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 .
Office Action: Notice
Any objection or rejection of record in the previous Office Action, mailed 6/4/2025, which is not addressed in this action has been withdrawn in light of Applicants' amendments and/or arguments. This action is FINAL.
Claim Status
Applicant amended claims 1, 10-11, 13, 15, 17 and 20-21 (9/4/2025). No new matter was added. Thus, claims 1-21 are under examination.
Priority
Claims 1-21 claim a priority date of 8/8/2019, the filing date of U.S. Provisional No. 62/884, 536.
Information Disclosure Statement
The listing of references in the specification is not a proper information disclosure statement. 37 CFR 1.98(b) requires a list of all patents, publications, or other information submitted for consideration by the Office, and MPEP § 609.04(a) states, "the list may not be incorporated into the specification but must be submitted in a separate paper." Therefore, unless the references have been cited by the examiner on form PTO-892, they have not been considered.
The Information Disclosure Statements from 7/2/2025, 9/23/2025 and 11/14/2025 are considered.
Objections Withdrawn
Specification:
The objections to the specification due to the use of a trademark or tradenames are withdrawn in view of Applicant’s amendments.
Claims:
The objections to write out abbreviations completely the first time used in claims 1, 13, 15 and 20 and to correct minor clerical issues to claim 20 are withdrawn due to Applicant’s amendments.
Rejections Withdrawn
Claim Rejections - 35 USC § 112(b)
The rejections of claims 10-11 and 17 under 35 U.S.C. 112(b) or pre-AIA 35 U.S.C. 112, 2nd paragraph, are withdrawn in view of Applicant’s amendments of claims 10-11 and 17 to address and further clarify indefiniteness.
Rejections Maintained
Claim Rejections - 35 USC § 102
Claims 1-20 are rejected under 35 U.S.C. 102 (a)(1) and 102 (a)(2) as being anticipated by Bauer et al. (WO 2016/182893 A1; published 11/17/2016).
Regarding claim 1, Bauer teaches a method of utilizing Cas9-based knockout screens in identifying essential genes and genes involved in drug resistance in various cell lines (Paragraph 10, lines 1-4). Further, Bauer teaches that expression of a gene of interest may be altered by said targeting by at least one guide RNA within the plurality of CRISPR-Cas system guide RNAs and that at least one continuous genomic region may comprise up to the entire genome including a functional element of the genome (i.e., within a coding gene, intronic region, promoter, or enhancer) (Paragraph 16, lines 1-5). Bauer also teaches that the DNA region may comprise an epigenetic insulator and may comprise two or more continuous genomic regions that physically interact or have separate signatures (i.e., histone acetylation, histone methylation, histone ubiquitination, histone phosphorylation, DNA methylation, or a lack thereof) (Paragraph 17, lines 5-10). Specifically, Bauer teaches that functional regions of the previously described DNA regions may comprise over 50 nucleotides (Paragraph 107, lines 1-5).
Additionally, Bauer teaches that targeting may comprise introducing into each cell in the population of cells a vector system of one or more vectors comprising an engineered, non-naturally occurring CRISPR-Cas system (Paragraph 20, lines 1-2). Further, Bauer teaches that specific sites are targeted through monitoring interaction strength with specified promoters via paired-end tag sequencing (Figure 41, Paragraph 72, lines 1-3), occurring via transduction with vectors (i.e., Cas9, sgRNAs) (Figures 8A-B; Paragraph 39, lines 1-3). Bauer further teaches the comparison of ranked enhancers or promoters via transcription levels through monitoring specifically signal intensity of H3K27ac (trait associated haplotypes) following normalization to GAPDH (control) (Figures 1A-1E; Paragraph 32, lines 1-5).
Regarding claim 2, Bauer teaches that the previously described identification method for promoter or enhancer sequences includes screening methods for identifying functional elements in the non-coding genome, more particularly using libraries via targeting Cas9 or other vectors to intergenic regions surrounding single genes and generating a library which flanks 100kb upstream and downstream of target gene with sgRNAs or polynucleotides (Paragraph 96, lines 1-5).
Regarding claim 3, Bauer teaches that the previously described identification method for promoter or enhancer sequences includes strategies to knock-in via homology-directed repair the fluorescent protein for further analyses (Figures 26A-26K; Paragraph 57, lines 1-10).
