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 .
Application Status and Election
Applicant’s amendments filed September 2, 2025, amending claim 1 and canceling claim 9 is acknowledged. It is noted that claim 1 is listed as “(Original)”; however, claim 1 is clearly amended and Applicant’s Remarks (page 9), states “the response amends claim 1 by incorporate the features from as-filed claim 9”. Additionally, claim 6 still has the mark-up “[[or 5]]” present in the previous claim set, which should have been removed from the current claim set.
Claims 1-8 and 10-24 are pending. Claims 17-24 remain withdrawn from further consideration pursuant to 37 CFR 1.142(b), as being drawn to nonelected groups, there being no allowable generic or linking claim.
Claims 1-8 and 10-16 are under examination.
Nucleotide and/or Amino Acid Sequence Disclosures
REQUIREMENTS FOR PATENT APPLICATIONS CONTAINING NUCLEOTIDE AND/OR AMINO ACID SEQUENCE DISCLOSURES
Items 1) and 2) provide general guidance related to requirements for sequence disclosures.
37 CFR 1.821(c) requires that patent applications which contain disclosures of nucleotide and/or amino acid sequences that fall within the definitions of 37 CFR 1.821(a) must contain a "Sequence Listing," as a separate part of the disclosure, which presents the nucleotide and/or amino acid sequences and associated information using the symbols and format in accordance with the requirements of 37 CFR 1.821 - 1.825. This "Sequence Listing" part of the disclosure may be submitted:
In accordance with 37 CFR 1.821(c)(1) via the USPTO patent electronic filing system (see Section I.1 of the Legal Framework for Patent Electronic System (https://www.uspto.gov/PatentLegalFramework), hereinafter "Legal Framework") as an ASCII text file, together with an incorporation-by-reference of the material in the ASCII text file in a separate paragraph of the specification as required by 37 CFR 1.823(b)(1) identifying:
the name of the ASCII text file;
ii) the date of creation; and
iii) the size of the ASCII text file in bytes;
In accordance with 37 CFR 1.821(c)(1) on read-only optical disc(s) as permitted by 37 CFR 1.52(e)(1)(ii), labeled according to 37 CFR 1.52(e)(5), with an incorporation-by-reference of the material in the ASCII text file according to 37 CFR 1.52(e)(8) and 37 CFR 1.823(b)(1) in a separate paragraph of the specification identifying:
the name of the ASCII text file;
the date of creation; and
the size of the ASCII text file in bytes;
In accordance with 37 CFR 1.821(c)(2) via the USPTO patent electronic filing system as a PDF file (not recommended); or
In accordance with 37 CFR 1.821(c)(3) on physical sheets of paper (not recommended).
When a “Sequence Listing” has been submitted as a PDF file as in 1(c) above (37 CFR 1.821(c)(2)) or on physical sheets of paper as in 1(d) above (37 CFR 1.821(c)(3)), 37 CFR 1.821(e)(1) requires a computer readable form (CRF) of the “Sequence Listing” in accordance with the requirements of 37 CFR 1.824.
If the "Sequence Listing" required by 37 CFR 1.821(c) is filed via the USPTO patent electronic filing system as a PDF, then 37 CFR 1.821(e)(1)(ii) or 1.821(e)(2)(ii) requires submission of a statement that the "Sequence Listing" content of the PDF copy and the CRF copy (the ASCII text file copy) are identical.
If the "Sequence Listing" required by 37 CFR 1.821(c) is filed on paper or read-only optical disc, then 37 CFR 1.821(e)(1)(ii) or 1.821(e)(2)(ii) requires submission of a statement that the "Sequence Listing" content of the paper or read-only optical disc copy and the CRF are identical.
Specific deficiencies and the required response to this Office Action are as follows:
Specific deficiency – Nucleotide sequences appearing in the drawings of Fig 1 and 2 and in the Amended Specification filed September 2, 2025 on pages 12-13 are not identified by sequence identifiers in accordance with 37 CFR 1.821(d). Sequence identifiers for nucleotide and/or amino acid sequences must appear either in the drawings or in the Brief Description of the Drawings. Specifically, the nucleotide sequences in rows 2-6 of the tables in Fig 1 and Fig 2, and on pages 12 and 13 of the Specification are not identified with SEQ ID NOs. According the sequence listing filed December 10, 2024, the sequences in rows 2-6 of Table 1 appear to be SEQ ID NOs 4-8, respectively. The sequences in rows 2-6 of Table 2 appear to be SEQ ID NOs 9-13, respectively.
