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 .
Applicant’s arguments and amendments have been thoroughly reviewed and considered. Claims 20-24, 28, and 33-36 have been canceled. Claims 39-45 have been added. Claims 18-19 remain withdrawn. Claims 15-17 and 37-45 are pending and are examined on the merits herein.
Response to Applicant’s Amendments
Claim Objections
Claim 37 was objected to due to a minor informality. In light of Applicant’s amendments to the claims submitted 12/11/2025, this objection has been withdrawn.
35 USC 112(a) Rejections
Claims 15-17, 33-34, and 36-38 were rejected for enablement issues. In light of Applicant’s amendments to the claims submitted 12/11/2025, these rejections have been withdrawn for all currently pending claims. Claims 33-34 and 36 have been canceled, and so these rejections have been rendered moot. However, see new grounds of rejection below.
35 USC 112(b) Rejections
Claims 15-17, 33-34, and 36-38 were rejected for various indefiniteness issues. In light of Applicant’s amendments to the claims submitted 12/11/2025, these rejections have been withdrawn for all currently pending claims. Claims 33-34 and 36 have been canceled, and so these rejections have been rendered moot.
35 USC 103 Rejections
Claims 15, 17, 34, and 37-38 were rejected over Alexandrescu et al. (Journal of Investigative Dermatology, 2012), in view of GenBank (Accession #s EU446494 and M36546.1), in view of van Kempen et al. (Int J Cancer, 2008), in view of Zaenker et al. (Oncotarget, 2009), in view of Ye et al. (BMC Bioinformatics, 2012), and in view of Godfrey (US 2006/0068433 A1). In light of Applicant’s amendments to the claims submitted 12/11/2025, these rejections have been withdrawn for all currently pending claims. Claim 34 has been canceled, and so this rejection has been rendered moot. However, see “Response to Applicant’s Arguments” and new grounds of rejection below.
Response to Applicant’s Arguments
Regarding the 35 USC 112(a) Rejections, Applicant argues that the use of the term “control primer set” provides structural meaning in the context of the claimed kit and should be given patentable weight. This is because the primer set is used to monitor for errors in the real time PCR assay that the kit is used for, where the control mRNA is not differentially present in the sample (Remarks, pages 8-9). Additionally, Applicant argues that the instant specification clearly teaches that SEQ ID NOs: 6-9 can be used as quality controls in the disclosed kits, and specifically that SEQ ID NOs: 6-7 may be used to detect the control mRNA histone H2 (Remarks, page 9).
Firstly, regarding the use of “control” primer sets, Applicant’s Remarks generally reflect a particular use of said primer sets. This use (detecting a control biomarker and monitoring for errors in a real time PCR assay) does not render the actual structure of the primers different than those of primers used for diagnostic purposes, aside from their use in targeting different biomarkers. Thus, prior art that reads on the claimed “control” primer sequences (for those claims which recite particular SEQ ID NOs) will also be considered to read on the claimed “control” primer sets. As to the claims that recite general “control” primer sets, prior art that teaches any specific primer set that differs from the claimed vGAF primer set may read on this limitation (see the “Claim Interpretation” section below).
In the newly amended claims, claims 39, 41, and 44 now recite the use of SEQ ID NOs: 6 and 7. In each of these claims, the use of these sequences to detect histone H2 mRNA is now recited, and gene hist1H3A is no longer recited. As this was the crux of the enablement rejections presented in the Non-Final Rejection, said rejections have been withdrawn for all currently pending claims. However, see new 35 USC 112(a) Rejections related to the use of SEQ ID NOs: 6-7 with histone H2 mRNA.
Regarding the 35 USC 103 Rejections, Applicant argues that the Zaenker reference, which is used to teach detecting vGAF in relation to early-stage melanoma via the use of autoantibodies, teaches vGAF in combination with other biomarkers, and states that the reference notes that the biomarkers used did not necessarily display the highest diagnostic potential (Remarks, pages 12-13). Applicant also argues that Zaenker states that most autoantibodies cannot act as standalone diagnostic tools, and only generate diagnostic power when used as a combination. Furthermore, as Zaenker uses sera samples, there would be no guidance to use vGAF as a marker in testing mRNA expression levels (Remarks, page 13). The reference also allegedly shows no significant difference between patients and controls when comparing early-stage melanoma (stage 0-1 melanoma). Applicant states that Zaenker teaches away from the claimed invention and that the Examiner is utilizing an obvious to try rationale utilizing Applicant’s own specification as a template (Remarks, pages 14-15).
Firstly, Applicant’s arguments are nearly entirely focused on the teachings and alleged deficiencies of Zaenker. In response to Applicant's arguments against the references individually, one cannot show nonobviousness by attacking references individually where the rejections are based on combinations of references. See In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981); In re Merck & Co., 800 F.2d 1091, 231 USPQ 375 (Fed. Cir. 1986). The claims of the Non-Final Rejection were rejected utilizing Alexandrescu, in view of GenBank, in view of van Kempen, in view of Zaenker, in view of Ye, and in view of Godfrey, and so do not rely on the teachings and methodologies of Zaenker alone.
In the Non-Final Rejection, the primary method used is that of Alexandrescu. This reference in particular teaches examining all stages of melanoma utilizing real time RT-PCR with control and target primer and probe sets. This reference also provides a general motivation for measuring biomarkers associated with skin cancer, as well as motivation for specifically utilizing PCR methods over histological/immunohistological methods (see para. 40 of the Non-Final Rejection). GenBank is used to teach the cited primer sequences (see para. 43 of the Non-Final Rejection). Para. 45 recites the teachings of Zaenker. Para. 46 specifically states that it would be prima facie obvious to detect the biomarkers suggested by Zaenker with the primers created using the sequences of GenBank. Para. 47 of the reference further discusses utilizing the method of Alexandrescu with other biomarkers, and specifically discusses the differences between protein analysis with antibody markers (as is taught in Zaenker) versus utilizing RNA and RT-PCR methods. Para. 48 then discusses specifically arriving at the claimed primer sequences using GenBank in view of primer design software, such as that used by Ye. Applicant does not appear to address the actual thrust of the rejections in view of the combination of references, nor does Applicant particularly point out specific objections or allegedly deficiencies in said thrust of the rejections.
