`
`
`
`
`Filed on behalf of: Sarepta Therapeutics, Inc.
`
`
`
`
`
`
`
`
`UNITED STATES PATENT AND TRADEMARK OFFICE
`______________________
`BEFORE THE PATENT TRIAL AND APPEAL BOARD
`______________________
`
`SAREPTA THERAPEUTICS, INC.
`Petitioner
`v.
`NIPPON SHINYAKU CO., LTD.
`&
`NATIONAL CENTER OF NEUROLOGY AND PSYCHIATRY
`Patent Owners
`______________________
`
`Case No. IPR2021-01136
`Patent No. 10,407,461
`______________________
`
`DECLARATION OF DR. DAVID R. COREY
`
`
`
`
`
`
`
`
`
`
`
`
`
`U.S. Patent No. 10,407,461
`
`TABLE OF CONTENTS
`INTRODUCTION ................................................................................ 1
`I.
`QUALIFICATIONS ............................................................................. 1
`II.
`III. MATERIALS CONSIDERED ............................................................. 3
`IV. SUMMARY OF MY OPINIONS ........................................................ 4
`V.
`BACKGROUND AND THE STATE OF THE ART .......................... 8
`A.
`From DNA to Protein ................................................................. 8
`B.
`Duchenne Muscular Dystrophy (DMD) ................................... 13
`C.
`Exon Skipping Antisense Oligomers (AOs) as DMD
`Therapies .................................................................................. 15
`1.
`Exon Skipping AOs ....................................................... 15
`2.
`Natural Precedents for Exon Skipping AOs in DMD .... 16
`3.
`Exon Skipping AOs in DMD ......................................... 17
`4.
`Exon 53 Targeting AOs ................................................. 25
`5.
`Known Techniques for Evaluating Exon Skipping
`AOs ................................................................................ 32
`The Asserted Prior Art References........................................... 33
`1.
`Popplewell ...................................................................... 33
`2.
`Sazani ............................................................................. 37
`VI. THE ’461 PATENT ............................................................................ 39
`A. Overview .................................................................................. 39
`B.
`The Claims of the ’461 Patent .................................................. 40
`C.
`The Disclosures of the ’461 Patent .......................................... 42
`
`D.
`
`
`
`i
`
`
`
`
`
`
`U.S. Patent No. 10,407,461
`
`AOs Disclosed in the Japanese Application .................. 42
`1.
`Additional AOs Disclosed in the ’461 Patent ................ 42
`2.
`The Disclosures of the ’461 Patent ................................ 43
`3.
`VII. LEVEL OF ORDINARY SKILL IN THE ART ................................ 44
`VIII. CLAIM 1 IS OBVIOUS OVER POPPLEWELL AND SAZANI ..... 45
`A. A POSA Would Have Been Motivated to Make and Test
`AOs of Claim 1 ......................................................................... 45
`1.
`A POSA Would Have Been Motivated to Make Exon
`53 Targeting AOs ........................................................... 45
`A POSA Would Have Been Motivated to Make and
`Test AOs Targeting the (+30+65) Hotspot Target
`Region of Exon 53 ......................................................... 46
`A POSA Would Have Been Motivated to Make and
`Test 25-mer AOs 100% Complementary to the
`(+36+60) Sequence of Exon 53 as Claimed in the
`’461 Patent ..................................................................... 50
`A POSA Would Have Been Motivated to Make and
`Test a 25-mer PMO 100% Complementary to the
`(+36+60) Sequence of Exon 53 ..................................... 53
`Popplewell Does Not Discourage AOs that Are 25
`Bases in Length .............................................................. 56
`A POSA Would Have Had a Reasonable Expectation of
`Making and Testing a 25-mer PMO 100% Complementary
`to the (+36+60) Sequence of Exon 53 ..................................... 57
`A POSA Would Have Had a Reasonable Expectation that a
`25-mer PMO 100% Complementary to the (+36+60)
`Sequence of Exon 53 Would Cause Exon 53 Skipping ........... 58
`IX. CLAIM 2 IS OBVIOUS OVER POPPLEWELL AND SAZANI ..... 60
`
`2.
`
`3.
`
`4.
`
`5.
`
`B.
`
`C.
`
`
`
`ii
`
`
`
`
`U.S. Patent No. 10,407,461
`
`
`X.
`
`THE ’461 PATENT DOES NOT EVIDENCE SUPERIORITY OF
`A CLAIMED AO OVER POPPLEWELL ......................................... 62
`XI. OTHER RESEARCHERS’ NEAR-SIMULTANEOUS ARRIVAL
`AT THE (+36+60) TARGET REGION OF EXON 53 ...................... 73
`XII. CONCLUSION ................................................................................... 74
`
`
`
`
`
`iii
`
`
`
`
`I.
`
`
`U.S. Patent No. 10,407,461
`
`INTRODUCTION
`I, David R. Corey, have been retained by Sarepta Therapeutics, Inc.
