Notice of Pre-AIA or AIA Status
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
DETAILED ACTION
Acknowledgement is hereby made of receipt and entry of the communication filed on Jun. 2, 2026. Claims 46, 49-50, 54-55, 60-62 and 69-71 are pending. Claims 50, 54-55 and 61-62 are withdrawn. Claims 46, 49, 60 and 69-71 are currently examined.
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.
(Previous Rejection – Withdrawn) Claims 46-49 and 60 were rejected under 35 U.S.C. 112(b) as being indefinite for failing to particularly point out and distinctly claim the subject matter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. This rejection has the following grounds.
This rejection is withdrawn in view of the amendment filed on Jun. 2, 2026.
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 of this title, 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.
(Previous Rejection – Withdrawn) Claims 46, 48-49 and 60 were rejected under 35 U.S.C. 103 as being unpatentable over CN 106706921 A (published on May 24, 2017; provided in IDS filed on 09/29/2022) in view GenBank: AXY93084.1 (BNLF2b [human gammaherpesvirus 4]. Dated Sep. 18, 2018).
This rejection is withdrawn in view of the amendments filed on Jun. 2, 2026 and in favor of the new rejection below.
(New Rejection) Claims 46, 49, 60 and 69-71 are rejected under 35 U.S.C. 103 as being unpatentable over CN 106706921 A (published on May 24, 2017; provided in IDS filed on 09/29/2022) in view GenBank: AXY93084.1 (BNLF2b [human gammaherpesvirus 4]. Dated Sep. 18, 2018)) and Van Berkel et al. (US 2022/0111065 A1, published on Apr. 14, 2022; PCT filing date May 22, 2019), and further in view of Taherkhani et al. (Ann Lab Med. 2014 Mar;34(2):118-26) and Schmitz (JOURNAL OF CLINICAL MICROBIOLOGY, Aug. 1982, vol. 16, No. 2, p. 361-366).
Base claim 46, as amended, is directed to a kit, which comprises a capture reagent, in which the capture reagent is a first isolated polypeptide; wherein the first isolated polypeptide consisting of 39 to 74 contiguous amino acid residues of wild-type BNLF2b protein of Epstein-Barr virus (EBV) having the sequence of SEQ ID NO: 101, and comprises amino acid residues 14-52, amino acid residues 1-52, amino acid residues 14-74, amino acid residues 11-65, amino acid residue 11-74, or amino acid residues 1-74, with reference to amino acid residue positions set forth in SEQ ID NO: 101;
and wherein the kit further comprises a detection reagent, wherein the detection reagent is a second isolated polypeptide bearing a detectable label; wherein the second isolated polypeptide consisting of 39 to 74 contiguous amino acid residues of wild-type BNLF2b protein of Epstein-Barr virus (EBV) having the sequence of SEQ ID NO: 101, and comprises amino acid residues 14-52, amino acid residues 1-52, amino acid residues 14-74, amino acid residues 11-65, amino acid residue 11-74, or amino acid residues 1-74, with reference to amino acid residue positions set forth in SEQ ID NO: 101.
CN 106706921 A discloses an invention relating to a chip and kit for detecting serum antibodies to the nasopharyngeal carcinoma related EBV. Nasopharyngeal carcinoma can be effectively detected by detecting the levels of the serum EBV antibodies BMRF-1-IgG, LF1-IgA, BNLF2b-IgA, BKRF4-IgG, BMRF1-IgA, BXLF1-IgG, BaRF1-IgG, BKRF3-IgG, BZLF1-E1-IgG and BNLF2b-IgG. The disclosed nasopharyngeal carcinoma test chip and kit have high specificity and sensitivity and can be used for effective screening, early diagnosis and prognosis. See Abstract (human translation is used here to replace the machine translation provided by Applicant).
A human translation of the claims 1-8 of CN 106706921 A is presented below:
1. A chip for detecting nasopharyngeal carcinoma, characterized in that: the chip contains EBV proteins or polypeptides that can detect at least 2 of the following 10 serum antibodies, the 10 serum antibodies being BMRF-1-IgG, LF1-IgA, BNLF2b-IgA, BKRF4-IgG, BMRF1-IgA, BXLF1-IgG, BaRF1-IgG, BKRF3-IgG, BZLF1-E1-IgG and BNLF2b-IgG.
2. The chip according to claim 1, characterized in that: the chip contains EBV proteins or polypeptides that can detect at least 3 out of the 10 serum anti-EBV antibodies.
