DETAILED ACTION
Notice of Pre-AIA or AIA Status
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Claims 1, 8-15, and 19-20 are pending.
Status of the Application
Applicant’s response and amendment filed 20 June 2025 are acknowledged and entered.
Applicant has amended Claims 1, 14, and 19-20.
Response to Amendment
The claim amendments have overcome the previous Objection.
Applicant has amended the claims to overcome the previous 112(b) rejection; the previous 112(b) rejection is withdrawn.
Applicant has amended Claims 1, 14, and 19 to overcome the 112(a) written description and enablement rejections; the 112(a) written description and enablement rejections are not withdrawn.
Claims 1, 8-15, and 19-20 are examined.
Arguments applicable to newly applied rejections to amended or newly presented claims are addressed below. Arguments that are no longer relevant are not addressed.
Rejections not reiterated here are withdrawn.
Claim Interpretation
Claims 1, 14, 19, and claims depending therefrom recite consists essentially of. The Spec. teaches (p. 6 L5-12) that the language “consists essentially of” and “consisting of” can be used in place of “comprising” and “including” to provide for more specific embodiments… Because what constitutes a more specific embodiment of comprising, the claims are interpreted as requiring that the shDNA comprises an ASO complementary to an endoglin (“ENG”) mRNA target sequence comprising SEQ ID NO 21.
Additionally, the claims recite several possibilities of ASOs that the shDNA comprises but then requires that the ASO complementary to an ENG target consists essentially of SEQ ID NO 21. See the 112(b) rejection below.
Claim 10 recites about. The instant Spec. teaches that (p. 12 L17-19) “about” is meant to encompass variations of ±20%... Therefore Claim 10 is interpreted as including ±20% of the recited values.
Claim Objections
Claims 1, 14, and 19 are objected to because of the following informalities: the claims recite the two or more shDNA [singular] but should recite the two or more shDNAs [plural]. 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, 8-15, and 19-20 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.
A claim may be considered indefinite if the resulting claim does not clearly set forth the metes and bounds of the patent protection desired. See MPEP § 2173. In the present instance, Claims 1, 14 and 19 recite wherein the two or more shDNAs are conjugated to a fluorescent dye. The claim(s) are considered indefinite because there is a question or doubt as to what are the metes and bounds of the claim. The claims are unclear because it is not clear whether the different shDNAs are conjugated to the same fluorescent dye or if each shDNA is conjugated to its own fluorescent dye. Therefore an artisan could not readily envision the physical structure of the claimed invention or that is used in the claimed method.
Claims 1, 14, and 19 are rejected for those reasons. Claims 8-13, 15, and 20 are rejected because they depend from Claims 1, 14, or 19 and do not remedy the issues.
In the interest of compact prosecution the claims are interpreted as requiring each shDNA is conjugated to its own fluorescent dye—rather than multiple shDNAs being conjugated to one shared fluorescent dye—because that is the interpretation that is consistent with the structure described in the Spec. and shown in Fig. 1, and with there being two or more shDNAs.
Claims 8-9 and 11 recite the limitation "the shDNA" in L1 or L2. The antecedent basis for this limitation in the claim is not clear because Claims 8 and 10 depend from Claim 1 which recites at least a first and second shDNA; it is not clear which shDNA is the shDNA.
Claims 8 and 10 are rejected for those reasons; Claim 9 is rejected because it depends from Claim 8 and doesn’t remedy the issues. In the interest of compact prosecution the claims are interpreted as if the shDNA refers to each shDNA, so the NP is conjugated to each shDNA by a linker and each shDNA comprises at least one chemically modified nt.
Claims 14-15 and 20 recite the limitation "inhibiting the expression of an mRNA involved in a retinal disease pathology" in L1-2, VCAM-1, VEGF, ENG, and HIF-1α mRNAs in L6-8 and 10, and “inhibits the expression of the mRNA” in the last line. There is insufficient antecedent basis for the limitation mRNA in the claim because the claim recites multiple species of mRNA (ENG, VCAM-1, VEGF, and HIF-1α) as well as a genus of mRNA (mRNA involved in a retinal disease pathology). Therefore, it is not clear which mRNA is the one whose expression is inhibited.
Claim 14 is rejected for those reasons and Claims 15 and 20 are rejected because they depend from Claim 14 and don’t remedy the issues. In the interest of compact prosecution, the claims are interpreted as: …wherein the binding of the ASO to the target sequence inhibits the expression of the mRNA involved in a retinal disease pathology.
Claims 14-15 and 20 recite an mRNA involved in a retinal disease pathology. The claim(s) are considered indefinite because there is a question or doubt as to what are the metes and bounds of the claim. Because “involved in” is a broad and nebulous term without a definition, what may be considered an mRNA involved in a retinal disease pathology can differ according to the eye of the beholder. Different artisans consider involve[ment] in a retinal disease pathology to begin and end at different points and there is no one art-accepted definition of what mRNA is or is not involved in a retinal disease pathology.
Claim 14 is rejected for those reasons and Claims 15 and 20 are rejected because they depend from Claim 14 and don’t remedy the issues. In the interest of compact prosecution, the term involved in a retinal disease pathology is interpreted as meaning any mRNA that is itself known to be involved in a retinal disease pathology or any mRNA encoding any protein that is known to be involved in a retinal disease pathology.
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 14-15 and 19-20 are rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, as failing to comply with the written description requirement. The claim(s) contains subject matter which was not described in the specification in such a way as to reasonably convey to one skilled in the relevant art that the inventor or a joint inventor, or for applications subject to pre-AIA 35 U.S.C. 112, the inventor(s), at the time the application was filed, had possession of the claimed invention. This is a written description rejection. This rejection is maintained and updated in response to the claim amendments.
