UNITED STATES PATENT AND TRADEMARK OFFICE
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`BEFORE THE PATENT TRIAL AND APPEAL BOARD
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`KELONIA THERAPEUTICS, INC.
`Petitioners
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`v.
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` INTERIUS BIOTHERAPEUTICS, INC.
`Patent Owner
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`U.S. Patent No. 11,767,366
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`DECLARATION OF PROFESSOR
`JOHN K. ROSE, PHD IN SUPPORT OF PETITION FOR
`POST-GRANT REVIEW OF U.S. PATENT NO. 11,767,366
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`Page 1 of 156
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`KELONIA EXHIBIT 1002
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`Declaration of Professor John K. Rose, Ph.D.
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`Patent No. 11,767,366
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`
`TABLE OF CONTENTS
`INTRODUCTION ........................................................................................... 1
`I.
`PROFESSIONAL BACKGROUND AND QUALIFICATIONS .................. 1
`II.
`III. MATERIALS REVIEWED ............................................................................ 5
`IV. PERSON OF ORDINARY SKILL IN THE ART .......................................... 6
`V.
`BACKGROUND OF TECHNOLOGY, THE ’366 PATENT AND
`PROSECUTION HISTORY, AND THE PRIOR ART .................................. 7
`A. Gene Therapy with Lentiviral Vectors .................................................. 7
`B. Vesicular Stomatitis Virus Glycoprotein .............................................. 9
`C. Making “Mutations” to Prepare Mutants of VSV-G .......................... 12
`D.
`Engineering of VSV-G Pseudotyped Lentiviral Vectors .................... 14
`E.
`Overview of the ’366 Patent ................................................................ 16
`F.
`Overview of the ’366 Patent Prosecution History .............................. 17
`G. Overview of Perkins ............................................................................ 18
`H. Overview of Perkins’ Provisional Application ................................... 19
`I.
`Overview of the RCSB PDB ................................................................ 21
`J.
`Overview of Hwang ............................................................................ 22
`VI. THE PRIOR ART DISCLOSED, TAUGHT, OR SUGGESTED
`CLAIMS 1-30 of THE ’366 PATENT .......................................................... 23
`A.
`Perkins Disclosed All of the Elements of Claims 1-3 and 10-25
`of the ’366 Patent ................................................................................ 23
`1.
`Claim 1 ...................................................................................... 24
`2.
`Claim 2 ...................................................................................... 43
`3.
`Claim 3: “The polypeptide of claim 1, wherein the
`polypeptide further comprises a mutation at position 214,
`a mutation at position 352, or a mutation at both position
`214 and position 352 as compared to SEQ ID NO: 2.” ............ 49
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`Declaration of Professor John K. Rose, Ph.D.
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`Patent No. 11,767,366
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`4.
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`5.
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`6.
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`Claim 10: “A nucleic acid molecule encoding the
`polypeptide of claim 1.” ............................................................ 55
`Claim 11: “A vector comprising the nucleic acid
`molecule of claim 10.” .............................................................. 58
`Claim 12: “A cell comprising the nucleic acid molecule
`of claim 10.” .............................................................................. 64
`Claim 13: “A viral particle comprising the polypeptide
`of claim 1.” ................................................................................ 70
`Claim 14: “The viral particle of claim 13, wherein the
`viral particle is a lentivirus comprising the polypeptide.” ........ 74
`Claim 15: “The viral particle of claim 13, wherein the
`viral particle further comprises a heterologous nucleic
`acid molecule encoding a heterologous molecule of
`interest.” .................................................................................... 77
`10. Claim 16: “The viral particle of claim 15, wherein the
`heterologous molecule of interest is an siRNA, an
`shRNA, a non-coding RNA, a polypeptide, a viral
`payload, a viral genome, or a combination thereof.” ................ 82
`11. Claim 17: “The viral particle of claim 15, wherein the
`heterologous molecule of interest is a chimeric antigen
`receptor (‘CAR’).” .................................................................... 84
`12. Claim 18: “The polypeptide of claim 1, wherein the
`polypeptide comprises an amino acid sequence that is at
`least 96% identical to the amino acid sequence of SEQ
`ID NO: 2.” ................................................................................. 87
`13. Claim 19: “The polypeptide of claim 1, wherein the
`polypeptide comprises an amino acid sequence that is at
`least 97% identical to the amino acid sequence of SEQ
`ID NO: 2.” ................................................................................. 88
`14. Claim 20: “The polypeptide of claim 1, wherein the
`polypeptide comprises an amino acid sequence that is at
`least 98% identical to the amino acid sequence of SEQ
`ID NO: 2.” ................................................................................. 88
`15. Claim 21: “The polypeptide of claim 1, wherein the
`polypeptide comprises an amino acid sequence that is at
`least 99% identical to the amino acid sequence of SEQ
`ID NO: 2.” ................................................................................. 89
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`7.
