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 .
Claim Status
Claims 1-20 are currently pending and under examination herein.
Claims 1-20 are rejected.
Claims 14 is objected to.
Priority
The instant application does not claim priority. In this action, claims 1-20 are examined as though they had an effective filing date of 27 January 2023. In future actions, the effective filing date of one or more claims may change, due to amendments to the claims, or further analysis of the disclosure(s) of the priority application(s).
Information Disclosure Statement
The information disclosure statement (IDS) submitted on 8 February 2023 is in compliance with the provisions of 37 CFR 1.97. The reference SIMON-LORIERE et al. (Rapid characterization of a Delta-Omicron SARSCoV-2 recombinant detected in Europe) was not found amongst the submitted NPL. Therefore, this reference was lined through on the IDS and not considered. The reference VARABYOU et al. (Rapid detection of inter-clade recombination in SARS-CoV-2 with Bolotie) was found to be submitted twice. Accordingly, the information disclosure statement is being considered by the examiner.
Drawings
The drawings filed 27 January 2023 are accepted.
Claim Objections
Claim 14 is objected to because it recites “identifying reads that start with the first mutation and send with the second mutation”. This is interpreted to be a typographical error where ends was replaced by send. Appropriate correction required.
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.
Claim 12 rejected under 35 U.S.C. 112(b) as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor regards as the invention.
Claim 12 recites “ The method of claim 1, wherein detecting a recombinant pathogen comprises”. Detecting a recombinant pathogen is not recited in claim 1 or any claim. Therefore, it is unclear what the wherein clause in claim 12 is referring to or limiting, as it is not currently recited as adding an additional active method step. The metes and bounds of the claim 12 are therefore unclear rendering the claim indefinite. For the purpose of examination, the limitation is interpreted as an additional step of the method of claim 1. This rejection can be overcome by changing the language to limit a limitation previously recited or making it add a sperate method step such as - The method of claim 1, further comprising detecting a recombinant pathogen.
Claim Rejections - 35 USC § 101
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 1-20 are rejected under 35 U.S.C. 101 because the claimed invention is directed to an abstract idea and a natural law without significantly more. In accordance with MPEP § 2106, claims found to recite statutory subject matter (Step 1: YES) are then analyzed to determine if the claims recite any concepts that equate to an abstract idea or natural law (Step 2A, Prong 1). Claims 1-16 are directed to a method and Claims 17-20 are directed to a system. In the instant application, the claims recite the following limitations that equate to an abstract idea or natural law:
Claim 1 recites the limitation - determining that the biological sample is a mixed sample of the first variant and the second variant based on an alternative allele fraction calculated according to the plurality of reads; searching individual reads of the plurality of reads for recombinant reads that include the first mutation and the second mutation; and determining whether the biological sample is indicative of a coinfection or a contamination, based on an amount of the recombinant reads that each indicate both the first variant and the second variant. Based on the broadest reasonable interpretation, determining the sample is mixed, searching individual reads, and determining the sample is a coinfection could practically be done by the human mind. This draws the limitation to a mental process, which classifies the limitation as an abstract idea. Searching is interpreted as identifying according to the specification (Page 20, Paragraph 0123). Additionally, determining if a biological sample contains a coinfection or contamination based on multiple variants within read data represents a natural correlation, which draws the limitation to a law of nature.
Claim 2 recites the limitation - wherein determining that the biological sample is a mixed sample comprises calling an alternative allele at a locus based on either the first mutation or the second mutation. Based on the broadest reasonable interpretation, calling an alternative allele could practically be done by the human mind. This draws the limitation to a mental process, which classifies the limitation as an abstract idea. Calling is interpreted as identifying according to the specification (Page 7, Paragraph 0052).
Claim 3 recites the limitation - wherein determining that the biological sample is a mixed sample comprises calculating an alternative allele fraction for the called alternative allele. Based on the broadest reasonable interpretation, calculating a fraction could include equations and could practically be done by the human mind. This draws the limitation to a mathematical concept and a mental process, which classifies the limitation as an abstract idea.
Claim 4 recites the limitation - wherein calculating an alternative allele fraction for one of the called alternative alleles is based on a number of reads of the called alternative allele divided by a total number of reads at the locus. Based on the broadest reasonable interpretation, calculating a fraction by dividing could include equations and could practically be done by the human mind. This draws the limitation to a mathematical concept and a mental process, which classifies the limitation as an abstract idea.
