DETAILED ACTION
Notice of Pre-AIA or AIA Status
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Please note: The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action.
Continued Examination Under 37 CFR 1.114
A request for continued examination under 37 CFR 1.114, including the fee set forth in 37 CFR 1.17(e), was filed in this application after final rejection. Since this application is eligible for continued examination under 37 CFR 1.114, and the fee set forth in 37 CFR 1.17(e) has been timely paid, the finality of the previous Office action has been withdrawn pursuant to 37 CFR 1.114. Applicant's submission filed on 3/9/2026 has been entered.
Election/Restrictions
In the reply filed on 03/28/2025, Applicants elected, without traverse, the particular marker that is SEQ ID NO: 17. As noted in the Office Action of 12/8/2025, the species election requirement (see the Requirement of 01/07/2025) as it applies to the elected marker (i.e.: SEQ ID NO: 17) and the combinations and subcombinations that include SEQ ID NOs: 28, 80, 108 and 193 was withdrawn.
Applicant has amended the claims to require a non-elected species (SEQ ID NO: 190, previously in claim 227). All claims now read on a non-elected invention. However, in the interest of compact prosecution, the examiner is rejoining SEQ ID NO: 190 with the elected marker and previously rejoined SEQ ID NOs. In light of the rejoinder, the examined subject matter is directed to two or more markers, wherein one of the at least two or more markers is at least a portion of a DMR selected from the group consisting of SEQ ID NO: 17, 28, 80, 108, and 193, and wherein one of the at least two markers is at least a portion of SEQ ID NO: 190.
Claim Status
Claims 227 and 238-239 are cancelled.
Claims 221-226, 228-237, and 240-243 are pending and being examined on the merits.
Duplicate Claims Warning
In light of Applicant’s cancellation of claim 238, the duplicate claims warning is withdrawn.
Claim Objections
Claim 231 is objected to because of the following informalities: Claim 231 reads “to capture two or more corresponding methylation locus / loci” and should read “to capture two or more corresponding methylation [[locus / ]]loci” given the plurality of “two or more” in the newly amended claim. Appropriate correction is required.
Claim Rejections - 35 USC § 112b - Indefiniteness
Withdrawn:
The rejection of claims 236-237 and 242 under 35 U.S.C. 112(b) is withdrawn in light of Applicant’s amendments to the claims.
New:
Claim 230 is rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
Claim 230 recites the limitation "the one or more markers" in line 2. There is insufficient antecedent basis for this limitation in the claim. Claim 230 depends from claim 221, which defines “at least two or more markers”. Claim 230 should be amended to recite “the at least two or more markers” to correct this lack of antecedent basis.
Claim Rejections - 35 USC § 103
Withdrawn:
The rejection of claims 221-226, 228-239, and 242-243 under 35 U.S.C. 103 as being unpatentable over Venn et al. (WO 2020163410 A1; published 8/13/2020, priority to 5/2/2019; cited on PTO-892 of 4/17/2025) in view of Ehrich et al. (Nucleic Acids Research 2007) is withdrawn in light of Applicant’s amendments to the claims.
The rejection of claims 240-241 under 35 U.S.C. 103 as being unpatentable over Venn et al. (WO 2020163410 A1; published 8/13/2020, priority to 5/2/2019) in view of Ehrich et al. (Nucleic Acids Research 2007) as applied to claims 221-226, 228-239, and 242-243 above, and further in view of Zeitoun et al. (WO 2019/222706 A1) is withdrawn in light of Applicant’s amendments to the claims.
New:
Claims 221-226, 228-237, and 242-243 are rejected under 35 U.S.C. 103 as being unpatentable over Venn (Venn et al., US 2022/0119890 A1; published 4/21/2022, priority to 1/25/2019) in view of Ehrich et al. (Nucleic Acids Research 2007; cited on PTO-892 of 12/8/2025).
Venn teaches a method of assaying for cancer through targeted assessment of methylation patterns (Abstract).
