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 .
Summary
Claims 1- 79 are pending.
Claims 1, 7, 10, 38, and 39 are objected.
Claims 1- 56, and 60- 79 are rejected.
Claims 17, 22- 34, 43, 46- 48, 50, and 57- 59 are withdrawn.
No claims are allowed.
Election/Restrictions
Claims 57- 59 withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim. Election was made without traverse in the reply filed on January 26, 2026.
Applicant’s election without traverse of:
Group I (Claims 1- 56 and 60- 79), drawn to an engineered molecular sensor and methods of using and making said sensor;
Species elections:
A. At Claims 1- 6, 8, 18, 22, and 79: TyrR (Phe & Tyr; claims 1, 2, 8, 79);
B. At Claims 2, 8, 18, 22, 43, 50, 54, 62-66, and 68: phenylalanine (Phe), tyrosine (Tyr), and tryptophan (Trp), and combinations thereof, encompassed by claims 8, 18, 43, 50, 54, and 62-63;
C. At Claim 7: R10F;
D. At Claim 10: a TyrR having at least 80% identity to TyrR comprising the mutation R10F;
E. At Claim 11: a Tyr repressible system;
F. At Claims 13-14: R10F (claim 14);
G. At Claim 17: a G415H mutation in TynA inducing reporter expression in the presence of Tyra;
H. At Claim 18: aromatic aldehyde and increased expression;
I. At Claims 19-20: overexpression of the reporter (claim 20);
J. At Claims 28-29: G415H (Tyra-selective; claim 29);
K. At Claim 35: TrpR SEQ ID NO: 11 and TrpR SEQ ID NO: 124;
L. At Claim 36: a polypeptide sequence that has at least 80% identity to the claimed SEQ ID NOs: 122, 123, 124, or 125;
M. At Claim 41: tyrosine
N. At Claim 44: TrpR variant O1;
O. At Claim 51: a polynucleotide encoding a polypeptide having TyrR activity;
P. At Claim 53: E. coli 1917 (EcN);
Q. At Claim 55: Tyra;
R. At Claim 57: aromatic amines (Applicants make this election for the sake of completeness in responding to the Restriction Requirement, however, it is noted that Group II (encompassing claims 57-59) was not elected herein);
S. At Claims 71-72: Tyr (claim 71 ); and
T. At Claim 74: Tyra.;
in the reply filed on January 26, 2026 is acknowledged.
It is noted for species election “A,” although not explicitly indicated in the initial Requirement of Election of Species dated October 24, 2025, the applicant is further limited to the engineered regulator protein TyrR with respect to claims 3- 6, 37, 56.
Furthermore, claims 17, 22- 34, 43, 46, 47, 50, are withdrawn from consideration based on the applicant’s election of TyrR for species election “A.”
Furthermore, claims 48, are withdrawn from consideration based on the applicant’s election of TyrR for species election “A” and “N.”
Regarding the possibly perceived inconsistency regarding withdrawn claims directed toward species election “A” and “N,” the requirement for restriction dated October 24, 2025 required an election of species for numerous claims. This examiner maintained consideration of such claims that were subject to further election of species, but would have otherwise been withdrawn due to inconsonance with another election of species(Claims 28 and 29 were excepted due to the withdrawal of claim 22). However, claims are withdrawn that are directed toward non-elected species.
It is noted for species election “B,” the applicant is limited to the single combination “phenylalanine (Phe), tyrosine (Tyr), and tryptophan (Trp)” concurrent with the requirement “One, singular, or combination of target aromatic molecules.” The additional election “and combinations thereof” is considered non-responsive.
It is noted for species election “K” and claim 35, the requirement for election of species was unclear and the applicant has carried the unclarity into their election. SEQ ID Nos 11 and 124 are representative of TyrR, not TrpR as indicated. This examiner recognizes the requirement for election of species “K” was unclear. However, the lack of clarity should have been noted and corrected. Claim 35 will be examined according to species election TyrR and the SEQ ID Nos 11 and 124 as consonant with applicant’s species election “A.”
It is noted for species election “L,” the applicant is limited to the single “SEQ ID No 124” concurrent with the requirement “One, singular, polypeptide sequence” and the applicant’s species election “A.” The additional elections “SEQ ID Nos 122, 123, or 125” are considered non-responsive.
Claim Objections
Claims 1, 7, 10, 38, and 39 are objected to because of the following informalities:
Claim 1 recites the limitation "ligand binding site " in lines 13 and 15 whereas this structure was referred to earlier in the claim on line 3 as “the ligand-protein binding site.” Use of consistent terminology is required throughout the claims.
Claim 7 recites the limitation "ligand binding site " in line 10 in reference to the “the ligand-protein binding site” cited on line 3 of claim 1. Use of consistent terminology is required throughout the claims.
Claim 10, line 2 recites “wild type (WT) TyrR (SEQ ID NO: 124) or having at least about 80% identity to.” This examiner interprets this line to read “wild type (WT) TyrR (SEQ ID NO: 124) or a sequence having at least about 80% identity to.”
Claim 10, line 6 recites “TyrR or TyrR having at least about 80% identity to TyrR comprising the mutation.” This examiner interprets this line to read “TyrR or TyrR having at least about 80% identity to TyrR comprising the mutations.”
Claim 10, line 8 recites “Phe+ Tyr and the reporter gene does not overexpress with Tyr in the absence of Phe.” This examiner objects to the inconsistent language shown in bold and interprets this line to read “Phe+ Tyr and the reporter gene does not overexpress in the presence of Tyr in the absence of Phe.” Similarly, lines 11 13 have the same issue. The claims will be better if they recite consistent terminology.
Regarding claim 38 recites “… wherein the presence of a target aromatic compound induces or represses reporter gene expression of the engineered regulator protein or enzyme compared to the wild type regulator protein or enzyme.” This examiner assumes the claim is intended to recite ““… wherein the presence of a target aromatic compound induces or represses reporter gene expression controlled by the engineered regulator protein or enzyme compared to the wild type regulator protein or enzyme.”
Regarding claim 39 recites “… wherein the engineered regulator protein or enzyme has a selectivity, induction, or repression response that is greater than wild type.” This examiner assumes the claim is intended to recite ““… wherein the engineered regulator protein or enzyme has a selectivity, induction, or repression response that is greater than wild type regulator protein.”
Appropriate correction is required.
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 1, 7, 8, 10, 11- 16, 18- 20, 36, 37, 44, 45, 49, 51, 52, 64, 69, and 79 are 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.
Those claims included in the statement of rejection but not otherwise discussed are rejected for depending from a rejected claim but failing to remedy the indefiniteness therein.
Regarding claim 1, the phrase "e.g." renders the claim indefinite because it is unclear whether the limitation(s) following the phrase are part of the claimed invention. See MPEP § 2173.05(d).
Furthermore, Claim 1, line 18 recites “wherein the engineered molecular sensor is optionally genomically integrated into a microorganism. ” This limitation limits the entire “engineered molecular sensor” to a DNA sequence because, proteins and protein sequences (as recited earlier in the claim) do not integrate into genomes.
Furthermore, claim 1, line 19 recites “optionally selected from a probiotic.” It is unclear whether the sensor is derived from a probiotic or is inserted into a probiotic.
Furthermore, claim 1, line 6 recites “a target aromatic compound bound to the engineered molecular sensor.” It is unclear whether the aromatic compound is a target of the sensor or whether the sensor is a target of the aromatic compound.
