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 is claims priority to United States Provisional Application No.
63/208,681 filed on June 9, 2021. As such, the earlier filing date of Jun. 9, 2021 is assigned to claims 1-47.
Claim Status
Claims 1-47 are pending and currently under examination.
Specification
The specification is objected to because the use of improperly demarcated trademarks has been noted in this application. Although the use of trademarks is permissible in patent applications, the proprietary nature of the marks should be respected and every effort made to prevent their use in any manner that might adversely affect their validity as trademarks. See MPEP §608. 01(v). 9. Some examples of such an improperly demarcated trademarks “MiSeq”, “NextSeq”, “HiSeq”, “NovaSeq”, “iSeq” which appear in the present specification in paragraph [0013]. Examiner notes that the provided examples are not meant to be a complete list of improperly demarcated trademarks found in the present specification. Applicant should review the entire specification and correct all instances of improperly demarcated trademarks. 10. Appropriate corrections required. Each letter of a trademark should be capitalized or otherwise the trademark should be demarcated with the appropriate symbol indicating its proprietary nature (e.g., ™ © ®) and accompanied by generic terminology. Applicants may identify trademarks using the USPTO's trademark database. Trademark Electronic Search System (TESS), on the Internet at https: / / www.uspto.gov7trademarks-application-process/search-trademark-database.
Claim Objections
Claims 6-12 are objected to because of the following informalities: typographical error.
Claim 6 recites “wherein said at least one primer comprises P5 and P7 primers” and should read “wherein said at least one primer comprises P5 or P7 primers”. Subsequent claims that depend on claim 6 (claims 7-12) should be amended to account for this updated verbiage as well.
Claim 8 recites “The method of claim 7, wherein said P7 primer is phosphorylated producing said functional sequence and said P5 primer is non-phosphorylated producing said non-functional sequence when 3' library is provided” and should read “The method of claim 7, wherein said P7 primer is phosphorylated producing said functional sequence and said P5 primer is non-phosphorylated producing said non-functional sequence when a 3' library is provided.”
Appropriate correction is required.
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 1-47 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 1 recites “both the artefactual dsDNA library member and the non-artefactual dsDNA library member comprises a first functional sequence at one end and a second functional sequence at the other end” however there is no definition of what structural or chemical attributes make a sequence “functional”, nor examples of alternative sequence types, primer chemistries or reaction conditions that would perform the same role. Additionally, term “functional sequence” has no common accepted meaning and is vague It is ambiguous whether any primer-binding site qualifies as a “functional sequence”. Under Ariad Pharm., Inc. v. Eli Lilly and Co., 598 F.3d 1336, 1341, 94 USPQ2d 1161, 1167 (Fed. Cir. 2010) and MPEP 2163, merely describing one or two species (P5/P7 adapters) does not demonstrate possession of a broad genus covering all unspecified “functional sequences”. Accordingly, the specification fails to show possession of the full breadth of “functional sequence” and is therefore rejected under 35 USC 112(a).
Further, the specification fails to reasonably convey to those skilled in the art that the inventor had possession of, or provided adequate description of the full scope of the limitation “neutralizing the second strand of the artefactual dsDNA library member” (and related passages referring to neutralizing a strand of the non-artefactual member).
The claim requires “neutralizing” a DNA strand, yet the specification does not define what constitutes “neutralizing” or provide objective criteria for when a strand is considered “neutralized.” In some places “neutralizing” appears to mean enzymatic digestion with Lambda or T7 exonuclease (see [0010], [0025]). Elsewhere it could include chemical modification (e.g., phosphorothioation) or other treatments that simply prevent sequencing or amplification (see [0057). Merely reciting two specific nucleases does not demonstrate possession of every possible means of “neutralizing” (chemical, photochemical, or physical) across the breadth of the claim.
Under MPEP 2163 and demonstrated in Ariad Pharm., Inc. v. Eli Lilly and Co., 598 F.3d 1336, 1341, 94 USPQ2d 1161, 1167 (Fed. Cir. 2010), disclosure of a few embodiments does not show possession of an undefined genus, which in this case would cover all “neutralizing” techniques.
