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 .
Claim Status
Claims 172-191 are currently pending in this application.
Election/Restrictions
Applicant’s election without traverse of Group I, claims 172-189, in the reply filed on Oct. 23, 2025 is acknowledged. Applicant’s election of the species of Oct3/4 comprising SEQ ID NO: 33 and the Oct3/4 having SEQ ID NO: 1 is acknowledged. Claims 190-191 have been withdrawn from consideration as being drawn to non-elected subject matter, and claims 172-189 have been considered on the merits.
Objections
Claim 174 is objected to because of the following informalities: claim 174 redundantly recites “wherein: (a) wherein.” Appropriate correction is required.
In claim 177, the term “M-6-methyladenosine” is abbreviated as “m6A;” however, the ordinary meaning of “m6A” in the art is for N6-methyladenosine (see Chen et al., Mol Cell 76: 96-109 (2019)). Appropriate correction is requested.
Claim Interpretation
In the claims, the term “induced pluripotent stem cell (iPSC)” is interpreted in light of the instant specification at [0073] as being limited to cells that must be derived from a differentiated somatic cell, which is defined to mean any cell that is not pluripotent (capable of producing cells from all three germ layers) ([0074]), using the terms “differentiated” and “somatic” interchangeably.
In the claims, the term “contacting” is interpreted under a broadest reasonable interpretation as encompassing contacting a cell with a single RNA molecule or plurality of identical RNA molecules (e.g., coincubation) as well as transfecting, electroporating, into the cell a single RNA molecule or a DNA encoding and expressing a recombinant circular RNA (see e.g., instant [0045], [0132], [0141], [0143]-[0144]).
In claim 174, the phrase “has the sequence of” with regard to a reprogramming factor and a recited SEQ ID NO. is interpreted as meaning the protein “consists” of a polypeptide with the recited sequence.
In claim 182, the phrase “contacting the somatic cell with at least 2, at least 3, at least 4, at least 5, at least 6 circular RNAs encoding Oct4, Sox2, Klf4, C-Myc, L-Myc, Lin28, or Nanog” is interpreted as encompassing contacting the cell with six copies of the same circular RNA or six different circular RNAs and intermediate permutations wherein each circular RNA may comprise one, some, or all of the sequences encoding Oct4, Sox2, Klf4, C-Myc, L-Myc, Lin28, or Nanog.
For claims 180-181, the lipid nanoparticle (LNP) complex is interpreted in view of [0083] to encompass any lipid molecule conjugated to a circular RNA wherein the lipid molecule is less than a micron in size, such as conjugation via a covalent bond, electrostatic adsorption, or a biotin-streptavidin interaction.
Claim Rejections - 35 USC § 112(a) - Written Description
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 172-173, 176-181, and 184-189 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.
The claimed invention as a whole is not adequately described if the claims require essential or critical elements that are not adequately described in the specification and that is not conventional in the art as of applicant’s effective filing date. Possession may be shown by actual reduction to practice, clear depiction of the invention in a detailed drawing, or by describing the invention with sufficient relevant identifying characteristics such that a person skilled in the art would recognize that the inventor had possession of the claimed invention. Pfaff v. Wells Electronics, Inc., 48 USPQ2d 1641,1646 (1998).
In making a determination of whether the application complies with the written description requirement under 35 U.S.C. 112(a) or 35 U.S.C. 112, first paragraph, it is necessary to understand what Applicant is claiming and what Applicant has possession of.
The claims are directed to methods comprising contacting a somatic cell with at least one of Oct3/4, Klf4, Sox2, Nanog, Lin28, c-Myc, L-Myc, or a fragment or variant thereof. The claims are broad in that neither “fragment” nor “variant” is defined in the claims or the specification and thus encompasses a wide genus of any fragment and any variant.
In analyzing whether the written description requirement is met for genus claims, it is first determined whether a representative number of species have been described. In the instant case, the specification fails to provide a single species of fragment of any of the recited factors. The specification describes variants as non-exclusively including factors having at least 90% identity to a factor listed in instant Table 1. Furthermore, nowhere does the application provide a working example using a fragment or variant.
The prior art teaches functional fragments of Oct3/4, Klf4, Sox2, Nanog, and c-Myc, albeit often exhibiting reduced activity, wherein each at least comprises a DNA binding domain and typically also requires a trans-activation and/or repressor domain (see e.g., Schuetz et al., Cell Mol Life Sci 68: 3121-31 (2011); Theunissen et al., Development 138: 4853-65 (2011); Zhang et al., Cell Stem Cell 19: 66-80 (2016)). The prior art also teaches variants of Oct3/4, Klf4, Sox2, Nanog, and c-Myc, such as chimeric fusions and sequence variants engineered to replace post-translationally modified residues, eliminate ubiquitination sites, replace post-translational redox-sensitive residues, e.g., Oct4(S151M), Klf4(K232R), Sox2(S248D), Nanog(L122A), and Myc(T58A) (Malik et al., Nat Commun 10: 3477 (2019); Wang et al., Stem Cell Res 25:88-97 (2017); Myers et al., Elife 7:5:e10647 (2016); Lim et al., BBRC 443: 1206-10 (2014); Navarro et al., Stem Cells 32: 436-46 (2014); WO2018214534A1; US10155929B2; US10508265B2).
The skilled artisan could not rely upon the disclosure in the specification such that the specification would sufficiently describe that Applicant was in possession of all the fragments and/or variants of Oct3/4, Klf4, Sox2, Nanog, Lin28, c-Myc, and L-Myc encompassed by the claims that are predictably capable of reprogramming a somatic cell into an IPSC beyond ones already validated in the prior art.
35 USC § 112(a), Scope of Enablement
Claims 172-189 are rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, because the specification does not enable any person, skilled in the art to which it pertains or with which it is most nearly connected to, to produce a reprogrammed iPSC merely by contacting any somatic cell with a circular RNA(s) encoding only one or two of Oct3/4, Klf4, Sox2, Nanog, Lin28, C-Myc, and L-Myc, or a fragment or variant thereof.
Although the vast majority of methods encompassed by the instant claims lack enablement, there are enabled embodiments for wherein a minimum of three factors are included selected from: (1) Oct4, Sox2, and Klf4 (OSK); (2) Oct4, Sox2, Nanog, and Lin28 (OSNL); and (3) Sox2, Klf4, and c-Myc (SKM) and wherein Lin28 is not a fragment or variant; and wherein the contacting comprises some means of introducing the RNA into the interior of the cell, i.e., to pass through the plasma membrane or be synthesized within the cell, and wherein each contacting comprises multiple RNA molecules and provides for sustained expression (whether continuous or noncontinuous) over a period of 7 days or more (which may require repeated contacting events depending on the half-life of the RNA).
Enablement is considered in view of the Wands factors (MPEP 2164.01 (a)). The court in Wands states that "Enablement is not precluded by the necessity for some experimentation such as routine screening. However, experimentation needed to practice the invention must not be undue experimentation. The key word is 'undue.' Not 'experimentation;" (Wands, 8 USPQ2d 104). Clearly, enablement of a claimed invention cannot be predicated on the basis of quantity of experimentation required to make or use the invention. "Whether undue experimentation is needed is not a single, simple factual determination, but rather is a conclusion reached by weighting many factual considerations." (Wands, 8 USPQ2d 1404).
