DETAILED ACTION
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Election/Restrictions
Applicant’s election without traverse of Group I, a solid support and a composition comprising a solid support, in the reply filed on 5/19/25 is acknowledged.
Claims 1-25 and 54 are pending. Claims 10-11 and 25 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim. Election was made without traverse in the reply filed on 5/19/25.
Claims 1-9, 12-24, and 54 are currently under examination.
Priority
The instant application 17/768,700 filed on 4/13/22 is a 371 US national phase of PCT/US20/58828 filed on 11/4/20, and claims domestic priority to provisional application 62/933,040 filed on 11/8/19. The priority date is determined to be 11/8/19.
Claim Rejections - 35 USC § 112 – Indefiniteness
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claim 22 is 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.
Claim 22 recites the limitation "the grid" in line 1. There is insufficient antecedent basis for this limitation in the claim. For purposes of compact prosecution, this claim is interpreted as depending from claim 16 instead of currently depending from claim 12.
Claim Rejections - 35 USC § 103
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
Claims 1, 5, 12-13, 15-24, and 54 are rejected under 35 U.S.C. 103 as being unpatentable over Fan et al. (2016; US 2016/0289669 A1; USPGPub citation 2 in IDS filed on 4/13/22; publication of US application 15/004,618, which claims the priority of provisional application 62/106,680 filed on 1/22/15, provisional application 62/121,361 filed on 2/26/15, and provisional application 62/217,274 filed on 9/11/15).
Fan et al. teaches “Devices and systems for molecular barcoding of nucleic acid targets in single cells” (Title).
Relevant to claim 1, Fan et al. teaches “Disclosed herein are methods, compositions, devices, systems, and kits for molecular barcoding of a plurality of target molecules in single cells” (page 10, paragraph 0153). Fan et al. teaches “Stochastic Labels May be Attached to Solid Supports:… In this embodiment, the stochastic labels comprise a plurality of 5’-amine modified oligonucleotides attached to a bead. The oligonucleotides comprise a 5’ amine group, a universal primer, a cellular label, a molecular label, and a target binding region… In some embodiments, the stochastic label oligonucleotide sequences may be attached to a solid support at their 5’ end” (page 12, paragraphs 0177-0178).
These teachings read on claim 1 A composition comprising: a) a solid support; b) a plurality of tethered oligonucleotides, wherein the tethered oligonucleotides are attached to the solid support via the 5’ termini of the oligonucleotides.
Further relevant to claim 1, Fan et al. teaches that “In some embodiments, each single bead of a plurality of beads contained within the at least 100 microwells comprises a plurality of tethered stochastic labels capable of attaching to a target nucleic acid molecule in a stochastic manner” (page 2, paragraph 0014). Fan et al. teaches that “In some embodiments, the plurality of microwells are distributed across a substrate in an ordered pattern, e.g. an ordered array” (page 19, paragraph 0238).
These teachings read on claim 1 in a patterned array.
Further relevant to claim 1, Fan et al. teaches “Attachment of Stochastic Labels to Target Nucleic Acid Molecules” (page 16, paragraph 0213) and that “the target binding region may comprise a nucleic acid sequence that is capable of specific hybridization to a restriction site overhang (e.g. an EcoRI sticky-end overhang)” (page 17, paragraph 0215).
These teachings read on claim 1 wherein each of the plurality of tethered oligonucleotides comprises a nucleotide sequence that, when hybridized to a complementary nucleotide sequence present in an untethered oligonucleotide, generates an enzyme cleavage site; and c) a plurality of untethered oligonucleotides that are hybridized to the plurality of tethered oligonucleotides in the patterned array, wherein the untethered oligonucleotides each comprise: i) the nucleotide sequence that generates an enzyme cleavage site, wherein the enzyme cleavage site is a restriction enzyme cleavage site…
Further relevant to claim 1, Fan et al. teaches that “In some embodiments of the disclosed methods, one or more nucleic acid amplification reactions may be performed to create multiple copies of the labeled target nucleic acid molecules… The amplification reactions may comprise amplifying at least a portion of the cellular and or molecular label. The amplification reactions may comprise amplifying at least a portion of a sample tag, a cellular label, a molecular label, a target nucleic acid, or a combination thereof” (page 17, paragraph 0220). Fan et al. teaches “A set of nucleic acid targets… may be chosen to identify, for example, cells undergoing apoptosis… rapid proliferation… or any other subpopulation of cells that can be defined on the basis of nucleic acid markers” (page 18, paragraph 0228).