With regard to the scope of the instant claims, independent claims 15 and 42 are drawn to kits that include a primer set for measuring vGAF, but are not limited to only utilizing this primer set in a diagnostic capacity, and evidenced by the kits comprising the listed components. Thus, additional primer sets, and additional diagnostic biomarkers, may be utilized with the kits. Thus, additional biomarkers as noted by Zaenker could be used in the combination of references described above, while still reading on the claimed invention. This was considered in the previous combination of references presented in the Non-Final Rejection - in fact, in para. 47 of the Non-Final Rejection, the specific disclosure of the use of “the biomarkers uncovered by Zaenker” is discussed.
With regard to the specific teachings of Zaenker, while the reference does state that single autoantibodies may not provide sufficient sensitivity or specificity, a panel assay, such as that used by the reference, increases both specificity and sensitivity (page 30546, column 2, para. 3). Additionally, though Applicant is correct that Zaenker did not find a significant relationship between the scores for the biomarkers in control and patient samples in the data shown in Figure 3B, there was a significant difference when looking at the larger biomarker panel of Figure 2B. It is also noted that according to Table 1 of the reference, Cohort 2 has a much smaller sample size than that of Cohort 1, including only 20 melanoma patients versus the 104 present in Cohort 1. The ordinary artisan would recognize that this difference in sample size, along with the disparity in comparing the results of Figures 2B and 3B, would not discount the biomarkers taught by Zaenker from being useful in detecting early stage melanoma. Additionally, the scope of the phrase “early stage melanoma” in the instant claims is not defined by Applicant, and so, in a broadest reasonable interpretation, the term can be interpreted as those stages that are not end stage disease. This would include stage 2 patients. Instant claim 38 does specifically note the detection of stage 0-1 patients only, but this still would not discount the results of Zaenker, particularly as Cohort 1 (which showed significant results in both Figure 2A and 3A) is comprised of over 80% stage 0-1 melanoma patients.
Regarding the argument that Zaenker teaches away from the instant invention, MPEP 2145 X (D) 1 states, “Furthermore, "the prior art’s mere disclosure of more than one alternative does not constitute a teaching away from any of these alternatives because such disclosure does not criticize, discredit, or otherwise discourage the solution claimed…." In re Fulton, 391 F.3d 1195, 1201, 73 USPQ2d 1141, 1146 (Fed. Cir. 2004). See also UCB, Inc. v. Actavis Labs, UT, Inc., 65 F.4th 679, 692, 2023 USPQ2d 448 (Fed. Cir. 2023) ("a reference does not teach away if it merely expresses a general preference for an alternative invention but does not criticize, discredit or otherwise discourage investigation into the invention claimed.") (internal quotations omitted) (quoting DePuy Spine, Inc. v. Medtronic Sofamor Danek, Inc., 567 F.3d 1314, 1327 (Fed. Cir. 2009)); and Schwendimann v. Neenah, Inc., 82 F.4th 1371, 1381, 2023 USPQ2d 1173 (Fed. Cir. 2023).” Zaenker is drawn to the use of autoantibodies, but nowhere in the reference is the use of mRNA expression discouraged. Furthermore, though the sensitivity and specificity restrictions for using autoantibodies alone may still apply to the analogous mRNA, Zaenker teaches a panel of biomarkers, and the use of such a panel would not be prohibited by the instant claims. MPEP 2141.03 I states, “‘A person of ordinary skill in the art is also a person of ordinary creativity, not an automaton.’ KSR Int'l Co. v. Teleflex Inc., 550 U.S. 398, 421, 82 USPQ2d 1385, 1397 (2007). "[I]n many cases a person of ordinary skill will be able to fit the teachings of multiple patents together like pieces of a puzzle." Id. at 420, 82 USPQ2d 1397. Office personnel may also take into account "the inferences and creative steps that a person of ordinary skill in the art would employ." Id. at 418, 82 USPQ2d at 1396.” Thus, the Examiner’s position is that the ordinary artisan could utilize the teachings of Zaenker with the additional cited references in the Non-Final Rejection, along with the knowledge, skill, and creativity available to someone of ordinary skill in the art, to arrive at the claimed invention, and that the teachings of Zaenker in combination with those of the additional cited references do not teach away from the claimed invention, nor render the scope of the combination outside of that stated by the instant claims.
Regarding the argument that the 35 USC 103 Rejections are utilizing an “obvious to try” rationale, the Examiner makes no such assertions in the Non-Final Rejection. The Examiner also makes no references to the Applicant’s disclosure in the rationale for the rejections. As noted above, the Examiner has provided a rationale/motivation and reasonable expectation of success for combining the cited references, and as Applicant does not specifically address said rationale, it is considered proper.
In light of Applicant’s amendments to the claims, new grounds of rejection are required to address each of the limitations of the instant claims. However, the relevant portions of the previously utilized rejections have been reiterated below.
Claim Interpretation
With regard to the prior art rejections below, the following interpretations have been made:
In claim 15, a “diagnostic” primer set as stated in (a) of instant claim 15 is not defined in the instant specification, and so this limitation is considered to be met if the desired primer sequences are generally added to the kit, regardless of whether or not they are intended to be used for diagnostic purposes. It is noted that the primer set is stated to be used for detecting vGAF expression in a sample, but this sample is not required to be present in the kit. As this use of the diagnostic primer set imparts no additional structural requirements on the diagnostic primer set, this will be considered intended use. See MPEP 211.02 II. The primer set will thus be interpreted to be a primer set for hybridizing to a vGAF mRNA.
