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 .
Claims Status
Claims 1-40 are canceled.
Claims 41-60 are pending and currently under examination
Priority
This application is a National Stage Application under 35 U.S.C. 371 and claims the
benefit of International Application No. PCT/US2021/065340, filed December 28, 2021, which
claims priority to U.S. Provisional Patent Application No. 63/132,112, filed on December 30, 2020 and U.S. Provisional Patent Application No. 63/146,815, filed on February 8, 2021.The priority date of claim set filed on June 26, 2023, is determined to be December 30, 2020.
Claim Objections
Claim 41 is objected to because of the following informalities: Claim 41 lacks a preamble comprising a general description of all the elements or steps of the claimed combination which are conventional or known. See MPEP 608.01(i).
Appropriate correction is required.
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 41-49 and 51-59 are rejected under 35 U.S.C. 103 as being unpatentable over Frisen et al. (“Frisen”; Patent App. Pub. No. WO 2012140224 A1, Oct. 18, 2012).
Frisen discloses “The present invention relates to methods and products for the localised or spatial detection of nucleic acid in a tissue sample and in particular to a method for localised detection of nucleic acid in a tissue sample comprising: (a) providing an array comprising a substrate on which multiple species of capture probes are directly or indirectly immobilized such that each species occupies a distinct position on the array and is oriented to have a free 3' end to enable said probe to function as a primer for a primer extension or ligation reaction, wherein each species of said capture probe comprises a nucleic acid molecule with 5' to 3': (i) a positional domain that corresponds to the position of the capture probe on the array, and (ii) a capture domain; (b) contacting said array with a tissue sample such that the position of a capture probe on the array may be correlated with a position in the tissue sample and allowing nucleic acid of the tissue sample to hybridize to the capture domain in said capture probes; (c) generating DNA molecules from the captured nucleic acid molecules using said capture probes as extension or ligation primers, wherein said extended or ligated DNA molecules are tagged by virtue of the positional domain; (d) optionally generating a complementary strand of said tagged DNA and/or optionally amplifying said tagged DNA; (e) releasing at least part of the tagged DNA molecules and/or their complements or amplicons from the surface of the array, wherein said part includes the positional domain or a complement thereof; and (f) directly or indirectly analyzing the sequence of the released DNA molecules.” (Abstract).
Regarding claim 41, Frisen teaches a method comprising “localized detection of nucleic acid in a tissue sample comprising:(a) providing an array comprising a substrate on which multiple species of capture probes … is oriented to have a free 3' end to enable said probe to function as a primer for a primer extension … wherein each species of said capture probe comprises a nucleic acid molecule with 5' to 3': ...(ii) a capture domain; (b) contacting said array with a tissue sample such that the position of a capture probe on the array may be correlated with a position in the tissue sample and allowing nucleic acid of the tissue sample to hybridize to the capture domain in said capture probes; (c) generating DNA molecules from the captured nucleic acid molecules using said capture probes as extension … primers…(d) optionally generating a complementary strand of said tagged DNA … (e) releasing at least part of the tagged DNA molecules and/or their complements or amplicons from the surface of the array… (f) directly or indirectly analyzing the sequence of the released DNA molecules” (Pg. 7 ln 15-33 Pg. 8 ln 5).
Frisen teaches a method comprising “the capture probe may also be synthesized on the array substrate using polymerase extension (similarly to as described above) and a terminal transferase enzyme to add a "tail" which may constitute the capture domain. This is described further in Example 7 below. The use of terminal transferases to add nucleotide sequences to the end of an oligonucleotide is known in the art, e.g. to introduce a homopolymeric tail e.g. a poly-T tail” (Pg. 26 ln 22-27).
Frisen teaches a method comprising the second strand synthesis may utilize, or be achieved by, template switching. In a particularly preferred embodiment, the template switching reaction, i.e. the further extension of the cDNA first strand to incorporate the complementary amplification domain, is performed in situ (whilst the capture probe is still attached, directly or indirectly, to the array” (Pg. 41 ln 18-20).
Frisen teaches “the marker may be detected using the same imaging conditions used to visualize the tissue sample. In a particularly preferred embodiment of the invention, the array comprises marker features to which a labelled, preferably fluorescently labelled, marker nucleic acid molecule, e.g. oligonucleotide, is hybridized” (Pg. 35 ln 14-18).
