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Filed on behalf of Senior Party

THE REGENTS OF THE UNIVERSITY OF CALIFORNIA,


UNIVERSITY OF VIENNA, AND EMMANUELLE CHARPENTIER
By:

Todd R. Walters, Esq.


Erin M. Dunston, Esq.
Travis W. Bliss, Ph.D., Esq.
Christopher L. North, Ph.D., Esq.
Buchanan Ingersoll & Rooney PC
1737 King Street, Suite 500
Alexandria, Virginia 22314-2727
Telephone (703) 836-6620
Facsimile (703) 836-2021
todd.walters@bipc.com
erin.dunston@bipc.com
travis.bliss@bipc.com
christopher.north@bipc.com

By:

Li-Hsien Rin-Laures, M.D., Esq.


Sandip H. Patel, Esq.
Greta Noland
Marshall Gerstein & Borun LLP
6300 Willis Tower
233 South Wacker Drive
Chicago, Illinois 60606
Telephone (312) 474-6300
Facsimile (312) 474-0448
lrinlaures@marshallip.com
spatel@marshallip.com
gnoland@marshallip.com

UNITED STATES PATENT AND TRADEMARK OFFICE


____________________
BEFORE THE PATENT TRIAL AND APPEAL BOARD
____________________
THE BROAD INSTITUTE, INC., MASSACHUSETTS INSTITUTE OF
TECHNOLOGY, and PRESIDENT AND FELLOWS OF HARVARD COLLEGE
Patents 8,697,359; 8,771,945; 8,795,965; 8,865,406; 8,871,445; 8,889,356;
8,895,308; 8,906,616; 8,932,814; 8,945,839; 8,993,233; 8,999,641; and Application 14/704,551,
Junior Party,
v.
THE REGENTS OF THE UNIVERSITY OF CALIFORNIA, UNIVERSITY
OF VIENNA, AND EMMANUELLE CHARPENTIER,
Application 13/842,859,
Senior Party.
____________________
Patent Interference 106,048 (DK)
____________________
UC et al. REPLY 4
(Seeking Benefit of Each of Senior Partys Three Provisional Applications for Count 1)

Interference No. 106,048


TABLE OF CONTENTS
Page
I.

INTRODUCTION ...............................................................................................................1

II.

THE EVIDENCE .................................................................................................................1

III.

STATEMENT OF MATERIAL FACTS.............................................................................1

IV.

ARGUMENT .......................................................................................................................1

V.

A.

Benefit Does Not Require a Working Example or an Actual Reduction


to Practice................................................................................................................ 1

B.

Broads Predictability Arguments Are Unavailing ................................................. 2


1.

Broads Incorrect Reading and Application of the Law ............................. 2

2.

Broads Allegation of Unpredictability Lacks Evidentiary Support........... 4

C.

Broad Ignores UCs Extensive Support for the Embodiments of Count 1 ............. 5

D.

Broads Additional Arguments Are Unavailing ..................................................... 7


1.

Broad Mischaracterizes the Early 2013 Art ................................................ 7

2.

Broad Mischaracterizes UCs Positive Results......................................... 10

3.

Broads PAM Arguments Are Unfounded ............................................... 11

4.

Broads Remaining Attacks Are Not Relevant to the Benefit Inquiry ..... 14

CONCLUSION ..................................................................................................................16

APPENDIX 1 - LIST OF EXHIBITS


APPENDIX 2 STATEMENT OF FACTS

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Interference No. 106,048


TABLE OF AUTHORITIES
Cases

Page(s)

Ariad Pharm., Inc. v. Eli Lilly & Co.,


598 F.3d 1336 (Fed. Cir. 2010)............................................................................................3
In re Borkowski,
422 F.2d 904 (C.C.P.A. 1970) .............................................................................................2
Capon v. Eshhar,
418 F.3d 1349 (Fed. Cir. 2005)..........................................................................................13
Falko-Gunter Falkner v. Inglis,
448 F.3d 1357 (Fed. Cir. 2006)............................................................................................2
Meitznzer v. Mindick,
549 F.2d 775 (C.C.P.A. 1977) .............................................................................................9
In re Wands,
858 F.2d 731 (Fed. Cir. 1998)..............................................................................................3
Rules
37 C.F.R. 41.158(a).......................................................................................................................9

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I.

INTRODUCTION
Senior Party (UC) Motion 4 points to several embodiments within the scope of Count 1

that are both described and enabled. Contrary to Broads assertions, nothing more is required

not working examples and not an actual reduction to practice. To obtain benefit, UC only

needed to show that its Provisionals evidence possession of at least one embodiment and that one

of ordinary skill in the art could carry out that embodiment. UC has satisfied that standard.

Apparently aware that it cannot attack the merits of UCs showing, Broad misstates UCs relied-

upon disclosures, attempts to add non-existent elements to Count 1, and offers immaterial

arguments and scientific theories. Such straw men warrant little attention.

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II.

11
12

THE EVIDENCE
A list of exhibits upon which this Reply relies is set forth in Appendix 1.

III.

13

STATEMENT OF MATERIAL FACTS


Material Facts 1-80 in support of UC Motion 4, additional Material Facts 81-188 alleged

14

in Broad Opposition 4, and the corresponding responses are repeated in Appendix 2.

15

IV.

16
17

ARGUMENT
Broads Opposition 4 (BO4) fails to rebut UCs showing that each of its Provisionals

provides a constructive reduction to practice of embodiments within Count 1.

18
19

A.

20

At page 2, lines 19-24; page 4, lines 14-17; page 5, line 25 to page 6, line 1; page 14,

Benefit Does Not Require a Working Example or an Actual Reduction to


Practice

21

lines 18-20; page 15, lines 9-10; page 16, lines 4-5; and page 23, lines 9-11 of BO4, it is argued

22

that UC does not deserve benefit to its First and Second Provisionals because they do not

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disclose a working example or an actual reduction to practice in eukaryotic cells. The response

24

is the law does not require disclosure of either.

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Broad fails to cite any legal source requiring a working example or actual reduction to

practice. BO4, at 4:19-5:18. And for good reason there is no such requirement under the law

of written description or enablement. See, e.g., In re Borkowski, 422 F.2d 904, 908 (C.C.P.A.

1970) ([A] specification need not contain a working example if the invention is otherwise

disclosed in such a manner that one skilled in the art will be able to practice it without an undue

amount of experimentation.); Falko-Gunter Falkner v. Inglis, 448 F.3d 1357, 1366 (Fed. Cir.

2006) ([E]xamples are not necessary to support the adequacy of a written description . . . [and]

the written description standard may be met . . . even where actual reduction to practice of an

invention is absent). Thus, Broads arguments based on a lack of a working example or an

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actual reduction to practice in eukaryotic cells are contrary to the law and should be disregarded.

11

B.

12

Broads arguments rely upon an alleged unpredictability in the art, which is based on an

13

incorrect reading and application of the law and an improper analysis of the art. When a proper

14

analysis is performed, it is clear that the art relating to the transfer of prokaryotic systems to

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eukaryotic cells was not unpredictable.

16
17

Broads Predictability Arguments Are Unavailing

1. Broads Incorrect Reading and Application of the Law


At page 2, line 22; page 5, lines 23-25; page 26, lines 13-14; and page 26, line 21 to page

18

27, line 1 of BO4, it is argued that the relevant field of art was nascent and unpredictable.

19

The response is that, while the Type II CRISPR-Cas cleavage system was newly elucidated, the

20

relevant aspect for the inquiry at hand transitioning a prokaryotic system to a eukaryotic cell

21

was not a nascent and unpredictable field, but rather had been successfully accomplished for

22

decades using techniques that were so well known as to be commonplace as of the First

23

Provisionals filing date.

24

Predictability in the art is one of the Wands factors that is taken into account when
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analyzing enablement. In re Wands, 858 F.2d 731, 737 (Fed. Cir. 1988). Similarly, the level of

detail required to satisfy the written description requirement varies depending on the nature and

scope of the claims and on the complexity and predictability of the relevant technology. Ariad

Pharm., Inc. v. Eli Lilly & Co., 598 F.3d 1336, 1351 (Fed. Cir. 2010). BO4 fails to analyze the

appropriate field of art for this inquiry. For biological subject matter, the Federal Circuit has

stated that the inquiry must be made not into the art in general, but rather the question is specific

for the predictability of the aspect at issue. Ariad, 598 F.3d at 1351 (emphasis added). The

aspect at issue in this benefit inquiry is not elucidation of the necessary and sufficient

components of the Type II CRISPR-Cas cleavage system, but rather application of that fully-

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11

elucidated system to eukaryotic cells. See, e.g., BO4, at 15:9-13, 16:4-6; 16:27-17:6.
By the time UCs First Provisional was filed, persons of ordinary skill in the art had been

12

successfully applying prokaryotic systems, including bacterial nuclease systems, to eukaryotic

13

cells for decades using known techniques. See Facts 42, 48, 50; Ex. 1022, 135-150; Ex.

14

1024, 128-142; Exs. 1161, 1302, 1327, 1329, 1335, 1336, 1502. There was nothing nascent

15

and unpredictable about the aspect at issue it was conventional and commonplace. Id.

16

Further, given this extensive knowledge, the Federal Circuits other factors for evaluating the

17

adequacy of the disclosure also weigh in favor of granting UCs motion. Ariad, 598 F.3d at

18

1351 ([W]e have set forth a number of factors for evaluating the adequacy of the disclosure,

19

including the existing knowledge in the particular field, the extent and content of the prior art,

20

the maturity of the science or technology, and the predictability of the aspect at issue.) (internal

21

citations omitted). Broads unpredictability arguments are therefore unavailing.

22
23

This wealth of existing knowledge and maturity of the technology is precisely why at
least seven groups were able to successfully use the Type II CRISPR-Cas system in eukaryotic

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cells, draft manuscripts, and have them peer reviewed and published all within a year of the

publication of Jinek 2012. See Facts 60-64, 76; Ex. 1022, 158-162; Ex. 1024, 150-154;

Exs. 1055-1060, 1371, 1372. The only aspect of Count 1 that was not previously known was the

Type II CRISPR-Cas cleavage system, but the UC inventors fully described and utilized that

system in working examples in each of UCs Provisionals. See, e.g., Ex. 1003, 00248-00252;

Ex. 1022, 342-349; Ex. 1024, 333-340.

2. Broads Allegation of Unpredictability Lacks Evidentiary Support

At page 20, line 17-page 23, line 11 of BO4, it is argued that skilled persons would have

recognized that an in vitro environment is very different from the environment of eukaryotic

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cells, and that various theoretical impediments (molecular crowding; temperature, pH, and ion

11

concentration differences; possible transcription or translation difficulties of prokaryotic proteins

12

in eukaryotic cells; possible interference by eukaryotic proteins; possible toxicity of the

13

prokaryotic components to the eukaryotic cell; and possible issues arising from the increased

14

complexity of eukaryotic cells) exist that could negatively impact the ability to move a

15

prokaryotic system into eukaryotic cells. The response is that neither Broad nor its witnesses

16

points to even one example of a prokaryotic system that could not be utilized in a eukaryotic cell

17

due to any of these theoretical impediments. See BO4, at 21:3-22:19; Facts 135-149; Ex. 2009,

18

11.28-11.31, 10.5, 10.7, 11.88.

19

Drs. Breaker and Simons admitted that they could not identify any prokaryotic system

20

that could not be applied to eukaryotic cells due to these theoretical impediments. See Ex. 1555,

21

at 179:8-180:9, 182:16-184:7, 193:17-194:4; Ex. 1638, at 122:11-17. Persons of ordinary skill

22

in the art would not have considered these theoretical impediments to be practical impediments,

23

and the rapid success of other independent research groups is objective evidence that these

24

theoretical impediments: (1) did not hinder transitioning the Type II CRISPR-Cas system to
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eukaryotic cells, and (2) were not considered sufficiently likely to cause persons of ordinary skill

in the art to lack a reasonable expectation of success. Ex. Facts 63, 64, 76; 1534, 75-77, 93-

109; Ex. 1535, 75-77, 93-109; Exs. 1055-1060, 1371, 1372.

At page 23, line 12 to page 24, line 9 of BO4, it is argued that targetrons, also known as

Group II introns, provide an example of a prokaryotic system that was difficult to apply to

eukaryotic cells and that [t]he history of prior failed attempts to transfer prokaryotic systems to

eukaryotes would have increased the insistence of persons of ordinary skill to see actual

experimental results in eukaryotic cells. The response is that targetrons were, as admitted by

Drs. Simons and Breaker, successfully utilized in eukaryotic cells many years before UCs First

10

Provisional was filed. See, e.g., Ex. 1534, 83; Ex. 1535, 83; Ex. 1556, at 217:15-218:2; Ex.

11

2010, 1.49 (citing Exs. 1293, 2274).

12

C.

13

At page 1, lines 13-15 of BO4, it is argued that Senior Party relies [only] on an in vitro

Broad Ignores UCs Extensive Support for the Embodiments of Count 1

14

experiment in combination with the general disclosure that the system could be used in any cells

15

of interest, including eukaryotic cells. The response is that UC provided ample support to

16

demonstrate that it is entitled to benefit of its Provisionals.

17

In addition to the successful in vitro experiment and a discussion of the cell types in

18

which the Type II CRISPR-Cas system could be used, which is alone sufficient, UC also cited

19

disclosure in its Provisionals explaining how to provide the Cas9 protein and/or DNA-targeting

20

RNA in the form of a nucleic acid encoding those components, which can be included in a

21

vector, and which can be operably linked to a control element, such as a promoter functional in a

22

eukaryotic cell. See UC Motion 4, at 15:5-21 (citing, inter alia, Ex. 1003, 00120-00123,

23

00126, 00127); Facts 37-40. UC also cited disclosures in its Provisionals explaining how one

24

can introduce those vectors into eukaryotic cells using techniques that had been well-known for
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decades. See UC Motion 4, at 15:22- 16:7 (citing, inter alia, Ex. 1003, 00121, 00129); Facts

41, 42. UC also cited additional disclosures in its Provisionals of conventional molecular

biology techniques that can be used in applying the Type II CRISPR-Cas system in eukaryotic

cells, such as the attachment of protein transduction domains or nuclear localization signals

(NLSs) to the Cas9 protein and the use of codon optimization. See UC Motion 4, at 16:8-17:6

(citing, inter alia, Ex. 1003, 0033, 00115, 00124, 00179); Facts 43-47.

