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
This action is written in response to applicant’s correspondence received on 2/4/2026. Claims 23, 25-26, 28-35, 37-39, 41-43, and 46-49 are pending. Claims 23 and 41 have been amended. Claim 34 is withdrawn from consideration. Claims 48-49 are newly added. Claims 23, 25-26, 28-33, 35, 37-39, 41-43, and 46-49 are addressed in this response.
Claim Rejections - 35 USC § 112 – Written Description Rejection (Withdrawn)
The written description rejection in the non-final rejection mailed 11/4/2025 is hereby withdrawn. The Applicant has made substantial amendments to the claims, which no longer recite the claim language which the 112(a) written description rejection was directed to. As such, the written description rejection is withdrawn.
Claim Rejections - 35 USC § 112
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
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 47 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.
Regarding claim 47, claim 47 recites “the subject has been administered a plurality of different sensors that comprise different substrates.” Thus, claim 47 is reciting “different substrates. However, claim 47 depends from claim 23, which recites that the substrates comprise SEQ ID NOs 1-5. It is unclear if the “different substrates” recited in claim 47 are referring to substrates other than SEQ ID NOs 1-5, or if claim 47 is drawn to different combinations of SEQ ID NOs 1-5. The term “different substrates” lacks clear antecedent basis, as it is unclear as to which substrates are being referred.
Claim Rejections - 35 USC § 112 – New Rejection Necessitated by Amendment
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 23, 25-26, 28-33, 35, 37-39, 41-43, and 46-49 are rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, as failing to comply with the enablement requirement. The claim(s) contains subject matter which was not described in the specification in such a way as to enable one skilled in the art to which it pertains, or with which it is most nearly connected, to make and/or use the invention.
Factors to be considered in determining whether a disclosure meets the enablement requirement of 35 U.S.C. 112, first paragraph, have been described by the court in In re Wands, 8 USPQ2d 1400 (Fed. Cir. 1988). Wands states, on page 1404:
Factors to be considered in determining whether a disclosure would require undue experimentation have been summarized by the board in Ex parte Forman. They 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 these in the art, (7) the predictability or unpredictability of the art, and (8) the breadth of the claims.
Nature of the Invention/Breadth of Claims
Regarding claim 23, claim 23 is drawn to a method of detecting cancer where a sample is obtained from a subject who has been administered a scaffold comprising a modified nucleic acid barcode attached to a substrate (one of SEQ ID NOs 1-5) which is cleaved in the presence of a protease present in the subject. Claim 23 further recites that the barcode is detected using a CRISPR assay and compared with a healthy control, where either an increase of decrease in the modified nucleic acid barcode is indicative of the presence of cancer in the subject. Thus, claim 23 is broadly drawn to detecting “cancer,” where the detection of cancer is reliant upon the cleavage of a specific peptide substrate (SEQ ID NOs 1-5) to indicate the presence of a protease that is associated with cancer. Claim 23 is problematic because the method depends upon the cleavage of protease substrates which are broadly cleaved by metallo, serine, and aspartic proteases (per the specification, page 63, lines 3-7, PAP7, PAP9, PAP11, PAP13, and PAP15, which comprise SEQ ID NOs 1-5). However, the activity of such proteases is also known to be elevated in disease states which are not related to cancer, as discussed further below. Thus, the method does not inherently detect the presence of cancer, as increased protease activity is known to be associated with other diseases (diabetes, cardiovascular disease, see below), where furthermore the practitioner is not enabled to “detect cancer” in the subject because the readout of the assay may not be associated with cancer.
Furthermore, the claims broadly encompass embodiments where only one substrate is used, and does not require the combination of SEQ ID NOs 1-5. This claim language is problematic because, as discussed further below in the section entitled “Guide Provided in the Specification” individual substrates recited are not necessarily capable of detecting cancer depending upon the cancer model used (see below).
Furthermore, claim 23 encompasses embodiments of the method where, for instance, either an increase or decrease in barcode is identified when a scaffold comprising SEQ ID NO 1-5 is administered. However, not all of the substrates of SEQ ID NOs 1-5 are associated with both an increase or decrease in modified barcode detection (see discussion of “Guidance Provided in the Specification,” below).
Furthermore, per Example 1 (see below), the functionality of the method to detect cancer or protease activity appears to be dependent upon nanobody targeting to a specific environment, where absent such a nanobody, the method and system failed to show a distinction between disease or healthy states (see discussion below). This is problematic because it casts doubt on the functionality and ability of a practitioner to practice the method as presently claimed because the present claims are broadly only drawn to a protease substrate attached to a DNA barcode with no recitation of an environment-targeting nanobody as required for functionality of certain embodiments of the method, per the specification (see below).
