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 .
Terminal Disclaimer
The terminal disclaimer filed on November 10, 2025 disclaiming the terminal portion of any patent granted on this application which would extend beyond the expiration date of any patent granted on Application Number 17/617,039 has been reviewed and is accepted. The terminal disclaimer has been recorded.
Application Status
This action is written in response to applicant' s correspondence received November 10, 2025. Claims 1-8 are currently pending.
Any rejection or objection not reiterated herein has been overcome by amendment. Applicant' s amendments and arguments have been thoroughly reviewed, but are not persuasive to place the claims in condition for allowance for the reasons that follow.
Claim Rejections - 35 USC § 103
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
Claims 1-8 are rejected under 35 U.S.C. 103 as being unpatentable over Hou Z., et. al., Pioneer Hi-Bred International, Inc., US 2019/0264232 A1, published Aug 29, 2019.
Regarding Claim 1, Hou teaches a method in Example 8 for “In Vivo Modification of a Human Cell Target Polynucleotide with Cas9 Ortholog Nucleases” (p. 56, Col. 2) for causing a double strand break in DNA in a cell. Hou teaches, “In some aspects, the Cas endonuclease is capable of creating a sticky-end overhang double strand break. In some aspects, the Cas endonuclease is capable of creating a blunt-end double strand break” (p. 4, [0011]), which reads on the use of Cas endonucleases to create double strand breaks. Merely to elaborate, Hou further teaches that native Cas9 cause double strand breaks: “Thus, a wild type Cas protein ( e.g., a Cas protein disclosed herein), or a variant thereof retaining some or all activity in each endonuclease domain of the Cas protein, is a suitable example of a Cas endonuclease that can cleave both strands of a DNA target sequence” (p. 23, [0238]). Hou recites, “In some aspects, the Cas endonuclease may not successfully recognize a target DNA sequence if the target DNA sequence is not adjacent to, or near, a PAM sequence. In some aspects, the PAM precedes the target sequence” (p. 15, [0155]), which reads as targeting a location preceding a PAM sequence in a DNA molecule. In Example 8, Hou teaches, “To validate the functional activity of novel Cas9 nucleases in human cells, RNP complexes are assembled using purified proteins and in vitro transcribed sgRNAs. RNPs are introduced into HEK293 cells by electroporation.” (p. 57, [0644]), which reads as introducing into a cell a Cas9 endonuclease. Together, Hou teaches a method of causing a double strand break in a DNA molecule immediately before a PAM sequence using a Cas9 endonuclease.
Hou teaches that “The Cas9 orthologs described herein further comprise endonuclease activity (p. 18, [0191])” and “A Cas9 ortholog protein is further defined as … native, full-length Cas9 ortholog protein of any of SEQ ID NO:86-170 and 511-1135. (p. 18, [0192])”. SEQ ID NO. 1047, taught by Hou, is a 100% identity match to SEQ ID NO. 1 from the instant application; an alignment is below.
Qy 1 MQNNPLNYILGLDLGIASIGWAVVEIDEESSPIRLIDVGVRTFERAEVAKTGESLALSRR 60
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1 MQNNPLNYILGLDLGIASIGWAVVEIDEESSPIRLIDVGVRTFERAEVAKTGESLALSRR 60
Qy 61 LARSSRRLIKRRAERLKKAKRLLKAEKILHSIDEKLPINVWQLRVKGLKEKLERQEWAAV 120
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 61 LARSSRRLIKRRAERLKKAKRLLKAEKILHSIDEKLPINVWQLRVKGLKEKLERQEWAAV 120
Qy 121 LLHLSKHRGYLSQRKNEGKSDNKELGALLSGIASNHQMLQSSEYRTPAEIAVKKFQVEEG 180
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 121 LLHLSKHRGYLSQRKNEGKSDNKELGALLSGIASNHQMLQSSEYRTPAEIAVKKFQVEEG 180
Qy 181 HIRNQRGSYTHTFSRLDLLAEMELLFQRQAELGNSYTSTTLLENLTALLMWQKPALAGDA 240
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 181 HIRNQRGSYTHTFSRLDLLAEMELLFQRQAELGNSYTSTTLLENLTALLMWQKPALAGDA 240
Qy 241 ILKMLGKCTFEPSEYKAAKNSYSAERFVWLTKLNNLRILENGTERALNDNERFALLEQPY 300