In claim 16, a control primer set is described, where the control primer is used for the detection of a control RNA. The claim then goes on to describe functions related to the control mRNA. However, claim 16 does not specifically require that the control mRNA be present in the kit. It is also noted that a “control” primer set is not specifically defined in the instant specification, and in the claim, the primer set is used for simply hybridizing to a sequence, with no additional structure of the primer itself being recited. As the detection of the control mRNA imparts no additional structural requirements on the control primer set, this will be considered intended use. See MPEP 211.02 II. This claim will therefore be interpreted as requiring at least one additional primer set compared to the kit described by instant claim 15, where the additional primer set hybridizes specifically to any additional sequence.
In claim 37, a control mRNA is required in the kit, where the control mRNA specifically must contain at least vGAF mRNA derived from a healthy individual. The claim goes on to recite a potential relationship between mRNA expression levels, but this relationship does not impart any additional structure to the control mRNA or to the other components of the kit of claim 15. The wherein clause describing this relationship is therefore considered non-limiting.
In claim 42, a “diagnostic” primer set is being used and is interpreted in the same manner as in instant claim 15. The “first control” primer set has a required forward and reverse sequence, and additional limitations are recited related to the “first control mRNA,” but this is considered intended use, similar to the interpretation of instant claim 16. The first control primer set will be interpreted to be particular primer sequences that simply hybridize specifically to an additional sequence other than vGAF mRNA.
In claim 43, the recited “collagen mRNA” is noted to not specifically be tied to the “first control mRNA” of (b) of instant claim 42, and so the limitations described by instant claim 42 regarding the “first control mRNA” are not required of the “collagen mRNA” of claim 43.
In claim 44, the “second control” primer set will be interpreted similarly to the “first control” primer set of instant claim 42, where it will interpreted to consist of particular primer sequences that simply hybridize specifically to an additional sequence.
In claim 45, the recited “histone H2 mRNA” is noted to not specifically be tied to the “second control mRNA” of instant claim 44, and so the limitations described by instant claim 44 regarding the “second control mRNA” are not required of the “histone H2 mRNA” of claim 45.
It is noted generally that a “histone H2 mRNA” includes any histone H2, and so may include variant or modified forms.
Claim Rejections - 35 USC § 112(a)
The following is a quotation of the first paragraph of 35 U.S.C. 112(a):
(a) IN GENERAL.—The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor or joint inventor of carrying out the invention.
The following is a quotation of the first paragraph of pre-AIA 35 U.S.C. 112:
The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor of carrying out his invention.
Claims 39-41 and 44-45 are rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, because the specification, while being enabling for primers for targeting vGAF and collagen mRNA, does not reasonably provide enablement for primers for targeting histone H2 mRNA. The specification does not enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and/or use the invention commensurate in scope with these claims.
Factors to be considered in determining whether a disclosure meets the enablement requirement of 35 USC 112, first paragraph, have been described by the court in In re Wands, 8 USPQ2d 1400 (CA FC 1988). These factors include (1) the quantity of experimentation necessary, (2) the amount of direction or guidance presented, (3) the presence or absence of working examples, (4) the nature of the invention, (5) the state of the prior art, (6) the relative skill of those in the art, (7) the predictability or unpredictability of the art, and (8) the breadth of the claims. Each of these factors is discussed below.
Nature of the Invention
Claims 39-41 and 44-45 are directed to kits adapted for detection of early stage skin cutaneous melanoma comprising particular primers, a probe, and a reverse transcriptase enzyme.
The claimed methods are classified in the unpredictable arts of molecular biology and biochemistry.
Breadth of the Claims
Claim 39 depends on claim 15 and requires the use of at least two (and up to three) specific primer pairs, but does not overall limit the aspects of the kit to a particular group of components (i.e. the kit comprises the listed components). The specific primers that must be used are listed in the accompanying Sequence Listing for the application, and are there labeled with their associated genes (vGAF for SEQ ID NOs: 4-5, hist1H3A for SEQ ID NOs: 6-7, and Col1A1 for SEQ ID NOs: 8-9).
Claim 40 depends on claim 39 and specifies two specific primer pairs that must be used, but does not further limit the broad aspects of claim 39.
Claim 41 specifies that all three primer pairs described in claim 39 (including that of the preceding claim 15) must be used, but does not further limit the broad aspects of claim 39.
Claim 44 depends on claim 42 and requires the use of at least three specific primer pairs, but does not overall limit the aspects of the kit to a particular group of components (i.e. the kit comprises the listed components). The specific primers that must be used are listed in the accompanying Sequence Listing for the application, and are there labeled with their associated genes (vGAF for SEQ ID NOs: 4-5, hist1H3A for SEQ ID NOs: 6-7, and Col1A1 for SEQ ID NOs: 8-9).
Claim 45 depends on claim 44 and adds a particular mRNA to the kit, but does not further limit the broad aspects of claim 44.
Level of Skill in the Art
The ordinary artisan typically holds at least a master’s degree has several years of experience.
State of the Prior Art & Unpredictability
Concerning the claimed invention, what was (and is) unpredictable in the art is whether the primers taught by the instant application to be associated with the histone H2 mRNA (SEQ ID NOs: 6-7) are actually able to hybridize to histone H2 mRNA. It is noted that this primer set is listed as a “control” primer set, though this phrase does not confer any particular structural meaning in the context of the kit, and so searching of the prior art is not limited to instances in which these primers of histone H2 mRNA act as a control biomarker.
