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
Acknowledgement is hereby made of receipt and entry of the communication filed on Nov. 10, 2022. Claims 1-20 are pending and are currently examined.
Priority
Acknowledgment is made of applicant's claim for foreign priority based on an application filed in CN on applications of CN202111329857.4, CN202210491741.9 and CN202210917136.3. It is noted, however, that applicant has not filed a certified copy of the application CN202210163969.5 as required by 37 CFR 1.55.
Claim Objections
Claims 1, 2 and 11-13 are objected to because of the following informalities:
The “step” in claim 1 should say “steps”.
Claim 1 recites abbreviation ‘Ig”, “and “S1-RBD” without spelling it out the first time it appears in the claim.
Claim 2 recites abbreviation “siRNA” without spelling it out the first time it appears in the claim. Also, the sentence “the targeting a conserved gene refers to that an siRNA” cited in claim 2 needs to be worded better. For improved clarity, claim 2 should recite “wherein targeting a conserved gene refers to selecting siRNA …”.
The phrase “that have a spacer arm” cited in claims 11-13 should be ““that has a spacer arm”.
Accordingly, appropriate correction is required.
Claim Rejections - 35 USC § 112
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.
Claims 1-20 are 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.
The base claim 1 recites the phrases “a short hairpin RNA (shRNA) region’ and “an angiotensin-converting enzyme 2 (ACE2) region”, where the “region” renders the claims indefinite. It is not clear if the region consists of the full length or a partial region of shRNA and ACE2. It is not clear what partial region is included in shRNA and ACE2.
The base claim 1 recites phrases “wherein the shRNA region is used for targeted silencing of a coronavirus mRNA”, “the ACE2 region is used for … and targeted delivery of the shRNA”, and the claim 2 recites a phrase “such that a synthesized siRNA and/or shRNA conducts targeted interference on the common gene”, where the term “targeted” renders the claims indefinite. The shRNA can be used to silence target gene expression via RNA interference (RNAi), and ACE2 acts as the receptor for the SARS-CoV-2 virus. It is unclear how the silencing or interference is targeted for shRNA/siRNA, and how the delivery is targeted for the ACE2 region.
The base claim1 recites a phrase “…the ACE2 region is used for neutralization of a coronavirus spike protein receptor-binding domain (S1-RBD) …”, where the term “neutralization” renders the claim indefinite. It is not clear how the ACE2 region can neutralize the S1-RBD.
The base claim 1 also recites that “…the artificial antibody binds to the coronavirus S1-RBD through the ACE2 in a same way that Ig specifically binds to an antigen through Fab”, where the “Ig” renders the claim indefinite. It is unclear what “Ig” referred here can bind to S1-RBD because a specific antibody is needed in order to bind a specific antigen such as binding to S1-RBD.
The base claims 1-2 and 5-7 recite a term “common gene” that renders the claim indefinite. It is unclear what the “common gene” is.
The base claim 1 recites a sentence “such that the artificial antibody binds to the coronavirus S1-RBD through the ACE2 in a same way that Ig specifically binds to an antigen through Fab”, where the phrase at “a same way that Ig specifically binds to an antigen through Fab” render the claim indefinite. Although they both are specific protein-protein interactions, ACE2 binds to RBD is a receptor-ligand binding that is fundamentally different from the antibody-antigen binding. Therefore, it is unclear “a same way” is referred to.
The base claim 1 recites the terms or phrases “shRNA-ACE2-RBD-virus”, “preventing virus infection”, “delivered to a target cell by the virus in the conjugate” and “an RNA interference (RNAi) effect on a virus-infected cell”, where the ‘virus” in each term/phrases renders the claim indefinite. It is not clear if these referred “virus” are the same or not. Also, it is not clear if the conjugate ““shRNA-ACE2-RBD-virus” only include the ‘RBD” domain or include the whole virus.
The base claim 1 recites the term “alternatively” that renders the claim indefinite. It is not clear what exact the method used for delivery and resulting in an RNA interference (RNAi) in a virus-infected cell.
Regarding claim 2, it recites a term “database” that renders the claim indefinite. It is not clear what database it refers to. Also, the claim 2 recites a term “various” to refer to the pathogenic coronaviruses, where the “various” renders the claim indefinite. It is unclear what the metes and bounds of the “various” is.
