Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`Filed on behalf of Patent Owners Genentech, Inc. and City of Hope by:
`Jeffrey P. Kushan
`Adam R. Brausa
`David L. Cavanaugh
`Reg. No. 43,401
`Reg. No. 60,287
`Reg. No. 36,476
`Peter S. Choi
`Daralyn J. Durie
`Heather M. Petruzzi
`Pro Hac Vice Application
`Reg. No. 54,033
`Reg. No. 71,270
`Sidley Austin LLP
`Pending
`Robert J. Gunther, Jr.
`Pro Hac Vice Application
`1501 K Street, N.W.
`Durie Tangri LLP
`Washington, D.C.
`217 Leidesdorff Street
`Pending
`20005
`San Francisco, CA 94111
`Wilmer Cutler Pickering
`Hale and Dorr LLP
`1875 Pennsylvania Ave., NW
`Washington, DC 20006
`
`UNITED STATES PATENT AND TRADEMARK OFFICE
`____________________________________________
`BEFORE THE PATENT TRIAL AND APPEAL BOARD
`____________________________________________
`SANOFI-AVENTIS U.S. LLC AND
`REGENERON PHARMACEUTICALS, INC.,
`Petitioners
`v.
`GENENTECH, INC. AND CITY OF HOPE
`Patent Owners
`____________________________________________
`Case IPR2015-01624
`Patent 6,331,415
`
`____________________________________________
`EXPERT DECLARATION OF JOHN FIDDES, PH.D.
`
`MERCK v. GENENTECH
`IPR2016-01373
`GENENTECH 2012
`
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`TABLE OF CONTENTS
`
`Page
`
`I.
`
`II.
`
`INTRODUCTION AND BACKGROUND............................................. 1
`A.
`Qualifications And Experience..................................................... 2
`B. Compensation ............................................................................ 4
`C.
`Prior Expert Testimony................................................................ 4
`LEGAL PRINCIPLES ON OBVIOUSNESS........................................... 4
`
`III. BACKGROUND OF THE TECHNOLOGY........................................... 7
`A.
`Genes, Proteins And Antibodies ................................................... 7
`
`B. Antibody Production Techniques As Of April 1983....................... 16
`C. Use of Recombinant Gene Expression To Produce Proteins............ 18
`1.
`Basic principles of recombinant gene expression.................. 18
`
`2.
`
`As of April 1983, Only a Small Number of Monomeric
`Eukaryotic Proteins Had Been Produced Recombinantly....... 20
`D. As of April 1983, Insulin Was The Only Multimeric Eukaryotic
`Protein Which Had Been Produced Recombinantly....................... 29
`
`F.
`
`E.
`
`In May 1981 (When Bujard Was Filed), The Speculative
`Possibility Of Using Recombinant Techniques To Produce
`Antibodies Was Highly Uncertain And Unpredictable ................... 37
`Research Involving Antibodies And Recombinant Gene
`Expression Between May 1981 And April 1983 Confirmed The
`Uncertainty And Unpredictability Of Whether Antibodies
`Could Be Produced Recombinantly............................................. 41
`G. As Of April 1983, Highly Acclaimed Scientists Were Still
`Uncertain Whether It Was Even Possible To Make Antibodies
`Using Recombinant Techniques.................................................. 46
`
`i
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`H. As Of April 1983, Nobody Had Produced An Antibody
`Recombinantly ......................................................................... 49
`
`I.
`
`As Of April 1983, Nobody Had Produced A Multimeric
`Eukaryotic Protein Recombinantly In A Single Host Cell............... 49
`THE CLAIMS UNDER CONSIDERATION AND THEIR
`INTERPRETATION .......................................................................... 53
`A.
`The Cabilly ’415 Patent............................................................. 53
`B.
`Summary Of Contested Claims................................................... 54
`C.
`The Person Of Ordinary Skill In The Art...................................... 56
`OPINIONS REGARDING THE ASSERTED PRIOR ART .................... 57
`
`IV.
