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 .
Application Status
This action is written in response to applicant’s correspondence received 02/13/2024. Claims 1-3, 5, 7-11, 13, 15, 17-20, 22-24, and 26-27 are currently pending.
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 1-3, 5, 7, 9-11, 13, 15, 17-20, 22-24, and 26-27 , 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 term “substantially identical” in claims 1, 3 and 13 is a relative term which renders the claim indefinite. The term is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree to which the sequences must be identical to be considered substantially identical, nor whether the substantial identity also encompasses function, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. The term “substantially” may mean a majority (i.e., >50%) or almost all (e.g., >/= 95%). While the specification provides some examples of percent identities which may be considered substantially identical (e.g., at least 95%, 96%, 97%, 98%, 99% or more; see page 32 ln 1-2, 15-16, 26-27), these are merely examples, not a clear definition, and do not indicate a clear intent to depart from the ordinary meaning or disavow the full scope of the claim term (MPEP 2111.01.IV).
While the term is vague and indefinite, in the interests of compact prosecution and customer service, and for comparison to the prior art, the term is interpreted as encompassing any sequence in which the bulk or majority of the sequence (i.e., >50%) is identical, and/or sequences which encode effectively the same functional and regulatory elements such as known protein coding sequences, known promoter sequences, etc.
Amending the claims to delete the term “or substantially identical to” would obviate this rejection.
Those claims identified in the statement of rejection but not explicitly referenced in the rejection are also rejected for depending from a rejected claim but failing to remedy the indefiniteness therein.
Claim Interpretation
The following is a quotation of 35 U.S.C. 112(f):
(f) Element in Claim for a Combination. – An element in a claim for a combination may be expressed as a means or step for performing a specified function without the recital of structure, material, or acts in support thereof, and such claim shall be construed to cover the corresponding structure, material, or acts described in the specification and equivalents thereof.
The following is a quotation of pre-AIA 35 U.S.C. 112, sixth paragraph:
An element in a claim for a combination may be expressed as a means or step for performing a specified function without the recital of structure, material, or acts in support thereof, and such claim shall be construed to cover the corresponding structure, material, or acts described in the specification and equivalents thereof.
The claims in this application are given their broadest reasonable interpretation using the plain meaning of the claim language in light of the specification as it would be understood by one of ordinary skill in the art. The broadest reasonable interpretation of a claim element (also commonly referred to as a claim limitation) is limited by the description in the specification when 35 U.S.C. 112(f) or pre-AIA 35 U.S.C. 112, sixth paragraph, is invoked.
As explained in MPEP § 2181, subsection I, claim limitations that meet the following three-prong test will be interpreted under 35 U.S.C. 112(f) or pre-AIA 35 U.S.C. 112, sixth paragraph:
(A) the claim limitation uses the term “means” or “step” or a term used as a substitute for “means” that is a generic placeholder (also called a nonce term or a non-structural term having no specific structural meaning) for performing the claimed function;
(B) the term “means” or “step” or the generic placeholder is modified by functional language, typically, but not always linked by the transition word “for” (e.g., “means for”) or another linking word or phrase, such as “configured to” or “so that”; and
(C) the term “means” or “step” or the generic placeholder is not modified by sufficient structure, material, or acts for performing the claimed function.
Use of the word “means” (or “step”) in a claim with functional language creates a rebuttable presumption that the claim limitation is to be treated in accordance with 35 U.S.C. 112(f) or pre-AIA 35 U.S.C. 112, sixth paragraph. The presumption that the claim limitation is interpreted under 35 U.S.C. 112(f) or pre-AIA 35 U.S.C. 112, sixth paragraph, is rebutted when the claim limitation recites sufficient structure, material, or acts to entirely perform the recited function.
Absence of the word “means” (or “step”) in a claim creates a rebuttable presumption that the claim limitation is not to be treated in accordance with 35 U.S.C. 112(f) or pre-AIA 35 U.S.C. 112, sixth paragraph. The presumption that the claim limitation is not interpreted under 35 U.S.C. 112(f) or pre-AIA 35 U.S.C. 112, sixth paragraph, is rebutted when the claim limitation recites function without reciting sufficient structure, material or acts to entirely perform the recited function.
Claim limitations in this application that use the word “means” (or “step”) are being interpreted under 35 U.S.C. 112(f) or pre-AIA 35 U.S.C. 112, sixth paragraph, except as otherwise indicated in an Office action. Conversely, claim limitations in this application that do not use the word “means” (or “step”) are not being interpreted under 35 U.S.C. 112(f) or pre-AIA 35 U.S.C. 112, sixth paragraph, except as otherwise indicated in an Office action.
Claim 17 recites a “means for mitigating the foreign body response (FBR)”. The term “means” is a generic placeholder for performing the function of mitigating the FBR. It is modified by the functional language, “mitigating the FBR”, but is not modified by sufficient structure, material, or acts for performing the claimed function.
The specification defines a means for mitigating FBR as follows:
a compound or polymer that mitigates the FBR (as defined herein) to the device (e.g., an afibrotic compound or afibrotic polymer). In an embodiment, an afibrotic polymer comprises a biocompatible, zwitterionic polymer, e.g., as described in WO 2017/218507, WO 2018/140834, or Liu et al…In an embodiment, the compound is a compound of Formula (I). (p. 3 ln 23-30)
"Afibrotic", as used herein, means a compound or material that mitigates the foreign body response (FBR)…(e.g., a hydrogel capsule comprising a polymer covalently modified with a compound listed in Table 4). (p. 8 ln 22-25)
On p. 47, the specification discloses Formula (I):
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67
308
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Greyscale
On pages 66-74, in Table 4, the specification discloses several exemplary afibrotic compounds for, as described above, mitigating the FBR
Therefore, a “means for mitigating the FBR” is interpreted as referring to the afibrotic compounds of Formula (I) and displayed in Table 4.
Claim Rejections - 35 USC § 112(a) – Written Description
The following is a quotation of the first paragraph of 35 U.S.C. 112(a):
IN GENERAL.—The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor or joint inventor of carrying out the invention.
The following is a quotation of the first paragraph of pre-AIA 35 U.S.C. 112:
The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor of carrying out his invention.
Claims 1-3, 5, 7-11, 13, 15, 17-20, 22-24, and 26-27 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 pre-AIA the inventor(s), at the time the application was filed, had possession of the claimed invention.
Claim 1 is drawn to an isolated polynucleotide which, among other characteristics, comprises a first expression cassette comprising a first polyA signal sequence that is identical or substantially identical to nucleotides 4304-4824 of SEQ ID NO: 15. While the specification provides support for the cassette comprising a polyA signal identical to positions 3175-3696 of SEQ ID NO: 15, neither the specification nor the art supports a polyA sequence identical to positions 4304-4824 of that sequence.
The specification provides the following information about SEQ ID NO: 15:
On page 2, the specification states that, “In an embodiment, the first exogenous expression cassette comprises nucleotide sequence [sic] comprises nucleotides 337 to 3696 of SEQ ID NO: 15.” This is consistent with what is shown in FIG. 5, which shows SEQ ID NO: 15 with the EF2A promoter sequence (SEQ ID NO: 16) and ARSB precursor sequence (SEQ ID NO: 17) underlined and in bold italics, respectively (see the brief description of the drawings, p. 5 ln 6-9). On page 32, lines 24-26, the specification states, “In an embodiment, the expression cassette further comprises a polyA sequence that consists essentially of, or consists of, nucleotides 3175-3696 of SEQ ID NO: 15”. This is further supported by Table 2A (pp. 33-34), which shows that positions 4303-4824 are primarily part of an EGFP/Puro coding sequence, while positions 3175-3696 are a rBG polyA signal sequence (table reproduced below and annotated for clarity):
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media_image2.png
252
591
media_image2.png
Greyscale
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media_image3.png
135
593
media_image3.png
Greyscale
On the other hand, Table 2B shows that it is actually positions 4303-4824 of SEQ ID NO: 18 which comprise a polyA sequence:
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media_image4.png
357
589
media_image4.png
Greyscale
Lastly, the specification also provides exemplary vector maps in FIGs. 8-10. However, it is not clear how SEQ ID NO: 15 relates to those maps, because only SEQ ID NOs: 18-20 are mentioned in the brief description of those drawings.
Evidence provided by the prior art:
The prior art supports the disclosures in the specification and contradicts the claims. The results of a sequence search for various portions of SEQ ID NO: 15 evidence that positions 3175-3696 of SEQ ID NO: 15, but not 4303-4824, most likely represent an EGFP sequence, as indicated by Tables 2A and 2B, not a polyA sequence as claimed.
For example, one of the top BLAST hits to positions 4303-4824 of SEQ ID NO: 15 is a “Cloning vector pABsv40-EG” (GenBank Accession No. MH290749.1, published 06/06/2018). Positions 4764-4824 of vector pABsv40-EG are a 100% match to positions 4334-4824 of SEQ ID NO: 15. However, the GenBank listing for the vector indicates that those positions are part of the coding sequence for EGFP (positions 4771-5490), not a polyA sequence (see ‘FEATURES’). Please see the below alignment, where the EGFP start codon at position 4771 of the vector, corresponding to position 4341 of SEQ ID NO: 15, is underlined (GenBank reference and alignment are attached to this office action):
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695
689
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This result is consistent with Table 2A, which shows that positions 4341-5657 of SEQ ID NO: 15 represent an EGFP coding sequence (see above).
A search of positions 3175-3696 of SEQ ID NO: 15, on the other hand, does return a clear hit to a rabbit beta globin polyA sequence in U.S. Patent to Little et al. (hereinafter ‘Little’, effective filing date 05/24/2017). Little teaches a rabbit beta globin polyA sequence identical to nucleotides 3175-3696 of SEQ ID NO: 15 (SEQ ID NO: 20, col 49-50):
GenCore version 6.5.2
Copyright (c) 1993 - 2026 Biocceleration Ltd.