These teachings read on claim 1 ii)… an effector molecule that affects an activity and/or a phenotype of a cell, wherein… the effector molecule is attached to the untethered oligonucleotide at the 5’ end of the untethered oligonucleotides.
Further relevant to claim 1, Fan et al. teaches that “individual beads are uniquely identified both by an optical code (e.g… by synthesis of an attached optical code through the use of solid-phase split-pool synthesis methodologies and a set of spectrally-distinct fluorescent building blocks)” (page 16, paragraph 0206).
This teaching reads on claim 1 iii) a fluorophore.
Relevant to claim 5, Fan et al. teaches “Attachment of Stochastic Labels to Target Nucleic Acid Molecules” (page 16, paragraph 0213) and that “the target binding region may comprise a nucleic acid sequence that is capable of specific hybridization to a restriction site overhang (e.g. an EcoRI sticky-end overhang)” (page 17, paragraph 0215).
This teaching reads on claim 5 the enzyme cleavage site is: a) a restriction enzyme cleavage site.
Relevant to claim 12, Fan et al. teaches that “Pre-synthesized stochastic labels may be attached to beads or other solid supports through any of a variety of immobilization techniques involving functional group pairs on the solid support and the oligonucleotide. In some embodiments, the oligonucleotide functional group and the solid support functional group are individually selected from the group consisting of… primary amine(s)… aldehyde(s)… A stochastic label oligonucleotide may be tethered to a solid support, for example, by coupling… a 5’ amino group on the oligonucleotide to the carboxyl group of the functionalized solid support” (page 14, paragraph 0194).
This teaching reads on claim 12 A solid support comprising: a) one or more patterns exposing an aldehyde-reactive substrate; b) one or more tethered oligonucleotides covalently attached to the exposed aldehyde-reactive substrate within the one or more patterns via an amine-modified terminus at the 5' end of the one or more tethered oligonucleotides.
Relevant to claims 12, 13, and 15, Fan et al. teaches “Attachment of Stochastic Labels to Target Nucleic Acid Molecules” (page 16, paragraph 0213) and that “the target binding region may comprise a nucleic acid sequence that is capable of specific hybridization to a restriction site overhang (e.g. an EcoRI sticky-end overhang)” (page 17, paragraph 0215).
These teachings read on claim 12 wherein the one or more tethered oligonucleotides comprises a nucleotide sequence that, when hybridized to a complementary nucleotide sequence present in an untethered oligonucleotide, generates an enzyme cleavage site; claim 13 the enzyme cleavage site is: a) a restriction enzyme cleavage site; and claim 15 the one or more tethered oligonucleotides comprises a nucleotide sequence that hybridizes to a complementary nucleotide sequence present in an untethered oligonucleotide, generates an enzyme cleavage site.
Further relevant to claim 15, Fan et al. teaches that “In some embodiments of the disclosed methods, one or more nucleic acid amplification reactions may be performed to create multiple copies of the labeled target nucleic acid molecules… The amplification reactions may comprise amplifying at least a portion of the cellular and or molecular label. The amplification reactions may comprise amplifying at least a portion of a sample tag, a cellular label, a molecular label, a target nucleic acid, or a combination thereof” (page 17, paragraph 0220). Fan et al. teaches “A set of nucleic acid targets… may be chosen to identify, for example, cells undergoing apoptosis… rapid proliferation… or any other subpopulation of cells that can be defined on the basis of nucleic acid markers” (page 18, paragraph 0228).
These teachings read on claim 15 the untethered oligonucleotide comprises an effector molecule bound to the 5' end of the untethered oligonucleotide.
Relevant to claim 16, Fan et al. teaches that “In some embodiments, the plurality of microwells are distributed across a substrate in an ordered pattern, e.g. an ordered array… In one embodiment of the present disclosure, the plurality of microwells (e.g. a microwell array) is a consumable component of the assay system” (page 19, paragraph 0238).
