DETAILED ACTION
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Applicant’s response to restriction requirement filed on April 27, 2026 have been received and entered. Claims 1-2, 10-11, 13, 18, 24, 27 have been amended, while claims 3-5, 7-9, 1216, 19-23, 26, 29, 33 and 27 have been canceled. Claims 1-2, 6, 10-11, 13-15, 17-18, 24- 25, 27-28, 30-32, 34-35 and 36 are pending in the instant application.
Election/Restrictions
Applicant’s election without traverse of claims 1, 2 and 6 (group I) in the reply filed on April 27, 2026 is acknowledged.
Claims 10-11, 13-15, 17-18, 24- 25, 27-28, 30-32, 34-35 and 36 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim.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 April 26, 2026.
Priority
This application is a 371 of PCT/US2022/027622 filed on 05/04/2022, which claims priority from US provisional application no 63/184,440 filed on 05/05/2021.
Information Disclosure Statement
The information disclosure statements (IDS) submitted on 05/28/2024, 11/05/2025 and 04/27/2026 are in compliance with the provisions of 37 CFR 1.97. Accordingly, the information disclosure statement has been considered by the examiner.
Claims 1, 2 and 6 are under consideration.
Claim Rejections - 35 USC § 112
The following is a quotation of the first paragraph of 35 U.S.C. 112(a):
(a) IN GENERAL.—The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor or joint inventor of carrying out the invention.
The following is a quotation of the first paragraph of pre-AIA 35 U.S.C. 112:
The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor of carrying out his invention.
Claims 1, 2 and 6 are rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, because the specification, while being enabling for:
A transgenic mouse whose genome comprises in its germline an endogenous mouse immunoglobulin variable gene locus is deleted and replaced with whole or in part equine immunoglobulin locus comprising equine immunoglobulin variable gene coding sequences, wherein the part of equine immunoglobulin locus of the transgenic mouse is functional and expresses immunoglobulin chains comprising equine variable domains and mouse constant domains,
does not reasonably provide enablement for any other rodent or wherein the non-coding regulatory sequence comprises promoters preceding individual V gene segments, splice sites, and recombination signal sequences for V(D)J recombination of the endogenous rodent immunoglobulin variable gene locus. The specification does not enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the invention commensurate in scope with these claims.
In determining whether Applicant’s claims are enabled, it must be found that one of skill in the art at the time of invention by applicant would not have had to perform “undue experimentation” to make and/or use the invention claimed. Such a determination is not a simple factual consideration, but is a conclusion reached by weighing at least eight factors as set forth in In re Wands, 858 F.2d at 737, 8 USPQ 1400, 2d at 1404. Such factors are: (1) The breadth of the claims; (2) The nature of the invention; (3) The state of the art; (4) The level of one of ordinary skill in the art; (5) The level of predictability in the art; (6) The amount of direction and guidance provided by Applicant; (7) The existence of working examples; and (8) The quantity of experimentation needed to make and/or use the invention.
The office has analyzed the specification in direct accordance to the factors outlines in In re Wands. MPEP 2164.04 states: “[W]hile the analysis and conclusion of a lack of enablement are based on factors discussed in MPEP 2164.01(a) and the evidence as whole, it is not necessary to discuss each factor in written enablement rejection.” These factors will be analyzed, in turn, to demonstrate that one of ordinary skill in the art would have had to perform “undue experimentation” to make and/or use the invention and therefore, applicant’s claims are not enabled.
Nature of the Invention:
Claims are directed to a transgenic rodent with a genome in which an endogenous rodent immunoglobulin variable gene locus is deleted and replaced with a partly equine immunoglobulin locus comprising equine immunoglobulin variable gene coding sequences and non-coding regulatory sequences based on the endogenous rodent immunoglobulin variable gene locus, wherein the partly equine immunoglobulin locus of the transgenic rodent is functional and expresses immunoglobulin chains comprising equine variable domains and rodent constant domains. Dependent claims limit the partly equine immunoglobulin locus comprises;{i) equine VH, DH and JH coding sequences; (ii) equine kappa VL and JL coding sequences; (iii) equine lambda VL and JL coding sequences; (iv) an ADAM6 gene: (v) Pax-5-Activated Intergenic Repeat (PAIR) elements; (vi) CTCF binding sites from a heavy chain intergenic control region 1; or (vii) any combination thereof. Claim 6 limits the non-coding regulatory sequences comprise promoters preceding individual V gene segments, splice sites, and recombination signal sequences for V(D)J recombination.
Breadth of the claims:
The claims as written broadly read on any rodent with a genome in which an endogenous rodent immunoglobulin variable gene locus is deleted and replaced with a partly equine immunoglobulin locus comprising equine immunoglobulin variable gene coding sequences and non-coding regulatory sequences based on the endogenous rodent immunoglobulin variable gene locus, wherein the partly equine immunoglobulin locus of the transgenic rodent is functional and expresses immunoglobulin chains comprising equine variable domains and rodent constant domains. Dependent claims limit the non-coding regulatory sequences comprise promoters preceding individual V gene segments, splice sites, and recombination signal sequences for V(D)J recombination.