`
`V.
`
`A. Bujard ..................................................................................... 58
`1. What is the focus of the Bujard reference? .......................... 58
`
`2.
`
`3.
`
`4.
`
`5.
`
`6.
`
`Do the references to “multimers” and “one or more structural
`genes” in Bujard suggest the use of the co-expression of
`multiple, distinct eukaryotic genes of interest in a single host
`cell? ............................................................................... 63
`
`Do the references to multiple “stop codons” in Bujard suggest
`the use of the strong promoter/terminator system for the
`expression of multiple, distinct genes in a single host cell?.... 72
`
`Does Bujard “at least suggest the coexpression of the heavy and
`light chains” of an immunoglobulin in a “single host cell,” as
`the Board found? ............................................................. 75
`
`Does Bujard “teach away from the production of light chains in
`one culture and heavy chains in another, to be combined
`chemically at a stage after their harvest and isolation,” as Dr.
`Foote argues?.................................................................. 79
`
`Does the statement in Bujard that the “proteins may be prepared
`as a single unit or as individual subunits and then joined
`
`ii
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`together in appropriate ways” suggest in vivo assembly of a
`multimeric protein in a single host cell, as Dr. Foote argues? . 84
`
`7.
`
`Do you agree with the Board’s finding that Bujard is “more
`specific and robust” than the Axel reference? ...................... 86
`B. Riggs & Itakura........................................................................ 88
`1. What is the focus of the Riggs & Itakura reference? ............. 88
`
`2.
`
`Does the Riggs & Itakura reference address the “same problem”
`as Bujard and would the skilled artisan have had “good reason”
`to combine these two references in April 1983?................... 89
`
`3. Would inferences gleaned from Riggs & Itakura have provided
`the person of ordinary skill with the motivation to “selectively
`apply the teachings of Bujard to the specific production of
`immunoglobulins” by means of co-expressing both the heavy
`and light chain in a single host cell? ................................... 91
`
`4.
`
`Do you agree with Dr. Foote’s opinion and the Board’s
`preliminary finding that Bujard in combination with Riggs &
`Itakura renders the claimed invention of the Cabilly ’415 patent
`obvious? ......................................................................... 98
`Southern ................................................................................ 100
`1. What is the focus of the Southern reference? ..................... 100
`
`C.
`
`2.
`
`Do you agree with Dr. Foote’s opinion that the skilled artisan
`would have been motivated to combine Bujard and Southern
`because “both have as a goal the expression of genes of interest
`in a single transformed host cell, whether by using one (Bujard)
`or two (Southern) vectors”?............................................. 102
`
`3. Would inferences gleaned from Southern have provided the
`person of ordinary skill with the motivation to “selectively
`apply the teachings of Bujard to the specific production of
`immunoglobulins” by means of co-expressing both the heavy
`and light chain in a single host cell? ................................. 104
`
`iii
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`1.
`
`Do you agree with Dr. Foote’s opinion and the Board’s
`preliminary finding that Bujard in combination with Southern
`renders the claimed invention of the Cabilly ’415 patent
`obvious? ....................................................................... 111
`
`iv
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`I.
`
`INTRODUCTION AND BACKGROUND
`1.
`I, John Fiddes, Ph.D., have been retained by counsel for
`
`Genentech, Inc. and City of Hope (collectively, “Patent Owners”) as an expert in
`
`this proceeding.
`
`2.
`
`I understand that, in a February 5, 2016 decision, the Patent
`
`Trial and Appeal Board (the “Board”) instituted inter partes review as to claims 1-
`
`4, 11, 12, 14, 18-20, and 33 of U.S. Patent No. 6,331,415 (“the Cabilly ’415
`
`patent”). I further understand that the references relied upon by the Board in
`
`instituting inter partes review include the Bujard patent (Ex. 1002), the Riggs &
`
`Itakura paper (Ex. 1003) and the Southern paper (Ex. 1004).