Furthermore, claim 1, line 5 recites “a promoter (e.g., PtyrP) capable of inducing or repressing the reporter in the presence or absence of a target aromatic compound.” It is unclear what conditions (absence or presence of aromatic compound) lead to promoter induction and repression.
The term “about” in claim 7 is a relative term which renders the claim indefinite. The term “about” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. Therefore, the limitation “at least… 80% identical” is rendered indefinite.
Furthermore, Claim 7, line 2 recites “one or more of the following mutations” and then later in line 12 recites “a polypeptide encoded by SEQ ID NO: 11 or the WT ligand binding site Sequence.” The sequences referenced in line 12 are both WT sequences according to the instant specifications. The sequences cannot simultaneously be mutations as recited in the claimed language and WT as recited in the specifications.
Regarding claim 8, the phrase "e.g." renders the claim indefinite because it is unclear whether the limitation(s) following the phrase are part of the claimed invention. See MPEP § 2173.05(d).
Claim 10, line 3 recites “the reporter gene overexpresses if Phe is present.” The extent of overexpression, among other parameters, of this limitation is unclear. Similarly, the quantity of Phe required for overexpression is unclear. The same indefiniteness is present in the similar limitations recited in lines 5, 7, 8, 10, 11, and 14.
The term “about” in claim 10 is a relative term which renders the claim indefinite. The term “about” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. Therefore, the limitation “at least… 80% identical” is rendered indefinite.
Claim 11, line 1 recites “a target aromatic compound.” It is unclear whether the aromatic compound is a target of the sensor or whether the sensor is a target of the aromatic compound.
Regarding claim 11, the phrase "e.g." renders the claim indefinite because it is unclear whether the limitation(s) following the phrase are part of the claimed invention. See MPEP § 2173.05(d).
Claim 12, line 2 recites “Phe-dysregulation without interference from intestinal Tyr.” It is unclear what structural and functional affects are meant by “interference.” In what manner, does Tyr interfere?
The term “sensitive” in claim 13 is a relative term which renders the claim indefinite. The term “sensitive” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. Therefore, the limitation of claim 8 from which the instant claim depends “A TyrR-based selective or specific sensor specifically detecting phenylalanine” is rendered indefinite.
The term “about” in claim 14 is a relative term which renders the claim indefinite. The term “about” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. Therefore, the limitation “a 12-fold repression in the presence of Tyr” is rendered indefinite.
Furthermore, the term “a 12-fold repression” in claim 14 is a relative term which renders the claim indefinite. The term “a 12-fold repression” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. Therefore, the limitation “a 12-fold repression in the presence of Tyr” is rendered indefinite.
The term “significant” in claim 15 is a relative term which renders the claim indefinite. The term “significant” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. Therefore, the limitation “TyrR sensor has no … response to Phe alone” is rendered indefinite.
Regarding claim 16, the phrase "preferably" renders the claim indefinite because it is unclear whether the limitation(s) following the phrase are part of the claimed invention. See MPEP § 2173.05(d).
The term “about” in claim 16 is a relative term which renders the claim indefinite. The term “about” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. Therefore, the limitations “a significant response is: under … 4000 au, under … 3000 au, or … at or under … 2000 au for Phe-inducible sensor; or above … 2000 au or … at or above 2000 au” is rendered indefinite.
Regarding claim 18, the phrase "e.g." renders the claim indefinite because it is unclear whether the limitation(s) following the phrase are part of the claimed invention. See MPEP § 2173.05(d).
Regarding claim 18, the use of parentheses in line 4, “(or functional fragments, conservative substitutions, mutants, or variants thereof)” in combination with the word “or” inside the parentheses renders the claim indefinite because it is unclear whether the limitation(s) following the word “or” withing the parentheses are part of the claimed invention.
Regarding claim 19, the phrase "e.g." renders the claim indefinite because it is unclear whether the limitation(s) following the phrase are part of the claimed invention. See MPEP § 2173.05(d).
The term “a repression” in claim 19 is a relative term which renders the claim indefinite. The term “a repression” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. Therefore, the limitation “the output response” is rendered indefinite.
Regarding claim 20, the phrase "e.g." renders the claim indefinite because it is unclear whether the limitation(s) following the phrase are part of the claimed invention. See MPEP § 2173.05(d).
The term “an overexpression” in claim 20 is a relative term which renders the claim indefinite. The term “an overexpression” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. Therefore, the limitation “the output response” is rendered indefinite.
Regarding claim 36, the phrase "e.g." renders the claim indefinite because it is unclear whether the limitation(s) following the phrase are part of the claimed invention. See MPEP § 2173.05(d).
Furthermore, Claim 36, line 2 recites “ the ligand binding site.” It is unclear, regarding SEQ ID No 122 what sequence portion of the protein is represented by this phrase. The limitation provide no indication as to what residues comprise the “ligand binding site.”
The term “about” in claim 37 is a relative term which renders the claim indefinite. The term “about” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. Therefore, the limitations “the KA value of the engineered TyrR sensor is between … 0.05 mM and … 0.3 mM” and “the KA value of the engineered TynA-FeaR is between about 0.001 mM and about 0.1 mM” are rendered indefinite.
The term “strong” in claim 44 is a relative term which renders the claim indefinite. The term “strong” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. Therefore, the limitation “the selective sensor having … repression” is rendered indefinite.
Regarding claim 45, the phrase "e.g." renders the claim indefinite because it is unclear whether the limitation(s) following the phrase are part of the claimed invention. See MPEP § 2173.05(d).
The term “about” in claim 49 is a relative term which renders the claim indefinite. The term “about” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. Therefore, the limitation “the O1 variant demonstrates … 60-fold and … 15-fold repression” is rendered indefinite.
Furthermore, The term “60-fold and about 15-fold” in claim 49 is a relative term which renders the claim indefinite. The term “60-fold and about 15-fold” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. Therefore, the limitation “the O1 variant demonstrates … 60-fold and … 15-fold repression … in response to Trp, 5-HTP, and Trpta, respectively” is rendered indefinite.
Regarding claim 51, the phrase "e.g." renders the claim indefinite because it is unclear whether the limitation(s) following the phrase are part of the claimed invention. See MPEP § 2173.05(d).
Claim 52 recites “engineered wild type or transgenic microorganism.” The microorganism may be engineered or wildtype, but not both.
The term “elevated” in claim 62 is a relative term which renders the claim indefinite. The term “elevated” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. Therefore, the limitation “levels of Phe” is rendered indefinite.
The term “elevated” in claim 63 is a relative term which renders the claim indefinite. The term “elevated” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. Therefore, the limitation “levels of Tyr” is rendered indefinite.
The term “elevated” in claim 64 is a relative term which renders the claim indefinite. The term “elevated” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. Therefore, the limitation “levels of PEA” is rendered indefinite.
The terms “manages” and “normalizes” in claim 69 are functional use terms which renders the claim indefinite. The terms “manages” and “normalizes” are not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. Therefore, the limitations “manages various debilitating neurological disorders” and “normalizes dysregulated metabolites” are rendered indefinite.
Regarding claim 79, the phrase "e.g." renders the claim indefinite because it is unclear whether the limitation(s) following the phrase are part of the claimed invention. See MPEP § 2173.05(d).