OM nucleic - nucleic search, using sw model
Run on: February 6, 2026, 15:13:15 ; Search time 1 Seconds
(without alignments)
6.623 Million cell updates/sec
Title: US-18-276-010-15
Perfect score: 6344
Sequence: 1 ttaaccctagaaagatagtc..........gattatctttctagggttaa 6344
Scoring table: IDENTITY_NUC
Gapop 10.0 , Gapext 1.0
Searched: 1 seqs, 522 residues
Total number of hits satisfying chosen parameters: 2
Minimum DB seq length: 0
Maximum DB seq length: inf
Post-processing: Minimum Match 0%
Maximum Match 100%
Listing first 2 summaries
Database : SEQIDNO_20_RBGPOLYA.seq:*
SUMMARIES
%
Result Query
No. Score Match Length DB ID Description
----------------------------------------------------------------------------
1 522 8.2 522 1 SEQIDNO_20_RBGPOLYA
c 2 20 0.3 522 1 SEQIDNO_20_RBGPOLYA
ALIGNMENTS
RESULT 1
SEQIDNO_20_RBGPOLYA
Query Match 8.2%; Score 522; DB 1; Length 522;
Best Local Similarity 100.0%;
Matches 522; Conservative 0; Mismatches 0; Indels 0; Gaps 0;
Qy 3175 TCCTCAGGTGCAGGCTGCCTATCAGAAGGTGGTGGCTGGTGTGGCCAATGCCCTGGCTCA 3234
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1 TCCTCAGGTGCAGGCTGCCTATCAGAAGGTGGTGGCTGGTGTGGCCAATGCCCTGGCTCA 60
Qy 3235 CAAATACCACTGAGATCTTTTTCCCTCTGCCAAAAATTATGGGGACATCATGAAGCCCCT 3294
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 61 CAAATACCACTGAGATCTTTTTCCCTCTGCCAAAAATTATGGGGACATCATGAAGCCCCT 120
Qy 3295 TGAGCATCTGACTTCTGGCTAATAAAGGAAATTTATTTTCATTGCAATAGTGTGTTGGAA 3354
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 121 TGAGCATCTGACTTCTGGCTAATAAAGGAAATTTATTTTCATTGCAATAGTGTGTTGGAA 180
Qy 3355 TTTTTTGTGTCTCTCACTCGGAAGGACATATGGGAGGGCAAATCATTTAAAACATCAGAA 3414
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 181 TTTTTTGTGTCTCTCACTCGGAAGGACATATGGGAGGGCAAATCATTTAAAACATCAGAA 240
Qy 3415 TGAGTATTTGGTTTAGAGTTTGGCAACATATGCCCATATGCTGGCTGCCATGAACAAAGG 3474
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 241 TGAGTATTTGGTTTAGAGTTTGGCAACATATGCCCATATGCTGGCTGCCATGAACAAAGG 300
Qy 3475 TTGGCTATAAAGAGGTCATCAGTATATGAAACAGCCCCCTGCTGTCCATTCCTTATTCCA 3534
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 301 TTGGCTATAAAGAGGTCATCAGTATATGAAACAGCCCCCTGCTGTCCATTCCTTATTCCA 360
Qy 3535 TAGAAAAGCCTTGACTTGAGGTTAGATTTTTTTTATATTTTGTTTTGTGTTATTTTTTTC 3594
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 361 TAGAAAAGCCTTGACTTGAGGTTAGATTTTTTTTATATTTTGTTTTGTGTTATTTTTTTC 420
Qy 3595 TTTAACATCCCTAAAATTTTCCTTACATGTTTTACTAGCCAGATTTTTCCTCCTCTCCTG 3654
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 421 TTTAACATCCCTAAAATTTTCCTTACATGTTTTACTAGCCAGATTTTTCCTCCTCTCCTG 480
Qy 3655 ACTACTCCCAGTCATAGCTGTCCCTCTTCTCTTATGGAGATC 3696
||||||||||||||||||||||||||||||||||||||||||
Db 481 ACTACTCCCAGTCATAGCTGTCCCTCTTCTCTTATGGAGATC 522
Based on the preponderance of evidence, the specification does not appear to provide support for a polyA sequence at positions 4304-4824 of SEQ ID NO: 15. However, as described above, the specification does provide support for a polyA sequence at positions 3175-3696 of SEQ ID NO: 15, in the first expression cassette.
Those claims identified in the statement of rejection but not explicitly referenced in the rejection are also rejected for depending from a rejected claim but failing to remedy the mischaracterization of the recited positions.
In the interests of customer service and compact prosecution, positions 3175-3696 will be searched for comparison to the prior art, since the preponderance of evidence suggests that this is the intended location of the polyA sequence in that cassette.
If this analysis is correct, then amending the claims to recite positions 3175-3696 would obviate this rejection.
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:
Determining the scope and contents of the prior art.
Ascertaining the differences between the prior art and the claims at issue.
Resolving the level of ordinary skill in the pertinent art.
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 1-3, 5, 7, 9-11, 13, and 15 are rejected under 35 U.S.C. 103 as being unpatentable over WIPO Publication 2018/046737 A1 to Auricchio et al. (hereinafter Auricchio) in view of U.S. Patent No. 11,492,644 to Little et al. (hereinafter ‘Little’, effective filing date 05/24/2017), VectorBuilder (Popular Promoters. 01/28/2021. https://web.archive.org/web/20210128103228/https://en.vectorbuilder.com/resources/vector-component/promoter.html. Accessed 02/06/2026 via web.archive.org.), Taylor-Parker (Plasmids 101: Terminators and PolyA signals. 03/31/2016. Addgene Blog. https://blog.addgene.org/plasmids-101-terminators-and-polya-signals. Accessed 02/06/2026.), NovoPro. (NovoPro. Commonly used leader peptide sequences for mammalian cells expression. 2018-04-21. https://www.novoprolabs.com/support/articles/commonly-used-leader-peptide-sequences-for-efficient-secretion-of-a-recombinant-protein-expressed-in-mammalian-cells-201804211337.html. Accessed 02/05/2026.) and Lalonde & Durocher (Therapeutic glycoprotein production in mammalian cells. Journal of Biotechnology 251 (2017) 128-140.).
Regarding claim 1:
Auricchio teaches a treatment of mucopolysaccharidosis type VI which comprises administration of a arylsulfatase B (ARSB) gene therapy (Abstract). The gene therapy vector is an isolated polynucleotide comprising a first expression cassette configured to express a mammalian precursor protein via a first promoter sequence and first polyA sequence operably linked to a coding sequence for the mammalian precursor ARSB protein, wherein the cassette is flanked by a pair of ITRs:
an "AAV vector" refers to nucleic acids, either single- stranded or double- stranded, having an AAV 5' inverted terminal repeat (ITR) sequence and an AAV 3' ITR flanking a polynucleotide encoding ARSB operably linked to transcription regulatory elements that are heterologous to the AAV viral genome, i.e., one or more promoters and/or enhancers and, optionally, a polyadenylation sequence (p. 9 ln 27-31)
AA V2/8.TBG.hARSB, which encodes human ARSB (hARSB) under the control of the liver-specific thyroxine-binding globulin (TBG) promoter (p. 35 ln 8-10)
Please see also e.g., SEQ ID NO: 8, paav2.1.TBG-hARSHB, which comprises 5’ and 3’ ITRs, a TBG promoter, a hARSB coding sequence, and BGH polyA sequence (pp. 20-22)
Auricchio further teaches that the purpose of the treatment is long-term systemic release (i.e.,secretion from the producing cell) of the therapeutic ARSB enzyme to treat MPS VI (p. 2 ln 21-22, 24-25, 26-27).
Auricchio does not teach that the first promoter sequence is substantially identical to SEQ ID NO: 16, or that the first polyA sequence is substantially identical to nucleotides 3175-3696 of SEQ ID NO: 15 (see the rejection under 112(a) above regarding nucleotides 3175-3696).
Little teaches an isolate nucleic acid (a transposase plasmid) with a first promoter that is identical to SEQ ID NO: 16 (SEQ ID NO: 12):
GenCore version 6.5.2
Copyright (c) 1993 - 2026 Biocceleration Ltd.
