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 4, 6, 8, 10, 12-13, 16-19, 21-23, 25, 29-32, 34-37, 39, 41, 44, 46-47, 49-51, and 53-57 are cancelled.
Claims 1-3, 5, 7, 9, 11, 14-15, 20, 24, 26-28, 33, 38, 40, 42-43, 45, 48, and 52 are currently pending and under exam herein.
Claims 1-3, 5, 7, 9, 11, 14-15, 20, 24, 26-28, 33, 38, 40, 42-43, 45, 48, and 52 are rejected.
Claims 1 is objected to.
Priority
Applicant’s claim for domestic benefit to the earlier filed international application PCT/US2022/025824, filed April 21, 2022, which claims the benefit of U.S. Provisional Application No. 63/177,903, filed April 21, 2021, is acknowledged. At this point in examination, the effective filing date of claims 1-3, 5, 7, 9, 11, 14-15, 20, 24, 26-28, 33, 38, 40, 42-43, 45, 48, and 52 is April 21, 2021.
Information Disclosure Statement
The information disclosure statement (IDS) submitted June 06, 2024, is in compliance with the provisions of 37 CFR 1.97. Accordingly, the IDS is being considered by the examiner.
Claim Objections
Claim 1 is objected to because of the following informalities: the acronym CGI is not defined and in step (e) quantifies should be quantifying. It is suggested to amend the claim to say “CpG islands (CGIs).” Appropriate correction is required.
Claim Interpretation
The claim limitation “Cellular Heterogeneity-Adjusted cLonal Methylation (CHALM)” analysis/score appearing throughout the claims is interpreted based on the explanation in paragraph 0067 of the published specification.
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.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claims 1-3, 5, 7, 9, 11, 45, and 48 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.
Regarding claim 1, the limitations “the gene expression and H3K4me3 level” in step (d), “the ratio” in step (e), and “the studied underlying mechanism” in step (f) lack sufficient antecedent basis. For purposes of examination, it will be interpreted as the first instance of the limitations. Additionally, the terms “more accurate” and “more relevant” in step (f) are relative terms which renders the claim indefinite. The terms are not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. It is not possible to determine which hypermethylated genes during oncogenesis and de novo differentially methylated regions are identified because there is no reference point to compare genes or regions to that allows for the determination of whether they are more accurate or more relevant than the reference point. For the purposes of examination, it will be interpreted that associated genes/regions with an underlying mechanism are more accurate and relevant than gene/regions that are not associated with the underlying mechanism.
Furthermore, step (c) recites “wherein the classification-rule is obtained by Cellular Heterogeneity-Adjusted cLonal Methylation (CHALM) analysis of a training-data-set, the training-data-set comprising pre-determined methylation data derived from multitude of pre-classified tumor species, wherein said pre-determined methylation data comprises the methylation status of said CpG positions in the genome of each of said pre-classified tumor species” in product-by-process claim language that renders the scope of the claim indefinite because it is not clear whether the recited limitation must be performed as part of the invention or if the limitation is describing how the classification-rule was obtained outside the metes and bounds of the invention.
Claims 2-3, 5, 7, 9, and 11 are also rejected because they depend from claim 1 and do not resolve the issue of indefiniteness.
Regarding claim 7, the limitations “the purest splitting rules,” “the number of boot strap samples,” “the minority class,” and “the lowest sample size” lack sufficient antecedent basis. Additionally, the meaning of the terms “the purest splitting rues” and “the number of boot strap samples” is not apparent, and the specification does not provide a definition which renders the claim indefinite (MPEP 2173.05(a)). For the purposes of examination, it will be treated as the first recitation of the term and splitting rules will be interpreted as being able to discriminate between various categories of tumor classification. Boot strap samples will be interpreted to mean samples included in the training data. The minority class will be interpreted to mean the tumor type with smallest sample size in the training data.
Regarding claim 45, the limitation “the individual” lacks sufficient antecedent basis. For the purposes of examination, the limitation will be treated as the first instance, and it will be interpreted as the sequencing information received is from an individual suspected of having cancer. Claim 48 is also rejected because it depends from claim 45 and does not resolve the issue of indefiniteness.
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 1-3, 5, 7, 9, 11, 14-15, 20, 24, 26-28, 33, 38, 40, 42-43, 45, 48, and 52 are rejected under 35 U.S.C. 101 because the claimed invention is directed to a judicial exception without significantly more.