Required response – Applicant must provide:
Replacement and annotated drawings in accordance with 37 CFR 1.121(d) inserting the required sequence identifiers;
AND/OR
A substitute specification in compliance with 37 CFR 1.52, 1.121(b)(3) and 1.125 inserting the required sequence identifiers into the Brief Description of the Drawings, consisting of:
A copy of the previously-submitted specification, with deletions shown with strikethrough or brackets and insertions shown with underlining (marked-up version);
A copy of the amended specification without markings (clean version); and
A statement that the substitute specification contains no new matter.
Response to Argument – Sequence Compliance
Applicant argues that the Application is now in compliance because the Specification was amended to incorporate FIGs 1 and 2 into the text of the Specification (Remarks, ¶ spanning pages 9-10). This argument has been fully considered but is not persuasive because there are still nucleotide sequences that are longer than 10 nucleotides that are not identified with a SEQ ID NO. Specifically, the sequences in rows 2-6 of the tables that are now in both the Drawings and the Specification. To remedy the noncompliance, Applicant should include “SEQ ID NO 4” in the table next to the sequence gCCATgAACgCCAAggT, and include the appropriate SEQ ID NO for each of the other sequences in the tables. Because the tables are now present in both the Drawings and the Specification (pages 12-13), the SEQ ID NOs must be entered twice, once for each appearance of the sequences.
Specification
The use of the terms LuxTM, TaqMan®, and Direct-zol®, which are trade names or marks used in commerce, have been noted in this application. The terms should be accompanied by the generic terminology; furthermore the terms should be capitalized wherever appearing or, where appropriate, include a proper symbol indicating use in commerce such as ™, SM , or ® following the terms.
Although the use of trade names and marks used in commerce (i.e., trademarks, service marks, certification marks, and collective marks) are permissible in patent applications, the proprietary nature of the marks should be respected and every effort made to prevent their use in any manner which might adversely affect their validity as commercial marks.
Response to Arguments - Specification
Applicant argues that Scorpion and Lux as used in the present application are not trademarks or trade names, but instead refer to well-recognized classes of PCR primers that are broadly used in the field of molecular biology (Remarks, page 10). This argument has been fully considered but is not persuasive as it applies to Lux, TaqMan, and Direct-zol. First, Applicant provides no argument for using the terms TaqMan and Direct-zol, which are also trademarks, but have not been designated so in the amended Specification. Second, the fact that a term is well-known and frequently used in the art does not negate the fact that it is a tradename or trademark. MPEP 608.01(v) states that trademarks or trade names may in fact be well known in the art; however, the MPEP still recognizes them as trademarks or trade names, and they should be treated as such with the appropriate designation and, if appropriate, an accompanying definition which is sufficiently descriptive. “Lux” is still used as a trademarked term, for instance at this chemical company website that originally trademarked the term: https://www.thermofisher.com/order/catalog/product/11743500?SID=srch-srp-11743500. As such “Lux” should still be marked the appropriate symbol and an accompanying definition like “fluorogenic primers”.
Claim Rejections - 35 USC § 112(b)
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.
Claim 15 is rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention. This is a maintained rejection.
Claim 15 contains the trademark/trade name Lux. Where a trademark or trade name is used in a claim as a limitation to identify or describe a particular material or product, the claim does not comply with the requirements of 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph. See Ex parte Simpson, 218 USPQ 1020 (Bd. App. 1982). The claim scope is uncertain since the trademark or trade name cannot be used properly to identify any particular material or product. A trademark or trade name is used to identify a source of goods, and not the goods themselves. Thus, a trademark or trade name does not identify or describe the goods associated with the trademark or trade name. In the present case, the trademark/trade name is used to identify/describe specific primers and, accordingly, the identification/description is indefinite.
Response to Arguments - §112(b)
Applicant argues that Scorpion and Lux as used in the present application are not trademarks or trade names, but instead refer to well-recognized classes of PCR primers that are broadly used in the field of molecular biology. Applicant argues that “Multiple manufacturers produce primers under this design principle, and the term ‘Scorpion’ has come to represent the primer class itself” and “the designation ‘LUX’ is widely understood in the art as a descriptive abbreviation of the underlying principle, and primers of this type are produced by multiple suppliers” (Remarks, page 12). This argument has been fully considered. As it applies to “Scorpion”, the argument is persuasive because 1) the trademarked name of the Scorpion primer is “Scorpions” (plural) and the art appears to use “Scorpion” in the singular form as an adjective to describe a type of primer (Thelwell et al., Nucleic Acids Research (2000), 28: 3752-3761). However, the argument is not persuasive as it is applied to “Lux”. The fact that a term is well-known and frequently used in the art does not negate the fact that it is a tradename or trademark. MPEP 2173.05(u) states “If the trademark or trade name is used in a claim as a limitation to identify or describe a particular material or product, the claim does not comply with the requirements of the 35 U.S.C. 112(b) or pre-AIA 35 U.S.C. 112, second paragraph.” In this case “Lux” is used to refer to a fluorogenic primer – which is a particular product. The term “Lux” is still used to refer to fluorogenic primers that a particular company produces. For instance, ThermoFisher currently uses “LuxTM” on their webpage for a qRT-PCR kit when referring to fluorogenic primers: https://www.thermofisher.com/order/catalog/product/11743500?SID=srch-srp-11743500 ([retrieved September 25, 2025]).
