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 .
Claims 1-20 are pending. Claim 14 is amended. Claims 19-20 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim. Claims 1-18 are currently under examination.
Response to Amendment
The Amendment filed 4/24/26 has been entered. Claims 1-20 are pending. Applicant’s amendment of claim 14 has overcome the 112(b) rejection previously set forth in the Non-Final Office Action mailed 1/27/26.
Response to Arguments
Applicant’s arguments, see pages 1-3, filed 4/24/26, with respect to the rejections of claims 1-18 under 35 USC 103 have been fully considered are found unpersuasive, and the rejections documented in the Non-Final mailed 1/27/26 have been revised to address claim amendments filed 4/24/26 in this Final Office Action. More detailed responses to Applicant’s arguments are provided at the end of each maintained rejection.
Claim Rejections - 35 USC § 103
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.
Claims 1-17 remain/are rejected under 35 U.S.C. 103 as being unpatentable over Weng et al. (2020; US 2020/0080141 A1; USPGPub citation A in PTO-892 filed 1/27/26).
This 103 rejection is revised/updated in response to claim amendments filed 4/24/26.
Relevant to claim 1, Weng et al. teaches "Recognized herein is a need for alternative and/or robust methods and compositions of detecting rare sequence variations, particularly rare sequence changes and gene fusion events. The compositions and methods of the present disclosure address this need, and provide additional advantages as well" (paragraph 0007).
This teaching reads on claim 1 A method for detecting a fusion gene in a sample from a subject, said method comprising.
Further relevant to claim 1, Weng et al. teaches "In an aspect, a method for enriching amplicons comprising a concatemer of at least two or more copies of a target polynucleotide is disclosed. The method comprises (a) generating a concatemer comprising a single-stranded polynucleotide from a circular target polynucleotide by extension of a first primer" (paragraph 0009).
This teaching reads on claim 1 circularizing a fusion linear nucleic acid molecule of the sample to form a fusion gene circular template polynucleotide, wherein said fusion linear nucleic acid molecule comprises a fusion gene… hybridizing a first primer to said fusion gene circular template polynucleotide and extending the first primer with a polymerase thereby generating a first fusion extension product.
Further relevant to claim 1, Weng et al. teaches "In another aspect, the disclosure provides a method of identifying a sequence variant, such as in a nucleic acid sample. In some embodiments, each polynucleotide of the plurality has a 5' end and a 3' end, and the method comprises: (a) circularizing individual polynucleotides of said plurality to form a plurality of circular polynucleotides, each of which having a junction between the 5' end and 3' end" (paragraph 0010).
The skilled artisan would recognize that not all of the polynucleotides of the plurality would each contain the target fusion gene, thus allowing for circularization of non-target (or non-fusion) nucleic acid molecules. This is supported by the Weng et al. teachings of "FIGS. 18A and 18B illustrate results of an analysis of ligation efficiency and on-target rate of an embodiment of the disclosure" (paragraph 0050) and "As shown in FIG. 18B, the amplification curves of 10 ng of WGA product and reference genomic DNA (gDNA) (12878, 10 ng) virtually overlap with each other. The average Ct for the WGA sample was 26.655, while that of the gDNA sample was 26.605, indicating a high on-target rate of over 96%" (paragraph 0215). The non-100% rate of 96% indicates that non-target (such as non-fusion) nucleic acid molecules are included within the methodology.
These teachings read on claim 1 circularizing a non-fusion linear nucleic acid molecule of the sample to form a non-fusion circular template polynucleotide, wherein said non-fusion linear nucleic acid molecule does not comprise the fusion gene.
Further relevant to claim 1, Weng et al. teaches "FIGS. 6A and 6B illustrate further non-limiting example methods of circularizing polynucleotides, such as single-stranded DNA. The adapter can be asymmetrically added to either the 5' or 3' end of a polynucleotide. As shown in FIG. 6A, the single-stranded DNA (ssDNA) has a free hydroxyl group at the 3' end, and the adapter has a blocked 3' end such that in the presence of a ligase, a preferred reaction joins the 3' end of the ssDNA to the 5' end of the adapter" (paragraph 0090).
This teaching reads on claim 1 binding a blocking element to said non-fusion circular template polynucleotide.
Further relevant to claim 1, Weng et al. teaches "(b) generating a plurality of extension products containing one or more copies of the target polynucleotide by extension of a second primer comprising a second 3' end that specifically hybridizes to the concatemer" (paragraph 0009).
