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 .
Election/Restrictions
Applicant’s election without traverse of the identified species in the reply filed on May 7, 2024, is acknowledged.
Applicant elected the following species:
a. A4V as the distinct mutation in SOD1
b. A targeting sequence complementary to a sequence of a SOD1 exon
c. An AAV vector for contacting
In light of the Applicant’s elected species, claims 175 (the elected A4V mutation causes a gain of function mutation and, as such, results in a protein with a function and not a non-functional protein) and 188 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim.
Rejoinder
The species election requirement for the species of mutation in SOD1 has been reconsidered in view of the prior art. The G93A mutation is rejoined.
DETAILED ACTION
The amended claims filed on December 19, 2023, have been acknowledged. Claims 1-168 were cancelled. Claims 169-193 are new. In light of the Applicant’s elected species, claims 175 and 188 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim. Claims 169-174, 176-187, and 189-193 are pending and examined on the merits.
Priority
The applicant claims domestic priority from U.S. provisional application No. 62/897,941, filed on September 9, 2019. Applicant’s claim for the benefit of a prior-filed application under 35 U.S.C. 119(e) or under 35 U.S.C. 120, 121, 365(c), or 386(c) is acknowledged. Claims 169-174, 176-187, and 189-193 receive domestic benefit from U.S. provisional application No. 62/897,941, filed on September 9, 2019.
Information Disclosure Statement
The information disclosure statement (IDS) filed on July 10, 2023, has been considered.
Specification
The use of the terms ZYMOCLEAN, QIAPREP, GIBSON ASSEMBLY, and LIPOFECTAMINE, which are trade names or marks used in commerce, has been noted in this application. The term should be accompanied by the generic terminology; furthermore the term should be capitalized wherever it appears or, where appropriate, include a proper symbol indicating use in commerce such as ™, SM , or ® following the term.
Although the use of trade names and marks used in commerce (i.e., trademarks, service marks, certification marks, and collective marks) are permissible in patent applications, the proprietary nature of the marks should be respected and every effort made to prevent their use in any manner which might adversely affect their validity as commercial marks.
Drawings
The drawings are objected to because the figures are not properly labeled.
37 CFR 1.84 (u)(1) states “The different views must be numbered in consecutive Arabic numerals, starting with 1, independent of the numbering of the sheets and, if possible, in the order in which they appear on the drawing sheet(s). Partial views intended to form one complete view, on one or several sheets, must be identified by the same number followed by a capital letter. View numbers must be preceded by the abbreviation "FIG." Where only a single view is used in an application to illustrate the claimed invention, it must not be numbered and the abbreviation "FIG." must not appear.”
The drawings are objected to because Figure 33 is improperly labeled. Figure 33 contains partial views on separate sheets. For example, Figure 33 spans multiple separate sheets and is labeled “Fig. 33” and “Fig. 33 (continued)” but should be labeled “Fig. 33A,” “Fig. 3B,” etc., for each new page of the drawing.
Corrected drawing sheets in compliance with 37 CFR 1.121(d) are required in reply to the Office action to avoid abandonment of the application. Any amended replacement drawing sheet should include all of the figures appearing on the immediate prior version of the sheet, even if only one figure is being amended. The figure or figure number of an amended drawing should not be labeled as “amended.” If a drawing figure is to be canceled, the appropriate figure must be removed from the replacement sheet, and where necessary, the remaining figures must be renumbered and appropriate changes made to the brief description of the several views of the drawings for consistency. Additional replacement sheets may be necessary to show the renumbering of the remaining figures. Each drawing sheet submitted after the filing date of an application must be labeled in the top margin as either “Replacement Sheet” or “New Sheet” pursuant to 37 CFR 1.121(d). If the changes are not accepted by the examiner, the applicant will be notified and informed of any required corrective action in the next Office action. The objection to the drawings will not be held in abeyance.
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 169-174, 176-187, and 189-193 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.
Claims 169-174, 176-187, and 189-193 encompass a genus of CasX variant proteins with at least 70% sequence identity to SEQ ID NO: 126 comprising protein domains from two or more different CasX proteins. Furthermore, the recited sequence is recited with the functionality of being a CasX protein that can modify SOD1, and therefore the claim requires functional limitations associated with the associated structure recited (the genus of proteins which are 70% identical to SEQ ID NO: 126 which also function as CasX proteins that can modify SOD1). This claim language is problematic because it recites a genus of proteins which are 70% identical to SEQ ID NO: 126 which are functionally CasX proteins. However, as discussed further below, the Applicant did not show possession commensurate in scope with what is being claimed, as it is known in the art that such large variations in uncharacterized Cas enzymes is known to cause unpredictable changes with regards to the protein’s functionality.
Regarding the genus of CasX variants of claim 169, under the new Written Description Guidelines for antigen binding proteins molecules, the Examiner is directed to determine whether one skilled in the art would recognize that the applicant was in possession of the claimed invention as a whole at the time of filing. The following considerations are critical to this determination: on 22 February 2018, the USPTO provided a Memorandum clarifying the Written Description Guidelines for claims drawn to antibodies, which can be found at www.uspto.gov/sites/default/files/documents/amgen_22feb2018.pdf. That Memorandum indicates that, in compliance with recent legal decisions, the disclosure of a fully characterized antigen no longer is sufficient written description of an antibody to that antigen. Accordingly, the instant claims have been re-evaluated in view of that guidance.
“[T]he purpose of the written description requirement is to ‘ensure that the scope of the right to exclude, as set forth in the claims, does not overreach the scope of the inventor’s contribution to the field of art as described in the patent specification.’” Ariad Pharm., Inc. v. Eli Lilly & Co., 598 F.3d 1336, 1353-54 (Fed. Cir. 2010) (en banc) (quoting Univ. of Rochester v. G.D. Searle & Co., 358 F.3d 916, 920 (Fed. Cir. 2004)). To satisfy the written description requirement, the specification must describe the claimed invention in sufficient detail that one skilled in the art can reasonably conclude that the inventor had possession of the claimed invention. Vas-Cath, Inc. v. Mahurkar, 935 F.2d 1555, 1562-63, 19 USPQ2d 1111 (Fed. Cir. 1991). See also MPEP 2163.04.
