DETAILED ACTION
Notice of Pre-AIA or AIA Status
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Status of Claims
Claims 1-6, 10-12, 15-16, 18-20, 22, 24-27 and 33 are pending in the application. Claims 24-27 and 33 are presently withdrawn. Claims 1 has been amended without adding new matter. Accordingly, claims 1-6, 10-12, 15-16, 18-20 and 22 presently considered.
Maintained Rejections and New Rejections Necessitated by Amendment
Claim Rejections - 35 USC § 103
The following is a quotation of the appropriate paragraphs of 35 U.S.C. 102 that form the basis for the rejections under this section made in this Office action:
A person shall be entitled to a patent unless –
(a)(1) the claimed invention was patented, described in a printed publication, or in public use, on sale, or otherwise available to the public before the effective filing date of the claimed invention.
(a)(2) the claimed invention was described in a patent issued under section 151, or in an application for patent published or deemed published under section 122(b), in which the patent or application, as the case may be, names another inventor and was effectively filed before the effective filing date of the claimed invention.
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.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
Claims 1-6, 10-12 and 15-16 are rejected under 35 U.S.C. 103 as being unpatentable over Nivala (US 20160032236 A1; previously cited) and further in view of Li et al. (US 20180080072 A1; previously cited), hereafter “Li”.
Regarding claim 1, Nivala teaches a device and method for translocating a protein through a nanopore while monitoring electronic changes caused by different amino acids in the protein (see Abstract). Nivala teaches that translocation of proteins through a nanopore (sensor)
offers a number of possible applications, including sequencing, structure/fold analysis, purification/separation, intracellular protein delivery, and insight into the mechanics of enzymes driving the translocating polypeptide (see pg. 5, para. [0061]). Nivala teaches an engineered protein “S2-35” used for translocation through a nanopore system having the following structure (see page 11, para. [0127]):
C-terminus>charged flexible tail>Smt3>flexible linker>Smt3>N-terminus
Nivala also teaches the engineered protein “S1” having the following structure (see pg. 11, para. [0127]):
C-terminus>charged flexible tail>Smt3>N-terminus
Nivala’s engineered proteins are also shown in Fig. 1C:
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Nivala teaches the engineered Smt3 protein (the folded domains in FIG. 1C above) was modified to facilitate nanopore analysis, wherein the protein is appended with a charged, flexible glycine/serine tail, or “polyanion”, that includes 13 interspersed negatively charged aspartate residues designed to promote capture and retention of the protein in the electric field across the nanopore (see pg. 10, para. [0118]; pg. 11, para. [0121]). Nivala teaches the charged flexible tail to be drawn to the positive side of the chamber by voltage gradient (see pg. 2, para. [0020]). Hence, the charged flexible tail is understood to have a net negative charge and satisfies the limitation of “a flexible tail domain”.
According to the instant specification (see pg. 11, lines 14-16) and in view of the instant claims (i.e., claims 5-6), SUMO-like protein Smt3 satisfies the limitation of a “blocking domain”. In regards to the limitation that the blocking domain has a “stably folded tertiary structure”, Nivala teaches that the nanopore is too narrow to permit passage of a folded protein (see pg. 7, para. [0081]) and both tertiary and secondary structures must be unfolded to allow the protein to thread through the nanopore sensor in single file order (see pg. 1, para. [0008]). Nivala teaches that at the point the charged, flexible tail is threaded through the nanopore, the folded Smt3 domain prevents complete translocation of the captured protein (see pg. 3, para. [0040], lines 24-27) as the folded Smt3 domain is predicted to sit on top of the nanopore vestibule (see pg. 11, para. [0121]). Hence, it is understood that the Smt3 domain has a stably folded tertiary structure in order to prevent translocation of the protein through the nanopore.
Regarding the limitation of a “flexible analyte domain”, Nivala teaches that “as a test” the above protein was engineered to include a linker in between two Smt3 domains at the N-terminus coupled to the charged flexible segment at its C-terminus (see pg. 10, para. [0118]; pg. 11, para. [0127]). Nivala teaches these artificial linkers show different current amplitudes and duration, and one may correlate observed changes in amplitude and RMS noise to the amino acid-dependent features of the translocated protein, including its amino acid sequence and post-translational modifications (see pg. 8, para. [0090]). According to the instant specification, the flexible analyte domain is a “variable region” (see pg. 20, lines 2-3) in which the system detects an ion current pattern associated with a structural characteristic of the flexible analyte domain when it is in the tunnel of the nanopore (see pg. 24, lines 14-17). Hence, as Nivala teaches the flexible linker domain to be a sequence which can be identified and measured in the same manner, it satisfies the limitation of a “flexible analyte domain”.
