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 .
Claims 7, 8 have been canceled. Claims 1-6, 9-18 are pending.
Election/Restrictions
Applicant's election with traverse of Group I, claims 1-6, 9-11 in the reply filed on 12-1-25 is acknowledged. The traversal is on the ground(s) that the amendment to claim 12 unifies the invention. Claim 12 as newly amended requires “the endogenous Cμ gene segment is inactive by a loss of function mutation, a deletion in part of the Cμ gene segment or by one or more mutations introducing one or more stop codons”. This encompasses an endogenous Cμ gene segment that is i) inactive by a loss of function mutation, ii) has a deletion in part of the Cμ gene segment, or iii) has one or more mutations introducing one or more stop codons”. Applicants’ argument is persuasive as it relates only to Group II. Claims 12-14 have been combined with group I. The requirement is otherwise deemed proper and is therefore made FINAL.
Claims 15-18 have been withdrawn from further consideration pursuant to 37 CFR 1.142(b), as being drawn to a nonelected inventions, there being no allowable generic or linking claim. Applicant timely traversed the restriction (election) requirement in the reply filed on 12-1-25.
Claims 1-6, 9-14 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.
Written Description
Claims 1-6, 9-14 are rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, as failing to comply with the written description requirement. The claim(s) contains subject matter which was not described in the specification in such a way as to reasonably convey to one skilled in the relevant art that the inventor or a joint inventor, or for applications subject to pre-AIA 35 U.S.C. 112, the inventor(s), at the time the application was filed, had possession of the claimed invention.
Claim 1 is drawn to a method of producing a mouse in which B cells express a diverse repertoire of heavy chain-only antibodies (HCAb), by incorporating within the endogenous immunoglobulin heavy chain constant region locus, a transgenic Cy gene segment upstream of the endogenous Cμ gene segment, wherein the Cy gene segment comprises a deletion of a nucleotide sequence encoding at least part of the CH1 domain, wherein the endogenous Cμ gene segment is inactive by a loss-of-function mutation, a deletion of part of the Cμ gene segment or by one or more mutations introducing one or more stop codons.
Claim 12 is drawn to a mouse obtainable by the method of claim 1, comprising within its endogenous immunoglobulin heavy chain constant region locus, a transgenic Cy gene segment upstream of the endogenous Cp gene segment, wherein the Cy gene segment comprises a deletion of a nucleotide sequence encoding at least part of the CH1 domain, wherein the endogenous Cu gene segment is inactive by aloss-of-function mutation, a deletion of part of the Cu gene segment or by one or more mutations introducing one or more stop codons.
The specification lacks written description for an Ig heavy chain constant region “locus” in claim 2 are unclear. First, “locus” is singular, and “loci” is plural (Wikipedia definition of “locus”, 2023; National Human Genome Res. Institute definition of “locus”, 2023). Second, locus (singular) is a position in space or an address on a chromosome. Third, the specification does not define an Ig heavy chain constant region “locus” (singular) as and an Ig heavy chain constant gene (which must have a plurality of nucleotides) or as a plurality of nucleotides encoding an Ig heavy chain constant domain. Fourth, the specification is limited to replacing a plurality of endogenous nucleotides (at a plurality of contiguous “loci” (plural)) with a plurality of exogenous nucleotides. The specification does not teach the exogenous nucleotides are present at a single locus, position, or address on the chromosome as claimed. Replacing pluralities of nucleotides is not replacing a nucleotide at a single locus as claimed. Fifth, the concept claimed does not accurately set forth the genetic modification because the addresses and positions of the nucleotides within the endogenous Ig heavy chain constant gene have changed because of differences in the length caused by the genetic modification. Accordingly, use of an Ig heavy chain constant region “locus” lacks written description.
The specification lacks written description for making a mouse having the features claimed by simply incorporating a transgenic Cγ gene segment upstream of a Cμ gene segment as broadly encompassed by claims 1 and 12. Claim 1 never clearly sets for incorporating the transgene into a mouse cell. Nor does it set forth any other active steps, i.e.
i) genetically modifying a mouse embryonic stem (ES) cell such that the genome of the ES cells comprises an endogenous immunoglobulin (Ig) heavy chain constant γ (Cγ) gene with a deletion of a nucleic acid sequence encoding at least part of the CH1 domain and an inactivated endogenous Ig heavy chain constant μ (Cμ) gene;
ii) introducing the ES cell into a recipient mouse embryo;
iii) implanting the mouse embryo into a recipient female mouse such that a genetically modified mouse whose genome comprises… …is obtained, wherein the genetically modified mouse is capable of expressing an endogenous Ig Cγ that is missing a CH1 domain and is incapable of expressing an endogenous Ig Cμ.
Claims 1 and 12 never result in a mouse having a genome comprising a genetic modification. Nor do they require the mouse is capable of expressing a diverse repertoire of heavy chain only antibodies.
