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 .
DETAILED ACTION
This action is in response to the reply of 29 April 2025.
Applicant’s arguments and amendments to the claims have been fully considered
but are found not persuasive.
The rejection of claims 1 and 18 under 35 U.S.C. § 112(b) or preAIA 35 U.S.C. § 112, second paragraph, for recitation of “a flat shape” has been overcome by the amendment to claims 1 and 18. The rejection of dependent claims 2-5 and 7-17 and 21 is therefore also withdrawn.
The rejection of claim 11 under 35 U.S.C. § 112(d) or pre-AIA 35 U.S.C. § 112, fourth paragraph, for failing to further limit the subject matter of the claim upon which it depends has been overcome by an amendment to delete the reference to a 28S ribosomal RNA subunit. The rejection is therefore withdrawn.
Claim Status
Claim 24 was newly added by an amendment on 29 April 2025. Subsequently, claims 1-21 and 24 are pending and currently under examination.
Specification
The objection to the Abstract has been overcome by correction of informalities. The objection is therefore withdrawn.
Drawings
Replacement drawings for figures 7 A-7D were received on 29 April 2025. These drawings are acceptable. The objection to the drawings is therefore withdrawn.
Maintained/Modified Rejections
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.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
Claims 1, 3-18, and 20-21 remain/are rejected under 35 U.S.C. 103 as being unpatentable over Woehrstein et al. (Sci. Adv., 3, 1-12, 2017, including Supplementary
Materials, page 9-30) in view of Grabmayr et al., 2019 (WO 2019/115801 Al) and Satokari et al. (Microbial Ecology, 50, 120-127, 2005).
Regarding claims 1 and 18, Woehrstein et al. teaches attaching first and second fluorophore binders to first and second DNA nanostructures by hybridization. In Woehrstein et al., the fluorophore binders are referred to as target-complementary 21-nt-long sequences and the DNA nanostructures are referred to as metafluorophores (page 7, paragraph 1 lines 6-13 and Figure 4). Woehrstein et al. teaches that the DNA nanostructures comprise parallel DNA double helices that form a rectangular shape with lengths less than 150 nm and maximum dimensions less than 300nm (abstract lines 6-10, page 2 column 2 lines 4-7 and Figure 4). Woehrstein et al. teaches that the DNA nanostructures have fluorophore combinations that result in different DNA nanostructures displaying different colors (page 4 column 2 paragraph 3, lines 4-6 and Figure 4). Woehrstein et al. teaches creating different colors for different nanostructures using a combination of 3 different colored organic fluorophores wherein first and second DNA nanostructures exhibit first and second fluorophore colors produced by a combination of intensities of the first color, second color and third color (Figure 3 and Figure 4). Further regarding claim 18, Woehrstein et al. further teaches creating different colors for different nanostructures using a combination of 2 different colors wherein first and second DNA nanostructures exhibit first and second fluorophore color produced by a combination of intensities of the first color and second colors (page 7, column 1, lines 1-5 and Figure 3).
Further regarding claims 1 and 18, Woehrstein et al. teaches a method that includes steps of selecting a first and second target molecule species, selecting a primary binding site on each target molecule species, selecting a secondary binding site on each target molecule species, and forming oligonucleotide probes that will hybridize to the first and second binding sites for each target molecule species (page 7 column 2 paragraph 1 lines 1-13 and Figure 4). The oligonucleotide probes in the reference are referred to as target-complementary 21-nt-long sequences and capture strands, and these correspond to the "immobilizing binders and the "fluorophore binders" of the instant claims.
Woehrstein et al. does not expressly use the language "selecting" and "forming" to refer to the process of identifying oligonucleotide probe binding sites, but it is inherent to the multiplex method taught by Woehrstein et al. that the target polynucleotides must be selected, and that the oligonucleotides which will hybridize to the targets must be formed (or synthesized).
Woehrstein et al., Figure 4 and page 7, paragraph 1 lines 10-13 and paragraph 2 lines 6-9 teaches attaching the immobilizing binders to a surface of a microscopy chamber and adding the target species to the microscopy chamber. Woehrstein et al., Figure 4A and 4B and page 7 paragraph 1, lines 6-13 and paragraph 2 lines 6-9, teaches performing hybridization reactions to bind the immobilizing binders to the primary binding site on the target species present in the microscopy chamber. In Woehrstein et al., a microscopy chamber is referred to as a “flow chamber”, immobilizing binders are referred to as “capture strands”, and the primary binding site is referred to as “a region on the target”. Woehrstein et al. teaches performing image analysis to detect the first and second fluorophore colors and thereby count each first and second DNA nanostructure present on the surface of a microscopy chamber (page 4, column 2, paragraph 3, lines 10-16 and Figure 3B). Woehrstein et al. further teaches determining the relative concentration of each target molecule species based on how many DNA nanostructures are present on the surface of a microscopy chamber (page 8, lines 4-7 and Figure 4C). In Woehrstein et al., the DNA nanostructures with attached fluorophores are referred to as “barcodes” and the microscopy chamber is referred to as “a surface”.
Regarding claim 3, Woehrstein et al., Figure S11, teaches no first organic fluorophore is attached to any of the first DNA nanostructures at a location within 5 nm of any second organic fluorophore or third organic fluorophore.
Regarding claim 4 and claim 5 - Woehrstein et al. page 5, column 2, lines 6-12 and Figure 3B and 3C C teaches DNA nanostructures have fluorophore combinations that result in different DNA nanostructures displaying 64 possible colors produced by 64 combinations of four intensities of the first color, the second color and the third color being emitted from fluorescent sites on each of the first DNA nanostructures.
