DETAILED ACTION
Status of the Claims
This application has been reassigned, and Christopher M. Babic is now the Examiner of record. Claims 203–213 remain pending in the application. Claims 203, 204, and 206–213 are under examination, and claim 205 remains withdrawn. Applicant’s response filed on August 4, 2025, has been entered and fully considered. The present Office action is non-final.
Abstract – Objection Withdrawn
The prior objection to the abstract is withdrawn in view of Applicant’s amendment.
Claim Rejections - 35 USC § 102 - Withdrawn
The prior rejection of claims 203, 204, and 206–213 under 35 U.S.C. §102(a)(1) as anticipated by Duchateau is withdrawn. Applicant persuasively argued that Duchateau does not expressly disclose contacting a cell with a preassembled non-viral Cas9:gRNA ribonucleoprotein complex as now recited in claim 203. Duchateau teaches delivery of CRISPR components generally, including electroporation of Cas9 mRNA and sgRNA plasmids, but does not expressly teach a naked preassembled (non-viral) RNP complex.
Claim Rejections - 35 USC § 103 - New Grounds
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.
Claim(s) 203, 204, and 206-213 are rejected under 35 U.S.C. 103 as being unpatentable over Duchateau (WO 2014/191128 A1) in view of NCBI Accession No. NG_001332 (revision history: first available June 10, 2002), Hsu et al. (Nat Biotechnol. 2013 Sep;31(9):827-32. Epub 2013 Jul 21), and Kim et al. (Genome Res. 2014 Jun;24(6):1012-9. Epub 2014 Apr 2).
Duchateau is drawn to an invention related to methods of developing genetically engineered, preferably non-alloreactive T-cells for immunotherapy and the use of CRISPR/Cas9 system, to specifically target a selection of key genes in T-cells. (Abstract).
Regarding Claims 203 and 209, Duchateau teaches methods of modifying T cells comprising the cells obtained from donor for making them suitable for immunotherapy purpose by using RNA-guided (or gRNA) endonuclease system or CRISPR/Cas9 gene editing system (pg. 4). Duchateau teaches that in CRISPR-based modification of T cells with respect to TCR gene, T cells are transfected with Cas9 and sgRNA where sgRNA specifically target TRAC gene (pg. 5). Duchateau teaches gRNA molecules that specifically hybridize a target sequence of TCR alpha (i.e. a targeting domain of TCR gene) (pg. 23, Table 2). Duchateau teaches that any means known in the art to allow delivery inside cells and molecules can be used (pg. 17).
Duchateau teaches that inactivation of TCR alpha and/or TCR beta is beneficial for generating engineered allogeneic T cells suitable for immunotherapy (see pg. 22-24, non-alloreactive T cells). Duchateau explains that “TCR is rendered not functional in the cells by inactivating TCR alpha gene and/or TCR beta gene(s)” and that such inactivation aims to prevent or reduce graft-versus-host disease (GvHD) by eliminating recognition of host alloantigens through the endogenous T-cell receptor (see pg. 22-24, non-alloreactive T cells). Duchateau therefore teaches that disruption of TCR alpha and/or beta provides a functional advantage in producing non-alloreactive therapeutic T cells for adoptive immunotherapy.
Duchateau does not appear to teach the full TRAC sequence, non-viral delivery of the CRISPR RNP complex, or the identification of the specific sequence targeting domains as claimed (e.g. SEQ ID NO: 49277).
Full TRAC Sequence
At the time of the invention, the TRAC locus and its nucleotide sequence were known in the art (see NCBI Accession No. NG_001332; revisionist history first available June 10, 2002), and there is no evidence presently of record indicating that Applicant targeted previously unknown or unelucidated TRAC sequences. Rather, the present record indicates that Applicant selected guide-targeting sequences from an already known genomic region. Accordingly, absent persuasive evidence to the contrary, the claimed targeting sequences are understood to have been chosen from a locus whose underlying sequence information was already available to persons of ordinary skill in the art.
Specific Sequence Targeting Domains
Hsu teaches experimentally validated guide RNA design principles for S. pyogenes Cas9 in mammalian cells, including that efficient targeting generally involves a 20-nucleotide guide sequence adjacent to an appropriate PAM, preferably NGG, and that cleavage specificity depends on the number, position, and distribution of mismatches between the guide RNA and the genomic target (see pg. 2-4). Hsu further teaches that off-target effects can be reduced by selecting guide sequences with favorable mismatch characteristics and by limiting the amount of Cas9/guide complex delivered to the cell (see pg. 6). Thus, Hsu is relevant for showing that guide selection and specificity optimization for Cas9-targeted genomic editing were governed by known and predictable design considerations.