Nivala does not explicitly teach wherein the flexible analyte domain is contiguous with the flexible tail domain, or separated from the flexible tail domain by an intervening linker domain.
The reason the flexible linker (“flexible analyte domain”) is not contiguous with the charged flexible tail (“flexible tail domain”) in Nivala’s S2-35, is because the protein comprises a second (intervening) Smt3 domain between the analyte domain and the tail domain, as shown above. This is because the nanopore taught by Nivala is dependent on an unfoldase, ClpX, which catalyzes unfolding of the Smt3 domains, facilitating translocation of the captured protein through the nanopore (see pg. 3, para. [0040]), rather than “blocking” further translocation of the captured protein. The examiner notes that the unfoldase, ClpX, is a feature of the nanopore system taught by Nivala, and is not part of the fusion proteins themselves. The examiner also notes that the “S1” protein taught by Nivala only has one N-terminal Smt3 domain, and while an analyte domain is not disclosed in S1, a skilled artisan would have envisaged variations of these engineered proteins in accordance with their desired application.
Nivala discloses that the “use of nanopores to sequence biopolymers was proposed more than a decade ago” (see pg. 1, para. [0009]), and since that time the most popular biosensing applications using nanopores have been for DNA sequencing (see pg. 1, para. [0008]). Nivala also teaches that the characteristics of the translocated protein can be determined in a manner analogous to the re-sequencing of DNA (see pg. 7, para. [0086]). Based on this disclosure, the prior art of Li is relied upon.
Li teaches methods of assaying the presence of target nucleic acid molecules comprising facilitating the flow of the sample through at least one nanopore in a membrane disposed adjacent or in proximity to an electrode that detects a current or current change upon movement of the target nucleic acid molecule through the nanopore (see Abstract). Hence, while Li’s disclosure is directed to nucleic acids, the principle of detection using nanopores is the same as Nivala’s. Further, Li teaches that the target nucleic acid molecule can have a tag coupled at a terminal end thereof that increases the dwell time of the target nucleic acid molecule in the nanopore (see Abstract). Li teaches that when using this method, the movement of the molecule through the nanopore takes a dwell time that is longer than when the molecule is not coupled to the tag (see pg. 1, para. [0005]). Li teaches the tag may be a polypeptide (see pg. 1, para. [0007]) which interacts with the nanopore to stop the flow of the target nucleic acid molecule through the nanopore (see pg. 3, para. [0038]), as shown in FIGURES 3A-3C:
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FIG. 3A
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FIG. 3B
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FIG. 3C
Li teaches that the tag can be a protein that has a cross-sectional size that is larger than the cross-sectional size of the nanopore, preventing the attached molecule from flowing through the nanopore (see pg. 4, para. [0054]) and by measuring current or current change with time and determining dwell times, the target nucleic acid molecule can be detected in the sample (see pg. 4, para. [0053]). Li teaches that this method overcomes various limitations that are associated with nucleic acid amplification and sequence identification, because other methods may not generate sequence information rapidly enough and/or at the accuracy necessary for the intended application (see pg. 1, para. [0003]).
Examiner notes that Li teaches that the longer dwell time is indicative of the presence of the target nucleic acid molecule (see pg. 2, para. [0015]), while Nivala teaches that a change in dwell time can be used to measure the secondary structure of a protein (see pg. 7, para. [0087]). A person of ordinary skill would have recognized that the Smt3 of Nivala’s disclosure effectively blocks translocation of the engineered protein through the nanopore, and that in the absence of the ClpX-mediated mechanism, a terminal Smt3 would remain in its folded state, preventing further translocation of the protein, analogous to what is shown in FIG. 3C of Li. As an intervening Smt3 domain would have prevented entry of the analyte domain into the nanopore, one would have had sufficient reason to remove this second Smt3 domain.
