 1323, 76 USPQ2d 1662, 1685 (Fed. Cir. 2005) (“One of ordinary skill in the art need not see the identical problem addressed in a prior art reference to be motivated to apply its teachings.”); In re Lintner, 458 F.2d 1013, 173 USPQ 560 (CCPA 1972); In re Dillon, 919 F.2d 688, 16 USPQ2d 1897 (Fed. Cir. 1990), cert. denied, 500 U.S. 904 (1991) (see MPEP 2144). Regardless, based on the combination of the prior art reference and evidentiary references provided above, applicant’s instant invention would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date of the instant invention. Specifically, Nicholas discloses all of Applicant’s active method steps (i.e., providing hPSCs in culture, differentiating into MGE-like cells, and that cells that have GABAergic interneuron lineage based on CXCR4/7 and other expression markers can be isolated. Nicholas 2 shows that cells that MGE-like cells that eventually develop into cortical GABAergic interneurons initially express NKX2.1 (as in instant claim 23), LHX6, DLX, LHX8, but gradually (i.e., a temporal element) lose NKX2.1 and LHX8 expression (as instantly claimed) and maintain CXCR4/7, ARX, and LHX6 (among other markers). Miyoshi evidences that markers that MGE-derived GABAergic cortical interneurons are enriched for markers such as LHX6, CXCR7, (both of these are also disclosed in evidentiary reference Nicholas 2) and MAFB (i.e., would provide one of ordinary skill that MGE-like cells that become GABAergic interneurons like the ones disclosed in Nicholas would express and subsequently enriched in at least LHX6, CXCR, and MAFB expression). Finally, evidentiary reference Mei shows that ERBB4 is critical for development of GABAergic circuitry and is present on progenitors in MGE (i.e., provides sufficient support that MGE-like cells (such as those discussed in the previous references) require and likely express ERBB4).
The examiner would also like to note that the Rubenstein declaration regarding the technical solution the instant claims purportedly improve upon are appreciated. Even if, arguendo, the markers as claimed are the reason for the improved engraftment, migratory ability, and faster development post-transplantation of NPCs, the prior art recognized the results before the effective filing date of the instantly claimed invention. Solely to rebut Applicant’s argument, the examiner directs Applicant’s attention to Cunningham et al (Cell Stem Cell, Volume 15, Issue 5, 559 - 573. November 2014), which describes that in vitro hPSC-derived maturing GABAergic interneurons (mGINs) migrate extensively and integrate into dysfunctional circuitry of the epileptic mouse brain (abstract). Human MGE cells were generated by in vitro differentiation of H7 human embryonic stem cells according to our optimized procedure and purified by fluorescence-activated cell sorting (FACS) with anti-ENCAM prior to transplantation (Figure 1A). Most of the FACS-sorted cells expressed the MGE markers Nkx2.1 (Results, para 1). Histological analysis showed that, 2 weeks post-transplantation (PT), cells were primarily clustered near the injection site (59,027 ± 18,724 total human nucleus+ cells per mouse, n = 3; Figures 1B and 1C). However, at 4 months PT, transplanted mGIN had extensively migrated, becoming well integrated within the host hippocampus (74,913 ± 15,417 total human nucleus+ cells per mouse, n = 8; Figures 1D–1J, S2, and S4) without significant difference in the total surviving cell numbers in comparison to 2 weeks PT (p = 0.58). Stereological analysis demonstrated migration of transplanted human mGIN greater than 1.6 mm from the site of injection (Figure 1K). At 2 weeks PT, most cells expressed GABA and Sox6 as well as Nkx2.1 (Figures 2A–2C), and a minority of cells expressed the more mature neuronal marker NeuN (Figure 2E). However, at 4 months PT, the majority of cells expressed NeuN and β-tubulin as well as GABA and Sox6 (Figures 2G, 2H, 2J, and 2R–2T). The expression of precursor marker Nkx2.1 was significantly diminished at 4 months PT in comparison to 2 weeks PT (Figures 2I and 2W), whereas the mature interneuron marker Lhx6 was significantly increased at 4 months PT in comparison to 2 weeks PT (Figures 2D, 2K, and 2W). In addition, proliferating cell marker Ki67 was significantly decreased after 4 months PT in comparison to 2 weeks PT (Figures 2F, 2L, and 2W). Furthermore, at 4 months PT, some GABAergic interneurons were found to express somatostatin, parvalbumin, calretinin, neuropeptide Y, and calbindin (Figures 2M–2Q, 2X, and S5A–S5F).