The specification teaches “free monoclonal H chains can be secreted in a rare B cell proliferative disorder called heavy chain disease (HCD) (Franklin, et al., Am. J. Med., 37:332, 1964). The H chains in HCD are truncated, and subsequent structural studies showed that CH1 domains are almost always deleted (Corcos, et al., Blood, 117:6991, 2011). Mechanistically, CH1 deletion frees the H chain from its restraining interaction with BiP, thus allowing its secretion, and also prevents disulfide bond-mediated covalent association with L chains, thus the HCD proteins are HH dimers. Heavy chain-only Abs (HCAbs) can also be found in non-disease contexts. Approximately 75% of serum IgG in normal camels consists of HCAbs, which lack a CH1 domain and also have structurally altered VH domains that prevent effective association with VL domains (de los Rios, et al., Cur. Opin. Struct. Biol., 33:27, 2015). ii) Mice in which both x and k L chain gene loci are inactivated still produce serum IgG, but production of this antibody requires errors in class switch recombination (CSR) that lead to deletion of the CH1 domain-encoding exon in the B cell DNA (Zou, et al., J. Exp. Med., 204:3271, 2007). HCAbs are attractive as therapeutics since they are highly stable and smaller than conventional immunoglobulins.” (pg 2-3)
“Reports of HCO antibodies in llamas, transgenic mice and rats, indicated that B cells, in which H chain does not pair with L chain, can develop. However, CH1 needs to be deleted, and in llamas, the H chains without L chains have a VH structure different from that of the canonical VH regions. The latter fact indicated that not all human VH domains would support HCO B cell development” (pg 4).
WO 2019018770 A1 discloses a single chain VH antibody comprising an antigen- binding part consisting of a VH domain and the immunoglobulin constant domains CL and CH1.
WO 2014141192 A2 discloses generation of heavy-chain only antibodies and
transgenic non-human animals producing the same. Such antibodies lack the CH1 domain.
US8754287B2 discloses mice producing heavy-chain antibodies that lack the CH1 domain, and transgenic mice comprising a germline modification to delete the nucleic acid encoding a CH1 domain.
Expression of heavy-chain only antibodies with no associated light chains in VJCL knockout chickens is described by Schusser et al. (Eur. J. Immunol., 46:2137, 2016).
Klein et al. (Biochemistry, 18:1473, 1979) describe the interaction of isolated variable and constant domains of light chain with the Fd' fragment of immunoglobulin G.
WO 2011072204 A1 discloses a transgenic mouse containing a functional Cμ segment. As a result, mature B cells, with canonical IgM as antigen receptor, develop normally. However, the downstream Cy segments of the genome lack the CH1 domain.
When during an immune response, the antibody class switches to the (desired) y chains, these chains cannot pair with L chain. In the H chain class switch, VH is kept, whereas CH is exchanged. This will have two effects: (i) Cells with antibodies in which specificity was defined by a combination of H plus L chain will no longer be stimulated and will die. And (ii), cells in which the specificity was mainly defined by the H chain may die because the unpaired VH is structurally not viable. Thus, in this mouse, VH selection is "backloaded," i.e. during the immune response.
WO 2007096779 A2 describes use of a transgenic mouse for the production of class-specific heavy chain-only antibodies, said mouse comprising more than one heterologous VH heavy chain locus, wherein each VH heavy chain locus comprises one or more V gene segments, one or more D gene segments, one or more J gene segments and a gene segment encoding a heavy chain constant region which, when expressed, does not include a CH1 domain.
WO 2009013620 A2 describes the production of fully-human, soluble VH domains by incorporating human V segments into a mouse heavy chain locus, wherein the mouse V, D and J gene segments are replaced by V, D and J segments of human origin, and the immunoglobulin heavy chain effector constant regions are replaced by immunoglobulin heavy chain effector constant regions which are devoid of CH1.
WO 2015143414 A2 describes a mouse comprising a deletion in an immunoglobulin constant region CH1 gene of a heavy chain constant region gene sequence, and replacement of one or all endogenous VH, DH and JH gene segments with at least one unrearranged VL gene segment and at least one unrearranged JL gene segment.
WO 2019184014A1 describes a mouse comprising a transgenic Cy gene segment which has been introduced into the immunoglobulin locus in place of the endogenous Cμ gene segment and which comprises a deletion of the CH1 domain (see Fig. 1).
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EP2411408B1 and US 9353179 B2 describe transgenic mice comprising randomly integrated VH and VHH loci. The problems of randomly integrated immunoglobulin (and other) loci have been documented since the Eighties of the last century: unphysiological levels of expression and instability of the inserted transgenes (loss or functional silencing).
Fig. 1 shows a mouse Ig heavy chain gene, Ig light chain kappa (κ) gene, and an Ig light chain lambda (λ) gene.