Guidance Provided in the Specification
Regarding the examples relevant to this 112(a) rejection, the Applicant offers Examples 1-2 with respect to in vivo testing of their methods in cancer models. Examples 1 and 2 are summarized below.
Example 1
With regards to guidance provided in the specification, the Applicant has identified peptide substrates of select proteases using proteomic data analysis, where 16 peptide sequences were screened and five were identified to be the top substrates (see entire page 58 and page 62, lines 20 through page 63 first paragraph). The Applicant has identified proteases that are associated with colorectal cancer, including MMP-1, -9, 12, and Cathepsin B and D (Example 1, page 58 first paragraph). The Applicant identifies SEQ ID NOs 1-5 (Q7, Q9, PQ2, PQ12, and PQ19, as defined on page 58, final paragraph) as substrates for metastatic states and their associated proteases, where SEQ ID NOs 1-5 are broadly cover mettalo and serine protease (page 58, final paragraph). The Applicant tested a combination of SEQ ID NOs 1-5 in vivo to differentiate between mice with lung metastases and healthy controls (Figure 4C). The Applicant reports that Q7, Q9, and Q19 could be used to differentiate between disease and healthy states in the metastatic model (page 59, first paragraph). Thus, protease substrates PQ2 and PQ12 (SEQ ID NOs 3 and 4) were not able to “detect cancer” in the model tested in Example 1, Figure 4C, alone or in combination together.
The Applicant has further tested PLAU-activated nanobodies conjugated with 20-mer DNA barcodes and tested them in vivo (page 61, Figure 5). The Applicant has therefore tested their invention in an in vivo tumor model, where the readout of the model was dependent on protease cleavage. However, it should be pointed out that the scaffold in Example 1 is conjugated to a nanobody which targets the modified nucleic acid to a specific site (see page 60, final paragraph to page 61, first paragraph). However, no such targeting nanobody is recited in the claim, which is broadly drawn to scaffolds consisting only of the peptide substrate and the modified nucleic acid. Indeed, the specification indicates that without a targeting nanobody the ability of the scaffold to detect protease activity was nonexistent (“In contrast, urine samples collected from tumor-bearing mice injected with GFP-Nb-DNA activated LbaCasl2a at a rate that was almost identical to the samples from healthy controls, indicating that the release of the DNA barcodes was triggered on-target,” page 62, first paragraph). Thus, the recited method appears to omit a critical aspect of the method, namely, a targeting nanobody (see Figures 5D and 5G, which show that non-specific targeting of the scaffolds, “GFP” results in a similar readout compared with the control), which appears to be required for specific instances of detection of cancer environments and tumors, per Example 1 because release of the DNA barcode “was triggered on-target” using a directed nanobody (above). Furthermore, there is no guidance regarding what types of cancers could be detected using a non-specific combination of the five substrates/scaffolds, which cancer/microenvironmets require nanobody targeting for DNA barcode release, or what cancer could be detected by, for instance, SEQ ID NOs 3 and 4 by themselves.
Example 2
In Example 2, the Applicant has administered systemic DNA barcodes associated with identified protease substrates in a non-specific conjugate (i.e., not targeted using nanobodies). The Applicant identified and tested 5 specific protease substrates (PAP7, PAP9, PAP11, PAP13, and PAP15, which comprise SEQ ID NOs 1-5, respectively, page 63, first paragraph). These substrates broadly cover mettalo, serine, an aspartic proteases (page 63, first paragraph.
The specification recites that transcriptomic data was acquired to predict proteases associated with colorectal cancer (pages 58 and 62). Protease substrates of proteases including metallo, serine, and aspartic acid proteases were screened, and the top five were identified (e.g., Figure 13D, page 62 final paragraph to 63, first paragraph). The Applicants tested a multiplexed in vivo model for five different protease-sensitive sensors, where differences in cleavage of the substrates was observed over time between each of the protease-sensitive constructs (page 63-64, Figures 6C, 12A, 15B). Thus, the Applicant has tested protease substrates in vivo to be used with their recited invention.
The Applicant selected five substrates for further testing (e.g., Figure 13). As seen in the heat map in Figure 13D, the selected substrate PAP15 appears to be cleaved across separate families of protease (e.g., MMP, or metalloproteases, CTSE an aspartic proteas, and CTSK, a serine protease). Thus, the substrates identified in the specification itself corroborate the idea that substrates for proteases can be promiscuous across families, where the inclusion of such a substrate would not clearly react to the protease activity of a specific protease family because the substrate itself is subject to cleavage across multiple families including MMPs, CTSE (aspartic protease) and a serine protease.