Claims 14 and 20 recite:
A method for inhibiting the expression of an mRNA involved in a retinal disease pathology, comprising:
introducing a nanoparticle into a cell or a tissue, the nanoparticle comprising:
two or more short hairpin DNA sequences (shDNAs), wherein a first shDNA comprises an anti-sense oligonucleotide complementary to a target sequence consisting of an endoglin mRNA, wherein a second shDNA comprises an anti-sense oligonucleotide complementary to a target sequence comprising a VCAM-1 mRNA, a VEGF mRNA, or a HIF-1a mRNA, wherein the two or more shDNA are conjugated to a fluorescent dye, and wherein the anti-sense oligonucleotide complementary to the target sequence of the endoglin mRNA consists essentially of SEQ ID NO:21; and
a colloidal gold nanoparticle conjugated to the two or more shDNA; and
allowing the anti-sense oligonucleotide to bind the target sequence;
wherein the binding of the anti-sense oligonucleotide to the target sequence inhibits the expression of the mRNA (Claim 14), and
wherein the retinal disease pathology is selected from AMD, ROP, DR, or BRVO (Claim 20).
Claim 19 recites:
A method for treating AMD, ROP, DR, or BRVO in a subject, comprising:
administering a therapeutically effective amount of a NP to the subject, wherein the NP comprises:
two or more short hairpin DNA sequences (shDNAs), wherein a first shDNA comprises an anti-sense oligonucleotide complementary to a target sequence consisting of an endoglin mRNA, wherein a second shDNA comprises an anti-sense oligonucleotide complementary to a target sequence comprising a VCAM-1 mRNA, a VEGF mRNA, or a HIF-1a mRNA, wherein the two or more shDNA are conjugated to a fluorescent dye, and wherein the anti-sense oligonucleotide complementary to the target sequence of the endoglin mRNA consists essentially of SEQ ID NO:21; and
a colloidal gold nanoparticle conjugated to the two or more shDNA.
Broad Claim 14 encompasses a method of inhibiting the expression of any mRNA involved in a retinal disease pathology which encompasses inhibiting expression of the broad genus of any mRNA(s) involved in any retinal disease pathology. Broad Claim 20 limits the retinal disease pathologies to AMD, ROP, DR, or BRVO. Broad Claim 19 encompasses a method of treating any AMD, ROP, DR, or BRVO by administering a nanoparticle (NP) that comprises shDNA comprising an ASO against endoglin mRNA and an ASO against one of VCAM-1, VEGF, and HIF-1α mRNAs.
An NP comprising an shDNA that comprises an ASO complementary to an ENG mRNA (wherein ENG-targeting ASO comprises SEQ ID NO 21), and a second shDNA that comprises any ASO complementary to any portion of a VCAM1 mRNA, a VEGF mRNA, or a HIF1α mRNA, wherein the NP can be used to inhibit expression of any mRNA involved in any retinal disease pathology (Claims 14-15) or the specific pathologies AMD, ROP, DR, or BRVO (Claim 20); or wherein the NP can be used to treat AMD, ROP, DR, or BRVO (Claim 19), would be encompassed by the claims as instantly presented.
An original claim may lack written description support when a broad genus claim is presented but the disclosure only describes a narrow species with no evidence that the genus is contemplated. See Ariad Pharms., Inc. v. Eli Lilly & Co., 598 F.3d 1336, 1349-50 (Fed. Cir. 2010) (en banc). The written description requirement for a claimed genus may be satisfied through sufficient description of a representative number of species by actual reduction to practice, reduction to drawings, or by disclosure of relevant, identifying characteristics, i.e., structure or other physical and/or chemical properties, by functional characteristics coupled with a known or disclosed correlation between function and structure, or by a combination of such identifying characteristics, sufficient to show the applicant was in possession of the claimed genus. See Eli Lilly, 119 F.3d at 1568, 43 USPQ2d at 1406. See MPEP 2163.
The Spec. discloses the claimed invention on pp. 19-20 L16-1:
In some embodiments, the nanoparticles comprise one type of antisense oligonucleotide contained with the shDNA sequence. In some embodiments, the nanoparticles comprise two types of antisense oligonucleotide contained with the shDNA sequence. In some embodiments, the nanoparticles comprise two or more types of antisense oligonucleotide contained with the shDNA sequence. For example, in some embodiments, the two types of gold-nanoparticles include sequences targeting VEGF mRNA with endoglin mRNA; sequences targeting VEGF mRNA with HIF-1α mRNA; and also sequences targeting endoglin mRNA with HIF-1α mRNA. In yet other embodiments, the endoglin, VEGF, or HIF-1α sequences can be combined with sequences targeting VCAM-1...
In some aspects, disclosed herein is a nanoparticle, comprising:
a short hairpin DNA (shDNA), wherein the shDNA comprises an anti-sense oligonucleotide complementary to a target sequence of an endoglin, HIF-1α, VCAM-1, or VEGF mRNA, or a combination thereof; and a colloidal gold nanoparticle conjugated to the shDNA.
That § discloses an shDNA sequence that comprises an ASO with two species of mRNA target (“or a combination thereof”) within the shDNA sequence.
The Spec. teaches an example of gold nanoparticles conjugated to (pp. 20-23 L10-8, Example 1) ASO against VCAM1 and a prophetic example of gold nanoparticles conjugated to (p. 23 L9-19, Example 2) ASO against endoglin. The specification teaches (p. 23 L20-25, Example 3) one example of combination nanoparticles. That § teaches that 15 nm hAuNP with two target sequences were made: sequences conjugated to 15 nm gold-nanoparticles include targeting VEGF mRNA with endoglin mRNA; and also sequences targeting VEGF mRNA with HIF-1α mRNA. Each nanoparticle contains about 48-50 of the hairpin DNAs. That § teaches that a combination nanoparticle comprises a nanoparticle with two separate target sequences.