Therefore, it would have been obvious for a person of ordinary skill to have arrived at the claimed invention by modifying the engineered protein of Nivala to have a single terminal Smt3 domain to effectively serve as a “blocking domain”, because Nivala teaches that the unfolded Smt3 domain prevents further translocation through the nanopore, as does the polypeptide tag taught by Li. One would have recognized that both references teach the linking of a polypeptide, larger than the nanopore, to a terminal region of a polymeric molecule which would be expected to stop translocation through the nanopore in order to detect and analyze the molecule while it is in the nanopore lumen. One would have immediately envisaged from Nivala’s disclosure that in the absence of ClpX-mediated unfolding, the intervening Smt3 domain of Nivala’s disclosure would need to be removed in order for the region of interest (i.e., “analyte domain”) to enter the nanopore lumen for detection. Thus, in further view of Li, one would have recognized that the analyte domain could be configured to be held static by a single blocking domain (Smt3), thereby allowing the analyte domain to provide a detectable signal. Furthermore, it is well within the ordinary skill in the art to have modified Nivala’s fusion protein by simply omitting the second Smt3 domain of S2-35 or by adding the analyte domain to S1. One would have also been motivated to combine these teachings, because Li suggests that using such methods may produce more rapid and accurate results when using nanopore sensing to analyze polymeric molecules. Hence, the combination would have been readily apparent and deemed to be a mere (A) combining of prior art elements according to known methods to yield predictable results (see MPEP 2143(I): Rationales to support rejections under 35 U.S.C. 103).
Regarding claim 2, Nivala teaches that the charged flexible tail is targeted to pass through a nanopore by being drawn to the positive side of the chamber by a voltage gradient (see pg. 2, para. [0020]). Nivala teaches that the nanopore is too narrow to permit passage of a folded protein (see pg. 7, para. [0081]) and both tertiary and secondary structures must be unfolded to allow the protein to thread through the nanopore sensor in single file order (see pg. 1, para. [0008]). This is why the negatively charged polyanion tail is “unstructured” (see pg. 11, para. [0121]) which is understood to mean lacking tertiary and secondary structure. Nivala teaches that at the point the charged, flexible tail is threaded through the nanopore, the folded Smt3 domain prevents complete translocation of the captured protein (see pg. 3, para. [0040], lines 24-27) as the folded Smt3 domain is predicted to sit on top of the nanopore vestibule (see pg. 11, para. [0121]). Hence, in view of Nivala’s teachings, it is understood that the tertiary structure of the folded Smt3 (i.e., “blocking domain”) exceeds the diameter of the nanopore which prevents further translocation of the engineered protein.
Regarding claim 3, Nivala teaches that the nanopores can have a diameter of 1.5 nm (see pg. 4, para. [0045]). As discussed regarding claim 2, it is understood from Nivala’s disclosure that the nanopore is too narrow to permit the passage of Smt3 in its folded state and that its tertiary structure must have a diameter greater than the nanopore in order for it to sit on top of the nanopore vestibule and prevent further translocation of the protein. The relative size of the unstructured tail, which is narrow enough to be threaded through the nanopore, and the folded Smt3 can also be seen in Figure 1C of Nivala. Hence, Nivala teaches the tertiary structure of Smt3 (i.e., the “blocking domain”) to have a diameter greater than the nanopore, which includes nanopores having a diameter of 1.5 nm.
Regarding claim 4, Nivala teaches the Smt3 domain is comprised of about 100 amino acids (see pg. 10, para. [0119]).
Regarding claim 5, as discussed above, Nivala teaches an N-terminal Smt3 domain which satisfies the limitation of a SUMO-like protein domain.
Regarding claim 6, as discussed above, Nivala teaches an N-terminal Smt3 domain.
Regarding claim 10, Nivala does not teach the flexible analyte domain has an amino acid sequence of between 15 and 25 amino acids. Instead, Nivala teaches the flexible linker (i.e., “flexible analyte domain”) to be 35 amino acid residues in length (see pg. 10, para. [0118], “S2-35”). Examiner notes that Nivala teaches nanopore sensing can be used to measure numerous characteristics of a protein, including its length, secondary characteristics, and modifications (see pg. 7, para. [0083]).
In view of the instant specification, “SEQ ID NO: 30 is an exemplary analyte domain sequence” (pg. 13, lines 12-14); however, this sequence is 31 amino acids in length (see instant Sequence Listing File) which is the same length as the analyte domain used in Applicant’s examples (see Table 1 on pgs. 31-33) and is also 20% higher than the upper limit of the claimed range. The instant specification also states that in some embodiments the analyte domain comprises “about 1 amino acid to about 30 amino acids” (see pg. 12, lines 10-11).