At page 15, lines 11-13; page 16, lines 5-6; page 26, lines 10-13; and page 26, lines 17-18

of BO4, it is argued that UC failed to identify structural modifications or other adaptations

necessary for practicing the Type II CRISPR-Cas system in eukaryotic cells. The response is

10

that Broad has not identified any structural modifications or adaptations that are necessary to

11

move the system into eukaryotic cells because no such necessary adaptations exist. As a further

12

response, Broad again ignores the numerous cited passages in UCs Provisionals that explain

13

which conventional techniques could be used and that indeed were used to apply the Type II

14

CRISPR-Cas system to eukaryotic cells.

15

Broad fails to identify any structural modifications or other adaptations that are actually

16

required for the Type II CRISPR-Cas system to operate in a eukaryotic cell because there are

17

none. See, e.g., Ex. 1022, 161-162, 416, 417; Ex. 1024, 153-154, 407, 408; Exs. 1012,

18

1058-1060. None of codon optimization, vectors, NLSs, and a longer tracrRNA sequence is

19

required to achieve cleavage of target DNA in eukaryotic cells. Id. Nonetheless, the UC

20

Provisionals describe all of these optional features. Ex. 1022, 387-404, 422; Ex. 1024,

21

378-395, 413; facts 36-47; see also UC Motion 4, at 15:5-17:6. Broad again ignores the

22

numerous cited passages in UCs Provisionals that explain which conventional techniques could

23

be used and that indeed were used to apply the Type II CRISPR-Cas system to eukaryotic

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cells. For example, UC cited to passages that discuss delivery of nucleic acid vectors encoding

the Cas9 protein and DNA-targeting RNA components, codon optimization of Cas9, and the

attachment of protein transduction domains to the Cas9 protein. See UC Motion 4, at 15:5-17:6

(citing, inter alia, Ex. 1003, 0033, 00115, 00120-00124, 00126, 00127, 00129, 00179); Facts

37-46. Not only are these structural aspect[s] that would make the system adapted for

eukaryotic cells, they are the same structural aspects that Broad and other research groups used

to apply the Type II CRISPR-Cas system to eukaryotic cells. UC Motion 4, at 20:2-5; Fact 62;

Ex. 1022, 423; Ex. 1024, 414; Exs. 1055-1059.

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10
11
12
13

D.

Broads Additional Arguments Are Unavailing

Broads additional arguments rely upon inaccurate characterizations of UCs


Provisionals, Count 1, and the underlying science, or are irrelevant to the issue of benefit.
1. Broad Mischaracterizes the Early 2013 Art
At page 27, line 5 to page 28, line 5 of BO4, it is argued that [t]he lack of enablement is

14

also demonstrated by the failures of other researchers using the CRISPR-Cas9 system with

15

Chimera A using routine techniques, citing specifically to U.S. App. No. 61/734,256 (Chen).

16

The response is that Chen shows that researchers successfully used Chimera A and the Type II

17

CRISPR-Cas system in eukaryotic cells. In Figure 1 of Chen, a Type II CRISPR-Cas system,

18

including a DNA-targeting RNA based on the Chimera A sequence from Jinek 2012, was

19

introduced into eukaryotic cells. Ex. 2009, 11.126-11.128. The level of expression of green

20

fluorescent protein (GFP) within those eukaryotic cells was then used to determine whether the

21

Type II CRISPR-Cas system was functioning. Id. As Dr. Simons admitted, Chen specifically

22

indicates that [t]he percent GFP detected in each of the four experimental treatments (A-D) was

23

greater than in the control treatments (E,F). Ex. 2009, 11.128 (quoting Ex. 2125, 0066). Dr.

24

Carroll agrees. See Ex. 2007, at 183:1-11. The higher percentage of GFP shown in Chen is
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objective evidence that Chen obtained positive results when applying a Type II CRISPR-Cas

system with a Chimera A-based DNA-targeting RNA to eukaryotic cells.

At page 26, lines 18-20 of BO4, it is argued that adapting Chimera A to function in

eukaryotic cells required extensive experimentation. The response is that there is no credible

evidence to support Broads assertion. While Broad relies upon a quote from Dr. Doudna that

her group encountered many frustrations when moving forward with the technology, Broad

fails to cite any evidence that these frustrations pertained to transitioning the system to

eukaryotic cells. BO4, at 26:18-20. In fact, numerous independent research groups moved the

system into eukaryotic cells very rapidly using only well-known, conventional molecular biology

10

techniques, and Broad fails to rebut this uncontroverted objective evidence. UC Motion 4, at

11

19:15-20:23; Fact 62; Ex. 1022, 158-162; Ex. 1024, 150-154; Ex. 1534, 75-76; Ex.

12

1535, 75-76; Exs. 1055-1059.

13

At page 7, line 23 to page 8, line 4 and at page 27, lines 13-17 of BO4, it is argued that

14

Broads scientists used special techniques or non-routine techniques when applying the Type

15

II CRISPR-Cas system to eukaryotic cells, including the use of two NLSs, the use of an

16

expression vector, and the use of a tracrRNA that was four nucleotides longer. The response is

17

that the techniques used by Broad are irrelevant to the benefit inquiry. Moreover, Drs. Breaker

18

and Simons admitted that the inclusion of NLSs and the use of expression vectors are not

19

special techniques, but rather these are conventional techniques that were well understood and

20

widely used by persons of ordinary skill in the art when applying a prokaryotic system to

21

eukaryotic cells. Ex. 1555, at 97:19-22, 134:14-20, 146:5-147:2; 147:17-148:16, 149:10-

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149:16, 150:8-13, 152:5-153:21, 168:4-169:11; Ex. 1556, at 238:8-16; Ex. 1638, at 179:24-

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180:2; see also Ex. 1022, 135-150; Ex. 1024, 128-142. Broad fails to present any

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evidence demonstrating that it used a tracrRNA that included an extra four nucleotides. BO4

at 8:3-5; Ex. 2009, 11.107; Ex. 1055, at 821 (Fig. 2B showing a chimeric RNA that has a

tracrRNA component that is identical to the one used in Jinek 2012); Ex. 1639, at 124:24-125:8.

Dr. Simons admitted that he is unaware of anything in Cong 2013 that shows the extra four

nucleotides or that even indicates extra nucleotides were added to the tracrRNA portion of the

chimeric DNA-targeting RNA. Ex. 1639, at 124:8-12, 126:22-127:7. Thus, this argument is

entitled to no weight as it is merely a bald assertion by Broad and Dr. Simons with no

evidentiary support in the underlying Cong 2013 publication or any other reference. See 37

C.F.R. 41.158(a) (Expert testimony that does not disclose the underlying facts or data on

10

which the opinion is based is entitled to little or no weight.); Meitznzer v. Mindick, 549 F.2d

11

775, 782 (C.C.P.A. 1977). Broads argument also lacks merit because UCs Provisionals

12

disclose each of the features to which Broad points (the use of NLSs, expression vectors, and

13

tracrRNA ranging from truncated versions to naturally occurring full length). See e.g. UC

14

Motion 4, at 15:5-17:6; Facts 36-47; Ex. 1003, 0012, 0033, 00115, 00121, Fig. 9; Ex. 1022,

15

387-404, 422; Ex. 1024, 378-395, 413.

16

At page 27, line 19 to page 28, line 1 of BO4, it is argued that the work of Dr. Kims

17

research group, published in Cho 2013 (Ex. 1059), was non-routine because those researchers

18

added 2-15 times more plasmids and RNA to the eukaryotic cells than was recommended by the

19

manufacturer of their nucleofection equipment. The response is that this assertion lacks

20

evidentiary support. Cho 2013 specifically states that the nucleofection procedure was done

21

according to the manufacturers protocol. Ex. 1059, at Supp. Info. p. 3. Broad relies only on

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Dr. Simonss opinions and provides no evidence of a manufacturers protocol from which Dr.

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Kims group allegedly deviated. See BO4, at 27:21-28:1 Ex. 2009, 11.132; Ex. 1059, at 6.

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Dr. Simonss Declaration does not include any citation to a manufacturers protocol and he

admitted that he was unaware of the date or version of the alleged protocol on which he was

relying, and he further admitted that the protocol likely changed over time. Ex. 2009, 11.132-

11.133; Ex. 1639, at 59:2-7, 59:11-21. Dr. Simons also admitted that these studies in fact show

the successful use of the Chimera A construct in eukaryotic cells. Ex. 1639, at 130:2-13.

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8
9

2. Broad Mischaracterizes UCs Positive Results


At page 28, line 21 to page 29, line 22 of BO4, it is argued that the eukaryotic working
examples reported in UCs Third Provisional failed. The response is that Broad is mistaken.
For the results shown in Figure 38B, Broad does not dispute that the lanes from samples

10

containing both DNA-targeting RNA and Cas9 protein (lanes 6, 8, and 9 from the left) possess

11

additional bands that could be cleavage product. Ex. 1005, at Fig. 38B; BO4, at 29:8-15.

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Instead, Broad focuses on the fact that the bands do not appear to be the proper size based on the

13

marker lane (lane 1), and concludes that any cleavage product is the wrong size so the

14

experiment must have failed. BO4, at 29:8-15. Yet, Dr. Simons admitted that the marker lane

15

upon which Broad relies could simply have been mislabeled. Ex. 1638, at 150:19-152:19.

16

Further, Broad ignores the fact that the cleavage product band in the lanes for the Type II

17

CRISPR-Cas samples is within the same size range as the cleavage product bands in the lanes for

18

the ZFN positive control sample. Ex. 1005, at Fig. 38B (compare the lower band in lanes 6, 8,

19

and 9 to the lower bands in lane 10). From this, one of ordinary skill in the art would conclude

20

that the Type II CRISPR-Cas samples had successfully cleaved the target DNA.

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Figure 36E of UCs Third Provisional clearly shows a cleavage product band in the lane

22

containing a DNA-targeting RNA and a Cas9 protein (lane 5) that is within the size range of the

23

cleavage product bands in the ZFN positive control sample (lane 7) and of the proper size

24

according to the marker lane (lane 1). Ex. 1003, at Fig. 36E. This cleavage band is not present
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in any of the negative control lanes. One of ordinary skill in the art would again conclude from

Figure 36E that the Type II CRISPR-Cas sample had successfully cleaved the target DNA. Id.

Furthermore, data presented in Figures 36E and 38B of UCs Third Provisional were also

presented in Jinek 2013. See Ex. 1057, at Figs. 1E and 3B. As Dr. Breaker admitted, Jinek 2013

was peer-reviewed and recommended for publication by persons of ordinary skill in the art, and

he is not aware of any publication criticizing its data. Ex. 1638, at 131:4-6, 131:24-132:6,

153:22-154:6. Further, Drs. Breaker and Simons both admitted that Jinek 2013 has been widely

cited in other publications by persons of ordinary skill in the art. Ex. 1638, at 153:13-16; 154:7-

172:24; Ex. 1639, at 101:18--121:1; see also Exs. 1604-1610, 1621-1627. This provides

10

objective evidence that one of ordinary skill in the art would have viewed and indeed did view

11

the same results presented in Figures 36E and 38B of UCs Third Provisional as successful.

12

3. Broads PAM Arguments Are Unfounded

13

At page 2, lines 4-6 and page 12, lines 6-8 of BO4, it is argued that Count 1 is directed to

14

a target DNA molecule in a eukaryotic cell, which is not a natural target of the Type II CRISPR-

15

Cas system. The response is that Count 1 does not require cleavage of a eukaryotic target or a

16

non-natural target.

17

There is nothing in Count 1 that precludes the cleavage of natural targets of the Type II

18

CRISPR-Cas system (DNA of viruses and plasmids that invade bacteria in nature) within a

19

eukaryotic cell. See, e.g., Ex. 1022, 48; Ex. 1024, 42. One could utilize a Type II CRISPR-

20

Cas system to cleave natural targets of the Type II CRISPR-Cas system within a eukaryotic cell,

21

e.g., by targeting in a eukaryotic cell the same viral or plasmid DNA sequences that the Type II

22

CRISPR-Cas system naturally targets in prokaryotic cells. This plainly falls within the scope of

23

Count 1 and, therefore, Broads arguments should be afforded no weight.

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At page 1, lines 17-18 of BO4, it is argued that UCs First Provisional fails to describe
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eukaryotic-specific considerations such as use of a target sequence comprising a protospacer

adjacent motif (PAM). At page 2, lines 7-13 of BO4, it is argued that it was known in the art by

May of 2012 that PAM sequences played a role in the ability of a CRISPR system in bacteria to

recognize natural DNA targets, and that the lack of discussion in the First Provisional of the

role of PAM would have indicated to persons of ordinary skill in the art that the inventors failed

to describe an invention configured for use against non-natural targets in a eukaryotic cell. At

page 12, lines 8-10 of BO4, it is argued that, [b]ased on the lack of any PAM discussion, skilled

persons would have concluded that the inventors had not begun to provide information needed to

use CRISPR systems with non-natural targets. The response is that Broads arguments about

10
11

the role of PAM in target cleavage are misleading.


First, Count 1 does not require cleavage of non-natural targets. Count 1 also does not

12

require a target sequence with a PAM. See, e.g., Ex. 1639, at 21:1-4. Count 1 only requires that

13

target DNA is cleaved or edited or transcription of at least one gene encoded by the target DNA

14

molecule is modulated. See Count 1. Cleavage of a target DNA encompasses cleavage of

15

both single-stranded and double-stranded target DNA. See, e.g., Ex. 1003, 0042; see also Ex.

16

1639, at 22:22-23:3. The Type II CRISPR-Cas system can target and cleave single-stranded

17

target DNA and such cleavage by Cas9 does not require a PAM sequence. See, e.g., Ex. 1155 at

18

p. 817; see also Ex. 1154 at E2584; Ex. 1639, at 36:20-23, 23:8-24:5, 27:4-9. Thus, Count 1

19

clearly encompasses embodiments that do not require a PAM sequence, so the presence or

20

absence of a PAM discussion in the UC Provisionals has no bearing on the benefit analysis.

21

As Broad admits, the existence of PAM sequences and their role in the Type II CRISPR-

22

Cas system were well-known in the art by May of 2012. See BO4 at 12:11-13; see also Ex. 1639

23

at 37:14-38:14, 39:6-40:20; Ex. 1298 at 736; Ex. 1022, 306-312; Ex. 102, 297-303. As

- 12 -

Interference No. 106,048


1

such, under well-settled law, the existence and role of the PAM sequence did not need to be

specifically recited in the Provisionals. Capon v. Eshhar, 418 F.3d 1349, 1357 (Fed. Cir. 2005).

