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 .
This action is in response to the amendment, filed 09/26/2025, in which claim 18 was cancelled; claims 1, 4-6, 24-25 were amended.
Applicant’s arguments have been thoroughly reviewed, but are not persuasive for the reasons that follow. Any rejections and objections not reiterated in this action have been withdrawn. This action is FINAL.
Priority
Acknowledgment is made of applicant’s claim for foreign priority under 35 U.S.C. 119 (a)-(d). The certified copy has been filed in parent Application No. PCT/EP2021/061999, filed on 05/06/2021, claiming a priority date 05/06/2020.
Rejections withdrawn
The rejection of claims 1, 4-7, 9-10, 12, 16, 19, 23-25, 35, under 35 U.S 102(a)(1), has been withdrawn in view of Applicant’s amendments and arguments to the claims in the reply in the reply filed 09/26/2025.
The rejection of claim 18 under 35 U.S 103, is moot in view of Applicant’s cancellation of the claim in the reply in the reply filed 09/26/2025.
The rejection of claims 16-17, 36, under 35 U.S 103, is withdrawn in view of Applicant’s arguments and amendments of the claims in the reply in the reply filed 09/26/2025.
New Rejection necessitated by Claim Amendment
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.
Claims 1, 4-7, 9-10, 12, 16-17, 19, 23-25, 35-36, are rejected under 35 U.S.C. 103 as being unpatentable over Fahrenkrug et al. (“Fahrenkrug”, US 2019/0323031 A1, cited as reference 2 on IDS filed 10/31/2022) in view of Busser et al. (“Busser”, WO 2018/073391 A1).
This rejection is made to address the amendment to the claims in the reply filed 09/26/2025.
Regarding claims 1, 4-6, 17, 19, 36, Fahrenkrug teaches method for altering the genome of an animal cell, the method comprising: identifying a target DNA region within the animal cell, the target region comprising a target cleavage site; contacting the animal cell with a targeted nuclease such that the nuclease cleaves the target DNA region at the target cleavage site, wherein the targeted nuclease comprises one or more binding domains that specifically bind to one or more sequences within the target DNA region. The targeted nuclease can be selected from the group consisting of a transcription-activator-like effector nuclease (TALEN), a CRISPR-based nuclease ( e.g., CRISPR/Cas9), and a zinc finger nuclease (e.g., paragraph 0005). Fahrenkrug teaches the method comprises exposing the embryos or cells to single stranded DNA (ssDNA) that contains an exogenous sequence, with the genetic modification comprising the exogenous sequence. The ssDNA can be introduced into the cell after a vector encoding a TALEN is introduced into the cell. The ssDNA can be introduced into the cell between about 8 hours and about 3 days after the vector expressing a TALEN is introduced into the cell (It reads on administration of DNA template between 8 and 20 hours after TALEN endonuclease) (e.g., paragraph 0037; Fig. 1 [see below]).
Fig. 1:
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Fahrenkrug teaches target sequence encodes at least part of an endogenous allele, wherein the HDR template DNA sequence encodes an allele that is homologous to the endogenous allele flanked by sequences homologous to the target sequence in the chromosomal DNA of the cell,
wherein the allele that is homologous to the endogenous allele replaces the endogenous allele (e.g., paragraph 0032). Fahrenkrug teaches an AAV-delivered single stranded DNA template for homologous recombination at the bovine GDFS locus. a) TALENs (btGDF83.1, blue arrow) and a rAAV homologous recombination template (AAV-BBHDR)
were designed to introduce an 11 bp deletion into exon-3 of the bovine GDFS gene (Belgium Blue mutation) by homologous recombination (the single stranded DNA template of 1623 bp targeting bovine exon 3 of GDF8 locus flank the TALEN cut site and comprise homology regions at both arms , see Fig. 8) (e.g., paragraphs [0085]-[0404-0405]; example 22; Fig. 18 [see below]).
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Regarding claims 7, 9-10, Fahrenkrug teaches targeted nuclease can be selected from the group consisting of a transcription-activator-like effector nuclease (TALEN), a CRISPR-based nuclease ( e.g., CRISPR/Cas9), and a zinc finger nuclease. The targeted nuclease comprises delivering mRNA encoding the TALEN into the animal cell such that the mRNA is expressed to produce the TALEN within the cell (e.g., paragraph 0005). Transfection of TALEN encoding plasmids, containing an 11 base pair Belgian Blue cattle mutation, into Wagyu cells. Single stranded oligodeoxynucleotides (ssODNs) were found to be an effective template for TALEN stimulated HR. The same loci as above (Examples 6 and 10) were targeted to introgress the 11 base pair Belgian Blue cattle mutation into Wagyu cells. Two 76 base pair ssODNs (SEQ ID NOS: 501
and 502) were designed to mimic either the sense or antisense strand of the BB GDFS gene including the 11 base pair deletion. Four micrograms of TALEN encoding plasmids were transfected into Wagyu cells, and 0.3 nMol of ssODNs were either co-transfected with TALENS (N) or delivered 24
hours after TALEN nucleofection by either MirusLTl (M) reagent or Lipofectamine LTX reagent (L) (e.g., paragraph 0390; Fig. 19 [see below]).
