DETAILED ACTION
Notice of Pre-AIA or AIA Status
1. The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
2. Applicants amended and response of 10/2/2025 are acknowledged. Claims 72, 74, 85-87, 89 and 91 has been amended. Claims 71-88 and 95-98 were previously pending in this application. As requested by the Examiner, the claims have been renumbered in the correct sequential order. Claims 84 and 89-92 were previously absent; accordingly, previous claims 85-88 have been renumbered as 84-87 and previous 95-98 have been renumbered as claims 88-91. No new claims have been added.
Status of Claims
3. Claims 72-91 are pending in this application. Claims 72, 74, 85-87, 89 and 91 has been amended. Claims 1-71 have been canceled previously.
Nucleotide and/or Amino Acid Sequence Disclosures
4. Specific deficiency - This application contains sequence disclosures in accordance with the definitions for nucleotide and/or amino acid sequences set forth in 37 CFR 1.821(a)(1) and (a)(2). However, this application fails to comply with the requirements of 37 CFR 1.821 - 1.825.
The sequence disclosures are located in drawing figure 10, page 15 and specification pages 3-6.
Required response – Applicant must provide:
A "Sequence Listing" part of the disclosure, as described above in item 1); as well as
An amendment specifically directing entry of the "Sequence Listing" part of the disclosure into the application in accordance with 1.825(b)(2);
A statement that the "Sequence Listing" includes no new matter in accordance with 1.825(b)(5); and
A statement that indicates support for the amendment in the application, as filed, as required by 37 CFR 1.825(b)(4).
If the "Sequence Listing" part of the disclosure is submitted according to item 1) a) or b) above, Applicant must also provide:
A substitute specification in compliance with 37 CFR 1.52, 1.121(b)(3) and 1.125 inserting the required incorporation-by-reference paragraph, consisting of:
A copy of the previously-submitted specification, with deletions shown with strikethrough or brackets and insertions shown with underlining (marked-up version);
A copy of the amended specification without markings (clean version); and
A statement that the substitute specification contains no new matter;
If the "Sequence Listing" part of the disclosure is submitted according to item 1) b), c), or d) above, Applicant must also provide:
A replacement CRF in accordance with 1.825(b)(6); and
Statement according to item 2) a) or b) above.
Specification
5. Specification has been objected by the examiner. Pages 3-6 contain sequences that are not identified by SEQ ID numbers.
Drawings
6. The drawings submitted by the applicant dated 5/15/2023 have been objected by the examiner. Figure 10, page 15 contains sequences that are not identified by SEQ ID numbers.
Information Disclosure Statement
7. Applicants’ information disclosure statement filed 1/11/2023 has been considered. Initialed copy of 1449 is enclosed.
Response to Election of Species Requirement
8. In response to the Election of Species Requirement set forth in the Office Action mailed July 2, 2025, Applicant hereby provisionally elects SEQ ID NO: 36 from claims 83, 86 (previously claim 87), 88 (previously claim 89), and 90 (previously claim 98), for searching purposes only, for continued examination. SEQ ID NO: 36 corresponds to a secretin which is GspD; accordingly, at least claims 72-87 encompass the elected species. Claims 88-91 has been withdrawn from further consideration as being drawn to non-elected inventions.
Claim Objections
9. Claims 82, 84, 85-87 are objected to because of the following informalities: The claims recite multiple abbreviations GspD, YseC, D153 etc. Full name of said abbreviations are required when appears in the claims for the first time. Appropriate correction is required.
Claim Rejections - 35 USC § 112
10. The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
11. Claim 72 is rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
Claim 72 recites:
Claim 72. A method of detecting the presence or absence of an analyte in a sample and/or characterizing an analyte in a sample, the method comprising contacting a secretin nanopore disposed in a membrane with the sample; applying a voltage gradient across the membrane; and detecting ionic current flow through the nanopore.
The claim missing a correlation step. The claim detects ionic current flow; but does not provide a correlation between detection and determining the presence or absence of an analyte. Does the method require a comparison, or require measuring a change in the current. Are other steps missing from the method.
Claim Rejections - 35 USC § 103
12. 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.
13. Claims 72-87 are rejected under 35 U.S.C. 103 as being un-patentable over Gyurcsanyi, (Trends in Analytical Chemistry, vol. 27, no. 7, pp627-639, 2008) in view of Korotkov et al. (PLoS Patogens, vol.7, no.9, pp. 1-15 , 2011) and Paas et al. (US 20130121915).
