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 .
Priority
The instant application is a national stage entry under 35 U.S.C. § 371 of PCT/IB2022/057587 (filed 08/13/2022). Acknowledgement is made of Applicants’ claim for benefit of U.S. Application 63/233,196 (filed 08/13/2021).
Election/Restrictions
Applicant’s election without traverse of Group I in the reply filed on 07/09/2026 in response to a restriction requirement is acknowledged.
Upon entry of the 07/09/2026 amendment, claims 79, 81-82, 140, and 144 are cancelled, and new claims 152-157 are added.
Claims 1-2, 4-9, 11-13, 15-16, 18-20, 22-24, 26, 32, 41, 45, 73, 127-128, 130, 135, 138, and 152-157 are now pending in the instant application, read on the elected invention, and are examined on the merits herein.
Claim Interpretation
The following comments are made to establish broadest reasonable interpretation for the record.
Regarding claims 22-23, 153: These claims recite limitations directed to the growth factor vascular endothelial growth factor (VEGF). For instance, claim 22 recites, “…wherein the VEGF comprises a polypeptide sequence that has at least at or about 90% sequence identity to the sequence set forth in SEQ ID NO: 7 or a sequence thereof that lacks a signal peptide.” The limitation or a sequence thereof that lacks a signal peptide is recited in each of the instant claims; the term thereof is interpreted as referring to the VEGF molecule per se, and not necessarily the SEQ ID NO: 7 limitation preceding each recitation.
Regarding claim 32: This claim recites, “…wherein cells of the stratified epidermis secrete the recombinant growth factor and the recombinant insulin at levels that result in greater improvement in one or more marker(s) of angiogenic reorganization relative to a further skin substitute comprising either the recombinant growth factor or the recombinant insulin alone.” For clarity of record, the word further is interpreted in this context as akin to another.
Regarding claim 127: This claim is directed to a skin substitute produced by a method comprising differentiating keratinocytes into a stratified epidermis, and transducing cells of said epidermis with an adenoviral vector which encodes a modified proinsulin and a growth factor.
The product as claimed is determined to be a product-by-process claim. Product-by-process limitations are considered only in as far as the method of production imparts distinct structural or chemical characteristics or properties to the product. Therefore, if the product, as claimed, is the same or obvious over a product of the prior art (i.e., is not structurally or chemically distinct), the claim is considered unpatentable over the prior art, even though the prior art product is made by a different process. See MPEP 2113.
In the instant case, the process recited in claim 127 is effective to produce a skin substitute wherein the cells thereof express a modified proinsulin and a growth factor.
Claim Rejections - 35 USC § 112
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 6-7, 13, and 153 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 claims 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 inventors, at the time the application was filed, had possession of the claimed invention.
The claims under rejection recite limitations directed to the recombinant insulin expressed by the cells of the stratified epidermis of a skin substitute, wherein the recombinant insulin comprises (i) the sequence of amino acids set forth in SEQ ID NO: 5 (claims 6, 7) or SEQ ID NO: 6 (claim 153); (ii) is a functional variant that has a sequence of amino acids that has at least 90% sequence identity to the sequence of amino acids set forth in SEQ ID NO: 5 (claims 6, 7) or SEQ ID NO: 6 (claim 153); or (iii) is a two-chain form of (i) or (ii) that comprises an A-chain and a B-chain (claims 6, 153).
The issue at present is the scope of a functional variant of recombinant insulin covered by the claims. The limitations set forth in part (ii) of these claims require the functional variant to have about 90% sequence identity to SEQ ID NOs: 5 or 6. However, as both of these sequences contain 86 residues, the number of possible sequences with 90% identity thereto is extraordinarily great at ~1.545 x 1011. Thus, a functional variant of recombinant insulin that has a sequence of amino acids that has at least 90% sequence identity to the sequence of amino acids set forth in SEQ ID NO: 5 or SEQ ID NO: 6 is a genus of molecules.
To satisfy the written description aspect of 35 U.S.C. 112(a) for a claimed genus of molecules, it must be clear that: (1) the identifying characteristics of the claimed molecules have been disclosed, e.g., structure or other physical and/or chemical properties, by functional characteristics coupled with a known or disclosed cor-relation between function and structure, or by a com-bination of such identifying characteristics; or (2) a representative number of species within the genus must be disclosed. See Eli Lilly, 119 F.3d at 1568, 43 USPQ2d at 1406.