Regarding claim 12, Fahrenkrug teaches the method can further comprise contacting the animal cell with a homology-dependent repair (HDR) template such that the HDR template is incorporated into
genomic DNA of the animal cell (e.g., paragraph 0014).
Regarding claim 16-17, Fahrenkrug teaches that the sequence of the homology-dependent repair template can be incorporated into the genomic DNA of the animal cell at a success rate of greater than 1 %. The HDR
template can be single-stranded DNA (e.g., paragraph 0014). Fahrenkrug teaches the use of single stranded oligonucleotides (ssOligos) as a template for homologous recombination at the bovine GDFS locus. TALENs (btGDF83.1, arrow) and two ssODNs were designed to introduce an 11 bp deletion into exon-3 of the bovine GDFS gene (Belgium Blue mutation) by homologous recombination. Each ssODN was 76 base pairs in length and were sense and antisense strands of the same target site Allele-specific PCR demonstrates that HDR induction is dependent on transfection btGDF83.1 TALENs and subsequent transfection of ssODNs using Lipofectamine LTX 24 hours later (e.g., paragraph 0086, Fig. 19 [see below]).
Fig. 19:
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Regarding claims 23-24 Fahrenkrug teaches the cell is selected from the group consisting of a primary cell, a primary somatic cell, a zygote,
a germ cell, a stem cell, an oocyte, and a sperm. CRISPR/Cas endonuclease can be introduced into the cell as mRNA. The cell can be homozygous for the allele or the gene introgression into the chromosomal DNA of the cell (e.g., paragraph 0030).
Fahrenkrug does not teach targeted insertion in a immune cell, as required by instant claims 1, 4-6, 25, 35. Fahrenkrug does not teach a DNA template with homology arms having 50-200 bp as required by instant claim 1. However, this is cured by Busser.
Busser teaches the method of the invention provides with the genetic insertion of exogenous coding sequence(s) that help the immune cells to direct their immune response against infected or malignant cells. These exogenous coding sequences are more particularly inserted under the transcriptional control of endogenous gene promoters that are up or downregulated upon immune cells activation, upon tumor microenvironment or life threatening inflammatory conditions or promoters that are insensitive to immune cells activation. The method of the invention contributes to improving the therapeutic potential and safety of engineered primary immune cells for their efficient use in cell therapy (e.g., lane 9, page 1). Busser teaches introducing the sequence specific endonuclease reagent and/or the donor template containing the gene of interest and sequences homologous to the target gene by transfecting ssDNA (oligonucleotides as non-limiting example), dsDNA (e.g., lane 3, page 8). Busser teaches Schematic representation of the donor sequences used in the experimental section to insert at the PD1 locus the exogenous sequences encoding IL-12 and gp130Fc. A: donor template (designated IL-12m-PD1) designed for site directed insertion of IL-12a and IL-12b coding sequences (SEQ ID NO:47 and 48) at the PD1 locus for obtaining co-transcription of IL-12a and IL-12b, while disrupting PD1 endogenous coding sequence. The right and left border sequences homologous to the PD1 locus sequences are at least 100pb long, preferably at least 200 pb long, and more preferably at least 300 pb long (e.g., lane 26, page 10; Fig 16 [see below]).
Fig. 16:
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Busser teaches "immune cell" is meant a cell of hematopoietic origin functionally involved in the initiation and/or execution of innate and/or adaptative immune response, such as typically CD3 or CD4 positive cells. The immune cell according to the present invention can be a dendritic cell, killer dendritic cell, a mast cell, a NK-cell, a B-cell or a T-cell selected from the group consisting of inflammatory T-lymphocytes, cytotoxic T-lymphocytes, regulatory T-lymphocytes or helper T-lymphocytes (e.g., lane 29, page13). Busser teaches that these cells form a population of cells, which preferably originate from a single donor or patient. These populations of cells can be expanded under closed culture recipients to comply with highest manufacturing practices requirements and can be frozen prior to infusion into a patient, thereby providing "off the shelf" or "ready to use" therapeutic compositions (e.g., lane 1, page 51). Busser teaches a method for treating a patient in need thereof, wherein said method comprises: preparing a population of engineered primary immune cells; optionally, purifying or sorting said engineered primary immune cells; activating said population of engineered primary immune cells upon or after infusion of said cells into said patient (e.g., lane 1, page 128; claim 69).