Guidance of the Specification and The Existence of Working Examples:
The specification prophetically contemplates replicating mouse VH, D and JH coding sequences in silico with equine VH, D and JH coding sequences, for example, using commercially available software. The specification contemplates replacing the VH, D and JH coding sequences while leaving the intervening mouse non-coding sequences intact. Similarly, a partly equine immunoglobulin light chain variable region locus can be generated in silico. Again, the VL and JL coding sequences can be replaced while leaving the intervening mouse non-coding sequences intact. The specification discloses methods are known for synthesizing a DNA sequence that includes the partly equine immunoglobulin locus based on the in-silico sequences (see para. 28 of the specification). The specification prophetically teaches introduction of a heterologous partly equine immunoglobulin locus into the genomic locus of mouse ES cell is illustrated in FIGS. 2-4 (see example 1). The specification discloses mouse embryonic stem (ES) cells (derived from C57B1/6NTac mice) are transfected by electroporation with the 5' vector (201) according to known procedures. Colonies of drug-resistant ES cells are physically extracted and these picked colonies are disaggregated, re-plated in micro-well plates, and cultured for several days. The primary screening procedure for the introduction of 5' vector can be carried out by Southern blotting, or by PCR with confirmations from secondary screening methods such as Southern blotting. DNA from the ES cell clones is screened by PCR using a widely practiced gene- targeting assay design. The assays detect DNA that would only be present in clones of ES cells that undergo homologous recombination between the 5' targeting vector and the endogenous mouse Igh locus. Clones of ES cells that have been mutated by both the 3' and the 5' vectors, i.e., doubly targeted cells carrying both engineered mutations, are isolated following vector targeting and analysis. The clones must have undergone gene targeting on the same chromosome, as opposed to homologous chromosomes (i.e., the engineered mutations created by the targeting vectors must be in cis on the same DNA strand rather than in trans on separate homologous DNA strands). ES cell clones carrying the sequence deletion in one of the two homologous copies of their immunoglobulin heavy chain locus are re-transfected with a Cre recombinase expression vector and a vector that includes a partly equine immunoglobulin heavy chain locus containing equine VH, DH and JH gene segment coding sequences embedded in mouse non-coding sequences. FIG. 4 illustrates introduction of the heterologous partly equine immunoglobulin heavy chain locus into a mouse genome in which the part of the endogenous immunoglobulin heavy chain locus that encodes the heavy chain variable region domains has been deleted, including the intervening sequences between the endogenous VH and JH gene loci. ES cell clones carrying the partly equine immunoglobulin heavy chain variable region (443) in the mouse heavy chain locus are microinjected into mouse blastocysts from strain DBA/2 to create ES cell-derived chimeric mice according to standard procedures. Example 2 teaches introduction of a heterologous partly equine immunoglobulin locus into the immunoglobulin Kappa chain gene locus of a mouse genome (see fig. 5). Example 3: teaches introduction of a heterologous partly equine immunoglobulin locus into the immunoglobulin lambda chain gene locus of a mouse genome (see fig. 6A,6B).
State of the Art and Predictability of the Art and the Amount of Experimentation Necessary:
The claims broadly embrace making any transgenic rodent species, where rodent comprises an extremely large genus of species including any of mouse, rat, gerbil or Guinea pig and includes such locus of the same species. The claim invention encompasses any species of rodent to be made via modification of ES cells (see para. 125-126 of the application, example 1). In this regard, the art teaches that creating genetically modified rodent from any species of animal using ES cells is unpredictable except for mice. It is further noted that the specification prophetically contemplates deletion of the endogenous heavy chain variable gene segment and its replacement with the equine variable gene segment gene cassette by recombinase-mediated cassette exchange (RMCE). However, the specification fails to provide any guidance as to other species of embryonic stem cells capable of contributing to the germline of an animal other than mouse. Before the time of filing, the skilled artisan did not consider the generation of knock-out or knock-in non-human animals other than the mouse as routine or predictable. As noted above, the specification only provides specific guidance for using mouse embryonic stem cells to generate a transgenic mouse. The specification provides no additional guidance for making any species of rodent. At the time of filing, the technology to produce animals with a gene-targeted knockout was considered limited to transgenic mice because the technology uses homologous recombination in embryonic stem (ES) cells. As the art teaches below, there is significant unpredictability in isolating, characterizing and using ES cells isolated from non-mouse species, and that pluripotency has not been validated or verified in any species of animal other than the mouse. Regarding ES cells, the art teaches that while mouse ES cells have been established, no validated Munoz et al. teaches that as of 2009, despite extensive investigation, ES cells lines from animals other than mouse and human had yet to be established due to difficulties in the isolation and maintenance of ESC lines from other species (Munoz et al. (2009) Stem Cell Rev. and Rep., Vol. 5, 6-9, pages 6 and 9). Ezashi et al (Annu. Rev. Anim. Biosci. 2016. 