Furthermore, The claim is not limited to nuclease digestion, but the specification gives no teaching or predictive guidance for alternative neutralization methods (e.g., chemical cross-linking, heat denaturation with selective protection, UV photolysis) that would satisfy the broad “neutralizing” requirement.
As such, independent claim 1 and dependent claims 2-47 are rejected under 35 USC 112(a) for failing to meet the written description requirements.
In regards to claim 6, nowhere in the specification does it describe any individual primer consisting of both P5 and P7 sequences. In contrast it describes them separately often with one being either phosphorylated or phosphorothioated to provide strand selectivity for neutralization.
In regards to claims 8, 9, 11, and 12, these claims recite, inter alia, “the method of claim _ when a 3’ / 5’ library is provided.” The specification, however, does not reasonably convey to those skilled in the art that applicant had possession of the invention as broadly claimed. They do not define the terms “3’ library” or “5’ library” nor do they identify structural, chemical, or process-based characteristics by which a skilled artisan could determine, with reasonable certainty, whether any given library falls within those categories. Consequently, the specification fails to provide representative species or common structural features commensurate with the full scope of the claims. See Ariad Pharm., Inc. v. Eli Lilly and Co., 598 F.3d 1336, 1341, 94 USPQ2d 1161, 1167 (Fed. Cir. 2010) and MPEP 2163. As such, claims 8, 9, 11, and 12 are rejected under 35 U.S.C. 112(a) for failing to meet the written description requirements.
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 1-47 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.
The term “functional sequence” lacks a clear definition and is used inconsistently in the specification. The claim recites that “both the artefactual dsDNA library member and the non-artefactual dsDNA library member comprises a first functional sequence at one end and a second functional sequence at the other end”. However, the specification does not provide an explicit definition of “functional sequence”. It variously associates the term with P5 and P7 primer sites, with 5’ phosphorylation or phosphorothioation, and with sequences that allow amplification or capture. (see [0008]-[0009], [0130]-[0133], [0149], and throughout). It is unclear whether “functional” refers to chemical modification that provides nuclease protection (phosphorothioation), chemical modification that signals nuclease activation (phosphorylation), primer-binding capacity (P5 and P7 binding sites), capture affinity (biotinylation), or some combination thereof. As these examples used do not have a consistent structure or function and there is no clear definition, one of ordinary skill in the art would not be able to determine, with reasonable certainty, the metes and bounds of the claims. For example, it is ambiguous whether any primer-binding site qualifies as a “functional sequence,” or whether it must have specific chemical modifications conferring nuclease sensitivity, or whether it is limited to particular adapters such as P5/P7. Where a claim term lacks an established meaning in the art or has a vague, subjective meaning, it is the applicant’s responsibility to clearly define and describe the term in the specification. See In re Morris, 127 F.3d 1048, 1054, 44 USPQ2d 1023, 1027 (Fed. Cir. 1997) and MPEP 2173.02 (claims must particularly point out and distinctly claim the invention).
When a term such as “functional sequence” has no generally accepted scientific definition, and no objective boundaries in the specification, the broadest reasonable interpretation cannot be ascertained with reasonable certainty. The burden is therefore on the applicant to prove an explicit definition or sufficient descriptive support to establish the scope of the term. Absent such clarification the claim fails to satisfy the requirements of 35 U.S.C. 112(b).
Claims 2-47 are likewise rejected due to their dependence on claim 1.
Claim 28 contains the trademark/trade name “MiSeq”, “NexSeq”, “HiSeq”, “NovaSeq”, “iSeq”, “PacBio”, and “SMRT”. 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 different high-throughput sequencing platforms and, accordingly, the identification/description is indefinite.
Claim Interpretation
For the purpose of examination the term “functional sequence” as recited in claim 1 (e.g., “wherein both the artefactual dsDNA library member and the non-artefactual dsDNA library member comprise a first functional sequence at one end and a second functional sequence at the other end”) is interpreted to mean:
A nucleic acid sequence within a library construct that provides a specific, demonstratable function essential to the claimed method – such as primer binding for amplification, adapter binding for sequencing, or chemical modification for selective strand protection – exemplified in the application by P5 and P7 primer/adaptor regions and their described 5’-end chemical modifications (e.g., phosphorylation or phoshorothioation).