This teaching reads on claim 16 the aldehyde-reactive substrate further comprises a grid that surrounds the one or more patterns.
Relevant to claim 17, Fan et al. teaches “In many embodiments of the automated assay system, the microwell substrate, with or without an attached flow cell, will be packaged within a consumable cartridge that interfaces with the instrument system and which may incorporate additional functionality… Design features of cartridges may include… (xi) optical alignment marks for determining the position of the cartridge or wells within the instrument” (page 27, paragraph 0308).
This teaching reads on claim 17 wherein the aldehyde-reactive substrate further comprises one or more alignment markers.
Relevant to claim 18, as seen in Fan et al. Fig. 1, the stochastic label contains “Universal”, “Cell Label”, “molecular label”, and “oligodT (17) V” regions. Fan et al. teaches nucleotide length limitations for the universal label (page 12, paragraph 0180), cellular label (page 13, paragraph 0182), molecular label (page 13, paragraph 0188).
Taken together, these numerical ranges for the stochastic label (or tethered oligonucleotides) read on claim 18 the one or more tethered oligonucleotides has a length of from 20 nucleotides to 50 nucleotides
Relevant to claims 19-20, Fan et al. page 27, paragraph 0313 continued through page 28, paragraph 0314 teaches that “the dimensions of fluid channels and the microwell chamber(s) in cartridge designs will be optimized to (i) provide uniform delivery of cells and beads to the microwells, and (ii) to minimize sample and reagent consumption” and provides dimensional limitations.
These teachings read on the diameter limitations recited in claims 19-20.
Relevant to claim 21, absent a limiting definition of micro-islands within the instant specification, the broadest reasonable interpretation of the recited limitations is read upon by Fan et al. Figs. 2A-B and associated description of “FIGS. 2A-B illustrate the loading of cells and beads into microwell arrays. FIG. 2A shows micrographs of a microwell array after loading with a dilute cell suspension (upper) and after loading with a bead suspension (lower). FIG. 2B shows magnified views of two microwells that contain both a single cell and a single bead” (page 6, paragraph 0036).
These teachings read on claim 21 the one or more patterns comprises one or more micro-islands.
Relevant to claim 22, Fan et al. teaches “Cartridge components may be fabricated using any of a number of suitable materials, including but not limited to silicon, fused-silica, glass, any of a variety of polymers” (page 28, paragraph 0317).
This teaching reads on claim 22 the grid is a polyacrylamide grid.
Relevant to claim 23, Fan et al. teaches “In many embodiments of the automated assay system, the microwell substrate will be packaged within a flow cell (FIG. 12) that provides for convenient interfacing with the rest of the fluid handling system and facilitate the exchange of fluids, e.g. cell and bead suspensions, lysis buffers, rinse buffers, etc., that are delivered to the microwells. In many embodiments, the flow cell may be designed to facilitate uniform distribution of cells and beads across the plurality of microwells” (page 25, paragraph 0292).
Relevant to claim 24, Fan et al. teaches “In some embodiments, the flow cell may constitute a fixed component of the instrument system. In some embodiments, the flow cell may be removable from the instrument system. In some embodiments, the flow cell may be a single use device. In other embodiments, the flow cell may be a multi-use device” (page 26, paragraph 0296). Embodiments are seen in Fan et al. Figs. 12 and 14.
These teachings read on claim 24 the one or more flow cells is positioned over the plurality of tethered oligonucleotides.
Relevant to claim 54, Fan et al. teaches that “Pre-synthesized stochastic labels may be attached to beads or other solid supports through any of a variety of immobilization techniques involving functional group pairs on the solid support and the oligonucleotide. In some embodiments, the oligonucleotide functional group and the solid support functional group are individually selected from the group consisting of… primary amine(s)… aldehyde(s)… A stochastic label oligonucleotide may be tethered to a solid support, for example, by coupling… a 5’ amino group on the oligonucleotide to the carboxyl group of the functionalized solid support” (page 14, paragraph 0194).
This teaching reads on claim 54 A solid support comprising: a) one or more patterns exposing an aldehyde-reactive substrate; b) one or more tethered oligonucleotides, covalently attached to the one or more patterns via an amine-modified termini at the 5' end of the plurality of tethered oligonucleotides.