To remedy the indefiniteness, it is recommended to remove “Lux” from the claim or to claim the Lux primers in terms of structure.
Claim Rejections - 35 USC § 101
35 U.S.C. 101 reads as follows:
Whoever invents or discovers any new and useful process, machine, manufacture, or composition of matter, or any new and useful improvement thereof, may obtain a patent therefor, subject to the conditions and requirements of this title.
Claims 1-8 and 10-16 are rejected under 35 U.S.C. 101 because the claimed invention is directed to a judicial exception (i.e., a law of nature, a natural phenomenon, or an abstract idea) without significantly more. The claims are drawn to methods of detection/diagnosing endometriosis and/or uterine adenomyosis by detecting expression of ESR2, CXCL12 and/or CXCR4. However, the claims do not include elements, when considered separately and in combination, that are sufficient to amount to significantly more than the judicial exceptions as outlined below. This is a maintained rejection. It is updated to reflect the incorporation of limitations for canceled claim 9 into claim 1.
Subject Matter Eligibility Test for Products and Processes – Claim 1
Step 1 - Is the Claim to a Process, Machine, Manufacture or Composition of Matter? YES
Claim 1 is directed to a method of disease detection. Thus, the claims are directed to a statutory category (e.g., a method).
Step 2A, Prong One - Does the Claim Recite an Abstract Idea, Law of Nature, or Natural Phenomenon? YES
Judicial exceptions have been identified by the courts by way of example, including natural correlations. Claim 1 recites one judicial exception: the expression of naturally occurring genes ESR2, CXCL12 and CXCR4 and their correlation to disease occurrence. MPEP 2106.04(b) states that one type of judicial exception is correlations between the presence of a natural product, such as the protein in a bodily fluid, and a disease risk. This is highly similar to claim 1 which is directed to the expression of genes, which can be measured as RNA or protein, in menstrual fluid and the risk of having endometriosis and/or uterine adenomyosis.
Step 2A, Prong Two - Does the Claim Recite an Additional Elements that Integrate the Judicial Exception into a Practical Application? NO
The Supreme Court has long distinguished between principles themselves, which are not patent eligible, and the integration of those principles into practical applications, which are patent eligible. The phrase "integration into a practical application" requires an additional element or a combination of additional elements in the claim to apply, rely on, or use the judicial exception in a manner that imposes a meaningful limit on the judicial exception, such that it is more than a drafting effort designed to monopolize the exception. MPEP 2106.04(d)(2) discusses the requirements for a natural phenomenon such as a correlation between gene expression and diagnosis to be considered to be integrated into a practical application. “One way to demonstrate such integration is when the additional elements apply or use the recited judicial exception to effect a particular treatment or prophylaxis for a disease or medical condition.” In this case, claim 1 does not recite any treatment or prophylactic step to be performed that may integrate the recited natural correlation into a practical application.
Step 2B - Does the Claim Recite Additional Elements that Amount to Significantly More than the Judicial Exception? NO
The Supreme Court has identified a number of considerations for determining whether a claim with additional elements amounts to "significantly more" than the judicial exception(s) itself. The claim as a whole is evaluated as to whether it amounts to significantly more than the recited exception, i.e., whether any additional element, or combination of additional elements, adds an inventive concept to the claim (MPEP 2106.05). However, the additional elements, individually and in combination, do not amount to "significantly more". Claim 1 recites the additional steps of 1) “providing menstrual blood to the patient to be tested” and 2) comparing the gene expression with a constitutively expressed housekeeping gene to normalize expression. However, these additional steps do amount to significantly more. First, it was already well known that gene expression could be detected from samples of menstrual blood. For instance, Villarreal teaches that biomarkers (i.e., gene expression measured as RNA or protein) can be detected in menstrual fluid (US 20170363641, [0006]; of record). Second, it is well known that in biomarker detection, the protein and mRNA expression in each sample is compared to a gene/protein that is not predicted to be different between samples, such as constitutively expressed housekeeping gene (See e.g., Ruiz et al., Reproductive Sciences (2010), 17(10): 894-903, Methods; of record). Therefore, the claim does not amount to something significantly more than the judicial exception.