Regarding claim 6, claim 7 and claim 20, Woehrstein et al. teaches DNA
nanostructures form a rectangular shape with dimensions of 50nm-70nm by 80nm-
100nm and lengths less than 110nm (abstract lines 6-10, page 2 column 2 lines 4-7 and
Figure 4).
Regarding claim 8 and claim 21, Woehrstein et al. teaches performing hybridization reactions to bind the first fluorophore binders to the secondary binding site on the first target is performed before the step of attaching the first fluorophore binders to the first DNA nanostructures. (Page 7, column 2, paragraph 1, lines 2-6 and 10-13 and paragraph 2 lines 1-2 and 7-9). In Woehrstein et al., the fluorophore binders are referred to as target-complementary 21-nt-long sequence.
Regarding claim 9, Woehrstein et al. page 3, Figure 1, teaches the first color
is red and the first organic fluorophore is Atta 647N, wherein the second color is green
and the second organic fluorophore is Cy3, and wherein the third color is blue and the
third organic fluorophore is Atta 488.
Regarding claim 10 Woehrstein et al. teaches that the second organic fluorophore has a wavelength of maximum absorption that is at least 25 nm larger than the third organic fluorophore and the first organic fluorophore has a wavelength of maximum absorption that is at least 25nm larger than the second organic fluorophore (Supplementary materials, page 29, Optical setup).
Regarding claim 13, claim 15 and claim 16, Woehrstein et al., figure 4A teaches that the target binding site includes more than 18 nucleotides and less than 26 nucleotides, wherein the primary binding site on the first target includes a sequence of no more than four repetitive nucleotides, and wherein the primary binding site on the first target includes nucleotides comprising 35%-55% Guanine and Cytosine.
Regarding claim 14, Woehrstein et al. teaches the claim requirement that the primary binding site includes 20 nucleotides "separated" into a first sequence having at least 10 nucleotides (the first 10 nucleotides bound to the biotinylated capture strand in figure 4) and a second sequence having at least 10 nucleotides (last 10 nucleotides bound to the "capture strand" in figure 4) is met because the target has a first ten nucleotides and a last 10 nucleotides. The two sets of 10 nucleotides are "separated" by a single nucleotide. There is no further structural element required. Woehrstein et al. further teaches the immobilizing binders include regions complementary to the first and second sequences.
Woehrstein et al. does not teach (i) attaching immobilizing oligonucleotides to a surface of a microscopy chamber and (ii) attaching first and second immobilizing binders to a first and second group of the immobilizing nucleotides as recited in claims 1 and 18.
However, Grabmayr et al., teaches a method for detecting target species using hybridization structures comprising DNA nanostructures attached to fluorophore binders wherein the fluorophore binders and immobilizing binders are hybridized to the target species. Grabmayr et al., page 36, paragraph 3, lines 7-11, teaches that (i) the hybridized target species can be bound to the microscopy chamber using (ii) one or more immobilizing oligonucleotides attached to the immobilizing binder as in the instant claims. Grabmayr et al. refers to the equivalent of the fluorophore binders as “target adapters” (page 42, lines 10- 11 and lines 22-27 and pg. 30, para. 1-2, lines 6-8), and the equivalent of immobilizing binders and immobilizing oligonucleotides as “carrier adapters” and “intermediate carrier adapters”, respectively (page 36, paragraph 3, lines 7-11).
Grabmayr et al. further teaches “Herein, host body refers to the structure to be analyzed, which may comprise the target structure to be analyzed. For example, the host body may be a cell, for example a prokaryotic, eukaryotic, bacterial” --- “The target structure to be analyzed can be localized on the surface of the host body and/or inside the host body” (pg. 5, 2nd full paragraph). Grabmayr et al. further teaches “In a preferred embodiment, the present invention is also directed to a method for the detection and/or the quantification of at least two different target structures ( e.g. two mRNAs, which are derived from different genes, i.e. which comprise a different nucleic acid sequence), preferably of a plurality of different target structures. The different target structures are pairwise distinguishable” (pg. 18, last full paragraph). Grabmayr et al. further teaches “However, a target structure that is a complex, which contains and/or consists of one or more DNAs (preferably at least an at least partially single-stranded DNA), one or more RNAs (preferably at least an at least partially single-stranded RNA), one or more LNAs (preferably at least an at least partially single-stranded LNA) and/or one or more proteins, is also applicable” --- “Preferably, the target structure comprises or is a polynucleotide (i.e., e.g., a DNA, an RNA or LNA), particularly preferably an at least partially single-stranded polynucleotide and particularly preferably a single-stranded polynucleotide. Particularly preferably, the target structure comprises or is a single-stranded DNA, a single-stranded RNA or a single-stranded LNA” (pg. 32 first paragraph, continued from page 31). Therefore, Grabmayr et al. teaches the method can be used for detecting rRNA subunits of bacterial species as recited in claims 1 and 18.
Regarding claim 17, Grabmayr et al., page 40, paragraph 1, lines 1-9 further
teaches that the immobilizing oligonucleotide can comprise biotin wherein the biotin
binds to streptavidin coated on the microscopy chamber surface. In Grabmayr et al., the
immobilizing oligonucleotide is referred to as an “intermediate carrier adapter” and the
microscopy chamber surface is referred to as the “carrier surface”.
Regarding claim 14, Grabmayr et al. teaches that the immobilizing binder
includes a third region that is not complementary to the target sequence because it is
complementary to the sequence on the support. In Grabmayr et al., the immobilizing
binder is referred to as a carrier adapter.
