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 .
Status of claims / Response to Amendment
This office action is in response to an amendment filed on October 28, 2025.
Claims 1-2, 4, 6, 9-12, 18-19, 23, 26, 28, 30-31, 52, 78-80 and 83-84 were previously pending. Applicant amended claims 52, 78-79 and 83-84. Claims 96-97 are newly added.
Claims 1-2, 4, 6, 9-12, 18-19, 23, 26, 28, 30-31, 52, 78-80, 83-84 and 96-97 are currently pending, with claims 1-2, 4, 6, 9-12 and 18-19, 23, 26, 28, 30-31 withdrawn.
Claim 52, 78-80, 83-84 and 96-97 are under examination.
Applicant's submission of the amendment to specification obviated the previously presented objection.
Applicant's claim amendments and arguments overcame the following objection and rejections:
Objection to claim 52;
Rejections to claims 52, 78-80 and 83-84 under 35 U.S.C. 112(b);
Rejections of claims 78 and 83-84 under 35 U.S.C. 103 as being unpatentable over Satija, in view of Stoeckius and Weber, as evidenced by Islam;
Rejections of claims 78 and 83-84 under 35 U.S.C. 103 as being unpatentable over McGinnis;
Provisional rejection of claim 78 on the ground of nonstatutory double patenting as being unpatentable over claims 1 and 5 of copending Application No. 18/762,349 (claims - 07/02/2024);
Provisional rejection of claim 78 on the ground of nonstatutory double patenting as being unpatentable over claims 12, 17 and 20 of copending Application No. 17/057,569 (Amended Claims - 09/18/2024).
All other previously presented rejections are maintained for reasons given in the "Response to Arguments" below. Applicant' s amendments and arguments have been thoroughly reviewed, but are not persuasive to place the claims in condition for allowance for the reasons that follow.
This office action contains new grounds for rejection necessitated by amendment.
Priority
The priority date of the instant claims 52, 78-80, 83-84 and 96-97 is 07/08/2019, filling date of the US provisional application NO. 62/871,702.
Claim Objections -- New Ground
Claim 79 is objected to because of the following informalities:
In claim 79, line 6: "a second second hybridization region" should read "a second hybridization region."
Claim Interpretation -- Updated
In evaluating the patentability of the claims presented in this application, claim terms have been given their broadest reasonable interpretation (BRI) consistent with the specification, as understood by one of ordinary skill in the art, as outlined in MPEP§ 2111.
For the purpose of applying prior art, both independent claims 52 and 78 recite the term "lipid-conjugated DNA oligonucleotide," which is not expressly defined in the application's disclosure. The specification provides definition for related term "lipid-modified oligonucleotide" as follows:
"The term "lipid-modified oligonucleotide", "lipid-DNA", "hydrophobic-anchored oligonucleotide" and similar terms are to be broadly construed to include any oligonucleotide or polynucleotide that is attached by any means to a hydrophobic, lipophilic, or amphiphilic region that can be inserted into a membrane, regardless of whether the "lipid-modified oligonucleotide" ,"lipid-DNA", "hydrophobic-anchored oligonucleotide", or portion thereof is actually inserted into a membrane. "(page 13)
Thus, under BRI and consistent with the specification, the term "lipid-conjugated DNA oligonucleotide" is interpreted to have the same meaning as the term ""lipid-modified oligonucleotide," defined above.
For the purpose of applying prior art, both independent claims 52 and 78 recite DNA oligonucleotides comprising “lipid moiety,” which is a term not expressly defined in the specification.
Thus, under BRI and based on the commonly understood meaning by one of ordinary skill in the art, the term "lipid moiety" is interpreted to encompass any element naturally occurring or synthetic that are generally hydrophobic in nature (See Fahy 1, introduction; see also Gartner 2, [0136]). This interpretation is consistent with the instant specification's description of "lipid moiety" (see page 36, lines 4-7 for example).