Subject Matter Eligibility Test for Products and Processes – Dependent Claims
Claim 2 recites the menstrual blood comes from a tampon. As this limitation is directed to the sample collection step, the judicial exception is still not integrated into a practical application. Additionally, Villarreal teaches that menstrual fluids can be collected from a tampon ([0065]). Thus, this additional limitation does not amount to something significantly more than the judicial exception.
Claim 3 recites the menstrual blood is frozen below -25o C after removal of the tampon from the body. As this limitation is directed to a sample preparation step, the judicial exception is still not integrated into a practical application. Additionally, Yang teaches that menstrual fluids can be collected from a tampon and frozen at -80o C (Yang et al., Molecular & Cellular Proteomics (2012), 11(10): 1024-1035; page 1025, ¶6; of record). Thus, this additional limitation does not amount to something significantly more than the judicial exception.
Claims 4-8 and 12 recite limitations that limit the genus of patients or controls, including by age, previous pregnancies, previous hormone therapy, and ones that suffer from dysmenorrhea that can be classified as secondary or tertiary dysmenorrhea and ICD 94-4 or 94-5 (medical coding number). Thus, only the judicial exception has been limited – the correlation between gene expression and a disease in a more specific patient cohort. As such the judicial exception is still not integrated into a practical application and there are no additional limitations to amount to something significantly more than the judicial exception.
Claims 10-11 recite limitations regarding the biomarker – simultaneous increase in expression of 2 or more the genes, or an increase in expression at least 1.5-fold. This limits the judicial exception to a more specific natural correlation. However, this more limited natural correlation is still a judicial exception. The judicial exception is not integrated into a practical application and no additional limitations are recited that could amount to something more than the judicial exception.
Claims 13-16 recites the expression is measured at the protein level by Western blotting or ELISA, or at the mRNA level by Northern, RNA microarray or by PCR. As these limitations are directed to a sample analysis steps, the judicial exception is still not integrated into a practical application. Additionally, these limitations are considered well understood, routine and conventional. Villarreal teaches that protein biomarkers can be detected by Western or ELISA (e.g., [0070]) and gene expression biomarkers can be detected by fluorogenic-based RT-PCR and microarray (e.g., [0077]).
Response to Arguments - §101
Applicant argues that the claims do not recite or describe any judicial exceptions because although the claims recite nature-based products like menstrual blood samples, the claims are not directed to the nature-based products (page 13, ¶4-5). This argument has been fully considered but is not persuasive. Examiner agrees that the claims are not directed to the nature-based products; however, that is/was not the identified judicial exception in the §101 rejection. The identified judicial exception recited in the claim is “an increased expression of one or more genes [ESR2, CXCL12, CXCR4] indicates endometriosis and/or uterine adenomyosis”, which is a natural correlation/phenomenon, and not a natural product.
Applicant argues that the claim is directed to an in vitro diagnostic method that requires specific technical steps. Applicant argues that unlike in Myriad, claim 1 recites a diagnostic process and requires laboratory steps that do not occur in nature (Remarks, page 14, ¶1). These arguments have been fully considered but are not persuasive.
First, the claim is directed to the identified judicial exception “an increased expression of one or more genes [ESR2, CXCL12, CXCR4] indicates endometriosis and/or uterine adenomyosis” because it is entirely a diagnostic method. The purpose of the method is to discover if an individual is at risk for endometriosis based on the level of the ESR2, CXCL12 and/or CXCR4. MPEP 2106.04(b) states that one type of judicial exception is correlations between the presence of a natural product, such as the protein in a bodily fluid, and a disease risk. This is precisely what the claim recites and is directed to.