Thus, Frisen teaches A method comprising:(a) a biological sample on an array comprising a plurality of capture probes, wherein a capture probe of the plurality of capture probes comprises a capture domain that hybridizes to an analyte in the biological sample; (b) extending a 3' end of the capture probe using the analyte as a template to generate an extended capture probe; (c) adding to a 3' end of the extended capture probe a first homopolymeric sequence; (d) contacting the extended capture probe with a template switching oligonucleotide comprising a strand of nucleic acid in a 5' to 3' direction: a primer binding sequence and a second homopolymeric sequence, where the second homopolymeric sequence hybridizes to the first homopolymeric sequence; (e) extending the 3' end of the extended capture probe using the template switching oligonucleotide as a template thereby generating an extended capture probe comprising a template-switching oligonucleotide sequence; (f) removing the analyte and the template-switching oligonucleotide from the extended capture probe comprising the template-switching oligonucleotide sequence; (g) hybridizing one or more labeled probes complementary to all or a portion of the template-switching oligonucleotide sequence of the extended capture probe comprising the template-switching oligonucleotide sequence; and (h) detecting the one or more labeled probes hybridized to the template-switching oligonucleotide sequence of the extended capture probe comprising the template-switching oligonucleotide sequence.
The teachings of Frisen are documented above in the rejection of claims 41 under 35 U.S.C. 103. Claims 42, 43, 44, 45, 46, 47, 48, 49, 51, 52, 56, 57, 58, 59 depends on claim 41 Claim 55 depends on claim 54. Claims 53 and 54 depend on claim 52, which depends on claim 41.
Regarding claim 42, “Figure 5 shows a fluorescent image captured after 99°C water mediated release of DNA surface probes from commercial arrays manufactured by Agilent. A fluorescent detection probe was hybridized after hot water treatment. Top array is an untreated control.” (Pg. 67 ln 23-25). “control” reads on a set of conditions. Thus, Frisen teaches a method wherein steps (a)-(h) are performed under a first set of conditions, and the method further comprises repeating steps (a)-(h) under a second set of conditions.
Regarding claim 43, Frisen teaches a method further comprising “the tissue sample is stained prior to visualization to provide contrast between the different regions, e.g. cells, of the tissue sample.” (Pg. 35 ln 21-23). Frisen also teaches a method further comprising “Hematoxylin” (Pg. 76 ln 24) and “Eosin” (Pg. 76 ln 27). Thus, Frisen teaches a method further comprising staining the biological sample, and optionally, wherein the staining comprises hematoxylin and eosin.
Regarding claim 44, Frisen teaches a method further comprising “The step of imaging the tissue may use any convenient histological means known in the art, e.g. light, bright field, dark field, phase contrast, fluorescence, reflection, interference, confocal microscopy or a combination thereof.” (Pg. 35 ln 19-21). Thus, Frisen teaches a method further comprising imaging the biological sample, wherein the imaging comprises one or more of light field microscopy, bright field microscopy, dark field microscopy, phase contrast microscopy, fluorescence microscopy, reflection microscopy, interference microscopy, and confocal microscopy.
Regarding claim 45, Frisen teaches a method wherein “arrays comprising "capture probes" for capturing and labelling transcripts from all of the single cells within a tissue sample …The resultant data may be correlated to images of the original tissue samples e.g. sections through so-called barcode sequences (or ID tags, defined herein as positional domains) incorporated into the arrayed nucleic acid probes.” (Pg. 12 ln 26-34 and Pg.13 ln 1-2). Thus, Frisen teaches a method wherein the capture probe of the plurality of capture probes further comprises a spatial barcode.
Regarding claim 46, Frisen teaches a method wherein “the capture probe comprises a cleavage domain ” (Pg. 23 ln 11-12). Thus, Frisen teaches a method wherein the capture probe of the plurality of capture probes further comprises one or more functional domains, a unique molecular identifier, a cleavage domain, and combinations thereof.
Regarding claim 47, Frisen teaches a method wherein “fluorescently labelled probes” (Pg. 67 ln 20-21). Thus, Frisen teaches a method wherein a labeled probe of the one or more labeled probes comprises a fluorescent label.
Regarding claim 48, Frisen teaches a method wherein “The capture probe may also be synthesised on the array substrate using polymerase extension (similarly to as described above) and a terminal transferase enzyme to add a "tail" which may constitute the capture domain. This is described further in Example 7 below. The use of terminal transferases to add nucleotide sequences to the end of an oligonucleotide is known in the art, e.g. to introduce a homopolymeric tail e.g. a poly-T tail.” (Pg. 26 ln 22-27). Thus, Frisen teaches a method wherein adding to the 3' end of the extended capture probe the first homopolymeric sequence in step (c) is performed using a terminal deoxynucleotidyl transferase.