`1.
`
`(“Sarepta”) as an independent expert in the design and evaluation of antisense
`
`oligomers for therapeutic purposes.
`
`2.
`
`I am being compensated for the time I spend on this matter, but my
`
`compensation is not contingent upon my opinions or the outcome of this or any
`
`other proceeding.
`
`II. QUALIFICATIONS
`I am a Professor of Pharmacology at the University of Texas
`3.
`
`Southwestern Medical Center (UTSWMC). I also hold the Rusty Kelley
`
`Professorship in Medical Science. For over 30 years, I have investigated various
`
`technologies for altering genetic materials and their therapeutic potentials as
`
`treatments of genetic diseases. My current research focuses on designing and
`
`testing antisense oligomers, peptide nucleic acids, antigene oligomers, and double
`
`stranded RNAs/single-stranded silencing RNAs (RNA interference) for treating
`
`rare genetic diseases such as Duchenne Muscular Dystrophy, Huntington’s
`
`Disease, Machado Joseph Disease, Dentatorubral-pallidoluysian atrophy,
`
`Friedreich’s ataxia, and Fuchs corneal endothelial dystrophy.
`
`4.
`
`I received my B.A. in Chemistry in 1985 from Harvard University. I
`
`received my Ph.D. in Chemistry in 1990 from the University of California,
`
`
`
`1
`
`
`
`
`Berkeley. My dissertation research focused on methods of using oligomer
`
`
`U.S. Patent No. 10,407,461
`
`conjugates for altering genetic materials such as DNA and RNA in a sequence-
`
`specific manner.
`
`5.
`
`From 1990 to 1992, I was a post-doctoral fellow at the University of
`
`California, San Francisco. In 1992, I joined the UTSWMC as an Assistant
`
`Professor in the Department of Pharmacology. I received a second appointment
`
`from the Department of Biochemistry in 1997. I was promoted to an Associate
`
`Professor in 1998 and a Professor in 2003. In 2014, I received the Rusty Kelley
`
`Professorship in Medical Science.
`
`6.
`
`I have been on the Board of Directors of the Oligonucleotide
`
`Therapeutics Society since 2009. In this role, I have helped to organize several
`
`Annual Oligonucleotide Therapeutics Society Meetings. I was a Chair of the 10th
`
`Annual Oligonucleotide Therapeutics Society Meeting held in 2014. Currently, I
`
`am President Elect of the Oligonucleotide Therapeutics Society.
`
`7.
`
`I have been on editorial boards for several journals including the
`
`Journal of RNAi and Gene Silencing (2005–2015), Artificial DNA (2009–2016),
`
`Methods (2012–2019), and Scientific Reports (2016). Currently, I am Executive
`
`Editor of Nucleic Acids Research, a prominent journal in the field of nucleic acids
`
`and related technologies. I am also a Senior Editor of Nucleic Acid Therapeutics. I
`
`have also recently served as a reviewer for numerous journals including Nature,
`
`
`
`2
`
`
`
`
`Science, Proceedings of the National Academy of Sciences USA, RNA,
`
`
`U.S. Patent No. 10,407,461
`
`Biochemistry, Journal of the American Chemical Society, Nature Neuroscience,
`
`Nature Medicine, and Cell Reports. I have reviewed grant applications for various
`
`institutes and foundations including the National Institutes of Health, the National
`
`Cancer Institute, the National Institute of General Medical Sciences, the National
`
`Science Foundation, the Wellcome Trust, and Muscular Dystrophy Campaign.
`
`8.
`
`I have authored over 170 peer-reviewed publications related to nucleic
`
`acids and related technologies such as antisense oligomers. I have also published
`
`several book chapters on designing and making antisense oligomers, including
`
`those that are chemically modified. I am regularly invited to present my work at
`
`conferences, and I have given more than 50 invited seminars since 2014.
`
`9. My professional qualifications are described in further detail in my
`
`curriculum vitae, which is attached as Appendix A.
`
`III. MATERIALS CONSIDERED
`In forming my opinions, I have considered the materials cited in this
`10.
`
`report, as well as those listed in Appendix B. In addition to these materials, I may
`
`consider additional documents and information in forming any supplemental
`
`opinions. To the extent I am provided with additional documents or information,
`
`including any expert declarations in this proceeding, I may offer further opinions.
`
`
`
`3
`
`
`
`
`IV. SUMMARY OF MY OPINIONS
`11. The claims of the ’461 patent are directed to antisense oligomers
`
`
`U.S. Patent No. 10,407,461
`
`(“AOs”) that induce skipping of exon 53 of the human dystrophin gene and consist
`
`of a “25-mer oligomer that is 100% complementary” to the (+36+60) sequence of
`
`exon 53 of the human dystrophin pre-mRNA. The claimed AOs would have been
`
`obvious to a person of ordinary skill in the art (“POSA”) as of August 31, 2011,
`
`the date that NS filed its PCT application which contained for the first time an AO
`
`complementary to the (+36+60) sequence of exon 53. As described herein, my
`
`opinions set forth in this declaration would not change if the September 1, 2010
`
`filing date of NS’s original Japanese application is used as the date for determining
`
`prior art.