For the purpose of applying prior art, both independent claims 52 and 78 recite DNA oligonucleotides comprising “barcode region, ” which is not expressed defined in the application's disclosure. The specification provides the following relevant description regarding barcode region:
"Barcode Regions
The barcode oligonucleotides comprise the second primer region operably linked ( e.g., covalently linked) to a barcode region which in turn is operably linked (e.g., covalently linked) to a capture sequence (described below), the barcode region comprising an oligonucleotide of 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32,33,34,35,36, 37,38,39,40,41,42,43,44,45,46,47,48,49, 50, 51,52,53, 54, 55, 56,57, 20 58, 59,60,61,62,63,64,65,66,67,68,69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80,81, 82,83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100 or more nucleotide bases. The oligonucleotide can be DNA, RNA, or modified or synthetic DNA or RNA. " (page 41)[emphasis added]
Thus, under BRI, the term "barcode region" without any additional modifier, is interpreted to encompass any oligonucleotide sequences that are at least 2 nucleotide bases long.
For the purpose of applying art, both independent claims 52 and 78 recite DNA oligonucleotides comprising "capture sequence," which is not expressly defined in the application's disclosure. The specification provides the following relevant description regarding capture sequence:
"In some embodiments, the barcode oligonucleotides comprise the second primer region operably linked (e.g., covalently linked) to the barcode region which in tum is operably linked (e.g., covalently linked) to a capture sequence (described below), the capture sequence comprising an oligonucleotide of 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25,26,27,28,29,30,31,32,33,34,35,36,37,38,39,40,41,42,43,44,45,46,47,48,49,50, 51, 52,53,54, 55, 56,57, 58, 59,60,61, 62,63,64,65,66,67,68,69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100 or more nucleotide bases. The oligonucleotide can be DNA, RNA, or modified or synthetic DNA or RNA. " (page 42-43)[emphasis added]
Thus, under BRI, the term "capture sequence" without any additional modifier, is interpreted to encompass any oligonucleotide sequences that are at least 2 nucleotide bases long.
For the purpose of applying prior art, claim 52 recites "each oligonucleotide unique for and corresponding to one of the plurality of vessels into which the one or plurality of cells are exposed."
Applicant's remarks filed on October 28, 2025, at page 14 provides the following clarification regarding claim interpretation on the record:
"With regard to ascertaining the intended relationship, the skilled artisan would have readily understood that each vessel contains unique oligonucleotides over other vessels, and that one or a plurality of cells were partitioned among the vessels. See, for example, step (a): "partitioning one or a plurality of cells from the sample or the tissue corresponding to a region of the sample into one of a plurality of vessels." The claims are clear, and one of ordinary skill would readily comprehend that, each vessel has a portion of the sample or the tissue comprising one or a plurality of cells and each vessel comprises unique oligonucleotides in comparison to the other vessels."
Accordingly, in view of Applicant's remarks, each vessel of the plurality of vessels is interpreted to comprise a plurality of oligonucleotides sharing common structure and sequence that specifically label the cells within the vessel with barcode (e.g., sample barcode), which also corresponds to the identity of the vessel (e.g., cells from a lung tissue sample specifically barcoded in a specific well position in a 24-well plate, thus the barcode corresponds to both the sample type and well-position).
For the purpose of applying prior art, claim 52 recites the term "isolating" in "isolating or identifying nucleic acid from the one or a plurality of cells according to the plurality of oligonucleotides."
The application's disclosure does not expressly define the term, but describes isolating as encompassing detection of oligonucleotide barcodes by methods such as sequencing, which may not involve physical isolation of nucleic acids:
"In some embodiments, the step of isolating the nucleic acid of the one or plurality of cells of the disclosed method comprises isolating the one or plurality of cells based upon the presence of one or plurality of labeling oligonucleotides, wherein the presence of the one or plurality of labeling oligonucleotides is determined by detection of one of more unique nucleotide sequences corresponding to the one or plurality of labeling oligonucleotides. " (page 6)
"The disclosure relates to a method of preparing a library of oligonucleotides expressed by a single cell or multiple cells in isolation, the method of generating the library comprising sequencing RNA or DNA from the one or multiple cells after the one or multiple cells are exposed to one or a plurality of lipid-modified oligonucleotides disclosed herein. In some embodiments, endogenous nucleotides from one or a plurality of cells can be isolated and/or identified by correlating a known signal or frequency of a probe bound to the lipid-modified oligonucleotide to the cell upon which the oligonucleotide was bound. The signal or frequency of the probe can be paired with the source of the endogenous DNA and/or RNA." (page 65) [emphasis added]
Therefore, under BRI and in light of the specification, the term "isolating" is interpreted to encompasses barcode sequence detection-based approaches (e.g., detecting specific barcode sequences in sequencing reads to identify transcripts from a specific source labeled by the barcode) that do not necessarily require physical separation.
