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 .
The amendment filed on 04/13/2026 has been entered.
Claims 1-6 were amended in the claim set filed on 03/20/2026 and presented in claim set filed on 04/13/2026.
Applicant’s election without traverse of Group I, claims 1-8, drawn to a cell detection system, in the reply filed on 12/08/2025 is acknowledged.
Claims 9-16 are withdrawn drawn to a nonelected Group II.
Claims 17-18 were added in the claim set filed on 04/13/2026. No new matter was added.
Claims 1-8 and 17-18 in the claim set filed on 04/13/2026 are currently under examination.
Response to the Arguments
Objections to the Drawings and Specification in the previously mailed non-final have been withdrawn in light of applicants Drawings amendments and Specification clarification.
As necessitated by amendment, the 35 U.S.C. 103 rejections of claim(s) 1-8 documented in the previously mailed non-final have been maintained and revised in light of applicants claim amendments and arguments. Applicants’ argument on Pg. 10, states that “Claim 1 and claims that depend therefrom are not obvious over the combination of cited references, Soderberg and Li, because both fails to teach or contemplate use of a single transmembrane nanosensor with a quenched fluorophore located in the cytosol and a masked HCR initiator domain located extracellularly, wherein the transmembrane nanosensor, upon binding to the intracellular targets, undergoes a conformational transition fluorescing the fluorophore located in the cytosol and unmasking of the HCR initiator to trigger the HCR reaction.” Revised rejections are made as documented below in the 35 U.S.C. 103 rejection in this office action on Pg. 3-10.
As necessitated by amendment, new grounds of rejection are made for claims 17-18 under U.S.C. 103 rejection in this office action on Pg. 10-12.
As necessitated by amendment, the Nonstatutory Double Patenting rejections of claim(s) 1 documented in the previously mailed non-final has been maintained and revised in light of applicants claim amendments and arguments. Revised rejections are made as documented below in the Nonstatutory Double Patenting rejections rejection in this office action on Pg. 13-15.
The revised and new grounds of rejections for claims 1-8 and 17-18 are documented below in this Final Office Action are necessitated by claim amendments filed on 04/13/2026.
Priority
This application is a continuation-in-part of International Application No. PCT/US2021/054861 with international filing date October 13, 2021. This application claims the benefit of International Application No. PCT/US2021/054861 and U.S. Application No. 63/091,113, filed on October 13, 2020, the content of each of which is incorporated herein by reference in its entirety.
Claim Rejections - 35 USC § 103
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
Claims 1-8 are rejected under 35 U.S.C. 103 as being unpatentable over Söderberg et al. (“Söderberg”; Patent App. Pub. EP 3102698 B1, Jan. 16, 2019) in view of Li et al. (“Li” :(2020). Lipid-oligonucleotide conjugates for bioapplications. National science review, 7(12), 1933–1953.).
Söderberg discloses “The present invention relates to a proximity-probe based detection assay ("proximity assay"), for an analyte in a sample. The analyte may be any molecule it is desired to detect, including proteins and nucleic acids, in any form or in any type of sample, e.g. whether in situ or in isolated form. The analyte is detected by the binding of at least two probes, which may bind to the analyte directly or indirectly and which, when brought into proximity by binding to the analyte, interact together to allow a signal to be generated. In the new method of the invention the signal is generated by a hybridization chain reaction (HCR), which is triggered when the probes interact upon analyte binding. Also provided are kits and reagents for performing such an assay.” (Abstract).
Regarding claim 1, Söderberg teaches a system comprising “the invention advantageously combines analyte detection by a proximity assay with signal generation by hybridization chain reaction (HCR) to provide a method which can be used to detect … analytes, selectively, or specifically, and sensitively, with strong signal amplification but without reliance on enzymes.” (Para. 10). Söderberg teaches a system comprising “proximity-probe based assays in which the nucleic acid domains of respective proximity probes interact to form a hairpin structure when the probes are bound in proximity to a target.” (Para. 8). Söderberg teaches a system comprising “the HCR monomers may be labelled, akin to conformationally selective probes such as molecular beacons, such that the signal (e.g. fluorescence) is quenched when the HCR monomers are in monomer form, but detectable when the monomers have been unfolded, i.e. when hybridized in the polymer. Thus, for example the HCR monomer may be labelled at one end of a stem with a quencher molecule and with a fluorophore at the opposing side of the duplex region, such that when the stem structure is present in the monomer the fluorescent signal is quenched. Upon polymerization the fluorophore and quencher are spatially separated in the polymer, and quenching is relieved, allowing a fluorescent signal to be detected.” (Para. 99). “Molecular beacons” reads on “a loop comprising a polynucleotide sequence which is complementary to the target. Söderberg teaches a system comprising “Typically the region of secondary structure will contain a loop of single stranded nucleic acid, more particularly a stem-loop or hairpin structure comprising a double-stranded "stem" region and a single stranded loop.” (Para. 21). Söderberg teaches a system comprising “This HCR initiator region is "protected" or shielded from being able to hybridize to the HCR monomer and thereby initiate an HCR reaction by being contained in a metastable secondary structure (more particularly a double stranded metastable secondary structure, e.g. a stem-loop structure akin to that of a HCR monomer).” (Para. 15).