`
`12. By August 31, 2011, exon skipping AOs were recognized as a
`
`promising therapeutic approach for the treatment of Duchenne muscular dystrophy
`
`(DMD). Numerous preclinical in vitro and in vivo studies had confirmed the safety
`
`and efficacy of exon skipping for DMD. By August 31, 2011, early results from
`
`human clinical trials of two AOs targeting exon 51 of the dystrophin gene were
`
`reported as encouraging. Both treatments were reported as of August 31, 2011 to
`
`cause exon skipping and to increase dystrophin levels in DMD patients. By
`
`August 31, 2011, researchers were also investigating and developing AOs targeting
`
`
`
`4
`
`
`
`
`exon 53 that were reported to have the potential to treat a large number of DMD
`
`
`U.S. Patent No. 10,407,461
`
`patients.
`
`13. By August 31, 2011, prior art references including Popplewell had
`
`identified a superior, hotspot target region within exon 53. It had been shown that
`
`many AOs targeting this hotspot target region effectively induced exon 53 skipping
`
`in in vitro and in vivo assays. Popplewell highlighted a number of these exon 53
`
`targeting AOs as “worthy of consideration for any upcoming clinical trial.”
`
`Among the most effective prior art AOs targeting this hotspot region was a 25-mer
`
`AO targeting the (+35+59) sequence in exon 53—just one nucleotide shifted from
`
`the target region of the AOs claimed in the ’461 patent—which was singled out by
`
`Popplewell as a “viable” candidate for clinical trials.
`
`14. Oligomers are synthetic molecules. During drug development of
`
`synthetic molecules, it is common to synthesize many variant compounds that are
`
`closely related to compounds that show promising activity. The purpose is to
`
`search for compounds that might have better properties and make successful pre-
`
`clinical and clinical development more likely. By August 31, 2011, it was
`
`common knowledge that testing dozens or even hundreds of different compounds
`
`was the norm for companies pursuing nucleic-acid-based drugs.
`
`15. By August 31, 2011, based on known effective exon 53 targeting
`
`compounds, a POSA would have been motivated to use routine screening
`
`
`
`5
`
`
`
`
`techniques to identify and test additional AOs that were close chemical relatives to
`
`
`U.S. Patent No. 10,407,461
`
`such known active AOs. Using the prior art 25-mer AO targeting the (+35+59)
`
`sequence in exon 53 as a starting point, a POSA would have designed a one-
`
`nucleotide stepped array and tested the resulting AOs for their ability to induce
`
`exon 53 skipping. Using such an approach, a POSA would have immediately
`
`identified and chosen for testing an AO 100% complementary to the (+36+60)
`
`sequence of exon 53 as claimed in the ’461 patent.
`
`16. A POSA would have reasonably expected that an AO 100%
`
`complementary to the (+36+60) sequence of exon 53 would induce skipping of
`
`exon 53. The (+36+60) target sequence falls within the hotspot target region of
`
`exon 53 identified in Popplewell. The numerous effective prior art AOs (including
`
`those identified in Popplewell) with target regions throughout the hotspot target
`
`region of exon 53 (+30+65) would have provided a reasonable expectation that an
`
`AO 100% complementary to the (+36+60) sequence of exon 53 would be similarly
`
`effective in causing skipping of exon 53. The (+36+60) AO lacks any
`
`distinguishing feature relative to the previously tested AOs that would suggest that
`
`it would have a significantly different activity from those previously tested AOs.
`
`17. A POSA would have expected that the (+36+60) AO would have
`
`skipping efficacy similar to the many closely related AOs that had been tested
`
`
`
`6
`
`
`
`
`previously and would have tested the (+36+60) AO because thorough testing of
`
`
`U.S. Patent No. 10,407,461
`
`related AOs reassures that no potentially better AO has been overlooked.
`
`18. The data disclosed in the ’461 patent demonstrates how similar in
`
`efficacy the claimed AOs are to the prior art AOs. For instance, the ’461 patent
`
`specification shows that an AO 100% complementary to the (+36+60) sequence of
`
`exon 53 was no better than an AO targeting the same (+35+59) sequence of exon
`
`53 that was disclosed in the prior art. The similar skipping efficiencies for these
`
`two AOs would have been unsurprising to a POSA, given that an AO 100%
`
`complementary to the (+36+60) sequence of exon 53 targets a region shifted by
`
`just one nucleotide as compared to the prior art AO targeting the (+35+59)
`
`sequence.