Claim Rejections - 35 USC § 112(b) -- New Grounds
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.
Claims 52, 78-80, 83-84 and 96-97 are rejected under 35 U.S.C. 112(b), 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.
A) Regarding claim 52, it recites a "plurality of oligonucleotides disclosed herein." The phrase "disclosed herein" is indefinite because it is ambiguous. It is unclear what specific oligonucleotides are being referenced. The claim does not provide sufficient antecedent basis or clearly identify or disclose the scope of the plurality of oligonucleotides. It is not clear whether this phrase refers to oligonucleotides disclosed in the specification, and whether it is limited to all of the specific oligonucleotides expressly recited in the specification, a subset thereof, or a broader group.
Claim 52 further recites "wherein each of the plurality of oligonucleotides disclosed herein in each of the plurality of vessels is independently selected," the meaning of "independently selected" within the context of the claimed method is unclear. Neither the claim nor the specification provide any guidance as to how "independently" modifies or distinguishes the selection process, or how it differs from the plain meaning of the term "selected." This introduces ambiguity to the scope of the claim.
Claim 96 is rejected for depending from claim 52 and not remedying the indefiniteness.
B) Claim 78 recites “correlating the sequenced nucleic acids from the one or plurality of cells to the spatial position of the one or plurality of cells within the tissue and/or relative to the tissue.”
However, the phrase "relative to the tissue" is indefinite and introduces ambiguity.
The preamble recites "[a] method of identifying a spatial expression pattern of a nucleic acid within a tissue," but it does not recite a spatial position "relative to the tissue."
The claim recites "nucleic acids from the one or plurality of cells" and "one or a plurality of cells from a sample corresponding to a region of the tissue," thus the origin of the nucleic acids is from cells that are derived from a sample that is part of the tissue. In other words, the origin of the nucleic acids is from within the tissue. It is unclear what "relative to the tissue" refers to in this context. If the cells and their nucleic acids originate from the tissue, their spatial position would inherently be "within the tissue." It is unclear how a spatial position of these cells could instead be "relative to the tissue," suggesting a location outside of the tissue. This creates uncertainty regarding the scope of "relative to the tissue," and whether it refers to a different tissue or spatial context outside the claimed tissue.
Claims 79-80, 83-84 and 97 are rejected for depending from claim 78 and not remedying the indefiniteness.
C) Claims 83 and 84 are indefinite for reciting "wherein the first lipid moiety comprises a compound of Formula I" and "wherein the second lipid moiety comprises a compound of Formula II," followed by "Formula I [or Formula II]or a salt thereof."
Specifically, the role of the recitation "or a salt thereof" in the claims is unclear. The claims only recite "wherein the first lipid moiety comprises a compound of Formula I" and "wherein the second lipid moiety comprises a compound of Formula II. Therefore, the first lipid moiety and the second lipid moiety are only recited to comprise a compound of Formula I or II.
However, "or a salt thereof" appears later in the claims as an alternative to Formula I or II. This creates ambiguity regarding the scope of the claimed invention because it is uncertain whether the "the first lipid moiety" and "the second lipid moiety" are recited to comprise salt of the Formulas in alternative embodiments or not. Accordingly, the scope of the claims are indefinite.
If the claims are intended to encompass the salt of the Formula I or II, the following potential amendment "wherein the first lipid moiety comprises a compound of Formula I or a salt thereof" and "wherein the second lipid moiety comprises a compound of Formula II or a salt thereof" could potentially resolve the indefiniteness issue.
Claim Rejections - 35 USC § 112(d) -- New Grounds
The following is a quotation of 35 U.S.C. 112(d):
(d) REFERENCE IN DEPENDENT FORMS.—Subject to subsection (e), a claim in dependent form shall contain a reference to a claim previously set forth and then specify a further limitation of the subject matter claimed. A claim in dependent form shall be construed to incorporate by reference all the limitations of the claim to which it refers.
Claim 80 is rejected under 35 U.S.C. 112(d) or pre-AIA 35 U.S.C. 112, 4th paragraph, as being of improper dependent form for failing to further limit the subject matter of the claim upon which it depends, or for failing to include all the limitations of the claim upon which it depends.