This construction is applied to give the broadest reasonable interpretation (BRI) consistent with the specification and to determine whether the prior art teaches or renders obvious the recited features. It is not limited to P5/P7 sequences per se if other art discloses sequences providing the same primer-binding or strand-protection function, but mere presence of a generic nucleic acid sequence without demonstrated function will not meet the limitation.
For purposes of examination, the term “artefactual dsDNA” is interpreted to mean:
Double-stranded DNA molecules present in a sequencing library that are products of library-preparation and do not faithfully correspond to the authentic sequence or orientation of the starting template nucleic acid.
For purposes of examination, the term “neutralizing” as recited in claim 1 (e.g., “neutralizing the second strand of the artefactual dsDNA library member”) is interpreted to mean:
Rendering a specified DNA strand non-functional for downstream amplification, capture, or sequencing, as exemplified in the application by selective nuclease digestion (e.g., lambda or T7 exonuclease) of a strand lacking protective chemical modification, and encompassing other strand-inactivation techniques only to the extent that they are functionally equivalent and would be understood by one of ordinary skill in the art as performing the same inactivating role.
This construction applies the BRI consistent with the specification: it covers any art-recognized method that irreversibly inactivates a DNA strand for library preparation (e.g., selective exonuclease digestion), but does not extend to generic denaturation or transient hybridization steps that fail to achieve permanent inactivation.
Examiners Note: These interpretations inform the subsequent rejections under 35 U.S.C. §§ 102 or 103. Applicant may overcome or narrow this construction by amending the claims or supplying an explicit definition in the specification clarifying the intended scope of “functional sequence” and/or “neutralizing”.
Claim Rejections - 35 USC § 103
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
Claims 1-7, 10, 13- is/are rejected under 35 U.S.C. 103 as being unpatentable over Lai et al. (US 2018/0216103 Al, published Aug. 2, 2018) in view of Agilent SureSelect Product Literature (published Apr. 21, 2017, and July 2020), and Pollner et al. (US 2013/0303382 Al, published Nov. 14, 2013)
The general goal of the present application is the appears to be enriching nucleic acid libraries to be used for sequencing reactions. The present application accomplishes this by combining steps to degrade unwanted strands of nucleic acid followed by using capture probes to pull down nucleic acids from genes of interest. The prior art teaches these methods. Lai is in the field of molecular biology and teaches methods and compositions for sequencing nucleic acids, and enriching libraries for genes of interest. While Pollner and the SureSelect platform marketed by Agilent and Illumina teach enrichment methods involving the bait capture systems for pulling down genes of interest.
Among the methods taught by Lai, they teach a method of preparing a library of nucleic acids, comprising two dsDNA molecules wherein a first strand of one dsDNA comprises an adapter sequence and a sense strand of at least one nucleic acid, and a second strand of the dsDNA molecule comprises a reverse complement of the adapter sequence and an antisense sequence of at least one nucleic acid. Further, they teach the second dsDNA molecule comprise a first stand that comprises the reverse complement of the adapter sequence and a sense sequence of at least one nucleic acid while the second strand comprises the adapter sequence and an anti-sense sequence of at least one nucleic acid (see Fig. 1). Furthermore, Lai teaches that both dsDNA comprise functional sequences, phosphorothioate at the 5’ end of one strand and a primer binding sequence on the 5’ end of the opposite strand. (see Fig. 1, [0016], [0055], [0112], and throughout). Lai also teaches neutralizing a strand of each dsDNA library member using nuclease digestion (see Fig. 1, [0009]-[0010], [0017], and throughout) and after digestion, contacting the nucleotides with a oligonucleotide probe, that has complementarity towards the remaining strands of nucleic acids (see Fig. 1, [0022], [0110], [0114], and throughout).