`
`3.
`
`I have been asked to review the challenged claims of the
`
`Cabilly ’415 patent and the references identified in the petition requesting inter
`
`partes review, and evaluate whether the cited references alone or in combination
`
`render the challenged claims unpatentable. As part of my review I have been
`
`asked to evaluate the prior art and scientific accuracy of the observations that the
`
`Board made in the decision instituting inter partes review. I also was asked to
`
`evaluate certain statements that Dr. Jefferson Foote made in his declaration (Ex.
`
`1006) submitted with the petition requesting inter partes review.
`
`1
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`4.
`
`A list of materials I have reviewed in preparation of this
`
`Declaration is attached as Exhibit B. I have also relied upon my scientific
`
`knowledge as of April 1983 when the Cabilly ’415 patent was filed.
`
`A.
`
`Qualifications And Experience
`5. My background is summarized below and in my curriculum
`
`vitae, which includes a list of my publications and patents and is attached as
`
`Exhibit A.
`
`6.
`
`I received a Bachelor of Science degree in Biological Sciences
`
`(Molecular Biology) with First Class Honors from the University of Edinburgh in
`
`1973. In 1977, I received my Ph.D. in Molecular Biology from Kings College,
`
`Cambridge University. My thesis advisor was Dr. Fred Sanger. The title of my
`
`thesis was “The Determination (cid:82)(cid:73)(cid:3)(cid:49)(cid:88)(cid:70)(cid:79)(cid:72)(cid:82)(cid:87)(cid:76)(cid:71)(cid:72)(cid:3)(cid:54)(cid:72)(cid:84)(cid:88)(cid:72)(cid:81)(cid:70)(cid:72)(cid:86)(cid:3)(cid:76)(cid:81)(cid:3)(cid:37)(cid:68)(cid:70)(cid:87)(cid:72)(cid:85)(cid:76)(cid:82)(cid:83)(cid:75)(cid:68)(cid:74)(cid:72)(cid:3)(cid:301)(cid:59)(cid:20)(cid:26)(cid:23)(cid:3)
`
`DNA.”
`
`7.
`
`From 1977 to 1980, I was a Postdoctoral Research Fellow at the
`
`University of California, San Francisco (“UCSF”), in the laboratory of Dr. Howard
`
`Goodman, where I worked on the human growth hormone, human chorionic
`
`somatomammotropin and human glycoprotein hormone genes.
`
`8.
`
`After my post-doc at UCSF, I became a Senior Staff
`
`Investigator at Cold Spring Harbor Laboratory (“CSHL”) in Cold Spring Harbor,
`
`New York, a position I held until January 1983.
`
`2
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`9. My research at CSHL focused on the structure, evolution and
`
`expression of the human glycoprotein hormone genes, specifically human
`
`chorionic gonadotropin and human luteinizing hormone, and on methods of
`
`making cDNA libraries suitable for immunological screening of expression
`
`products. I was also an instructor at the CSHL Advanced Cloning Course in the
`
`summers of 1982-1983.
`
`10.
`
`Following my academic career, I entered industry and spent
`
`over twenty years in drug discovery and development. In January 1983, just
`
`shortly before the filing date of the Cabilly ’415 patent, I took a position at
`
`California Biotechnology Inc. (later renamed Scios Inc.) in Mountain View,
`
`California. The primary interest of this company was in applying recombinant
`
`DNA technologies to the production of therapeutically useful proteins.
`
`11. Among other things, I was involved in the development of
`
`systems for the production of recombinant forms of basic fibroblast growth factor,
`
`and the isolation of cDNA and genomic clones for atrial natriuretic peptide,
`
`vascular endothelial growth factor variant and heparin-binding, EGF-like growth
`
`factor.
