DETAILED ACTION
Notice of Pre-AIA or AIA Status
1. The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
2. This action is in response to the amendment filed on 02 February 2026. Applicant's arguments and amendments to the claims have been fully considered but do not place the application in condition for allowance. All rejections not reiterated herein are hereby withdrawn.
The previous rejection of claim 16 under 35 U.S.C. 101 has been obviated by the amendment of claim 16 to recite “amplifying human genomic DNA treated with bisulfite using primers SEQ ID NO: 9-12.” This step was not well-understood, routine or conventional in the prior art and thereby adds something significantly more to the recited judicial exceptions.
The previous rejection of claim 16 under 35 U.S.C. 102(a)(1) as being anticipated by Tao et al (PLoS ONE, 6(5): e19862, 11 pages, 2011), as evidenced by GenBank Accession No AY128659 has been obviated by the amendment to the claim. Tao does not teach or suggest the claimed method for detecting one or more tumors in a subject wherein the method comprises (in part) detecting a modification of a CpG site of a polynucleotide obtained by amplifying bisulfite treated genomic DNA from a tumor (from a subject) with the primers of SEQ ID NO: 9-12 and detecting the modification of a CpG in the polynucleotide as indicative of tumor in the subject wherein the tumor is selected from colorectal cancer, liver cancer and head and neck cancer. Similarly, the previous rejection of claim 16 under 35 U.S.C. 102(a)(1) as being anticipated by Esteller Badosa (WO 2014/173905 A2), as evidenced by Dong et al has been obviated by the amendment of claim 16.
Claim Status
3. Claims 14-16 and 20-28 are pending.
Claims 14, 15 and 20-27 are withdrawn from further consideration pursuant to 37 CFR 1.142(b), as being drawn to a nonelected invention, there being no allowable generic or linking claim.
Claims 16 and 28 read on the elected invention and have been examined herein.
Claim Interpretation
4. Claim 16 has been amended to recite “detecting a modification of a CpG site of a polynucleotide using a tumor detection agent or kit wherein said polynucleotide is a fragment obtained by amplifying human genomic DNA treated with bisulfite using primers SEQ ID NO: 9-12.”
The response states:
“Amended claim 16 recites a step of treating DNA extracted from one or more samples comprising tumor cells with a reagent that modifies DNA in a methylation-specific manner. Furthermore, amended claim 16 recites a polynucleotide wherein the polynucleotide is "a fragment obtained by amplifying human genomic DNA treated with bisulfite using primers SEQ ID NO: 9- 12." Thus, the claimed method recites specific steps for treating extracted DNA using specific reagents to amplify a particular polynucleotide and detect CpG modifications within that polynucleotide.”
The response also states “the polynucleotide obtained by amplification with primers SEQ ID NO: 9-12 in amended claim 16 consists only of residues 240-296 of SEQ ID NO: 1, accounting for only a small portion of the overall length of the sequence in Tao.” (Emphasis added)
Accordingly, claim 16 is considered to require obtaining the polynucleotide by amplifying the bisulfite-treated human genomic DNA with the primers of SEQ ID NO: 9-12 and requires that the polynucleotide consists only of nucleotides 240-296 of SEQ ID NO: 1.
New Claim Rejections - 35 USC § 112(b) - Indefiniteness
5. 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 16 and 28 are rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
Claim 16 is indefinite over the recitation of “treating DNA extracted from one or more samples comprising tumor cells with a reagent that modifies DNA in a methylation-specific manner; detecting a modification of a CpG site of a polynucleotide using a tumor detection agent or kit wherein said polynucleotide is a fragment obtained by amplifying human genomic DNA treated with bisulfite using primers SEQ ID NO: 9-12” because it is unclear as to the relationship between the DNA extracted from the samples and the polynucleotide in which modification of a CpG site is detected. If the polynucleotide is not from the treated DNA extracted from one or more samples comprising tumor cells, then it is unclear as to the source of the polynucleotide. It is also unclear as to the relationship between the subject and DNA extracted from one or more samples, as well as the polynucleotide since the claim does not require that the DNA or polynucleotide are from a sample comprising tumor cells from the subject.
Claim 16 is further indefinite over the recitation of “wherein the detection methods include pyrosequencing” because “the detection methods” lacks proper antecedent basis since the claim does not previously recite the particular language of “a detection method. It is unclear as to how this recitation is intended to further limit the claim and particularly how it relates to detecting a modification of a CpG site of a polynucleotide.
