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 .
Continued Examination Under 37 CFR 1.114
A request for continued examination under 37 CFR 1.114, including the fee set forth in 37 CFR 1.17(e), was filed in this application after final rejection. Since this application is eligible for continued examination under 37 CFR 1.114, and the fee set forth in 37 CFR 1.17(e) has been timely paid, the finality of the previous Office action has been withdrawn pursuant to 37 CFR 1.114. Applicant's submission filed on 04/09/2026 has been entered.
Priority
Applicant’s claim for the benefit of a prior-filed application under 35 U.S.C. 119(e) or
under 35 U.S.C. 120, 121, 365(c), or 386(c) is acknowledged. This application claims benefit of
the U.S. application 63/130,545 filed 12/24/2020. Based on the filing receipt, the
effective filing date of this application is December 24, 2020 which is the filing date of U.S. application 63/130,545 from which the benefit of priority is claimed.
Information Disclosure Statements
The information disclosure statements (IDS) filed 09/03/2025, 09/11/2025, and 03/18/2026 have been considered by the examiner.
Election/Restrictions
The species election requirement of cargo molecules, dated 03/10/2025, has been withdrawn. Upon reconsideration, the species do not impart a serious search and/or examination burden.
Status of Claims
Claims 5-7, 9-10, 12-17, 20-24, 27-29, 31, 34-35, 37, 41-45, 47-59, 63-75, 78, 83-94, 96, 98-99, 101-107, 109-114, 116-126, 128-135, and 139 are cancelled.
Claims 1-4, 8, 11, 18-19, 25-26, 30, 32-33, 36, 38-40, 46, 60-62, 76-77, 79-82, 95, 97, 100, 108, 115, 127, 136-138, and 140 are pending.
Claims 33, 36, 38-40, 46, 60-62, 76-77, 79-82, 95 are withdrawn due to the claims being drawn to a nonelected invention in the restriction and species elections filed 04/30/2025.
Claims 1-4, 8, 11, 18-19, 25-26, 30, 32, 97, 100, 108, 115, 127, 136-138, and 140 are examined herein.
Withdrawn Objections/Rejections
The rejection of claims 10 and 139 as rejected under 35 U.S.C. 102(a)(1) has been withdrawn, due to the cancellation of the claims in amendments filed 04/09/2026.
Claim Objections
Claim 25 is objected to because of the following informalities: the claim recites, “wherein releasing the isolated biological particle step (d) comprises”. However, the claim should recite, “wherein releasing the isolated biological particle in step (d) comprises”. Appropriate correction is required.
Claim Interpretation
Claim 1 recites, “wherein the method: (i) minimizes nonspecific binding of the biological particles to the low non-specific small RNA binding solid support without the binding agent; (ii) minimizes background binding of small RNAs to the low non-specific small RNA binding solid support; and (iii) detects low abundance small-RNA cargos”. The claim limitations are functional limitations without structure. Furthermore, the claim limitations are indefinite due to the use of relative terms, as discussed below. However, para. [0323] of the applicant’s specification discloses that magnetic beads have the capacity to bind both small-RNAs and exosomes non-specifically, which contributes to non-specific RNA background. Therefore, the use of solid supports other than magnetic beads are interpreted as meeting the limitations above.
Claim 3 recites, “The method of claim 1, further comprising an initial ultrafiltration or ultracentrifugation step to provide a pooled heterogeneous population of biological particles and to increase specificity of exosome capture”. The initial ultrafiltration or ultracentrifugation step used with the method of claim 1 will inherently lead to a pooled heterogeneous population of biological particles and increased specificity of exosome capture. Therefore, any initial ultrafiltration or ultracentrifugation step with the steps of claim 1 will meet the limitations of claim 3.
Claim 97 recites, “wherein the method: (i) minimizes nonspecific binding to the low non-specific small RNA binding solid support without the binding agent; and (ii) minimizes background binding of small RNAs to the low non-specific small RNA binding solid support”. The claim limitations are functional limitations without structure. Furthermore, the claim limitations are indefinite due to the use of relative terms, as discussed below. However, para. [0323] of the applicant’s specification discloses that magnetic beads have the capacity to bind both small-RNAs and exosomes non-specifically, which contributes to non-specific RNA background. Therefore, the use of solid supports other than magnetic beads are interpreted as meeting the limitations above.
New Rejections
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.
Claims 1, 97, and 140 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.
The term “low” in claims 1 and 97 is a relative term which renders the claims indefinite. The term “low” 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. The term “low” describes the non-specific small-RNA binding solid support. However, it is unclear what degree of non-specific small-RNA binding is “low”. Therefore, the metes and bounds of the claims cannot be ascertained.
The term “minimizes” in claims 1 and 97 is a relative term which renders the claims indefinite. The term “minimizes” 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. The term “minimizes” describes the background binding of small RNAs to the solid support. However, it is unclear what degree of small-RNA background binding constitutes minimized binding. Therefore, the metes and bounds of the claims cannot be ascertained.
The term “low abundance” in claim 1 is a relative term which renders the claim indefinite. The term “low abundance” 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. The term “low abundance” describes the small-RNA cargos. However, it is unclear what degree of small-RNA cargo abundance is “low”. Therefore, the metes and bounds of the claims cannot be ascertained.
The term “about” in claim 140 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. The term “about” describes the diameter of the extracellular vesicles of claim 140. The specification does not provide any indication as to what range of specific diameters is covered by the term "about". Therefore, the metes and bounds of the claims cannot be ascertained.
Modified Rejections
Claim Rejections - 35 USC § 102
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, 8, 25-26, 30, 32, 97, 100, 127, 136, 138, and 140 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Routenberg (WO 2020/086751 A1, published 2020-04-30, cited in PTO-892 dated 2026-01-14). The following rejections have been modified, necessitated by amendments filed 04/09/2026.