OM nucleic - nucleic search, using sw model
Run on: February 6, 2026, 14:42:11 ; Search time 1 Seconds
(without alignments)
21.583 Million cell updates/sec
Title: US-18-276-010-16
Perfect score: 1179
Sequence: 1 ggctccggtgcccgtcagtg..........ttccatttcaggtgtcgtga 1179
Scoring table: IDENTITY_NUC
Gapop 10.0 , Gapext 1.0
Searched: 1 seqs, 9153 residues
Total number of hits satisfying chosen parameters: 2
Minimum DB seq length: 0
Maximum DB seq length: inf
Post-processing: Minimum Match 0%
Maximum Match 100%
Listing first 2 summaries
Database : SEQIDNO_12_PTMSES.seq:*
SUMMARIES
%
Result Query
No. Score Match Length DB ID Description
----------------------------------------------------------------------------
1 1179 100.0 9153 1 SEQIDNO_12_PTMSES
c 2 23 2.0 9153 1 SEQIDNO_12_PTMSES
ALIGNMENTS
RESULT 1
SEQIDNO_12_PTMSES
Query Match 100.0%; Score 1179; DB 1; Length 9153;
Best Local Similarity 100.0%;
Matches 1179; Conservative 0; Mismatches 0; Indels 0; Gaps 0;
Qy 1 GGCTCCGGTGCCCGTCAGTGGGCAGAGCGCACATCGCCCACAGTCCCCGAGAAGTTGGGG 60
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 337 GGCTCCGGTGCCCGTCAGTGGGCAGAGCGCACATCGCCCACAGTCCCCGAGAAGTTGGGG 396
Qy 61 GGAGGGGTCGGCAATTGAACCGGTGCCTAGAGAAGGTGGCGCGGGGTAAACTGGGAAAGT 120
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 397 GGAGGGGTCGGCAATTGAACCGGTGCCTAGAGAAGGTGGCGCGGGGTAAACTGGGAAAGT 456
Qy 121 GATGTCGTGTACTGGCTCCGCCTTTTTCCCGAGGGTGGGGGAGAACCGTATATAAGTGCA 180
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 457 GATGTCGTGTACTGGCTCCGCCTTTTTCCCGAGGGTGGGGGAGAACCGTATATAAGTGCA 516
Qy 181 GTAGTCGCCGTGAACGTTCTTTTTCGCAACGGGTTTGCCGCCAGAACACAGGTAAGTGCC 240
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 517 GTAGTCGCCGTGAACGTTCTTTTTCGCAACGGGTTTGCCGCCAGAACACAGGTAAGTGCC 576
Qy 241 GTGTGTGGTTCCCGCGGGCCTGGCCTCTTTACGGGTTATGGCCCTTGCGTGCCTTGAATT 300
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 577 GTGTGTGGTTCCCGCGGGCCTGGCCTCTTTACGGGTTATGGCCCTTGCGTGCCTTGAATT 636
Qy 301 ACTTCCACCTGGCTGCAGTACGTGATTCTTGATCCCGAGCTTCGGGTTGGAAGTGGGTGG 360
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 637 ACTTCCACCTGGCTGCAGTACGTGATTCTTGATCCCGAGCTTCGGGTTGGAAGTGGGTGG 696
Qy 361 GAGAGTTCGAGGCCTTGCGCTTAAGGAGCCCCTTCGCCTCGTGCTTGAGTTGAGGCCTGG 420
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 697 GAGAGTTCGAGGCCTTGCGCTTAAGGAGCCCCTTCGCCTCGTGCTTGAGTTGAGGCCTGG 756
Qy 421 CCTGGGCGCTGGGGCCGCCGCGTGCGAATCTGGTGGCACCTTCGCGCCTGTCTCGCTGCT 480
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 757 CCTGGGCGCTGGGGCCGCCGCGTGCGAATCTGGTGGCACCTTCGCGCCTGTCTCGCTGCT 816
Qy 481 TTCGATAAGTCTCTAGCCATTTAAAATTTTTGATGACCTGCTGCGACGCTTTTTTTCTGG 540
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 817 TTCGATAAGTCTCTAGCCATTTAAAATTTTTGATGACCTGCTGCGACGCTTTTTTTCTGG 876
Qy 541 CAAGATAGTCTTGTAAATGCGGGCCAAGATCTGCACACTGGTATTTCGGTTTTTGGGGCC 600
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 877 CAAGATAGTCTTGTAAATGCGGGCCAAGATCTGCACACTGGTATTTCGGTTTTTGGGGCC 936
Qy 601 GCGGGCGGCGACGGGGCCCGTGCGTCCCAGCGCACATGTTCGGCGAGGCGGGGCCTGCGA 660
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 937 GCGGGCGGCGACGGGGCCCGTGCGTCCCAGCGCACATGTTCGGCGAGGCGGGGCCTGCGA 996
Qy 661 GCGCGGCCACCGAGAATCGGACGGGGGTAGTCTCAAGCTGGCCGGCCTGCTCTGGTGCCT 720
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 997 GCGCGGCCACCGAGAATCGGACGGGGGTAGTCTCAAGCTGGCCGGCCTGCTCTGGTGCCT 1056
Qy 721 GGTCTCGCGCCGCCGTGTATCGCCCCGCCCTGGGCGGCAAGGCTGGCCCGGTCGGCACCA 780
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1057 GGTCTCGCGCCGCCGTGTATCGCCCCGCCCTGGGCGGCAAGGCTGGCCCGGTCGGCACCA 1116
Qy 781 GTTGCGTGAGCGGAAAGATGGCCGCTTCCCGGCCCTGCTGCAGGGAGCTCAAAATGGAGG 840
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1117 GTTGCGTGAGCGGAAAGATGGCCGCTTCCCGGCCCTGCTGCAGGGAGCTCAAAATGGAGG 1176
Qy 841 ACGCGGCGCTCGGGAGAGCGGGCGGGTGAGTCACCCACACAAAGGAAAAGGGCCTTTCCG 900
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1177 ACGCGGCGCTCGGGAGAGCGGGCGGGTGAGTCACCCACACAAAGGAAAAGGGCCTTTCCG 1236
Qy 901 TCCTCAGCCGTCGCTTCATGTGACTCCACGGAGTACCGGGCGCCGTCCAGGCACCTCGAT 960
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1237 TCCTCAGCCGTCGCTTCATGTGACTCCACGGAGTACCGGGCGCCGTCCAGGCACCTCGAT 1296
Qy 961 TAGTTCTCGAGCTTTTGGAGTACGTCGTCTTTAGGTTGGGGGGAGGGGTTTTATGCGATG 1020
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1297 TAGTTCTCGAGCTTTTGGAGTACGTCGTCTTTAGGTTGGGGGGAGGGGTTTTATGCGATG 1356
Qy 1021 GAGTTTCCCCACACTGAGTGGGTGGAGACTGAAGTTAGGCCAGCTTGGCACTTGATGTAA 1080
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1357 GAGTTTCCCCACACTGAGTGGGTGGAGACTGAAGTTAGGCCAGCTTGGCACTTGATGTAA 1416
Qy 1081 TTCTCCTTGGAATTTGCCCTTTTTGAGTTTGGATCTTGGTTCATTCTCAAGCCTCAGACA 1140
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1417 TTCTCCTTGGAATTTGCCCTTTTTGAGTTTGGATCTTGGTTCATTCTCAAGCCTCAGACA 1476
Qy 1141 GTGGTTCAAAGTTTTTTTCTTCCATTTCAGGTGTCGTGA 1179
|||||||||||||||||||||||||||||||||||||||
Db 1477 GTGGTTCAAAGTTTTTTTCTTCCATTTCAGGTGTCGTGA 1515
Little further teaches that a plasmid with a polyA sequence identical to nucleotides 3175-3696 of SEQ ID NO: 15 (SEQ ID NO: 20, rabbit beta globin polyA, col 49-50):
GenCore version 6.5.2
Copyright (c) 1993 - 2026 Biocceleration Ltd.
OM nucleic - nucleic search, using sw model
Run on: February 6, 2026, 15:13:15 ; Search time 1 Seconds
(without alignments)
6.623 Million cell updates/sec
Title: US-18-276-010-15
Perfect score: 6344
Sequence: 1 ttaaccctagaaagatagtc..........gattatctttctagggttaa 6344
Scoring table: IDENTITY_NUC
Gapop 10.0 , Gapext 1.0
Searched: 1 seqs, 522 residues
Total number of hits satisfying chosen parameters: 2
Minimum DB seq length: 0
Maximum DB seq length: inf
Post-processing: Minimum Match 0%
Maximum Match 100%
Listing first 2 summaries
Database : SEQIDNO_20_RBGPOLYA.seq:*
SUMMARIES
%
Result Query
No. Score Match Length DB ID Description
----------------------------------------------------------------------------
1 522 8.2 522 1 SEQIDNO_20_RBGPOLYA
c 2 20 0.3 522 1 SEQIDNO_20_RBGPOLYA
ALIGNMENTS
RESULT 1
SEQIDNO_20_RBGPOLYA
Query Match 8.2%; Score 522; DB 1; Length 522;
Best Local Similarity 100.0%;
Matches 522; Conservative 0; Mismatches 0; Indels 0; Gaps 0;
Qy 3175 TCCTCAGGTGCAGGCTGCCTATCAGAAGGTGGTGGCTGGTGTGGCCAATGCCCTGGCTCA 3234
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1 TCCTCAGGTGCAGGCTGCCTATCAGAAGGTGGTGGCTGGTGTGGCCAATGCCCTGGCTCA 60
Qy 3235 CAAATACCACTGAGATCTTTTTCCCTCTGCCAAAAATTATGGGGACATCATGAAGCCCCT 3294
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 61 CAAATACCACTGAGATCTTTTTCCCTCTGCCAAAAATTATGGGGACATCATGAAGCCCCT 120
Qy 3295 TGAGCATCTGACTTCTGGCTAATAAAGGAAATTTATTTTCATTGCAATAGTGTGTTGGAA 3354
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 121 TGAGCATCTGACTTCTGGCTAATAAAGGAAATTTATTTTCATTGCAATAGTGTGTTGGAA 180
Qy 3355 TTTTTTGTGTCTCTCACTCGGAAGGACATATGGGAGGGCAAATCATTTAAAACATCAGAA 3414
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 181 TTTTTTGTGTCTCTCACTCGGAAGGACATATGGGAGGGCAAATCATTTAAAACATCAGAA 240
Qy 3415 TGAGTATTTGGTTTAGAGTTTGGCAACATATGCCCATATGCTGGCTGCCATGAACAAAGG 3474
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 241 TGAGTATTTGGTTTAGAGTTTGGCAACATATGCCCATATGCTGGCTGCCATGAACAAAGG 300
Qy 3475 TTGGCTATAAAGAGGTCATCAGTATATGAAACAGCCCCCTGCTGTCCATTCCTTATTCCA 3534
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 301 TTGGCTATAAAGAGGTCATCAGTATATGAAACAGCCCCCTGCTGTCCATTCCTTATTCCA 360
Qy 3535 TAGAAAAGCCTTGACTTGAGGTTAGATTTTTTTTATATTTTGTTTTGTGTTATTTTTTTC 3594
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 361 TAGAAAAGCCTTGACTTGAGGTTAGATTTTTTTTATATTTTGTTTTGTGTTATTTTTTTC 420
Qy 3595 TTTAACATCCCTAAAATTTTCCTTACATGTTTTACTAGCCAGATTTTTCCTCCTCTCCTG 3654
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 421 TTTAACATCCCTAAAATTTTCCTTACATGTTTTACTAGCCAGATTTTTCCTCCTCTCCTG 480
Qy 3655 ACTACTCCCAGTCATAGCTGTCCCTCTTCTCTTATGGAGATC 3696
||||||||||||||||||||||||||||||||||||||||||
Db 481 ACTACTCCCAGTCATAGCTGTCCCTCTTCTCTTATGGAGATC 522
Little also provides a teaching, suggestion or motivation to use a transposase plasmid for gene therapy instead of an AAV vector, as taught by Auricchio, by stating, “PiggyBac transposons offer several advantages over traditional gene delivery systems, including a large cargo capacity…multiplexed gene delivery…flexibility of target cell type…suitability for in vivo applications…and ability to be excised from the genome” (col 28 ln 12-19).