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`8.
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`9.
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`ii
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`Declaration of Professor John K. Rose, Ph.D.
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`Patent No. 11,767,366
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`2.
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`3.
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`4.
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`16. Claim 22: “The polypeptide of claim 2, wherein the
`polypeptide comprises an amino acid sequence that is at
`least 96% identical to the amino acid sequence of SEQ
`ID NO: 2.” ................................................................................. 90
`17. Claim 23: “The polypeptide of claim 2, wherein the
`polypeptide comprises an amino acid sequence that is at
`least 97% identical to the amino acid sequence of SEQ
`ID NO: 2.” ................................................................................. 91
`18. Claim 24: “The polypeptide of claim 2, wherein the
`polypeptide comprises an amino acid sequence that is at
`least 98% identical to the amino acid sequence of SEQ
`ID NO: 2.” ................................................................................. 91
`19. Claim 25: “The polypeptide of claim 2, wherein the
`polypeptide comprises an amino acid sequence that is at
`least 99% identical to the amino acid sequence of SEQ
`ID NO: 2.” ................................................................................. 92
`Perkins and Hwang Taught or Suggested All of the Elements of
`Claims 3-9 and 26-30 of the ’366 Patent ............................................ 93
`1.
`Claim 3: “The polypeptide of claim 1, wherein the
`polypeptide further comprises a mutation at position 214,
`a mutation at position 352, or a mutation at both position
`214 and position 352 as compared to SEQ ID NO: 2.” ............ 93
`Claim 4: “The polypeptide of claim 1, wherein the
`polypeptide further comprises a T214N mutation, a
`T352A mutation, or a combination thereof, as compared
`to SEQ ID NO: 2.” ..................................................................104
`Claim 5: “The polypeptide of claim 1, wherein the
`polypeptide further comprises a T214N mutation and a
`T352A mutation as compared to SEQ ID NO: 2.” .................104
`Claim 6: “The polypeptide of claim 2, wherein the
`polypeptide further comprises a mutation at position 214,
`a mutation at position 352, or a mutation at both position
`214 and position 352, as compared to SEQ ID NO: 2.” .........105
`Claim 7: “The polypeptide of claim 2, wherein the
`polypeptide further comprises a T214N mutation, a
`T352A mutation, or a combination thereof, as compared
`to SEQ ID NO: 2.” ..................................................................105
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`5.
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`iii
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`B.
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`Declaration of Professor John K. Rose, Ph.D.
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`Patent No. 11,767,366
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`6.
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`7.
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`8.
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`9.