The claims 2-20 recite a term “not limited to” that render the claims indefinite. It is not clear what exact elements that these claims claimed. It is also unclear if the claimed limitations are required or not. In addition, claims 5-7 and 9-10 recite different SEQ ID NOs in their claims and also recite “not limited to” to these SEQ ID NOs, it is not clear what exact the SEQ ID NO is claimed or if these SEQ ID NOs are required. For purposes of compact prosecution and applying prior art, these claims were interpreted herein to encompass any one of these SEQ ID NOs.
Based on the description above, one of ordinary skill in the art will not know the metes and bounds of the claims.
Regarding claims 3 and 4, they recite the limitation “ligating an ACE2 polypeptide or an RBD polypeptide to each single strand of the shRNA duplex” in reference to claim 1. There is insufficient antecedent basis for this limitation in the claim.
Also, claim 3 recites “synthesizing two complementary oligonucleotide polypeptide siRNAs of 21 nt to 25 nt…”, where the “polypeptide” renders the claim indefinite. It is not clear how the siRNA becomes a “polypeptide”.
The term “middle part” in claims 3-4, 9-10 and 14-17 is a relative term which renders the climes indefinite. The term: middle part” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. For example, the “middle part” is used to refer a shRNA loop in claims 3-4, the “middle part” term is also used to refer to ACE2 polypeptides in claims 9-10 and then the “middle part” term again is used to refer to a transmembrane ACE2-extracellular ACE2 and intracellular ACE2-extracellular ACE2 in claims 14-17. Therefore, one of ordinary skill in the art will not know the metes and bounds of the claims.
Regarding claims 6-7, the term " preferably" renders the claim indefinite because it is unclear whether the limitations following the phrase are parts of the claimed invention.
Regarding claim 8, it recites that an extracellular ACE2 with amino acid sequences “1 to 740”, a transmembrane ACE2 with amino acid sequences “741 to 763” and an intracellular ACE2 with amino acid sequences “764 to 805”, where the amino acids sequences render the claim indefinite because they lack a reference sequence for the cited amino acids numbers. One of ordinary skill in the art would not be reasonably apprised of the metes and bounds of the invention without a reference sequence. Therefore, a sequence of ACE2 referred by SEQ ID NO: should be recited in the claims as a reference for the amino acid sequence numbers cited the claims. Also, it is unclear which region of ACE2 at “1 to 740” (extracellular ACE2), “741 to 763” (transmembrane ACE2) and “764 to 805” (intracellular ACE2) in claim 8 contains the S1-RBD binding domain, which is limited in the base claim 1. This rejection is also extended to the claims 14-17 for their extracellular ACE2, transmembrane ACE2 and intracellular ACE2 claims.
Regarding claims 11-13, they recite a “spacer arm” that renders the claims indefinite. It is unclear if all bonds have the “spacer arm” or only the maleimide-thiol bond has the” spacer arm”.
Regarding claims 18-20, it is not clear what the numbers of shRNA5, 16, 21 and 30 mean in 2ACE2-shRNA5, 2ACE2-shRNA16, 2ACE2-shRNA21, and 2ACE2-shRNA30 as claimed. It is also not clear what the “2” means in 2ACE2, and if the ACE2 is a full-length, extracellular ACE2, transmembrane ACE2 or intracellular ACE2.
For purposes of compact prosecution and applying prior art, these claims were interpreted herein to encompass any shRNA and any ACE2.
Claims 1-20 are rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being incomplete for omitting essential steps, such omission amounting to a gap between the steps. See MPEP § 2172.01.
The omitted steps are: if the alternative method of “the shRNA is delivered to a target cell by the virus in the conjugate resulting in an RNA interference (RNAi) effect on a virus-infected cell” is used for RNA interference, then the method and steps for how to deliver the construct into cells is missed.
Claim Rejections - 35 USC § 112 (Scope of Enablement)
The following is a quotation of the first paragraph of 35 U.S.C. 112(a):
(a) IN GENERAL.—The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor or joint inventor of carrying out the invention.
The following is a quotation of the first paragraph of pre-AIA 35 U.S.C. 112:
The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor of carrying out his invention.
Claims 1-20 are rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, because the specification, while being enabling the effect of ACE-targeted delivery of 2ACE2-shRNA5, 2ACE2-shRNA16 and 2ACE2-shRNA21 into Vero E6 cell and reduce the infection with a low TCID50 (See Tables 7, 10; [0117], [0080], does not reasonably provide enablement for constructing an artificial antibody with any short hairpin RNA (shRNA) region and any angiotensin-converting enzyme 2 (ACE2) region as claimed.