This teaching reads on claim 1 hybridizing a second primer to said first extension product and extending said second primer with a polymerase thereby generating a second extension product.
Further relevant to claim 1, Weng et al. teaches "In some embodiments, the method further comprises sequencing the plurality of amplicons produced in step (c)" (paragraph 0009).
This teaching reads on claim 1 sequencing the second extension product, or a complement thereof, thereby detecting the fusion gene.
Relevant to claims 2-3, Weng et al. teaches "According to some embodiments, polynucleotides among the plurality of polynucleotides from a sample are circularized. Circularization can include joining the 5' end of a polynucleotide to the 3' end of the same polynucleotide, to the 3' end of another polynucleotide in the sample, or to the 3' end of a polynucleotide from a different source (e.g. an artificial polynucleotide, such as an oligonucleotide adapter)" (paragraph 0083).
Relevant to claim 2, Weng et al. teaches "(b) a first primer comprising sequence A', wherein the first primer specifically hybridizes to sequence A of the target sequence via sequence complementarity between sequence A and sequence A'…(d) a polymerase that extends the first primer and the second primer to produce amplified polynucleotides; wherein the distance between the 5' end of sequence A and the 3' end of sequence B of the target sequence is 75 nt or less" (paragraph 0012).
The skilled artisan would recognize that the blocking element (Weng et al. oligonucleotide adapter with a blocked end, see rejection of claim 1) bound to the end of the sequence would be within 75 nt or less distance from the first primer (Weng et al. A/A' region).
Relevant to claim 4, Weng et al. teaches "(c) amplifying the plurality of extension products of step (b) under conditions to generate a plurality of amplicons, wherein amplicons comprising at least 2 or more copies of the target polynucleotide are enriched" (paragraph 0009).
Further relevant to claim 4, Weng et al. teaches "In some embodiments, amplifying is effected by using a polymerase having strand-displacement activity, such as in rolling-circle amplification (RCA)" (paragraph 0011).
Relevant to claim 5, Weng et al. teaches "The target sequence may be a portion of a gene, a regulatory sequence, genomic DNA…" (paragraph 0070).
Relevant to claim 6, Weng et al. teaches "Where polynucleotides are isolated from a sample without a cellular extraction step, polynucleotides will largely be extracellular or 'cell-free' polynucleotides, which may correspond to dead or damaged cells" (paragraph 0080).
Relevant to claims 7 and 9, Weng et al. teaches "A sample may be a fresh sample, or a sample subjected to one or more storage processes (e.g. paraffin-embedded samples, particularly formalin-fixed paraffin-embedded (FFPE) sample)" (paragraph 0158).
Relevant to claim 8, Weng et al. teaches "Examples of cancers that may be diagnosed based on calling one or more sequence variants in accordance with a method disclosed herein include… Hodgkin Lymphoma, Hodgkin's lymphoma…" (paragraph 0174).
Relevant to claims 10-12, Weng et al. teaches "A variety of methods for circularizing polynucleotides are available. In some embodiments, circularization comprises an enzymatic reaction, such as use of a ligase (e.g. an RNA or DNA ligase). A variety of ligases are available, including, but not limited to… [various ligases]" (paragraph 0087).
Relevant to claims 13-14, Weng et al. teaches "According to some embodiments, circularized polynucleotides (or amplification products thereof, which may have optionally been enriched) are subjected to a sequencing reaction to generate sequencing reads… In some embodiments, sequencing comprises a sequencing by synthesis process, where individual nucleotides are identified iteratively, as they are added to the growing primer extension product. Pyrosequencing is an example of a sequence by synthesis process that identifies the incorporation of a nucleotide by assaying the resulting synthesis mixture for the presence of by-products of the sequencing reaction, namely pyrophosphate. In particular, a primer/template/polymerase complex is contacted with a single type of nucleotide. If that nucleotide is incorporated, the polymerization reaction cleaves the nucleoside triphosphate between the a and 13 phosphates of the triphosphate chain, releasing pyrophosphate. The presence of released pyrophosphate is then identified using a chemiluminescent enzyme reporter system that converts the pyrophosphate, with AMP, into ATP, then measures ATP using a luciferase enzyme to produce measurable light signals. Where light is detected, the base is incorporated, where no light is detected, the base is not incorporated" (paragraph 0102).