Actual Reduction to Practice
In regard to the genus of CasX varaints of claim 169, the specification does not provide any examples of CasX variants that fall within 70% sequence identity to SEQ ID NO: 126 and comprise protein domains from two or more different CasX proteins that were shown to modify SOD1. Example 5 identifies that CasX variants 397, 395, 485-491, and 494 are CasX variants that comprise protein domains from two or more different CasX proteins. These CasX variants corresponds to amino acid sequences SEQ ID NOs: 240, 242, 244, 246, 248, 250, 252, 254, 256, and 258, respectively. These SEQ ID NOs are at least 96.8% similar to SEQ ID NO: 126. However, each of these variants were only discussed as being created but were not used to assess genetic modification of the SOD1 gene (Example 5). Instead, Examples 10 and 15 discuss using CasX2 with guide 2, CasX119 with guide 64, or CasX119 with guide 174. Although each of these combinations were shown to genetically modify SOD1, there were distinct differences in editing efficiency as CasX119 with guide 64 achieved 35-fold higher editing than CasX2 with guide 2 in HEK293T cells (Example 15). Furthermore, none of these CasX variants or and guide RNAs, or cells comprising both, were administered to a subject to assess treatment efficacy of a SOD1 disease. Therefore, the Applicant has not reduced to practice that any of their CasX variants that fall in the genus of claim 169 are capable of treating a SOD1-related disease by administering a composition comprising the CasX variant to a subject.
Disclosure of structure
The Applicant has provided multiple SEQ ID NOs as identified above that correspond to CasX variants that fall within the genus of claim 169. However, the specification does not provide a nexus between the structure of the claimed genus of CasX variants and the ability to treat SOD1 related diseases.
Furthermore, the Applicant has offered CasX variants with different domain substitutions; however, the Applicant has not recited or elucidated any key structural-functional relationships between the recited genus of protein sequences and their ability to function as CasX proteins. For instance, the Applicant has not performed structural or mutagenesis studies to identify enzymatic domains which may be critical for functionality regarding the recited genus. For instance, the recited genus allows for proteins which have 30% of their amino acids deleted from the final protein structure; however, the Applicant has not identified such regions that are associated with the functionality of the CasX proteins.
Additionally, the Applicant has not demonstrated possession of a method to treat a SOD1-related disease, as they have only offered a prophetic example of the compositions be tested in an animal model in the final Example (Example 19). Thus, given the unpredictability of the components of the method, which includes the highly unpredictable genus of CasX variants, the Applicant has not shown possession of any treatment method to treat a SOD1-associated disease, where furthermore they were not in possession of recited claim limitations such as any improvement in a clinically relevant end-point (claim 193) because 1) the Applicant has not tested any such method and 2) the components which are unpredictable and uncharacterized (i.e., the genus of CasX variant with 70% identity to SEQ ID NO:126) have furthermore not been characterized in vivo to treat any subject.
The present specification provides no guidance nor description to any rational in choosing the sequences that may be mutated as the Applicant has provided limited examples of CasX variants that fall within the genus of claim 169 and each of those sequences have at least 96.8% sequence identity, as well. Therefore the skilled artisan would not know what rational approach to take to make modifications with any predictable outcome on CasX gene editing function of SOD1. Therefore, it is incumbent on the applicant to provide this nexus between structure and function, in order to be given credit for possession of a larger genus of antibodies related to those comprising these individual species. Otherwise, the Written Description guidelines suggest that the applicant is entitled to only the species specifically recited as having this activity. Moreover, even when several species are disclosed, these are not necessarily representative of the entire genus. AbbVie Deutschland GMBH v. Janssen Biotech, 111 USPQ2d 1780, 1790 (Fed. Cir. 2014) (“The ’128 and ’485 patents, however, only describe species of structurally similar antibodies that were derived from Joe-9. Although the number of the described species appears high quantitatively, the described species are all of the similar type and do not qualitatively represent other types of antibodies encompassed by the genus.”). Thus, when there is substantial variation within the genus, one must describe a sufficient variety of species to reflect the variation within the genus to provide a "representative number” of species.
An applicant may show that an invention is complete by disclosure of sufficiently detailed, relevant identifying characteristics which provide evidence that applicant was in possession of the claimed invention, i.e., complete or partial structure, other physical and/or chemical properties, functional characteristics when coupled with a known or disclosed correlation between function and structure, or some combination of such characteristics. Enzo Biochem, 323 F.3d at 964, 63 USPQ2d at 1613.
Accordingly, if the skilled artisan sought to generate the claimed genus of CasX varinats, they would first need to know which sequences/domains could be chosen for mutation and combined and still be able to predictably produce a functional CasX protein. Hence, based on the new written description guidelines, the Examiner should conclude that the applicant was not in possession of the claimed genus of CasX variants
STATE OF THE ART & QUANTITY OF EXPERIMENTATION
Regarding the state of the art, it was known in the art at the time of filing that CasX proteins were novel Cas enzymes which had not been fully characterized. For instance, Yang et al. (Cell Res. 2019 May;29(5):345-346; referenced in IDS) – which was published the same year as the instantly filed invention and therefore is an accurate representation of the knowledge of the state of the art – teaches that CasX proteins were a novel class of enzyme discovered at the time of filing (Title, Abstract). Yang teaches that CasX was identified by metagenomic analysis of bacteria from groundwater and characterized as an RNA-guided DNA nuclease. It recognizes a 5′-TTCN PAM and is capable of plasmid interference in E. coli when presenting sgRNA (covalently linked crRNA-tracrRNA). It shares no similarity to other reported Cas endonucleases except for a RuvC domain located at the C-terminus. The above features of CasX correlate with those of type V Cas12; however, the size of CasX (~980 aa) is smaller than those of reported Cas12 (~1200 aa) (Intro, second paragraph).
Thus, Yang teaches that at the time of filing CasX was a novel enzyme which was not known to share similarity with other Cas enzymes. Yang further teaches that CasX is significantly smaller than other known Cas enzymes. Therefore, at the time of filing, the structural domains were still in the stages of being elucidated as CasX was known to have a novel structure. Yang concludes by saying that further structural analysis with higher resolution is required to guide CasX engineering for future applications (final sentence). As such, it was known in the art at the time of filing that CasX was a novel protein.
Applicant has claimed a genus of CasX variants, yet the specification has only managed to identify a limited set of example variants that are at most 3.2% different from SEQ ID NO: 126. Furthermore, Applicant did not assess whether any of these variants can edit SOD1, let alone whether they can edit SOD1 at sufficient levels to treat a SOD1 related disease when administered to a subject. Because Applicant has no manner a priori to predict which sequences of the CasX can be modified and used to make a functioning CasX protein that can modify SOD1, the genus of CasX variants claimed by Applicant cannot be predictably made or used by the ordinary artisan.
CONCLUSION
Therefore, the examiner concludes that there is insufficient written description of the instantly claimed genus. Specifically, there is limited description of the structure-function relationship between the claimed genus of CasX variants with as little as 70% sequence identity and their ability to produce functional CasX variants that can edit SOD1 and treat SOD1 related diseases, and the Examiner further concludes a skilled artisan would find the specification inadequately describes the claimed genus of CasX variants.