Thus, taken together, the Applicant’s data appears to show that in certain cancer models the recited substrates can or can not be cleaved (Examples 1 and 2 show discrepancies between SEQ ID NOs 3 and 4 and their ability to be cleaved and used to detect cancer). Furthermore, Example 1 indicates that certain cancers and cancer models required specific targeting of the protease scaffolds using antibodies while Example 2 indicated that broad, non-specific administration without antibody/nanobody targeting is sufficient to detect certain cancers/cancer models. The data in the specification is therefore ambiguous with respect to which cancers can be detected by using the assay, and by what method (targeted vs. non-targeted) is required to use the method to detect cancer. Additionally, the specification acknowledges that the substrates are broadly targeted by classes of protease such as MMPs, serine proteases, and aspartic proteases.
Furthermore, claim 23 broadly encompasses an embodiment where a scaffold using SEQ ID NO: 1 is used, where a “decrease” in the readout of the assay recited in claim 23 is used to detect cancer. However, SEQ ID NO: 1 has only been demonstrated to be associated with an increase in the readout of the nucleic acid barcode (see Figures 4C and 6C, “Q7” and “PAP7,” both of which are increased in the tumor sample). Thus, a practitioner is a priori not enabled for using SEQ ID NO: 1 to detect a “decrease” in the level of modified nucleic acid barcode to detect cancer because the application itself only shows that an increase in nucleic acid barcode using SEQ ID NO: 1 as the substrate is associated with cancer. The trends are true for the remaining SEQ ID NOs (2-5), where at the very least the applicant must specify which change (i.e., increase or decrease) is associated with detecting cancer using each of the recited substrates (SEQ ID NOs 1-5).
Additionally, the practitioner is a priori not enabled to use, for instance, SEQ ID NOs 3 or 4 alone in a method to detect cancer, as these substrates have failed to detect cancer in in vivo tumor models when used separately (see Figure 4C, PQ2 and PQ12, which did not demonstrate an ability to distinguish cancer vs. healthy states).
State of the Art
Regarding the state of the art, there is known unpredictability with respect to using the presently recited method in order to detect cancer. In general, the recited method relies on the cleavage of protease specific substrates in order to release a detectable marker (a nucleic acid). The protease specific substrate is a substrate for MMP9 (e.g., Figure 13D, “PAP7”, i.e., SEQ ID NO: 1). However, increased expression of MMP9 is not a physiological or disease state that is unique to cancer. For instance, Yabluchanskiy (Yabluchanskiy A et al. Physiology (Bethesda). 2013 Nov;28(6):391-403) is a research article which focuses on MMP9 and its role in cardiovascular disease (Title, Abstract, and throughout). Yabluchanskiy teaches that increased expression of MMP9, the protease which targets the substrate encoded by SEQ ID NO: 1, is associated with disease which are not related to cancer or cancer progression, such as cardiovascular disease:
“the C-1562T polymorphism has been reported to increase gene expression
of MMP-9 and was associated with severity of coronary atherosclerosis,” (page 396, left column, final paragraph)
and
“polymorphism in the MMP9 gene promoter is associated with an elevated MMP9
expression and increased susceptibility to MI,” (page 396, right column, final paragraph).
Thus, as taught by Yabluchanskiy, increased expression of MMP-9, the protease which targets and cleaves SEQ ID NO: 1, is known to be associated with increased expression in cardiovascular diseases. However, the specification does not offer guidance with respect to how a scaffold comprising SEQ ID NO: 1 alone would be able to distinguish the detection of cancer from a different disease such as cardiovascular disease.
Furthermore, per instant Figure 13D, the substrate PAP7 was a target substrate for other metalloproteases such as MMP-13. However, it is known in the art that increased expression levels of MMP-13 are associated with other disease states aside from cancer such as diabetes. For instance, Lopez (Lopez J et al. Metabolites. 2023 Sep 14;13(9):1014) is an article which focuses on differential expression of genes in diabetic mouse models (Title, Abstract, and throughout). Lopez teaches that diabetic mouse models show an increased expression of MMP-13, which is a protease which targets instant SEQ ID NO: 1 (per Figure 13D of the specification). Thus, Lopez teaches that common diseases such as diabetes are also associated with an increased expression of metalloproteases such as MMP-13, which the specification indicates are proteases which would target the presently recited scaffolds.
The art therefore teaches unpredictability with respect the presently recited method’s ability to identify cancer in a subject because it is known in the art that diseases such as cardiovascular disease and diabetes are also associated with variable expression levels of MMPs with respect to healthy controls. Thus, the practitioner using the method is faced with uncertainty with respect to whether or not the method could be used to specifically identify cancer using, for instance, SEQ ID NO: 1 as the substrate in the scaffold recited in claim 23.