Regarding claim 49, Frisen teaches a method wherein “Excess oligonucleotides may be removed by washing the array under standard hybridization conditions.” (Pg. 26 ln 30-31) and “washing step directly following cDNA synthesis” ( Pg. 77 ln 13-14). Thus, Frisen teaches a method wherein removing the analyte and the template-switching oligonucleotide from the extended capture probe comprising the template-switching oligonucleotide in step (f) comprises one or more washing steps.
Regarding claim 51, Frisen teaches a method wherein “High density nucleic acid arrays or microarrays are a core component of the spatial transcriptome labelling method described herein… A typical microarray consists of an arrayed series of microscopic spots of oligonucleotides” (Pg. 13 ln 3-5). Frisen also teaches a method wherein “Such arrays may also be used as the array surface in the context of the present invention e.g. an lllumina bead array.” (Pg. 15 ln 32-33). Thus, Frisen teaches a method wherein the array comprises one or more features selected from the group consisting of: a bead, a well, and a spot.
Regarding claim 52, Frisen teaches a method wherein “tissue sample” (Pg. 1 ln 5) and “tissue sections” (Pg. 5 ln 33). Thus, Frisen teaches a method wherein the biological sample is a tissue sample or a tissue section.
Regarding claim 53-55, Frisen teaches a method wherein “the tissue may be prepared using standard methods of formalin-fixation and paraffin-embedding (FFPE), which are well-established in the art. Following fixation of the tissue sample and embedding in a paraffin or resin block, the tissue samples may be sectioned” (Pg. 30 ln 9-12). Thus, Frisen teaches a method wherein the tissue sample is a fixed tissue sample; wherein the tissue section is a fixed tissue section; and wherein the fixed tissue section is a formalin-fixed paraffin- embedded tissue section.
Regarding claim 56-57, Frisen teaches a method wherein “array technology coupled with high throughput DNA sequencing technologies, which allows the nucleic acid molecule (e.g. RNA or DNA molecules) in the tissue sample, particularly mRNA or DNA, to be captured and labelled with a positional tag.” (Pg. 1 ln 22-25.). Thus, Frisen teaches a method wherein the analyte is DNA; and wherein the analyte is mRNA.
Regarding claim 58, Frisen teaches a method wherein “capture domain comprises a poly-T DNA oligonucleotide” (Pg. 18 ln 26-27; Fig. 1.). Thus, Frisen teaches a method wherein the capture domain comprises a poly(T) sequence.
Regarding claim 59, Frisen teaches a method wherein “terminal transferases to add nucleotide sequences to the end of an oligonucleotide is known in the art, e.g. to introduce a homopolymeric tail e.g. a poly-T tail. Accordingly, in such a synthesis an oligonucleotide that corresponds to the universal domain of the capture probe may be contacted with the array and allowed to hybridize to the complementary domain of the surface probes” (Pg. 26 ln 25-30; Pg. 61 ln 12-13). “homopolymeric tail” reads on a poly(C) or poly (G) sequence. Thus, Frisen teaches a method wherein the first homopolymeric sequence is a poly(C) sequence and the second homopolymeric sequence is a poly(G) sequence.
Therefore, the invention as recited in claims 41-49 and 51-59 is prima facie obvious over the prior art Frisen et al. One of ordinary skill in the art would have had a reasonable expectation of success given the lack of novelty. It would have been obvious to provide a method according to the limitations of the instant application claims 41-49 and 51-59 based on Frisen et al. (Patent App. Pub. No. WO 2012140224 A1).
Claim 41 and 49-50 are rejected under 35 U.S.C. 103 as being unpatentable over Frisen et al. (“Frisen”; Patent App. Pub. No. WO 2012140224 A1, Oct. 18, 2012) in view of Jacobson et al. (“Jacobson”; Patent App. Pub. No. US 20130244884 A1, Sept. 19, 2023).
The teachings of Frisen are documented above in the rejection of claims 41-49 and 51-59 under 35 U.S.C. 103. Claim 50 depends on claim 49, which depends on claim 41. Frisen does not explicitly teach the limitations of claim 50.
Jacobson discloses “Methods of obtaining sequence information about target polynucleotide having a predefined sequence are disclosed. The methods include sequencing by ligation and sequencing by polymerase.” (Abstract)
Regarding claim 50, Frisen teaches a method wherein “regenerated under alkali conditions for example 0.3 M NaOH or KOH” (Para. 46). “regenerated” reads on washing step. Thus, Frisen teaches a method wherein the one or more washing steps comprises potassium hydroxide.
