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 .
Please note: The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action.
Interview Summary
Applicant’s submission of a summary of the interview of 10/20/2025 on page 11 of the Remarks of 1/8/2026 is acknowledged.
Claim Status
Claims 1, 3, 4, 7, 12, 14, 17, 23, 30, 45, 53, 56, 58, 79-81, and 83-86 are pending and being examined on the merits.
Abstract
The objection to the abstract of the disclosure is withdrawn in light of Applicant’s amendment to the Abstract to correct a typo.
Specification
The objection to the disclosure is withdrawn in light of Applicant’s amendments to the specification.
Applicant’s amendments to the specification to properly denote trade names or marks used in commerce is acknowledged.
Claim Objections
The objection of claim 79 as being a substantial duplicate thereof of claim 1 is withdrawn given Applicant’s amendments to the claims.
New Claim Rejections - 35 USC § 101
Necessitated by Amendments
35 U.S.C. 101 reads as follows:
Whoever invents or discovers any new and useful process, machine, manufacture, or composition of matter, or any new and useful improvement thereof, may obtain a patent therefor, subject to the conditions and requirements of this title.
Claims 1, 3, 4, 7, 12, 14, 17, 23, 30, 45, 53, 56, 58, 79-81, and 83-86 are rejected under 35 U.S.C. 101 because the claimed invention is directed to abstract ideas (e.g.: mental processes, mathematical calculations) without significantly more.
The claim(s) recite(s) methods of verifying a specific mutation (as recited in claims 1, 79, and 84), applying a noise filter (claim 79), and excluding a specific mutation (claim 84). The claims are thus directed to the assessment of collected data, which is an abstract idea that is a mental process (e.g.: MPEP 2106.04(a (2)(III)(A)); it is the observation and evaluation of information to reach a judgment or conclusion.
This judicial exception is not integrated into a practical application because there are no practical steps related to the verification/exclusion of the presence of a specific mutation. There are no additional steps of the claims that are directed to applying or using the judicial exception(s) noted above (e.g.: MPEP 2106.04(d)(I)). The dependent claims further relate to extra solution activity regarding lengths and composition of probes, temperatures of annealing, threshold requirements, and further steps of the method that are not practical integration of the judicial exceptions.
The claim(s) does/do not include additional elements that are sufficient to amount to significantly more than the judicial exception because the claims only broadly recite steps of performing duplex sequencing on an enriched sample (attaching UMIs to 5’ and 3’ ends of DNA duplexes in a pool of duplexes, amplifying the tagged duplexes, denaturing the tagged duplexes, capturing target DNA with target-specific probes, and sequencing said captured targets. However, such steps were well understood, routine and convention in the prior art (e.g.: MPEP 2106.05(d)). For example: Kennedy (2014) teaches using duplex sequencing to examine mutations in enriched sequences involving the above listed steps; Jeffreys (2003) and Zhang (2019) teach the use of allele-specific probe hybridization to enrich samples for specific mutations. Additionally, the specification teaches that the process of sequencing using UMIs is “well established in the art” (paragraph [0099]), as are “methods for determining a consensus sequence” (paragraph [0108]) and amplification by PCR (paragraph [0100]).
Claim Rejections - 35 USC § 112b - Indefiniteness
Withdrawn: The rejection of claims 12, 17, and 23 under 35 U.S.C. 112(b) are withdrawn in light of Applicant’s amendments to the claims.
New (necessitated by amendments): Claims 79-81, 83, and 86 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 79 is directed to a method wherein a specific mutation is verified by “applying a locus specific noise filter that includes a threshold for a double-stranded consensus to single-stranded consensus ration (DSC to SSC ratio)”. However, it is unclear how this threshold is being applied to verify or exclude specific mutations. Does the DSC to SSC ratio need to be above or below said threshold? Additionally, it is unclear what all a locus specific noise filter entails in addition to the included threshold. Paragraph [0184] indicates that the locus specific noise filter is the application of a threshold regarding the DSC to SSC ratio, not that it merely includes said ratio threshold.
Claims 80-81, 83, and 86 depend from claim 79, inherit these deficiencies, and are rejected on the same basis. While claim 86 corrects the deficiency regarding the application of the threshold, it does not rectify the indefiniteness of the locus specific noise filter.