Claim 80 depends from claim 79 and recites "wherein the first lipid-conjugated DNA oligonucleotide further comprises a first hybridization region."
This dependent claim is improper because it does not add any limitation to the subject matter of claim 79, which already recites that the first lipid-conjugated DNA oligonucleotide comprises a first hybridization region. Therefore, claim 80 fails to further limit the subject matter of claim 79, from which it depends.
Applicant may cancel the claim(s), amend the claim(s) to place the claim(s) in proper dependent form, rewrite the claim(s) in independent form, or present a sufficient showing that the dependent claim(s) complies with the statutory requirements.
Claim Rejections - 35 USC § 112(a) -- Maintained with Update
The following is a quotation of the first paragraph of 35 U.S.C. 112(a):
(a) IN GENERAL.—The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor or joint inventor of carrying out the invention.
Claims 52 and 96 are rejected under 35 U.S.C. 112(a) as failing to comply with the written description requirement. The claims contains subject matter which was not described in the specification in such a way as to reasonably convey to one skilled in the relevant art that the inventor or a joint inventor, or for applications subject to pre-AIA 35 U.S.C. 112, the inventor(s), at the time the application was filed, had possession of the claimed invention.
Claim 52 recites "[a] method of identifying a spatial position of a pattern of nucleic acid expression within a sample or tissue of a subject." However, the specification does not provide sufficient written description support to reasonably convey to one skilled in the art that the inventors were in possession of the full scope of the claimed invention at the time of filling, particularly with the respect to the broader genus of "sample."
The term "sample" is broadly defined in the specification:
"As used herein, the term "sample" refers generally to a limited quantity of something which is intended to be similar to and represent a larger amount of that thing. In the present disclosure, a sample is a collection, swab, brushing, scraping, biopsy, removed tissue, or surgical resection that is to be tested for an assay or method disclosed herein."(page 20)
Thus, the genus "sample" includes both tissue and non-tissue species. Under BRI, the term "sample" encompasses a wide range of materials, including those that do not or may not contain intact cells ꟷ such as environmental or inorganic samples, waste water collection, surface swabs or brushing within minimal or no cellular content, or homogenized samples in lysis buffer in which cells have already lysed.
However, the disclosure only supports methods of identifying a spatial position of a pattern of nucleic acid expression when performed using tissue. All working examples and prophetic examples require tissue samples (see Figures 1 and 2; see also examples on pages 66-68). The disclosure lacks sufficient detail to support the genus "sample" for use in the claimed method, because disclosure of one species (tissue) is not sufficient to support the full scope of the genus, especially where other species encompassed by the genus (e.g., swabs, brushings, scrapings) may differ significantly in cellular content and structure, raising doubts in operability.
While the specification mentions that the sample may be "a brushing of an environmental are or location, such as a lab bench or medical device" (page 20, lines 10-11), it does not provide any detail on how such a sample would be suitable for use in the methods disclosed. A tissue sample typically contains a high number of viable cells, whereas a brushing or swab may contain far fewer cells, potentially making them unsuitable for identifying a spatial pattern of nucleic acid expression in the same manner. Therefore, a skilled artisan would not reasonably predict that the methods disclosed for tissues would also apply to these non-tissue sample types. In re Curtis, 354 F.3d 1347, 1358 (Fed. Cir. 2004) (“[A] patentee will not be deemed to have invented species sufficient to constitute the genus by virtue of having disclosed a single species when, as is the case here, the evidence indicates ordinary artisans could not predict the operability in the invention of any species other than the one disclosed.”)
Accordingly, the specification fails to provide written description support for the claimed method as applied to the full scope of "sample."
Given the broad scope of the term "sample," encompassing all tissue and non-tissue sample types, and the lack of written description support for practicing the claimed method of identifying a spatial position of a pattern of nucleic acid expression within a sample using any non-tissue sample type, the specification, at best, presents an unsubstantiated wish or a research plan, “leaving it to others to explore the unknown contours of the claimed genus.” AbbVie Deutschland GmbH v. Janssen Biotech, Inc., 759 F.3d 1285, 1300 (Fed. Cir. 2014); see also Centocor, 636 F.3d at 1351.