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`Claim 8: “The polypeptide of claim 2, wherein the
`polypeptide further comprises a T214N mutation and a
`T352A mutation as compared to SEQ ID NO: 2.” .................105
`Claim 9: “The polypeptide of claim 1, wherein the
`polypeptide comprises the amino acid sequence of SEQ
`ID NO: 23.” .............................................................................106
`Claim 26: “The polypeptide of claim 2, wherein the
`polypeptide comprises the amino acid sequence of SEQ
`ID NO: 22.” .............................................................................107
`Claim 27: “A viral particle comprising the polypeptide
`of claim 9.” ..............................................................................108
`10. Claim 28: “The viral particle of claim 27, wherein the
`viral particle further comprises a nucleic acid molecule
`encoding a chimeric antigen receptor.” ..................................109
`11. Claim 29: “A viral particle comprising the polypeptide
`of claim 26.” ............................................................................110
`12. Claim 30: “The viral particle of claim 29, wherein the
`viral particle further comprises a nucleic acid molecule
`encoding a chimeric antigen receptor.” ..................................111
`Perkins and the RCSB PDB Taught or Suggested All of the
`Elements of Claims 1-3 and 10-25 of the ’366 Patent ......................111
`1.
`Claim 1 ....................................................................................112
`2.
`Claim 2 ....................................................................................134
`3.
`Claim 3: “The polypeptide of claim 1, wherein the
`polypeptide further comprises a mutation at position 214,
`a mutation at position 352, or a mutation at both position
`214 and position 352 as compared to SEQ ID NO: 2.” ..........136
`Claim 10: “A nucleic acid molecule encoding the
`polypeptide of claim 1.” ..........................................................136
`Claim 11: “A vector comprising the nucleic acid
`molecule of claim 10.” ............................................................137
`Claim 12: “A cell comprising the nucleic acid molecule
`of claim 10.” ............................................................................137
`Claim 13: “A viral particle comprising the polypeptide
`of claim 1.” ..............................................................................137
`Claim 14: “The viral particle of claim 13, wherein the
`viral particle is a lentivirus comprising the polypeptide.” ......137
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`4.
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`5.
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`6.
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`7.
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`8.
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`Declaration of Professor John K. Rose, Ph.D.
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`Patent No. 11,767,366
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`9.
`
`Claim 15: “The viral particle of claim 13, wherein the
`viral particle further comprises a heterologous nucleic
`acid molecule encoding a heterologous molecule of
`interest.” ..................................................................................138
`10. Claim 16: “The viral particle of claim 15, wherein the
`heterologous molecule of interest is an siRNA, an
`shRNA, a non-coding RNA, a polypeptide, a viral
`payload, a viral genome, or a combination thereof.” ..............138
`11. Claim 17: “The viral particle of claim 15, wherein the
`heterologous molecule of interest is a chimeric antigen
`receptor (‘CAR’).” ..................................................................138
`12. Claim 18: “The polypeptide of claim 1, wherein the
`polypeptide comprises an amino acid sequence that is at
`least 96% identical to the amino acid sequence of SEQ
`ID NO: 2.” ...............................................................................138
`13. Claim 19: “The polypeptide of claim 1, wherein the
`polypeptide comprises an amino acid sequence that is at
`least 97% identical to the amino acid sequence of SEQ
`ID NO: 2.” ...............................................................................139
`14. Claim 20: “The polypeptide of claim 1, wherein the
`polypeptide comprises an amino acid sequence that is at
`least 98% identical to the amino acid sequence of SEQ
`ID NO: 2.” ...............................................................................139
`15. Claim 21: “The polypeptide of claim 1, wherein the
`polypeptide comprises an amino acid sequence that is at
`least 99% identical to the amino acid sequence of SEQ
`ID NO: 2.” ...............................................................................139
`16. Claim 22: “The polypeptide of claim 2, wherein the
`polypeptide comprises an amino acid sequence that is at
`least 96% identical to the amino acid sequence of SEQ
`ID NO: 2.” ...............................................................................139
`17. Claim 23: “The polypeptide of claim 2, wherein the
`polypeptide comprises an amino acid sequence that is at
`least 97% identical to the amino acid sequence of SEQ
`ID NO: 2.” ...............................................................................140
`18. Claim 24: “The polypeptide of claim 2, wherein the
`polypeptide comprises an amino acid sequence that is at
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`Patent No. 11,767,366
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`least 98% identical to the amino acid sequence of SEQ
`ID NO: 2.” ...............................................................................140
`19. Claim 25: “The polypeptide of claim 2, wherein the
`polypeptide comprises an amino acid sequence that is at
`least 99% identical to the amino acid sequence of SEQ
`ID NO: 2.” ...............................................................................140
`Perkins, the RCSB PDB, and Hwang Taught or Suggested All
`of the Elements of Claims 3-9 and 26-30 of the ’366 Patent ............141
`VII. CONCLUSION ............................................................................................142
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`D.