It would have been obvious before the effective filing date of the instant application to have modified the method taught by Woehrstein et al. to have attached
the hybridization structure to the microscopy chamber using the intermediate carrier
adapter taught by Grabmayr et al. because both Woehrstein et al. and Grabmayr et al. teach methods for attaching hybridization structures to microscopy chamber surfaces.
It would have been obvious to one skilled in the art to substitute one method for the other to achieve the predicted result of attaching the hybridization structure to the microscopy chamber to allow for further analysis of the hybridization structures.
Woehrstein et al. in view of Grabmayr et al. does not teach identifying two or
more bacterial species by detecting 16S rRNA subunits extracted from a gastrointestinal
microbiomic sample of a patient.
However, Satokari et al., abstract lines 13-21, figure 4 and table 1, teaches a multiplex hybridization assay to detect two or more bacterial species using 16S rRNA -specific oligonucleotide probes hybridizing to RNA extracted from a gastrointestinal microbiomic sample of a patient. In Satokari et al., a gastrointestinal microbiomic sample is referred to as a “fecal sample” and a patient is referred to as a “human adult”.
Regarding claim 11 and 12, Satokari et al, abstract and throughout, teaches
the rRNA subunit is a 16S rRNA subunit. With regard to claim 11, the first rRNA subunit
is considered to be comprised of more than 1000 nucleotides and less than 5000
nucleotides since approximately 1500 base pairs is the typical size of the subunit for
most bacteria due to the conserved nature of the 16S rRNA gene.
It would have been obvious before the effective filing date of the instant
application to have used the method taught by Woehrstein et al. in view of Grabmayr et
al. to detect two or more bacterial species from a gastrointestinal microbiomic sample
using 16S rRNA-specific oligonucleotide probes because both Woehrstein et al. in view
of Grabmayr et al. and Satokari et al. teach multiplex detection of nucleic acid targets using oligonucleotide probes.
It would have been obvious to one of ordinary skill in the art to use the method to detect ribosomal ribonucleic acids from bacterial species to achieve the predictable result of detecting the two more bacterial species based on hybridization of the 16S rRNA-specific oligonucleotide probes. It would have been obvious to have modified the method taught by Woehrstein et al. in view of Grabmayr et al. so as to apply the technique taught by Satokari et al. to a gastrointestinal microbiomic sample in order to hypothesize the possible role of intestinal bacteria in gas production in the colon since this is linked with some intestinal imbalance states (page 126, column 2 lines 8-11 ).
Response to Remarks:
Applicant contends:
“A. Independent claim 1: The combination of Woehrstein, Grabmayr and Satokari does not form the basis of a valid rejection of claim 1 under § 103 because none of Woehrstein, Grabmayr or Satokari teaches either (i) the recited attaching of immobilizing oligonucleotides to immobilizing binders to primary binding sites on rRNA subunits having secondary binding sites that are attached to fluorophore binders, or (ii) determining the relative concentrations of two bacterial species by counting DNA nanostructures.
(i) None of Woehrstein, Grabmayr or Satokari teaches attaching immobilizing oligonucleotides to immobilizing binders to binding sites to fluorophore binders. Claim 1 recites the attaching of immobilizing oligonucleotides to immobilizing binders to primary binding sites on rRNA subunits having secondary binding sites that are attached to fluorophore binders as illustrated in Applicant's annotated FIG. 3 below” --- “Applicant disputes that Grabmayr teaches attaching immobilizing oligonucleotides to immobilizing binders to primary binding sites on rRNA subunits having secondary binding sites that are attached to fluorophore binders” on the basis of:
“First, the Grabmayr WIPO publication WO 2019/115801 is in the German language and has a 108-page specification. The German text at page 30, paragraph 2, lines 7-8, and at page 36, paragraph 3, lines 7-11 does not correspond to the disclosure that the examiner describes above. Second, the passage of the English translation of WO 2019/115801 (from Google Patents) that appears to correspond to the examiner's description does not teach attaching immobilizing oligonucleotides to immobilizing binders to primary binding sites on rRNA subunits having secondary binding sites that are attached to fluorophore binders”--- “Applicant contends that that this short passage from the 108-page German specification of WO 2019/115801 would not have disclosed the limitations listed above to one of ordinary skill in the art at the time of the invention. The examiner is improperly attempting to use hindsight to interpret the passage from WO 2019/115801 quoted above” (emphasis added),
Applicant further contends:
“C. Independent claim 18: The combination of Woehrstein, Grabmayr and Satokari
does not form the basis of a valid rejection of claim 18 under § 103 because
none of Woehrstein, Grabmayr or Satokari teaches determining the relative concentrations of two bacterial species by counting DNA nanostructures.”
Regarding (i) “Claim 1 recites the attaching of immobilizing oligonucleotides to immobilizing binders to primary binding sites on rRNA subunits having secondary binding sites that are attached to fluorophore binders as illustrated in Applicant's annotated FIG. 3 below---” (emphasis added), attention is drawn to the MPEP 904.01 Analysis of Claims [R-08.2012]:
The breadth of the claims in the application should always be carefully noted; that is, the examiner should be fully aware of what the claims do not call for, as well as what they do require. During patent examination and reexamination, the claims are given the broadest reasonable interpretation consistent with the specification. See In re Morris, 127 F.3d 1048, 44 USPQ2d 1023 (Fed. Cir. 1997) and In re NTP Inc., 654 F3d 1279, 99 USPQ 1481 (Fed. Cir. 2011). See MPEP § 2111 - § 2116.01 for case law pertinent to claim analysis.