Claim Rejections - 35 USC § 102
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
The following is a quotation of the appropriate paragraphs of 35 U.S.C. 102 that form the basis for the rejections under this section made in this Office action:
A person shall be entitled to a patent unless –
(a)(1) the claimed invention was patented, described in a printed publication, or in public use, on sale, or otherwise available to the public before the effective filing date of the claimed invention.
Claims 1- 4, 6, 9, 10- 12, 18- 21, and 41- 43 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Chou et al. (Chou. 2015. US 2015/0353614 A1).
Regarding Claims 1- 3, 11, 18- 21, 41, and 42 Chou discloses “The present invention provides for a synthetic transcription factor (TF) comprising a first peptide capable of binding a target ligand, a second peptide capable of binding a target DNA, and a peptide linker linking the first and second peptides. In some embodiments, the target DNA is an activator or repressor site of a gene of interest. Depending on the target DNA and the gene of interest, the binding of target DNA by the synthetic TF can either activate or repress transcription of the gene of interest from a target promoter.” [0007]. Here, Chou’s disclosure teaches an engineered regulator protein comprising a ligand binding site and teaches an operably linked promoter capable of inducing or repressing a reporter (gene of interest). Chou continues to disclose embodiments of their invention “The Sensor is … a reporter gene, such as monomeric red fluorescence protein (mRFP) genes, are under the control of constitutive promoters,” [0145]. Here, Chou’s disclosure teaches the invention is a molecular sensor wherein the gene of interest a reporter. Chou furthermore, discloses “we implemented FREP [the invention] … using the tyrosine-responsive TF TyrR to regulate expression” [0082]. Here, Chou’s disclosure teaches TyrR is tyrosine-specific and the molecular sensor capable of activating (inducing or repressing) the reporter in the presence or absence of a target aromatic compound bound to the sensor. Here, Chou’s disclosure also teaches the WT protein, ligand-protein binding site or 80% identical sequence thereof. Chou further discloses “In some embodiments, the system is an in vitro or cell-free system”[0055] and dedicates a section to teach “Construction of FREP Vectors.” [0158]- [0160]. Here, Chou’s disclosure teaches the molecular sensor is optionally genomically integrated into a microorganism, optionally selected from a probiotic or is a purified cell-free sensor.
Therefore, Chou teaches at least one embodiment of all of the elements of claims 1, 2, and 3, 11, and therefore, Chou anticipates claims 1- 3, 11, 18- 21, 41, and 42.
Regarding claims 4 and 8, Chou discloses a functional TyrR mutant, TyrR E274Q, in figures 18, 19, and 20. According to the instant specifications, the TyrR ligand binding site is residues 7- 274 (page 3, line 14).
Therefore, Chou teaches all of the elements of claims 4 and independent claim 8, and therefore, Chou anticipates claims 4 and the independent claim 8.
Regarding claim 6, Chou teaches TyrR of the TyrR-based sensor is tyrosine specific as described above. Therefore, the ligands of claim 6 do not activate TyrR to induce or repress expression linked to its promoter.
Therefore, Chou teaches the embodiment “indole-3-acetic acid (IAA) do not induce reporter expression” of claim 6, and therefore anticipates claim 6.
Regarding claim 12, the limitation “for use to kinetically diagnose or treat disorders…” is simply “intended use” and not given any patentable weight.
Therefore, Chou anticipates claim 12.
Regarding claim 43, Chou discloses the use of engineered microorganisms capable of expressing TyrR in the section, “Construction of tyrR Sensor Module,” “tyrR was amplified from E.coli using the following primers…”[0155].
Therefore, Chou teaches all of claim 43, and therefore anticipates claim 43.
Claims 5, 9, 15 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Chou et al. (Chou. 2015. US 2015/0353614 A1) as evidenced by Pittard (Pittard et al. The TyrR regulon. Mol Microbiol. 2005 Jan;55(1):16-26.)
Regarding Claims 5 and 9, merely states a scientific truism that tyrosine may either repress or induce expression via the TyrR-based sensor as evidenced by Pittard “TyrR protein can mediate both activation and repression and can interact with all three aromatic amino acids [phenylalanine, tyrosine or tryptophan].” [Introduction, 2nd column, 1st paragraph].
Therefore, Chou teaches the embodiments “Tyr represses or induces reporter expression” and “the inducible promoter comprises … a repressing response to Tyr.”
Therefore, Chou teaches all of the elements of claims 5 and 9, and therefore, Chou anticipates claims 5 and 9.
Regarding claim 15, Pittard teaches TyrR-based systems may be non-responsive toward phenylalanine “TyrR protein in the presence of phenylalanine, tyrosine or tryptophan can interact with the α-subunit of RNA polymerase to activate transcription. In one unusual case, activation of a nonproductive promoter is used to repress transcription from a promoter on the opposite strand.” (Summary). This is simply another scientific truism of the limitation “wherein the TyrR sensor has no significant response to Phe alone.”.
Therefore, Chou teaches all of the elements of claim 15, and therefore, Chou anticipates claim 15.
Claim 79 is rejected under 35 U.S.C. 102(a)(1) as being anticipated by Chou et al. (Chou. 2015. US 2015/0353614 A1) as evidenced by Kwok (Kwok et al. Mol Microbiol. 1995 Aug;17(3):471-81.).
Regarding Claim 79, Chou discloses “The present invention also provides for a genetically modified host cell comprising the synthetic TF [transcription factor] or system useful for the methods of the present invention” [0009]. Here, Chou teaches a method of protein engineering.
Chou discloses their method of constructing their mutant TyrR protein and the specific point mutation in the known ligand binding site, “pCtl-RFP-SaroF1 was constructed by mutating TyrR with the primers TyrR-E274Q-F and TyrR-E274Q-R to make TyrR E274Q” [0214]. Here, Chou teaches mutagenizing specific amino acids in and around a ligand-binding site of a protein or enzyme wherein the mutagenizing enables changes in ligand-protein binding specificity. The latter teaching is evidenced by Kwok two decades prior to Chou. Kwok shows the differential ligand binding specificity of WT TyrR compared to mutant E274Q TyrR, “this observation suggested that the E274Q mutation lowered the affinity of TyrR for tyrosine more than fourfold” (page 475, 2nd column).
Chou further discloses their synthetic transcription factor system, “The present invention provides for a synthetic transcription factor (TF) comprising a first peptide capable of binding a target ligand, a second peptide capable of binding a target DNA, and a peptide linker linking the first and second peptides. In some embodiments, the target DNA is an activator or repressor site of a gene of interest.” [0007]. Here, Chou teaches the element ligand-protein binding while maintaining protein-DNA interaction and thus downstream gene expression control and the element linking ligand-protein binding to output response.
Therefore, Chou, as evidenced by Kwok, teaches all of the elements of claim 79, and therefore anticipates claim 79.
Claim Rejections - 35 USC § 103
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
Claim 10 is rejected under 35 U.S.C. 103 as being unpatentable over Chou as applied to claim 1 above, and further in view of Fukui (Fukui. US 2018/0230441 A1) as evidenced by Pittard (Pittard et al. The TyrR regulon. Mol Microbiol. 2005 Jan;55(1):16-26.).
Regarding claim 10, Chou teaches all of the elements of claim 8.
Chou do not teach wherein TyrR is selected from: wild type (WT) TyrR (SEQ ID NO: 124) or having at least about 80% identity to WT TyrR (SEQ ID NO: 124) and the reporter gene overexpresses if Phe is present.