`
`19. PMOs were one of two prominent classes of AOs as of August 31,
`
`2011. This class of AOs binds its targets by Watson-Crick base pairing under
`
`physiological conditions. Indeed, one of the two exon 51 targeting AOs
`
`undergoing clinical trials for DMD as of August 31, 2011 was a PMO (being
`
`developed by Sarepta) and the other was a 2’-OMePS (being developed by
`
`Prosensa, now BioMarin). By August 31, 2011, a POSA would have been aware
`
`that Sazani provides the structure of Sarepta’s exon 51 PMO (including its 5’
`
`triethylene glycol moiety) and reports on its favorable safety profile. Because an
`
`exon skipping PMO was showing promise in the clinic for the treatment of DMD,
`
`
`
`7
`
`
`
`
`a POSA making an AO for the treatment of DMD would have been motivated to
`
`
`U.S. Patent No. 10,407,461
`
`generate PMOs and would have had reasonable expectation that these AOs could
`
`be used to safely and effectively induce skipping of exon 53.
`
`20. Thus, as discussed below, it is my opinion that the challenged claims
`
`of the ’461 patent would have been obvious to a POSA as of August 31, 2011.
`
`V. BACKGROUND AND THE STATE OF THE ART
`From DNA to Protein
`A.
`21. Deoxyribonucleic acid (“DNA”) is the hereditary material in humans
`
`and other organisms. (Ex. 1068 [Alberts], 191–192.) DNA comprises smaller
`
`portions (“genes”), each of which encodes a protein that carries out various
`
`functions in the body. (Id., 301.) The process of encoding a protein from a gene
`
`involves three main steps: transcription, splicing, and translation. (Id.) Each step
`
`is explained below.
`
`22. DNA is a long polymer made up of four different building blocks
`
`(referred to as “nucleotides”). (Id., 193.) Each nucleotide comprises a five-carbon
`
`sugar (deoxyribose), a phosphate group, and one of four nitrogen-containing bases.
`
`(Id.) The base is selected from adenine (A), guanine (G), cytosine (C), and
`
`thymine (T). (Id.) The four base symbols, A, G, C, and T commonly denote the
`
`four nucleotides as well. (Id.) Figure 1 depicts the four nucleotides found in
`
`DNA:
`
`
`
`8
`
`
`
`
`U.S. Patent No. 10,407,461
`
`O
`
`N
`
`NH
`
`NH2
`
`Guanine (G)
`
`N
`
`N
`
`H
`
`O
`
`O
`
`HH
`
`H
`
`OH
`
`H
`
`-
`
`O
`
`O
`
`P
`
`-
`
`O
`
`
`
`
`
`NH
`
`O
`
`Thymine (T)
`
`O
`
`N
`
`H
`
`
`
`-
`
`O
`
`O
`
`P
`
`-
`
`O
`
`O
`
`O
`
`H
`
`OH
`
`H
`
`H
`
`H
`
`NH2
`
`N
`
`N
`
`Adenine (A)
`
`
`
`N
`
`N
`
`H
`
`NH2
`
`N
`
`N
`
`H
`
`H
`
`H
`
`O
`
`O
`
`H
`
`OH
`
`H
`
`O
`
`Cytosine (C)
`
`
`
`-
`
`O
`
`O
`
`P
`
`-
`
`O
`
`O
`
`O
`
`H
`
`OH
`
`H
`
`H
`
`H
`
`-
`
`O
`
`O
`
`P
`
`-
`
`O
`
`
`
`Figure 1. Nucleotides in DNA
`
`23.
`
`In DNA, nucleotides are covalently linked via phosphodiester
`
`linkages between the 5’ carbon of one nucleotide and the 3’ hydroxyl group of the
`
`next nucleotide. (Id.) The sugar-phosphate “backbone” forms the structural
`
`framework of DNA. As shown in Figure 2, one end of DNA has a free phosphate
`
`group and is referred to as the 5’-end of DNA. (Id.) The other end has a free -OH
`
`group and is referred to as the 3’-end of DNA. (Id.)
`
`
`
`9
`
`
`
`
`U.S. Patent No. 10,407,461
`
`O
`
`N
`
`NH
`
`NH2
`
`NH2
`
`N
`
`O
`
`N
`
`H
`
`H
`
`H
`
`O
`
`H
`
`H
`
`H
`
`H
`
`OH
`3'-end
`
`O
`
`O
`
`O
`
`N
`
`H
`
`NH
`
`O
`
`
`
`N
`
`N
`
`N
`
`H
`
`NH2
`
`N
`
`HH
`
`O
`
`O
`
`H
`
`O
`
`H
`
`O
`
`P
`
`-
`
`O
`
`5'-end
`
`O
`
`P
`
`-
`
`O
`
`O
`
`-
`
`O
`
`H
`
`O
`
`H
`
`O
`
`P
`
`-
`
`O
`
`N
`
`N
`
`H
`
`H
`
`H
`
`O
`
`O
`
`H
`
`O
`
`H
`
`O
`
`P
`
`-
`
`O
`
`
`
`
`
`Figure 2. Linked Nucleotides in DNA
`
`24. One chain of polynucleotides can form a double helix with another
`
`chain of polynucleotides because a base from one chain can pair with its
`
`complementary base from the other chain (also referred to as “Watson-Crick” base
`
`pairing). (Id., 194.) As shown in Figure 3 below, in DNA, A pairs with T while G
`
`pairs with C.