Regarding the German language document, the corresponding passage with the page and paragraph numbers indicated is from an English translation of the WO 2019/115801 publication, which is attached to this office action.
Page 30, paragraph 2, lines 7-8 teaches “For the target adapter or carrier adapter, an oligomer can be selected which is incorporated in a helix that is located at the rim of the hollow cylinder” (emphasis added).
Page 36, 3rd full paragraph, lines 7-10 further teaches “The binding of the carrier adapter to the carrier and/or the first carrier surface can again be direct ( e.g. by a covalent or non-covalent bond) or mediated by an intermediate carrier adapter which specifically binds the carrier adapter and which specifically binds or is directly or indirectly bound to the carrier and/or the first surface thereof itself” (emphasis added).
Therefore, the text in the English translation corresponds to the passages recited in the office action. A copy of the English language translation of the WO 2019/115801 publication is attached to this office action.
Regarding improper hindsight, the MPEP (¶ 7.37.03) states:
“In response to applicant’s argument that the examiner’s conclusion of obviousness is based upon improper hindsight reasoning, it must be recognized that any judgment on obviousness is in a sense necessarily a reconstruction base upon hindsight reasoning. But so long as it takes into account only knowledge which was within the level of ordinary skill at the time the claimed invention was made, and does not include knowledge gleaned only from the applicant’s disclosure, such a reconstruction is proper. See In re McLaughlin, 443 F.2d 1392, 170 USPQ 209 (CCPA 1971).”
In response to applicant's argument that “The combination of Woehrstein, Grabmayr and Satokari does not form the basis of a valid rejection of claim 1 under § 103 because none of Woehrstein, Grabmayr or Satokari teaches either (i) the recited attaching of immobilizing oligonucleotides to immobilizing binders to primary binding sites on rRNA subunits having secondary binding sites that are attached to fluorophore binders” the test for obviousness is not whether the features of a secondary reference may be bodily incorporated into the structure of the primary reference; nor is it that the claimed invention must be expressly suggested in any one or all of the references. Rather, the test is what the combined teachings of the references would have suggested to those of ordinary skill in the art. See In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981).
The claims are rejected based on obviousness of the combined teachings of Woehrstein et al. and Grabmayr et al. in view of Satokari et al. as documented in the 103 rejection of claims 1 and 18. Woehrstein et al. teaches a method that includes the metafluorophores attached to metafluorophore binders, which are hybridized to the target species (e.g. a rRNA), immobilizing binders which are hybridized to the target species, and attaching this structure to a surface. Grabmayr further teaches that the carrier adapter binds the target specifically and the binding of the carrier adapter to the surface was mediated by an “intermediate carrier adapter” that hybridizes to the carrier adapter. Therefore, the combined teachings of Woehrstein et al. and Grabmayr et al. as documented in the 103 rejection of claims 1 and 18 teaches the limitations of attaching immobilizing oligonucleotides to immobilizing binders to primary binding sites on rRNA subunits having secondary binding sites that are attached to fluorophore binders, as illustrated below:
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In response to applicant's argument that:
“The combination of Woehrstein, Grabmayr and Satokari does not form the basis of a valid rejection of claim 1 under § 103 because none of Woehrstein, Grabmayr or Satokari teaches” --- “(ii) determining the relative concentrations of two bacterial species by counting DNA nanostructures.” (emphasis added),
the claims are rejected based on obviousness of the combined teachings of Woehrstein et al. and Grabmayr et al. in view of Satokari et al. as documented in the 103 rejection of claims 1 and 18. Woehrstein et al. in view of Grabmayr et al. teaches performing image analysis to detect the first and second fluorophore colors and thereby count each first and second DNA nanostructure present on the surface of the microscopy chamber (Woehrstein et al., pg. 4, col. 2, para. 3, lines 10-16 and fig. 3B, emphasis added). Woehrstein et al. in view of Grabmayr et al. further teaches determining the relative concentration of each target molecule species based on how many DNA nanostructures are present on the surface of a microscopy chamber (page 8, lines 4-7 and Figure 4C, emphasis added).
Woehrstein et al. in view of Grabmayr et al. does not teach identifying two or more bacterial species by detecting 16S rRNA subunits extracted from a gastrointestinal microbiomic sample of a patient. Satokari et al. teaches a multiplex hybridization assay for detecting two or more bacterial species using 16S rRNAspecific oligonucleotide probes extracted from a gastrointestinal microbiomic sample of a patient ( abstract lines 13-21, figure 4 and table 1, emphasis added). Therefore Woehrstein et al. in view of Grabmayr et al. and Satokari et al. teaches (ii) determining the relative concentrations of two bacterial species by counting DNA nanostructures.
Applicant further contends:
“B. Dependent claims 3-17
Claim 16 recites, "the primary binding site on the first rRNA subunit includes nucleotides comprising 35%-55% Guanine and Cytosine." The examiner contends that FIG. 4A of Woehrstein teaches this limitation. (Office Action, p. 9, lines 6-10) However, FIG. 4A of Woehrstein does not teach that a binding site comprises 35%-55% Guanine and Cytosine. In addition to the reasons stated above, claims 3-17 depend from claim 1
and are allowable for at least the same reasons for which claim 1 is allowable.”
The test for obviousness is not whether the features of a secondary reference may be bodily incorporated into the structure of the primary reference; nor is it that the claimed invention must be expressly suggested in any one or all of the references. Rather, the test is what the combined teachings of the references would have suggested to those of ordinary skill in the art. See In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981). The claims are rejected based on obviousness of the combined teachings of Woehrstein et al. and Grabmayr et al. in view of Satokari et al. as documented in the 103 rejection of claims 1 and 18.
