DETAILED ACTION
A request for continued examination under 37 CFR 1.114, including the fee set forth in 37 CFR 1.17(e), was filed in this application after final rejection. Applicant’s Request for Continued Examination, Amendment and Arguments/Remarks received on 19 August 2026 have been entered.
Claims 4-5, 7-8, 10, 12, 23, 25, 28, and 36-37 were previously pending in the application. Claims 4 and 37 have been cancelled, and new claims 38-43 have been added by Applicant. Claims 5, 7-8, 10, 12, 23, 25, 28, 36, and 38-43 are currently pending in the application. Claims 8 and 23 are independent claims.
The election of Group I, drawn to a T7 RNA polymerase (T7 RNAP) variant, a nucleic acid molecule encoding a T7 RNAP variant, and a method of synthesizing a 2’ modified polynucleotide using a T7 RNAP variant, remains in effect in the instant application.
The following election of species remains in effect in the instant application:
2) nucleic acid aptamer target of interest: protein.
Claims 5, 7-8, 10, 12, 23, 25, 28, 36, and 38-43 are currently pending and under examination in the instant application. An action on the merits follows.
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA . The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action.
Priority
The present application is a 35 U.S.C. 371 national stage filing of International Application No. PCT/US2019/050777, filed 12 September 2019, which claims priority to U.S. Provisional Application No. 62/730,226, filed 12 September 2018.
Thus, the earliest possible priority for the instant application is 12 September 2018.
37 CFR 1.121(c)
The objection to the claim amendment filed 05 February 2026 under 37 CFR 1.121(c) for Applicant’s claim listing not being in compliance with 37 CFR 1.121(c) is withdrawn in view of the filing of a replacement claim amendment on 19 August 2026.
Claim Objections
The objection to cancelled claim 37 for reciting “SELEX” without first writing out the term for which it is an abbreviation is withdrawn.
Amended claims 10 and 12 are newly objected to because of the following informalities: claim 10 has an extra space between “condition” and the period “.” at the end of the claim. Claim 12 has an extra space between “modified” and “polynucleotide” in line 2. Appropriate correction is required.
Claim Rejections - 35 USC § 112(b)
The rejection of amended, previously presented, and cancelled claims 4-5, 7-8, 10, 12, 23, 25, 28, and 36-37 under 35 U.S.C. 112(b) as failing to particularly point out and distinctly claim the subject matter which the inventor(s) regards as the invention for reciting “wherein the T7 RNA polymerase variant does not have a transcription bias to incorporate 2’-modified nucleotides to a polynucleotide” in amended claim 8 and “SELEX” in cancelled claim 37, is withdrawn in view of Applicant’s cancellation of claim 37 and amendment to claim 8 clarifying that the bias is referring to the relative incorporation of each of the 2’-O-methyl modified bases A, T, G or C.
Amended, previously presented, and new claims 5, 7-8, 10, 12, 23, 25, 28, 36, and 38-43 are newly rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention. Amended, previously presented, and new claims 5, 7-8, 10, 12, 23, 25, 28, and 38-43 are included in this rejection due to their dependence on and/or encompassing of claims 8, 10, 23, 36 and/or 38-40.
Amended independent claim 8 recites, “wherein the T7 RNA polymerase variant does not have a bias in incorporating 2’-O-methyl modified A, T, G, or C nucleotides into a polynucleotide” in lines 2-4, which is indefinite because RNA polymerases incorporate uridine nucleotides and not thymidine nucleotides. Therefore, it is unclear whether Applicant intends to recite a T7 RNA polymerase that incorporates 2’-O-methyl modified thymidine nucleotides at the same frequency as 2’-O-methyl modified adenosine, cytidine, and guanosine, or whether Applicant intended to recite a T7 RNA polymerase that incorporates 2’-O-methyl modified uridine nucleotides at the same frequency as 2’-O-methyl modified adenosine, cytidine, and guanosine. Applicant indicates Table 3 in Example 3 for support for equal incorporation of A, T, G, or C. However, Table 3 presents sequence data obtained after reverse transcription of the RNA product transcribed by the mutant T7 RNA polymerases, and as such is presenting the DNA counterparts to the RNA produced by the T7 RNA polymerase mutants. As such, the metes and bounds of the claim cannot be determined.
