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 .
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 247-250 and 261 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 247 is directed to the modified RNA of claim 242, further comprising (iii) one or more copies of a structural sequence comprising at least two nucleotides that are “capable of forming a secondary structure”, wherein the one or more copies of the structural sequence are 3' to the poly-A region, and wherein the modified RNA comprises a secondary structure, wherein the secondary structure comprises one or more copies of the structural sequence.
However, the specification does not adequately describe the structure required for the at least two nucleotides to be “capable of forming a secondary structure”. Without further description of the structure required for the function, one would not be able to readily envision which types of nucleotides are necessarily included or excluded from the genus of nucleotides that are capable of forming a secondary structure.
Claim 250 is directed to the modified RNA of claim 247, wherein the secondary structure of the RNA is an aptamer that is “capable of binding to a target molecule”. The specification does not adequately describe the structure required for the aptamer to be capable of binding at any level to any target molecule. Without further description of the structure required for the function, one would not be able to readily envision which aptamers with which specific structures are necessarily included or excluded from the genus of aptamers that are “capable of binding to a target molecule”.
Claim 261 is directed to the method of claim 256, wherein the structural sequence is an aptamer sequence comprising at least two nucleotides that are “capable of interacting to form an aptamer”, wherein the aptamer is a secondary structure that is “capable of binding to a target molecule”. The specification does not adequately describe the structure required for the at least two nucleotides that are “capable of interacting to form an aptamer”; and does not adequately describe the aptamer structure that is capable of binding at any level to any target molecule. Without further description of the structure required for the function, one would not be able to readily envision which nucleotides are capable of interacting to form an aptamer. Claim 261 requires for as little as two nucleotides to interact in any manner and form an aptamer, a genus that has not been adequately described in the specification.
Response to Arguments
Applicant argues that the specification describes in detail the structural sequences that are "capable of forming a secondary structure," including RNA G-quadruplex sequences (e.g., SEQ ID NO: 2), DNA G- quadruplex sequences (e.g., SEQ ID NO: 3), telomeric repeat sequences (e.g., SEQ ID NO: 4), and aptamers. These species are not representative of the entire claimed genus and are not a closed definition. Without further description of the structure required for the function, one would not be able to readily envision the structure required for the two nucleotides to be capable of forming any possible secondary structure. Applicant argues that the specification describes aptamers that are "capable of binding to a target molecule" in paragraphs [0024]-[0026] and [0301]-[0304], including specific examples of aptamers and their targets. Again, the species are not representative of the entire claimed genus. The aptamer is recited as being capable in any manner to bind to any target molecule, but does not require any specific structural requirement in comparison to the target.
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.
Claim(s) 242, 243, and 245-262 is/are rejected under 35 U.S.C. 103 as being unpatentable over Strzelecka et al. (RNA, 2020, 26:1815–1837), in view of Bolen et al. (WO 2015/048744 A2), and Petkovic et al. (Nucleic Acids Research, 2015, Vol. 43, No. 4, 2454-2465),
Strzelecka et al. teach: Chemical modifications enable preparation of mRNAs with augmented stability and translational activity. In this study, we explored how chemical modifications of 5′,3′-phosphodiester bonds in the mRNA body and poly(A) tail influence the biological properties of eukaryotic mRNA. To obtain modified and unmodified in vitro transcribed mRNAs, we used ATP and ATP analogs modified at the α-phosphate (containing either O-to-S or O-to-BH3 substitutions) and three different RNA polymerases—SP6, T7, and poly(A) polymerase. To verify the efficiency of incorporation of ATP analogs in the presence of ATP, we developed a liquid chromatography–tandem mass spectrometry (LC–MS/MS) method for quantitative assessment of modification frequency based on exhaustive degradation of the transcripts to 5′-mononucleotides. The method also estimated the average poly(A) tail lengths, thereby providing a versatile tool for establishing a structure-biological property relationship for mRNA. We found that mRNAs containing phosphorothioate groups within the poly(A) tail were substantially less susceptible to degradation by 3′-deadenylase than unmodified mRNA and were efficiently expressed in cultured cells, which makes them useful research tools and potential candidates for future development of mRNA-based therapeutics (Abstract).
Strzelecka et al. teach that the poly(A) tail/region is 3’ to the ORF (Table 1) (instant claims 242 and 245).
Strzelecka et al. teach: We investigated various properties of mRNAs with modified poly(A) tails, including poly(A) tail length, composition, susceptibility to deadenylation, and protein expression efficiency in mammalian cells. The DNA templates for in vitro transcription used in these experiments encoded a short 5′ UTR sequence, followed by the Gaussia luciferase ORF and either a single or double H. sapiens β-globin 3′ UTR sequence (RNA H and RNA G, respectively), and in the case of the reference mRNA (RNA I), an A128 poly(A) tail was encoded (page 1823) (instant claims 246, 247, 251, and 257). The RNA being non-coding would be a matter of design choice depending on the intended use. The nucleotides are considered to meet the instant limitation of claim 247 of being capable of forming secondary structure. Therefore, the ORF encodes a protein (instant claims 242 and 245).
