Ex Parte Manoharan et al

Board of Patent Appeals and Interferences

May 17, 2012

10899912 (B.P.A.I. May. 17, 2012)

UNITED STATES PATENT AND TRADEMARKOFFICE
UNITED STATES DEPARTMENT OF COMMERCE
United States Patent and Trademark Office
Address: COMMISSIONER FOR PATENTS
P.O. Box 1450
Alexandria, Virginia 22313-1450
www.uspto.gov
APPLICATION NO. FILING DATE FIRST NAMED INVENTOR ATTORNEY DOCKET NO. CONFIRMATION NO.
10/899,912 07/26/2004 Muthiah Manoharan 29520-0523 7208
84717 7590 05/18/2012
LeClairRyan (Alnylam - ALX)
2318 Mill Road
Suite 1100
Alexandria, VA 22314
EXAMINER
SHIN, DANA H
ART UNIT PAPER NUMBER
1635
MAIL DATE DELIVERY MODE
05/18/2012 PAPER
Please find below and/or attached an Office communication concerning this application or proceeding.
The time period for reply, if any, is set in the attached communication.
PTOL-90A (Rev. 04/07)
UNITED STATES PATENT AND TRADEMARK OFFICE
__________

BEFORE THE BOARD OF PATENT APPEALS
AND INTERFERENCES
__________

Ex parte MUTHIAH MANOHARAN and
KALLANTHOTTATHIL G. RAJEEV
__________

Appeal 2011-005870
Application 10/899,912
Technology Center 1600
__________

Before LORA M. GREEN, MELANIE L McCOLLUM, and STEPHEN
WALSH, Administrative Patent Judges.

McCOLLUM, Administrative Patent Judge.

DECISION ON APPEAL
This is an appeal under 35 U.S.C. § 134 involving claims to a method
of reducing off-target gene silencing. The Examiner has rejected the claims
as obvious. We have jurisdiction under 35 U.S.C. § 6(b). We affirm.
STATEMENT OF THE CASE
Claims 1, 4-13, and 19-24 are pending and on appeal (App. Br. 2).
Claim 1 is representative and reads as follows:
1. A method of reducing off-target gene silencing in a cell
comprising contacting a duplex RNA with the cell, wherein the duplex RNA
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comprises a sense strand and an antisense strand, wherein each of said
strands consists of 15-30 ribonucleotides and said sense strand comprises
one or more 2’-arabino-fluorodeoxyuridine nucleotides; and wherein the
sense strand facilitates the off-target gene silencing.
Claims 1, 4-13, and 19-24 stand rejected under 35 U.S.C. § 103(a) as
obvious over Jackson1 in view of Leake,2 Chiu,3 Baker,4 and Manoharan5
(Ans. 4).
The Examiner relies on Jackson for teaching “that off-target effects by
siRNA molecules have been recognized in the art and that . . . off-target
RNAi-mediated gene silencing can be ‘directed by the sense strand’” (id.).
In addition, the Examiner finds that Jackson teaches “that there is an art-
recognized need and demand for target-specific siRNA agents and that
‘siRNA design can be improved to convincingly reduce this off-target
activity’” (id.).
The Examiner relies on Leake for teaching “that one can induce RNAi
in a mammalian cell with increased efficiency and specificity with an siRNA
wherein the sense strand is made non-functional by incorporating a
chemically modified uracil” (id.). In particular, the Examiner finds that
Leake teaches that, “by making the sense strand non-functional with