Claim 6 recites the limitation - wherein determining that the biological sample is a mixed sample comprises calculating at least one of a median alternative allele fraction, a mean alternative allele fraction, a weighted alternative allele fraction, or an average alternative allele fraction, based on one or more calculated alternative allele fractions. Based on the broadest reasonable interpretation, the calculations could include equations and could practically be done by the human mind. This draws the limitation to a mathematical concept and a mental process, which classifies the limitation as an abstract idea.
Claim 7 recites the limitation - wherein calculating at least one of a median alternative allele fraction, a mean alternative allele fraction, a weighted alternative allele fraction, or an average alternative allele fraction is based on loci that have alternative allele fractions of at least a threshold minimum fraction. This limitation specifies the possible fractions going into the calculation in the judicial exception of claim 6. The refined calculations indicated by this limitation still represents a judicial expectation.
Claim 8 recites the limitation - wherein the threshold minimum fraction is about 0.15. This limitation specifies the possible fractions going into the calculation in the judicial exception of claim 6. The refined calculations indicated by this limitation still represents a judicial expectation.
Claim 9 recites the limitation - wherein if the calculation based on the alternative allele fraction is below a threshold alternative allele fraction, the sample is a mixed sample. Based on the broadest reasonable interpretation, determining if the sample is mixed based on the calculation could practically be done by the human mind. This draws the limitation to a mental process, which classifies the limitation as an abstract idea.
Claim 10 recites the limitation - wherein if the calculation based on the alternative allele fraction is above a threshold alternative allele fraction, the sample is dominant in one of the first variant and the second variant. Based on the broadest reasonable interpretation, determining if the sample is dominant based on the calculation could practically be done by the human mind. This draws the limitation to a mental process, which classifies the limitation as an abstract idea.
Claim 11 recites the limitation - wherein the threshold alternative allele fraction is about 0.80. This limitation specifies the possible calculations used to determine in the judicial exception of claim 10. The refined determination indicated by this limitation still represents a judicial expectation.
Claim 12 recites the limitation - wherein detecting a recombinant pathogen comprises identifying reads or read pairs straddling at least one of the first mutation and the second mutation. Based on the broadest reasonable interpretation, identifying reads could practically be done by the human mind. This draws the limitation to a mental process, which classifies the limitation as an abstract idea.
Claim 13 recites the limitation - wherein determining whether the biological sample is indicative of a coinfection or a contamination comprises walking across the sequence from a 5′ end to a 3′ end to determine whether the first mutation, the second mutation, or combinations thereof are present. Based on the broadest reasonable interpretation, reviewing a sequence and determining if a sequence is present could practically be done by the human mind. This draws the limitation to a mental process, which classifies the limitation as an abstract idea. Walking is interpreted as reviewing based on the specification (Paragraph 0064).
Claim 14 recites the limitation - wherein determining whether the biological sample is indicative of a coinfection or a contamination comprises identifying reads that start with the first mutation and send with the second mutation. Based on the broadest reasonable interpretation, identifying reads could practically be done by the human mind. This draws the limitation to a mental process, which classifies the limitation as an abstract idea.
Claim 15 recites the limitation - wherein determining whether the biological sample is indicative of a coinfection or a contamination comprises identifying reads that include mutations from both the first variant and the second variant. Based on the broadest reasonable interpretation, identifying reads could practically be done by the human mind. This draws the limitation to a mental process, which classifies the limitation as an abstract idea.
Claim 16 recites the limitation - wherein determining whether the biological sample is indicative of a coinfection or a contamination comprises identifying one or more breakpoints. Based on the broadest reasonable interpretation, identifying breakpoints could practically be done by the human mind. This draws the limitation to a mental process, which classifies the limitation as an abstract idea. The interpretation of identifying breakpoints as a mental process is based on Figure 4.
Claim 17 recites the limitation - identifying a first variant and a second variant of the biological sample, the first variant corresponding to a first mutation and the second variant corresponding to a second mutation different than the first mutation; determining that the biological sample is a mixed sample of the first variant and the second variant based on an alternative allele fraction calculated according to the plurality of reads; searching individual reads of the plurality of reads for recombinant reads that include the first mutation and the second mutation; and determining whether the biological sample is indicative of a coinfection or a contamination, based on an amount of the recombinant reads that each indicate both the first variant and the second variant. Based on the broadest reasonable interpretation, identifying variants, determining the sample is mixed, searching individual reads, and determining the sample is a coinfection could practically be done by the human mind. This draws the limitation to a mental process, which classifies the limitation as an abstract idea. Searching is interpreted as identifying according to the specification (Page 20, Paragraph 0123). Additionally, determining if a biological sample contains a coinfection or contamination based on multiple variants within read data represents a natural correlation, which draws the limitation to a law of nature.