Fig. 1 teaches that the mRNA recognition sequence is the hairpin’s loop region. The Fig. 1 description discloses (p. 4 L9-31) how the invention works. This § teaches:
In probe type-1, 15 nm spherical gold nanoparticles were designed and functionalized with single or multiple targeted hairpin-DNA/RNA oligonucleotides incorporating anti-sense sequence specific for mouse Endoglin (ENG) mRNA, or VEGFA mRNA, or VCAM-1 mRNA, or HIF-1α mRNA (AS-hAuNP) allowing multi-targeted therapy, or a scrambled version of this sequence (NS-hAuNP). (FIG. 1B) In probe type-2, 1.4 nm gold nanoparticles were functionalized with a single hairpin-DNA/RNA oligonucleotide incorporating anti-sense sequence specific for mouse ENG mRNA, or VEGFA mRNA, or VCAM-1 mRNA, or HIF-1α mRNA (AS-hAuNP) or a scrambled version of this sequence (NS-hAuNP). The DNA hairpin-loops are modified on their 5' ends with a thiol (SH) group and coupled to a maleimide group, which is connected to the gold surface through a phosphine-gold (Au-P) bond. The dye is linked to the 3' end of the oligonucleotide through an O-(CH2)7-amide linkage and is quenched. Hybridization of the target mRNA to the anti-sense recognition sequence causes the hairpin to open increasing the distance between the fluorophore and the gold surface, resulting in fluorescence emission. (FIG. 1C) The AS-mENG hAuNP is highly specific for its complementary sequence. [emphases added.]
Those probe types comprise either a nanoparticle functionalized with multiple shDNAs, each of which each comprises a specific ASO sequence specific for ENG mRNA (whose sequence comprises SEQ ID NO 21) or VEGF mRNA or VCAM-1 mRNA or HIF-1α mRNA.
The art of Mukamal (2021. Genetics and Age-Related Macular Degeneration. Am. Acad. Ophthalmol. Available online at aao.org. Accessed on 27 August 2025, “Mukamal”) and Advani (et al. 2024. QTL mapping of human retina DNA methylation identifies 87 gene-epigenome interactions in age-related macular degeneration. Nat. Comm. 15:1972, “Advani”) respectively teach (Mukamal) there are over 30 genes associated with the risk of developing AMD and (Advani) there are 87 gene-epigenome interactions in AMD. Swan (et al. 2018. The genetics of retinopathy of prematurity: a model for neovascular retinal disease. Ophthalmol. Retina 2[9]:949-962, “Swan”) teaches (Table 2) genes other than those recited in the claims that are found to have a significant association with ROP (e.g., AGTR1, BDNF, CETP). Searches for the causes of DR and BRVO revealed involvement of other genes besides those recited in the claims. Those teachings indicate that it would not be possible to inhibit expression of any mRNA involved in a retinal disease pathology (or even the specific pathologies AMD, ROP, DR, and BRVO) by inhibiting expression of solely ENG mRNA and one of VCAM-1, VEGF, and HIF-1α mRNAs.
Searches revealed that inhibiting VEGF is a treatment for wet AMD, DR, ROP, and BRVO. However, the art of National Eye Institute (2021. Age-Related Macular Degeneration [AMD]. Available online at nei.nih.gov. Accessed on 27 August 2025, “NEI”) teaches (§What’s the treatment for AMD?) there’s currently no treatment for late dry AMD. Those teachings indicate inhibiting VEGF can treat some—but not all—retinal diseases wherein the disease is AMD, ROP, DR, or RBVO. Those teachings, together with the above teachings of Mukamal, Advani, and Swan, indicate it would not be possible to treat AMD, ROP, DR, or BRVO in any subject by administering the claimed NPs that comprise ASOs to ENG and one of VEGF, VCAM-1, or HIF-1α.
Regarding shDNAs that comprise any ASO complementary to a target mRNA that is an ENG (wherein the ENG-targeting ASO comprises SEQ ID NO 21), VCAM1, VEGF, or HIF1α mRNA, the Spec. discloses (pp. 25-30) some shDNA sequences complementary to the following targets: four sequences that target ENG (ENG1-3 and human ENG1), including SEQ ID NO 21; three sequences that target HIF1α, one sequence that targets VCAM, and one sequence that targets VEGF. Those few species do not demonstrate possession of any species of ASO complementary to any mRNA target of any mRNA involved in any retinal disease pathology. Nowhere does the Spec. disclose the structure of the claimed shDNAs that can perform the functions of (1) inhibiting expression of any mRNA involved in a retinal disease pathology or the specific pathologies AMD, DR, ROP, or BRVO, or (2) treating any AMD, any ROP, any DR, or any BRVO.
The Specification does not provide specific guidance for producing an shDNA that inhibits expression of any mRNA involved in a retinal disease pathology or which can treat the recited diseases. Although Claims 14-15 and 19-20 claim functional characteristics (i.e., inhibiting expression of an mRNA involved in a retinal disease pathology and treating AMD, ROP, DR, or BRVO), the functional characteristic is not coupled with a known structure of the shDNA.
Although the Specification teaches the examples discussed above, it does not identify a core structure for an shDNA that inhibits any retinal disease pathology-associated mRNA or that treats any AMD/ROP/DR/BRVO. No core structure, partial structure, physical or chemical property, or functional characteristic coupled with a known or disclosed structure/function relationship responsible for inhibiting any retinal disease pathology-associated mRNA or treating any AMD/ROP/DR/BRVO is disclosed in such a way to demonstrate possession of the full invention as claimed at time of filing. The Spec. does not identify a core structure for inhibiting any mRNA involved in a retinal disease pathology or for treating any kind of AMD/ROP/DR/BRVO. That is relevant because the art indicates those conditions can be caused by various genes and that there is no treatment for dry AMD.
