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 .
Status of the Claims
Claims 26 and 27 are newly added,
Claim 25 is newly cancelled,
Claims 4, 8, 14, 15, 17 and 20 are newly amended.
Claims 1-24 and 26-27 are pending.
Claims 21-25 were withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim. Election was made without traverse in the reply filed on 03/27/2025.
Claims 1-20 and 26-27 have been examined on the merits.
Withdrawn Objections & Rejections
Applicant's response filed 8/28/2025 has been considered. Rejections and/or objections not reiterated from the previous Office action mailed 05/30/2025 are hereby withdrawn.
The objections and rejections presented herein represent the full set of objections and rejections currently pending in the application.
Claim Objections
This is a new objection
Regarding claim 8: The claim is objected to because of the following informalities: the claim recites “determining from the imaging aggregated CART cells”. This is grammatically incorrect. Replacing the phrase “determining from the imaging” with a phrase such as “evaluating the imaging for” or “observing the imaging for” would remedy this objection.
Appropriate correction is required.
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.
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, 4-9, 11, 16, 20 and 26-27 are rejected under 35 U.S.C. 103 as being unpatentable over Roth et al (Nature (2018)559:405-435; as cited in the IDS filed 05/07/2024) in view of Kriegsmann et al (BJC (2018)38-44) and Jenkins et al (Current Opinion in Chemical Engineering (2018) 22;26-33).
This is a new rejection, substantially similar to a previous rejection, however any aspect of Applicants traversal that is relevant to the rejection as newly written is addressed below.
Regarding claims 1, 4, 5, 7, 16 and 26-27: Claim 1 recites “chimeric antigen receptor”. The instant specification defines chimeric antigen receptor (CAR) as “a recombinant fusion protein that has an antigen-specific extracellular domain coupled to an intracellular domain that directs the cell to perform a specialized function upon binding of an antigen to the extracellular domain” (p8 ¶0022).
The claim recites “CART cells”. The instant claim 1 defines a CART cell as a T cell with a transgene containing a chimeric antigen receptor (claim 1).
The claims are interpreted with the definitions as stated supra.
Roth teach reprogramming human T cell function for therapeutic purposes and specificity using non-viral genome targeting (title).
Roth teach electroporation of human primary T cells with CRISPR-Cas9 ribonucleoprotein (RNP) complexes using linear dsDNA templates for homology-directed repair (HDR) (p405 col 1 para 2).
Extended data figure 9a shows the dsDNA HDR template comprises a transgene flanked by homology arms that are complementary to sequences on both sides of the cleavage site in the T cell expressed gene (TRAC), and that the transgene is inserted into EXON 1 of TRAC (p 427).
Roth confirm the transgene is specifically integrated into the cleavage site of the T cell expressed gene locus created by the Cas9 RNP in the CART cells by observing expression of the inserted transgene (Fig 4, p 408 col 1 para 1).
The T cells are cultured in XVivo15 medium with IL-2 (an ex vivo method using Xeno-free media, as evidence by XVivo15 product sheet) (p 410 col 1 para 3).
Roth replace the endogenous T cell receptor (TCR; a T cell expressed gene) by targeting the TCR-alpha constant region (TRAC) (p407 col 2 para 5), to insert engineered receptors, TCR-beta and TCR-alpha, that recognize (binds) the NY-ESO-1 tumor antigen, to generate NY-ESO-1 cells (p408 col 1 para 1).
Fig 4h shows solid tumor volume is reduced in a mouse tumor model when mice are injected with the NY-ESO-1 CART cells (Fig 4h), demonstrating the CART cells have activity against an antigen on a solid tumor in vivo.
While Roth teach a non-viral method of site-specifically inserting a transgene encoding an antigen specific extracellular domain into a T cell, however Roth do not explicitly teach that the transgene comprises a fusion protein with the antigen specific extracellular domain coupled to an intracellular domain by a transmembrane domain.
Kriegsmann teach that chimeric antigen receptor (CAR) T cells are genetically modified T cells that express a CAR directed against specific tumor antigens (abstract). Kriegsmann also teach that CART immunotherapy shows promising results in early clinical trials as an anti-cancer therapy (p38 col2 ¶1). Kriegsmann teach CARs are directed against specific cell-surface antigens (p38 col2 ¶2). Figure 1c shows a CAR comprises an antigen specific extracellular domain coupled to an intracellular domain by a transmembrane domain, and that the antigen specific extracellular domain comprises an antibody variable region (p 42).