Claim 18 recites the limitation - wherein determining whether the biological sample is a mixed sample comprises calling an alternative allele at a locus based on either the first mutation or the second mutation. Based on the broadest reasonable interpretation, calling an alternative allele could practically be done by the human mind. This draws the limitation to a mental process, which classifies the limitation as an abstract idea. Calling is interpreted as identifying according to the specification (Page 7, Paragraph 0052).
Claim 19 recites the limitation - wherein determining whether the biological sample is a mixed sample comprises calculating an alternative allele fraction for the called alternative allele. Based on the broadest reasonable interpretation, calculating a fraction could include equations and could practically be done by the human mind. This draws the limitation to a mathematical concept and a mental process, which classifies the limitation as an abstract idea.
Claim 20 recites the limitation - wherein calculating an allele fraction for one of the called alternative alleles comprises taking a number of reads of the called alternative allele divided by a total number of reads at the locus. Based on the broadest reasonable interpretation, calculating a fraction by dividing could include equations and could practically be done by the human mind. This draws the limitation to a mathematical concept and a mental process, which classifies the limitation as an abstract idea.
These limitations recite concepts of determining, identifying and comparing information and calculating values that are so generically recited that they can be practically performed in the human mind as claimed, which falls under the “Mental processes” and “Mathematical concepts” grouping of abstract ideas. Therefore, these limitations fall under the “Mental process” and “Mathematical concepts” groupings of abstract ideas. Additionally, the limitations describe natural correlations between genotypes (mutations) and their corresponding phenotypes (pathogenic variants), which fall under natural laws. As such, claims 1-20 recite an abstract idea and law of nature (Step 2A, Prong 1: YES).
Claims found to recite a judicial exception under Step 2A, Prong 1 are then further analyzed to determine if the claims as a whole integrate the recited judicial exception into a practical application or not (Step 2A, Prong 2). These judicial exceptions are not integrated into a practical application because the claims do not recite an additional element that reflects an improvement to technology (MPEP § 2106.04(d)(1)). Rather, the claims provide insignificant extra-solution activity (MPEP § 2106.05(g)) and provide mere instructions to apply a judicial exception (MPEP § 2106.05(f)). Specifically, the claims recite the following additional elements:
Claim 1 recites the biological sample including a first variant of a pathogen and a second variant of the pathogen, wherein the first variant corresponds to a first mutation and the second variant corresponds to a second mutation different than the first mutation; and acquiring sequencing data for the biological sample, the sequencing data including a plurality of reads;
Claim 5 recites further comprising selecting the first variant and the second variant of the pathogen, wherein selecting the first variant and the second variant comprises retrieving information from a database.
Claim 17 recites a non-transitory machine-readable medium including instructions that, when executed by a processor of a machine, cause the machine to perform operations comprising: acquiring sequencing data for a biological sample, the sequencing data including a plurality of reads;
There are no limitations that indicate that the claimed determining, identifying and comparing information and calculating values require anything other than generic computing systems, including a non-transitory machine-readable medium. As such, these limitations equate to mere instructions to implement the abstract idea on a generic computer that the courts have stated does not render an abstract idea eligible in Alice Corp., 573 U.S. at 223, 110 USPQ2d at 1983. See also 573 U.S. at 224, 110 USPQ2d at 1984. There is no indication that these steps are affected by the judicial exception in any way and thus do not integrate the recited judicial exception into a practical application. As such, claims 1-20 are directed to an abstract idea and natural law (Step 2A, Prong 2: NO).
Claims found to be directed to a judicial exception are then further evaluated to determine if the claims recite an inventive concept that provides significantly more than the judicial exception itself (Step 2B). The claims do not include additional elements that are sufficient to amount to significantly more than the judicial exception because the claims recite conventional additional elements that equate to mere instructions to apply the recited exception in a generic way or in a generic computing environment. The claims also recite conventional additional elements that represent insignificant extra-solution activities.
As discussed above, there are no additional limitations to indicate that the claimed determining, identifying and comparing information and calculating values require anything other than generic computer components in order to carry out the recited abstract idea in the claims. Claims that amount to nothing more than an instruction to apply the abstract idea or natural law using a generic computer do not render an abstract idea or natural law eligible. MPEP 2106.05(f) discloses that mere instructions to apply the judicial exception cannot provide an inventive concept to the claims. As specified in MPEP 2106.05(g), extra-solution activities can be understood as incidental to the primary process or product that are merely a nominal or tangential addition to the claim. Insignificant extra-solution activities include mere data gathering, selecting a particular data source or type of data to be manipulated, and displaying information. Additionally, Sommers et al. (2021, Annual Review of Virology, Vol. 8: 133-158) teach the use of computers (Page 146, Paragraph 3: Development of computational tools such as DRAM-v) as part of acquiring sequence data with multiple reads (Page 140, Paragraph 1: the length of reads obtained through metagenomic sequencing has advantages and disadvantages for addressing questions) related to samples with coinfection of a pathogen (Page 146, Paragraph 4: RNA sequencing moreover has shown the extent of infection and coinfection) and retrieving variant information from databases (Page 136, Paragraph 2: uncultivated virus-derived genome sequence further increase the power of these databases by providing ecological context) is well understood, routine, and conventional.