While Little does not specifically discuss their rationale for using an EF1A promoter and polyA sequence, the prior art shows that these were commonly known, commonly used, and effective regulatory elements for plasmid vectors. VectorBuilder teaches that the EF1A promoter was considered a “strong promoter” (p. 1), and AddGene teaches that the rabbit beta globin polyA sequence, together with the SV40 late polyA sequence, “are thought to be more efficient in terminating transcription due to the presence of additional helper sequences”. Therefore, the prior art provides a teaching, suggestion or motivation to try using those the regulatory sequences in an expression cassette.
The above references do not teach a heterologous signal peptide linked to the N-terminus of the ARSB amino acid sequence, nor do they teach a first and/or second cassette comprising a coding sequence for a sialytransferase (ST) protein operably linked to a promoter and polyA sequence, either in the first (bicistronic) or a second expression cassette.
NovoPro teaches a variety of commonly used leader peptide sequences for mammalian cell expression (title). Novopro further teaches that any of these leader peptide sequences are appropriate for use in a recombinant protein, for efficient secretion of the protein expressed from mammalian cells (p. 1). As noted above, Auricchio teaches a gene therapy treatment for MPS VI whose goal is the long-term systemic release of ARSB from edited cells. Based on Auricchio’s disclosures, the ordinary artisan would have been motivated to add a leader peptide to the expressed ARSB protein to facilitate its secretion from the cell for systemic distribution.
The above references do not teach a sialytransferase protein expressed from the plasmid.
Lalonde & Durocher teach that “addition of terminal sialic acids on the glycans of protein therapeutics helps to maintain them into the blood circulation” and “it has become attractive for the industry to produce proteins with optimal sialylation, as it could generate substantial savings and confer a therapeutical advantage for patients” (p. 134 § Increasing sialylation). They further teach that transient expression of different sialylation enzymes, including ST3GALII, ST3GALIV, and ST6GAL1, enhanced sialylation in mammalian cells (Id.). Based on these teachings, the ordinary artisan would have been motivated to co-express a sialytransferase with the ARSB protein to help maintain the protein in blood circulation, per Auricchio’s stated goal of systemic distribution.
It would have been prima facie obvious to a person having ordinary skill in the art before the effective filing date of the claimed invention to have combined the teachings of the above references to produce an isolated nucleic acid comprising an ARSB coding sequence to generate genetically edited cells capable of producing and releasing systemic ARSB to treat MPS VI, as taught by Auricchio. Knowing that Auricchio’s model relies on effective secretion of ARSB to achieve systemic distribution, the ordinary artisan would have recognized the benefits of fusing the enzyme to one of the leader sequences commonly known in the art for such a purpose, as taught by NovoPro. Similarly, knowing that Auricchio’s model requires systemic distribution of ARSB, the ordinary artisan would have turned to co-expression of a sialytransferase, as taught by Lalonde & Durocher, in the same cells, to better maintain the secreted enzyme in the blood circulation. Given that this approach requires expression of multiple transgenes, the ordinary artisan would have turned to Little’s transposon plasmid for multiplexed delivery of both ARSB and the ST transgenes, as taught by Little. Regarding the regulatory elements of the vector, as taught by VectorBuilder and Taylor-Parker, the EFA1 promoter and rabbit beta globin polyA were well known and commonly used regulatory elements for mammalian expression vectors. Their advantages – strong and efficient expression of a transgene – were widely recognized in the art and had already been used by Little in their expression plasmids, showing that they were suitable for that application. If the use of those well-known regulatory sequences in a mammalian expression vector, leads to success, it is likely the product not of innovation but of ordinary skill and common sense to provide routine optimization.
Regarding claim 2, NovoPro teaches SEQ ID NO: 9, METDTLLLWVLLLWVPGSTGD, is one of the known and commonly used leader peptide sequences for mammalian cell expression of proteins for secretion from the cell.
Regarding claim 3, Little teaches a multicistronic expression cassette where the transgenes are separated by a 2A sequence (col 28 ln 39-43)
Regarding claim 5, Lalonde & Durocher teach the expression of the recited ST proteins to maintain recombinant proteins in blood circulation, as discussed above.
Regarding claim 7:
Little teaches a plasmid which is identical to positions 1-1548 of SEQ ID NO: 15, comprising an EFA1 promoter, as discussed above and shown again here:
GenCore version 6.5.2
Copyright (c) 1993 - 2026 Biocceleration Ltd.
OM nucleic - nucleic search, using sw model
Run on: February 9, 2026, 08:57:58 ; Search time 1 Seconds
(without alignments)
116.133 Million cell updates/sec
Title: US-18-276-010-15
Perfect score: 6344
Sequence: 1 ttaaccctagaaagatagtc..........gattatctttctagggttaa 6344
Scoring table: IDENTITY_NUC
Gapop 10.0 , Gapext 1.0
Searched: 1 seqs, 9153 residues
Total number of hits satisfying chosen parameters: 2
Minimum DB seq length: 0
Maximum DB seq length: inf
Post-processing: Minimum Match 0%
Maximum Match 100%
Listing first 2 summaries
Database : LITTLE_SEQIDNO12_PLASMID_EFA1.seq:*
SUMMARIES
%
Result Query
No. Score Match Length DB ID Description
----------------------------------------------------------------------------
1 1548 24.4 9153 1 LITTLE_SEQIDNO12_PLASMID
c 2 48.8 0.8 9153 1 LITTLE_SEQIDNO12_PLASMID
ALIGNMENTS
RESULT 1
LITTLE_SEQIDNO12_PLASMID_EFA1
Query Match 24.4%; Score 1548; DB 1; Length 9153;
Best Local Similarity 100.0%;
Matches 1548; Conservative 0; Mismatches 0; Indels 0; Gaps 0;
Qy 1 TTAACCCTAGAAAGATAGTCTGCGTAAAATTGACGCATGCATTCTTGAAATATTGCTCTC 60
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1 TTAACCCTAGAAAGATAGTCTGCGTAAAATTGACGCATGCATTCTTGAAATATTGCTCTC 60
Qy 61 TCTTTCTAAATAGCGCGAATCCGTCGCTGTGCATTTAGGACATCTCAGTCGCCGCTTGGA 120
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 61 TCTTTCTAAATAGCGCGAATCCGTCGCTGTGCATTTAGGACATCTCAGTCGCCGCTTGGA 120
Qy 121 GCTCCCGTGAGGCGTGCTTGTCAATGCGGTAAGTGTCACTGATTTTGAACTATAACGACC 180
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 121 GCTCCCGTGAGGCGTGCTTGTCAATGCGGTAAGTGTCACTGATTTTGAACTATAACGACC 180
Qy 181 GCGTGAGTCAAAATGACGCATGATTATCTTTTACGTGACTTTTAAGATTTAACTCATACG 240
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 181 GCGTGAGTCAAAATGACGCATGATTATCTTTTACGTGACTTTTAAGATTTAACTCATACG 240
Qy 241 ATAATTATATTGTTATTTCATGTTCTACTTACGTGATAACTTATTATATATATATTTTCT 300
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 241 ATAATTATATTGTTATTTCATGTTCTACTTACGTGATAACTTATTATATATATATTTTCT 300
Qy 301 TGTTATAGATATCATCAACTTTGTATAGAAAAGTTGGGCTCCGGTGCCCGTCAGTGGGCA 360
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 301 TGTTATAGATATCATCAACTTTGTATAGAAAAGTTGGGCTCCGGTGCCCGTCAGTGGGCA 360
Qy 361 GAGCGCACATCGCCCACAGTCCCCGAGAAGTTGGGGGGAGGGGTCGGCAATTGAACCGGT 420
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 361 GAGCGCACATCGCCCACAGTCCCCGAGAAGTTGGGGGGAGGGGTCGGCAATTGAACCGGT 420
Qy 421 GCCTAGAGAAGGTGGCGCGGGGTAAACTGGGAAAGTGATGTCGTGTACTGGCTCCGCCTT 480
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 421 GCCTAGAGAAGGTGGCGCGGGGTAAACTGGGAAAGTGATGTCGTGTACTGGCTCCGCCTT 480
Qy 481 TTTCCCGAGGGTGGGGGAGAACCGTATATAAGTGCAGTAGTCGCCGTGAACGTTCTTTTT 540
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 481 TTTCCCGAGGGTGGGGGAGAACCGTATATAAGTGCAGTAGTCGCCGTGAACGTTCTTTTT 540
Qy 541 CGCAACGGGTTTGCCGCCAGAACACAGGTAAGTGCCGTGTGTGGTTCCCGCGGGCCTGGC 600
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 541 CGCAACGGGTTTGCCGCCAGAACACAGGTAAGTGCCGTGTGTGGTTCCCGCGGGCCTGGC 600
Qy 601 CTCTTTACGGGTTATGGCCCTTGCGTGCCTTGAATTACTTCCACCTGGCTGCAGTACGTG 660