The specification teaches only NPs comprising either one shDNA with one target or two shDNAs with one target each. Altogether the number of species disclosed by complete structure is not sufficient to provide the written description support for the huge genera and subgenera of nanoparticles claimed, namely NPs comprising shDNA against ENG and one of VCAM-1, VEGF, or HIF1-1α, wherein the NP can inhibit expression of any mRNA involved in a retinal disease pathology or can treat any AMD/ROP/DR/BRVO. The number of species disclosed by complete structure is not sufficient to provide the written description support for the huge genera of NPs that can inhibit expression of any mRNA involved in a retinal disease pathology or can treat any AMD/ROP/DR/BRVO .
While none of these elements is specifically required to demonstrate possession, in combination their absence means that one skilled in the art at the time of filing would conclude that the inventors lacked possession of the full breadth of what is claimed: an invention a method for inhibiting expression of any mRNA involved in a retinal disease pathology or treating any AMD/ROP/DR/BRVO by administering the claimed NP comprising a colloidal gold NP conjugated to an shDNA that comprises two ASOs, wherein one ASO is complementary to ENG mRNA and comprises SEQ ID NO 21 and wherein the second ASO is complementary to a target sequence selected from a VCAM1 mRNA, a VEGF mRNA, and a HIF1α mRNA. Claims 14 and 19-20 are rejected for failing to demonstrate possession of the claimed invention. Claims 15 and 20 are rejected because they depend from Claims 14 or 19 and do not remedy the issues.
Claims 14-15 and 19-20 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 for:
A method for inhibiting the expression of an mRNA involved in a retinal disease pathology, wherein the retinal disease pathology is selected from age-related macular degeneration (AMD), retinopathy of prematurity (ROP), diabetic retinopathy (DR), or branch retinal vein occlusion (BRVO) and wherein the mRNA involved in a retinal disease pathology is either VCAM-1 mRNA or VEGF mRNA, the method comprising:
introducing a nanoparticle into a cell or a tissue, the nanoparticle comprising:
two or more short hairpin DNA sequences (shDNAs), wherein a first shDNA comprises an anti-sense oligonucleotide complementary to a target sequence consisting of an endoglin mRNA, wherein a second shDNA comprises an anti-sense oligonucleotide complementary to a target sequence comprising a VCAM-1 mRNA, wherein the ASO comprises SEQ ID NO:66, or a VEGF mRNA, wherein the ASO comprises [the ASOs whose results are shown in Fig. 4], wherein each of the two or more shDNA is conjugated to its own fluorescent dye, and wherein the anti-sense oligonucleotide complementary to the target sequence of the endoglin mRNA consists essentially of SEQ ID NO:21;
and a colloidal gold nanoparticle conjugated to the two or more shDNAs;
and allowing the anti-sense oligonucleotide to bind the target sequence;
wherein the binding of the anti-sense oligonucleotide to the target sequence inhibits the expression of the mRNA that is targeted by the ASO,
and
A method for treating age-related macular degeneration (AMD), wherein the AMD is caused by overexpression of VCAM-1 or VEGF, retinopathy of prematurity (ROP), diabetic retinopathy (DR), or branch retinal vein occlusion (BRVO) in a subject, comprising:
administering a therapeutically effective amount of a nanoparticle to the subject, wherein the nanoparticle comprises:
two or more short hairpin DNA sequences (shDNAs), wherein a first shDNA comprises an anti-sense oligonucleotide complementary to a target sequence consisting of an endoglin mRNA, wherein a second shDNA comprises an anti-sense oligonucleotide complementary to a target sequence comprising a VEGF mRNA, wherein the ASO comprises [the ASOs whose results are shown in Fig. 4], and wherein an additional shDNA comprises an antisense oligonucleotide complementary to a VCAM-1 mRNA or a HIF-1a mRNA, wherein the two or more shDNA are each conjugated to a separate fluorescent dye, and wherein the anti-sense oligonucleotide complementary to the target sequence of the endoglin mRNA consists essentially of SEQ ID NO:21;
and a colloidal gold nanoparticle conjugated to the two or more shDNAs.
The specification does not enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to use the invention commensurate in scope with these claims. This is a scope of enablement rejection. This is a new rejection.
The factors to be considered in determining whether a disclosure would require undue experimentation include: (A) The breadth of the claims; (B) The nature of the invention; (C) The state of the prior art; (D) The level of one of ordinary skill; (E) The level of predictability in the art; (F) The amount of direction provided by the specification; (G) The existence of working examples; and (H) The quantity of experimentation needed to make or use the invention based on the content of the disclosure. In re Wands, 8 USPQ2d, 1400 (CAFC 1988) and MPEP 2164.01.
The breadth of the claims and the nature of the invention: With respect to claim breadth, the standard under 35 U.S.C. §112(a) entails determining what the claims recite and what the claims mean as a whole. Claims 14-15 and 19-20 recite
(Claims 14-15 and 20) a method of inhibiting the expression of any mRNA involved in a retinal disease pathology by administering to a cell or tissue…,
(Claim 20) wherein the retinal disease pathology is AMD/ROP/DR/BRVO, or
(Claim 19) a method of treating any AMD/ROP/DR/BRVO by administering to a subject…
…a nanoparticle comprising (1) two or more shDNAs wherein a first shDNA comprises an ASO that consists essentially of SEQ ID NO 21 and is complementary to an ENG mRNA and wherein a second shDNA comprises an ASO complementary to a target sequence comprising a VCAM1 mRNA, a VEGF mRNA, or a HIF1α mRNA; wherein the shDNAs are conjugated to a fluorescent dye and (2) a colloidal gold nanoparticle conjugated to the two or more shDNA [emphasis added].