`
`12. My last industry position was at Genencor International Inc. in
`
`Palo Alto, California where I served as Vice President Research, Health Care from
`
`2003 to 2005. Since 2005, I have been an independent consultant on
`
`3
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`biopharmaceutical matters for a variety of organizations, including the California
`
`Antiviral Foundation and the Institute for One World Health.
`
`13. Based on my academic and early industry experience, I was
`
`well aware of the birth of recombinant DNA technology and followed the
`
`developments that eventually led to the production of recombinant forms of
`
`medically important proteins. This, in my view, is the art to which the Cabilly
`
`’415 patent pertains, and I believe I am well-positioned to understand and address
`
`the skills and mindset of a person of ordinary skill in this field circa 1982-1983.
`
`B.
`
`Compensation
`14.
`I am being compensated at my normal consulting rate for my
`
`work, which is $650 per hour. My compensation is not dependent on and in no
`
`way affects the substance of my statements in this Declaration.
`
`C.
`
`Prior Expert Testimony
`I provided expert reports and deposition testimony in Bristol-
`15.
`
`Myers Squibb Co. v. Genentech, Inc. & City of Hope, 2:13-cv-05400-MRP-JEM
`
`(C.D. Cal), and Eli Lilly & Co. v. Genentech, Inc. 2:13-cv-07248-MRP-JEM (C.D.
`
`Cal.).
`
`II.
`
`LEGAL PRINCIPLES ON OBVIOUSNESS
`16.
`I have been informed and understand that in order to invalidate
`
`a patent claim as obvious in the context of an inter partes review, it must be shown
`
`4
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`by a preponderance of the evidence that the claim would have been obvious to a
`
`person of ordinary skill at the time the invention was made. The prior art does not
`
`need to render obvious every possible embodiment within the scope of the claim:
`
`the prior art renders the claim obvious if the combined teachings disclose an
`
`embodiment that is within the scope of the claim.
`
`17.
`
`I have been informed and understand that factors relevant to the
`
`determination of obviousness include the scope and content of the prior art, the
`
`level of ordinary skill in the art at the time of the invention, differences between
`
`the claimed invention and the prior art and “secondary considerations” or objective
`
`evidence of non-obviousness.
`
`18.
`
`I have been informed and understand that obviousness can be
`
`established by combining or modifying the teachings of the prior art to produce the
`
`claimed invention where there is some teaching, suggestion or motivation to do so;
`
`and that a reasonable expectation of success in achieving the subject matter of the
`
`claim at issue must also be shown. Further, I have been informed and understand
`
`that the teaching, suggestion or motivation test is flexible and that an explicit
`
`suggestion to combine the prior art is not necessary – the motivation to combine
`
`may be implicit and may be found in the knowledge of one of ordinary skill in the
`
`art, from the nature of the problem to be solved, market demand or common sense.
`
`5
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`19.
`
`I have been informed and understand that a patent claim
`
`composed of several limitations is not obvious merely because each limitation was
`
`independently known in the prior art. Hindsight reasoning is not an appropriate
`
`basis for combining references to form an obviousness combination. I also have
`
`been informed and understand that it can be important to identify a reason that
`
`would have prompted a person of ordinary skill in the relevant field to combine the
`
`limitations in the way the claimed new invention does.
`
`20.
`
`In undertaking an obviousness analysis, I have been informed
`
`and understand that I may take into account the inferences and creative steps that a
`
`person of ordinary skill would have employed in reviewing the prior art at the time
`
`of the invention. If the claimed invention combines elements known in the prior
`
`art and the combination yields results that would have been predictable to a person
`
`of ordinary skill at the time of the invention, then this evidence would make it
`
`more likely that the claim was obvious.
`
`21.
`
`I have also been informed and understand that obviousness may
`
`be established if the combination of prior art elements was obvious to try, even if
`
`no one attempted the combination. For a combination to be obvious to try,
`
`however, a solution must be among a finite number of identified, predictable
`
`solutions. Where the art is uncertain or unpredictable, a person of ordinary skill in
`
`the art will not have a reasonable expectation of success.