Regarding (1): A review of the specification fails to identify any clear guidance for what structural characteristics the claimed molecules must have. For instance, par. 0097 states (emphasis added):
In some embodiments, the recombinant insulin is a variant of a human insulin, such as a functional variant or species or allelic variant, or is a truncated form of human insulin that has activity. In some embodiments, variants of insulin, including allelic and species variants, variants encoded by splice variants and other functional variants, such as insulin analogs or other derivatized or modified forms, including polypeptides that have at least 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or more sequence identity to human insulin set forth in SEQ ID NO: 5 or to a processed insulin thereof that contains an A and B chain, so long as the insulin binds to the human insulin receptor to initiate a signaling cascade that results in an increase of glucose uptake and storage and/or a decrease of endogenous glucose production.
The bolded guidance seemingly directs the skilled artisan as to the required functional characteristics the recombinant insulin functional variant must possess; i.e., it must bind to the human insulin receptor to initiate the described signaling cascade. However, there isn’t a clear disclosure of any particular core chemical or physical structure which correlates to this function. This highlighted section recites guidance regarding sequence identity, but it’s unclear if this is in reference to the structure required for binding and initiation of the signaling cascade. Thus, Applicants have not disclosed the required identifying characteristics for the claimed genus of molecules.
Regarding (2): Applicants have disclosed human insulin proprotein (SEQ ID NO: 5), modified human insulin proprotein (SEQ ID NO: 6), bovine insulin (SEQ ID NO: 17), and porcine insulin (SEQ ID NO: 18). For clarity of record, it is noted that SEQ ID NOs: 34-41 are directed to insulin, but these sequence are for the individual A and B chains of the insulin molecules, and not considered a species of recombinant insulin. Thus, Applicants have only disclosed four species of recombinant insulin functional variants; this does not constitute a representative number for such a broad genus as is encompassed by the breadth of the functional variant limitation. Therefore, Applicants have not disclosed a representative number of species, as would be required to support description and to show possession of the entire genus.
Thus, one of ordinary skill in the art, in looking to the instant specification, would not be able to determine that Applicants were in possession of the invention, as claimed, at the time the invention was made. Accordingly, the claims are considered to lack sufficient written description and are properly rejected under 35 USC 112(a).
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 22-23 and 153 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.
Regarding claims 22-23, 153: Claims 22 and 153 recite, “…the VEGF comprises a polypeptide sequence that has at least at or about 90% sequence identity to the sequence set forth in SEQ ID NO: 7 or a sequence thereof that lacks a signal peptide”; claim 23 recites, “…the VEGF comprises the sequence of amino acids to the sequence set forth in SEQ ID NO: 7 or a sequence thereof that lacks a signal peptide.”
As set forth above, under broadest reasonable interpretation, the term thereof is interpreted as referring to the VEGF molecule. However, as currently written, it is unclear exactly what is meant by the limitation in this claim. Is this a Markush group of useable alternatives for the VEGF molecule? Using claim 23 for illustrative purposes, is the limitation satisfied by either a VEGF molecule which comprises SEQ ID NO: 7 or a VEGF molecule which comprises a sequence lacking a signal peptide? Or does the limitation or a sequence thereof that lacks a signal peptide refer specifically to residues 27-191 of SEQ ID NO: 7, excluding residues 1-26 (the signal peptide)?
As currently written, the metes and bounds of the instant claims are not clearly or precisely defined, rendering claims 22, 23, and 153 indefinite.
Claim Rejections - 35 USC § 102
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 1-2, 4-5, 18, 41, and 135 are rejected under 35 U.S.C. 102(a)(1) and (a)(2) as being anticipated by Hermonat, et al. (US 2003/0104575), as evidenced by Roberts and Horsley (Wiley Interdiscip Rev Dev Biol. 2014) and Treins, et al. (J Biol Chem. 2001).
Hermonat, et al. (hereinafter Hermonat) teaches the transfection of epithelial cells with an AAV useful for preparing recombinant skin (Abstract).