In utilizing the National Center for Biotechnology Information (NCBI) BLAST system, SEQ ID NO: 6 appears to align with histone H3, with no results showing alignment with histone H2. One result shows a “histone H2A-like protein gene,” but is not specifically a histone H2A protein. See BLAST results below (note that these results are limited to those for Homo sapiens):
PNG
media_image1.png
892
1110
media_image1.png
Greyscale
In utilizing this software for SEQ ID NO: 7, similar results are shown. See BLAST results below:
PNG
media_image2.png
1013
1146
media_image2.png
Greyscale
In searching the sequences in typical search engines, Lim et al. (Mechanisms of Aging and Development, 2025) was found to teach instant SEQ ID NOs: 6 and 7 as the forward and reverse primer for performing PCR on H3 (see Table 1). In searching the sequences in sequencing databases, nowhere except in Applicant’s own work are SEQ ID NOs: 6 and 7 connected with histone H2.
Guidance in the Specification and Examples
In the instant specification, SEQ ID NOs: 6 and 7 are briefly discussed on page 3, para. 1 and page 23, para. 7, where it is stated that these sequences are used to amplify the histone H2. In the working examples, a histone variant H2AX is examined in conjunction with vGAF (see Examples 2 and 4, which discuss skin UV damage and DNA repair). H2AX is also shown in Figures 1 and 5. In these examples, SEQ ID NOs: 6 and 7 are not described, and their use is not stated.
In view of the unpredictability in the art discussed above, it is not clear that SEQ ID NOs: 6 and 7 can actually be used with histone H2 mRNA.
Quantity of Experimentation
The ordinary artisan would have to conduct a very large quantity of highly unpredictable experimentation before being able to successfully practice the full scope of the claimed methods. Specifically, the ordinary artisan would have to determine that SEQ ID NOs: 6 and 7 are actually capable of use with histone H2 mRNA. Based on the lack of teachings in the prior art and in the instant specification, this would be an inventive and unpredictable undertaking, requiring extensive experimentation in which there is no guarantee of success. The large quantity of experimentation and its unpredictability constitute undue experimentation.
Conclusion
In view of the foregoing, it is clear that the specification fails to enable the full scope of the claimed methods, and claims 39-41 and 44-45 are rejected under 35 U.S.C. 112(a) for failing to comply with the enablement requirement.
Claim Rejections - 35 USC § 103
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 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
Claims 15-16 and 37-38 are rejected under 35 U.S.C. 103 as being unpatentable over Alexandrescu et al. (Journal of Investigative Dermatology, 2012), in view of GenBank (Accession #M36546.1), in view of Zaenker et al. (Oncotarget, 2009), in view of Ye et al. (BMC Bioinformatics, 2012), and in view of Godfrey (US 2006/0068433 A1).
Alexandrescu teaches measuring the expression levels of biomarkers in melanoma (Abstract). The types of melanomas examined ranged from early stage to advanced (Table 1). A quantitative real-time RT-PCR was performed involving a control gene marker and specific primer and probes sets (pages 1888-1889, “Reverse transcription-PCR assay”). This RT-PCR utilized a reverse transcriptase (page 1892, column 1, para. 6). Expression levels were compared between melanoma and non-cancerous skin samples (Tables 3-4 and Figure 1). Samples were all taken from skin biopsies (page 1887, “Patient clinical and pathological characteristics”). The reference states that, “Cutaneous melanoma is the malignancy with the fastest growing incidence in the United States, with an annual percentage increase of 2.8% in the last two decades (SEER, 2008), and reaching epidemic proportions in some parts of the world. It is estimated that 62,000 persons will be diagnosed with melanoma in 2008 in the United States, among which 8,200 will die of this disease (SEER, 2008),” and notes that current diagnostic tools primarily rely on histological and immunohistological examinations (page 1887, column 1, para. 1 and column 2, para. 2). The authors conclude that, “…we have shown the potential utility of this RT-PCR assay using melanoma-specific markers that can reliably differentiate melanoma from non-melanoma lesions.” (page 1891, column 2, para. 5).
However, the primers taught by Alexandrescu do not comprise the sequences taught by the instant claims.
As noted in the Sequence Listing, SEQ ID NOs: 4-5 are associated with vGAF.
GenBank Accession #EU446494 comprises SEQ ID NOs: 4-5 and is a construct for the ZBTB7B gene (also known as vGAF).
Zaenker teaches the development of diagnostic tests for early-stage melanoma (Abstract). This test utilizes antibodies for antigens that can differentiate between primary melanoma patients and healthy individuals (Abstract). The patients examined were specifically diagnosed with early-stage melanoma, and these patients were compared to healthy controls (page 30540, column 2, para. 1). ZBTB7B (the gene encoding vGAF) was shown to be in a combination of sensitive and specific biomarkers for detecting melanoma (page 30544, columns 1-2 joining para.). Generally, the biomarker intensity in the melanoma patients was higher than that of the normal samples (Figure 3). Blood samples from patients were used and underwent protein profiling (page 30547).
Prior to the effective filing date of the claimed invention, it would have been prima facie obvious for one of ordinary skill in the art to use the method of Alexandrescu to detect the biomarkers suggested by Zaenker with primers created using the sequences taught by GenBank. Alexandrescu provides rationale for wanting to study melanoma biomarkers (as melanoma detection may prevent metastases and/or improve patient outcomes) and for using real-time RT-PCR with primers and probes to perform biomarker detection, as this method can analyze a large number and variety of melanoma tissues from biopsy samples, which are often taken when a patient presents with an unknown and/or suspicious lesion. As these biopsies are already part of routine care, analyzing them via RT-PCR allows for a relatively quick way to measure biomarker expression that can be compared with results of histological analyses. In particular, RT-PCR may reveal expression levels that can indicate very early stages of melanoma that would be difficult or impossible to detect in histological slides, which would allow for earlier disease interventions.
