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 .
DETAILED ACTION
Status of Claims
Claims 13-25 and 27 are pending and the subject of this NON-FINAL Office Action. This is the first office action on the merits.
Election/Restrictions
Applicant’s election without traverse of Invention II (claims 13-25 and 27) in the reply filed on 05/13/2026 is acknowledged. Applicants canceled all other claims.
Claim Interpretations
The “ending sequences” in Claims 13 and 27 encompass any sequence of any size at or near a sequence end. Claims 13 and 27 state “identifying a set of the sequence reads from the aligned sequence reads, wherein each sequence read of the set of the sequence reads includes an ending sequence corresponding to a set of one or more sequence end signatures.” Paragraph 0040 states “[t]he term “ending sequence” refers to an end of a sequence read” and “can correspond to the outermost N bases of the fragment, e.g., 2-30 bases at the end of the fragment,” and include paired end sequence reads. However, “end signature” is never defined. Instead, only “end motif” (para. 0041), which is not claimed. It is “a sequence motif [short, recurring pattern of bases in nucleic acid fragments] for an ending sequence that preferentially occurs at ends of nucleic acid, e.g., DNA, fragments, potentially for DNA molecules originating from pathogenic microbes.”
In light of these differing meanings, prior art is applied below as to “ending sequence” that is any length and sequence, at any location around a sequence end. This encompasses a lot of prior art. For example, as explained in WO2020125709, “it was reported that a subset of human genomic locations (e.g., positions on a reference genome) are preferentially cut, thereby generating plasma DNA fragment having end positions that bear a relationship with the tissue of origin (Chan et al, Proc Natl Acad Sci USA. 2016; 113: E8159-8168; Jiang et al, Proc Natl Acad Sci USA. 2018; doi: 10.1073/pnas.1814616115). Chandrananda et al (BMC Med Genomics. 2015; 8: 29) used the de novo discovery software DREME (Bailey, Bioinformatics. 2011; 27: 1653-9) to mine the cell-free DNA data for motifs related to nuclease cleavage, irrespective of tissue type” (Background). As to paired-end sequencing, “Paired-end sequencing is a technique that obtains sequence information for both ends of each DNA molecule. By finding the coordinates of the 2 sequences on the genome through sequence alignment, one can deduce the length of the DNA fragment” (Fan et al, Analysis of the Size Distributions of Fetal and Maternal Cell-Free DNA by Paired-End Sequencing, Clinical Chemistry 56:8, 1279–1286 (2010), pg. 1279, col. 2).
An “end motif” is not much better due to the expansive definition above. Yet, the specification only discloses a very specific “end motif” in Figure 1. Prior art is applied to this as well in the Obviousness rejection, below.
Claim Rejections - 35 USC § 102
The following is a quotation of the appropriate paragraphs of 35 U.S.C. § 102 that form the basis for the rejections under this section made in this Office action:
A person shall be entitled to a patent unless –
(1) the claimed invention was patented, described in a printed publication, or in public use, on sale, or otherwise available to the public before the effective filing date of the claimed invention.
Claims 13-14, 18 and 27 are rejected under 35 U.S.C. § 102(a)(1) as being anticipated by LO (US 20200002770).
As to claims 13 and 27, LO teaches a method of analyzing a biological sample to determine a level of infection in a subject, the biological sample including a mixture of cell-free DNA molecules from the subject and microbes, the method comprising:
analyzing a plurality of cell-free DNA molecules from the biological sample to obtain sequence reads, wherein the sequence reads include ending sequences corresponding to ends of the plurality of cell-free DNA molecules (“determining the pathogen integration index comprises determining an amount of the plurality of cell-free nucleic molecules that comprise a first end from the genome of the pathogen and a second end from the genome of the pathogen”; para. 0004);
aligning the sequence reads to one or more reference microbe genomes to identify aligned sequence reads, each of the one or more reference microbe genomes corresponding to a particular species of microbes (aligning to reference genome; para. 0004);
identifying a set of the sequence reads from the aligned sequence reads, wherein each sequence read of the set of the sequence reads includes an ending sequence (id.);
determining a parameter for the set of the sequence reads based at least in part on a first amount of the set of sequence reads (id.); and
determining a classification of a level of infection using the parameter (amount of pathogen infection; id.)
As to claim 14, LO teaches the parameter is a frequency determined based on the first amount of the set of sequence reads (claim 1, above).
As to claim 18, LO teaches the determination of the classification of the level of infection is based on a comparison between the parameter and a reference value (para. 0004).
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.
Claim(s) 13-27 is/are rejected under 35 U.S.C. 103 as being unpatentable over KARIUS (WO 2018045359), in view of GRAIL (WO2020125709).
This rejection is presented in the interest of compact prosecution to the extent the claims encompass end motifs.
The prior art as a whole demonstrates that it would have been obvious to a skilled artisan at the time of filing to apply familiar end motif cfDNA quantification analysis to the quantification analysis of KARIUS with a reasonable expectation of success.