Furthermore, there is no apparent support showing the criticality of the claimed range. There is zero discussion in the present disclosure as to what considerations are taken into account when selecting a flexible analyte domain of, for example, 1 amino acid as opposed to 31 amino acids, and there is no evidence of unexpected results presented in Applicant’s disclosure using the claimed range. Furthermore, a person of ordinary skill would have recognized from Nivala and Li that nanopore analysis could be used to analyze molecules of various lengths and sizes and would have had a reasonable expectation of success in detecting a protein domain of 15 to 25 amino acids. One would have been motivated to do so, because Nivala teaches that the translocation of proteins through a nanopore offers a number of possible applications and can be used to measure a number of different protein characteristics, including sequence length, secondary structures, and modifications, which would be expected to vary amongst proteins of interest and their respective domains. See MPEP 2144.05(II)(B) which states that “[w]hen there is a design need or market pressure to solve a problem and there are a finite number of identified, predictable solutions, a person of ordinary skill has good reason to pursue the known options within his or her technical grasp. If this leads to the anticipated success, it is likely the product not of innovation but of ordinary skill”. Hence, in the absence of any evidence to show the claimed range is critical, the further limitation of claim 10 would have been obvious.
Regarding claim 11, according to the instant specification, the term “identifiable barcode” refers to the ability to detect and differentiate a particular unique barcode sequence in relation to different barcode sequences in other analyte domains using, e.g., a nanopore detection platform (see pg. 12, lines 23-27). The instant specification states that the barcode is a region of the protein that is held within the sensitive region of the nanopore lumen upon which the changes to the barcode sequence manifest changes to the nanopore ionic current signal (see pg. 4, lines 12-14).
Nivala teaches the method and device may involve measuring several characteristics of the protein including the identity of the protein (see pg. 7, para. [0083]). Nivala teaches the identity of the protein may be measured by measuring the sequence of the protein itself or by measuring the presence of a particular motif in the protein (see pg. 7, para. [0085]). In Nivala’s examples, Nivala teaches that the differences in levels of current measured between two engineered proteins containing two different linker regions during translocation was due to differences in linker amino acid composition (see pg. 12, para. [0135]). Nivala teaches that the amino acid sequence may be read in blocks of amino acids, which need not resolve each individual amino acid, but rather one could just resolve “words” or blocks/chunks of amino acids that still enable identification of the protein/polypeptide sequence (see pg. 7, para. [0086]). Hence, Nivala teaches fusion proteins containing amino acid sequences (e.g., in the flexible linker region of Nivala’s exemplified fusion proteins) that can be measured to identify the engineered protein (e.g., like a barcode). Accordingly, this disclosure meets the limitation of “an amino acid sequence containing a uniquely identifiable barcode”.
Regarding claim 12, Nivala teaches the method may involve measuring whether or not the protein is modified (see pg. 7, para. [0083]) and preferably comprises determining whether or not the protein is modified by methylation, phosphorylation, oxidation, or glycosylation (see pg. 7, para. [0088]). Nivala teaches that the artificial linkers used in the examples showed different current amplitudes and duration which may be correlated to post-translational modifications (see pg. 8, para. [0090]) which can also be determined using one or more labels, tags or spacers (see pg. 7, para. [0088]).
In view of the instant specification, exemplary post-translation modifications to be targeted include phosphorylation (see pg. 13, lines 5-12), and the analyte domain may contain a casein II (CKII) domain based on the motif “SXXD”, which can result in phosphorylation of the serine in the motif (see pg. 7, para. 12-14).
Nivala teaches a fusion protein, as described regarding claim 1, wherein the artificial linker comprises the following sequence (see pg. 11, para. [0123]) which has this motif (underlined and in bold):
GGSGSAGSGASGSSGSEGSGASGSAGSGSAGSRGSGASGSAGSGSAGSGGAEAAKEAAKEAAKEAAKEAAKAGGSGSAGSAGSASSGSDGSGASGSAGSGSAGSKGSGASGSAGSGSSGS
Hence, Nivala teaches the fusion protein wherein the flexible linker domain (i.e., “flexible analyte domain”) contains a target sequence for a post-translation modification.
Regarding claim 15, Nivala teaches the charged flexible tail domain to be 65 amino acids long (see pg. 10, para. [0118]; pg. 11, para. [0121]).
Regarding claim 16, Nivala teaches the 65-amino-acid-long, charged tail to be comprised of glycine/serine residues with 13 interspersed aspartate residues (pg. 11, para. [0121])
Claims 18-20 and 22 are rejected under 35 U.S.C. 103 as being unpatentable over Nivala and Li, as applied to claims 1-6, 10-11 and 15-16 above, and further in view Le (previously cited).