Regarding claims 1-2, 5, 18: Hermonat teaches a recombinant skin (r-skin) that is an intact stratified squamous epithelium composed of AAV-transfected keratinocyte cells which secrete a heterologous protein (pars. 0008, 0021); human GM-CSF, insulin, and factor VIII are transgenes secreted by the r-skin (pars. 0023, 0076). As evidenced by Roberts and Horsley, stratified squamous epithelium comprises a basal layer, a spinous layer, a suprabasal layer, a granular layer, and a cornified layer (i.e., stratum corneum) (pg. 1; par. 1). Thus, the r-skin of Hermonat anticipates:
the skin substitute comprising a stratified epidermis comprising a basal layer, a spinous layer, a granular layer and a stratum corneum, wherein cells of the stratified epidermis express a recombinant growth factor and a recombinant insulin limitations recited in claim 1;
the wherein the recombinant growth factor and recombinant insulin are secretable from cells of the stratified epidermis limitation recited in claim 2;
the wherein the recombinant insulin is or comprises a recombinant human insulin limitation recited in claim 5; and
the wherein the recombinant growth factor is GM-CSF limitation recited in claim 18.
Regarding claim 4: Following the above discussion, Hermonat teaches the proteins are secreted through the basal cell layer (par. 0034); this anticipates the wherein the cells of the stratified epidermis that express the recombinant growth factor and the recombinant insulin comprise cells of the basal layer limitation recited in claim 4.
Regarding claim 41: As evidenced by Treins, et al., hyperglycemia induces the production of advanced glycation end products (AGEs) generated by non-enzymatic reactions between glucose and free amino reactive group of proteins and lipids (pg. 43836; col. 1, par. 1). Thus, the insulin-secreting r-skin of Hermonat would necessarily reduce AGEs by reducing glucose levels, anticipating the limitation recited in the instant claim.
Regarding claim 135: Following the above discussion, Hermonat teaches the treatment of a subject with the r-skin (par. 0009); thus, Hermonat implicitly discloses a container comprising the r-skin, as the r-skin would necessarily need to be transported to the subject. This anticipates the container comprising the skin substitute of claim 1 limitation recited in claim 135.
Claims 45, 73, and 154-155 are rejected under 35 U.S.C. 102(a)(1) and (a)(2) as being anticipated by Bancel, et al. (US 2013/0259924).
Bancel, et al. (hereinafter Bancel) teaches compositions and methods for the preparation, manufacture and therapeutic use of polynucleotides, primary transcripts and modified mRNA (mmRNA) molecules (Abstract).
Regarding claims 45, 73, 154-155: Bancel teaches a two polynucleotide construct (see e.g., pars. 0113-0114), wherein two polynucleotides of the disclosure are incorporated into an mRNA construct and translated independently via inclusion of an internal ribosome entry site (IRES) for generation of multicistronic nucleic acid molecules (par. 0233); in an embodiment, this construct is transformed into a viral vector (par. 0257). In an embodiment, the two polynucleotides encode VEGF and insulin (Table 6; pgs. 55 and 71). Thus, the bicistronic vector of Bancel anticipates:
the bicistronic expression cassette comprising a polynucleotide encoding a recombinant human growth factor and a recombinant insulin limitation recited in claim 45;
the vector comprising the bicistronic expression cassette of claim 45 limitations recited in claim 73; and
the wherein the recombinant growth factor is VEGF limitations recited in claims 154 and 155.
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.
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 1-2, 4-7, 13, 18-20, 22, 24, 26, 41, 45, 73, 127, 135, and 152-155 are rejected under 35 U.S.C. 103 as being unpatentable over Hermonat, et al. (US 2003/0104575) in view of Supp, et al. (J Invest Dermatol. 2000) and Bancel, et al. (US 2013/0259924); as evidenced by Roberts and Horsley (Wiley Interdiscip Rev Dev Biol. 2014).
The teachings of Hermonat and Bancel are set forth above. Claims 1-2, 4-5, 18, 41, and 135 are anticipated by Hermonat; claims 45, 73, and 154-155 are anticipated by Bancel.
Supp, et al. (hereinafter Supp) teaches genetically-modified cultured skin substitutes with enhanced vascularization (Abstract).
Regarding claims 6-7, 13, 19, 22, 24, 26, 152-153: Following the above discussions, Hermonat does not teach the (1) VEGF or (2) sequence limitations recited in the instant claims.
Regarding (1), Supp, et al. teaches a cultured skin substitute (CSS) comprising keratinocytes transduced with retrovirus encoding VEGF and engrafted onto wound beds of mice (“Materials and Methods”; pgs. 6-7). Healing CSS were excised from mice at 3, 7, and 14 days after surgery; VEGF was quantified via Northern blot, with greatly elevated levels of VEGF detected in the VEGF-modified grafts compared to the control grafts (pg. 9; par. 1). Further, VEGF-modified CSS had improved vascularization compared to control CSS, with better adherence to the wound beds and increased bleeding (pg. 9; par. 2).