Relevant to claims 15-17, Weng et al. teaches "In some embodiments of any of the various aspects disclosed herein, the methods, compositions, and systems have therapeutic applications, such as in the characterization of a patient sample and optionally diagnosis of a condition of a subject. Therapeutic applications may also include informing the selection of therapies to which a patient may be most responsive (also referred to as 'theranostics'), and actual treatment of a subject in need thereof, based on the results of a method described herein… In some embodiments, a subject is monitored for treatment efficacy. For example, by monitoring ctDNA over time, a decrease in ctDNA can be used as an indication of efficacious treatment, while increases can facilitate selection of different treatments or different dosages" (paragraph 0154).
Weng et al. does not teach a specific embodiment having all the claimed elements. That being said, however, it must be remembered that "[w]hen a patent simply arranges old elements with each performing the same function it had been known to perform and yields no more than one would expect from such an arrangement, the combination is obvious." KSR v. Teleflex, 127 S.Ct. 1727, 1740 (2007) (quoting Sakraida v. AG. Pro, 425 U.S. 273, 282 (1976)). "[W]hen the question is whether a patent claiming the combination of elements of prior art is obvious," the relevant question is "whether the improvement is more than the predictable use of prior art elements according to their established functions." (Id.). Addressing the issue of obviousness, the Supreme Court noted that the analysis under 35 USC 103 "need not seek out precise teachings directed to the specific subject matter of the challenged claim, for a court can take account of the inferences and creative steps that a person of ordinary skill in the art would employ." KSR at 1741. The Court emphasized that "[a] person of ordinary skill is... a person of ordinary creativity, not an automaton." Id. At 1742.
Consistent with this reasoning, it would have been prima facie obvious to have selected various combinations of various disclosed elements — including blocking elements, ligases, amplification, and sequencing — for a method for detecting a fusion gene in a sample from a subject, to arrive at compositions "yielding no more than one would expect from such an arrangement."
Applicant’s Arguments and Response to Applicant’s Arguments
Applicant argues that “Weng does not teach or suggest selectively circularizing fusion vs. non-fusion molecules separately, or binding a blocking element to non-fusion circles. Weng also does not suggest a strategy to suppress non-fusion circular templates in order to enrich fusion molecules” (Remarks 4/24/26, page 2, paragraph 1).
The Examiner respectfully disagrees with these assertions.
In response to applicant's argument that the references fail to show certain features of the invention, it is noted that the features upon which applicant relies (i.e., selectively circularizing and enriching for fusion molecules) are not recited in the rejected claim(s). Although the claims are interpreted in light of the specification, limitations from the specification are not read into the claims. See In re Van Geuns, 988 F.2d 1181, 26 USPQ2d 1057 (Fed. Cir. 1993).
The claim, as written, does not explicitly require the argued “selectively circularizing” or enrichment of fusion molecules. Thus, Weng et al. disclosure of circularizations of both fusion and non-fusion linear nucleic acid molecules (FIGS. 18A and 18B; paragraphs 0050 and 0215) fulfills the claim 1 limitation of circularizing a fusion linear nucleic acid molecule of the sample to form a fusion gene circular template polynucleotide… circularizing a non-fusion linear nucleic acid molecule of the sample to form a non-fusion circular template polynucleotide.
As stated in the above rejection of claim 1 (and repeated from the Non-Final rejection mailed 1/27/26) “The non-100% rate of 96% indicates that non-target (such as non-fusion) nucleic acid molecules are included within the methodology.” Thus, claim 1 limitation of binding a blocking element to said non-fusion circular template polynucleotide would be read upon the Weng et al. methodology which teaches blocking elements (FIG. 6A and paragraph 0090), which would also be performed upon the non-fusion circular template polynucleotides present within the workflow.
Applicant further argues that “Weng likewise fails to teach or suggest a two-primer sequential extension scheme specific to fusion circles, or any approach for enriching fusion sequences by differential primer-specific extension before sequencing… each hybridization [of first and second primers to circularized polynucleotides] is not followed by extension” (Remarks 4/24/26, page 2, paragraph 2).
The Examiner respectfully disagrees with these assertions.