With respect to claim 1, Routenberg teaches a method of preparing a purified population of biological particles from a biological sample and for evaluating a cargo of the purified population of biological particles (see, e.g., method - sheet 9/72, under “FIG. 7”; biological sample – para. [0098]; evaluating the purified population of biological particles – para. [0302] and sheet 9/72, under “FIG. 7”). Routenberg teaches (a) obtaining a biological sample comprising biological particles, such as extracellular vesicles (EVs) (see, e.g., obtaining biological sample comprising biological particles - para. [0102]-[0103], and sheet 9/72, under “FIG. 7”). Routenberg teaches (b) contacting the biological sample comprising the biological particles from the subject with a binding agent directed to a biological particle surface antigen, wherein one or more biological particle surface antigens comprises CD63, wherein the binding agent is immobilized directly or indirectly on a low non-specific small-RNA binding solid support, wherein, when the binding agent is immobilized indirectly on the low non-specific small-RNA binding solid support, it is coupled to a nucleic acid, and the nucleic acid is immobilized on the low non-specific small RNA binding solid support; and wherein the low non-specific small RNA binding solid support is a well plate (see, e.g., contacting particles with a binding agent - sheet 9/72, under “FIG. 7”; particle surface antigen comprises CD63 – para. [00110]; wherein the binding agent is immobilized directly or indirectly on a low non-specific small-RNA binding solid support, wherein, when the binding agent is immobilized indirectly on the low non-specific small-RNA binding solid support, it is coupled to a nucleic acid, and the nucleic acid is immobilized on the low non-specific small RNA binding solid support - sheet 9/72, under “FIG. 7”, and para. [00322]; wherein the low non-specific small RNA binding solid support is a well plate – para. [0087], and para. [00322], and sheet 9/72, under “FIG. 7). In “FIG. 7” of Routenberg, the capture antibodies (equivalent to binding agents) are attached to dsDNA linkers (equivalent to a nucleic acid coupled to the binding agent for indirectly immobilizing the binding agent to the solid support) which tether the capture antibodies to a solid support, such as a plate. Routenberg teaches wherein the method: (i) minimizes nonspecific binding of the biological particles to the low non-specific small RNA binding solid support without the binding agent; (ii) minimizes background binding of small RNAs to the low non-specific small RNA binding solid support; and (iii) detects low abundance small-RNA cargos (see, e.g., para. [00419], and para. [00312], and para. [00322]). It is understood that the solid phase, specifically the plate, of “FIG. 7” is equivalent a well plate. It is also understood that the method: (i) minimizes nonspecific binding of the biological particles to the low non-specific small RNA binding solid support without the binding agent; (ii) minimizes background binding of small RNAs to the low non-specific small RNA binding solid support; and (iii) detects low abundance small-RNA cargo is taught by the streptavidin-coated well plates of Routenberg because the applicant’s specification discloses that streptavidin-coated well plates are a low non-specific nucleic acid binding platform. See para. [0323] of the applicant’s specification. Routenberg teaches (c) isolating the biological particle bound by the binding agent from the biological sample by washing (see, e.g., para. [0089], and sheet 9/72, under “FIG. 7”, and para. [00563], and para. [00143]: “In embodiments, the unwanted components are soluble in the sample and/or the washing fluid”, and para. [00232], and para. [00375]). Routenberg teaches (d) releasing, the biological particle from the low non-specific small RNA binding solid support, bound to the binding agent (see, e.g., sheet 9/72, under “FIG. 7”, under step “2.”: “Cleave linker or denature dsDNA to elute EVs”). Routenberg teaches (e) eluting the bound biological particle from the binding agent to form a population of free purified biological particles (see, e.g., sheet 9/72, under “FIG. 7”, under step “3a.”: “elute antibodies from EV surface using low-pH”). Routenberg teaches evaluating surface molecules, specifically protein CD63 of the purified population of biological particles, further comprising extracting RNA from the purified population of biological particles and identifying the quantifying expression of small non-coding RNAs derived from the cell of origin comprising microRNAs encapsulated by the purified population of biological particles, wherein the isolated biological particles are derived from a subject (see, e.g., CD63 - para. [00161]; subjects - para. [00152]; miRNA – para. [0312]).
With respect to claim 2, Routenberg teaches step (f) evaluating cargo, further comprising identifying protein and/or lipid cargos by mass spectrometry (see, e.g., para. [0071]-[0072], and para. [0089]).
With respect to claim 3, Routenberg teaches an initial ultrafiltration step to provide a pooled heterogeneous population of biological particles and to increase specificity of exosome capture (see, e.g., para. [00143]). It is noted that the benefits of the ultrafiltration step follow naturally from the step itself.
With respect to claim 4, Routenberg teaches the biological sample comprises a body fluid comprising plasma (see, e.g., para. [00148]).
With respect to claim 8, Routenberg teaches the binding agent is an antibody (see, e.g., sheet 9/72, under “FIG. 7”).
With respect to claim 25, Routenberg teaches releasing the isolated biological particle comprises enzymatically cleaving the nucleic acid bound to the binding agent (see, e.g., para. [00258], and sheet 9/72, under “FIG. 7”, under step “2.”: “Cleave linker or denature dsDNA to elute EVs”). It is understood that the capture reagent is equivalent to the binding agent.
With respect to claim 26, Routenberg teaches the enzymatically cleaving is with an endonuclease (see, e.g., para. [00258]: “the labile linker […] is an oligonucleotide comprising a restriction site cleavable by a restriction endonuclease”).
With respect to claim 30, Routenberg teaches identifying the one or more small non-coding RNAs comprising miRNAs encapsulated in the one or more biological particles by next generation sequencing (see, e.g., para. [00291], and para. [00312]).
With respect to claim 32, Routenberg teaches the disease is cancer (see, e.g., para. [0028], and para. [0098], and para. [00111], and para. [00128], and para. [00149]).