Fukui discloses “the amino acid sequence of the TyrR protein encoded by this gene is shown as SEQ ID NO: 94.” [0118]. Fukui’s SEQ ID No: 94 shares 100% identity with the instant claim’s SEQ ID No 124.
It would have been obvious to a person having ordinary skill in the art (PHOSITA) at the time of filing of the instant application to have taken the Chou’s TyrR described above, and simply used Fukui’s TyrR SEQ ID No 94 because it is a simple substitution of one known element for another to obtain predictable results. A PHOSITA would have known the TyrR of Chou is structurally and functionally identical to Fukui’s TyrR SEQ ID No 94. Therefore a PHOSITA would have substituted Fukui’s SEQ ID No 94 into Chou’s molecular sensor and had a reasonable expectation of success of arriving at the instant invention.
Pittard teaches the reporter gene overexpresses if Phe is present in the following disclosure “TyrR protein in the presence of phenylalanine, tyrosine or tryptophan can interact with the α-subunit of RNA polymerase to activate transcription.” (Summary, 1st column, 1st paragraph).
Therefore, Chou and Fukui teach the embodiment “wherein TyrR is selected from: wild type (WT) TyrR (SEQ ID NO: 124) or having at least about 80% identity to WT TyrR (SEQ ID NO: 124) and the reporter gene overexpresses if Phe is present” and therefore, Chou and Fukui teach all of the elements of claim 10.
Claims 35 and 36, are rejected under 35 U.S.C. 103 as being unpatentable over Chou as applied to claim 1 above, and further in view of Fukui (Fukui. US 2018/0230441 A1).
Regarding claims 35 and 36, Chou teaches all of the elements of claim 1.
Chou does not teach the reference nucleotide sequence encoding TyrR is SEQ ID NO: 11 and the reference amino acid sequence for TyrR is SEQ ID NO: 124 (claim 35) nor does Chou teach wherein the engineered TyrR protein comprises at least 80$ identity to SEQ ID No 124 (TyrR WT) (claim 36).
Fukui discloses “the amino acid sequence of the TyrR protein encoded by this gene is shown as SEQ ID NO: 94.” [0118]. Fukui’s SEQ ID No: 94 shares 100% identity with the instant claim’s SEQ ID No 124.
It would have been obvious to a person having ordinary skill in the art (PHOSITA) at the time of filing of the instant application to have taken the Chou’s TyrR described above, and simply used Fukui’s TyrR SEQ ID No 94 because it is a simple substitution of one known element for another to obtain predictable results. A PHOSITA would have known the TyrR of Chou is structurally and functionally identical to Fukui’s TyrR SEQ ID No 94. Therefore a PHOSITA would have substituted Fukui’s SEQ ID No 94 into Chou’s molecular sensor and had a reasonable expectation of success of arriving at the instant invention.
Therefore, Chou and Fukui teach the embodiment “the reference nucleotide sequence encoding TyrR is SEQ ID NO: 11 and the reference amino acid sequence for TyrR is SEQ ID NO: 124” and therefore, Chou and Fukui teach all of the elements of claim 35.
Similarly, Chou and Fukui teach the embodiment “at least 80% identity to SEQ ID NO: SEQ ID NO: 124 (tyrR WT)” and therefore, Chou and Fukui teach all of the elements of claim 36.
Claims 37 and 38, are rejected under 35 U.S.C. 103 as being unpatentable over Chou as applied to claim 1 above, and further in view of Wilson (Wilson et al. Molecular Microbiology, 17: 483-492.).
Regarding claims 37 and 38, Chou teaches all of the elements of claim 1.
Chou does not teach wherein the KA value of the engineered regulator protein is less than the KA value of the wild type regulator protein in the presence of a target aromatic compound (claim 37) nor does Chou teach wherein the presence of a target aromatic compound induces or represses reporter gene expression of the engineered regulator protein or enzyme compared to the wild type regulator protein or enzyme (claim 38).
Wilson discloses TyrR WT binding constants “dissociation constants for this site in the dimeric and hexameric forms of TyrR were determined to be 330 [Symbol font/0x6D]M and 24 [Symbol font/0x6D]M, respectively, for tyrosine, and 55 mM and 3.7 mM, respectively, for phenylalanine.” [Abstract]. Furthermore, Wilson explains the binding constants of the engineered TyrR mutants are lower compared to WT while simultaneously showing comparative regulatory properties of WT TyrR versus TyrR mutants “TyrR G237D, TyrR E302R, TyrR G324C, TyrR G285Q and TyrR E274Q… With all five mutants, phenylalanine-mediated repression at aroG was similar to that seen with wild-type TyrR, while tyrosine-mediated repression at other operons was either partial or not detectable.” [page 489, 2nd column, last paragraph].
It would have been obvious to a person having ordinary skill in the art (PHOSITA) at the time of filing of the instant application to have taken the Chou’s TyrR described above, and simply used Wilson’s TyrR or TyrR mutants because it is a simple substitution of one known element for another to obtain predictable results. A PHOSITA would have known the TyrR of Chou is structurally and functionally identical to Wilson’s TyrR. Therefore a PHOSITA would have substituted Wilson’s TyrR or engineered TyrR mutants into Chou’s molecular sensor and had a reasonable expectation of success of arriving at the instant invention.
Therefore, Chou and Wilson teach the embodiment “wherein the KA value of the engineered regulator protein is less than the KA value of the wild type regulator protein in the presence of a target aromatic compound” and therefore, Chou and Wilson teach all of the elements of claim 37.
Similarly, Chou and Wilson teach the embodiment “wherein the presence of a target aromatic compound induces or represses reporter gene expression of the engineered regulator protein or enzyme compared to the wild type regulator protein or enzyme” and therefore, Chou and Wilson teach all of the elements of claim 38.
Claim 39 is rejected under 35 U.S.C. 103 as being unpatentable over Chou as applied to claim 1 above, and further in view of Kwok (Kwok et al. Mol Microbiol. 1995 Aug;17(3):471-81.).
Regarding claims 39, Chou teaches all of the elements of claim 1.
Chou does not teach wherein the engineered regulator protein or enzyme has a selectivity, induction, or repression response that is greater than wild type.
Kwok discloses “With this promoter the β-galactosidase levels in minimal medium were slightly higher in the TyrR E274Q strain than in wild-type TyrR,” [page 474, 2nd column, 1st paragraph].
It would have been obvious to a person having ordinary skill in the art (PHOSITA) at the time of filing of the instant application to have taken the Chou’s TyrR described above, and simply used Kwok’s TyrR or TyrR mutants because it is a simple substitution of one known element for another to obtain predictable results. A PHOSITA would have known the TyrR of Chou is structurally and functionally identical to Kwok’s TyrR. Therefore a PHOSITA would have substituted Kwok’s TyrR or engineered TyrR mutants into Chou’s molecular sensor and had a reasonable expectation of success of arriving at the instant invention.
Therefore, Chou and Kwok teach the embodiment “wherein the engineered regulator protein or enzyme has a selectivity, induction, or repression response that is greater than wild type” and therefore, Chou and Kwok teach all of the elements of claim 39.
Claim 40 is rejected under 35 U.S.C. 103 as being unpatentable over Chou as applied to claim 1 above, and further in view of Koyanagi (Koyanagi et al. J Bacteriol. 2008 Dec;190(24):8238-43.).
Regarding claim 40, Chou teaches all of the elements of claim 1.