3. The chip according to claim 1, characterized in that: the chip contains EBV proteins or polypeptides that can detect at least 5 out of the 10 serum anti-EBV antibodies.
4. The chip according to claim 1, characterized in that: the chip contains EBV proteins or polypeptides that can detect all 10 out of the 10 serum anti-EBV antibodies.
5. A kit for detecting nasopharyngeal carcinoma, comprising EBV proteins or polypeptides for quantifying at least 2 of BMRF-1-IgG, LF1-IgA, BNLF2b-IgA, BKRF4-IgG, BMRF1-IgA, BXLF1-IgG, BaRF1-IgG, BKRF3-IgG, BZLF1-E1-IgG and BNLF2b-IgG.
6. The kit for detecting nasopharyngeal carcinoma according to claim 5, characterized in that: the kit comprises EBV proteins or polypeptides for quantifying at least 3 of BMRF-1-IgG, LF1-IgA, BNLF2b-IgA, BKRF4-IgG, BMRF1-IgA, BXLF1-IgG, BaRF1-IgG, BKRF3-IgG, BZLF1-E1-IgG and BNLF2b-IgG.
7. The kit for detecting nasopharyngeal carcinoma according to claim 5, characterized in that: the kit comprises EBV proteins or polypeptides for quantifying at least 5 of BMRF-1-IgG, LF1-IgA, BNLF2b-IgA, BKRF4-IgG, BMRF1-IgA, BXLF1-IgG, BaRF1-IgG, BKRF3-IgG, BZLF1-E1-IgG and BNLF2b-IgG.
8. The kit for detecting nasopharyngeal carcinoma according to claim 5, characterized in that: the kit comprises EBV proteins or polypeptides for quantifying BMRF-1-IgG, LF1-IgA, BNLF2b-IgA, BKRF4-IgG, BMRF1-IgA, BXLF1-IgG, BaRF1-IgG, BKRF3-IgG, BZLF1-E1-IgG and BNLF2b-IgG.
Paragraph [0032] of CN 106706921 A teaches preparation of a protein chip, a human translation is presented below:
“[0032] Dotting of Protein Chips
Tween-20 and protein solution expressed in 6 were added to a 384-well plate, and dotted onto a 16-pad Fast slide coated with NC by a dotting device. In the dotting process, temperature was kept at 20oC-22oC; humidity was kept at 30%-35%; 16 independent and identical arrays were made on one slide, with two duplicated dots for each protein. Refer to the instruction of Boao Smart Arrayer 136 printer for details.”
Paragraphs [0033] and [0034] of CN 106706921 A teach a hybridization process of a protein chip. A human translation is presented below:
“[0033] Protein chip hybridization
1) Put dotted protein chips in 16-well array incubation chamber on FASTrameslide holders;
2) Sealing: adding 100 ul blocking buffer to each pad, incubating at room temperature in a humidity box (putting 3 wet paper tissue on the bottom of the iron box) for 30 min, shaking at 60 rpm/min;
3) add to two 200 ul-PCR tubes each 100 ul low cross buffer-mild; adding mouse anti-His5 to one tube, and rat anti-HA to the other tube, the antibody concentrations being 1:100 and 1:250, respectively, incubating at room temperature for 1 hour, shaking at 150 rpm/min;
4) Serum pretreatment: after thawing serum on ice, add to a 200 ul-PCR tube: 0.5ul serum, 89.5ul low cross buffer-mild, 10ul E.coli lysate, concentration of serum being maintained at 1:200; incubate at RT for 1 hr, shaking at 150 rpm/min.
[0034] 5) suck off the blocking buffer, add 100 ul of pretreated (3) and (4), and put the FASTFrame slide holders in a humidity box, incubate at 4oC overnight, 60 rpm/min.
6) secondary antibody pretreatment: separately add low cross buffer-mild to biotinated-horse-anti-mouse IgG(H+L), biotinated rabbit anti-rat IgG(H+L), biotin conjugated goat anti-human IgG(H+L) and biotin conjugated goat anti-human IgA to make secondary antibody concentrations of 1:400, 1:400, 1:400 and 1:200, respectively; incubate at RT, shaking at 150rpm/min;
7) suck off the liquid from the pad, wash with TBST for three times, 5 mins each;
8) add 100ul of pretreated secondary antibody, incubate in humidity box at RT for 1 hr, 60 rpm/min;
9) repeat step (7);
10) add Cy5-conjugated streptavidin to low cross buffer-mild to make the antibody concentration at 1:200; incubate in dark at RT for 1 hr, 60 rpm/min;
11) transfer the slides to a staining jar, wash twice with TBST, 5 min each wash, wash twice with TBST, 5 min each wash, wash twice with dH2O, 1 min each wash;
12) dry the slide by spinning.”