The BRI of Claims 14-15 and 20 is that administering the nanoparticles comprising the first shDNA comprising an ASO consisting essentially of SEQ ID NO 21 (which targets ENG mRNA) and the second shDNA comprising an ASO complementary to any target mRNA wherein the target mRNA is a VCAM1 mRNA, a VEGF mRNA, or a HIF1α mRNA will inhibit expression of any mRNA involved in a retinal disease pathology, and that the ASO will bind to any of those target sequences and inhibit mRNA expression.
The BRI of Claim 19 is that the method of administering the nanoparticles comprising the first shDNA comprising an ASO consisting essentially of SEQ ID NO 21 (which targets ENG mRNA) and the second shDNA comprising an ASO complementary to any target mRNA wherein the target mRNA is a VCAM1 mRNA, a VEGF mRNA, or a HIF1α mRNA will treat any AMD, ROP, DR, or BRVO in a subject.
A skilled artisan would not be able to use the method as claimed with a reasonable expectation of success based on guidance provided in the specification and art at the time of the filing of the application. Specifically, a skilled artisan would not be able to use the method to:
inhibit any mRNA involved in a retinal disease pathology by administering an shDNA comprising any ASO that is complementary to an ENG mRNA (wherein the ENG-targeting ASO comprises SEQ ID NO 21), or to any portion of a VCAM1 mRNA, a VEGF mRNA, or a HIF1α mRNA (i.e., Claims 14-15 and 20);
inhibit the expression of the any mRNA involved in a retinal disease pathology wherein the retinal disease pathology is AMD/ROP/DR/BRVO (i.e., Claim 20); or
treat any of the diseases recited in Claim 19.
The state of the art and prior art, the level of one of ordinary skill, and the level of predictability in the art: The art and prior art indicate that:
An ASO must be specific to its target, so it would not be possible to inhibit expression of any mRNA involved in a retinal disease pathology by introducing to a cell or tissue an ASO complementary to only an ENG mRNA (wherein the ENG-targeting ASO comprises SEQ ID NO 21), a VCAM1 mRNA, a VEGF mRNA, or a HIF1α mRNA.
An ASO is not guaranteed to bind and downregulate its target unless binding has been shown.
It is not possible to treat any AMD/ROP/DR/BRVO by targeting for downregulation an ENG mRNA (wherein the ENG-targeting ASO comprises SEQ ID NO 21) and a VCAM1 mRNA or a HIF1α mRNA.
Regarding point (1), the art of Crooke (et al. 2008. Mechanisms of Antisense Drug Action, an Introduction. Chapter 1 in Antisense Drug Technology: Principles, Strategies, and Applications, Crooke, ed. Boca Raton, FL: CRC Press, “Crooke”, of record on previous 892) teaches (§1.1.1 The Opportunity ¶2) ASOs allow for creating gene-selective reagents and drugs and (§1.1.2 The Challenge ¶1) the “receptor” for ASO drugs is a specific sequence of nucleotides in a target RNA. Crooke teaches (§1.1.3 Phases of Antisense Drug Action ¶3) there is evidence that ASOs can specifically hybridize to many specific RNA species and cause reduction of the target RNA but little is understood either about the process by which hybridization takes place or the kinetics of intracellular hybridization in cells. Those teachings indicate that an ASO must be specific for its target.
Furthermore, the art of Hagedorn (et al. 2017. Managing the sequence-specificity of antisense oligonucleotides in drug discovery. Nuc. Ac. Res. 45[5]:2262-2282, “Hagedorn”, of record) teaches (§Abstract) all drugs perturb the expression of many genes in the cells that are exposed to them and such gene expression changes can be divided into effects resulting from engaging the intended target and effects resulting from engaging unintended targets. Although it could be possible that any particular mRNA is downregulated as a result of the target being downregulated, that is not demonstrated to be the case for any mRNA involved in a retinal disease pathology.
In addition, the art of Mukamal (2021. Genetics and Age-Related Macular Degeneration. Am. Acad. Ophthalmol. Available online at aao.org. Accessed on 27 August 2025, “Mukamal”) and Advani (et al. 2024. QTL mapping of human retina DNA methylation identifies 87 gene-epigenome interactions in age-related macular degeneration. Nat. Comm. 15:1972, “Advani”) respectively teach (Mukamal) there are over 30 genes associated with the risk of developing AMD and (Advani) there are 87 gene-epigenome interactions in AMD. Swan (et al. 2018. The genetics of retinopathy of prematurity: a model for neovascular retinal disease. Ophthalmol. Retina 2[9]:949-962, “Swan”) teaches (Table 2) genes other than those recited in the claims that are found to have a significant association with ROP (e.g., AGTR1, BDNF, CETP). Searches for the causes of DR and BRVO revealed involvement of other genes besides those recited in the claims. Those teachings indicate that it would not be possible to inhibit expression of any mRNA involved in a retinal disease pathology (or even the specific retinal disease pathologies AMD, ROP, DR, and BRVO) by inhibiting expression of solely ENG mRNA and one of VCAM-1, VEGF, and HIF-1α mRNAs.
Those teachings indicate that it would not be possible to inhibit the expression of any mRNA involved in a retinal disease pathology by administering a nanoparticle comprising an ASO complementary to the specific set of mRNAs recited in the claims: an ENG mRNA (wherein the ENG-targeting ASO comprises SEQ ID NO 21), a VCAM1 mRNA, a VEGF mRNA, or a HIF1α mRNA.