Further, Broads arguments attempting to confine this knowledge to so-called natural targets

lack merit. Broad fails to provide evidence of any actual differences between natural and

non-natural targets of the Type II CRISPR-Cas system likely because no such differences

exist. See BO4, at 11:22-14:8. The knowledge in the art was not that the PAM sequence plays a

role in cleavage of natural double-stranded targets, as Broad states, but rather that it played a

role in all double-strand cleavage, regardless of whether the target was natural or not. See Ex.

1033, at 9276, 9278-9; Ex. 1022, 225-226, 310; Ex. 1024, 217-218, 301. For example,

10

studies performed by Sapranauskas et al. in 2011 showed that the PAM sequence played the

11

same role in cleavage of a non-natural (recombinantly constructed, man-made plasmid) target as

12

it was previously known to play in the cleavage of natural bacteriophage targets. Id.

13

To the extent that Broad is arguing that the target of the method of Count 1 is non-natural

14

merely because the Type II CRISPR-Cas system and/or target DNA is being introduced into a

15

cell in which it does not naturally occur, Sapranauskas is objective evidence that refutes this

16

argument as it demonstrates that PAM played a role even when an S. thermophilus Type II

17

CRISPR locus was introduced into an E. coli host cell, which does not naturally possess a Type

18

II CRISPR locus, and used to cleave a heterologously introduced target DNA. Id.

19

Thus, the knowledge in the art as of May of 2012 regarding the role of the PAM sequence

20

was not limited to natural targets, but rather it was well known that a PAM sequence played the

21

same role in cleavage of both natural and non-natural targets.

22
23

At page 12, line 18 to page 13, line 4 of BO4, it is argued that the experiment set forth in
Figure 5 of the First Provisional illustrates a failure to appreciate the significance of PAM.

- 13 -

Interference No. 106,048


1

The response is that Broad is clearly mistaken. The DNA-targeting RNA in Figure 5B is from S.

pyogenes, as can be seen by comparing the sequence in Figure 5B to those in Figures 6 and 7.

Ex. 1003, at Figs. 5B, 6, 7. Dr. Simons admitted that the target DNA in Figure 5B included the

appropriate PAM sequence for S. pyogenes. Ex. 1639, at 42:5-43:20; Ex. 1611. If UCs

inventors truly fail[ed] to appreciate the significance of PAM, as Broad argues, they would not

have utilized a target DNA that included the appropriate PAM sequence. Ex. 1003, at Fig. 5B.

At page 13, line 5 to page 14, line 5 of BO4, it is argued that the absence of a label for the

PAM sequence in Figure 1 of the First Provisional shows that UC chose to conceal that

information [about the PAM sequence] for some reason. The response is that this grossly

10

mischaracterizes Figure 1. Broad compares Figure 1A of Jinek 2012 (PAM labeled) and Figure

11

1 of UCs First Provisional (PAM not labeled) and alleges that the figure in the First Provisional

12

had been scrubbed to remove any reference to PAM sequences. BO4, at 13:10-14:5. Broads

13

allegation conveniently ignores the fact that all of the labels from the figure in the First

14

Provisional are different from those in Jinek 2012 and that the two figures were used in different

15

contexts. Compare Ex. 1003, Fig. 1, with Ex. 1155, Fig. 5A. Broad has no evidence to support

16

its allegation that UC chose to conceal the PAM sequences.

17
18

4. Broads Remaining Attacks Are Not Relevant to the Benefit Inquiry


At page 6, lines 19-21 and at page 6, line 24 to page 7, line 14 of BO4, it is argued that

19

Dr. Zhang began his efforts to use CRISPR systems in eukaryotic cells before Jinek 2012

20

published. The response is that Broads alleged priority evidence has nothing to do with UCs

21

request for benefit. To the extent it is considered, the Zhang NIH grant application actually

22

supports UCs assertion that Broad was not able to utilize the Type II CRISPR-Cas system in

23

eukaryotic cells until after Jinek 2012 because it shows that Dr. Zhang did not understand the

24

components of the Type II CRISPR cleavage system Dr. Zhang did not understand the
- 14 -

Interference No. 106,048


1

necessity of tracrRNA for cleavage. See Ex. 2411, at 16.

At page 11, lines 3-12 of BO4, it is argued that [t]he industry has recognized the

pioneering nature of Dr. Zhangs work as evidenced by the receipt of one award by Dr. Zhang

for his application of the Type II CRISPR-Cas system to eukaryotic cells. The response is that

this is again irrelevant to the issue of benefit. To the extent it is considered, UC notes that Broad

conveniently omits from its discussion that the award Dr. Zhang shared with Drs. Doudna and

Charpentier was for development of CRISPR-Cas as a genome editing tool for eukaryotes. Ex.

2419, p. 2 (emphasis added). Further, as Dr. Simons admitted, Drs. Doudna and Charpentier

have received numerous other prizes for their contributions to the CRISPR gene-editing field.

10

See Ex. 1639, at 143:22-155:23; see also Exs. 1629-1635. Thus, the eukaryotic work of Drs.

11

Doudna and Charpentier has been as celebrated, if not more celebrated, as that of Dr. Zhang. Id.

12

At page 16, line 7 to page 17, line 2 of BO4, it is argued that the rejection of the

13

Sontheimer patent application is relevant to whether UCs Provisionals disclose an embodiment

14

within Count 1. The response is that the Patent Offices rejection of an unrelated application

15

regarding different claimed subject matter is not relevant to the benefit issue. Broad suggests

16

that the Patent Office rejected Sontheimer for lack of written description and enablement based

17

on the absence in that application of a single working example, and analogizes that to UCs

18

Provisionals by stating that the Doudna P1 and P2 applications are entirely prophetic. BO4, at

19

16:15-27. Broads analogy lacks merit. In contrast to Broads characterization of Sontheimer,

20

the UC Provisionals include working examples of the Type II CRISPR-Cas system cleaving a

21

target DNA outside of the prokaryotic cellular environment. See, e.g., Ex. 1003, 00248-

22

00252; Facts 10-21; UC Motion 4, at 4:11-9:16. Those examples teach one of ordinary skill in

23

the art the components that are necessary and sufficient to operate the system in a heterologous

- 15 -

Interference No. 106,048


1

environment, which includes in vitro and eukaryotic cellular environments. Id.

At page 18, line 5 to page 20, line 16 of BO4, it is argued that Jinek 2012 is not sufficient

to demonstrate that the inventors were in possession of the invention of Count 1. The response is

that it is each of UCs Provisionals that must describe and enable an embodiment within the

scope of Count 1, not Jinek 2012. As a further response, Broad mischaracterizes UCs

discussion of Jinek 2012 by alleging that UC asserts that the Jinek 2012 paper is a proxy for

Doudna P1 and P2, because it contains the same in vitro tests set forth in Doudna P1 and P2 with

the same general disclosure of eukaryotic cells. BO4, at 18:5-8. In reality, UCs Motion states

that Jinek 2012 contains much of the same information as the First Provisional, albeit with

10

much less detail about how one would apply the system in other organisms. UC Motion 4, at

11

18:14-16 (emphasis added). As explained in UC Motion 4, even this far less detailed disclosure

12

was sufficient to allow multiple research groups to rapidly move the system into eukaryotic cells.

13

See id., at 19:13-20:23

14

V.

15

CONCLUSION
The absence in BO4 of any rebuttal to UCs positive benefit case is telling. Broads

16

arguments all rely upon an improper legal standard for written description and enablement and

17

mischaracterizations of both the record evidence and the knowledge in the art. Because each of

18

UCs Provisionals describes and enables an embodiment within the scope of Count 1, UC

19

Motion 4 should be granted.

20

Respectfully submitted,
By /Todd R. Walters/
Todd R. Walters, Esq. (Reg. No. 34,040)
BUCHANAN INGERSOLL & ROONEY PC
1737 King Street, Suite 500
Alexandria, Virginia 22314
todd.walters@bipc.com
Counsel for UC and Vienna
Date: September 28, 2016

By: / Sandip H. Patel /


Sandip H. Patel, Esq. (Reg. No. 43,848)
MARSHALL GERSTEIN & BORUN LLP
6300 Willis Tower, 233 South Wacker Dr.
Chicago, Illinois 60606
spatel@marshallip.com
Counsel for EC
Date: September 28, 2016
- 16 -

Interference No. 106,048


APPENDIX 1 LIST OF EXHIBITS
EXHIBIT

DESCRIPTION

NO.
1001

U.S. Patent Application No. 13/842,859, filed on March 15, 2013, to Jennifer
Doudna et al. (the 859 Application)

1002

U.S. Patent Application Publication No. 2014/0068797, published on March 6,


2014, to Jennifer Doudna et al. (the 797 Publication)

1003

U.S. Provisional Application No. 61/652,086, filed on May 25, 2012, to


Martin Jinek et al. (the 086 Provisional or the First 086 Provisional)

1004

U.S. Provisional Application No. 61/716,256, filed on October 19, 2012, to


Martin Jinek et al. (the 256 Provisional or the Second 256 Provisional)

1005

U.S. Provisional Application No. 61/757,640, filed on January 28, 2013, to


Martin Jinek et al. (the 640 Provisional or the Third 640 Provisional)

1007

U.S. Patent No. 8,697,359, issued on April 15, 2014, to Feng Zhang (the 359
Patent)

1011

U.S. Patent No. 8,932,814, issued on January 13, 2015, to Le Cong and Feng
Zhang (the 814 Patent)

1012

U.S. Patent No. 8,795,965, issued on August 5, 2014, to Feng Zhang (the
965 Patent)

1013

U.S. Patent No. 8,871,445, issued on October 28, 2014, to Le Cong and Feng
Zhang (the 445 Patent)

1-1

Interference No. 106,048


APPENDIX 1 LIST OF EXHIBITS
EXHIBIT

DESCRIPTION

NO.
1020

Diagram depicting the DNA-Binding Domain and DNA-Cleaving Domains of


two ZFNs, http://www.sigmaaldrich.com/life-science/zinc-finger-nucleasetechnology/learning-center/what-is-zfn.html (downloaded on Feb. 3, 2015)

1022

Declaration of Carol Greider, Ph.D.

1023

Curriculum Vitae of Carol Greider, Ph.D.

1024

Declaration of Dana Carroll, Ph.D.

1025

Curriculum Vitae of Dana Carroll, Ph.D.

1029

Dai et al., The Transcription Factors GATA4 and dHAND Physically Interact
to Synergistically Activate Cardiac Gene Expression Through a p300dependent Mechanism, 277(27) J. BIOL. CHEM. 24390-24398 (2002) (Dai)

1030

Gustafsson et al., Codon Bias and Heterologous Protein Expression, 22(7)


TRENDS BIOTECHNOL. 346-353 (2004) (Gustafsson)

1031

43 METHODS IN CELL BIOLOGY, Protein Expression In Animal Cells, Chapters


2, 3, 6, 9 (Michael G. Roth ed., 1994) (Roth)

1032

Deltcheva et al., CRISPR RNA maturation by trans-encoded small RNA and


host factor RNase III, 471 NATURE 602-607 (2011) with Supplementary
Materials (Deltcheva)

1033

Sapranauskas et al., The Streptococcus thermophilus CRISPR/Cas system


provides immunity in Escherichia coli, 39(21) NUCL. ACIDS RES. 9275-9282
(2011) (Sapranauskas)

1-2

Interference No. 106,048


APPENDIX 1 LIST OF EXHIBITS
EXHIBIT

DESCRIPTION

NO.
1035

Park et al., Regulation of Ribosomal S6 Kinase 2 by Mammalian Target of


Rapamycin, 277(35) J. BIOL. CHEM. 31423-31429 (2002) (Park)

1039

Boden et al., Efficient Gene Transfer of HIV-1-Specific Short Hairpin RNA


into Human Lymphocytic Cells Using Recombinant Adeno-associated Virus
Vectors, 9(3) MOL. THER. 396-402 (2004) (Boden)

1040

MOLECULAR CLONING: A LABORATORY MANUAL, Chpt. 16 (J. Sambrook & D.


Russell, 3rd ed. 2001) (Sambrook)

1055

Cong et al., Multiplex Genome Engineering Using CRISPR/Cas Systems,


339(6121) SCIENCE 819-823 (2013) with Supplemental Material (Cong)

1056

Mali et al., RNA-Guided Human Genome Engineering via Cas9, 339(6121)


SCIENCE 823-826 (2013) with Supplemental Materials (Mali)

1057

Jinek et al., RNA-Programmed Genome Editing in Human Cells, 2 ELIFE


e00471 (2013) (Jinek 2013)

1058

Hwang et al., Efficient In Vivo Genome Editing Using RNA-Guided Nucleases,


31(3) NAT. BIOTECHNOL. 227-229 (2013) (Hwang)

1059

Cho et al., Targeted Genome Engineering in Human Cells With the Cas9
RNA-Guided Endonuclease, 31(3) NAT. BIOTECHNOL. 230-232 (2013) with
Supplemental Information (Cho)

1060

Shen et al., Generation of gene-modified mice via Cas9/RNA-mediated gene


targeting, 23(5) CELL RES. 720-723 (2013) (Shen)

1-3

Interference No. 106,048


APPENDIX 1 LIST OF EXHIBITS
EXHIBIT

DESCRIPTION

NO.
1125

Diagram depicting the DNA-Binding Domains (an array of TAL effector


subunits) and DNA-Cleaving Domains of two TALENs,
http://www.systembio.com/services_tales (downloaded on Feb. 3, 2015)

1126

Nam et al., Cas5d Protein Processes Pre-crRNA and Assembles into a


Cascade-like Interference Complex in Subtype I-C/Dvulg CRISPR-Cas
System, 20 STRUCTURE 1574-1584 (2012) (Nam)

1152

Carroll, A CRISPR approach to gene targeting, 20(9) MOLECULAR THERAPY


1658-60 (2012)

1153

Garneau et al., The CRISPR/Cas bacterial immune system cleaves


bacteriophage and plasmid DNA, 468(7320) NATURE 67-71 (2010)

1154

Gasiunas et al., Cas9-crRNA ribonucleoprotein complex mediates specific


DNA cleavage for adaptive immunity in bacteria, 109(39) PNAS e2579-86
(2012)

1155

Jinek et al., A programmable dual-RNA-guided DNA endonuclease in adaptive


bacterial immunity, 337(6096) SCIENCE 816-821 (2012) (Jinek 2012)

1156

Makarova et al., Evolution and classification of the CRISPR-Cas systems, 9(6)


NAT. REV. MICROBIOL. 467-477 (2011)

1161

U.S. Patent Publication No. 2010/0076057, published on March 25, 2010 to


Sontheimer et al.