Claim 16 is indefinite over the recitation of “wherein detecting modification of the polynucleotide” because this phrase lacks proper antecedent basis. While the claim previously recites “detecting a modification of a CpG site of a polynucleotide,” the claim does not previously recite a more general step of detecting any modification of a polynucleotide. Claim 28 is indefinite over the recitation of “the polynucleotide amplified in (iii)” because step (iii) does not recite amplifying a polynucleotide. Rather, step (iii) recites “amplifying the genomic DNA treated in (ii).” Claim 28 does not recite a nexus between the genomic DNA and the polynucleotide and thereby it is unclear as to what constitutes the polynucleotide amplified in (iii).
Modified Claim Rejections - 35 USC § 102
6. 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.
Claim(s) 28 is/are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Tao et al (PLoS ONE, 6(5): e19862, 11 pages, 2011), as evidenced by GenBank Accession No AY128659 (NCBI Database, National Library of Medicine, available via URL: <ncbi.nlm.nih.gov/nuccore/AY128659.1/>, 2002).
Tao teaches a method comprising assaying a sample from a subject to determine the methylation status of CpGs in the HIST1H4F gene, wherein the method comprises obtaining a tumor tissue sample from a human subject; extracting genomic DNA from the sample; treating the genomic DNA with bisulfite; amplifying the bisulfite treated genomic DNA; performing combined bisulfite restriction analysis (COBRA) and bisulfite sequencing of the amplified DNA to detect hypermethylation of CpG sites in the amplified HIST1H4F gene sequences (e.g., abstract; p. 8, col. 2 to p. 9; Figure 4; and Tables 3, 4 and 8). Tao teaches amplifying a 380bp sequence of the HIST1H4F gene which includes positions -130 to +250 (relative to the transcription start site; see Table 8 and p. 9, col. 2).
The region amplified in the method of Tao has the property that it includes nucleotides 240 to 296 of present SEQ ID NO: 1. Note that, as evidenced by GenBank Accession No. AY128659, the start site of the HIST1H4F gene is at nucleotide position 312 of the GenBank Accession Number. As shown in the alignment at the end of this Office action, SEQ ID NO: 1 consists of nucleotides 163-861 of the HIST1H4F gene in GenBank Accession No. AY128659 and nucleotides 240-296 of SEQ ID NO: 1 are present at positions 402-459 of GenBank Accession No. AY128659. Thereby, nucleotides 240-296 of SEQ ID NO: 1 are present within the 380bp amplified sequence of Tao that consists of positions -130 to +250 of the HIST1H4F gene.
Note that GenBank Accession No AY128659 is cited only to establish what is inherent to the teachings of Tao.
As shown in Figure 4, HIST1H4F gene sequences (i.e., sequences within SEQ ID NO: 1, which include positions 240-296 of SEQ ID NO: 1) from hepatocytes derived from hepatitis B virus-related hepatocellular carcinoma (HBHC) samples were hypermethylated as compared to HIST1H4F gene sequences from control liver tissue:
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Note that in the above Figure 4 of Tao, black circles represent methylated CpG dinucleotides and white circles represent unmethylated CpG dinucleotides (see legend for Figure 4). Methylated CpG dinucleotides are shown in red in the original article for the sequence of nucleotides -130 to +250 of the HIST1H4F gene.
Thereby, Tao teaches methods of detecting the methylation profile of the 380bp fragment of the HIST1H4F gene which includes nucleotides -130 to +250 of HIST1H4Fand includes nucleotides 240 to 296 of SEQ ID NO: 1. The method of Tao comprises: providing a sample and extracting genomic DNA from the sample; treating the genomic DNA with bisulfite; amplifying the bisulfite treated DNA; and analyzing the amplified, bisulfite-treated DNA by performing bisulfite conversion sequencing or a simplified bisulfite sequencing technology to detect a modified CpG between nucleotides 240-296, with respect to present SEQ ID NO: 1. As discussed above, the 380 bp amplified region of Tao includes numerous CpGs, including CpGs within present nucleotides 240-296, whose methylation status - i.e., modification - is detected in the method of Tao.