"A greater than expected result is an evidentiary factor pertinent to the legal conclusion of obviousness ... of the claims at issue." In re Corkill, 771 F.2d 1496, 226 USPQ 1005 (Fed. Cir. 1985). However, a greater than additive effect is not necessarily sufficient to overcome a prima facie case of obviousness because such an effect can either be expected or unexpected. Applicants must further show that the results were greater than those which would have been expected from the prior art to an unobvious extent, and that the results are of a significant, practical advantage. Ex parte The NutraSweet Co., 19 USPQ2d 1586 (Bd. Pat. App. & Inter. 1991) (see MPEP § 716.02(a)). Thus, neither the arguments posited nor the Rubenstein Declaration provide any evidence that the instantly claimed method provides any benefits/unexpected results not previously contemplated by the prior art. The combination of the applied references render the claims prima facie obvious in light of their disclosures.
Claim(s) 27 and 32-33 remain rejected under 35 U.S.C. 103 as being unpatentable over Nicholas et al as evidenced by Miyoshi et al, Mei et al, and Nicholas et al as applied to claims 17, 20-21, 23-24, 26, 28-29, 31, 54, 58-59, 62, 64, 66-68, and 70 above, and further in view of Cameron et al (BMC Neuroscience, 2012; 13:90, 1-23; previously cited).
The teachings of Nicholas (as evidenced by Miyoshi, Mei, and Nicholas) were recited in the above 35 U.S.C. 103 rejection as applied to claim 17 of which claim 27 and 32-33 depend. The teachings will not be repeated here.
The difference between the combined teachings and the invention as instantly claimed is that they do not teach that the neural precursor cells express PLEXINA4 (instant claim 27), further depleting the population of NPCs with low expression of cortical interneuron markers upregulated in NPCs (claim 32), and depleting the population of NPCs of cells expressing ATP1A2, BCAN, CD271, CD98, CNTFR, EGFR, FGFR1, FGFR2, FGFR3, GJA1, MLC1, NOTCH1, NOTCH3, PDGFRB, PDPN, PLXNA4, PROM1, PTPRZ1, SLC1A3, SLC1A5, TMEM158, or TTYH1.
Cameron teaches Clustered QISPs of transcripts expressed in excitatory cortical neurons and discloses expression of plexin A4 (PLXNA4) (see figure 6). Cameron teaches this study provides a rigorously validated list of transcripts expressed by immature excitatory neurons that have been grouped by in situ expression pattern for understanding how specific genes, and functional networks of such genes, control neuronal development and contribute to pathology (pg. 19, col. 2, last par.). Cameron teaches Plxna4 mRNA expression functional groupings of QISPs based on Gene Ontology molecular function and biological process annotations (graphical representation in Figure 4). Following transcriptional profiling, genes are sorted by RMA (robust multichip average) and their level of fold up- or down-regulation are compared to the FACS GFP-population. Verification of up-regulated genes is accomplished with quantified in situ hybridization (QISP), and the verified genes hierarchically clustered (see Methods) (Figure 1). This shows that excitatory cortical neurons express PLXNA4, which have opposite function to GABAergic cortical neurons (i.e, inhibitory cortical neurons), which provides one of ordinary skill to perform the depleting step as claimed in instant claim 32 and 33 with a reasonable expectation of success to ensure production of inhibitory cortical neurons.
Therefore, it would have been obvious prior to the effective filing date of the instantly claimed invention to create MGE precursor cells as taught by Nicholas, Miyoshi, Mei and Nicholas in combination, where the population is depleted of cells expressing PLXNA4 as taught by Cameron, to arrive at the instantly claimed invention. As Cameron shows PLXNA4 is expressed by excitatory cortical interneurons, one of ordinary skill would have been motivated to further isolate the cells of Nicholas, Miyoshi, Mei, and Nicholas in combination to exclude cells expressing PLXNA4 (depleting PLXNA4+ cells) as taught by Cameron with a reasonable expectation of advantageously having only cells capable of becoming cortical inhibitory neurons (i.e., GABAergic cortical neurons) as taught by the prior art.
Response to Arguments
Applicant has not provided any arguments against previously cited reference Cameron.
Conclusion
No claim is 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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/G.R./Examiner, Art Unit 1632
/KARA D JOHNSON/Primary Examiner, Art Unit 1632