Fig. 2 shows a targeting vector that removes the CH1 coding sequence in the Ig heavy chain μ gene and was described in 20130219535 (pg 30, line 16). It does not show a “transgenic C [light chain]λ gene segment upstream of the endogenous Cμ gene segment” “wherein the Cλ gene segment comprises a deletion of a nucleotide sequence encoding at least part of the CH1 domain” as required in claim 1. Pg 30, which describes Fig. 2, says the vector is for modifying the CH1 domain of a heavy chain Cμ gene and not the CH1 domain of a light chain Cλ as claimed.
Fig. 3 shows amouse Ig heavy chain gene after integration of the targeting vector. It does not show a “transgenic C [light chain]λ gene segment upstream of the endogenous Cμ gene segment” “wherein the Cλ gene segment comprises a deletion of a nucleotide sequence encoding at least part of the CH1 domain” as required in claim 1. Pg 30, which describes Fig. 2, says the vector is for modifying the CH1 domain of a heavy chain Cμ gene and not the CH1 domain of a light chain Cλ as claimed.
Example 1 (pg 63) contemplates inactivating a CH1 domain in an Ig heavy chain μ gene using the targeting vector in Fig. 2. The targeting vector in Fig. 2 shows a “[heavy chain] Cγ gene segment upstream of [an] endogenous [heavy chain] Cμ gene segment” “wherein the Cγ gene segment comprises a deletion of a nucleotide sequence encoding at least part of the CH1 domain” (“Ighg1 ΔCH1” in Fig. 2) as required in claim 1. Pg 30 and Example 1 (pg 63) describe Fig. 2, but the Ig Cμ gene starts in the 5’ intron and terminates at the 3’ intron to for a homology arm. Pg 64, lines 26-30, says stop codons were introduced into the CH1, CH2, and CH23 exons of the Ig Cμ gene to inactivate the Ig Cμ gene as required in claim 1. Overall, the specification is limited to a mouse/mouse cell whose genome comprises an endogenous heavy chain Ig gene with a deletion of a nucleic acid sequence encoding a CH1 domain of an Ig Cγ gene upstream of an inactivated Ig Cμ gene, wherein the endogenous heavy chain Ig gene is capable of expressing a heavy chain Igγ that is missing a CH1 domain but is not capable of expressing a heavy chain Igμ. The specification does not correlate this limited embodiment to incorporating a transgenic Cγ gene segment with a deletion of a nucleic acid encoding at least part of the CH1 domain that is upstream of an inactivated Cμ gene by loss-of-function, a deletion of part of the Cμ gene segment, or by introducing one or more stop codons as broadly claimed. Nor does the specification correlate the mere incorporation of the transgene without qualifying the functional expression of the genetically modified heavy chain Ig gene, i.e. the gene must be capable of expressing a heavy chain Igγ that is missing a CH1 domain but it cannot express a heavy chain Igμ.
The specification lacks written description for when the mouse expresses a “diverse” repertoire of heavy chain only antibodies (HCAbs). It is unclear whether three antibody types are adequate or whether more are required. It is unclear how the genetic modification implied in claim 1 or 12 confers the ability to express a diverse repertoire of heavy chain only antibodies as required in the preamble.
The specification lacks written description for a mouse that produces HCAbs without inactivating the endogenous light chain Ig genes. The claim as written encompasses making a mouse with or without their endogenous light chain Ig genes. However, pg 65, Example 2, says the HCAbs are produced in mice lacking functional light chain Ig genes described in WO 2011/072204. The specification and the art do not correlate leaving the endogenous light chain Ig gene wild-type to obtaining HCAbs as broadly encompassed by claim 1.
The specification lacks written description for when an Eμ enhancer is “major” as required in claim 3. Ariizumi (PNAS, 1993, Vol. 90, pg 3695-3699) taught “The immunoglobulin heavy chain (Igh) intronic [heavy chain join ing region (JH-A chain constant region (C,u)] enhancer (E,u) is required for tissue-specific transcription of Igh genes and is essential for somatic recombination of diversity (D) and J segments” (abstract). The specification and the art do not teach when the Eμ enhancer is “major”. Therefore, the concept lacks written description.
The specification lacks written description for an endogenous Cγ gene segment “comprising” a Cγ1 gene as required in claim 4 other than an endogenous Cγ gene that IS a Cγ1 gene. An endogenous Cγ gene may be a Cγ1, Cγ2, Cγ3, et al. gene, but it does not “comprise” a Cγ1 gene along with other things as broadly encompassed by claim 4. The claim should be limited to an endogenous Cγ gene that IS a Cγ1 gene.
The specification lacks written description for a BiP chaperone-binding domain as required in claim 5. The specification and the art at the time of filing do not teach the meaning of the phrase. Accordingly, the concept lacks written description.