Therefore, the claims encompass subject matters that extend beyond the content disclosed in the application, failing to meet the written description requirement of 35 U.S.C. 112(a).
Claim 96 is rejected because it depends from claim 52 and inherit the deficiencies of the base claim.
Response to Argument
The rejection of claim 52 under 35 U.S.C. 112(a) as failing to comply with the written description requirement is maintained in this Office Action.
Regarding the 112(a) rejection previously set forth in the Non-Final Office Action, Applicant did not amend the claim to specifically address the issue presented, and instead argues that the rejection is improper (Remarks, page 16-17).
Specifically, Applicant contends that "the written description need only be so specific as to lead one having ordinary skill in the art to envisage the class," citing two case laws in support.
Applicant further asserts that "the skilled artisan would have readily understood that Applicant was in possession of and intended to claim the full scope of claim 52 at the time of the original specification" and "one of ordinary skill in the art, using the specification as prepared, would know how to extract a tissue sample or a plurality of cells from a sample and place them in the vessels to perform the claimed method steps."
Applicant also argues that the Office Action "impermissibly conflates the concepts of possession with the concepts of enablement and ignores the listed examples of a sample - a tissue, a collection, a swab, a brushing, a scraping, a biopsy, removed tissue, or surgical resection, which leads the skilled artisan to the class of 'sample.' "
Applicant's arguments have been fully considered but are not found persuasive.
First, while the cited case law address certain aspects of the written description requirement, but the arguments make only generalizations not tied to the specific facts of the cases.
Second, the applicant's entire argument relies on interpretations that a skilled artisan would recognize the entire scope of the claimed genus as supported by the disclosure. However, no objective evidence has been provided to support these assertations or to establish what would be considered general knowledge in the art. As the arguments of counsel cannot take the place of evidence in the record, assertions made without objective evidence from relevant references are insufficient to bridge the gap between the content disclosed in the application, and the breadth of the claims. The arguments offered by the applicant, though detailed, amount to speculation without corroborative support from the what is known in the field.
The assertion that a skilled artisan would know how to extract tissue or cells from a sample presumes that the sample necessarily contains a sufficient amount of intact tissue or cells for extraction. However, under BRI, the term "sample" encompasses a wide range of materials, including those that do not or may not contain intact cells ꟷ such as environmental or inorganic samples, waste water collection, surface swabs or brushing within minimal or no cellular content, or homogenized samples in lysis buffer in which cells have already lysed.
Thus, as clearly articulated in the rejection, the issue lies with the breadth of the term "sample." In view of the above, it remains the Examiner's position that a written description rejection for lack of possession is appropriate and proper 3.
The applicant is advised to provide objective evidence, such as technical literature, review articles or other references, to support the assertions made regarding the knowledge of a skilled artisan and the support for the claimed invention in the disclosed embodiments.
Therefore, the 112(a) written description rejection is maintained and updated to reflect the recent claim amendment.
Claim Rejections - 35 USC § 103 -- New Grounds
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
Claims 52, 78-80, 83-84 and 96-97 are rejected under 35 U.S.C. 103 as being unpatentable over McGinnis (McGinnis et al. MULTI-seq: sample multiplexing for single-cell RNA sequencing using lipid-tagged indices. Nat Methods 16, 619–626 (2019). doi.org/10.1038/s41592-019-0433-8; Published 17 June 2019; cited as NPL # 4 on IDS filed 10/03/2023), as evidenced by
Satija (Satija et al. Spatial reconstruction of single-cell gene expression data. Nat Biotechnol 33, 495–502 (2015);
Ocqueteau (Ocqueteau et al. Three-dimensional morphometry of mammalian cells. I. Diameters. Arch Biol Med Exp. 1989 Jul;22(2):89-95. PMID: 269496), and
Weber (Weber, Robert J., et al. "Efficient targeting of fatty-acid modified oligonucleotides to live cell membranes through stepwise assembly." Biomacromolecules 15.12 (2014): 4621-4626.).
A) McGinnis teaches methods for sample multiplexing labeling for single-cell RNA sequencing using lipid-conjugated DNA oligonucleotides, referred to as MULTI-seq (entire document; Fig. 1 for example).