Söderberg teaches a system comprising “In the simplest form of HCR, two different stable hairpin monomers undergo a chain reaction of hybridization events to form a long nicked double-stranded DNA molecule when an "initiator" nucleic acid molecule is introduced. In the absence of the initiator the hairpin monomers are stable, or kinetically trapped ("metastable"), and remain as monomers (i.e. preventing the system from rapidly equilibrating). However, once introduced, the initiator is able to hybridize to the first hairpin monomer, and invades it, causing it to open up and hybridize to and invade the second hairpin monomer, in turn opening this up and allowing it to hybridize to and invade another molecule of the first monomer, and so on, until the monomers are exhausted… The hybridization and invasion by the initiator thus triggers the HCR. … when a triggered conformational change allows the single stranded bases in the loops to hybridize with a complementary strand.” (Para.12; Para. 18).
Furthermore, Söderberg teaches a system comprising “Advantageously, the nucleic acid may be detected in situ” (Para. 54). Söderberg teaches a system comprising “read-out of the internalization of AlexaFluor488-EGF/EGFR complex in A431 cells after stimulation. The prox-HCR assay was used to detect AlexaFluor488-EGF/EGFR at the cell the membrane” (Para. 139). Thus, one of skill in the art would be motivated to monitor localized "in situ" detection of analytes in the cytosol.
Thus, Söderberg suggests a cell detection system comprising: (i) membrane associated nanosensor comprising a lipid-conjugated DNA comprising a first hairpin stem-loop comprising: a loop comprising a polynucleotide sequence which is complementary to the target polynucleotide, a stem comprising complementary 5' and 3' domains, a fluorophore, a quencher paired to the fluorophore, and a masked hybridization chain reaction (HCR) initiator domain; and (ii) components for HCR comprising: a set of metastable hairpins configured for HCR, wherein at least one of the metastable hairpins hybridizes to the HCR initiator domain of the nanosensor and initiates an HCR reaction when the HCR initiator is unmasked; wherein upon binding of the first hairpin stem-loop binding to the target polynucleotide in the cytosol of the cell, the nanosensor transitions from a closed conformation to an open conformation unmasking the HCR initiator domain and allowing for fluorescence the fluorophore without quenching thereby detecting the target polynucleotide in the cell.
Söderberg does not explicitly teach the limitations comprising “a transmembrane nanosensor comprising a lipid-conjugated DNA”.
Li discloses “Lipid–oligonucleotide conjugates (LONs) are powerful molecular-engineering materials for various applications ranging from biosensors to biomedicine. Their unique amphiphilic structures enable the self-assembly and the conveyance of information with high fidelity. In particular, LONs present remarkable potential in measuring cellular mechanical forces and monitoring cell behaviors. LONs are also essential sensing tools for intracellular imaging and have been employed in developing cell-surface-anchored DNA nanostructures for biomimetic-engineering studies. When incorporating therapeutic oligonucleotides or small-molecule drugs, LONs hold promise for targeted therapy. Moreover, LONs mediate the controllable assembly and fusion of vesicles based on DNA-strand displacements, contributing to nanoreactor construction and macromolecule delivery. In this review, we will summarize the general synthesis strategies of LONs, provide some characterization analysis and emphasize recent advances in bioanalytical and biomedical applications. We will also consider the relevant challenges and suggest future directions for building better functional LONs in nanotechnology and materials-science applications” (Abstract)
Regarding claim 1, Li teaches a system comprising “We recently developed varied LON structures in which the lipid moiety allows facile and efficient engineering of DNA-based sensors onto the cell phospholipid bilayer” (Pg. 1939, Col 1, Para. 2). LON reads on Lipid–oligonucleotide conjugate. Thus, Söderberg and Li suggest a system comprising a transmembrane nanosensor comprising a lipid conjugated DNA.
