Ex Parte Yang et al

Board of Patent Appeals and Interferences

Sep 20, 2011

10448612 (B.P.A.I. Sep. 20, 2011)

UNITED STATES PATENT AND TRADEMARK OFFICE
__________

BEFORE THE BOARD OF PATENT APPEALS
AND INTERFERENCES
__________

Ex parte DUN YANG and J. MICHAEL BISHOP
__________

Appeal 2011-003581
Application 10/448,612
Technology Center 1600
__________

Before ERIC GRIMES, JEFFREY N. FREDMAN, and
STEPHEN WALSH, Administrative Patent Judges.

FREDMAN, Administrative Patent Judge.

DECISION ON APPEAL
This is an appeal under 35 U.S.C. § 134 involving claims to a method
of inhibiting mammalian gene expression. The Examiner rejected the claims
as obvious. We have jurisdiction under 35 U.S.C. § 6(b). We reverse.

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Statement of the Case
Background
“Small interfering RNA (siRNA) has bec[o]me a powerful tool to
selectively silence gene expression in cultured mammalian cells” (Spec. 2, ll.
2-3). “In the present invention, E. coli RNase III is used to cleave double-
stranded RNA into esiRNA (endoribonuclease-prepared siRNA) that can
target multiple sites within an mRNA. The invention therefore provides an
RNA duplex pool that can recognize multiple sites in any particular RNA to
silence gene expression” (Spec. 2, ll. 6-9).
The Claims
Claims 11, 13-19, and 22-24 are on appeal. Claim 11 is representative
and reads as follows:
11. A method of inhibiting expression of a mammalian
gene, the method comprising the steps of:
(i) providing double stranded RNA having a
sequence at least 95% identical to a sequence of mRNA
transcribed from the mammalian gene over a comparison
window of 200 continuous residues;
(ii) digesting the double stranded RNA with E. coli
RNAse III in vitro to form a plurality of siRNAs capable
of interacting with multiple sites of the mRNA, wherein
the siRNAs are -20-25 nucleotides in length; and
(iii) transfecting mammalian cells with the
plurality of siRNAs, thereby specifically inhibiting
expression of the mammalian gene by at least 50%.

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The issues

A. The Examiner rejected claims 11, 13-19, 22, and 24 under 35 U.S.C. §
103(a) as obvious over Elbashir #1,
1
Amarasinghe,
2
and Elbashir #2
3
(Ans.
4-6).
B. The Examiner rejected claims 11, 13-19, and 22-24 under 35 U.S.C. §
103(a) as obvious over Elbashir #1, Amarasinghe, Elbashir #2, and Corn
4

(Ans. 6-7).
A. 35 U.S.C. § 103(a) over Elbashir #1, Amarasinghe, and Elbashir #2
The Examiner finds that “Elbashir et al. #1 teach that in Drosophila
long dsRNAs are processed into short RNAs 21-22 nucleotides in length. . . .
The processing is postulated to proceed via a mechanism similar to that of E.
coli RNase III” (Ans. 4). The Examiner finds that Amarasinghe et al. teach
that E. coli RNase III is an enzyme that cleaves double stranded RNAs into
shorter fragments that may be as short as 12 nucleotides in length.
“Amarasinghe et al. further teach that an RNase III homolog is involved in