The factors to be considered when determining whether there is sufficient evidence to support a determination that a disclosure does not satisfy the enablement requirement and whether any necessary experimentation required is “undue” include, but are not limited to:
(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 inventor;
(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.
Furthermore, the USPTO does not have laboratory facilities to test if an invention will function as claimed when working examples are not disclosed in the specification. Therefore, enablement issues are raised and discussed based on the state of knowledge pertinent to an art at the time of the invention. And thus, skepticism raised in the enablement rejections are those raised in the art by artisans of expertise.
All of the Wands factors have been considered with regard to the instant claims, with the most relevant factors discussed below.
Breadth of claims 172-182 and 184-189:
The claims are directed to a method of producing an iPSC whereby the method comprises contacting any type of somatic cell with one or more recombinant circular RNA encoding at least one of Oct3/4, Klf4, Sox2, Nanog, Lin28, c-Myc, and L-Myc, or a fragment or variant thereof.
Claims 172-182 and 184-189 are broad in that only one of Oct3/4, Klf4, Sox2, Nanog, Lin28, c-Myc, and L-Myc, or a fragment or variant thereof, is required to be present in the method. Furthermore, claims 172-183 and 185-189 are broad in that the somatic cell may be any somatic cell.
Breadth of claims 172-179 and 182-189:
The claims are directed to a method of producing an induced pluripotent stem cell (iPSC) whereby the method comprises (1) contacting a somatic cell with one or more recombinant circular RNA as recited in the claims and, then, (2) maintaining the cell under conditions to achieve said producing. The claims are broad in that the contacting of the somatic cell with the RNA encompasses any type of contacting without limitation (see e.g., instant [0141]-[0144], [0170], [0199]).
Further in view of claims 180 or 181, all the claims are directed to a method of producing an induced pluripotent stem cell (iPSC) whereby the method comprises contacting a somatic cell with one or more recombinant circular RNA conjugated with an LNP to form a complex wherein the RNA is as recited in the claims (e.g., claim 172). The claims are broad in that the complex may consist of a single lipid conjugated to a single circular RNA so long as the lipid is less than 1 micron in size but not requiring the RNA to be encapsulated by any lipid structure.
The state of the art:
The prior art teaches that at a minimum of Oct4, Sox2, and Klf4 (OSK); or Oct4, Sox2, Nanog, and Lin28 (OSNL); or Sox2, Klf4, and cMyc (SKM) are required to predictably reprogram a non-pluripotent cell into an iPSC, with adding c-Myc to OSKM or valproic acid to the OSK cocktail increasing efficiency (Preskey et al., BBRC 473(3):743-51 (2016), IDS ref; at pg. 746, left col., last para.).
Certain subtypes of somatic cells can predictably be reprogrammed with other combinations of factors: e.g., human fibroblasts with valproic acid along with Oct4 and Sox2 (OS-v), and the combination Oct4, Sox2 and c-Myc along with kenpaullone (OSM-k) can reprogram fibroblasts and mouse neural progenitor cells, at least inefficiently (Lyssiotis et al., PNAS 106: 8912-7 (2009) at pg.8912, left col., las para., to right col.; abstract). While using only Oct4 for mouse or Oct4 and Klf4/c-Myc (OK/OM) for human can reprogram neural progenitor cells, this is not applicable generally to any somatic cell type (Kim et al., Cell 136: 411-9 (2009); Shi et al., Cell Stem Cell 3: 568-74 (2008)). If any of these required factors are removed reprogramming often only occurs if other small molecule inhibitors are included (e.g., RepSox, SB431542, PD0325901, CHIR99021) or in rare situations where the cell being reprogrammed endogenously expresses the missing exogenous factor (e.g., Oct4 or Sox2) (see e.g., Shi et al., 2008; Maherali et al., Curr Biol 19: 1718-23 (2009)). The prior art teaches some cells, e.g., mature red blood cells or neurons, cannot be reprogrammed, and that senescent cells are resistant to reprogramming. Also, Nanog, Lin28, Sall4, and Esrrb can reprogram mouse embryonic fibroblasts (Buganim et al., Cell Stem Cell 15: 295-309 (2014)), but again this is not generally applicable to any somatic cell type, such as endothelial progenitors, keratinocytes, melanocytes, adipose-derived and urine-derived cells.
The prior art teaches a limited set of functional fragments and variants of Oct3/4, Klf4, Sox2, c-Myc, and Nanog but not any functional fragment of Lin28 or L-Myc, as noted above.
The prior art teaches specific RNA-based reprogramming of human somatic cells to iPSCs requires RNA transfection or viral infection and expression of at least the 4 factors OSKM (Preskey et al., at pg. 746, left col., last para.; Warren et al., Cell Stem Cell 7: 18-30 (2010), IDS ref.; Yoshioka et al., Cell Stem Cell 13: 246-54 (2013); Sarkar et al., Nat Commun 11: 1545 (2020)) and reprogramming a senescent fibroblasts required RNA encoding 6 factors OSKMNL (Kogut et al., Nat Commun 9: 745 (2018); IDS ref.). Thus, the prior art teaches RNA generally, or naked circular RNA, is not stable in extracellular environments and is not readily absorbed into cells without more direct interventions, such as chemical disruption of membranes, electroporating, transfecting and/or targeting with protective lipid nanoparticles encapsulating the RNA.
The prior art teaches the expression of reprogramming factors via RNA-based methods can take at least up to 10 days (Annand, R., Methods Mol Biol 2239: 163-74 (2021)), but that sometimes reprogramming is accomplished in as few as 7 days (Yoshioka et al., Cell Stem Cell 13: 246-54 (2013) at pg. 248, left col., 3rd para.).
Thus, these aspects of using must be shown to a reasonable extent so that one of the ordinary skills in the art would be able to practice the invention without any undue or unreasonable burden being on such artisan: (1) using only one of Oct3/4, Klf4, Sox2, Nanog, Lin28, c-Myc, and L-Myc; (2) using only fragments/variants of Nanog, Lin28, and/or L-Myc; (3) reprogramming any somatic cell type with said combination of factors; (4) using mere contacting of the somatic cell with the circular RNA is sufficient to transport sufficient RNA inside the cell to result in expression of the factors required; and (5) mere complexes of nanolipid and circular RNA can deliver and express the RNA in the cell.
The amount of direction and guidance as well as any working examples provided:
The instant specification notes that the term “contacting” may encompass transfecting a circular RNA (circRNA), or a vector comprising a nucleic acid (e.g., a DNA molecule) encoding the same, into the cell, such as using lipid-mediated transfection, Lipofectamine®, and/or other transfection reagents ([0141]). The instant application notes that the contacting can be performed multiple times, which is also known in the art. The instant specification notes prophetically that circRNA-LNP complexes can be introduced into cells by merely contacting with the cells in the absence of any transfection reagent ([0294]).