Claims 170-174, 176-187, and 189-193 depend from claim 169 and do not resolve this 112(a) issue. Claims 170-174, 176-187, and 189-193 are therefore also rejected.
Furthermore, claim 179 contains an additional 112(a) written description issue in that it recites a gNA with a scaffold sequence that is at least 70% identical to SEQ ID NO: 2238. Regarding this claim, the authors have not offered additional gNA scaffold structures tested with CasX variants with 70% identity to SEQ ID NO: 126 to show that such variants of both the CasX and scaffold are permitted. Recited together, the Applicant is reciting an enormous genus of CasX variant with 70% identity to SEQ ID NO: 126 and a gNA scaffold with 70% identity to SEQ ID NO: 2238, such that it is unknown which combinations of such scaffolds and CasX variants yield the functional requirements of the claims (i.e., that the molecules function as gNA scaffolds and CasX enzymes, where furthermore they function to “target” a SOD1 gene and treat a SOD1 related disease).
For instance, Yang teaches that the domain composition of CasX-sgRNA-DNA ternary complexes showed some similarity to that of Cas12; however, each structural element adopts distinct folds. They also made efforts at structure determination of apo CasX and CasX-sgRNA and obtained CasX-sgRNA maps at low resolution (7.5 Å). In combination with mass spectrometry data, they found that sgRNA assembly and DNA loading trigger domain rearrangements (page 345, right column, second paragraph).
Thus, Yang teaches that structural elements of the novel CasX-gRNA complex are unique, comprising unique folds, and that such structures must be elucidated empirically in order to determine functional structures. The Applicant has not characterized gNA scaffolds with up to 30% variation which can function as gNA with the similarly uncharacterized genus of CasX protein.
Claim Rejections - 35 USC § 103
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
Claims 169-174, 176-187, and 189-193 are rejected under 35 U.S.C. 103 as being unpatentable over World Intellectual Property Organization Patent Application No. 2019051278 (Qi; referenced in IDS), World Intellectual Property Organization Patent Application No. 2018064371 (Doudna; referenced in IDS), and Gaj et al. (Sci. Adv. 3: 1-10. 2017; referenced in IDS), as evidenced by MacLean et al. (Communications Biology 5: 1-16. 2022).
Regarding claims 169, 186-187, and 192, Qi teaches a method of treating SOD1 related ALS by genetically modifying a target sequence in cells of the subject. Qi teaches that CasX nucleases and gRNAs targeting a sequence involved in disease, such also SOD1 related ALS, can be used for genetic editing of the disease causing gene. Qi teaches that viral vectors, such as AAV vectors, can be used to deliver the Cas nuclease and gRNA to a cell for in vivo modification of target cells (paragraphs 0183, 200, 228-230, 277-284, and 302-307).
Qi is silent as to the sequence of the CasX protein.
However, Doudna teaches a composition comprising a CasX polypeptide and a CasX guide RNA. Doudna teaches that their CasX nuclease can be encoded by a viral vector for delivery to a eukaryotic cell. Doudna teaches that the CasX protein can have protein domains from two separate CasX proteins, an N-terminal domain (amino acids 1-650 of SEQ ID NO: 2) from CasX2 derived from Planctomycetes and a C-terminal domain (nucleotides 664-986 of SEQ ID NO: 1) from CasX1 derived from Deltaproteobacter (paragraphs 0068-00107). Doudna teach that the guide RNA contains a “guide sequence” or “targeting sequence” that can be modified so that the CasX guide RNA can target a CasX protein to any desired sequence of any desired target nucleic acid, with the exception that the PAM sequence can be taken into account (paragraphs 147-150).
The combined chimeric CasX protein disclosed by Doudna (bottom row) has 86.8% sequence identity to SEQ ID NO: 126 (top row), as shown below.
1 QEIKRINKIRRRLVKDSNTKKAGKTGPMKTLLVRVMTPDLRERLENLRKKPENIPQPISN 60
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
2 QEIKRINKIRRRLVKDSNTKKAGKTGPMKTLLVRVMTPDLRERLENLRKKPENIPQPISN 61
61 TSRANLNKLLTDYTEMKKAILHVYWEEFQKDPVGLMSRVAQPASKKIDQNKLKPEMDEKG 120
||||||||||||||||||||||||||||||||||||||||||| | ||| || | |
62 TSRANLNKLLTDYTEMKKAILHVYWEEFQKDPVGLMSRVAQPAPKNIDQRKLIPVKDGNE 121
121 NLTTAGFACSQCGQPLFVYKLEQVSEKGKAYTNYFGRCNVAEHEKLILLAQLKPEKDSDE 180
||::||||||| |||:|||||||::||| :|||||||||:|||:||||: ||| :||
122 RLTSSGFACSQCCQPLYVYKLEQVNDKGKPHTNYFGRCNVSEHERLILLSPHKPEA-NDE 180
181 AVTYSLGKFGQRALDFYSIHVTKESTHPVKPLAQIAGNRYASGPVGKALSDACMGTIASF 240
|||||||||||||||||||||:|| |||||| || || ||||||||||||||| :|||
181 LVTYSLGKFGQRALDFYSIHVTRESNHPVKPLEQIGGNSCASGPVGKALSDACMGAVASF 240
241 LSKYQDIIIEHQKVVKGNQKRLESLRELAGKENLEYPSVTLPPQPHTKEGVDAYNEVIAR 300
|:||||||:|||||:| |:||| :|:::| | :| :|||||||||||::||| |:|:
241 LTKYQDIILEHQKVIKKNEKRLANLKDIASANGLAFPKITLPPQPHTKEGIEAYNNVVAQ 300
301 VRMWVNLNLWQKLKLSRDDAKPLLRLKGFPSFPLVERQANEVDWWDMVCNVKKLINEKKE 360
: :|||||||||||: ||:|||| ||||||||||||||||||||||||||||||||||||
301 IVIWVNLNLWQKLKIGRDEAKPLQRLKGFPSFPLVERQANEVDWWDMVCNVKKLINEKKE 360
361 DGKVFWQNLAGYKRQEALRPYLSSEEDRKKGKKFARYQLGDLLLHLEKKHGEDWGKVYDE 420
|||||||||||||||||| ||||||||||||||||||| |||||||||||||||||||||
361 DGKVFWQNLAGYKRQEALLPYLSSEEDRKKGKKFARYQFGDLLLHLEKKHGEDWGKVYDE 420
421 AWERIDKKVEGLSKHIKLEEERRSEDAQSKAALTDWLRAKASFVIEGLKEADKDEFCRCE 480
