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 .
Priority
This application claims priority to provisional application 63/357,843. Therefore, it is entitled to the 1st Jul 2022 priority date of the parent application.
Status of Claims
Claims 1-3, 5-7, 9-12, 14-17,and 22-27 are under consideration.
Claims Objections
Claim 1 recites, ethylenediaminetratracetic, instead of ethylenediaminetetraacetic.
Claim 25 recites in lines 2-4, 1 to 25 g of single stranded transcribed RNA/ L of liquid of the transcription reaction prior to the addition of DNase in step (b) is produced.
The in step (b) is probably a typo because the addition of DNase happens in step (e) and not step (b), and must be corrected.
Claim 27 recites, otherwise identical transcription reaction wherein the yield is less than about 5 g/L.28.
The .28 is probably a typo and must be corrected.
Appropriate correction is required.
Specification
The abstract of the disclosure is objected to because:
The abstract is less than 50 words in length. A corrected abstract of the disclosure is required and must be presented on a separate sheet, apart from any other text. See MPEP § 608.01(b).
Applicant is reminded of the proper content of an abstract of the disclosure.
A patent abstract is a concise statement of the technical disclosure of the patent and should include that which is new in the art to which the invention pertains. The abstract should not refer to purported merits or speculative applications of the invention and should not compare the invention with the prior art.
If the patent is of a basic nature, the entire technical disclosure may be new in the art, and the abstract should be directed to the entire disclosure. If the patent is in the nature of an improvement in an old apparatus, process, product, or composition, the abstract should include the technical disclosure of the improvement. The abstract should also mention by way of example any preferred modifications or alternatives.
Where applicable, the abstract should include the following: (1) if a machine or apparatus, its organization and operation; (2) if an article, its method of making; (3) if a chemical compound, its identity and use; (4) if a mixture, its ingredients; (5) if a process, the steps.
Extensive mechanical and design details of an apparatus should not be included in the abstract. The abstract should be in narrative form and generally limited to a single paragraph within the range of 50 to 150 words in length.
See MPEP § 608.01(b) for guidelines for the preparation of patent abstracts.
In the case of instant, words such as “disclosed” should be avoided.
Claim Rejections - 35 USC § 112
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claims 2, 24-25, and 27 are rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
Claim 2 recites “wherein the L.DNA template is in a solution comprising 50 mM to 1200 mM NaCl”. This language is considered indefinite because it is not clear how this product limitation in claim 2 relates to the method of claim 1. The specification in [00208] discloses, “For the self-replicating mRNAs made with unmodified NTPs, any salt spike level contributed to dsRNA level decrease with significant effects seen with> 200 mM NaCl salt spike (Fig. 8). It was observed that salt spikes > 1200 mM NaCl will result in IVT yield inhibition”. However, the claim remains indefinite because this disclosure still does not limit a step or steps of claim 1.
The following recitation may be remedial:
“The method of claim 1 further comprising a step of producing a transcription reaction mixture by introducing each of the elements into the mixture, wherein the L.DNA template is in a solution comprising 50-1200 mM NaCL prior to introducing the L.DNA into the transcription reaction mixture”
Claim 25 recites parenthetical expression “(define as…)” and “(define in spec…)”. The metes and bounds of claim 25 are rendered vague and indefinite by various parenthetical recitations. Such parenthetical recitations are indefinite because it is unclear as to whether the limitations are part of the instantly claimed subject matter.
Claims 24 and 27 recite, otherwise identical transcription reaction wherein the yield is less than about 5 g/L, in the second to third lines, which is indefinite. The term “identical" is not defined by the claim, the specification does not provide a clear definition for the term, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. If the reaction to which it is to be compared is identical then the yield should be same. However, the recitation includes a different yield. It’s confusing to know that the yield is different but the reaction that resulted in that yield is identical.
Claim Interpretation
Claim 1 is being interpreted as a method of producing a transcribed RNA product comprising the following steps:
i. transcription reaction wherein a yield of 1 g to 25 g of single stranded transcribed RNA per liter of liquid of the transcription reaction prior to the addition of DNase in step (ii) is produced.
ii. stopping the transcription reaction
The transcription reaction mixture of a. comprises:
a buffer solution comprising Mg2+, molar concentration of Mg2+ is 2 to 15 mM above the total molar concentration of all rNTPs plus the optional RNA capping reagent,
linear DNA (L.DNA) template,
ribonucleoside tri-phosphate (rNTPs),
optionally an RNA capping reagent,
and RNA polymerase; wherein RNA polymerase/L. DNA template mass ratio is between 0.25 and 3;
The stopping transcription reaction comprises:
deoxyribonuclease (DNase) or ethylenediaminetetraacetic acid (EDTA).