The instant specification provides working examples using transfecting of circRNA or circRNA-LNP complexes according to a ThermoFisher® Lipofectamine® RNAiMax commercial product’s instructions, Neon® nucleofection electroporation system, or DOTAP liposomal transfection technique (Examples 3-9). Furthermore, all the working examples rely on using the 6 factors in combination (OSKLMN), use repeating daily transfections for at least 4-6 days, and are limited to human fibroblasts.
Nowhere does the instant specification show how to reprogram a somatic cell into an iPSC by a single contacting with a single recombinant circular RNA molecule. Instead, 25,000-75,000 cells were transfected with 30 ng circular RNA according to Lipofectamine® manufacturer’s instructions. This is equivalent to about 0.5-1 pg per cell, or tens of thousands of RNA molecules per cell per transfection. Therefore, in all the working examples at least a subset of the cells are contacted thousands of times by thousands of RNA molecules (and LNP complexes thereof) in a single transfection event. Similarly, at least a subset of the cells are contacted at least dozens or hundreds of times by copies of the same circular RNA.
Thus, there is no evidence in the instant application or the prior art that the scope of contacting encompassed by the claims could predictably result in a reprogrammed iPSC by performing the methods as recited in the claims.
Undue experimentation would be required to fill these gaps and unpredictability. Undue experimentation is required to reprogram any type of somatic cell using only one of Oct3/4, Klf4, Sox2, Nanog, Lin28, c-Myc, and L-Myc, or a fragment or variant thereof. Undue experimentation is required to reprogram a somatic cell using only a single contacting with a single circular RNA or whereby the RNA does not enter the cell’s interior but merely contacts an outer surface.
In summary, the claims are rejected under 35 U.S.C. 112(a) because the specification does not reasonably provide enablement to a person skilled in the art to which it pertains or with which it is most nearly connected to produce an iPSC via merely contacting any somatic cell with the circular RNA(s) as recited in the claims. Given the lack of working examples, the limited guidance provided in the specification, the lack of guidance in the prior art, and the broad scope of the claims, undue and/or unreasonable experimentation would have been required for one skilled in the art to produce the recited product over the full scope of the methods of claims 172-189.
Claim Rejections - 35 USC § 112(b)
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 172-189 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 claims are interpreted as set forth in a previous section.
Claim 172 recites the relative term “variant” regarding a reprogramming factor. A person of ordinary skill in the art reading the claim would not understand the metes and bounds of “variant” because neither the claim nor the instant application provides any definition or standard for the term “variant” nor any anchoring/canonical/consensus sequences for any of Oct3/4, Klf4, Sox2, Nanog, Lin28, c-Myc, and L-Myc. See MPEP 2173.05(b). Although the prior art often uses variant to mean a sequence variation from a reference sequence, this is not a definite standard without a sequence similarity limitation. Furthermore, the prior art less frequently uses variant to also encompass modified derivates having identical sequences, such as side chain modified, dimerized, and/or fusions with additional polypeptides/proteins. As used in the instant case, the term variant has no boundary and, thus, ambiguously might encompasses any factor comprising 25% or lower sequence homology as identifiable by a software analysis known in the prior art.
In claims 172, 174 and 175, the term “Oct3/4” is not clear as to whether what is before and after the slash forms a single limitation, recites an optional feature or alternative limitations. Claims 173 and 176-189 are included in this rejection for depending from an indefinite claim.
In claim 176, the term “substantially” regarding non-immunogenic is a relative term which renders the claim indefinite because neither the claim nor the specification provides a standard for ascertaining a requisite minimum degree of substantiality and, thus, one of ordinary skill in the art would not be reasonably appraised of the scope of this claim limitation. Although the specification discusses examples of substantially non-immunogenic include not inducing the expression or activity of one or more interferon-regulated genes, such as IFN-alpha, IFN-beta and TNF-alpha, which may be a result of an RNA modification(s) ([0124], [0080]), the instant application is silent as to a definition of “substantially.”
In claim 178, the term “about” is a relative term which renders the claim indefinite because neither the claim nor the specification provides a standard or requisite minimum range for “about” and, thus, one of ordinary skill in the art would not be reasonably appraised of the scope of this term. Although the specification discusses examples of “about” as encompassing ±20%, ±10%, ±5%, ±1%, ±0.5%, or ±0.1% ([0068]), the instant application lacks a clear range for “about”, such as regarding the number of nucleotides in a circular RNA molecule.
In claims 180-181, the term lipid nanoparticle (LNP) is used to mean any lipid-based particle in the submicron size range, including non-bilayer structures ([0083]). However, the ordinary meaning of “lipid nanoparticle” means a spherical, ellipsoid, or rod-shaped micelle or bicelle-like structure due to the assembly of amphipathic lipids in a polar solvent. If the applicant acts as her own lexicographer to specifically define a term of a claim contrary to its ordinary meaning, the written description must clearly redefine the claim term and set forth the uncommon definition so as to put one reasonably skilled in the art on notice that the applicant intended to so redefine that claim term. Process Control Corp. v. HydReclaim Corp., 190 F.3d 1350, 1357, 52 USPQ2d 1029, 1033 (Fed. Cir. 1999). In the instant case, there is no such redefinition, and, thus, the term “lipid nanoparticle (LNP)” as used in the claims is indefinite.
In claim 189, the terms “end” of culture, “culture,” “rate” of maturation, and “mesenchymal-to-epithelial transition (MET)” each lacks proper antecedent basis within in the claim or in claim 172.
Claim Rejections - 35 USC § 112(d)
The following is a quotation of 35 U.S.C. 112(d):
(d) REFERENCE IN DEPENDENT FORMS.—Subject to subsection (e), a claim in dependent form shall contain a reference to a claim previously set forth and then specify a further limitation of the subject matter claimed. A claim in dependent form shall be construed to incorporate by reference all the limitations of the claim to which it refers.
The following is a quotation of pre-AIA 35 U.S.C. 112, fourth paragraph:
Subject to the following paragraph [i.e., the fifth paragraph of pre-AIA 35 U.S.C. 112], a claim in dependent form shall contain a reference to a claim previously set forth and then specify a further limitation of the subject matter claimed. A claim in dependent form shall be construed to incorporate by reference all the limitations of the claim to which it refers.
Claims 178, 186 and 189 are rejected under 35 U.S.C. 112(d) or pre-AIA 35 U.S.C. 112, 4th paragraph, as being of improper dependent form for failing to further limit the subject matter of the claim upon which it depends, or for failing to include all the limitations of the claim upon which it depends.
Claim 178 limits the circular RNA to comprising about 200 to about 5,000 nucleotides but as the transitional term “comprises” is open-ended (see MPEP 2111.03), this phrase only places a minimum size of the circular RNA at 200 nucleotides but as the RNA must inherently be at least 600 nucleotides long just to comprise a protein coding sequence encoding at least one of Oct3/Oct4, Sox2, Klf4, C-Myc, L-Myc, Lin28, and Nanog; claim 178 fails to further limit the subject matter of claim 172.