The base claim 1 is directed to a method for preparing an artificial antibody comprising shRNA region and an ACE2 region. The artificial antibody is prepared by ligating sense and antisense strands of the shRNA to an ACE2 polypeptide separately, such that the artificial antibody binds to the coronavirus S1-RBD through the ACE2 in a same way that Ig specifically binds to an antigen through Fab, to constitute an shRNA-ACE2-RBD-virus conjugate, thereby preventing virus infection through the RBD; alternatively, the shRNA is delivered to a target cell by the virus in the conjugate, resulting in an RNA interference (RNAi) effect on a virus-infected cell.
Based on the claims, an artificial antibody with a generic conjugation of shRNA-ACE2-RBD-virus is used for RNA interference (RNAi). However, the instant specification only discloses that 2ACE2-shRNA5, 2ACE2-shRNA16 and 2ACE2-shRNA21 are able to reduce the infection in Vero E6 cells (See [0117]), and all these conjugations are constructed by a specific shRNA SEQ ID NO and ACE2 SEQ ID NO. The specification does not provide support and evidence to demonstrate that any artificial antibody with any conjugations can performance RNA interference (RNAi) effect on a virus-infected cell. Therefore, the specification does not enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to practice the invention commensurate in scope with these claims.
To be enabling, the specification of the patent must teach those skilled in the art how to make and use the full scope of the claimed invention without undue experimentation. In re Wriqht, 999 F.2d 1557, 1561 (Fed. Cir. 1993). Explaining what is meant by "undue experimentation," the Federal Circuit has stated:
The test is not merely quantitative, since a considerable amount of experimentation is permissible, if it is merely routine, or if the specification in question provides a reasonable amount of guidance with respect to the direction in which the experimentation should proceed to enable the determination of how to practice a desired embodiment of the claimed invention. PPG v. Guardian, 75 F.3d 1558, 1564 (Fed. Cir. 1996).1
The factors that may be considered in determining whether a disclosure would require undue experimentation are set forth by In re Wands, 8 USPQ2d 1400 (CAFC 1988) at 1404 where the court set forth the eight factors to consider when assessing if a disclosure would have required undue experimentation. Citing Ex parte Forman, 230 USPQ 546 (BdApls 1986) at 547 the court recited eight factors:1) the nature of the invention, 2) the state of the prior art, 3) the breadth of the claims, 4) the amount of guidance in the specification, 5) the presence or absence of working examples, 6) the relative skill of those in the art, 7) the predictability or unpredictability of the art, and 8) and the quantity of experimentation necessary. Id. While it is not essential that every factor be examined in detail, those factors deemed most relevant should be considered.
M.P.E.P. §2164.03 [R-2] states: [I]n applications directed to inventions in arts where the results are unpredictable, the disclosure of a single species usually does not provide an adequate basis to support generic claims. In re Soil, 97 F.2d 623,624, 38 USPQ 189, 191 (CCPA 1938). In cases involving unpredictable factors, such as most chemical reactions and physiological activity, more may be required. In re Fisher, 427 F.2d 833,839, 166 USPQ 18, 24 (CCPA 1970). See also In re Wright, 999 F.2d 1557, 1562, 27 USPQ2d 1510, 1513 (Fed. Cir. 1993); In re Vaeck, 947 F.2d 488,496, 20 USPQ2d 1438, 1445 (Fed. Cir. 1991 ). A conclusion of lack of enablement means that, based on the evidence regarding each of the above factors, the specification, at the time the application was filed, would not have taught one skilled in the art how to make and/or use the full scope of the claimed invention without undue experimentation. In re Wright, 999 F.2d 1557,1562, 27 USPQ2d 1510, 1513 (Fed. Cir. 1993).
Accordingly, the specification does not provide sufficient guidance to allow one skilled in the art to practice the claimed invention on the full scope with a reasonable expectation of success and without undue experimentation. In the absence of such guidance and evidence of working examples, the specification fails to provide an enabling disclosure commensurate in scope with the claim.
Claim Rejections - 35 USC § 103
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
Claims 1-4, 11, 14-15 and 18-20 are rejected under 35 U.S.C. 103 as being unpatentable over Gu et al. (Virus Genes. 2019 Dec;55(6):795-801) in view of Mesias et al. (Chem Commun (Camb). 2021 Jul 15;57(57):6979-6982) as evidenced by Traube et al. (Angew Chem Int Ed Engl. 2022 Sep 19;61(38):e202204556), Thermfisher (https://rnaidesigner.thermofisher.com/rnaiexpress/help/convert_sirna_to_shrna.htm), and Tai et al. (Molecules. 2019 Jun 13;24(12):2211).
