DETAILED ACTION
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
This action is in response to an amendment filed 9/15/ 2025.
Claims 26-54 are pending. Claims 53 and 54 are newly added.
This application is a § 371 National Stage Application of PCT/US2020/037740, filed June 15, 2020, which claims the benefit of U.S. Patent Application Serial No. 62/861,186, filed on June 13, 2019.
Information Disclosure Statement
An information disclosure statement filed 9/15/2025 has been identified and the documents considered. The corresponding signed and initialed PTO Form 1449 has been mailed with this action. Initials indicate that the document has been considered even if the reference is lined through.
Response to Amendments
The amendments to the claims are effective in overcoming the objections to the claims. As well, the amendments are sufficient to overcome the previous rejection under 35 USC 112, second paragraph. However, the amendments have elicited a new such rejection set forth below.
The terminal disclaimers filed on 9/15/2025 disclaiming the terminal portion of any patent granted on this application which would extend beyond the expiration date of any patent granted on U.S. 12,315,814 and 12,351,815 have been reviewed and is accepted. The terminal disclaimer has been recorded.
Claim Rejections - 35 USC § 112, second paragraph
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.
Claim 26 is unclear for reciting in part (d) that the particle does not comprise “any” non-human viral gag, pol, “or” virally derived envelope protein. Use of the term “any” with “or” makes the claim unclear as “any” means all of them and “or” means some of them. As well, the term “derived” is unclear in that the metes and bounds of the term “derived” are unclear. It is unclear the nature and number of steps required to obtained a “derivative” of a viral envelope protein. The term implies a number of different steps that may or may not result in a change in the functional characteristics of the component from the source that it is “derived from”. This is true of claim 40. These are new rejections necessitated by applicants’ amendment.
Claim Rejections - 35 USC § 112, first paragraph
The following is a quotation of the first paragraph of 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 26-52 are rejected under 35 U.S.C. 112, first paragraph, because the specification, while being enabling for a method of delivering a therapeutic cargo to a target cell in vitro, by contacting the target cell with a particle comprising a phospholipid bilayer membrane comprising a HERV glycoprotein, a Pleckstrin homology domain (PH) or a human activity regulated cytoskeleton associated protein (hArc) and the therapeutic cargo wherein the cargo is delivered by the particle to the target cell after uptake of the particle by the cell, does not reasonably provide enablement for any other embodiment. The specification does not enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the invention commensurate in scope with these claims. This rejection is maintained.
The test of enablement is whether one skilled in the art could make and use the claimed invention from the disclosures in the patent coupled with information known in the art without undue experimentation (United States v. Telectronics, Inc., 8 USPQ2d 1217 (Fed. Cir. 1988)). Whether undue experimentation is required is not based on a single factor but is rather a conclusion reached by weighing many factors (See Ex parte Forman, 230 USPQ 546 (Bd. Pat. App. & Inter, 1986) and In re Wands, 8USPQ2d 1400 (Fed. Cir. 1988); these factors include the following:
1) Nature of invention. The instant claims are drawn to a method of delivering cargo to a cell (claim 26) and further modifying a cell (claim 40) by contacting a particle comprising a phospholipid bilayer membrane comprising a HERV glycoprotein and a PH domain or an hARC a therapeutic cargo. The therapeutic cargo in claim 40 is “a genome modulating reagent”.
2) Scope of the invention. The scope of the invention is extremely broad in that the method is neither limited as to target cell type or whether the method is in vitro except in newly added claims 53 and 54. In addition to the lack of characterization of the target cell, the cargo is also not-limited in the base claim 26 and the genome modulating reagent in claim 40.
3) Number of working examples and guidance. The specification teaches only in vitro methods of producing heVLP in HEK 293 cells by PEI transfection of plasmids expressing 1) hPLCd1 PH Cas9 fusion, 2) HERVK GAG, 3) ENV. Produced heVLPs were harvested and purified which were then used in vitro to deliver the cargo to cells in culture.
4) State of the art. The disclosure is directed to methods of making heVLP or HERV derived viral like particles. HERV are endogenous retroviral sequences found in the human genome. They make up 8% of the total human genome and comprises gene sequences retained through evolution. Some do produce active viral particles that are replication defective. The glycoproteins encoded by HERV are highly fusogenic (Song, abstract).