`
`6
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`22.
`
`I have been informed and understand that an obviousness
`
`analysis must take into account any “secondary considerations” or, as they are
`
`sometimes called, objective indicia of non-obviousness. These secondary
`
`considerations can include the invention’s commercial success, long-felt but
`
`unresolved needs, licenses showing industry respect, the failure of others,
`
`skepticism by experts, praise by others, teaching away by others, recognition of a
`
`problem and copying of the invention by competitors. Such secondary
`
`considerations, when present, offer objective information as to the state of the art at
`
`the time of the invention and provide a check to hindsight analysis.
`
`III. BACKGROUND OF THE TECHNOLOGY
`23.
`To place the importance and innovation of the Cabilly ’415
`
`patent into context, I have been asked to provide some background on the relevant
`
`technology.
`
`A.
`
`Genes, Proteins And Antibodies
`
`24. A deoxyribonucleic acid (DNA) molecule encodes the genetic
`
`instructions that a living organism uses for a wide variety of critical functions.
`
`Sequences of DNA nucleotides are organized into discrete structures, called genes,
`
`which a cell’s machinery “reads” to make proteins. Proteins, comprised of a string
`
`of units called amino acids, are biomolecules that perform many of the functions of
`
`cells and organisms.
`
`7
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`25.
`
`The basic process of making a protein from a gene is called
`
`“gene expression.” First, the cell copies the gene of interest (a DNA sequence)
`
`into messenger ribonucleic acid (mRNA) via a process called transcription.
`
`Second, the mRNA is converted into the corresponding sequence of amino acids
`
`(called a polypeptide) via a process called translation. Finally, the translated
`
`polypeptide undergoes folding and possibly post-translational modifications to
`
`assemble as the active protein structure.
`
`26.
`
`This step-wise process is reflected in the following illustration:
`
`(Ex. 2081, Bruce Alberts et al., Essential Cell Biology, Chapter 1 (3rd ed. 2009).)
`
`8
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`27.
`
`Transcription: In transcription, an enzyme in cells called RNA
`
`polymerase synthesizes single-stranded mRNA in a linear fashion from the
`
`doubled-stranded genomic DNA template.
`
`28.
`
`Translation: In translation, complex structures in cells called
`
`ribosomes bind to specific sites on the mRNA transcript and translate the mRNA
`
`sequence into a polypeptide chain of amino acids. The mRNA sequence is read
`
`three nucleotides at a time, with each triplet of nucleotides – called a codon –
`
`specifying one amino acid. A start codon initiates translation, and stop codons
`
`signal termination of translation. Codons, therefore, provide the information that
`
`dictates the order and arrangement of amino acids in a polypeptide chain, and
`
`when the formation of the chain begins and ends.
`
`29. An exemplary coding region (from protein B of bacteriophage
`
`G4) is shown in the table below (see Ex. 2082, G. N. Godson et al., Nucleotide
`
`Sequence of Bacteriophage G4 DNA, Nature 276:236-47 (1978)):
`
`Codon
`Amino Acid
`
`TAC
`TYR
`
`GGA
`GLY
`
`TAT
`TYR
`
`TTC
`PHE
`
`TGA
`STOP
`
`TGA
`STOP
`
`30. As seen in this table above, multiple stop codons (written for
`
`ease of reference as DNA as opposed to mRNA) can signal the end of translation
`
`of a single polypeptide. A coding region may end with multiple stop codons to
`
`9
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`ensure termination of translation. A start codon would be found upstream of the
`
`region shown above.
`
`31. Depending on where translation begins, each mRNA sequence
`
`can be translated in any one of several “reading frames,” i.e., the various ways in
`
`which the sequence may be divided into sets of nucleotide triplets.
`
`32. Folding: As part of the process of translation, the polypeptide
`
`chain folds to take on its final structure and to become an assembled, active
`
`protein. Folding allows the polypeptide to form its three dimensional structure and
`
`occurs as amino acids within a polypeptide chain interact with one another.
