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 .
Continued Examination Under 37 CFR 1.114
A request for continued examination under 37 CFR 1.114, including the fee set forth in 37 CFR 1.17(e), was filed in this application after final rejection. Since this application is eligible for continued examination under 37 CFR 1.114, and the fee set forth in 37 CFR 1.17(e) has been timely paid, the finality of the previous Office action has been withdrawn pursuant to 37 CFR 1.114. Applicant's submission filed on 11/14/2025 has been entered.
Application Status
This action is written in response to applicant’s correspondence received on 11/14/2025. Claims 1-4, 6-7, 10-11, 34-39, 41-42, 44-46, and 109-112 are pending. Claims 1, 3-4, and 110 have been amended. Claims 111-112 have been newly added. All pending claims are currently under examination.
Claim Rejections - 35 USC § 103 – Updated in Response to Amendment
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.
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 1-4, 6-7, 34-36, 39, 41, and 109-110 are rejected under 35 U.S.C. 103 as being unpatentable over Hartl (Hartl D et al. Nucleic Acids Res. 2017 Nov 16;45(20):11607-11621, submitted in applicant’s IDS filed 5/21/2024) in view of Tasic (Tasic B et al. Nat Neurosci. 2016 Feb;19(2):335-46) and Nguyen (Nguyen A et al. Front Immunol. 2018 Jul 4;9:1553).
Regarding claim 1, Hartl is a research publication that teaches the use of a parallel reporter assay to identify regulatory elements with differential expression in different cell types (Title, Abstract, and see document). Hartl teaches a method of identifying a regulatory element that provides selective expression in a given cell type, comprising obtaining a library of candidate regulatory elements (page 11608, left column, third paragraph, “we tested hundreds of CREs [cis-regulatory elements]”). Hartl teaches creating a library of vectors, where each vector of the library of vectors comprises a candidate regulatory element of the library of candidate regulatory elements operably linked to a transgene (page 11609, right column, first paragraph). Hartl teaches that the transgene comprises a barcode unique to the candidate regulatory element (Figure 3, and caption below Figure 3, which teaches that GFP (a transgene) was attached to a unique barcode for each regulatory element tested). Hartl teaches contacting a population of cells with the library of vectors (e.g., Figure 3A). Hartl teaches identifying the cell type of the individual cells (“cell populations are FACS sorted,” Figure 3 and Figure 3 caption). Hartl teaches assessing the expression of the barcoded transgenes in each of the cell types to identify a barcoded transgene which is selectively expressed in the given cell type compared to other cell types of the population of cells (page 11613, right column final paragraph to page 11615, entirety, and Figures 3 and 4). Specifically, Hartl teaches that the parallel reporter assay that they used, which included a transgene with unique barcodes for the library of regulatory elements described in Figure 3 is applied to unique cell types to identify selective expression in each cell type (see page 11615, section entitled “Autonomous CRE activities in four retinal cell types,” and Figure 4). Hartl teaches correlating the identified barcode to a candidate regulatory element to identify a regulatory element that provides selective expression in the given cell type (e.g., “this effort let[sic] to the identification of a small set of short sequences showing preferential activity in different cellular subsets of the retina,” page 11618, left column, first paragraph).
Furthermore, Hartl teaches the use of a commercially available kit for single cell RNA sequencing which was used in their RNA-seq library preparation (Norgen 51800, page 11608, right column, second paragraph).
Hartl, while teaching the identification of regulatory elements expressed differentially in different cell types, does not teach that single-cell RNA sequencing was performed to obtain transcriptome data for a plurality of single cells, where the cell type is identified by the transcriptome data and not sorted by reporter or natural marker, as Hartl appears to have differentiated the cells based on FACS (e.g., Figure 3A).