Early experimental systems used baculovirus to produce HERV-K VLP (Tonjes et al, figure 1).
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Raguram et al teach that art derived methods of encapsulating proteins or RNP into VLP requires gag and cargo linked by short peptide sequences (page 19, past ¶). Other methods fused the cargo to VPR and related molecules.
5) Unpredictability of the art. The claims are drawn to a method of delivering a therapeutic cargo to a target cell. Hence, the desired goal is that the cargo is delivered to the cell which based on the disclosure is intended that this cargo is delivered internally to the cell. The figure 2 below depicts applicants design.
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[0041] FIG. 2: Depiction of purified heVLPs entering a target cell and delivering cargo to the cytosol. Importantly, the human-endogenous GAG or other phospholipid bilayer recruitment domain allows cargo to enter the target cell nucleus as long as cargo possesses a nuclear localization sequence.
The art (Raguram et al, page 6) teaches that there are several obstacles to delivering the therapeutic cargo to a target cell as broadly claimed in terms of cell type and in vivo delivery. First, the route of administration is critical for the target cell and secondly the cell type is restricted as the VLP must engage specific receptors on the surfaces of the target cells to promote entry. Finally,
Therefore, successful vehicles must escape endosomes to release their cargos outside of endosomes and into the cell cytosol. Many successful delivery vehicles exploit the acidic environment of endosomes to trigger changes in the vehicle’s structure that promote endosomal escape and cargo release (Mitchell et al., 2021; Staring et al., 2018). Importantly, efficient delivery vehicles must be stable enough to protect their cargo while outside of cells, but must be able to disassemble and release their cargo after entering cells and escaping endosomes.
To this end, Raguram teaches that the cargo for gene editing are mRNA and protein (also called RNPs). This seems to be understood in the art.
The in vitro systems did not recapitulate well in vivo.
While various Cas9 RNP-packaging VLPs exhibited promising efficiencies in vitro, all of the systems described above were either not tested in vivo or exhibited low in vivo efficacy (<10% editing). We recently developed engineered VLPs (eVLPs) based on Moloney MLV (MMLV) that package Cas9 nuclease or base editor RNPs and mediate potent, therapeutic levels of gene editing across multiple organs in mice (Banskota et al., 2022).
The main obstacle for in vivo delivery to a target cell. Reference is made to Banskota et al (see pages 251-254) as to fourth generation eVLP and means to work through these obstacles. Some limited success in delivery was achieved. However, applicants have not demonstrated the viability of delivery to target cells with the instant particles.
6) Undue experimentation. The claims have been evaluated in light of the art at the time of filing and found not to be commensurate in scope with the specification. MPEP 2164.05 teaches, “However, the examiner should carefully compare the steps, materials, and conditions used in the experiments of the declaration with those disclosed in the application to make sure that they are commensurate in scope; i.e., that the experiments used the guidance in the specification as filed and what was well known to one of skill in the art. Such a showing also must be commensurate with the scope of the claimed invention, i.e., must bear a reasonable correlation to the scope of the claimed invention. The invention recites use of a broad group of target cells to be delivered the heVLP with a broad set of cargos and genome modulating reagents. Given the unpredictability of the art, the poorly developed state of the art with regard to predicting the cellular uptake, the lack of adequate working examples and the lack of guidance provided by applicants, the skilled artisan would have to have conducted undue, unpredictable experimentation to practice the claimed invention.”
Response to Arguments
HERV glycoproteins are the proteins that make up the HERV envelope. The phospholipid bilayer of the instant particle comprises a HERV glycoprotein. Applicants point to provided for references as evidence of the success of in vivo delivery. Applicants argue that Blaise et al teaches the fusogenic nature of HERVW and HERV-FRD. This paper does identify the first two HERV proteins and demonstrates that they are fusogenic. Analysis also indicates that the HERVW receptor is the D type retrovirus receptor and that HERV-FRD receptors are not known (page 13017, col 2 and page 13018, col 2).