More specifically, Venn teaches a method of determining a methylation status of markers in enriched DNA fragments…wherein one of the at least two or more markers is a methylation locus comprising at least a portion of a differentially methylated region (DMR) selected from the group consisting of SEQ ID NOs: 2, 6, 15, 24, 25, 26, 28, 35, 41, 49, 55, 60, 69, 80, 86, 90, 92, 99, 108, 124, 159, 193, and 17 (claim 221; paragraph [0007]) and wherein one of the at least two or more markers is a methylation locus comprising at least of portion of DMR chr19:53254542-53254962 (SEQ ID NO: 190). The “target genomic regions” that Venn is assessing read on methylation locus/loci (claim 221; paragraph [0186]). The instant application’s specification defines “at least a portion of” as “at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, or at least 95%” of a DMR selected (paragraph [0104]). Using sequence alignments, the methylation loci that Venn determine the methylation status of are comprised of at least a portion of SEQ ID NOs.: 17, 28, 80, 108, 193, and 190 (relevant to claims 221-226; see SEQ ID NOs.: 337588 (44.3%), 228552 (83.9%), 232583 (100%), 333511 (100%), 12874 (100%), and 191195 (97.6%), respectively). These portions all compris[e] at least (3) CpGs. Given the 100% overlap with SEQ ID NOs: 80, 108, and 193, and the 97.6% overlap with SEQ ID NO: 90, Venn necessarily teaches that the at least two or more markers is a methylation locus comprising at least 80% of the CpGs in a differentially methylated region in the indicated SEQ ID NOs (claim 243).
Each of the above-mentioned methylation loci from Venn are less than or equal to 5000 bp (relevant to claim 221) and less than or equal to 3000 bp (relevant to claim 232).
Venn teaches that the DNA from a human subject can be obtained through a blood sample, a stool sample, or a blood product sample and can comprise cell-free DNA (cfDNA) (claims 221 and 228-229; paragraphs [0150]). Venn teaches that the method can be used for “determining that a test subject has a type of cancer”, one of which is “colorectal cancer” (paragraph [0024, 0027, 0029, and 0114]). This reads on wherein the subject is suspected of having colorectal cancer (claim 233). Additionally, Venn teaches that the method may be used for “staging cancer” and compare various methods of predicting cancer based on methylation pattern differences in early- versus late-stage colorectal cancer, which reads on suspected of having early stage colorectal cancer (claim 234; paragraph [0195, 0233, and 0313] and Table 27).
Venn teaches a method in which the target genomic regions (methylation loci) are enriched by a plurality of bait oligonucleotides (capture baits; claim 221 and 231; paragraph [0007]). Venn teaches converting the DNA and then enriching the converted DNA by contacting the enriched sample with baits (claim 221 and 231; paragraphs [0033, 0281, and 0316]). Upon target region enrichment, Venn et al. teach sequencing the enriched DNA sequences using next generation sequencing for determining the methylation status (claim 230; paragraph [0155]).
Venn teaches determining a read-wise methylation value for each sequence read corresponding to a particular marker by determining a percentage of methylated CpG sites in a sequence read and comparing it to a threshold percentage of methylated CpG sites (claim 242; in order to classify whether it is abnormally methylated/hypermethylated/hypomethylated; paragraphs [0202]).
Venn teaches using a commercial kit such as EZ DNA Methylation-Direct for determining DNA methylation (paragraph [0151]). Venn does not teach modifications to the protocol wherein the denaturation is performed at 93ºC-97ºC (claim 221), conversion is performed at 58ºC-62ºC (claim 235) or wherein the denaturing and conversion steps are repeated at least 5 times (claim 236) or at least 10 times (claim 237). However, optimization of this protocol for bisulfite treatment of DNA for methylation analysis according to these parameters is known in the art, as taught by Ehrich et al.