`
`
`
`10
`
`
`
`
`
`
`U.S. Patent No. 10,407,461
`
`
`
`Figure 3. Complementary Base Pairing in DNA (Ex. 1068, 194, Fig. 4-4)
`
`25. To encode a protein, cells copy (or “transcribe”) a portion of DNA
`
`into a precursor messenger RNA (“pre-mRNA”). (Id., 302, 317.) The pre-mRNA
`
`transcript contains the same genetic information as its DNA counterpart, except
`
`that it is in a different chemical form. (Id., 302.) Like DNA, RNA is a polymer
`
`made up of nucleotides containing a five-carbon sugar (ribose) and one of four
`
`nucleobases. RNA has the same bases as DNA except that it contains uracil (U)
`
`instead of thymine (T). (Id.) Like T in DNA, U in RNA makes a complementary
`
`base pairing with A. (Id.)
`
`26. Each pre-mRNA transcript contains segments that ultimately encode a
`
`protein (“exons”), interspersed with additional segments that are deleted (or
`
`
`
`11
`
`
`
`
`“spliced”) from the transcript (“introns”). (Id., 317.) Concurrently with
`
`
`U.S. Patent No. 10,407,461
`
`transcription of pre-mRNA, a complex referred to as the spliceosome recognizes
`
`and binds to certain parts of the transcript, cuts out introns, and stitches exons
`
`together. (Id., 319–320.) The resulting product is referred to as messenger RNA
`
`(“mRNA”). (Id., 304, 317.)
`
`27. The spliceosome binds to two ends of each intron (referred to as
`
`“donor” and “acceptor” regions). Other spicing components such as serine and
`
`arginine rich proteins (“SR proteins”) bind to select regions within exons known as
`
`exonic splicing enhancers (“ESEs”). (Ex. 1057, 1611–1612.) Binding of SR
`
`proteins to ESEs was understood to be essential for exon inclusion and interfering
`
`with this binding with AOs would have been expected to result in exon skipping.
`
`(Id., 1612.)
`
`28.
`
`In translating an mRNA molecule to a protein, proteins within cells
`
`read the sequence of nucleotides in the mRNA molecule in groups of three
`
`nucleotides (“codons”). (Ex. 1068, 336.) Each group of three nucleotides
`
`corresponds to (1) one of twenty amino acids or (2) a stop codon (i.e., a signal that
`
`terminates the translation of mRNA to protein). (Id.) In principle, an mRNA
`
`molecule can be read in three possible reading frames. (Id.) In reality, only one of
`
`the three reading frames has the ability to encode a correct protein (“open reading
`
`frame”). (Id.) Generally, each mRNA molecule contains a special signal that
`
`
`
`12
`
`
`
`
`allows for cells to read the mRNA molecule in a correct reading frame. (Id., 348–
`
`
`U.S. Patent No. 10,407,461
`
`349.) If, however, the mRNA molecule contains a mutation such as a deletion, the
`
`reading frame may be disrupted (“out-of-frame” or “frameshift” mutations). In
`
`that case, the translation of mRNA to protein may terminate prematurely due to
`
`inadvertent introduction of a stop codon and lead to a truncated protein or no
`
`protein at all. In contrast, an “in-frame” mutation can result from, for example, the
`
`addition or deletion of three nucleotides (or a multiple of three nucleotides). In this
`
`case, the open reading frame is preserved and subsequent codons after the mutation
`
`will not have a disrupted reading frame.
`
`B. Duchenne Muscular Dystrophy (DMD)
`29. DMD is an X-linked recessive muscular dystrophy that affects 1 in
`
`3,500–6,000 newborn males and causes progressive muscle weakness,
`
`cardiomyopathy, and respiratory failure. (Ex. 1027 [Bushby 2010], 77; Ex. 1028
`
`[Kinali 2009], 918; Ex. 1067 [Mann 2002], 644–645.) DMD patients lack
`
`functional dystrophin, a protein which is required to maintain the membrane
`
`integrity of muscle fibers. (Id.)
`
`30.
`
`In DMD, dystrophin protein is not produced, typically because a
`
`frame-shift mutation in the dystrophin (or DMD) gene disrupts the triplet reading
`
`frame, leading to premature termination of translation. (Ex. 1028 [Kinali 2009],
`
`918–919; Ex. 1029 [Hoffman 2007], 2721.) As shown in Figure 4 below,
`
`
`
`13
`
`
`
`
`dystrophin plays an important role in the muscle fiber’s structure and function,
`
`
`U.S. Patent No. 10,407,461
`
`forming a complex with other proteins that stabilizes the membrane of muscle
`
`cells. (Ex. 1040 [Nakamura 2009], 495.)