It would be obvious to the ordinary artisan that the method of Alexandrescu could be used with other biomarkers. Zaenker discusses measuring antibody markers through a protein analysis, but this analysis came from blood sample determinations, which occurred after an initial tumor diagnosis and excision (page 30547, columns 1-2 joining para.). Thus, as Alexandrescu teaches measuring biomarkers in tumor biopsies, and the methods of Zaenker uncovered additional melanoma biomarkers, it would be obvious to analyze the biomarkers uncovered by Zaenker earlier in the melanoma discovery and analysis process (i.e. in the initial biopsy sample), to measure all potential biomarkers at the same time, and in the same manner (i.e. utilizing RNA and RT-PCR). As the ZBTB7B gene is known, as evidenced by GenBank and Zaenker, such RNA targeting would be possible. It is also noted that Alexandrescu and Zaenker both teach analysis of early stage melanoma samples, and the ordinary artisan would be particularly interested in diagnosing such samples, as early diagnosis would produce the best patient outcomes.
GenBank teaches the sequences of the claimed biomarker (i.e. ZBTB7B) in humans, and methods of appropriate primer design using publicly available software would be available to the ordinary artisan, such as NCBI Primer-BLAST (Ye Abstract). In view of the above, the ordinary artisan would have been motivated to use the publicly available software disclosed in Ye, the teachings of Zaenker, and the known vGAF sequence described by GenBank by to design primers for use in the method of Alexandrescu to detect melanoma biomarkers with real-time RT-PCR, and in the absence of unexpected results, the claimed polynucleotides (instant SEQ ID NOs: 4-5) simply represent the result of an obvious series of steps.
Additionally, including PCR reagents (including reverse transcriptase) and the recited primers in a kit for performing the method of Alexandrescu, in view of GenBank, in view of Zaenker, and in view of Ye would have been obvious to the ordinary artisan at the time of the invention in view of the teachings of Godfrey. In particular, since Godfrey teaches that kits can be “particularly useful” for commercializing nucleic acid detection methods that make use of reagents that include oligonucleotides such as nucleic acid primers (para. 112), the ordinary artisan would have recognized that the detection methods disclosed by Alexandrescu, in view of GenBank, in view of Zaenker, and in view of Ye could also be commercialized by providing the reagents used to practice the methods (e.g., primers, probes, dNTPs, etc.) in a packaged kit. Accordingly, the ordinary artisan would have been motivated to so provide kits containing the detection reagents to obtain the ability to commercialize the disclosed detection method. The ordinary artisan would have had a reasonable expectation of success in view of the guidance provided by Godfrey in paras. 112-115.
Therefore, claim 15 is prima facie obvious over Alexandrescu, in view of GenBank, in view of Zaenker, and in view of Ye, and in view of Godfrey.
Regarding claim 16, Alexandrescu and Zaenker teach the use of controls with their methods, and Alexandrescu in particular teaches using control, benign samples from healthy individuals when comparing expression levels of target biomarkers, as well as using a control gene expression marker (page 1889, column 1, para 2, Table 3, and Figure 1, for example). Page 1892, column 1, paras. 5-6 of Alexandrescu specifically notes that this control gene was used in the RT-PCR assay, where primers were used for each gene. As noted above, Godfrey teaches that primers can be included in kits, as well as control nucleic acids (para. 112), and the reference specifically teaches that these control nucleic acids can be control mRNAs (claim 20). Thus, it would be prima facie obvious to the ordinary artisan that controls from healthy (i.e. non-melanoma) samples could be included in the kit, where the samples would contain the mRNA for vGAF, as well as control primers for analyzing these healthy controls in a PCR assay. Providing these controls in the kit would eliminate the need for the user of the kit to solicit donations from volunteers for control samples or to design their own control primers, thus simplifying the overall protocol, enhancing the ease of use of the kit, and keeping control/normalization procedures similar across different uses and users of the kit. As Godfrey already teaches that nucleic acids can be included in the kit, and teaches that kits can also comprise reagents for extracting mRNAs from samples (para. 115) there would be a reasonable expectation of success.
Regarding claim 37, as described above in the rejection of claim 16, it would be prima facie obvious to include control samples and control gene markers in the kit of Alexandrescu, in view of GenBank, in view of Zaenker, and in view of Ye, and in view of Godfrey. In Alexandrescu, the same biomarkers are compared for melanoma and benign samples (e.g. Table 3). Thus, in the method of Alexandrescu, in view of GenBank, in view of Zaenker, and in view of Ye, and in view of Godfrey, it would also be prima facie obvious to compare the same markers, including vGAF, between control and target samples. Using the control samples in this way would ensure they provide a useful and accurate comparison to target samples, which would lead to the most accurate diagnostics.
Regarding claim 38, as noted above, Alexandrescu and Zaenker both consider early-stage melanoma. Alexandrescu examines all tumor stages (Table 1). Zaenker specifically examines Stage 0-II tumors, with a focus on Stage 0 and Stage 1 (Table 1, note the number of Stage II patients compared to those of the other stages). Thus, it would be prima facie obvious to use the kit on early melanoma stages, such as Stage I.
Claim 17 is rejected under 35 U.S.C. 103 as being unpatentable over Alexandrescu et al. (Journal of Investigative Dermatology, 2012), in view of GenBank (Accession #M36546.1), in view of Zaenker et al. (Oncotarget, 2009), in view of Ye et al. (BMC Bioinformatics, 2012), in view of Godfrey (US 2006/0068433 A1), and further in view of Latham et al. (WO 2012/109326 A2).
Regarding claim 17, Alexandrescu, in view of GenBank, in view of Zaenker, and in view of Ye, and in view of Godfrey teaches the kits of claims 15-16 and 37-38, as described above. However, none of these references teach the use of histone H2 mRNA or collagen mRNA.