Step 1:
The first part of the eligibility analysis evaluates whether a claim falls within any statutory category (MPEP 2106.03). Claims 1-3, 5, 7, 9, 11, 14-15, 20, 24, 26-28, 33, 38, 40, 42-43, 45, and 48 recite a series of steps to analyze methylation sequence data. The claims are directed to a process and fall within one of the statutory categories of invention (Step 1: YES). Although claim 52 is not directed to statutory subject matter, the eligibility analysis will continue in the interest of compact prosecution.
Step 2A, prong 1:
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, law of nature, or natural phenomenon (Step 2A, prong 1). In the instant application, the claims recite the following limitations that equate to those concepts:
Claim 1 recites: (b) determining a methylation profile from a DNA methylation status of a multitude of independent genomic CpG positions in the genome of said tumor-sample; (c) classifying the tumor species of the tumor-sample based on the methylation levels as determined in using a classification-rule, wherein the classification-rule is obtained by Cellular Heterogeneity-Adjusted cLonal Methylation (CHALM) analysis of a training-data-set, the training-data-set comprising pre-determined methylation data derived from multitude of pre-classified tumor species, wherein said pre-determined methylation data comprises the methylation status of said CpG positions in the genome of each of said pre-classified tumor species; (d) predicting the gene expression and H3K4me3 level in promoter CGIs; (e) quantifies the ratio of methylated reads; and (f) identifying more accurate hypermethylated genes during oncogenesis and de novo differentially methylated regions (DMRs) that are more relevant to the studied underlying mechanisms.
Claim 2 recites: wherein the methylation profile comprises any CpG DNA methylation site.
Claim 7 recites: wherein training of the classification-rule comprises: (i) a preceding step of selecting CpG position which of all CpG positions used provide the purest splitting rules, and using said selected CpG positions as a training-data-optimization-set to train the classification-rule; and/or (ii) a step of down-sampling for each tumor species which may include downsampling of the number of boot strap samples to the minority class, the minority class being the lowest sample size of a tumor species in the training-data-set.
Claim 9 recites: including the pre-determined methylation data of the tumor sample as classified in (b) into the training-data-set to obtain an enhanced-training-data-set, and computing an enhanced classification-rule by CHALM analysis based on the enhanced-training-data-set.
Claim 14 and 42 recite: … determining a number of methylated sequence reads associated with the genomic region, or a portion thereof, from the sequencing information, wherein the methylated sequence reads each comprise methylation of at least one qualified CpG site mapped to the genomic region, or the portion thereof; and determining a number of unmethylated sequence reads associated with the genomic region, or the portion thereof, from the sequencing information, wherein the unmethylated sequence reads each comprise at least one qualified CpG site mapped to the genomic region, or the portion thereof, and wherein none of the qualified CpG sites of the unmethylated sequence reads are methylated; determining the CHALM score for the genomic region based on the number of methylated sequence reads associated with the genomic region, or the portion thereof, divided by the sum of the numbers of methylated sequence reads and unmethylated sequence reads associated with the genomic region, of the portion thereof.
Claim 15 recites: wherein the qualified CpG site comprises at least one sequence read covering the CpG site from the sequencing information.
Claim 33 recites: wherein the received sequencing information is subjected to informatics pre-processing prior to determining the number of methylated and/or unmethylated sequence reads.
Claim 38 recites: determining differential methylation associated with the genomic region, or the portion thereof, based on the CHALM score for the genomic region.
Claim 40 recites: correlating the CHALM score for the genomic region with a level of expression of an associated gene and/or an associated H3K4me3 level.
Claim 42 recites: … generating a methylation profile based on the determined CHALM score(s).
Claim 43 recites: determining differential methylation of the one or more genomic regions based on the associated CHALM score.
Claim 48 recites: wherein the CHALM score is indicative of the individual having the cancer.
Claim 52 recites: … determining a number of methylated sequence reads associated with the genomic region, or a portion thereof, from the sequencing information, wherein the methylated sequence reads each comprise methylation of at least one qualified CpG site mapped to the genomic region, or the portion thereof; determining a number of unmethylated sequence reads associated with the genomic region, or the portion thereof, from the sequencing information, wherein the unmethylated sequence reads each comprise at least one qualified CpG site mapped to the genomic region, or the portion thereof, and wherein none of the qualified CpG sites are methylated; and determining a CHALM score for the genomic region based on the number of methylated sequence reads associated with the genomic region, or the portion thereof, divided by the sum of the numbers of methylated sequence reads and unmethylated sequence reads associated with the genomic region, of the portion thereof.