Regarding the relevant teaching of Woehrstein et al., the figure legend of Woehrstein et al. teaches “Fig. 4 Quantitative nucleic acid detection. (A and B) Schematic of the hybridization reaction. A metafluorophore is programmed to hybridize to part (green) of a specific nucleic acid target. A biotinylated capture strand binds to a second region (red)---" (emphasis added). The sequence of the target region is ATGGGATAGACTCACTCATCG-TTT-CGGTTGTACTGTGACCGATTC, as shown below. The sequence contains 45 bases, of which 11 are G and 10 are C, which corresponds to 46.7% G~C. The red region contains 21 bases, of which 5 are G and 5 are C, which corresponds to 47.6% G~C. The green region contains 21 bases, of which 6 are G and 5 are C, which corresponds to 52.4% G~C. One of these regions will inherently bind the primary binding site of the target (e.g. a rRNA subunit). Therefore, Woehrstein teaches the primary binding site on the first rRNA subunit includes nucleotides comprising 35%-55% Guanine and Cytosine as documented in the 103 rejection of claim 16. The rejection of dependent claims 3-17 is also therefore maintained.
Woehrstein et al. Fig. 4:
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Applicant further contends:
“Claims 20-21 depend from claim 18 and are allowable for at least the same reasons for which claim 18 is allowable.”
The rejection of claims 1 and 18 is therefore maintained. The rejection of dependent claims 3-17, 20 and 21 is therefore also maintained as responded above.
Claims 2, 19 and claim 24 remain/are rejected under 35 U.S.C. 103 as being unpatentable over Woehrstein et al. (Sci. Adv., 3, 1-12, 2017) in view of Grabmayr et al., 2019 (WO 2019/115801 Al) and Satokari et al. (Microbial Ecology, 50, 120-127, 2005) as applied to claims 1, 3-18, 20 and 21 above, and further in view of Berkell et al. (WO 2021 /123387).
Woehrstein et al. in view of Grabmayr et al. and Satokari et al. does not teach determining an optimal relative concentration of the first bacterial species compared to the second bacterial species to improve a medical condition of a patient from whom the gastrointestinal microbiomic sample was taken and administering a dietary supplement to the patient that changes the relative concentration of the first bacterial species compared to the second bacterial species so as to move closer to the optimal relative concentration.
Regarding Claims 24, 2 and 19, Berkell et al., page 4 lines 1-5 and 13-15, teaches “The present inventors have shown that the risk of developing clinical manifestations of dysbiosis, such as CDI, can be correlated to the presence, absence, quantity or relative abundance of certain bacteria in the gut microbiota (also known as the gut flora or gut microflora). These bacteria can thus serve as biomarkers to detect subjects potentially at risk of developing CDI, from a gut microbiota sample or fecal sample of said subjects.” Berkell et al., further teaches “measure of the abundance of a first bacteria and a second bacteria, and the abundance of the first bacteria compared with a first threshold (or first cutoff value) and the abundance of the second bacteria compared with a second threshold (or second cutoff value)” (pg. 4, lines 23-25). This reads on claims 2, 19 and 24, which recite determining an optimal relative concentration of the first bacterial species compared to the second bacterial species to improve a medical condition of a patient from whom the gastrointestinal microbiomics sample was taken.
Berkell et al., page 3 lines 25-28 further teaches “Another aspect of the invention relates to a method for the prevention of CDI in a subject, the method comprising administering a live biotherapeutic product, such as a probiotic” --- wherein decision to proceed with these procedures is based on the prediction of said subject to potentially be at risk of developing CDI according to the methods described herein---“. This reads on claims 2, 19 and 24, which recite administering a dietary supplement to the patient that changes the relative concentration of the first bacterial species compared to the second bacterial species so as to move closer to the optimal relative concentration. In Berkell et al., a dietary supplement is referred to as a “probiotic”, a gastrointestinal microbiomic sample is referred to as a “gut microbiota sample”, the relative concentration is referred to as “relative abundance” and a patient is referred to as a “subject.”
Further regarding claim 24, Berkell et al. further teaches “As such, in certain aspects, the potential of a subject to be at risk of developing CDI is determined based on a detection of the presence or absence, or on the measure of the quantity or relative abundance of bacteria from at least one bacterial species from a bacterial genus selected in the group consisting of” --- “in the gut microbiota sample or a fecal sample of the subject. For example, the measured quantity or relative abundance of said bacteria can be compared to a reference value, wherein a decreased or increased measured value as compared to the reference value can be indicative of a subject potentially at risk of developing CDI” (pg. 4, lines 8-15, emphasis added). Berkell et al. further teaches “Another aspect of the invention relates to a method for the prevention of CDI in a subject, the method comprising administering a live biotherapeutic product, such as a probiotic” --- “wherein decision to proceed with these procedures is based on the prediction of said subject to potentially be at risk of developing CDI according to the methods described herein” (pg. 23 lines 26-29). This reads on claim 24, which recites determining an optimal quantity of the first bacterial species per amount of gastrointestinal microbiomic sample, and administering a dietary supplement to the patient that changes the relative concentration of the first bacterial species compared to the second bacterial species so as to move closer to the optimal relative concentration and to move closer to the optimal quantity of the first bacterial species per amount of gastrointestinal microbiomic sample.