Tasic is a research article which focuses on the identification of cell by taxonomical classification using single cell transcriptomics (Title, Abstract, and throughout). Furthermore, Tasic is generally drawn to methods of classifying cell characteristics of small cell populations within larger cell populations (Abstract, page 335, left column, second paragraph). Tasic and Hartl therefore overlap in field of endeavor and subject matter because both concern determining the expression patterns of specific cell populations within larger, more complex cell populations. To further corroborate the overlap between Tasic and Hartl, Hartl in fact directly references Tasic, where Hartl teaches that retina cells and neuronal cells can be challenging samples in the same way, in the sense that they are both cells that are rare within tissue cells and therefore require unique approaches to analyze cell data (page 11614, left column, first paragraph). Tasic teaches that single-cell transcriptomic signatures can be associated with specific cellular properties (Abstract). Tasic teaches and reduces to practice methods of single cell isolation and RNA transcriptomic profiling (“Online Methods” page 347, right column beginning second paragraph to final page of the document). Furthermore, Tasic teaches that molecular markers such as surface markers can be used to differentiate cell types, but that it is more useful to further characterize cells based not only on cell surface markers but single cell sequencing, as such in-depth analysis can further elucidate heterogeneity within a cell population (Introduction, second paragraph). Furthermore, Tasic teaches that such single-cell RNA sequencing to define transcriptomics of cell types is robust and scalable, has emerged as a powerful tool to further classify cell types, and that such methods have been reduced to practice in rare cell types within tissues, such as neurons (Introduction, second paragraph). Furthermore, Tasic teaches that such single-cell transcriptomic sequencing methods can lead to a clearer understanding of cell-type subdivisions, where unique cell lines offer unique expression markers (see Discussion, particularly the first paragraph). Tasic therefore teaches a strong motivation to perform single-cell transcriptomic sequencing to identify cell types, such that unique subdivision of cell types can be elucidated from complex tissues for the purposes of discovering new expression markers (Discussion entirety, and specifically first paragraph). Furthermore, similar to the methods of Hartl, Tasic’s method also relies on the use of cell separation by FACS (e.g., page 344, right column, second paragraph). Thus, the methods of both Hartl and Tasic appear to be compatible with one another.
As discussed above, Tasic teaches that molecular markers such as surface markers can be used to differentiate cell types, but that it is more useful to further characterize cells based not only on cell surface markers but single cell sequencing, as such in-depth analysis can further elucidate heterogeneity within a cell population (Introduction, second paragraph). To further elaborate upon this point, Tasic teaches that, while their methods and research focused on CRE line generated cells isolated by FACS, their study in fact compliments another study (see “Discussion,” fifth paragraph, bottom of page 344 to the first paragraph of page 345). When discussing the previous alternate study, Tasic teaches that:
“In addition, our study differed from the previous one in the genetic background (mostly C57BL/6J versus CD-1) and age of analyzed mice, as well as the cell isolation procedures (FACS versus mostly Fluidigm C1 microfluidics). Overall, the two studies overlap in their identification of some transcriptomic types, but differ in their focus: the previous study offers deeper insight into non-neuronal transcriptomic types, hippocampal excitatory cells and cells from brain ventricles, whereas our results provide a more comprehensive classification of adult neocortical neurons,” (page 345, left column first paragraph, emphasis added).
Thus, Tasic teaches that similar methods which do not rely upon FACS separation (i.e., Fluidigm C1 microfluidics, see discussion of Nguyen, below) can be used for the same purposes as the method taught by Tasic, where furthermore the results of using FACS and non-sorting methods are similar but also broader in scope for non-sorting methods (i.e. “offers deeper insight into non-neuronal transcriptomic types”) than using cell sorting based upon FACS (page 345, first paragraph). Thus, Tasic, by referencing this previous study in the Discussion, also teaches non-sorting methods using microfluidics such as Fluidigm, where such approaches predictably correlate with results using sorting-based methods such as FACS and furthermore offer advantages such as deeper classification and broader understanding of non-target cells/heterogenous populations (Discussion fifth paragraph).
With regards to the Fluidigm C1 microfluidic separation method taught by Tasic, Nguyen is a review article that teaches various single-cell RNA sequencing methods. Nguyen teaches the Fluidigm C1 approach taught by Tasic (page 4 left column final paragraph). Furthermore, Nguyen teaches that such microfluidic methods of cell separation are advantageous, much like Tasic taught, because they are high-throughput and low-cost (page 4, left column, final paragraph). Furthermore, Nguyen teaches that such microfluidic methods of cell sorting do not rely on natural markers or fluorescent reporters, as recited in the claim (page 4, right column, first paragraph). Thus, the Fluidigm C1 microfluidic cell separation method taught by Tasic/Nguyen does not rely on sorting of a reporter or marker.