The limitations in claim 1 of determining a methylation profile, classifying the tumor species, obtaining a classification-rule by CHALM analysis, predicting the gene expression and H3K4me3 level in promoter CGIs, quantifying a ratio of methylated reads, and identifying hypermethylated genes and differentially methylated regions recite the mental process of evaluating sequence data, forming conclusions based on the evaluation, and mathematical calculations involved in the data analysis. The limitation in claim 2 further limits the mental process of evaluating sequence data to obtain a methylation profile by specifying that any CpG site DNA methylation site can be included.
The limitations in claim 7 of selecting CpG positions providing the purest splitting rules, using selected CpG positions to train the classification-rule, and down-sampling samples to the lowest tumor sample size in the data encompass the mental process of observing data and deciding which information to exclude from further evaluation and mathematical calculations to form the classification-rule. Lastly, the limitations in claim 9 of including the determined methylation profile in step (b) in claim 1 and computing an enhanced classification-rule by CHALM analysis recite a mental process of deciding to use data in an analysis and mathematical calculations to retrain the classification-rule; respectively.
The limitations in claims 14, 42, and 52 of determining a number of methylated reads, determining a number of unmethylated reads, and determining a CHALM score recite mathematical calculations in the analysis of sequence data. The limitations of claim 15 further limit the mathematical calculations in claim 14 by indicating criteria about which CpG sites are used in the calculation. The limitation in claim 33 of informatics pre-processing recites mathematical calculations to obtain high quality data from raw data. The limitation in claims 38 and 43 of determining differential methylation of a genomic region recites the mental process of evaluating CHALM scores to detect differential methylation.
The limitations in claim 40 of correlating CHALM scores of a genomic region with associated gene expression or H3K4me3 level encompasses the mental process of observing data to discover trends and mathematical calculations to quantify correlation. The limitation in claim 42 of generating a methylation profile encompasses the mental process of grouping and organizing methylation values for genomic regions. Lastly, the limitation of claim 48 further limits the mathematical calculations to obtain a CHALM score in claim 14 by specifying the value of the score pertains to individuals with cancer.
Therefore, these limitations fall under the “Mathematical concepts” and “Mental processes” groupings of abstract ideas (Step 2A, prong 1: YES).
Step 2A, prong 2:
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 (Step 2A, prong 2). The claims recite the following additional elements:
Claim 1 recites: (a) providing a tumor-sample to be classified obtained from a tumor of a patient.
Claim 3 recites: wherein the methylation profile comprises one or more biomarkers obtained from whole-genome bisulfite sequencing (WGBS) or one or more biomarkers obtained from whole-genome enzymatic sequencing.
Claim 5 recites: wherein determining DNA methylation comprises a bisulfite treatment of the DNA or an enzymatic conversion of the DNA.
Claim 11 recites: wherein the biological sample comprises a blood sample, a tissue biopsy sample, or a liquid biopsy sample.
Claim 14 and 42 recite: receiving sequencing information comprising sequence reads…
Claim 20 recites: wherein the sequencing information is obtained from a next-generation sequencing technique.
Claim 24 recites: performing the sequencing technique, wherein the sequencing technique comprises sequencing nucleic acids obtained from a sample from an individual.
Claim 26 recites: wherein the sample is a blood sample comprising cell-free DNA.
Claim 27 recites: wherein the nucleic acids obtained from the sample are subjected to processing prior to sequencing, wherein the processing enables determination of a methylation status of one or more CpG sites of the nucleic acids.
Claim 28 recites: wherein the processing is an enzyme-based technique for the conversion of unmethylated cytosines to enable the determination of the methylation status of one or more CpG sites.
Claim 45 recites: wherein the individual is suspected of having a cancer.
Claim 52 recites: one or more processors; and memory storing one or more programs, the one or more programs configured to be executed by the one or more processors…, receiving sequencing information comprising sequencing reads…
The additional element in claim 1 of providing a tumor-sample from a patient amounts to insignificant extra-solution activity because it is necessary data gathering for use with the judicial exception (MPEP 2106.05.(g)). Similarly, the additional elements: in claim 3 of biomarkers obtained from whole-genome bisulfite sequencing or whole-genome sequencing, in claim 5 of determining DNA methylation with a bisulfite treatment of the DNA or an enzymatic conversion of the DNA, in claim 24 of performing the sequencing technique, and in claims 14, 42, and 52 of receiving sequencing information are data gathering steps necessary to obtain information used by the judicial exception.