In the interest of compact prosecution, amended independent claim 8 is being interpreted to encompass wherein lack of bias is for incorporating 2’-O-methyl A, U, G, or C.
Amended claim 10 recites, “the identical reaction condition” in lines 3-4. There is insufficient antecedent basis for this limitation in the claim. Claim 10 is dependent on independent claim 8. Neither claims 10 nor claim 8 have any prior recitation of any reaction condition. As such, the metes and bounds of the claim cannot be determined.
Amended claim 23 contains the trademark/trade name “Systematic Evolution of Ligands by Exponential Enrichment (SELEX)” in lines 6-7. New claims 39 and 40 additionally each recite the trademark/trade name “SELEX” in lines 1 and 2, respectively. "The terms "trade name" and "commercial name" mean any name used by a person to identify his or her business or vocation" (as defined by 15 U.S.C. 1127, see MPEP 608.01 (v)). Where a trademark or trade name is used in a claim as a limitation to identify or describe a particular material or product, the claim does not comply with the requirements of 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph. See Ex parte Simpson, 218 USPQ 1020 (Bd. App. 1982). The claim scope is uncertain since the trademark or trade name cannot be used properly to identify any particular material or product. A trademark or trade name is used to identify a source of goods, and not the goods themselves. Thus, a trademark or trade name does not identify or describe the goods associated with the trademark or trade name. In the present case, the trademark/trade name is used to identify/describe a method for the in vitro evolution of nucleic acid molecules with highly specific binding to target molecules, and, accordingly, the identification/description is indefinite.
Previously presented claim 36 recites, “The T7 RNA polymerase variant of claim 8, wherein the T7 RNA polymerase variant has at least 2 times more modified transcription production as compared to a T7 RNA polymerase polypeptide having the mutations of: i. Y639L and H784A substitutions; or ii. K378R, Y639L, and H784A substitutions” in lines 1-5, which is indefinite because the T7 RNA variant of claim 8 itself comprises the mutations of K378R, Y639L, and H784A substitutions. Therefore, it is unclear how the T7 RNA polymerase variant can have at least 2 times the activity relative to a T7 RNA polymerase polypeptide which comprises mutations comprised by the T7 RNAP variant itself in that the comparison encompasses a comparison with itself. The T7 RNAP variant cannot have 2 times the activity level as itself. As such, the metes and bounds of the claim cannot be determined.
New claim 38 recites, “The T7 RNA polymerase variant of claim 8, wherein the polypeptide further comprises the P266L substitution”, which has multiple issues of indefiniteness.
Firstly, neither claim 38 nor claim 8 have any prior recitation of any P266L substitution, and as such “the P266L substitution” lacks antecedent basis.
Secondly, independent claim 8 recites “A T7 RNA polymerase variant comprising a polypeptide sequence of SEQ ID NO: 2”, and so the limitation “wherein the polypeptide further comprises the P266L substitution” is indefinite because it conflicts with the requirement of claim 8 that the T7 RNAP comprise the sequence of SEQ ID NO: 2 which comprises a Pro and does not comprise a Leu at position 266. To the extent that claim 38 intends for “wherein the polypeptide further comprises the P266L substitution” to indicate a sequence in addition to the sequence of SEQ ID NO: 2 which is comprised by the T7 RNAP as recited in claim 8, then it is indefinite in that the P266L substitution has no context for any additional sequence in which the substitution can be present.
As such, the metes and bounds of the claim cannot be determined.
New claim 40 recites, “wherein the T7 RNA polymerase variant incorporates 2’ modified nucleoside triphosphates consistently in each round of the SELEX process without base incorporation bias”, which is indefinite because it is unclear what constitute “consistent” incorporation of 2’ modified nucleoside triphosphates. The specification does not provide a limiting definition for the term “consistently”. As such, the metes and bounds of the claim cannot be determined.
Claim Rejections - 35 USC § 112(a)- New Matter
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.