1-4

Interference No. 106,048


APPENDIX 1 LIST OF EXHIBITS
EXHIBIT

DESCRIPTION

NO.
1194

Anderson et al., A simple method for the rapid generation of recombinant


adenovirus vectors, 7 GENE THERAPY 1034-1038 (2000)

1199

Barrangou et al., CRISPR provides acquired resistance against viruses in


prokaryotes, 315(5819) SCIENCE 1709-1712 (2007)

1200

Behr et al., Efficient gene transfer into mammalian primary endocrine cells
with lipopolyamine-coated DNA, 86 PNAS 6982-6986 (1989)

1203

Bergemann et al., Excision of specific DNA-sequences from integrated


retroviral vectors via site-specific recombination, 23(21) NUCL. ACIDS RES.
4451-4456 (1995)

1208

Bolotin et al., Clustered regularly interspaced short palindrome repeats


(CRISPRs) have spacers of extrachromosomal origin, 151(Pt 8)
MICROBIOLOGY 2551-2561 (2005)

1211

Brouns et al., Small CRISPR RNAs guide antiviral defense in prokaryotes, 321
SCIENCE 960-964 (2008)

1213

Carney and Morgan, Induction of DNA Double-Strand Breaks by


Electroporation of Restriction Enzymes into Mammalian Cells, 113 METHODS
IN MOL. BIOL.

1214

465-471 (1999)

Carroll, Genome Engineering With Zinc-Finger Nucleases, 188 GENETICS


773-782 (2011)

1-5

Interference No. 106,048


APPENDIX 1 LIST OF EXHIBITS
EXHIBIT

DESCRIPTION

NO.
1217

Chiu et al., Engineered GFP as a vital reporter in plants, 6(3) CURR. BIO.
325-330 (1996)

1218

Choulika, et al., Transfer of single gene-containing long terminal repeats into


the genome of mammalian cells by a retroviral vector carrying the cre gene
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1225

Davis and Cui, Zinc Finger Nucleases for Genome Editing, 30(13) GEN. ENG.
& BIOTECH NEWS 1-3 (2010)

1227

Deveau et al., Phage Response to CRISPR-Encoded Resistance in


Streptococcus thermophiles, 190(4) J BACTERIOL 1390-1400 (2008)
(Deveau)

1229

Dykxhoorn et al., Killing the Messenger: Short RNAs That Silence Gene
Expression, 4 NATURE REV. 457-467 (2003)

1235

Fieck et al., Modifications of the E.coli Lac repressor for expression in


eukaryotic cells: effects of nuclear signal sequences on protein activity and
nuclear accumulation, 20(7) NUCL. ACIDS RES. 1785-1791 (1992)

1236

Fischer-Fantuzzi and Vesco, Cell-Dependent Efficiency of Reiterated Nuclear


Signals in a Mutant Simian Virus 40 Oncoprotein Targeted to the Nucleus,
8(12) MOL. CELL BIOL. 5495-5503 (1988)

1237

Foecking and Hofstetter, Powerful and versatile enhancer-promoter unit for


mammalian expression vectors, 45(1) GENE 101-105 (1986)

1-6

Interference No. 106,048


APPENDIX 1 LIST OF EXHIBITS
EXHIBIT

DESCRIPTION

NO.
1248

Gorman et al., High efficiency gene transfer into mammalian cells, B307
PHIL. TRANS. R. SEC. LAND. 343-346 (1984)

1254

Haft et al., A guild of 45 CRISPR-associated (Cas) protein families and


multiple CRISPR/Cas subtypes exist in prokaryotic genomes, 1(6) PLOS
COMPUTATIONAL BIOLOGY 477-483 (2005)

1255

Hale et al., RNA-guided RNA cleavage by a CRISPR RNA-Cas protein


complex, 139(5) CELL 945-956 (2009) (Hale 2009)

1260

Hashimoto et al., A novel method for transformation of intact yeast cells by


electroinjection of plasmid DNA, 21 APPLIED MICROBIOL. BIOTECHNOL. 336339 (1985)

1261

Hatoum-Aslan et al., Mature clustered, regularly interspaced, short


palindromic repeats RNA (crRNA) length is measured by a ruler mechanism
anchored at the precursor processing site, 108(52) PNAS 21218-21222
(2011)

1262

Haurwitz et al., Sequence- and structure-specific RNA processing by a


CRISPR endonuclease, 329 SCIENCE 1355-1358 (2010)

1270

Ishino et al., Nucleotide sequence of the iap gene, responsible for alkaline
phosphates isozyme conversion in Escherichia coli, and identification of the
gene product, 169(12) J. BACTERIOL. 5429-5433 (1987)

1-7

Interference No. 106,048


APPENDIX 1 LIST OF EXHIBITS
EXHIBIT

DESCRIPTION

NO.
1271

Jansen et al., Identification of genes that are associated with DNA repeats in
prokaryotes, 43(6) MOLECULAR MICROBIOLOGY 1565-1575 (2002)

1283

Li et al., In vivo genome editing restores haemostasis in a mouse model of


haemophilia, 475 NATURE 217-223 (2011)

1285

Lombardo et al., Gene editing in human stem cells using zinc finger nucleases
and integrase-defective lentiviral vector delivery, 25(11) NAT. BIOTECHNOL.
1298-1306 (2007)

1288

Makarova et al., A putative RNA-interference-based immune system in


prokaryotes: computational analysis of the predicted enzymatic machinery,
functional analogies with eukaryotic RNAi, and hypothetical mechanisms of
action, 1(7) BIOL. DIRECT. 1-26 (2006)

1293

Mastroianni et al., Group II intron-based gene targeting reactions in


eukaryotes, 3(9) PLOS ONE. e3121 (2008)

1298

Mojica et al., Short motif sequences determine the targets of the prokaryotic
CRISPR defence system, 155(3) MICROBIOLOGY 733-740 (2009) with
Supplementary Data

1299

Mojica et al., Biological significance of a family of regularly spaced repeats in


the genomes of Archaea, Bacteria and mitochondria, 36(1) MOL. MICROBIOL.
244-246 (2000)

1-8

Interference No. 106,048


APPENDIX 1 LIST OF EXHIBITS
EXHIBIT

DESCRIPTION

NO.
1301

Mojica et al., Intervening sequences of regularly spaced prokaryotic repeats


derive from foreign genetic elements, 60(2) J. MOL EVOL 60(2) 174-182
(2005)

1302

Morgan et al., Inducible Expression and Cytogenetic Effects of the EcoRI


Restriction Endonuclease in Chinese Hamster Ovary Cells, 8(10) MOL. CELL.
BIOL. 4204-4211 (1988)

1304

Mussolino et al., A novel TALE nuclease scaffold enables high genome editing
activity in combination with low toxicity, 39(21) NUCL. ACIDS RES. 9283-9293
(2011)

1307

Neumann et al., Gene transfer into mouse lyoma cells by electroporation in


high electric fields, 1(7) EMBO JOURNAL 841-845 (1982)

1310

OHare et al., Transformation of mouse fibroblasts to methotrexate resistance


by a recombinant plasmid expressing a prokaryotic dihydrofolate reductase,
78(3) PNAS 1527-1531 (1981)

1319

Planey et al., Inhibition of Glucocorticoid-induced Apoptosis in 697 Pre-B


Lymphocytes by the Mineralocorticoid Receptor N-terminal Domain, 277(44)
J. BIOL. CHEM. 42188-42196 (2002)

1322

Pourcel et al., CRISPR elements in Yersinia pestis acquire new repeats by


preferential uptake of bacteriophage DNA, and provide additional tools for
evolutionary studies, 151 MICROBIOL. READ. ENGL. 653-663 (2005)

1-9

Interference No. 106,048


APPENDIX 1 LIST OF EXHIBITS
EXHIBIT

DESCRIPTION

NO.
1327

Raymond and Soriano, High-efficiency FLP and C31 site-specific


recombination in mammalian cells, 2(1) PLOS ONE. e162 (2007)

1329

Reiss et al., RecA protein stimulates homologous recombination in plants, 93


PROC. NATL. ACAD. SCI. USA 3094-3098 (1996)

1332

Sandy et al., Mammalian RNAi: a practical guide, 39 BIOTECHNIOUES 215224 (2005)

1334

Sato et al., Generation of Adeno-Associated Virus Vector Enabling Functional


Expression of Oxytocin Receptor and Fluorescence Marker Genes Using the
Human elF4G Internal Ribosome Entry Site Element, 73(9) BIOSCI.
BIOTECHNOL. BIOCHEM. 2145-2148 (2009)

1335

Sauer, Functional expression of the cre-lox site-specific recombination system


in the yeast saccharomyces cerevisiae, 7(6) MOL. CELL. BIOL. 2087-2096
(1987) (Sauer 1987)

1336

Sauer and Henderson, Site-specific DNA recombination in mammalian cells by


the Cre recombinase of bacteriophage P1, 85 PROC. NATL. ACAD. SCI. USA
5166-5170 (1988)

1337

Schramm and Hernandez, Recruitment of RNA polymerase III to its target


promoters, 16(20) GENES DEV. 2593-2620 (2002)

1-10

Interference No. 106,048


APPENDIX 1 LIST OF EXHIBITS
EXHIBIT

DESCRIPTION

NO.
1353

Tiscornia et al., Development of Lentiviral Vectors Expressing siRNA in Gene


Transfer Delivery and Expression of DNA and RNA, Chpt. 3 (Friedmann &
Rossi eds.) (2007)

1370

Rouillon et al., Structure of the CRISPR Interference Complex CSM Reveals


Key Similarities with Cascade, 52(1) MOL. CELL. 124134 (2013)
(Rouillon)

1371

Gratz et al., Genome Engineering of Drosophila with the CRISPR RNAGuided Cas9 Nuclease, 194 GENETICS 1029-1035 (2013) (Gratz)

1372

DiCarlo et al., Genome engineering in Saccharomyces cerevisiae using


CRISPR-Cas systems, 41(7) NUCL. ACIDS RES. 4336-4343 (2013) (DiCarlo)

1502

M. Kido et al., Escherichia coli RecA protein modified with a nuclear


localization signal binds to chromosomes in living mammalian cells, 198
EXPERIMENTAL CELL RESEARCH 107-114 (1992)

1508

Kim et al., Highly efficient RNA-guided genome editing in human cells via
delivery of purified Cas9 ribonucleoproteins, 24 GENOME RES. 1012-1019
(2014) with Supplemental Information

1509

Cho et al., Heritable Gene Knockout in Caenorhabditis elegans by Direct


Injection of Cas9-sgRNA Ribonucleoproteins, 195 GENETICS 1177-1180
(2013) with Supplemental Materials and Methods

1-11

Interference No. 106,048


APPENDIX 1 LIST OF EXHIBITS
EXHIBIT

DESCRIPTION

NO.
1510

Sung et al., Highly efficient gene knockout in mice and zebrafish with RNAguided endonucleases, 24 GENOME RES. 125-131 (2014) with Supplemental
Material

1512

Supplemental Amendment filed on January 9, 2014 in U.S. Patent Application


No. 14/054,414

1513

Office Action dated January 13, 2014 in U.S. Patent Application No.
14/054,414

1514

Claims filed on December 12, 2013 in U.S. Patent Application No. 14/105,017

1515

Office Action dated November 13, 2014 in U.S. Patent Application No.
14/105,017

1516

Claims filed on February 18, 2014 in U.S. Patent Application No. 14/183,429

1517

Office Action dated April 9, 2014 in U.S. Patent Application No. 14/183,429

1518

Second Supplemental Amendment and Interview Summary and Request for


Interview filed on October 9, 2014 in U.S. Patent Application No. 14/256,912

1519

Final Office Action dated November 18, 2014 in U.S. Patent Application No.
14/256,912

1520

Supplemental Amendment, Interview Summary, and Request for Interview


filed on October 9, 2014 in U.S. Patent Application No. 14/226,274

1521

Final Office Action dated November 18, 2014 in U.S. Patent Application No.
14/226,274

1-12

Interference No. 106,048


APPENDIX 1 LIST OF EXHIBITS
EXHIBIT

DESCRIPTION

NO.
1522

Preliminary Amendment filed on July 27, 2015 in U.S. Patent Application No.
14/704,551

1523

Office Action dated August 14, 2015 in U.S. Patent Application No.
14/704,551

1524

Claims filed on February 18, 2014 in U.S. Patent Application No. 14/183,471

1525

Office Action dated July 1, 2014 in U.S. Patent Application No. 14/183,471

1526

Claims filed on April 22, 2014 in U.S. Patent Application No. 14/258,458

1527

Office Action dated July 14, 2014 in U.S. Patent Application No. 14/258,458

1528

Truant and Cullen, The Arginine-Rich Domains Present in Human


Immuodeficiency Virus Type 1 Tat and Rev Function as Direct Importin Dependent Nuclear Localization Signals, 19(2) MOLECULAR AND CELLULAR
BIOLOGY 1210-1217 (1999)

1531

Zuris et al., Efficient delivery of genome-editing proteins in vitro and in vivo,


33(1) NAT. BIOTECHNOL. 73-80 (2015)

1532

Gagnon et al., Efficient mutagenesis by Cas9 protein-mediated oligonucleotide


insertion and large-scale assessment of single-guide RNAs, 9(5) PLOS ONE
e98186 (2014)

1533

Corrected Filing Receipt, dated December 11, 2015 in U.S. Patent Application
No. 13/842,859

1534

Second Declaration of Carol Greider, Ph.D.

1-13

Interference No. 106,048


APPENDIX 1 LIST OF EXHIBITS
EXHIBIT

DESCRIPTION

NO.
1535

Second Declaration of Dana Carroll, Ph.D.

