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 .
Election/Restrictions
Claims 26 and 32-33 were previously withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected groups, there being no allowable generic or linking claim. Election was made without traverse in the reply filed on 8/13/2024.
Therefore, claims remain withdrawn.
Status of the Claims
This action is in response to papers filed 10/16/2025 in which no claims were canceled or added and claim 1 was amended.
Claims 1-4,6,8,11,13-15,17-18,20,23,25, and 29 are under prosecution.
Amendments & Response to Arguments
Applicant has amended claim 1, with a limitation of an intended function, to overcome the 103 rejection; the rejection is not withdrawn.
Rejections not reiterated here are withdrawn.
Applicant' s arguments, see Pgs. 7 - 12, filed 16th October 2025, with respect to:
rejections of claims 1-4,6,8,11,13-15,17-18,20,23,25, and 29 under 35 USC § 103 have been fully considered but are not persuasive for the reasons discussed in this office action. The 103 rejection is maintained.
Claim Interpretation
The phrase, “promoter operably linked” in claim 1 is being interpreted as per specification to mean a promoter whose activity controls a gene’s expression and not to mean a promoter located directly upstream of the gene whose activity it controls (A promoter is said to be operably linked to a gene if the promoter controls the degree to which the gene is expressed, Pg. 8, 2nd paragraph).
Wherein clause:
It is noted that the subject matter of a properly construed claim is defined by the terms that limit its scope. It is this subject matter that must be examined. As a general matter, the grammar and intended meaning of terms used in a claim will dictate whether the language limits the claim scope. Language that suggests or makes optional but does not require steps to be performed or does not limit a claim to a particular structure does not limit the scope of a claim or claim limitation. “Wherein” clauses are examples of language that may raise a question as to the limiting effect of the language in a claim. See MPEP 2103 I.C. and MPEP § 2111.04.
It is also noted that a “wherein” clause, such as that in (claim 1), must give “meaning and purpose to the manipulative steps.” See, MPEP § 2111.04.
In instant claim 1, the wherein clauses are followed by functional language: reduction or absence of the repressor results in increased expression of the output molecule and the amended limitation of the repressor…inhibits translation of a constitutively expressed RNA.
Thus, the wherein clauses recite inherent functions which will flow from the structural limitations of the claims they depend from. See, MPEP § 2112.01: II. COMPOSITION CLAIMS — IF THE COMPOSITION IS PHYSICALLY THE SAME, IT MUST HAVE THE SAME PROPERTIES.
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.
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 1-4, 6, 8, 11, 13-15, 17-18, 20, 23, 25, and 29 remain rejected under 35 U.S.C. 103 as being unpatentable over Benenson (US 2017 /0159135, Jun. 8, 2017) in view of Borchardt et al. (2015, RNA, 21: 1921-1930) for reasons of record set forth in the office action of 07-16-2025, and reiterated below.
Regarding claims 1 and 11, Benenson teach a cellular biosensor system comprising: (i) a sensor circuit comprising a first constitutive promoter (CMV), operably linked to a nucleic acid sequence encoding: (a) a repressor (LacI); and (b) one or more target sequences for a first set of one or more miRNAs (miR-21 binding region); and (ii) a signal circuit comprising a second constitutive promoter (CAG) operably linked to a nucleic acid sequence encoding: (a) a repressor recognition sequence that is capable of being bound by the repressor of (i)(a); and (b) a nucleic acid sequence encoding an output molecule (DSRed) (Benenson, Fig. 10 (rtTA-Standard Sensor). See Examiner-annotated Fig. 10 from Benenson in previous office action dated 8/21/24. Further, Benenson teach that their system is modular, i.e., allows for swapping components or circuits. See recitation below:
[0154] FIG. 11 shows the modularity of sensor design.
Circuit schemes with swapped promoter (left) or output coding
sequence (right) are shown. Performance characterization
is shown, comparing the standard and the delayed
architectures. Transfection details are given in Table 2.7
Further, in Fig. 4 legend, Fig. 4 shows an example of a preferred embodiment of Benenson’s biosensor, Benenson teach that genetic control can be either transcription or translation (para [0147]).