4:223–53) reviewed the state of the art and states “authentic ESC homologous to those described for rodent have not been established conclusively in any of these species (see page 227, para.1). Ezashi et al continue to teach that “the persistent failure in generating ESC from these same species may stem from a shared problem, namely, instability of the gene networks necessary to maintain pluripotency under the culture conditions employed” (see page 231, para. 1). Hong et al. (Stem Cells and Development, 2012, Vol. 21(9), pgs. 1571-1586) teaches that there is unpredictability even among ES cells from different strains of rat (see abstract). Specifically, Hong teaches that while ES cells from either F344 or dark agouti rats could contribute to generate a chimera, only ES cells from dark agouti (DA) were determined to be germline competent (see Abstract and pg. 1584 col. 2 para. 2). Further Hong teaches that "In other laboratories, F344 ESCs have not produced chimeric rats after an injection into DA or SD blastocysts [references therein]. Tong et al. (2010, Nature, Vol. 467(7312), pgs. 211-213) teaches: "Failure of ES cells to contribute to the germline is often caused by chromosomal abnormalities in ES cells. This is also likely to be true for rat ES cells. In the instant case, Tong et al or any other prior art do not provide information about the PCR primer sequences by which a gerbil or Guinea pig or any other rodent genomic library can be screened for BACs containing the relevant sequences. One of skill in the art would require more information and undue experimentation to identify suitable BAC clones, to make and use the invention in ES cells derived from different species of rodent, without reasonable expectation of success. The disclosure of the specification is prophetic and provides no actual rodent whose genome comprises an endogenous rodent immunoglobulin variable gene locus is deleted and replaced with a partly equine immunoglobulin locus comprising equine immunoglobulin variable gene coding sequences and non-coding regulatory sequences based on the endogenous rodent immunoglobulin variable gene locus. It is unclear performing ES cell clones derived from other species of rodent through multiple rounds of manipulation would results in maintaining the germ line potential of the rodent ES cell line other than mouse ES cells (see Liu et al Developmental Dynamics, 1997, 209, 85-91). No such data is presented to show that a transgenic rodent genome comprises the deletion of endogenous variable gene segment that is replaced with a partly equine immunoglobulin locus comprising equine immunoglobulin variable gene coding sequences and non-coding regulatory sequences based on the endogenous rodent immunoglobulin variable gene locus as broadly claimed. The evidence in prior art clearly establishes the fact that use of rodent ES cells other than making transgenic mouse was still evolving and unpredictable before the effective filing date of the claimed invention. An artisan would have to perform undue experimentation to make and use the invention, without reasonable expectation of success.
The claims embrace rodent immunoglobulin variable gene locus that is deleted and replaced with a partly equine immunoglobulin locus comprising equine immunoglobulin variable gene coding sequences and non-coding regulatory sequences comprising promoters preceding individual V gene segments, splice sites, and recombination signal sequences for V(D)J recombination embedded at the endogenous rodent immunoglobulin variable gene locus. Thus, the claims as written require mining of the gene segments from equine and any species of rodent genome. The sequences are then required to be analyzed in silico such that critical noncoding regulatory components are rodent (gerbil, rat, mouse or guinea pig), while equally crucial coding sequence of equine variable gene segment. The resulting chimeric gene segment sequences are optimized for expression in the mouse because of the presence of the mouse noncoding sequences comprising promoters preceding individual V gene segments, splice sites, and recombination signal sequences for V(D)J recombination, yet they express equine IgH/IgL.
Montalbano et al (J Immunol 2003 Nov 15;171(10):5296-304) teaches VDJ recombination frequencies could be affected by changes in the spacer sequence even without change in coding sequence identity. This is especially important since claim requires retaining rodent intervening non-coding DNA (see abstract). McMurry et al (Mol Cell Biol. 1997 Aug;17(8):4553-61) reported that enhancer context determines whether recombination proceeds efficiently and correct order (see abstract). McMurry concludes that enhancers regulate V(D)J recombination by providing local accessibility to the recombinase. cis-acting elements other than Edelta must maintain Ddelta3 in an accessible state in the absence of Edelta. The analysis of DSB formation in a single-copy minilocus integrant indicates that efficient DSB formation at the accessible RSS 3' of Ddelta3 requires an accessible partner RSS, arguing that RSS synapsis is required for DSB formation in chromosomal substrates in vivo. In view of foregoing , it is apparent that species mismatched coding and regulatory sequence may also not work predictably even if RSS remain functional. In the instant case, neither the specification nor the prior art teaches how to combine equine and mouse sequences accurately in silico such that critical noncoding regulatory components are rodent (i.e mouse), while equally crucial coding components are equine. The claimed transgenic rodent further require optimization of non-coding sequence for expression in the rodent because of the presence of the rodent noncoding intervening sequences, which are embedded with equine variable gene segment. There is no teaching of any optimization that would enable or produce the claimed transgenic mouse such that a functional equine immunoglobulin is expressed and produced. The skilled artisan can simply not practice the claimed method to produce the claimed transgenic rodent without specific guidance with respect to in silico design of the chimeric variable gene segments and the further optimization to make them functional such that a reverse chimeric equine antibody is produced. One of skill in the art would have to perform undue experimentation to make and use the invention, without reasonable expectation of success.