It would have been prima facie obvious to a person having ordinary skill in the art to have modified the r-skin of Hermonat by modifying the AAV vector to encode for VEGF in addition to insulin. This conclusion of obviousness is based on the ‘teaching, suggestion, or motivation rationale’. Supp, et al. teaches aberrant regulation of VEGF expression is associated with defective wound repair, such as the abnormal wound healing seen in diabetic mice (pg. 6; col. 1, par. 1); enhanced expression of VEGF in skin substitutes is useful for diabetic wound healing, which is associated with reduced VEGF expression (pg. 12; par. 3). As Hermonat teaches the insulin-expressing r-skin is useful for the treatment of diabetes (par. 0028), it would have therefore been obvious to incorporate the teachings of Supp to reduce the risk of impaired wound healing for a diabetic subject. Additionally, such a modification is well within the purview of the skilled artisan, as evidenced by both of the cited disclosures.
Regarding (2), the bicistronic vector of Bancel is set forth above. In an embodiment, the insulin molecule comprises an amino acid sequence as set forth in SEQ ID NO: 1083 (Table 6; pg. 55), which shares 100% sequence identity to instant SEQ ID NO: 5, and 95.35% sequence identity to instant SEQ ID NO: 6; please see Office Action (OA) Appendix I for sequence alignments (pg. 2). In an embodiment, the VEGF molecule comprises an amino acid sequence as set forth in SEQ ID NO: 1369 (Table 6; pg. 71), which shares 100% sequence identity to instant SEQ ID NOs: 7 and 44 (pg. 4; OA Appendix I). Bancel teaches the polynucleotide constructs of the disclosure have been designed to improve stability, cellular access by the compositions, engagement with translational machinery, mRNA half-life, translation efficiency, secretion efficiency (par. 0093).
Therefore, it would have been prima facie obvious to a person having ordinary skill in the art to have further modified the r-skin of Hermonat by using a bicistronic vector encoding an insulin molecule comprising SEQ ID NO: 1083 and a VEGF molecule comprising SEQ ID NO: 1369, as taught by Bancel. This conclusion of obviousness is based on the ‘teaching, suggestion, or motivation rationale’. One would be motivated to do so for the improved stability, cellular access by the compositions, engagement with translational machinery, mRNA half-life, translation efficiency, and secretion efficiency of the construct as disclosed by Bancel (par. 0093). Further, the skilled artisan would have more than a reasonable expectation of success in doing so, as Bancel teaches the construct can be transformed into a viral vector (par. 0257) and can be used with bioengineered skin substitutes (par. 0883). As the IRES element inserted into the construct of Bancel for independent translation of VEGF and insulin reads on the bicistronic element recited in claim 26, the modified r-skin of Hermonat renders obvious:
the wherein the recombinant insulin comprises the sequence of amino acids set forth in SEQ ID NO: 5 limitations recited in claims 6 and 7;
the wherein the recombinant insulin is a functional variant that has a sequence of amino acids that has at least 90% sequence identity to the sequence of amino acids set forth in SEQ ID NO: 6 limitation recited in claim 13;
the wherein the recombinant growth factor is a VEGF limitation recited in claim 19;
the wherein the VEGF comprises a polypeptide sequence that has at least at or about 90% sequence identity to the sequence set forth in SEQ ID NO: 7 limitation recited in claim 22;
the wherein the VEGF comprises a polypeptide sequence that has at least at or about 90% sequence identity to the sequence of amino acids set forth in SEQ ID NO: 44, and the recombinant insulin comprises a polypeptide sequence that has at least at or about 90% sequence identity to the sequence of amino acids set forth in SEQ ID NO: 6 limitations recited in claim 24;
the wherein the recombinant growth factor and the recombinant insulin are encoded by a bicistronic expression cassette comprising a polynucleotide encoding the recombinant growth factor and a polynucleotide encoding the recombinant insulin separated by a bicistronic element limitations recited in claim 26;
the wherein cells of the stratified epidermis express the recombinant growth factor that is VEGF and the recombinant insulin limitations recited in claim 152; and
the wherein the recombinant insulin is a functional variant that has a sequence of amino acids that has at least 90% sequence identity to the sequence of amino acids set forth in SEQ ID NO: 6; and the VEGF comprises a polypeptide sequence that has at least at or about 90% sequence identity to the sequence set forth in SEQ ID NO: 7 limitations recited in claim 153.