Once Duchateau taught targeting TCR loci in T cells and provided targetable TCR sequences/windows, one of ordinary skill would have looked to known CRISPR guide-design principles to choose among available candidate target sites. Hsu supplies those principles. Hsu teaches that guide selection depends on: a compatible PAM, sequence similarity to potential off-target sites, mismatch position and density, and dosage/specificity considerations.
Thus, Hsu would have directed a POSA to evaluate candidate TRAC/TRBC guides according to known specificity criteria and to choose those expected to be effective and selective. This is the ordinary application of known guide-design rules to a disclosed target locus. Selection of particular 20-mer guides from a finite, target-locus-defined set of candidates is therefore an obvious optimization of a result-effective variable (e.g. the claimed sequence target sites). That conclusion is especially supported where the claim also extends to guides that “differ[] by no more than 3 nucleotides,” which reflects routine guide-variant exploration rather than a sharply delimited, singular discovery.
Non-viral CRISPR RNP Complex Delivery
Kim teach delivery of purified Cas9 protein preassembled with guide RNA into human cells (see pg. 1013, col. 1-2). Kim et al. reports that their methods enable efficiency and precise editing while avoiding unwanted integration of plasmids and reducing off-target effect. (see pg. 1017).
A person of ordinary skill in the art would have been motivated to substitute Kim’s preassembled Cas9:gRNA RNP format into Duchateau’s non-viral T-cell CRISPR method because Duchateau expressly identifies prolonged Cas9 expression as undesirable and seeks transient delivery approaches to reduce toxicity. Kim provides exactly such a transient format: direct delivery of active Cas9 protein already complexed with guide RNA, without requiring transcription or translation of CRISPR components in the cell.
Replacing Duchateau’s transient Cas9 mRNA + sgRNA plasmid delivery format with Kim’s transient preassembled Cas9:gRNA RNP format would have been an obvious substitution of one known transient CRISPR delivery format for another to obtain the predictable benefit of reducing prolonged Cas9 expression. A POSA would have had a reasonable expectation of success because Kim demonstrates that non-viral preassembled Cas9:gRNA RNPs work in human cells.
Conclusion
Taken as a whole, it would have been prima facie obvious to a person of ordinary skill in the art at the time of the invention to modify Duchateau’s CRISPR-based T-cell engineering methods by using the non-viral preassembled Cas9:gRNA ribonucleoprotein delivery approach taught by Kim and by selecting TRAC/TRBC-targeting guide sequences using the known guide-design and specificity principles taught by Hsu, with a reasonable expectation of success. Applicant has not presented substantiated evidence that the claimed target regions or guide sequences yield unexpected results relative to the closest prior art.
Other Teachings
Regarding claim 204, Duchateau teaches that in type II CRISPR/Cas system the Cas9 is derived from S. pyogenes for use in mammalian cells (pg. 3, lines 22-23). Duchateau teaches that the Cas9 nuclease cuts the DNA only if a specific sequence known as PAM is present immediately downstream of the protospacer sequence, whose canonical sequence in S. pyogenes is 5' - NGG -3', where N refers to any nucleotide (pg. 3, lines 16-19).
Regarding claim 206, Duchateau teaches that the genetic modification of a target sequence relies on the expression in provided cells to engineer when treated with gRNA-endonuclease (pg. 22, lines 31-33). Duchateau teaches a method more particularly of comprising the steps of modifying T-cells by inactivating at least one component of the T-cell receptor (TCR) by using an RNA guided endonuclease associated with a specific gRNA (pg. 22, lines 24-26).
Regarding claim 207, Duchateau teaches that the inactivation of TCR alpha (i.e. TRAC) or TCR beta (i.e. TRBC) by using a RNA guided endonuclease associated with a specific gRNA can result in the elimination of the TCR from the surface of T cells (pg. 22, lines 17-18 and lines 24-26). Duchateau teaches that electroporating T cells with Cas9 and gRNA targeted for TCR alpha (or TRAC) results in inactivated or engineered T cells with a deletion of the TCR alpha gene (i.e. introducing mutations) (pg 50, lines 4-20).