As seen in the table below, Claim 1 and claim 25 recite same subject matter except for the bolded limitation and the order of steps presented in claim 25.
Claim 1 is being interpreted as a method of producing a transcribed RNA product comprising the following steps: i. transcription reaction, wherein the transcription reaction mixture of i. comprises:
Claim 25 is A method of producing a transcribed RNA product comprising:
a buffer solution comprising Mg2+, molar concentration of Mg2+ is 2 to 15 mM above the total molar concentration of all rNTPs plus the optional RNA capping reagent
a. loading a reactor vessel with a buffer solution comprising Mg2+ and nuclease free water, molar concentration of Mg2+ is 2 to 15 mM above the total molar concentration
of all rNTPs plus the optional RNA cap after steps (a)-(b) are completed
linear DNA (L.DNA) template
b. adding a solution comprising L.DNA template spiked with 200-1000 mM NaCl;
ribonucleoside tri-phosphate (rNTPs)
c. adding rNTPs and optionally RNA cap
RNA polymerase; wherein RNA polymerase/L. DNA template mass ratio is between 0.25 and 3;
d. adding RNA polymerase and mixing the resulting transcription solution wherein the RNA polymerase/DNA template mass ratio is between 0.25 and 3; and,
yield of 1 g to 25 g of single stranded transcribed RNA per liter of liquid of the transcription reaction prior to the addition of DNase in step (ii) is produced.
1 to 25 g of
single stranded transcribed RNA/ L of liquid of the transcription reaction prior to the addition of
DNase in step (b) is produced.
ii. The stopping transcription reaction comprises:
deoxyribonuclease (DNase) or ethylenediaminetetraacetic acid (EDTA).
e. adding DNase to the transcription solution;
Claims 24 and 27 are being interpreted as methods wherein when yields of greater than 5 g/ L of RNA transcripts occur, these are accompanied by reduced amounts of dsRNA impurities.
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) 1, 3, 5-7, 9-12, 14, 17, and 22-24 are rejected under 35 U.S.C. 103 as being obvious over Dias (US 2018/0258423 A1, IDS) in view of Piao (WO 2021/158789 A1).
Regarding claim 1, Dias teaches methods of producing large scale RNA in a buffered transcription reaction mix using RNA polymerase (whole document). In an embodiment with T7 polymerase, 10g of CFTR mRNA is prepared([0200] a 10 gram batch of CFTR mRNA; Fig. 24B). See two pertinent citations that read on instant claim 1 limitations:
[0165]: The T7 transcription reaction consisted of 1 xT7
transcription buffer (80 mM HEPES pH 8.0, 2 mM Spermidine,
and 25 mM MgCl2 with a final pH of7.7), 10 mM
DTT, 7.25 mM each ATP, GTP, CTP, and UTP, RNAse
Inhibitor, Pyrophosphatase, and T7 Polymerase.
a) Scaled up reaction [0166]: 20 mg of a linearized double stranded
DNA plasmid with an RNA polymerase-specific promoter,
10 mg RNA polymerase, RNase inhibitor, pyrophosphatase,
5 mM NTPs, 10 mM DTT and a reaction buffer (l0x-250
mM Tris-HCI, pH 7.5, 20 mM spirmidine, 50 mM NaCl,)
was used and quantity sufficient (QS) to 200 ml with
RNase-free water then incubated at 37 C for 60 min.
The amounts in [0166] are for each gm of mRNA produced. Therefore, if 1gm is produced in 200mL, when an yield of 10gm is obtained as in [0200], this must be 10g/2000mL, which is 5g/L, which is within the recited range of 1g/L to 25g/L.
As seen in the citation, ratio of DNA to polymerase is 20:10. Therefore, ratio of polymerase to DNA is 1:2, which is 0.5; i.e., within the required ratio of 0.25 to 3.
Regarding claim 3, Dias teaches wherein the transcription reaction mixture of step (a) further comprises one or more of the group consisting of RNase inhibitor and inorganic pyrophosphatase [0167].
Regarding claim 5, Dias teaches wherein the temperature during step a) and b) is in a range from about 30 °C to about 40 °C [0167].