It would have been obvious to one of ordinary skill in the art to adapt the methods of Roth drawn to a non-viral method of site-specifically inserting a transgene encoding an antigen specific extracellular domain into a T cell by inserting a transgene comprising a fusion protein with the antigen specific extracellular domain coupled to an intracellular domain by a transmembrane domain.
One would have been motivated to modify the method of Roth to insert an antigen specific extracellular domain coupled to an intracellular domain by a transmembrane domain because Roth teach that the need for recombinant viral vectors to genetically reprogram T cells has slowed down research and clinical because of lengthy and expensive manufacturing and testing (abstract) and Kriegsmann teach CAR T cells are promising anti-cancer therapies.
One would have had a reasonable expectation of success because both methods are drawn to genetic modification of T cells and T cell receptors for the purposes of targeting tumor antigen expressing cells.
The teachings of Roth are discussed supra. While Roth teach culture in xeno-free medium (XVivo15), Roth do not teach culturing cells in the absence of serum.
Jenkins teach serum content is a significant hurdle in T cell cultivation, and that serum-free T cell cultivation will reduce potential harmful elements and simplify T cell purification steps (p29 col1 ¶2). Jenkins further teach antigen-specific T cell cultivation has been undertaken with xeno-free serum replacement (SR), resulting in comparable T cell growth kinetics with FBS and reduced risk of adventitious infectious pathogens (p29 col1 ¶2).
It would have been obvious to one of ordinary skill in the art to adapt the methods of Roth drawn to non-viral insertion of a transgene by culturing the cells in xeno-free conditions by using SR in place of serum as taught by Jenkins. Jenkins teach that cultivation in serum-free conditions reduces potential harmful elements and simplifies T cell purifications steps.
Accordingly, one of ordinary skill in the art would have been motivated to modify the method as taught by Roth with the teachings of Jenkins for the purposes of a more efficient and protocol with reduced harmful elements.
One would have had a reasonable expectation of success because both inventions are directed to T cell cultivation under defined conditions.
Regarding claim 6: The teachings of Roth are discussed supra. Roth teach NY-ESO-1 cells are co-incubated with human melanoma cell lines and show robust killing of the target cells (Fig9n, p 427).
Regarding claim 8: The claim recites “imaging the population of CART cells and determining from the imaging aggregated CART cells”. The broadest reasonable interpretation of this limitation is imaging (collecting an image) of a population of cells such that aggregation can be observed (if aggregation occurs). It is noted that observing aggregated cells is not required by this claim limitation.
The claim recites optional components “optionally selecting a population of aggregated CART cells” which are interpreted as not limiting the claim in any way.
The claims are interpreted as discussed supra.
The teachings of Roth are discussed supra. Roth also teach imaging the population of modified T cells (Figure 2b) and the images can be used to determine (identify) aggregated cells.
Regarding claim 9: The teaching of Roth are discussed supra. Roth also teach electroporation of HDR template at a concentration of 2ug/ul (2000 ng/ul) (p410 col2 para 3).
Roth do not teach the double-stranded HDR template has an OD260/OD280 and/or OD260/OD230 of a specific ratio.
With regard to the OD260/OD280 and/or OD260/OD230 of the HDR template, generally, differences in concentration or temperature will not support the patentability of subject matter encompassed by the prior art unless there is evidence indicating such concentration or temperature is critical. “[W]here the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation.” In re Aller, 220 F.2d 454, 456, 105 USPQ 233, 235 (CCPA 1955) (MPEP 2144.05).
The selection of a specific level of purity of the HDR template (which would result in a more accurate concentration of HDR template introduced to the target cells, and the exclusion of impurities from the target cells) clearly would have been a routine matter of optimization and experimentation on the part of the artisan of ordinary skill, said artisan recognizing that the amount and purity of the HDR template introduced to the cultured cells would have been affected by these parameters.
Regarding claim 11: The teachings of Roth are discussed supra. Roth teach a dsDNA HDR template designed to introduce an N-terminal green fluorescent protein GFP into human primary T cells to generate CD4-GFP CART cells (p405 col 1/2 para 2/1-2). Extended data figure 4 shows the HDR template, the GFP tag, is flanked by a 5’ and 3’ homology arm (p417).
Regarding claim 20: The teachings of Roth are discussed supra. Roth also teach Figure 4b, 12.3% of the non-viral genome targeted cells express the transgene NY-ESO-1, reading on the instant claim of more than 4% of the population of unmodified T cells have the transgene inserted into their genomes (p408).