Regarding claim 18, as discussed above, Nivala teaches a fusion protein comprising, in order, an Smt3 (“blocking”) domain, a flexible linker (“analyte”) domain, and a flexible, negatively charged tail domain.
Nivala does not teach the fusion protein further comprising a secretion domain functional in a cell type of interest.
Le teaches that Escherichia coli is one of the most extensively used prokaryotic organisms for the industrial production of enzymes (see pg. 1, col. 1, para. 1), but it is a common problem that some recombinant proteins will aggregate to form inclusion bodies and only a small proportion of the protein produced is soluble and active (see pg. 1, col. 1, para. 2). Le teaches that many attempts have been made to improve the soluble expression of recombinant proteins in E. coli and the formation of inclusion bodies could be decreased by fusing the target gene to another gene (see pg. 1, col. 2, para. 1). Le teaches OsmY has been used as a fusion partner to excrete target proteins into the medium, and when fused to OsmY, E. coli alkaline phosphatase, Bacillus subtilis a-amylase, and human leptin could be secreted into the medium at high levels (see pg. 1, col. 2, para. 2). Le reports the fusion of the xynA enzyme with the protein OsmY, and the finding that cells harboring the recombinant plasmid pET-OsmY-xynA expressed an activity level about 24 times higher than that expressed from pET-20b-xynA (see Abstract).
It would have been obvious at the time of filing for a person of ordinary skill to have combined the teachings of Nivala and Le to fuse the secretion domain taught by Le to the fusion protein taught by Nivala for expression in E. coli cells, because Le teaches that fusion with OsmY results in high levels of expression using other proteins. One would have recognized that in order to apply the teachings of Nivala, a supply of the proteins being studied may be required in order to utilize the proteins and study their characteristics using a nanopore system. One would have been motivated to select OsmY, because addition of the OsmY signal sequence for secretion of the fusion protein would allow for high levels of production of the protein in soluble form, thus providing a better means for producing the fusion proteins and a good supply for applications using a nanopore system. As Le teaches fusions of an OsmY domain have been successful using several other proteins, one would have recognized the results of the combination to be predictable. Hence, the combination would have been readily apparent and deemed to be a mere (A) combining of prior art elements according to known methods to yield predictable results (see MPEP 2143(I): Rationales to support rejections under 35 U.S.C. 103). Furthermore, it is well within the ordinary skill in the art to design and make fusion proteins using peptides of known structure and function for secretion in E. coli cells.
Regarding claim 19, Le teaches the fusion protein was designed to carry an N-terminal OsmY signal sequence (see pg. 2, col. 2, para. 1).
Regarding claim 20, Le teaches the overexpression of a soluble fusion protein in E.coli using OsmY as a fusion partner (see pg. 1, col. 2, para. 3). As discussed above, E. coli is a prokaryotic organism and, hence, satisfies the limitation of a “prokaryotic cell”.
Regarding claim 22, Le teaches the fusion of OsmY to recombinant proteins to improve their soluble expression in E. coli, as discussed above.
Response to Declaration
The declaration under 37 CFR 1.132 filed 04/06/2026 is insufficient to overcome the rejection of claims 1-6, 10-12, 15-16, 18-20 and 22 under 35 U.S.C. 103 as set forth in the last Office action.
Note: Although factual evidence is preferable to opinion testimony, such testimony is entitled to consideration and some weight so long as the opinion is not on the ultimate legal conclusion at issue. See MPEP 716.01(c). In assessing the probative value of an expert opinion, the examiner must consider the nature of the matter sought to be established, the strength of any opposing evidence, the interest of the expert in the outcome of the case, and the presence or absence of factual support for the expert’s opinion. Ashland Oil, Inc. v. Delta Resins & Refractories, Inc., 776 F.2d 281, 227 USPQ 657 (Fed. Cir. 1985), cert. denied, 475 U.S. 1017 (1986).
(1) The declaration sets forth the inventor’s opinion that the proposed “reconfiguration” of Nivala would not be a simple or routine modification, as the key distinction between the claimed invention and the prior art of Nivala is the operating architecture and sensing mechanism, which would require changing Nivala from a translocation-based readout architecture to a held-state analyte sensing architecture, which depends on different structural constraints, different signal-generating principles, and different expectations regarding reproducibility and sequence discrimination.