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 601 CTCTTTACGGGTTATGGCCCTTGCGTGCCTTGAATTACTTCCACCTGGCTGCAGTACGTG 660
Qy 661 ATTCTTGATCCCGAGCTTCGGGTTGGAAGTGGGTGGGAGAGTTCGAGGCCTTGCGCTTAA 720
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 661 ATTCTTGATCCCGAGCTTCGGGTTGGAAGTGGGTGGGAGAGTTCGAGGCCTTGCGCTTAA 720
Qy 721 GGAGCCCCTTCGCCTCGTGCTTGAGTTGAGGCCTGGCCTGGGCGCTGGGGCCGCCGCGTG 780
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 721 GGAGCCCCTTCGCCTCGTGCTTGAGTTGAGGCCTGGCCTGGGCGCTGGGGCCGCCGCGTG 780
Qy 781 CGAATCTGGTGGCACCTTCGCGCCTGTCTCGCTGCTTTCGATAAGTCTCTAGCCATTTAA 840
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 781 CGAATCTGGTGGCACCTTCGCGCCTGTCTCGCTGCTTTCGATAAGTCTCTAGCCATTTAA 840
Qy 841 AATTTTTGATGACCTGCTGCGACGCTTTTTTTCTGGCAAGATAGTCTTGTAAATGCGGGC 900
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 841 AATTTTTGATGACCTGCTGCGACGCTTTTTTTCTGGCAAGATAGTCTTGTAAATGCGGGC 900
Qy 901 CAAGATCTGCACACTGGTATTTCGGTTTTTGGGGCCGCGGGCGGCGACGGGGCCCGTGCG 960
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 901 CAAGATCTGCACACTGGTATTTCGGTTTTTGGGGCCGCGGGCGGCGACGGGGCCCGTGCG 960
Qy 961 TCCCAGCGCACATGTTCGGCGAGGCGGGGCCTGCGAGCGCGGCCACCGAGAATCGGACGG 1020
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 961 TCCCAGCGCACATGTTCGGCGAGGCGGGGCCTGCGAGCGCGGCCACCGAGAATCGGACGG 1020
Qy 1021 GGGTAGTCTCAAGCTGGCCGGCCTGCTCTGGTGCCTGGTCTCGCGCCGCCGTGTATCGCC 1080
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1021 GGGTAGTCTCAAGCTGGCCGGCCTGCTCTGGTGCCTGGTCTCGCGCCGCCGTGTATCGCC 1080
Qy 1081 CCGCCCTGGGCGGCAAGGCTGGCCCGGTCGGCACCAGTTGCGTGAGCGGAAAGATGGCCG 1140
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1081 CCGCCCTGGGCGGCAAGGCTGGCCCGGTCGGCACCAGTTGCGTGAGCGGAAAGATGGCCG 1140
Qy 1141 CTTCCCGGCCCTGCTGCAGGGAGCTCAAAATGGAGGACGCGGCGCTCGGGAGAGCGGGCG 1200
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1141 CTTCCCGGCCCTGCTGCAGGGAGCTCAAAATGGAGGACGCGGCGCTCGGGAGAGCGGGCG 1200
Qy 1201 GGTGAGTCACCCACACAAAGGAAAAGGGCCTTTCCGTCCTCAGCCGTCGCTTCATGTGAC 1260
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1201 GGTGAGTCACCCACACAAAGGAAAAGGGCCTTTCCGTCCTCAGCCGTCGCTTCATGTGAC 1260
Qy 1261 TCCACGGAGTACCGGGCGCCGTCCAGGCACCTCGATTAGTTCTCGAGCTTTTGGAGTACG 1320
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1261 TCCACGGAGTACCGGGCGCCGTCCAGGCACCTCGATTAGTTCTCGAGCTTTTGGAGTACG 1320
Qy 1321 TCGTCTTTAGGTTGGGGGGAGGGGTTTTATGCGATGGAGTTTCCCCACACTGAGTGGGTG 1380
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1321 TCGTCTTTAGGTTGGGGGGAGGGGTTTTATGCGATGGAGTTTCCCCACACTGAGTGGGTG 1380
Qy 1381 GAGACTGAAGTTAGGCCAGCTTGGCACTTGATGTAATTCTCCTTGGAATTTGCCCTTTTT 1440
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1381 GAGACTGAAGTTAGGCCAGCTTGGCACTTGATGTAATTCTCCTTGGAATTTGCCCTTTTT 1440
Qy 1441 GAGTTTGGATCTTGGTTCATTCTCAAGCCTCAGACAGTGGTTCAAAGTTTTTTTCTTCCA 1500
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1441 GAGTTTGGATCTTGGTTCATTCTCAAGCCTCAGACAGTGGTTCAAAGTTTTTTTCTTCCA 1500
Qy 1501 TTTCAGGTGTCGTGACAAGTTTGTACAAAAAAGCAGGCTGCCACCATG 1548
||||||||||||||||||||||||||||||||||||||||||||||||
Db 1501 TTTCAGGTGTCGTGACAAGTTTGTACAAAAAAGCAGGCTGCCACCATG 1548
Auricchio’s SEQ ID NO: 8, the coding sequence for human ARSB, is also identical or substantially identical to positions 1549-3152 in SEQ ID NO: 15:
GenCore version 6.5.2
Copyright (c) 1993 - 2026 Biocceleration Ltd.
OM nucleic - nucleic search, using sw model
Run on: February 9, 2026, 09:04:55 ; Search time 1 Seconds
(without alignments)
82.104 Million cell updates/sec
Title: US-18-276-010-15
Perfect score: 6344
Sequence: 1 ttaaccctagaaagatagtc..........gattatctttctagggttaa 6344
Scoring table: IDENTITY_NUC
Gapop 10.0 , Gapext 1.0
Searched: 1 seqs, 6471 residues
Total number of hits satisfying chosen parameters: 2
Minimum DB seq length: 0
Maximum DB seq length: inf
Post-processing: Minimum Match 0%
Maximum Match 100%
Listing first 2 summaries
Database : AURICCHIO_SEQID8_HARSB.seq:*
SUMMARIES
%
Result Query
No. Score Match Length DB ID Description
----------------------------------------------------------------------------
1 1607.6 25.3 6471 1 AURICCHIO_SEQID8_HARSB
c 2 30 0.5 6471 1 AURICCHIO_SEQID8_HARSB
ALIGNMENTS
RESULT 1
AURICCHIO_SEQID8_HARSB
Query Match 25.3%; Score 1607.6; DB 1; Length 6471;
Best Local Similarity 99.4%;
Matches 1613; Conservative 0; Mismatches 9; Indels 0; Gaps 0;
Qy 1531 AAGCAGGCTGCCACCATGGGTCCGCGCGGCGCGGCGAGCTTGCCCCGAGGCCCCGGACCT 1590
| || | ||| |||||||||||||||||||||||||||||||||||||||||||||||
Db 1088 AGGCCCGGAGCCGCCATGGGTCCGCGCGGCGCGGCGAGCTTGCCCCGAGGCCCCGGACCT 1147
Qy 1591 CGGCGGCTGCTCCTCCCCGTCGTCCTCCCGCTGCTGCTGCTGCTGTTGTTGGCGCCGCCG 1650
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1148 CGGCGGCTGCTCCTCCCCGTCGTCCTCCCGCTGCTGCTGCTGCTGTTGTTGGCGCCGCCG 1207
Qy 1651 GGCTCGGGCGCCGGGGCCAGCCGGCCGCCCCACCTGGTCTTCTTGCTGGCAGACGACCTA 1710
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1208 GGCTCGGGCGCCGGGGCCAGCCGGCCGCCCCACCTGGTCTTCTTGCTGGCAGACGACCTA 1267
Qy 1711 GGCTGGAACGACGTCGGCTTCCACGGCTCCCGCATCCGCACGCCGCACCTGGACGCGCTG 1770
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1268 GGCTGGAACGACGTCGGCTTCCACGGCTCCCGCATCCGCACGCCGCACCTGGACGCGCTG 1327
Qy 1771 GCGGCCGGCGGGGTGCTCCTGGACAACTACTACACGCAGCCGCTGTGCACGCCGTCGCGG 1830
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1328 GCGGCCGGCGGGGTGCTCCTGGACAACTACTACACGCAGCCGCTGTGCACGCCGTCGCGG 1387
Qy 1831 AGCCAGCTGCTCACTGGCCGCTACCAGATCCGTACAGGTTTACAGCACCAAATAATCTGG 1890
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1388 AGCCAGCTGCTCACTGGCCGCTACCAGATCCGTACAGGTTTACAGCACCAAATAATCTGG 1447
Qy 1891 CCCTGTCAGCCCAGCTGTGTTCCTCTGGATGAAAAACTCCTGCCCCAGCTCCTAAAAGAA 1950
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1448 CCCTGTCAGCCCAGCTGTGTTCCTCTGGATGAAAAACTCCTGCCCCAGCTCCTAAAAGAA 1507
Qy 1951 GCAGGTTATACTACCCATATGGTCGGAAAATGGCACCTGGGAATGTACCGGAAAGAATGC 2010
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1508 GCAGGTTATACTACCCATATGGTCGGAAAATGGCACCTGGGAATGTACCGGAAAGAATGC 1567
Qy 2011 CTTCCAACCCGCCGAGGATTTGATACCTACTTTGGATATCTCCTGGGTAGTGAAGATTAT 2070
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1568 CTTCCAACCCGCCGAGGATTTGATACCTACTTTGGATATCTCCTGGGTAGTGAAGATTAT 1627
Qy 2071 TATTCCCATGAACGCTGTACATTAATTGACGCTCTGAATGTCACACGATGTGCTCTTGAT 2130
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1628 TATTCCCATGAACGCTGTACATTAATTGACGCTCTGAATGTCACACGATGTGCTCTTGAT 1687
Qy 2131 TTTCGAGATGGCGAAGAAGTTGCAACAGGATATAAAAATATGTATTCAACAAACATATTC 2190
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1688 TTTCGAGATGGCGAAGAAGTTGCAACAGGATATAAAAATATGTATTCAACAAACATATTC 1747
Qy 2191 ACCAAAAGGGCTATAGCCCTCATAACTAACCATCCACCAGAGAAGCCTCTGTTTCTCTAC 2250
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1748 ACCAAAAGGGCTATAGCCCTCATAACTAACCATCCACCAGAGAAGCCTCTGTTTCTCTAC 1807
Qy 2251 CTTGCTCTCCAGTCTGTGCATGAGCCCCTTCAGGTCCCTGAGGAATACTTGAAGCCATAT 2310
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1808 CTTGCTCTCCAGTCTGTGCATGAGCCCCTTCAGGTCCCTGAGGAATACTTGAAGCCATAT 1867
Qy 2311 GACTTTATCCAAGACAAGAACAGGCATCACTATGCAGGAATGGTGTCCCTTATGGATGAA 2370
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1868 GACTTTATCCAAGACAAGAACAGGCATCACTATGCAGGAATGGTGTCCCTTATGGATGAA 1927
Qy 2371 GCAGTAGGAAATGTCACTGCAGCTTTAAAAAGCAGTGGGCTCTGGAACAACACGGTGTTC 2430
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1928 GCAGTAGGAAATGTCACTGCAGCTTTAAAAAGCAGTGGGCTCTGGAACAACACGGTGTTC 1987
Qy 2431 ATCTTTTCTACAGATAACGGAGGGCAGACTTTGGCAGGGGGTAATAACTGGCCCCTTCGA 2490
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1988 ATCTTTTCTACAGATAACGGAGGGCAGACTTTGGCAGGGGGTAATAACTGGCCCCTTCGA 2047
Qy 2491 GGAAGAAAATGGAGCCTGTGGGAAGGAGGCGTCCGAGGGGTGGGCTTTGTGGCAAGCCCC 2550
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 2048 GGAAGAAAATGGAGCCTGTGGGAAGGAGGCGTCCGAGGGGTGGGCTTTGTGGCAAGCCCC 2107
Qy 2551 TTGCTGAAGCAGAAGGGCGTGAAGAACCGGGAGCTCATCCACATCTCTGACTGGCTGCCA 2610
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 2108 TTGCTGAAGCAGAAGGGCGTGAAGAACCGGGAGCTCATCCACATCTCTGACTGGCTGCCA 2167
Qy 2611 ACACTCGTGAAGCTGGCCAGGGGACACACCAATGGCACAAAGCCTCTGGATGGCTTCGAC 2670
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 2168 ACACTCGTGAAGCTGGCCAGGGGACACACCAATGGCACAAAGCCTCTGGATGGCTTCGAC 2227
Qy 2671 GTGTGGAAAACCATCAGTGAAGGAAGCCCATCCCCCAGAATTGAGCTGCTGCATAATATT 2730
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 2228 GTGTGGAAAACCATCAGTGAAGGAAGCCCATCCCCCAGAATTGAGCTGCTGCATAATATT 2287