The additional elements in claim 11 further limit the proving a tumor-sample step in claim 1 by specifying particular types of samples. Likewise, the additional elements in claims 20 and 45 further limit the receiving steps in claim 14 by specifying how the sequencing information was obtained and from which particular individual. The additional elements in claims 26-27 further limit the sequencing technique step in claim 24 by characterizing the type of sample used, processing the sample prior to sequencing, and specifying the particular processing of an enzyme-based technique.
The additional elements in claim 52 of one or more processors and memory storing one or more programs are invocations of a computer to be used to perform the judicial exception which amounts to instructions to apply the judicial exception in a generic computer environment (MPEP 2106.05(f)).
Therefore, the judicial exception is not integrated into a practical application because the claims do not recite an additional element that reflects an improvement to technology or applies/uses the recited judicial exception in some other meaningful way and the claims are directed to the judicial exception (Step 2A, prong 2: NO).
Step 2B:
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). Claim 52 recite additional elements that equate to mere instructions to apply the recited judicial exception in a generic computing environment. Claims that amount to nothing more than instructions to apply the judicial exception using a generic computer do not render an abstract idea eligible. Alice Corp., 576 U.S. at 223, 110 USPQ2d at 1983. See also 573 U.S. at 224, 110 USPQ2d at 1984. The additional elements recited in claims 1, 3, 5, 11, 24, and 26-28 amount to well-understood, routine and conventional (WURC) activity because paragraph 0023 of the published specification states that a plurality of methods for determining the methylation status in a biological sample are known in the art and paragraph 0101 states sequencing techniques for non-disruptive methylation sequencing are well known in the art. Additionally, paragraph 0110 lists several commercially available next generation sequencing technologies.
Claims 14, 20, 42, 45, and 52 also recite computer functions that the courts have ruled to be WURC such as receiving or transmitting data over a network, e.g., using the Internet to gather data, Symantec, 838 F.3d at 1321, 120 USPQ2d at 1362 (utilizing an intermediary computer to forward information); OIP Techs., Inc., v. Amazon.com, Inc., 788 F.3d 1359, 1363, 115 USPQ2d 1090, 1093 (Fed. Cir. 2015) (sending messages over a network); buySAFE, Inc. v. Google, Inc., 765 F.3d 1350, 1355, 112 USPQ2d 1093, 1096 (Fed. Cir. 2014) (computer receives and sends information over a network).
As such, the combination of additional elements recited in the claims is well-understood, routine, and conventional. The additional elements do not comprise an inventive concept when considered individually or as an ordered combination that transform 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) and claims 1-3, 5, 7, 9, 11, 14-15, 20, 24, 26-28, 33, 38, 40, 42-43, 45, 48, and 52 are not patent eligible.
Claim Rejections - 35 USC § 102
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
The following is a quotation of 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 –
(a)(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.
(a)(2) the claimed invention was described in a patent issued under section 151, or in an application for patent published or deemed published under section 122(b), in which the patent or application, as the case may be, names another inventor and was effectively filed before the effective filing date of the claimed invention.
Claims 14-15, 20, 33, 38, 40, 42-43, and 52 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Xu et al. (bioRxiv preprint, pp. 1-25; IDS document 6/6/2024), as evidenced by Gene Expression Omnibus accession number GSM1186660. The italicized text corresponds to the instant claim limitations.
Regarding claims 14 and 52, Xu et al. teach a methylation quantification method called Cell Heterogeneity-Adjusted cLonal Methylation (CHALM) which quantifies a methylation ratio as methylated reads to total reads mapped to a region (p. 3-4, Main text, paragraph 2) which was demonstrated on several datasets of sequence reads (p. 10-11, Methods, RNA-seq analysis, WGBS data pre-processing) which discloses the claim 14 limitations of a method for determining a Cellular Heterogeneity-Adjusted cLonal Methylation (CHALM) score for a genomic region, the method comprising: receiving sequencing information comprising sequence reads; determining a number of methylated sequence reads associated with the genomic region, or a portion thereof, from the sequencing information, wherein the methylated sequence reads each comprise methylation of at least one qualified CpG site mapped to the genomic region, or the portion thereof; and determining a number of unmethylated sequence reads associated with the genomic region, or the portion thereof, from the sequencing information, wherein the unmethylated sequence reads each comprise at least one qualified CpG site mapped to the genomic region, or the portion thereof, and wherein none of the qualified CpG sites of the unmethylated sequence reads are methylated; determining the CHALM score for the genomic region based on the number of methylated sequence reads associated with the genomic region, or the portion thereof, divided by the sum of the numbers of methylated sequence reads and unmethylated sequence reads associated with the genomic region, of the portion thereof and the same limitations in claim 52 for the steps in program instructions of a system.