Latham generally teaches measuring expression levels of genes in test samples, and comparing those expression levels to that of controls. Para. 78 teaches that expression level comparisons can be normalized against the levels of reference genes, where one such reference gene may be histone H2A. This normalization allows for test sample to control sample comparisons. In para. 126, Latham specifically notes a kit that may include internal standards, such as histone H2A mRNA. The reference also teaches the use of quantitative RT-PCR (e.g. paras. 30, 78, and 83).
Prior to the effective filing date of the claimed invention, it would have been prima facie obvious for one of ordinary skill in the art to substitute the reference gene for normalization already described by Alexandrescu, in view of GenBank, in view of Zaenker, in view of Ye, and in view of Godfrey with that described by Latham. As noted above in the rejection of claims 15-16, Alexandrescu teaches the use of a control gene marker, TYR (page 1889, column 1, para. 2). This gene is explicitly used for normalization of target gene marker expression levels within the reference, as described in Tables 3 and 4, for example. Latham describes the same process with different reference genes, and in particular with histone H2A mRNA. Substituting TYR for histone H2A would therefore amount to simple substitution. MPEP 2143 I (B) states, “The rationale to support a conclusion that the claim would have been obvious is that the substitution of one known element for another yields predictable results to one of ordinary skill in the art.” As both TYR and histone H2A RNA are shown being successfully utilized by Alexandrescu and Latham, respectively, for the same purpose, their substitution is considered to generate predictable results.
Therefore, claim 17 is prima facie obvious over Alexandrescu, in view of GenBank, in view of Zaenker, and in view of Ye, in view of Godfrey, and further in view of Latham.
Claims 16 and 39-40 are rejected under 35 U.S.C. 103 as being unpatentable over Alexandrescu et al. (Journal of Investigative Dermatology, 2012), in view of GenBank (Accession #s EU446494 and M36546.1), in view of Zaenker et al. (Oncotarget, 2009), in view of Ye et al. (BMC Bioinformatics, 2012), in view of Godfrey (US 2006/0068433 A1), and further in view of van Kempen et al. (Int J Cancer, 2008).
Regarding claims 39-40, Alexandrescu, in view of GenBank, in view of Zaenker, and in view of Ye, and in view of Godfrey teaches the kits of claims 15-16 and 37-38, as described above. However, none of these references teach the use of histone or collagen primer sets.
As noted in the Sequence Listing, SEQ ID NOs: 8-9 are associated with Col1A1 (collagen type I alpha 1 chain).
GenBank Accession #M36546.1 comprises SEQ ID NOs: 8-9 in association with alpha-1 collagen type 1.
van Kempen teaches Col1a1 mRNA expression in melanoma cells (Figure 1b-c and Figure 2). They conclude that, “In early stages of melanoma development and progression, Col(I) synthesis contributes to the development of a tumor vasculature, whereas its inhibition in advanced stages after a proper vasculature is formed, can contribute to further progression,” (page 1028, column 2, para. 1), indicating that levels of Col1a1 may be predictive of the presence and progression of melanoma. van Kempen also states that controls were used to compare expression levels (e.g. page 1020, column 2, paras. 5-6, page 1021, column 1, para. 1, and page 1023, column 1, para. 2).
Prior to the effective filing date of the claimed invention, it would have been prima facie obvious for one of ordinary skill in the art to use the method of Alexandrescu, in view of GenBank, in view of Zaenker, in view of Ye, and in view of Godfrey to detect the biomarker suggested by van Kempen with primers created using the sequences taught by GenBank. Alexandrescu provides rationale for wanting to study melanoma biomarkers (as melanoma detection may prevent metastases and/or improve patient outcomes) and for using real-time RT-PCR with primers and probes to perform biomarker detection, as this method can analyze a large number and variety of melanoma tissues from biopsy samples, which are often taken when a patient presents with an unknown and/or suspicious lesion. As these biopsies are already part of routine care, analyzing them via RT-PCR allows for a relatively quick way to measure biomarker expression that can be compared with results of histological analyses. In particular, RT-PCR may reveal expression levels that can indicate very early stages of melanoma that would be difficult or impossible to detect in histological slides, which would allow for earlier disease interventions.
It would be obvious to the ordinary artisan that the method of Alexandrescu, in view of GenBank, in view of Zaenker, in view of Ye, and in view of Godfrey could be used with other biomarkers. van Kempen teaches that Col(I) expression differs between early and late stage melanoma (see Figure 7 for example), where expression is increasing during early stage disease and decrease in later stage disease. These distinct patterns would render this marker useful to include in a kit for detecting early-stage melanoma. For example, if Col(I) expression was measured repeatedly in a patient over time, if expression began decreasing, this could indicate an increase in severity of or a relapse of disease, and would provide an early and non-invasive indicator that more aggressive treatment may be required. In another example, if Col(I) expression levels were measured in a population of melanoma patients, general disease stage categories could be made based on the expression levels, allowing for quicker comparisons between patients. It is also noted that Alexandrescu, van Kempen, and Zaenker all teach analysis of early-stage melanoma samples, and the ordinary artisan would be particularly interested in diagnosing such samples, as early diagnosis would produce the best patient outcomes.
GenBank teaches the sequence of the biomarker target for the claimed primer sequences (i.e. Col1a1) in humans, and methods of appropriate primer design using publicly available software would be available to the ordinary artisan, such as NCBI Primer-BLAST (Ye Abstract). In view of the above, the ordinary artisan would have been motivated to use the publicly available software disclosed in Ye, the teachings of van Kempen and the known Col1a1 sequence described by GenBank by to design primers for use in the method of Alexandrescu, in view of GenBank, in view of Zaenker, in view of Ye, and in view of Godfrey to detect melanoma biomarkers with real-time RT-PCR, and in the absence of unexpected results, the claimed polynucleotides (instant SEQ ID NOs: 8-9) simply represent the result of an obvious series of steps.