It would have been obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention to modify the regulatory element detection method of Hartl to include single cell transcriptomic profiling for cell classification as taught by Tasic because such a combination is simply the combination of known prior art elements to yield predictable results. In the present case, a practitioner would modify the known method of single-cell transcriptomic profiling as taught by Tasic with the regulatory element/expression profiling technique taught by Hartl. Furthermore, the combination is not simply a combination of known elements; a practitioner would be motivated to combine the teachings of Tasic with Hartl because Tasic teaches that such single-cell transcriptomic profiling is not only robust, scalable, and feasible, but that it offers the advantage of further refining the classification of subdivisions of cell types with unique expression profiles (Tasic, Discussion). Thus, a practitioner would acquire more informative data when classifying regulatory elements of a complex tissue such as retina and/or neuronal cells, as taught by Hartl and Tasic, respectively, by performing single-cell transcriptomic sequencing/ classification as taught by Tasic.
Furthermore, the results are predictable because Tasic has reduced to practice such a method in cortical cells, and Hartl has already taught that neuronal and retina cells are challenging in the same way in that they comprise rare cell populations in complex tissues (Hartl, page 11614, left column).
Furthermore, aside from the obviousness of single cell transcriptomic profiling as taught by Tasic and its application to Hartl, it is further obvious that the cells are not sorted by reporter or marker, as Tasic further teaches such microfluidic cell separation approaches in their Discussion (fifth paragraph). Furthermore, Tasic teaches a motivation to use this known technique as taught by Nguyen, because Tasic teaches that using such a non-marker sorting strategy gives additional information about a given complex cell population, where furthermore the substitution of FACS sorting with Fluidigm C1 is predictable because Tasic teaches that the results compliment each other (Discussion, fifth paragraph), where the non-FACS separation method is further useful because it can provide additional data about other cell types, is high-throughput, and cost-effective (Tasic Discussion Fifth paragraph and Nguyen page 4 left column final paragraph). The practitioner is therefore motivated to use such a separation method as taught by Tasic/Nguyen where the results are predictable.
Regarding claim 2, Hartl teaches that the regulatory element selectively increases expression of the transgene in the cell type (e.g., page 11615, right column, fourth paragraph, “revealed differences over three orders of magnitude”).
Regarding claims 3-4, Hartl teaches that a fraction of the tested regulatory elements drives detectable activity in rod cells this inherently means that some regulatory elements measured by Hartl did not drive detectable activity while some did (page 11615, right column, second paragraph). Thus, Hartl teaches that the regulatory element provides selective expression of the transgene that is at least 2-fold (claim 3) and/or 50% (claim 4) greater or less compared to expression driven by a control regulatory element in the same cell type (page 11615, right column, second paragraph). The term “control” regulatory element is not specifically restricted in the claim, and could thus reasonably be interpreted to be a negative control, where the increased expression of the above regulatory elements that are at least 2-fold or 50% greater could be compared with a negative control, and therefore read on claims 3-4.
Regarding claims 6-7, Hartl teaches that SAC and HC cells, when compared to other cell types, revealed differences of expression over three orders of magnitude (page 11615, right column, fourth paragraph). Thus, Hartl teaches that the regulatory element provides selective expression of the transgene at least 2-fold (claim 6) and/or 2% (claim 7) greater or less compared to expression of the transgene from the same regulatory element in a different cell type (page 11615, right column, fourth paragraph).
Regarding claims 34-36, Hartl teaches the viral vector AAV (Figure 3A). Furthermore, regarding claim 36, Hartl teaches that constructs were packaged in AAV serotype 8, i.e., AAV8 (page 11614, right column, final paragraph).
Regarding claim 39, Hartl teaches that they generated a catalogue of hundreds of putative regulatory elements in their methods (Abstract). Thus, Hartl teaches that the library of candidate regulatory elements comprise at least 16 candidate regulatory elements (Abstract).