The base claim 1 is directed to a preparation method of an artificial antibody, comprising the following step:
synthesizing an artificial antibody comprising a short hairpin RNA (shRNA) region and an angiotensin-converting enzyme 2 (ACE2) region, wherein the shRNA region is used for targeted silencing of a coronavirus mRNA, and the ACE2 region is used for neutralization of a coronavirus spike protein receptor-binding domain (S1-RBD) and targeted delivery of the shRNA; the shRNA targets a conserved gene or a common gene of a coronavirus variant strain, and the ACE2 is a receptor of a coronavirus receptor- binding domain (RBD);
the artificial antibody is prepared by ligating sense and antisense strands of the shRNA to an ACE2 polypeptide separately, such that the artificial antibody binds to the coronavirus S1-RBD through the ACE2 in a same way that Ig specifically binds to an antigen through Fab, to constitute an shRNA-ACE2-RBD-virus conjugate, thereby preventing virus infection through the RBD;
alternatively, the shRNA is delivered to a target cell by the virus in the conjugate, resulting in an RNA interference (RNAi) effect on a virus-infected cell.
Gu et al. describes a short hairpin RNAs (shRNA) targeting M and N genes reduce replication of porcine deltacoronavirus in ST cells, and teaches that in order to study the potential of RNA interference (RNAi) as a strategy against PDCoV infection, two short hairpin RNA (shRNA)-expressing plasmids (pGenesil-M and pGenesil-N) that targeted the M and N genes of PDCoV were constructed and transfected separately into swine testicular (ST) cells, which were then infected with PDCoV strain HB-BD. The cytopathogenicity assays demonstrated that pGenesil-M and pGenesil- N protected ST cells against pathological changes with high specificity and efficacy. The 50% tissue culture infective dose showed that the PDCoV titers in ST cells treated with pGenesil-M and pGenesil-N were reduced 13.2- and 32.4-fold, respectively. Real-time quantitative RT-PCR also confirmed that the amount of viral RNA in cell cultures pre-transfected with pGenesil-M and pGenesil-N was reduced by 45.8 and 56.1%, respectively. This is believed to be the first report to show that shRNAs targeting the M and N genes of PDCoV exert antiviral effects in vitro, which suggests that RNAi is a promising new strategy against PDCoV infection (See Abstract).
Accordingly, Gu et al. teaches a shRNA targeting the coronavirus genes that can be delivered into cells to silence the coronavirus and reduce the virus infection in cells. However, Gu et al. is silent on a conjugation of the ACE2 polypeptide and shRNA.
Mesias et al. discloses an effective ACE2 peptide–nanoparticle conjugation and its binding with the SARS-Cov-2 RBD quantified by dynamic light scattering, and teaches that they construct truncated ACE2 peptide-conjugated gold nanoparticles as antiviral scaffolds and study their binding with the SARS-CoV-2 RBD using dynamic light scattering (DLS). Systematic DLS analysis identifies the effective peptide–nanoparticle conjugation and its efficient, specific, and long-lasting multivalent binding towards the RBD with a binding affinity of 41 nM, indicating the potential of this antiviral platform to compete with natural ACE2–RBD interactions for viral blocking and showcasing an accessible approach to measure the binding constants and kinetics (See Abstract). Mesias et al. also teaches that they chemically conjugate the modified tACE2 to the surface of AuNPs to develop tACE2–AuNPs as antiviral scaffolds, aiming to sequester SARS-CoV-2 by binding to its RBD, and discloses that previous studies demonstrated the enhanced antiviral performance of peptide-conjugated nanoparticles over soluble peptide inhibitors (See page 6979, right column; page 6980, left column, paragraph 2), wherein it is a common knowledge in the art that the nanoparticles can be made from either RNA or DNA. Therefore, Mesias et al. indicates that ACE2 peptide can be chemically conjugated to an RNA such as shRNA/siRNA to enhance the antiviral performance. This can be evidenced by Traube’s study. Traube et al. teaches that they report the synthesis of chemically stabilized small interfering RNA (siRNA) against SARS-CoV-2. The siRNA can be further modified with receptor ligands such as peptides using Cu-catalysed click-chemistry. They demonstrate that optimized siRNAs can reduce viral loads and virus induced cytotoxicity by up to five orders of magnitude in cell lines challenged with SARS-CoV-2. Furthermore, they show that an ACE2-binding peptide-conjugated siRNA is able to reduce virus replication and virus induced apoptosis in 3D mucociliary lung microtissues. The adjustment of the siRNA sequence allows a rapid adaptation of their antiviral activity against different variants of concern (See Abstract).