Regarding claim 6, Dias teaches wherein the L.DNA template is from about 0.01 mg/mL to about 0.3 mg/mL in the transcription reaction mixture [0167]. As seen in the citation, DNA is 20mg/200mL, which is 1mg/10mL, which is 0.1mg/1mL.
Regarding claim 7, Dias teaches wherein the RNA polymerase is T7 polymerase [0165].
Regarding claim 9, Dias teaches wherein the transcription reaction mixture of step (a) is allowed to react for at least 20 minutes prior to step (b) stopping the transcription reaction [0167].
Regarding claim 10, Dias teaches the Cap and Tail (C/7') Reaction is carried out on purified DNA [0167].
Regarding claim 12, Dias teaches wherein the RNA polymerase is 0.0125 to 0.15 μg/μL T7 polymerase [0167]. As seen in the citation, T7 polymerase is 10mg/200mL, which is 0.05μg/1 μL.
Regarding claim 14, Dias teaches wherein the amount of single stranded transcribed RNA is measured after purification of the transcription mixture after step (b) via a silica column (the purification via silica column various ways to purify, [0120]; and Qiagen RNA maxi column, [0165]; 1 to 2 μg of RNA was treated…[0170]). Since Dias runs 1-2 μg of purified RNA on a gel, a measurement has to be done to further treat 1-2 μg.
Regarding claim 22, Dias teaches wherein the L.DNA template comprises an open reading frame that encodes a vaccine antigen, enzyme, antibody, receptor, tRNA, and/ or a protein (antibody, para [0150]-[0151]; endonuclease, para [0153]; CFTR mRNA, [0200]).
Regarding claim 23, Dias teaches wherein the total concentration of rNTPs is at least 8 mM (5 mM NTPs, [0167]). If 5 mM of each NTPs are used, then for four of the NTPs, this would amount to 20mM. Since the MgCl2 has not been scaled up, the MgCl2 remains at 25mM. 25mM is greater than the total of all rNTPs.
Dias is silent on whether Mg2+ is carried forward in the large scale reaction, as required by instant claims 1 and 11.
Dias lacks a teaching on a solvent from the recited group in claim 17.
However, before the effective filing date of instant invention, Piao had taught a method of producing a transcribed RNA product comprising reacting a transcription reaction mixture comprising a RNA polymerase, resulting in increased yield and lacking dsRNA impurities (abstract). Piao taught wherein the reaction mixture in the method of producing a transcribed RNA product contains Mg2+ ([0089]: Magnesium/magnesium ions and DTT (or some other reducing agent) are often included in transcription buffers). Magnesium in Piao’s transcription reaction mixture is at 165 mM Mg2+ (The starting reaction mixture comprises, in certain embodiments, 50 ng/μL DNA plasmid template, rNTPs (5 mM), CleanCap AG (4 mM), reaction buffer (l0X buffer: 400 mM HEPES (pH 7.2-7.5), 100 mM DTT, 20 mM spermidine, 0.02% triton X-100, 165 mM Mg2+), T7 RNA polymerase (about 4000 U/mL), RNase inhibitor (about 1000 U/mL), and inorganic pyrophosphatase (about 2 U/mL), [0090]). The transcribed RNA yields without any additives are at least 2.8 mg/mL (Tables 2 - 14).
Thus, Piao teach that Mg2+ is essential for the transcription reaction when an yield of 1g/L to 25g/L of transcribed RNA is produced. The recited molar concentration of Mg2+ is 5 to 15 mM above the total molar concentration of all rNTPS plus the molar concentration of any optional RNA capping reagent: In Piao’s reaction, if 5 mM of each NTPs are used, then for four of the NTPs, this would amount to 20mM, plus 4 mM of the capping reagent = 24mM total. To be 2 to 15 mM and 5 to 15 mM above this total molar concentration, as required by claims 1 and 11 respectively, one would require a minimum of 26mM and 29mM respectively. Piao’s MgCl2 is greater than this required molarity.