Regarding claims 4-5, Bauer teaches a method of prior library design with specified sgRNAs cloned into lentiviral vectors following synthesized onto an array prior to the transduction of BRAF mutant and control cells that are again sequenced via deep-sequencing readout for further identification for early time point comparisons (Figures 30A-30D; Paragraph 61, lines 1-20).
Regarding claims 6-7, Bauer teaches that the previously described identification method for promoter or enhancer sequences includes a DNA region that may comprise an epigenetic insulator and may comprise two or more continuous or similar genomic regions that physically interact or have completely separate signatures (i.e., histone acetylation, histone methylation, histone ubiquitination, histone phosphorylation, DNA methylation, or a lack thereof) (Paragraph 17, lines 5-10). Specifically, Bauer teaches that functional regions of the previously described DNA regions may comprise over 50 nucleotides (Paragraph 107, lines 1-5).
Regarding claims 8-9, Bauer teaches that the previously described identification method for promoter or enhancer sequences includes methods that allow for the identification of targets in the 5’ and 3’ region of a gene which may affect a phenotypic change only under particular circumstances or only for particular cells or tissues in an organism (Paragraph 79, lines 1-10).
Regarding claim 10, Bauer teaches that the previously described identification method for promoter or enhancer sequences includes methods that specifically target a multitude of sequences or DNA fragments within the continuous genomic region of interest via introducing into each cell of a population of cells a vector system of one or more vectors comprising an engineered, non-naturally occurring CRISPR-Cas system comprising: at least one Cas protein, and one or more guide RNAs of the guide RNA library where the Cas protein and the one or more guide RNAs may be on the same or on different vectors of the system and are integrated into each cell, whereby each guide sequence targets a sequence within the continuous genomic region in each cell in the population of cells (Paragraph 89, lines 1-10).
Regarding claim 11, Bauer teaches that the previously described identification method for promoter or enhancer sequences includes regulatory sequences and are intended to include promoters, enhancers and other expression control elements (i.e., polyadenylation signals) (Paragraph 119, lines 1-5). For example, Bauer teaches that these promoters include such genes (HbSC, HbS/P+, HbS/PO) that cause a reduction in the amount of hemoglobin produced as compared to a normal or standard condition (Paragraph 134, lines 1-15).
Regarding claims 12-15, Bauer teaches that the previously described identification method for promoter or enhancer sequences includes the identification of targets in the 5’ and 3’ region of a gene which may affect a phenotypic change only under particular circumstances or only for particular cells or tissues in an organism where the genomic region of interest comprises a transcription factor binding site, a region of DNase I hypersensitivity, a transcription enhancer or repressor element, including the genomic region of interest comprises an epigenetic signature for a particular disease or disorder (Paragraph 79, lines 5-10). Further Bauer teaches that additionally or alternatively the genomic region of interest may comprise an epigenetic insulator where the guide RNA library is directed to a genomic region which comprises two or more continuous genomic regions or cell types (Paragraph 79, lines 10-15).
Bauer further teaches that to calculate the fold enrichment of the sgRNAs in proximity to other molecular hallmarks for hypersensitivity (DNAse-seq, ATAC-seq, conservation), the average sgRNA enrichment of sgRNAs near the peaks of these molecular hallmarks were examined (Paragraph 323, lines 1-5).
Bauer also teaches that further hallmarks for comparison between cell types using the previously described method are induced via enhancer sequences that bind transcription factors and chromatin regulators and are correlated with specific chromatin features including reduced DNA methylation, characteristic histone modifications, heightened chromatin accessibility, long-range promoter interactions, and bidirectional transcription. Recent chromatin mapping has demonstrated the abundance of distal regulatory elements bearing an enhancer chromatin signature (Paragraph 232, lines 1-10). Bauer teaches that specifically histone modifications in human H3K27ac can be monitored via ChIP-seq (Paragraph 252, lines 1-2).
Regarding claim 16, Bauer teaches that the previously described identification method for promoter or enhancer sequences includes the comparison of ranked enhancers or promoters via transcription levels through monitoring specifically signal intensity of H3K27ac (trait associated haplotypes) following normalization to GAPDH (control) (Figures 1A-1E; Paragraph 32, lines 1-5). Specifically, Bauer teaches that quantitative sequencing or qPCR can be used to assess the specified gene expression described previously (Figures 14A-14D; Paragraph 45, lines 1-5).