Söderberg and Li are both considered to be analogous to the claimed invention because they are in the same field of sensor detection systems. Therefore, it would have been obvious to someone of ordinary skill in the art before the effective filing date of the claimed invention to have modified the detection system comprising hairpin stem loop and components for HCR as taught by Söderberg to incorporate the transmembrane detection system comprising a lipid conjugated DNA as taught by Li and provide transmembrane nanosensor and components for HCR according to the limitations according to claim 1. These claim elements were known in the art and one of skill in the art could have combined these elements by known methods with no change in their respective functions, and the combination would have yielded the predictable outcome according to the limitations of claim 1. Furthermore, the teachings of Söderberg would have motivated one of ordinary skill in the art to monitor localized "in situ" detection of analytes through membrane associated sensors including the cytosol. Doing so would allow for the molecular sensors to be anchored onto the cell membrane, resulting in the performance of live-cell studies with high spatio-temporal resolution (Pg. 1939, Col 1, Para. 2).
The teachings of Söderberg and Li are documented above in the rejection of claim 1 under 35 U.S.C. 103. Claims 2,4-7 depends on claim 1. Claim 3 depends on claim 2, which depends on claim 1. Claim 8 depend on claim 7, which depends on claim 1.
Regarding claims 2-3, Li depicts a system wherein the LON spans the lipid bilayer and the lipid bilayer is an cellular outer membrane (See Figure 3B below). Thus, Söderberg and Li suggest a system wherein the lipid-conjugated DNA spans a lipid bilayer; and wherein the lipid bilayer is a cellular outer membrane or episome.
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Regarding claims 4-5, Söderberg teaches a system wherein “the analyte may be DNA (e.g. genomic, mitochondrial) or RNA (e.g. messenger RNA, ribosomal RNA, microRNA etc.).” (Para. 54). Thus, Söderberg and Li suggest a system wherein the target polynucleotide is an RNA or a DNA; and wherein the target polynucleotide is a messenger RNA (mRNA) or microRNA (miRNA).
Regarding claim 6, Li teaches a system wherein “DNA tweezers with a FRET pair as the signal reporter and cholesterol as the anchoring unit on the cell surface. Li also teaches a system wherein “. A cholesterol tail was attached to an aptamer probe and the two ends of the hairpin structure were separately labeled with a fluorophore and a quencher for recognition and sensing” (Pg. 1940, Col. 1, Para. 1; Fig 3B). Thus, Söderberg and Li suggest a system wherein the lipid comprises a cholesterol molecule.
Regarding claims 7-8, Söderberg teaches a system comprising “HCR monomer may be labelled at one end of a stem with a quencher molecule and with a fluorophore at the opposing side of the duplex region, such that when the stem structure is present in the monomer the fluorescent signal is quenched. Upon polymerization the fluorophore and quencher are spatially separated in the polymer, and quenching is relieved, allowing a fluorescent signal to be detected. (Para. 99). Thus, Söderberg and Li suggest a system wherein the metastable hairpins configured for HCR comprise a detectable label; and wherein the metastable hairpins configured for HCR comprises a quencher.
Response to Arguments
Applicant's arguments filed 04/13/2026 (Pg. 7-10) with respect to claims 1-8 have been fully considered but they are not persuasive. To clarify some instances argued in the response filed 04/13/2026 see responses to each argument made by Applicant below:
Applicants’ argument: “Claim 1 and claims that depend therefrom are not obvious over the combination of cited references, Soderberg and Li, because both fails to teach or contemplate use of a single transmembrane nanosensor with a quenched fluorophore located in the cytosol and a masked HCR initiator domain located extracellularly, wherein the transmembrane nanosensor, upon binding to the intracellular targets, undergoes a conformational transition fluorescing the fluorophore located in the cytosol and unmasking of the HCR initiator to trigger the HCR reaction” (Pg. 10)
Response: Applicant’s arguments have been fully considered and found unpersuasive because in response to applicant's arguments against the references individually, one cannot show nonobviousness by attacking references individually where the rejections are based on combinations of references. See In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981); In re Merck & Co., 800 F.2d 1091, 231 USPQ 375 (Fed. Cir. 1986).