New claim 43 is 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. This is a new matter rejection.
The Applicant is reminded that an amendment to the claims or the addition of a new claim must be supported by the description of the invention in the application as filed. In re Wright, 866 F.2d 422, 9 USPQ2d 1649 (Fed. Cir. 1989). New or amended claims which introduce elements or limitations which are not supported by the as-filed disclosure violate the written description requirement. See, e.g., In re Lukach, 442 F.2d 967, 169 USPQ 795 (CCPA 1971); In re Smith, 458 F.2d 1389, 1395, 173 USPQ 679, 683 (CCPA 1972).
New claim 43 recites, “wherein the modified polynucleotides are modified aptamers comprising 25-100 nucleotides”. However, this limitation is not taught in the instant disclosure.
The specification recites, “A suitable nucleotide length for an aptamer ranges from about 15 to about 100 nucleotide (nt), and in various other preferred embodiments, 15-30 nt, 20-25 nt, 30-100 nt, 30-60 nt, 25-70 nt, 25-60 nt, 40-60 nt, 25-40 nt, 30-40 nt, any of 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39 or 40 nt or 40-70 nt in length” [0144], but does not recite the specific range of 25-100 nucleotides. The drawings do not present any aptamers have a range of lengths of 25-100 nt.
Therefore, the disclosure does not provide support for the limitation newly added to new claim 43 of an aptamer “comprising 25-100 nucleotides”, and the limitation constitutes new matter.
Claim Rejections - 35 USC § 112(a)- Scope of Enablement
Amended, previously presented, and new claims 23, 25, 28, and 39-43 are newly rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, because the specification, while being enabling for:
A method of preparing a library of 2'-O-methyl modified polynucleotides comprising synthesizing the 2'-O-methyl modified polynucleotides via in vitro transcription with the T7 RNA polymerase variant of claim 8 in the presence of 2'-O-methyl modified nucleoside triphosphates under conditions that allow synthesis of the 2'--O-methyl modified polynucleotide by the polymerase activity of the T7 RNA polymerase variant, wherein the method comprises in vitro evolution of nucleic acid molecules with highly specific binding to target molecules, and wherein the T7 RNA polymerase variant incorporates 2’-O-methyl modified nucleoside triphosphates with reduced base incorporation biases compared to T7 RNA polymerase variants having mutations relative to the wild-type T7 RNA polymerase according to SEQ ID NO: 1 under the same reaction conditions, wherein the mutations consist of one of the following sets of mutations relative to a wild-type T7 RNA polymerase according to SEQ ID NO: 1:
Y639L and H784A substitutions (LA),
K378R, Y639L, and H784A substitutions (LAR), or
E593G, Y639V, V685A, H784G, S430P, N433T, S633P, F8491, and F880Y substitutions (RGVG-M5).
does not reasonably provide enablement for:
A method of preparing a library of any 2' modified polynucleotides comprising synthesizing the any 2' modified polynucleotides via in vitro transcription with the T7 RNA polymerase variant of claim 8 in the presence of any 2' modified nucleoside triphosphates under conditions that allow synthesis of the any 2'-modified polynucleotide by the polymerase activity of the T7 RNA polymerase variant, wherein the method comprises a Systematic Evolution of Ligands by Exponential Enrichment (SELEX) process, and wherein the T7 RNA polymerase variant incorporates any 2’ modified nucleoside triphosphates consistently in each round of the process without base incorporation biases.
The specification does not enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make, use, and practice the invention commensurate in scope with these claims.
The rejection comprises the issue of an absence of an enabling disclosure for a method or preparing a library of any 2’ modified polynucleotides with any 2’ modifications other than 2’-O-methyl modifications, wherein the any 2’ modified nucleoside triphosphates are incorporated without an incorporation bias.
This issue was identified by the Office after analysis of the disclosure provided by the specification. The Office has analyzed the specification in direct accordance to the factors outlined in In re Wands, namely 1) the nature of the invention, 2) the state of the prior art, 3) the predictability of the art, 4) the amount of direction or guidance present, and 5) the presence or absence of working examples, and presented detailed scientific reasons supported by publications from the prior art for the finding of a lack of enablement for the scope of the instant methods. The Wands analysis and supporting specific evidence are presented below for each of the identified issues.