Regarding claim 208, Duchateau teaches that the preferred target sequences are optimally located within the 3 exons encoding TCR gene i.e. modification is within the first 500 bp of the coding sequence from the start codon (see Table 2, pg. 23), allowing a significant reduction of toxicity, while retaining cleavage efficiency (pg. 23, lines 10-12).
Regarding claims 210-212, Duchateau teaches that modifying T cells are derived from the group consisting of CD4+ T- lymphocytes and CD8+ T-lymphocytes (pg. 30, lines 6-8). Duchateau teaches that these T cells are obtained from a non-limiting sources including PBMCs, bone marrow, tissues from lymph node, thymus, or spleen, i.e. indicating various sources of CD8+ naïve T cells (pg. 30, lines 10-12).
Regarding claim 213, Duchateau teaches that engineering T cells can be derived from stem cells such as adult stem cells, embryonic stem cells, non-human stem cells, cord blood stem cells, progenitor cells, bone marrow stem cells, or induced pluripotent stem cells (pg. 30, lines 1-4).
Response to Arguments
Applicant argues that Duchateau does not teach a non-viral RNP complex and that the presently claimed guide sequences were selected according to particular criteria and shown to have activity in the application.
These arguments have been considered. As stated above, the Examiner agrees that Duchateau alone does not expressly disclose a preassembled naked Cas9:gRNA RNP complex; therefore, the anticipation rejection has been withdrawn.
Applicant’s arguments regarding the alleged special selection and activity of the claimed guide sequences have also been considered but are not persuasive on the present record. Applicant has not provided substantiated evidence that the claimed target regions or claimed guide sequences yield unexpected results relative to the closest prior art target regions or guide sequences. Evidence that certain claimed guides are active does not, without more, establish nonobviousness where the prior art already teaches targeting the same loci and selecting guides using known design principles.
If Applicant contends that the claimed target regions or guide sequences provide unexpected advantages, Applicant is invited to submit objective comparative evidence demonstrating unexpectedly superior results, such as improved editing efficiency, specificity, or reduced off-target activity, relative to the closest prior art comparators, together with evidence of nexus to the full scope of the claims.
Double Patenting – New Grounds
The nonstatutory double patenting rejection is based on a judicially created doctrine grounded in public policy (a policy reflected in the statute) so as to prevent the unjustified or improper timewise extension of the “right to exclude” granted by a patent and to prevent possible harassment by multiple assignees. A nonstatutory double patenting rejection is appropriate where the conflicting claims are not identical, but at least one examined application claim is not patentably distinct from the reference claim(s) because the examined application claim is either anticipated by, or would have been obvious over, the reference claim(s). See, e.g., In re Berg, 140 F.3d 1428, 46 USPQ2d 1226 (Fed. Cir. 1998); In re Goodman, 11 F.3d 1046, 29 USPQ2d 2010 (Fed. Cir. 1993); In re Longi, 759 F.2d 887, 225 USPQ 645 (Fed. Cir. 1985); In re Van Ornum, 686 F.2d 937, 214 USPQ 761 (CCPA 1982); In re Vogel, 422 F.2d 438, 164 USPQ 619 (CCPA 1970); In re Thorington, 418 F.2d 528, 163 USPQ 644 (CCPA 1969).
A timely filed terminal disclaimer in compliance with 37 CFR 1.321(c) or 1.321(d) may be used to overcome an actual or provisional rejection based on nonstatutory double patenting provided the reference application or patent either is shown to be commonly owned with the examined application, or claims an invention made as a result of activities undertaken within the scope of a joint research agreement. See MPEP § 717.02 for applications subject to examination under the first inventor to file provisions of the AIA as explained in MPEP § 2159. See MPEP § 2146 et seq. for applications not subject to examination under the first inventor to file provisions of the AIA . A terminal disclaimer must be signed in compliance with 37 CFR 1.321(b).
The filing of a terminal disclaimer by itself is not a complete reply to a nonstatutory double patenting (NSDP) rejection. A complete reply requires that the terminal disclaimer be accompanied by a reply requesting reconsideration of the prior Office action. Even where the NSDP rejection is provisional the reply must be complete. See MPEP § 804, subsection I.B.1. For a reply to a non-final Office action, see 37 CFR 1.111(a). For a reply to final Office action, see 37 CFR 1.113(c). A request for reconsideration while not provided for in 37 CFR 1.113(c) may be filed after final for consideration. See MPEP §§ 706.07(e) and 714.13.