With respect to claim 97, Routenberg teaches a method of preparing a purified population of cells from a biological sample and for evaluating the purified population of biological particles (see, e.g., cells – para. [0070]: “Examples of surface marker displaying agents include cells […] Although the present specification may refer to EVs in certain embodiments, the disclosure contemplates that such aspects also apply to any surface marker displaying agent provided herein without limitation”; biological sample – para. [0098]; evaluating the purified population of biological particles – para. [0302] and sheet 9/72, under “FIG. 7”). Note: Routenberg explicitly discloses that even when the specification refers to EVs, the aspects also apply to any surface marker displaying agent, such as cells. Routenberg teaches (a) obtaining a biological sample comprising cells (see, e.g., para. [0098]: “For example, the methods provided herein can be used to isolate abnormal cells, e.g., a cancer cell, from a sample of tissue or bodily fluid, for example, blood (e.g., comprising a mixture of cancer and non-cancer cells)”). Routenberg teaches (b) contacting the biological sample comprising cells from the subject with a binding agent directed to one or more cell surface antigens, wherein the one or more cell surface antigens comprises CD63; wherein the binding agent, specifically an antibody, is immobilized indirectly on a low non-specific small-RNA binding solid support, wherein the binding agent is immobilized indirectly on the low non-specific small-RNA binding solid support and coupled to a nucleic acid, and the nucleic acid is immobilized on a low non-specific small-RNA binding solid support, such as a well plate (see, e.g., contacting a biological sample with a binding agent directed to a cell surface antigen, specifically CD63 - para. [0089], para. [00126], and para. [0070]: “Examples of surface marker displaying agents include cells […] Although the present specification may refer to EVs in certain embodiments, the disclosure contemplates that such aspects also apply to any surface marker displaying agent provided herein without limitation”; wherein the binding agent, specifically an antibody, is linked to a nucleic acid, and wherein the nucleic acid is immobilized indirectly on a low non-specific small-RNA binding solid support and coupled to a nucleic acid – sheet 9/72, under “FIG. 7”). It is understood that the solid phase, specifically the plate, of “FIG. 7” is equivalent to a low non-specific small-RNA binding solid support. Routenberg teaches isolating the cells bound by the binding agent from the biological sample by washing (see, e.g., para. [0089], and sheet 9/72, under “FIG. 7”, and para. [00143]: “In embodiments, the unwanted components are soluble in the sample and/or the washing fluid”, and para. [00232], and para. [00375]). Routenberg teaches releasing, from the low non-specific small-RNA binding solid support, the cells bound to the binding agent (see, e.g., sheet 9/72, under “FIG. 7”, under step “2.”: “Cleave linker or denature dsDNA to elute EVs”). Routenberg teaches eluting the bound cells from the binding agent to form a population of purified cells, wherein the disease comprises cancer; and wherein the method: minimizes nonspecific binding to the low non-specific small-RNA binding solid support without the binding agent; and minimizes background binding of small RNAs to the low non-specific small-RNA binding solid support; and is tailorable for selecting specific populations of cells for analysis of their contents (see, e.g., eluting - sheet 9/72, under “FIG. 7”, under step “3a.”: “elute antibodies from EV surface using low-pH”; cells - para. [0070]: “Examples of surface marker displaying agents include cells […] Although the present specification may refer to EVs in certain embodiments, the disclosure contemplates that such aspects also apply to any surface marker displaying agent provided herein without limitation; cancer - para. [0028], and para. [0098], and para. [00111], and para. [00128], and para. [00149]; solid support - sheet 9/72, under “FIG. 7”, “solid phase”). It is understood that the method: minimizes nonspecific binding to the low non-specific small-RNA binding solid support without the binding agent; and minimizes background binding of small RNAs to the low non-specific small-RNA binding solid support; and is tailorable for selecting specific populations of cells for analysis of their contents because the solid support of a well plate meets this limitation as discussed in the claim interpretation section.
With respect to claim 100, Routenberg teaches the biological sample comprises a body fluid (see, e.g., para. [0098]: “For example, the methods provided herein can be used to isolate abnormal cells, e.g., a cancer cell, from a sample of tissue or bodily fluid, for example, blood”).
With respect to claim 127, Routenberg teaches the population of purified cells comprises eukaryotic cells, specifically human cells (see, e.g., para. [0098]).
With respect to claim 136, Routenberg teaches the population of purified cells is homogenous (see, e.g., para. [00147]: “purified mammalian fluid, secretion, or excretion”, and para. [00151]: “tissue homogenates”).
With respect to claim 138, Routenberg teaches the biological sample comprises a heterogeneous population of cells, and the surface antigens are cell surface antigens, and the binding agent is directed to one or more cell surface antigens (see, e.g., para. [0096], and para. [00148]).
With respect to claim 140, Routenberg teaches the extracellular vesicles actively secreted extracellular bilayered membrane-bound vesicles of endosomal origin in a size range of about 100 nm on average generated by cells (see, e.g., para. [0567]).
Claim Rejections - 35 USC § 103
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
Claims 11, 18-19, 108, and 115 are rejected under 35 U.S.C. 103 as being unpatentable over Routenberg (cited above), as applied to claims 1-4, 8, 25-26, 30, 32, 97, 100, 127, 136, 138, and 140 above, and further in view of Lof, et al. (“Detection of Extracellular Vesicles Using Proximity Ligation Assay with Flow Cytometry Readout—ExoPLA”, published 2017-07-05, cited in PTO-892 filed 06/17/2025) as evidenced by the product sheet for restriction enzyme RsaI from Thermo Scientific™ - Catalog number ER1121 (https://www.thermofisher.com/order/catalog/product/ER1121, cited in PTO-892 filed 06/17/2025).
Routenberg teaches as set forth above. The reference further teaches the nucleic acid comprises DNA, as in claim 11 (see, e.g., sheet 9/72, under “FIG. 7”). Routenberg further teaches the nucleic acid comprises a binding moiety, specifically biotin, on a first end and a different binding moiety, as in claims 18-19 and 115 (see, e.g., sheet 58/72, under “FIG. 49”, panel “A)”). It is understood that the anchor of “FIG. 49”, panel “A)” of Routenberg has biotin on one end and a different binding moiety, an antibody, on the other end.
Routenberg fails to teach the DNA comprises one or more ribonucleic acid nucleotides, wherein the ribonucleic acid nucleotide is uracil, and wherein the DNA comprises a restriction enzyme recognition site, as in claims 11 and 108.