1
Elbashir et al., RNA interference is mediated by 21- and 22-
nucleotide RNAs, 15 GENES AND DEVELOPMENT 188-200 (2001)
(We will refer to this as Elbashir #1, consistent with the usage of
the Examiner and Appellants).
2
Amarasinghe et al., Escherichia coli Ribonuclease III: Affinity
Purification of Hexahistidine-Tagged Enzyme and Assays for
Substrate Binding and Cleavage, 342 METHODS ENZYMOLOGY
143-158 (2001).
3
Elbashir et al., Duplexes of 21-nucleotide RNAs mediate RNA
interference in cultured mammalian cells, 411 NATURE 494-498
(2001) (We will refer to this as Elbashir #2, consistent with the
usage of the Examiner and Appellants).
4
Corn et al., US 2003/0100127 A1, published May 29, 2003.
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RNA in RNA interference” (Ans. 4). The Examiner finds that “Elbashir et
al. #2 teach that long RNA duplexes are incapable of inhibiting gene
expression via RNA interference in mammalian cells, probably due to
stimulation of an immune response, while RNA duplexes 21 nucleotides in
length corresponding to a mammalian gene sequence do mediate RNA
interference in mammalian cells in culture” (Ans. 5).
The Examiner finds it obvious to
use an RNase III enzyme to process double-stranded RNAs
to short lengths capable of inhibiting gene expression in
mammalian cells and to use them for this purpose. Elbashir
et al. #1 provide a motivation by teaching that long double
stranded RNAs are processed into shorter RNAs by an
enzyme with activity similar to that of E. coli RNase III.
From the teachings of Amarasinghe et al., one of ordinary
skill in the art would recognize that the enzyme taught by
Elbashir et al. #1 and E. coli RNase III have equivalent
activities and that determining the conditions needed to
produce RNA of 20-25 base pairs is a matter of routine
optimization.

(Ans. 5.)
The issue with respect to this rejection is: Does the evidence of record
support the Examiner’s conclusion that Elbashir #1, Amarasinghe, and
Elbashir #2 render obvious the method of claim 11?
Findings of Fact
1. Elbashir #1 teaches that “[f]ormation of 21-23-nt fragments in
Drosophila lysate (Fig. 2) was readily detectable for 39-501 bp dsRNAs”
(Elbashir #1 189, col. 2).
2. Elbashir #1 teaches that “[a]nalysis of the products of dsRNA
processing indicated that the ~21-nt fragments are generated by a reaction
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with all the characteristics of an RNase III cleavage reaction” (Elbashir #1
193, col. 1).
3. Elbashir #1 teaches that “the siRNA duplexes may represent a
new alternative to antisense or ribozyme therapeutics” (Elbashir #1 198, col.
1).
4. Amarasinghe teaches that the “original assays for E. coli RNase
III took advantage of the ability of the enzyme to cleave polymeric dsRNA
to acid solubility” (Amarasinghe 151).
5. Amarasinghe teaches that “[e]xhaustive cleavage of polymeric
dsRNA provides duplex products averaging 12-15 bp in length, which
corresponds to slightly greater than one turn of the A-helix (11 bp)”
(Amarasinghe 144).
6. Elbashir #2 teaches that “siRNA duplexes only reduced the
expression of their cognate reporter gene, while the longer dsRNAs strongly
and nonspecifically reduced reporter-gene expression” (Elbashir #2 496, col.
1).
7. Elbashir #2 teaches “siRNA-mediated gene silencing in
mammalian cells. The use of exogenous 21-nucleotide siRNAs holds great
promise for analysis of gene function in human cell culture and the
development of gene-specific therapeutics” (Elbashir #2 497, col. 2).
8. Blaszczyk
5
teaches that the “active form of Ec-RNase III is a
dimer of two identical 25-kDa polypeptides . . . which degrades both natural