Applicants’ argument: “a person of ordinary skill in the art would not have been motivated to modify Soderberg in view of Li to arrive at the claimed invention because doing so would require a substantial reconstruction and redesign of the elements shown in Soderberg' s proximity probes as well as a change in the basic principle under which the Soderberg' s proximity probes were designed to operate.” (Pg. 10)
Response: Applicant’s arguments have been fully considered and found unpersuasive because as stated above in the Final office action, Söderberg teaches a system comprising “Advantageously, the nucleic acid may be detected in situ” (Para. 54). Söderberg teaches a system comprising “read-out of the internalization of AlexaFluor488-EGF/EGFR complex in A431 cells after stimulation. The prox-HCR assay was used to detect AlexaFluor488-EGF/EGFR at the cell the membrane” (Para. 139). Thus, one of skill in the art would be motivated to monitor localized "in situ" detection of analytes in the cytosol. Furthermore, the teachings of Söderberg would have motivated one of ordinary skill in the art to monitor localized "in situ" detection of analytes through membrane associated sensors including the cytosol.
Claims 17-18 are rejected under 35 U.S.C. 103 as being unpatentable over Söderberg et al. (“Söderberg”; Patent App. Pub. EP 3102698 B1, Jan. 16, 2019) in view of Li et al. (“Li” :(2020). Lipid-oligonucleotide conjugates for bioapplications. National science review, 7(12), 1933–1953.) and Zhou et al. (“Zhou”; (2012). Reversible regulation of protein binding affinity by a DNA machine. Journal of the American Chemical Society, 134(3), 1416-1418, as cited in the IDS.).
The teachings of Söderberg and Li are documented above in the rejection of claims 1-8 under 35 U.S.C. 103. Claims 17-18 depend on claim 1. However, the teaching of Söderberg and Li do not explicitly teach the limitations of claims 17 and 18.
Zhou discloses “We report a DNA machine that can reversibly regulate target binding affinity on the basis of distance-dependent bivalent binding. It is a tweezer-like DNA machine that can tune the spatial distance between two ligands to construct or destroy the bivalent binding. The DNA machine can strongly bind to the target protein when the ligands are placed at an appropriate distance but releases the target when the bivalent binding is disrupted by enlargement of the distance between the ligands. This “capture–release” cycle could be repeatedly driven by single-stranded DNA without changing the ligands and target protein.” (Abstract)
Regarding claim 17, Zhou depicts a system in Figure 1 wherein the first hairpin stem-loop is formed by a hairpin with a single-stranded loop and a first arm and a second arm linked by a crossover hinge, and wherein the distance between the tip of the first arm and the tip of the second arm is reversibly or irreversibly controlled by binding and release of the target polynucleotide to the single-stranded loop of the hairpin. (Figure 1, shown below) .
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Thus, Söderberg, Li and Zhou suggest a system wherein the first hairpin stem-loop is formed by a hairpin with a single-stranded loop and a first arm and a second arm linked by a crossover hinge, and wherein the distance between the tip of the first arm and the tip of the second arm is reversibly or irreversibly controlled by binding and release of the target polynucleotide to the single-stranded loop of the hairpin.
Regarding claim 18, Zhou depicts a system on the right in Figure 1 wherein the first arm and second arm are double-crossover tile arms. (Figure 1, shown above). Thus, Söderberg, Li and Zhou suggest a system wherein the first arm and second arm are double-crossover tile arms.
Söderberg, Li and Zhou are considered to be analogous to the claimed invention because they are in the same field of sensor detection systems. Therefore, it would have been obvious to someone of ordinary skill in the art before the effective filing date of the claimed invention to have modified the detection system suggested by Söderberg and Li to incorporate wherein the first hairpin stem-loop is formed by a hairpin with a single-stranded loop and a first arm and a second arm linked by a crossover hinge, and wherein the distance between the tip of the first arm and the tip of the second arm is reversibly or irreversibly controlled by binding and release of the target polynucleotide to the single-stranded loop of the hairpin and wherein first arm and second arm are double-crossover tile arms as taught by Zhou and provide transmembrane nanosensor and components for HCR according to the limitations according to claims 17 and 18. These claim elements were known in the art and one of skill in the art could have combined these elements by known methods with no change in their respective functions, and the combination would have yielded the predictable outcome according to the limitations of claims 17-18. Doing so would allow for the molecular sensors to be anchored onto the cell membrane, resulting in the performance of live-cell studies with high spatio-temporal resolution (Pg. 1939, Col 1, Para. 2).
Response to Arguments
Applicant's arguments filed 04/13/2026have been fully considered but they do not apply to the new grounds of rejection. Arguments against Söderberg, Li and Zhou on Pg. 7-10 are not persuasive as discussed above.