The additional elements do not comprise an inventive concept when considered individually or as an ordered combination that transforms the claimed judicial exception into a patent-eligible application of the judicial exception. Therefore, the claims do not amount to significantly more than the judicial exception itself (Step 2B: No). As such, Claims 1-20 are not patent eligible.
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.
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.
Claims 1-20 are rejected under 35 U.S.C. 103 as being unpatentable over Tell et al. (US 20210343372 A1), in view of Combes et al. (2022, Clinical Microbiology and Infection, Vol. 28: 1-8). Italicized text from reference art.
The applicable claims include:
Claim 1. A method of distinguishing between coinfection and contamination for a biological sample, the biological sample including a first variant of a pathogen and a second variant of the pathogen, wherein the first variant corresponds to a first mutation and the second variant corresponds to a second mutation different than the first mutation, the method comprising: (i) acquiring sequencing data for the biological sample, the sequencing data including a plurality of reads; (ii) determining that the biological sample is a mixed sample of the first variant and the second variant based on an alternative allele fraction calculated according to the plurality of reads; (iii) searching individual reads of the plurality of reads for recombinant reads that include the first mutation and the second mutation; and (iv) determining whether the biological sample is indicative of a coinfection or a contamination, based on an amount of the recombinant reads that each indicate both the first variant and the second variant.
Claim 2. The method of claim 1, wherein determining that the biological sample is a mixed sample comprises calling an alternative allele at a locus based on either the first mutation or the second mutation.
Claim 3. The method of claim 2, wherein determining that the biological sample is a mixed sample comprises calculating an alternative allele fraction for the called alternative allele.
Claim 4. The method of claim 3, wherein calculating an alternative allele fraction for one of the called alternative alleles is based on a number of reads of the called alternative allele divided by a total number of reads at the locus.
Claim 5. The method of claim 1, further comprising selecting the first variant and the second variant of the pathogen, wherein selecting the first variant and the second variant comprises retrieving information from a database.
Claim 6. The method of claim 1, wherein determining that the biological sample is a mixed sample comprises calculating at least one of a median alternative allele fraction, a mean alternative allele fraction, a weighted alternative allele fraction, or an average alternative allele fraction, based on one or more calculated alternative allele fractions.
Claim 7. The method of claim 6, wherein calculating at least one of a median alternative allele fraction, a mean alternative allele fraction, a weighted alternative allele fraction, or an average alternative allele fraction is based on loci that have alternative allele fractions of at least a threshold minimum fraction.
Claim 8. The method of claim 7, wherein the threshold minimum fraction is about 0.15.
Claim 9. The method of claim 7, wherein if the calculation based on the alternative allele fraction is below a threshold alternative allele fraction, the sample is a mixed sample.
Claim 10. The method of claim 7, wherein if the calculation based on the alternative allele fraction is above a threshold alternative allele fraction, the sample is dominant in one of the first variant and the second variant.
Claim 11. The method of claim 10, wherein the threshold alternative allele fraction is about 0.80.
Claim 12. The method of claim 1, wherein detecting a recombinant pathogen comprises identifying reads or read pairs straddling at least one of the first mutation and the second mutation.
Claim 13. The method of claim 1, wherein determining whether the biological sample is indicative of a coinfection or a contamination comprises walking across the sequence from a 5′ end to a 3′ end to determine whether the first mutation, the second mutation, or combinations thereof are present.
Claim 14. The method of claim 1, wherein determining whether the biological sample is indicative of a coinfection or a contamination comprises identifying reads that start with the first mutation and send with the second mutation.
Claim 15. The method of claim 1, wherein determining whether the biological sample is indicative of a coinfection or a contamination comprises identifying reads that include mutations from both the first variant and the second variant.
Claim 16. The method of claim 1, wherein determining whether the biological sample is indicative of a coinfection or a contamination comprises identifying one or more breakpoints.