Regarding claim 17, Bauer teaches that the previously described identification method for promoter or enhancer sequences includes a “3C”design template that utilizes a target sequence or ‘bait’ (i.e., a restriction fragment of choice that encompasses a selected gene) ligated to many different nucleotide sequences of interest or sequence tags (representing this gene's genomic environment) where the template is cleaved by another, secondary or different, restriction enzyme and subsequently re-ligated to form small DNA circles and advantageously, the one or more nucleotide sequences of interest that are ligated to the target nucleotide sequence are amplified using at least two oligonucleotide primers or promoters, wherein at least one primer or promoter hybridizes to the target sequence to flank the nucleotide of interest labeled and optionally hybridized to an array, typically against a control sample containing genomic DNA digested with the same combination of restriction enzymes (Paragraph 102, lines 1-30). Bauer teaches that this previously described method can be used to search the complete genome for interacting DNA elements (Paragraph 102, lines 25-30).
Regarding claim 18, Bauer teaches that the previously described identification method for promoter or enhancer sequences includes the comparison of ranked enhancers or promoters via transcription levels through monitoring specifically signal intensity of specified sequences of interest or tagged sequences involving H3K27ac (trait associated haplotypes) following normalization to GAPDH (control) (Figures 1A-1E; Paragraph 32, lines 1-5). Specifically, Bauer teaches that quantitative sequencing or qPCR can be used to assess the specified gene expression described previously (Figures 14A-14D; Paragraph 45, lines 1-5). For example, Bauer teaches that the previously described methods of monitoring transcription levels can be used for motif analysis to evaluate the human and mouse enhancer regions across different cell types for potential binding sites for known transcription factors (Paragraph 258, lines 1-3).
Regarding claim 19, Bauer teaches a method of prior library design with specified sgRNAs cloned into lentiviral vectors following synthesized onto an array prior to the transduction of BRAF mutant and control cells that are again sequenced via deep-sequencing readout for further identification for early time point comparisons (Figures 30A-30D; Paragraph 61, lines 1-20).
Regarding claim 20, Bauer teaches that the previously described method of library design for promoters or enhancers can be applied to analyze multiple populations of eukaryotic cells including embryonic stem (ES) cells, neuronal cells, epithelial cells, immune cells, endocrine cells, muscle cells, erythrocytes, lymphocytes, plant cells, or yeast cells (Paragraph 90, lines 1-5).
Bauer teaches each and every limitation of claims 1-20, and therefore Bauer anticipates claims 1-20.
Applicant’s Response: The Applicant argues that the primary reference, Bauer, does not anticipate the amended instant claims because Bauer is limited to CRISPR/Cas-based knockout or mutagenesis screens, whereas the claimed methods do not use CRISPR/Cas proteins, guide RNAs, or genomic modification, and instead rely on synthetic DNA fragments derived from epigenetically defined regions (i.e., DNAse hypersensitivity, histone marks) that are barcoded and assayed via transcriptional readout. Further, the Applicant asserts that Bauer fails to teach key claim limitations, including constructing promoter sequences from combinatorial DNA fragments with sequence tags, inserting those constructs into vectors to drive reporter expression, and determining promoter activity by measuring transcription of the sequence tag rather than endogenous gene expression, and therefore does not teach each and every element of the claimed methods.
Examiner’s Response to Traversal: Applicant’s arguments have been carefully and fully considered but are not found persuasive, as discussed below.
The Applicant’s argument that Bauer is limited to CRISPR/Cas knockout or mutagenesis is not persuasive because Bauer expressly teaches identifying promoter and enhanced sequences using epigenetic features, transcriptional output, and comparative analyses across cell types (Paragraph 10, lines 1-4; Paragraph 16, lines 1-5; Paragraph 17, lines 5-10; Paragraph 107, lines 1-5). As of note, anticipation does not require the reference to exclude additional or alternative techniques (see MPEP 2131.01).
Further, the Applicant’s assertion that Bauer fails to teach the use of synthetic DNA fragments, barcodes or sequence tags is unavailing, as Bauer teaches libraries of regulatory sequences introduced via viral vectors and assessed by sequencing-based transcriptional readouts (Paragraph 119, lines 1-5; Paragraph 134, lines 1-15), which reasonably correspond to the claimed sequence tag-based measurement of transcription. Since the instant amended claims do not require the absence of CRISPR, Cas proteins or guide RNAs, such limitations cannot be read into the claims to distinguish the prior art (see MPEP 2111).