Claims 2-3 are rejected under 35 U.S.C. 103 as being unpatentable over Roth et al (Nature(2018)559:405-435; as cited in the IDS filed 05/07/2024) in view of Kriegsmann et al (BJC (2018)38-44) and Jenkins et al (Current Opinion in Chemical Engineering (2018) 22;26-33) as applied to claim 1, and further in view of Li et al (PLOS one(2014)9(8) 1-10) and Ran et al (Nature Protocols (2013)8; 2281-2308).
This is a new rejection, substantially similar to a previous rejection, however any aspect of Applicants traversal that is relevant to the rejection as newly written is addressed below.
Regarding claims 2 and 3: The teachings of Roth are discussed above. Roth also teach that the 5’ and 3’ homology arms for the dsHDR template are ~300bp (Fig 4a p408).
Roth do not teach the homology arms have a length of 400-1000bp or 450-750bp.
Li teach optimization of genome engineering with CRISPR/CAS9 (title). Li disclose increasing the length of the homology arms of the homology-directed repair template strongly enhances targeting efficiency in CRISPR/Cas9 editing systems (Abstract). Li teach that, while larger insert sizes can have reduced efficiency of genome modification, increasing the homology arm size increases modification frequency (p9/10 col 2/1 para 3/1).
Ran et al teach genome engineering using CRISPR-Cas9 (title). Ran disclose homology arms on each side of the repair template can vary in length, but are typically longer than 500 bp (p2284 col2 para 2).
It would have been prima facie obvious to one of ordinary skill in the art to adapt the methods of Roth with the teachings of Li and Ran by using longer homology arms that can improve editing efficiency of HDR templates with larger inserts.
One having ordinary skill in the art would have understood that more efficient gene editing would enable a CRISPR/Cas9 system to generate a potentially therapeutic population of modified T cells more efficiently and therefor with less expense.
One would have had a reasonable expectation of success because Ran disclose typical homology arms are longer than 500 bp and Li disclose longer homology arms are beneficial to HDR template insertion efficiency.
Claim 10 is rejected under 35 U.S.C. 103 as being unpatentable over Roth et al (Nature(2018)559:405-435; as cited in the IDS filed 05/07/2024) in view of Kriegsmann et al (BJC (2018)38-44) and Jenkins et al (Current Opinion in Chemical Engineering (2018) 22;26-33) as applied to claim 1, and further in view of Sjoukje et al (Nat Rev Drug Discov(2015)14(7):499-509).
This is a new rejection, substantially similar to a previous rejection, however any aspect of Applicants traversal that is relevant to the rejection as newly written is addressed below.
The teachings of Roth and Kriegsmann are discussed supra. Kriegsmann also teach the intracellular domain comprises a CD3 zeta intracellular domain (Fig 1c). Kriegsmann is silent on the nature of the transmembrane domain.
Sjoukje teach CAR intracellular domains comprise an intracellular CD3 zeta domain (Figure 1c) and that the first fusion receptors were coupled to the CD3 intracellular domain (Figure 1). Sjoukje also teach that the transmembrane domain of the CAR CD3 zeta facilitates enhanced anti-tumor function (p25 box1, Figure 1d).
It would have been prima facie obvious to one of ordinary skill in the art to adapt the methods of Roth and Kriegsmann with the teachings of Sjoukje, to include a CD3 zeta intracellular and transmembrane domain in the receptor.
One having ordinary skill in the art would have been motivated to modify the method of Roth and Kriegsmann to include a CD3 zeta intracellular and transmembrane domain because Sjoukje teach the CD3 zeta intracellular domain is one of the first intracellular domains used, and as such one of ordinary skill in the art would understand that the function of the intracellular domain is well studied and understood, and the CD3 zeta transmembrane domain facilitates enhanced antitumor function and cytokine production (p503 Box 1).
One would have had a reasonable expectation of success because the methods are drawn to genetically engineering T cells to express chimeric T cell receptors.
Claims 12-13 are rejected under 35 U.S.C. 103 as being unpatentable over Roth et al (Nature(2018)559:405-435; as cited in the IDS filed 05/07/2024) in view of Kriegsmann et al (BJC (2018)38-44) and Jenkins et al (Current Opinion in Chemical Engineering (2018) 22;26-33) as applied to claim 1, and further in view of Lu et al (RNA(2003)9:1-13) and Lanoix et al (EMBP(1988)7(8)1-8).