The specification lacks written description for a mouse with an inactivated or deleted endogenous Ig light chain gene, preferably a “loss-of-function mutations within, or deletion of, any of the endogenous kappa or lambda chain loci, or both” as required in claim 6 other than a mouse whose genome comprises an inactivated endogenous Ig light chain kappa gene and an inactivated endogenous Ig light chain lambda gene, wherein the mouse is incapable of producing an endogenous Ig light chain. The specification does not teach or suggest partial deletion of either light chain gene will allow production of heavy chain only antibodies as required in claim 1. The specification does not teach or suggest inactivation of one but not the other light chain gene will allow production of heavy chain only antibodies as required in claim 1. Both light chain genes must be completely inactivated. Accordingly, the concept lacks written description as broadly claimed.
The specification lacks written description for a “locus” in claims 6, 9, 11-14 for reasons set forth above.
The specification lacks written description for the genetic modification in claim 9 other than a mouse whose genome comprises a replacement of all endogenous Ig variable heavy chain (VH) gene segments with a plurality of human Ig VH gene segments. The claim encompasses putting one or more human VH gene segment anywhere in any cell of a mouse cell. The specification and the art at the time of filing are limited to a mouse whose genome comprises a replacement of all endogenous Ig VH gene segments with a plurality of human Ig VH gene segments. Accordingly, the concept lacks written description as broadly written.
The specification lacks written description for putting the human Ig VH gene segments under the control of any “expression control sequences” in a mouse as broadly encompassed by claim 9 other than operably linking the human Ig VH gene segments to an endogenous Ig heavy chain promoter. The claim encompasses inserting the human Ig VH gene segments anywhere in the mouse randomly or targeting integration into the endogenous Ig heavy chain gene. The control sequences may be of any species. The term “murine” encompasses mice and rats. However, the specification and the art at the time of filing is limited to operably linking the human Ig VH gene segments to an endogenous Ig heavy chain promoter. The specification does not correlate the mouse control sequences to any species of control sequences as broadly claimed. The specification does not correlate the mouse control sequences to any rat control sequences as inferred by “murine” in the “preferred” embodiment. Accordingly, the concept lacks written description as broadly written.
The specification lacks written description for claim 10 as broadly written. The claim requires the V, D, J sequences “are recombined to for a VDJ coding sequence expressing a VH binding site specifically recognizing the antigen, thereby obtaining a recombined VHcoding sequence in a given B cell, which upon differentiating into a plasma cell is capable of secreting an HCAb of the IgG type comprising an antigen-specific VH binding domain encoded by the recombined VH coding sequence”. It does not require it occurs in any mouse cell or in any mouse. Nor does it require the sequences are recombined in any B-cell. Recombination of Ig gene segments only occurs in B-cells. The phrase “the antigen” in claim 10 lacks antecedent basis, but it would only occur if an antigen were administered to the mouse after it was born and it would only occur in B-cells. Plasma cells are terminally differentiated B-cells, so it is unclear how/why/what applicants are attempting to further limit by saying “which upon differentiating into a plasma cell is capable of secreting an HCAb of the IgG type comprising an antigen-specific VH binding domain encoded by the recombined VH coding sequence” – the metes and bounds and intent of this phrase will require extensive clarification because it is unclear how it modifies the structure/function of the mouse beyond being able to produce a variety of HCAbs with different human VH domains that bind an antigen that has been introduced into the mouse.
Claim 11 lacks written description for reasons set forth in claim 1. The specification does not teach “providing one or more vectors” as claimed other than the single vector in Fig. 2. The phrase “murine” ES cell encompasses mouse and rat; however, the specification and the art are limited to making genetically modified mice using mouse ES cells. The specification does not teach making the Cμ “optionally inactive” in claim 11 because it is “inactive by loss-of-function, a deletion… …, or by one or more mutations” in claim 1. The specification is limited to “utilizing the transgenic cell” for reasons set forth above.
Claim 13 lacks written description for reasons set forth above. The claim encompasses putting 2 more VH, DH, JH gene segments into a VH gene; however, the specification and the art is limited to replacing all endogenous VH gene segments with a plurality of human VH, DH, and JH gene segments.
Claim 14 lacks written description for reasons set forth above. Both lambda and kappa Ig light chain genes must be inactivated for producing a heavy chain only antibody.
Enablement
Claims 1-6, 9-14 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 genetically modified mouse whose genome comprises an endogenous heavy chain Ig gene with a deletion of a nucleic acid sequence encoding a CH1 domain of an Ig Cγ gene upstream of an inactivated Ig Cμ gene, wherein the endogenous heavy chain Ig gene is capable of expressing a heavy chain Igγ that is missing a CH1 domain but is not capable of expressing a heavy chain Igμ, does not reasonably provide enablement for making/using a mouse as broadly encompassed by claims 1 and 12. 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/use the invention commensurate in scope with these claims.