`
`Disulfide bonds or bridges (which are covalent bonds formed between cysteine
`
`amino acids) form a scaffolding that helps maintain a protein’s proper three
`
`dimensional structure. (Disulfide bonds may also be referred to as S-S bonds.)
`
`Such scaffolding may be present in both simple and more complex proteins, and
`
`where present is essential to the activity of a protein.
`
`33.
`
`Post-translational modification: This generally refers to
`
`processes by which cells may modify a polypeptide after it has been produced to
`
`achieve a mature product, and can include any number of processes.
`
`34. Monomeric and Multimeric Proteins: The most simple
`
`proteins are monomeric, meaning they are formed by only a single polypeptide
`
`chain. Other proteins exist as a complex of multiple polypeptide chains called
`
`10
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`“multimeric proteins.” Multimeric proteins can be made up of multiple identical
`
`polypeptide chains or a combination of different polypeptide chains. For
`
`multimeric proteins, amino acid interactions between individual chains and
`
`disulfide bonding between individual chains are critical to correct folding.
`
`35.
`
`Insulin: There are many different types of multimeric proteins.
`
`One example is insulin, which is a relatively simple and small (i.e., ~ 5,800
`
`Daltons) multimeric protein. (A Dalton is a standard unit of measurement used to
`
`characterize the mass of a protein.) The insulin molecule contains two different
`
`polypeptide chains – an “A” chain consisting of 21 amino acids and a “B” chain
`
`consisting of 30 amino acids – that are chemically attached via two intra-chain
`
`disulfide (S-S) bonds.
`
`36.
`
`The relatively simple configuration of the insulin protein
`
`(including its disulfide bonding scheme) is shown in the illustration below:
`
`11
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`(Ex. 2083, Protein Structure, Boundless,
`
`https://www.boundless.com/biology/textbooks/boundless-biology-
`
`textbook/biological-macromolecules-3/proteins-56/protein-structure-304-11437/.
`
`(last visited May 12, 2016).)
`
`37. As can be seen above, the two chains that comprise insulin are
`
`joined together by two inter-chain disulfide bonds; there is also one intra-chain
`
`disulfide bond.
`
`38. Antibodies: Antibodies, also known as immunoglobulins, are
`
`large tetrameric proteins (which may form even larger complexes) that are
`
`expressed and secreted by B cells (a type of white blood cell made in the bone
`
`marrow). There are five classes of antibodies (IgG, IgD, IgE, IgA, and IgM), each
`
`of which is further divided into multiple different types called isotypes.
`
`39. A naturally occurring tetrameric antibody is composed of four
`
`polypeptide chains – two identical “heavy” chains (or “H” chains) and two
`
`identical “light” chains (or “L” chains). The heavy and light chains differ in their
`
`size, and thus, their respective molecular weights. By way of example, in
`
`antibodies of the immunoglobulin G (“IgG1”) class, the longer H chains are
`
`naturally comprised of about 450 amino acids and each have a molecular weight of
`
`about 50,000 Daltons, whereas the shorter L chains are naturally comprised of
`
`about 212 amino acids and each have a molecular weight of about 25,000 Daltons.
`
`12
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`40.
`
`The heavy and light chains of an antibody form what is often
`
`depicted schematically as a Y-shaped molecule via multiple intra- and inter-chain
`
`disulfide bonds. For example, in the case of an antibody of the IgG1 class, there
`
`are 12 intra-chain disulfide bonds and 4 inter-chain disulfide bonds that together
`
`provide inter- and intra-chain “scaffolding” within the structure of an antibody, as
`
`discussed above.
`
`41.