Regarding claim 52, McGinnis teaches a method comprising:
(a) partitioning one or a plurality of cells from the sample or the tissue corresponding to a region of the sample into one of a plurality of vessels (Fig. 3a and 3b; page 623, left-hand col, lines 1-3; page 627, left-hand col, para 6, cells that correspond to tissues in primary tumors and lungs are partitioned for individual sample barcoding prior to pooling);
(b) exposing the one or plurality of cells corresponding to a region of the sample with a plurality of oligonucleotides disclosed herein to incorporate the plurality of oligonucleotides into the one or plurality of cells (Fig. 1a; Fig 3a), wherein each oligonucleotide is unique for and corresponding to one of the plurality of vessels into which the one or plurality of cells are exposed (page 623, left hand col, lines 1-3, 9 distinct samples are barcoded; page 619, right-hand col, para 2, lines 7-9, “MULTI-seq sample barcodes include a 3'poly-A capture sequence, an 8-bp sample barcode and a 5' PCR handle necessary for library preparation and anchor hybridization.”; page 627, left-hand col, para 5, barcoding of different samples is done using different wells of a 24 well plate, thus the barcode is unique to each well and sample type );
(c) isolating or identifying nucleic acid from the one or a plurality of cells according to the plurality of oligonucleotides (Fig 3b; page 628, left hand col, para 6-10, sequenced transcripts in cells were identified per barcode);
(d) quantifying expression of nucleic acids and/or sequencing the nucleic acid from the one or a plurality of cells (Fig. 3) ;
(e) normalizing the expression of nucleic acid in an expression profile (page 628, left-hand col, para 10, “MULTI-seq sample classification” lines 4-6); and
wherein each of the plurality of oligonucleotides disclosed herein in each of the plurality of vessels is independently selected from one or a combination of:
(x) a first lipid-conjugated DNA oligonucleotide comprising a first lipid moiety, a first hybridization region, and a first primer region (Fig 1a);
wherein the step of partitioning comprises placing a slice or three-dimensional preparation of the sample from about 0.1 to about 99 microns in thickness into one of a plurality of vessels (Fig. 3a and 3b; page 623, left-hand col, lines 1-3; page 627, left-hand col, para 6, mouse cells that correspond to tissues in primary tumors and lungs are partitioned for individual sample barcoding prior to pooling)).
McGinnis teaches "placing a three-dimensional preparation of the sample from about 0.1 to about 99 microns in thickness into one of a plurality of vessels " by teaching partitioning mouse cells for separate barcoding (Id.). As evidenced by Ocqueteau (Table II), mouse cells are three-dimensional preparations having mean thickness (diameter) range from 3.93-21.42 microns.
Regarding claim 78, McGinnis teaches a method comprising:
(a) partitioning one or a plurality of cells from a sample corresponding to a region of the tissue into one of a plurality of vessels (Fig. 3a and 3b; page 623, left-hand col, lines 1-3; page 627, left-hand col, para 6, cells that correspond to tissues in primary tumors and lungs are partitioned for individual sample barcoding prior to pooling);
(b) exposing the one or plurality of cells with lipid-conjugated DNA oligonucleotides comprising a first lipid moiety, a barcode region ,and a capture sequence (Fig. 1a; page 619, right-hand col, para 2, lines 7-9, “MULTI-seq sample barcodes include a 3'poly-A capture sequence, an 8-bp sample barcode and a 5' PCR handle necessary for library preparation and anchor hybridization.”) to embed the lipid-conjugated DNA oligonucleotides within cell membrane of the one or plurality of cells,
wherein the barcode region of each of the lipid-conjugated DNA oligonucleotides is unique for each of the one of plurality of vessels in which the one or plurality of cells are exposed (Fig. 1d, legend, unique molecular identifiers; page 621,right-hand col, lines 1-4; );
(c) sequencing nucleic acids captured by the capture sequence of the lipid- conjugated DNA oligonucleotide in the one or plurality of cells (Fig 3).
With regard to both independent claims 52 and 78, while McGinnis does not explicitly teach a step of correlating sequenced nucleic acids from cells to the spatial position of the cells within the tissue, based on the barcode in the sequenced nucleic acids, it suggests that its MULTI-seq method has the capability to do so (page 625, left-hand col, para 1-2), thus providing motivations for modification.