Chou does not teach wherein the regulator protein or enzyme or the regulator protein or enzyme binding site is modified to increase selectivity.
Koyanagi discloses multiple experiments directed against TyrR mutants. These experiments alter concentrations of phenylalanine and tyrosine and measure the protein’s promoter induction/repression response. “In the present study, we tried to isolate second-site suppressors of this mutant protein and identified asparagine-316 as a critical residue in the fine-tuning of the oligomeric state of TyrR,” [page 8242, 2nd column, 2nd paragraph]. Kwok notes “fine tuning” the protein to achieve optimum oligomerization.
It would have been obvious to a person having ordinary skill in the art (PHOSITA) at the time of filing of the instant application to have taken the Chou’s TyrR described above, and simply used Koyanagi’s oligomer optimization method of TyrR mutants because it is a choosing from a finite number of identified, predictable solutions, with a reasonable expectation of success. A PHOSITA would have known the TyrR of Chou is structurally and functionally identical to Koyanagi’s WT TyrR. Furthermore, they would be aware of the N316 ligand binding site highlighted by Koyanagi. Therefore a PHOSITA would have substituted Koyanagi’s N316 mutant TyrR, or variants into Chou’s molecular sensor and had a reasonable expectation of success. Therefore, It would be obvious to extend Koyanagi’s oligomerization optimization and interpret the results toward the instant applications ligand selectivity optimization and arrived at the instant invention.
Therefore, Chou and Koyanagi teach all of the elements of 40.
Claim 45, is rejected under 35 U.S.C. 103 as being unpatentable over Chou as applied to claim 1 above, and further in view of Yu (Yu et al. 2008 May;24(5):844-50. Article in Chinese).
Regarding claims 45, Chou teaches all of the elements of claim 1.
Chou does not teach wherein a genomic copy of wildtype trpR is knocked out from the engineered microorganism selected from an E. coli strain.
Yu discloses “To improve tryptophan production in Escherichia coli, key genes in the tryptophan biosynthesis pathway -aroG, trpED, trpR and tnaA were manipulated. TrpR gene was knocked out…” [Abstract].
It would have been obvious to a person having ordinary skill in the art (PHOSITA) at the time of filing of the instant application to have taken the Chou’s E.coli described above, and simply used Yu’s TrpR knockout E.coli because it is a simple substitution of one known element for another to obtain predictable results. A PHOSITA would have known the E.coli of Chou is structurally and functionally identical to Yu’s WT E.coli. Therefore a PHOSITA would have substituted Yu’s method of TrpR knockout into Chou’s E.coli based molecular sensor and had a reasonable expectation of success of arriving at the instant invention.
Therefore, Chou and Yu teach all of the elements of claim 45.
Claim 51, is rejected under 35 U.S.C. 103 as being unpatentable over Zhu (Zhu et al. Protein Expr Purif. 1997 Jul;10(2):237-46.) as evidenced by SnapGene (SnapGene_pET-3a Sequence and Map_2026 May 12. www.snapgene.com/plasmids/pet_and_duet_vectors_(novagen)/pET-3a.), and further in view of Cornish (Cornish et al. J Biol Chem. 1986 Jan 5;261(1):403-10.) as evidenced by Ruteshouser (Ruteshouser et al. J Mol Biol. 1989 Jul 5;208(1):23-43.).
Regarding claims 51, Zhu discloses transforming E. coli with plasmid pZZ “pZZ contains an insertion of H. influenzae tyrR+ (in anticlockwise orientation) between the BamHI and NdeI sites of the pET-3a vector… The ligation mixture was used to transform E. coli DH5α.”. (page 239, 2nd column). As evidenced by SnapGene, inspection of the pET-3a vector shows from 5’ to 3’ a T7 promoter, the restriction site where tyrR gene was inserted and the ampicillin-based reporter (SnapGene pET-3a website).
Here, Zhu teaches an artificial DNA construct comprising, as operably associated components in the 5' to 3' direction of transcription comprising a promoter functional in a microorganism, a polypeptide having TyrR activity, wherein the microorganism is capable of expressing non-native TyrR, and a reporter gene.
Zhu does not teach a termination sequence downstream of the reporter.
However, SnapGene does evidence a transcriptional termination sequence on the pET-3a plasmid downstream of the gene insertion site.
Cornish discloses plasmid pMU364 showing from 5’ to 3’ the tyrR gene and the lacZα reporter “FIG. 1. Physical map of relevant plasmids. The DNA is represented as follows: solid regions, pBR322; diagonal stripes, tyrR; broken vertical stripes, lac' I' lac' Z; dotted, lac Y' X fusion from pLG400 ( 15 ); intact vertical stripes, pMC489… The thick arrow shows the direction of transcription of tyrR.” (Fig. 1 legend; see figure below). The lacZα reporter comprises a 3’ transcription termination sequence as evidenced by Ruteshouser “These termination stop points correspond to five of the 11 transcriptional pause sites” (Abstract).
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It would have been obvious for a person having ordinary skill in the art (PHOSITA) to place the post-reporter transcriptional termination sequence taught by Cornish and inserted the sequence into Zhu’s pZZ plasmid downstream of the ampicillin reporter because it is simply combining prior art elements according to known methods to yield a predictable result.
A PHOSITA would look for the most efficient means of gene expression and manipulation. A PHOSITA starting with Zhu’s pZZ would recognize the problems associated with aberrant transcription and therefore would want to terminate gene transcription at their preferred location. Therefore, a PHOSITA would have included Cornish’s transcriptional termination sequence downstream of Zhu’s pZZ plasmid’s ampicillin-based reporter sequence and had a reasonable expectation of success of arriving at the instant invention.
Regarding the limitation “the microorganism specifically expresses or represses reporter gene expression compared to a microorganism not comprising the artificial DNA construct in the presence or absence of aromatic compounds." The microorganism not having the reporter gene construct would be incapable of expression or repression. Therefore, it is obvious that a microorganism having a reporter gene construct would express or repress the gene compared to a microorganism not comprising that same gene.
Therefore, Zhu and Cornish and Ruteshouser teach all of the elements of claim 51.
Claim 52 is rejected under 35 U.S.C. 103 as being unpatentable over Zhu (Zhu et al. Protein Expr Purif. 1997 Jul;10(2):237-46.) as evidenced by SnapGene (SnapGene_pET-3a Sequence and Map_2026 May 12. www.snapgene.com/plasmids/pet_and_duet_vectors_(novagen)/pET-3a.), and Cornish (Cornish et al. J Biol Chem. 1986 Jan 5;261(1):403-10.) as evidenced by Ruteshouser (Ruteshouser et al. J Mol Biol. 1989 Jul 5;208(1):23-43.) as applied to claim 51 above, and further in view of Lin (Lin et al. Anal Chim Acta. 2018 Dec 24;1041:108-113.).
Regarding claim 52, Zhu and Cornish teach all of the elements of claim 51.
Zhu and Cornish do not teach the DNA construct is a sensor.
Lin discloses developing TyrR biosensors “we developed a whole-cell biosensor for the simultaneous detection of Phe and Tyr based on the transcriptional regulation of TyrR in E. coli,” (page 109, 1st paragraph). Furthermore, Lin discloses their biosensor DNA construct comprising 5’ to 3’ a promoter, TyrR, and a reporter, similar to Zhu’s pZZ (Fig. 1A, see insert below). Additionally, Lin teaches a motivation for the use of their biosensor DNA construct “Clinicians require a simple quantitative method for the detection of both phenylalanine and tyrosine to facilitate the diagnosis of phenylketonuria” (Abstract).