Regarding claim 16: Following the above discussion, Bancel teaches the insulin molecule which comprises SEQ ID NO: 1038 is encoded by a nucleic acid sequence as set forth in SEQ ID NO: 465 (Table 6; pg. 55). SEQ ID NO: 465 shares 98.2% sequence identity to instant SEQ ID NO: 2 (pg. 3 of OA Appendix I); this reads on the wherein the recombinant insulin is encoded by a polynucleotide that comprises a sequence that has at least at or about 90% sequence identity to the sequence set forth in SEQ ID NO: 2 limitation recited in claim 16.
Regarding claim 20: Following the above discussion, Bancel teaches the VEGF molecule which comprises SEQ ID NO: 1369 is encoded by a nucleic acid sequence as set forth in SEQ ID NO: 745 (Table 6; pg. 71). SEQ ID NO: 745 shares 100% sequence identity to instant SEQ ID NO: 4 (pgs. 5, 10 of OA Appendix I); this reads on the wherein the VEGF is encoded by a polynucleotide sequence that has at least at or about 90% sequence identity to the sequence set forth in SEQ ID NO: 4 limitation recited in claim 20.
Regarding claim 127: It is set forth above the skin substitute of the instant claim is a product-by-process limitation. The instant specification discloses the insulin molecule comprising the amino acid sequence of SEQ ID NO: 5 is a human insulin proprotein; i.e., human proinsulin (pg. 113). As SEQ ID NO: 1083 of Bancel shares 100% sequence identity to instant SEQ ID NO: 5 (i.e., the insulin molecule of Bancel comprising SEQ ID NO: 1083 is a human proinsulin molecule), the modified r-skin of Hermonat reads on the skin substitute of claim 127.
Claims 1-2, 4-5, 8, 15, 18, 23, 41, 45, 73, 135, and 154-155 are rejected under 35 U.S.C. 103 as being unpatentable over Hermonat, et al. (US 2003/0104575) in view of Supp, et al. (J Invest Dermatol. 2000) and Bancel, et al. (US 2013/0259924), further in view of Gupta, et al. (US 2024/0279300); as evidenced by Roberts and Horsley (Wiley Interdiscip Rev Dev Biol. 2014) and Lipkind and Steiner (Biochemistry. 1999).
The teachings of Hermonat, Supp, and Bancel are set forth above. Claims 1-2, 4-5, 18, 41, and 135 are anticipated by Hermonat; claims 45, 73, and 154-155 are anticipated by Bancel.
Gupta, et al. (hereinafter Gupta) teaches modified nucleic acid sequences encoding insulin and glucokinase (Abstract).
Regarding claims 8, 15, 23: Following the above discussion, Hermonat does not teach the limitations recited in the instant claims.
However, Gupta teaches a proinsulin molecule comprising an amino acid sequence as set forth in SEQ ID NO: 144 (par. 0014), which shares 100% sequence identity to instant SEQ ID NOs: 6, 36, and 41; additionally, and as seen in the sequence alignment, the residue of SEQ ID NO: 144 which corresponds to residue 10 of the B chain of the modified proinsulin is aspartic acid (pg. 11 of OA Appendix I).
It would have been prima facie obvious to a person having ordinary skill in the art to have further modified the r-skin of Hermonat by substituting the nucleic acid sequence encoding the insulin molecule comprising an amino acid sequence as set forth in SEQ ID NO: 1038 with a nucleic acid sequence encoding an insulin molecule comprising an amino acid sequence as set forth in SEQ ID NO: 144, as taught by Gupta. The use of a nucleic acid sequence encoding SEQ ID NO: 144 in place of a nucleic acid sequence encoding SEQ ID NO: 1038 is a predictable use of prior art elements according to their established functions as coding nucleic acid sequences, leading to the predictable result of a translated insulin molecule. This rationale aligns with the principle of a simple substitution of one known element for another to obtain predictable results; see MPEP 2143(I)(B). This renders obvious:
the wherein the recombinant insulin is an AspB10 insulin analog comprising a histidine to aspartic acid mutation at position 10 in the B chain of the modified human proinsulin compared to wild-type insulin set forth in SEQ ID NO: 5 limitation recited in claim 8;
the wherein the recombinant insulin comprises an A chain set forth in SEQ ID NO: 36 and a B chain set forth in SEQ ID NO: 41 limitation recited in claim 15; and
the wherein the VEGF comprises the sequence of amino acids set forth in SEQ ID NO: 7 or a sequence thereof that lacks the signal peptide, and the recombinant insulin comprises the sequence of amino acids set forth in SEQ ID NO: 6 limitations recited in claim 23.