Double Patenting
The nonstatutory double patenting rejection is based on a judicially created doctrine grounded in public policy (a policy reflected in the statute) so as to prevent the unjustified or improper timewise extension of the “right to exclude” granted by a patent and to prevent possible harassment by multiple assignees. A nonstatutory double patenting rejection is appropriate where the conflicting claims are not identical, but at least one examined application claim is not patentably distinct from the reference claim(s) because the examined application claim is either anticipated by, or would have been obvious over, the reference claim(s). See, e.g., In re Berg, 140 F.3d 1428, 46 USPQ2d 1226 (Fed. Cir. 1998); In re Goodman, 11 F.3d 1046, 29 USPQ2d 2010 (Fed. Cir. 1993); In re Longi, 759 F.2d 887, 225 USPQ 645 (Fed. Cir. 1985); In re Van Ornum, 686 F.2d 937, 214 USPQ 761 (CCPA 1982); In re Vogel, 422 F.2d 438, 164 USPQ 619 (CCPA 1970); In re Thorington, 418 F.2d 528, 163 USPQ 644 (CCPA 1969).
A timely filed terminal disclaimer in compliance with 37 CFR 1.321(c) or 1.321(d) may be used to overcome an actual or provisional rejection based on nonstatutory double patenting provided the reference application or patent either is shown to be commonly owned with the examined application, or claims an invention made as a result of activities undertaken within the scope of a joint research agreement. See MPEP § 717.02 for applications subject to examination under the first inventor to file provisions of the AIA as explained in MPEP § 2159. See MPEP § 2146 et seq. for applications not subject to examination under the first inventor to file provisions of the AIA . A terminal disclaimer must be signed in compliance with 37 CFR 1.321(b).
The filing of a terminal disclaimer by itself is not a complete reply to a nonstatutory double patenting (NSDP) rejection. A complete reply requires that the terminal disclaimer be accompanied by a reply requesting reconsideration of the prior Office action. Even where the NSDP rejection is provisional the reply must be complete. See MPEP § 804, subsection I.B.1. For a reply to a non-final Office action, see 37 CFR 1.111(a). For a reply to final Office action, see 37 CFR 1.113(c). A request for reconsideration while not provided for in 37 CFR 1.113(c) may be filed after final for consideration. See MPEP §§ 706.07(e) and 714.13.
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Claim 1 is provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claim 1 of copending Application No. 18/298922 (reference application), filed 04/11/2023 in view of Söderberg et al. (“Söderberg”; Patent App. Pub. EP 3102698 B1, Jan. 16, 2019) and Li et al. (“Li” :(2020). Lipid-oligonucleotide conjugates for bioapplications. National science review, 7(12), 1933–1953.). Although the claims at issue are not identical, they are not patentably distinct from each other because the instantly claimed invention is made obvious over the claims of copending Application No. 18/298922 in view of Söderberg and Li.
The claims of copending Application No. 18/298922 are drawn to:
“1. A transmembrane nanosensor device comprising a lipid conjugated DNA tweezer comprising a hairpin loop complementary to a target polynucleotide trigger strand; a fluorophore and a quencher paired to the fluorophore, or a FRET pair; wherein when the hairpin loop is bound by the target polynucleotide trigger strand, the DNA tweezer transitions from a closed conformation to an open conformation, the quencher is separated from the fluorophore, and the fluorophore fluoresces.”
The teachings of Söderberg and Li are documented above in the rejection of claims 1-8 under 35 U.S.C. 103.
Therefore, the invention as recited in claims 1 is prima facie obvious over the copending Application No. 18/298922 in view of Söderberg and Li. One of ordinary skill in the art would have had a reasonable expectation of success given the obviousness of the claims in view of Söderberg and Li. It would have been obvious to use a detection system comprising a transmembrane nanosensor and HCR components according to the limitations recited in claim 1 of the instant application based on claim 1 of copending Application No. 18/298922 (Patent App. No. US 20230374607 A1, filed 04/11/2023) in view of Söderberg and Li.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Response to Arguments
Applicant's arguments filed 04/13/2026have been fully considered but are not persuasive. Arguments against Söderberg, Li and Zhou on Pg. 7-10 are not persuasive as discussed above.
Conclusion
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
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/KENDRA R VANN-OJUEKAIYE/Examiner, Art Unit 1682
/WU CHENG W SHEN/Supervisory Patent Examiner, Art Unit 1682
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