Further relevant to claim 54, Fan et al. teaches “Attachment of Stochastic Labels to Target Nucleic Acid Molecules” (page 16, paragraph 0213) and that “the target binding region may comprise a nucleic acid sequence that is capable of specific hybridization to a restriction site overhang (e.g. an EcoRI sticky-end overhang)” (page 17, paragraph 0215).
These teachings read on claim 54 wherein the one or more of tethered oligonucleotides comprises a nucleotide sequence that, when hybridized to a complementary nucleotide sequence present in an untethered oligonucleotide, generates a restriction enzyme cleavage site.
Further relevant to claim 54, Fan et al. teaches “In some embodiments, the downstream analysis of sequence data may be focused on selected subpopulations of cells by performing an amplification using one or more target-specific primers, wherein the one or more target-specific primers are capable of specific hybridization with, for example, one or more genes or gene products that define a subpopulation of cells. A set of nucleic acid targets… may be chosen to identify, for example, cells undergoing apoptosis… rapid proliferation… or any other subpopulations of cells that can be defined on the basis of nucleic acid markers” (page 18, paragraph 0228).
This teaching reads on claim 54 c) one or more untethered oligonucleotides.
Fan et al. does not teach a specific embodiment having all the claimed elements. That being said, however, it must be remembered that "[w]hen a patent simply arranges old elements with each performing the same function it had been known to perform and yields no more than one would expect from such an arrangement, the combination is obvious." KSR v. Teleflex, 127 S.Ct. 1727, 1740 (2007) (quoting Sakraida v. AG. Pro, 425 U.S. 273, 282 (1976)). "[W]hen the question is whether a patent claiming the combination of elements of prior art is obvious," the relevant question is "whether the improvement is more than the predictable use of prior art elements according to their established functions." (Id.). Addressing the issue of obviousness, the Supreme Court noted that the analysis under 35 USC 103 "need not seek out precise teachings directed to the specific subject matter of the challenged claim, for a court can take account of the inferences and creative steps that a person of ordinary skill in the art would employ." KSR at 1741. The Court emphasized that "[a] person of ordinary skill is... a person of ordinary creativity, not an automaton." Id. At 1742.
Consistent with this reasoning, it would have been prima facie obvious to have
selected various combinations of various disclosed elements — including tethered/untethered oligonucleotides, restriction enzyme cleavage sites, and substrates — for a solid support composition, to arrive at compositions "yielding no more than one would expect from such an arrangement."
Claims 2-4, 6-9, and 14 are rejected under 35 U.S.C. 103 as being unpatentable over Fan et al. (2016; US 2016/0289669 A1; USPGPub citation 2 in IDS filed on 4/13/22; publication of US application 15/004,618, which claims the priority of provisional application 62/106,680 filed on 1/22/15, provisional application 62/121,361 filed on 2/26/15, and provisional application 62/217,274 filed on 9/11/15), as applied to claims 1, 5, 12-13, 15-24, and 54 above, and further in view of Hsiao et al. (2016; US 9,327,036 B2).
The teachings of Fan et al. are applied to instantly rejected claims 2-4, 6-9, and 14 as they were previously applied to claims 1, 5, 12-13, 15-24, and 54 as rendering obvious a solid support composition.
Fan et al. is silent to specifics regarding effector molecule limitations (claims 2-4, and 9) and bound target cells (claims 6-8 and 14). However, these limitations were known in the prior art and taught by Hsiao et al.
Hsiao et al. teaches “DNA-cell conjugates” (Title).
Relevant to claims 2-4 and 9, Hsiao et al. teaches “In general, any affinity molecule useful in the prior art, in combination with a known ligand to provide specific recognition of a detectable substance will find utility in the attachment of nucleic acid groups of the invention. Examples of such biological molecules which can then be attached to these functional groups include linker molecules having a known binding partner, or affinity molecule, include but are not limited to, polysaccharides, lectins, selectins, nucleic acids (both monomeric and oligomeric), proteins, enzymes, lipids, antibodies, and small molecules such as sugars, peptides, aptamers, drugs, and ligands” (column 19, lines 50-60).