Regarding claim 41, Hartl teaches that the transgene comprises a reporter assay (e.g., GFP, Figure 3).
Regarding claim 109, Hartl teaches that the cells are in vivo cells in a mouse (e.g., Figure 3).
Regarding claim 110, Hartl teaches separating cells by FACS, and further that FACS-sorting relies on sorting cells based on fluorescent labeling of markers (e.g., Figure 3, page 11609, left column, third paragraph, and page 11618, left column, second paragraph). Furthermore, Tasic also teaches that single-cell transcriptomics can be used to classify/identify cells based on unique marker genes (Discussion, first and second paragraph).
Claims 10-11 are rejected under 35 U.S.C. 103 as being unpatentable over Hartl (Hartl D et al. Nucleic Acids Res. 2017 Nov 16;45(20):11607-11621, submitted in applicant’s IDS filed 5/21/2024) in view of Tasic (Tasic B et al. Nat Neurosci. 2016 Feb;19(2):335-46) and Nguyen (Nguyen A et al. Front Immunol. 2018 Jul 4;9:1553) as applied to claims 1-4, 6-7, 34-36, 39, 41, and 109-110 above, and further in view of Cohen (Cohen SM et al. Neuron. 2016 Apr 20;90(2):292-307).
Regarding claims 10 and 11, a discussion of the teachings of Hartl/Tasic is discussed above in the rejection of claim 1. Furthermore, Hartl teaches that their method was successful, and should further be expanded to other cell types (page 11619, right column, first paragraph). Hartl further teaches that their method can contribute to the goal of providing more insight into expression drivers in vivo (page 11619, left column, final paragraph). Thus, Hartl teaches that their method is useful, practical, and can target disease relevant cell types (page 11619, left and right columns).
Furthermore, Tasic is directed to methods involving neuronal tissues, and teach that their method was reduced to practice in neuronal cells (Abstract, Discussion).
Hartl and Tasic do not teach that the regulatory element provides selective expression in PV neurons compared to non-PV neurons such as excitatory neurons.
Cohen is a research article which teaches transcription profiling in cell types, specifically neurons (Title, Abstract, and see document). Cohen and Hartl therefore overlap in subject matter and field of endeavor because both research articles teach methods of investigating transcriptional profiles in different cell types (see documents). Cohen teaches that understanding excitation-transcription profiling is relevant to understanding disease pathology states (Abstract). Cohen further teaches that gene expression as it relates to morphological development in neurons is poorly understood, and thus teaches that there is a motivation to further understand transcription/gene expression in neurons, as this understanding would help to understand disease states (Abstract). Furthermore, Cohen teaches that expression profiles and morphological/regulatory differences exist between PV neurons and excitatory neurons (Abstract, page 2 final paragraph). Furthermore, Cohen teaches that relatively little is known about gene expression changes in interneurons (page 2, second paragraph). Additionally, Cohen teaches that understanding gene expression/excitation-transcription coupling in PV cells is critical for understanding disease states (page 13, first paragraph). Thus, Cohen teaches both PV cells and excitatory neurons, that these two cells have different transcriptional responses to external stimuli, and further that understanding transcriptional/gene expression regulation is critical to understanding disease states that are not entirely understood (Abstract, page 2 paragraphs 2-3, page 13 first paragraph).
It would have been obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention to adapt the transcript profiling method rendered obvious by Hartl/Tasic to the PV/excitatory neurons taught by Cohen because such a combination is the simple substitution of one known element for another to obtain predictable results. In the present case, a practitioner would have simply substituted the study of the retina and interneurons of Hartl with PV/excitatory neurons taught by Cohen. Furthermore, a practitioner would be motivated to apply the method of Hartl/Tasic to PV/excitatory neurons because Cohen teaches that there is a need to understand transcription states in these two different cell populations as it would help to understand disease states (e.g., Abstract, page 2, page 13). Additionally, there is a reasonable expectation of success because Hartl has already reduced their method to practice and furthermore the method relies on elements which would also function in PV/excitatory cells (promoters, regulatory elements, reporter genes, vectors) and furthermore Tasic has already reduced to practice such RNA sequencing transcriptomic profiling in neuronal cells (Abstract).