As to claims 13 and 27, KARIUS teaches a method of analyzing a biological sample to determine a level of infection in a subject, the biological sample including a mixture of cell-free DNA molecules from the subject and microbes, the method comprising:
analyzing a plurality of cell-free DNA molecules from the biological sample to obtain sequence reads, wherein the sequence reads include ending sequences corresponding to ends of the plurality of cell-free DNA molecules (Fig. 1 and claims 1-3);
aligning the sequence reads to one or more reference microbe genomes to identify aligned sequence reads, each of the one or more reference microbe genomes corresponding to a particular species of microbes (Fig. 1 and paras. 0137 & 0199);
identifying a set of the sequence reads from the aligned sequence reads, wherein each sequence read of the set of the sequence reads includes an ending sequence (id.);
determining a parameter for the set of the sequence reads based at least in part on a first amount of the set of sequence reads (quantity; claim 3); and
determining a classification of a level of infection using the parameter (detecting pathogen, and amounts/quantity; claims 1-2).
As to claim 14, KARIUS teaches the parameter is a frequency determined based on the first amount of the set of sequence reads (claim 1, above).
As to claim 18, KARIUS teaches the determination of the classification of the level of infection is based on a comparison between the parameter and a reference value (claims 4-7).
As to claim 19, KARIUS teaches the level of infection indicates a presence of sepsis (claim 54).
As to claim 20, KARIUS teaches for each of the plurality of cell-free DNA molecules in the biological sample:
measuring a size of the cell-free DNA molecule; and
determining that the cell-free DNA molecule is from the one or more reference microbe genomes;
determining a statistical value of the measured sizes of the plurality of cell-free DNA molecules;
comparing the statistical value to a cutoff value; and
further determining the level of infection in the subject based on the comparison (“To estimate the abundance of each pathogen, DNA and/or RNA molecules of various lengths, GC content and at known concentrations can be spiked into the sample before conducting an assay such as a sequencing assay”; para. 0060).
As to claim 21, KARIUS teaches aligning the sequence reads to one or more reference microbe genomes includes:
aligning the sequence reads to a reference human genome;
identifying one or more non-aligned sequence reads by filtering out, from the sequence reads, a plurality of sequence reads that align to the reference human genome; and
realigning the one or more non-aligned sequence reads to the one or more reference microbe genomes to identify the aligned sequence reads (“In the first, two samples are prepared with a known template (Lambda gDNA, Pacbio Part no: 001-119-535), and purified DNA for sequencing using the above-described workflow (lllumina Miseq, 3.4 and 3.5 million reads). Lambda-derived sequences are removed and the remaining sequences (0.4%) are aligned to reference database using BLAST”; para. 0199).
As to claim 22, KARIUS teaches enriching the set of sequence reads (id.)
As to claim 25, KARIUS teaches the subject is a pregnant female, and wherein the classification of the level of infection includes an infection conducive to preterm labor (claim 54).
KARIUS does not explicitly teach ending sequences corresponding to a set of one or more sequence end signatures or end motifs; or claims 15-17 and 23-24.
However, end motif analysis was a known option to increase quantification accuracy. Specifically, GRAIL teaches
The present disclosure describes techniques for measuring quantities (e.g., relative frequencies) of sequence end motifs of cell-free DNA fragments in a biological sample of an organism for measuring a property of the sample (e.g., fractional concentration of clinically-relevant DNA) and/or determining a condition of the organism based on such measurements. Different tissue types exhibit different patterns for the relative frequencies of the sequence end motifs. The present disclosure provides various uses for measures of the relative frequencies of sequence end motifs of cell-free DNA, e.g., in mixtures of cell-free DNA from various tissues. DNA from one of such tissue may be referred to as clinically-relevant DNA
(Abstract). This technique is shown in Figure 1, which is identical to Figure 1 here:
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The property or condition of the sample or organism includes levels of pathologies (claim 1, for example) such as infection with a pathogen such as a virus (e.g. Fig. 22, HBV). One advantage of this quantification technique is “as the number of end motifs may be smaller than the number of preferred end positions in a reference genome, greater statistics can be gathered for each end motif, potentially increasing accuracy” (para. 0085). A skilled artisan reading GRAIL in light of KARIUS would have immediately recognized the application of the cfDNA end motif analysis method of GRAIL to detect other known pathologies such as infections with a pathogen as in KARIUS. The method of GRAIL would have yielded increased quantification accuracy for familiar cfDNA pathogen detection and quantification. In other words, GRAIL provides motivation to apply its end motif cfDNA quantification technique to other familiar cfDNA-based detections that use sequencing to detect pathologies such as infections.
As to claims 15-17, GRAIL also teaches to compare observed amount/frequency of sequence reads to expected to generate ratios or combined values (paras. 0110 & 0125).
As to claims 23-24, GRAIL also teaches processing the first amount of the set of the sequence reads using a machine-learning model such as SVM (3. Machine Learning (SVM, Regression, and Clustering)).
Prior Art
The following prior art is also pertinent: US20190153512.
Conclusion
No claims are allowed.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to Aaron Priest whose telephone number is (571)270-1095. The examiner can normally be reached 8am-6pm.
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/AARON A PRIEST/Primary Examiner, Art Unit 1681
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