Withdrawn Claim Rejections – 35 USC § 103
The rejections of claims 1, 3-4, 7, 14, 53, 56, 58 and 79-85 under 35 U.S.C. 103 as being unpatentable over Kennedy (Kennedy et al.; Nature Protocols, 2014) in view of Jeffreys (Jeffreys et al.; Genome Research, 2003), claims 12, 17, 23, and 30 under 35 U.S.C. 103 as being unpatentable over Kennedy (Kennedy et al.; Nature Protocols, 2014) in view of Jeffreys (Jeffreys et al.; Genome Research, 2003) as applied to claims 1, 3-4, 7, 14, 53, 56, 58 and 79-85 above, and further in view of Zhang (Zhang et al., US 2019/0185933 A1), and claim 45 under 35 U.S.C. 103 as being unpatentable over Kennedy (Kennedy et al.; Nature Protocols, 2014) in view of Jeffreys (Jeffreys et al.; Genome Research, 2003) as applied to claims 1, 3-4, 7, 14, 53, 56, 58 and 79-85 above, and further in view of Maguire (Maguire et al., US 2017/0275689 A1) are withdrawn in light of Applicant’s amendments to the claims.
New Claim Rejections - 35 USC § 103
Necessitated by Amendments
Claims 1, 3-4, 7, 14, 53, 56, 58 and 79-81, and 83-86 are rejected under 35 U.S.C. 103 as being unpatentable over Kennedy (Kennedy et al.; Nature Protocols, 2014; cited on PTO-892 of 8/8/2025) in view of Wan (Wan et al., bioRxiv 2019; cited on IDS of 3/13/2025) and Jeffreys (Jeffreys et al.; Genome Research, 2003; cited on PTO-892 of 8/8/2025).
Claims 1, 4, 79, and 83-86: Kennedy teaches obtaining a pool of DNA duplexes and attaching UMIs to the 5’ and 3’ ends of the duplex DNA such that the UMIs are unique to each tagged duplex (claims 1a-b, claim 79 lines 2-5, claim 84 lines 2-5; “label each fragmented DNA molecule with its own unique DNA sequence” reads on UMIs are unique to each tagged duplex; The concept of DS, Figure 1). Kennedy teaches amplifying by polymerase chain reactions the tagged duplexes to generate amplified tagged duplexes (claim 1c, claim 79 line 6, claim 84 line 6; Figure 1b). Kennedy teaches sequencing the amplified sample and identifying the presence of a mutation is the mutation is observed in both strands of a given tagged duplexes identified by the UMIs (claim 1f-g, claim 79 lines 12-15, claim 84 lines 12-14; The concept of DS, Figure 1).
Kennedy teaches that the theoretical ideal is that in which the double-stranded consensus to single-stranded consensus ratio is 0.5 (“The SSCS:DCS ratio is the other key factor. A ratio of two is the theoretically ideal and would indicate that every SSCS can find its partner and form a DCS”, Experimental Design - Quality Metrics; SSCS (single-strand consensus sequence) and DCS (duplex consensus sequence) read on single-stranded consensus and double-stranded consensus, respectively). Kennedy teaches that they regularly achieve an SSCS:DCS ratio of 4-10, with an average of 6 (which would be a range of double-stranded consensus to single-stranded consensus ratio of 0.25-0.1 and an average of 0.17). Kennedy teaches that the “optimal peak family size” is six SSCSs to one DCS, which is a DCS:SSCS ratio of 0.17.
Kennedy does not teach using this optimal peak family size ratio as a threshold for discriminating between artifacts and true mutations (“verifying”) to distinguish noisy loci from non-noisy loci (claims 1h, 79, and 83-86). However, use of locus specific noise filters in addition to DCS to remove artifactual errors is known in the art, as taught by Wan.
Wan teaches a method of identifying mutations in a DNA sample through patient-specific mutation hybridization panels (pg 3 ln 87-91 – pg 4 ln 92-95 and Abstract). Wan teaches a method of error-suppression in which mutation signal must be present in both the forward and reverse read of the read pair (double-strand consensus; pg 5, ln 148-150). Wan teaches that in addition to this, using targeted hybridization can enrich for “technically noisy sites” and thus uses an additional locus noise filter threshold, wherein only loci that were below the threshold were included in further analysis to remove any artefactual mutations (pg 6, ln 153-162).