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Applicants argue that Coquin was submitted with the response, it was found in the IDS filed 2/13/2023. Applicants argue that this reference demonstrates in vivo delivery with lentivirus pseudotyped with either of these proteins. The specific results shown are pseudotyped lentiviral particles wherein the pseudotyped particles were made with Syn1 (HEnvW) and Syn2 (ERVFRD-1). This is a limited scope of the instant claims which require any phospholipid bilayer and a glycoprotein from any HERV. SynA and SynB are mouse versions of Syn 1 and 2 wherein Coquin teaches “In mice, syncytin-A (SynA) and syncytin-B (SynB) which are phylogenetically unrelated to human Syn1 and Syn2 have entered the Muridae lineage more than 20 million years ago”. Coquin teaches that the use of this limited version of the instant claims was not possible to use as a transduction vector (see page 4, first full ¶. To this end, Coquin adds an inventive concept to improve use of pseudotyped LV by addition of Vectofusion, “a peptidic transduction additive identified in our laboratory”. As to in vivo analysis of the particles of Coquin et al, these are studies performed in mice with LV-SynA and LV-SynB in the presence of VF1. The effect was transduction of spleen cells and specifically spleen B cells which matches as the receptors are expressed on these cells. It is further known that transduction of cells in mice does not provide evidence of delivery in humans. This is a well known fact in gene therapy. Issues include organ barriers, failure to persist, side-effects in other organs, virus neutralizing antibodies, humoral immunity, normal tropism of the vector to other organs and more. In fact, applicants own arguments in response to the art rejections confirms this was well known (see reference to this in the response to applicants arguments regarding art). The challenge is to maintain the efficiency of delivery and expression while minimizing any pathogenicity of the sequences formulating the particle. These issues are touched on in Raguram, see page 19 and 23 ¶2. Raguram also teaches that human usage requires further study (see page 22).
Applicants argue that these results are extended by demonstration that HERV-K is also fusogenic. As an aside, neither HERV-W nor HERV-FRD were shown to successfully provide delivery to target cells in vivo in humans. Considering the effect with HERV-K, applicants refer to Laderoute and Contraras-Galindo. Laderoute and Contreras-Galindo do not demonstrate activity of an HERV-K particle in vivo in humans. Hence, there is no demonstration that these particles are effective in vivo in a manner that overcome the art demonstrated obstacles
Applicants argue that the lack of delivery to all cells doesn’t mean the methods aren’t enabled. However, the issue is there is no evidence that the particles deliver the cargo to the target cell. By calling it a target cell it is a specific cell. However, no cells in vivo are shown to be delivered of cargo. Hence, applicants provide no guidance for the claimed invention except for the ability to transduce cells in vitro with a pseudotyped particle. The only results are presented in ¶0242.
[0242] HEK 293T cells were transduced with T1heVLPs containing PLC PH fused to spCas9, hGAGKcon fused to spCas9, or hArc fused to spCas9 targeted to VEGF site #3. T1heVLPs were pseudotyped with either hENVW (left chart) or hENVFRD (right chart). Gene modification was measured by amplicon sequencing. Particle purification and concentration was performed by PVDF filtration and ultracentrifugation at 100,000×g for 2 hours. Results are shown in FIG. 3. Importantly, if HERV-derived GAG (hGAGKcon) was not overexpressed by itself in producer cells, then efficient delivery was not achieved.
This is simply in vitro transduction of HEK293 cells with a very differently described particle that is pseudotyped with either hENV-W or hENV-FRD wherein internal to the pseudotyped coat is a plasma recruitment domain i.e. PH or hARC is fused to cargo i.e. Cas9. The claims to the contrary are drawn to a particle comprising a HERV glycoprotein (which is a single envelope protein) wherein internal to the particle is a plasma recruitment domain and the cargo. To distinguish from the art, the claims have been amended to omit non-human viral proteins i.e. gag, pol and env. Given the predictability of transducing cells in vitro one would expect this to be proof of principle for the claimed invention in vitro. However, the art of transducing “target” cells in vivo is highly unpredictable. For example, Raguram teaches on page 6,
In all cases, successful in vivo delivery of gene editing agents requires overcoming several biological and molecular barriers to the intracellular delivery of macromolecules. Specifically, an efficient in vivo delivery vehicle must (1) package and protect its cargo from sequestration or destruction before it enters cells, (2) bind desired cells, (3) traverse the target cell membrane to access the cellular interior, and (4) release its cargo into the appropriate intracellular compartment (Figure 2).