1 Jackson et al., Expression profiling reveals off-target gene regulation by
RNAi, 21 NATURE BIOTECHNOLOGY 635-637 (2003).
2 Leake et al., US 2004/0266707 A1, Dec. 30, 2004.
3 Chiu et al., RNAi in Human Cells: Basic Structural and Functional
Features of Small Interfering RNA, 10 MOLECULAR CELL 549-561 (2002).
4 Baker et al., US 2004/0161777 A1, Aug. 19, 2004.
5 Manoharan, Oligonucleotide Conjugates as Potential Antisense Drugs with
Improved Uptake, Biodistribution, Targeted Delivery, and Mechanism of
Action, 12 ANTISENSE & NUCLEIC ACID DRUG DEVELOPMENT 103-128
(2002).
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chemical modifications, one can reduce ‘nonspecific RNA interference’”
and “that modified nucleotides that can be used . . . include modified
nucleotides having sugar moieties such as arabinoses” (id.).
The Examiner relies on Chiu for teaching
a chemical modification at the 3’ or 5’ terminus of the sense
strand or a bulge in the internal region of the sense strand has
little or no effect on RNAi activity in cells, whereas a chemical
modification at the 3' terminus of the antisense strand slightly
lowers the RNAi efficacy; a chemical modification at the
5' terminus of the antisense strand or a bulge in the internal
region of the antisense strand significantly lowers the RNAi
activity in cells.
(Id. at 5.) Hence, the Examiner finds that Chiu suggests “that modifications
on the sense strand are well-tolerated and do not interfere with the RNAi
activity, where[as] modifications on the antisense strand are not well-
tolerated and significantly abolish the normal RNAi activity in cells when
the siRNA duplex is transfected into the cells” (id.).
The Examiner relies on Baker for teaching “that preferred 2’-sugar
position modifications for siRNA molecules include 2’-arabino
modifications on fluoro nucleotides” (id.).
The Examiner relies on Manoharan for teaching features recited in
dependent claims (id. at 5-6).
The Examiner concludes that it would have been obvious
to reduce siRNA-mediated off-target silencing effects by
disrupting the activity of the sense strand sequence of an siRNA
by incorporating 2’-arabino-fluorouridines that help inactivate
the sense strand activity, thereby allowing the activity of the
antisense strand for RNAi-mediated target silencing, thereby
inhibiting potential off-target silencing effects of the sense
strand oligonucleotide in cells.
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(Id. at 6.)
FINDINGS OF FACT
1. The Specification discloses that, “[t]o minimize the effect of an
iRNA agent on off-target silencing, the sense strand of the iRNA agent can
be modified, such as at the 5’ or 3’ ends or at an internal site in the sense
strand” (Spec. 3).
2. The Specification also discloses: “In one embodiment, the
modification is a DNA modification, e.g., a deoxynucleotide replaces a
ribonucleotide. . . . In another embodiment, a ribonucleotide is modified.
For example, a uridine can be replaced with 2’-arabino-fluorodeoxyuridine.”
(Id. at 6.)
3. Jackson discloses:
Interestingly, for one of these siRNAs, the off-target gene
silencing was directed by the antisense strand of the siRNA,
whereas for the other siRNA the off-target gene silencing
appeared to be directed by the sense strand. This suggests that
both the sense and antisense strands of an siRNA duplex can
contribute to transcript silencing.
(Jackson 636.)
4. Jackson also discloses:
Given the small degree of similarity implicated in off-target
gene regulation, it may be difficult to select an siRNA sequence
that will be absolutely specific for the target of interest. Until
siRNA design can be improved to convincingly reduce this off-
target activity, incorporating multiple siRNA duplexes to
silence a target gene of interest will increase the confidence
with which an observed phenotype and expression pattern can
be linked to target gene silencing.
(Id. at 637.)
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5. Leake discloses “compositions and methods for performing
RNA interference, including siRNA-induced gene silencing” (Leake,
¶ [0071]).
6. In particular, Leake discloses “a double stranded polynucleotide
having a sense strand comprising a polynucleotide comprised of at least one
orthoester modified nucleotide, and an antisense strand comprising a
polynucleotide comprised of at least one 2' modified nucleotide unit” (id. at
¶ [0012]).
7. Leake also discloses that “[n]ucleotide analogs include
nucleotides having modifications in the chemical structure of the base, sugar
and/or phosphate, including, but not limited to, . . . 2'-position sugar
modifications, including but not limited to, sugar-modified ribonucleotides
in which the 2'-OH is replaced by a group such as . . . halo” (id. at ¶ [0110]).
8. In addition, Leake discloses that modified nucleotides “include
those nucleotides that are modified with respect to the sugar moiety, as well
as nucleotides having sugars or analogs thereof that are not ribosyl. For
example, the sugar moieties may be . . . arabinoses.” (Id. at ¶ [0111].)
9. Leake also discloses that, in “addition to the orthoester
modification, any of the above described other modifications may also be
present when using this method” (id. at ¶ [0206]).
10. In addition, Leake discloses that “[e]ach of the aforementioned
embodiments permits the conducting of efficient RNAi interference because
the polynucleotide is more stable than naked polynucleotides” (id. at
¶ [0214]).
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11. Leake also discloses:
An additional surprising benefit of the present invention is that