The specification does not provide an enabling disclosure for a method or preparing a library of any 2’ modified polynucleotides with any 2’ modifications other than 2’-O-methyl modifications, wherein the any 2’ modified nucleoside triphosphates are incorporated without an incorporation bias. The broadest independent claim, claim 8, recites a T7 RNA polymerase variant comprising a polypeptide sequence of SEQ ID NO: 2, wherein the T7 RNA polymerase variant does not have a bias in incorporating 2’-O-methyl modified A, T, G, or C nucleotides into a polynucleotide. Additionally, claim 23 encompasses the T7 RNAP of claim 8 in a “method of preparing a library of 2’ modified polynucleotides comprising synthesizing the 2’ modified polynucleotides via in vitro transcription with the T7 RNA polymerase variant of claim 8 in the presence of 2’ modified nucleoside triphosphates under conditions that allow synthesis of the 2’-modified polynucleotide by the polymerase activity of the T7 RNA polymerase variant”. Claims 25, 28, and 39-43 depend on claim 23. Claim 40 additionally recites “wherein the T7 RNA polymerase variant incorporates 2’ modified nucleoside triphosphates consistently in each round of the SELEX process without base incorporation biases”.
The specification teaches generically “In some embodiments, a T7 RNA polymerase variant of the present disclosure does not have a bias in incorporating a 2’-modifeid ATP, CTP, UTP, or GTP” and “In one embodiment, a T7 RNA polymerase variant of the present disclosure does not have a bias in incorporating 2'-O-methyl ATP, 2'-O-methyl CTP, 2'-O-methyl UTP, or 2'-O-methyl GTP” [0083].
However, the examples teach only a single experiment assessing the incorporation bias of the T7 RNAPs of the present invention in comparison to prior art T7 RNAPs. The transcription bias for incorporation of only 2’-O-methyl nucleotides by the LA, LAR, LAR-M5 (SEQ ID NO: 2), RGVG-M5, or LAR-M6 (SEQ ID NO: 2 modified with the P266L substitution) variants are presented in Table 3, wherein the 2’-O-methyl nucleotide incorporation bias for the LAR-M5 and LAR-M6 variants was reduced by at least about half compared to the biases for the LA, LAR, and RGVG-M5 variants. In reference to the data presented in Table 3, the specification includes a conclusory statement “It appears that while RGVG-M5 has a bias against incorporating 2'-O-methyl-ATP, all other variants including LAR-M5 do not have transcription biases” [0161].
Neither the specification nor the drawings provide any data for any assessment of T7 RNAP variant incorporation bias for any modifications other than 2’-O-methyl modified nucleotides.
The art at the time of filing teaches a consistent and early bias against inclusion of adenine in nontargeted SELEX library selections, resulting in aptamers with lower predicted minimum free energies [Thiel et al. 2011, Nucleic Acid Therapeutics, 21(4), 253-263, abstract, pg 256 col 2 ¶ 3, pg 259 col 2 ¶ 1, Figure 4, 5]. Thiel teaches that explanations for the nucleotide bias include biases in nucleotide misincorporations by the Y639F mutant T7 RNA polymerase and/or reverse transcriptase [pg 261 col 2 ¶ 1]. Additionally, Thompson teaches that T7 RNAP variants comprising LAR mutations have no bias for incorporating 2’-O-methyl modified A, U, G, or C nucleotides, wherein the percentage of nucleotide composition after transcription was very similar to the percentage of nucleotide composition of the starting library [Thompson, WO2008078180A2, published 2 July 2008, IDS, cited in a prior action, 00189-00190, Figure 8].