Claim 186 limits the somatic cell to being adherent or in suspension meaning claim 172 encompasses a situation wherein the cell is neither adherent nor in suspension. Absent evidence to the contrary, claim 186 fails to further limit the subject matter of claim 172 because there is no other coherent and unambiguous status for the cell beyond adherent or in suspension.
Claim 189 further recites only language regarding a result(s) of the claimed method without any further active step meaning the method as recited in claim 172 inherently produces these results absence evidence to the contrary. Thus, this dependent claims fails to further limit the subject matter of claim 172.
Applicant may cancel the claim(s), amend the claim(s) to place the claim(s) in proper dependent form, rewrite the claim(s) in independent form, or present a sufficient showing that the dependent claim(s) complies with the statutory requirements.
Claim Rejections - 35 USC § 102
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 the appropriate paragraphs of 35 U.S.C. 102 that form the basis for the rejections under this section made in this Office action:
A person shall be entitled to a patent unless –
(a)(1) the claimed invention was patented, described in a printed publication, or in public use, on sale, or otherwise available to the public before the effective filing date of the claimed invention.
(a)(2) the claimed invention was described in a patent issued under section 151, or in an application for patent published or deemed published under section 122(b), in which the patent or application, as the case may be, names another inventor and was effectively filed before the effective filing date of the claimed invention.
Claims 172-174, 176, 178, 184, 186, and 189 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Stadtfeld (US 20160326497 A1).
Regarding claim 172, Stadtfeld discloses a method comprising contacting a somatic cell (introducing into) with an exogenous circular RNA(s) encoding the exogenous reprogramming factors Oct4, Klf4, Sox2, and myc (e.g., c-Myc) (as well as Nanog and/or Lin28), and culturing the cell in a medium for an amount of time sufficient to generate iPSCs, e.g., 3-8 days or about 9 days ([0008]-[0011], [0050], [0087]).
Regarding claim 173, Stadtfeld discloses wherein the first four reprogramming factors are human ones ([0011]).
Regarding claim 174, Stadtfeld discloses wherein the Oct4 has a sequence (SEQ ID NO: 2) identical to instant SEQ ID NO: 1, as show below.
Query Match 100.0%; Score 1948; Length 361;
Matches 360; Conservative 0; Mismatches 0; Indels 0; Gaps 0;
Qy 1 MAGHLASDFAFSPPPGGGGDGPGGPEPGWVDPRTWLSFQGPPGGPGIGPGVGPGSEVWGI 60
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1 MAGHLASDFAFSPPPGGGGDGPGGPEPGWVDPRTWLSFQGPPGGPGIGPGVGPGSEVWGI 60
Qy 61 PPCPPPYEFCGGMAYCGPQVGVGLVPQGGLETSQPEGEAGVGVESNSDGASPEPCTVTPG 120
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 61 PPCPPPYEFCGGMAYCGPQVGVGLVPQGGLETSQPEGEAGVGVESNSDGASPEPCTVTPG 120
Qy 121 AVKLEKEKLEQNPEESQDIKALQKELEQFAKLLKQKRITLGYTQADVGLTLGVLFGKVFS 180
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 121 AVKLEKEKLEQNPEESQDIKALQKELEQFAKLLKQKRITLGYTQADVGLTLGVLFGKVFS 180
Qy 181 QTTICRFEALQLSFKNMCKLRPLLQKWVEEADNNENLQEICKAETLVQARKRKRTSIENR 240
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 181 QTTICRFEALQLSFKNMCKLRPLLQKWVEEADNNENLQEICKAETLVQARKRKRTSIENR 240
Qy 241 VRGNLENLFLQCPKPTLQQISHIAQQLGLEKDVVRVWFCNRRQKGKRSSSDYAQREDFEA 300
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 241 VRGNLENLFLQCPKPTLQQISHIAQQLGLEKDVVRVWFCNRRQKGKRSSSDYAQREDFEA 300
Qy 301 AGSPFSGGPVSFPLAPGPHFGTPGYGSPHFTALYSSVPFPEGEAFPPVSVTTLGSPMHSN 360
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 301 AGSPFSGGPVSFPLAPGPHFGTPGYGSPHFTALYSSVPFPEGEAFPPVSVTTLGSPMHSN 360
Regarding claim 176, Stadtfeld anticipates wherein the circular RNA is substantially non-immunogenic by virtue of anticipating claim 172 as set forth fully above because there are no structural feature(s) recited or implied by the language of claim 176, and because circular RNAs are inherently less immunogenic than linear RNAs with regard to inducing the expression and/or activity of an interferon-regulated gene(s).
Regarding claim 178, as shown above Stadtfeld discloses wherein the RNA encodes Oct4 having the sequence according to instant SEQ ID NO: 1, which is inherently larger than 200 nucleotides and further must be over 1080 nucleotides to provide expression of the Oct4.
Regarding claim 184, Stadtfeld discloses wherein the cell is a fibroblast, a peripheral blood-derived cell (mature T or B cell), a cord-blood derived cell (blood stem cell or blood progenitor cell, immature blood cell, T cell or B cell), or a keratinocyte ([0013], [0018]).
Regarding claim 186, Stadtfeld discloses wherein the cell (MEF) is adherent or in suspension (Example I).
Regarding claim 189, Stadtfeld discloses the same active method steps as required by claim 189 as set forth fully above regarding claim 172 because there is no additional step or feature implied in claim 189.
Thus, Stadtfeld anticipates the claimed invention.
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.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
Claims 172-174, 176, 178, 182, 184, 186, and 188-189 are rejected under 35 U.S.C. 103 as being unpatentable over Stadtfeld (US 20160326497 A1) in view of Warren (Warren et al., Cell Stem Cell 7: 18-30 (2010), IDS ref.).
The claims are interpreted as set forth in a previous section. As set forth above, Stadtfeld anticipates claims 172-174, 176, 178, 184, 186, and 189 and, thus, Stadtfeld renders obvious the subject matter of claims 172-174, 176, 178, 184, 186, and 189.
Regarding claim 182, Stadtfeld does not expressly teach wherein the cell is contacted with at least 4 circular RNAs encoding the reprogramming factor(s).
However Warren teaches RNA-based methods of iPSC generation comprising daily consecutive transfections of different RNAs together encoding four different reprogramming factors, one RNA type per reprogramming factor (Fig. 2; pg. 621, left col., last para., to pg. 624, left col., 1st para.).
Therefore, it would have been prima facie obvious to one of ordinary skill in the art before the effective time of filing to perform the method taught by Stadtfeld using a different circular RNA per exogenous reprogramming factor (i.e., Oct4, Klf4, Sox2, and C-Myc) such that the cell is transfected with 4 circular RNAs in view of Warren. One of ordinary skill in the art would be motivated by Warren teaching RNA-based methods of iPSC generation comprising transfections of 4 different RNAs, one per reprogramming factor. Note, an overlapping range between the prior art and a claim provides a prima facie case of obviousness (see MPEP 2144.05).
Regarding claim 188, as set forth above for claim 182, Warren teaches wherein a combination of different RNAs (one per reprogramming factor) are repeatedly transfected at least 4 times (pg. 623, left col., 1st para.). Note, an overlapping range between the prior art and a claim provides a prima facie case of obviousness (see MPEP 2144.05).