Additionally, the Applicant’s argument that Bauer measures endogenous gene expression rather than promoter activity is also unpersuasive, as Bauer teaches ranking promoters and enhancers based on transcriptional levels and signal intensity derived from tagged or targeted sequences, which meets the claimed set of determining transcription to identify tissue- or cell-specific regulatory activity (Paragraph 232, lines 1-10; Paragraph 252, lines 1-2). Notably, In re Gleave, 560 F.3d 1331 (Fed. Cir. 2009) holds that a reference anticipates even if it does not recognize the same utility or emphasize the same advantages as the Applicant. Therefore, regardless of Bauer’s broader CRISPR-based context, Bauer’s teaching of promoter and enhancer identification using transcriptional and epigenetic readouts anticipates the instant amended claims.
Accordingly, Bauer discloses each and every element of the claimed methods arranged as claimed, and the anticipation rejection for claims 1-20 is proper and therefore maintained (see MPEP 2131.02).
Claim Rejections - 35 USC § 103
Claim 21 is rejected under 35 U.S.C. 103 as being unpatentable over Bauer et al. (WO 2016/182893 A1; published 11/17/2016), as applied to claims 1-20 above, in view of Ahfeldt et al. (WO 2018/129486 A2; published 7/12/2018).
As previously discussed, teaches a method of utilizing Cas9-based knockout screens in identifying essential genes and genes involved in drug resistance in various cell lines (Paragraph 10, lines 1-4). Further, Bauer teaches that expression of a gene of interest may be altered by said targeting by at least one guide RNA within the plurality of CRISPR-Cas system guide RNAs and that at least one continuous genomic region may comprise up to the entire genome including a functional element of the genome (i.e., within a coding gene, intronic region, promoter, or enhancer) (Paragraph 16, lines 1-5). Bauer also teaches that the DNA region may comprise an epigenetic insulator and may comprise two or more continuous genomic regions that physically interact or have separate signatures (i.e., histone acetylation, histone methylation, histone ubiquitination, histone phosphorylation, DNA methylation, or a lack thereof) (Paragraph 17, lines 5-10). Specifically, Bauer teaches that functional regions of the previously described DNA regions may comprise over 50 nucleotides (Paragraph 107, lines 1-5).
Regarding claim 21, Bauer teaches that the previously described method of library design for promoters or enhancers can be applied to analyze multiple populations of eukaryotic cells including embryonic stem (ES) cells, neuronal cells, epithelial cells, immune cells, endocrine cells, muscle cells, erythrocytes, lymphocytes, plant cells, or yeast cells (Paragraph 90, lines 1-5).
Bauer does not teach or suggest that the specific neuronal cell sub-type includes dopaminergic, gabaergic or glutamatergic neurons.
Ahfeldt teaches techniques for enhanced expression of the reporter gene when a gene of interest is important for continued biomedical research (Paragraph 2, lines 1-5) via the detection of co-expression of reporter genes (i.e., dopaminergic neurons) to study Parkinson’s Disease (Paragraph 2, lines 1-5). Further, Ahfeldt teaches that differentiation of the embryonic stem cells into dopaminergic neurons can then be tracked or differentiated cells can be isolated using the properties of the reporter (i.e., fluorescence) (Figure 5E; Paragraph 27, lines 1-10) via CRISPR mediated knock-out mutagenesis to create specified lines from a WT donor line (Figures 4B and 9 A-D; Paragraph 129, lines 1-5).
It would have been obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention to combine Bauer’s promoter identification method with Ahfeldt’s specific neuronal subtypes because both references are directed towards the same field of identifying and utilizing cell-specific gene expression systems. Moreso, Ahfeldt specifically teaches the importance of dopaminergic neurons for biomedical research, and one of ordinary skill in the art would recognize that Bauer’s general method for identifying tissue-specific promoters would be directly applicable to the specific neuronal subtypes taught by Ahfeldt. Further, a person of ordinary skill in the art would have a reasonable expectation of success in combining these reference because Ahfeldt demonstrates that dopaminergic neurons can be successfully differentiated and tracked using reporter systems, confirming these cells are amenable to the expression vector approaches taught by Bauer and therefore it would have been obvious to apply Bauer’s promoter identification methodology to the well-characterized neuronal subtypes specifically disclosed by Ahfeldt.