However, Lof teaches a DNA-linker conjugated to a bead and a binding agent, specifically an antibody for a surface marker of extracellular vesicles (see, e.g., Lof, p. 4.8.2, under “Figure 4.8.1”). The DNA-linker of Lof is digested by an enzyme to release the extracellular vesicles (see, e.g., Lof, p. 4.8.2, under “Figure 4.8.1”). The DNA-linker comprises one or more ribonucleic acid nucleotides, specifically uracil, and wherein the DNA comprises a restriction enzyme recognition site, as in claims 11 and 108 (see, e.g., Lof, p. 4.8.4, under “Table 4.8.1”, under “Capturing antibody/ UNG digestion oligonucleotide 1”). The 5’ GTAC 3’ potion of the sequence comprises a restriction enzyme recognition site as evidenced by the product sheet for restriction enzyme RsaI from Thermo Scientific™ - Catalog number ER1121.
Routenberg and Lof are analogous to the field of the claimed invention because they are all in the field of biological particle analysis. One of ordinary skill in the art before the effective filing date of the application would have found it obvious to incorporate the DNA-linker of Lof into the methods of Routenberg. The artisan would have been motivated to do so because “detection and characterization of EVs is challenging due to their small size” (see, e.g., Lof, p. 4.8.1, under abstract). To overcome the challenge, Lof “established a method, called ExoPLA, that allows individual EVs to be detected and characterized at high specificity and sensitivity. Based on the in situ proximity ligation assay (in situ PLA), proximal oligonucleotide-conjugated antibodies bound to their targets on the surfaces of the EVs allow formation of circular products that can be fluorescently labeled by rolling circle amplification” (see, e.g., Lof, p. 4.8.1, under abstract). Further, the extracellular vesicle-complex is “released from the beads by digestion of the uracil-containing oligonucleotides by treatment with uracil N-glycosylase” (see, e.g., p. 4.8.2, under the caption of “Figure 4.8.1”). Consequently, the extracellular vesicles are “released from the beads prior to the RCA reaction, to allow subsequent detection of individual EVs by flow cytometry” (see, e.g., p. 4.8.3, para. 2). An artisan would have a reasonable expectation of success based on the given disclosures.
Claim 137 is rejected under 35 U.S.C. 103 as being unpatentable over Routenberg (cited above), and Lof, et al. (cited above) as evidenced by the product sheet for restriction enzyme RsaI from Thermo Scientific™ - Catalog number ER1121 (https://www.thermofisher.com/order/catalog/product/ER1121, cited in PTO-892 filed 06/17/2025), as applied to claims 11, 18-19, 108, and 115 above, and further in view of Han, et al. (“An Approach to Multiplexing an Immunosorbent Assay with Antibody-Oligonucleotide Conjugates”, published 2011, cited in PTO-892 dated 2026-01-14).
Routenberg and Lof teach as set forth above, but fail to teach the DNA/RNA duplex is degraded by RNase H, as in claim 137.
Han rectifies this in a journal article on immunosorbent assays with antibody-oligonucleotide conjugates. Han teaches the DNA/RNA duplex is degraded by RNase H, as in claim 137 (see, e.g., p. 2190, under abstract).
Routenberg, Lof, and Han are analogous to the field of the claimed invention because they are all in the field of antibody-oligonucleotide conjugates. One of ordinary skill in the art before the effective filing date of the application would have found it obvious to add the DNA/RNA duplex with RNase H of Han to the assay of Routenberg and Lof because “the DNA-RNA duplex by RNase H is exploited for fluorescent signal generation. Iterative cycles of DNA-RNA duplexation and subsequent degradation of the RNA in the duplex by RNase H further lead to amplification of the detection signal in OLISA” (see, Han, p. 2190, under abstract). An artisan would have had a reasonable expectation of success.
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.
The USPTO Internet website contains terminal disclaimer forms which may be used. Please visit www.uspto.gov/patent/patents-forms. The actual filing date of the application in which the form is filed determines what form (e.g., PTO/SB/25, PTO/SB/26, PTO/AIA /25, or PTO/AIA /26) should be used. A web-based eTerminal Disclaimer may be filled out completely online using web-screens. An eTerminal Disclaimer that meets all requirements is auto-processed and approved immediately upon submission. For more information about eTerminal Disclaimers, refer to www.uspto.gov/patents/apply/applying-online/eterminal-disclaimer.
Claims 1-4, 8, 11, 18-19, 25-26, 30, 138, and 140 are provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claims 1, 4, and 7 of copending Application No. 19/071,303 (referred to as ‘303 hereto) in view of Routenberg and Lof as evidenced by the product sheet for restriction enzyme RsaI from Thermo Scientific™ - Catalog number ER1121 (https://www.thermofisher.com/order/catalog/product/ER1121.
‘303 teaches a method for selectively purifying EVs expressing a surface marker, wherein the EVs comprise miRNA cargo, and determining a cargo profile for the EVs by extracting then quantifying miRNAs encapsulated in the EVs, as in claim 1 (see, claim 1 of ‘303). ‘303 teaches the method comprises an initial ultrafiltration step, an ultracentrifugation step or both to provide a pooled heterogeneous population of biological particles, as in claim 3 (see, claim 4 of ‘303). ‘303 teaches the samples is a body fluid, as in claim 4 (see, claim 1 of ‘303). ‘303 teaches the selective purifying is by antibody captures, as in claim 8 (see, claim 7 of ‘303).
‘303 fails to teach one of the surface markers is CD63 and the binding agent is linked to a low non-specific small RNA binding solid support by a nucleic acid, as in claim 1. ‘303 fails to teach identifying protein cargos by mass spectrometry, as in claim 2. ‘303 fails to teach the nucleic acid comprises DNA with one or more ribonucleic acid nucleotides and a restriction enzyme recognition site, as in claim 11. ‘303 fails to teach the nucleic acid has different binding moieties on each end, as in claim 18. ‘303 fails to teach the binding moiety is biotin, as in claim 19. ‘303 fails to teach releasing the biological particles by enzymatically cleaving the nucleic acid, as in claim 25. ‘303 fails to teach the enzymatic cleaving is with uracil glycosylase, as in claim 26. ‘303 fails to teach identifying the miRNAs by next generation sequencing, as in claim 30. ‘303 fails to teach the biological sample comprises a heterogeneuous population of cells, as in claim 138. ‘303 fails to teach the extracellular vesicles in a size range of about 40 nm to about 160 nm, as in claim 140.