Claims 1-2, 4-5, 9, 11, 18, 41, 45, 73, 135, and 154-155 are rejected under 35 U.S.C. 103 as being unpatentable over Hermonat, et al. (US 2003/0104575) in view of Supp, et al. (J Invest Dermatol. 2000) and Bancel, et al. (US 2013/0259924), further in view of Gupta, et al. (US 2024/0279300) and Brown (US 2007/0202568); as evidenced by Roberts and Horsley (Wiley Interdiscip Rev Dev Biol. 2014) and Lipkind and Steiner (Biochemistry. 1999).
The teachings of Hermonat, Supp, Bancel, and Gupta are set forth above. Claims 1-2, 4-5, 18, 41, and 135 are anticipated by Hermonat; claims 45, 73, and 154-155 are anticipated by Bancel.
Brown teaches cleavage sites for the protease furin (Abstract).
Regarding claim 9: Following the above discussion, Gupta teaches an embodiment wherein the proinsulin molecule comprises a furin endoprotease cleavage site (par. 0106). Gupta does not teach where the furin sequence is inserted in the proinsulin molecule.
However, Brown teaches furin cleavage sequences are inserted into a region that divides the protein into separate domains; e.g., a region with predominant polar residues (par. 0030). As evidenced by Lipkind and Steiner, the intracellular conversion of proinsulin to insulin occurs via cleavage at the two dibasic sites of the molecule: Arg31-Arg32, B-chain—-C-peptide junction; and Lys64-Arg-65, A-chain—C-peptide junction (Abstract).
Therefore, it would have been prima facie obvious to a person having ordinary skill in the art to have modified the proinsulin molecule comprising a furin endoprotease cleavage site of Gupta by inserting a furin cleavage sequence into the Arg31-Arg32 (B-chain—-C-peptide junction) or the Lys64-Arg-65 (A-chain—C-peptide junction), as taught by Brown. This conclusion of obviousness is based on the ‘obvious to try rationale’. There are only two dibasic sites in proinsulin, and Brown teaches furin cleavage sequences should be inserted into a region with polar residues (par. 0030). Therefore, it would have been obvious for the skilled artisan to try both the Arg31-Arg32 and Lys64-Arg-65 sites, which is well within the purview of the skilled artisan, as evidenced by the Brown disclosure. This rationale aligns with choosing from a finite number of identified, predictable solutions with a reasonable expectation of success; see MPEP 2143(I)(E).
This renders obvious the wherein the skin substitute comprises a polynucleotide encoding a proinsulin comprising at least one furin recognition sequence in place of the endopeptidase Arg31-Arg32 cleavage site and/or the endopeptidase Lys64-Arg65 cleavage site limitations recited in claim 9.
Regarding claim 11: Following the above discussion, Brown teaches furin consensus sequence RXRR, where X is any amino acid (par. 0017). Therefore, it would have been prima facie obvious to a person having ordinary skill in the art to have used the RXRR sequence of Brown for the furin cleavage sequence, as Brown teaches furin recognizes this consensus sequence; it would have been obvious to the skilled artisan to use a consensus sequence recognized by furin. Further, the use of furin sequences to introduce cleavage sites in proteins is a well-known technique in the art, as evidenced by both the Brown and Gupta disclosures; thus, one in the art would have more than a reasonable expectation of success. This renders obvious the wherein the at least one furin recognition sequence comprises the consensus sequence R-X-R-R, where X is any amino acid (SEQ ID NO: 8) limitation recited in claim 11.
Claims 1-2, 4-5, 11-12, 18, 41, 45, 73, 135, and 154-155 are rejected under 35 U.S.C. 103 as being unpatentable over Hermonat, et al. (US 2003/0104575) in view of Supp, et al. (J Invest Dermatol. 2000) and Bancel, et al. (US 2013/0259924), further in view of Gupta, et al. (US 2024/0279300) and Hosaka, et al. (J Biol Chem. 1991); as evidenced by Roberts and Horsley (Wiley Interdiscip Rev Dev Biol. 2014) and Lipkind and Steiner (Biochemistry. 1999).