Accordingly, CN 106706921 A teaches an array chip or kit thereof containing EBV proteins or polypeptides for the detection of serum antibodies specific the EBV proteins, including antibodies specific to BNLF2b (BNLF2b-IgA and BNLF2b-IgG). It teaches that the EBV proteins/polypeptides are dotted onto slides to produce the arrays (attached to surface of a solid support), it also teaches biotin-labeled secondary antibodies. However, it is silent on the sequence information about the EBV BNLF2b gene used in the invention, even though its teachings imply that any EBV BNLF2b protein or polypeptide that detects serum BNLF2b-IgG and BNLF2b-IgA can be used.
GenBank: AXY93084.1 discloses the amino acid sequence of BNLF2b protein of an EBV, isolate “P2-1213”. An alignment between instant SEQ ID NO: 101 and GenBank: AXY93084.1 is shown below:
Score Identities Positives Gaps
196 bits(497) 96/98(98%) 97/98(98%) 0/98(0%)
SEQ1 1 MRPGRPLAGFYATLRRSFRRMSKRSKNKAKKERVPVEDRPPTPMPTSQRLIRRNALGGGV 60
MRPGRPLAGFY+TLRRSFRRMSKRSKNKAKKERVPVEDRPPTPMPTSQRLIRRNALGGGV
Genba 1 MRPGRPLAGFYSTLRRSFRRMSKRSKNKAKKERVPVEDRPPTPMPTSQRLIRRNALGGGV 60
SEQ1 61 RPDAEDCIQRFHPLEPALGVSTKNFDLLSLRCELGWCG 98
RPDAEDCIQR HPLEPALGVSTKNFDLLSLRCELGWCG
Genba 61 RPDAEDCIQRCHPLEPALGVSTKNFDLLSLRCELGWCG 98
Van Berkel teaches the amino acid sequence of an Epstein-Barr virus antigen, SEQ ID NO: 320, that is 100% identical to SEQ ID NO: 101 of the instant application, indicating that the EBV BNLF2b sequence as claimed is publicly known at the time of invention.
Accordingly, both GenBank: AXY93084.1 and Van Berkel teach a full-length BNLF2b of an EBV strain, with teachings of Van Berkel indicate that the EBV BNLF2b sequence as claimed is publicly known at the time of invention.
Taherkhani teaches a study on development of Enzyme-Linked Immunosorbent Assays using 2 truncated ORF2 proteins for detection of IgG antibodies against Hepatitis E Virus (HEV). Taherkhani teaches that two truncated forms of the ORF2 protein were expressed in E coli and were purified by Ni2+-chelate-affinity chromatography, that two ELISAs were developed using these proteins and were compared with DIA.PRO HEV IgG ELISA kit (DIA.PRO. Italy) in 220 serum samples. See Abstract. These teachings indicate that it is a known practice in the art of antibody detection to use truncated antigen polypeptides.
Schmitz teaches that immunoglobulin M (IgM) antibodies to Epstein-Barr virus were detected by using microtiter plates coated with anti-u-chain antiserum and enzyme-labeled Epstein-Barr virus antigen. Optimum conditions for labeling were determined. The addition of unlabeled control antigen to the enzyme-labeled antigen was effective in reducing background reactions. Rheumatoid factor no longer interfered. Blocking of specific IgM antibody by IgG also was no longer observed. Four different methods for the detection of acute Epstein-Barr virus infections were compared. A combination of the enzyme-labeled antigen-IgM test with the detection of antibodies to either Epstein-Barr nuclear antigen or heterophile antigen was highly sensitive and specific. By changing the solid phase, IgA antibodies to Epstein-Barr virus could be detected. See Abstract. Teachings of Schmitz indicate that enzyme-labeling of EBV viral antigens is known and practiced in the field of serum antibody detections, even though Schmitz is silent on the EBV BNLF2b antigen.