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
421 AWERIDKKVEGLSKHIKLEEERRSEDAQSKAALTDWLRAKASFVIEGLKEADKDEFCRCE 480
481 LKLQKWYGDLRGKPFAIEAENSILDISGFSKQYNCAFIWQKDGVKKLNLYLIINYFKGGK 540
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
481 LKLQKWYGDLRGKPFAIEAENSILDISGFSKQYNCAFIWQKDGVKKLNLYLIINYFKGGK 540
541 LRFKKIKPEAFEANRFYTVINKKSGEIVPMEVNFNFDDPNLIILPLAFGKRQGREFIWND 600
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
541 LRFKKIKPEAFEANRFYTVINKKSGEIVPMEVNFNFDDPNLIILPLAFGKRQGREFIWND 600
601 LLSLETGSLKLANGRVIEKTLYNRRTRQDEPALFVALTFERREVLDSSNIKPMNLIGVDR 660
||||||||||||||||||||||||||||||||||||||||||||||||||||:|||||||
601 LLSLETGSLKLANGRVIEKTLYNRRTRQDEPALFVALTFERREVLDSSNIKPVNLIGVDR 660
661 GENIPAVIALTDPEGCPLSRFKDSLGNPTHILRIGESYKEKQRTIQAKKEVEQRRAGGYS 720
|||||||||||||||||| |||| | || |||||| |||||| ||| ||||||||||||
661 GENIPAVIALTDPEGCPLPEFKDSSGGPTDILRIGEGYKEKQRAIQAAKEVEQRRAGGYS 720
721 RKYASKAKNLADDMVRNTARDLLYYAVTQDAMLIFENLSRGFGRQGKRTFMAERQYTRME 780
||:|||::|||||||||:|||| |:||| ||:|:||||||||||||||||| |||||:||
721 RKFASKSRNLADDMVRNSARDLFYHAVTHDAVLVFENLSRGFGRQGKRTFMTERQYTKME 780
781 DWLTAKLAYEGL-SKTYLSKTLAQYTSKTCSNCGFTITSADYDRVLEKLKKTATGWMTTI 839
|||||||||||| |||||||||||||||||||||||||:|||| :| :||||: || ||:
781 DWLTAKLAYEGLTSKTYLSKTLAQYTSKTCSNCGFTITTADYDGMLVRLKKTSDGWATTL 840
840 NGKELKVEGQITYYNRYKRQNVVKDLSVELDRLSEESVNNDISSWTKGRSGEALSLLKKR 899
| |||| ||||||||||||| | |:|| ||||||||| ||||| ||||| ||| |||||
841 NNKELKAEGQITYYNRYKRQTVEKELSAELDRLSEESGNNDISKWTKGRRDEALFLLKKR 900
900 FSHRPVQEKFVCLNCGFETHADEQAALNIARSWLFLRSQ--EYKKYQTNKTTGNTDKRAF 957
||||||||:||||:|| | ||||||||||||||||| | |:| |:: |: |
901 FSHRPVQEQFVCLDCGHEVHADEQAALNIARSWLFLNSNSTEFKSYKSG-------KQPF 953
958 VETWQSFYRKKLKEVWKP 975
| ||:||:::|||||||
954 VGAWQAFYKRRLKEVWKP 971
It would have been obvious that this sequence could have been used as the sequence for the CasX as it was previously identified by Doudna as being a chimeric CasX that can be used as part of their composition and that their CasX proteins can be encoded by a viral vector for delivery to a eukaryotic cell Furthermore, the successful cloning and sequencing of a known protein/DNA encoding a known protein and making an amino acid sequence is obvious, and thus unpatentable, if (1) there was some suggestion or motivation in the prior art to clone the DNA, and (2) there was a “reasonable expectation of success,” based on "detailed enabling methodology" in the prior art. Ex parte Kubin, 83 U.S.P.Q.2d (BNA) 1410 (B.P.A.I. 2007), aff'd, 561 F.3d 1351 (Fed. Cir. 2009). Because the prior art teaches all of the elements of the claimed invention, there is a reasonable expectation of success.
Although the combined teachings of Qi and Doudna identify that gRNAs targeting the SOD1 gene related to ALS, they do not specifically identify the mutations responsible for ALS nor modifying the SOD1 gene in a CNS or PNS cell of the subject.
However, Gaj teaches a method of gene editing G93A-SOD1 mutant mice. They used Cas9 nuclease from Staphylococcus aureus (SaCas9) to target the mutant SOD1 gene in the G93A-SOD1 mouse model of ALS (5), which carries ~25 tandem repeat copies of the hSOD1G93A transgene and recapitulates many aspects of the disease, including progressive muscle atrophy and impaired motor function. Gaj injected AAV vectors encoding the Cas9 and a sgRNA targeting exon 2 of the SOD1 gene. Gaj found the presence of indels in ~94% of hSOD1G93A transgenes analyzed from these cells (fig. S2B), confirming that mutant SOD1 expression was disrupted by SaCas9-mediated genome editing, including in PNS cells of the spinal cord. Furthermore, the treated mice exhibited delayed disease onset and increased survival time (page 1, column 2, paragraph 2-page 4, column 2, paragraph 1, page 6, column 1, paragraph 1-page 7, column 2, paragraph 2, and Figures 1-3).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of treating SOD1 related ALS of the combined teachings of Qi and Doudna by targeting the G93A mutation and administering an AAV encoding the CasX protein and gRNA targeting exon 2, as identified by Gaj, to arrive at the instantly claimed invention. One of ordinary skill in the art would have a reason to modify with a reasonable expectation of success because Qi and Gaj discuss treating SOD1 related ALS by delivering AAV vectors encoding Cas nucleases and gRNAs for gene editing of the mutant target gene and Gaj successfully reduces to practice that a Cas nuclease and sgRNA targeting exon 2 of SOD1 can successfully delay disease onset of ALS from the G93A gene mutation and increase survival time through successful gene editing, including in PNS cells. Therefore, it would have been obvious that an AAV vector could encode the CasX nuclease of the combined teachings of Qi and Doudna a sgRNA targeting exon 2 and be administered to a subject comprising the G93A SOD1 mutation to treat ALS. Because the prior art teaches all of the elements of the claimed invention, there is a reasonable expectation of success.
Regarding claims 170, Gaj teaches that the G93A mutation is in exon 4 (Figure 1)
Regarding claim 171, the G93A mutation is a substitution mutation.
Regarding claim 172, MacLean evidences that the G93A mutation is considered a gain of function mutation (page 2, column 1, paragraph 1).
Regarding claims 173-174, Gaj, as stated supra, teaches that the mutation is the G93A mutation (Figure 1).