1555

Simons Deposition Transcript, July 18, 2016 with errata

1556

Simons Deposition Transcript, July 19, 2016 with errata

1604

Bikard et al., Programmable repression and activation of bacterial gene


expression using an engineered CRISPR-Cas system, 41(15) NUCL. ACIDS
RES. 7429-7439 (2013)

1605

Wu et al., Genome-wide binding of the CRISPR endonuclease Cas9 in


mammalian cells, 32(7) NAT. BIOTECHNOL. 670-676 (2014)

1606

Tzur et al., Heritable Custom Genomic Modifications in Caenorhabditis


elegans via a CRISPR-Cas9 System, 195 GENETICS 1181-1185 (2013)

1607

Mali et al., CAS9 transcriptional activators for target specificity screening


and paired nickases for cooperative genome engineering, 31(9) NAT.
BIOTECHNOL. 833-838 (2013)

1608

Koo et al., Measuring and Reducing Off-Target Activities of Programmable


Nucleases Including CRISPR-Cas9, 38(6) MOL. CELLS 475-481 (2015)

1609

Ding et al., Enhanced efficiency of human pluripotent stem cell genome


editing through replacing TALENs with CRISPRs, 12(4) CELL STEM CELL
393-394 (2013)

1610

Gaj et al., ZFN, TALEN and CRISPR/Cas-based methods for genome


engineering, 31(7) TRENDS BIOTECHNOL. 397-405 (2013)

1-14

Interference No. 106,048


APPENDIX 1 LIST OF EXHIBITS
EXHIBIT

DESCRIPTION

NO.
1611

Figures 5 and 6 of U.S. Provisional Application No. 61/652,086 (Ex. 1003)


annotated by Dr. Simons on September 15, 2016

1621

Zhang et al., Processing-Independent CRISPR RNAs Limit Natural


Transformation in Neisseria meningitidis, 50(4) MOL. CELL 488-503 (2013)

1622

Katic and Grohans, Targeted Heritable Mutation and Gene Conversion by


Cas9-CRISPR in Caenorhabditis elegans, 195 GENETICS 1173-1176 (2013)

1623

Li et al., Multiplex and homologous recombination-mediated plant genome


editing via guide RNA/Cas9, 31(8) NAT. BIOTECHNOL. 688-691 (2013)

1624

Musunuru, Genome editing of human pluripotent stem cells to generate human


cellular disease models, published online June 10, 2013, downloaded from
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3701209/?report=printable
on September 10, 2016

1625

Ousterout et al., Multiplex CRISPR/Cas9-Based Genome Editing for


Correction of Dystrophin Mutations that Cause Duchenne Muscular
Dystrophy, 6 NAT. COMMUN. 6244 (2015)

1626

Perez-Pinera et al., RNA-guided gene activation by CRISPR-Cas9-based


transcription factors, 10(10) NAT. METHODS 973-976 (2013)

1627

Richter et al., Exploiting CRISPR/Cas: Interference Mechanisms and


Applications, 14 INT. J. MOL. SCI 14518-14531 (2013)

1-15

Interference No. 106,048


APPENDIX 1 LIST OF EXHIBITS
EXHIBIT

DESCRIPTION

NO.
1629

Recipients of the 2015 Breakthrough Prizes in Fundamental Physics and Life


Sciences Announced, downloaded from https://breakthroughprize.org/News/21
on September 7, 2016

1630

Alpert Prize Recognizes CRISPR Pioneers downloaded from


https://hms.harvard.edu/news/alpert-prize-recognizes-crispr-pioneers on
September 7, 2016

1631

Crispr: Scientist who awarded patents on gene-editing excluded from Warren


Alpert Foundation Prize, Steve Connor Science Editor (March 9, 2016)
downloaded from http://www.independent.co.uk/news/science/crispr-scientistwho-awarded-patents-on-gene-editing-excluded-from-warren-alpertfoundation-prize-a6921541.html on September 7, 2016

1632

Press Release, FNIH Awards Lurie Prize to Jennifer Doudna (February 24,
2014),
available at http://www.fnih.org/news/press-releases/lurie-prize-in-thebiomedical-sciences-to-jennifer-doudna

1633

Press Release, The Gruber Foundation, Yale University, 2015 Gruber Genetics
(June 16, 2015) available at http://gruber.yale.edu/genetics/press/2015-grubergenetics-press-release

1-16

Interference No. 106,048


APPENDIX 1 LIST OF EXHIBITS
EXHIBIT

DESCRIPTION

NO.
1634

Pioneering research laying the foundations for revolutions in modern


biochemistry, downloaded from https://www.knaw.nl/en/awards/heinekenprizes/jennifer-doudna on September 7, 2016

1635

Time Magazine 100 Pioneers: Emmanuelle Charpentier & Jennifer Doudna


Creators of gene-editing technology by Mary-Claire King (April 16, 2015)
available at http://time.com/3822554/emmanuelle-charpentier-jenniferdoudna-2015-time-100/

1638

Deposition Transcript of Ronald Breaker, Ph.D., September 13, 2016, with


errata

1639

Deposition Transcript of Paul Simons, Ph.D., September 15, 2016, with errata

2007

Deposition Transcript of Dr. Dana Carroll, July 21, 2016.

2009

Third Declaration of Dr. Paul Simons, executed August 15, 2016.

2010

Declaration of Dr. Ronald Breaker, executed August 15, 2016.

2125

Chen et al., U.S. Patent Application No. 61/734,256, filed December 18, 2013.

2207

College of Chemistry, University of California, Berkeley, 9 Catalyst 1-32


(Spring/Summer 2014).

2213

Grens, Enzyme Improves CRISPR A smaller Cas9 protein enables in vivo


genome engineering via viral vectors, The Scientist (April 1, 2015),
http://www.the-scientist.com//?articles.view/articleNo/42580/title/EnzymeImproves-CRISPR/.

1-17

Interference No. 106,048


APPENDIX 1 LIST OF EXHIBITS
EXHIBIT

DESCRIPTION

NO.
2230

Pandika, Rising Stars: Jennifer Doudna, CRISPR Code Killer, OZY (Jan. 7,
2014), http://www.ozy.com/rising-stars/jennifer-doudna-crispr-codekiller/4690.

2274

Guo et al. Group II Introns Designed to Insert into Therapeutically Relevant


DNA Target Sites in Human Cells, 289 Science 452-457 (2000).

2290

Max E. Perutz Laboratories, Programmable RNA Complex Could Speed


Genome Editing in the Lab (June 28, 2012), https://www.mfpl.ac.at/aboutus/news/article/news-detail/programmable-rna-complex-could-speed-genomeediting-in-the-lab.html.

2411

SN 14/704,551 Excerpt, NIH grant application.

2412

SN 14/054,414 Excerpt, Cong notebook.

2419

Gairdner, 2016 Canada Gairdner Awards Honour CRISPR-Cas Researchers


and HIV/Aids Leaders.

2420

Science Magazine 2014 Media Kit.

2422

SN 14/685,510 File History Excerpt.

2423

Edge, Conversation: Life, The Augmented Human Being: A Conversation with


George Church (Mar. 30, 2013),
https://www.edge.org/conversation/george_church-the-augmented-humanbeing

1-18

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
UCs Material Facts 1-80 with Broads Responses
1.

The U.S. Provisional Application No. 61/652,086 (the First Provisional) titled

Methods and Compositions for RNA-Directed Site-Specific DNA Modification, was filed on
May 25, 2012, and lists Martin Jinek, Jennifer Doudna, Emmanuelle Charpentier, and Krzysztof
Chylinski as co-inventors. See Ex. 1003.
Broads response: ADMIT
2.

U.S. Provisional Application No. 61/716,256 (the Second Provisional), titled,

Methods and Compositions for RNA-Directed Site-Specific DNA Modification, was filed on
October 19, 2012, and lists Martin Jinek, Jennifer Doudna, Emmanuelle Charpentier, Krzysztof
Chylinski, and James Harrison Doudna Cate as co-inventors. See Ex. 1004.
Broads response: ADMIT
3.

U.S. Provisional Application No. 61/757,640 (the Third Provisional), titled, Methods

and Compositions for RNA-Directed Site-Specific DNA Modification, was filed on January 28,
2013, and lists Martin Jinek, Jennifer Doudna, Emmanuelle Charpentier, Krzysztof Chylinski,
and James Harrison Doudna Cate as co-inventors. See Ex. 1005.
Broads response: ADMIT
4.

The technology behind Count 1 is explained in the Declarations of Drs. Carroll and

Greider. See Ex. 1022, 36-67, 332-337, Ex. 1024, 30-61, 323-328 (both citing Exs. 1007,
1020, 1032, 1033, 1125, 1126, 1153, 1154, 1155, 1156, 1199, 1208, 1211, 1214, 1225, 1227,
1254, 1255, 1261, 1262, 1270, 1271, 1288, 1298, 1299, 1301, 1304, 1322, 1370).
Broads response: DENY
5.

Each of the First Provisional, Second Provisional, and Third Provisional describes and

enables an anticipation of Count 1. See Exs. 1003-1005; see also Ex. 1022, 35, 329, 332-363,

2-1

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
373-448, Ex. 1024, 29, 320, 323-354, 3640439.
Broads response: DENY
6.

The First Provisional describes and enables methods within the scope of Count 1. See

Ex. 1003; see also Ex. 1022, 341-363, 373-431, Ex. 1024, 332-354, 364-422.
Broads response: DENY
7.

Example 1 of the First Provisional sets forth all elements of Count 1 other than the

eukaryotic cell element. See Ex. 1003, 00248-00252, Figures 3, 5; see also Ex. 1022, 341349, Ex. 1024, 332-340.
Broads response: DENY
8.

The eukaryotic cell element of Count 1 is described and enabled throughout the First

Provisional, including at Paragraph 00165. See Ex. 1003, 00165; see also 00167, 00177,
00178, 00201, 00215, 00216; see also Ex. 1022, 341-363, 373-431, Ex. 1024, 332-354,
364-422.
Broads response: DENY
9.

Example 1 in combination with Paragraph 00165 describes and enables an anticipation of

Count 1 because one of ordinary skill in the art as of May 25, 2012, would have recognized that
the listed inventors were in possession of the methods of Count 1 and could have performed
those methods using only routine and predictable techniques. See Ex. 1003, 00165, 0024800252; see also Ex. 1022, 350-351, Ex. 1024, 341-342.
Broads response: DENY
10.

Example 1 in the First Provisional is an actual working example of a method of cleaving

target DNA, albeit not in a eukaryotic cell. See Ex. 1003, 00248-00252, Figs. 3, 5; see also
see also Ex. 1022, 350-352, Ex. 1024, 341-343.

2-2

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
Broads response: DENY
11.

Example 1 of the First Provisional satisfies all aspects of the second element of Count 1,

except for contacting in a eukaryotic cell, because it employs Cas9, which is necessarily a Type
II CRISPR-associated Cas protein, and specifies that the [t]arget DNAs were obtained by
chemical synthesis and that the target DNA was contacted with a Type II CRISPR-Cas system:
[t]he DNA-targeting RNA/polypeptide complexes were assembled . . . . [and] the assembled
complex was added to target DNA and incubated. See Ex. 1003, 00248-00249; see also Ex.
1156, pp. 4-6; see also Ex. 1022, 350-353, Ex. 1024, 341-344.
Broads response: DENY
12.

Included in the Type II CRISPR-Cas system of Example 1 of the First Provisional were

engineered and non-naturally-occurring single-molecule DNA-targeting RNAs. See Ex. 1003,


00248; see also Ex. 1022, 353, Ex. 1024, 344.
Broads response: ADMIT
13.

Contacting a target DNA with the system of Example 1 of the First Provisional in

eukaryotic cells is taught throughout the First Provisional. See Ex. 1003, 00165; see also Ex.
1003, 00167, 00177, 00178, 00201, 00215, 00216; see also Ex. 1022, 352-353, 376, Ex.
1024, 343-344, 367.
Broads response: DENY
14.

Example 1 of the First Provisional satisfies all aspects of the third, fourth, fifth, and sixth

elements of Count 1 because it uses the required system and specifies that the DNA-targeting
polypeptide was based on the sequence of Streptococcus pyogenes Cas9 and that a complex
was assembled from [t]he DNA-targeting RNA and the [Cas9] polypeptide. See Ex. 1003,
00248-00249; see also Appendix 3; see also Ex. 1022, 354, Ex. 1024, 345.

2-3

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
Broads response: DENY
15.

Figure 3B of the First Provisional illustrates the structure of the engineered and non-

naturally occurring single-molecule DNA-targeting RNAs, which combined the targeter-RNA


and activator-RNA into a single molecule. See Ex. 1003, 00251, Figs. 1, 3B; see also
Appendix 3; see also Ex. 1022, 354, Ex. 1024, 345.
Broads response: DENY
16.

Example 1 of the First Provisional satisfies all aspects of element 7 of Count 1 because

the DNA-targeting RNA formed a complex with the Cas9 protein during incubation [with each
other] in the cleavage buffer for 15 min[utes] at room temperature. See Ex. 1003, 00249; see
also Ex. 1022, 355, Ex. 1024, 346.
Broads response: DENY
17.

Example 1 of the First Provisional explains how the Cas9 protein was targeted to the

target DNA by adding the assembled DNA-targeting RNA/Cas9 complex to target DNA and
incubated for 1 h[ou]r at 37C. See Ex. 1003, 00249; see also Ex. 1022, 355, Ex. 1024,
346.
Broads response: DENY
18.

Figure 1 of the First Provisional supports the seventh element of Count 1 because it

depicts the DNA-targeting RNA forming a complex with the site-directed modifying polypeptide
(such as Cas9) and that cleavage (depicted by scissors) occurs because the Cas9 protein was
targeted to the target DNA. See Ex. 1003, Fig. 1; see also Appendix 3; see also Ex. 1022, 355,
Ex. 1024, 346.
Broads response: DENY
19.

Figures 3A and 5A of the First Provisional show that cleavage of the target DNA

2-4

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
molecules did occur, i.e., the methods worked. See Ex. 1003, 00248-00252, Figs. 3A, 5A; see
also Ex. 1022, 355, Ex. 1024, 346.
Broads response: DENY
20.

The First Provisional states the tested single-molecule DNA-targeting RNAs (RNA

chimera A) supported efficient target DNA cleavage. See Ex. 1003, 00251, Fig. 3B.
Broads response: ADMIT
21.

Example 1 of the First Provisional demonstrates the necessary and sufficient components

of the CRISPR-Cas9 system. See Ex. 1003, 00248-00252; see also Ex. 1022, 342-349, Ex.
1024, 333-340.
Broads response: DENY
22.

Extensive guidance in the First Provisional, such as that provided in Paragraph 00165,

explained that the methods of Example 1 could be used in host cells, including eukaryotic cells.
See Ex. 1003; see also Ex. 1022, 341-358, 373-407, Ex. 1024, 332-349, 364-398.
Broads response: DENY
23.