`
`The below figure (taken from FIG. 1 of the Cabilly ’415 patent)
`
`provides a representation of an antibody’s Y-shape and how the various heavy and
`
`light chains assemble via disulfide bonds to form a functional antibody:
`
`42. As shown above, both the heavy and light chains have a
`
`“constant” region and a “variable” region. The “constant” regions may be the
`
`same between different types of antibodies. By contrast, the “variable” regions
`
`13
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`differ from one individual antibody to the next; these variable regions are
`
`responsible for identifying and binding to a particular antigen (i.e., a substance that
`
`causes the immune system of an organism to generate antibodies that bind the
`
`antigen in order to provoke an immune response).
`
`43.
`
`In sum, an antibody is a much larger and more complex protein
`
`than insulin (and most other proteins). The molecular weight of a typical antibody
`
`is approximately 150,000 Daltons, or more than 25 times the size of insulin.
`
`Further, the three-dimensional shape of an antibody is more complex and requires
`
`much more post-translational processing.
`
`[Continued on next page]
`///
`///
`///
`///
`///
`///
`///
`
`///
`///
`///
`
`14
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`44.
`
`The table below provides a to-scale comparison that illustrates
`
`the relative size of insulin compared to an antibody.
`
`Structure
`
`Insulin
`dimer (ab)
`
`Immunoglobulin G
`tetramer (a2b2)
`
`Size
`
`51 amino acids
`5,800 Daltons
`
`Disulfide
`Bonds
`
`3
`
`1324 amino acids
`150,000 Daltons
`
`16
`
`45. At my request, the space-filling models in the above figure
`
`were generated by scientists at Genentech using PyMOL software (available for
`
`download at https://www.pymol.org/). Each sphere represents an atom, with gray
`
`representing carbon, red representing oxygen, blue representing nitrogen, yellow
`
`representing sulfur, and green representing zinc. The size difference between
`
`insulin and an antibody represented above is consistent with my general
`15
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`understanding regarding the relative overall shape and size of each. (See also Ex.
`
`2084, RCSB Protein Databank, Molecular Machinery: A Tour of the Protein Data
`
`Bank, http://cdn.rcsb.org/pdb101/learn/resources/2014-mol-mach-poster.pdf (last
`
`visited May 10, 2016) (illustrating the diversity of proteins, with an antibody
`
`shown as protein 13 and insulin as protein 16).)
`
`B.
`
`Antibody Production Techniques As Of April 1983
`Polyclonal Antibodies: As of April 1983, it was well known
`46.
`
`that antibodies could be produced by immunizing an animal with a foreign antigen,
`
`and then recovering the antibodies that had been produced by the animal. Under
`
`this approach, a “polyclonal” serum is generated, i.e., a mixture of antibodies with
`
`varying specificities.
`
`47. As of April 1983, polyclonal antibodies were being widely used
`
`to study the structure and function of antibodies. Due to their varying specificities,
`
`however, polyclonal antibodies had limited usefulness for therapeutic and non-
`
`therapeutic applications. (Ex. 1001 at 1:51-63, 2:40-43.)
`
`48. Hybridomas: In the 1970s, Drs. Georges Köhler and César
`
`Milstein pioneered a technique for producing “monoclonal” antibodies, i.e.,
`
`antibodies that have the same amino acid sequence and bind to the same location
`
`on the antigen (called an epitope) in the same way. Their technique involved the
`
`use of fused cells known as “hybridomas.” (See Ex. 2013, G. Kohler & C.
`
`16
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`Milstein, Continuous Cultures of Fused Cells Secreting Antibody of Predefined
`
`Specificity, Nature 256:495-497 (Aug. 7, 1975).)
`
`49.
`
`Like the production of polyclonal antibodies, the production of
`
`a hybridoma starts with the immunization of an animal with a particular antigen of
`
`interest. B cells from the animal are then isolated and fused to an immortalized
`
`cell, called a myeloma, which are blood cell-derived cancer cells that have the
`
`ability to grow indefinitely in a cell culture. The fused cells, i.e., hybridomas,
`
`grow continuously in culture and produce the desired antibody.