Claim 17. A non-transitory machine-readable medium including instructions that, when executed by a processor of a machine, cause the machine to perform operations comprising: (i) acquiring sequencing data for a biological sample, the sequencing data including a plurality of reads; (ii) identifying a first variant and a second variant of the biological sample, the first variant corresponding to a first mutation and the second variant corresponding to a second mutation different than the first mutation; (iii) determining that the biological sample is a mixed sample of the first variant and the second variant based on an alternative allele fraction calculated according to the plurality of reads; (iv) searching individual reads of the plurality of reads for recombinant reads that include the first mutation and the second mutation; and (v) determining whether the biological sample is indicative of a coinfection or a contamination, based on an amount of the recombinant reads that each indicate both the first variant and the second variant.
Claim 18. The non-transitory machine-readable medium of claim 17, wherein determining whether the biological sample is a mixed sample comprises calling an alternative allele at a locus based on either the first mutation or the second mutation.
Claim 19. The non-transitory machine-readable medium of claim 18, wherein determining whether the biological sample is a mixed sample comprises calculating an alternative allele fraction for the called alternative allele.
Claim 20. The non-transitory machine-readable medium of claim 19, wherein calculating an allele fraction for one of the called alternative alleles comprises taking a number of reads of the called alternative allele divided by a total number of reads at the locus.
Regarding Claim 1, Tell et al. teach (Claim 1.ii) determining that the biological sample is a mixed sample of variants based on an alternative allele fraction calculated according to the plurality of reads (Page 3, Paragraph 0022: the present disclosure solves this and other needs in the art by providing improved somatic variant identification methodology that better accounts for locus-specific and/or sample specific considerations to more accurately identify true somatic mutations in a liquid biopsy sample. the variant allele fraction for the variant being evaluated; Page 3, Paragraph 0024: Each respective sequence read in the first plurality of sequence reads is aligned to a reference sequence for the species of the subject, thus identifying a variant allele fragment count for a candidate variant that maps to a locus in the reference sequence; Page 6, Paragraph 0061: sequence signals originating from cancerous cells, which may constitute multiple sub-clonal populations, must be computationally deconvoluted from signals originating from germline and hematopoietic origins). If it is determined that multiple variants or multiple populations are in a sample, it is obvious that the sample is considered to be mixed. Multiple variants (i.e. mixed) of a pathogen are also detected within a sample when present as taught by Combes et al. (see below Combes et al. section below).
Regarding Claim 4 and 20, Tell et al. teach calculating an alternative allele fraction for one of the called alternative alleles is based on a number of reads of the called alternative allele divided by a total number of reads at the locus (Page 8, Paragraph 0077: the term “variant allele fraction” refers to the number of times a variant or mutant allele was observed divided by the total number of times the position was sequenced). The terms variant and alternative are considered synonymous. Claim 20 recites the limitations of claim 4 directed to a CRM.
Regarding Claim 5, Tell et al. teach selecting variants of the pathogen, wherein selecting comprises retrieving information from a database (Page 18, Paragraph 0163: one or more genomic variant analysis algorithms evaluate various genomic features by querying a database).
Regarding Claim 6, Tell et al. teach determining that the sample is mixed comprises calculating a median alternative allele fraction, a mean alternative allele fraction, a weighted alternative allele fraction, or an average alternative allele fraction, based on one or more calculated alternative allele fractions (Page 31, Paragraph 0271: a baseline variant threshold is influenced by the sequencing depth of the reaction, e.g., a locus-specific sequencing depth and/or an average sequencing depth; Page 10, Paragraph 0097: Alternatively, read-depth, sequencing depth, or depth can refer to a measure of central tendency (e.g., a mean or mode) of the number of unique nucleic acid fragments that encompass one of a plurality of loci or regions of the genome of a subject that are sequenced in a particular sequencing reaction). The art defines sequencing depth as a total number of unique nucleic acid fragments encompassing a particular locus or region of the genome (Page 10, Paragraph 0097), which is involved in the alternative allele fraction calculation.
Regarding Claim 7, Tell et al. teach calculating a median alternative allele fraction, a mean alternative allele fraction, a weighted alternative allele fraction, or an average alternative allele fraction is based on loci that have alternative allele fractions of at least a threshold minimum fraction (Page 3, Paragraph 0025: The method further includes comparing the variant allele fragment count for the candidate variant against a dynamic variant count threshold for the locus in the reference sequence that the candidate variant maps to). The variant allele fragment count is involved in the calculation of the alternative allele fraction, which Tell et al. teaches as related to an average/mean calculation (see Regarding claim 6 above).