This is a new rejection, substantially similar to a previous rejection, however any aspect of Applicants traversal that is relevant to the rejection as newly written is addressed below.
Regarding claims 12 and 13: The teachings of Roth are discussed supra. Roth also teach the double stranded HDR template is targeted to the 5’ end of the first exon of TRAC to generate NY-ESP-1 CART cells (Extended Data Figure 9a, p427). Figure 9a also teaches the HDR template sequentially comprises a 5’ (left) homology arm- the self-cleaving peptide T2A- the CAR gene (NY-ESO-1)-a 3’ (right) homology arm.
Roth do not explicitly teach a splice acceptor site 3’ adjacent to the 5’ homology arm or a poly A terminator 5’ adjacent of the 3’ homology arm (fig 9a pa 427).
Lu et al (RNA(2003)9:1-13) teach mRNA splicing invariably enhances the level of gene expression (abstract).
Lanoix et al (EMBP(1988)7(8)1-8) teach rabbit beta-globin poly A terminator directs efficient termination of transcription of polyomavirus DNA.
It would have been prima facie obvious for one of ordinary skill in the art at the time of the effective filing date to incorporate a splice acceptor and a rabbit beta-globin poly a terminator with the non-viral double-stranded HDR template taught by Roth because Lu teach mRNA splicing enhances the level of gene expression and Lanoix teach the poly A terminator directs efficient termination of transcription.
One would have been motivated to modify the method of Roth with the teachings of Lu and Lanoix to enhance the levels of correct transgene expression. One of ordinary skill in the art would understand that efficient termination of transcription reduces improperly transcribed transgenes due to read-through errors and that improved transgene expression would result in a more efficient and therefore less expensive method.
One would have had a reasonable expectation of success because the teachings are all directed to methods of transgenes expression in human cells.
Claim 17 is rejected under 35 U.S.C. 103 as being unpatentable over Roth et al (Nature (2018)559:405-435; as cited in the IDS filed 05/07/2024) in view of Kriegsmann et al (BJC (2018)38-44) and Jenkins et al (Current Opinion in Chemical Engineering (2018) 22;26-33) as applied to claim 1, and further in view of Terrett et al (US 2018/0325955 A1).
This is a new rejection as necessitated by the claim amendments.
Regarding claim 17: The teachings of Roth are discussed above. Roth do not teach a guide RNA which comprises Seq ID NO: 9.
Terrett teach genome-edited cells engineered to express a chimeric antigen receptor construct (abstract). Terrett disclose the gRNA sequence Seq ID: 51, which comprises 100% sequence identity with the instant Seq ID NO:9 (p3 ¶0029).
It would have been prima facie obvious for one of ordinary skill in the art at the time of the effective filing date to combine the gRNA sequence, seq ID 51, as taught by Terrett with genome editing method taught by Roth because it would have been obvious to combine prior art elements according to known methods to yield predictable results. Combining the gRNA disclosed by Terrett with non-viral editing method of Roth would have led to predictable results with a reasonable expectation of success because both inventions teach using an gRNA to direct genome editing in the native TRAC locus of the T-cell.
Claim 18 is rejected under 35 U.S.C. 103 as being unpatentable over Roth et al (Nature(2018)559:405-435; as cited in the IDS filed 05/07/2024) in view of Kriegsmann et al (BJC (2018)38-44) and Jenkins et al (Current Opinion in Chemical Engineering (2018) 22;26-33) as applied to claim 1, and further in view of Phelan et al (Stem Cell Investigation (2018)5:221-4) and as evidenced by ThermoFisher (Difference Between: 96-Well Plate Bottom Shapes [online]. COMPANY:ThermoFisher [retrieved on 01/04/2026]. Retrieved from the Internet: <URL: https://www.fishersci.com/shop/products/abgene-96-well-1-2ml-polypropylene-deepwell-storage-plate/AB1127)
This is a new rejection.
Regarding claim 18: The teachings of Roth are discussed above. Roth do not teach culturing in round bottom culture wells or culture in a low culture volume (20% standard) of cell culture medium.
As evidenced by ThermoFisher, round bottom wells provide optimal sample recovery (p1, downloaded from the internet 01/04/2026).
Phelan teach cell culture in low volumes exhibit similar growth as traditional protocols with improved consistency (p2 col2 ¶2). Phelan further teach that low-volume multi-well setups provide a wide-range of options possible through customization.