Applicants have not demonstrated that the particles avoid recognition by the immune system (obstacle 1 detailed by Raguram, page 6, ¶3). Applicants have not demonstrated that the vehicle can bind to “a target cell” in the body particularly as these are not specifically claimed. The issues are organ barriers, sequestration and inability to enter the cell (obstacle 2, ¶4 above). The third obstacle provided by Raguram is that once the vehicle can cross the cell membrane, endosomal trapping becomes an issue unless the vehicle disassemble in the cell (¶5, above).
Applicants have demonstrated identification of two HERV glycoproteins and demonstrated that they have fusogenic activity. Applicants have not demonstrated what target cell is targeted by the two such that the target cell is defined in the claims. Binding of glycoproteins is receptor specific and hence cell specific but the disclosure of the two glycoproteins without knowledge of what target cells applicants hope to deliver and modify, and in view of the clear obstacles set forth above of the ability to target cells in humans in vivo, one would have to perform undue, unpredictable experimentation to practice the claimed invention.”
Claim Rejections - 35 USC § 103
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
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 26-29, 34, 36, 38, 40-42 and 52-54 are rejected under 35 U.S.C. 103 as being unpatentable over King et al (US 11,028,383) in view of Maltzahn et al (US 20210228627). This rejection is updated based upon applicants amendment.
As regards claim 26, King et al teach construction of multimeric assemblies comprising a lipid bilayer enveloping cargo that includes proteins and nucleic acids (abstract, col 2, line 47-50). The outer layer is made of cell membrane.
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It can comprise envelope proteins from a virus but does not require this. It copmris3es what they call M domains.
In one embodiment, the one or more M domains comprise a polypeptide having an acylation motif (including but not limited to N-terminal myristoylation motifs, palmitoylation motifs, farnesylation motifs, and geranylgeranylation motifs), a polar headgroup-binding domain (including but not limited to those described herein and in the attached appendices), envelope proteins of enveloped viruses, membrane protein transporters, membrane protein channels, B-cell receptors, T-cell receptors, transmembrane antigens of human pathogens, growth factors receptors, G-protein coupled receptors (GPCRs), complement regulatory proteins including but not limited to CD55, CD59, and transmembrane protein domains. In a further embodiment, the one or more M domains are selected from the group consisting of SEQ ID NOS: 52-151 and 280-300.
As to GAG, there is no requirement for GAG in the complex either. It is used in an assay to demonstrate budding and a domain from GAG is optionally part of the
(134) Any viral structural protein that is known to be defective in budding can be used in the budding assay, including but not limited to budding-defective versions of HIV-1 Gag, RSV Gag, MuMoLV Gag, SV5 M, Ebola VP40 and other structural proteins from different families of enveloped viruses including retroviruses, filoviruses, rhabdoviruses, arenaviruses, and paramyxoviruses. In addition, as the inventors describe below, the multimeric assemblies of the invention can be used to test the ability of an L domain to effect membrane scission and release of an enveloped multimeric assembly in a similar manner.
This M domain can include pleckstrin (see e.g. col 73, line 3-13).
(199) In another embodiment (SEQ ID 228; Late2-4GS-I3-01-10GS-PH-flag), 22 residues of the Ebola VP40 protein encompassing the polypeptide motif PTAPPEY, which is known to recruit the ESCRT pathway to facilitate the budding and release of Ebola from host cells, were fused to the N terminus of I3-01 to provide an L domain and the pleckstrin homology (PH) domain of the rat phospholipase C-δ1 protein was fused to the C terminus of I3-01 to provide an M domain; the construct also includes a FLAG tag to facilitate specific detection of the protein using anti-FLAG antibodies.
As shown in the figure, the particle is delivered to a target cell.
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King et al did not teach that the HERV is composed of HERV glycoproteins.
However, lipid bilayers comprising HERV glycoproteins as fusogens were known in the art (see von Maltzahn ¶ 0207). Von Maltzahn et al teach delivery of Zn finger nucleases as well as TRALEN and CRISPR plus gRNA (see e.g. ¶0472-0473) as taught in claim 34, 40-42.