Regarding point (2), Hagedorn teaches (§Determinants for RNase H-activity on unintended target RNA, Fig. 2) target site accessibility and hybridization between an ASO and its target affect ASO activity. Hagedorn teaches (§Target site accessibility, entire §) an RNA target can fold into complex secondary and tertiary structures, and/or can be bound to RNA-binding proteins, which affects the ability of an ASO to access its target site. Hagedorn teaches (same §, ¶3) competing interactions from higher order structures in RNA and protein binding should be assessed for every potential target site to evaluate the likelihood of oligonucleotide binding at that site. Those teachings indicate that an ASO targeting any ENG mRNA (wherein the ENG-targeting ASO comprises SEQ ID NO 21), or any portion of any VCAM1 mRNA, any VEGF mRNA, or any HIF1α mRNA would not necessarily bind to the intended target or inhibit expression of those mRNAs, let alone expression of any mRNA involved in a retinal disease pathology. Therefore, without presentation of evidence to the contrary, the teachings of Crooke, Hagedorn, Mukumal, Advani, and Swan indicate that it would not be possible to inhibit the expression of any mRNA involved in a retinal disease pathology—or the specific pathologies AMD/ROP/DR/BRVO—by administering ASOs against ENG and VCAM-1/VEGF/HIF-1α.
Further regarding points (1) and (2), the art of Uddin (et al. 2018. Real-time imaging of VCAM-1 mRNA in TNF-α activated retinal microvascular endothelial cells using antisense hairpin-DNA functionalized gold nanoparticles. Nanomedicine: Nanotechnol. Biol. Med. 14:63-71, “Uddin Nano”, of record on IDS filed 18 April 2023) indicates that the claimed strategy would not necessarily inhibit target mRNA expression. Uddin Nano teaches (§Abstract, §Specificity of AS-VCAM-1 hAuNP for mRNA target, Fig. 3) the compound of the instant claims (comprising the VCAM-1–targeting ASO) but shows that applying the AS-VCAM-1-hAuNP does not reduce VCAM-1 mRNA expression after 2 hours of incubation with the AS-VCAM-1-hAuNP. Although that experiment was intended to detect the effect of TNF-α treatment on VCAM-1 expression using the AS-VCAM-1 hAuNP and was not intended to inhibit VCAM-1 expression, results in Fig. 3 show that applying the AS-VCAM-1 hAuNP did not inhibit VCAM-1 expression after 2 hours. The experiment observed a 6.25-fold increase in reactions with AS-VCAM-1-hAuNP compared with NS hAuNP. Figs 1BCD show the AS-VCAM-1-hAuNP only makes a signal when it binds its mRNA target. Those data indicate that using the method as claimed to inhibit target mRNA expression is unpredictable.
Regarding point (3), the art indicates that it would not be possible to treat any retinal disease by administering a therapeutically effective amount of a nanoparticle comprising an shDNA comprising an ASO complementary to an ENG mRNA (wherein the ENG-targeting ASO comprises SEQ ID NO 21), any VCAM1 mRNA, any VEGF mRNA, or any HIF1α mRNA because some disease are caused mutations in genes other than those genes. A search of the art and prior art revealed no relationship between overexpression of ENG and the specific retinal disease BRVO. Searches revealed that inhibiting VEGF is a treatment for wet AMD, DR, ROP, and BRVO. However, the art of Mayo Clinic (2024. Dry macular degeneration. Available online at mayoclinic.org. Accessed on 27 August 2025, “Mayo) teaches (§Causes) no one knows exactly what causes dry macular degeneration… it may be a combination of genes and other factors, including smoking, obesity and diet, and National Eye Institute (2021. Age-Related Macular Degeneration [AMD]. Available online at nei.nih.gov. Accessed on 27 August 2025, “NEI”) teaches (§What’s the treatment for AMD?) there’s currently no treatment for late dry AMD. Those teachings indicate inhibiting VEGF can treat some—but not all—retinal diseases wherein the disease is AMD, ROP, DR, or BRVO. Those teachings, together with the above teachings of Mukamal, Advani, and Swan, indicate it would not be possible to treat any AMD, ROP, DR, or BRVO in any subject by administering the claimed NPs that comprise ASOs to ENG and one of VEGF, VCAM-1, or HIF-1α.
The art of Shen (et al. 2018. A Combination Therapy Targeting Endoglin and VEGF-A Prevents Subretinal Fibro-Neovascularization Caused by Induced Muller Cell Disruption. Invest. Ophthalmol. Vis. Sci. 59:6075–6088, “Shen”, of record) teaches (§Abstract. Fig. 4) anti-VEGF and anti-ENG therapy treat AMD and the combination of both works better.
Therefore, without presentation of evidence to the contrary, nothing in the art indicates that it would be possible to treat any AMD or AMD/ROP/DR/BRVO with the full scope of the methods of Claim 19.
Altogether, although the skill level of a person of ordinary skill is quite high, an artisan would conclude that, unless evidence is presented showing otherwise, the art and prior art indicate that inhibiting the expression of any mRNA involved in a retinal disease pathology, allowing the ASO to bind the target mRNAs of an ENG mRNA (wherein the ENG-targeting ASO comprises SEQ ID NO 21), any VCAM1 mRNA, any VEGF mRNA, or any HIF1α mRNA; and treating any AMD/ROP/DR/BRVO with the claimed methods is unpredictable.
The amount of direction provided by the specification and the existence of working examples: What is enabled by the working examples is narrow compared to the breadth of the claims.
The Spec. teaches an example of gold nanoparticles conjugated to (pp. 20-23 L10-8, Example 1) VCAM1 and a prophetic example of gold nanoparticles conjugated to (p. 23 L9-19, Example 2) endoglin. The specification teaches (p. 23 L20-25, Example 3) one example of combination nanoparticles. That § teaches that 15 nm hAuNP with two target sequences were made: sequences conjugated to 15 nm gold-nanoparticles include targeting VEGF mRNA with endoglin mRNA; and also sequences targeting VEGF mRNA with HIF-1α mRNA. Each nanoparticle contains about 48-50 of the hairpin DNAs. That § teaches that a combination nanoparticle comprises a nanoparticle with two separate target sequences and (§Example 3) teaches production of combination particles but does not show that they binding their target sequence and inhibit its expression, let alone that they inhibit the expression of any mRNA involved in a retinal disease pathology, even if the pathology is limited to AMD/ROP/DR/BRVO. Applicant has provided no data showing their method inhibits expression of the broad spectrum of genes involved in retinal disease pathology.