It would have been obvious for one of ordinary skill in the art to modify the method of Roth with the teaching of ThermoFisher and Phelan, to culture cells in round bottom plates using a low culture volume.
One would have been motivated to modify the method of Roth to use round bottom plates with a low culture volume to improve optimize sample recovery and to improve consistency of cell cultures. Furthermore, one of ordinary skill in the art would recognize that culturing cells in a low culture volume would reduce the amount of reagents required for cell culture and thus decrease cost of the cell culture.
One would have had a reasonable expectation of success because one of ordinary skill in the art would know round bottom plates are commonly used for sample processing and culture of cells in suspension and that cell culture volumes would have been a matter of routine optimization based on optimization of the cell culture protocol and cell culture protocols in the art.
Claim 19 is rejected under 35 U.S.C. 103 as being unpatentable over Roth et al (Nature(2018)559:405-435; as cited in the IDS filed 05/07/2024) in view of Kriegsmann et al (BJC (2018)38-44) and Jenkins et al (Current Opinion in Chemical Engineering (2018) 22;26-33) as applied to claim 1, and further in view of Sadelain et al (WO 2019/157454) and Zhang et al (Blood Advances (2018) 2:14;1691-1696).
Regarding claim 19: The teachings of Roth are discussed above. Roth also teaches T cells are isolated from healthy human donors (p409 col 1 para 1).
Roth does not teach the T cells are autologous or allogenic T cells.
Sadelain teach novel design of T cell receptors and engineered immunoresponsive cells comprising the same (abstract). Sadelain disclose genetic modifications at the TRAC locus to permit generation of T cells expressing a chimeric antigen receptor (abstract).
Sadelain disclose the immunoresponsive cell is an autologous T cell (claims 42, 43) and that the method could advance both autologous and allogenic T cell therapies (Example 1, p76 ln 1-5).
Zhang teach allogeneic and autologous CAR-T cell therapy to treat a leukemic patient (title). Zhang disclose CAR-T studies usually use autologous CAR-T cells, however allogeneic CAR-T cells may be more robust, though could result in graft-versus-host disease (abstract).
It would have been prima facie obvious to one of ordinary skill in the art to adapt the methods of Roth with the teachings of Sadelain and Zhang to engineer autologous or allogeneic T cells.
One having ordinary skill in the art would have understood that using autologous T cells would reduce the likelihood of the patient developing graft vs host disease, while allogeneic T cells may provide a source of T-cells for those whom use of allogeneic T cells are not possible.
One would have had a reasonable expectation of success because Zhang teaches that CAR-T therapies in humans are almost always conducted with allogeneic CAR-T cells, but autologous CAR-T cell therapy results in complete remission for a patient that failed standard treatment.
Allowable Subject Matter
Claims 14-15 are objected to as being dependent upon a rejected base claim, but would be allowable if rewritten in independent form including all of the limitations of the base claim and any intervening claims.
The claims recite amplifying a sequence from Seq ID NO: 1 using primer sets of Seq ID NOs:11/12 or Seq ID NOs:17/18. Primer pairs with the sequences of Seq ID NOs: 11/12 and 17/18 appear free from the art.
Response to Arguments
The responses are directed to the Arguments filed 8/28/2025 and are directed to Arguments relevant to the rejection as newly written and discussed supra.
Regarding the Sequence Disclosures/Drawings:
Applicant argues that the amended figures 1A and 1I, filed on 08/28/2025 overcome the rejection to the drawings in the office filed 05/30/2025.
The amended figures filed on 08/28/2025 comprise Seq ID NO: identifiers for the relevant sequences and thus the argument is persuasive. The rejections/objections over the Drawings and Sequence Disclosures are withdrawn.
Regarding Arguments directed to 35 USC § 112(a):
Applicant's arguments filed on 8/28/2025 have been fully considered and they are persuasive.
Applicant submits amended the claim to replace SEQ ID NOs. 2 and 3 with the gRNA of SEQ ID NO: 9.
This overcomes the rejection under 35 USC § 112(a) in the office action filed 05/30/2025 and the rejection is withdrawn.
Regarding Arguments directed to 35 USC § 112(b):
Applicant's arguments filed on 8/28/2025 have been fully considered and they are persuasive.
Regarding claim 4: Applicant has deleted the “such as” phrase and has thus overcome the rejection
Regarding claim 8: The amendment filed 8/28/2025 renders the rejection moot.