The teachings of Hermonat, Supp, Bancel, and Gupta are set forth above. Claims 1-2, 4-5, 18, 41, and 135 are anticipated by Hermonat; claims 45, 73, and 154-155 are anticipated by Bancel.
Hosaka, et al. (hereinafter Hosaka) teaches the Arg-X-Lys/Arg-Arg motif as a signal for precursor cleavage by furin (Title).
Regarding claim 11: Following the above discussion, Hosaka teaches furin consensus sequence Arg-X-Lys/Arg-Arg (RXK/RR); when expressed in endocrine and non-endocrine cells, a precursor with the RXKR sequence was cleaved in both types of cells, whereas a precursor with the RXRR sequence was cleaved only in the endocrine cells (Abstract).
Therefore, it would have been prima facie obvious to a person having ordinary skill in the art to have used the RXKR furin consensus sequence for the proinsulin molecule comprising a furin endoprotease cleavage site of Gupta. This conclusion of obviousness is based on the ‘teaching, suggestion, or motivation rationale’; one would be motivated to use RXKR for the proinsulin molecule of Gupta, as the proinsulin is transfected into keratinocytes (i.e., non-endocrine cells) for the r-skin of Hermonat. Further, the skilled artisan would have more than a reasonable expectation of success in doing so, as Hosaka teaches this furin consensus sequence results in precursor cleavage for non-endocrine cells as well as endocrine cells (Abstract). This renders obvious the wherein the at least one furin recognition sequence comprises the consensus sequence R-X-K-R, where X is any amino acid (SEQ ID NO: 9) limitation recited in claim 11.
Regarding claim 12: Following the above discussion, it would have been prima facie obvious to a person having ordinary skill in the art to have further modified the proinsulin molecule by using RTKR or RQKR as the furin cleavage sequence. This conclusion of obviousness is based on the ‘obvious to try rationale’. There are only twenty amino acids possible for the X residue of RXKR; therefore, it would have been obvious for the skilled artisan to try all twenty. Further, as evidenced by the Gupta and Hosaka disclosures, the use of furin sequences to introduce cleavage sites in proteins is a well-known technique in the art; thus, the one skilled in the art would have more than a reasonable expectation of success. This renders obvious the wherein the at least one furin recognition sequence is RTKR (SEQ ID NO: 10) or RQKR (SEQ ID NO: 42) limitations recited in claim 12.
Claims 1-2, 4-5, 18, 32, 41, 45, 73, 135, and 154-155 are rejected under 35 U.S.C. 103 as being unpatentable over Hermonat, et al. (US 2003/0104575) in view of Supp, et al. (J Invest Dermatol. 2000) and Bancel, et al. (US 2013/0259924), further in view of Yamagishi, et al. (Microvasc Res. 1999); as evidenced by Roberts and Horsley (Wiley Interdiscip Rev Dev Biol. 2014).
The teachings of Hermonat, Supp, and Bancel are set forth above. Claims 1-2, 4-5, 18, 41, and 135 are anticipated by Hermonat; claims 45, 73, and 154-155 are anticipated by Bancel.
Yamagishi, et al. (hereinafter Yamagishi) teaches the effects of insulin on the growth and tube formation of microvascular endothelial cells (Abstract).
Regarding claim 32: Following the above discussion, Hermonat does not explicitly teach the limitations of the instant claim.
However, Yamagishi teaches human microvascular endothelial cells (ECs) exposed to insulin (pg. 331; col. 2, par. 1). Angiogenesis was induced in the ECs via hypoxic treatment; VEGF mRNA levels increased as atmospheric O2 concentration decreased (pg. 333; col. 1, pars. 2-3). The effects of insulin and hypoxia on the in vitro tube formation of microvascular EC were examined (pg. 333; col. 1, par. 3); the effects of insulin and hypoxia were additive in the tube formation assay (pg. 335; col. 2, par. 1). As supported by the instant specification, markers of angiogenic reorganization can be evaluated in a tube formation assay (par. 0111).