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`I, John K. Rose, declare as follows:
`
`Declaration of Professor John K. Rose, Ph.D.
`
`Patent No. 11,767,366
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`
`I.
`
`INTRODUCTION
`1.
`I have been retained by counsel for the Petitioners as an independent
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`expert consultant in this proceeding before the United States Patent and Trademark
`
`Office, which I understand involves U.S. Patent No. 11,767,366 (“the ’366 patent”)
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`(Ex. 1001).1
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`2.
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`I am being compensated at my normal consulting rate of $800 per hour
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`for my work.
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`3. My compensation is in no way contingent on the nature of my findings,
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`the presentation of my findings in testimony, or the outcome of this or any other
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`proceeding. I have no other interest in this proceeding.
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`4.
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`I have been asked to consider whether certain references disclose or
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`suggest the features recited in claims 1-30 of the ’366 Patent. My opinions are set
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`forth below.
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`II.
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`PROFESSIONAL BACKGROUND AND QUALIFICATIONS
`5.
`I am an independent consultant. All of my opinions stated in this
`
`declaration are based on my own personal knowledge and professional judgment. In
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`1 Where appropriate, I refer to exhibits that I understand to be attached to the petition
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`for post-grant review of the ’366 patent.
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`
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`1
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`Declaration of Professor John K. Rose, Ph.D.
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`Patent No. 11,767,366
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`forming my opinions, I have relied on my knowledge and experience in the structure
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`and function of glycoproteins and the specific targeting of viral vectors.
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`6.
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`I am over 18 years of age and, if called upon to do so, I would be
`
`competent to testify as to the matters set forth herein. I understand that a copy of
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`my current curriculum vitae (Ex. 1003), which details my education and professional
`
`and academic experience, is being submitted in this proceeding. The following
`
`provides an overview of some of my experience that is relevant to the matters set
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`forth in this declaration.
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`7.
`
`I have significant experience and familiarity with the structure and
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`function of viral glycoproteins, including the vesicular stomatitis virus (VSV)
`
`glycoprotein (VSV-G). In 1981 my laboratory reported the first sequence of VSV-
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`G mRNA and protein determined from a cDNA clone. See J. Rose and C. Gallione,
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`Nucleotide Sequences of the mRNAs Encoding the Vesicular Stomatitis Virus G and
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`M Proteins Determined from cDNA Clones Containing the Complete Coding
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`Regions, 39 J. VIROLOGY 519 (1981) (EX. 1028). We then showed that cells
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`expressing VSV-G protein in the absence of other VSV proteins would fuse at low
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`pH. See R. Florkiewicz and J. Rose, A Cell Line Expressing Vesicular Stomatitis
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`Virus Glycoprotein Fuses at Low pH, 225 SCIENCE 721 (1984) (Ex. 1029). This was
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`the first direct evidence that VSV-G protein has membrane fusion activity. We later
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`Declaration of Professor John K. Rose, Ph.D.
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`Patent No. 11,767,366
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`performed numerous studies using VSV-G mutants to study the signals involved in
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`protein transport from the endoplasmic reticulum to the cell surface.
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`8.
`
`I am currently Professor Emeritus of Pathology and Senior Research
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`Scientist at Yale University School of Medicine in New Haven, CT. I have held this
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`role since 2019, and to this day operate a laboratory engaged in research on VSV.