It would have been obvious before the effective filing date of the instant
application to have used the method taught by Woehrstein et al. in view of Grabmayr et
al. and Satokari et al. to determine an optimal relative concentration of the first bacterial
species compared to the second bacterial species and to administer a dietary
supplement that alters the relative concentrations of each bacterial species to improve a
medical condition of a patient from whom the gastrointestinal microbiomic sample was
taken because both Satokari et al. and Berkell et al. teach gut microbiome profiling as
applied to monitoring human gastrointestinal health. It would have been obvious to one
of ordinary skill in the art to have modified the method taught by Woehrstein et al. in
view of Grabmayr et al. and Satokari et al. so as to apply the method taught by Berkell
et al. to determine an optimal concentration of each bacterial species and to administer
a dietary supplement that alters the relative concentrations of each bacterial species to
improve a medical condition of a patient from whom the gastrointestinal microbiomic
sample was taken in order to improve the health of a patient with gut microbiota
dysbiosis.
Response to Remarks:
Applicant contends:
“V. Section 103 rejection of claims 2 and 19” ---
“A. Dependent claim 2”
---“Berkell also does not teach either (i) the recited attaching of immobilizing
oligonucleotides to immobilizing binders to primary binding sites on rRNA
subunits having secondary binding sites that are attached to fluorophore binders,
or (ii) determining the relative concentrations of two bacterial species by counting
DNA nanostructures. Berkell's method does not depend on quantifying two bacterial species by counting associated RNA or DNA structures. Because none of Woehrstein, Grabmayr, Satokari or Berkell teaches either (i) attaching immobilizing oligonucleotides to immobilizing binders to primary binding sites on rRNA subunits having secondary binding sites that are attached to fluorophore binders, or (ii) determining the relative concentrations of two bacterial species by counting DNA nanostructures, reconsideration of the § 103 rejection and allowance of claim 2 are requested.” ---
" B. Dependent claim 19”
---" Berkell also does not teach determining the relative concentrations of two
bacterial species by counting DNA nanostructures. Berkell's method does not
depend on quantifying two bacterial species by counting associated RNA or DNA
structures. Because none of Woehrstein, Grabmayr, Satokari or Berkell teaches
determining the relative concentrations of two bacterial species by counting DNA
nanostructures, reconsideration of the § 103 rejection and allowance of claim 19
are requested.”
Applicant further contends:
“VI. New Claim 24” ---“New claim 24 recites quantifying the number of rRNA subunits of first and second bacterial species by counting DNA nanostructures, determining an optimal quantity of the first bacterial species per amount of sample, and administering a supplement that changes the relative concentrations of the first and second bacterial species so as to move closer to the optimal quantity of the first bacterial species per amount of sample. Berkell does not teach administering a supplement that changes the relative concentrations of each bacterial species so as to move closer to the optimal quantity of a first bacterial species per amount of sample. Berkell only concerns the relative abundance of bacteria species and attempting to achieve a predetermined "control ratio" of bacterial species. The method of Berkell only attempts to achieve a control ratio of bacterial species and not also an optimal quantity of each bacterial species. Berkell's treatment method might achieve the desired control ratio of two bacterial species, but yet both species could still be present in an overabundance or deficit in the patient's gastrointestinal microbiome. (See Berkell, 13:14-24)”
Regarding new claim 24, Berkell further teaches the method used either “the measured quantity or relative abundance of said bacteria” ---” compared to a reference value, wherein a decreased or increased measured value as compared to the reference value can be indicative of a subject potentially at risk of developing CDI” (pg. 4, lines 8-15, emphasis added). Berkell et al. further teaches “A patient enters hospital and is presented to a physician. A fecal swab is taken at hospital admission. The swab is discharged in a sterile tube on 0.5 ml water, and one drop is inserted in the analysis cassette of a diagnostic multiplex PCR machine, containing DNA primers specific for several bacterial taxa, among which Enterococcus faecium, Blautialuti and Blautia wexlerae. Quantitative detection of the amplified DNA is performed within the machine either using a fluorescent taqman probe for each of these taxa, or by analysis on a microarray slide. The relative abundance of the various identified bacteria are compared to thresholds. In particular: The ratio Enterococcus faecium I Blautialuti is compared with a threshold of 26, when Enterococcus faecium I Blautialuti >= 26, the patient is deemed to be at high risk of CDI; The relative abundance of Blautia wexlerae and of Enteroccus faecium are compared respectively to 0.093% and 0.086%. If Blautia wexlerae < 0.093% and Enteroccus faecium >= 0.086%, the patient is deemed to be at high risk of CDI” (Example 3, pg. 28-29).
The teachings by Berkell read on the limitations of claim 24 regarding determining an optimal quantity of the first bacterial species per amount of sample, and administering a supplement that changes the relative concentrations of the first and second bacterial species so as to move closer to the optimal quantity of the first bacterial species per amount of sample.
Regarding “V. Section 103 rejection of claims 2 and 19”, the combined teachings of Woehrstein, Grabmayr, Satokari and Berkell, the test for obviousness is not whether the features of a secondary reference may be bodily incorporated into the structure of the primary reference; nor is it that the claimed invention must be expressly suggested in any one or all of the references. Rather, the test is what the combined teachings of the references would have suggested to those of ordinary skill in the art. See In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981). The claims are rejected based on obviousness of the combined teachings of Woehrstein et al. and Grabmayr et al. in view of Satokari et al. and further in view of Berkell et al. as documented in the 103 rejection of claims 2 and 19.
The rejection of claims 2 and 19 is therefore maintained.
Claim 24 is rejected under 35 U.S.C. 103 as being unpatentable over Woehrstein et al. (Sci. Adv., 3, 1-12, 2017) in view of Grabmayr et al., 2019 (WO 2019/115801 Al) and Stintzi et al. (WO 2014/138999 Al).