Regarding claim 17, Piao had taught a method of producing a transcribed RNA product comprising reacting a transcription reaction mixture comprising a RNA polymerase, wherein the method comprises adding a solvent (chaotropic agent) such as EtOH to the transcription reaction mixture which acts to reduce the formation of double-stranded ribonucleic acid (dsRNA) during in vitro transcription (claims 1-6 A method of reducing, minimizing, or inhibiting the formation of double-stranded ribonucleic acid (dsRNA) during in vitro transcription, comprising adding at least one chaotropic agent to an in vitro transcription reaction mixture ... wherein the transcription yields ribonucleic acid (RNA) ... wherein the at least one chaotropic agent is selected from the group consisting of urea, formamide, sodium salicylate, ethanol, sodium perchlorate, arginine, n-butanol, thiourea, and 2-propanol). Piao teaches the composition comprising EtOH in a concentration of 1 to 10 % v/v (claim 18 wherein the ethanol is at a concentration of from about 0.4M to less than about 1.6M). Piao teaches addition of the solvent maintains mRNA yield while reducing the formation of unwanted dsRNA during in vitro transcription (claims 1, 5 wherein the amount or yield of RNA is not significantly reduced by the addition of the at least one chaotropic agent).
It would have been obvious to one of ordinary skill, in the art at the time, to modify the method of Dias by scaling up Mg2+ ions and including at least one chaotropic agent as taught by Piao to gain the advantage of reducing the formation of unwanted dsRNA during in vitro transcription. Further, it would have been obvious to one of ordinary skill in the art to have applied the transcription reaction mixture components taught by Piao to the mRNA transcription methods taught by Dias, in order to improve the overall quality of the mRNA transcription products generated. One would have had reasonable expectation of success in doing so because both references taught methods of improving yield of IVT RNA. See MPEP 2143 I (A) and 2144 II.
Regarding claim 24, the method of producing a transcribed RNA product comprising reacting a transcription reaction mixture comprising a RNA polymerase, wherein the method comprises adding a solvent (chaotropic agent) such as EtOH to the transcription reaction mixture which acts to reduce the formation of double-stranded ribonucleic acid (dsRNA) during in vitro transcription (claims 1-6), wherein a yield of about 5 g/L of RNA transcript results in a reduced amount of dsRNA as compared to a transcription reaction wherein the yield is less than about 5 g/L (para [0121]-[0122]).
Thus, Dias in view of Piao make obvious instant claims 1, 3, 5-7, 9-12, 14, 17, and 22-24.
Claims 2, 15-16, 25 and 27 are rejected under 35 U.S.C. 103 as being unpatentable over Dias (US 2018/0258423 A1, IDS) in view of Piao (WO 2021/158789 A1) as applied to claims 1, 3, 5-7, 9-12, 14, 17, and 22-24 above in view of Cavac (Cavac et al., J. Biol. Chem. (2021) 297(3) 100999). Claims 15-16 are evidenced by Martins (R. Martins et al. / J. Chromatogr. A 1355 (2014) 1–14).
Regarding claim 2, the method of claim 1 is discussed above. Neither Dias nor Piao teach wherein the L.DNA template is in a solution comprising 50 mM to 1200 mM NaCl to produce a salt-spiked L.DNA template prior to introduction to the transcription reaction mixture.
However, before the effective filing date of instant invention, Cavac had taught a method of producing transcribed RNA comprising reacting T7 RNA polymerase with linear DNA in the presence of high salt to generate increased yields of highly pure RNA (title, abstract). The taught method results in significantly lower yields of dsRNA impurities (abstract). Regarding amounts of added NaCl, Cavac teach at pg. 3, l col, “At 0.3 M added NaCl, most of the primer extension activity is inhibited, leading directly to an increase in the encoded RNA yield.”; at pg. 6, r col, 1st para “a practical optimum of about 0.3 M added NaCl provides a good trade-off of purity versus yield”. Also see Fig. 2. Cavac further teach the concentrations of higher salt will vary with the needs of the user (For practical consideration, users can determine the optimal concentration of added salt, depending on their RNA sequence, targeted degree of purity, and desired yield, at pg. 6, r col, 3rd para).
It would have been obvious to one of ordinary skill, in the art at the time, to modify the method of Dias and Piao, to include a higher concentration of salt in the reaction mix, for the advantage of obtaining high purity yield as demonstrated by Cavac. With respect to the order of steps, this is a matter of design choice. See MPEP 2144.05 II and In re Williams, 36 F.2d 436, 438, 4 USPQ 237 (CCPA 1929) ("It is a settled principle of law that a mere carrying forward of an original patented conception involving only change of form, proportions, or degree, or the substitution of equivalents doing the same thing as the original invention, by substantially the same means, is not such an invention as will sustain a patent, even though the changes of the kind may produce better results than prior inventions."). With respect to the order of steps, in particular, it is noted that the courts have held that any order of performing process steps is prima facie obvious in the absence of new or unexpected results (In re Gibson, 39 F.2d 975, 5 USPQ 230 (CCPA 1930); Ex parte Rubin, 128 USPQ 440 (Bd. App. 1959)). See MPEP §2144.04 IV C.