Strzelecka et al. teach estimation of polyA tail lengths that are more than 25 nucleotides (Table S4) (instant claims 242 and 243).
Therefore, Strzelecka et al. teach that phosphorothioate groups within the poly(A) tail of mRNA comprising an ORF encoding a protein, Gaussia luciferase, wherein the poly(A) tail is 3’ to the ORF with the result of substantially less susceptibility to degradation by 3′-deadenylase than unmodified mRNA (instant claim 242). In Figure 6A, Strzelecka et al. teach enzymatically adding the poly(A) region to the 3’ end of the mRNA using poly(A) polymerase (5’ cap, 5’-UTR, ORF, 3’-UTR, polyA 3’) and depict two phosphorothioates in the Figure at the end (Figure 1A) (instant claims 253 and 257).
Although Strzelecka et al. do not specifically teach that 3 or more or 4 or more of the last 25 nucleotides of the poly-A region are modified and wherein at least 5% of the nucleotides of the poly-A region are modified, Strzelecka et al.: Further increases in the phosphorothioate content in the poly(A) tail to 13%, 15%, and 23% corresponded to a deadenylation rate reduced by 4.4-fold, 6.7-fold, and almost 20-fold, respectively (Supplemental Table S3) (pages 1824-1825) (instant claims 242 and 244).
Since Strzelecka et al. teach that increasing phosphorothioate content progressively decreases susceptibility to 3’ deadenylation, it would have been obvious as a matter of design choice to provide at least 3 or at least 4 phosphorothioate linkages at the 3’ end instead of the 2 depicted and a total of at least 5% in the poly(A) region or up to full modification (instant claims 242, 243, and 262).
The benefit of increasing the percentage of phosphorothioate modifications in the poly(A) tail of an mRNA was known, as taught by Strzelecka et al. therefore, increasing the percentage would have been obvious with the expectation of a progressive decrease in susceptibility to 3’ deadenylation as taught by Strzelecka et al.
Strzelecka et al. demonstrate that increasing phosphorothioate content in the poly(A) tail decreased CNOT7-mediated deadenylation. It is noted that PAP (poly(A) polymerase utilized by Strzelecka et al.) builds the tail at the 3’ terminus. Therefore, the phosphorothioate containing segment is part of the newly created 3’-terminal poly(A) region. At the percentages of phosphorothioate incorporation taught by Strzelecka et al., there would be 5.75 phosphorothioates at 23% of a 25 nucleotide region of the poly(A) tail.
With regards to the method of making, Strzelecka et al. teach: To study the effects of poly(A) tail modification on mRNA stability and translation, we prepared unmodified mRNAs (RNA G and RNA H) and enzymatically added a poly(A) tail onto the 3′ end using PAP and phosphate-modified ATP analogs (Fig. 6A) (page 1823).
Strzelecka et al. teach: The 3′ ends of Gaussia luciferase encoding transcripts were tailed with a poly(A) sequence using an enzymatic approach with E. coli PAP(Lucigen) (page 1833) (instant claims 252 and 256). Additionally, Bolen et al.t each 4 RNA ligase may enzymatically link nucleic acids.
Strzelecka et al. do not teach incorporation of DNA G-quadruplex sequences, as required by instant claims 249 and 260. It is noted that the claims do not require any specific sequence as the sequences are recited as optional.
It would have been obvious to incorporate one or more DNA G-quadruplex sequences into the mRNA of Strzelecka et al. because Bolen et al. teach: [000377] In one embodiment, the polynucleotides of the present invention are designed to include a polyA-G Quartet region. The G-quartet is a cyclic hydrogen bonded array of four guanine nucleotides that can be formed by G-rich sequences in both DNA and RNA. In this embodiment, the G-quartet is incorporated at the end of the poly-A tail. The resultant polynucleotide is assayed for stability, protein production and other parameters including half- life at various time points. It has been discovered that the polyA-G quartet results in protein production from an mRNA equivalent to at least 75% of that seen using a poly-A tail of 120 nucleotides alone (SEQ ID NO: 1646). One would reasonably expect the benefits taught by Bolen et al. when incorporated into the mRNA of Strzelecka et al.
Additionally, it was known in the art that mRNA can be designed in a circular form, as evidenced by Bolen et al. (figure 6). It would have been obvious to design the mRNA of Strzelecka et al. to be circular as a matter of design choice (instant claims 248, 254, 258, and 259).
Bolen et al. teach: [0001147] According to the present invention, a polynucleotide may be cyclized, or concatemerized, to generate a translation competent molecule to assist interactions between poly- A binding proteins and 5 '-end binding proteins. The mechanism of cyclization or concatemerization may occur through at least 3 different routes: 1) chemical, 2) enzymatic, and 3) ribozyme catalyzed. The newly formed 5 '-/3 '-linkage may be intramolecular or intermolecular (instant claims 254 and 255).