Examiner has interpreted Benenson’s biosensor system as a sequestron as it meets all the structural requirements as instantly claimed. Benenson do not specifically teach wherein the repressor is an endoribonuclease (claims 1 and 11), or teach the amended limitation wherein the repressor is an endoribonuclease, an RNAi molecule, or a ribozyme (claim 1).
However, at the time of the effective filing date, Borchardt teach making and using endoribonucleases in genetic circuits (abstract). Borchardt teach the bacterial CRISPR endoribonuclease Cys4 cleaves an RNA hairpin stem. Borchardt disclose this about the mechanism of action of the repressor: endoribonuclease Csy4 binds its substrate RNA through a specific 28-nt hairpin sequence and rescues the target mRNA substrate from degradation, resulting in protein expression (abstract). Non-binding mutants were unable to exert any of these effects. Thus, binding of the repressor (resulting in the absence of a free repressor) increased expression of the output.
Borchardt discloses the study providing the ability of CRISPR endoribonuclease Csy4 to control the mRNA stability and translation by transfecting two plasmid vectors in HEK293 cells; one vector containing a reporter transcript of a GFP or Gaussia Luciferase (GLuc) reporter genes with a Csy4 target hairpin (HP) inserted within the 5’ UTR and second vector containing a Csy4 encoding transcript (pg. 1922). Both the vectors were under the control of the constitutive promoter, chicken-beta actin (CBA) promoter (pg. 1927-1928).
Thus Borchardt’s first vector with Csy4 encoding transcript and second vector with reporter transcripts (i.e. the output with endoribonuclease target site within the 5’UTR) reads on sensor and signal circuit, respectively, with identical CBA promoter in each vector. When the repressor binds to its cognate sequence on the signal circuit it is sequestered, resulting in an absent repressor, the output molecule is expressed.
It would have been prima facie obvious to a person of ordinary skill in the art before the effective filing date to combine the teachings of Benenson taken with Borchardt to use as endoribonuclease as a repressor in place of LacI in Benenson’s biosensor system.
A person of ordinary skill in the art would have been motivated to combine the teachings for the advantage of creating a sequestron capable of increased control of post-transcriptional expression of intended targets, specifically binding of an RNA repressor (e.g., endoribonuclease) by its cognate sequence to result in high output expression to arrive at the instant claim 1. It would be a simple substitution for one of ordinary skill in the art to replace generic repressor or recombinase in the biosensor system of Benenson with the endoribonuclease taught by Borchardt. See MPEP 2143 I.(B) and 2144 II. One of ordinary skill in the art and would be able to make such a substitution with reasonable expectation of success because both references are in the field of genetic circuits designed to effectuate modulation of a target gene.
Thus, Benenson in view of Borchardt make obvious instant claims 1 and 11.
Regarding claim 2, Benenson teach the biosensor system of claim 1, wherein the one or more target sequences for the first set of one or more miRNAs of (i)(b) are downstream from the nucleic acid sequence encoding the repressor (Fig. 10).
Regarding claim 3, Benenson teach the biosensor system of claim 1, wherein the nucleic acid sequence encoded by the signal circuit of (ii) further comprises one or more target sequences for a second set of one or more miRNAs (Fig. 10).
Regarding claim 4, Benenson teach the biosensor system of claim 1, wherein the nucleic acid sequence encoded by the signal circuit of (ii) further comprises one or more target sequences for a second set of one or more miRNAs and wherein these sequences are downstream from the nucleic acid sequence encoding the output molecule (Fig. 10).
Regarding claim 6, Benenson teach various embodiments of their circuits. Benenson Fig. 1b shows a repressor module comprising genes in their cumulative gene action exert repressing and/or inhibitory effect(s) on an output module (Benenson Abstract, Fig. 1b). Thus, Benenson teach a cellular biosensor system comprising (i) a sensor circuit, made up of two expression cassettes, that cumulatively exert repressing and/or inhibitory effect(s) on a (ii) a signal circuit, that read on components of instantly claimed sequestron. As evidenced by Benenson, Pg. 4, lines bridging 1st and 2nd columns: CMV and CAG are constitutive promoters. See Examiner-annotated Fig. 1b from Benenson showing the discussed embodiment of a biosensor system in previous office action dated 8/21/24.