Additionally, including the recited primers in a kit for performing the method of Alexandrescu, in view of GenBank, in view of Zaenker, in view of Ye, in view of Godfrey, and further in view of van Kempen would have been obvious to the ordinary artisan at the time of the invention in view of the teachings of Godfrey. In particular, as noted above, Godfrey already teaches the use of a kit with primers, and renders such a kit obvious in the method of Alexandrescu, in view of GenBank, in view of Zaenker, in view of Ye, and in view of Godfrey. As adding the primers described by van Kempen to this method would be prima facie obvious, it would also be prima facie obvious to include the primers described by van Kempen in the kit for performing this method. The ordinary artisan would have been motivated to so provide kits containing the detection reagents to obtain the ability to commercialize the disclosed detection method. The ordinary artisan would have had a reasonable expectation of success in view of the guidance provided by Godfrey in paras. 112-115.
Furthermore, as noted above in the “Claim Interpretation” section, the use of a “control” primer pair amounts to the use of a primer pair generally. In the rejection of claim 16 above, a control primer pair as taught in Alexandrescu is used, where this primer pair falls in line with this interpretation. However, the primer pair of SEQ ID NOs: 8-9 to target Col1a1 expression also would fall in line with this interpretation, as this is a primer pair hybridizing to a particular sequence. This combination of references thus meets the limitations of claim 16 in addition to claims 39-40.
Thus, claims 16 and 39-40 are prima facie obvious over Alexandrescu, in view of GenBank, in view of Zaenker, and in view of Ye, in view of Godfrey, and further in view of van Kempen.
Claims 42-43 are rejected under 35 U.S.C. 103 as being unpatentable over Alexandrescu et al. (Journal of Investigative Dermatology, 2012), in view of GenBank (Accession #s EU446494 and M36546.1), in view of van Kempen et al. (Int J Cancer, 2008), in view of Zaenker et al. (Oncotarget, 2009), in view of Ye et al. (BMC Bioinformatics, 2012), and in view of Godfrey (US 2006/0068433 A1).
Alexandrescu teaches measuring the expression levels of biomarkers in melanoma (Abstract). The types of melanomas examined ranged from early stage to advanced (Table 1). A quantitative real-time RT-PCR was performed involving a control gene marker and specific primer and probes sets (pages 1888-1889, “Reverse transcription-PCR assay”). This RT-PCR utilized a reverse transcriptase (page 1892, column 1, para. 6). Expression levels were compared between melanoma and non-cancerous skin samples (Tables 3-4 and Figure 1). Samples were all taken from skin biopsies (page 1887, “Patient clinical and pathological characteristics”). The reference states that, “Cutaneous melanoma is the malignancy with the fastest growing incidence in the United States, with an annual percentage increase of 2.8% in the last two decades (SEER, 2008), and reaching epidemic proportions in some parts of the world. It is estimated that 62,000 persons will be diagnosed with melanoma in 2008 in the United States, among which 8,200 will die of this disease (SEER, 2008),” and notes that current diagnostic tools primarily rely on histological and immunohistological examinations (page 1887, column 1, para. 1 and column 2, para. 2). The authors conclude that, “…we have shown the potential utility of this RT-PCR assay using melanoma-specific markers that can reliably differentiate melanoma from non-melanoma lesions.” (page 1891, column 2, para. 5).
However, the primers taught by Alexandrescu do not comprise the sequences taught by the instant claims.
As noted in the Sequence Listing, SEQ ID NOs: 4-5 are associated vGAF and SEQ ID NOs: 8-9 are associated with Col1A1 (collagen type I alpha 1 chain).
GenBank Accession #EU446494 comprises SEQ ID NOs: 4-5 and is a construct for the ZBTB7B gene, (also known as vGAF). GenBank Accession #M36546.1 comprises SEQ ID NOs: 8-9 in association with alpha-1 collagen type 1.
van Kempen teaches Col1a1 mRNA expression in melanoma cells (Figure 1b-c and Figure 2). They conclude that, “In early stages of melanoma development and progression, Col(I) synthesis contributes to the development of a tumor vasculature, whereas its inhibition in advanced stages after a proper vasculature is formed, can contribute to further progression,” (page 1028, column 2, para. 1), indicating that levels of Col1a1 may be predictive of the presence and progression of melanoma. van Kempen also states that controls were used to compare expression levels (e.g. page 1020, column 2, paras. 5-6, page 1021, column 1, para. 1, and page 1023, column 1, para. 2).
Zaenker teaches the development of diagnostic tests for early stage melanoma (Abstract). This test utilizes antibodies for antigens that can differentiate between primary melanoma patients and healthy individuals (Abstract). The patients examined were specifically diagnosed with early stage melanoma, and these patients were compared to healthy controls (page 30540, column 2, para. 1). ZBTB7B (the gene encoding vGAF) was shown to be in a combination of sensitive and specific biomarkers for detecting melanoma (page 30544, columns 1-2 joining para.). Generally, the biomarker intensity in the melanoma patients was higher than that of the normal samples (Figure 3). Blood samples from patients were used and underwent protein profiling (page 30547).
Prior to the effective filing date of the claimed invention, it would have been prima facie obvious for one of ordinary skill in the art to use the method of Alexandrescu to detect the biomarkers suggested by van Kempen and Zaenker with primers created using the sequences taught by GenBank. Alexandrescu provides rationale for wanting to study melanoma biomarkers (as melanoma detection may prevent metastases and/or improve patient outcomes) and for using real-time RT-PCR with primers and probes to perform biomarker detection, as this method can analyze a large number and variety of melanoma tissues from biopsy samples, which are often taken when a patient presents with an unknown and/or suspicious lesion. As these biopsies are already part of routine care, analyzing them via RT-PCR allows for a relatively quick way to measure biomarker expression that can be compared with results of histological analyses. In particular, RT-PCR may reveal expression levels that can indicate very early stages of melanoma that would be difficult or impossible to detect in histological slides, which would allow for earlier disease interventions.