Second, Myriad is not an analogous case/decision that should be used when analyzing the diagnostic claims for subject matter eligibility. Instead, the Vanda decision should be used: Vanda Pharmaceuticals Inc. v. West-Ward Pharmaceuticals, 887 F.3d 1117, 1135-36, 126 USPQ2d 1266, 1281 (Fed. Cir. 2018). MPEP 2106.04(b) states “The courts have identified the following concepts and products as examples of laws of nature or natural phenomena… (xi) the natural relationship between a patient’s CYP2D6 metabolizer genotype and the risk that the patient will suffer QTc prolongation after administration of a medication called iloperidone, Vanda Pharmaceuticals Inc. v. West-Ward Pharmaceuticals”. Again, as stated in the previous paragraph, the claims are directed to the natural relationship between a genetic phenotype – the presence of at least one of three gene products in menstrual blood – and the risk the subject has endometriosis. Thus, the claims recite and are directed to a judicial expression. MPEP 2106.04(d)(2) then continues to discuss Vanda under the Step 2A, prong 2 analysis. “Vanda’s claims used the recited law of nature to more safely treat the patients with the drug, thereby reducing the patient’s risk of QTc prolongation. 887 F.3d at 1135, 126 USPQ2d at 1280.” Thus, the judicial exception of a natural correlation in Vanda was directed to a practical application, namely reducing the risk of a drug based on the genotype. MPEP 2106.04(d)(2) provides examples of practical applications including “limitations that treat or prevent a disease or medical condition, including, e.g., acupuncture, administration of medication, dialysis, organ transplants, phototherapy, physiotherapy, radiation therapy, surgery, and the like”. However, in the case of the instant claims, there are no limitations as to how the natural correlation – once determined by the technical/laboratory steps – is to be applied. Therefore, the natural correlation is not integrated into a practical application and the subject matter eligibility analysis must proceed to Step 2B.
Applicant argues that the normalization process now required for claim 1 integrates the discovery of a correlation into a structured diagnostic process which constitutes “significantly more” than the judicial exception itself (Remarks, page 14, ¶1). This argument has been fully considered but is not persuasive. First, a normalization process is merely calculating a ratio between the expression of two genes, which is itself could be interpreted as part of the natural correlation. Second, MPEP 2106.05 discusses the analysis for Step 2B and how to evaluate additional elements in the claim to determine if a claim is directed to “significantly more” than the judicial exception. Limitations that have been found not to amount to “significantly more” include “ii) simply appending well-understood, routine, conventional activities previously known to the industry, specified at a high level of generality, to the judicial exception” and “iii) adding insignificant extra-solution activity to the judicial exception, e.g., mere data gathering in conjunction with a law of nature or abstract idea such as a step of obtaining information about credit card transactions so that the information can be analyzed by an abstract mental process.” In the instant case, the normalization process could be considered as either falling under (ii) or (iii). As indicated in the current and previous rejections, it is well known and conventional in the art that gene expression levels in each sample should be compared to a gene that is not predicted to be different between samples, such as a constitutively expressed housekeeping gene, which is the process of normalization. Second, gathering data about the housekeeping gene so that the expression of the biomarker can be analyzed and compared across samples could be considered “mere data gathering in conjunction with the law of nature” because without the normalization process, the biomarker may not be informative due to variation between samples of the same groups. In either event the normalization process would not mount to significantly more than the judicial exception of the natural correlation between ESR2, CXCL12 and/or CXCR4 expression and the risk of having endometriosis.
Claim Rejections - 35 USC § 103
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
Claims 1-2, 4-8 and 10-16 are rejected under 35 U.S.C. 103 as being unpatentable over Villarreal (US 20170363641 A1, published December 21, 1017; of record) in view of Ruiz (Ruiz et al., Reproductive Sciences (2010), 17(10): 894-903; of record) and Evans (Evans et al., Nature Reviews Endocrinology (2016), 12(11): 654-667; of record). Claims 5 and 6 are evidenced by ACOG (Dysmenorrhea and endometriosis in the adolescent. ACOG Committee Opinion No. 760. American College of Obstetricians and Gynecologists. Obstet Gynecol (2018), 132:e249–58; of record) and ICD10 (N94.5, Secondary dysmenorrhea, https://www.icd10data.com/ICD10CM/Codes/N00-N99/N80-N98/N94-/N94.5, [retrieved April 29, 2025]; of record). This is a maintained rejection. It is updated to reflect the incorporation of limitations for canceled claim 9 into claim 1.
Regarding claims 1-2, Villarreal teaches endometriosis is a gynecological disease that is defined as the presence of endometrial tissue outside the uterine cavity, such as the peritoneum, ovary or fallopian tubes ([0049]). Villarreal teaches evaluating endometriosis (i.e., detection) by measuring one or more of… CXCR4… using the samples and method of the present invention ([0049]). Villarreal teaches biomarker expression can be detected by (a) obtaining a sample of the female subject’s menstrual fluid, and (b) measuring the presence, absence or level of a disease-related biomarker ([0006]). Villarreal teaches the menstrual fluid can be collected using a tampon ([0065]) and on day 2 of the menstrual period (i.e., day 2 of the cycle) ([0066]). Villarreal teaches the biomarkers can be detected using RT-PCR and quantitative PCR ([0076]-[0077]).
Villarreal does not teach in a single embodiment measuring CXCR4 in menstrual fluid from a tampon or on day 1-7 to diagnose endometriosis. Villarreal does not teach normalization to expression of a housekeeping gene.