Applicant’s Response: The Applicant argues that the primary reference, Bauer, as well as the secondary reference, Ahfeldt, relate to fundamentally different methods than those claimed and, even in combination, fail to teach or suggest identifying promoter sequences using DNA fragments with sequence tags and determining promoter activity by measuring transcription of the sequence tag. The Applicant further asserts that Ahfeldt’s teachings of knock-in reporter cells for assessing endogenous gene expression (including neuronal subtypes) does not cure Bauer’s alleged deficiencies and does not teach tissue-or cell-specific promoter identification as claimed. Accordingly, the Applicant asserts that Bauer in view of Ahfeldt does not render claim 21 obvious.
Examiner’s Response to Traversal: Applicant’s arguments have been carefully and fully considered but are not found persuasive, as discussed below.
Firstly, the Applicant’s argument that Bauer and Ahfeldt relate to completely different methods is not persuasive because Bauer broadly teaches methods for identifying promoters and enhancers based on functional genomic and epigenetic features across multiple eukaryotic cell populations, expressly including neuronal cells (Paragraphs 16-17, 90, 107), while Ahfeldt teaches the differentiation, identification, and reporter-based tracking of specific neuronal subtypes, including dopaminergic neurons (Abstract; Paragraphs 2, 27, 129). As of note, a reference need not disclose the identical purpose or end use of the claimed invention, so long as it teaches the claimed elements and a rationale exists to combine them (see MPEP 2141, 2143).
Further, the Applicant’s assertion that Ahfeldt does not cure Bauer’s alleged deficiencies because Ahfeldt measures endogenous gene expression rather than promoter activity is unavailing. Ahfeldt teaches reporter-based systems confirming successful differentiation and expression in defined neuronal subtypes (Paragraph 2, lines 1-5; Figure 5E; Paragraph 27, lines 1-10), thereby establishing that such neuronal subtypes are amenable to expression vector and reporter methodologies of the type taught by Bauer. A person of ordinary skill in the art would have reasonably expected success in applying Bauer’s promoter identification techniques to the well-characterized neuronal subtypes disclosed by Ahfeldt, given their shared focus on cell-specific gene expression systems (see MPEP 2143; KSR Int’l Co. v. Teleflex Inc., 550 US 298 (2007)).
More so, the Applicant’s argument that Bauer does not explicitly list dopaminergic, GABAergic, or glutamatergic neurons is not persuasive, as Bauer teaches applicability to neuronal cells generally, and Ahfeldt expressly identifies specific neuronal subtype[es of biomedical importance. Selecting a known neuronal subtype taught by Ahfeldt as a target population for Bauer’s known promoter identification methods represents a predictable variation using known techniques to improve or tailor results, which is prima facie obvious (see MPEP 2144.04).
Accordingly, Bauer in view of Ahfeldt, teaches or suggests all limitations of claim 21, and therefore the 103 rejection is properly maintained.
Conclusions
No claim is allowed.
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). 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 extension fee 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 date of this final action.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to ELIZABETH ROSE LAFAVE whose telephone number is (703)756-4747. The examiner can normally be reached Compressed Bi-Week: M-F 7:30-4:30.
Examiner interviews are available via telephone, in-person, and video conferencing using a USPTO supplied web-based collaboration tool. To schedule an interview, applicant is encouraged to use the USPTO Automated Interview Request (AIR) at http://www.uspto.gov/interviewpractice.
If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Heather Calamita can be reached on 571-272-2876. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
Information regarding the status of published or unpublished applications may be obtained from Patent Center. Unpublished application information in Patent Center is available to registered users. To file and manage patent submissions in Patent Center, visit: https://patentcenter.uspto.gov. Visit https://www.uspto.gov/patents/apply/patent-center for more information about Patent Center and https://www.uspto.gov/patents/docx for information about filing in DOCX format. For additional questions, contact the Electronic Business Center (EBC) at 866-217-9197 (toll-free). If you would like assistance from a USPTO Customer Service Representative, call 800-786-9199 (IN USA OR CANADA) or 571-272-1000.
/ELIZABETH ROSE LAFAVE/ Examiner, Art Unit 1684
/HEATHER CALAMITA/ Supervisory Patent Examiner, Art Unit 1684
Prosecution Insights