McGinnis highlights the benefits of enabling direct interrogation of tumor heterogeneity and indicates that its method can be further utilized to obtain spatial coordinate information:
"Since we isolated immune cells from the whole lung in this study, we could not
discern whether Cd14-high and Cd14-low states were spatially correlated with metastatic sites. However, MULTI-seq could be employed to spatially barcode distinct regions of a single metastatic lung, enabling direct interrogation of classical monocyte spatial heterogeneity.
In summary, MULTI-seq broadly enables users to incorporate additional layers of information into scRNA-seq experiments. We anticipate that in the future, more diverse types of information will be targeted, including spatial coordinates, time-points, species-of origin and sub-cellular structures (for example, nuclei from multinucleated cells). We also anticipate that increasing LMO membrane residency time using alternative oligonucleotide conjugate designs may enable MULTI-seq applications for non-genetic lineage tracing and/or cellular competition assays." (page 625, left-hand col, para 1-2)[emphasis added]
Therefore, it would have been prima facie obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention to apply the method taught by McGinnis to analyze spatial coordinates of nucleic acids by correlating the sequenced nucleic acids to the spatial position of the cells containing these nucleic acids within the tissue, using barcodes, as suggested by McGinnis.
The person of ordinary skill would have had a reasonable expectation of success in further analyzing nucleic acids spatial information while applying McGinnis's method, because the data analysis approach described in McGinnis directly supports obtaining nucleic acid spatial information, as evidenced by Satija.
McGinnis performs gene expression analysis for the sequenced RNA transcripts coupled with their corresponding UMI information, using ‘Seurat’ R package, as disclosed in Satija (see McGinnis, page 628, para 6 "Expression library analysis"). Satija provides detailed teaching of spatial localization methods comprising single cell RNA sequencing with data analysis approach using the ‘Seurat’ R package, to accurately perform spatial mapping of single cells from tissues and generate a transcriptome-wide map of spatial patterning (entire document, abstract for example).
Therefore, a skilled artisan, considering McGinnis in light of Satija, would possess all the necessary tools and knowledge, with a reasonable expectation of success to obtain spatial information using barcoded RNA sequencing data and the data analysis tools disclosed in the references.
B) Regarding claims 79-80, McGinnis teaches wherein the plurality of lipid-conjugated DNA oligonucleotides comprise:
(i) a first lipid-conjugated DNA oligonucleotide comprising the first lipid moiety, the barcode region, the capture sequence, a first hybridization region and a first primer region (Fig. 1a, longer lipid-conjugated oligo complex marked with R1, hybridized to barcode oligo); and
(ii) a second lipid-conjugated DNA oligonucleotide comprising a second lipid moiety, a second second hybridization region, wherein the second hybridization region is the reverse complement of the first primer region (Fig. 1a, shorter lipid-conjugated oligo marked with R2).
Regarding claims 83-84, McGinnis teaches Formula I and Formula II by disclosing that Anchor LMO and co-anchor LMO were synthesized as described in Weber (page 627, left-hand col, para 1).
Weber specifically teaches the structures in Formula I and II as anchor and co-anchor oligo strands in Scheme 1.
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Regarding claim 96, it is obvious in view of the teachings in McGinnis because it is not required by the claimed method.
MPEP§ 2111.04 states: "Claim scope is not limited by claim language that suggests or makes optional but does not require steps to be performed, or by claim language that does not limit a claim to a particular structure."
Claim 96 depends from claim 52 and recites a wherein clause further describing the barcode region and capture sequence. However, this wherein clause does not distinguish the claims from the prior art. This is because part (z) in claim 52, which introduces barcode region and capture sequence, is not a required element in every embodiment of the claim but is instead presented as an alternative structure for the oligonucleotide.
Regarding claim 97, McGinnis teaches wherein the barcode region comprises 18, 19, 20, 21, 22, 23, or 24 nucleotides (page 628, left-hand col, para 1-3, “The sample barcode was designed to have three components (as in Stoeckius et al.52): (1) a 5′ PCR handle for barcode amplification and library preparation, (2) an 8-bp barcode with Hamming distance greater than three relative to all other utilized barcodes and (3) a 30-bp poly-A tail necessary for hybridization to the oligo-dT region of mRNA capture bead oligonucleotides”) and the capture sequence comprises 18, 19, 20, 21, 22, 23, or 24 nucleotides ( page 628, left-hand col, para 1-3, 30-bp poly-A tail).