It would have been prima facie obvious to one having ordinary skill in the art, before the effective filing date of the instant application, to have modified the method of Kennedy to use a locus specific noise filter as taught by Wan. One would be motivated to use a locus specific noise filter given the assertion by Wan that combining this with a DCS produces a synergistic effect in reducing background error rates. Given the teaching by Kennedy that an optimal peak family size of six SSCS to one DCS indicates high quality, one would be motivated to use this methodology to call true mutations in the context of DCS:SSCS ratios only for loci that are at or above this quality threshold (0.17, which reads on at least 0.15; claims 1, 4, 79 and 83-86).
Kennedy in view of Wan teach enriching the DNA via hybridization-based target capture to analyze specific regions of the genome (Targeted DNA capture). However, Kennedy in view of Wan does not explicitly teach the use of an allele-specific probe for capturing a specific mutation to perform this enrichment step (claim 1d-e, claim 79 lines 7-11, and claim 84 lines 7-11). However, use of allele-specific probes that anneal to specific mutations to enrich a sample for target mutations is known in the art, as taught by Jeffreys.
Jeffreys teaches a method in which a single nucleotide variant (mutation) is enriched by hybridization using a biotinylated allele-specific oligonucleotide and hybrid capture with streptavidin-coated beads to generate an enriched sample (Abstract and Figure 1). Jeffreys teaches that the target DNA is denatured prior to hybridization with the allele-specific probes (Figure 1a and Results-The Principles of DEASH).
It would have been prima facie obvious to one having ordinary skill in the art, before the effective filing date of the instant application, to have modified the method of Kennedy in view of Wan to use allele-specific probes for target enrichment as taught by Jeffreys. One would be motivated to use allele-specific probes given the assertion by Jeffreys that their method allows detection of “DNA molecules containing a single base change at a frequency of 10-5 per cell” and allows for detection of “very rare de novo base substitutions” (Abstract). One would have a reasonable expectation of success given that Kennedy in view of Wan accounts for an enrichment step via hybridization to complementary probes (Targeted DNA capture) and Jeffreys demonstrates successful enrichment of target SNPs in human DNA samples using allele-specific probes (Figure 4).
Claims 3, 7, and 80: Jeffreys teaches hybridizing the allele-specific probes to target DNA at an “optimal annealing temperature determined empirically” and specifically demonstrates results of annealing temperatures of 35˚C-50˚C (Table 1). Jeffreys also teaches that enriching the sample using the allele-specific probes comprises recovering the allele-specific probe (The Principles of DEASH, Figure 1). Jeffreys teaches using allele-specific probes that are 18 nucleotides long (reads on about 10 to about 60, about 15 to about 50, about 20 to about 40).
It is noted that the courts have held that optimization through routine experimentation, and in particular, when related to differences in temperature, do not support patentability (In re Aller, 220 F.2d 454, 456, 105 USPQ 233, 235 (CCPA 1955)). Thus, the claimed temperature range merely represents routine optimization of the teachings of the cited prior art. Additionally, the claimed length ranges merely represent routine optimization of the teachings of the cited prior art.
Applicant is advised that MPEP 716.01(c) makes clear that “[t]he arguments of counsel cannot take the place of evidence in the record” (In re Schulze, 346 F.2d 600, 602, 145 USPQ 716, 718 (CCPA 1965)). Thus, Applicant should not merely rely upon counsel’s arguments in the place of evidence in the record.
Claim 14: Jeffreys teaches performing multiple rounds of enrichment on the enriched sample (“re-enrichment”). Jeffreys demonstrates a 100-fold enrichment with two rounds of enrichment with allele-specific probes and teaches that there is “no reason why additional rounds of purification [enrichment] should not increase sensitivity even further” (page 2322, col 2). Therefore, it is reasonable for this art to read on “repeated on the enriched sample at least 10 times” relative to a control (in which enrichment is not performed or is only performed once).
It is noted that the courts have found that “where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation.” In re Aller, 220 F.2d 454, 456, 105 USPQ 233, 235 (CCPA 1955). See MPEP 2144.05 II. Applicant is again advised to not merely rely upon counsel’s arguments in place of evidence of the record.