As for synthesizing shRNA, Thermofisher teaches that Short-hairpin RNA (shRNA) is an RNA molecule that contains sense and antisense sequences connected by a short spacer of nucleotides that enables the molecule to form a loop structure. If one has identified a synthetic siRNA that is active in triggering knockdown of your target gene, one can generate a corresponding shRNA molecule using the RNAi Designer.
Accordingly, based on the combination teachings from Mesias as evidenced by Traube and Thermofisher, the ACE2 peptide can be chemically conjugated to nanoparticles or siRNA for RNA interference to reduce coronavirus infection, where the siRNA can be extended to shRNA based on the teaching of Thermofisher.
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As for the claimed limitation “…the artificial antibody is prepared by ligating sense and antisense strands of the shRNA to an ACE2 polypeptide separately…” in the instant application, Gu et al. and Thermofisher both teach that the shRNA contains sense and antisense sequences of siRNA (See Gu et al., Table 1, page 796 and below). Therefore, it would be obvious that the ACE2 peptide can be conjugated with both the strands of the shRNA separately.
In addition, the base claim is directed to a method for making an artificial antibody, which is “in a same way that Ig specifically binds to an antigen through Fab”, which indicates an antibody/construct is defined and then to make it. Claim 1 recites “the artificial antibody is prepared by…” in line 7 of the claim, wherein only one active step is for “synthesizing an artificial antibody …”. Such language is interpreted as a product by process. The method steps recited to produce the antibody/artificial antibody are not considered when determining patentability of the product (MPEP § 2113). In In re Thorpe, the court stated, “even though product-by-process claims are limited by and defined by the process, determination of patentability is based on the product itself. The patentability of a product does not depend on its method of production. If the product in the product-by-process claim is the same as or obvious from a product of the prior art, the claim is unpatentable even though the prior product was made by a different process.” In re Thorpe, 777 F.2d 695, 698, 227 USPQ 964, 966 (Fed. Cir. 1985). (Emphasis added)
Nevertheless, Tai reviews the recent developments of siRNA bioconjugate, including the conjugation with antibody, peptide, aptamer, small chemical, lipidoid, cell-penetrating peptide polymer, and nanoparticle, and teaches that these siRNA bioconjugate, either administrated alone or formulated with other agents, could significantly improve pharmacokinetic behavior, enhance the biological half-life, and increase the targetability while maintaining sufficient gene silencing activity, with a concomitant improvement of the therapeutic outcomes and diminishment of adverse effects (See Abstract). Table 1 of Tai teaches that siRNA can be conjugated with many peptides such as antibody and Fab (See page 17, Table 1). Also, Tai teaches that siRNA is the duplex of two complementary strands of RNAs (sense and antisense strand). Structurally, siRNA has four terminal phosphate groups available for conjugation. Previous studies have demonstrated that the 5′ end of the antisense strand is essential for the initiation of RNAi machinery and will not tolerate any chemical modification or conjugation. The 5′ and 3′ ends of the sense strand and the 3′ end of the antisense strand have been considered as potential sites for conjugation. It is worthy notice that conjugation sites can also be located on the backbone of siRNA such as the 2′-OH of ribose, phosphate groups and the bases. The chemical groups on backbone provide better flexibility to the selection of conjugate position and site number, having potential as a major strategy of siRNA conjugation in future (See page 3, paragraphs 2-3; Figure 2 and below), which indicates that both the sense and antisense of siRNA or the backbone of siRNA can be conjugated to a peptide if needed. Because shRNA can be synthesized from siRNA, the shRNA can also be ligated to a peptide through both the sense and antisense strands of the shRNA or through the backbone of the shRNA to form a structure as claimed in the instant application.
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It would have been prima facie obvious for one having ordinary skill in the art before the effective filing date of the claimed invention to combine the teachings of Gu, Mesias, Thermofisher and Tai to arrive at an invention as claimed. Gu teaches a shRNA targeting M and N genes to reduce replication of the coronavirus in the host cells. Mesias teaches that the ACE2 peptide can be chemically conjugate to a nanoparticle to enhance its antiviral performance, which is evidenced by Traube for developing an ACE-binding peptide-conjugated siRNA targeting of multiple SARS-COV-2 viral genomic sites. At the same time, Thermofisher teaches synthesizing the shRNA from siRNA, and Tai teaches that the benefit of the siRNA bioconjugate, either administrated alone or formulated with other agents, can significantly improve pharmacokinetic behavior, enhance the biological half-life, and increase the targetability while maintaining sufficient gene silencing activity, with a concomitant improvement of the therapeutic outcomes and diminishment of adverse effects (See Tai et al., abstract). Tai also teaches there are multiple sites for the peptide conjugation to the siRNA/shRNA. Therefore, one of skill in the art would have been motivated to introduce the siRNA/shRNA bioconjugate taught by Mesias, Thermofisher and Tai into Gu’s study and add the ACE2 peptide conjugation with the shRNA of Gu in order to enhanced a RNA interference (RNAi) effect on a virus-infected cell, and there would be a reasonable expectation of success to develop such a method to constitute an shRNA-ACE2-RBD-virus conjugate to prevent virus infection as claimed in the instant application.