Furthermore, the descriptions of those Examples do not show data or call out to figures. The examples do not disclose the specific mRNA sequences that were targeted or the specific ASOs that were used.
Fig. 1 teaches that the mRNA recognition sequence is the hairpin’s loop region. The Fig. 1 description discloses (p. 4 L9-31) how the invention works and discloses probe types -1 and -2. This § teaches designing 15 nm spherical gold nanoparticles that were designed and functionalized with single or multiple targeted hairpin-DNA/RNA oligonucleotides incorporating ASO sequence specific for mouse ENG mRNA, or VEGF mRNA, or VCAM-1 mRNA, or HIF-1α mRNA (AS-hAuNP), or a scrambled version of this sequence (NS-hAuNP). The Fig. 1B caption discloses 1.4 nm gold nanoparticles functionalized with a shDNA/RNA oligonucleotide incorporating ASO sequence specific for mouse ENG mRNA, or VEGF mRNA, or VCAM-1 mRNA, or HIF-1α mRNA (AS-hAuNP) or a scrambled version of this sequence (NS-hAuNP). The caption indicates that the sequences shown are SEQ ID NOs 67-68. The Fig. 1C caption discloses the AS-mENG hAuNP is highly specific for its complementary sequence. However, that caption does not disclose the SEQ ID NO of either the ASO or the target used in the ASO-mRNA binding experiment. The Fig. 1D caption discloses imaging of AS-hAuNP in highly ionic media. All of the ENG-targeting ASOs are mouse ENG (i.e., “mENG”), not SEQ ID NO 21 (which the Spec. teaches on p. 26 is human ENG), but none of them is shown to inhibit expression of ENG mRNA. Yet, Claims 14-15 recite binding and inhibiting expression of any mRNA.
Fig. 2 and its caption disclose retinal cells treated with hypoxia and then administered antisense gold nanoparticles (AS-hAuNP) and identifies that the AS-hAuNP interact with ENG mRNA in the cell and fluoresce which indicates those ASOs bind target ENG mRNA. Fig. 3 and its caption disclose that a VCAM1-targeting AS-hAuNP was administered to VCAM1 mRNA and the target was depleted by over 67%. The Spec. discloses only a single VCAM-1–targeting AS-hAuNP—SEQ ID NO 66—so presumably that was the ASO used.
The Fig. 4 caption discloses that two AS-hAuNP were administered to choroidal tissue and show that the lesion size was reduced to a similar extent as tissue administered positive control anti-VEGF antibodies. The caption teaches those results show that it’s possible that the VEGF-targeting ASOs can treat AMD. However, that caption does not disclose the SEQ ID NO of either of those ASOs or their targets.
Fig. 1 shows that SEQ ID NO 67 binds ENG mRNA comprising SEQ ID NO 68. The Spec. does not show or describe using ASOs targeting ENG mRNA (wherein the ASO comprises SEQ ID NO 21) or HIF-1α mRNA to inhibit expression of any mRNA involved in a retinal disease pathology. The Spec. does not show or describe using ASOs targeting ENG mRNA wherein the sequence comprises SEQ ID NO 21, any VEGF mRNA (aside from the unidentified ASOs used in the experiment whose results are shown in Fig. 4), any VCAM-1 mRNA (aside from that targeted by the ASO comprising SEQ ID NO 66), or any HIF-1α mRNA, wherein the ASO binds the target sequence and the binding of the ASO to the target sequence inhibits RNA expression. No data showing any binding between any mRNA involved in a retinal disease pathology or involved in specifically AMD/ROP/DR/BRVO and SEQ ID NO 21 are shown. No data showing any binding between any ASO and any HIF-1α mRNA are shown. The only VCAM-1 mRNA-targeting ASO (results shown in Fig. 3) disclosed is SEQ ID NO 66. The only VEGF mRNA-targeting ASOs are shown in Fig. 4 but the SEQ ID NOs used in those experiments are not identified.
Only Fig. 4 shows results from using VEGF-targeting ASOs to reduce choroidal lesions. No details about whether that is a model of the single retinal disease AMD are provided. No other models of any retinal diseases are shown.
However, aside from the VCAM mRNA targeting ASO SEQ ID NO 66 (Fig. 3), and the unidentified VEGF mRNA-targeting ASOs (Fig. 4), the Specification provides no evidence of using nanoparticles comprising ASOs targeting ENG mRNA (wherein the ASO comprises SEQ ID NO 21), or VEGF mRNA, or VCAM-1 mRNA, or HIF-1α mRNA to inhibit expression of any mRNA involved in a retinal disease pathology. Aside from the limited data described, the Spec. shows no other ASOs binding any ENG mRNA, or VEGF mRNA, or VCAM-1 mRNA, or HIF-1α mRNA target sequences and no evidence of treating any retinal disease using the unidentified VEGF-targeting ASOs—aside from AMD, although details of the model are not provided. Yet, Claims 14-15 and 19-20 claim methods directed to inhibiting any mRNA, using shDNAs comprising ASOs to bind ENG mRNA (specifically using an ASO comprising SEQ ID NO 21), or VEGF mRNA, or VCAM-1 mRNA, or HIF-1α mRNA target mRNAs and inhibit mRNA expression, and treating any retinal disease including any AMD, ROP, DR, and BRVO.