Neither the specification nor the art at the time of filing teaches a lack of 2’-modified nucleotide incorporation bias by T7 RNAP variants comprising mutations encompassed by the instant claims for any 2’-modified nucleotides other than 2’-O-methyl modified nucleotides. Thus, in view of the multiple alternative 2’ modifications possible for RNA nucleotides, Applicant’s limitation to assessing bias for incorporation of 2’-O-methyl modified nucleotides, the limitations of teaching in the art for a lack of bias for LAR-comprising variants only for incorporation of 2’-O-methyl modified nucleotides, and the breadth of the claims, the skilled artisan would have considered preparing a library of any 2’ modified polynucleotides with any 2’ modifications other than 2’-O-methyl modifications, wherein the any 2’ modified nucleoside triphosphates are incorporated without an incorporation bias, as highly unpredictable. As such, it would have required undue experimentation to practice the scope of Applicant’s invention as claimed.
Claim Rejections - 35 USC § 103
The rejection of amended, previously presented, and cancelled claims 4-5, 7-8, 10, 12, 23, 25, 28, and 36-37 under 35 U.S.C. 103 as being unpatentable over Thompson [WO2008078180A2, published 2 July 2008, IDS]; in view of Ellington [WO2015143318A1, published 24 September 2015]; Addgene [Plasmid: pQE-80L, archived 21 May 2014, retrieved on 22 August 2024 from the Internet: <https://web.archive.org/web/20140521214512/https://www.addgene.org/vector-database/3883/>]; and Elena et al. [2014, Frontiers in Microbiology, Vol. 5, Article 21, 1-8]; is withdrawn over cancelled claims 7 and 37, maintained over amended and previously presented claims 5, 7-8, 10, 12, 23, 25, 28, and 36-37, and newly applied to new claims 38-43. Applicant's amendments to the claims and arguments have been fully considered but have not been found persuasive in overcoming the rejection for reasons of record as discussed in detail below.
Applicant amended independent claim 8 to recite “wherein the T7 RNA polymerase variant does not have a bias in incorporating 2’-O-methyl modified A, T, G, or C nucleotides into a polynucleotide” in claim 8 lines 2-4, which was addressed in the prior rejection, wherein Thompson was cited for teaching that the triple mutant comprising Y639L/H784A/K378R has higher fidelity than wildtype T7 RNAP and no 2’-O-methyl nucleotide bias [00189-00190].
Applicant amended claim 10 to recite that the enhanced ability to incorporate a 2’-O-methyl modified nucleotide compared to a wild-type T7 RNA polymerase is “under the identical reaction condition”, which limitation was taught by Thompson [0016, 0053, 0057, 0094, 0096-0097, 0108, Figure 7 and 11].
Regarding amended claim 12 and new claim 41, Ellington teaches wherein the “M5” mutations arose in a T7 RNA polymerase selection for transcriptional activity at higher temperatures, wherein the mutations increase the half-life of enzyme at 50oC and allow for transcription at that temperature (i.e., 50oC) [0040]. Ellington also teaches that for all the T7 RNAP variants tested, addition of the M5 mutations increased their thermal stability [0044, Figure 11]. Figure 11 further shows that all variants with added M5 mutations had Tm values above 45oC, wherein only the RGVG-M5 variant Tm was lower than 50 oC. Thompson teaches that the LAR mutant is active at 37oC [00234]. Therefore, given the teachings that the M5 mutations allow for transcription at 50oC and the teachings that the M5 mutations increase the Tm of all T7 RNAP variants to which they are added, an ordinarily skilled artisan at the time of filing the instant application would expect that addition of the M5 mutations to the LAR T7 RNA polymerase variants would increase the Tm to allow for transcription at 50oC with a reasonable expectation of success.
Regarding amended claim 23 and new claims 39-40, note that Thompson was cited for teaching a method of preparing a 2’-O-methyl modified polynucleotides comprising synthesizing the 2’ modified polynucleotides via in vitro transcription using a SELEX process, as discussed in the prior action [0059, 0085-0088, 00182-00196]. Thompson also teaches preparing a library of 2’-O-mehtyl modified polynucleotides [0190]. Thompson further teaches carrying out 10 or 12 rounds of SELEX selection [00205-00206, 00216, 00237, Table 4, 5, 6, 7, 8]. As discussed above, Thompson teaches wherein the LAR variant does not have a base transcription bias [00189-00190].