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 241 ILKMLGKCTFEPSEYKAAKNSYSAERFVWLTKLNNLRILENGTERALNDNERFALLEQPY 300
Qy 301 EKSKLTYAQVRAMLALSDNAIFKGVRYLGEDKKTVESKTTLIEMKFYHQIRKTLGSAELK 360
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 301 EKSKLTYAQVRAMLALSDNAIFKGVRYLGEDKKTVESKTTLIEMKFYHQIRKTLGSAELK 360
Qy 361 KEWNELKGNSDLLDEIGTAFSLYKTDDDICRYLEGKLPERVLNALLENLNFDKFIQLSLK 420
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 361 KEWNELKGNSDLLDEIGTAFSLYKTDDDICRYLEGKLPERVLNALLENLNFDKFIQLSLK 420
Qy 421 ALHQILPLMLQGQRYDEAVSAIYGDHYGKKSTETTRLLPTIPADEIRNPVVLRTLTQARK 480
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 421 ALHQILPLMLQGQRYDEAVSAIYGDHYGKKSTETTRLLPTIPADEIRNPVVLRTLTQARK 480
Qy 481 VINAVVRLYGSPARIHIETAREVGKSYQDRKKLEKQQEDNRKQRESAVKKFKEMFPHFVG 540
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 481 VINAVVRLYGSPARIHIETAREVGKSYQDRKKLEKQQEDNRKQRESAVKKFKEMFPHFVG 540
Qy 541 EPKGKDILKMRLYELQQAKCLYSGKSLELHRLLEKGYVEVDHALPFSRTWDDSFNNKVLV 600
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 541 EPKGKDILKMRLYELQQAKCLYSGKSLELHRLLEKGYVEVDHALPFSRTWDDSFNNKVLV 600
Qy 601 LANENQNKGNLTPYEWLDGKNNSERWQHFVVRVQTSGFSYAKKQRILNHKLDEKGFIERN 660
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 601 LANENQNKGNLTPYEWLDGKNNSERWQHFVVRVQTSGFSYAKKQRILNHKLDEKGFIERN 660
Qy 661 LNDTRYVARFLCNFIADNMLLVGKGKRNVFASNGQITALLRHRWGLQKVREQNDRHHALD 720
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 661 LNDTRYVARFLCNFIADNMLLVGKGKRNVFASNGQITALLRHRWGLQKVREQNDRHHALD 720
Qy 721 AVVVACSTVAMQQKITRFVRYNEGNVFSGERIDRETGEIIPLHFPSPWAFFKENVEIRIF 780
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 721 AVVVACSTVAMQQKITRFVRYNEGNVFSGERIDRETGEIIPLHFPSPWAFFKENVEIRIF 780
Qy 781 SENPKLELENRLPDYPQYNHEWVQPLFVSRMPTRKMTGQGHMETVKSAKRLNEGLSVLKV 840
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 781 SENPKLELENRLPDYPQYNHEWVQPLFVSRMPTRKMTGQGHMETVKSAKRLNEGLSVLKV 840
Qy 841 PLTQLKLSDLERMVNRDREIALYESLKARLEQFGNDPAKAFAEPFYKKGGALVKAVRLEQ 900
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 841 PLTQLKLSDLERMVNRDREIALYESLKARLEQFGNDPAKAFAEPFYKKGGALVKAVRLEQ 900
Qy 901 TQKSGVLVRDGNGVADNASMVRVDVFTKGGKYFLVPIYTWQVAKGILPNRAATQGKDEND 960
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 901 TQKSGVLVRDGNGVADNASMVRVDVFTKGGKYFLVPIYTWQVAKGILPNRAATQGKDEND 960
Qy 961 WDIMDEMATFQFSLCQNDLIKLVTKKKTIFGYFNGLNRATSNINIKEHDLDKSKGKLGIY 1020
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 961 WDIMDEMATFQFSLCQNDLIKLVTKKKTIFGYFNGLNRATSNINIKEHDLDKSKGKLGIY 1020
Qy 1021 LEVGVKLAISLEKYQVDELGKNIRPCRPTKRQHVR 1055
|||||||||||||||||||||||||||||||||||
Db 1021 LEVGVKLAISLEKYQVDELGKNIRPCRPTKRQHVR 1055
Hou does not teach a method to use SEQ ID NO. 1 of the instant application to cause a double-strand break immediately before the sequence 5’-NNNN(A/G)TT-3’, the PAM sequence.
Hou does teach a method to determine the PAM sequences from Cas9 endonucleases in Example 2, titled “Determination of the Protospacer Adjacent Motif Requirement and Target Cleavage Pattern for Cas9 Orthologs” (p. 51, Col. 1), which would identify the PAM sequence 5’-NNNN(A/G)TT-3’. Hou teaches, “In this example, methods for the rapid characterization of the protospacer adjacent motif (PAM) requirement and the position and type … of double-stranded DNA target cleavage for orthologous Cas9 proteins are described” (p. 51, [0582]). Hou teaches that the method found “a diverse range of PAM sequence preferences were obtained” (p. 52, [0593]) providing predictability to the method. Hou also provides motivation for using the method: “Most CRISPR gene editing is based almost entirely the Cas9 system derived from Streptococcus pyogenes …, which leaves a blunt-end overhang and effects gene editing via the recognition of a Protospacer Adjacent Motif (PAM)sequence of "NGG" on the target polynucleotide. A greater diversity of Cas9 proteins with different biophysical and biochemical characteristics, including different PAM recognition sequences, is desirable.” (p. 1, [0006]).