Additionally, as discussed above regarding Example 1 of the specification, the presently recited method requires targeting via antibodies or nanobodies in certain embodiments, while other embodiments detected protease cleavage without specific targeting. The specification does not specifically address what nanobodies or when such nanobodies would be required to target the scaffolds to specific tumors or tissues (see discussion of Example 1, above, “Guidance Provided in the Specification”). However, it is known in the art that tumor targeting using specific antibodies is unpredictable. For instance, Sanchez-Martin (Sánchez-Martín D. et al. Trends Biotechnol. 2015 May;33(5):292-301) is a research article that focuses on anti-cancer antibody targeting (Title, Abstract, and throughout). Sanchez-Martin teaches that:
“there is no technology for antigen profiling of intact tumors to identify tumor markers targetable with antibodies. Such constraints have led to a relative paucity of tumor-associated antigens for antibody targeting in oncology,” (Abstract).
Thus, Sanchez-Martin teaches that, with respect to specifically targeting tumors or tumor environments, specific antibodies to direct the scaffolds to tumor-specific cells is highly unpredictable, as technology does not exist to predict such antigens (Abstract). Thus, the practitioner is not enabled to practice the recited method where the scaffold is attached to an antibody, as presently recited in claim 38 because cancer or tumor specific antibodies are unpredictable and not well characterized, per Sanchez-Martin.
Undue Experimental Burden
In the present case, the practitioner faces undue experimental burden in order to practice the recited method. As an initial matter, the practitioner appears to be a priori not enabled to practice the recited method because the method includes embodiments where an “decrease” in nucleic acid barcode detection is observed using SEQ ID NO: 1 was used as the substrate. Such a method was not demonstrated or reduced to practice, and the practitioner would be forced to identify specific cancer conditions where a scaffold comprising SEQ ID NO: 1 yielded a “decrease” in nucleic acid barcode detection. Furthermore, a practitioner is not enabled to use, for instance, SEQ ID NO: 3 or 4 alone in order to detect cancer, as Figure 4C shows that such substrates were not indicative of disease/cancer states when tested vs. healthy controls (see Figure 4D).
Furthermore, the practitioner is not enabled to use the method to detect cancer, as the recited substrates are cleaved by proteases such as MMP9 and MMP-13 (see Figure 13D, PAP7). It is known in the art that other disease also demonstrate an increase in the expression of such metalloproteases, including cardiovascular disease or common diseases such as diabetes. As such, a practitioner would be saddled with undue experimental burden because they would be required to develop a method which could decouple the activity of proteases associated with cancer from those associated with common disease states such as diabetes.
Additionally, the practitioner is not enabled for the newly recited claims 48 and 49 because these claims depend from claim 23 and are directed to a method of detecting cancer. Thus, newly recited claims 48 and 49 are rejected because they suffer the same enablement problems associated with the detection of “cancer,” specifically, where the substrates recited in claims 48 and 49 are known targets of proteases associated with other diseases such as cardiovascular disease and diabetes.
Finally, even if the recited scaffolds are coupled with specific antibodies, as recited in claim 38, the practitioner would be unfairly burdened with discovering antigen targets associated with tumors, where such antigens are not characterized and further technologies to predict such antigens are not known for such tumors, per Sanchez-Martin.
In conclusion, the specification is not enabling for the scope of what is being claimed because it encompasses embodiments which are non-functional (individual use of SEQ ID NOs 3 and 4 to detect certain cancer/tumor conditions, where guidance is not provided for when such cancer could or could not be detected), and/or embodiments which include the use of protease substrates which are broadly targeted by proteases that are overexpressed in other disease states such as cardiovascular disease and diabetes. The practitioner is not enabled to use the method because there is no guidance or predictability with decoupling other diseases that are also associated with dysregulation of proteases from the dysregulation of proteases associated with cancer.
Response to Arguments
The Applicant’s arguments filed 2/4/2026 have been considered but are not persuasive. The Applicant argues that their amendments obviate the 112(a) Written Description rejection. This argument is persuasive, and the original 112(a) rejection is withdrawn. However, the Applicant has substantially amended the claims, where the amendments have prompted a new rationale and rejection (see 112(a) enablement rejection, above). As such, the present claims are not in condition for allowance for the reasons outlined above.
Conclusion
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to DOUGLAS CHARLES RYAN whose telephone number is (571)272-8406. The examiner can normally be reached M-F 8AM - 5PM.
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/D.C.R./Examiner, Art Unit 1635
/RAM R SHUKLA/Supervisory Patent Examiner, Art Unit 1635
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