Qy 2731 GACCCGAACTTCGTGGACTCTTCACCGTGTCCCAGGAACAGCATGGCTCCAGCAAAGGAT 2790
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 2288 GACCCGAACTTCGTGGACTCTTCACCGTGTCCCAGGAACAGCATGGCTCCAGCAAAGGAT 2347
Qy 2791 GACTCTTCTCTTCCAGAATATTCAGCCTTTAACACATCTGTCCATGCTGCAATTAGACAT 2850
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 2348 GACTCTTCTCTTCCAGAATATTCAGCCTTTAACACATCTGTCCATGCTGCAATTAGACAT 2407
Qy 2851 GGAAATTGGAAACTCCTCACGGGCTACCCAGGCTGTGGTTACTGGTTCCCTCCACCGTCT 2910
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 2408 GGAAATTGGAAACTCCTCACGGGCTACCCAGGCTGTGGTTACTGGTTCCCTCCACCGTCT 2467
Qy 2911 CAATACAATGTTTCTGAGATACCCTCATCAGACCCACCAACCAAGACCCTCTGGCTCTTT 2970
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 2468 CAATACAATGTTTCTGAGATACCCTCATCAGACCCACCAACCAAGACCCTCTGGCTCTTT 2527
Qy 2971 GATATTGATCGGGACCCTGAAGAAAGACATGACCTGTCCAGAGAATATCCTCACATCGTC 3030
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 2528 GATATTGATCGGGACCCTGAAGAAAGACATGACCTGTCCAGAGAATATCCTCACATCGTC 2587
Qy 3031 ACAAAGCTCCTGTCCCGCCTACAGTTCTACCATAAACACTCAGTCCCCGTGTACTTCCCT 3090
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 2588 ACAAAGCTCCTGTCCCGCCTACAGTTCTACCATAAACACTCAGTCCCCGTGTACTTCCCT 2647
Qy 3091 GCACAGGACCCCCGCTGTGATCCCAAGGCCACTGGGGTGTGGGGCCCTTGGATGTAGACC 3150
||||||||||||||||||||||||||||||||||||||||||||||||||||||||| |
Db 2648 GCACAGGACCCCCGCTGTGATCCCAAGGCCACTGGGGTGTGGGGCCCTTGGATGTAGCTC 2707
Qy 3151 CA 3152
|
Db 2708 GA 2709
Little further teaches a rabbit beta globin polyA which is identical to positions 3175-3696 of SEQ ID NO: 15, as already discussed above. In combination, the specific sequences amount to an expression cassette encoding an ARSB protein operably linked to a promoter (EFA1) and polyA sequence (rabbit beta globin) as taught by the prior art and already discussed above, which is substantially and functionally identical to positions 1-3696 of SEQ ID NO: 15. It is relevant to note that the differences between SEQ ID NO: 15 and the prior art sequences amount to 23 nucleotides at positions 3153-3174, which is a less than 1% difference in overall identity.
Regarding claim 9, Little teaches that the nucleic acid (plasmid) may be double-stranded (col 10 ln 36-37).
Regarding claim 10, Auricchio and Little teach engineered mammalian cells capable of expressing and secreting mammalian ARSB and an ST protein from an exogenous nucleotide sequence, as discussed above.
Regarding claim 11, Little teaches that the exogenous sequence may be extrachromosomal or integrated in the genome (col 14 ln 1-7).
Regarding claim 13, Auricchio and Little render obvious the first promoter which is identical to SEQ ID NO: 16.
Regarding claim 15, Little teaches that a population of cells was monoclonal, i.e., derived from an individual clone (col 20 ln 60-64).
Claims 17-20, 22-23,4 and 26-27 are rejected under 35 U.S.C. 103 as being unpatentable over Auricchio, Little, VectorBuilder, Taylor-Parker, NovoPro, and Lalonde & Durocher as applied to claims 1-3, 5, 7, 9-11, 13, and 15 above, further in view of WIPO Publication 2020/069429 A1 to Barney et al. (published 04/02/2020, effective filing date 09/28/2018, hereinafter ‘Barney’).
The applied reference has a common assignee with the instant application. Based upon the earlier effectively filed date of the reference, it constitutes prior art under 35 U.S.C. 102(a)(2).
This rejection under 35 U.S.C. 103 might be overcome by: (1) a showing under 37 CFR 1.130(a) that the subject matter disclosed in the reference was obtained directly or indirectly from the inventor or a joint inventor of this application and is thus not prior art in accordance with 35 U.S.C.102(b)(2)(A); (2) a showing under 37 CFR 1.130(b) of a prior public disclosure under 35 U.S.C. 102(b)(2)(B); or (3) a statement pursuant to 35 U.S.C. 102(b)(2)(C) establishing that, not later than the effective filing date of the claimed invention, the subject matter disclosed and the claimed invention were either owned by the same person or subject to an obligation of assignment to the same person or subject to a joint research agreement. See generally MPEP § 717.02.
Auricchio, Little, VectorBuilder, Taylor-Parker, NovoPro, and Lalonde & Durocher render obvious the composition comprising a plurality of engineered cells of claims 10 and/or 15, from which the instantly rejected claims depend.
Auricchio, Little, VectorBuilder, Taylor-Parker, NovoPro, and Lalonde & Durocher do not teach an implantable device comprising at least one cell-containing compartment which comprises the composition of claim 15 and at least one means for mitigating the FBR when the device is implanted into the subject, as recited in claim 17.
Barney teaches an implantable device comprising at least one cell-containing compartment that comprises the plurality of engineered cells (p. 1 ln 29-32, p. 2 ln 1-4, relevant to claim 17 and claim 23 in part):
a device comprising (i) at least one cell-containing compartment which comprises a polymer composition comprising a first cell-binding substance (CBS) and a plurality of cells…the device is implanted in or otherwise administered to a test subject).
Barney teaches that the device is suitable for implanting cells in a subject which are engineered to produce therapeutic agents to treat chronic and genetic diseases (Abstract), and that the device .
Barney further teaches the device comprising an afibrotic compound that mitigates the FBR when the device is implanted into a subject (p. 4 ln 26-28), which is an afibrotic compound (p. 5 ln 6-7) of generic Formula (I) (p. 5 ln 16-17, relevant to claim 19 in part), including one of the compounds shown in Table 4 (p. 4 ln 21-22), and is the specific compound 101 as shown in claims 20 and 23 (pp. 79-80, Table 4, relevant portions shown below):
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Regarding claim 18, Barney teaches that the cells are derived from APRE-19 cells, wherein the composition comprises an alginate covalently modified with GRGDSP (p. 3 ln 17-18, see also claim 10):
the cells are engineered ARPE-19 cells, the linker-RGD moiety in the CBP-polymer is GRGDSP (SEQ ID NO: 60), the polymer is an alginate (e.g., sodium alginate)
Regarding claim 19, Barney teaches that the cell-containing compartment is surrounded by a barrier compartment comprising an alginate hydrogel and a compound of Formula (I) dispersed on the outer surface (p. 4 ln 31, p. 5 ln 1-2, ln 5, ln 11-12; also relevant to claim 23):
the means comprises a barrier compartment formed of a biocompatible polymer that surrounds the cell-containing compartment…the biocompatible polymer is an alginate….an afibrotic compound is covalently attached to the exterior device surface and to a polymer within the barrier compartment…
Regarding claim 20, Barney teaches the two-compartment device comprising an alginate modified with GRGDSP and compound 101, as already discussed, and further teaches that the device is a spherical hydrogel capsule between 0.75 and 2 mm in diameter (p. 6 ln 11-13):
the hydrogel capsule of the disclosure has two compartments and is spherical in shape, has a diameter of about…0.8 mm to about 1.2 mm, or about 1.0 mm to about 1.5 mm
Regarding claim 23, Barney teaches a hydrogel capsule comprising an inner compartment which comprises a plurality of engineered cells encapsulated in a hydrogel-forming polymer, and a barrier compartment surrounding the inner compartment and comprising an alginate covalently modified with Compound 101 (see above for discussion of Compound 101):
A device of the disclosure is a 2-compartment hydrogel capsule…in which a cell-containing compartment (e.g., the inner compartment) comprising a plurality of cells…and one or more CBP-polymer(s) is surrounded by a barrier compartment…comprising an afibrotic polymer…the afibrotic compound is a compound of Formula (I) (p. 5 ln 24-30)
the first CBP-polymer is a polysaccharide or other hydrogel-forming polymer (p. 2 ln 28-29)
Regarding claim 24, Barney teaches the selected compound as already discussed, and further teaches that the concentrated of the engineered cell in the inner compartment is at least 40 million cells per ml of the first polymer composition (see claim 10).