`
`
`
`
`
`Figure 4. Dystrophin (Ex. 1069 [Wood 2013], 170, Fig. 1)
`
`31. The dystrophin (or DMD) gene contains 79 exons. (Ex. 1030 [Wilton
`
`2008], 255.) Although mutations can occur anywhere in the dystrophin gene, it
`
`was known by August 31, 2011 that mutations in DMD patients are concentrated
`
`
`
`14
`
`
`
`
`in regions between exon 43 and exon 55. (Ex. 1036 [Muntoni 2010], 358.)
`
`
`U.S. Patent No. 10,407,461
`
`Researchers by August 31, 2011 understood that AOs targeting exon 51 or exon 53
`
`had the potential to treat relatively large numbers of DMD patients. (Ex. 1021
`
`[Popplewell], 102–103; Ex. 1036 [Muntoni 2010], 358, Table 1.)
`
`C. Exon Skipping Antisense Oligomers (AOs) as DMD Therapies
`Exon Skipping AOs
`1.
`In the 1990s, Kole and colleagues demonstrated that AOs can induce
`
`32.
`
`alternative splicing. (Ex. 1070 [Dominski and Kole 1993].) For instance, certain
`
`mutations to β-globin genes generate alternative splice sites. These result in an
`
`incorrect, alternatively spliced transcript. (Id. [Dominski and Kole 1993], 8673.)
`
`AOs complementary to these aberrant splice sites masked the alternative sites,
`
`forcing the splicing machinery to recognize normal splice sites, thereby restoring
`
`correct splicing. (Id. [Dominski and Kole 1993], 8675; see also Ex. 1071
`
`[Dominski 1994], Abstract; Ex. 1072 [Sierakowska 1996], Abstract.)
`
`33.
`
`Inducing exon skipping is similar in principle (Figure 5). A sequence
`
`in the pre-mRNA is targeted by an AO such that the binding of the AO to the pre-
`
`mRNA interferes with splicing. This interference results in one or more exons in
`
`the pre-mRNA being excluded from the mRNA transcript. (Ex. 1030 [Wilton
`
`2008], 256.)
`
`
`
`15
`
`
`
`
`
`
`U.S. Patent No. 10,407,461
`
`
`
`Figure 5. AO Inducing Skipping of Exon 3
`
`Natural Precedents for Exon Skipping AOs in DMD
`2.
`34. Truncated but semi-functional dystrophins are expressed in so-called
`
`“revertant fibers” in many DMD patients. (Ex. 1031 [Arechavala-Gomeza 2007],
`
`798–799.) These individual dystrophin positive fibers were believed to arise via
`
`alternative splicing of dystrophin pre-mRNAs. (Id.)
`
`35. Mutations in the dystrophin gene were also known to cause a milder
`
`form of muscular dystrophy called Becker muscular dystrophy (BMD). (Ex. 1028
`
`[Kinali 2009], 919; Ex. 1030 [Wilton 2008], 254; Ex. 1031 [Arechavala-Gomeza
`
`2007], 798; Ex. 1067 [Mann 2002], 645.) In BMD patients (unlike in DMD
`
`patients), in-frame mutations allow the expression of internally deleted, but largely
`
`functional, dystrophin. (Id.) BMD patients experience milder symptoms than
`
`DMD patients, ranging from borderline DMD to no symptoms at all. (Id.)
`
`36. Exon skipping therapy in DMD has a natural precedent in the
`
`revertant fibers found in many DMD patients and in the truncated but functional
`
`dystrophin of BMD patients. (Ex. 1031 [Arechavala-Gomeza 2007], 798–799.)
`16
`
`
`
`
`
`
`
`
`U.S. Patent No. 10,407,461
`
`Exon Skipping AOs in DMD
`3.