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`9.
`
`For over thirty years my laboratory at Yale has been concerned with
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`novel approaches to vaccine development based on recombinant viruses and on
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`specific targeting of viral vectors. In particular, my laboratory has developed
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`methodology for generating recombinants of VSV starting from plasmid DNA.
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`VSV is a simple membrane-enveloped, negative-strand RNA virus that grows to
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`high titers in most animal cells. These recombinant VSVs expressing foreign viral
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`proteins induce potent cellular and humoral immune responses to the foreign
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`proteins in animals and protect from infection or disease caused by other viruses
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`such as influenza, measles, respiratory syncytial virus, SARS, and a monkey AIDS
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`virus. My laboratory has also performed research into the mechanisms by which the
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`recombinants of VSV generate such strong immune responses and in ways to
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`enhance these responses further while improving vector safety. My laboratory has
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`also developed novel priming and boosting vaccine vectors based on propagating
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`replicons of positive-strand RNA viruses.
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`3
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`Declaration of Professor John K. Rose, Ph.D.
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`Patent No. 11,767,366
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`10. Prior to my current role at Yale University School of Medicine, I was a
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`Professor of Pathology and Cell Biology at Yale School of Medicine from 1986 to
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`2019. Before that position, from 1982 to 1986, I was an Associate Professor at the
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`Salk Institute. From 1979 to 1982, I was an Assistant Professor at the Salk Institute.
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`11. Previously I was a Research Associate at the Massachusetts Institute of
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`Technology with Dr. David Baltimore from 1976 to 1978. From 1974 to 1975, I
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`was a Postdoctoral Fellow at the Massachusetts Institute of Technology in the
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`laboratories of Drs. Harvey Lodish and Dr. David Baltimore. From 1969 to 1973, I
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`was a Predoctoral trainee in the U.S. Public Health Service with Dr. Charles
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`Yanofsky at Stanford University.
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`12.
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`I was awarded a B.A in Biology from Brandeis University in 1969, and
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`a Ph.D. in Biology and Biochemical Genetics in 1973 from Stanford University.
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`13.
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`14.
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`I have published over 200 peer-reviewed papers and reviews.
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`I have served on the Editorial Board of the Journal of Virology from
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`1980 to 2011 and Virology from 1994 to 2014.
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`15.
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`I have also held various titles in professional activities at Yale. From
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`1987 to 1992 I was the Director of Graduate Studies in Experimental Pathology.
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`From 1992 to 2000 I was the Director of the Yale Medical School HIV research
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`facility. From 1994 to 1999 I was the Co-director of the Yale Graduate Program in
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`Declaration of Professor John K. Rose, Ph.D.
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`Patent No. 11,767,366
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`Microbiology. From 2005 to 2015 I was the Director of the Program in Virology
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`and Vaccine Development at Yale University.
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`16.
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`I am not an attorney and offer no legal opinions, but in the course of
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`my work, I have had experience studying and analyzing patents and patent claims
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`from the perspective of a person of ordinary skill in the art (which I define in Section
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`IV below).
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`III. MATERIALS REVIEWED
`17. The opinions in this Declaration are based on the documents I reviewed,
`
`my knowledge and experience, and professional judgment. In forming my opinions
`
`expressed in this Declaration, I have reviewed the documents and other materials
`
`referred to herein. I understand that the documents and materials I reviewed and
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`refer to herein are being submitted as exhibits attached to the petition for Post Grant
`
`Review of the ’366 patent, and are listed in Appendix A (Table of Cited Exhibits).
`
`18. My opinions have been guided by my understanding of how one of
`
`ordinary skill in the art would have understood claims 1-30 and the specification of
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`the ’366 patent at the time of the alleged invention, which I have been asked to
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`consider as of the 2021 timeframe (including December 15, 2021, the filing date of
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`U.S. Provisional Application No. 63/289,888 (“the ’888 application”) and U.S.