Claims 37 is rejected under 35 U.S.C. 103 as being unpatentable over Hartl (Hartl D et al. Nucleic Acids Res. 2017 Nov 16;45(20):11607-11621, submitted in applicant’s IDS filed 5/21/2024) and Tasic (Tasic B et al. Nat Neurosci. 2016 Feb;19(2):335-46) and Nguyen (Nguyen A et al. Front Immunol. 2018 Jul 4;9:1553) as applied to claims 1-4, 6-7, 34-36, 39, 41, and 109-110 above, and further in view of Inagaki (Inagaki K et al. Mol. Ther. 2006 Jul;14(1):45-53).
Regarding claim 37, a discussion of the teachings of Hartl/Tasic as they relate to claims 1 and 34-36 is given above. Hartl teaches the use of AAV serotype 8 (AAV8) vectors in their methods (e.g., page 11614, right column, first paragraph). Furthermore, Hartl teaches that their library vectors comprising AAV vectors were used in mice (Figure 3).
Hartl, while teaching AAV vectors and AAV8, does not specifically teach AAV9.
Inagaki is a research article that teaches robust systemic transduction of AAV9 vectors into mice (Title, Abstract, and see document). Inagaki teaches that AAV9 are as robust and can be more robust than AAV8 vectors in mice (Abstract). Thus, Inagaki teaches that AAV9 vectors to be used for transduction in mice is a viable and ready alternative to AAV8 vectors, and further that such AAV9 vectors may be superior to AAV8 vectors.
It would have been obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention to substitute the AAV8 vectors used in mice taught by Hartl/Tasic with the AAV9 vectors taught by Inagaki because such a combination is the simple substitution of one known prior art element for another with predictable results. In the present case, a practitioner would have substituted the known AAV8 vector of Hartl with the AAV9 vector of Inagaki. Furthermore, Inagaki teaches that AAV9 vectors can be superior to AAV8 vectors, a teaching that would motivate a practitioner to adopt the AAV9 vectors of Inagaki (Inagaki, Abstract and see the entire document). Furthermore, because both Hartl and Inagaki concern AAV vectors used in mice to deliver vectors a practitioner would therefore have a reasonable expectation of success when combining the teachings of Hartl and Inagaki.
Claims 38 is rejected under 35 U.S.C. 103 as being unpatentable over Hartl (Hartl D et al. Nucleic Acids Res. 2017 Nov 16;45(20):11607-11621, submitted in applicant’s IDS filed 5/21/2024) and Tasic (Tasic B et al. Nat Neurosci. 2016 Feb;19(2):335-46) and Nguyen (Nguyen A et al. Front Immunol. 2018 Jul 4;9:1553) as applied to claims 1-4, 6-7, 34-36, 39, 41, and 109-110 above, and further in view of Savy (Savy A et al. Hum Gene Ther Methods. 2017 Oct;28(5):277-289).
Regarding claim 38, the teachings of Hartl/Tasic as they relate to claims 35-36 are discussed above. Hartl teaches the use of AAV vectors (Figure 3). Hartl teaches the use of AAV8 vectors (page 11614, right column, first paragraph).
Hartl does not specifically state that the AAV vectors comprise inverted terminal repeats.
Savy is a research article that teaches the impact of inverted terminal repeats (ITRs) on AAV vectors, specifically AAV8 vectors (Title, Abstract, see document). Savy teaches that ITRs are key elements of AAVs, and that these sequences are the only sequences conserved in recombinant AAV vectors, as they allow the AAV replication, encapsidation, and long-term maintenance and expression in target cells (Abstract). Thus, Savy teaches that recombinant AAV vectors routinely use ITR sequences (Abstract).
It would have been obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention to use ITR sequences as taught by Savy with the AAV vectors taught by Hartl. A practitioner would have been motivated to include Savy’s ITR sequences in order to render functional AAV vectors, as ITRs were already known in the art to be critical elements to be included with AAV vectors a taught by Savy, and specifically AAV8 vectors taught by both Savy and Hartl (Savy Abstract, Hartl page 11614, right column, first paragraph and Figure 3).