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It would have been obvious to a person having ordinary skill in the art (PHOSITA) at the time of filing of the instant application to have taken the Zhu-based DNA construct described above, and simply used it as a biosensor as described by Lin because it is a simple substitution of one known element for another to obtain predictable results. A PHOSITA familiar with the DNA construct teachings of Zhu would also be familiar with the similar DNA construct and biosensor teachings of Lin. The PHOSITA would recognize the opportunity of the practical application of Zhu’s DNA construct toward clinical use. Therefore a PHOSITA would have applied Zhu’s DNA construct toward the use of Lin’s biosensor and had a reasonable expectation of success of arriving at the instant invention.
Therefore, Zhu and Cornish and Lin teach all of the elements of claim 52.
Claim 53 is rejected under 35 U.S.C. 103 as being unpatentable over Zhu (Zhu et al. Protein Expr Purif. 1997 Jul;10(2):237-46.) as evidenced by SnapGene (SnapGene_pET-3a Sequence and Map_2026 May 12. www.snapgene.com/plasmids/pet_and_duet_vectors_(novagen)/pET-3a.), and Cornish (Cornish et al. J Biol Chem. 1986 Jan 5;261(1):403-10.) as evidenced by Ruteshouser (Ruteshouser et al. J Mol Biol. 1989 Jul 5;208(1):23-43.) and Lin (Lin et al. Anal Chim Acta. 2018 Dec 24;1041:108-113.) as applied to claim 52 above, and further in view of Chen (Chen et al. ACS Synth. Biol. 2021, 10, 10, 2566–2578.).
Regarding claim 53, Zhu, Cornish, and Lin teach all of the elements of claim 52.
Zhu, Cornish, and Lin do not teach the microorganism is selected from Escherichia coli Nissie 1917 (EcN).
Chen discloses developing Escherichia coli Nissie 1917 biosensors “we engineered a set of NO-responsive biosensors based on the PnorV promoter and its NorR regulator in the norRVW operon; the circuits were characterized and optimized in probiotic Escherichia coli Nissle 1917” (Abstract).
It would have been obvious to a person having ordinary skill in the art (PHOSITA) at the time of filing of the instant application to have taken the Zhu-Lin-based biosensor in E. coli DH5α described above, and instead of using DH5α substituted in Escherichia coli Nissie 1917 (EcN) as described by Chen because it is a simple substitution of one known element for another to obtain predictable results. A PHOSITA familiar with the molecular biology and biosensor teachings of Zhu, and Cornish, and Lin would also be familiar with the similar molecular biology and biosensor teachings of Chen. The PHOSITA would recognize the limited clinical capabilities of Zhu’s DH5α. Therefore a PHOSITA would have applied Chen’s EcN toward the use of Zhu-Lin’s biosensor and had a reasonable expectation of success of arriving at the instant invention.
Therefore Zhu, Cornish, Lin, and Chen teach all of the elements of claim 53.
Claim 54 is rejected under 35 U.S.C. 103 as being unpatentable over Zhu (Zhu et al. Protein Expr Purif. 1997 Jul;10(2):237-46.) as evidenced by SnapGene (SnapGene_pET-3a Sequence and Map_2026 May 12. www.snapgene.com/plasmids/pet_and_duet_vectors_(novagen)/pET-3a.), and Cornish (Cornish et al. J Biol Chem. 1986 Jan 5;261(1):403-10.) as evidenced by Ruteshouser (Ruteshouser et al. J Mol Biol. 1989 Jul 5;208(1):23-43.) and Lin (Lin et al. Anal Chim Acta. 2018 Dec 24;1041:108-113.) as applied to claim 52 above, and further evidenced by Pittard (Pittard et al. The TyrR regulon. Mol Microbiol. 2005 Jan;55(1):16-26.).
Regarding claims 54 and 56, Zhu, Cornish, and Lin teach all of the elements of claim 52.
The further claim limitation “the selective sensor is selective for an aromatic compound and the aromatic compound is an aromatic amino acid selected from phenylalanine (Phe), tyrosine (Tyr), and tryptophan (Trp)” merely states a scientific truism, evidenced by Pittard and repeated from above, “TyrR protein can mediate both activation and repression and can interact with all three aromatic amino acids [phenylalanine, tyrosine or tryptophan].” [Introduction, 2nd column, 1st paragraph].
Therefore, Zhu, Cornish, and Lin teach all of the elements of claim 54 and 56.
Claims 60- 62, 67, 70, 71, 75, and 76, are rejected under 35 U.S.C. 103 as being unpatentable over Chou as applied to claim 1 above, and further in view of Lin (Lin et al. Anal Chim Acta. 2018 Dec 24;1041:108-113.).
Regarding claims 60, Chou teaches all of the elements of claim 1. Furthermore, Chou teaches their method in E. coli cells “FIG. 18. Fluorescent output from tyrosine sensors using ParoF E. coli DJ106 and DJ166 with one of seven sensors consisting of the promoter ParoF and a variant of TyrR” [0028].
Chou do not teach obtaining a biological sample from a subject and contacting the biological sample with the engineered molecular sensor.
Lin discloses “we developed a novel approach to the analysis of Phe/Tyr in urine samples using cell-based biosensor technology.”(Conclusion). Furthermore, Lin suggests “This could yield performance models useful to the development of new applications for the tyrR regulon” (Conclusion). Lin further teaches in their Experimental methods, “E. coli DH5α cells were used as the hosts” (page 109, 1st column).
It would have been obvious to a person having ordinary skill in the art (PHOSITA) at the time of filing of the instant application to take the engineered molecular biosensor DJ106 cells of Chou and substitute them for the DH5α biosensor cells of Lin and then contacted them with a urine biological sample as performed by Lin because it is a simple substitution of one known element for another to obtain predictable results. A PHOSITA would recognize E. coli biosensor cells used in the methods of Lin would exhibit similar fluorescent profiles as the E. coli biosensor cells of Chou. Therefore a PHOSITA would have applied Chou’s DJ106 cells toward the use of Lin’s DH5α cell-based biosensor and had a reasonable expectation of success of arriving at the instant invention.
Therefore, Chou, and Lin teach all of the elements of claim 60.
Regarding claims 61, 62, 67, 70, 71, 75, and 76, Chou, and Lin teaches all of the elements of claim 60. (Claim 67, 70, 71, and 76 do not depend from claim 60 but the foregoing analysis applies).
Furthermore, Lin immediately discloses the motivations for their biosensor in the first sentence “Clinicians require a simple quantitative method for the detection of both phenylalanine and tyrosine to facilitate the diagnosis of phenylketonuria, a common inherited disorder of amino acid metabolism.” (Abstract). “We developed a whole-cell biosensor for the simultaneous detection of Phe and Tyr based on the transcriptional regulation of TyrR in E. coli” (page 109, 1st column, 1st paragraph). Lin further discloses “the proposed method could provide potential applications and simple solutions to the analysis of Phe/Tyr content in urine of PKU [phenylketonuria] patients” (Conclusion).
Therefore, Chou, and Lin teach all of the elements of claim 61 and 62, 67, 70, 71, 75, 76.
Regarding claims 77, Chou, and Lin teaches all of the elements of claim 60. (Claim 77 does not depend from claim 60 but the foregoing analysis applies).