`
`Provisional Application No. 63/289,977 (“the ’977 patent”), which is the earliest
`
`filing date to which the ’366 patent claims priority). My opinions reflect how one
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`Declaration of Professor John K. Rose, Ph.D.
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`Patent No. 11,767,366
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`of ordinary skill in the art would have understood the ’366 patent, the prior art to
`
`that patent, and the state of the art at the time of the alleged invention.
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`19. Based on my review of the materials in view of my experience and
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`expertise, it is my opinion that the references identified below taught a mutant
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`VSV-G envelope protein with a I182E or I182D mutation, including all of the
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`elements recited in claims 1-30 of the ’366 patent, as I discuss in detail below.
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`IV. PERSON OF ORDINARY SKILL IN THE ART
`20.
`I have been asked to assume that the relevant timeframe for the alleged
`
`inventions of the ’366 patent is 2021, including the time period up to and including
`
`December 15, 2021, the filing date of the earliest applications (the ’888 and the ’977
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`applications) to which the ’366 patent claims priority (referred to herein as “the
`
`relevant timeframe”). I applied that understanding in my analysis.
`
`21.
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`I am familiar with the level of ordinary skill in the art with respect to
`
`the technology disclosed and claimed in the ’366 patent during the relevant
`
`timeframe. Based on my review of the ’366 patent, the technology, the educational
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`level and experience of active workers in the field, the types of problems faced by
`
`professionals in the field, the solutions found to those problems, the sophistication
`
`of the technology in the field, and drawing on my own experience, I believe one of
`
`ordinary skill in the art would have had an advanced degree in molecular biology,
`
`biochemistry, or an equivalent field, as well as one to three years of experience in
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`
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`Declaration of Professor John K. Rose, Ph.D.
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`Patent No. 11,767,366
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`retroviruses and the modification of envelope proteins associated with retroviruses.
`
`All of my opinions in this declaration are from the perspective of one of ordinary
`
`skill in the art as I have defined it here. My opinions expressed in this declaration
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`would be the same if this definition was altered to some extent to account for a
`
`slightly greater or lesser level of skill in the art, at least because of the clear prior art
`
`disclosures discussed herein.
`
`V. BACKGROUND OF TECHNOLOGY, THE ’366 PATENT AND
`PROSECUTION HISTORY, AND THE PRIOR ART
`A. Gene Therapy with Lentiviral Vectors
`22. By the 1990s, gene therapies involving the insertion of a DNA molecule
`
`(“a retroviral vector”) carrying a gene of interest into a cell had become appealing
`
`for the treatment of patients with immunodeficiencies or cancer. (See, e.g., Ex. 1008
`
`at 1.)2 In particular, the use of lentiviral vectors (i.e., retroviral vectors that come
`
`from lentiviruses, a type of virus that includes HIV), was known as early as 1996 to
`
`provide stable and effective delivery of genes into cells. (See, e.g., Ex. 1009 at 263;
`
`Ex. 1008 at 1.)
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`2 Unless otherwise noted, this Petition cites the original page numbering of any non-
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`patent exhibits.
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`23. By the 2020s, lentiviral vectors were known to be “among the most
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`promising vectors” for cellular therapies and, for certain cells, “the tools of choice.”
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`(See, e.g., Ex. 1010 at 1; Ex. 1008 at 1.) This stems not only from their stable and
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`effective delivery of genes into cells, also known as cell “transduction,” but also a
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`lower probability of inducing an innate immune response during cell transduction.
`
`(See, e.g., Ex. 1010 at 2.) Lentiviral vectors were known to be able to carry an
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`envelope protein that mediates cell transduction. (See, e.g., Ex. 1010 at 1-2.) The
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`nature of the envelope protein carried by a lentivirus vector “is the major determinant
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`for the specificity of transduction.” (See, e.g., Ex. 1010 at 1.)