Ehrich et al. teaches performing denaturation of DNA at 95ºC and conversion at 55ºC for a total of 20 cycles (claims 221, and 235-237; 20 repetitions, Methods-Bisulfite treatment). Ehrich et al. teaches conversion at 50ºC, 55ºC, and 65ºC and demonstrated similar results for smaller amplicons in the cycled version of the bisulfite treatment between the temperatures, with a preference for lower incubation temperatures shown for longer amplicon lengths (Figure 5). The range tested by Ehrich et al. (50ºC to 65ºC) fully encompasses the claimed range of 58ºC-62ºC. It is noted that the courts have held that optimization through routine experimentation, and in particular, when related to differences in temperature, do not support patentability (In re Aller, 220 F.2d 454, 456, 105 USPQ 233, 235 (CCPA 1955)). Thus, the claimed temperature range merely represents routine optimization of the teachings of the cited prior art. In addition, regarding the temperature range claimed in claim 235, it is noted that the courts have stated where the claimed ranges “overlap or lie inside the ranges disclosed by the prior art” and even when the claimed ranges and prior art ranges do not overlap but are close enough that one skilled in the art would have expected them to have similar properties, a prima facie case of obviousness exists (see In re Wertheim, 541 F.2d 257, 191 USPQ 90 (CCPA 1976); In re Woodruff, 919 F.2d 1575, 16 USPQ2d 1934 (Fed. Cir. 1990); Titanium Metals Corp. of America v. Banner, 778 F2d 775. 227 USPQ 773 (Fed. Cir. 1985) (see MPEP 2144.05.01).
It would have been prima facie obvious to one having ordinary skill in the art, before the effective filing date of the instant application, to have modified the method of Venn with Ehrich et al. One would be motivated to do so given the assertion by Ehrich et al. that lower temperatures and cycling through denaturing and conversion “result in higher amplification success for longer amplicons and lower standard deviations on the determination of methylation ratios” (Figure 5 legend). One would have a reasonable expectation of success given that Venn teaches using commercial EZ DNA Methylation kits for bisulfite conversion, and Ehrich et al. teaches using bisulfite commercial kits from Zymo as well and modifying the protocol for optimizing of DNA integrity for downstream applications (Methods-Bisulfite treatment and Abstract).
Claims 240-241 are rejected under 35 U.S.C. 103 as being unpatentable over Venn (Venn et al., US 2022/0119890 A1; published 4/21/2022, priority to 1/25/2019) in view of Ehrich et al. (Nucleic Acids Research 2007; cited on PTO-892 of 12/8/2025) as applied to claims 221-226, 228-237, and 242-243 above, and further in view of Zeitoun et al. (WO 2019/222706 A1; cited on PTO-892 of 12/8/2025).
The teachings of Venn in view of Ehrich et al. as they apply to claims 221-226, 228-237, and 242-243 are detailed above. Relevant to the instantly rejected claims, Venn in view of Ehrich et al. teach determining the methylation status of one or more differentially methylated regions in enriched DNA fragments.
Venn in view of Ehrich et al. do not teach determining a GC dropout rate for the plurality of sequence reads (claim 240) or that the GC dropout rate is less than 6% (claim 241). However, determination of GC dropout rates of a plurality of sequencing reads is known in the art, as taught by Zeitoun et al.
Zeitoun et al. teach determining GC dropout rates of sequencing reads produced from enriched target DNA using polynucleotide bait libraries that perform enrichment through hybridization (Abstract, paragraph [0025, 0082, 0137, 0177, 0194]). Zeitoun et al. teaches achieving less than 6% GC dropout (paragraph [0137, 0194]).
It would have been prima facie obvious to one having ordinary skill in the art, before the effective filing date of the instant application, to have modified the method of Venn in view of Ehrich et al. with that of Zeitoun et al. One would be motivated to calculate GC dropout rate in sequencing reads given the assertion by Zeitoun et al. that calculation of GC dropout rate allows for assessment of the performance of a polynucleotide library for capturing targets (paragraph [0177]). Additionally, Zeitoun et al. teaches that lower GC dropout rates indicate improved capture uniformity (paragraph [0194]). One would have a reasonable expectation of success given that Zeitoun et al. teaches calculating GC dropout rates on enriched DNA that undergoes next generation sequencing, which is the methodology employed for examining differentially methylated regions in Venn.
Response to Remarks
In the Remarks of 3/9/2026 Applicant traversed the rejections as set forth in the Office Action of 12/8/2025. Applicant has amended independent claim 221 and cancelled claim 227 in response to these rejections. In light of the new amendments, new art rejections have been made above.
Conclusion
No claims are allowed.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to KAILEY E CASH whose telephone number is (571)272-0971. The examiner can normally be reached Monday-Friday 8:30am-6pm ET.
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/KAILEY ELIZABETH CASH/Examiner, Art Unit 1683
/STEPHEN T KAPUSHOC/Primary Examiner, Art Unit 1683