Specifically, with respect to claim 6, Benenson further teach wherein the biosensor system comprises a plurality of the signal circuit (Fig. 4), wherein the individual circuit was shown in Fig. 1b . The optional limitations following the wherein clause are not considered because they are recited as optional; i.e., not required.
Regarding claim 8, the biosensor system of claim 1 is taught above. Benenson further teach wherein the biosensor system comprises a plurality of the sensor circuit (Fig. 4). The optional limitations following the wherein clause are not considered because they are recited as optional; i.e., not required.
Regarding claim 13, the biosensor system of claim 1 is taught above. Benenson further teach wherein the first and/or second constitutive promoter is an hEF1-alpha promoter (Pg.18, column 6, line 10; and Pg.19, column 7, line 56).
Regarding claim 14, the biosensor system of claim 1 is taught above. Benenson further teach a plurality of biosensor systems of claim 1 (Fig. 4), wherein (A) the biosensor system encodes a combination of different repressors (Pg.19, column 7, line 21; and Fig.4: Gene1 depicting Activator, and Gene2 depicting Recombinase 1), (C) the repressor encoded by the sensor circuit of each of the plurality of biosensor systems is capable of binding or cleaving the repressor recognition sequence of the signal circuit of the same biosensor system (Fig.4: Activator binds to Gene2, and Recombinase 1 acts on Gene3).
Benenson do not teach (B) the repressor recognition sequence of (ii)(a) of each of the plurality of biosensor systems comprises a different nucleic acid sequence, and (D) the repressor encoded by the sensor circuit of each biosensor system is not capable of binding or cleaving the repressor recognition sequence of a different biosensor system, in the above described embodiment.
However, Benenson does describe embodiments wherein the sensor circuit comprises (B) the repressor recognition sequence of (ii)(a) of each of the plurality of biosensor systems comprises a different nucleic acid sequence (the repressor encoded by the transcriptional repressor sequence (TR) of gene (2) is selected from the group consisting of a (poly)peptide repressor, a non-coding RNA repressor, preferably a small or long non-coding RNA repressor, a combination of different repressors, paragraph [0055]); Further, Benenson teaches (D) the repressor encoded by the sensor circuit of each biosensor system is not capable of binding or cleaving the repressor recognition sequence of a different biosensor system (Negative interference with repressor components, Fig. 13).
It would have been prima facie obvious to a person of ordinary skill in the art before the effective filing date to have tried to combine the various embodiments of Benenson and obvious variations of the same into a biosensor system, to arrive at the limitations in the claimed invention. A person of ordinary skill in the art would be motivated by looking at various embodiments of Benenson and make such a combination because such a combination would be within the scope of one of ordinary skill in the art given that Benenson explicitly disclose that standard molecular biology techniques were used in the construction of plasmid DNA constructs (Benenson Pg. 26, 1st column, 1st line of example 5). See MPEP 2143 I.(A, D, and G).
Regarding claims 15 and 29, the biosensor system of claim 1 is taught above. Benenson further teach the biosensor system of claim 1 is delivered using a suitable delivery method, e.g., liposomal vesicles Benenson, Pg.16, column 2, line 18 – 26, for intended use; and Pg.21, column 2, para [0138] for delivery). As evidenced by O'Driscoll, liposomes are an excipient for delivery of nucleic acids.
Regarding claim 17, the biosensor system of claim 1 is taught above. Benenson further teach wherein the biosensor system is in a cell (Benenson abstract, line 1: a cellular biosensor system).
Regarding claim 18, the biosensor system of claim 1 is taught above. Benenson further disclose that standard molecular biology techniques were used in the construction of plasmid DNA constructs (Benenson Pg. 26, 1st column, 1st line of example 5). As evidenced by Green & Sambrook, such techniques used in the construction of the biosensor system of claim 1 are performed in a bacterial cell. Thus, the biosensor system of claim 1 in a bacterial cell is rendered obvious by Benenson.
Regarding claim 20, the cell comprising the biosensor system of claim 17 is taught above. Benenson further teach wherein the cell is a eukaryotic cell (Benenson, Pg.17, 1st column, para [0042]).