Double Patenting- Obvious Type -- New Grounds
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.
Claim 78 is provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claims 1 and 5 of copending Application No. 18/762,349 (reference application). Although the claims at issue are not identical, they are not patentably distinct from each other because the instant claims are anticipated by the claims (claims - 07/02/2024) of the '349 application.
Instant claim 78 recites:
A method of identifying a spatial expression pattern of a nucleic acid within a tissue of a subject, the method comprising:
(a) partitioning one or a plurality of cells from a sample corresponding to a region of the tissue into one of a plurality of vessels (‘349 Application, claim 1);
(b) exposing the one or plurality of cells with a plurality of lipid-conjugated DNA oligonucleotides comprising a first lipid moiety, a barcode region, and a capture sequence to embed the lipid-conjugated DNA oligonucleotides within cell membrane of the one or plurality of cells,
wherein the barcode region of each the plurality of lipid-conjugated DNA oligonucleotides is unique for each of the one of plurality of vessels in which the one or plurality of cells are exposed (‘349 Application, claim 1 and 5);
(c) sequencing nucleic acids captured by the capture sequence of the lipid-conjugated DNA oligonucleotides (‘349 Application, claim 1); and
(d) correlating the sequenced nucleic acids from the one or plurality of cells to the spatial position of the one or plurality of cells within the tissue and/or relative to the tissue according to the barcode region contained in each of the sequenced nucleic acids (‘349 Application, claim 1 and 5).
Therefore, instant claim 78 is anticipated by claims 1 and 5 of the ‘349 application.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Claim 78 is provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claims 12, 17 and 20 of copending Application No. 17/057,569 (reference application). Although the claims at issue are not identical, they are not patentably distinct from each other because the instant claims are anticipated by the claims (Amended Claims - 07/23/2025) of the '569 application.
Instant claim 78 recites:
A method of identifying a spatial expression pattern of a nucleic acid within a tissue of a subject, the method comprising:
(a) partitioning one or a plurality of cells from a sample corresponding to a region of the tissue into one of a plurality of vessels (‘569 Application, claim 12);
(b) exposing the one or plurality of cells with a plurality of lipid-conjugated DNA oligonucleotides comprising a first lipid moiety, a barcode region, and a capture sequence to embed the lipid-conjugated DNA oligonucleotides within cell membrane of the one or plurality of cells (‘569 Application, claim 12 and 20),
wherein the barcode region of each the plurality of lipid-conjugated DNA oligonucleotides is unique for each of the one of plurality of vessels in which the one or plurality of cells are exposed (‘569 Application, claim 12);
(c) sequencing nucleic acids captured by the capture sequence of the lipid-conjugated DNA oligonucleotides (‘569 Application, claim 17); and
(d) correlating the sequenced nucleic acids from the one or plurality of cells to the spatial position of the one or plurality of cells within the tissue and/or relative to the tissue according to the barcode region contained in each of the sequenced nucleic acids (‘569 Application, claim 12).
Therefore, instant claim 78 is anticipated by claims 12, 17 and 20 of the ‘569 application.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Prior Art
Other prior art also teach cell labeling using lipid conjugated oligonucleotides:
Gartner (US20140294782A1 - Reprogramming of Cellular Adhesion; Published on 2014-10-02);
McGinnis (McGinnis, C. S. et al. MULTI-seq: scalable sample multiplexing for single-cell RNA sequencing using lipid-tagged indices 1–34 (2018). doi: https://doi.org/10.1101/387241; Published August 08, 2018).
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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/TIAN NMN YU/Examiner , Art Unit 1681 /AARON A PRIEST/Primary Examiner, Art Unit 1681
1 Fahy et al. A comprehensive classification system for lipids. J Lipid Res. 2005 May;46(5):839-61. doi: 10.1194/jlr.E400004-JLR200. Epub 2005 Feb 16
2 Gartner et al. (US20140294782A1 - Reprogramming of Cellular Adhesion; Published on 2014-10-02; hereinafter "Gartner")
3 However, Applicant's suggestion regarding lack of enablement is noted. If Applicant wishes to pursue this line of argument, the Examiner is willing to consider it ꟷ provided that Applicant can further elaborate why a person of ordinary skill in the art would not be able to make and use the full scope of the claimed method without undue experimentation.
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