Further regarding claim 52, since Xu et al. teach downloading datasets from the Gene Expression Omnibus (GEO) database and using bioinformatics programs/python scripts to pre-process the data (p. 10-11, Methods, RNA-seq analysis, WGBS data pre-processing), it is inherent that a computer was used that contains at least one processor and memory to store the data and programs used. Therefore, these teachings disclose the limitations a system for determining a Cellular Heterogeneity-Adjusted cLonal Methylation (CHALM) score for a genomic region, the system comprising: one or more processors; and memory storing one or more programs, the one or more programs configured to be executed by the one or more processors.
Regarding claim 15, Xu et al. teach that CHALM required an average CpG depth of more than 7x for optimal performance (p. 6, Main text, paragraph 5) which discloses wherein the qualified CpG site comprises at least one sequence read covering the CpG site from the sequencing information.
Regarding claim 20, Xu et al. downloaded raw bisulfite sequencing data of CD3 primary cells from GEO; accession number GSM1186660 (p. 11, WGBS data pre-processing). In the GEO record of the data, the instrument model used to obtain the reads was an Illumina HiSeq 2000 (GSM1186660, Instrument model field) which discloses wherein the sequencing information is obtained from a next-generation sequencing technique.
Regarding claim 33, Xu et al. teach trimming low-quality bases and sequencing adapters from the sequencing data used in their CHALM analysis and aligning reads to a human reference genome before calculating methylation ratios (p. 11, WGBS data pre-processing) which discloses wherein the received sequencing information is subjected to informatics pre-processing prior to determining the number of methylated and/or unmethylated sequence reads.
Regarding claims 38 and 43, Xu et al. teach comparing the CHALM method with traditional methods to identify differentially methylated genes between cancerous and normal tissue samples (p. 7, Main text paragraph 8) which discloses the claim 38 limitation of determining differential methylation associated with the genomic region, or the portion thereof, based on the CHALM score for the genomic region and the claim 43 limitation of determining differential methylation of the one or more genomic regions based on the associated CHALM score.
Regarding claim 40, Xu et al. teach correlating CHALM methylation levels with gene expression (p. 4, Main text, paragraph 3) and H3K4me3 level (p. 5, Main text, paragraph 4) which discloses correlating the CHALM score for the genomic region with a level of expression of an associated gene and/or an associated H3K4me3 level.
Regarding claim 42, Xu et al. generated a methylation profile by virtue of the CHALM analysis performed to examine the relationship of DNA methylation and gene expression and DNA methylation and H3K4me3 level on a CD3 primary cell dataset (p. 4-5, Main text, paragraphs 3-4) show in Figure 2 (p. 22) which discloses a method of generating a methylation profile of one or more biomarkers from a sample from an individual, wherein the one or more biomarkers comprise one or more genomic regions, the method comprising: determining a CHALM score for each of the one or more genomic regions according to claim 14; and generating a methylation profile based on the determined CHALM score(s).
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.
Claim 24 is rejected under 35 U.S.C. 103 as being unpatentable over Xu et al. (bioRxiv preprint, pp. 1-25; IDS document 6/6/2024) as applied to claims 14 and 20, and further in view of Gene Expression Omnibus accession number GSM1186660. The italicized text corresponds to the instant claim limitations.
The limitations of claims 14 and 20 were taught by Xu et al.
Xu et al. appears to be silent on the limitations of claim 24 of … performing the sequencing technique, wherein the sequencing technique comprises sequencing the nucleic acids obtained from a sample from an individual. However, these limitations were known in the prior art before the effective filing date of the invention as taught by the GEO entry GSM1186660.
Regarding claim 24, GEO entry GSM1186660 discloses performing library sequencing using 101 BP PE Illumina technology in the extraction protocol section which discloses … performing the sequencing technique, wherein the sequencing technique comprises sequencing the nucleic acids obtained from a sample from an individual.