This is insufficient to overcome the rejection, because the "architecture" that the inventor refers to is not commensurate in scope with that of the rejected claims. The claims are directed to the fusion protein itself, which does not include, for example, the system comprising a nanopore or the device used to detect an ion current. The rejection under 35 U.S.C. 103 remains proper because the cited prior art discloses all the structural features of the claims. Furthermore, the declaration fails to set forth any objective evidence to support that the proposed modification to the fusion protein would have been more than a “simple or routine modification” (e.g., the removal of a blocking domain). Although an affidavit or declaration which states only conclusions may have some probative value, such an affidavit or declaration may have little weight when considered in light of all the evidence of record in the application. In re Brandstadter, 484 F.2d 1395, 179 USPQ 286 (CCPA 1973). See MPEP 176.01(c).
(2) The declaration sets forth the inventor’s opinion that Li does not provide a straightforward path to the claimed invention because nucleic-acid dwell-time detection does not translate predictably to protein reporter readout. This is because nucleic acids have a relatively uniform, strongly negative backbone and more predictable electrophoretic behavior, whereas proteins are heterogeneous in charge, chemistry, folding, and conformational flexibility.
This is insufficient to overcome the rejection, because no objective evidence is presented to consider the alleged lack of predictability. There is also no discussion as to why the Nivala reference would not have overcome this alleged lack of predictability, as Nivala teaches methods demonstrating the detection of amino acids using ionic current in a nanopore-based system. Although an affidavit or declaration which states only conclusions may have some probative value, such an affidavit or declaration may have little weight when considered in light of all the evidence of record in the application. In re Brandstadter, 484 F.2d 1395, 179 USPQ 286 (CCPA 1973). See MPEP 176.01(c).
(3) The declaration sets forth the inventor’s opinion that adding a secretion domain as suggested from Le would not have been a predictable “bolt-on” modification in this setting, because secretion domains are “context dependent” and can alter folding, processing, heterogeneity, and the exact molecular species presented to the nanopore.
This is insufficient to overcome the rejection, because the declaration does not set forth any objective evidence to support the lack of predictability of using the secretion domain taught by Le. There is also no further discussion as to why the use of this particular secretion domain would have been unpredictable in the present “context” or “setting”. Therefore, in light of all the evidence of record, the general conclusions set forth in the declaration regarding secretion domains, stated generically, are insufficient to overcome the rejection. See MPEP 176.01(c).
(4) The declaration sets forth the inventor’s opinion that the 10-25 amino acid range of the flexible analyte domain in claim 10 represents a critical range for usefulness of the flexible analyte domain and briefly describes the detriments of having a domain that is too long or too short.
This is insufficient to overcome the rejection, because the declaration does not provide any objective evidence to show the criticality of the claimed range. As noted in the present rejection, the specification discloses analyte domains that are outside the claimed range and there has not been any showing that this particular range was critical. Therefore, in light of all the evidence of record, the general conclusions set forth in the declaration regarding amino acid length, which do not specifically address the criticality of the claimed range, are insufficient to overcome the rejection. See MPEP 176.01(c).
In view of the foregoing, when all of the evidence is considered, the totality of the rebuttal evidence of nonobviousness fails to outweigh the evidence of obviousness.
Response to Arguments
Regarding the rejections under 35 U.S.C. 103, Applicant argues that while Claim 1 positively recites that "the flexible analyte domain is configured to be held static in a nanopore tunnel by the blocking domain, thereby influencing current passing through the tunnel to provide detectable signals," Nivala expressly forecloses such a structure-function relationship by noting that current influence is present only during translocation of Nivala's device. Nivala notes that "[i]n the absence of ClpX/ATP, these ramping states were never observed for S2-35 (1.7 hours of experimentation), nor for S2-148 (1.2 hours of experimentation).” Nivala, paragraph [0132].
Applicant’s arguments have been fully considered but they are not persuasive.
While Nivala discloses the observation of a “ramping state” during translocation of the proteins, Nivala’s disclosure is not limited thereto. Applicant is reminded that “[t]he use of patents as references is not limited to what the patentees describe as their own inventions or to the problems with which they are concerned. They are part of the literature of the art, relevant for all they contain." In re Heck, 699 F.2d 1331, 1332-33, 216 USPQ 1038, 1039 (Fed. Cir. 1983) (quoting In re Lemelson, 397 F.2d 1006, 1009, 158 USPQ 275, 277 (CCPA 1968)). Further, “[a] reference may be relied upon for all that it would have reasonably suggested to one having ordinary skill in the art, including nonpreferred embodiments. Merck & Co. v. Biocraft Labs., Inc. 874 F.2d 804, 10 USPQ2d 1843 (Fed. Cir. 1989), cert. denied, 493 U.S. 975 (1989). See MPEP 2123. In the instant case, Nivala discloses that even before the ClpX-mediated ramping state, changes in ionic current can be measured which represent the capture of the protein (see pg. 10, para. [0114]; pg. 11, para. [0124]). The prior art of Li also teaches the measuring of current or current change, wherein the protein tag stops the flow of the target molecule, without any mention of “ramping” (see pg. 1, para. [0005]; see pg. 4, para. [0054]). Hence, a person of skill would have recognized that a nanopore system does not require ClpX-mediated ramping in order to detect a useful signal.