Regrading Claim 8, Tell et al. teach the threshold minimum fraction is about 0.15 (Page 22, Paragraph 0193: For each section, background tissue may be excluded or removed such that the section meets a tumor purity threshold, e.g., where at least 20% of the nuclei in the section are tumor nuclei). This indicates if at least 20% of sequences are not associated with a specific mutation (i.e., not tumor DNA), the data is excluded from analysis (i.e. is a threshold). Tell et al. also directly teaches considering a threshold in calculations (see regarding claim 6). 20% is interpreted as about 15%.
Regarding Claim 9, Tell et al. teach if a calculation based on the alternative allele fraction is below a threshold alternative allele fraction, the sample is a mixed sample (See Regarding Claims 1 and 7 above for teachings on the considering the sample mixed given a particular allele fraction considered against a threshold; Page 12, Paragraph 0109: A threshold value can be a value above or below which a particular classification applies). Additionally, the limitation - if the calculation based on the alternative allele fraction is below a threshold alternative allele fraction, the sample is a mixed sample - is interpreted as a CONTINGENT LIMITATION, which indicates the broadest reasonable interpretation of a method (or process) claim requires only those steps that must be performed and does not include steps that are not required to be performed because the condition(s) precedent are not met (MPEP 211.04). Therefore, the limitation is taught by Tell et al.
Regarding Claim 10, the limitation - if the calculation based on the alternative allele fraction is above a threshold alternative allele fraction, the sample is dominant in one of the first variant and the second variant - is interpreted as a CONTINGENT LIMITATION, which indicates the broadest reasonable interpretation of a method (or process) claim requires only those steps that must be performed and does not include steps that are not required to be performed because the condition(s) precedent are not met (MPEP 211.04). Therefore, the limitation is taught by Tell et al.
Regarding Claim 11, Tell et al. teach the threshold alternative allele fraction is about 0.80 (Page 44, Paragraph 400: the threshold value for calling the patient as MSI-H is at least 80% probability). MSI-H indicates there are multiple mutations/variants present (i.e. a threshold for calling) (Page 44, Paragraph 0398: microsatellite instability-high (MSI-H) tumors, defects in DNA mismatch repair (MMR) can cause a hypermutated phenotype where alterations accumulate in the repetitive microsatellite regions of DNA), therefore it would be obvious to use that threshold for establishing the sample as not mixed (i.e. dominant). See above for Tell et al. teaching thresholds related to alternative allele fraction.
Regarding Claim 12, Tell et al. teach detecting a recombinant pathogen comprises identifying reads or read pairs straddling at least one of the first mutation and the second mutation (Page 3, Paragraph 0024: thus identifying a variant allele fragment count for a candidate variant that maps to a locus; Page 7, Paragraph 0071: a locus is defined by a gene, a sub-genic structure, or a predefined span of a chromosome). A locus (i.e. a read) associated with variants, whether recombinant or not, are identified and can be any size including those that span the mutations. Applying sequence analysis methods to recombinant pathogen detection is directly taught by Combes et al. (see below Combes et al. section below).
Regarding Claim 14, Tell et al. determining whether the biological sample is indicative of a multiple variants comprises identifying reads that start with the first mutation and send with the second mutation (Page 28, Paragraph 0249: The alignment position information may indicate a beginning position and an end position of a region in the reference genome that corresponds to a beginning nucleotide base and end nucleotide base of a given sequence read) This would include those that are used to identify a variant. It would be obvious to use this to detect coinfection as in Combes et al. (see reason to combine).
Regarding Claim 17, Tell et al. teach (Claim 17.iii) determining that the biological sample is a mixed sample variants based on an alternative allele fraction calculated according to the plurality of reads (Page 3, Paragraph 0022: the present disclosure solves this and other needs in the art by providing improved somatic variant identification methodology that better accounts for locus-specific and/or sample specific considerations to more accurately identify true somatic mutations in a liquid biopsy sample. the variant allele fraction for the variant being evaluated; Page 3, Paragraph 0024: Each respective sequence read in the first plurality of sequence reads is aligned to a reference sequence for the species of the subject, thus identifying a variant allele fragment count for a candidate variant that maps to a locus in the reference sequence; Page 6, Paragraph 0061: sequence signals originating from cancerous cells, which may constitute multiple sub-clonal populations, must be computationally deconvoluted from signals originating from germline and hematopoietic origins). Multiple variants of a pathogen are also detected within a sample when present as taught by Combes et al.(see below Combes et al. section below). Tell et al. also teach the methods are conducted at a generic computer which inherently contain computer readable media performing the method (Page 3, Paragraph 0023: The method is performed at a computer system; also see paragraph 0124).
