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 .
Withdrawn Objections/Rejections
The objection to claim 11 for informalities is withdrawn, necessitated by amendments filed 09/23/2025.
The rejection of claim 108 on the grounds of 35 U.S.C. 112(b) is withdrawn, necessitated by amendments filed 09/23/2025.
The rejection of claims 1-4, 8, 10, 25-26, 30 and 32 on the grounds of 35 U.S.C. 103 is withdrawn, necessitated by amendments filed 09/23/2025.
The rejection of claims 11, 18, and 19 on the grounds of 35 U.S.C. 103 is withdrawn, necessitated by amendments filed 09/23/2025.
The rejection of claims 97-98, 100, 108, 115, and 127 on the grounds of 35 U.S.C. 103 is withdrawn, necessitated by amendments filed 09/23/2025.
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.
Status of Claims
Claims 5-7, 9, 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, and 128-135 are cancelled.
Claims 33, 36, 38-40, 46, 60-62, 76-77, 79-82, and 95 are withdrawn due to restriction and species election.
Claims 1-4, 8, 10-11, 18-19, 25-26, 30, 32, 97, 100, 108, 115, 127, and 136-140 are examined herein.
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, 10, 25-26, 30, 32, 97, 100, 127, 136, and 138-140 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Routenberg (WO 2020/086751 A1, published 2020-04-30).
With respect to claim 1, Routenberg teaches a method of preparing a purified population of biological particles from a biological sample and for evaluating the purified population of biological particles (see, e.g., 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”). It is understood that extracellular vesicles (EVs) are equivalent to biological particles. Routenberg teaches preparing a purified population of biological particles by (1) obtaining a biological sample comprising biological particles (2) contacting the biological sample comprising biological particles from the subject with a binding agent directed to a biological particle surface antigen; wherein the binding agent is linked to a nucleic acid, and wherein the nucleic acid is immobilized on a well plate format, and wherein the method: (i) minimizes nonspecific binding of the biological particles to the solid support without the binding agent; (ii) minimizes background binding of small RNAs to the solid support; and (iii) detects low abundance small-RNA cargos (see, e.g., obtaining biological sample comprising biological particles - para. [0098]; contacting a biological sample with a binding agent directed to a biological particle antigen, wherein the binding agent is linked to a nucleic acid, and wherein the nucleic acid is immobilized on a solid support – sheet 9/72, under “FIG. 7”). 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 solid support without the binding agent; (ii) minimizes background binding of small RNAs to the solid support; and (iii) detects low abundance small-RNA cargos because the method steps naturally result in the effects enumerated. Routenberg teaches isolating the biological particle bound by the binding agent from the biological sample (see, e.g., para. [0089], and sheet 9/72, under “FIG. 7”). Routenberg teaches releasing the biological particle 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 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 the protein CD63, of the purified population of biological particles, wherein the isolated biological particles are derived from a subject (see, e.g., CD63 - para. [00161]; subjects - para. [0028]).
With respect to claim 2, Routenberg teaches extracting RNA from the biological particles and identifying and quantifying expression of microRNAs (miRNAs) encapsulated by the purified population of biological particles (see, e.g., para. [00312]).
With respect to claim 3, Routenberg teaches an initial ultrafiltration step to provide a starting pooled heterogeneous population of biological particles (see, e.g., para. [00143]).
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 10, Routenberg teaches the biological particle surface antigen is CD63 and the solid support is a multiwell format well plate (see, e.g., surface antigen - para. [00161]; solid support is a multiwell format well plate – sheet 9/72, under “FIG. 7” and para. [0087]: “the methods use a high throughput 96-well plate format using a single immunoaffinity capture step”).
With respect to claim 25, Routenberg teaches releasing the isolated biological particle comprises enzymatically cleaving the nucleic acid (see, e.g., para. [00258]: “the labile linker […] is an oligonucleotide comprising a restriction site cleavable by a restriction endonuclease”).
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 [00312]).
With respect to claim 32, Routenberg teaches identifying the sample is from a subject with a disease and the disease is cancer, as in claim 32 (see, e.g., para. [0028]).
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 preparing a purified population of cells by (1) obtaining a biological sample comprising cells (2) contacting the biological sample comprising cells from the subject with a binding agent directed to a biological particle surface antigen; wherein the binding agent is linked to a nucleic acid, and wherein the nucleic acid is immobilized on a well plate (see, e.g., obtaining biological sample comprising cells - 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)”; contacting a biological sample with a binding agent directed to a cell antigen, wherein the binding agent is linked to a nucleic acid, and wherein the nucleic acid is immobilized on a solid support – sheet 9/72, under “FIG. 7”). It is understood that the solid phase, specifically the plate, of “FIG. 7” is equivalent to a well plate format. Routenberg teaches isolating the cells bound by the binding agent from the biological sample (see, e.g., para. [0089], and sheet 9/72, under “FIG. 7”). Routenberg teaches releasing 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 free purified biological particles; and wherein the method: minimizes nonspecific binding to the solid support without the binding agent; and minimizes background binding of small RNAs to the solid support; and is tailorable for selecting specific populations of cells for analysis of their contents (see, e.g., sheet 9/72, under “FIG. 7”, under step “3a.”: “elute antibodies from EV surface using low-pH”). It is understood that the method: minimizes nonspecific binding to the solid support without the binding agent; and minimizes background binding of small RNAs to the solid support; and is tailorable for selecting specific populations of cells for analysis of their contents because the methods steps naturally result in the described effects. Routenberg teaches evaluating surface molecules, specifically the protein CD63, of the purified population of cells, wherein the isolated cells are derived from a subject (see, e.g., CD63 - para. [00161]; subjects - para. [0028]).
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]).
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]).
With respect to claim 139, Routenberg teaches the biological particles include “cells (including prokaryotic cells such as bacterial cells or archaeal cells; eukaryotic cells such as mammalian cells, insect cells, or plant cells); viruses and viral particles; cellular organelles such as nucleus, endoplasmic reticulum, Golgi apparatus, mitochondria, vacuoles, or chloroplast; vesicles such as lysosome, endosome, peroxisome, and liposome; and extracellular vesicles (EVs) or exosomes” (see para. [0070]).
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.
Claims 11, 18, 19, 108, 115 are rejected under 35 U.S.C. 103 as being unpatentable over Routenberg (cited above), as applied to claims 1-4, 8, 10, 25-26, 30, 32, 97, 100, 127, 136, and 138-140 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 teach 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 also teaches the population of purified cells comprises eukaryotic cells, specifically human cells, as in claim 127 (see, e.g., para. [0098]).
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 rectifies these deficiencies in a journal article entitled “Detection of Extracellular Vesicles Using Proximity Ligation Assay with Flow Cytometry Readout—ExoPLA”. 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).
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, Hang, p. 2190, under abstract). An artisan would have had a reasonable expectation of success.
Conclusion
No claims are allowed.
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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/MICHAEL CAMERON SVEIVEN/Examiner, Art Unit 1678
/GREGORY S EMCH/Supervisory Patent Examiner, Art Unit 1678