Regarding the preamble of claim 28 of “detecting a fragment of SEQ ID NO: 1, said fragment comprising residues 240-296 of SEQ ID NO: 1,” this limitation is only an intended use limitation and does not distinguish the claimed method over that of Tao. Claim 28 does not include any steps of detecting a fragment of SEQ ID NO: 1, said fragment comprising residues 240-296 of SEQ ID NO: 1. Rather, claim 28 recites a final step of analyzing the amplification product to detect methylation of a CpG, wherein the CpG is located between nucleotides 240-296 of SEQ ID NO: 1. As discussed above, Tao teaches this step since the method of Tao detects the methylation status of each of the CpGs in the 380 bp amplification product, including CpGs that correspond to nucleotides 240-296 of present SEQ ID NO: 1.
Response to Remarks:
The response states:
“The Examiner also cites Tao with regard to claim 28, alleging that Tao "teaches that the methylation profile of the 380bp fragment of the HIST1H4F gene which includes nucleotides -130 to +250 of HIST1H4F ... includes nucleotides 240-296 of SEQ ID NO: 1." Office Action, at p. 18. For the same reasons outlined supra for claim 16, Applicant respectfully disagrees..”
These arguments have been fully considered but are not persuasive. Applicant’s arguments regarding claim 16 are directed to limitations recited in claim 16 which are not recited in claim 28. For example, with respect to claim 16, Applicant argues “Tao discloses a 380 base pair portion of the HIST1H4F gene. In contrast, the polynucleotide obtained by amplification with primers SEQ ID NO: 9-12 in amended claim 16 consists only of residues 240-296 of SEQ ID NO: 1, accounting for only a small portion of the overall length of the sequence in Tao.” Claim 28 does not require performing a step in which amplification is performed using the primers of SEQ ID NO: 9-12 to generate an amplification product consisting of only nucleotides 240-296 of SEQ ID NO: 1. The response further argued “amended claim 16 is directed to a method that detects the presence of "one or more tumors from a group consisting of colorectal cancer, liver cancer, and head and neck cancers." However, claim 28 does not recite this limitation.
The rejection is maintained for the reasons set forth above.
7. Claim(s) 28 is/are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Esteller Badosa (WO 2014/173905 A2), as evidenced by Dong et al (Cancer Research. 2019. 79: 6101-6112; published online 01 Oct 2019; cited in the IDS of 01 August 2025).
Esteller Badosa teaches a method for detecting lung cancer, and particularly non-small cell lung cancer (NSCLC), the method comprising obtaining a sample from a subject; extracting genomic DNA from the sample; treating the genomic DNA with bisulfite; amplifying the bisulfite treated genomic DNA; and performing bisulfite sequencing or pyrosequencing of the amplified DNA to detect hypermethylation of CpG sites in the amplified HIST1H4F gene sequences (e.g., p. 4, lines 7-18; p. 18 line 24 to p. 19, line 3; p. 13, lines 1-8; p. 19, line 25 to p. 20 line 19; p. 23 line 31 to p. 24, line 14; p. 28; p. 38, line 21 to p. 39 line 9; and Table 1 and 3).
The reference teaches detecting methylation of CpG dinucleotides in the promoter and adjoining region of the HIST1H4F gene. It is stated (p. 14, lines 21-28) that:
“HISTJH4F promoter, it starts at position 26239153 (according to TSS1500, as in
Infinium HumanMethylation450 BeadChip, Manifest vl.2 or according to UCSC
database, as in Genome Reference Consortium Human Build 37 (GRCh37) and
25 UCSC hg19 as released on February 2009) and ends at position 26241021
( according to 1st exon, as in Infinium HumanMethylation450 BeadChip, Manifest
vl.2 or according to UCSC database, as in Genome Reference Consortium
Human Build 37 (GRCh37) and UCSC hg19 as released on February 2009).”
It is disclosed that the methods therein detect the methylation status of CpG islands located at 26240697-26240951 and CpG shore/island/shore of positions 26238697 to 26242951, which are located within the promoter which starts at 26239153 and exons 1 at 26241021. See Table 1 and p. 18:
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The reference (p. 18 line 24 to p. 19, line 3) further states:
“In a preferred embodiment, the methods according to the invention involve
determining methylation at some specific CpG site( s) which are significantly associated
with a bad prognosis and selection of a stage I NSCLC in a subject. Accordingly, in
another preferred embodiment of the method of the first and/or the second aspect of the
invention, the methylation pattern of said gene(s) is determined at the CpG site(s) located at positions
-26240782 (cg10723962), 26240528 (cg22723502), 26240519 (cg12260798),
26240762, 26240771, 26240774, 26240776, 26240779, 26240789 and/or
26240796 in HIST1H4F.”