Claims 42 and 44-46 are rejected under 35 U.S.C. 103 as being unpatentable over Hartl (Hartl D et al. Nucleic Acids Res. 2017 Nov 16;45(20):11607-11621, submitted in applicant’s IDS filed 5/21/2024) and Tasic (Tasic B et al. Nat Neurosci. 2016 Feb;19(2):335-46) and Nguyen (Nguyen A et al. Front Immunol. 2018 Jul 4;9:1553) as applied to claims 1-4, 6-7, 34-36, 39, 41, and 109-110 above, and further in view of Swiech (Swiech L et al. Nat Biotechnol. 2015 Jan;33(1):102-6, filed with applicant’s IDS filed 5/21/2024).
Regarding claim 42, the teachings of Hartl/Tasic are discussed above. Hartl teaches that the transgene comprises a reporter gene (“GFP,” Figure 3). Hartl also teaches separating cell types transduced with AAV vectors using FACS (e.g., Figure 3).
Regarding claims 44 and 46, Hartl teaches that the barcodes are encoded immediately beside the transgene, upstream of the poly(A) tail of the mRNA, which is within the coding region of the gene (Figure 3 and caption beneath Figure 3).
Regarding claim 45, claim 45 recites “the barcode comprises alternative codons.” According to the specification, alternative codons are simply synonymous codons that are redundant with respect to an amino acid they encode (paragraph 74 of specification). Thus, the broadest reasonable interpretation of claim 45 includes any barcode sequence that comprises codons that have redundant coding with other codons in the genetic code. Hartl teaches 15 bp barcode sequences that are random sequences (page 11608, right column, fourth paragraph). Hartl therefore inherently teaches that the barcodes comprise “alternative codons” because such barcode codons would have redundancy with other amino-acid encoding codons.
Hartl does not directly teach that the reporter gene sequence comprises the barcode (claim 44) or that the sequence encoding the nuclear binding domain comprises the barcode (claim 46).
Hartl does not teach that the reporter gene is operably linked to a sequence encoding a nuclear binding domain.
Swiech is a research article which teaches the delivery of transgenes using AAV vectors of neuronal cells (Abstract). Thus, Swiech and Hartl overlap in subject matter and field of endeavor. Swiech teaches that transgenes such as GFP are fused to nuclear binding domains in AAV-directed vector delivery, and that such domains (”KASH” domains) direct the fused GFP transgene protein to the nuclear membrane, which enables the identification of neurons which have been transduced by AAV vectors (page 102, left column, third paragraph into right column first paragraph). Swiech teaches that GFP-KASH fusions can be used in FACS cell sorting to identify cells with AAV vectors 9 (page 102, right column, final paragraph).
It would have been obvious to a person of ordinary skill in the art the effective filing date of the claimed invention to modify the GFP transgenes taught by Hartl with nuclear binding domain fusions, as taught by Swiech, because such a combination is the simple combination of known prior art elements with predictable results. In the present case, tagging transgenes with nuclear binding domains is already a known method taught by Swiech which can be used to help identify AAV-vector delivered cells using FACS in neuronal cells (Abstract, page 102). It was therefore predictable that the methods taught by Swiech would work with Hartl because Hartl taught methods to be used with AAV vectors encoding GFP which can be sorted using FACS (Figure 3).
It would have been obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention to modify the barcode position taught by Hartl to be comprised either within the transgene taught by Hartl or the nuclear binding domain taught by Swiech, where such a combination would be obvious to try, where a practitioner would be choosing from a finite number of identified, predictable solutions, with a reasonable expectation of success. In the present case, a practitioner would be choosing from three regions: within the transgene, within the nuclear binding domain, or outside of these domains. The placement of a barcode is predictable in any of these locations because a barcode sequence would still function as a barcode regardless of its placement within a sequence. There is therefore a reasonable expectation that a barcodes function would not be lost if moving its location to one of the three regions of the sequence rendered obvious by Hartl and Swiech.