It would be obvious to the ordinary artisan that the method of Alexandrescu could be used with other biomarkers. van Kempen teaches that Col(I) expression differs between early and late stage melanoma (see Figure 7 for example), where expression is increasing during early stage disease and decrease in later stage disease. These distinct patterns would render this marker useful to include in a kit for detecting early stage melanoma. For example, if Col(I) expression was measured repeatedly in a patient over time, if expression began decreasing, this could indicate an increase in severity of or a relapse of disease, and would provide an early and non-invasive indicator that more aggressive treatment may be required. In another example, if Col(I) expression levels were measured in a population of melanoma patients, general disease stage categories could be made based on the expression levels, allowing for quicker comparisons between patients. Zaenker discusses measuring antibody markers through a protein analysis, but this analysis came from blood sample determinations, which occurred after an initial tumor diagnosis and excision (page 30547, columns 1-2 joining para.). Thus, as Alexandrescu teaches measuring biomarkers in tumor biopsies, and the methods of Zaenker uncovered additional melanoma biomarkers, it would be obvious to analyze the biomarkers uncovered by Zaenker earlier in the melanoma discovery and analysis process (i.e. in the initial biopsy sample), to measure all potential biomarkers at the same time, and in the same manner (i.e. utilizing RNA and RT-PCR). As the ZBTB7B gene is known, as evidenced by GenBank and Zaenker, such RNA targeting would be possible. It is also noted that Alexandrescu, van Kempen, and Zaenker all teach analysis of early stage melanoma samples, and the ordinary artisan would be particularly interested in diagnosing such samples, as early diagnosis would produce the best patient outcomes.
GenBank teaches the sequences of the claimed biomarkers (i.e. Col1a1 and ZBTB7B) in humans, and methods of appropriate primer design using publicly available software would be available to the ordinary artisan, such as NCBI Primer-BLAST (Ye Abstract). In view of the above, the ordinary artisan would have been motivated to use the publicly available software disclosed in Ye, the teachings of van Kempen and Zaenker and the known vGAF and Col1a1 sequences described by GenBank by to design primers for use in the method of Alexandrescu to detect melanoma biomarkers with real-time RT-PCR, and in the absence of unexpected results, the claimed polynucleotides (instant SEQ ID NOs: 4-5 and 8-9) simply represent the result of an obvious series of steps.
Additionally, including PCR reagents (including reverse transcriptase) and the recited primers in a kit for performing the method of Alexandrescu, in view of GenBank, in view of van Kempen, in view of Zaenker, and in view of Ye would have been obvious to the ordinary artisan at the time of the invention in view of the teachings of Godfrey. In particular, since Godfrey teaches that kits can be “particularly useful” for commercializing nucleic acid detection methods that make use of reagents that include oligonucleotides such as nucleic acid primers (para. 112), the ordinary artisan would have recognized that the detection methods disclosed by Alexandrescu, in view of GenBank, in view of van Kempen, in view of Zaenker, and in view of Ye could also be commercialized by providing the reagents used to practice the methods (e.g., primers, probes, dNTPs, etc.) in a packaged kit. Accordingly, the ordinary artisan would have been motivated to so provide kits containing the detection reagents to obtain the ability to commercialize the disclosed detection method. The ordinary artisan would have had a reasonable expectation of success in view of the guidance provided by Godfrey in paras. 112-115.
Furthermore, Alexandrescu, van Kempen, and Zaenker teach the use of controls with their methods, and Alexandrescu in particular teaches using control, benign samples from healthy individuals when comparing expression levels of target biomarkers, as well as using a control gene expression marker (page 1889, column 1, para 2) and Table 3. Godfrey teaches that control nucleic acids can be included in created kits (para. 112) and specifically teaches that these can be control mRNAs (claim 20). Thus, it would be prima facie obvious to the ordinary artisan that controls from healthy (i.e. non-melanoma) samples could be included in the kit, where the samples would contain the mRNAs for vGAF and/or Col1a1. Providing these controls in the kit would eliminate the need for the user of the kit to solicit donations from volunteers for control samples, thus simplifying the overall protocol and enhancing the ease of use of the kit. As Godfrey already teaches that nucleic acids can be included in the kit, and teaches that kits can also comprise reagents for extracting mRNAs from samples (para. 115) there would be a reasonable expectation of success.
Therefore, claims 42-43 are prima facie obvious over Alexandrescu, in view of GenBank, in view of van Kempen, in view of Zaenker, and in view of Ye, and in view of Godfrey.
Conclusion
No claims are currently allowable.
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 FRANCESCA F GIAMMONA whose telephone number is (571)270-0595. The examiner can normally be reached M-Th, 7-5pm.
Examiner interviews are available via telephone, in-person, and video conferencing using a USPTO supplied web-based collaboration tool. To schedule an interview, applicant is encouraged to use the USPTO Automated Interview Request (AIR) at http://www.uspto.gov/interviewpractice.
If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Gary Benzion can be reached at (571) 272-0782. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
Information regarding the status of published or unpublished applications may be obtained from Patent Center. Unpublished application information in Patent Center is available to registered users. To file and manage patent submissions in Patent Center, visit: https://patentcenter.uspto.gov. Visit https://www.uspto.gov/patents/apply/patent-center for more information about Patent Center and https://www.uspto.gov/patents/docx for information about filing in DOCX format. For additional questions, contact the Electronic Business Center (EBC) at 866-217-9197 (toll-free). If you would like assistance from a USPTO Customer Service Representative, call 800-786-9199 (IN USA OR CANADA) or 571-272-1000.
/F.F.G./Examiner, Art Unit 1681
/GARY BENZION/Supervisory Patent Examiner, Art Unit 1681