`37. As of August 31, 2011, AOs investigated for exon skipping therapy
`
`were typically single stranded and were 20–30 bases in length. (Ex. 1035 [Wilton
`
`2007], 1290, Table 1; Ex. 1040 [Nakamura 2009], 495; Ex. 1041 [Yokota 2009],
`
`32; Ex. 1021 [Popplewell], 109; Ex. 1042 [Aartsma-Rus 2009], 548.) Exon
`
`skipping AOs can bind (or “hybridize”) to a target sequence in the pre-mRNA
`
`under physiological conditions via Watson-Crick base paring. Binding of the AO
`
`to the pre-mRNA alters splicing of the pre-mRNA such that one or more exon(s) in
`
`the pre-mRNA are “skipped” in the mRNA transcript. (Ex. 1029 [Hoffman 2007],
`
`2719–2720.) The resulting restoration of the reading frame produces truncated,
`
`semi-functional dystrophin protein. (Ex. 1031 [Arechavala-Gomeza 2007], 799;
`
`Ex. 1032 [Ginjaar 2000], 796; see generally Ex. 1033 [Wilton 2005].) Figure 6
`
`below depicts the mechanism of skipping exon 53 in the dystrophin gene:
`
`
`
`
`
`
`
`17
`
`
`
`
`
`
`U.S. Patent No. 10,407,461
`
`
`
`Figure 6. Mechanism of Exon Skipping AO (adapted from Ex. 1043
`
`[Aartsma-Rus 2009], 873, Fig. 1)
`
`Preclinical and Clinical Studies
`a)
`38. By August 31, 2011, numerous preclinical studies, including DMD
`
`animal models, had confirmed exon skipping AOs to be a viable treatment strategy
`
`for DMD. (See Ex. 1031 [Arechavala-Gomeza 2007], 800–801, Table 1
`
`(summarizing various in vitro and in vivo studies evaluating the exon skipping
`
`efficacy of PMOs and 2’-OMePSs).)
`
`39. Following the 2007 publication of the pre-clinical testing and
`
`selection of Sarepta’s exon 51 targeting AO (Ex. 1031 [Arechavala-Gomeza 2007],
`
`and by August 31, 2011, two companies had initiated clinical trials evaluating two
`
`AOs targeting exon 51. Prosensa was developing drisapersen (f/k/a PRO051), a
`
`2’-OMePS, and Sarepta was developing eteplirsen (f/k/a AVI-4658), a PMO. In
`
`2007, it was reported that in a proof-of-concept study PRO051 induced skipping of
`
`
`
`18
`
`
`
`
`exon 51 and resulted in production of truncated dystrophin protein in four DMD
`
`
`U.S. Patent No. 10,407,461
`
`patients. (Ex. 1037 [van Deutekom 2007], Abstract, Figure 4.) Similarly
`
`encouraging results were reported in 2009 for AVI-4658. (Ex. 1028 [Kinali 2009],
`
`Abstract, Figure 3.) A study published in April 2011 reported that PRO051
`
`resulted in functional improvement in DMD patients. (Ex. 1038 [Goemans 2011],
`
`Abstract.) Shortly thereafter, on August 13, 2011, positive results from a Phase II
`
`trial evaluating AVI-4658 were published, further establishing exon skipping
`
`therapy as a viable approach for treating DMD. (Ex. 1039 [Cirak 2011], Abstract.)
`
`b) Design of Exon Skipping AOs Targeting the
`Dystrophin Transcript
`40. By August 31, 2011, researchers had identified length as important in
`
`the design of AOs. For some AOs, increasing their length improved skipping
`
`efficiency, whereas for other AOs, increasing AO length decreased skipping
`
`efficiency. (Ex. 1046 [Adams 2007], 3 (“While ‘longer is better’ in many
`
`instances of single AO-induced exon skipping, this does not hold true in all
`
`cases.”), 5; Ex. 1057 [Aartsma-Rus 2007], 1613 (“Our own recent studies indicate
`
`that the efficiency of some AONs that induce very low levels of exon skipping can
`
`be enhanced by increasing AON length, whereas increasing the length of an
`
`already efficient AON did not enhance AON efficiency and occasionally even
`
`
`
`19
`
`
`
`
`reduced exon skipping levels.”); Ex. 1042 [Aartsma-Rus 2009], 552; Ex. 1030
`
`
`U.S. Patent No. 10,407,461
`
`[Wilton 2008], 258.)
`
`41.
`
`It was understood as of August 31, 2011 that as AO length increased,
`
`so too did the possibility of specific and non-specific off-target effects.
`
`Researchers were also mindful of considerations such as ease of synthesis and
`
`manufacturing costs. (See, e.g., Ex. 1040 [Nakamura 2009], 495 (“Development
`
`of appropriate AOs requires consideration of several characteristics of AOs, such
`
`as the chemical specificity, affinity, nuclease resistance, stability, safety, and ease
`
`of synthesis[.]”).) As a general matter, as AO length increases, so do
`
`manufacturing costs and synthesis challenges. For these reasons, when two AOs
`
`showed similar skipping efficiency, the shorter one was often favored.
`
`42. By August 31, 2011, researchers routinely used modified nucleobases,
`
`chemical backbones, and intersubunit linkages to enhance the stability and potency
`
`of AOs, while preserving their ability to bind to their target pre-mRNA sequences
`
`via Watson-Crick base pairing. (Ex. 1044 [Chan 2006], 535–536; Ex. 1040
`
`[Nakamura], 495–496; Ex. 1045 [Summerton 1997].) Two of the most common
`
`classes of AOs used as of August 31, 2011 were “2’-OMePSs” and “PMOs,”
`
`shown in Figure 7 below.