Regarding claim 20: Applicant has amended the claim to provide clarification and thus overcome the rejection.
This is found persuasive and the rejections over 35 USC § 112(b) in the office action filed 05/30/2025 have been withdrawn.
Regarding Arguments directed to 35 USC § 101:
Applicant’s arguments filed on 8/28/2025 with respect to claim 8 under 35 USC § 101 have been fully considered and are persuasive.
The rejection over 35 USC § 101 in the office action filed 05/30/2025 is withdrawn.
Regarding Arguments directed to 35 USC § 102:
Applicant's arguments filed on 8/28/2025 have been fully considered and they are found persuasive.
Applicant submits that Roth includes FBS in the culture medium and thus the method of Roth does not comprise xeno-free medium. Applicant also submits the genetically modified T cells do not explicitly comprise an intracellular domain and thus do not read on the limitations as set forth in claim 1.
Roth does disclose use of FBS with the culture medium and Roth does not disclose an intracellular domain and thus the arguments are found persuasive.
The rejection over 35 USC § 102 in the office action filed 05/30/2025 has been withdrawn. A new rejection over 35 USC § 103 has been entered.
Applicant submits claim 14 is not anticipated by Roth. This argument is moot in view of the withdrawn rejection under 35 USC § 102 in the office action filed 05/30/2025.
Regarding Arguments directed to 35 USC § 103:
Applicant's arguments filed on 8/28/2025 have been fully considered and arguments which pertain to the newly entered rejection over 35 USC § 103 are addressed below.
Regarding claims 2-3:
Applicant’s arguments filed on 8/28/2025 have been fully considered and are unpersuasive.
Applicant argues that increasing the homology arms to 588 and 499 base pairs, compared to 383 and 391 base pairs in Example 2 of the instant specification increased CAR knock in percentages, which could not be predicted based on the prior art.
The argument of an unexpected result (improved function) claimed by Applicant is not commiserate with the scope of the claim. The claim limitations allows for the homology arms to be in the range of 400-1000bp and in the range of 450-750bp. Applicant provides a single example of homology arms with increased length (588bp and 499bp) which produce the unexpected result.
The rejection over 35 USC § 103 is maintained.
Regarding claim 8:
Applicant’s arguments filed on 8/28/2025 have been fully considered and are unpersuasive.
In response to applicant’s argument that the references fail to show certain features of the invention, it is noted that the features upon which applicant relies (i.e., improvement to cell health/genome editing based on level of cellular aggregation) are not recited in the rejected claim(s). Although the claims are interpreted in light of the specification, limitations from the specification are not read into the claims. See In re Van Geuns, 988 F.2d 1181, 26 USPQ2d 1057 (Fed. Cir. 1993).
The rejection over 35 USC § 103 is maintained.
Regarding claim 9:
Applicant’s arguments filed on 8/28/2025 with have been fully considered and are unpersuasive.
Applicant argues the additional round of purification to provide the claimed concentration and purity of the HDR template improved the efficiency of site-specifically inserting a CAR could not have been predicted based on the prior art.
In response to applicant’s argument that the references fail to show certain features of the invention, it is noted that the features upon which applicant relies (i.e., an additional round of purification, changed efficiency of site-specifically inserted transgene) are not recited in the rejected claim(s). Although the claims are interpreted in light of the specification, limitations from the specification are not read into the claims. See In re Van Geuns, 988 F.2d 1181, 26 USPQ2d 1057 (Fed. Cir. 1993).
The rejection over 35 USC § 103 is maintained.
Regarding claims 10:
Applicant’s arguments filed on 8/28/2025 have been fully considered and are unpersuasive.
Applicant submits the combination of Roth and Sjoukje do not render the claim obvious. As discussed supra and reiterated below, the instant rejection relies upon the combined teachings of Roth, Kriegsman and Jenkins (in the instant rejection for claim 1, upon which claim 10 depends) and further relies upon the teachings of Sjoukje for the instant rejection of claim 10.
Regarding claim 1: Roth teach reprogramming human T cell function for therapeutic purposes and specificity using non-viral genome targeting (title).
Roth teach electroporation of human primary T cells with CRISPR-Cas9 ribonucleoprotein (RNP) complexes using linear dsDNA templates for homology-directed repair (HDR) (p405 col 1 para 2).