Therefore, the claimed order of steps of spiking the stock template DNA solution or incorporating the high salt in the reaction mix, is an obvious variant of the steps of the cited prior art.
Regarding claims 15 and 16, Cavac teach in vitro T7 RNA polymerase transcription is often followed by extensive purification methods (abstract). Cavac teach reliance on different purification methods depends upon the length of RNA obtained (At long RNA lengths, purification methods do not have the resolution to eliminate all impurities (for example, separating a 2000 base transcript from a 2040 base impurity) and purifications lead to a loss in yield, (pg. 1, r col, 2nd para). As evidenced by Martins cited by Cavac at pg. 1, r col, 2nd para, one such method is affinity purification. Martin further evidences, more than one purification step is required to achieve higher RNA enrichments (pg. 9, l col, first line). Martin further evidences, that the major advantage of these systems is the broad applicability to any RNA of interest, even though limited to the need of several design issues, which may lead to longer optimization processes. Thus, Cavac teaches that the choice of purification and order of use is a results-effective variable that is readily optimized by one of ordinary skill in the art as needed or desired.
It would have been obvious to one of ordinary skill in the art apprised of the teachings of Dias and Piao to also come across the teachings of Cavac and incorporate an affinity purification method because Cavac indicates that extensive purification is often required and many methods exist for the same, and the choice of which method to pick depends upon the RNA that needs purification. Therefore, for any given IVT procedure, the skilled artisan would recognize that the purification methods to follow are subject to optimization via routine methods. One would have had reasonable expectation of success in optimizing an affinity purification method after silica column purification to obtain a highly pure IVT RNA, as these were well-known methods, well-within the grasp of the ordinary skilled artisan. See MPEP §2143 A and 2144.05 II.
Regarding claim 25, as discussed in claim interpretation, the method recited in claim 25 is substantially identical to the method recited in claim 1, except for the limitation of L.DNA in salt solution. Therefore, the discussion of claim 1 in the §103 rejection in view of Dias and Piao and the discussion of claim 2 in the §103 rejection in view of Cavac is incorporated herein.
Regarding claim 27 that depends upon claim 25, Cavac further teach double-stranded impurities are significantly reduced when a “high-salt transcription” method is followed (abstract). This reads on the limitation of claim 24, wherein a yield of greater than about 5 g/ L of RNA transcript results in a reduced amount of dsRNA as compared to a reaction which is not carried out in high-salt concentration.
It would have been obvious to one of ordinary skill, in the art at the time, to modify the method of Dias and Piao, to include a higher concentration of salt in the reaction mix, for the advantage of obtaining high purity yield as demonstrated by Cavac. One would have had reasonable expectation of success in optimizing a high-salt step, as Cavac provides a detailed methodology to do so. See MPEP §2143 A and 2144.05 II.
Regarding claim 26, the discussion of claim 1 and 17 in the §103 rejection in view of Dias and Piao, the discussion of claim 2 in the §103 rejection in view of Cavac, and the rationale for combining references are incorporated herein.
The combination of the limitations taught by Dias, Piao, and Cavac put together will result in the limitations of claim 26. With respect to the order of steps, again, it is noted that the courts have held that any order of performing process steps is prima facie obvious in the absence of new or unexpected results (In re Gibson, 39 F.2d 975, 5 USPQ 230 (CCPA 1930); Ex parte Rubin, 128 USPQ 440 (Bd. App. 1959)). See MPEP §2144.04 IV C.
Thus, Dias and Piao in view of Cavac make obvious instant claims 2, 15-16, and 25 - 27.
Therefore the invention as a whole would have been prima facie obvious to one ordinary skill in the art before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains.
Conclusion
No claims are allowed.
Correspondence
Any inquiry concerning this communication or earlier communications from the examiner should be directed to SHABANA MEYERING, Ph.D. whose telephone number is (703)756-4603. The examiner can normally be reached M - F: 9am to 5pm EST.
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SHABANA S. MEYERING, Ph.D.
Examiner
Art Unit 1635
/SHABANA S MEYERING/ Examiner, Art Unit 1635
/CATHERINE KONOPKA/ Primary Examiner, Art Unit 1635
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