An invention would have been prima facie obvious to one of ordinary skill in the art at the time of the effective filing date of the invention if some motivation in the prior art would have led that person to combine the prior art teachings to arrive at the claimed invention. Xu et al. teaches using the sequence data from the GSM1186660 GEO entry and other datasets in order to utilize their method of DNA methylation quantification. Therefore, one of ordinary skill in the art would have been motivated to perform nucleic acid sequencing in order to obtain more data from other samples and study methylation patterns. Furthermore, one of ordinary skill in the art would predict performing sequencing could be readily added to the methylation analysis of Xu et al. with a reasonable expectation of success because the data used in Xu et al. was obtained by sequencing technology. The invention of claim 24 is therefore prima facie obvious.
Claims 26-27, 45, and 48 are rejected under 35 U.S.C. 103 as being unpatentable over Xu et al. (bioRxiv preprint, pp. 1-25; IDS document 6/6/2024) and Gene Expression Omnibus accession number GSM1186660 as applied to claims 14, 20, 24, and 40 above, and further in view of Zhang et al. (US10544467B2; IDS document 6/6/2024). The italicized text corresponds to the instant claim limitations.
The limitations of claims 14, 20, 24, and 40 have been taught by Xu et al. and GEO accession number GSM1186660.
Regarding claim 27, the Extraction protocol section of GEO entry GSM1186660 shows that a bisulfite-seq library preparation protocol was conducted in order to obtain the sequence data (GSM1186660, Extraction protocol) which discloses wherein the nucleic acids obtained from the sample are subjected to processing prior to sequencing, wherein the processing enables determination of a methylation status of one or more CpG sites of the nucleic acids.
Xu et al. and the GEO entry GSM1186660 appear to be silent on the limitations of claims 26, 45, and 48. However, these limitations were known in the prior art at the effective filing date of the invention as taught by Zhang et al.
Regarding claim 26, Zhang et al. teach the biological sample can be blood and contain cell-free DNA (p. 17, col. 12, lines 5 – 15) which discloses wherein the sample is a blood sample comprising cell-free DNA.
Regarding claim 45, Zhang et al. teach identifying when a subject has cancer (p. 17, col. 11, line 64 – col. 12, line 4) which discloses wherein the individual is suspected of having a cancer.
Regarding claim 48, Zhang et al. teach a biomarker indicates a patient has a disease when it is hypo- or hypermethylated respective to a normal sample by a certain percentage (p. 17-18, col. 12, line 60 – col. 13, line 6) which discloses wherein the CHALM score is indicative of the individual having the cancer.
An invention would have been prima facie obvious to one of ordinary skill in the art at the time of the effective filing date of the invention if some motivation in the prior art would have led that person to combine the prior art teachings to arrive at the claimed invention. Zhang et al. teach that high cancer morbidities and mortalities are associated with late diagnosis (p. 12, col. 1, line 20-31). Xu et al. teach CHALM methods provide better identification of hypermethylated promotor CpG islands (CGIs) during tumorigenesis (p. 7-8, Main text, paragraph 8; p. 24, Figure 4). Therefore, one of ordinary skill in the art would have been motivated to utilize the CHALM method in analyzing methylation sequence data in order to accurately diagnose patients suspected of having cancer. Furthermore, one of ordinary skill in the art would predict the CHALM method could be readily added to Zhang et al.’s diagnosis technique with a reasonable expectation of success because both utilize methylation sequence data. The invention of claims 24, 26-27, 45, and 48 is therefore prima facie obvious.
Claim 28 is rejected under 35 U.S.C. 103 as being unpatentable over Xu et al. (bioRxiv preprint, pp. 1-25; IDS document 6/6/2024), Gene Expression Omnibus accession number GSM1186660, and Zhang et al. (US10544467B2; IDS document 6/6/2024) as applied to claims 14, 20, 24, and 27 above, and further in view of Williams et al. (NEB expressions, pp. 1-4). The italicized text corresponds to the instant claim limitations.
The limitations of claims 14, 20, 24, and 27 have been taught by Xu et al., GSM1186660, and Zhang et al.
Xu et al., GSM1186660, and Zhang et al. appear to be silent on the limitations of claim 28. However, these limitations were known in the prior art before the effective filing date of the invention as taught by Williams et al.