Regarding claim 26, Barney teaches a composition comprising a plurality of the hydrogel capsules (p. 6 ln 22-29):
the present disclosure features a preparation (e.g., a composition) comprising a plurality (at least any of 3, 6, 12, 25, 50 or more) of cell-containing devices described herein. In one embodiment, one or more of the devices in the plurality is a device comprising (i) a cell-containing compartment which comprises a plurality of cells (e.g., live cells) encapsulated in a polymer composition comprising a first polymer covalently modified with a first cell-binding peptide (first CBP-polymer), wherein the cells are capable of expressing a therapeutic agent when the device is implanted into a subject; and means for mitigating the foreign body response (FBR) when the device is implanted into the subject.
Regarding claim 27, Barney teaches a method of treating a human subject comprising providing the plurality of hydrogel capsules and disposing the composition in the intraperitoneal space of the subject (p. 126, ln 18-19).
It would have been prima facie obvious to a person having ordinary skill in the art before the effective filing date of the claimed invention to have combined the teachings of Auricchio, Little, VectorBuilder, Taylor-Parker, NovoPro, and Lalonde & Durocher with those of Barney to produce an implantable device suitable for implanting the cells engineered to stably produce ARSB into a subject to treat MPS VI. As discussed above, Auricchio teaches administering engineered cells producing ARSB to a subject in need thereof to treat MPS VI. The ordinary artisan would have been motivated to place Auricchio’s engineered cells in Barney’s implant and administer it to a subject to treat MPS VI, with a reasonable expectation of success, based on Barney’s disclosures that the implant was suitable for long-term release of therapeutic proteins to treat genetic diseases such as MPS VI.
Claims 17, 22, 23, 24, 26, and 27 are rejected under 35 U.S.C. 103 as being unpatentable over Auricchio, Little, VectorBuilder, Taylor-Parker, NovoPro, and Lalonde & Durocher as applied to claims 1-3, 5, 7, 9-11, 13, and 15 above, further in view of WIPO Publication 2019/195055 A1 to Miller et al. (published 10/10/2019, hereinafter ‘Miller’).
Auricchio, Little, VectorBuilder, Taylor-Parker, NovoPro, and Lalonde & Durocher render obvious the composition comprising a plurality of engineered cells of claims 10 and/or 15, from which the instantly rejected claims depend.
Auricchio, Little, VectorBuilder, Taylor-Parker, NovoPro, and Lalonde & Durocher also render obvious a method of treating a human subject for MPS VI disease, comprising providing the plurality of engineered cells and disposing the composition in the intraperitoneal space of the subject (relevant to claim 27).
Auricchio, Little, VectorBuilder, Taylor-Parker, NovoPro, and Lalonde & Durocher do not teach an implantable device comprising at least one cell-containing compartment which comprises the composition of claim 15 and at least one means for mitigating the FBR when the device is implanted into the subject, as recited in claim 17.
Regarding claim 17, Miller teaches implants encapsulating engineered cells, wherein the device comprises a means for mitigating the FBR:
Described herein are particles comprising a first compartment, a second compartment, and a compound of Formula (I), as well as compositions and methods of making and using the same. The particles may comprise a cell capable of expressing a therapeutic agent useful for the treatment of a disease, disorder, or condition described herein. (Abstract)
the particle is capable of modulating the immune response (e.g., FBR) or the effect of an immune response (e.g., FBR) in a subject. (p. 2 ln 4-5)
the particle comprises an engineered cell (e.g., an engineered RPE cell or an engineered MSC).(p. 4 ln 8-9)
Regarding claims 22 and 26, Miller further teaches a preparation of particles (i.e., a pharmaceutical composition) comprising a plurality of particles:
the present disclosure features a preparation of a plurality of particles…the
preparation is a pharmaceutically acceptable preparation. (p. 4 ln 17, ln 22-23)
Regarding claims 23 and 24, Miller teaches a hydrogel capsule comprising an inner compartment comprising the cells encapsulated in a first hydrogel-forming polymer composition, and a compartment surrounding the inner compartment comprising an alginate covalently modified with at least of the compounds shown (see claims 2, 3, 28, 32, 67, 77:
the present disclosure features a preparation of a plurality of particles, wherein one or more of the particles in the plurality comprises: a) a first compartment; b) a second compartment; and c) a compound of Formula (I) as described herein. (p. 4 ln 17-19)
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421
609
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first and second polymer solutions that comprise a hydrogel forming polymer or a mixture of hydrogel forming polymers. In some embodiments, the polymer or mixture of polymers is modified with a compound of Formula (I). In some embodiments, the polymer is an alginate. (p. 4 ln 27-30)
The polymer in the first compartment (Inner) is an unmodified high molecular weight alginate (p. 6 ln 21-22)
the particle comprises alginate, and the compound of Formula (I) is covalently attached to some or all the monomers in the alginate. (p. 45 ln 30-31)
A polymer of a particle described herein may be modified with a compound of Formula (I) or a pharmaceutically acceptable salt thereof on one or more monomers of the polymer. The modified polymer of the particle may be present in the first (inner) compartment of the particle, the second (outer) compartment of the particle, or both the first (inner) and second (outer) compartments of the particle. In some embodiments, the modified polymer is present only in the second compartment (which includes the exterior particle surface). (p. 46 ln 14-19)
When a particle (e.g., a first compartment or second compartment therein) comprises alginate, the alginate can be chemically modified with a compound of Formula (I) using any suitable method known in the art. (p. 49 ln 1-3)
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Miller further teaches that conjugated compounds of Formula (I) in the outer compartment contributed to the afibrotic properties of the capsules (p. 144 ln 5-7), that two-compartment capsules are effective at containing cells (p. 146 ln 12-14), and that therapeutic proteins expressed by cells in the inner compartment are released from the capsules (p. 146 ln 27-29). Therefore, Miller provides a reasonable expectation that the capsules would have been effective at delivering engineered cells to provide therapeutic proteins without inducing fibrosis.
It would have been prima facie obvious to a person having ordinary skill in the art before the effective filing date of the claimed invention to have encapsulated the engineered cells as taught by Auricchio, Little, VectorBuilder, Taylor-Parker, NovoPro, and Lalonde & Durocher in a two-compartment hydrogel capsule comprising an alginate hydrogel and a compound of Formula (I) to mitigate the FBR, as taught by Miller. The ordinary artisan would have both have had a reasonable expectation of success and been motivated by Miller’s teachings that the capsules were effective at delivering engineered cells and therapeutic proteins while minimizing the FBR.
Claims 18-20 are rejected under 35 U.S.C. 103 as being unpatentable over Auricchio, Little, VectorBuilder, Taylor-Parker, NovoPro, Lalonde & Durocher and Miller, as applied to claims 1-3, 5, 7, 9-11, 13, 15, 17, 22-24 and 26-27 above, further in view of Sayyar (Sayyar et al. Cell-matrix Interactions of Factor IX (FIX)-engineered human mesenchymal stromal cells encapsulated in RGD-alginate vs. Fibrinogen-alginate microcapsules. Artificial Cells, Nanomedicine, and Biotechnology, 2014; 42: 102–109.)
Auricchio, Little, VectorBuilder, Taylor-Parker, NovoPro, Lalonde & Durocher and Miller render obvious the implantable device of claim 17, from which the instantly rejected claims depend.
Miller further teaches wherein the cells are derived from APRE-19 cells (relevant to claim 18) (see claim 57 and Table 3 on page 72, which lists exemplary cell lines which may be used in Miller’s hydrogel capsule and where they may be obtained commercially).
Miller further teaches wherein the cell-containing compartment is surrounded by a barrier compartment comprising an alginate hydrogel and a compound of Formula (I) (including the compound shown in claim 20) disposed on the outer surface of the barrier compartment, as already discussed above (relevant to claims 19 and 20).
Miller does not teach wherein the polymer composition encapsulating the cells comprises an alginate covalently modified with a GRGDSP peptide.
Sayyar teaches that an RGD-alginate (a GRGDSP peptide cross-linked to MCG alginate,p. 103 § Incorporation of fibrinogen into alginate microcapsules) significantly enhances the viability of encapsulated cells (Abstract), and was known to prolong the cells’ long-term in vitro and in vivo viability (p. 103).
It would have been prima facie obvious to a person having ordinary skill in the art before the effective filing date of the claimed invention to have combined the implantable hydrogel capsules comprising the engineered cells to produce ARSB, as taught by Auricchio, Little, VectorBuilder, Taylor-Parker, NovoPro, Lalonde & Durocher and Miller, with the RGD-alginate as taught by Sayyar. The ordinary artisan would have been motivated to do so and would have had a reasonable expectation of success based on Sayyar’s teachings that RGD-alginates enhance the viability and long-term survival of encapsulated cells.
Allowable Subject Matter
The prior art does not teach or suggest isolated polynucleotides comprising SEQ ID NOs: 15, 18, 19, or 20 in their entirety, with 100% identity, as required by claim 8.