Altogether, the Spec. does not demonstrate successfully using the methods as claimed and does not provide guidance for doing so. In summary, the guidance present in the specification does not address the enablement issues raised in view of the state of art discussion presented above. The specification does not describe how an ASO complementary to the recited targets could inhibit expression of any mRNA involved in any retinal disease pathology, and does not describe sequences that are demonstrated to bind and inhibit the recited targets (aside from SEQ ID NO 66 inhibiting VCAM-1 and the unidentified VEGF-targeting sequences inhibiting VEGF). The Spec. does not demonstrate that the claimed methods could be used to treat any retinal disease besides for the unidentified VEGF-targeting sequences treating AMD.
As described in the discussion of the state of the art, the art regarding inhibiting expression of any RNA by administering an ASO targeting an ENG mRNA (wherein ASO comprises SEQ ID NO 21), VEGF mRNA, VCAM-1 mRNA, or HIF-1α mRNA; the art of having an ASO bind and inhibit those targets wherein binding or inhibition have not been demonstrated; and the art of treating any retinal disease by administering the claimed NP are unpredictable. Note that the art does teach inhibiting VEGF is used to treat wet AMD, ROP, DR, and BRVO, so a method that requires a VEGF-targeting ASO to treat those diseases (i.e., wet AMD, ROP, DR, and BRVO) could be considered enabled. However, the art indicates there is no treatment for dry AMD. Furthermore, the claims do not require the VEGF-targeting ASO to be included in the NP. Therefore an artisan would determine that the methods as currently recited could not be used as claimed.
The quantity of experimentation needed to make or use the invention: The standard of an enabling disclosure is not the ability to make and test if the invention works but one of the ability to make and use with a reasonable expectation of success. A patent is granted for a completed invention, not the general suggestion of an idea (MPEP 2164.03 and Chiron Corp. v. Genentech Inc., 363 F.3d 1247, 1254, 70 USPQ2d 1321, 1325-26 (Fed. Cir. 2004). The instant specification is not enabling because one cannot follow the guidance presented therein or within the art at the time of filing, and practice the claimed method without first making a substantial inventive contribution. Given the teachings described above, an artisan of ordinary skill would not be able to use the invention as claimed with a reasonable expectation of success. The amount of experimentation required for enabling guidance commensurate in scope with what is claimed goes beyond what is considered “routine” within the art and constitutes undue further experimentation in order to successfully use the methods of inhibiting expression of any mRNA involved in any retinal disease pathology or treating any AMD, DR, ROP, or BRVO by administering a nanoparticle comprising two shDNAs: an shDNA comprising an ASO that consists essentially of SEQ ID NO 21 and targets ENG mRNA and an shDNA comprising an ASO that targets VCAM-1 mRNA, VEGF mRNA, or HIF-1α mRNA; wherein the shDNAs are conjugated to a fluorescent dye, and wherein a colloidal gold nanoparticle is conjugated to the shDNAs.
Claims 14 and 19-20 are rejected for lack of enablement. Claims 15 and 20 are rejected because they depend from Claim 14 and don’t remedy the issues.
In conclusion, because the Spec. shows (Figs. 3-4) inhibiting expression of VCAM-1 and VEGF with specific ASOs, the specification provides enablement for:
A method for inhibiting the expression of an mRNA involved in a retinal disease pathology, wherein the retinal disease pathology is selected from age-related macular degeneration (AMD), retinopathy of prematurity (ROP), diabetic retinopathy (DR), or branch retinal vein occlusion (BRVO) and wherein the mRNA involved in a retinal disease pathology is either VCAM-1 mRNA or VEGF mRNA, the method comprising:
introducing a nanoparticle into a cell or a tissue, the nanoparticle comprising:
two or more short hairpin DNA sequences (shDNAs), wherein a first shDNA comprises an anti-sense oligonucleotide complementary to a target sequence consisting of an endoglin mRNA, wherein a second shDNA comprises an anti-sense oligonucleotide complementary to a target sequence comprising a VCAM-1 mRNA, wherein the ASO comprises SEQ ID NO:66, or a VEGF mRNA, wherein the ASO comprises [the ASOs whose results are shown in Fig. 4], wherein each of the two or more shDNA is conjugated to its own fluorescent dye, and wherein the anti-sense oligonucleotide complementary to the target sequence of the endoglin mRNA consists essentially of SEQ ID NO:21;
and a colloidal gold nanoparticle conjugated to the two or more shDNAs;
and allowing the anti-sense oligonucleotide to bind the target sequence;
wherein the binding of the anti-sense oligonucleotide to the target sequence inhibits the expression of the mRNA that is targeted by the ASO.
Because the art teaches inhibiting VEGF to treat ROP/DR/BRVO/wet AMD and the specification shows (Figs. 3-4) inhibition of VEGF, the following is enabled:
A method for treating age-related macular degeneration (AMD), wherein the AMD is caused by overexpression of VCAM-1 or VEGF, retinopathy of prematurity (ROP), diabetic retinopathy (DR), or branch retinal vein occlusion (BRVO) in a subject, comprising:
administering a therapeutically effective amount of a nanoparticle to the subject, wherein the nanoparticle comprises:
two or more short hairpin DNA sequences (shDNAs), wherein a first shDNA comprises an anti-sense oligonucleotide complementary to a target sequence consisting of an endoglin mRNA, wherein a second shDNA comprises an anti-sense oligonucleotide complementary to a target sequence comprising a VEGF mRNA, wherein the ASO comprises [the ASOs whose results are shown in Fig. 4], and wherein an additional shDNA comprises an antisense oligonucleotide complementary to a VCAM-1 mRNA or a HIF-1a mRNA, wherein the two or more shDNA are each conjugated to a separate fluorescent dye, and wherein the anti-sense oligonucleotide complementary to the target sequence of the endoglin mRNA consists essentially of SEQ ID NO:21;
and a colloidal gold nanoparticle conjugated to the two or more shDNAs.
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