Regarding claim 23, the cell comprising the biosensor system of claim 17 is taught above. Benenson further teach wherein the cell is a diseased cell (Benenson, Pg.20, 2nd column, para [0111]).
Regarding claim 25, the cell comprising the biosensor system of claim 17 is taught above. Benenson further teach wherein the cell expresses any one of the first set of microRNAs (Benenson, Claim 4 recitation, “The cellular biosensor system of claim 1, wherein the inputs are … selected from the group consisting of endogenously expressed microRNA molecules”).
Regarding claim 26, claim 26 is being interpreted as a cell comprising the biosensor system, i.e., a cell of claim 17. Regarding claim 26, the cell comprising the biosensor system of claim 17 is taught above. Benenson further teach a method comprising maintaining the cell comprising the biosensor system in culture (paragraph [0115]).
Thus, Benenson in view of Borchardt make obvious instant claims 2-4, 6, 8, 13-15, 17-18, 20, 23, 25, and 29.
Therefore the invention as a whole would have been prima facie obvious to one ordinary skill in the art before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains.
Response to Arguments
Applicant's arguments filed 10/16/2025 to obviousness rejection of claims under 35 USC § 103 have been fully considered but are not found persuasive. Applicant's argue by:
stating a) MPEP § 2143.01 (V), if a proposed modification would render the prior art invention being modified unsatisfactory for its intended purpose, there may be no suggestion or motivation to make the proposed modification and b) MPEP § 2143.01 (VI), if the proposed modification or combination of the prior art would change the principle of operation of the prior art invention being modified, then the teachings of the references are not sufficient to render the claims prima facie obvious (Remarks, pg. 8 , middle para).
Applicants argue the "repressor" taught by the primary reference of Benenson works by inhibiting transcription of the DNA molecule and instant application defines repressors as "protein[s] or nucleic acid molecule[s] that [are] capable of inhibiting translation of an RNA, and the endoribonuclease described by Borchardt, the secondary reference, is an "RNA processing tool" and regulates expression of an output molecule by targeting translation, not transcription (Remarks, bridging pg. 8-9).
regulation of output molecule expression via transcription or translation results in distinct dynamics. Instant application is drawn to translation inhibition and not transcription inhibition; translational inhibition requires a baseline level of expression of the transcript which needs repression (Remarks, pg. 9, 2nd para).
Applicants allege that the intended purpose according to Benenson, is to “minimize the leakage of a biosensor system in the non-activated state, i.e. prior to the commencement of output expression”. and they allege that “Benenson also require use of "recombinase modules" that "enable gene rearrangement in the output module" such that transcription does not occur in the absence of a recombinase (Remarks, pg. 9, last para).
Applicant allege that Fig. 10 of Benenson, cited by the Examiner in the Office Action, is used in combination with the recombinase module to further minimize leakage of gene expression in the output module, hence the name given by Benenson to their circuit as a "double inversion" module. One of ordinary skill in the art would thus understand that the purpose of the combined system is primarily to prevent gene expression prior to transcription (Remarks, pg. 10, middle para). Applicants allege that Examiner's proposed modification of Benenson - substitution of LacI with Csy4- thus conflicts with both the intended purpose and principle operation of Benenson's biosensors …such substitution would render Benenson’s sensor inoperable and such proposed modification would require one to change the principle of operation of Benenson such operation being both transcription-repression and translation-repression, therefore, Examiner is flawed in equating the transcription repressor of Benenson (LacI) with a translation repressor (endoribonuclease of Borchardt) (Remarks, pg. 11). .
Neither Benenson nor Borchardt disclose or suggest modifying the "double inversion module" or biosensors of Benenson for alternative uses, nor has any rationale to so modify Benenson been provided by the Examiner.
Applicants arguments have been fully considered but are unpersuasive. Regarding the MPEP citations, these have been fully considered and the arguments considered and analyzed in accordance with the stated statutes.
In response, the amendments made to the claims have overcome the previous rejection argued about in ii., above.
Regarding iii., the relevance of this fact as pertinent to the instant rejection is unclear since instant composition claims do not recite dynamics. Therefore, Applicant’s arguments of transcription or translation resulting in distinct dynamics is unpersuasive.