`
`
`
`20
`
`
`
`
`
`
`U.S. Patent No. 10,407,461
`
`Phosphorothioate-linked 2’-O-methyl
`
`Phosphorodiamidate morpholino
`
`ribose oligomer (“2’-OMePS”)
`
`oligomer (“PMO”)
`
`
`
`
`
`
`
`Figure 7. 2’-OMePS and PMO Modifications (adapted from Ex. 1040
`
`[Nakamura 2009], 496, Fig. 1)
`
`43. For example, Nakamura 2009 noted that “[2’-OMePS] and PMO are
`
`the most frequently utilized because of their suitable properties.” (Ex. 1040
`
`[Nakamura 2009], 495). Similarly, Aartsma-Rus 2009 stated: “The most
`
`commonly used chemistries for exon-skipping antisense oligonucleotides are 2’-O-
`
`methyl RNA with a negatively charged phosphorothioate backbone (2’-OMePS)
`
`and uncharged phosphorodiamidate morpholino oligomers (PMO) . . . .” (Ex. 1043
`
`[Aartsma-Rus 2009], 873.) Sazani further recognized: “The favorable safety
`
`profile of the uncharged PMOs has been remarkably consistent and predictable, as
`
`
`
`21
`
`
`
`
`more than 460 patients have been safely dosed with PMO in clinical trials for a
`
`
`U.S. Patent No. 10,407,461
`
`variety of indications.” (Ex. 1022 [Sazani 2010], 155.)
`
`44.
`
`2’-OMePSs have a structure similar to RNA but the 2’-OH position of
`
`the ribose ring has been methylated, and instead of using a phosphodiester link
`
`between nucleotides, one of the non-bridging oxygen atoms of the phosphate group
`
`is substituted with a sulfur atom to create a phosphorothioate linkage. PRO051,
`
`one of two exon 51 clinical candidates being evaluated for the treatment of DMD
`
`as of August 31, 2011, was a 2’-OMePS that was 20 bases in length. (Ex. 1037
`
`[van Deutekom 2007], 2679; Ex. 1043 [Aartsma-Rus 2009], 874, Table 1.)
`
`45. As of August 31, 2011, it was understood that 2’-OMePSs could
`
`contain either uracil or thymine bases, and that either base would be
`
`complementary to an adenine base found in pre-mRNA and would pair with
`
`adenine in the same way via Watson-Crick base pairing. (Ex. 1048 [Aartsma-Rus
`
`2005], 286, Table 1 (describing 2’-OMePSs with U’s); Ex. 1055 [Summerton
`
`1999], 144, Fig. 1 (showing 2’-OMePS modification and indicating bases are
`
`selected from A, C, G, T, and U).)
`
`46. PMOs, a type of “morpholino,” have a six-membered morpholinyl
`
`moiety instead of a ribose. PMO subunits are linked through uncharged
`
`phosphorodiamidate linkages. (Ex. 1040 [Nakamura 2009], 496.) The second
`
`exon 51 targeting AO undergoing clinical trials as of August 31, 2011, AVI-4658,
`
`
`
`22
`
`
`
`
`was a PMO that was 30 bases in length. (Ex. 1031 [Arechavala-Gomeza 2007],
`
`
`U.S. Patent No. 10,407,461
`
`808; Ex. 1043 [Aartsma-Rus 2009], 874, Table 1.) AVI-4658 was ultimately
`
`approved in 2016 by the U.S. Food and Drug Administration as Exondys 51®, the
`
`first disease-modifying treatment for DMD.
`
`47. As of August 31, 2011, it was understood that PMOs could contain
`
`either uracil or thymine bases, and that eith
Accessing this document will incur an additional charge of $.
After purchase, you can access this document again without charge.
Accept $ Charge
Still Working On It
This document is taking longer than usual to download. This can happen if we need to contact the court directly to obtain the document and their servers are running slowly.
Give it another minute or two to complete, and then try the refresh button.
A few More Minutes ... Still Working
It can take up to 5 minutes for us to download a document if the court servers are running slowly.
Thank you for your continued patience.
This document could not be displayed.
We could not find this document within its docket. Please go back to the docket page and check the link. If that does not work, go back to the docket and refresh it to pull the newest information.
Your account does not support viewing this document.
You need a Paid Account to view this document. Click here to change your account type.
Your account does not support viewing this document.
Set your membership
status to view this document.
With a Docket Alarm membership, you'll
get a whole lot more, including:
- Up-to-date information for this case.
- Email alerts whenever there is an update.
- Full text search for other cases.
- Get email alerts whenever a new case matches your search.
One Moment Please
The filing “” is large (MB) and is being downloaded.
Please refresh this page in a few minutes to see if the filing has been downloaded. The filing will also be emailed to you when the download completes.
Your document is on its way!
If you do not receive the document in five minutes, contact support at support@docketalarm.com.
Sealed Document
We are unable to display this document, it may be under a court ordered seal.
If you have proper credentials to access the file, you may proceed directly to the court's system using your government issued username and password.
Access Government Site