However, with respect to claim 1, Routenberg teaches a method of preparing a purified population of biological particles from a biological sample and for evaluating a cargo of the purified population of biological particles (see, e.g., method - sheet 9/72, under “FIG. 7”; biological sample – para. [0098]; evaluating the purified population of biological particles – para. [0302] and sheet 9/72, under “FIG. 7”). Routenberg teaches (a) obtaining a biological sample comprising biological particles, such as extracellular vesicles (EVs) (see, e.g., obtaining biological sample comprising biological particles - para. [0102]-[0103]). Routenberg teaches (b) contacting the biological sample comprising the biological particles from the subject with a binding agent directed to a biological particle surface antigen, wherein one or more biological particle surface antigens comprises CD63, wherein the binding agent is immobilized directly or indirectly on a low non-specific small-RNA binding solid support, wherein, when the binding agent is immobilized indirectly on the low non-specific small-RNA binding solid support, it is coupled to a nucleic acid, and the nucleic acid is immobilized on the low non-specific small RNA binding solid support; and wherein the low non-specific small RNA binding solid support is a well plate (see, e.g., contacting particles with a binding agent - sheet 9/72, under “FIG. 7”; particle surface antigen comprises CD63 – para. [00161]; wherein the binding agent is immobilized directly or indirectly on a low non-specific small-RNA binding solid support, wherein, when the binding agent is immobilized indirectly on the low non-specific small-RNA binding solid support, it is coupled to a nucleic acid, and the nucleic acid is immobilized on the low non-specific small RNA binding solid support- sheet 9/72, under “FIG. 7”, and para. [00322]; wherein the low non-specific small RNA binding solid support is a well plate – para. [0087], and para. [00322], and sheet 9/72, under “FIG. 7). In “FIG. 7” of Routenberg, the capture antibodies (equivalent to binding agents) are attached to dsDNA linkers (equivalent to a nucleic acid coupled to the binding agent for indirectly immobilizing the binding agent to the solid support) which tether the capture antibodies to a solid support, such as a plate. Routenberg teaches wherein the method: (i) minimizes nonspecific binding of the biological particles to the low non-specific small RNA binding solid support without the binding agent; (ii) minimizes background binding of small RNAs to the low non-specific small RNA binding solid support; and (iii) detects low abundance small-RNA cargos (see, e.g., para. [00419], and para. [00312], and para. [00322]). It is understood that the solid phase, specifically the plate, of “FIG. 7” is equivalent a well plate. It is also understood that the method: (i) minimizes nonspecific binding of the biological particles to the low non-specific small RNA binding solid support without the binding agent; (ii) minimizes background binding of small RNAs to the low non-specific small RNA binding solid support; and (iii) detects low abundance small-RNA cargo is taught by the streptavidin-coated well plates of Routenberg because the applicant’s specification discloses that streptavidin-coated well plates are a low non-specific nucleic acid binding platform. See para. [0323] of the applicant’s specification. Routenberg teaches (c) isolating the biological particle bound by the binding agent from the biological sample by washing (see, e.g., para. [0089], and sheet 9/72, under “FIG. 7”, and para. [00563], and para. [00143]: “In embodiments, the unwanted components are soluble in the sample and/or the washing fluid”, and para. [00232], and para. [00375]). Routenberg teaches (d) releasing, the biological particle from the low non-specific small RNA binding solid support, bound to the binding agent (see, e.g., sheet 9/72, under “FIG. 7”, under step “2.”: “Cleave linker or denature dsDNA to elute EVs”). Routenberg teaches (e) eluting the bound biological particle from the binding agent to form a population of free purified biological particles (see, e.g., sheet 9/72, under “FIG. 7”, under step “3a.”: “elute antibodies from EV surface using low-pH”). Routenberg teaches evaluating surface molecules, specifically protein CD63 of the purified population of biological particles, further comprising extracting RNA from the purified population of biological particles and identifying the quantifying expression of small non-coding RNAs derived from the cell of origin comprising microRNAs encapsulated by the purified population of biological particles, wherein the isolated biological particles are derived from a subject (see, e.g., CD63 - para. [00161]; subjects - para. [00152]; miRNA – para. [0312]).
With respect to claim 2, Routenberg teaches step (f) evaluating cargo, further comprising identifying protein and/or lipid cargos by mass spectrometry (see, e.g., para. [0071]-[0072], and para. [0089]).
With respect to claim 3, Routenberg teaches an initial ultrafiltration step to provide a pooled heterogeneous population of biological particles and to increase specificity of exosome capture (see, e.g., para. [00143]). It is noted that the benefits of the ultrafiltration step follow naturally from the step itself.
With respect to claim 4, Routenberg teaches the biological sample comprises a body fluid comprising plasma (see, e.g., para. [0028]).
With respect to claim 8, Routenberg teaches the binding agent is an antibody (see, e.g., sheet 9/72, under “FIG. 7”).
With respect to claim 25, Routenberg teaches releasing the isolated biological particle comprises enzymatically cleaving the nucleic acid bound to the binding agent (see, e.g., para. [00258]). It is understood that the capture reagent is equivalent to the binding agent.
With respect to claim 26, Routenberg teaches the enzymatically cleaving is with an endonuclease (see, e.g., para. [00258]: “the labile linker […] is an oligonucleotide comprising a restriction site cleavable by a restriction endonuclease”).
With respect to claim 30, Routenberg teaches identifying the one or more small non-coding RNAs comprising miRNAs encapsulated in the one or more biological particles by next generation sequencing (see, e.g., para. [00291], and para. [00312]).
With respect to claim 32, Routenberg teaches the disease is cancer, as in claim 32 (see, e.g., para. [0028], and para. [0098], and para. [00111], and para. [00128], and para. [00149]).
With respect to claim 138, Routenberg teaches the biological sample comprises a heterogeneous population of cells, and the surface antigens are cell surface antigens, and the binding agent is directed to one or more cell surface antigens (see, e.g., para. [0096], and para. [00148]).