Regarding new claim 38, Thompson was cited for teaching the inclusion of the P266L mutation [0050-0054, 0094, 0096, 00104, 00177].
Regarding new claim 42, Thompson teaches wherein the library of 2’ modified polynucleotides is used to identify modified polynucleotides that specifically bind to a target of interest [00201-00205], and wherein the modified polynucleotides are modified aptamers comprising 76 nucleotides (i.e., starting with the first transcribed nucleotide of the T7 RNAP promoter (TAATACGACTCACTATAG(+1)GG) within the DNA template according to SEQ I D NO 7 through the end of the DNA template) [00203].
Therefore, given the teachings of Ellington that the M5 mutations allow for transcription at 50oC and that the M5 mutations increase the Tm of all T7 RNAP variants to which they are added, such that an ordinarily skilled artisan at the time of filing the instant application would expect that addition of the M5 mutations to the LAR T7 RNA polymerase variants would increase the Tm to allow for transcription at 50oC, it would have been prima facie obvious to an ordinarily skilled artisan that modifying the T7 RNA polymerase variant of Thompson with the M5 mutations of Ellington would produce a T7 RNA polymerase variant capable of producing a modified polynucleotide at 45-50 oC with a reasonable expectation of success.
Applicant argues that:
a PHOSITA would not have had a reasonable expectation of success in combining the specific sets of mutations taught by Thompson and Ellington because introducing different mutations into a single enzyme is notorious for having unpredictable, non-additive, or synergistic effects. In the present case, combining 3 mutations from Thompson ("LAR') with 5-6 mutations from Ellington ("M5" or "M6") in T7 RNA polymerase requires the simultaneous introduction of eight or nine distinct amino acid alterations into a single enzyme framework, which can create an entirely new structural landscape, such that stacking these independent sets of mutations introduces a severe risk of disrupting the delicate tertiary or quaternary folding of the polymerase. A PHOSITA would reasonably expect that such structural disruption would partially or completely silence catalytic activity or compromise enzyme stability. Because none of Thompson, Ellington, Addgene and Elena provide any structural or functional assurance that these two disparate sets of mutations would be structurally compatible - let alone achieve superior integrity and high-yield transcriptional activity when combined, the Examiner's rejection relies on an impermissible assumption of additive predictability.
the claimed LAR-M5 and LAR-M6 variants exhibit unexpected results that completely distinguish them from the prior art. Although Thompson and Ellington teach the "LAR" variant and the "M5" variant, respectively, neither reference discloses or suggests that the claimed "LAR-M5" and "LAR-M6" variants exhibit an unbiased ability to incorporate 2'-O-methyl modified ATP, CTP, GTP, and TTP during transcription. Table 3 reveals that the relative distributions of 2'-O-methyl modified A, T, C, and G nucleotides in transcripts produced by the LAR-M5 and LAR-M6 variants closely track the initial distribution in the naive pool, demonstrating a lack of incorporation bias. As shown in Table 3, testing under identical reaction conditions reveals that the prior art variants exhibit significant incorporation bias. These data suggest the RGVG-M5 variant has a bias against modified A and in favor of modified T incorporation. Transcripts produced by the LA and LAR variants demonstrate a decrease in 2'-O-methyl modified G and a concurrent increase in 2'-O-methyl modified C, suggesting a bias against modified G and in favor of modified C. In contrast, the LAR-M5 variant demonstrates exceptional fidelity across all four bases, restricting deviations to nominal baseline shifts. The LAR-M6 variant demonstrates a similarly unbiased profile. These data collectively demonstrate that the LAR-M5 and LAR-M6 variants unexpectedly eliminate incorporation bias across all four nucleotide bases, which a PHOSITA could neither predict nor derive from the prior art. Because the claimed variants unexpectedly eliminate the systematic incorporation biases of the prior art T7 RNA polymerases, the claimed invention is non- obvious. This unbiased incorporation is of critical operational significance when the T7 RNA polymerase is utilized in the preparation of libraries of 2'- modified polynucleotides, such as in Systematic Evolution of Ligands by Exponential Enrichment (SELEX). This operational divergence proves that the claimed variants achieve an unexpected, non-obvious breakthrough that solves an inherent bias in the prior art variants. A PHOSITA would have had no reasonable expectation that combining them would successfully eliminate these biases. Accordingly, the objective evidence of unexpected results completely overcomes the prima facie allegation of obviousness.