Applicant further argues that Nivala's flexible linker domain shows different current amplitudes only when unfoldase ClpX is present to promote translocation through the nanopore. Thus, in order to satisfy the teachings of amended Claim 1, Nivala would need to teach the presence of "a blocking domain with a stably folded tertiary structure" and "a flexible analyte domain ... configured to be held static in a nanopore tunnel by the blocking domain, thereby influencing current passing through the tunnel to provide detectable signals." The Office Action at pages 19-20 states that "a person of skill would have recognized that the additional (intervening) Smt3 could be excluded in order to allow for translocation, and the terminal Smt3 would then function in the manner disclosed by Li." But amended Claim 1 explicitly recites that the signal arises when the protein is held static. Thus, the Office Action's logic is that one of ordinary skill would remove a blocking feature to promote translocation to arrive at the claimed system which generates results when there is no translocation. The Office Action fails to explain why one of ordinary skill would be motivated to remove a feature of Nivala in the interest of promoting translocation, when the claimed system performs sensing when translocation is blocked.
Applicant’s arguments have been fully considered but they are not persuasive.
Applicant’s argument appears to regard “translocation” as the complete translocation of the protein through the nanopore using a ClpX-mediated system without any blocking mechanism, while the plain meaning of the term “translocation” includes any amount of movement through the nanopore. To clarify the examiner’s statement, the movement of the molecule into the nanopore is clearly required by both Nivala and Li, and it would have been apparent to a person of skill that the region of interest (e.g., analyte) must enter the nanopore before translocation can be halted by the folded Smt3 protein. See MPEP 2144.04 II(A) which states that the omission of an element and its function is obvious if the function of the element is not desired. In the instant case, the omission of this intervening Smt3 domain would have been obvious because its function would not have been desired in a system that does not rely on ClpX.
Applicant further argues that the alleged modification of Nivala to have the structure defined in the amended claim would render Nivala unsatisfactory for its intended purpose and would change the principle of operation of Nivala. Nivala's protein functions as an enzyme driven controlled-translocation system, whereas the claimed reporter protein is a motor-free hold architecture. Modifying Nivala to be held static would mean it could no longer function as an enzyme-driven controlled-translocation system. "If a proposed modification would render the prior art invention being modified unsatisfactory for its intended purpose, there may be no suggestion or motivation to make the proposed modification." M.P.E.P. § 2143.0l(V).
Applicant’s arguments have been fully considered but they are not persuasive.
In response to applicant's argument that the references fail to show certain features of the invention, it is noted that the features upon which applicant relies (i.e., “motor-free hold architecture”) are not recited in the rejected claim(s). Although the claims are interpreted in light of the specification, limitations from the specification are not read into the claims. See In re Van Geuns, 988 F.2d 1181, 26 USPQ2d 1057 (Fed. Cir. 1993). In this particular case, the “architecture” Applicant is referring to (absence of an “enzyme-driven controlled-translocation system”) is directed to the system used for analyzing the fusion protein, not the claimed fusion protein itself. It should also be noted that the proposed modification (i.e., removal of an Smt3 protein) is within the ordinary skill in the art (as demonstrated by Nivala’s S1 protein discussed in the rejection) and would not require “a substantial reconstruction and redesign” of said fusion protein (see MPEP 2143.01(VI). Hence, the proposed modification would be expected to still allow for the fusion protein to enter the nanopore and for the region of interest (analyte) to be analyzed, and therefore, this modification does not fundamentally change the principle of operation described by Nivala.
In response to applicant's arguments against the references individually, one cannot show nonobviousness by attacking references individually where the rejections are based on combinations of references. See In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981); In re Merck & Co., 800 F.2d 1091, 231 USPQ 375 (Fed. Cir. 1986). In the instant case, Nivala and Li both teach the attachment of folded proteins to polymeric molecules for nanopore analysis, wherein the attached polypeptides (Smt3 or “tag”) block further translocation after the molecules enter the nanopore, as discussed in the rejection. Further, Li suggests that using such methods may produce more rapid and accurate results when using nanopore sensing, and Nivala teaches that the use of nanopores to identify structural features in a protein are analogous to previous methods using nucleic acids. Here, there would have been a reasonable expectation that the structural features of the reporter protein could still be identified using the modified protein, so long as the region of interest (analyte) was within the nanopore. Therefore, the proposed modification would not have rendered the prior art of Nivala unsatisfactory for its intended purpose in view of MPEP 2143.01(V).