Extended data figure 9a shows the dsDNA HDR template comprises a transgene flanked by homology arms that are complementary to sequences on both sides of the cleavage site in the T cell expressed gene (TRAC), and that the transgene is inserted into EXON 1 of TRAC (p 427).
Roth confirm the transgene is specifically integrated into the cleavage site of the T cell expressed gene locus created by the Cas9 RNP in the CART cells by observing expression of the inserted transgene (Fig 4, p 408 col 1 para 1).
The T cells are cultured in XVivo15 medium with IL-2 (an ex vivo method using Xeno-free media, as evidence by XVivo15 product sheet) (p 410 col 1 para 3).
Roth replace the endogenous T cell receptor (TCR; a T cell expressed gene) by targeting the TCR-alpha constant region (TRAC) (p407 col 2 para 5), to insert engineered receptors, TCR-beta and TCR-alpha, that recognize the NY-ESO-1 tumor antigen, to generate NY-ESO-1 cells (p408 col 1 para 1).
Fig 4h shows solid tumor volume is reduced in a mouse tumor model when mice are injected with the NY-ESO-1 CART cells (Fig 4h), demonstrating the CART cells have activity against an antigen on a solid tumor in vivo.
While Roth teach a non-viral method of site-specifically inserting a transgene encoding an antigen specific extracellular domain into a T cell, however Roth do not explicitly teach that the transgene comprises a fusion protein with the antigen specific extracellular domain coupled to an intracellular domain by a transmembrane domain.
Kriegsmann teach that chimeric antigen receptor (CAR) T cells are genetically modified T cells that express a CAR directed against specific tumor antigens (abstract). Kriegsmann also teach that CART immunotherapy shows promising results in early clinical trials as an anti-cancer therapy (p38 col2 ¶1). Kriegsmann teach CARs are directed against specific cell-surface antigens (p38 col2 ¶2). Figure 1c shows a CAR comprises an antigen specific extracellular domain coupled to an intracellular domain by a transmembrane domain, and that the antigen specific extracellular domain comprises an antibody variable region (p 42).
It would have been obvious to one of ordinary skill in the art to adapt the methods of Roth drawn to a non-viral method of site-specifically inserting a transgene encoding an antigen specific extracellular domain into a T cell by inserting a transgene comprising a fusion protein with the antigen specific extracellular domain coupled to an intracellular domain by a transmembrane domain.
One would have been motivated to modify the method of Roth to insert an antigen specific extracellular domain coupled to an intracellular domain by a transmembrane domain because Roth teach that the need for recombinant viral vectors to genetically reprogram T cells has slowed down research and clinical because of lengthy and expensive manufacturing and testing (abstract) and Kriegsmann teach CAR T cells are promising anti-cancer therapies.
One would have had a reasonable expectation of success because both methods are drawn to genetic modification of T cells and T cell receptors for the purposes of targeting tumor antigen expressing cells.
The teachings of Roth are discussed supra. While Roth teach culture in xeno-free medium (XVivo15), Roth do not teach culturing cells in the absence of serum.
Jenkins teach serum content is a significant hurdle in T cell cultivation, and that serum-free T cell cultivation will reduce potential harmful elements and simplify T cell purification steps (p29 col1 ¶2). Jenkins further teach antigen-specific T cell cultivation has been undertaken with xeno-free serum replacement (SR), resulting in comparable T cell growth kinetics with FBS and reduced risk of adventitious infectious pathogens (p29 col1 ¶2).
It would have been obvious to one of ordinary skill in the art to adapt the methods of Roth drawn to non-viral insertion of a transgene by culturing the cells in xeno-free conditions by using SR in place of serum as taught by Jenkins. Jenkins teach that cultivation in serum-free conditions reduces potential harmful elements and simplifies T cell purifications steps.
Accordingly, one of ordinary skill in the art would have been motivated to modify the method as taught by Roth with the teachings of Jenkins for the purposes of a more efficient and protocol with reduced harmful elements.
One would have had a reasonable expectation of success because both inventions are directed to T cell cultivation under defined conditions.
Regarding claim 10: The teachings of Roth and Kriegsmann are discussed supra. Kriegsmann also teach the intracellular domain comprises a CD3 zeta intracellular domain (Fig 1c). Kriegsmann is silent on the nature of the transmembrane domain.
Sjoukje teach CAR intracellular domains comprise an intracellular CD3 zeta domain (Figure 1c) and that the first fusion receptors were coupled to the CD3 intracellular domain (Figure 1). Sjoukje also teach that the transmembrane domain of the CAR CD3 zeta facilitates enhanced anti-tumor function (p25 box1, Figure 1d).