Claims 1 and 12 are recited above.
The specification does not enable making/using an Ig heavy chain constant region “locus” as required in claim 2. First, “locus” is singular, and “loci” is plural (Wikipedia definition of “locus”, 2023; National Human Genome Res. Institute definition of “locus”, 2023). Second, locus (singular) is a position in space or an address on a chromosome. Third, the specification does not define an Ig heavy chain constant region “locus” (singular) as and an Ig heavy chain constant gene (which must have a plurality of nucleotides) or as a plurality of nucleotides encoding an Ig heavy chain constant domain. Fourth, the specification is limited to replacing a plurality of endogenous nucleotides (at a plurality of contiguous “loci” (plural)) with a plurality of exogenous nucleotides. The specification does not teach the exogenous nucleotides are present at a single locus, position, or address on the chromosome as claimed. Replacing pluralities of nucleotides is not replacing a nucleotide at a single locus as claimed. Fifth, the concept claimed does not accurately set forth the genetic modification because the addresses and positions of the nucleotides within the endogenous Ig heavy chain constant gene have changed because of differences in the length caused by the genetic modification. Accordingly, making/using an Ig heavy chain constant region “locus” with a genetic modification in a mouse is not enabled as written; genetically modifying a mouse gene IS enabled.
The specification does not enable making/using a mouse having the features claimed by simply incorporating a transgenic Cγ gene segment upstream of a Cμ gene segment as broadly encompassed by claims 1 and 12. Claim 1 never clearly sets for incorporating the transgene into a mouse cell. Nor does it set forth any other active steps, i.e.
i) genetically modifying a mouse embryonic stem (ES) cell such that the genome of the ES cells comprises an endogenous immunoglobulin (Ig) heavy chain constant γ (Cγ) gene with a deletion of a nucleic acid sequence encoding at least part of the CH1 domain and an inactivated endogenous Ig heavy chain constant μ (Cμ) gene;
ii) introducing the ES cell into a recipient mouse embryo;
iii) implanting the mouse embryo into a recipient female mouse such that a genetically modified mouse whose genome comprises… …is obtained, wherein the genetically modified mouse is capable of expressing an endogenous Ig Cγ that is missing a CH1 domain and is incapable of expressing an endogenous Ig Cμ.
Claims 1 and 12 never result in a mouse having a genome comprising a genetic modification. Nor do they require the mouse is capable of expressing a diverse repertoire of heavy chain only antibodies.
The teachings in the specification and the art at the time of filing are summarized above.
Fig. 1 shows a mouse Ig heavy chain gene, Ig light chain kappa (κ) gene, and an Ig light chain lambda (λ) gene.
Fig. 2 shows a targeting vector that removes the CH1 coding sequence in the Ig heavy chain μ gene and was described in 20130219535 (pg 30, line 16). It does not show a “transgenic C [light chain]λ gene segment upstream of the endogenous Cμ gene segment” “wherein the Cλ gene segment comprises a deletion of a nucleotide sequence encoding at least part of the CH1 domain” as required in claim 1. Pg 30, which describes Fig. 2, says the vector is for modifying the CH1 domain of a heavy chain Cμ gene and not the CH1 domain of a light chain Cλ as claimed.
Fig. 3 shows a mouse Ig heavy chain gene after integration of the targeting vector. It does not show a “transgenic C [light chain]λ gene segment upstream of the endogenous Cμ gene segment” “wherein the Cλ gene segment comprises a deletion of a nucleotide sequence encoding at least part of the CH1 domain” as required in claim 1. Pg 30, which describes Fig. 2, says the vector is for modifying the CH1 domain of a heavy chain Cμ gene and not the CH1 domain of a light chain Cλ as claimed.
Example 1 (pg 63) contemplates inactivating a CH1 domain in an Ig heavy chain μ gene using the targeting vector in Fig. 2. The targeting vector in Fig. 2 shows a “[heavy chain] Cγ gene segment upstream of [an] endogenous [heavy chain] Cμ gene segment” “wherein the Cγ gene segment comprises a deletion of a nucleotide sequence encoding at least part of the CH1 domain” (“Ighg1 ΔCH1” in Fig. 2) as required in claim 1. Pg 30 and Example 1 (pg 63) describe Fig. 2, but the Ig Cμ gene starts in the 5’ intron and terminates at the 3’ intron to for a homology arm. Pg 64, lines 26-30, says stop codons were introduced into the CH1, CH2, and CH23 exons of the Ig Cμ gene to inactivate the Ig Cμ gene as required in claim 1. Overall, the specification is limited to a mouse/mouse cell whose genome comprises an endogenous heavy chain Ig gene with a deletion of a nucleic acid sequence encoding a CH1 domain of an Ig Cγ gene upstream of an inactivated Ig Cμ gene, wherein the endogenous heavy chain Ig gene is capable of expressing a heavy chain Igγ that is missing a CH1 domain but is not capable of expressing a heavy chain Igμ. The specification does not correlate this limited embodiment to incorporating a transgenic Cγ gene segment with a deletion of a nucleic acid encoding at least part of the CH1 domain that is upstream of an inactivated Cμ gene by loss-of-function, a deletion of part of the Cμ gene segment, or by introducing one or more stop codons as broadly claimed. Nor does the specification correlate the mere incorporation of the transgene without qualifying the functional expression of the genetically modified heavy chain Ig gene, i.e. the gene must be capable of expressing a heavy chain Igγ that is missing a CH1 domain but it cannot express a heavy chain Igμ.