Claims 54, 58, 61, 66, and 72-75 of the First Provisional describe and enable all elements

of Count 1, including the eukaryotic limitation. See Ex. 1003, pp. 76-78; see also Ex. 1022,
359-363, 380-381, Ex. 1024, 350-354, 371-372.
Broads response: DENY
24.

Claim 54, whose DNA-targeting RNA can be a single-molecule DNA-targeting RNA as

illustrated in Figure 1B, provides the framework of Count 1s second, third, fourth, and fifth
elements of relying upon a DNA-targeting RNA comprising i) a targeter-RNA or guide
sequence that hybridizes with the target sequence, and ii) an activator-RNA, or a trans-activating
CRISPR RNA (tracrRNA), that hybridizes with the targeter-RNA to form a double-stranded

2-5

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
RNA duplex of a protein binding segment. See Ex. 1003, p. 76, Fig. 1B; see also Ex. 1022,
360, Ex. 1024, 351.
Broads response: DENY
25.

Claims 58, 61, and 66 of the First Provisional make clear that the methods for cleaving

target DNA set forth in Example 1 may take place in a eukaryotic cell, such as in animal cells.
See Ex. 1003, pp. 76-77; see also Ex. 1022, 361, Ex. 1024, 352.
Broads response: DENY
26.

Claim 72 of the First Provisional satisfies the second element of Count 1 because it

specifies that the DNA-modifying polypeptide used in the method of Claim 54 comprises an
amino acid sequence having at least about 75% amino acid sequence identity to amino acids 7166 or 731-1003 of the Cas9/Csn1 amino acid sequence depicted in Figure 2, or to the
corresponding domains in any of the amino acid sequences depicted in Figure 12. See Ex.
1003, p. 78; see also Ex. 1022, 362, Ex. 1024, 353.
Broads response: DENY
27.

Because Cas9 proteins are necessarily part of a Type II CRISPR system and because the

Cas9 protein may be mutated up to 25% from a naturally-occurring Cas9, the second and sixth
elements of Count 1 are satisfied by Claim 72 of the First Provisional. See Ex. 1003, pp. 76-78;
see Ex. 1156, pp. 4-6; see also Ex. 1022, 362, Ex. 1024, 353.
Broads response: DENY
28.

As used in Claim 75 of the First Provisional, nuclease activity is synonymous with

cleavage. See Ex. 1003, p. 78; see also Ex. 1022, 363, Ex. 1024, 354.
Broads response: DENY
29.

Claims 54 and 73-75 of the First Provisional, especially in view of Figure 1, satisfy

2-6

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
element seven of Count 1. See Ex. 1003, p.78; see also Ex. 1022, 363, Ex. 1024, 354.
Broads response: DENY
30.

From at least Claims 54, 58, 61, 66, and 72-75 of the First Provisional, one of ordinary

skill in the art would have recognized that the inventors were in possession of the methods of
Count 1 and would have been able to perform those methods from the enabling disclosures
throughout the First Provisional. Ex. 1003, pp. 76-78; see also Appendix 3; see also Ex. 1022,
380, Ex. 1024, 371.
Broads response: DENY
31.

Many disclosures in the First Provisional other than Example 1 in combination with

Paragraph 00165 and Claims 54, 58, 61, 66, and 72-75 demonstrate that by May 25, 2012, the
inventors were in possession of methods that include each and every element of Count 1. See
Ex. 1003; see also Ex. 1022, 356-358, Ex. 1024, 347-349.
Broads response: DENY
32.

The First Provisional uses Cas9, formerly known as Csn1, as an example of a site-

directed modifying polypeptide. See Ex. 1003, 0006, 0096; see also Ex. 1022, 356, Ex.
1024, 347.
Broads response: ADMIT
33.

The First Provisional explains that the target DNA used in the disclosed methods may

include a cell from any organism and specifically identifies, among others, a cell of a singlecell eukaryotic organism. See Ex. 1003, 0165; see also Ex. 1022, 356, Ex. 1024, 347.
Broads response: DENY
34.

Figure 1B of the First Provisional illustrates a single-molecule DNA-targeting RNA,

demonstrating that the inventors were in possession of an engineered and non-naturally occurring

2-7

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
Type II CRISPR-Cas system, because the natural system uses a DNA-targeting RNA comprising
two RNA molecules (illustrated in Figure 1A). See Ex. 1003, 0004, Fig. 1; see also Ex. 1022,
357, Ex. 1024, 348.
Broads response: DENY
35.

The First Provisional makes clear that the inventors possessed the methods of Count 1 by

explaining that crRNA, tracrRNA, and the Cas9 protein were both necessary and sufficient for
Cas9 cleavage of target DNA and that such cleavage would occur in a eukaryotic cell. See Ex.
1003, 0002-0004, 0006, 0096, 0165, Fig. 1A; see also Ex. 1022, 358, Ex. 1024, 349.
Broads response: DENY
36.

Guidance provided in Example 1 and Paragraph 00165 of the First Provisional, Claims

54, 58, 61, 66, and 72-75, and numerous passages in the Specification demonstrate that one of
ordinary skill in the art as of May 25, 2012, would have readily performed the methods of Count
1 without undue experimentation. See Ex. 1003, 00248-00252; see also Ex. 1022, 373407, Ex. 1024, 364-398.
Broads response: DENY
37.

The First Provisional teaches that the Cas9 protein and/or the DNA-targeting RNAs can

be provided in the form of a nucleic acid containing a nucleotide sequence encoding those
components, and further explains that the nucleotide sequences can be contained on an
expression vector, such as a viral vector, as would be commonly done when performing the
method in a eukaryotic cell. See Ex. 1003, 00120-00123; see also Ex. 1022, 387, Ex. 1024,
378.
Broads response: DENY
38.

Suitable expression vectors are specifically identified in the First Provisional and it was

2-8

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
known as of May 25, 2012, that vectors, such as viral vectors, were used to deliver prokaryotic
polynucleotides into eukaryotic cells. See Ex. 1003, 00120-00123; see Exs. 1031, 1039,
1194, 1203, 1218, 1283, 1285, 1334, 1353; see also Ex. 1022, 388-389, Ex. 1024, 379380.
Broads response: DENY
39.

The First Provisional explains that [i]n some embodiments, a nucleotide sequence

encoding a DNA-targeting RNA and/or a site-directed modifying polypeptide is operably linked


to a control element, e.g., a transcriptional control element, such as a promoter, and that the
transcriptional control element may be functional in [] a eukaryotic cell. See Ex. 1003,
00126; see also Ex. 1022, 390, Ex. 1024, 381.
Broads response: ADMIT
40.

The First Provisional provides numerous examples of promoters that were known by May

25, 2012, to be functional in eukaryotic cells. See Ex. 1003, 00127; see also Exs. 1229, 1237,
1310, 1332, 1337; see also Ex. 1003, 00127; see also Ex. 1022, 390-392, Ex. 1024, 381383.
Broads response: DENY
41.

The First Provisional guides one to carry out the methods in eukaryotic cells by stating

that the methods can involve introducing into a cell (or a population of cells) one or more
nucleic acids comprising nucleotide sequences encoding a DNA-targeting RNA and/or a sitedirected modifying polypeptide, and explains that there are numerous well-known methods that
can be used to accomplish this introduction. See Ex. 1003, 00121, 00129; see also Ex. 1022,
393, Ex. 1024, 384.
Broads response: DENY

2-9

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
42.

Methods for introducing a nucleic acid into a eukaryotic cell were known in the art for

over 30 years before the First Provisional was filed. See Exs. 1040, 1200, 1248, 1260, 1307; see
also Ex. 1022, 393-394, Ex. 1024, 384-385.
Broads response: DENY
43.

The First Provisional explains the use of elements to target the Cas9 protein to a desired

cellular compartment, such as Protein Transduction Domains (PTDs) or nuclear localization


signals (NLSs). See Ex. 1528; see also Ex. 1022, 395, Ex. 1024, 386.
Broads response: DENY
44.

PTDs and NLSs are helpful, but not always necessary, to target eukaryotic genomic DNA

(residing in the nucleus of eukaryotic cells). See Ex. 1022, 395, Ex. 1024, 386.
Broads response: DENY
45.

The First Provisional discloses that the Cas9 protein can include a conjugate to facilitate

traversing a cell or organelle membrane (a PTD). See Ex. 1003, 00115; see also Ex. 1022,
395, Ex. 1024, 386.
Broads response: ADMIT
46.

Some of the PTDs discussed in the First Provisional were well known NLSs, including

the sequence RKKRRQRRR. See Ex. 1003, 00115, 00179; see also Ex. 1022, 395, Ex.
1024, 386.
Broads response: DENY
47.

The First Provisional also provides examples of expression vectors that contain an NLS,

such as the vector pSVK3. See Ex. 1003, 00124; see also Ex. 1022, 395, Ex. 1024, 386.
Broads response: DENY
48.

For decades prior to the filing of the First Provisional, researchers had used nuclear

2-10

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
localization signals to target prokaryotic peptides expressed in eukaryotic cells. See Ex. 1528;
see also Ex. 1022, 395, Ex. 1024, 386.
Broads response: DENY
49.

The First Provisional explains that standard recombinant nucleic acid manipulation

techniques, such as codon optimization, can be used to practice the methods in eukaryotic cells.
See Ex. 1003, 0033; see also Exs. 1029, 1030, 1035, 1213, 1217, 1235, 1236, 1293, 1302,
1319, 1327, 1329, 1335, 1336, 1502; see also Ex. 1003, 0033; see also Ex. 1022, 396-404,
Ex. 1024, 387-395.
Broads response: DENY
50.

For many years prior to the filing of the First Provisional, researchers had used codon

optimization to enhance expression of prokaryotic polypeptides in eukaryotic cells. See Ex.


1003, 0033; see also Ex. 1022, 397, Ex. 1024, 388.
Broads response: DENY
51.

The First Provisional would have allowed persons of ordinary skill in the art to carry out

the methods of Count 1 without undue experimentation. See Ex. 1003; see also Ex. 1022,
405-407, 431, Ex. 1024, 396-398, 422.
Broads response: DENY
52.

In terms of adapting in vitro results to a eukaryotic environment, the relative skill of those

in the art as of May 25, 2012, was high and the art associated with biotechnology was vast and
highly developed. See Ex. 1022, 406, Ex. 1024, 397.
Broads response: DENY
53.

Numerous passages throughout the First Provisional set forth each and every element of

Count 1 and the application explains how to perform the methods. See Ex. 1003; see also Ex.

2-11

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
1022, 406, Ex. 1024, 397.
Broads response: DENY
54.

Because the First Provisional provides detailed descriptions and the reagents and

techniques necessary to perform the methods were well-known, minimal experimentation was
needed to perform the methods of Count 1. See Ex. 1003; see also Ex. 1022, 406, Ex. 1024,
397.
Broads response: DENY
55.

That the First Provisional describes and enables embodiments within the scope of Count

1 is further evidenced by the U.S. Patent and Trademark Office rejecting Junior Partys claims as
anticipated by the First Provisional. See Exs. 1003, 1512-1527; see also Ex. 1022, 430a-430f,
Ex. 1024, 421a-421f.
Broads response: DENY
56.

Example 1 in the First Provisional is an actual, working example of all but the eukaryotic

element of Count 1, and one of ordinary skill in the art could readily and predictably have
transitioned Example 1 to eukaryotic cells based upon the guidance provided in the application.
See Ex. 1003; see also Ex. 1022, 406, Ex. 1024, 397.
Broads response: DENY
57.

The rapid success of numerous other research groups in applying the system of Example

1 of the First Provisional to eukaryotic cells immediately after the invention was published is
empirical evidence that the First Provisional describes and enables embodiments within the
scope of Count 1. See Exs. 1003, 1055, 1056, 1058, 1059, 1508-1510; see also Ex. 1022,
408-430, Ex. 1024, 399-421.
Broads response: DENY

2-12

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
58.

Jinek 2012 publicly disclosed the components that are necessary and sufficient for

cleavage by a Type-II CRISPR-Cas system (crRNA, tracrRNA, and Cas9) and demonstrated
successful reconstitution of the system outside of its natural bacterial environment. See Ex.
1155; see also Ex. 1022, 410, Ex. 1024, 401.
Broads response: DENY
59.

Jinek 2012 explained the basic components required for the CRISPR system to operate,

demonstrated its effectiveness outside of a cell, and discussed its usefulness within a eukaryotic
cell. See Ex. 1155; see also Ex. 1022, 413, Ex. 1024, 404.
Broads response: DENY
60.

Within just seven months after Jinek 2012 published, the Cong (Ex. 1055), Mali (Ex.

1056), Cho (Ex. 1059), and Hwang (Ex. 1058) groups had successfully performed experiments,
prepared and submitted manuscripts for review, and published papers on their results showing
that the system published in Jinek 2012 (disclosed in the First Provisional) worked in eukaryotic
cells. See, e.g., Ex. 1055, p. 819, middle col.; Ex. 1056, p. 1, Ex. 1059, p. 230, left col, and Ex.
1058, pp. 1-2; see Exs. 1002, 1011-1013, 1057, 1371, 1372; see also Ex. 1022, 414-423, Ex.
1024, 405-414.
Broads response: DENY
61.

Cong, Mali, Cho, and Hwang used the single molecule DNA-targeting RNA from the

First Provisional and Jinek 2012 and attribute Jinek 2012 as the inspiration that led to the rapid
and successful demonstration of DNA cleavage and gene editing in eukaryotes. See Ex. 1155,
Fig. 5; Ex. 1055, p. 819 and Fig. 2; Ex. 1056, p. 1, and Fig. 1; Ex. 1059, p. 230, left col, and Ex.
1058, pp. 1-2; see also Ex. 1022, 416-418, Ex. 1024, 407-409.
Broads response: DENY

2-13

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
62.

The ordinary, well-known, and routine techniques (including particular promoters,

codon-optimization, and nuclear localization signals) employed by Cong, Mali, Cho, and Hwang
to successfully move the system of Jinek 2012 into eukaryotic cells are disclosed in the First
Provisional. See Exs. 1003, 1055, 1056, 1058, 1059; see also Ex. 1022, 423, Ex. 1024, 414.
Broads response: DENY
63.

Less than two years after Jinek 2012 published, at least three more groups Kim (Ex.