The claims are drawn to:
Claim 72. A method of detecting the presence or absence of an analyte in a sample and/or characterizing an analyte in a sample, the method comprising contacting a secretin nanopore disposed in a membrane with the sample; applying a voltage gradient across the membrane; and detecting ionic current flow through the nanopore.
Claim 73. The method of claim 72, wherein the secretin nanopore is a modified secretin nanopore.
Claim 74. The method of claim 73, wherein the modified secretin nanopore comprises a lumenal surface defining a lumen that extends through the membrane between a cis-opening and a trans-opening of the modified secretin nanopore, wherein the lumenal surface of the modified secretin nanopore comprises one or more amino acid modifications relative to a lumenal surface of a wild-type secretin nanopore.
Gyurcsanyi teaches a method of detecting an analyte in a sample contacting nanopores dispersed in a membrane and applying voltage gradient to the membrane and detect the analyte ( see title, abstract, figure 1 and 2). Gyurcsanyi do not teach secretin nanopores. Gyurcsanyi recites: “Sensing with chemically-modified nanopores is an emerging field that is expected to have major impact on bioanalysis and fundamental understanding of nanoscale chemical interactions down to the single-molecule level. The main strength of nanopore sensing is that it implies the prospect of label free single-molecule detection by taking advantage of the built-in transport modulation-based amplification mechanism.”
Korotkov et al. teach Type II secretion systems (T2SSs) are critical for secretion of many proteins from Gram-negative bacteria. In the T2SS, the outer membrane secretin GspD forms a multimeric pore for translocation of secreted proteins. GspD and the inner membrane protein GspC interact with each other via periplasmic domains (see abstract). Korotkov et al., teach that the inner membrane platform consisting of multiple copies each of GspC, GspF, GspL and GspM with an associated cytoplasmic
secretion ATPase; (ii) the pseudopilus, a filamentous arrangement of multiple copies of five different pseudopilins; and (iii) a large, pore-forming outer membrane complex, mainly consisting of the secretin GspD (see page 10). Korotkov et al., teach that secretins are multimeric outer membrane proteins composed of 50–70 kDa subunits and are among the largest outer membrane proteins known. The secretin superfamily has representatives in several other multi-protein complexes engaged in transport of large macromolecular substrates across the outer membrane including the (type II) T2SS, the filamentous phage extrusion machinery, the type IV pilus system (T4PS)
and the type III secretion system (T3SS) see page 1. It is common knowledge nanopore is present it automatically extends through the raw opening the cis and trans opening.
Korotkov et al., teach hydrophobic and N1, N0 domains (see pages 3, 8 and fig 1). Korotkov et al., teach modified and mutants of GspD (pages 10-11) and deletions (page 10). Korotkov et al., teach different cell dimensions from 7-156 Ao (see table 1). Korotkov et al., teach do not teach SEQ ID 36.
Paas et al. teach an isolated pore-forming polypeptide is disclosed which comprises a naturally-occurring plugging module and a naturally-occurring pore domain, wherein at least one amino acid of the pore-forming polypeptide is mutated to generate a protease cleavage site, serving to at least partially remove the plugging module from the pore domain ( see abstract). Paas et al. teach a protease-sensitive pore-forming polypeptide comprises a modified pore protein that has been engineered such that the pore opens in the presence of a disease-associated protease (see para 0088-0089). Paas et al. teach cis opening (see para 0050-0051). Paas et al., teach sequences having 100% identity to SEQ ID 36. According to instant specification these sequences represents secretins and modified secretins from InvG and GspD from different bacteria. Paas et al. teach sequences that are identical to secretins from InvG and GspD.
Therefore, it would have been prima facie obvious at the time of applicants’ invention to combine the methods of references to obtain the instant invention.
It would have been obvious to one of ordinary skill in the art that the method of Korotkov et al., because T2SSs are critical for secretion of many proteins from Gram-negative bacteria. In the T2SS, the outer membrane secretin GspD forms a multimeric pore for translocation of secreted protein. One of skill in the art wants to exchange the nanopores of Gyurcsanyi for the secretin of Korotkov is that Gyurcsanyi teaches: sensing with chemically-modified nanopores is an emerging field that is expected to have major impact on bioanalysis and fundamental understanding of nanoscale chemical interactions down to the single-molecule level. Additionally, the benefit of using the secretin nanopores is that they are biological nanopores and these multi-protein complexes engaged in transport of large macromolecular substrates across the outer membrane.