Regarding claim 28, William et al. teach alternative approaches for investigating methylomes using enzymatic modification steps (p. 1, Alternative methods for detecting 5mC and 5hmC, paragraph 1; Table 1). Williams et al. teach an enzymatic methyl-seq approach that relies on APOBEC to deaminate cytosines to uracils (p. 2, Enzymatic methyl-seq – a new approach; Figure 3) which discloses wherein the processing is an enzyme-based technique for the conversion of unmethylated cytosines to enable the determination of the methylation status of one or more CpG sites.
An invention would have been prima facie obvious to one of ordinary skill in the art at the time of the effective filing date of the invention if some motivation in the prior art would have led that person to combine the prior art teachings to arrive at the claimed invention. Williams et al. teach that bisulfite libraries have uneven genome coverage and suffer from incomplete representation of cytosine methylation across genomes due to resulting DNA damage and fragmentation from bisulfite conversion (p. 1, Bisulfite sequencing) and that the ability to analyze complete methylomes is important in studying diseases like cancer (Abstract). Therefore, one of ordinary skill in the art would have been motivated to use the enzymatic methyl-seq technique taught in Williams et al. to obtain sequence data in order to obtain more genome coverage of relevant methylation data to determine the presence of cancer. Furthermore, one of ordinary skill in the art would predict that enzymatic methyl-seq could be readily added to the combined teachings of Xu et al., GSM1186660, and Zhang et al. with a reasonable expectation of success because Willaims et al. teach the enzymatic methyl-seq is an alternative to bisulfite sequencing which was the technique used in Zhang et al., Xu et al., and GSM1186660. The invention of claim 28 is therefore prima facie obvious.
Claims 1-3, 5, 7, 9, and 11 are rejected under 35 U.S.C. 103 as being unpatentable over Zhang et al. (US10544467B2; IDS document 6/6/2024) in view of Xu et al. (bioRxiv preprint, pp. 1-25; IDS document 6/6/2024). The italicized text corresponds to the instant claim limitations.
Regarding claim 1, Zhang et al. teach processing extracted genomic DNA obtained from a biological sample of a subject suspected of having a solid tumor to identify if they have cancer (p. 14, col. 5, lines 7-14) disclosing a method for the classification, stratification, and/or diagnosis of a tumor species, the method comprising:(a) providing a tumor-sample to be classified obtained from a tumor of a patient… Zhang et al. teach generating a methylation profile from the extracted genomic DNA (p. 14, col. 5, lines 14-16); the methylation profile may comprise a plurality of CpG data for one or more biomarker (p. 14, col. 5, lines 24-26) disclosing (b) determining a methylation profile from a DNA methylation status of a multitude of independent genomic CpG positions in the genome of said tumor-sample…
Zhang et al. teach inputting training data into a machine learning/classification program to make predictions of the sample based on a methylation profile (p. 14, col. 5 line 40 – 51) disclosing (c) classifying the tumor species of the tumor-sample based on the methylation levels as determined in using a classification-rule, …, the training-data-set comprising pre-determined methylation data derived from multitude of pre-classified tumor species, wherein said pre-determined methylation data comprises the methylation status of said CpG positions in the genome of each of said pre-classified tumor species… Zhang et al. teach in some embodiments, CpG methylation sites are located in promoter regions which leads to downregulation of its’ corresponding gene expression (p. 15, col. 8, lines 12 – 23) and linking differentially methylated markers to gene expression by calculating a beta value (M/(M+U)) (p. 25-26, col. 28, line 63 – col. 29, line 30) which discloses (d) predicting the gene expression … in promoter CGIs; (e) quantifies the ratio of methylated reads; and (f) identifying more accurate hypermethylated genes during oncogenesis and de novo differentially methylated regions (DMRs) that are more relevant to the studied underlying mechanisms.
Regarding claim 2, the above cited teaching of Zhang et al. of generating a methylation profile from the extracted genomic DNA (p. 14, col. 5, lines 14-16) discloses wherein the methylation profile comprises any CpG DNA methylation site.