1508), Cho (Ex. 1509), and Sung (Ex. 1510) successfully transitioned the methods of the First
Provisional to eukaryotic cells. See Exs. 1508-1510, 1531, 1532; see also Ex. 1022, 424-430,
Ex. 1024, 415-421.
Broads response: DENY
64.

The Cong, Mali, Cho, Hwang, Kim, Cho, and Sung groups were not conducting

extensive research, nor were they engaged in undue experimentation but were instead performing
well-known, routine experiments with a new technology that had been fully described in the First
Provisional. See Exs. 1003, 1055, 1056, 1058, 1059, 1508-1510; see also Ex. 1022, 430, Ex.
1024, 421.
Broads response: DENY
65.

The Second Provisional provides at least one constructive reduction to practice of Count

1. See Ex. 1004; see also Ex. 1022, 432-439, Ex. 1024, 423-430.
Broads response: DENY
66.

The Second Provisional includes all relevant content from the First Provisional to show at

least one constructive reduction to practice of Count 1. See Ex. 1004; see also Ex. 1022, 434436, Ex. 1024, 425-427.
Broads response: DENY

2-14

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
67.

Claims 60, 65, 66, 69-72, and 98-100 of the Second Provisional disclose embodiments

within the scope of Count 1. See Ex. 1004, pp. 111, 112, 115-117; see also Appendices 6-8; see
also Ex. 1022, 435, Ex. 1024, 426.
Broads response: DENY
68.

All of the pertinent disclosure found to describe and enable an embodiment within the

scope of Count 1 in the First Provisional is carried through and included in the Second
Provisional. Compare Ex. 1003 with Ex. 1004; see also Ex. 1022, 437-439, Ex. 1024,
428-430.
Broads response: DENY
69.

That in vitro working Example 1 of the First Provisional could have readily been

transitioned to a eukaryotic cell is evidenced by Cho, Mali, Cong, and Hwang and their reliance
on Jinek 2012. See Exs. 1003, 1004, 1055, 1056, 1058, 1059; see also Ex. 1022, 437, Ex.
1024, 428.
Broads response: DENY
70.

The information in the Second Provisional that is not in the First Provisional provides

more guidance to one of ordinary skill in the art for carrying out the methods of Count 1. See Ex.
1004, 0016-0018, 0053-0061, 0084-0089, 00178, 00255-00256, 00289-00359, Figs. 13-35;
see also Ex. 1022, 437-439, Ex. 1024, 428-430.
Broads response: DENY
71.

The Third Provisional contains a working example of the methods of Count 1, which is

one of several constructive reductions to practice of Count 1 in that application. See Ex. 1005;
see also Ex. 1022, 440-448, Ex. 1024, 431-439.
Broads response: DENY

2-15

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
72.

The Third Provisional includes all relevant content from the First Provisional to describe

and enable an embodiment within the scope of Count 1. Compare Ex. 1003 to Ex. 1005; see also
Ex. 1022, 442, Ex. 1024, 433.
Broads response: DENY
73.

Claims 64, 69, 70, 73-76, and 102-104 of the Third Provisional disclose embodiments

within the scope of Count 1. See Ex. 1005, pp. 152, 153, 155-160; see also Ex. 1022, 443, Ex.
1024, 434.
Broads response: DENY
74.

All of the pertinent disclosure found in the First Provisional and the Second Provisional

to describe and enable an embodiment within the scope of Count 1 is carried through and
included in the Third Provisional. Compare Ex. 1003 with Ex. 1004 and Ex. 1005; see also Ex.
1022, 445-448, Ex. 1024, 436-439.
Broads response: DENY
75.

Example 1 of the First Provisional could have readily been transitioned to a eukaryotic

cell even as early as May 25, 2012, and certainly by the Third Provisionals filing date of
January 28, 2013. See Exs. 1003, 1005; see also Ex. 1022, 445, Ex. 1024, 436.
Broads response: DENY
76.

The rapid successes at performing the methods of Example 1 in eukaryotic cells by Mali,

Cho, Cong, Hwang, Kim, Cho, and Sung further evidence that one of ordinary skill in the art was
readily and predictably able to transition those methods to eukaryotic cells. See Exs. 1055, 1056,
1058, 1059, 1508-1510; see also Ex. 1022, 445, Ex. 1024, 436.
Broads response: DENY
77.

Additional information included in the Third Provisional not included in the First

2-16

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
Provisional provides guidance to one of ordinary skill in the art for carrying out the methods of
Count 1. See Ex. 1005, 0005-0028, 0064-0074, 00119-00120, 00240-00244, Figs. 36-46; see
also Ex. 1022, 445, Ex. 1024, 436.
Broads response: DENY
78.

The Third Provisional includes working Examples 2 and 3. See Ex. 1005, 00408-

00450; see also Ex. 1022, 446, Ex. 1024, 437.


Broads response: DENY
79.

Not only does Example 2 of the Third Provisional describe and enable embodiments

within Count 1, it further demonstrates that one of ordinary skill in the art would have applied
the methods disclosed in the First Provisional in eukaryotic cells without undue experimentation.
See Ex. 1005, 00408-00423; see also Ex. 1022, 447, Ex. 1024, 438.
Broads response: DENY
80.

U.S. Patent Application Serial No. 13/842,859 currently shares four inventors from each

of the Provisional Applications, i.e., Martin Jinek, Jennifer Doudna, Emmanuelle Charpentier,
and Krzysztof Chylinski. See Ex. 1001, pp. 346-347; see also Exs. 1003-1005, 1533.
Broads response: ADMIT

2-17

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
Broads Alleged Facts 81-188 with UCs Responses
81.

Neither Doudna P1 nor P2 includes an experiment that falls within the scope of Count 1.

Ex. 1003; Ex. 1004; Ex. 2009 11.22. Response: Insufficient information, therefore unable
to admit or deny.
82.

P1 lacks any discussion of PAM sequences. Ex. 1003; Ex. 2009 11.43. Response:

Denied.
83.

Prior to P1s May 2012 filing date, the role of PAM sequences in CRISPR system

targeting of natural DNA targets in bacteria was known. Ex. 2009 11.12-11.13. Response:
Admitted.
84.

Persons of ordinary skill (POSITA) in 2012 would have expected the role of PAM to

be addressed when adapting a CRISPR-Cas9 system for non-natural target DNA. Ex. 2009
11.43. Response: Denied.
85.

The state of the art in May 2012 relating to CRISPR-Cas9 systems was nascent and

unpredictable. Ex. 2009 11.117. Response: Denied.


86.

Dr. Doudna conceded that even after Jinek 2012, she did not know if the Jinek system

would work in a eukaryotic cell. Ex. 2230. Response: Denied.


87.

After its in vitro tests, Senior Party experienced many frustrations in eukaryotic cells.

Ex. 2009 11.100. Response: Denied.


88.

By early 2011, Junior Partys Feng Zhang had begun experiments adapting CRISPR-

Cas9 systems to function in eukaryotic cells. See Ex. 2412; Paper 53; Ex. 2009 11.102.
Response: Denied.
89.

By 2011, publications had identified certain components that potentially played role in

CRISPR-Cas9 system in prokaryotes. Ex. 1032; Ex. 1153; Ex. 1227; Ex. 2009 11.98-11.99.

2-18

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
Response: Admitted.
90.

Dr. Zhang filed a grant application with the NIH on January 12, 2012, relating to his

development of a CRISPR-Cas9 system showing his development of a single vector delivery of a


multiplexed system with dual molecule guide RNA and Cas9. Ex. 2411 at 16; Ex. 2009 11.97.
Response: Denied.
91.

Using four componentsCas9 nuclease and bacterial RNase III genes, a tracrRNA

element, and the guide RNA arrayDr. Zhangs group successfully adapted a CRISPR-Cas9
system to function in eukaryotic cells using a dual molecule guide RNA. Ex. 1055 at 2.
Response: Denied.
92.

The Cong et al. 2013 article reported Dr. Zhangs groups work adapting a CRISPR-Cas

system to function in eukaryotic cells. Ex. 1055 at 2. Response: Admitted.


93.

Senior Partys inventors created two versions of chimeric RNA (Chimera A and Chimera

B), with shorter tracrRNA components than the wild-type tracrRNA. Ex. 1003 at 2, 0006.
Response: Denied.
94.

Jinek 2012 reported that Chimera A worked efficiently in vitro but the even shorter

Chimera B did not. Ex. 1155 at 820. Response: Denied.


95.

The Jinek 2012 paper did not teach any longer tracrRNA components for their chimeric

RNA, but instead suggested to those of ordinary skill the use of Chimera A with its relatively
short tracrRNA component as compared to wild-type tracrRNA. Ex. 1155; Ex. 2009 11.66.
Response: Denied.
96.

Dr. Zhangs group at the Broad compared a modified version of Chimera A of Jinek 2012

to the CRISPR-Cas9 system previously developed in their laboratory. Ex. 1155 at 20;
Response: Denied.

2-19

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
97.

Dr. Zhangs group at the Broad adapted CRISPR-Cas9 with Chimera A-type RNA to

function in eukaryotic cells, using non-routine techniques, adding two NLSs to target the Cas9
protein to the nucleus, and a vector to express Cas9 and crRNA, with four nucleotides not
present on Chimera A. Ex. 1055 at 2. Response: Denied.
98.

Eight of the 14 Chimera A-type samples tested by the Broad scientists did not work in a

eukaryotic cell. Ex. 1055 at 20; Ex. 2009 11.130. Response: Denied.
99.

Dr. Churchs group at Harvard published the Mali 2013 article in January 2013 using

longer tracrRNA. Ex. 1056 at 824; Ex. 2009 11.109. Response: Insufficient information,
therefore unable to admit or deny.
100.

The Junior Party includes both the Broad and Harvard, which is affiliated with the Broad,

and some Broad scientists have advisors at both institutions. Ex. 2423; Ex. 2009 11.90.
Response: Insufficient information, therefore unable to admit or deny.
101.

Dr. Zhangs group at the Broad included Dr. Cong and Dr. Church at Harvard was the co-

advisor to Dr. Cong. Simons Tr. 89:4-15. Response: Insufficient information, therefore
unable to admit or deny.
102.

Dr. Hwangs group thanked Dr. Church for sharing unpublished results. Ex. 1058 at 6.

Response: Admitted.
103.

Senior Party does not assert actual reduction to practice until October 29, 2012. Paper 58.

Response: Denied.
104.

The Senior Partys in vitro work published in Jinek 2012. UC Motion 4 at 18:14-19:5

Response: Admitted.
105.

The Jinek system with Chimera A did not work in eukaryotic cells using routine

techniques. Ex. 2009 11.74. Response: Denied.

2-20

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
106.

Dr. Doudnas laboratory was surprised when they learned that Dr. Churchs laboratory at

Harvard had achieved success in eukaryotes. Ex. 2230. Response: Denied.


107.

After Dr. Churchs laboratory shared unpublished results, Dr. Doudnas laboratory

published Jinek 2013 paper with attempt to adapt CRISPR-Cas9 system to function in eukaryotic
cells. Ex. 2009 11.141. Response: Denied.
108.

Senior Partys inventors filed P3 containing the same eukaryotic test results as in Jinek

2013 using a chimeric RNA with longer tracrRNA components than Chimera A. Ex. 1005 at
Figures 36-38; Ex. 1055 at Figures 3-5. Response: Denied.
109.

Dr. Luciano Marraffini, who studies CRISPR/Cas systems at Rockefeller University,

stated that Its not trivial to make CRISPR/Cas systems work in eukaryotic cells, and that
[o]ne thing is to have them in silico and have a sequence and realize theyre smaller, and
another thing is to do the [eukaryotic] experiments and make it work. Ex. 2213. Response:
Insufficient information, therefore unable to admit or deny.
110.

Drs. Zhang, Horvath, Doudna, Charpentier and Barrangou jointly received the 2016

Gairdner Award for their work with CRISPR-Cas systems. Ex. 2419. Response: Denied.
111.

The Gairdner Foundation noted Dr. Zhangs work in the development of the microbial

CRISPR-Cas system as a genome editing tools for function in eukaryotic cells. Ex. 2419 at 2.
Response: Admitted.
112.

Count 1 recites a target DNA molecule in a eukaryotic cell that is not a natural target of

the prokaryotic CRIPSR-Cas9 system. Ex. 2009 11.17. Response: Denied.


113.

Figure 5 of Doudna P1 shows results using six Cas9 orthologs from different species of

bacteria, of which three did not show any cleavage. Ex. 1003 at 95; Ex. 2009 11.49. Response:
Denied.

2-21

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
114.

Persons of ordinary skill would have understood that the Doudna P1 inventors did not use

the correct PAM for all six orthologs in Figure 5 of Doudna P1. Ex. 2009 11.49. Response:
Denied.
115.

Example 1 of Doudna P1 and P2 describes only in vitro testing and not testing in

eukaryotic cells. Ex. 1003 at 248-253; Carroll Tr. 156:20-157:8; Ex. 2009 11.5. Response:
Admitted.
116.

Dr. Church characterized the move of CRISPR-Cas from bacteria to eukaryotic cells as a

huge jump. Ex. 2423. Response: Denied.


117.

Example 1 and Paragraph 00165 of P1 and P2 do not provide any evidence that the

CRISPR-Cas9 system of Example 1 actually works in eukaryotic cells. Ex. 2009 11.22.
Response: Denied.
118.

None of the portions of P1 or P2 cited by Senior Party provide evidence that the

CRISPR-Cas9 actually worked in eukaryotic cells, or identify a structural aspect of a CRISPRCas9 system that would make it adapted for eukaryotic cells. Ex. 2009 11.25. Response:
Denied.
119.

Persons of ordinary skill in the art in 2012 would not predict the operability of a

CRISPR-Cas system in a eukaryotic cell based on in vitro experiments. Ex.2009 11.6, 11.92.
Response: Denied.
120.

Skilled persons in 2012 would have required positive eukaryotic tests results conclude

that the Doudna inventors provided sufficient information to show possession of an invention of
an embodiment of a CRISPR-Cas system functioning in eukaryotic cells. Ex.2009 11.6, 11.92.
Response: Denied.
121.

Senior Party did not make an embodiment of Count 1 until more than a year after the

2-22

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
Senior Partys UC and University of Vienna inventors asserted conception date for Count 1 of
October 18, 2011. Paper 58 at 1. Response: Denied.
122.

Senior Party did not make an embodiment of Count 1 until more than two years after Dr.

Charpentiers asserted conception date for Count 1 of July 30, 2010. Paper 58 at 1. Response:
Denied.
123.