Tell et al. does not teach acquiring sequencing data for the biological sample, including a plurality of reads (Claim 1.i). Tell also does not teach searching individual reads for recombinant reads that include the first mutation and the second mutation (Claim 1.iii). Tell also does not teach (Claim 1.iv) determining whether the biological sample is indicative of a coinfection, based on an amount of the recombinant reads that each indicate both the first variant and the second variant. Tell also does not teach determining that the biological sample is a mixed sample comprises calling an alternative allele at a locus based on either the first mutation or the second mutation (Claims 2 and 18). Tell also does not teach determining that the biological sample is a mixed sample comprises calculating an alternative allele fraction for the called alternative allele (Claims 3 and 9). Tell also does not teach determining whether the biological sample is indicative of a coinfection comprises walking across the sequence from a 5′ end to a 3′ end to determine whether the first mutation, the second mutation, or combinations thereof are present (Claim 13). Tell et al. also does not teach determining whether the biological sample is indicative of a coinfection comprises identifying reads that include mutations from both the first variant and the second variant (Claim 15). Tell et al. also does not teach determining whether the biological sample is indicative of a coinfection or a contamination comprises identifying one or more breakpoints (Claim 16). Tell et al. also does not teach acquiring sequencing data for a biological sample, the sequencing data including a plurality of reads (Claim 17.i). Tell et al. also does not teach identifying a first variant and a second variant of the biological sample, the first variant corresponding to a first mutation and the second variant corresponding to a second mutation different than the first mutation (Claim 17.ii). Tell et al. also does not teach searching individual reads of the plurality of reads for recombinant reads that include the first mutation and the second mutation (Claim 17.iv). Tell et al. also does not teach determining whether the biological sample is indicative of a coinfection or a contamination, based on an amount of the recombinant reads that each indicate both the first variant and the second variant (Claim 17.v).
Regarding Claim 1, Combes et al. teach (Claim 1.i) acquiring sequencing data for the biological sample, including a plurality of reads (Page 2, Column 1, Paragraph 3: All samples underwent whole genome sequencing (WGS). Raw reads were deposited). Additionally, a plurality of reads can be seen in figures S3 (Page 7) and S4 (Page 8). At least one sample included multiple variants caused by multiple mutations (Page 3, Column 2, Paragraph 2: This work shows that co-infection with two SARS-CoV-2 can occur; Page 7, Figure S3: Visualisation of DNA inserts with Delta and Omicron specific mutations for the P6 sample). Combes et al. also teach (Claim 1.iii) searching individual reads for recombinant reads that include the first mutation and the second mutation (Page 7, Figure S3: This representation showing 8 out of the 70 observed individual DNA fragments carrying both the Delta specific deletion 22029_22034 as well as the Omicron specific deletion 22194_22196 (*) is an additional argument for a recombination event between both strains). Combes et al. also teach (Claim 1.iv) determining whether the biological sample is indicative of a coinfection, based on an amount of the recombinant reads that each indicate both the first variant and the second variant (Page 7, Figure S3: This representation showing 8 out of the 70 observed individual DNA fragments carrying both the Delta specific deletion 22029_22034 as well as the Omicron specific deletion 22194_22196 (*) is an additional argument for a recombination event between both strains).
Regarding Claims 2 and 18, Combes et al. teach determining that the biological sample is a mixed sample comprises calling an alternative allele at a locus based on one of the mutations (Page 8, Figure S4: Single Molecule Real Time sequencing from Pacific Bioscience technology allows to individualise the mutations from individual molecules and generate a consensus for each population. This method applied to the spike coding sequence showed the presence of 3 different haplotypes). Within Figure S4, different letters/colors in the figures represent different calls. Figures S2 and S3 also show identifying reads with mutations as specific locations. Claim 18 recites the limitations of claim 2 directed to a CRM.
Regarding Claim 3 and 19, Combes et al. teach determining that the biological sample is a mixed sample comprises calculating an alternative allele fraction for the called alternative allele (Page 6, Figure S2: Representation of Variant Allele Frequency clade defining mutations for 21J Delta and 21K Omicron along the SARS-CoV-2 genome using the Midnight 1200 primer set. Co-infections for patients P1-D1 (A.) , P5-D1 (B.) and P6-D1 (C.) were confirmed with another WGS method using independent RNA extraction and xGen™ SARS-CoV-2-Midnight-1200 Amplicon (IDT) sequencing on a NovaSeq Illumina Platform). Variant Allele Frequency is interpreted as equivalent to alternative allele fraction. Whenever a sample is confirmed to have multiple variants present, it would be considered mixed. Claim 19 recites the limitations of claim 3 directed to a CRM.