Thus, the claimed invention as a whole is prima facie obvious prior to the earliest effective filing date in the absence of evidence to the contrary.
Claims 172-174, 176-179, 184, 186, and 188-189 are rejected under 35 U.S.C. 103 as being unpatentable over Stadtfeld as applied above, and further in view of Yang (Yang et al., Cell Res 27: 626-41 (2017)).
Regarding claim 177, Stadtfeld does not teach wherein the circular RNA comprises one or more M-6-methyladenosine (m6A) residues.
However Yang teaches incorporating m6A residues promotes translation of circular RNAs (Abstract; Fig. 1-2).
It would have been prima facie obvious to one of ordinary skill in the art before the effective time of filing to make the circular RNA used in the method taught by Stadtfeld such that it incorporates m6A nucleotides. One of ordinary skill in the art with the goal of achieving strong expression of the reprogramming factors in a human cell would be motivated by Yang teaching improved expression for circular RNAs comprising m6A nucleotides.
Regarding claim 179, Stadtfeld does not teach wherein the circular RNA comprises an IRES operably linked to the coding sequence.
However Yang teaches using an IRES in a circular RNA to provide protein expression (Fig. 1; pg. 626, right col.) and that m6A nucleotides function as an IRES (pg. 627, left col., last para., Fig. 1).
It would have been prima facie obvious to one of ordinary skill in the art before the effective time of filing to make the circular RNA used in the method taught by Stadtfeld to have an IRES in front of the reprogramming factor coding sequence(s). One of ordinary skill in the art with the goal of achieving strong expression of the reprogramming factors would be motivated by Yang teaching constructs engineered with various IRES sequences all successful drove protein expression from circular formats. Thus, the claimed invention as a whole is prima facie obvious prior to the earliest effective filing date in the absence of evidence to the contrary.
Claims 172-174, 176, 178, 180-181, 184, 186, and 188-189 are rejected under 35 U.S.C. 103 as being unpatentable over Stadtfeld as applied above, and further in view of Wesselhoeft (Wesselhoeft et al., Molecular Cell 74(3): 508-20 (2019); IDS ref.).
Regarding claims 180-181, Stadtfeld does not teach wherein the circular RNA is complexed with a lipid nanoparticle (LNP), either covalently or non-covalently.
However Wesselhoeft teaches wherein expression vectors consisting of circular RNAs are encapsulated into lipid nanoparticles for deliver to target cells (pg. 515, left col., last para., to pg. 516, left col., 2nd para.)
It would have been prima facie obvious to one of ordinary skill in the art before the effective time of filing to contact the circular RNA in the method taught by Stadtfeld using an LNP delivery system taught by Wesselhoeft that non-covalently complexes circular RNA to LNP. One of ordinary skill in the art would be motivated by Wesselhoeft teaching LNP delivery was equivalent to electroporation but provides an advantage for being less immunogenic than linear RNAs (unrecognized by cellular RNA sensors), such as for in vivo use or with TLR-competent somatic cells (Abstract; pg. 515, left col., last para., to pg. 516, left col., 2nd para.; Fig. 4-6).
Claims 172-174, 176, 178, 182-184, 186, and 188-189 are rejected under 35 U.S.C. 103 as being unpatentable over Stadtfeld and Warren as applied above, and further in view of Wang2 (Wang et al., Biol Open 8: bio047225 (2019)).
Regarding claim 183, although Stadtfeld and Warren does not expressly teach wherein the cell is contacted with exactly 5 or 6 circular RNAs encoding the recited reprogramming factors; Stadtfeld teaches further including exogenous nucleic acids encoding Nanog and/or Lin28 ([0008]-[0011]) and Wang2 teaches the combination of Oct4, Klf4, Sox2, c-Myc and Lin28 or the combination of Oct4, Klf4, Sox2, c-Myc, Lin28 and Nanog more efficiently reprograms iPSCs (e.g., 76-fold) than using the four factors OSKM alone (Abstract; Fig. 1; pg. 2 left col., 3rd para., to right col., 1st para.; Fig. 4B; pg. 8, left col., last para, to right col.). Thus, it would have been prima facie obvious to one of ordinary skill in the art before the effective time of filing to perform the method taught by Stadtfeld and Wang2 using 6 or 7 circular RNAs, with one per factor respectively encoding Oct4, Klf4, Sox2, C-Myc and Lin28, or Oct4, Klf4, Sox2, C-Myc, Lin28 and Nanog. One of ordinary skill in the art would be motivated by Wang2 teaching the beneficial synergistic effect on increasing reprogramming efficiency.
Claims 172-174, 176, 178, 184-186, and 188-189 are rejected under 35 U.S.C. 103 as being unpatentable over Stadtfeld as applied above, and further in view of Ye (Ye et al., Blood 114: 5473-80 (2009)).
Regarding claim 185, Stadtfeld does not teach wherein the somatic cell is CD34+.
However Ye teaches reprogramming adult CD34+ cells from patient samples into iPSCs using expression of Oct4, Sox2, Klf4, and c-Myc from viral vectors in order to study hematopoiesis and cancer (abstract; pg. 5474, left col., 3rd para.). It would have been prima facie obvious to one of ordinary skill in the art before the effective time of filing to perform the method taught by Stadtfeld on a CD34+ somatic cell. One of ordinary skill in the art with the goal of studying cancer would be motivated to avoid using viral vectors in their study to avoid confounding transformation events and, thus, would find non-viral alternatives like RNA transfection as taught by Stadtfeld ([0009]; [0067]; [0111]-[0113]), e.g., RNA electroporation, lipofection, PEG fusion, etc. Thus, the claimed invention as a whole is prima facie obvious prior to the earliest effective filing date in the absence of evidence to the contrary.
Claims 172-174, 176, 178, 184, and 186-189 are rejected under 35 U.S.C. 103 as being unpatentable over Stadtfeld in view of Ye as applied above, and further in view of Mali (Mali et al., Stem Cells 26: 1998-2005 (2008)).
Regarding claim 187, Stadtfeld does not teach wherein the somatic cell is a CD34+ cell in suspension. However Mali, as used by Xu (pg. 5474, left col., 2nd para.), teaches culturing CD34+ cells in suspension and during reprogramming (pg. 1999, left col., last para., to right col. 2nd para.). Thus, it would have been prima facie obvious to one of ordinary skill in the art before the effective time of filing to perform the method taught by Stadtfeld and Ye on a CD34+ somatic cell in suspension.
Claims 172-173, 176, 178-179, 182, 184, 186, and 188-189 are rejected under 35 U.S.C. 103 as being unpatentable over Yoshioka (Yoshioka et al., Cell Stem Cell 13: 246-54 (2013)) in view of Wesselhoeft2 (Wesselhoeft et al., Nat Commun 9: 2629 (2018); IDS ref.).