Regarding claims 176-177, Qi (paragraphs 0183, 200, 228-230, 260, 277-284, and 302-307), Doudna (paragraphs 0068-00107 and 00148), and Gaj (Figure 1) use their Cas nucleases in combination with a gRNA/sgRNA.
Regarding claim 178, Qi is silent as to the scaffold stem loop sequence of their guide RNAs to be used with the CasX nuclease.
However, Doudna teaches that the activator-RNA (e.g., in dual or single guide RNA format) comprises the tracrRNA sequence (bolded sequences are the same nucleotides as found in SEQ ID NO: 20 of the instant application) UUAUCUCAUUACUUUGAGAGCCAUCACCAGCGACUAUGUCGUAUGGGUAAAGCGCUUAUUUAUCGG (SEQ ID NO: 27) (paragraph 00190). As can be seen above, SEQ ID NO: 27 of Doudna only has two mismatches compared to SEQ ID NO: 20 of the instant application.
It would have been obvious that this sequence could have been used as the sequence for the gRNA scaffold stem loop sequence as it was previously identified by Doudna as being a viable scaffold sequence for a gRNA to be used with a CasX. Furthermore, the successful cloning and sequencing of a known RNA sequence/DNA encoding a known RNA sequence is obvious, and thus unpatentable, if (1) there was some suggestion or motivation in the prior art to clone the DNA, and (2) there was a “reasonable expectation of success,” based on "detailed enabling methodology" in the prior art. Ex parte Kubin, 83 U.S.P.Q.2d (BNA) 1410 (B.P.A.I. 2007), aff'd, 561 F.3d 1351 (Fed. Cir. 2009). Because the prior art teaches all of the elements of the claimed invention, there is a reasonable expectation of success.
Regarding claim 179, as an initial matter, the claim language “at least about 70% sequence identity” is broadly interpreted to include sequences that are at least 56% similar to SEQ ID NO: 2238 as applicant has not defined the terms “about”, “at least”, or “at least about” and ±20% is a common definition for about. The about is considered to modify the lower bounds of the at least term and allow for values less than 70%.
Qi is silent as to the scaffold sequence of their guide RNAs to be used with the CasX nuclease.
However, Doudna teaches a CasX single guide RNA comprises the sequence
UUAUCUCAUUACUUUGAGAGCCAUCACCAGCGACUAUGUCGUAUGGGUAAAGCGCUUAUUUAUCGGgaaaUCUCCGAUAAAUAAGAAGCAUCAAAG (SEQ ID NO: 43) that has 56.25% sequence identity to SEQ ID NO: 2238 of the instant application (as shown below).
1 ACTGGCGCTTTTATCTGATTACTTTGAGAGCCATCACCAGCGACTATGTCGTAGTGGGTA 60
|||||| |||||||||||||||||||||||||||||||||||| || |
1 UUAUCUCAUUACUUUGAGAGCCAUCACCAGCGACUAUGUCGUAUGGGUAA 50
61 AAGCTCCCTCTTCGGAGGGAGCATCAAAG 89
| | | | | | | | |
51 AGCGCUUAUUUAUCGGGAAAUCUCCGAUAAAUAAGAAGCAUCAAAG 96
It would have been obvious that this sequence could have been used as the sequence for the gRNA scaffold sequence as it was previously identified by Doudna as being a viable scaffold sequence for a gRNA to be used with a CasX. Furthermore, the successful cloning and sequencing of a known RNA sequence/DNA encoding a known RNA sequence is obvious, and thus unpatentable, if (1) there was some suggestion or motivation in the prior art to clone the DNA, and (2) there was a “reasonable expectation of success,” based on "detailed enabling methodology" in the prior art. Ex parte Kubin, 83 U.S.P.Q.2d (BNA) 1410 (B.P.A.I. 2007), aff'd, 561 F.3d 1351 (Fed. Cir. 2009). Because the prior art teaches all of the elements of the claimed invention, there is a reasonable expectation of success.
Regarding claim 180, as stated supra, Gaj teaches a sgRNA that targets exon 2 of SOD1 to treat SOD1 G93A based ALS (Figure 1).
Regarding claim 181, as an initial matter, the term language is complementary to a target nucleic acid sequence encoding G93A is interpreted to not require that the gRNA specifically targets and binds to a region of a nucleic acid that comprises a G93A mutation. Instead, this claim language is broadly interpreted to include any gRNA that binds to a target nucleic acid sequence that comprises a G93A mutation within the nucleic acid sequence but does not specifically need to bind to a region containing the G93A mutation. As the sgRNA of Gaj binds to exon 2 of a SOD1 nucleic acid that comprises the G93A in exon 4, this is considered to fall within the broadly interpreted gRNA of claim 181.
Regarding claim 182, Qi teaches that the Cas can comprise an NLS (paragraph 0306).
Regarding claim 183, Qi teaches that the genetic modification can occur through single or double strand breaks (paragraph 0092) and Doudna teaches that their CasX proteins can generate single or double strand breaks (paragraph 00343).
Regarding claim 184, Qi teaches that the genetic modification can occur through single or double strand breaks that cause a deletion, insertion, and/or mutation (paragraph 0092) and Gaj teaches that their editing led to insertions and deletions (Figure 1).
Regarding claim 185, as can be seen in Gaj, the insertions/deletions led to reduced expression of the hSOD1 gene in the mice. As such, a reduction would also occur in the combined method of Qi, Doudna, and Gaj.
Regarding claim 189, Qi teaches that vectors can be delivered in vivo by administration of to an individual subject through intravenous administration (paragraph 0307) and Gaj teaches that they used intravenous administration of the AAV vector encoding the Cas and sgRNA to treat their mice (Figure 1).
Regarding claim 190, Qi teaches that their method can be used to treat non-human mammals or humans (paragraph 00200) and Gaj teaches that they treated mice (Figure 1).
Regarding claim 191, although Qi teaches that their method can be used to express their CasX protein and sgRNA in neurons, they do not explicitly state targeting neurons in vivo.
However, Gaj teaches that their AAV9 vectors encoding the Cas and sgRNA transduced motor neurons (page 2, column 2, paragraph 2-page 4, column 2, paragraph 1 and Figure 1).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have used an AAV9 vector to target motor neurons in the combined method of treating ALS of Qi, Doudna, and Gaj to arrive at the instantly claimed invention. One of ordinary skill in the art would have a reason to use an AAV9 vector with a reasonable expectation of success because ALS is known to impact motor neurons and Gaj successfully reduces to practice that their AAV9 vectors encoding the Cas and sgRNA transduced motor neurons and delayed disease occurrence and lengthened survival time. Because the prior art teaches all of the elements of the claimed invention, there is a reasonable expectation of success.