Furthermore, Lin discloses their biosensor’s ligand-protein binding selectivity, “we evaluated the selectivity of the proposed sensor by examining the fluorescence induction for twenty amino acids, PPA, and PA” (page 110, 2nd column, section 3.2). Lin further discloses this selectivity while maintaining protein-DNA (TyrR-promoter) interaction and downstream gene expression control (gfp), “We began by selecting five promoters (aroF, tyrP, aroP, mtr, and aroG) for use in examining the expression of fluorescent protein(s) in the presence of Phe and Tyr, respectively.” (page 109, 2nd column, section 3.1).
Therefore, Chou, and Lin teach all of the elements of claim 77.
Claims 63 is rejected under 35 U.S.C. 103 as being unpatentable over Chou and Lin 2018 as applied to claim 60 above, and further in view of Lin 2019 (Lin et al. Anal. Methods,2019,11,1400.).
Regarding claim 63, Chou and Lin 2018 teach all of the elements of claim 60.
Chou and Lin 2018 do not teach wherein elevated levels of Tyr detected by the sensor indicate the subject has type 2 tyrosinemia.
Lin 2019 discloses “we developed a fully integrated reagentless platform in which biocompatible agarose is used as an entrapment agent with cell sensors… based on a Tyr sensor prototype uses the native regulatory system in E. coli to monitor variations in Tyr concentrations”(Introduction, 2nd column). Furthermore, Lin 2019 describes their strains and methods were “performed in E. coli DH5a” (page 1400, 2nd column) and “urine analysis, 10% artificial urine solution” page 1401, 1st column).
It would have been obvious to a person having ordinary skill in the art (PHOSITA) at the time of filing of the instant application to take the phenylketonuria biosensor of Chou and Lin 2018 and substitute the detection of phenylketonuria for the biosensor detector of tyrosinemia of Lin 2019. In doing so, a PHOSITA would continue by substituting an artificial urine biological sample as performed by Lin 2019 because it is a simple substitution of one known element for another to obtain predictable results. A PHOSITA would recognize artificial urine used in the methods of Lin 2019 would exhibit similar fluorescent profiles as human urine of Chou and Lin 2018. Therefore a PHOSITA would have applied Lin’s 2019 artificial urine toward the use of Chou and Lin’s 2018 human urine biosensor and had a reasonable expectation of success of arriving at the instant invention.
Therefore, Chou, Lin 2018 and Lin 2019 teach all of the elements of claim 63.
Claim 64, is rejected under 35 U.S.C. 103 as being unpatentable over Chou and Lin 2018 as applied to claim 60 above, and further in view of Myojin (Myojin et al. Jpn J Psychiatry Neurol. 1989 Jun;43(2):171-6.).
Regarding claim 64, Chou and Lin 2018 teach all of the elements of claim 60.
Chou and Lin 2018 do not teach wherein elevated levels of PEA detected by the sensor indicate the subject has a psychological disorder.
Myojin discloses “The plasma PEA levels of the schizophrenics were significantly higher than those of the normals, and the platelet PEA levels of the schizophrenics were lower than those of the normals”(Abstract). Furthermore, Myojin describes PEA as a biomarker in urine “Fischer et al. (1968, 1972) first reported elevated concentrations of urinary PEA in schizophrenic patients using a fluorescence method.” (page 174, Discussion, 1st column). Finally, Myojin provides a suggestion to study PEA levels in biological samples, “Therefore, more precise data on the PEA concentrations in CSF, the blood and urine of schizophrenic patients were earnestly desired” (page 174, Discussion, 2nd column).
It would have been obvious to a person having ordinary skill in the art (PHOSITA) at the time of filing of the instant application to use the urine biosensor of Chou and Lin 2018 and apply Myojin’s technique of PEA detection as a means to indicate a psychological disorders because it is simply applying a known technique to a known device (method, or product) ready for improvement to yield predictable results.
A PHOSITA would recognize urine sample-based methods of Chou and Lin 2018 would exhibit similar fluorescent profiles as urine sample-based methods of Myojin. Furthermore, a PHOSITA would have followed the suggestion of Myojin and applied the suggestion toward Chou and Lin’s 2018 biosensor. Furthermore, as shown by Myojin, a PHOSITA would recognize PEA detection as a means to indicate a psychological disorder.
Therefore a PHOSITA would have applied Myojin’s PEA detection as a means to indicate a psychological disorders toward the use of Chou and Lin’s 2018 human urine biosensor and had a reasonable expectation of success of arriving at the instant invention.
Therefore, Chou, Lin 2018 and Myojin teach all of the elements of claim 64.
Claim 65 is rejected under 35 U.S.C. 103 as being unpatentable over Chou and Lin 2018 as applied to claim 60 above, and further in view of Rafehi (Rafehi et al. Front Pharmacol. 2019 Oct 30;10:1297.).
Regarding claim 65, Chou and Lin 2018 teach all of the elements of claim 60.
Chou and Lin 2018 do not teach wherein the presence of Tyra detected by the sensor indicates catecholamine release and an increase in blood pressure.
Rafehi discloses “Tyramine functions as an indirect sympathomimetic by promoting the release of noradrenaline from synaptic vesicles, thereby having hypertensive effects (referred to as the tyramine pressor response;”(Introduction, 1st column).
Furthermore, Rafehi describes their method of using tyramine as a biomarker in urine “Urine samples were taken from 3 h to 10 min before tyramine administration.” (Materials and Methods, page 3, 1st column 2nd paragraph).
It would have been obvious to a person having ordinary skill in the art (PHOSITA) at the time of filing of the instant application to use the urine biosensor of Chou and Lin 2018 and apply Rafehi’s technique of tyramine detection as a means to indicate catecholamine release and an increase in blood pressure because it is simply applying a known technique to a known device (method, or product) ready for improvement to yield predictable results.
A PHOSITA would recognize urine sample-based biosensors of Chou and Lin 2018 would exhibit similar measurements as urine sample-based methods of Rafehi. Furthermore, as shown by Rafehi, a PHOSITA would recognize that urinal tyramine to indicates catecholamine release and an increase in blood pressure.
Therefore a PHOSITA would have applied Rafehi’s tyramine detection as a means to indicate catecholamine release and an increase in blood pressure toward the use of Chou and Lin’s 2018 human urine biosensor and had a reasonable expectation of success of arriving at the instant invention.
Therefore, Chou, Lin 2018 and Rafehi teach all of the elements of claim 65.
Claim(s) 66, is/are rejected under 35 U.S.C. 103 as being unpatentable over Chou and Lin 2018 as applied to claim 60 above, and further in view of Takaki (Takaki et al. Neuroscience. 1985 Sep;16(1):223-40.).
Regarding claim 66, Chou and Lin 2018 teach all of the elements of claim 60.
Chou and Lin 2018 do not teach wherein the presence of Trypta detected by the sensor causes serotonin release and stimulation of gastrointestinal motility.
Takaki discloses “Measurements of the release of preloaded [3H]serotonin or [3H]norepinephrine revealed that tryptamine is a potent releaser of these labeled amines… Moreover, incubation with tryptamine depleted the myenteric plexus of endogenous serotonin.”(Abstract).
Furthermore, Takaki discloses contacting the sample to sensors, “Intracellular recordings showed that administration of pulses of tryptamine mimicked one of the actions of serotonin…” (Abstract).