Sontheimer application stated that its CRISPR system could be used in eukaryotes and

taught techniques for eukaryotic experiments. Ex. 1161 at 0054-0060; Ex. 2009 11.33.
Response: Admitted.
124.

The Sontheimer application did not include actual eukaryotic experiments. Ex. 1161.

Response: Insufficient information, therefore unable to admit or deny.


125.

The claims in the Sontheimer application to Type III CRISPR system for use in

eukaryotic cells were rejected by the USPTO. Ex. 2413 at 23. Response: Admitted.
126.

The lack of tests in eukaryotic cells in Doudna P2 four months after Jinek 2012 would

have conveyed to persons of ordinary skill lack of possession of an invention of a CRISPR-Cas9


system adapted to function in a eukaryotic cell. Ex. 2009 11.74. Response: Denied.
127.

Dr. Carroll admitted that all of the information in P1 and P2 either appears in Jinek 2012

or was known. Carroll Tr. at 255:4-258:17; 305:6-338:10; Ex. 2009 11.86-87. Response:
Denied.
128.

Dr. Carroll observed in a September 2012 article that [a]ll the experiments described in

Jinek 2012 were performed in vitro with purified components and expressed doubt about
whether the CRISPR-Cas system would function in eukaryotic cells. Ex. 1152 at 1660.
Response: Denied.
129.

Dr. Carroll claims to have knowledge of one of ordinary skill in the art. Ex. 1024 at 19.

2-23

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
Response: Admitted.
130.

Dr. Carroll did not dispute that his 2012 article does not state any expectation of success

for CRISPR-Cas9 systems in eukaryotic cells. Carroll Tr. at 117:5-118:2. Response: Denied.
131.

In 2012, Dr. Carroll stated in reference to Jinek 2012 that [o]nly attempts to apply the

system in eukaryotes will address concerns he had raised. Ex. 1152 at 1660. Response:
Admitted.
132.

In a July 2014 article, Dr. Doudna confirmed that after publication of Jinek 2012, she did

not know if the CRISPR-Cas9 system would work in eukaryotic cells. Ex. 2207. Response:
Denied.
133.

An article from July 28, 2012 stated that [t]he next steps according to Dr. Charpentier

and Dr. Doudna were to test the single-RNA construct with Cas9 to find out whether it works in
eukaryotic organisms. Ex. 2290. Response: Admitted.
134.

A POSITA in May 2012 would have recognized that an in vitro environment is very

different from the environment of eukaryotic cells. Ex. 2009 11.114. Response: Denied.
135.

Differences between eukaryotic and prokaryotic cells could have unpredictable effects on

Cas9 and RNA expression, Cas9 folding, and CRISPR-Cas9 complex formation and function,
even if it worked in an in vitro environment. Ex. 2009 11.30. Response: Denied.
136.

It was known in May 2012 that it may not be possible to deliver components of CRISPR

systems to eukaryotic cells so as to be properly transcribed and translated. Ex. 2009 11.31.
Response: Denied.
137.

It was known in May 2012 that proteins or nucleic acids present in eukaryotic cells could

interfere with interactions of components of CRISPR-Cas9. Ex. 2009 11.31. Response:


Denied.

2-24

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
138.

It was known in May 2012 that systems in eukaryotic cells developed may degrade RNA

of CRISPR-Cas9 system. Ex. 2009 11.31. Response: Denied.


139.

P1 and P2 did not discuss any of the unique features of eukaryotic cells that might

prevent CRISPR-Cas9 from functioning in eukaryotic cells. Ex. 2009 11.31. Response:
Denied.
140.

It was known in May 2012 that CRISPR-Cas9 evolved in prokaryotes. Ex. 2009 11.88.

Response: Insufficient information, therefore unable to admit or deny.


141.

Count 1 requires engineering a CRISPR-Cas9 system to function in a manner that is

antithetical to its natural function in prokaryotic cells. Ex. 2009 10.5. Response: Denied.
142.

A POSITA in May 2012 understood that the human genome is at least a thousand-fold

larger than that of bacteria. Ex. 2009 10.7. Response: Insufficient information, therefore
unable to admit or deny.
143.

It was understood in May 2012 that CRISPR had never been found in eukaryotes in

nature. Ex. 2009 10.7. Response: Admitted.


144.

It was known in May 2012 that the proteins and RNA molecules of a CRISPR-Cas9

system may be toxic to eukaryotic cells. Ex. 2009 11.31. Response: Denied.
145.

Toxic effects include the effect of genome editing in locations other than the target DNA,

which are called off-target effects. Ex. 2009 11.31. Response: Denied.
146.

Dr. Carroll testified that off-target effects are an important concern for genome editing

and that off-target effects could, in fact, be lethal. Carroll Tr. at 227:3-6, 229:2-12. Response:
Denied.
147.

Dr. Greider testified that POSITA would have had no expectation regarding off-target

effects from CRISPR-Cas9 in eukaryotes without doing experiments. Greider Tr. at 405:14-22.

2-25

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
Response: Denied.
148.

It was known in May 2012 that it may not be possible to localize the components of the

CRISPR-Cas9 system to target DNA in the environment of a eukaryotic cell. Ex. 2009 11.31.
Response: Denied.
149.

P1 and P2 do not teach using an NLS in eukaryotic cells. Ex. 2009 11.31; Ex. 1003.

Response: Denied.
150.

Dr. Greider testified DNA target of interest is in nucleus. Greider Tr. at 285:20-286:1.

Response: Denied.
151.

Dr. Greider provided no opinion that the Senior Party inventors possessed an

embodiment of Count 1 for delivering a functioning CRISPR-Cas9 system, with the requisite
RNA, into a mitochondria or chloroplast. Greider Tr. (7/20/16) 422:4-18. Response: Denied.
152.

A POSITA reading P1 and P2 at the times they were filed would have known of prior

difficulties adapting a prokaryotic system to eukaryotic cells. Ex. 2010 1.45-1.54. Response:
Insufficient information, therefore unable to admit or deny.
153.

A POSITA in May 2012 would have also been aware of the history of research relating to

targetrons, gene editing tools based on group II self-splicing ribozymes. Ex. 2010 1.54
Response: Admitted.
154.

Like CRISPR-Cas9, targetrons originated in prokaryotic cells, and researchers attempted

to adapt them to function in eukaryotic cells. Ex. 2010 1.45-1.48 Response: Denied.
155.

Targetrons include protein and RNA components, like CRISPR systems. Ex. 2010 1.45.

Response: Admitted.
156.

Group II self-splicing ribozymes were initially described in 1986, and targetrons were

developed that could act on bacterial genes by 2003. Ex. 2010 1.46. Response: Admitted.

2-26

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
157.

After nine more years of research, researchers had not been able to adapt targetrons to

function effectively in eukaryotic cells. Ex. 2010 1.47-1.54 Response: Denied.


158.

A POSITA in May 2012 reading the P1 and P2 applications would have understood that

similar challenges could face researchers attempting to adapt the CRISPR-Cas9 system described
in Example 1 to function in eukaryotic cells. Ex. 2010 1.54. Response: Denied.
159.

History of targetrons taught that ability of prokaryotic system to edit genes in other

environments did not show adaptation to function in eukaryotic cells. Ex. 2010 1.45-1.54.
Response: Denied.
160.

A POSITA in May 2012 would have known that the level of Mg2+ ions could be a

problem for CRISPR-Cas9 as with targetrons, since both systems evolved to function in
prokaryotic cells. Ex. 2010 1.51-1.53. Response: Denied.
161.

Given the history of prior failed attempts to transfer prokaryotic systems to eukaryotes,

POSITA would have needed experimental results in eukaryotic cells to show possession of a
CRISPR-Cas9 invention for eukaryotic cells. Ex. 2009 11.35. Response: Denied.
162.

Dr. Zhang achieved success with a CRISPR-Cas9 system in eukaryotic cells prior to the

publication of Jinek 2012. Ex. 1055 at 2; Ex. 2411; Ex. 2009 11.97. Response: Denied.
163.

Dr. Zhang showed that his CRISPR-Cas9 system with longer tracrRNA worked better

than modified Chimera A-type system for eukaryotic cells. Ex. 1055 at 20; Ex. 2009 11.130.
Response: Denied.
164.

Senior Party has not shown that the research groups responsible for the Cong 2013, Mali

2013, Cho 2013 and Hwang 2013 articles were inspired by Jinek 2012 or that those groups were
independent with respect to CRISPR research. Ex. 2009 11.6. Response: Denied.
165.

The research groups responsible for the Cong 2013, Mali 2013, Cho 2013 and Hwang

2-27

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
2013 articles include individuals with more than the ordinary level of skill, including Drs.
George Church, Keith Joung and, Jin-Soo Kim. Ex. 2009 11.112. Response: Insufficient
information, therefore unable to admit or deny.
166.

Dr. Carroll confirmed that Dr. Kim and Dr. Doudna, one of the authors on Jinek 2013,

had higher than the ordinary level of skill. Carroll Tr. 263:18-264:20. Response: Insufficient
information, therefore unable to admit or deny.
167.

P1 and P2 provide no guidance for practicing Count 1 in eukaryotes. Ex. 2009 11.114.

Response: Denied.
168.

Chen P1 reported testing of Chimera A and modified Chimera A, all of which generated

negative results. Ex. 2125 at 32, 34; Carroll Tr. at 178:6-18. Response: Denied.
169.

The testing in Chen P1 of Chimera A or modified Chimera A was inconclusive or failed.

Ex. 2009 11.123-11.129. Response: Denied.


170.

Dr. Church acknowledged that his graduate student Dr. Cong worked with Dr. Zhang on

the eukaryotic experiments at the Broad. Ex. 2423. Response: Denied.


171.

The Kim group filed a patent application and recently submitted arguments, and a

supporting declaration of Bryan Cullen, stating that their eukaryotic work was not obvious over
Jinek 2012. Ex. 2422 at 11, 20-24. Response: Admitted.
172.

Cho 2013 added more plasmids and RNA to eukaryotic cells than recommended by

manufacturer of their nucleofection equipment, which was not routine. Ex. 2009 11.132.
Response: Denied.
173.

The testing in Cho 2013 confirms that a POSITA would have needed extensive

experimentation to make unmodified Chimera A function in eukaryotic cells. Ex. 2009 11.133.
Response: Denied.

2-28

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
174.

In later papers from Cho laboratory, researchers abandoned Chimera A and used a longer

tracr. Ex. 1509; Ex. 1508; Carroll Tr. at 279:20-291:7; Ex. 2009 11.134. Response: Denied.
175.

The journal SCIENCE has large reach in scientific community, with 570,400 readers

each week and over five million monthly visits to Science website. Ex. 2420; Ex. 2009 11.113.
Response: Insufficient information, therefore unable to admit or deny.
176.

Only Dr. Chos group using non-routine techniques showed that a CRISPR-Cas9 system

with unmodified chimera A could function in eukaryotic cells. Ex. 2009 11.113. Response:
Denied.
177.

The Doudna P3 application was filed January 29, 2013. Ex. 1005. Response: Denied.

178.

P3 includes testing in eukaryotic cells, and also adds a description of new CRISPR-Cas9

systems with tracrRNA components that differ from Chimera A. Ex. 1005. Response: Denied.
179.

Two experiments in P3 purport to show cleavage in living eukaryotic cells. Ex. 1005.

Response: Admitted that at least two experiments in P3 show cleavage in living eukaryotic
cells.
180.

According to the methods section in P3, the experiment reported in Figure 38B of P3

should show bands of 360 bp for the PCR product in all samples. Ex. 1005 at 412. Response:
Admitted.
181.

In P3, Figure 38B, the gel should show bands between 160 and 200 bp if the CRISPR-

Cas9 successfully cleaved the target DNA. Ex. 2009 11.140. Response: Admitted.
182.

The samples in P3 containing Cas9 and sgRNA do not show any diagnostic bands

between 160 and 200 bp. Ex. 1005 at 36E; Ex. 2009 11.140. Response: Denied.
183.

A POSITA would have concluded from Figure 38B of P3 that there was no successful

cleavage in eukaryotic cells. Ex. 2009 11.140. Response: Denied.

2-29

Interference No. 106,048


APPENDIX 2 STATEMENT OF MATERIAL FACTS
184.

A POSITA reviewing Fig. 36E of P3 would have concluded that necessary experimental

details are missing. Ex. 2009 11.151-11.159. Response: Denied.


185.

A POSITA reviewing Fig. 36E of P3 would have concluded that the authors did not

purify their PCR product as evidenced by the additional bands visible in lanes 2 and 4 and that
the test results were inconclusive at best. Ex. 2009 11.151-11.159. Response: Denied.
186.

Figure 36B reports results of an experiment that according to P3, revealed abundant

Cas9 expression and nuclear localization. Ex. 1005 at 416. Response: Admitted.
187.

To a POSITA, the results in Fig. 36B of P3 would not have indicated that the Senior

Partys inventors obtained nuclear localization. Ex. 2009 11.154. Response: Denied.
188.

The failure in Fig. 36B of P3 to localize the Cas9 protein in the nucleus could account for

the poor results shown in Figures 36E and 38B. Ex. 2009 11.162. Response: Denied.

2-30

Interference No. 106,048


CERTIFICATE OF FILING AND SERVICE
I hereby certify that on this 28th day of September, 2016, the foregoing UC et al.
REPLY 4 is being filed, via the Interference Web Portal, by 5:00 PM Eastern Time. Pursuant to
agreement by the parties, service copies are being sent, via electronic mail by 8:00 PM Eastern
Time today, to Junior Partys counsel as follows:
Steven R. Trybus, Esq.
Harry J. Roper, Esq.
Paul D. Margolis, Esq.
Jenner & Block LLP
353 North Clark Street
Chicago, Illinois 60654
(312) 222-9350
strybus@jenner.com
hroper@jenner.com
pmargolis@jenner.com
Raymond N. Nimrod, Esq.
Quinn Emanuel Urquhart & Sullivan, LLP
51 Madison Avenue, 22nd Floor
New York, NY 10010
(212) 849-7000
raynimrod@quinnemanuel.com
Date: September 28, 2016

/Todd R. Walters/
Todd R. Walters, Esq.
Registration No. 34,040
Buchanan Ingersoll & Rooney PC
1737 King Street, Suite 500
Alexandria, Virginia 22314
Telephone (703) 836-6620
Facsimile (703) 836-2021
todd.walters@bipc.com
Counsel for UC and Vienna

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