Yoshioka teaches methods of making iPSCs by contacting (introducing into via transfection) a somatic cell (human fibroblast) with an exogenous recombinant RNA encoding the exogenous protein reprogramming factors Oct4, Klf4, Sox2, and c-Myc (OSKM) along with B18R mRNA (Fig. 1A, VEE-OMKS or VEE-OKS-iM) and culturing the cell in media (Advanced DMEM or ES medium) for an amount of time sufficient to generate iPSCs, e.g., at least 7-10 days (pg. 248, left col., 3rd para., to pg. 249, left col., last para.; Fig. 1-2; Table 1; pg. 252, right col., 3rd para.). Yoshioka teaches the benefits of expressing all four reprogramming factors at consistent levels and ratios over time in the same somatic cell on increasing the efficiency of reprogramming (Abstract).
Regarding claim 172, Yoshioka does not teach wherein the RNA is circular, instead relying on two different linear RNAs (pg. 252, left col., last para.).
However Wesselhoeft2 teaches increased translation for circular RNAs compared to their linear parental forms and how to circularize linear RNAs generally (Abstract; Fig. 6; pg. 4, left col., 2nd para.; pg. 6, left col., 1st para. to pg. 7; Fig. 1 and 3).
It would have been prima facie obvious to one of ordinary skill in the art before the effective time of filing to modify the method taught by Yoshioka to use a circular RNA(s) instead of linear ones to express OSKM in a somatic cell and reprogram it into an iPSC. One of ordinary skill in the art with the goal of ensuring robust reprogramming factor expression over long durations as taught by Yoshioka (pg. 248, left col., 3rd para.) would be motivated by Wesselhoeft2 teaching this a general effect for circularized RNA (circRNA) expression vectors (pg. 7, left col.; abstract).
Regarding claim 173, Yoshioka teaches wherein each of the OSKM are human ones (Supplemental Experimental Procedures, pg. 12).
Regarding claim 176, the circular RNAs taught by the combination of Yoshioka and Wesselhoeft2 are substantially non-immunogenic because there are no structural feature(s) recited or implied by the language of claim 176 that is not in claim 172, and because circular RNAs are inherently less immunogenic than linear RNAs with regard to inducing the expression and/or activity of an interferon-regulated gene(s).
Regarding claim 178, Wesselhoeft2 teaches how to circularize linear RNAs as large as 5 kb (Abstract), which accommodates at most up to 3 of the 4 factors in a single circular RNA. Furthermore by definition, each circular RNA taught by the combination of Yoshioka and Wesselhoeft2 must comprise over 300 nucleotides just to encode one of human Oct4, Klf4, Sox2, or c-Myc.
Regarding claim 179, Yoshioka teaches wherein the RNA construct comprises an IRES operably linked to the coding sequence for Sox2 and/or c-Myc (Fig. 1A).
Regarding claim 182, Yoshioka teaches wherein the contacting must be with at least two different circular RNAs due Yoshioka teaching using all four factors be used and Wesselhoeft2 teaching a maximum size (5 kb) (abstract) that cannot accommodate all 4 human versions of Oct4, Klf4, Sox2, and c-Myc in a single circular RNA.
Regarding claims 184 and 186, Yoshioka teaches wherein the somatic cell is an adherent human fibroblast (HFF or BJ) (Fig. 2).
Regarding claim 188, Yoshioka teaches repeating transfections to the same cell population two times or more than four times (pg. 248, right col., 3rd para.; Fig. 2B-C). Note, an overlapping range between the prior art and a claim provides a prima facie case of obviousness (see MPEP 2144.05).
Regarding claim 189, the combination of Yoshioka and Wesselhoeft2 teach the same active method steps as required by claim 189 in teaching claim 172 as set forth above because there is no additional step or feature implied in claim 189 and performing the method of claim 172 inherently produces these results.
Claims 172-174, 176, 178-179, 182, 184, 186, and 188-189 are rejected under 35 U.S.C. 103 as being unpatentable over in view of Wesselhoeft2 as applied above, and further as evidenced by NM002701 (NCBI locus NM_002701, updated NOV 2018).
Regarding claim 174, Yoshioka (Supplemental Experimental Procedures, Plasmids, pg. 12) as evidenced by NM002701 teaches wherein the Oct4 has a sequence (translation of NM002701) identical to instant SEQ ID NO: 1, as show below.
100.0% identity in 360 residues overlap; Score: 1948.0; Gap frequency: 0.0%
Sequence1 1 MAGHLASDFAFSPPPGGGGDGPGGPEPGWVDPRTWLSFQGPPGGPGIGPGVGPGSEVWGI
Trans002701 1 MAGHLASDFAFSPPPGGGGDGPGGPEPGWVDPRTWLSFQGPPGGPGIGPGVGPGSEVWGI
************************************************************
Sequence1 61 PPCPPPYEFCGGMAYCGPQVGVGLVPQGGLETSQPEGEAGVGVESNSDGASPEPCTVTPG
Trans002701 61 PPCPPPYEFCGGMAYCGPQVGVGLVPQGGLETSQPEGEAGVGVESNSDGASPEPCTVTPG
************************************************************
Sequence1 121 AVKLEKEKLEQNPEESQDIKALQKELEQFAKLLKQKRITLGYTQADVGLTLGVLFGKVFS
Trans002701 121 AVKLEKEKLEQNPEESQDIKALQKELEQFAKLLKQKRITLGYTQADVGLTLGVLFGKVFS
************************************************************
Sequence1 181 QTTICRFEALQLSFKNMCKLRPLLQKWVEEADNNENLQEICKAETLVQARKRKRTSIENR
Trans002701 181 QTTICRFEALQLSFKNMCKLRPLLQKWVEEADNNENLQEICKAETLVQARKRKRTSIENR
************************************************************
Sequence1 241 VRGNLENLFLQCPKPTLQQISHIAQQLGLEKDVVRVWFCNRRQKGKRSSSDYAQREDFEA
Trans002701 241 VRGNLENLFLQCPKPTLQQISHIAQQLGLEKDVVRVWFCNRRQKGKRSSSDYAQREDFEA
************************************************************
Sequence1 301 AGSPFSGGPVSFPLAPGPHFGTPGYGSPHFTALYSSVPFPEGEAFPPVSVTTLGSPMHSN
Trans002701 301 AGSPFSGGPVSFPLAPGPHFGTPGYGSPHFTALYSSVPFPEGEAFPPVSVTTLGSPMHSN
************************************************************
Thus, the claimed invention as a whole is prima facie obvious prior to the earliest effective filing date in the absence of evidence to the contrary.
Claims 172-174, 176-179, 184, 186, and 188-189 are rejected under 35 U.S.C. 103 as being unpatentable over Stadtfeld as applied above, and further in view of Yang (Yang et al., Cell Res 27: 626-41 (2017)).
Regarding claim 177, Stadtfeld does not teach wherein the circular RNA comprises one or more M-6-methyladenosine (m6A) residues.
However Yang teaches incorporating m6A residues promotes translation of circular RNAs (Abstract; Fig. 1-2).
It would have been prima facie obvious to one of ordinary skill in the art before the effective time of filing to make the circular RNA used in the method taught by Stadtfeld such that it incorporates m6A nucleotides. One of ordinary skill in the art with the goal of achieving strong expression of the reprogramming factors in a human cell would be motivated by Yang teaching improved expression for circular RNAs comprising m6A nucleotides.