Thus, the invention as a whole was clearly prima facie obvious to one of ordinary skill in the art before the effective filing date of the claimed invention.
Regarding claim 2, Gu et al. discloses that the predicted and analyzed shRNAs were obtained by predicting the M and N gene shRNAs of PDCoV using RNAi target finder software (www.Ambit ion.com/techl ib/misc/siRNA/ (See page 796, left column, paragraph 4 and Table 1, also see below), which teaches that the shRNA is obtained from the siRNA as claimed and the siRNA is targeted to the conserved M or N gene that can be considered as commonly shared gene among the coronaviruses. Also, Gu et al. teaches that the Porcine deltacoronavirus (PDCoV) is a pathogenic virus that causes intestinal diseases in neonatal piglets with diarrhea, vomiting, dehydration, and post-infection mortality of 50–100% (See Abstract).
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Regarding claims 3 and 4, Gu et al. teaches the shRNA at follows: Gu et al. discloses that the shRNA is obtained from the sense siRNA (21-23 nt) and antisense siRNA (21-23 nt) with a loop sequence (See Table 1 and below, the bold italic letters; page 799, right column, paragraph 2), where the sense siRNA and antisense siRNA is complementary oligonucleotide. At the same time, Thermofisher also teaches that the short-hairpin RNA (shRNA) is an RNA molecule that contains sense and antisense sequences connected by a short spacer of nucleotides that enables the molecule to form a loop structure.
Although Gu does not teach the ligation of shRNA to the ACE2, the combination references of Mesias as evidenced by Traube and Tai teach that the ACE2 peptide can be chemically conjugated to shRNA through the sense strand at the 5’ and 3’ ends and the antisense at 3’ end (See Tai, page 3, paragraph 2). It would be obvious for one of ordinary skill in the art to introduce the teachings of Mesias and Tai into Gu’s study to ligate ACE2 into the shRNA and the result would be predictable based on the method and motivation Mesias and Tai taught (See the description regarding Mesias and Tai above).
Regarding claim 11, Tai et al. teaches that a targeting peptide ligand cRGD was conjugated with siRNA by noncleavable thioester bond (See page 5, Table 3). It would be obvious for one of ordinary skill in the art to introduce the bond of Tai into Gu’s study to ligate a peptide of ACE2 to the shRNA and the result would be predictable based on the method Tai taught.
Regarding claims 14-15 and 18-20, they require a different combination of the claimed artificial antibody among ACE2, full-length ACE2, extracellular ACE2, transmembrane ACE2, intracellular ACE2, shRNA and shRNA/siRNA.
Based on the description above, Gu, Mesias, Thermofisher and Tai teaches synthesizing the shRNA from siRNA that targets a M or N gene of a coronavirus, and then the shRNA is ligated to the ACE2 to form an “artificial antibody” as claimed in the instant application. Mesias, Thermofisher and Tai teaches that the ACE2 peptide can be chemically conjugated to shRNA /siRNA through the sense strand at the 5’ and 3’ ends and the antisense at 3’ end (See Tai, page 3, paragraph 2). Here all the teachings indicate that the claimed “artificial antibody” can be formed by ligating the sense and antisense strands of the shRNA to an ACE2 polypeptide separately, thus a different combination among ACE2 and shRNA can be achieved. Therefore, if providing with an ACE2 and shRNA/siRNA sequences, one of skilled in the art can make a construct such as ACE-shRNA/siRNA-ACE through routine experimental optimization in various combinations depending the application needs and the sequences used for ACE2 or shRNA/siRNA. At the same time, the acquiring ACE2 sequence and domain information are routine in the art. This can be evidenced by the ACE2 protein sequence search through NCBI (https://www.ncbi.nlm.nih.gov/protein/?term=ACE2) and literature searches. For example, Bian et al. (Acta Pharm Sin B. 2021 Jan;11(1):1-12. doi: 10.1016/j.apsb.2020.10.006. Epub 2020 Oct 13.) teaches that ACE2 domain structure with the extracellular N-terminal domain and an intracellular C-terminal tail and the transmembrane domain (See page 4, Figure 2 and below). Therefore, these claims would have been obvious with the teachings.