Based on such teachings, it would have prima facie been obvious to one of ordinary skill in the art at the time the invention was made to combine the teachings of King et al with that of von Maltzahn. Such a modification would have resulted in a method encompassed by claim 26 and 40. Each are drawn to analogous art of creating delivery agents comprised of lipid bilayers and glycoproteins to mediate cell entry. Further 1) King teaches design of lipid bilayers wherein the particle comprises the cargo, a glycoprotein and a pleckstrin domain 2) von Maltzahn et al teaches a related lipid bilayer particle wherein the glycoprotein is from HERV which is a fusogen and mediates targeted delivery to cells by facilitating a connection between the VLP and the cell (see e.g. col 9, line 44-62). Thus, a person of ordinary skill in the art, absent evidence to the contrary, would have reasonably expected that the combination.
The cargo can be fused to the M domain. The non-covalent linkage between the M domain and the cargo can be present and hence allows for no linkage as recited in claim 27.
As recited in claims 53 and 54, King teaches the that the compositions can be used in vitro (see figure 8).
(15) In one embodiment, the M domain is capable of noncovalently- covalently interacting with a lipid bilayer. In another embodiment, the L domain is capable of non-covalently interacting with one or more proteins in the ESCRT machinery or proteins known to recruit the ESCRT machinery to the site of budding by binding to one or more ESCRT proteins directly or indirectly.
Claims 30-33, 35, 37, 39 and 43-51 are rejected under 35 U.S.C. 103 as being unpatentable over King et al (US 11,028,383) in view of Maltzahn et al (US 20210228627) as applied to claims 26-29, 34, 36, 38, 40-42 and 52 above, and further in view of Zhang (US 20140256046), Haugwitz et al (WO 2015199111) and Lin et al (US20170065588).
As regards claims 30-33, 35, 37, 39 and 43-51, Zhang teaches that dCas9 is a catalytically inactive Cas9 to be used in therapeutic approaches and furthermore can be fused to transcriptional activators and repressors (KRAB) as well as methylation domains (see ¶0240).
Cargo proteins in such constructs are shown to have improved loading when used with A/C heterodimers such as DmrC or DmrA (see figure 2 of Haugwitz).
An improved PH domain from Akt1 leads to improve membrane translocation as shown by Lin et al (¶0530).
Based on such teachings, it would have prima facie been obvious to one of ordinary skill in the art at the time the invention was made to. Such a modification would have resulted in a method encompassed by claims 30-33, 35, 37, 39 and 43-51. As set forth above 1) King teaches design of lipid bilayers wherein the particle comprises the cargo, a glycoprotein and a pleckstrin domain 2) von Maltzahn et al teaches a related lipid bilayer particle wherein the glycoprotein is from HERV which is a fusogen and mediates targeted delivery to cells by facilitating a connection between the VLP and the cell (see e.g. col 9, line 44-62), 3) Zhang teaches dCas9 improves gene editing methods and can be improved further by coupling with epigenetic modulators 4) Haugwitz teaches improved cargo loading with A/C heterodimers which is the way that the particles are made in the instant claims and 5) Lin et alt each that6 Akt1 and mutant Akt1 is improved for membrane translocation. Thus, a person of ordinary skill in the art, absent evidence to the contrary, would have reasonably expected that the improvements would have been preferable to the ordinary practitioner in building a particle for delivery to target cells.
Response to Applicants arguments
Applicants argue that King and von Maltzahn teach use of non-human viral gag, pol or virally derived envelope proteins and this are occluded by the new claim limitation. As well, applicants argue that the art would not have used HERV glycoproteins as they elicit immune responses unsuitable for therapy. This is supported by applicants with reference to Latzahn and Holst and Chatterjee.
As to the first issue, the claims are unclear as set forth above. The claim requires in one interpretation that there is a lack of one of gag, pol or env from virus that are non-human. King does not teach use of any pol. The only mention of pol is its use to derive a packaging domain (bridging ¶, col 81-81). Hence, King et al do meet the newly amended claims as interpreted above. Considering the omission of all three proteins, the structure need not comprise any of these proteins. The inclusion of envelope and gag are optional. The membrane is cell membrane, PH and cargo are transported and optionally there is involvement of viral env and gag but these are not part of the construct. As to the lack of support for the use of these delivery agents, immune reactions are not relevant in in vitro settings. King teaches cell culture use for these constructs.
Conclusion
Copending application 18/351,800 is directed to cells comprising a nucleic acid encoding a HERV glycoprotein and a cargo fused to a plasma recruitment domain. This cell is used to produce particles. The compositions and steps of the methods are distinct from the instant claims in that
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
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/MARIA MARVICH/ Primary Examiner, Art Unit 1631