Claims 111-112 are rejected under 35 U.S.C. 103 as being unpatentable over Hartl (Hartl D et al. Nucleic Acids Res. 2017 Nov 16;45(20):11607-11621, submitted in applicant’s IDS filed 5/21/2024) in view of Tasic (Tasic B et al. Nat Neurosci. 2016 Feb;19(2):335-46) and Nguyen (Nguyen A et al. Front Immunol. 2018 Jul 4;9:1553) as applied to claims 1-4, above, and further in view of Sakaguchi (Sakaguchi M et al. Mol Biotechnol. 2014 Jul;56(7):621-30).
A discussion of Hartl, Tasic, and Nguyen is given above concerning claims 1-4.
Regarding claims 111-112, Hartl teaches the known CAG promoter (11608, right column, first paragraph) and controls used in their experiments (Figure 2, caption) but does not teach that it is the control promoter used.
Sakaguchi is a research article that focuses on the expression of transgenes from various promoters, and therefore directly overlaps in subject matter of Hartl, Tasic, and Nguyen. Furthermore, Sakaguchi teaches that the CAG promoter is a known, highly potent promoter (Abstract). Furthermore, given these properties, Sakaguchi teaches that the CAG promoter can be used as a control reference for the expression of transgenes (page 626, left column, third paragraph). Thus, the use of CAG as a reference/control promoter was already a known technique owing to the high potency/consistency of expression of the CAG promoter (page 626, left column, third paragraph, Figure 3A).
It would have been obvious to a person of ordinary skill in the art before the effective filing date to modify the method rendered obvious by Hartl, Tasic, and Nguyen, to include CAG as a control promoter because the use of CAG as a control/reference promoter for transgene expression is already a known technique as taught by Sakaguchi. Furthermore, a practitioner would be motivated to use CAG as a control regulatory element because it is a highly potent and therefore consistent expression driver (Sakaguchi).
Response to Arguments
The Applicant’s arguments submitted 11/14/2025 have been considered but are not persuasive. The Applicant argues that their amendments overcome the 103 rejection, as the teachings of Hartl, Tasic, Cohen, Inagaki, Savy, and Swiech do not teach that the cells are “not sorted based on the presence of a reporter gene or a natural cell-specific marker prior to sequencing.” This argument is not persuasive. As discussed above, Tasic directly references and teaches a prior study which relied upon an alternative sorting method: the microfluidic Fluidigm C1 separation technique, as discussed above. As evidenced by Nguyen, the Fluidigm C1 separation of cells does not rely upon sorting based on a reporter or marker gene. Thus, Tasic, by referencing and teaching the prior study (see Discussion of Tasic, fifth paragraph), does in fact teach a method wherein sorting not based on a reporter gene or natural marker (i.e., Fluidigm C1 separation, Discussion of Tasic, fifth paragraph). Furthermore, Tasic further teaches that such a separation method appears to expand the types of cells which can be identified and characterized using transcriptomic profiling (Discussion, fifth paragraph). Tasic further teaches that using such a method produces results which align with and in a sense expand upon data that is derived from cell-sorting based upon FACS sorting (Discussion, fifth paragraph). Thus, Tasic not only teaches such a method, where cells are not sorted based on the presence of a reporter gene or a natural cell marker, but also teaches a motivation to use such a method, where furthermore the results are predictable because Tasic teaches that the data of their study compliment the data of the fluidigm C1 study (Discussion, fifth paragraph). The Applicant’s argument is therefore not persuasive, as Tasic does appear to teach such a separation method which is not based on reporters/markers, and that there are benefits and predictability with using such a method.
The Applicant argues that Cohen, Inagaki, Savy, and Swiech do not cure the deficiencies of for instance Hartl and Tasic. However, the present rejection based upon Hartl, Tasic, and evidentiary teachings of Nguyen do not appear to be deficient with regards to rendering obvious the claim limitations of claim 1.
Conclusion
Any inquiry concerning this communication or earlier communications from the examiner should be directed to DOUGLAS CHARLES RYAN whose telephone number is (571)272-8406. The examiner can normally be reached M-F 8AM - 5PM.
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/D.C.R./Examiner, Art Unit 1635
/KIMBERLY CHONG/Primary Examiner, Art Unit 1636