Bolen et al. teach: [000180] According to the present invention, the polynucleotides may be administered with, conjugated to or further encode one or more of RNAi agents, siRNAs, shRNAs, miRNAs, miRNA binding sites, antisense RNAs, ribozymes, catalytic DNA, tRNA, RNAs that induce triple helix formation, aptamers or vectors, and the like. Therefore, it would have been obvious to conjugate the mRNA to an aptamer as a matter of design choice (instant claims 250 and 261).
With regards to instant claim 255, Petkovic et al. teach enzymatic circularization requiring phosphorylation of the 5’ terminus followed by the circularization reaction. Petkovic et al. teaches cleavage the generates a 3’-OH. The 5’ phosphate and 3’-OH ends participate in circularization. This process renders instant claim 255 obvious and would have been an obvious method for circularization of the modified mRNA.
The claims are obvious combinations of known features for mRNA design and production.
Response to Arguments
Applicant argues that Strzelecka does not disclose 3 or more consecutive modified nucleotides at any specific position in the poly(A) tail. Strzelecka uses E. coli poly(A) polymerase ("PAP") to incorporate ATPaS (a phosphorothioate ATP analog) randomly throughout the poly(A) tail in the presence of a mixture of ATP and ATPaS. See Strzelecka at pages 1820-1823. Since PAP incorporates modified and unmodified nucleotides stochastically, the resulting poly(A) tails contain phosphorothioate modifications at random, non-predetermined positions, with modification frequencies of 6-23%. See Strzelecka at pages 1824-1825 and Supplemental Table S3. At these modification frequencies, the probability that any three adjacent nucleotide positions will all contain phosphorothioate modifications is exceedingly low (e.g., at 23% modification, the probability of 3 consecutive modified positions is approximately 0.233 1.2%).
Contrary to applicant’s argument, this is a rejection under 35 USC 102 rather than 103 and therefore the instant claims are obvious in view of the cited references rather than anticipated.
Strzelecka et al.: Further increases in the phosphorothioate content in the poly(A) tail to 13%, 15%, and 23% corresponded to a deadenylation rate reduced by 4.4-fold, 6.7-fold, and almost 20-fold, respectively (Supplemental Table S3) (pages 1824-1825). Since Strzelecka et al. teach that increasing phosphorothioate content progressively decreases susceptibility to 3’ deadenylation, it would have been obvious as a matter of design choice to provide at least 3 or at least 4 phosphorothioate linkages at the 3’ end instead of the 2 depicted and a total of at least 5% in the poly(A) region or up to full modification.
Applicant argues that Bolen does not remedy the deficiencies of Strzelecka. Bolen teaches general polynucleotide designs, including poly-A G-quartet regions and circularization of polynucleotides. See Bolen at paragraphs [000377] and [001147]. However, Bolen does not teach or suggest 3 or more consecutive modified nucleotides at the 3' end of the poly-A region, nor does Bolen teach modified nucleotides as recited in independent claim 242. Bolen et al. was not relied upon for teaching 3 or more consecutive modified nucleotides at the 3' end of the poly-A region, but rather is relied upon for the teachings set forth in the rejection directed to general polynucleotide design that is recited in the instant claims.
Applicant argues that the present specification demonstrates unexpected and surprising results for the claimed modifications. Specifically, Example 2 (para. [0381]) shows that inclusion of only 5% modified ATP increased expression of reporter proteins encoded by mRNAs including a modified poly-A region, as claimed. Moreover, Example 4 of the specification (paragraph [0393]) demonstrates that ligation of oligonucleotides containing 3 sequential phosphorothioate linkages (3xSrA_ddC, 3xSrA_InvdT, and 3xSrG_InvdT) showed 140%-210% increased GFP production compared with the control oligonucleotide at each timepoint. Example 5 (paragraph [0469]) demonstrates that DNA G-quadruplex sequences at the 3' terminus significantly enhanced protein translation (150%-170%). Example 6 (paragraphs [0562]-[0564]) further demonstrates that mocRNA containing nuclease-resistant groups at the 3' terminus increases expression of a luciferase reporter in both cell culture and in vivo.
The results argued by applicant are not commensurate in scope with the instant claims. The poly-A region of the instant claims is of any length and comprises at least 3 or more consecutive modifications at any location within the last 25 nucleotides wherein the entire length of the poly-A region has at least 5% modification and ranges to full modification of any combination of deoxyribonucleotides, 2’-modified nucleotides (any possible 2’-modification), and phosphorothioates. Applicant has not demonstrated any unexpected result commensurate in scope with the claims. Additionally, one would have expected that increasing phosphorothioate content would progressively decreases susceptibility to 3’ deadenylation, as specifically taught by Strzelecka et al.
Conclusion
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). 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 Amy R Hudson whose telephone number is (571)272-0755. The examiner can normally be reached M-F 8:00am-6:00pm.
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/AMY ROSE HUDSON/Primary Examiner, Art Unit 1636
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