With respect to claim 140, Routenberg teaches the extracellular vesicles actively secreted extracellular bilayered membrane-bound vesicles of endosomal origin in a size range of about 100 nm on average generated by cells (see, e.g., para. [0567]).
‘303 and Routenberg are analogous to the field of the claimed invention because they are both in the field of EV analysis. One of ordinary skill in the art before the effective filing date of the application would have found it obvious to incorporate the nucleic acid linker of Routenberg into the method of ‘303. An artisan would have been motivated to do so because Routenberg discloses, “EVs are captured by labeled capture antibodies immobilized on a solid surface. The EV-capture antibody complex is first eluted from the solid surface, and the capture antibody can then optionally be released from the EV for further characterization, or the EV-capture antibody complex can be used directly in detection assays” (see, para. [0019]). An artisan would have had reasonable expectation of success based on the given disclosures.
This is a provisional nonstatutory double patenting rejection.
Claims 1-3, 8, 11, 18-19, 25-26, 30, 32, and 140 are provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claims 1, 4-9, and 12-27 of copending Application No. 18/777,426 (hereto referred to as ‘426) in view of Routenberg and Lof as evidenced by the product sheet for restriction enzyme RsaI from Thermo Scientific™ - Catalog number ER1121 (https://www.thermofisher.com/order/catalog/product/ER1121.
‘426 teaches a method for selectively purifying EVs expressing a surface marker, specifically CD63, wherein the EVs comprise miRNA cargo, contacting the EVs with a binding agent that is immobilized by a nucleic acid on a solid support with low non-specific small-RNA binding, and determining a cargo profile for the EVs by extracting then quantifying miRNAs encapsulated in the EVs, as in claim 1 (see, claims 1, 4, 9, and 24 of ‘426). ‘426 teaches evaluating cargo further comprises identifying protein and/or lipid cargos by mass spectrometry, as in claim 2 (see, claims 4-5 of ‘426). ‘426 teaches the method comprises an initial ultrafiltration step, an ultracentrifugation step or both to provide a pooled heterogeneous population of biological particles, as in claim 3 (see, claim 6 of ‘426). ‘426 teaches the selective purifying is by antibody captures, as in claim 8 (see, claim 7 of ‘426). ‘426 teaches the nucleic acid comprises DNA with one or more ribonucleic acid nucleotides and a restriction enzyme recognition site, as in claim 11 (see, claims 8, 15, and 26 of ‘426). ‘426 teaches the nucleic acid has different binding moieties on each end, as in claim 18 (see, claim 8 of ‘426). ‘426 teaches the binding moiety is biotin, as in claim 19 (see, claim 8 of ‘426). ‘426 teaches releasing the biological particles by enzymatically cleaving the nucleic acid, as in claim 25 (see, e.g., claim 25 of ‘426). ‘426 teaches the enzymatic cleaving is with uracil glycosylase, as in claim 26 (see, e.g., claim 26 of ‘426). ‘426 teaches identifying the miRNAs by next generation sequencing, as in claim 30 (see, e.g., claim 27 of ‘426). ‘426 teaches where in the disease comprises a cancer, as in claim 32 (see, e.g., claim 1 of ‘426).
‘426 fails to teach the extracellular vesicles in a size range of about 40 nm to about 160 nm, as in claim 140.
With respect to claim 140, Routenberg teaches the extracellular vesicles actively secreted extracellular bilayered membrane-bound vesicles of endosomal origin in a size range of about 100 nm on average generated by cells (see, e.g., para. [0567]).
‘426 and Routenberg are analogous to the field of the claimed invention because they are both in the field of EV analysis. One of ordinary skill in the art before the effective filing date of the application would have found it obvious to incorporate the nucleic acid linker of Routenberg into the method of ‘426. An artisan would have been motivated to do so because Routenberg discloses, “EVs are captured by labeled capture antibodies immobilized on a solid surface. The EV-capture antibody complex is first eluted from the solid surface, and the capture antibody can then optionally be released from the EV for further characterization, or the EV-capture antibody complex can be used directly in detection assays” (see, para. [0019]). An artisan would have had reasonable expectation of success based on the given disclosures.
This is a provisional nonstatutory double patenting rejection.
Claims 1-3, 8, 11, 18-19, 25-26, 30, 32, and 140 are provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claims 1 and 4-5 of copending Application No. 18/661,353 (hereto referred to as ‘353) in view of in view of Routenberg and Lof as evidenced by the product sheet for restriction enzyme RsaI from Thermo Scientific™ - Catalog number ER1121 (https://www.thermofisher.com/order/catalog/product/ER1121.
‘353 teaches a method for selectively purifying EVs expressing a surface marker, wherein the EVs comprise miRNA cargo, contacting the EVs with a binding agent that is immobilized by a nucleic acid on a solid support with low non-specific small-RNA binding, and determining a cargo profile for the EVs by extracting then quantifying miRNAs encapsulated in the EVs, as in claim 1 (see, claims 1 and 4-5 of ‘353). ‘353 teaches the selective purifying is by antibody captures, as in claim 8 (see, claims 1 and 5 of ‘353). ‘353 teaches the nucleic acid comprises DNA with one or more ribonucleic acid nucleotides and a restriction enzyme recognition site, as in claim 11 (see, claim 5 of ‘353). ‘353 teaches releasing the biological particles by enzymatically cleaving the nucleic acid, as in claim 25 (see, e.g., claim 5 of ‘353). ‘353 teaches the enzymatic cleaving is with uracil glycosylase, as in claim 26 (see, e.g., claim 5 of ‘353). ‘353 teaches wherein the disease comprises a cancer, as in claim 32 (see, e.g., claim 10 of ‘353).
‘353 fails to teach one of the surface markers is CD63 and the binding agent is linked to a low non-specific small RNA binding solid support by a nucleic acid, as in claim 1. ‘353 fails to teach evaluating cargo further comprises identifying protein and/or lipid cargos by mass spectrometry, as in claim 2. ‘353 fails to teach the method comprises an initial ultrafiltration step, an ultracentrifugation step or both to provide a pooled heterogeneous population of biological particles, as in claim 3. ‘353 fails to teach the nucleic acid has different binding moieties on each end, as in claim 18. ‘353 fails to teach the binding moiety is biotin, as in claim 19. ‘353 fails to teach identifying the miRNAs by next generation sequencing, as in claim 30. ‘353 fails to teach the extracellular vesicles in a size range of about 40 nm to about 160 nm, as in claim 140.