The specification does not enable those of skill to determine when the mouse expresses a “diverse” repertoire of heavy chain only antibodies (HCAbs). It is unclear whether three antibody types are adequate or whether more are required. It is unclear how the genetic modification implied in claim 1 or 12 confers the ability to express a diverse repertoire of heavy chain only antibodies as required in the preamble.
The specification does not enable those of skill to determine how to make a mouse that produces HCAbs without inactivating the endogenous light chain Ig genes as broadly encompassed by claims 1 and 12. The claims as written encompass making a mouse with or without their endogenous light chain Ig genes. However, pg 65, Example 2, says the HCAbs are produced in mice lacking functional light chain Ig genes described in WO 2011/072204. The specification and the art do not correlate leaving the endogenous light chain Ig gene wild-type to obtaining HCAbs as broadly encompassed by claims 1 and 12.
The specification does not enable those of skill to determine when an Eμ enhancer is “major” as required in claim 3. Ariizumi (PNAS, 1993, Vol. 90, pg 3695-3699) taught “The immunoglobulin heavy chain (Igh) intronic [heavy chain join ing region (JH-A chain constant region (C,u)] enhancer (E,u) is required for tissue-specific transcription of Igh genes and is essential for somatic recombination of diversity (D) and J segments” (abstract). The specification and the art do not teach when the Eμ enhancer is “major”.
The specification does not enable making/using an endogenous Cγ gene segment “comprising” a Cγ1 gene as required in claim 4 other than an endogenous Cγ gene that IS a Cγ1 gene. An endogenous Cγ gene may be a Cγ1, Cγ2, Cγ3, et al. gene, but it does not “comprise” a Cγ1 gene along with other things as broadly encompassed by claim 4. The claim should be limited to an endogenous Cγ gene that IS a Cγ1 gene.
The specification does not enable making/using for a BiP chaperone-binding domain as required in claim 5. The specification and the art at the time of filing do not teach the meaning of the phrase.
The specification does not enable making/using a mouse with an inactivated or deleted endogenous Ig light chain gene, preferably a “loss-of-function mutations within, or deletion of, any of the endogenous kappa or lambda chain loci, or both” as required in claim 6 other than a mouse whose genome comprises an inactivated endogenous Ig light chain kappa gene and an inactivated endogenous Ig light chain lambda gene, wherein the mouse is incapable of producing an endogenous Ig light chain. The specification does not teach or suggest partial deletion of either light chain gene will allow production of heavy chain only antibodies as required in claim 1. The specification does not teach or suggest inactivation of one but not the other light chain gene will allow production of heavy chain only antibodies as required in claim 1. Both light chain genes must be completely inactivated.
The specification does not enable a “locus” in claims 6, 9, 11-14 for reasons set forth above.
The specification does not enable the genetic modification in claim 9 other than a mouse whose genome comprises a replacement of all endogenous Ig variable heavy chain (VH) gene segments with a plurality of human Ig VH gene segments. The claim encompasses putting one or more human VH gene segment anywhere in any cell of a mouse cell. The specification and the art at the time of filing are limited to a mouse whose genome comprises a replacement of all endogenous Ig VH gene segments with a plurality of human Ig VH gene segments.
The specification does not enable putting the human Ig VH gene segments under the control of any “expression control sequences” in a mouse as broadly encompassed by claim 9 other than operably linking the human Ig VH gene segments to an endogenous Ig heavy chain promoter. The claim encompasses inserting the human Ig VH gene segments anywhere in the mouse randomly or targeting integration into the endogenous Ig heavy chain gene. The control sequences may be of any species. The term “murine” encompasses mice and rats. However, the specification and the art at the time of filing is limited to operably linking the human Ig VH gene segments to an endogenous Ig heavy chain promoter. The specification does not correlate the mouse control sequences to any species of control sequences as broadly claimed. The specification does not correlate the mouse control sequences to any rat control sequences as inferred by “murine” in the “preferred” embodiment.