It would have been obvious to a person having ordinary skill in the art (PHOSITA) at the time of filing of the instant application to use the cellular biosensor of Chou and Lin 2018 and apply Takaki’s technique of tryptamine detection as a means to indicate serotonin release and stimulation of gastrointestinal motility because it is simply applying a known technique to a known device (method, or product) ready for improvement to yield predictable results.
A PHOSITA would recognize cellular sample-based biosensors of Chou and Lin 2018 would exhibit similar measurements as intracellular sample-based methods of Takaki. Furthermore, as shown by Takaki, a PHOSITA would recognize that cellular tryptamine causes serotonin release and gastrointestinal motility.
Therefore a PHOSITA would have applied Takaki’s tryptamine detection as a means to indicate serotonin release and stimulation of gastrointestinal motility toward the use of Chou and Lin’s 2018 cellular biosensor and had a reasonable expectation of success of arriving at the instant invention.
Therefore, Chou, Lin 2018 and Takaki teach all of the elements of claim 66.
Claim(s) 68, is/are rejected under 35 U.S.C. 103 as being unpatentable over Chou and Lin 2018 as applied to claim 67 above, and further in view of Marcobal (Marcobal et al. Critical Reviews in Food Science and Nutrition, 52(5), 448–467.).
Regarding claim 68, Chou and Lin 2018 teach all of the elements of claim 67.
Marcobal discloses “Tyramine poisoning is caused by the ingestion of food containing high levels of tyramine, a biogenic amine. The molecular knowledge of the genes involved in tyramine production has led to the development of molecular methods for the detection of bacteria able to produce tyramine and PEA.” (Abstract). Marcobal further specifies microbial contamination, “Decarboxylase-positive bacteria may constitute part of the associated population of the food or may be introduced by contamination”(page 449, 1st column, 2nd paragraph). Marcobal suggests using methods to detect microbial contamination, “These rapid and simple methods could be used for the analysis of the ability to form tyramine by bacteria in order to evaluate the potential risk of tyramine biosynthesis in food products” (Abstract).
It would have been obvious to a person having ordinary skill in the art (PHOSITA) at the time of filing of the instant application to use the cellular biosensor of Chou and Lin 2018 and apply Marcobal’s technique of PEA, tyramine, or tryptamine detection as a means to indicate microbial contamination because it is simply applying a known technique to a known device (method, or product) ready for improvement to yield predictable results.
A PHOSITA would recognize cellular sample-based biosensors of Chou and Lin 2018 would exhibit similar measurements as the methods of Marcobal. Furthermore, as shown by Marcobal, a PHOSITA would recognize that levels of PEA, tyramine, or tryptamine in food indicate microbial contamination.
Therefore a PHOSITA would have applied Marcobal’s PEA, tyramine, or tryptamine detection as a means to indicate microbial contamination toward the use of Chou and Lin’s 2018 cellular biosensor to detect the PEA, tyramine, or tryptamine and had a reasonable expectation of success of arriving at the instant invention.
Therefore, Chou, Lin 2018 and Marcobal teach all of the elements of claim 68.
Claim(s) 69 and 78, is/are rejected under 35 U.S.C. 103 as being unpatentable over Chou as applied to claim 1 above, and further in view of Ali (Ali AA, Altemimi et al. Biosensors (Basel). 2020 May 30;10(6):58.).
Regarding claims 69 and 78, Chou teaches all of the elements of claim 1.
Chou does not teach the sensor dynamically identifies microbial contamination in consumable products, manages various debilitating neurological disorders, or normalizes dysregulated metabolites associated with metabolic disorders.
Ali broadly discloses biosensor applications for detection of pathogenic food bacteria in their section titled “Bioluminescence Methods for Detection of Food Contamination” (page 8, section 5). Ali discloses the use of fluorescence and bioluminescence to “Bioluminescence plays an important role in real-time process monitoring due to the emission of bright light by living microorganisms.” (page 9, paragraph 1).
It would have been obvious for a person having ordinary skill in the art (PHOSITA) to use the biosensor taught by Chou to dynamically detect food contamination according to Ali’s bioluminescent biosensors because it is simply combining prior art elements according to known methods to yield a predictable result.
A PHOSITA in the field of biosensors would look for multiple applications. This would lead a PHOSITA, beginning at Chou’s biosensor, to find the teachings of Ali’s bioluminescent food contaminant biosensor. The PHOSITA would recognize the opportunity of the application of Chou’s biosensor toward detection of microbial contamination. Therefore a PHOSITA would have applied Chou’s biosensor toward the application of Ali’s bioluminescent food contaminant biosensor and had a reasonable expectation of success of arriving at the instant invention.
Therefore, Chou and Ali teach the embodiment “the sensor dynamically identifies microbial contamination in consumable products” (claim 69) and the embodiment “monitoring food quality” (claim 78) and therefore, Chou and Ali teach all of the elements of claim 69 and 78.
Claim(s) 72, is/are rejected under 35 U.S.C. 103 as being unpatentable over Chou and further in view of Lin (Lin et al. Anal Chim Acta. 2018 Dec 24;1041:108-113.) as applied to claim 70 above, and further evidenced by Mitchell (Mitchell et al. N Engl J Med. 1990 Feb 15;322(7):432-7.).
Regarding claim 72, Chou and Lin 2018 teach all of the elements of claim 70.
Chou and Lin do not teach wherein the aromatic compound [tyrosine] are neurochemicals.
Mitchell teaches tyrosine is a neurochemical. Mitchell discloses “Hereditary tyrosinemia results from an inborn error in the final step of tyrosine metabolism… Neurologic manifestations have been reported but not emphasized as a common problem”(Abstract).
Therefore, Chou, and Lin teach all of the elements of claim 72.
Claim(s) 73, is/are rejected under 35 U.S.C. 103 as being unpatentable over Chou and further in view of Lin (Lin et al. Anal Chim Acta. 2018 Dec 24;1041:108-113.), as evidenced by Mitchell (Mitchell et al. N Engl J Med. 1990 Feb 15;322(7):432-7.) as applied to claim 72 above, and further evidenced by Bloemendaal (Bloemendaal et al. eNeuro. 2018 Apr 30;5(2):ENEURO.0035-17.2018.).
Regarding claim 73, Chou and Lin 2018 teach all of the elements of claim 72.
Chou and Lin do not teach wherein the neurochemicals are selected from aromatic neurotransmitters or neuromodulators.
Bloemendaal teaches tyrosine is a neuromodulators. Bloemendaal discloses “It is unclear whether tyrosine-induced modulation of catecholaminergic signaling in older adults will affect reactive and/or proactive response inhibition.”(page 2, 2nd column, 1st paragraph).
Therefore, Chou, and Lin teach all of the elements of claim 74.
Allowable Subject Matter
Claim 55 objected to as being dependent upon a rejected base claim, but would be allowable if rewritten in independent form including all of the limitations of the base claim and any intervening claims.
Regarding claim 49, this examiner has been unable to identify the claimed “wherein the O1 variant demonstrates about 5-fold and about 7-fold repression in response to Trp and 5-HTP, respectively” in the prior art.
Regarding claim 55, this examiner has been unable to identify the claimed “polypeptide having TyrR activity” that selectively binds tyramine in the prior art.
Conclusion
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/AARON DUREL WARD/Examiner, Art Unit 1636
/NEIL P HAMMELL/Supervisory Patent Examiner, Art Unit 1636
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