Regarding iv., Benenson is broadly interested in solving the problem of synthetic circuits being “leaky”; i.e., prior to the commencement of output expression [0007], as Applicants have pointed out. The skill in the art of cellular biosensor gene regulatory circuits was very high before the effective filing date of the presently claimed invention. One of skill in the art knows that output expression is the end-result of both transcription and translation. Further, and as discussed in previous OA, one of skill in the art would recognize the modularity of Benenson’s design would substitute the RNA-binding repressor protein(s) (LacI) with an endoribonuclease which would result in a circuit functionally identical to the claimed circuit, and still be within the scope of Benenson’s objective. To this end, several endoribonucleases from the Cas endoribonuclease family and their respective recognition sites were known in the art (Borchardt, L column, middle para, pg. 1922).
Indeed, Benenson’s preferred embodiment is [0009] (A) a repressor module comprising one or more genes, which in their cumulative gene action exert repressing and/or inhibitory effect(s) on [0010] (B) an output module comprising at least one gene comprising at least one output sequence generating one or more output signals (i) in the absence of repressing and/or inhibitory effect(s) of the repressor module (A) and (ii) in the presence of at least one recombinase expressed by [0011] (C) a recombinase module… wherein the repressing and/or inhibitory effects of the repressor module are controlled by one or more inputs that negatively affect the repressing and/or inhibitory effects of the repressor module (A). Benenson’s FIG. 4 legend describes this well [0147] while simultaneously stating, “Filled arrows indicate transcription or translation, unless indicated otherwise”.
Regarding v., applicant’s arguments pertaining to Fig. 10, which was the embodiment used by the Examiner in the rejection, are not persuasive because:
Applicants are conflating principle of operation of Benenson's biosensors with its mechanism of action. The principle of operation of Benenson's biosensors is inhibiting output expression [0007] while its mechanism of action, transcription-repression and translation-repression as stated in Remarks (Remarks, pg. 10 , last para) is improved upon by a recombinase module (Benenson 0003, 0007, 0162).
a mechanism of action is merely an intended use. The prior art (in combination) is not required to teach the same intended use of the compound, but is rather required to teach the structural limitations of the instant claim. Benenson teaches a circuit and Borchardt teaches circuit components. Any or all of taught embodiments or components are available for further consideration by skilled artisans as they see fit. See MPEP 2114 II.
Applicant does not provide evidence that modification of Benenson’s constructs to substitute DNA-binding proteins and their target sites with endonucleases with their cleavage sites would render the construct inoperable with respect to the end-goal of inhibiting output expression. In the circuit shown in Figure 10A, Benenson use LacI, a DNA-binding protein, as Applicants have pointed out, to ‘toggle’ the switch between the two constructs i.e. use the presence of miRNA to ‘turn off’ a construct which orthogonally turns on the other construct. This does not imply that the circuits are limited to DNA-binding protein-based toggle or absolutely require a recombinase module i.e., intend to only function with a recombinase; especially considering teachings in Benenson regarding other recombinase module is to improve dynamic range of ON/OFF states. See the following recitation from Benenson para 0162:
To summarize, strong repression can be achieved
without transcriptional activator but the latter is essential for
the sensor 'de-repression' to high On state and for high
dynamic range.
Critically, this does not imply that an miRNA-detecting sensor could not be used in combination with a circuit wherein the DNA-binding protein repressors are replaced with endonucleases, that result in the same end-goal with respect to output expression. Thus, allegations that the circuit of Figure 10A could not modified in the way noted in OA are unfounded.
Regarding vi., See MPEP 2143 G. The courts have made clear that the teaching, suggestion, or motivation test is flexible and an explicit suggestion to combine the prior art is not necessary.
Conclusion
No claim is allowed.
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.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to SHABANA MEYERING, Ph.D. whose telephone number is (703)756-4603. The examiner can normally be reached M - F: 9am to 5pm EST.
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/SHABANA S MEYERING/Examiner, Art Unit 1635
/RAM R SHUKLA/Supervisory Patent Examiner, Art Unit 1635
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