In regards to claims 2, 13-16, 18-19 , Lai teaches neutralizing a strand from each dsDNA library member using nuclease digestion, such as with lambda exonuclease (see Fig. 1, [0017], [0113]). One of ordinary skill in the art would recognize that lambda exonuclease prefers a phosphorylated 5’ end to initiate digestion, and that this is a function of the enzyme (see NEB Lambda Exonuclease). Lai teaches using phosphorothioate modified P5 and P7 adapters as well as phosphorylated P5 and P7 adapters (see Fig. 1, [0009], [0016], [0112], and throughout). It would have been obvious to one of ordinary skill in the art to modify which adapter was phosphorylated or phosphorothioated in order to preserve and degrade desired strands as taught by Lai.
In regards to claim 3 and 4, Lai teaches adapters may have any sequence and would base-pair with the complementary sequence. (see [0031]). Lai does not expressly state the adapter is poly-A or poly-G as required by claims 3 and 4, respectively. However, Pollner teaches using homopolymeric adapters such a poly-A, or poly-G (see [0010], [0030], [0045], [0069], and throughout). It would have been obvious to one of ordinary skill in the art to utilize homopolymeric adapters sequences in nucleic acid library preparation. Homopolymers tails were a long-established tool in molecular biology, used to facilitate ligation, primer annealing, capture and to protect or cushion junctions. Homopolymeric adapters has been described to aid hybridization, amplification, and allow for sequencing regions of repeats in the genome. Selecting homopolymeric adapters of any desired length or composition would therefore have represented no more than the predictable use of a well-known element for its established function. See MPEP §§ 2141, 2143, 2144; KSR International Co. v. Teleflex Inc. (KSR), 550 U.S. 398, 82 USPQ2d 1385 (2007) (combining familiar elements according to known methods is likely to be obvious when it yields no more than predictable results).
In regards to claim 5, Lai teaches amplifying the dsDNA strands using primer adapters that produce functional sequences at the ends of the strands, specifically siting using phosphorylated primers and those with phosphorothioate modified nucleotides (see Fig. 1, [0009], [0012], [0016], and throughout).
In regards to claim 6-12, Lai teaches using primers that comprise P5 and P7 sequences (see Fig. 1, [0008]-[0013], [0110], and throughout). Furthermore, Lai teaches using these primers configured so that one phosphorylated of phophorothioated to produce the functional end chemistries desired to either signal nuclease digestion or protect the strand from digestion. (see Fig. 1, [0016], [0110]-[0112]). This can be adjusted to preserve either 3’ or 5’ end chemistries of the original dsDNA molecules as desired based on adapter binding sequence of interest.
In regards to claim 17 and 20, while Lai teaches exonuclease digestion from 5’ – 3’, they recited using lambda exonuclease and do not specifically teach using T7 exonuclease. However, one of ordinary skill in the art would recognize that both lambda and T7 function similarly, with T7 exonuclease having a wider temperature range and different salt concentration requirements. Further, one of ordinary skill in the art would recognize that phosphorothioate modified residues adjacent to the 5’ end protect the end from degradation and that the opposite strand would be the one degraded by exonuclease. It would have been prima facie obvious to one of ordinary skill in the art at the time of filing to substitute T7 exonuclease for lambda exonuclease in enriched library development. One would be motivated to use T7 because it is less active on single stranded DNA, which is important if the single-stranded DNA is the desired product. One of ordinary skill in the art would swap these exonucleases based on assay conditions and availability and expect similar or improved result with the use T7 exonuclease (NEB “Properties of Exonucleases and Non-specific Endonucleases”).
In regards to claims 21 and 22, Lai teaches generating an enriched library using capture probe oligonucleotides comprise an amplification sequence (S3) and a sequence of corresponding to the DNA of interest but does not specify the length of the sequence, and that these may be of any length (see [0014], [0054]). However, varying the lengths of capture oligonucleotides is well known in the art, Albert teaches use of capture oligonucleotide that are greater than >60 nucleotides in length (see pg. 903, right col. 2nd para.). It would have been obvious to a person of ordinary skill in the art to select capture oligonucleotides of about 120 nt in length utilizing Lai’s method, because Albert, and contemporary commercial kits such as Agilent’s SureSelect® and Roche’s HyperCap® established that capture probes of approximately 120 bp were widely adopted before the effective filing date. (see Agilent and Roche manuals).