`
`24. To expand the use of lentiviral vectors, researchers have commonly
`
`replaced the envelope protein of a lentivirus with those of other viruses, a process
`
`called “pseudotyping.” (See, e.g., Ex. 1010 at 2; Ex. 1012 at 130.) Pseudotyping
`
`lentiviral vectors with envelope proteins of different viruses allows for the properties
`
`of lentiviruses to be combined with the viral entry properties of other viruses. (See,
`
`e.g., Ex. 1010 at 7.) For instance, some envelope proteins possess a broad tropism,
`
`also called “pantropism,” which means they have the ability to infect a wide gamut
`
`of cell types. (See, e.g., Ex. 1010 at 1.) This property, however, also makes their
`
`use more challenging for in vivo treatments, where “a high specificity for the target
`
`cells is required to avoid off-target transduction.” (See, e.g., id.)
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`Declaration of Professor John K. Rose, Ph.D.
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`Patent No. 11,767,366
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`25. By the 2020s, it was known that pseudotyping lentiviral vectors by the
`
`vesicular stomatitis virus envelope glycoprotein G (“VSV-G”) was the most
`
`common approach for pseudotyping lentiviral vectors. (See, e.g., Ex. 1010 at 9.)
`
`For instance, it was reported in 2013 that “VSV-G-pseudotyped lentiviruses exhibit
`
`the same broad tropism as VSV, excellent stability, and high transduction efficiency,
`
`rendering them the gold standard for experimental gene transfer procedures.” (See,
`
`e.g., Ex. 1016 at 7307.)
`
`B. Vesicular Stomatitis Virus Glycoprotein
`26. VSV-G is a transmembrane glycoprotein (G) that is incorporated into
`
`the lipid bilayer in VSV viral particles and mediates attachment of VSV to host cells
`
`through VSV-G-host cell receptor binding, and catalyzes fusion of viral and cellular
`
`membranes to initiate infection. (See, e.g., Ex. 1011 at 291-292, 296.) A schematic
`
`illustration of a VSV viral particle is reproduced below for illustration:
`
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`Declaration of Professor John K. Rose, Ph.D.
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`Patent No. 11,767,366
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`27. The interior of VSV viral particles were known to include an “N-RNA”
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`ribonucleoprotein core associated with two viral components: the large polymerase
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`protein (“L”), and the accessory phosphoprotein (“P”), which form the replication
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`machinery of the virus, and a matrix (“M”) protein that mediates particle assembly
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`and is efficiently incorporated into virions during the process of viral assembly.
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`(See, e.g., Ex. 1011 at 291-92, 295-96, FIG. 1.) The exterior of a VSV viral particle
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`was known to contain a lipid envelope (“lipid bilayer”) decorated with trimeric
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`spikes of the VSV-G (G) protein. (See, e.g., id.)
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`28. VSV-G was known to have “a critical role during the initial steps of
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`virus infection,” including both recognition and binding of the receptor at the cell
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`surface, and subsequently in mediating “the fusion between the viral and endosomal
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`Declaration of Professor John K. Rose, Ph.D.
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`Patent No. 11,767,366
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`membranes.” (See, e.g., Ex. 1012 at 118.) In particular, after receptor binding, VSV
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`was understood to enter a cell by the endocytic pathway. (See, e.g., Ex. 1012 at 120-
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`121; Ex. 1015 at 1717.) Subsequently, “the viral envelope fuses with a cellular
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`membrane within the acidic environment of the endosome.” (See, e.g., Ex. 1015 at
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`1717.) Fusion was known to be triggered by the low pH of the endosome and
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`mediated by the VSV-G glycoprotein, with optimal fusion occurring at pH 6. (See,
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`e.g., Ex. 1015 at 1717; Ex. 1012 at 120.)