The combined teachings of Woehrstein et al. and Grabmayr et al. as they relate to Claim 24 are documented above in the 103 rejection of Claims 1, 3-18, and 20-21.
Grabmayr et al. further teaches “Herein, host body refers to the structure to be analyzed, which may comprise the target structure to be analyzed. For example, the host body may be a cell, for example a prokaryotic, eukaryotic, bacterial, plant and/or animal cell, a virus, an exosome, a vesicle and/or a droplet. The target structure to be analyzed can be localized on the surface of the host body and/or inside the host body. Thus, the invention is also directed to a variant of the method in which the target structure is present inside a host body, wherein the method further comprises: e) disruption of the host bodies, preferably by liquid exchange with a disruption buffer in order to release the target structure from the host body. (pg. 5, 2nd full paragraph – 3rd paragraph). Therefore, Grabmayr et al. teaches the method applied to analyzing bacterial species, as recited in claim 24.
Grabmayr et al. further teaches “Preferably, the method of the invention for the detection of a target structure is also configured for the detection of further target structures that are different from the first target structure (wherein the different target structures are pairwise different). Consequently, the method can also be referred to as a method for the detection of at least two different target structures, wherein the at least two different target structures are pairwise distinguishable from each other” (pg. 44, first full paragraph)” and “If the method for the detection of a target structure is designed for the detection of further target structures which differ from the first target structure, or if the method is for the detection of at least two different target structures, the target structures can be target structures of the same type ( e.g. several mRNAs) with pairwise different structure/sequence, or the target structures comprise several different target structures of the same type. Alternatively, also target structures of different types (e.g., at least one partially single-stranded DNA or single-stranded DNA and at least one partially single-stranded or single-stranded RNA; or at least one partially single-stranded or single-stranded RNA) are conceivable” (pg. 46, last full paragraph). This reads on claim 24, quantifying a first number of rRNA subunits of the first bacterial species and a second number of rRNA subunits of the second bacterial species.
Grabmayr further teaches “In a preferred embodiment, the present invention is also directed to a method for the detection and/or the quantification of at least two different target structures (e.g. two mRNAs, which are derived from different genes, i.e. which comprise a different nucleic acid sequence), preferably of a plurality of different target structures. The different target structures are pairwise distinguishable” (pg. 18 last full paragraph). Grabmayr et al. further teaches “In particular, the method of the invention is also suited for the quantification of the target structure. In other words, the detection of the target structure preferably comprises the quantification of a target structure (e.g. in a sample solution). The quantification may be relative, i.e. in relation to another component (e.g. a second structure/target structure in a sample solution) or absolute (i.e. in form of a concentration or absolute number). For example, it may be quantified in absolute terms when the detection of the identification structure(s) is in solution, such as in flow cytometry, FCS or light sheet microscopy-based measurement geometries. Then, all identification structures in a given sample volume can be measured and an absolute number and/or concentration can be indicated” (pg. 34, first full paragraph), and “The quantification can also be carried out on the basis of an internal standard or on empirical data. Preferably, the internal standard defines a comparative value with a known concentration” (pg. 34 last full paragraph). Grabmayr et al. further teaches “It is also possible to use said method mutatis mutandis for other target structures than mRNA” (pg. 39, last full paragraph). Grabmayr et al. further teaches “Depending on the selection of the fluorescence dyes on the DNA nanostructures, the target structures are distinguishable by their color and/or by their intensity” (pg. 50, last paragraph). Grabmayr et al. further teaches “Another aspect of the present invention relates to a microwell array. The microwell array isolates the host bodies from one another and localizes the majority”--- “of the target structures of the individual host bodies into the associated microwell. The microwell array may be a preferred embodiment of a carrier” (pg. 6, 2nd full paragraph). Grabmayr further teaches “The introduction of the 3D DNA nanostructures can take place before, at the same time and/or after the introduction of the host bodies” (pg. 14, last paragraph). This reads on claim 24, quantifying a first number of rRNA subunits of the first bacterial species and a second number of rRNA subunits of the second bacterial species by counting how many first DNA nanostructures and how many second DNA nanostructures are present on the surface of the microscopy chamber.
Woehrstein et al. in view of Grabmayr et al. does not teach (i) identifying two or
more bacterial species by detecting 16S rRNA subunits extracted from a gastrointestinal
microbiomic sample of a patient. Woehrstein et al. in view of Grabmayr et al. further does not teach (ii) determining an optimal relative concentration of the first bacterial species compared to the second bacterial species to improve a medical condition of a patient from whom the gastrointestinal microbiomic sample was taken, (iii) determining an optimal quantity of the first bacterial species per amount of gastrointestinal microbiomic sample; and (iv) administering a dietary supplement to the patient that changes the relative concentration of the first bacterial species compared to the second bacterial species so as to move closer to the optimal relative concentration and to move closer to the optimal quantity of the first bacterial species per amount of gastrointestinal microbiomic sample.
However, Stintzi et al. teaches “Samples containing gut microbiota collected as described above can be assayed for determining their microbial composition---” (page 14, lines 26-27), and “metagenomic DNA can be subjected to multiplexed massively parallel sequencing on the hypervariable V6 region of the 16S rRNA gene. It is appreciated that the sequencing of regions other than the hypervariable V6 region of the 16S rRNA gene can be used provided that such regions provide discriminating power (taxonomic resolution) for at least some bacterial taxa or operational taxonomic units” --- “It will also be appreciated that other methods can be used to identify bacteria from the gut samples including but not limited to microscopy, metabolites identification, Gram staining, flow cytometry, immunological techniques (antibodies), culture-based methods such a colony forming unit counting and the like as would be known to a person skilled in the art” (pg. 15, lines 1-5 and 8-11). Therefore, Stintzi et al. teaches (i) identifying two or more bacterial species by detecting 16S rRNA subunits extracted from a gastrointestinal microbiomic sample of a patient.