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Claims 203, 204, and 206-213 are rejected on the ground of nonstatutory obviousness-type double patenting as being unpatentable over Claims 1-9 of U.S. Patent No. 12,325,854 in view of Duchateau (WO 2014/191128 A1), Hsu et al. (Nat Biotechnol. 2013 Sep;31(9):827-32. doi: 10.1038/nbt.2647. Epub 2013 Jul 21), and Kim et al. (Genome Res. 2014 Jun;24(6):1012-9. Epub 2014 Apr 2).
Claims 1-9 of U.S. Patent No. 12,325,854 recite populations of T cells or immune cells comprising a CRISPR nuclease-guide complex, including an RNP complex, wherein the guide RNA targets the TRAC gene. Although the claims recite Cpf1 rather than Cas9, and do not recite the presently claimed guide sequence limitations, these differences do not render the instant claims patentably distinct.
As set forth in the present obviousness rejection, Duchateau teaches CRISPR-mediated engineering of T cells for immunotherapy, including disruption of TCR alpha and/or TCR beta loci and targeting of TRAC in T cells; Kim teaches non-viral delivery of preassembled Cas9:gRNA RNP complexes into human cells; and Hsu teaches known and predictable guide-selection principles for SpCas9. It therefore would have been obvious to a person of ordinary skill in the art to use Cas9 as an alternative CRISPR nuclease in the TRAC-targeting RNP framework and to select TRAC-targeting guide sequences using routine design principles, with a reasonable expectation of success.
Claims 1-9 of U.S. Patent No. 12,325,854 recite CD8+ T cells, including CD8+ naive T cells, central memory T cells, and effector memory T cells, which substantially overlap the presently claimed T-cell subtype limitations. In view of the teachings of the patent claims together with Duchateau’s disclosure of CRISPR-based T-cell engineering and Kim and Hsu’s teachings regarding RNP delivery and guide optimization, the presently claimed subject matter would have been an obvious variation and is not patentably distinct.
Accordingly, Claims 203, 204, and 206-213 are rejected for nonstatutory obviousness-type double patenting over Claims 1-9 of U.S. Patent No. 12,325,854 in view of Duchateau, Kim, and Hsu.
Claims 203, 204, and 206-213 are provisionally rejected on the ground of nonstatutory obviousness-type double patenting as being unpatentable over claims 162, 393, 408, and 417 of copending Application No. 16/098,845 in view of Duchateau (WO 2014/191128 A1), Kim et al. (Genome Research, 2014), and Hsu et al. (Nat. Biotechnol., 2013). This rejection is provisional because the conflicting claims are in a copending application and have not yet issued.
Claims 162, 393, 408, and 417 of Application No. 16/098,845 recite methods of altering a T cell by contacting the T cell with Cas9/gRNA complexes, including complexes in which a gRNA targets the TRAC gene, and further recite Streptococcus pyogenes Cas9. Although those claims do not expressly recite the presently claimed non-viral RNP complex language, the NGG target-site limitation, or the presently recited guide-sequence framework, these differences do not render the instant claims patentably distinct.
As set forth in the present §103 rejection, Duchateau teaches CRISPR/Cas9-mediated engineering of T cells for immunotherapy, including disruption of TCR alpha and/or TCR beta loci, and further teaches that the canonical PAM recognized by S. pyogenes Cas9 is 5′-NGG-3′. Kim teaches non-viral delivery of preassembled Cas9:gRNA RNP complexes into human cells, and Hsu teaches known and predictable guide-selection principles for SpCas9, including targeting 20-nt sequences adjacent to NGG PAMs and optimizing specificity through routine guide design. Thus, it would have been obvious to a person of ordinary skill in the art to implement the TRAC-targeting Cas9 methods using a non-viral Cas9 RNP format and NGG-adjacent target sites as taught by Duchateau, Kim, and Hsu, with a reasonable expectation of success.
Accordingly, Claims 203, 204, and 206-213 are provisionally rejected for nonstatutory obviousness-type double patenting over claims 162, 393, 408, and 417 of copending Application No. 16/098,845 in view of Duchateau, Kim, and Hsu.
Conclusion
No claims are allowed.
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/CHRISTOPHER M BABIC/ Supervisory Patent Examiner, Art Unit 1633
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