Claim 53: Jeffreys teaches an 18 nucleotide long allele-specific probe in which the nucleotide complementary to the specific mutation is 11 nucleotides from the 5’ end. Therefore, this reads on the complementary nucleotide being in the middle 50% and in the middle 34%.
It is again noted that the courts have held that optimization through routine experimentation does not support patentability. Thus, the claimed locations of the complementary nucleotides merely represent routine optimization of the teachings of the cited prior art. Applicant is again advised to not merely rely upon counsel’s arguments in place of evidence of the record.
Claim 56: Jeffreys teaches 18 nucleotide long allele-specific probes (see rejections of claims 7 and 82 above) which exhibit a range of annealing temperatures between 35˚C-50˚C (Table 1). Given that Jeffreys teaches allele-specific probes with overlapping annealing temperatures (as noted in the rejection of claims 3 and 80 above) and similar lengths it would be routine optimization to determine an ideal range of Gibbs free energy and these probe structures would likely inherently have a similar range of Gibbs free energy given their overlap with the structures taught by the claims.
It is again noted that the courts have held that optimization through routine experimentation does not support patentability. Thus, the claimed free energies of the allele-specific probes merely represents routine optimization of the teachings of the cited prior art. Applicant is again advised to not merely rely upon counsel’s arguments in place of evidence of the record.
Claim 58: While Kennedy in view of Wan and Jeffreys do not explicitly teach that hybridization probes (in the case of Kennedy) or allele-specific probes (in the case of Jeffreys) are 100% homologous with less than 10 or less than 5 sequences of a reference genome, it would be obvious to one skilled in the art to optimize probe sequences in a way that would only enrich target sequences, especially target sequences containing a specific mutation. Ensuring lack of homology with other regions of a reference genome amounts to routine optimization without demonstration or evidence of unexpected results.
Claim 81: Jeffreys teaches that enriching the sample using the allele-specific probes comprises recovering the allele-specific probe (The Principles of DEASH, Figure 1).
Claims 12, 17, 23, and 30 are rejected under 35 U.S.C. 103 as being unpatentable over Kennedy (Kennedy et al.; Nature Protocols, 2014; cited on PTO-892 of 8/8/2025) in view of Wan (Wan et al., bioRxiv 2019; cited on IDS of 3/13/2025) and Jeffreys (Jeffreys et al.; Genome Research, 2003; cited on PTO-892 of 8/8/2025) as applied to claims 1, 3-4, 7, 14, 53, 56, 58 and 79-81, and 83-86 above, and further in view of Zhang (Zhang et al., US 2019/0185933 A1; cited on PTO-892 of 8/8/2025).
The teachings of Kennedy in view of Wan and Jeffreys as applied to claims 1, 3-4, 7, 14, 53, 56, 58 and 79-81, and 83-86 are detailed above. Relevant to the instantly rejected claims, Kennedy in view of Wan and Jeffreys teaches using duplex consensus sequencing combined with allele-specific probe enrichment to accurately detect specific mutations in DNA.
Kennedy in view of Wan and Jeffreys does not teach that the specific mutation is identified with at least 10 times or at least 100 times fewer sequencing reads as compared with conventional duplex sequencing methods (claim 12), that the pool of duplex DNA is from a liquid biopsy that contains cell-free DNA (claims 17 and 23), or that there is at least one additional allele-specific probe targeted to a distinct specific mutation (claim 30). However, each of these features are known in the art and are taught by Zhang.
Zhang teaches a method of sequencing through the use of allele-specific enrichment for detection of rare alleles (Abstract). Zhang teaches that allele-specific probe enrichment enables a 36-fold reduction in reads needed for identifying a variant allele (claim 12, reads on at least 10 times fewer…sequencing reads; Figure 3B, paragraph [0024]). Zhang teaches that their method allows for analysis of duplex DNA from liquid biopsies which contain cfDNA (claims 17 and 23; paragraphs [0005 and 0053]). Zhang teaches that this method can be multiplexed (reads on at least one additional allele-specific probe in claim 30; paragraph [0024]).