Stintzi et al. further teaches “There is provided assays and methods to diagnose and treat IBD as well as to classify gut samples into IBD, UC or CD samples. There is also provided a device for classifying gut samples into IBD, UC or CD samples. In an embodiment there is provided an assay comprising the steps of measuring a level of proteobacteria or H2S producing bacteria or both in a gut microbiota sample from a human subject to identify the likelihood of the human subject having inflammatory bowel disease (IBD), and comparing the level of proteobacteria or H2S producing bacteria or both to a reference level of proteobacteria or H2S producing bacteria or both from gut microbiota samples of healthy human subjects, wherein a level of proteobacteria or H2S producing bacteria or both higher than the reference level is indicative of disease” (pg. 4 lines 29-31, continued on pg. 5, lines 1-5). Stintzi et al. further teaches “By level or abundance of bacteria or bacterial taxa it is meant a level or abundance obtained by a means to quantify bacteria such as culture-based methods, flow cytometry, microscopy, quantitative DNA analysis and any other means that would be obvious to a person skilled in the art” (pg. 13, lines 13-16). Stintzi et al. further teaches “In an aspect of the invention the relative abundance of certain bacterial taxa namely phylum, class, order, family, genus or species or combination thereof in the gut (gut microbiota profile) of patients is used to assess the presence or absence of IBD disease” --- “For example, increase in the levels of H2S producers such as Fusobacterium nucleatum, Veillonella parvula, and Atopobium parvulum is indicative of disease. Assessment of the presence of CD and UC disease in a human subject can be achieved by measuring the relative abundance of taxa as exemplified in table 1” (pg. 15, lines 12-22). Therefore, Stintzi et al. teaches (ii) determining an optimal relative concentration of the first bacterial species compared to the second bacterial species to improve a medical condition of a patient from whom the gastrointestinal microbiomic sample was taken; and (iii) determining an optimal quantity of the first bacterial species per amount of gastrointestinal microbiomic sample.
Stintzi et al. further teaches “To test the causative role of H2S-producing microbes in colitis, we assessed whether an H2S scavenger (bismuth) could alleviate Atopobium-induced colitis in 1110-/- mice”--- “This result indicates that the gut microbiota is required for the development of Atopobium-induced colitis” and “bismuth treatment prevented GALT neogenesis in mice mono-associated with A. parvulum” --- “due to a potential antimicrobial activity of bismuth on A. parvulum, as evidenced by a reduced colonization level (P=0.0001; Fig.12B)” (pg. 79 lines 12-14, 30-33, continued on pg. 80 lines 1-2 and Figure 12B). Stintzi et al., further teaches “Figure 12 B represents levels of chromosomal DNA was extracted from stool pellets obtained 6 week after mono-association or not of 129/SvEv 1110-/- mice with A. parvulum, colonization level was estimated using real-time qPCR and reported as the number of 16S rDNA gene copies per mg of stool (pg. 11, lines 16-19). Stintzi et al. further teaches “Bismuth (Ill) subsalicylate” --- “was incorporated to the chow” --- “at a concentration of 7 g/kg” (pg. 93 lines 18-20). Stintzi et al. further teaches “In another embodiment there is provided an assay comprising the steps of measuring a level of A. parvulum in a gut microbiota sample from a human subject to identify the likelihood of the human subject having IBD, and comparing the level of A. parvulum to a reference level of A. parvulum from gut microbiota samples of healthy human subjects, wherein a level of A. parvulum higher than the reference level is indicative of disease” (pg. 5, lines 6-10). Therefore, Stintzi et al. teaches (iv) administering a dietary supplement to the patient to move closer to the optimal quantity of the first bacterial species per amount of gastrointestinal microbiomic sample.
Stintzi et al. further teaches “Because both A. parvulum and the gut microbiota are required for colitis-development and because bismuth exhibits antimicrobial properties, we assessed the effect of bismuth on the gut microbiota composition of our SPF and Atopobium-associated mice” --- "Concomitantly to colitis prevention, bismuth administration altered the microbiota composition” --- “these results indicate that (1) A. parvulum colonization altered the composition of the gut microbiota (with a significant decrease in abundance of the major butyrate producers including Eubacterium and Faecalibacterium (P<0.02)) analogously to the microbiota composition of pediatric IBD patients; (2) the aberrant composition of the gut microbiota in Atopobium-associated mice is a major inducer of colitis; and (3) bismuth restores the microbiota of these mice toward a healthier community (with an increase abundance of butyrate-producers)” (pg. 80, lines 2-15). Stintzi et al. further teaches “Figure 18 A-F shows the quantitative PCR analysis of key butyrate producers. Eubacterium rectale and Faecalibacterium prausnitzii were quantified using BCoAT and 16S rRNA primers” (pg. 12, lines 25 -
Therefore Stintzi et al. teaches (iv) administering a dietary supplement to the patient that changes the relative concentration of the first bacterial species compared to the second bacterial species so as to move closer to the optimal relative concentration and to move closer to the optimal quantity of the first bacterial species per amount of gastrointestinal microbiomic sample.
Conclusion
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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/MELISSA S. POMPILIUS/Examiner, Art Unit 1682
/WU CHENG W SHEN/Supervisory Patent Examiner, Art Unit 1682
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