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Claims 5-7, 9, 12 and 16 are rejected under 35 U.S.C. 103 as being unpatentable over Gu et al. (Virus Genes. 2019 Dec;55(6):795-801) in view of Mesias et al. (Chem Commun (Camb). 2021 Jul 15;57(57):6979-6982) as evidenced by Traube et al. (Angew Chem Int Ed Engl. 2022 Sep 19;61(38):e202204556), Thermfisher (https://rnaidesigner.thermofisher.com/rnaiexpress/help/convert_sirna_to_shrna.htm), and Tai et al. (Molecules. 2019 Jun 13;24(12):2211) as applied to claims 1-4, 11-12, 14-16 and 18-20 above, and further in view of Khvorova et al. (US 2023/0021431 A1, published on Jan. 26, 2023; Filed on May 28, 2021).
Claims 5-7 and 9 require a specific SEQ ID NO for the siRNA/shRNA that targets a conserved gene or a common gene of a coronavirus.
Based on the description above, Gu, Mesias, Thermofisher and Tai teaches synthesizing the shRNA from siRNA that targets a gene of a coronavirus, and then the shRNA is ligated to the ACE2 to form an artificial antibody as claimed. However, they are silent on a siRNA/shRNA with a specific SEQ ID NO.
Khvorova et al. teaches using the novel SARS-CoV-2 targeting oligonucleotides for the treatment of SARS-CoV-2 infection, and discloses a second oligonucleotide compound comprises a sequence substantially complementary to the 45-nucleotide target gene region M_27032, as recited in Table 6A (See e.g., page 152, left column and Table 6A , page 80, SEQ ID NO: 188), where the SEQ ID NO: 188 is identical to one of the claimed SEQ ID NOs, the SEQ ID NO: 10, in these claims (See Table B below).
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It would have been prima facie obvious for one having ordinary skill in the art before the effective filing date of the claimed invention to combine the teachings of Gu, Mesias, Thermofisher, Tai and Khvorova to arrive at an invention as claimed. One of skill in the art would have been motivated to do so to introduce the known siRNA/shRNA sequence into Gu’s study and treating SARS-COV-2 infection, and there would be a reasonable expectation of success to develop such a method to construct an artificial antibody as claimed.
Regarding claim 12, Tai et al. teaches that a targeting peptide ligand cRGD was conjugated with siRNA by noncleavable thioester bond (See page 5, Table 3). It would be obvious for one of ordinary skill in the art to introduce the bond of Tai into Gu’s study to ligate a peptide of ACE2 to the shRNA and the result would be predictable based on the method Tai taught.
Regarding claim 16, it requires a different combination of the claimed artificial antibody among ACE2, full-length ACE2, extracellular ACE2, transmembrane ACE2, intracellular ACE2, shRNA and shRNA/siRNA.
Based on the description above, Gu, Mesias, Thermofisher and Tai teaches synthesizing the shRNA from siRNA that targets a M or N gene of a coronavirus, and then the shRNA is ligated to the ACE2 to form an “artificial antibody” as claimed in the instant application. Mesias, Thermofisher and Tai teaches that the ACE2 peptide can be chemically conjugated to shRNA /siRNA through the sense strand at the 5’ and 3’ ends and the antisense at 3’ end (See Tai, page 3, paragraph 2). Here all the teachings indicate that the claimed “artificial antibody” can be formed by ligating the sense and antisense strands of the shRNA to an ACE2 polypeptide separately, thus a different combination among ACE2 and shRNA can be achieved. The descriptions in the rejection on claims 14-15 and 18-20 above are also applicable to claim 16.
Claims 8, 10, 13 and 17 are rejected under 35 U.S.C. 103 as being unpatentable over Gu et al. (Virus Genes. 2019 Dec;55(6):795-801) in view of Mesias et al. (Chem Commun (Camb). 2021 Jul 15;57(57):6979-6982) as evidenced by Traube et al. (Angew Chem Int Ed Engl. 2022 Sep 19;61(38):e202204556), Thermfisher (https://rnaidesigner.thermofisher.com/rnaiexpress/help/convert_sirna_to_shrna.htm), and Tai et al. (Molecules. 2019 Jun 13;24(12):2211) as applied to claims 1-4, 11-12, 14-16 and 18-20 above, and further in view of Li et al. (Science. 2005 Sep 16;309(5742):1864-8) and Khvorova et al. (US 2023/0021431 A1, published on Jan. 26, 2023; Filed on May 28, 2021).