Regarding Claim 1, it would be obvious to try the method of Hou for determining Cas9 PAM sequences with SEQ ID NO. 1, taught by Hou as SEQ ID NO. 1047, find the PAM sequence of 5’-NNNN(A/G)TT-3’ and to use the method provided by Hou of causing a double strand break in DNA immediately before 5’-NNNN(A/G)TT-3’ by introducing into a cell a SEQ ID NO. 1 because Hou teaches that there is a recognized need in the art for Cas9 proteins with specific and diverse biochemistry. To target 5’-NNNN(A/G)TT-3’ , Hou provides a finite number of identified Cas9 proteins, which when used with the method of determining Cas9 PAM sequences, would produce the consensus 5’-NNNN(A/G)TT-3’ when applied to SEQ ID NO. 1. One of ordinary skill could have pursued using Hou’s methods if they needed to cause a double strand break in a DNA molecule in a cell in a specific sequence and find that SEQ ID NO. 1 causes a break 5’-NNNN(A/G)TT-3’ with a reasonable expectation of success because Hou was successfully able to use the method to identify many PAM sequences across a diverse set of Cas9 orthologs and use those orthologs to cause double strand breaks in DNA in cells. Therefore, Claim 1 is obvious over Hou.
Regarding Claim 2, Hou teaches, “Electroporated cells were incubated at 37° C.” (p. 57, [0645]) of the Cas9 ortholog, which is between 35° C and 45° C. Therefore, Claim 2 is obvious over by Hou.
Regarding Claim 3, Hou teaches SEQ ID NO. 1 of the instant application as SEQ ID NO. 1047. Therefore, Claim 3 is obvious over Hou.
Regarding Claim 4, Hou teaches the use of Cas9 orthologs in HEK293 cells, human embryonic kidney cells, which are mammalian cells (p. 57, [0645]). Hou also teaches, “Targeting endogenous genes allows evaluation of the activity of the selected Cas9s on chromosomal DNA” (p. 56, [0643]), which reads as genomic DNA of a mammalian cell. Therefore, Claim 4 is obvious over Hou.
Regarding Claim 5, Hou teaches the use of Cas 9 orthologs to cause a double strand break leading to a modification in the genomic DNA: “The genome editing activity of selected Cas9 proteins is measured” (p. 56, [0643]). Therefore, Claim 5 is obvious over Hou.
Regarding Claim 6, Hou teaches a method for modifying a genomic DNA sequence in a cell comprising DNA comprising introducing a nucleic acid encoding SEQ ID NO. 1 and a gRNA that forms a complex to target a region of the cell’s genomic DNA directly adjacent to a PAM sequence to form a double stranded break immediately adjacent to the nucleotide sequence by using Hou’s SEQ ID NO. 1047 and Example 8 (p. 85). Hou also teaches a method for determining the PAM sequence of a Cas9 ortholog (p. 51, Example 2). As described above for Claim 1, it would be obvious to try the method for determining the PAM sequence of a Cas9 ortholog taught by Hou with the Cas9 sequences taught by Hou because there is a need in the art for diversity in Cas9 biochemistry and target selection, Hou teaches a finite list of Cas9 sequences, and one of ordinary skill in the art would have a reasonable expectation of success at finding SEQ ID NO. 1’s PAM sequence of 5’-NNNN(A/G)TT-3’ because the method described by Hou was successful for a diverse set of Cas9 orthologs. Therefore, Claim 6 is obvious over Hou.
Regarding Claim 7, Hou teaches, in Example 7, a method of modifying a genomic DNA sequence in a cell of a multicellular organism by introducing plasmids containing “different optimized Cas9 endonuclease gene and nuclear localization signal variants” (p. 54, [0618]) with “the guide RNA … added to the 5' end of the sequence to be transcribed” (p. 54, [0619]) into maize embryos. Hou combined the Cas9/gRNA containing plasmids with exogenous DNA: “The plasmids and DNA of interest are precipitated onto … gold pellets using a water-soluble cationic lipid transfection reagent” and were transfected into maize embryos (p. 54, [0623]). SEQ ID NO. 1 is a possible Cas9 endonuclease gene, described in Hou as SEQ ID NO. 1047. This reads as a method of introducing a nucleic acid encoding SEQ ID NO. 1 of the instant application, a gRNA and an exogenous DNA sequence simultaneously to modify the genomic DNA sequence in a cell of a multicellular organism to cause a double strand break in DNA. As described above for Claim 6, it would be obvious to try the method of Hou to determine the PAM sequence of SEQ ID NO. 1 and use the methods of Hou to introduce a nucleic acid encoding SEQ ID NO. 1 into a cell to cause a double strand break immediately before 5’-NNNN(A/G)TT-3’. Therefore, Claim 7 is obvious over Hou.