Regarding claim 3 and 5, Zhang et al. teach methylation analysis procedures include bisulfite treatments and DNA sequencing of untreated and bisulfite-treated DNA (p. 18, col. 13, line 65 – col. 14, line 18); including kits for whole genome amplification to quantify DNA regions (p. 24, col. 26, lines 47 – 54) which discloses the claim 3 limitation of wherein the methylation profile comprises one or more biomarkers obtained from whole-genome bisulfite sequencing (WGBS)… and the claim 5 limitation of wherein determining DNA methylation comprises a bisulfite treatment of the DNA…
Regarding claim 7, Zhang et al. teach in some cases, the machine learning method comprises identifying a plurality of markers and a plurality of weights based on a top score and using those markers and weights to classify samples (p. 14, col. 6, lines 15 – 19) which discloses wherein training of the classification-rule comprises: (i) a preceding step of selecting CpG position which of all CpG positions used provide the purest splitting rules, and using said selected CpG positions as a training-data- optimization-set to train the classification-rule…
Regarding claim 9, Zhang et al. teach generating biological samples and inputting the data into the machine learning/classification program (p. 14, col. 5, lines 43-45). Zhang et al. teach the set of biological samples can comprise 2-30 or more samples (p. 14, col. 5, lines 45-47) which discloses including the pre-determined methylation data of the tumor sample as classified in (b) into the training-data-set to obtain an enhanced-training-data-set...
Regarding claim 11, Zhang et al. teach the biological sample can be blood, tissue, or a variety of liquid biopsy samples (p. 17, col. 12, lines 5 – 40) which discloses wherein the biological sample comprises a blood sample, a tissue biopsy sample, or a liquid biopsy sample.
Zhang et al. appear to be silent on the claim 1 limitations of … wherein the classification-rule is obtained by Cellular Heterogeneity-Adjusted cLonal Methylation (CHALM) analysis of a training-data-set…, predicting H3K4me3 level in promoter CGIs, the claim 7 limitation of … (ii) a step of down-sampling for each tumor species which may include downsampling of the number of boot strap samples to the minority class, the minority class being the lowest sample size of a tumor species in the training-data-set, and the claim 9 limitation of … computing an enhanced classification-rule by CHALM analysis based on the enhanced-training-data-set. However, these limitations were known in the prior art before the effective filing date of the invention as taught by Xu et al.
Regarding claim 1 and 9, Xu et al. teach a methylation quantification method called Cell Heterogeneity-Adjusted cLonal Methylation (CHALM) which quantifies a methylation ratio as methylated reads to total reads mapped to a region (p. 3-4, Main text, paragraph 2) which was demonstrated on several datasets of sequence reads (p. 10-11, Methods, RNA-seq analysis, WGBS data pre-processing) which discloses the claim 1 limitation of … wherein the classification-rule is obtained by Cellular Heterogeneity-Adjusted cLonal Methylation (CHALM) analysis of a training-data-set… and the claim 9 limitation of … computing an enhanced classification-rule by CHALM analysis based on the enhanced-training-data-set. Xu et al. teach correlating CHALM methylation levels with H3K4me3 level (p. 5, Main text, paragraph 4) which discloses the claim 1 limitation of predicting H3K4me3 level in promoter CGIs.
Regarding claim 7, Xu et al. teach down-sampling the original dataset to generate a relatively even-distributed dataset (p. 16, Deep learning prediction, paragraph 5) which discloses … (ii) a step of down-sampling for each tumor species which may include downsampling of the number of boot strap samples to the minority class, the minority class being the lowest sample size of a tumor species in the training-data-set.
An invention would have been prima facie obvious to one of ordinary skill in the art at the time of the effective filing date of the invention if some motivation in the prior art would have led that person to combine the prior art teachings to arrive at the claimed invention. Zhang et al. teach that high cancer morbidities and mortalities are associated with late diagnosis (p. 12, col. 1, line 20-31). Xu et al. teach CHALM methods provides better identification of hypermethylated promotor CpG islands (CGIs) during tumorigenesis (p. 7-8, Main text, paragraph 8; p. 24, Figure 4). Therefore, one of ordinary skill in the art would have been motivated to utilize the CHALM method in analyzing methylation sequence data in order to accurately diagnose patients suspected of having cancer. Furthermore, one of ordinary skill in the art would predict the CHALM method could be readily added to Zhang et al.’s diagnosis technique with a reasonable expectation of success because both utilize methylation sequence data and Xu et al. teach training a convolutional neural network and a linear model on CHALM-determined methylation levels (p. 6-7, Main text, paragraph 7). The invention of claims 1-3, 5, 7, 9, and 11 is therefore prima facie obvious.
Conclusion
No claims are allowed.
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/T.Y.O./Examiner, Art Unit 1685
/OLIVIA M. WISE/Supervisory Patent Examiner, Art Unit 1685
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