However, this is not agreed
In response to Applicant’s arguments against the references individually, it is noted that the test for obviousness is not whether the features of a secondary reference may be bodily incorporated into the structure of the primary reference; nor is it that the claimed invention must be expressly suggested in any one or all of the references. Rather, the test is what the combined teachings of the references would have suggested to those of ordinary skill in the art. See In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981). One cannot show nonobviousness by attacking references individually where the rejections are based on combinations of references. See In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981); In re Merck & Co., 800 F.2d 1091, 231 USPQ 375 (Fed. Cir. 1986). Further, the Examiner recognizes that obviousness may be established by combining or modifying the teachings of the prior art to produce the claimed invention where there is some teaching, suggestion, or motivation to do so found either in the references themselves or in the knowledge generally available to one of ordinary skill in the art. See In re Fine, 837 F.2d 1071, 5 USPQ2d 1596 (Fed. Cir. 1988), In re Jones, 958 F.2d 347, 21 USPQ2d 1941 (Fed. Cir. 1992), and KSR International Co. v. Teleflex, Inc., 550 U.S. 398, 82 USPQ2d 1385 (2007). In addition, it must be recognized that any judgment on obviousness is in a sense necessarily a reconstruction based upon hindsight reasoning. But so long as it takes into account only knowledge which was within the level of ordinary skill at the time the claimed invention was made, and does not include knowledge gleaned only from the applicant's disclosure, such a reconstruction is proper. See In re McLaughlin, 443 F.2d 1392, 170 USPQ 209 (CCPA 1971).
Specifically, regarding Applicant’s argument 1), Thompson teaches wherein the LAR mutations themselves do not compromise the structure of the T7 RNAP, in that the LAR variant itself is active in generating RNA transcripts []. Additionally, Ellington teaches wherein the M5 mutations themselves do not compromise the structure of the T7 RNAP by teaching the molecular structure of the M5 variant [0017, Figure 2], along with teaching that both the M5 variant and variants comprising the M5 mutations in addition to other mutations are active in generating RNA transcripts [0039-0040]. Applicant has not provided any evidence that the specific mutations taught by Thompson and Ellington, and making up the LAR-M5 or LAR-M6 T7 RNAP variants, would be expected to interact with each other structurally so as to disrupt the intramolecular or protein-solvent interactions contributing to the structural folding and stability of the T7 RNAP polymerase and is merely making generalization about the possibility of structural disruption by the generic introduction of mutations.
As discussed in the prior actions, Ellington was cited for teaching the motivation to add the M5 variants to a T7 RNA polymerase variant with substrate broadening amino acid substitutions that confer enhanced ability to incorporate 2’ modified nucleotides to further enhance the activity of the resulting T7 RNA polymerase variants. Specifically, Ellington teaches that the M5-FA and M5-RGVG variants dramatically increased the yield for transcripts comprising all 2’-O-methyl modified nucleotides [Figure 6]. Although Figure 6 does not have quantitation, the band intensities presented in the gel indicate much greater than 2-fold increases in yield for the FA-M5 and RGVG-M5 variants compared to the parental FA and RGVG variants, respectively.