However, with respect to claim 1, Routenberg teaches a method of preparing a purified population of biological particles from a biological sample and for evaluating a cargo of the purified population of biological particles (see, e.g., method - sheet 9/72, under “FIG. 7”; biological sample – para. [0098]; evaluating the purified population of biological particles – para. [0302] and sheet 9/72, under “FIG. 7”). Routenberg teaches (a) obtaining a biological sample comprising biological particles, such as extracellular vesicles (EVs) (see, e.g., obtaining biological sample comprising biological particles - para. [0102]-[0103]). Routenberg teaches (b) contacting the biological sample comprising the biological particles from the subject with a binding agent directed to a biological particle surface antigen, wherein one or more biological particle surface antigens comprises CD63, wherein the binding agent is immobilized directly or indirectly on a low non-specific small-RNA binding solid support, wherein, when the binding agent is immobilized indirectly on the low non-specific small-RNA binding solid support, it is coupled to a nucleic acid, and the nucleic acid is immobilized on the low non-specific small RNA binding solid support; and wherein the low non-specific small RNA binding solid support is a well plate (see, e.g., contacting particles with a binding agent - sheet 9/72, under “FIG. 7”; particle surface antigen comprises CD63 – para. [00161]; wherein the binding agent is immobilized directly or indirectly on a low non-specific small-RNA binding solid support, wherein, when the binding agent is immobilized indirectly on the low non-specific small-RNA binding solid support, it is coupled to a nucleic acid, and the nucleic acid is immobilized on the low non-specific small RNA binding solid support- sheet 9/72, under “FIG. 7”, and para. [00322]; wherein the low non-specific small RNA binding solid support is a well plate – para. [0087], and para. [00322], and sheet 9/72, under “FIG. 7). In “FIG. 7” of Routenberg, the capture antibodies (equivalent to binding agents) are attached to dsDNA linkers (equivalent to a nucleic acid coupled to the binding agent for indirectly immobilizing the binding agent to the solid support) which tether the capture antibodies to a solid support, such as a plate. Routenberg teaches wherein the method: (i) minimizes nonspecific binding of the biological particles to the low non-specific small RNA binding solid support without the binding agent; (ii) minimizes background binding of small RNAs to the low non-specific small RNA binding solid support; and (iii) detects low abundance small-RNA cargos (see, e.g., para. [00419], and para. [00312], and para. [00322]). It is understood that the solid phase, specifically the plate, of “FIG. 7” is equivalent a well plate. It is also understood that the method: (i) minimizes nonspecific binding of the biological particles to the low non-specific small RNA binding solid support without the binding agent; (ii) minimizes background binding of small RNAs to the low non-specific small RNA binding solid support; and (iii) detects low abundance small-RNA cargo is taught by the streptavidin-coated well plates of Routenberg because the applicant’s specification discloses that streptavidin-coated well plates are a low non-specific nucleic acid binding platform. See para. [0323] of the applicant’s specification. Routenberg teaches (c) isolating the biological particle bound by the binding agent from the biological sample by washing (see, e.g., para. [0089], and sheet 9/72, under “FIG. 7”, and para. [00563], and para. [00143]: “In embodiments, the unwanted components are soluble in the sample and/or the washing fluid”, and para. [00232], and para. [00375]). Routenberg teaches (d) releasing, the biological particle from the low non-specific small RNA binding solid support, bound to the binding agent (see, e.g., sheet 9/72, under “FIG. 7”, under step “2.”: “Cleave linker or denature dsDNA to elute EVs”). Routenberg teaches (e) eluting the bound biological particle from the binding agent to form a population of free purified biological particles (see, e.g., sheet 9/72, under “FIG. 7”, under step “3a.”: “elute antibodies from EV surface using low-pH”). Routenberg teaches evaluating surface molecules, specifically proteins CD63 of the purified population of biological particles, further comprising extracting RNA from the purified population of biological particles and identifying the quantifying expression of small non-coding RNAs derived from the cell of origin comprising microRNAs encapsulated by the purified population of biological particles, wherein the isolated biological particles are derived from a subject (see, e.g., CD63 - para. [00161]; subjects - para. [00152]; miRNA – para. [0312]).
With respect to claim 2, Routenberg teaches step (f) evaluating cargo, further comprising identifying protein and/or lipid cargos by mass spectrometry (see, e.g., para. [0071]-[0072], and para. [0089]).
With respect to claim 3, Routenberg teaches an initial ultrafiltration step to provide a pooled heterogeneous population of biological particles and to increase specificity of exosome capture (see, e.g., para. [00143]). It is noted that the benefits of the ultrafiltration step follow naturally from the step itself.
With respect to claim 4, Routenberg teaches the biological sample comprises a body fluid (see, e.g., para. [0028]).
With respect to claim 30, Routenberg teaches identifying the one or more small non-coding RNAs comprising miRNAs encapsulated in the one or more biological particles by next generation sequencing (see, e.g., para. [00291], and para. [00312]).
With respect to claim 140, Routenberg teaches the extracellular vesicles actively secreted extracellular bilayered membrane-bound vesicles of endosomal origin in a size range of about 100 nm on average generated by cells (see, e.g., para. [0567]).
‘353 and Routenberg are analogous to the field of the claimed invention because they are both in the field of EV analysis. One of ordinary skill in the art before the effective filing date of the application would have found it obvious to incorporate the nucleic acid linker of Routenberg into the method of ‘353. An artisan would have been motivated to do so because Routenberg discloses, “EVs are captured by labeled capture antibodies immobilized on a solid surface. The EV-capture antibody complex is first eluted from the solid surface, and the capture antibody can then optionally be released from the EV for further characterization, or the EV-capture antibody complex can be used directly in detection assays” (see, para. [0019]). An artisan would have had reasonable expectation of success based on the given disclosures.
This is a provisional nonstatutory double patenting rejection.