Regarding claim 193, Gaj, as stated supra, teaches that targeting exon 2 with their Cas-sgRNA AAV vector led to delayed disease onset and increased survival time in their mice Figure 2). It would have been obvious that using a different Cas (CasX) as used in the combined method of Qi, Doudna, and Gaj would still lead to similar results by targeting the same region of exon 2 with the gRNA.
Claim 169-174, 181, and 192 are rejected under 35 U.S.C. 103 as being unpatentable over World Intellectual Property Organization Patent Application No. 2019051278 (Qi), World Intellectual Property Organization Patent Application No. 2018064371 (Doudna), and Gaj et al. (Sci. Adv. 3: 1-10. 2017), as evidenced by MacLean et al. (Communications Biology 5: 1-16. 2022) as applied to claim 169 above, and further in view of Kiskinis et al. (Cell Stem Cell 14: 781–795. 2014; referenced in IDS), as evidenced by Brasil et al. (Mol Neurobiol 55: 5269-5281. 2018).
The teachings of Qi, Doudna, and Gaj are as discussed above. Gaj teaches that a CRISPR system configured to target disease causing SOD1 mutations, including the A4V mutation, or a gene knockout and replace therapy can be used (page 5, column 1, paragraph 4).
Although Gaj discusses targeting the A4V mutation, the combined teachings of Qi, Doudna, and Gaj do not teach a method of targeting the A4V mutation to treat ALS.
However, Kiskinis teaches combined reprogramming and stem cell differentiation approaches with genome engineering and RNA sequencing (RNA-seq) technologies to identify the transcriptional and functional changes induced by the SOD1 A4V (a substitution) mutation in human motor neurons (MNs) (e.g., page 783, left column, paragraph 3). Kiskinis teach the use of a zinc finger nuclease (ZFN)-mediated gene targeting strategy to correct the A4V mutation in an ALS patient-derived iPSC cell line and provide an isogenic control. Kiskinis teaches that the A4V mutation is in Exon 1 of the SOD1 gene, and the double-strand break induced by the ZFN is upstream of exon 1 (Fig. 2). Kiskinis teaches that they were able to correct the mutant allele and replace it with a wildtype gene (page 783, column 1, paragraph 4-page 785, column 1, paragraph 2 and Fig. 2).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to target the A4V mutation instead of the G93A mutation, as identified by Gaj and Kiskinis, to arrive at the instantly claimed invention. One of ordinary skill in the art would have a reason to substitute with a reasonable expectation of success because Gaj specifically identifies that other mutations can be targeted for treating ALS using CRISP based therapies, including the A4V mutation, and Kiskinis successfully reduces to practice that the A4V mutation can be treated by using a nuclease to disrupt the mutant gene. Furthermore, Doudna teaches that CasX guide RNAs can target a CasX protein to any desired sequence of any desired target nucleic acid (paragraph 00147). Therefore, it would have been obvious that one could use a CasX with a gRNA targeting the same region as identified by Kiskinis to disrupt the A4V mutant gene and treat ALS.
Brasil evidences that the A4V gene mutation is considered a gain of function mutation (page 5278, column 2, paragraph 2).
Regarding claim 181, as an initial matter, the term language is complementary to a target nucleic acid sequence encoding A4V is interpreted to not require that the gRNA specifically targets and binds to a region of a nucleic acid that comprises a A4V mutation. Instead, this claim language is broadly interpreted to include any gRNA that binds to a target nucleic acid sequence that comprises an A4V mutation within the nucleic acid sequence but does not specifically need to bind to a region containing the A4V mutation. As the resulting gRNA based on the target of Kiskinis binds to a region upstream of exon 1 of a SOD1 nucleic acid that comprises the A4V in exon 1, this is considered to fall within the broadly interpreted gRNA of claim 181.
Double Patenting
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 169-174, 180-187, and 189-193 are provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claims 169-192 of copending Application No. 17/641,426 (reference application), as evidenced by Gaj et al. (Sci. Adv. 3: 1-10. 2017), Kiskinis et al. (Cell Stem Cell 14: 781–795. 2014), and Brasil et al. (Mol Neurobiol 55: 5269-5281. 2018). Although the claims at issue are not identical, they are not patentably distinct from each other.
As an initial matter, the method claims of ‘426 (claims 185-192) are dependent on the composition claim 169. Therefore, all limitations associated with the composition claims are considered to be imported into the method claims, as well.
Regarding claims 169, 173-174, 185-187, 189-191, and 193, ‘426 claims a method of modifying a SOD1 target nucleic acid sequence in a population of cells in vivo in a rodent, a mouse, a rat, a non-human primate, or a human, the method comprising introducing into cells of the population the vector of claim 184 (A vector comprising the nucleic acid of claim 183 (A nucleic acid comprising a sequence that encodes the CasX variant protein and/or the gNA of claim 169 (a composition comprising: a. a CasX variant protein comprising the sequence of SEQ ID NO: 126, or a sequence having at least about 70% sequence identity thereto; and b. a first guide nucleic acid (gNA), wherein the gNA comprises a targeting sequence complementary to a superoxide dismutase 1 (SOD1) gene target nucleic acid sequence, wherein the SOD1 gene comprises one or more mutations)), wherein the vector is selected from the group consisting of a retroviral vector, a lentiviral vector, an adenoviral vector, an adeno-associated viral (AAV) vector, a herpes simplex virus (HSV) vector, a virus-like particle (VLP), a plasmid, a minicircle, a nanoplasmid, a DNA vector, and an RNA vector)), wherein the SOD1 target nucleic acid sequence of the cells targeted by the first gNA is modified by the CasX variant protein, wherein the modifying results in a knocking down of the SOD1 gene expression in the cells of the population such that expression of a non-functional SOD 1 is decreased by at least about 10%, in comparison to a cell where the SOD1 gene has not been modified (claims 169, 183-185, and 190).
‘426 claims that the CasX protein is a chimeric protein comprising protein domains from two or more different CasX proteins (claim 180).
‘426 claims that the SOD1 gene can include one or more mutations selected from a group of a multitude of mutations, including A4V and G93A (claim 172).
‘426 claims that this genetic modification can occur in rodent cells, mouse cells, rat cells, non-human primate cells, or human cells, specifically identifying central nervous system (CNS) cells, and/or peripheral nervous system (PNS) cells, such as spinal motor neurons (claims 186-188).
‘426 claims wherein the vector is administered to the subject by a route of administration selected from intraparenchymal, intravenous, intra- arterial, intracerebroventricular, intracisternal, intrathecal, intracranial, lumbar, intraperitoneal, or combinations thereof (claim 191).