It would have been prima facie obvious for one of ordinary skill in the art before the effective filing date of the current invention to arrive, from the teachings of CN 106706921 A, at the sequence characteristics of the polypeptides as claimed through routine experimental optimization, in testing polypeptides containing different lengths and/or regions out of a full-length “wild-type” EBV BNFL2b protein, such as the one disclosed in GenBank: AXY93084.1 and Van Berkel, for immunogenicity as well as feasibility in the assays as disclosed in CN 106706921 A, unless there is evidence that the specified sequence limitations are critical. Since CN 106706921 A already teaches a test for screening or diagnosing nasopharyngeal carcinomas by detecting antibodies against a panel of EBV antigens including the BNFL2b and amino acid sequence of BNFL2b is known in the art, there is a reasonable expectation of success that sequence regions of BNFL2b that are suitable for antibody assay can be identified through routine experimental optimization.
As for the limitation of the truncated BNFL2b polypeptides, teachings of Taherkhani indicate that it is known and practiced in the art of antibody detection that truncated antigen polypeptides can be used. As to the limitation that the claimed kit further comprises as a detect reagent a second polypeptide of the same sequence feature of the first polypeptide bearing a detectable label, teachings of Schmitz indicate that enzyme-labeling of EBV viral antigens is known and practiced in the field of antibody detections associated with EBV infection. It would have been prima facia obvious for one of skill in the art at the time of invention to prepare an enzyme-labeled EBV BNLF2b polypeptide, as claimed, based on the teachings of Schmitz that enzyme-labeled EBV antigens can be used in the detection of antibodies. One would have motivated to do so, e.g., to evaluate an additional detection method known at the time of invention for the assays disclosed in CN 106706921 A. There is a reasonable expectation of success that EBV BNLF2b polypeptides as claimed can be produced and labelled based on the teachings of the cited references.
Response to Applicant’s Arguments
Applicant’ arguments filed on Jun. 2, 2026 have been fully considered. Arguments regarding withdrawn rejections are moot. Applicant’s arguments relevant to the current rejection are addressed on follows.
Applicant argues that CN 106706921 A teaches using full-length protein of BNLF2b-IgG or BNLF2b-IgA, instead of using BNLF2b fragments; Applicant argues that the Coghill ref (Annex A) provided in the previous response did not identify BNLF2b (IgA/IgG) as a promising marker to discriminate between cases and controls. Applicant argues that, therefore, the state of the art when this invention was made was limited to IgA/IgG detection using full-length BNLF2b, which provided only a limited diagnostic performance.
Applicant refers to a prior art reference, Coghill et al. (Clin Cancer Res., 2018; 24(6): 1305-1314; Annex A), stating that IgA and IgG antibody responses against 199 EBV protein sequences in a cross-sectional study of patients with NPC and community controls were measured using a protein microarray (the same method used in CN 106706921), and that the authors did not identify BNLF2b (IgA/IgG) as a promising marker to discriminate between cases and controls. Applicant argues that, therefore, the state of the art when this invention was made was limited to IgA/IgG detection using full- length BNLF2b, which provided only a limited diagnostic performance. Applicant argues that it is well-known that full-length proteins usually contain multiple epitopes, and a person skilled in the art would normally expect full-length proteins to have better or equivalent antigenicity compared to fragments. Applicant argues that since full-length BNLF2b already showed unsatisfactory diagnostic performance in the art, a person of ordinary skill in the art possessing this background knowledge would have lacked any reason or reasonable expectation of success to further develop BNLF2b fragments for improved diagnosis.
Applicant argues that the diagnostic performance of the present application surpasses the state-of-the art serological detection panel (EBNA1/IgA+VCA/IgA for NPC screening, which represents a significant advance over the prior art, and that achieving this advance could not have been predicted based on prior knowledge.
Applicant argues that the research of Lin Ma, et al. (Int J Cancer. 2024;155(10):1874-1885) showed that the detection results based on full-length BNLF2b protein did not demonstrate significant advantages compared to EBNA1/IgA (Table 1). This indirectly demonstrates the advantages of detection based on BNLF2b fragments over full- length BNLF2b as taught by CN 106706921.
Applicant argues that the relevant research based on aa1-74 submitted with the previous response (Li T, et al., NEngl JMed. 2023;389(9):808-819) showed that P85 (aa1- 74) total antibody (P85-Ab) detection provides superior diagnostic results compared to P85-IgA and P85-IgG with a specificity of up to 100%, see Figure 1C-1D and Table S5 in the Supplementary appendix thereof, and that this further supports the forgoing argument that the claimed subject matter achieves unexpectedly high diagnostic accuracy, which is not obvious to a person skilled in the art. Applicant thus argues that the amended claim 46 possesses unexpected advantages and is non-obvious and inventive over the cited documents.