As discussed above, the prior art (Auricchio, Little) discloses elements of e.g., SEQ ID NO: 15, such as the ITR sequences, ARSB coding sequence, EFA1 promoter, rabbit beta globin polyA sequence, etc. However, these known sequences do not span the entirety of SEQ ID NO: 15 with 100% identity. For example, the prior art does not teach the span of SEQ ID NO: 15 between positions 3152 and 3175. Additionally, while the prior art teaches the inclusion of a sequence encoding a sialytransferase, it does not teach the entirety of positions 3175-6344 of SEQ ID NO: 15 with 100% identity. GenBank Accession No. LT726799.1 teaches an expression vector comprising a CMV promoter and EGFP coding sequence at positions 372-1700, which match positions 3735-5057 of SEQ ID NO: 15 with 97.5% identity:
RESULT 43
LT726799
LOCUS LT726799 6448 bp DNA circular SYN 06-FEB-2017
DEFINITION Vector pGFP-N-GW, complete sequence.
ACCESSION LT726799
VERSION LT726799.1
KEYWORDS .
SOURCE Vector pGFP-N-GW
ORGANISM Vector pGFP-N-GW
other sequences; artificial sequences; vectors.
REFERENCE 1
AUTHORS De Schamphelaire,W., Olbrechts,A., Meert,J., Verhelst,K., Roggeman
Fonseca,M., Vanhoucke,M. and Beyaert,R.
TITLE BCCM/LMBP Plasmid collection
JOURNAL Unpublished
REFERENCE 2 (bases 1 to 6448)
AUTHORS De Schamphelaire,W.
TITLE Direct Submission
JOURNAL Submitted (02-FEB-2017) BCCM/LMBP, Universiteit Gent,
Technologiepark 927, 9052, BELGIUM
FEATURES Location/Qualifiers
source 1..6448
/organism="Vector pGFP-N-GW"
/mol_type="other DNA"
/db_xref="taxon:1945317"
/note="BCCM/LMBP Plasmid collection (Ghent
University,Belgium) accession number LMBP 8367."
rep_origin 39..365
/note="pMB1 ori"
regulatory 372..939
/regulatory_class="promoter"
/note="hCMV-IE promoter and enhancer"
CDS 984..>1700
/note="unnamed protein product; EGFP"
/codon_start=1
/transl_table=11
/protein_id="SJL86518.1"
/translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLT
LKFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFK
DDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKN
GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH
MVLLEFVTAAGITLGMDELYK"
regulatory complement(1719..1843)
/regulatory_class="attenuator"
/note="attR1"
regulatory 1851..1903
/regulatory_class="promoter"
/note="lacUV5 promoter"
CDS 1952..2611
/note="unnamed protein product; chloramphenicol
resistance"
/codon_start=1
/transl_table=11
/protein_id="SJL86519.1"
/translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAF
LKTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETF
SSLWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNV
ANMDNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGG
A"
CDS 2953..3258
/note="unnamed protein product; ccdB"
/codon_start=1
/transl_table=11
/protein_id="SJL86520.1"
/translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSD
KVSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
regulatory 3299..3423
/regulatory_class="attenuator"
/note="attR2"
regulatory 3585..3681
/regulatory_class="terminator"
/note="SV40 polyA early"
regulatory complement(3585..3681)
/regulatory_class="terminator"
/note="SV40 polyA late"
rep_origin complement(3735..4190)
/note="f1 ori"
regulatory 4191..4286
/regulatory_class="promoter"
/note="ampicillin resistance promoter"
misc_feature 4287..4321
/note="5' UTR of amp, incl. RBS"
regulatory 4342..4623
/regulatory_class="promoter"
/note="SV40 early promoter"
rep_origin 4580..4655
/note="SV40 ori"
CDS 4715..5509
/note="unnamed protein product; Tn5 neomycin resistance"
/codon_start=1
/transl_table=11
/protein_id="SJL86521.1"
/translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGR
PVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDL
LSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
EHQGLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRY
QDIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
regulatory 5700..5769
/regulatory_class="terminator"
/note="HSV-TK polyA
Query Match 20.0%; Score 1265.8; Length 6448;
Best Local Similarity 97.5%;
Matches 1301; Conservative 0; Mismatches 22; Indels 12; Gaps 1;
Qy 3735 ATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTCATAGCCCATATATGGA 3794
|| | ||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 366 ATGCATTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTCATAGCCCATATATGGA 425
Qy 3795 GTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCGCCCAACGACCCCCG 3854
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 426 GTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCGCCCAACGACCCCCG 485
Qy 3855 CCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGGGACTTTCCATTG 3914
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 486 CCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGGGACTTTCCATTG 545
Qy 3915 ACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATCAAGTGTATCA 3974
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 546 ACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATCAAGTGTATCA 605
Qy 3975 TATGCCAAGTACGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGGCATTATGC 4034
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 606 TATGCCAAGTACGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGGCATTATGC 665
Qy 4035 CCAGTACATGACCTTATGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTCATCGC 4094
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 666 CCAGTACATGACCTTATGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTCATCGC 725
Qy 4095 TATTACCATGGTGATGCGGTTTTGGCAGTACATCAATGGGCGTGGATAGCGGTTTGACTC 4154
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 726 TATTACCATGGTGATGCGGTTTTGGCAGTACATCAATGGGCGTGGATAGCGGTTTGACTC 785
Qy 4155 ACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAAA 4214
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 786 ACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAAA 845
Qy 4215 TCAACGGGACTTTCCAAAATGTCGTAACAACTCCGCCCCATTGACGCAAATGGGCGGTAG 4274
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 846 TCAACGGGACTTTCCAAAATGTCGTAACAACTCCGCCCCATTGACGCAAATGGGCGGTAG 905
Qy 4275 GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCTCTGGCTAACTAGAGA--------- 4325
|||||||||||||||||||||||||||||||||| | | | | |
Db 906 GCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTGGTTTAGTGAACCGTCAGATCCGCTA 965
Qy 4326 ---ACCCACTGCGCCACCATGGTGAGCAAGGGCGAGGAGCTGTTCACCGGGGTGGTGCCC 4382
| |||||||||||||||||||||||||||||||||||||||||||||||||
Db 966 GCGCTACCGGTCGCCACCATGGTGAGCAAGGGCGAGGAGCTGTTCACCGGGGTGGTGCCC 1025
Qy 4383 ATCCTGGTCGAGCTGGACGGCGACGTAAACGGCCACAAGTTCAGCGTGTCCGGCGAGGGC 4442
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1026 ATCCTGGTCGAGCTGGACGGCGACGTAAACGGCCACAAGTTCAGCGTGTCCGGCGAGGGC 1085
Qy 4443 GAGGGCGATGCCACCTACGGCAAGCTGACCCTGAAGTTCATCTGCACCACCGGCAAGCTG 4502
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1086 GAGGGCGATGCCACCTACGGCAAGCTGACCCTGAAGTTCATCTGCACCACCGGCAAGCTG 1145
Qy 4503 CCCGTGCCCTGGCCCACCCTCGTGACCACCCTGACCTACGGCGTGCAGTGCTTCAGCCGC 4562
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1146 CCCGTGCCCTGGCCCACCCTCGTGACCACCCTGACCTACGGCGTGCAGTGCTTCAGCCGC 1205
Qy 4563 TACCCCGACCACATGAAGCAGCACGACTTCTTCAAGTCCGCCATGCCCGAAGGCTACGTC 4622
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1206 TACCCCGACCACATGAAGCAGCACGACTTCTTCAAGTCCGCCATGCCCGAAGGCTACGTC 1265
Qy 4623 CAGGAGCGCACCATCTTCTTCAAGGACGACGGCAACTACAAGACCCGCGCCGAGGTGAAG 4682
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1266 CAGGAGCGCACCATCTTCTTCAAGGACGACGGCAACTACAAGACCCGCGCCGAGGTGAAG 1325
Qy 4683 TTCGAGGGCGACACCCTGGTGAACCGCATCGAGCTGAAGGGCATCGACTTCAAGGAGGAC 4742
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1326 TTCGAGGGCGACACCCTGGTGAACCGCATCGAGCTGAAGGGCATCGACTTCAAGGAGGAC 1385
Qy 4743 GGCAACATCCTGGGGCACAAGCTGGAGTACAACTACAACAGCCACAACGTCTATATCATG 4802
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1386 GGCAACATCCTGGGGCACAAGCTGGAGTACAACTACAACAGCCACAACGTCTATATCATG 1445
Qy 4803 GCCGACAAGCAGAAGAACGGCATCAAGGTGAACTTCAAGATCCGCCACAACATCGAGGAC 4862
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1446 GCCGACAAGCAGAAGAACGGCATCAAGGTGAACTTCAAGATCCGCCACAACATCGAGGAC 1505
Qy 4863 GGCAGCGTGCAGCTCGCCGACCACTACCAGCAGAACACCCCCATCGGCGACGGCCCCGTG 4922
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1506 GGCAGCGTGCAGCTCGCCGACCACTACCAGCAGAACACCCCCATCGGCGACGGCCCCGTG 1565
Qy 4923 CTGCTGCCCGACAACCACTACCTGAGCACCCAGTCCGCCCTGAGCAAAGACCCCAACGAG 4982
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1566 CTGCTGCCCGACAACCACTACCTGAGCACCCAGTCCGCCCTGAGCAAAGACCCCAACGAG 1625
Qy 4983 AAGCGCGATCACATGGTCCTGCTGGAGTTCGTGACCGCCGCCGGGATCACTCTCGGCATG 5042
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1626 AAGCGCGATCACATGGTCCTGCTGGAGTTCGTGACCGCCGCCGGGATCACTCTCGGCATG 1685
Qy 5043 GACGAGCTGTACAAG 5057
|||||||||||||||
Db 1686 GACGAGCTGTACAAG 1700
However, the prior art does not teach or suggest how to arrive from LT726799.1 to a sequence with 100% identity to SEQ ID NO: 15. Likewise, a thorough search and examine of SEQ ID NOs: 18, 19 and 20 yielded the same results: while the above elements are present in the claimed sequences, they are not present in the same configuration with 100% identity for the full length of the claimed sequences, and the prior art does not clearly teach or suggest how to arrive from these elements to the claimed sequences.
Conclusion
No claims are allowed at this time.
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/A.M.Z./Examiner, Art Unit 1636
/NEIL P HAMMELL/Supervisory Patent Examiner, Art Unit 1636