Regarding claim 179, Stadtfeld does not teach wherein the circular RNA comprises an IRES operably linked to the coding sequence.
However Yang teaches using an IRES in a circular RNA to provide protein expression (Fig. 1; pg. 626, right col.) and that m6A nucleotides function as an IRES (pg. 627, left col., last para., Fig. 1).
It would have been prima facie obvious to one of ordinary skill in the art before the effective time of filing to make the circular RNA used in the method taught by Stadtfeld to have an IRES in front of the reprogramming factor coding sequence(s). One of ordinary skill in the art with the goal of achieving strong expression of the reprogramming factors would be motivated by Yang teaching constructs engineered with various IRES sequences all successful drove protein expression from circular formats. Thus, the claimed invention as a whole is prima facie obvious prior to the earliest effective filing date in the absence of evidence to the contrary.
Claims 172-173, 176, 178-179, 180-182, 184, 186, and 188-189 are rejected under 35 U.S.C. 103 as being unpatentable over Yoshioka in view of Wesselhoeft2 as applied above, and further in view of Wesselhoeft (Wesselhoeft et al., Molecular Cell 74(3): 508-20 (2019); IDS ref.).
Regarding claims 180-181, Yoshioka teaches transfecting RNA encoding reprogramming factors into somatic cells using Lipofectamine® 2000 (Invitrogen®) (pg. 252, left col., para. 2-3), and Wesselhoeft2 teaches efficiently introducing recombinant circular RNAs into somatic cells in vitro using cationic lipid transfection reagent (pg. 8, left col., last para, to right col.); however, the combination of Yoshioka and Wesselhoeft2 does not expressly teach wherein the circular RNA is complexed with a lipid nanoparticle (LNP), either covalently or non-covalently.
However Wesselhoeft teaches wherein expression vectors consisting of circular RNAs are encapsulated into lipid nanoparticles for deliver to target cells (pg. 515, left col., last para., to pg. 516, left col., 2nd para.)
It would have been prima facie obvious to one of ordinary skill in the art before the effective time of filing to transfect the circular RNAs in the method taught by Yoshioka and Wesselhoeft2 using an LNP delivery system taught by Wesselhoeft that non-covalently complexes the circular RNAs to LNPs. One of ordinary skill in the art would be motivated by Wesselhoeft teaching LNP delivery was equivalent to electroporation but provides an advantage for being less immunogenic than linear RNAs (unrecognized by cellular RNA sensors), such as for in vivo use or with TLR-competent somatic cells (Abstract; pg. 515, left col., last para., to pg. 516, left col., 2nd para.; Fig. 4-6).
Claims 172-173, 176, 178-179, 182-184, 186, and 188-189 are rejected under 35 U.S.C. 103 as being unpatentable over Yoshioka in view of Wesselhoeft2 as applied above, and further in view of Wang2 (Wang et al., Biol Open 8: bio047225 (2019)).
Regarding claim 183, the combination of Yoshioka and Wesselhoeft2 does not teach wherein the cell is contacted with exactly 5 or 6 circular RNAs encoding the recited reprogramming factors.
However Wang2 teaches the combination of Oct4, Klf4, Sox2, c-Myc and Lin28 or the combination of Oct4, Klf4, Sox2, c-Myc, Lin28 and Nanog more efficiently reprograms iPSCs (e.g., 76-fold) than using the four factors OSKM alone (Abstract; Fig. 1; pg. 2 left col., 3rd para., to right col., 1st para.; Fig. 4B; pg. 8, left col., last para, to right col.). Thus, it would have been prima facie obvious to one of ordinary skill in the art before the effective time of filing to perform the method taught by Yoshioka and Wesselhoeft2 using 6 or 7 circular RNAs, with one per factor respectively encoding Oct4, Klf4, Sox2, C-Myc and Lin28, or Oct4, Klf4, Sox2, C-Myc, Lin28 and Nanog. One of ordinary skill in the art would be motivated by Wang2 teaching the beneficial synergistic effect on increasing reprogramming efficiency by including Lin28 or Lin28+Nanog with the foundational OSKM.
Claims 172-173, 176, 178-179, 182, 184-186, and 188-189 are rejected under 35 U.S.C. 103 as being unpatentable over Yoshioka in view of Wesselhoeft2 as applied above, and further in view of Ye (Ye et al., Blood 114: 5473-80 (2009)).
Regarding claim 185, Stadtfeld does not teach wherein the somatic cell is CD34+.
However Ye teaches reprogramming adult CD34+ cells from patient samples into iPSCs using expression of Oct4, Sox2, Klf4, and c-Myc from viral vectors in order to study hematopoiesis and cancer (abstract; pg. 5474, left col., 3rd para.). It would have been prima facie obvious to one of ordinary skill in the art before the effective time of filing to perform the method taught by the combination of Yoshioka and Wesselhoeft2 on a CD34+ somatic cell. One of ordinary skill in the art with the goal of studying cancer would be motivated to avoid using viral vectors in their study to avoid confounding transformation events and, thus, would find non-viral alternatives like RNA transfection as taught by Yoshioka and Wesselhoeft2.
Claims 172-173, 176, 178-179, 182, 184, and 186-189 are rejected under 35 U.S.C. 103 as being unpatentable over Yoshioka in view of Wesselhoeft2 and Ye as applied above, and further in view of Mali (Mali et al., Stem Cells 26: 1998-2005 (2008)).
Regarding claim 187, the combination of Yoshioka, Wesselhoeft2, and Ye does not teach wherein the somatic cell is a CD34+ cell in suspension. However Mali, as used by Xu (pg. 5474, left col., 2nd para.), teaches culturing CD34+ cells in suspension and during reprogramming (pg. 1999, left col., last para., to right col. 2nd para.). Thus, it would have been prima facie obvious to one of ordinary skill in the art before the effective time of filing to perform the method taught by Stadtfeld and Ye on a CD34+ somatic cell in suspension.
Double Patenting
The nonstatutory double patenting rejection is based on a judicially created doctrine grounded in public policy (a policy reflected in the statute) so as to prevent the unjustified or improper timewise extension of the “right to exclude” granted by a patent and to prevent possible harassment by multiple assignees. A nonstatutory double patenting rejection is appropriate where the conflicting claims are not identical, but at least one examined application claim is not patentably distinct from the reference claim(s) because the examined application claim is either anticipated by, or would have been obvious over, the reference claim(s). See, e.g., In re Berg, 140 F.3d 1428, 46 USPQ2d 1226 (Fed. Cir. 1998); In re Goodman, 11 F.3d 1046, 29 USPQ2d 2010 (Fed. Cir. 1993); In re Longi, 759 F.2d 887, 225 USPQ 645 (Fed. Cir. 1985); In re Van Ornum, 686 F.2d 937, 214 USPQ 761 (CCPA 1982); In re Vogel, 422 F.2d 438, 164 USPQ 619 (CCPA 1970); In re Thorington, 418 F.2d 528, 163 USPQ 644 (CCPA 1969).