The specification does not enable claim 10 as broadly written. The claim requires the V, D, J sequences “are recombined to for a VDJ coding sequence expressing a VH binding site specifically recognizing the antigen, thereby obtaining a recombined VHcoding sequence in a given B cell, which upon differentiating into a plasma cell is capable of secreting an HCAb of the IgG type comprising an antigen-specific VH binding domain encoded by the recombined VH coding sequence”. It does not require it occurs in any mouse cell or in any mouse. Nor does it require the sequences are recombined in any B-cell. Recombination of Ig gene segments only occurs in B-cells. The phrase “the antigen” in claim 10 lacks antecedent basis, but it would only occur if an antigen were administered to the mouse after it was born and it would only occur in B-cells. Plasma cells are terminally differentiated B-cells, so it is unclear how/why/what applicants are attempting to further limit by saying “which upon differentiating into a plasma cell is capable of secreting an HCAb of the IgG type comprising an antigen-specific VH binding domain encoded by the recombined VH coding sequence” – the metes and bounds and intent of this phrase will require extensive clarification because it is unclear how it modifies the structure/function of the mouse beyond being able to produce a variety of HCAbs with different human VH domains that bind an antigen that has been introduced into the mouse.
Claim 11 is not enabled for reasons set forth in claim 1. The specification does not teach “providing one or more vectors” as claimed other than the single vector in Fig. 2. The phrase “murine” ES cell encompasses mouse and rat; however, the specification and the art are limited to making genetically modified mice using mouse ES cells. The specification does not teach making the Cμ “optionally inactive” in claim 11 because it is “inactive by loss-of-function, a deletion… …, or by one or more mutations” in claim 1. The specification is limited to “utilizing the transgenic cell” for reasons set forth above.
Claim 13 is not enabled for reasons set forth above. The claim encompasses putting 2 more VH, DH, JH gene segments into a VH gene; however, the specification and the art is limited to replacing all endogenous VH gene segments with a plurality of human VH, DH, and JH gene segments.
Claim 14 is not enabled for reasons set forth above. Both lambda and kappa Ig light chain genes must be inactivated for producing a heavy chain only antibody.
Given the lack of guidance in the specification taken with the art at the time of filing, it would have required those of skill undue experimentation to determine how to make/use the mouse as broadly encompassed by claims 1 and 12.
Indefiniteness
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claims 1-6, 9-14 are rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
The metes and bounds of an Ig heavy chain constant region “locus” in claim 2 cannot be determined. First, “locus” is singular, and “loci” is plural (Wikipedia definition of “locus”, 2023; National Human Genome Res. Institute definition of “locus”, 2023). Second, locus (singular) is a position in space or an address on a chromosome. Third, the specification does not define an Ig heavy chain constant region “locus” (singular) as and an Ig heavy chain constant gene (which must have a plurality of nucleotides) or as a plurality of nucleotides encoding an Ig heavy chain constant domain. Fourth, the specification is limited to replacing a plurality of endogenous nucleotides (at a plurality of contiguous “loci” (plural)) with a plurality of exogenous nucleotides. The specification does not teach the exogenous nucleotides are present at a single locus, position, or address on the chromosome as claimed. Replacing pluralities of nucleotides is not replacing a nucleotide at a single locus as claimed. Fifth, the concept claimed does not accurately set forth the genetic modification because the addresses and positions of the nucleotides within the endogenous Ig heavy chain constant gene have changed because of differences in the length caused by the genetic modification. Accordingly, the concept of an Ig heavy chain constant region “locus” with a genetic modification in a mouse is in indefinite; a mouse with a genetically modified Ig heavy chain gene WOULD be definite.
Claim 1 is rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being incomplete for omitting essential steps, such omission amounting to a gap between the steps. See MPEP § 2172.01. It never clearly sets for incorporating the transgene into a mouse cell. Nor does it set forth any other active steps, i.e.
i) genetically modifying a mouse embryonic stem (ES) cell such that the genome of the ES cells comprises an endogenous immunoglobulin (Ig) heavy chain constant γ (Cγ) gene with a deletion of a nucleic acid sequence encoding at least part of the CH1 domain and an inactivated endogenous Ig heavy chain constant μ (Cμ) gene;
ii) introducing the ES cell into a recipient mouse embryo;
iii) implanting the mouse embryo into a recipient female mouse such that a genetically modified mouse whose genome comprises… …is obtained, wherein the genetically modified mouse is capable of expressing an endogenous Ig Cγ that is missing a CH1 domain and is incapable of expressing an endogenous Ig Cμ.
Claims 1 and 12 never result in a mouse having a genome comprising a genetic modification. Nor do they require the mouse is capable of expressing a diverse repertoire of heavy chain only antibodies.
The metes and bounds of when the mouse expresses a “diverse” repertoire of heavy chain only antibodies (HCAbs) cannot be determined. It is unclear whether three antibody types are adequate or whether more are required. It is unclear how the genetic modification implied in claim 1 or 12 confers the ability to express a diverse repertoire of heavy chain only antibodies as required in the preamble.