Based on these teachings, it would have been prima facie obvious to one of ordinary skill in the art, before the effective filing date of the claimed invention, to combine the teachings of Fahrenkrug -a method for altering the genome of an animal by sequential transfection of targeted nuclease selected from the group consisting of a transcription-activator-like effector nuclease (TALEN), a CRISPR-based nuclease and HDR template DNA sequence that encodes an allele that is homologous to the endogenous allele flanked by sequences homologous to the target sequence, where the DNA template can be introduced into the cell between about 8 hours and about 3 days after transfecting the vector expressing a specific-endonuclease into the cell with the teachings of Busser -genetic insertion of exogenous coding sequence into immune cells, using specific-endonuclease and a DNA template consisting of a right and left border sequences homologous to the gene locus sequences are at least 100pb long, preferably at least 200 pb long, and more preferably at least 300 pb long; for someone skilled in the art would have been obvious to use both teachings to achieve the predictable result of obtaining a method for altering the genome of immune cells suitable for using short single-stranded oligodeoxynucleotide for delivering as DNA exogenous sequences for targeting specific genes in immune cells.
One of ordinary skill in the art before the effective filing date of the invention would have been motivated to do so in order to develop a method for altering the genome of an engineered primary immune cells by insertion of exogenous coding sequence that help the immune cells to direct their immune response against infected or malignant cells by using site-specific endonucleases and a DNA template by targeting a specific gene locus.
The MPEP states “In the case where the claimed ranges "overlap or lie inside ranges disclosed by the prior art" a prima facie case of obviousness exists. In re Wertheim, 541 F.2d 257, 191 USPQ 90 (CCPA 1976); In re Woodruff, 919 F.2d 1575, 16 USPQ2d 1934 (Fed. Cir. 1990) (The prior art taught carbon monoxide concentrations of "about 1-5%" while the claim was limited to "more than 5%." The court held that "about 1-5%" allowed for concentrations slightly above 5% thus the ranges overlapped.); In re Geisler, 116 F.3d 1465, 1469-71, 43 USPQ2d 1362, 1365-66 (Fed. Cir. 1997) (Claim reciting thickness of a protective layer as falling within a range of "50 to 100 Angstroms" considered prima facie obvious in view of prior art reference teaching that "for suitable protection, the thickness of the protective layer should be not less than about 10 nm [i.e., 100 Angstroms]." The court stated that "by stating that ‘suitable protection’ is provided if the protective layer is ‘about’ 100 Angstroms thick, [the prior art reference] directly teaches the use of a thickness within [applicant’s] claimed range."). See also In re Bergen, 120 F.2d 329, 332, 49 USPQ 749, 751-52 (CCPA 1941) (The court found that the overlapping endpoint of the prior art and claimed range was sufficient to support an obviousness rejection, particularly when there was no showing of criticality of the claimed range).
The claimed range of 5-20 hours substantially overlaps with the disclosed range by Fahrenkrug of 8 hours to 3 days, specifically the 8-20 hour segment is common. The claimed range therefore, is largely a selection of a subrange from the prior art known operable range. The motivation for narrowing the range is clear, a person of ordinary skill in the art is motivated to optimize a therapeutic or cellular process to maximize efficiency. The person of ordinary skill in the art knows that the endonuclease must be first expressed, reach a peak activity and cleave the DNA, and that the subsequent repair process (Homology directed repair) is transient and active in the early phases of the cell cycle. Given that Fahrenkrug broad window of 8h to 3 days, the person of ordinary skill in the art would have been motivated to conduct routine experimentation to test the earliest effective time points to synchronize the DNA template with the peak cleavage and repair activity. The selection of 5h-20h window is thus a predictable refinement of the know teaching.
Conclusion
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 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.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to JULIO GOMEZ RODRIGUEZ whose telephone number is (571)270-0991. The examiner can normally be reached Monday - Friday 8:00 am - 5:00 pm.
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/JULIO WASHINGTON GOMEZ RODRIGUEZ/Examiner, Art Unit 1637
/J. E. ANGELL, Ph.D./Primary Examiner, Art Unit 1637