Note that the nucleotide positions in chromosome 6 are with respect to GRCh37 / hg19 (p. 19, lines 13-17). See also Table 3 at p. 45 for the location of CpG sites in the HIST1H4F gene that were found to be hypermethylated in NSCLC.
At p. 28, lines 24-32, Esteller Badosa teaches using the following primers to amplify a 253bp region of HIST1H4F to detect the methylation status of 8 CpG dinucleotides present within the amplified region, including the cg10723962 site.
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The region amplified in the method of Esteller Badosa includes nucleotides 240 to 296 of present SEQ ID NO: 1.
This finding is evidenced by the post-filing date art of the present inventors - Dong et al. Dong teaches the occurrence of CpGs that are methylated in cancers in the region of nucleotides chr6: 26,240,743 to 26,240,800, which consists of 57 bp and is the same region as nucleotides 240-296 of present SEQ ID NO :1 (p. 6102, col. 2, final para).
This region (nucleotides 240-296) includes the following CpG sites detected in the method of Esteller Badosa: 26,240,782 (cg10723962), 26,240,762, 26,240,771, 26,240,774, 26,240,776, 26,240,779, 26,240,789 and 26,240,796.
Accordingly, the 253bp region of HIST1H4F amplified in the method of Esteller Badosa includes CpGs within nucleotides 240-296 of SEQ ID NO: 1.
Note that Dong et al is cited only to establish what is inherent to the teachings of Esteller Badosa
Esteller Badosa teaches that the methylation profile of the region of the 253 bp fragment of the HIST1H4F gene is detected. Again, this 253bp region includes CpG dnucleotides within the region of nucleotides 240-296 of SEQ ID NO: 1. Thus, the method of Esteller Badosa includes the step of analyzing an amplified polynucleotide to detect methylation of a CpG, wherein the CpG is located between nucleotides 240-296 of SEQ ID NO: 1.
Regarding the preamble of claim 28 of “detecting a fragment of SEQ ID NO: 1, said fragment comprising residues 240-296 of SEQ ID NO: 1,” this limitation is only an intended use limitation and does not distinguish the claimed method over that of Esteller Badosa. Claim 28 does not include any steps of specifically detecting a fragment of SEQ ID NO: 1, said fragment comprising residues 240-296 of SEQ ID NO: 1. Rather, claim 28 recites a final step of analyzing the amplification product to detect methylation of a CpG, wherein the CpG is located between nucleotides 240-296 of SEQ ID NO: 1. As discussed above, Esteller Badosa teaches this step since the method of Esteller Badosa detects the methylation status of each of the CpGs in the 253 bp amplification product, including CpGs that correspond to nucleotides 240-296 of present SEQ ID NO: 1.
Response to Remarks:
The response states:
“as amended claim 28 recites steps of both "amplifying the genomic DNA treated in (ii) to ampify the fragment of SEQ ID NO:1" and "analyzing the polynucleotide amplified in (iii) to detect methylation of a CpG, wherein the CpG is located between nucleotides 240-296 of SEQ ID NO: 1." Thus, the amended claim incorporates steps for specifically amplifying the fragment in question and for detecting methylation of CpGs on said fragment.”
These arguments have been fully considered but are not persuasive. As set forth in the rejection, the method of Esteller Badosa comprises amplifying a 253bp region of HIST1H4F to detect the methylation status of 8 CpG dinucleotides present within the amplified region, which amplified region includes CpGs located within the region of nucleotides 240-296 of present SEQ ID NO: 1.
8. The prior art made of record and not relied upon is considered pertinent to applicant's disclosure.
GenBank Accession No AY128659 (NCBI Database, National Library of Medicine, available via URL: <ncbi.nlm.nih.gov/nuccore/AY128659.1/>, 2002; previously cited) discloses the nucleotide sequence of the Homo sapiens HIST1H4F gene which consists of a 930bp sequence (“Db” below) that includes a 699bp sequence that has 100% identity with present SEQ ID NO: 1 (“Qy” below) and which as mRNA start site at position 312 (i.e., “mRNA <312..>623 /product="histone H4">:
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Conclusion
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to CARLA J MYERS whose telephone number is (571)272-0747. The examiner can normally be reached M-Th 6:30-5:00 EST.
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If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Wu-Cheng (Winston) Shen can be reached on 571-272-0731. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
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/CARLA J MYERS/Primary Examiner, Art Unit 1682