`
`50.
`
`In the early 1980s, the hybridoma technique was being widely
`
`used to produce monoclonal antibodies, and researchers were focusing on
`
`expanding their use even further. (Ex. 1039, Milstein, Monoclonal Antibodies
`
`from Hybrid Myelomas, Proceedings of the Royal Society of London, 211:393-412,
`
`407 (1981) (“use of hybrid myelomas to define the complete repertoire of
`
`antibodies to single antigens . . . is now expanding very rapidly” and “many
`
`commercial companies are beginning to market them”); Ex. 2020, Foote Dep. 37,
`
`48 (describing hybridoma technique as a “very big” deal in the early 1980s due to
`
`“significant achievements”); Ex. 1001 at 1:64-2:11.)
`
`17
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`C.
`
`Use of Recombinant Gene Expression To Produce Proteins
`1.
`Basic principles of recombinant gene expression
`
`51. Recombinant gene expression is a method that allows for the
`
`production and isolation of a protein of interest in a foreign, i.e., “heterologous”
`
`host organism, usually a cell or “host cell.”
`
`52. As explained in the Cabilly ’415 patent, this process involves
`
`four fundamental steps: (1) identification and isolation of a particular gene of
`
`interest (Ex. 1001 at 4:10-12), (2) insertion of that DNA sequence into a vector or
`
`plasmid (id. at 4:17-21), (3) insertion of that vector or plasmid into a suitable host
`
`cell (id. at 4:20), and (4) expression of the sequence, i.e., transcribing the gene of
`
`interest into mRNA, and then translating the mRNA into a polypeptide, by the host
`
`cell. (Id. at 4:23-29.) Host cells transformed with the vector are grown in culture
`
`to express the gene of interest in high quantities and the resulting protein is then
`
`isolated. (Id. at 4:21-24.)
`
`53. Vectors: A vector, comprised of DNA, includes a number of
`
`components to assist with the insertion and expression of the DNA sequence of
`
`interest. For example, a vector will generally include a number of “restriction
`
`enzyme sites,” which are locations in DNA that can be cut by certain proteins
`
`known as restriction enzymes. Restriction enzyme sites allow for the insertion of
`
`the DNA sequence of interest into the vector. (E.g., Ex. 2085, Vectors: A survey
`
`18
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`of molecular cloning vectors and their uses, Chapter 1: The Plasmid pBR322,
`
`Rodriguez, R.L. and Denhardt, D.T. eds. 1988).)
`
`54.
`
`Promoters/Terminators: A vector also includes regulatory
`
`sequences such as promoters and terminators to signal the initiation and
`
`termination of transcription. (Ex. 1001 at 8:57-9:15, 9:56-10:3.) A promoter is a
`
`specific DNA sequence that signals where RNA polymerase should bind the
`
`template DNA and where mRNA synthesis should begin. A transcription
`
`terminator is a specific DNA sequence that signals where mRNA synthesis should
`
`end.
`
`55. Markers: A vector or plasmid will also often include a
`
`“marker” (also called a “marker gene”) that allows for the identification and
`
`isolation of host cells that have taken up the vector. A “selectable” marker (usually
`
`based on antibiotic resistance) is used to select only the bacterial cells that have
`
`taken up the vector. The antibiotic will kill all of the other cells that do not have
`
`the vector. A “reporter” marker does not provide selection, but provides the ability
`
`to distinguish between cells that do or do not contain the marker. For example, a
`
`particular marker might turn cells containing the reporter a particular color in the
`
`presence of an indicator dye. (Id. at 8:11-15.)
`
`56. A marker is different from a gene of interest, i.e., the gene
`
`encoding for the protein sought to be expressed, as the marker is not intended to be
`
`19
`
`

`
`Case No. IPR2015-01624
`Declaration of John Fiddes, Ph.D.
`
`produced in large amounts, isolated, or studied. Rather, the only function of a
`
`marker is to select or identi