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`29. By the mid to late 2000s, crystal structures of the pre-fusion and post-
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`fusion states of VSV-G had been published. (See, e.g., Ex. 1013 at 843; Ex. 1014 at
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`187.) Armed with this structural characterization of VSV-G, researchers
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`investigated the regions of VSV-G’s structure that were responsible for the fusion
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`characteristics of VSV-G, including identifying which mutations potentially
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`impacted the fusion properties of VSV-G. (See, e.g., Ex. 1015 at 1722 (Table 1).)
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`30.
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`In 2013, the native receptor for VSV-G binding—which had until then
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`remained elusive—was determined to be the low-density lipoprotein receptor
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`(“LDL-R”). (See, e.g., Ex. 1016 at 7306, 7310.) This helped to explain the broad
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`tropism of VSV-G, as the LDL-R is a highly ubiquitous receptor found in many cell
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`types. (See, e.g., Ex. 1016 at 7306.) It also explained why VSV-G pseudotyped
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`lentiviral vectors did not provide efficient transfer of genetic materials in certain
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`Declaration of Professor John K. Rose, Ph.D.
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`Patent No. 11,767,366
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`desirable gene-therapy targets, such as quiescent T cells, as these cells had low
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`expression of LDL-R. (See, e.g., Ex. 1017 at 1422-23.)
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`C. Making “Mutations” to Prepare Mutants of VSV-G
`31. A person of ordinary skill in the art would have understood that
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`nucleotides (or “nucleic acids”)—the building blocks of genetic information—can
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`be
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`joined
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`together
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`to form polymers of nucleic acids, also known as
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`“polynucleotides.” (See, e.g., Ex. 1022 at 39-44.) A person of ordinary skill in the
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`art also would have understood that polynucleotides are nucleic acid molecules that
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`include both deoxyribonucleic acid (“DNA”) and ribonucleic acid (“RNA”). (See,
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`e.g., Ex. 1022 at 39-44.)
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`32. DNA contains genes that encode for the production of amino acid
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`polymers, or “polypeptides,” which includes proteins. (See, e.g., Ex. 1022 at 49,
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`78.) A person of ordinary skill in the art would have understood that changes to the
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`DNA sequence, also known as “mutations,” could result in proteins with altered
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`amino acids, structures, functions, and expression levels. (See, e.g, id.) This can
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`occur, for instance, when one or more altered DNA nucleotides results in an altered
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`amino acid sequence for the encoded protein. (See, e.g., id.)
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`33. Proteins are composed of 20 “standard” amino acids. (See, e.g., Ex.
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`1022 at 74-77.) A person of ordinary skill in the art was also well aware of the
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`abbreviated one-letter symbols for amino acids. (See, e.g., Ex. 1022 at 76-77, 81-
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`Declaration of Professor John K. Rose, Ph.D.
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`Patent No. 11,767,366
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`82.) For example, the amino acid “isoleucine” using its one-letter symbol is referred
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`to as “I.” (See, e.g., Ex. 1022 at 76.)
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`34. Wild-type VSV-G includes a 16 amino acid signal peptide (also
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`referred to as a “signal sequence”) that is cleaved in the endoplasmic reticulum of a
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`eukaryotic cell. (See, e.g., Ex. 1001 at 1:36-39, 17:32-39, 17:31-19:23; Ex. 1005 at
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`12:16-29, 20:15-18, 74 (SEQ ID NO: 88); Ex. 1006 at 9-10, 17, 50 (SEQ ID NO:
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`72), 51 (SEQ ID NO: 78). In the wild-type amino acid sequence of VSV-G absent
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`its signal peptide, the amino acid at position 182 is isoleucine, which in one-letter
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`code is represented by “I182.” (See, e.g., 1005 at 74-75 (SEQ ID NO: 90).) In the
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`amino acid sequence of VSV-G with its signal sequence still present, the same
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`isoleucine would be represented by I198, which is shifted forward 16 positions to
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`account for the signal peptide. (See, e.g., id.)
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`35. A person of ordinary skill