In conclusion, in view of breadth of the claims and absence of a strong showing by Applicant, in the way of specific guidance and direction, and/or working examples demonstrating the same, such invention as claimed by Applicant is not enabled for the claimed inventions. An artisan of skill would have required undue experimentation to practice the invention, without reasonable expectation of success as supported by the observations in the art record.
Claim Rejections - 35 USC § 102
The following is a quotation of the appropriate paragraphs of 35 U.S.C. 102 that form the basis for the rejections under this section made in this Office action:
A person shall be entitled to a patent unless –
(a)(1) the claimed invention was patented, described in a printed publication, or in public use, on sale, or otherwise available to the public before the effective filing date of the claimed invention.
Claims 1 and 2 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Martin et al (WO2018/189520, dated 10/18/2018).
Claims are directed to a transgenic rodent with a genome in which an endogenous rodent immunoglobulin variable gene locus is deleted and replaced with a partly equine immunoglobulin locus comprising equine immunoglobulin variable gene coding sequences and non-coding regulatory sequences based on the endogenous rodent immunoglobulin variable gene locus, wherein the partly equine immunoglobulin locus of the transgenic rodent is functional and expresses immunoglobulin chains comprising equine variable domains and rodent constant domains.
With respect to claims 1-2, Bradley teaches a transgenic rodent whose genome comprising; i) one or more companion animal IGH V region genes, one or more companion animal D region genes and one or more companion animal J region genes; and (ii) one or more companion animal IGL kappa V region genes and one or more companion animal IGL kappa J region genes; and/or one or more companion animal IGL lambda V region genes and one or more companion animal IGL lambda J region genes, wherein the companion animal gene(s) are located in the genome upstream of the rodent constant region, suitably upstream of the heavy chain constant region for inserted companion animal heavy chain variable region genes and suitably upstream of a light chain constant region for inserted companion animal light chain variable region genes, such that the rodent is able to produce a chimeric antibody chain resulting from expression of the inserted variable region gene and the host constant region, wherein the rodent is a mouse and the companion animal is a horse (equine) (See claims 1-2, 8, 13). It is further disclosed that rodent is a mouse and the genome comprises a deletion of one or some or all of the mouse IGH V region genes, a deletion of one or some or all of the mouse IGL kappa V region genes. Martin teaches rodent genome is modified to reduce or prevent expression of fully rodent antibodies that have both variable and constant regions from the rodent. This may be by a deletion of, or an insertion into, the endogenous rodent VDJ or VJ region of the genome (see page 12, para. 4-7). It is further disclosed that at least one endogenous regulatory sequence (rodent enhancer or other control sequence, such as a switch region), is maintained in functional arrangement with the rodent constant region (see page 13, para. 4-6).
Accordingly, Martin anticipates claims 1-2.
Conclusion
No claims allowed.
The prior art made of record and not relied upon is considered pertinent to applicant's disclosure. Wabl (US20130219535, dated 08/22/2013) teaches a transgenic mouse whose genome comprises an entire endogenous mouse immunoglobulin locus variable region which has been deleted and replaced with an engineered immunoglobulin locus variable region comprising at least one of each of a chimeric V, D and J immunoglobulin variable region gene segment at the immunoglobulin heavy chain locus, and/or at least one of each of a chimeric V and J variable gene segment at the immunoglobulin light chain loci, wherein each chimeric gene segment comprises human V, D, or J immunoglobulin variable region coding sequences embedded in mouse immunoglobulin variable region non-coding gene segment sequences, wherein the engineered immunoglobulin locus of the transgenic mouse is functional and expresses immunoglobulin chains comprised of human variable domains and mouse constant domains.
Bradley (WO-2020074874, IDS) teaches the transgenic mouse with same genotype and phenotype as disclosed in Martin of record.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to ANOOP K. SINGH whose telephone number is (571)272-3306. The examiner can normally be reached Monday-Friday, 8AM-5PM.
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/ANOOP K SINGH/ Primary Examiner, Art Unit 1632
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