Ruiz teaches CXCR4 has increased gene expression of CXCR4 in the rat model of endometriosis (Abstract). Ruiz teaches is CXCR4 is localized in the uterine endometria (Fig 1A-B) and has increased expression in patients with endometriosis than in controls (Fig 1D). Ruiz teaches higher CXCR4 expression was observed in the epithelium from endometriotic tissues (page 897, ¶5). Ruiz teaches detection of CXCR4 expression by quantitative PCR (qPCR) from the normal and endometrial cells lines (Fig 2). Ruiz teaches in qPCR, total RNA was isolated and the expression of CXCR4 and CXCL12 was normalized (i.e., compared) to the expression of GAPDH (i.e., a constitutively expressed housekeeping gene) (page 896, ¶3).
Evans teaches that during the mid-to-late secretory phase of each menstrual cycle, endometrial stromal cells terminally differentiate into epithelial, secretory cells (Fig 1; page 2, ¶7). Evans teaches that in a menstrual cycle that does not result in conception, the terminally differentiated cells are shed during menses (page 2, ¶7). Evans teaches that cytokines produced by the endometrium are secreted into the uterine cavity (page 6, ¶5).
If would have been obvious to one skilled in the art before the effective filing date of the claimed invention to have used Villarreal’s generic method of detecting biomarkers in menstrual fluid with Ruiz’s qPCR method to detect mRNA expression of CXCR4 by measuring CXCR4 mRNA levels normalized to GAPDH mRNA levels in the menstrual blood/fluid. It would have amounted to detecting a known biomarker for endometriosis in a bodily fluid that is known be useful for biomarker detection. The skilled artisan would have expected that CXCR4 expression could be detected in menstrual fluid as taught in Villarreal because Ruiz teaches it is located in endometrial epithelia, which Evans teaches is shed each cycle. The skilled artisan would have been motivated to detect CXCR4 as a biomarker using qPCR, which involves normalization to a housekeeping gene, for endometriosis from menstrual fluid because Villarreal suggests it and qPCR is a well-known way to measure differences in gene expression between disease and healthy tissue
Regarding claims 4-6, Villarreal teaches endometriosis occurs in 5-20% of females with pelvic pain (i.e., dysmenorrhoea) ([0049]). Villarreal is silent to as to the ICD code (claim 5) or classification of the pelvic pain (claim 6). However, ICD10 teaches that ICD94-5 is the code for Secondary dysmenorrhea. ACOG teaches secondary dysmenorrhea refers to painful menses due to pelvic pathology or recognized medical conditions and whose most common cause endometriosis.
Regarding claim 7 and 12, Villarreal teaches that endometriosis occurs in 20-50% of infertile females (i.e., nulligravida) and in 6-10% of females of reproductive age (i.e., the patient is 18-35 years of age) ([0049]).
It would have been obvious to one skilled in the art for the only difference between the test patient and the control subject was for the control subject to have no detectable endometriosis and no history of pelvic pain because Villarreal teaches pelvic pain is a main symptom of endometriosis. The skilled artisan would have been motivated to have the subject to be the same age range and also nulligravida to reduce the number of variables between the text subject and control, according to standard scientific study design principles.
Regarding claim 8, Evans teaches that the endometrium undergoes substantial remodeling under the influence of ovarian steroid hormones (i.e., progesterone and estrogen) (page 2, ¶4). Evans teaches that progesterone induces decidualization, which causes the stromal cells to produce higher levels of cytokines (page 3, ¶2; Fig 1). Ruiz teaches that treatment with estradiol, progesterone or both hormones decreases CXCR4 and CXCL12 expression in endometrial cells lines (Fig 3).
It would have been obvious to one skilled in the art before the effective filing date of the claimed invention to have excluded patients that have undergone hormone therapy within 3 months. Evans teaches that development and shedding of the endometrium is controlled by hormones as well as the production of cytokines. The skilled artisan would have recognized that the addition of hormones could affect the expression of CXCR4 such that it would be unreliable biomarker for endometriosis in patients undergoing hormone therapy.
Regarding claim 10, Ruiz also teaches that CXCL12 has high expression in stromal endometrial cells lines (Fig 2). Ruiz also teaches that CXCL12 has been previously shown to be expressed at higher levels in epithelium of ovarian endometriosis tissue when compared to normal ovary (page 897, ¶5).