Regarding Claim 8, Hou teaches the use of Cas9 orthologs in HEK293 cells, human embryonic kidney cells, which are mammalian cells (p. 57, [0646]). Therefore, Claim 8 is obvious over Hou.
Response to Arguments - 35 USC § 103
Applicant argues “Hou only discloses the sequence SEQ ID NO: 1047 as one of a very lengthily list of sequences (over 1000 sequences, but does not provide any specific teaching on the use of this sequence.” This argument is not persuasive because Hou anticipates SEQ ID NO: 1047 as a native full-length Cas9 ortholog (p. 18, [0192]) and teaches the use of native Cas9 orthologs as described in the rejection above.
Applicant further argues, “Hou discloses methods for introducing double strand breaks using proteins, but specifically for variants of its SEQ ID NO: 1125, and does not describe that SEQ ID NO: 1047 can be used for this purpose.” This argument is not persuasive because Hou teaches SEQ ID NO: 1047 as a wild-type Cas9 (p. 18, [0192]) and wild-type Cas9 can be used to create double strand breaks (p. 23, [0238]) as described in the rejection above.
Applicant further argues, “Claim 1 of the present application specifically relates to the PAM sequence 5’-NNNN(A/G)TT-3’ which is a functional feature of claim 1 … Therefore, the claimed method relates to a protein that is combined with a respective DNA nucleotide sequence, generating a specific double strand break. However, such a method is not disclosed in Hou.” This argument is not persuasive because Hou teaches that a method using a PAM sequence for a Cas9 to recognize a DNA nucleotide site for double strand cleavage and a method for determining the PAM sequence of a Cas9 as described in the rejection above. One skilled in the art would recognize Hou teaches SEQ ID NO: 1 of the instant application as a Cas9 that may be used to cause double strand breaks, and it would be obvious to try the method of Hou to determine PAM sequences in order to use the Cas9 sequence.
Applicant further argues, “due to the significant amount of sequences disclosed in Hou (over 1000 sequences, where SEQ ID NO: 1047 … is but one of these hundreds of sequences, the skilled person would not readily appreciate, without extensive trial and error, that SEQ ID NO: 1047 would be adapted to preformed specific types of double-stranded breaks, breaks following the PAM nucleotide sequence 5'- NNNN(A/G)TT-3'. This is because Hou never discloses that SEQ ID NO: 1047 is capable of
performing double-stranded breaks, nor of interacting with the PAM nucleotide sequence 5'-NNNN(A/G)TT-3'. The skilled person would have to test all of these sequences disclosed in Hou, with different combinations of PAM sequences, to ascertain that SEQ ID NO: 1047 is capable of performing double-strand breaks, and more particularly, perform double-stranded breaks related to the PAM nucleotide sequence 5'-NNNN(A/G)TT-3.” This argument is not persuasive because Hou teaches SEQ ID NO: 1047, a wild-type Cas9, causes double strand breaks before a PAM sequence, Hou teaches the PAM sequence of 80 Cas9 orthologs, and Hou teaches a method to screen Cas9 proteins for their PAM sequence, which would easily traverse the finite number of solutions required. MPEP 2143.I.E recites in Example 5: “ ’a finite number’ of predictable solutions when an obvious to try rationale is applied: under the Federal Circuit’s case law "finite" means "small or easily traversed" in the context of the art in question”. Hou teaches 710 Cas9 orthologs (p. 18, [0192]) and provides the PAM sequence for 80 sequences (p. 65-85, Tables 4-83), and therefore not all the Cas9 orthologs would need to be tested. For the remaining 630 Cas9 orthologs, one skilled in the art would easily traverse trying the method of Hou to determine the PAM sequence because Hou teaches a detailed method in Example 2 (p. 51), and the method is predictable as shown in the listed PAM sequences.
Conclusion
No claims are allowed.
THIS ACTION IS MADE FINAL. Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to Krishna Nuggehalli Ravindra whose telephone number is (571)272-2758. The examiner can normally be reached M-Th, alternate F, 8a-5p est.
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/K.N.R./Examiner, Art Unit 1636
/NEIL P HAMMELL/Supervisory Patent Examiner, Art Unit 1636