Frisen and Jacobson are both considered to be analogous to the claimed invention because they are in the same field of detection of labelled probes. Therefore, it would have been obvious to someone of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method as taught by Frisen according to the limitations of claims 41 and 49 to incorporate the method of washing comprising KOH as taught by Jacobson and provide a washing step comprising KOH. Doing so would provide a stringent wash and/ or regeneration.
Claim 60 is rejected under 35 U.S.C. 103 as being unpatentable over
Frisen et al. (“Frisen”; Patent App. Pub. No. WO 2012140224 A1, Oct. 18, 2012).
The teachings of Frisen are documented above in the rejection of claims 41-49 and 51-59 under 35 U.S.C. 103. Claim 60 is independent of claim 41.
Regarding claim 60, Frisen teaches a method comprising “localized detection of nucleic acid in a tissue sample comprising:(a) providing an array comprising a substrate on which multiple species of capture probes … is oriented to have a free 3' end to enable said probe to function as a primer for a primer extension … wherein each species of said capture probe comprises a nucleic acid molecule with 5' to 3': ...(ii) a capture domain; (b) contacting said array with a tissue sample such that the position of a capture probe on the array may be correlated with a position in the tissue sample and allowing nucleic acid of the tissue sample to hybridize to the capture domain in said capture probes; (c) generating DNA molecules from the captured nucleic acid molecules using said capture probes as extension … primers…(d) optionally generating a complementary strand of said tagged DNA … (e) releasing at least part of the tagged DNA molecules and/or their complements or amplicons from the surface of the array… (f) directly or indirectly analyzing the sequence of the released DNA molecules” (Pg. 7 ln 15-33 Pg. 8 ln 5).
Frisen teaches a method comprising “the capture probe may also be synthesized on the array substrate using polymerase extension (similarly to as described above) and a terminal transferase enzyme to add a "tail" which may constitute the capture domain. This is described further in Example 7 below. The use of terminal transferases to add nucleotide sequences to the end of an oligonucleotide is known in the art, e.g. to introduce a homopolymeric tail e.g. a poly-T tail” (Pg. 26 ln 22-27).
Frisen teaches a method comprising the second strand synthesis may utilize, or be achieved by, template switching. In a particularly preferred embodiment, the template switching reaction, i.e. the further extension of the cDNA first strand to incorporate the complementary amplification domain, is performed in situ (whilst the capture probe is still attached, directly or indirectly, to the array” (Pg. 41 ln 18-20).
Frisen teaches “the marker may be detected using the same imaging conditions used to visualize the tissue sample. In a particularly preferred embodiment of the invention, the array comprises marker features to which a labelled, preferably fluorescently labelled, marker nucleic acid molecule, e.g. oligonucleotide, is hybridized” (Pg. 35 ln 14-18).
Thus, Frisen teaches a method comprising: (a) contacting a biological sample with an array comprising a plurality of capture probes, wherein a capture probe of the plurality of capture probes comprises a capture domain that hybridizes to an analyte in the biological sample; (b) staining and imaging the biological sample; (c) extending a 3' end of the capture probe using the analyte as a template to generate an extended capture probe; (d) adding to a 3' end of the extended capture probe a first homopolymeric sequence; (e) contacting the extended capture probe with a template-switching oligonucleotide comprising in a 5' to 3' direction: a primer binding sequence and a second homopolymeric sequence, wherein the second homopolymeric sequence hybridizes to the first homopolymeric sequence; (f) extending the 3' end of the extended capture probe using the template-switching oligonucleotide as a template thereby generating an extended capture probe comprising a template-switching oligonucleotide sequence; (g) removing the analyte and the template-switching oligonucleotide from the extended capture probe comprising the template-switching oligonucleotide sequence; (h) hybridizing one or more labeled probes comprising a sequence complementary to the template-switching oligonucleotide sequence, or a portion thereof, of the extended capture probe comprising the template-switching oligonucleotide sequence; and (i) detecting the one or more labeled probes hybridized to the template-switching oligonucleotide sequence of the extended capture probe comprising the template-switching oligonucleotide sequence.
Conclusion
No claims are in condition for allowance.
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/KENDRA R VANN-OJUEKAIYE/Examiner, Art Unit 1682
/WU CHENG W SHEN/Supervisory Patent Examiner, Art Unit 1682