This teaching reads on claim 2 the effector molecule is a polypeptide; claim 3 the effector molecule comprises a lipid; claim 4 the effector molecule comprises an oligosaccharide; and claim 9 the effector molecule is…
Relevant claims 6-7, Fan et al. teaches that “In some embodiments, each single bead of a plurality of beads contained within the at least 100 microwells comprises a plurality of tethered stochastic labels capable of attaching to a target nucleic acid molecule in a stochastic manner… In some embodiments, the plurality of tethered stochastic labels attached to a single bead further comprises a diverse set of molecular labels...” (page 2, paragraph 0014).
This teaching reads on claim 6 the composition comprises at least a first, a second, and a third plurality of hybridized, untethered oligonucleotides that are bound to the tethered oligonucleotides in the patterned array, wherein: a) the first plurality of hybridized, untethered oligonucleotides comprises, bound to the 5' end of the oligonucleotides, a first effector molecule, wherein the first plurality of hybridized, untethered oligonucleotides generates a first enzyme cleavage site; and b) the second plurality of hybridized, untethered oligonucleotides comprises, bound to the 5' end of the oligonucleotides, a second effector molecule, wherein the second plurality of hybridized, untethered oligonucleotides generates a second enzyme cleavage site; and claim 7 further comprising a fourth plurality of hybridized, untethered oligonucleotides, wherein the fourth plurality of hybridized, untethered oligonucleotides bound to the 5' end of the oligonucleotides, a third effector molecule, wherein the fourth plurality of hybridized, untethered oligonucleotides generates a third enzyme cleavage site.
Relevant to claims 6 and 14, Hsiao et al. column 13, line 43 onwards teaches “III. Cell-DNA Conjugates”, wherein “In some embodiments, the present invention provides a conjugate of a cell and a nucleic acid moiety, wherein the cell has a surface including a native functional group, the cell has no cell wall, and the nucleic acid moiety is covalently linked to the native functional group” (column 13, lines 56-60). Hsiao et al. teaches that “The nucleic acid moiety can be any suitable nucleic acid moiety having a nucleic acid or nucleotide. Exemplary nucleic acid moieties include, but are not limited to, an oligonucleotide…” (column 16, lines 23-26).
These teachings read on claim 6 c) the third plurality of hybridized, untethered oligonucleotides comprises, bound to the 5’ end of the oligonucleotides, a target cell; and claim 14 the untethered oligonucleotide comprises a target cell bound to the 5’ end of the untethered oligonucleotide.
Relevant claim 8, Hsiao et al. teaches that “In some embodiments, the cell is a primary cell. In other embodiments, the cell is a mammalian cell. In some other embodiments, the cell is a stem cell” (column 16, lines 12-14).
This teaching reads on claim 8 the cell is a stem cell or a progenitor cell.
Although Fan et al. does not explicitly teach the Hsiao et al. oligonucleotide-bound target cells, it would have been prima facie obvious to the skilled artisan. Fan et al. and Hsiao et al. are analogous disclosures in the field of oligonucleotide tethered moieties.
The skilled artisan would be motivated to combine the analogous art. Hsiao et al. teaches that “This procedure can be carried out in some embodiments in less than 1 hour, and leads to equivalent levels of cell surface functionalization with any oligonucleotide sequence of interest. The present method can be applied to various embodiments including, the capture of single cells for RT-PCR analysis, the attachment of living cells to a solid substrate for force measurement or cell patterning techniques. In the examples, we demonstrate the use of this new labeling method for the capture of red blood cells, primary T-cells, and myoblasts, which are all types of cells that are difficult to pattern using other methods. This new technique greatly expands the scope of the DNA-based adhesion strategy and is sufficiently straightforward to be used in labs that do not specialize in organic synthesis” (column 10, lines 7-20).
Thus, the skilled artisan would be motivated to include the Hsiao et al. oligonucleotide-bound target cells because Hsiao et al. teaches that their methodology “greatly expands the scope of the DNA-based adhesion strategy” and is straightforward and applicable to various embodiments. The skilled artisan would have a reasonable expectation of success based on the disclosures of Fan et al., and further in view of Hsiao et al.
Conclusion
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/SARAH JANE KENNEDY/Examiner, Art Unit 1682
/WU CHENG W SHEN/Supervisory Patent Examiner, Art Unit 1682