Applicant’s arguments are not persuasive. First, there is no indication in CN 106706921 A that full-length BNFL2b protein is used. Secondly, the claims do not recite any limitations relating the claimed BNLF2b fragments to the detection of EBV-specific antibodies or NPC diagnosis. Therefore, any rationale for the production and labeling of BNFL2b fragments may be used to support an obviousness rejection.
CN 106706921 A teaches an assay for the detection of serum antibodies against a panel of EBV antigens, including BNFL2b, implying that at least one form of BNFL2b antigen polypeptide is produced and used. GenBank: AXY93084.1 and Van Berkel teach the amino acid sequence of EBV BNLF2b that is 98% or 100% identical to SEQ ID NO: 101 of the instant application. Teachings of Taherkhani and Schmitz indicate that it is known and practiced in the art of antibody detection that truncated antigen polypeptides can be used, and that enzyme-labeling of EBV viral antigens is known and practiced in the field of antibody detections. Therefore, it would have been prima facie obvious to combine the teachings of CN 106706921 A, GenBank: AXY93084.1, Van Berkel, Taherkhani and Schmitz to arrive at the invention as claimed, i.e., production of truncated EBV BNLF2b fragments, labeling them and putting the labeled and unlabeled fragments in a “kit”.
As to the argument that the claimed BNFL2b fragments perform better in the diagnosis of nasopharyngeal cancer than the full-length counterpart, to come up with the conclusion that BNFL2b fragments are superior, Applicant relied on results of two separate studies comparing separately a fragment of BNFL2b (P85 (aa 1- 74)) with EBNA1 and a full-length BNFL2b with EBNA1. This comparison is not persuasive since the two cited studies may be performed and/or accessed under different conditions. It would have been more persuasive if Applicant provides results comparing directly the performance of the claimed fragments and the performance of the full-length counterpart.
As to Applicant’s argument about unexpectedness of the claimed invention, as an initial matter, “the burden of showing unexpected results rests on he who asserts them. Thus it is not enough to show that results are obtained which differ from those obtained in the prior art: that difference must be shown to be an unexpected difference.” In re Klosak, 455 F.2d 1077, 1080 (CCPA 1972) (citation omitted). Moreover, “[i]t is well settled that unexpected results must be established by factual evidence. Mere argument or conclusory statements in the specification does not suffice.” In re De Blauwe, 736 F.2d 699, 705 (Fed. Cir. 1984) (citation omitted). Applicant’s attention is directed to MPEP 716.02(b)-(e) for how unexpected results can be established. E.g., to evaluate if the claimed invention produces unexpected results, one must consider if the results produced by the claimed invention are commensurate in scope with the claims and how the results compare with the closest prior art. See MPEP Section 716.02(d) and (e). Here, Applicants provide results for sensitivity and specificity of diagnosing nasopharyngeal cancer by detection of IgAs against three antigens, BNLF2b, EBNA1 or VCA (see Examples 6-7). These results are not commensurate in scope of the invention as claimed, which encompasses a kit comprising a BNLF2b antigen as capture agent. Additionally, comparison between the diagnostic sensitivity and specificity between the processes using BNLF2b as antigen and EBNA1/VCA as antigen cannot be considered as a comparison with the closest prior art, since CN 106706921 A discloses a test for nasopharyngeal carcinoma by detection of IgA/IgG’s against a panel of EBV antigens including BNLF2b. A comparison with the test method disclosed in CN 106706921 A would be more persuasive.
Conclusion
No claims are allowed.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to NIANXIANG (NICK) ZOU whose telephone number is (571)272-2850. The examiner can normally be reached on Monday - Friday, 8:30 am - 5:00 pm, EST. If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, MICHAEL ALLEN, on (571) 270-3497, can be reached. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300. Information regarding the status of an application may be obtained from the Patent Application Information Retrieval (PAIR) system. Status information for published applications may be obtained from either Private PAIR or Public PAIR. Status information for unpublished applications is available through Private PAIR only. For more information about the PAIR system, see http://pair-direct.uspto.gov. Should you have questions on access to the Private PAIR system, contact the Electronic Business Center (EBC) at 866-217-9197 (toll-free). If you would like assistance from a USPTO Customer Service Representative or access to the automated information system, call 800-786-9199 (IN USA OR CANADA) or 571-272-1000.
/NIANXIANG ZOU/
Primary Examiner, Art Unit 1671
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