Regarding Claim 13, Combes et al. teach determining whether the biological sample is indicative of a coinfection comprises walking across the sequence from a 5′ end to a 3′ end to determine whether the first mutation, the second mutation, or combinations thereof are present (Page 7, Figure S3: Pairs are materialised by a line between each forward read (red) and reverse read (blue) assigning both reads to the same cluster and therefore to the same DNA fragment). This indicates that both the forward and reverse strands were analyzed (i.e. walked) to determine whether the variants were present.
Regarding Claim 14, Combes et al. determining whether the biological sample is indicative of a multiple variants comprises identifying reads that start with the first mutation and send with the second mutation (Page 7, Figure S3: Reads are identified as starting or ending in the 21J Delta deletion region or 21K Omicron deletion region).
Regarding Claim 15, Combes et al. teach determining whether the biological sample is indicative of a coinfection comprises identifying reads that include mutations from both the first variant and the second variant (Page 7, Figure S3: This representation showing 8 out of the 70 observed individual DNA fragments carrying both the Delta specific deletion 22029_22034 as well as the Omicron specific deletion 22194_22196 (*) is an additional argument for a recombination event between both strains). Reads with the two variants present are identified.
Regarding Claim 16, Combes et al. teach determining whether the biological sample is indicative of a coinfection or a contamination comprises identifying one or more breakpoints (Page 3, Paragraph 3: Three haplotypes were detected including two Delta-Omicron recombinant variants with distinct breaking points).
Regarding Claim 17, Combes et al. teach (Claim 17.i) acquiring sequencing data for a biological sample, the sequencing data including a plurality of reads (Page 2, Column 1, Paragraph 3: All samples underwent whole genome sequencing (WGS). Raw reads were deposited). A plurality of reads can be seen in figures s3 (Page 7) and s4 (Page 8). At least one sample included multiple variants caused my multiple mutations (Page 3, Column 2, Paragraph 2: This work shows that co-infection with two SARS-CoV-2 can occur, particularly during a major epidemic wave marked by a cocirculation of two highly transmissible variants in the same region). Combes et al. teach (Claim 17.ii) identifying a first variant and a second variant of the biological sample, corresponding to a first mutation and a second mutation different than the first mutation (Page 2, Column 1, Paragraph 2: SARS-CoV-2 was detected in respiratory samples; Page 3, Column 2, Paragraph 2: This work shows that co-infection with two SARS-CoV-2 can occur). Combes et al. teach (Claim 17.iv) searching individual reads of the plurality of reads for recombinant reads that include the first mutation and the second mutation (Page 7, Figure S3: This representation showing 8 out of the 70 observed individual DNA fragments carrying both the Delta specific deletion 22029_22034 as well as the Omicron specific deletion 22194_22196 (*) is an additional argument for a recombination event between both strains). Combes et al. teach (Claim 17.v) determining whether the biological sample is indicative of a coinfection or a contamination, based on an amount of the recombinant reads that each indicate both the first variant and the second variant (Page 7, Figure S3: This representation showing 8 out of the 70 observed individual DNA fragments carrying both the Delta specific deletion 22029_22034 as well as the Omicron specific deletion 22194_22196 (*) is an additional argument for a recombination event between both strains). If both variants are present, it would be considered coinfection.
It would have been obvious to one of ordinary skill in the art at the time of the effective filing date to combine the methods of Tell et al. with the methods of Combes et al. because Tell et al. teach their methods are efficacious for considering mutations across samples (Page 3, Paragraph 0022: improved somatic variant identification methodology that better accounts for locus-specific and/or sample specific considerations to more accurately identify true somatic mutations in a liquid biopsy sample), which is a major focus of the Combes et al. Additionally, Tell et al. teach their methods are should be applied to pathogenic organisms (Page 10, Paragraph 100: In some embodiments, in addition to loci that are informative for precision oncology, a targeted panel includes one or more probes for sequencing one or more of a loci associated with a different medical condition or a loci from a pathogenic organism). This would include viruses, which are the focus of the methods of Combes et al. Therefore, it would have been obvious to someone of ordinary skill in the art at the time of the effective filling date to combine the methods from the references indicated above. Furthermore, one of ordinary skill in the art would predict that the methods could be readily combined with a reasonable expectation of success because both are within the same technical field – the use of pathogenic sequence data to analyze genetic variants and mutations. Accordingly, Claims 1-20 taken as a whole would have been prima facie obvious before the effective filing date and are rejected under 35 U.S.C. 103.
Double Patenting
No double patenting was identified.
Conclusion
No claims are allowed.
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/B.H.E./Examiner, Art Unit 1687
/Karlheinz R. Skowronek/Supervisory Patent Examiner, Art Unit 1687
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