It would have been obvious to one skilled in the art before the effective filing date of the claimed invention to have also assayed CXCL12 as a simultaneous biomarker for endometriosis in Villarreal’s method. It would have amounted to assaying a second gene that is also known to be upregulated in endometriosis. The skilled artisan would have predicted that a simultaneous increase in CXCR4 and CXCL12 would be predictive of endometriosis because Ruiz teaches that both genes are upregulated in endometriosis. The skilled artisan would have been motivated to do so for providing a second correlative biomarker, which could reduce the possibility of false positives compared to measuring a single biomarker alone.
Regarding claim 11, Ruiz teaches comparing gene expression of endometrial tissue and cell lines to normal control tissue and cell lines (Figures 2-3). Ruiz teaches reporting the expression data as an average of data from three experiments (Figures 2-3). Ruiz teaches an increase of ~ 1.5-fold in expression of CXCR4 in endometrial epithelia compared to normal tissue (Fig 1D).
It would have been obvious to one skilled in the art before the effective filing date of the claimed invention to have used a cut-off of 1.5-fold increase to make a diagnosis of endometriosis in Villarreal’s method. It would have amounted to using a known difference in CXCR4 gene expression between endometrial and normal tissues. The skilled artisan would have predicted that a 1.5-fold cut-off could be used as the diagnostic cut-off and been motivated to do so because Ruiz teaches that is the difference in CXCR4 expression between endometrial and normal tissues.
Regarding claims 13-15, Villarreal teaches the biomarkers can be detected using RT-PCR and quantitative PCR ([0076]-[0077]). Ruiz teaches detection of CXCR4 expression by quantitative PCR (qPCR) from the normal and endometrial cells lines (Fig 2). Ruiz teaches in qPCR, total RNA was isolated (i.e., expression is determined on the basis of mRNA determination) and the expression of CXCR4 and CXCL12 was normalized (i.e., compared) to the expression of GAPDH (i.e., a constitutively expressed housekeeping gene) (page 896, ¶3). Ruiz also teaches in qPCR, mRNA is reverse transcribed into cDNA and TaqMan Gene Expression Assay Mix (i.e., FRET probes) was used in the qPCR reaction (page 896, ¶3).
As indicated above for claim 1, it would have been obvious to one skilled in the art before the effective filing date of the claimed invention to have used Ruiz’s qPCR method to detect mRNA expression of CXCR4 in the obvious diagnostic method of detecting CXCR4 expression in menstrual fluid. It would have amounted to applying a well-known method of measuring gene expression to Villarreal’s method of detecting biomarkers in menstrual fluid. The skilled artisan would have predicted that Ruiz’s qPCR method could be used and been motivated to do so because 1) qPCR is a known proven way to measure differences in gene expression between disease and healthy tissue and 2) Villarreal suggests it.
Regarding claims 13 and 16, Villarreal teaches biomarkers can be detected as protein using western blotting and ELISA (i.e., an immunoassay) ([0071]). Ruiz teaches detecting CXCR4 protein from endometrial cells lines using a western blot (Figure 4).
It would have been obvious to one skilled in the art before the effective filing date of the claimed invention to have used Ruiz’s western blot method to detect CXCR4 protein levels in the obvious diagnostic method of detecting CXCR4 expression in menstrual fluid. It would have amounted to applying a well-known method of measuring gene expression to Villarreal’s method of detecting biomarkers in menstrual fluid. The skilled artisan would have predicted that Ruiz’s western blot method could be used and been motivated to do so because 1) western blot is a known and proven way to measure differences in gene expression between disease and healthy tissue, and 2) Villarreal suggests it.
Claim 3 is rejected under 35 U.S.C. 103 as being unpatentable over Villarreal (US 20170363641 A1, published December 21, 1017; of record), Ruiz (Ruiz et al., Reproductive Sciences (2010), 17(10): 894-903; of record) and Evans (Evans et al., Nature Reviews Endocrinology (2016), 12(11): 654-667; of record), as applied to claims 1-2, 4-8 and 10-16 above, and further in view of Yang (Yang et al., Molecular & Cellular Proteomics (2012), 11(10): 1024-1035; of record). This is a maintained rejection. It was updated to reflect the incorporation of limitations for canceled claim 9 into claim 1.
The teachings of Villarreal, Ruiz and Evans are recited above and applied as for claims 1-2, 4-8 and 10-16.
Villarreal, Ruiz and Evans do not teach how to handle, preserve and/or store menstrual blood/fluid samples.