Gaj evidences that disrupting hSOD1G93A transgenes and reducing expression of the mutant gene/protein in mouse motor neuron cells caused the treated mice to exhibit delayed disease onset and increased survival time (page 1, column 2, paragraph 2-page 4, column 2, paragraph 1, page 6, column 1, paragraph 1-page 7, column 2, paragraph 2, and Figures 1-3).
‘426 claims a CasX composition further comprising a donor template nucleic acid, wherein the donor template comprises a nucleic acid comprising at least a portion of a wild-type SOD1 gene (claim 178).
Kiskinis evidences that using a nuclease to correct the mutant allele and replace it with a wild type gene improved the function of A4V patient derived motor neurons (page 783, column 1, paragraph 4-page 785, column 1, paragraph 2 and Fig. 2).
Therefore, although the method of claim 185-192 only identifies modifying a cell, Gaj and Kiskinis evidence that modification can also lead to alleviation of symptoms associated with A4V or G93A based ALS. Therefore, this method is also considered to treat ALS.
Regarding claims 170-171, the A4V mutation is in exon 1, as evidenced by Figure 2 of Kiskinis and the G93A mutation is in exon 4, as evidenced by Figure 1 of Gaj and are substitutions.
Regarding claim 172, Brasil evidences that the A4V and G93A gene mutations are considered gain of function mutations (page 5278, column 2, paragraph 2)
Regarding claim 180, ‘426 claims wherein the targeting sequence of the gNA is complementary to a sequence of a SOD1 exon (claim 170).
Regarding claim 181, as an initial matter, the term language is complementary to a target nucleic acid sequence encoding A4V or G93A is interpreted to not require that the gNA specifically targets and binds to a region of a nucleic acid that comprises a A4V or G93A mutation. Instead, this claim language is broadly interpreted to include any gNA that binds to a target nucleic acid sequence that comprises an A4V or G93A mutation within the nucleic acid sequence but does not specifically need to bind to a region containing the A4V or G93A mutation. As the gNA of ‘426 binds to an exon of a SOD1 nucleic acid that comprises the A4V mutation in exon 1 or the G93A mutation in exon 4, this is considered to fall within the broadly interpreted gNA of claim 181.
Regarding claim 182, ‘426 claims wherein the CasX variant protein comprises one or more nuclear localization signals (NLS) (claim 176).
Regarding claim 183, CasX proteins are known to cause single or double stranded breaks.
Regarding claim 184, ‘426 claims the CasX composition further comprising a donor template nucleic acid, wherein the donor template comprises a nucleic acid comprising at least a portion of a wild-type SOD1 gene (claim 178). It is well understood that including a donor template will lead to homologous recombination, leading to insertion of the homologous wild type gene but that NHEJ can also occur, leading to a deletion.
Regarding claim 192, the A4V and G93A gene mutations are known to be associated with ALS, as evidenced by Kiskinis and Gaj (whole documents), respectively.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claims 169 and 176-179 provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claims 169-192 of copending Application No. 17/641,426 as applied to claim 169 above and further in view of World Intellectual Property Organization Patent Application No. 2018064371 (Doudna).
The teachings of ‘426 are as discussed above.
‘426 does not teach that the gNA can be a gRNA or sgRNA.
However, Doudna teaches a composition comprising a CasX polypeptide and a CasX guide RNA. Doudna teaches that their CasX nuclease can be encoded by a viral vector for delivery to a eukaryotic cell. Doudna teaches that the CasX protein can have protein domains from two separate CasX proteins, an N-terminal domain (amino acids 1-650 of SEQ ID NO: 2) from CasX2 derived from Planctomycetes and a C-terminal domain (nucleotides 664-986 of SEQ ID NO: 1) from CasX1 derived from Deltaproteobacter (paragraphs 0068-00107). Doudna teach that the guide RNA contains a “guide sequence” or “targeting sequence” that can be modified so that the CasX guide RNA can target a CasX protein to any desired sequence of any desired target nucleic acid, with the exception that the PAM sequence can be taken into account (paragraphs 147-150).
Doudna (paragraphs 0068-00107 and 00148) use their Cas nucleases in combination with a gRNA or sgRNA.
Therefore, it would have been obvious that the gNA used in the method of ‘426 could be a gRNA or sgRNA as Doudna has identified that CasX enzymes can be used with gRNAs or sgRNAs for gene editing.
Regarding claim 178-179, ‘426 claims the gNA comprises a scaffold sequence comprising the sequence of SEQ ID NO: 2238 which also comprises SEQ ID NO: 20 of the instant application (claim 181).
This is a provisional nonstatutory double patenting rejection.
Claims 169-174, 180-187, and 189-193 are provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claims 169-184 and 194-196 of copending Application No. 17/641,426 in view of Gaj et al. (Sci. Adv. 3: 1-10. 2017), as evidenced by Kiskinis et al. (Cell Stem Cell 14: 781–795. 2014) and Brasil et al. (Mol Neurobiol 55: 5269-5281. 2018).
As an initial matter, the method claims of ‘426 (claims 194-196) are dependent on the composition claim 169. Therefore, all limitations associated with the composition claims are considered to be imported into the method claims, as well.
Regarding claims 169, 173-174, 186-187, 189-191, and 193, ‘426 claims a method of treating a SOD1-related disease in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of the vector of claim 184 (A vector comprising the nucleic acid of claim 183 (A nucleic acid comprising a sequence that encodes the CasX variant protein and/or the gNA of claim 169 (a composition comprising: a. a CasX variant protein comprising the sequence of SEQ ID NO: 126, or a sequence having at least about 70% sequence identity thereto; and b. a first guide nucleic acid (gNA), wherein the gNA comprises a targeting sequence complementary to a superoxide dismutase 1 (SOD1) gene target nucleic acid sequence, wherein the SOD1 gene comprises one or more mutations)), wherein the vector is selected from the group consisting of a retroviral vector, a lentiviral vector, an adenoviral vector, an adeno-associated viral (AAV) vector, a herpes simplex virus (HSV) vector, a virus-like particle (VLP), a plasmid, a minicircle, a nanoplasmid, a DNA vector, and an RNA vector)), wherein the subject is selected from the group consisting of a rodent, mouse, rat, non-human primate and human, and wherein the method results in improvement in at least one clinically-relevant endpoint selected from the group consisting of Amyotrophic Lateral Sclerosis Functional Rating Scale, combined assessment of function and survival, time to death, time to tracheostomy or persistent assisted ventilation, forced vital capacity, manual muscle test, maximum voluntary isometric contraction, duration of response, progression-free survival, time to progression of disease, and time-to-treatment failure (claims 194-196).