Therefore, it would have been prima facie obvious for a person having ordinary skill in the art to have expected the cells of the modified r-skin of Hermonat to have secreted the recombinant VEGF and recombinant insulin at levels that result in greater improvement in angiogenic reorganization relative to a skin substitute comprising either the recombinant VEGF or recombinant insulin alone. This conclusion of obviousness is based on the ‘teaching, suggestion, or motivation rationale’. Yamagishi teaches hypoxia-induced VEGF expression combined with insulin treatment results in an additive effect of tube formation in microvascular endothelial cells (pg. 335); i.e., insulin combined with increased VEGF expression results in an additive effect of markers of angiogenic reorganization. Further, the skilled artisan would have more than a reasonable expectation of success in the additive effect of VEGF and insulin for the modified r-skin of Hermonat, as the microvascular ECs of Yamagishi are from human skin (pg. 330; col. 2, par. 1).
This renders obvious the wherein cells of the stratified epidermis secrete the recombinant growth factor and the recombinant insulin at levels that result in greater improvement in one or more marker(s) of angiogenic reorganization relative to a further skin substitute comprising either the recombinant growth factor or the recombinant insulin alone limitation recited in claim 32.
Claims 1-2, 4-5, 18, 41, 45, 73, 128, 130, 135, 138, and 154-157 are rejected under 35 U.S.C. 103 as being unpatentable over Hermonat, et al. (US 2003/0104575) in view of Supp, et al. (J Invest Dermatol. 2000) and Bancel, et al. (US 2013/0259924), further in view of Koepsel and Gatz (US 2020/0128815); as evidenced by Roberts and Horsley (Wiley Interdiscip Rev Dev Biol. 2014).
The teachings of Hermonat, Supp, and Bancel are set forth above. Claims 1-2, 4-5, 18, 41, and 135 are anticipated by Hermonat; claims 45, 73, and 154-155 are anticipated by Bancel.
Koepsel and Gatz (hereinafter Koepsel) teaches tissue container systems (Abstract).
Regarding claims 128, 156: Following the above discussion, Hermonat teaches the treatment of a subject with the r-skin (par. 0009); thus, Hermonat implicitly discloses storage and/or transportation of the r-skin. Still, Hermonat does not teach the limitations recited in the instant claims.
However, Koepsel teaches current engineered tissues must often be stored and shipped under carefully controlled conditions to maintain viability and function, wherein engineered tissue products take weeks or months to produce but must be used within hours or days after manufacture (par. 0003). As a result, tissue engineering companies must continually operate with their production facilities at top capacity and absorb the costs of unsold product which must be discarded (par. 0003). Koepsel teaches a cryopreservation process for skin equivalents comprising culturing the skin equivalent in a cryoprotectant solution (par. 0045), as well as a tissue container system that supports the transport, thawing, and use of cryopreserved human skin equivalents (par. 0021); the cryopreserved skin equivalents enclosed within the container are storable for up to 12 or 24 months without a substantial loss of viability (par. 0044).
Therefore, it would have been prima facie obvious to a person having ordinary skill in the art to have further modified the r-skin of Hermonat by cryopreserving the r-skin in a cryoprotectant solution as well as use the tissue container system of Koepsel for needed transportation. This conclusion of obviousness is based on the ‘teaching, suggestion, or motivation rationale’. One would be motivated to use the cryopreservation process and tissue container system of Koepsel for the maintained viability during long-term storage and/or transportation (pars. 0044-0045). Further, as Koepsel teaches the system and cryopreservation process is suitable for skin equivalents (par. 0021), the skilled artisan would have more than a reasonable expectation of success for use with the modified r-skin of Hermonat. This renders obvious the cryopreserved skin substitute, comprising the skin substitute and a cryoprotectant limitations recited in claims 128 and 156.
Regarding claims 130, 157: Following the above discussion, Koepsel teaches an embodiment wherein the cryopreserved skin equivalent is removed from the tissue container and placed on an absorbent medium, wherein the absorbent medium is a gauze pad (par. 0051); this renders obvious the skin substitute dressing comprising the skin substitute and an absorbent gauze, wherein the skin substitute is overlaid on the absorbent gauze limitations recited in claims 130 and 157.
Regarding claim 138: Following the above discussion, Koepsel teaches an embodiment wherein the tissue container system is sealed in a sterile bag (par. 0011); this renders obvious the package comprising the container of claim 135, wherein the package is a bag limitation recited in claim 138.
Conclusion
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/GINA PRONZATI/Examiner, Art Unit 1633
/ALLISON M FOX/Primary Examiner, Art Unit 1633
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