Yang teaches menstruation is the expulsion of the endometrial lining of the uterus, which is increasingly being recognized as a critical factor in female fertility (Abstract). Yang teaches that the proteomes and transcriptomes from endometrial biopsies have been studied for both diagnostic and therapeutic purposes (Abstract). Yang teaches that menstrual fluids can be collected from a tampon and frozen at -80o C for downstream proteomic analysis (page 1025, ¶6)
It would have been obvious to one skilled in the art before the effective filing date of the claimed invention to have stored the menstrual blood/fluid at -80o C immediately upon collection of the tampon in Villarreal’s method. It would have amounted to using known methods of sample handling and storage in the obvious method of detecting CXCR4 in menstrual fluid as a biomarker for endometriosis. The skilled artisan would have predicted that the menstrual blood could be frozen and stored at -80o C for preservation for downstream analysis, and been motivated to do so, because Yang teaches the step can preserve proteomic features of menstrual fluid.
Response to Arguments - §103
Applicant recites the text of amended claim 1 and explains the invention in terms of a solution to the disclosed technical problem of using invasive procedures of conventional diagnostic methods. Applicant cites the solution from the Specification ([0021]) as “the eco-friendly colloid lithography process… may be easily implemented to reproduce micropillars with high precision” and the benefits of the invention (pages 15-17). Examiner thanks Applicant for the explanation; however, the above quote is not recited anywhere in the Specification. It is not clear where Applicant originated the quote. Nevertheless, the cited solution “colloid lithography” is not recited in the claims. The claims encompass any means to determine gene expression from menstrual fluid samples including batch analysis.
Applicant reviews the teachings from each of the prior art references and concludes 1) Villarreal does not provide specific gene markers and comparative analysis, 2) Ruiz requires invasive procedures and does not address the non-invasive menstrual blood-based test, 3) Evans does not disclose CXCL12 or CXCR4 for endometriosis, and 4) AOCG is not concerned with molecular diagnostics. Applicant argues that the cited reference taken along or in combination fail to disclose or suggest the claimed invention (pages 19 to 20). This argument has been fully considered but is not persuasive. First, 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). Second, Applicant’s conclusion that the technical combination was not disclosed or suggested in the prior art is merely arguments of counsel, which cannot take the place of factually supported objective evidence. MPEP 716.01(c). As indicated in the rejection above, CXCR4 and CXCR12 were known biomarkers for endometriosis and the means to detect biomarkers in general from menstrual fluid was previously developed. Additionally, Villarreal even suggests detecting biomarkers for endometriosis from menstrual fluid, including from a tampon, which requires normalization to a housekeeping gene, as taught in both Villarreal and Ruiz.
Applicant argues that the cited prior art references fail to specifically teach “determining expression of the gene ESR2 and/or CXCL12 and/or CXCR4 in comparison to a control sample… to normalize expression” (page 21-22). This argument has been fully considered, but it not persuasive. As indicated in the §103 rejections in this and the previous office action, Ruiz teaches during qRT-PCR assays, the expression of CXCR4 and CXCL12 was normalized to the expression of GAPDH (i.e., a constitutively expressed housekeeping gene) (page 896, ¶3). Thus, the prior art teaches normalizing expression of CXCR4 and CXCL12 to a constitutively expressed housekeeping gene.
Applicant again argues that the prior art references do not include all of the claim limitations and the rejections do not explain why the elements missing from the proposed combination would have been obvious (Remarks, ¶ spanning pages 22-23). This argument has been fully considered but is not persuasive because, as indicated above, the prior art references do teach every limitation recited in the claimed invention.
Applicant argues that MPEP 2143.03 requires “consideration” of every claim feature in an obviousness determination, but that the office must do more than merely “consider” each and every feature for the claim. Applicant 1) argues that the asserted combination must also teach or suggest each and every claim feature and 2) cites In re Royka 490 F.2d 981, 180 USPQ 580 (CCPA 1974) (Remarks, page 23, ¶2). This argument has been fully considered but is not persuasive. The case law and standard Applicant is referring to “all elements test” was removed from MPEP 2143 over 15 years before the effective filing date of the claimed invention. As such, In re Royka is not relevant to examination of the present claims. In any event, Examiner contends that all claimed elements are indeed taught and/or suggested by the prior art references. CXCR4 and CXCR12 were known biomarkers for endometriosis and detected by normalization to a housekeeping gene; the means to detect biomarkers in general from menstrual fluid was previously developed; and Villarreal suggests detecting biomarkers for endometriosis from menstrual fluid, including from a tampon.
Applicant argues that claim 3 is nonobvious because it depends from claim 1, which is nonobvious for the reasons argued above (Remarks, page 24). This argument has been fully considered but is not persuasive because a prima facie case of obviousness is established for claim 1 for the reasons set for in the §103 rejections of record and the response to Applicant' s arguments in paragraphs 61-65 above.
Conclusion
No claims are allowable.
THIS ACTION IS MADE FINAL. 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.
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/CATHERINE KONOPKA/Examiner, Art Unit 1635