Ellington was also cited for teaching that the M5 or M6 polymerase substitutions increase the thermal stability of the T7 RNAP variants to increase the transcription yield [0044, Figure 11]. As discussed above, Ellington teaches wherein the “M5” mutations arose in a T7 RNA polymerase selection for transcriptional activity at higher temperatures, wherein the mutations increase the half-life of enzyme at 50oC and allow for transcription at that temperature (i.e., 50oC) [0040]. Ellington also teaches that for all the T7 RNAP variants tested, addition of the M5 mutations increased their thermal stability [0044, Figure 11]. Figure 11 further shows that all variants with added M5 mutations had Tm values above 45oC, wherein only the RGVG-M5 variant Tm was lower than 50 oC. Therefore, given the teachings that the M5 mutations increase thermal stability and allow for transcription at 50oC and the teachings that the M5 mutations increase the Tm of all T7 RNAP variants to which they are added, an ordinarily skilled artisan at the time of filing the instant application would expect that addition of the M5 mutations to the LAR T7 RNA polymerase variant would actually increase the structural stability of the T7 RNAP variant. Accordingly, the skilled artisan would not expect that combining the M5 variants with the LAR variants would disrupt the delicate tertiary or quaternary folding of the polymerase nor would the ordinarily skilled artisan expect that addition of the M5 mutations would introduce such structural disruption so as to partially or completely silence catalytic activity or compromise enzyme stability.
Regarding Applicant’s argument 2), it is noted that any evidence of unexpected results must be commensurate in scope with the claimed invention, and that a greater, or greater than additive, effect is not necessarily sufficient to overcome a prima facie case of obviousness because such an effect can either be expected or unexpected MPEP 716.02 (a) and (d). Whether the unexpected results are the result of unexpectedly improved results or a property not taught by the prior art, the "objective evidence of nonobviousness must be commensurate in scope with the claims which the evidence is offered to support." In other words, the showing of unexpected results must be reviewed to see if the results occur over the entire claimed range. In re Clemens, 622 F.2d 1029, 1036, 206 USPQ 289, 296 (CCPA 1980).
Specifically, note that the data presented in Table 3 demonstrate a reduced base incorporation bias for incorporating 2’-O-methyl nucleotides, but does not demonstrate that the instantly claimed LAR-M5 or LAR-M6 have no bias in that the nucleotide proportions following transcription are not identical to the nucleotide proportions of the initial dataset. Accordingly, although the reduction in nucleotide incorporation bias presented in instant Table 3 may have been unexpected, the data is not commensurate in scope with the claims because the claims require that the recited T7 RNAP does not have a bias in incorporating 2’-O-methyl modified A, T, G, or C nucleotides.
Alternatively, to the extent that the reduced transcription bias presented in Table 3 is sufficiently low to approximate no bias in incorporating 2’-O-methyl modified A, U, G, or C nucleotides, a lack of bias in incorporating 2’-O-methyl modified nucleotides is not unexpected given the teachings of the cited prior art. Thompson was cited for teaching that the triple mutant comprising Y639L/H784A/K378R has no 2’-O-methyl nucleotide bias [00189-00190]. Even the instant application itself, contrary to Applicant’s arguments, teaches wherein the LA and LAR mutants do not have a bias in incorporating 2’-O-methyl modified A, U, G, or C nucleotides. In reference to the data presented in Table 3, the instant specification concludes “It appears that while RGVG-M5 has a bias against incorporating 2'-O-methyl-ATP, all other variants including LAR-M5 do not have transcription biases” [0161]. Note that both LA and LAR are included among the “all other variants” presented in Table 3 and referenced in [0161]. Nothing in Thompson, Ellington, nor the instant application suggests that addition of the M5 mutations would be expected to increase the transcriptional bias of a T7 variant. Therefore, given that both Thompson and the instant specification teach wherein the LAR variant does not have a transcription bias for incorporating 2’-O-methyl modified nucleotides, and given that a lower than zero bias is still a zero bias, it follows that a lack of transcription bias by the LAR-M5 variant for incorporating 2’-O-methyl modified nucleotides cannot be unexpected in view of Thompson and Ellington.
Accordingly, Applicant’s arguments do not overcome a finding of obviousness under 35 U.S.C. 103 over Thompson, Ellington, Addgene, and Elena, and the rejection is maintained.
Conclusion
No claim is allowed.
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DR. KATIE L. PENNINGTON
Examiner
Art Unit 1634
/KATIE L PENNINGTON/Examiner, Art Unit 1634
Dr. A.M.S. Wehbé
/ANNE MARIE S WEHBE/Primary Examiner, Art Unit 1634