5
Blaszczyk et al., Crystallographic and Modeling Studies of
RNase III Suggest a Mechanism for Double-Stranded RNA
Cleavage, 9 STRUCTURE 1225-1236 (2001).
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and synthetic dsRNA to small duplex products ~10-18 nucleotides (nt) in
length” (Blaszczyk 1225, col. 2).
9. Blaszczyk teaches that on “a long duplex RNA substrate, each
dimeric RNase III would have the potential to make double-stranded
cleavages at each of its two compound active centers, i.e., four cleavages per
dimer, producing a 9-bp dsRNA segment with a 2-base 3’ overhang at each
end and containing two strands of 11 nt (i.e., a total length of 13 nt)”
(Blaszczyk 1233, col. 2).
Principles of Law
“In rejecting claims under 35 U.S.C. § 103, the examiner bears the
initial burden of presenting a prima facie case of obviousness. Only if that
burden is met, does the burden of coming forward with evidence or
argument shift to the applicant.” In re Rijckaert, 9 F.3d 1531, 1532 (Fed.
Cir. 1993).
“[R]ejections on obviousness grounds cannot be sustained by mere
conclusory statements; instead, there must be some articulated reasoning
with some rational underpinning to support the legal conclusion of
obviousness.” KSR Int’l Co. v. Teleflex Inc., 550 U.S. 398, 418 (2007).
Analysis
The Examiner finds that Amarasinghe et al. teach that E. coli RNase
III is an enzyme that cleaves double stranded RNAs into shorter fragments
that may be as short as 12 nucleotides in length. “Amarasinghe et al. further
teach that an RNase III homolog is involved . . . in RNA interference” (Ans.
4).
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Appellants contend that, because “the 11-mers that were then taught
in the art to be routinely produced upon each cut with E. coli RNase III
would be of the wrong size for RNAi, there would have been no reasonable
expectation of success for the use of E. coli RNase III to significantly
produce a plurality of competent siRNAs” (App. Br. 8).
We find that Appellants have the better position. All of the art cited
regarding E. coli RNase III suggests that the enzyme cleaves double
stranded RNA into pieces smaller than the 20-25 nucleotides required by
claim 11. In particular, Amarasinghe, cited by the Examiner, teaches that
“[e]xhaustive cleavage of polymeric dsRNA provides duplex products
averaging 12-15 bp in length, which corresponds to slightly greater than one
turn of the A-helix (11 bp)” (Amarasinghe 144; FF 5). Blaszczyk, cited by
Appellants, teaches that the “active form of Ec-RNase III is a dimer of two
identical 25-kDa polypeptides . . . which degrades both natural and synthetic
dsRNA to small duplex products ~10-18 nucleotides (nt) in length”
(Blaszczyk 1225, col. 2; FF 8).
While the Examiner attempts to rebut Appellants’ arguments (see
Ans. 7-10), the Examiner never identifies any teaching in the art that E. coli
RNase III will produce double stranded RNAs which are 20-25 nucleotides
in length. At best, the Examiner cites Elbashir #2 as providing a reason to
make 21 nucleotide double stranded RNAs (FF 7; Ans. 5), but this does not
provide any expectation that E. coli RNase III is capable of cleaving a
double-stranded RNA into a 21 nucleotide long fragment.
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We therefore conclude that the Examiner has not established a prima
facie case of obviousness. We therefore need not address the asserted
secondary considerations, as we will reverse the rejection.
Conclusion of Law
The evidence of record does not support the Examiner’s conclusion
that Elbashir #1, Amarasinghe, and Elbashir #2 render obvious the method
of claim 11.
B. 35 U.S.C. § 103(a) over Elbashir #1, Amarasinghe, Elbashir #2 and
Corn
This rejection relies upon the underlying obviousness rejection over
Elbashir #1, Amarasinghe, and Elbashir #2 which we reversed above.
Having reversed the obviousness rejection over Elbashir #1, Amarasinghe,
and Elbashir #2 for a failure to a prima facie case of obviousness for Claim
11, we necessarily reverse this obviousness rejection further including Corn
as this rejection does not provide any teachings of the missing elements.
SUMMARY
In summary, we reverse the rejection of claims 11, 13-19, 22, and 24
under 35 U.S.C. § 103(a) as obvious over Elbashir #1, Amarasinghe, and
Elbashir #2.
We reverse the rejection of claims 11, 13-19, and 22-24 under 35
U.S.C. § 103(a) as obvious over Elbashir #1, Amarasinghe, Elbashir #2, and
Corn.

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REVERSED

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