The metes and bounds of when an Eμ enhancer is “major” cannot be determined as required in claim 3. Ariizumi (PNAS, 1993, Vol. 90, pg 3695-3699) taught “The immunoglobulin heavy chain (Igh) intronic [heavy chain join ing region (JH-A chain constant region (C,u)] enhancer (E,u) is required for tissue-specific transcription of Igh genes and is essential for somatic recombination of diversity (D) and J segments” (abstract). The specification and the art do not teach when the Eμ enhancer is “major”.
The concept of an endogenous Cγ gene segment “comprising” a Cγ1 gene as required in claim 4 cannot be determined. While an endogenous Cγ gene may BE a Cγ1, Cγ2, Cγ3, et al. gene, it does not “comprise” a Cγ1 gene along with other things as broadly encompassed by claim 4. The claim should be limited to an endogenous Cγ gene that IS a Cγ1 gene.
The specification metes and bounds of a BiP chaperone-binding domain cannot be determined as required in claim 5. The specification and the art at the time of filing do not teach the meaning of the phrase. It is unclear what BiP stands for or what structures/functions are associated with BiP. It is unclear whether the part of the CH1 domain that has been deleted contains the BiP or whether the coding sequence for the CH1 domain must include a BiP chaperone-binding domain (whether it is deleted or not).
The concept of a mouse with an “inactivated or deleted endogenous Ig light chain gene, preferably a loss-of-function mutations within, or deletion of, any of the endogenous kappa or lambda chain loci, or both” as required in claim 6 makes the claim indefinite other than a mouse whose genome comprises an inactivated endogenous Ig light chain kappa gene and an inactivated endogenous Ig light chain lambda gene, wherein the mouse is incapable of producing an endogenous Ig light chain. The phraseology fails to clearly set forth that one or both Ig light genes have been inactivated or that the mice are incapable of expressing one or both Ig light chains.
The metes and bounds of “locus” in claims 6, 9, 11-14 cannot be determined for reasons set forth above.
The concept in claim 10 is indefinite. The claim requires the V, D, J sequences “are recombined to for a VDJ coding sequence expressing a VH binding site specifically recognizing the antigen, thereby obtaining a recombined VHcoding sequence in a given B cell, which upon differentiating into a plasma cell is capable of secreting an HCAb of the IgG type comprising an antigen-specific VH binding domain encoded by the recombined VH coding sequence”. It does not require it occurs in any mouse cell or in any mouse. Nor does it require the sequences are recombined in any B-cell. Recombination of Ig gene segments only occurs in B-cells. The phrase “the antigen” in claim 10 lacks antecedent basis, but it would only occur if an antigen were administered to the mouse after it was born and it would only occur in B-cells. Plasma cells are terminally differentiated B-cells, so it is unclear how/why/what applicants are attempting to further limit by saying “which upon differentiating into a plasma cell is capable of secreting an HCAb of the IgG type comprising an antigen-specific VH binding domain encoded by the recombined VH coding sequence” – the metes and bounds and intent of this phrase will require extensive clarification because it is unclear how it modifies the structure/function of the mouse beyond being able to produce a variety of HCAbs with different human VH domains that bind an antigen that has been introduced into the mouse.
Claim 11 is indefinite because phrase “murine” ES cell encompasses mouse and rat; however, the “endogenous” Ig genes in a genetically modified mice must use mouse ES cells.
The concept of making the Cμ “optionally inactive” in claim 11 is broader than the concept in claim 1 because it clearly requires the Cμ is “inactive by loss-of-function, a deletion… …, or by one or more mutations”. Therefore, the concept makes the claim indefinite.
W02019184014A1 describes a mouse comprising a transgenic Cy gene segment which has been introduced into the immunoglobulin locus in place of the endogenous Cμ gene segment and which comprises a deletion of the CH1 domain (see Fig. 1).
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Janssens (PNAS, 2006, Vol. 103, No. 41, pg 15130-15135) taught genetically modified mice whose genomes comprised inactivated Ig light chain kappa and lambda genes and two llama VHH exons linked to human HC D, and J regions with ΔCH1s.
The art at the time of filing did not reasonably teach or suggest a mouse with an Ig heavy chain Cγ gene with a deletion of a CH1 domain upstream of an inactivated Ig heavy chain Cμ gene as required in claims 1 and 12.
Conclusion
No claim is allowed.
Inquiry concerning this communication or earlier communications from the examiner should be directed to Michael C. Wilson who can normally be reached at the office on Monday through Friday from 9:30 am to 6:00 pm at 571-272-0738.
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Michael C. Wilson
/MICHAEL C WILSON/
Primary Examiner, Art Unit 1638