It would have been prima facie obvious to one having ordinary skill in the art, before the effective filing date of the instant application, to have modified the method of Kennedy in view of Wan and Jeffreys with the methodology of Zhang. One would be motivated to use allele-specific probes that enable a 36-fold reduction in sequencing reads given the assertion by Zhang that 36-fold reduction in sequencing reads means a 36-fold reduction in sequencing cost (paragraph [0024]). One would have a reasonable expectation of success given that Jeffreys and Zhang exhibit successful usage of allele-specific probes for enriching specific mutations and Kennedy and Wan exhibit successful use of a method that includes an enrichment step before consensus calling. One would be motivated to perform this methodology on liquid biopsies that contain cfDNA given the assertion by Zhang that this could be used to assay for detection of cancer mutations (paragraph [0005]). One would have a reasonable expectation of success given that Zhang successfully detects cancer mutations in liquid biopsies with allele-specific probe enrichment (Figure 6). One would be motivated to use additional allele-specific probes for distinct specific mutations, as taught by Zhang, in order to increase the throughput/multiplex the assay to detect multiple variants at one time.
Claim 45 is rejected under 35 U.S.C. 103 as being unpatentable over Kennedy (Kennedy et al.; Nature Protocols, 2014; cited on PTO-892 of 8/8/2025) in view of Wan (Wan et al., bioRxiv 2019; cited on IDS of 3/13/2025) and Jeffreys (Jeffreys et al.; Genome Research, 2003; cited on PTO-892 of 8/8/2025) as applied to claims 1, 3-4, 7, 14, 53, 56, 58 and 79-81, and 83-86 above, and further in view of Maguire (Maguire et al., US 2017/0275689 A1; cited on PTO-892 of 8/8/2025).
The teachings of Kennedy in view of Wan and Jeffreys as applied to claims 1, 3-4, 7, 14, 53, 56, 58 and 79-81, and 83-86 are detailed above. Relevant to the instantly rejected claims, Kennedy in view of Wan and Jeffreys teaches using duplex consensus sequencing combined with allele-specific probe enrichment to accurately detect specific mutations in DNA. Jeffreys teaches an allele-specific probe that is biotinylated on the 5’ end.
Kennedy in view of Wan and Jeffreys does not teach that at least one nucleotide of the allele-specific probe comprises a modification. However, modified nucleotides in oligonucleotide probes is known in the art, as taught by Maguire.
Maguire teaches a method of using probes specific to a “segregating marker” (reads on specific mutation) in order to enrich target cfDNA fragments from a patient sample (paragraph [0006-0007 and 0229]). Maguire teaches that in order to recover the target-hybrid probe, purification can be performed by a binding moiety and its moiety binding partner (paragraph [0229]). Maguire teaches that the binding moiety can be “a modified nucleotide comprising the binding partner” (paragraph [0220]).
It would have been prima facie obvious to one having ordinary skill in the art, before the effective filing date of the instant application, to have modified the method of Kennedy in view of Wan and Jeffreys with the methodology of Maguire. One would be motivated to include a modified nucleotide in the allele-specific probe given the assertion by Maguire that by incorporating a binding partner moiety as a modification to the nucleotide of the allele-specific probe, this would enable the target-hybrid complex to be isolated from a sample (Maguire, paragraph [0219]).
Response to Remarks
Applicant's arguments filed 1/8/2026 have been fully considered but they are not persuasive for the following reasons. Arguments regarding the rejection of claim 1 over Kennedy in view of Jeffreys are moot given the amendment to claim 1 and the withdrawal of said rejections under 35 USC 103. New rejections have been made in light of said amendments above.
Regarding Applicant’s arguments against Kennedy (pages 13-15 of Remarks of 1/8/2026), the application of a quality metric (“optimal peak family size”) needed to reliably form a DCS, while being used in Kennedy as a metric for yield, is noted as an “appropriate peak family size” for high quality depth. And as noted by Wan, and referenced above, enriching for specific mutation sites in a patient can lead to accumulation of “technically noisy sites” that may yield false mutation data, and thus some form of quality metric in addition to DCS should be employed to reduce background error. Given that Kennedy already teaches an optimal peak family size of six SSCS to one DCS, one would be motivated to use this as a quality cut off to reduce said noise as taught by Wan.
Conclusion
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to KAILEY E CASH whose telephone number is (571)272-0971. The examiner can normally be reached Monday-Friday 8:30am-6pm ET.
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/KAILEY ELIZABETH CASH/Examiner, Art Unit 1683
/STEPHEN T KAPUSHOC/Primary Examiner, Art Unit 1683
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