As stated in above rejection of claim 1 and reiterated from the 1/27/26 Non-Final rejection, Weng et al. teaches "In an aspect, a method for enriching amplicons comprising a concatemer of at least two or more copies of a target polynucleotide is disclosed. The method comprises (a) generating a concatemer comprising a single-stranded polynucleotide from a circular target polynucleotide by extension of a first primer" (paragraph 0009). This teaching provides the argued extension from the first primer. Additionally, Weng et al. teaches "(b) generating a plurality of extension products containing one or more copies of the target polynucleotide by extension of a second primer comprising a second 3' end that specifically hybridizes to the concatemer" (paragraph 0009). This teaching provides the argued extension from the second primer. The argued primer-based enrichment limitation is not explicitly required in the claim 1 language, similarly to the above response to selectivity/enrichment argument.
The Applicant further argues that “To render the present method obvious, Weng would at the very least need to provide motivation to 1) distinguish fusion vs. non fusion molecules, 2) selectively block non fusion circles, and, 3) perform sequential primer extensions to isolate a fusion-specific junction” (Remarks 4/24/26, page 2, paragraph 3).
The Examiner respectfully disagrees with these assertions. As written, the claims do not require distinguishing/selectivity/enrichment (as discussed above). Instead, as stated in the above rejection and reiterated from the 1/27/26 Non-Final Rejection, the instant patent “simply arranges old elements with each performing the same function it had been known to perform” from the disclosure of Weng et al. The skilled artisan is “a person of ordinary creativity, not an automaton” and would find it obvious and reasonable to select previously disclosed elements from the Weng et al. disclosure to arrive at the instantly rearranged combination of old elements.
Claim 18 remains/is rejected under 35 U.S.C. 103 as being unpatentable over Weng et al. (2020; US 2020/0080141 A1; USPGPub citation A in PTO-892 filed 1/27/26) as applied to claims 1-17 above, and further in view of Gu et al. (2016; NPL citation U in PTO-892 filed 1/27/26; "Depletion of Abundant Sequences by Hybridization (DASH): using Cas9 to remove unwanted high-abundance species in sequencing libraries and molecular counting applications"; Genome Biology 17:41; DOI 10.1186/s13059-016-0904-5).
The teachings of Weng et al. are applied to instantly rejected claim 18 as they were applied to claims 1-17 as rendering obvious a method for detecting a fusion gene in a sample from a subject.
Weng et al. is silent to specifics regarding a CRISPR-Cas9 complex with a guide RNA molecule blocking element. However, these limitations were known in the prior art and taught by Gu et al.
Relevant to claim 18, Gu et al. Abstract teaches "Next-generation sequencing has generated a need for a broadly applicable method to remove unwanted high-abundance species prior to sequencing. We introduce DASH (Depletion of Abundant Sequences by Hybridization). Sequencing libraries are ‘DASHed’ with recombinant Cas9 protein complexed with a library of guide RNAs targeting unwanted species for cleavage, thus preventing them from consuming sequencing space… We also demonstrate an application of DASH in cancer. This simple method can be adapted for any sample type and increases sequencing yield without additional cost."
Although Weng et al. does not include a CRISPR-Cas9 complex with a guide RNA molecule blocking element, it would have been prima facie obvious to the skilled artisan. It is noted that Weng et al. and Gu et al. are analogous disclosures to the instant method of nucleic acid detection.
The skilled artisan would be motivated to include the Gu et al. CRISPR-Cas9 complex with a guide RNA molecule blocking element within the Weng et al. methodology because Weng et al. allows for enzymatic removal of blocking moieties ("Once the linear ligation is accomplished, the ligated pieces can be treated with an enzyme to remove the blocking moiety, such as through the use of a kinase or other suitable enzymes or chemistries" (Weng et al. paragraph 0090)) and Gu et al. Abstract teaches that the CRISPR-Cas9 complex with a guide RNA molecule blocking element is a “simple method can be adapted for any sample type and increases sequencing yield without additional cost.”
The skilled artisan would have a reasonable expectation of success based on the disclosure of Weng et al., and further in view of Gu et al., as discussed in the preceding paragraphs.
Applicant’s Arguments and Response to Applicant’s Arguments
Applicant argues that “Gu only applies to dependent claim 18 and does not correct any of the above-noted deficiencies” (Remarks 4/24/26, page 2, paragraph 4). The Examiner does not require Gu et al. to remedy the argued deficiencies of Weng et al., and directs Applicant to the response to the argued deficiencies above.
Conclusion
THIS ACTION IS MADE FINAL. Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
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/SARAH JANE KENNEDY/Examiner, Art Unit 1682
/WU CHENG W SHEN/Supervisory Patent Examiner, Art Unit 1682
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