In regards to claim 23, Lai teaches that the capture probe may be biotinylated (see [0055]).
In regards to claim 24, while Lai teaches that the capture probe is biotinylated, they never detail the actual isolation steps involved with the capture probe, only causally referencing the capture step (see Fig. 2). The Agilent SureSelect literature teaches biotin pulldown. It would have been obvious to a person of ordinary skill in the art to immobilize or “pull down” biotinylated capture oligonucleotides using avidin/streptavidin or other functional analogs (e.g. neutravidin) or binding partners (e.g. carboxylases) on a solid support (e.g. magnetic bead or coated surface). The biotin-(strept)avidin affinity pair has been well-established routinely used capture chemistry for decades in nucleic acid and protein workflows, providing specific, high-affinity binding and facile separation under standard conditions. Using the known, art-accepted binding partner (avidin/streptavidin) to capture biotinylated oligonucleotides amounts to the predictable use of prior-art elements according to their established functions to obtain predictable results (immobilization or separation), and thus would have been obvious next step with the capture oligonucleotides to generate an enriched library. See MPEP §§ 2141, 2143, 2144; KSR International Co. v. Teleflex Inc. (KSR), 550 U.S. 398, 82 USPQ2d 1385 (2007) (combining familiar elements according to known methods is likely to be obvious when it yields no more than predictable results).
In regards to claim 25-28, Lai teaches sequencing the enriched library of nucleic acids using next generation sequencing methods (Abstract, [0050], and throughout). Lai further teaches using P5 and P7 adapters, which are used as priming sites for paired-end sequencing in Illumina platforms, such as MiSeq®, NexSeq®, and HiSeq® platforms (see Fig. 1, [0008]-[0012], and throughout).
In regards to claim 29-32, Lai teaches using libraries that comprise either a single library or pooled libraries with different barcodes to separate them after sequencing (see [0033]). It would have been obvious to a person of ordinary skill in the art to combine any number of separately prepared libraries for sequencing into a single mixture for downstream processing or sequencing. At the time of the invention, pooling multiple libraries, commonly called multiplexing, was a routine and well-understood practice in next-generation sequencing. Standard methods allowed researchers to mix libraries in any desired quantity, provided that each library carried unique index or barcode sequences to permit demultiplexing. Whether a researcher chose to pool two, ten, or hundreds of libraries would have been a matter of obvious optimization of throughput, cost, and experimental design, yielding only predictable results. See MPEP §§ 2141, 2143, 2144; KSR International Co. v. Teleflex Inc. (KSR), 550 U.S. 398, 82 USPQ2d 1385 (2007). Accordingly, it would have been obvious to combine sequencing libraries in any quantity for simultaneous processing or analysis.
In regards to claim 33-40, while Lai references examining different genes associated with many different diseases, they do not specifically recite looking specifically at a set predetermined panel of genes. However, Agilent SureSelect offers both pre-designed panels and customizable panels of genes. The pre-designed panels from Agilent cover cancer associated variants, gene signature panels, and includes their exome kit covering ~4800 genes while their cancer All-in-one assay panel consisted of 118 genes. Agilent SureSelect customizable panels allow user to choose up to 60K unique 120-mer probes. User can easily customize that to comprise between 100 and 2000 genes or more than 1500 genes. See SureSelect NGS Panels pamphlet that outlines various panels between 100 to 2000 genes or more than 1500 genes.
In regards to claims 41-47, the kits merely recite packaging the reagents (e.g. primers, adapters, enzymes, buffers, capture probes) necessary to practice the rejected methods together in a commercial kit. It would have been obvious toa person of ordinary skill in the art at the time of filing to provide such reagents as a kit since supplying reagents needed to carry out a known method in a kit form is nothing more than routine product packaging and marketing, well established in biotechnology and sequencing arts. See MPEP 2111.05(III), providing the known reagents together as a kit does not impart patentable distinction over the method itself and therefore would have been obvious.
Conclusion
No claim is allowed.
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/MATTHEW HAROLD RAYMONDA/Examiner, Art Unit 1684 /AARON A PRIEST/Primary Examiner, Art Unit 1681