It would have been prima facie obvious to one of ordinary skill in the art to adapt the methods of Roth and Kriegsmann with the teachings of Sjoukje, to include a CD3 zeta intracellular and transmembrane domain in the receptor.
One having ordinary skill in the art would have been motivated to modify the method of Roth and Kriegsmann to include a CD3 zeta intracellular and transmembrane domain because Sjoukje teach the CD3 zeta intracellular domain is one of the first intracellular domains used, and as such one of ordinary skill in the art would understand that the function of the intracellular domain is well studied and understood, and the CD3 zeta transmembrane domain facilitates enhanced antitumor function and cytokine production (p503 Box 1).
One would have had a reasonable expectation of success because the methods are drawn to genetically engineering T cells to express chimeric T cell receptors.
The rejection over 35 USC § 103 is maintained.
Regarding claims 12-13:
Applicant’s arguments filed on 8/28/2025 have been fully considered and are unpersuasive.
Applicant submits there is a lack of motivation for combination of relied upon references.
One would have been motivated to modify the method of Roth with the teachings of Lu and Lanoix to enhance the levels of correct transgene expression. One of ordinary skill in the art would understand that efficient termination of transcription reduces improperly transcribed transgenes due to read-through errors and that improved transgene expression would result in a more efficient and therefore less expensive method.
One would have had a reasonable expectation of success because the teachings are all directed to methods of transgenes expression in human cells.
The rejection over 35 USC § 103 is maintained.
Regarding claim 17:
Applicant’s arguments filed on 8/28/2025 have been fully considered and are unpersuasive.
Applicant does not provide arguments to support the assertation that claim 17 is not obvious over the cited prior art, and thus there are no arguments to answer.
The rejection over 35 USC § 103 is maintained.
Regarding claim 18:
Applicant’s arguments filed on 8/28/2025 have been fully considered and are unpersuasive.
Applicant submits claim 18 is not obvious over Roth, which does not specify the shape of the culture wells.
The teachings of Roth are discussed above. Roth do not teach culturing in round bottom culture wells or culture in a low culture volume (20% standard) of cell culture medium.
As evidenced by ThermoFisher, round bottom wells provide optimal sample recovery (p1, downloaded from the internet 01/04/2026).
Phelan teach cell culture in low volumes exhibit similar growth as traditional protocols with improved consistency (p2 col2 ¶2). Phelan further teach that low-volume multi-well setups provide a wide-range of options possible through customization.
It would have been obvious for one of ordinary skill in the art to modify the method of Roth with the teaching of ThermoFisher and Phelan, to culture cells in round bottom plates using a low culture volume.
One would have been motivated to modify the method of Roth to use round bottom plates with a low culture volume to improve optimize sample recovery and to improve consistency of cell cultures. Furthermore, one of ordinary skill in the art would recognize that culturing cells in a low culture volume would reduce the amount of reagents required for cell culture and thus decrease cost of the cell culture.
One would have had a reasonable expectation of success because one of ordinary skill in the art would know round bottom plates are commonly used for sample processing and culture of cells in suspension and that cell culture volumes would have been a matter of routine optimization based on optimization of the cell culture protocol and cell culture protocols in the art.
Furthermore, Applicant recites unexpected results attributed to culturing in round bottom wells at a 20% culture volume. While the instant specification does recite “It was unexpectedly found that by using round bottom culture wells rather than flat, for example, improved recovery was observed”, the claimed unexpected result is drawn only to round bottom culture wells, and improved sample recovery is a property of round bottom wells that is known in the art. Furthermore, the unexpected result recited in the instant specification is an assertation of an unsupported result which is not commensurate in scope with the claims.
The rejection over 35 USC § 103 is maintained.
Regarding claim 19:
Applicant’s arguments filed on 8/28/2025 have been fully considered and are unpersuasive.
Applicant submits that the teaching of Sadelain regarding autologous and allogenic T cells does not cure the deficiencies of Roth, however Applicant does not provide specific arguments, and thus there are no arguments to answer.
The rejection over 35 USC § 103 is maintained.
Conclusion
No claims are allowed.
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/ANDREA LYNNE MORRIS SPENCER/Examiner, Art Unit 1631
/LAURA SCHUBERG/Primary Examiner, Art Unit 1631