It should also be noted that although a modification of a prior art element may alter certain functions of a prior art invention, the courts have stated that “[a] given course of action often has simultaneous advantages and disadvantages, and this does not necessarily obviate motivation to combine" (Medichem, S.A. v. Rolabo, S.L., 437 F.3d 1157, 1165, 77 USPQ2d 1865, 1870 (Fed. Cir. 2006). See MPEP 2143.01(V).
Applicant argues that the Nivala Declaration highlights that "Nivala is directed to an enzyme-driven controlled-translocation architecture in which signal is generated as a protein substrate is actively moved relative to the nanopore," whereas Claim 1 describes "a blocking-domain architecture that holds a flexible analyte domain in a defined sensing position so that the analyte domain itself generates signal while in that held state." Thus, the "proposed reconfiguration of Nivala would not be a simple or routine modification. It would move Nivala away from its original controlled-translocation mode and toward a different sensing mechanism with different structural constraints and different sources of variability."
Applicant’s arguments have been fully considered but they are not persuasive.
In response to applicant's argument that the references fail to show certain features of the invention, it is noted that the features upon which applicant relies (i.e., the absence of an “enzyme-driven controlled-translocation architecture”) are not recited in the rejected claim(s). Although the claims are interpreted in light of the specification, limitations from the specification are not read into the claims. See In re Van Geuns, 988 F.2d 1181, 26 USPQ2d 1057 (Fed. Cir. 1993). As discussed under Response to Declaration, the "architecture" refers to the system in which the fusion protein is used, not to the claimed fusion protein itself.
Applicant further argues that the Nivala Declaration establishes that "nucleic-acid dwell-time detection does not translate predictably to protein reporter readout" because whereas "[n]ucleic acids have a relatively uniform, strongly negative backbone and more predictable electrophoretic behavior, ... proteins are heterogeneous in charge, chemistry, folding, and conformational flexibility."
Applicant’s arguments have been fully considered but they are not persuasive.
As discussed under Response to Declaration, no objective evidence is presented to consider the alleged lack of predictability. There is also no discussion as to why the Nivala reference would not overcome this lack of predictability, as Nivala teaches methods involving the detection of amino acids using ionic current in a nanopore system.
Applicant further argues, with respect to Claim 18, the Nivala Declaration establishes that "secretion domains are context dependent and can alter folding, processing, heterogeneity, and the exact molecular species presented to the nanopore," and thus the allegations that "[a]s Le teaches fusions of an OsmY domain have been successful using several other proteins, one would have recognized the results of the combination to be predictable" fail because of the inherent unpredictability of providing secretion domains to a reporter protein.
Applicant’s arguments have been fully considered but they are not persuasive.
As discussed under Response to Declaration, the declaration does not set forth any objective evidence to support the lack of predictability of using the secretion domain taught by Le. There is also no further discussion as to why the use of this particular secretion domain would have been unpredictable in the present context.
Applicant further argues that, with respect to Claim 10, the Nivala Declaration discusses why the 10-25 amino acid length is not an arbitrarily determined range, but rather seeks to balance competing interests: "[w]hen the amino acid flexible analyte-domain is too short, the analyte provides too little information content to be useful as a flexible analyte domain"; and "[w]hen the amino acid flexible analyte-domain is too long, excessive conformational averaging or other undesirable effects on overall reporter properties like poor solubility occur."
Applicant’s arguments have been fully considered but they are not persuasive.
As discussed under Response to Declaration, the declaration does not set forth any objective evidence nor does it discuss why the 10-25 amino acid length is critical. While the inventor discusses the effects of the domain being “too short” or “too long”, there is no discussion as to how the claimed range is arrived upon or why this specific range is critical when compared to, for example, an analyte domain comprising 31 or 35 amino acids, as discussed in the rejection.
Conclusion
No claims are allowed.
THIS ACTION IS MADE FINAL. 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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/DENNIS IGNATIUS ARMATO JR/Examiner, Art Unit 1651
/MELENIE L GORDON/Supervisory Patent Examiner, Art Unit 1651
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