‘426 claims that the CasX protein is a chimeric protein comprising protein domains from two or more different CasX proteins (claim 180).
‘426 claims that the SOD1 gene can include one or more mutations selected from a group of a multitude of mutations, including A4V and G93A (claim 172).
‘426 does not explicitly state that they modified PNS or CNS cells as part of this method.
However, Gaj teaches a method of gene editing G93A-SOD1 mutant mice. They used Cas9 nuclease from Staphylococcus aureus (SaCas9) to target the mutant SOD1 gene in the G93A-SOD1 mouse model of ALS (5), which carries ~25 tandem repeat copies of the hSOD1G93A transgene and recapitulates many aspects of the disease, including progressive muscle atrophy and impaired motor function. Gaj injected AAV vectors encoding the Cas9 and a sgRNA targeting exon 2 of the SOD1 gene. Gaj found the presence of indels in ~94% of hSOD1G93A transgenes analyzed from these cells (fig. S2B), confirming that mutant SOD1 expression was disrupted by SaCas9-mediated genome editing, including in PNS cells of the spinal cord. Furthermore, the treated mice exhibited delayed disease onset and increased survival time (page 1, column 2, paragraph 2-page 4, column 2, paragraph 1, page 6, column 1, paragraph 1-page 7, column 2, paragraph 2, and Figures 1-3).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of treating SOD1 related disease of ’426 by targeting motor neurons, as identified by Gaj, to arrive at the instantly claimed invention. One of ordinary skill in the art would have a reason to modify with a reasonable expectation of success because Qi and Gaj discuss treating SOD1 related ALS by delivering AAV vectors encoding Cas nucleases and gNAs for gene editing of the mutant target gene and Gaj successfully reduces to practice that a Cas nuclease and sgRNA targeting exon 2 of SOD1 can successfully delay disease onset of ALS from the G93A gene mutation and increase survival time through successful gene editing, including in PNS motor neuron cells. Furthermore, it is well known that ALS mutations, such as the A4V and G93A mutations affect motor neuron cells. Therefore, it would have been obvious to transduce motor neuron cells with the vector of ‘426 to treat ALS. Because the prior art teaches all of the elements of the claimed invention, there is a reasonable expectation of success.
Regarding claims 170-171, the A4V mutation is in exon 1, as evidenced by Figure 2 of Kiskinis and the G93A mutation is in exon 4, as identified by Figure 1 of Gaj and are substitutions.
Regarding claim 172, Brasil evidences that the A4V and G93A gene mutations are considered gain of function mutations (page 5278, column 2, paragraph 2)
Regarding claim 180, ‘426 claims wherein the targeting sequence of the gNA is complementary to a sequence of a SOD1 exon (claim 170).
Regarding claim 181, as an initial matter, the term language is complementary to a target nucleic acid sequence encoding A4V or G93A is interpreted to not require that the gNA specifically targets and binds to a region of a nucleic acid that comprises a A4V or G93A mutation. Instead, this claim language is broadly interpreted to include any gNA that binds to a target nucleic acid sequence that comprises an A4V or G93A mutation within the nucleic acid sequence but does not specifically need to bind to a region containing the A4V or G93A mutation. As the gNA of ‘426 binds to an exon of a SOD1 nucleic acid that comprises the A4V mutation in exon 1 or the G93A mutation in exon 4, this is considered to fall within the broadly interpreted gNA of claim 181.
Regarding claim 182, ‘426 claims wherein the CasX variant protein comprises one or more nuclear localization signals (NLS) (claim 176).
Regarding claim 183, CasX proteins are known to cause single or double stranded breaks.
Regarding claim 184, ‘426 claims the CasX composition further comprising a donor template nucleic acid, wherein the donor template comprises a nucleic acid comprising at least a portion of a wild-type SOD1 gene (claim 178). It is well understood that including a donor template will lead to homologous recombination, leading to insertion of the homologous wild type gene but that NHEJ can also occur, leading to a deletion.
Regarding claim 185, Regarding claim 185, as can be seen in Gaj, the insertions/deletions led to reduced expression of the hSOD1 gene in the mice. As such, a reduction would also occur in the combined method of ‘426 and Gaj.
Regarding claim 189, ‘426 is silent as to the method of administering to the subject a therapeutically effective amount of the vector.
However, Gaj teaches that they used intravenous administration of the AAV vector encoding the Cas and sgRNA to treat their mice and this method successfully targeted motor neurons and treated the mice (Figures 1-2).
Therefore, it would have been obvious that one could administer the vectors intravenously as Gaj has already shown that this can work.
Regarding claim 192, the A4V and G93A gene mutations are known to be associated with ALS, as evidenced by Kiskinis and identified by Gaj (whole documents), respectively.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claims 169 and 176-179 provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claims 169-184 and 194-196 of copending Application No. 17/641,426and Gaj et al. (Sci. Adv. 3: 1-10. 2017), as applied to claim 169 above and further in view of World Intellectual Property Organization Patent Application No. 2018064371 (Doudna).
The teachings of ‘426 and Gaj are as discussed above.
‘426 and Gaj do not teach that the gNA can be a gRNA or sgRNA for use with the CasX enzyme.
However, Doudna teaches a composition comprising a CasX polypeptide and a CasX guide RNA. Doudna teaches that their CasX nuclease can be encoded by a viral vector for delivery to a eukaryotic cell. Doudna teaches that the CasX protein can have protein domains from two separate CasX proteins, an N-terminal domain (amino acids 1-650 of SEQ ID NO: 2) from CasX2 derived from Planctomycetes and a C-terminal domain (nucleotides 664-986 of SEQ ID NO: 1) from CasX1 derived from Deltaproteobacter (paragraphs 0068-00107). Doudna teach that the guide RNA contains a “guide sequence” or “targeting sequence” that can be modified so that the CasX guide RNA can target a CasX protein to any desired sequence of any desired target nucleic acid, with the exception that the PAM sequence can be taken into account (paragraphs 147-150).
Doudna (paragraphs 0068-00107 and 00148) use their Cas nucleases in combination with a gRNA or sgRNA.
Therefore, it would have been obvious that the gNA used in the method of ‘426 and Gaj could be a gRNA or sgRNA as Doudna has identified that CasX enzymes can be used with gRNAs or sgRNAs for gene editing.
Regarding claim 178-179, ‘426 claims the gNA comprises a scaffold sequence comprising the sequence of SEQ ID NO: 2238 which also comprises SEQ ID NO: 20 of the instant application (claim 181).
This is a provisional nonstatutory double patenting rejection.
Conclusion
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/KEENAN A BATES/Examiner, Art Unit 1631