Paas et al. teach a protease-sensitive pore-forming polypeptide comprises a modified pore protein that has been engineered such that the pore opens in the presence of a disease-associated protease. Paas et al., teach cis opening and sequences having 100% identity to SEQ ID 36.
The benefit of using Paas et al. is that Paas et al. teach an isolated pore-forming polypeptide is disclosed which comprises a naturally-occurring plugging module and a naturally-occurring pore domain.
Additionally, KSR International Co. v. Teleflex Inc., 127 S. Ct. 1727, 1741 (2007), discloses combining prior art elements according to known methods to yield predictable results, thus the combination is obvious unless its application is beyond that person's skill. KSR International Co. v. Teleflex Inc., 127 S. Ct. 1727, 1741 (2007) also discloses that "The combination of familiar element according to known methods is likely to be obvious when it does no more than yield predictable results". It is well known to combine known methods which function in a predictable manner to yield a reasonable expectation of success along with predictable results to one of ordinary skill in the art at the time of the invention. Thus, it would have been obvious to a person of ordinary skill in the art to combine prior art elements according to known methods that is ready for improvement to yield predictable results. The claimed invention is prima facie obvious in view of the teachings of the prior art, absent any convincing evidence to the contrary.
For SEQ ID NO;36
RESULT 6
US-13-261-466A-1221
(NOTE: this sequence has 2 duplicates in the database searched.
See complete list at the end of this report)
Sequence 1221, US/13261466A
Publication No. US20130121915A1
GENERAL INFORMATION
APPLICANT: Bar-Ilan University
APPLICANT: Paas, Yoav
APPLICANT: Pittel, Ilya
APPLICANT: Nir, Uri
APPLICANT: Ben-David, Barak
TITLE OF INVENTION: PROTEASE-ACTIVATABLE PORE-FORMING POLYPEPTIDES
FILE REFERENCE: 54889
CURRENT APPLICATION NUMBER: US/13/261,466A
CURRENT FILING DATE: 2012-10-04
PRIOR APPLICATION NUMBER: US 61/282,809
PRIOR FILING DATE: 2010-04-05
NUMBER OF SEQ ID NOS: 23004
SEQ ID NO 1221
LENGTH: 674
TYPE: PRT
ORGANISM: Alteromonadales bacterium TW-7
Query Match 100.0%; Score 1341; Length 674;
Best Local Similarity 100.0%;
Matches 272; Conservative 0; Mismatches 0; Indels 0; Gaps 0;
Qy 1 RAQVLIEALIVEMAEGDGINLGVQWGSLESGSVIQYGNTGASIGNVMIGLEEAKDTTQTK 60
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 341 RAQVLIEALIVEMAEGDGINLGVQWGSLESGSVIQYGNTGASIGNVMIGLEEAKDTTQTK 400
Qy 61 AVYDTNNNFLRNETTTTKGDYTKLASALSSIQGAAVSIAMGDWTALINAVSNDSSSNILS 120
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 401 AVYDTNNNFLRNETTTTKGDYTKLASALSSIQGAAVSIAMGDWTALINAVSNDSSSNILS 460
Qy 121 SPSITVMDNGEASFIVGEEVPVITGSTAGSNNDNPFQTVDRKEVGIKLKVVPQINEGNSV 180
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 461 SPSITVMDNGEASFIVGEEVPVITGSTAGSNNDNPFQTVDRKEVGIKLKVVPQINEGNSV 520
Qy 181 QLNIEQEVSNVLGANGAVDVRFAKRQLNTSVMVQDGQMLVLGGLIDERALESESKVPLLG 240
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 521 QLNIEQEVSNVLGANGAVDVRFAKRQLNTSVMVQDGQMLVLGGLIDERALESESKVPLLG 580
Qy 241 DIPLLGQLFRSTSSQVEKKNLMVFIKPTIIRD 272
||||||||||||||||||||||||||||||||
Db 581 DIPLLGQLFRSTSSQVEKKNLMVFIKPTIIRD 612
Conclusion
14. No claims are allowed.
15. Any inquiry concerning this communication or earlier communications from the examiner should be directed to KHATOL S SHAHNAN SHAH whose telephone number is (571)272-0863. The examiner can normally be reached on Mon-Tue, Thurs-Fri 12pm-8pm.
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/KHATOL S SHAHNAN SHAH/Examiner, Art Unit 1645
January 28, 2026
/JANA A HINES/Primary Examiner, Art Unit 1645