it minimizes nonspecific RNA interference. Nonspecific RNA
interference occurs when a sense strand silences or partially
silences the function of untargeted genes. Orthoester
modifications and the other modifications described herein,
alone or in combination with one another, can be employed in
the sense strand to reduce or prevent such nonspecific RNA
interference.
(Id. at ¶ [0215].)
12. In addition, Leake discloses “a method of performing RNA
interference, said method comprising exposing a double stranded
polynucleotide to a target nucleic acid, wherein said double stranded
polynucleotide is comprised of a sense strand and an antisense strand, and
wherein said sense strand is substantially nonfunctional” (id. at ¶ [0217]).
13. Leake also discloses that the “modifications described herein
are an inexpensive, reliable, and non-toxic method of modifying siRNA
duplexes such that a sense strand will be substantially unable to function as
an antisense strand” (id. at ¶ [0226]).
14. Baker discloses “modified oligonucleotides useful in the RNAi
pathway,” specifically oligonucleotides “modified by having 3' terminal
cap” (Baker, ¶ [0032]).
15. Baker also discloses that the oligonucleotides “can also have
additional modifications that modulate pharmacokinetic properties” (id.).
16. In particular, Baker discloses that “preference for the 3'-endo
conformation can be achieved by deletion of the 2'-OH as exemplified by
2'deoxy-2'-F-nucleosides” (id. at ¶ [0034]).
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17. In addition, Baker discloses: “A 2'-substituent group on a
furanosyl ring can be in the ribo (down) or arabino (up) position. Preferred
2'-arabino modifications include fluoro.” (Id. at ¶ [0143].)
ANALYSIS
Jackson discloses that the sense strand of an siRNA duplex can
contribute to off-target transcript silencing (Findings of Fact (FF) 3-4). In
view of the foregoing findings of fact, we conclude that the Examiner has set
forth a prima facie case that it would have been obvious to reduce this
problem by incorporating one or more 2'-arabino-fluorodeoxyuridine
nucleotides in the sense strand of an siRNA duplex.
Appellants argue: “Leake teaches that the 2’-O-alkyl modification is
responsible for the sense strand being made non-functional. There is no
teaching or suggestion in Leake that any other modification besides the
2’-O-alkyl modification could accomplish this.” (App. Br. 8.) We are not
persuaded.
As noted by the Examiner (Ans. 11), Leake discloses that
“[o]rthoester modifications and the other modifications described herein,
alone or in combination with one another, can be employed in the sense
strand to reduce or prevent such nonspecific RNA interference” (FF 11
(emphasis added)). As also noted by the Examiner (Ans. 12), Leake
discloses that the “modifications described herein are an inexpensive,
reliable, and non-toxic method of modifying siRNA duplexes such that a
sense strand will be substantially unable to function as an antisense strand”
(FF 13). We agree with the Examiner that Leake suggests that modifications
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in the sense strand other than orthoester modifications reduce or prevent
nonspecific RNA interference.
Moreover, Leake clearly discloses including modifications other than
orthoester modifications in an RNA sense strand that facilitates off-target
gene silencing (FF 11), Baker specifically discloses that “[p]referred
2'-arabino modifications include fluoro” (FF 17), and Appellants’
specification teaches that these modifications would inherently reduce off-
target gene silencing (FF 1-2). “Mere recognition of latent properties in the
prior art does not render nonobvious an otherwise known invention.” In re
Baxter Travenol Labs., 952 F.2d 388, 392 (Fed. Cir. 1991).
Appellants also argue that they “have surprisingly found that presence
of one or more 2’-arabino-fluorodeoxyuridines in the sense strand of a
duplex RNA reduces the gene silencing activity of that strand” (App.
Br. 12). We are not persuaded.
Appellants have not provided evidence demonstrating that the
reduction in gene silencing would have been surprising in view of the
applied references. “[I]t is well settled that unexpected results must be
established by factual evidence. ‘Mere argument or conclusory statements
in the specification does not suffice.’” In re Geisler, 116 F.3d 1465, 1470
(Fed. Cir. 1997) (quoting In re De Blauwe, 736 F.2d 699, 705 (Fed. Cir.
1984)).
In addition, Appellants argue: Baker’s “disclosure of a 2’-arabino
modification . . . is not the same thing as 2’-arabino-fluorodeoxyuridine, as
recited in Appellants’ claims. There is no suggestion in Baker that this
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modification should contain a uridine nucleobase.” (Reply Br. 6.) We are
not persuaded.
We agree that the Examiner has not pointed to a specific teaching of a
2'-arabino-fluorodeoxyuridine. However, in the absence of evidence that
modifying a uridine, in particular, would provide unexpectedly superior
results, we conclude that the Examiner has set forth a prima facie case of
obviousness.
CONCLUSION
The evidence supports the Examiner’s conclusion that the method of
claim 1 would have been obvious. We therefore affirm the obviousness
rejection of claim 1. Claims 4-13 and 19-24 are not separately argued and
therefore fall with claim 1. 37 C.F.R. § 41.37(c)(1)(vii).
TIME PERIOD FOR RESPONSE
No time period for taking any subsequent action in connection with
this appeal may be extended under 37 C.F.R. § 1.136(a).

AFFIRMED

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