A timely filed terminal disclaimer in compliance with 37 CFR 1.321(c) or 1.321(d) may be used to overcome an actual or provisional rejection based on nonstatutory double patenting provided the reference application or patent either is shown to be commonly owned with the examined application, or claims an invention made as a result of activities undertaken within the scope of a joint research agreement. See MPEP § 717.02 for applications subject to examination under the first inventor to file provisions of the AIA as explained in MPEP § 2159. See MPEP § 2146 et seq. for applications not subject to examination under the first inventor to file provisions of the AIA . A terminal disclaimer must be signed in compliance with 37 CFR 1.321(b).
The USPTO Internet website contains terminal disclaimer forms which may be used. Please visit www.uspto.gov/patent/patents-forms. The filing date of the application in which the form is filed determines what form (e.g., PTO/SB/25, PTO/SB/26, PTO/AIA /25, or PTO/AIA /26) should be used. A web-based eTerminal Disclaimer may be filled out completely online using web-screens. An eTerminal Disclaimer that meets all requirements is auto-processed and approved immediately upon submission. For more information about eTerminal Disclaimers, refer to www.uspto.gov/patents/process/file/efs/guidance/eTD-info-I.jsp.
Claims 172-179 and 182-189 are provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claims 1, 4-5, 25, 38, 50, 52-53, 55, 57, 77, 80, 93, 97, 103, 110-111, 116, 137, 142, 144-145, and 150 of copending Application No. 18/745761 (reference application).
Although the claims at issue are not identical, they are not patentably distinct from each other because claims 1 and 4 of the reference application teaches a method of producing an induced pluripotent stem cell (iPSC) from a blood cell and/or CD34+ cell in suspension whereby the method comprises contacting the cell with one or more circular RNAs encoding at least one of Oct3/4, Klf4, Sox2, Nanog, Lin28, c-Myc, and L-Myc; and maintaining the cell under conditions under which a reprogrammed iPSC is obtained. Furthermore, reference claims 52-53, 142, 144-145, and 150 requires performing such a method to obtain an iPSC, and reference claims 110-111, 116, and 137 disclose an intermediate product of such a method.
Regarding instant claim 173 and 176-179, reference claim 5 teaches wherein the factor is a human or humanized factor (a), is non-immunogenic (p), comprises one or more M-6-methyladenosine residues (q), comprises from about 200-5000 nucleotides (r), or comprises an IRES operably linked to the protein coding sequence (s).
Regarding instant claims 174-175, reference claims 5 and 57 each teaches the method wherein the Oct3/4 has the amino acid sequence (SEQ ID NO: 7) of instant SEQ ID NO: 1 as shown below.
100.0% identity in 360 residues overlap; Score: 1948.0; Gap frequency: 0.0%
SEQ ID 1 1 MAGHLASDFAFSPPPGGGGDGPGGPEPGWVDPRTWLSFQGPPGGPGIGPGVGPGSEVWGI
Sequence7 1 MAGHLASDFAFSPPPGGGGDGPGGPEPGWVDPRTWLSFQGPPGGPGIGPGVGPGSEVWGI
************************************************************
SEQ ID 1 61 PPCPPPYEFCGGMAYCGPQVGVGLVPQGGLETSQPEGEAGVGVESNSDGASPEPCTVTPG
Sequence7 61 PPCPPPYEFCGGMAYCGPQVGVGLVPQGGLETSQPEGEAGVGVESNSDGASPEPCTVTPG
************************************************************
SEQ ID 1 121 AVKLEKEKLEQNPEESQDIKALQKELEQFAKLLKQKRITLGYTQADVGLTLGVLFGKVFS
Sequence7 121 AVKLEKEKLEQNPEESQDIKALQKELEQFAKLLKQKRITLGYTQADVGLTLGVLFGKVFS
************************************************************
SEQ ID 1 181 QTTICRFEALQLSFKNMCKLRPLLQKWVEEADNNENLQEICKAETLVQARKRKRTSIENR
Sequence7 181 QTTICRFEALQLSFKNMCKLRPLLQKWVEEADNNENLQEICKAETLVQARKRKRTSIENR
************************************************************
SEQ ID 1 241 VRGNLENLFLQCPKPTLQQISHIAQQLGLEKDVVRVWFCNRRQKGKRSSSDYAQREDFEA
Sequence7 241 VRGNLENLFLQCPKPTLQQISHIAQQLGLEKDVVRVWFCNRRQKGKRSSSDYAQREDFEA
************************************************************
SEQ ID 1 301 AGSPFSGGPVSFPLAPGPHFGTPGYGSPHFTALYSSVPFPEGEAFPPVSVTTLGSPMHSN
Sequence7 301 AGSPFSGGPVSFPLAPGPHFGTPGYGSPHFTALYSSVPFPEGEAFPPVSVTTLGSPMHSN
************************************************************
Regarding instant claim 182-183, reference claim 25 teaches wherein the contacting comprises six separate circular RNAs encoding the six factors.
Regarding instant claims 185-187, reference claims 1 and 4 teach wherein the cell is CD34+ and/or in suspension.
Regarding instant claim 188, reference claim 38 teaches wherein the cell is contacted with a circular RNA multiple times, including 2-4 times or more than 2, 3, or 4 times.
Regarding instant claim 189, reference claim 50 teaches whereby the method results in more reprogrammed iPSCs than using linear RNA(s) and/or a decrease in toxicity compared to using linear RNA(s).
This is a provisional nonstatutory double patenting rejection because the reference application claims have not in fact been patented.
Claims 172-189 are provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claims 1, 4-5, 25, 38, 50, 52-53, 55, 57, 77, 80, 93, 97, 103, 110-111, 116, 137, 142, 144-145, and 150 of copending Application No. 18/745761 (reference application) in view of Wesselhoeft (Wesselhoeft et al., Molecular Cell 74(3): 508-20 (2019); IDS ref.).
Although the reference claims do not teach wherein the circular RNA is complexed with a lipid nanoparticle (LNP), either covalently or non-covalently, it was known in the prior art to use LNPs for RNA delivery to a cell. Wesselhoeft teaches wherein expression vectors consisting of circular RNAs are encapsulated into lipid nanoparticles for deliver to target cells (pg. 515, left col., last para., to pg. 516, left col., 2nd para.). Thus, it would have been prima facie obvious to one of ordinary skill in the art to contact the circular RNA(s) in the methods of the reference claims using an LNP delivery system taught by Wesselhoeft that non-covalently complexes circular RNA to LNP. One of ordinary skill in the art would be motivated by Wesselhoeft teaching LNP delivery was equivalent to electroporation but provides an advantage for use in vivo for being less immunogenic but provides an advantage for being less immunogenic than linear RNAs (unrecognized by cellular RNA sensors) for use or with TLR-competent blood cells (Abstract; pg. 515, left col., last para., to pg. 516, left col., 2nd para.; Fig. 4). This is a provisional nonstatutory double patenting rejection because the reference application claims have not in fact been patented.
Conclusion
No claim is allowed.
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/ERIC J ROGERS/Examiner, Art Unit 1638
/KEVIN K HILL/Primary Examiner, Art Unit 1638