Claim 8 requires the ACE2 is selected from but not limited to the group consisting of an extracellular ACE2 with amino acid sequences 1 to 740, a transmembrane ACE2 with amino acid sequences 741 to 763, an intracellular ACE2 with amino acid sequences 764 to 805, a full-length ACE2, and an amino acid codon-optimized ACE2 and a polypeptide thereof.
Based on the description above, Gu, Mesias, Thermofisher and Tai teaches synthesizing the shRNA from siRNA that targets a M or N gene of a coronavirus, and then the shRNA is ligated to the ACE2 to form an artificial antibody. Mesias et al. teaches that the modified tACE2 is designed from the 22–50 residues in the N-terminal helix of ACE2 that interact with the RBD, highlighted in green in Fig. 1A (See page 6980, left column, paragraph 3), where the 22–50 residues are the parts of the extracellular domain.
Nevertheless, Li et al. studies the structure of SARS Coronavirus Spike Receptor-Binding Domain Complexed with Receptor and teaches that the spike protein (S) of SARS coronavirus (SARS-CoV) attaches the virus to its cellular receptor, angiotensin-converting enzyme 2 (ACE2). A defined receptor binding domain (RBD) on S mediates this interaction (See Abstract). Li et al. further discloses that they express the soluble ACE2, residues 19 to 615 in Sf9 cells and then determines the structure of the ACE2/SARS-CoV/RBD complex, and the final model contains residues 19 to 615 of the N-terminal peptidase domain of human ACE2 and residues 323 to 502 (except for 376 to 381) of the RBD (See page 1865, right column, paragraph 2 and page 1866, left column, paragraph 1). Here Li’s study teaches that a N-terminal domain of ACE2 contains the RBD binding epitope, which teaches the claimed “extracellular ACE2 with amino acid sequences 1 to 740” in claim 8.
It would have been prima facie obvious for one having ordinary skill in the art before the effective filing date of the claimed invention to combine the teachings of Gu, Mesias, Thermofisher, Tai and Li to arrive at an invention as claimed. One of skill in the art would have been motivated to do so to introduce the extracellular domain of ACE2, which bind to the S-RBD taught by Li, into Gu’s study, and there would be a reasonable expectation of success to develop an artificial antibody with a ACE2 that can bind to S1_RBD as claimed.
Regarding claim 10, it requires a specific SEQ ID NO for the siRNA/shRNA that targets a conserved gene or a common gene of a coronavirus based on claim 8.
Based on the description in claims 5-7 and 9 above, Khvorova et al. teaches using the novel SARS-CoV-2 targeting oligonucleotides for the treatment of SARS-CoV-2 infection, and discloses a second oligonucleotide compound comprises a sequence substantially complementary to the 45-nucleotide target gene region M_27032, as recited in Table 6A (See e.g., page 152, left column and Table 6A , page 80, SEQ ID NO: 188), where the SEQ ID NO: 188 is identical to one of the claimed SEQ ID NOs, the SEQ ID NO: 10, in these claims (See Table B above).
It would have been prima facie obvious for one having ordinary skill in the art before the effective filing date of the claimed invention to combine the teachings of Gu, Mesias, Thermofisher, Tai, Li and Khvorova to arrive at an invention as claimed. One of skill in the art would have been motivated to do so to introduce the known siRNA/shRNA sequence into Gu’s study and treating SARS-COV-2 infection, and there would be a reasonable expectation of success to develop such a method to construct an artificial antibody as claimed.
Regarding claim 13, Tai et al. teaches that a targeting peptide ligand cRGD was conjugated with siRNA by noncleavable thioester bond (See page 5, Table 3). It would be obvious for one of ordinary skill in the art to introduce the bond of Tai into Gu’s study to ligate a peptide of ACE2 to the shRNA that can be obtained from the SEQ ID NO: 188 of Khvorova and the result would be predictable based on the method Tai taught.
As for the claim 17, this claim is obvious based on the teachings from Gu, Mesias, Thermofisher, Tai, Li and Khvorova, and the descriptions regarding the rejections above in claims 14-15 is also applicable to the claim 17.
Conclusion
No claims are allowed.
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/RUIXUE WANG/Examiner, Art Unit 1672
/M FRANCO G SALVOZA/ Primary Examiner, Art Unit 1672
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