Response to Arguments
The applicant’s arguments filed 04/09/2026 have been fully considered but have been found not persuasive.
35 U.S.C. 102 Rejection
The applicant argues that Routenberg (cited above) does not teach:
The solid support is a low non-specific small RNA binding solid support, which minimizes nonspecific and small RNA background binding; and
The evaluation of cargo and surface molecules comprising identifying and quantifying expression of small non-coding RNAs comprising miRNAs encapsulated by the purified population of biological particles.
As discussed in the claim interpretation section above, the low non-specific RNA binding solid support is described only in terms of benefits, not method steps or material constraints, except that the solid support is not a magnetic bead. See para. [0323] of the applicant’s specification. Routenberg discloses using multi-well plates with their methods (see, e.g., para. [00170], para. [00322]). The applicant continues arguments by pointing to the multiple examples of Routenberg that do not use magnetic beads. Therefore, the methods of Routenberg meet the limitations of the low non-specific small RNA binding solid support. Regarding the identification and quantification of miRNAs encapsulated by the purified population of biological particles, Routenberg also discloses these limitations (see, e.g., para. [00312]: “the encapsulated molecules within the EV of interest are subjected to an assay, e.g., an ultrasensitive assay, that comprises lysing the EV, and conducting an assay on the lysate […] the lysate is detected, analyzed or both using an assay. Assays for proteins, oligonucleotides and lipids are well known in the art. In embodiments, the EV of interest encapsulates a protein, a nucleic acid, a lipid, or a combination thereof. In embodiments, the nucleic acid is RNA. In embodiments the RNA is a mature miRNA”). While the applicant argues that Routenberg does not disclose an enabling disclosure for identifying and quantifying expression of miRNA, Routenberg teaches that assays for oligonucleotides are well known in the art and explicitly discloses miRNA encapsulated in the biological particles.
35 U.S.C. 103 Rejections
The applicant argues that Routenberg does not teach that the solid support is a low non-specific small RNA binding solid support. However, as discussed above, the applicant claims the disclosed benefits of the non-specific small RNA binding solid support by not using magnetic beads, and Routenberg teaches the use of multi-well plates (see, e.g., para. [00170], para. [00322]). The applicant again argues that Routenberg does not teach the evaluation of miRNA cargo. This argument has been addressed above.
The applicant continues arguments by arguing that while Lof is directed to an extracellular vesicle detection assay, the method is a bead-based capture method and does not use a low non-specific small RNA binding support as recited in the instant claims. The office agrees that Lof teaches a bead-based capture method and does not use a low non-specific small RNA binding solid support. However, Routenberg teaches the low non-specific small RNA binding solid support while disclosing in “Fig. 7” that well plates or beads are a design choice for solid phases, and Lof is merely relied on for disclosing the use of DNA linkers that include uracil. The applicant also argues that the DNA linker including uracil of Lof is used to enable probe attachment, circularization, ligation, and amplification. However, “Figure 4.8.1”, panel “E” of Lof clearly shows that the DNA linker is digested to release the beads from the capturing antibody-beads. Finally, the applicant argues that Lof does not disclose isolating EVs, releasing them as a purified population and then extracting and analyzing RNA derived from those particles by recovering RNA cargo from the EVs or analyzing the expression of small non-coding RNAs. However, as stated before, Lof is relied upon merely for the DNA linker comprising uracil. Lof is used for detecting EVs by capturing them and analyzing them, therefore, an artisan of ordinary skill in the art would understand the benefit that the DNA linkers bring, as discussed above.
The applicant continues arguments by stating that the Thermo Scientific Product Sheet for Restriction Enzyme RsaI does not pertain to the specific context of the claimed method. However, the product sheet is merely relied upon to provide evidence that the DNA linker of Lof, specifically the oligonucleotide disclosed on p. 4.8.4, under “Table 4.8.1”, under “Capturing antibody/ UNG digestion oligonucleotide 1”, includes a restriction enzyme site, as in claims 11 and 108. Teaching regarding EV or cell capture, low non-specific small RNA binding solid supports, preservation of endogenous RNA EV cargo, or extraction and quantification of small non-coding RNAs is disclosed by Routenberg.
The applicant continues by arguing that a skilled artisan would not have modified Routenberg by using the DNA linker of Lof to arrive at the claimed invention. However, the artisan would have been motivated to do so because “detection and characterization of EVs is challenging due to their small size” (see, e.g., Lof, p. 4.8.1, under abstract). To overcome the challenge, the extracellular vesicle-complex is “released from the beads by digestion of the uracil-containing oligonucleotides by treatment with uracil N-glycosylase” (see, e.g., Lof, p. 4.8.2, under the caption of “Figure 4.8.1”). Consequently, the extracellular vesicles are “released from the beads prior to the RCA reaction, to allow subsequent detection of individual EVs by flow cytometry” (see, e.g., Lof, p. 4.8.3, para. 2). An artisan would have understood that the release of EVs from the solid substrate allows for the subsequent processing of captured EVs. An artisan would have had a reasonable expectation of success based on the given disclosures.
The applicant continues by arguing that Routenberg emphasizes surface phenotyping strategies that rely on amplification of signals derived from binding events, rather than on recovery and downstream biological analyses of purified particles or cells. However, “Fig. 7” of Routenberg clearly discloses a method where EVs are purified then recovered for downstream by biological analyses.
The applicant continues with arguments regarding the low non-specific small RNA binding solid supports and the Thermo Scientific product sheet that have already been addressed above.
The applicant continues by arguing that their specification demonstrates that magnetic beads are inferior to multi-well plates, regarding non-specific binding of RNA. However, as discussed above, Routenberg teaches using multi-well plates.
Regarding claim 137, the applicant argues that Han (cited above) does not cure the deficiencies noted above for Routenberg, Lof, and the Thermo Scientific product sheet. However, Han is merely relied upon because Han teaches the DNA/RNA duplex is degraded by RNase H, as in claim 137 (see, e.g., p. 2190, under abstract).
Conclusion
No claims are allowed.
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/MICHAEL CAMERON SVEIVEN/Examiner, Art Unit 1678
/GREGORY S EMCH/Supervisory Patent Examiner, Art Unit 1678
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