France Behr et al.

Patent Trials and Appeals Board

Jul 14, 2020

2020000108 (P.T.A.B. Jul. 14, 2020)

UNITED STATES PATENT AND TRADEMARK OFFICE
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.
12/735,595 12/15/2010 France Jean-Paul Behr ES-0571-0014 8997
23117 7590 07/14/2020
NIXON & VANDERHYE, PC
901 NORTH GLEBE ROAD, 11TH FLOOR
ARLINGTON, VA 22203
EXAMINER
SHIN, DANA H
ART UNIT PAPER NUMBER
1635
NOTIFICATION DATE DELIVERY MODE
07/14/2020 ELECTRONIC
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.
Notice of the Office communication was sent electronically on above-indicated "Notification Date" to the
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PTOL-90A (Rev. 04/07)
UNITED STATES PATENT AND TRADEMARK OFFICE
__________

BEFORE THE PATENT TRIAL AND APPEAL BOARD
__________

Ex parte FRANCE JEAN-PAUL BEHR, MITSUHARU KOTERA,
NADEGE PFENDER, and JEAN-SERGE REMY
__________

Appeal 2020-000108
Application 12/735,595
Technology Center 1600
__________

Before DONALD E. ADAMS, ERIC B. GRIMES, and
JEFFREY N. FREDMAN, Administrative Patent Judges.

FREDMAN, Administrative Patent Judge.

DECISION ON APPEAL
This is an appeal1,2 under 35 U.S.C. § 134 involving claims to a
cationic siRNA. The Examiner rejected the claims as obvious. We have
jurisdiction under 35 U.S.C. § 6(b). We reverse.

1 We use the word “Appellant” to refer to “applicant” as defined in 37
C.F.R. § 1.42. Appellant identifies the Real Parties in Interest as Centre
National De La Recherche Scientifique, Universite De Strasbourg, and
Polyplus Transfection (see Appeal Br. 3).
2 We have considered and refer to the Specification of July 30, 2010
(“Spec.”); Final Action of Mar. 29, 2016 (“Final Act.”); Appeal Brief of Jan.
27, 2017 (“Appeal Br.”); and Examiner’s Answer of May 3, 2017 (“Ans.”).
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Statement of the Case
Background
“RNA interference is an intracellular mechanism that allows the
sequence-specific control of expression of an endogenous or exogenous
gene” (Spec. 1:9–11). “The mediators of this mechanism are small helices
of bicatenary ribonucleic acid of about 2 turns, a fragment of the siRNAs
[small interfering ribonucleic acids], of identical sequence to target gene”
(id. at 1:11–14). “[T]he most immediate effect of the introduction of an
siRNA in a cell is the inactivation of the target messenger RNA” (id. at
1:16–18). “The disappearance of a protein or the selective destruction of a
viral RNA may have therapeutic applications” (id. at 1:25–27).
“Unfortunately, intact animal cells are impermeable to nucleic acids”
(Spec. 1:25–30). “Transfection of siRNA into the cytoplasm of cells -
without rupturing the cell membrane - may be obtained after mixing with an
excess of cationic transporter” (id. at 2:4–6). However, when “injected into
the blood, the particles are taken up and digested by the cells that
phagocytose the microbial particles. When injected into a tissue or a tumor,
their size (100-1000 nm) prevents their diffusion” (id. at 2:17–20).
“These considerations led the inventors to envision the synthesis of
intrinsically cationic siRNAs” (Spec. 2:28–29).
The Claims
Claims 16, 17, 19–24, and 40 are on appeal. Claim 16 is
representative and reads as follows:
16. A cationic siRNA comprising a fragment having 15 to 30
ribonucleotides of double-stranded RNA at the ends of which
are grafted via a phosphate group on a ribonucleotide on the
siRNA by covalent bonding 20 to 30 linear oligocations, the
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number of grafted cationic charges being greater than that of the
anionic charges borne by the internucleoside phosphates of the
RNA strands and wherein the cationic siRNA is a molecule and
active at a concentration of 1 nM to 10 nM without any
transfection agent.

The issues
The Examiner rejected claims 16, 17, 19–24, and 40 under 35 U.S.C.
§ 103(a) as obvious over Behr3 and Neuberg4 (Final Act. 3–6).
The Examiner rejected claims 16, 17, 19–24, and 40 on the ground of
nonstatutory double patenting as being unpatentable over claims 1–12, 15–
19, and 17–22 of Neuberg in view of Behr (Final Act. 9–11).
Because the same issue is involved, and same references are used, in
both of these rejections, we will consider them together.
The Examiner finds “Behr teaches that the cationic oligonucleotides
of their invention are useful for ‘siRNA degradation of messenger RNA’”
(Final Act. 4; citing Behr 3:32–34). The Examiner finds Behr teaches that
“a cationic oligonucleotide-spermine molecule (S18N19) comprising 18
spermines and 19-mer oligonucleotide is able to penetrate into the cytoplasm
and nucleus of cells without any additional transfection agent” (id.; citing
Behr 19).
The Examiner finds “Neuberg teaches that ‘cationic’ delivery agents
have been commonly used for introducing oligonucleotides including siRNA
into cells” (Final Act. 4). The Examiner finds that Neuberg teaches “a
double-stranded siRNA molecule of about 19 bps having a 3'-overhang in
each strand” (id.). The Examiner finds “Neuberg teaches that an siRNA

3 Behr et al., WO 2007/069092 A2, published June 21, 2007.
4 Neuberg et al., US 8,399,422 B2, issued Mar. 19, 2013.
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molecule at concentration of 10 nM provides about 90% silencing and
provides about 70% silencing at 1 nM when conjugated to a cationic carrier
agent without other transfection agent” (id. at 5).
The Examiner finds it obvious that an
siRNA molecule at 1 nM–10 nM would be active when
conjugated to at least 20 spermines without additional
transfection agent, because even the siRNA conjugated with a
carrier having less cationic charges was shown to be active at 1
nM . . . and because a cationic oligonucleotide comprising
spermines was shown to successfully enter into the
cytoplasm/nucleus of cells without additional agent as taught by
Behr.
(Final Act. 6).
The issue with respect to this rejection is: Does a preponderance of
the evidence of record support the Examiner’s finding that Behr and
Neuberg render the claims obvious?

Findings of Fact
1. Behr teaches “cationic oligonucleotides, i.e, oligonucleotide-
oligocation molecules, also called cationic oligonucleotides in the
description (irrespective of their global charge) that can be synthetized
stepwise on an oligonucleotide synthesizer” (Behr 1:6–9).
2. Behr teaches
the cationic oligonucleotides AiBjH of the invention have
oligonucleotides moieties Ai and oligocations moieties Bj,
wherein
. Ai is an i-mer oligonucleotide residue, with i = 5 to 50, where
nucleotide A is an oligomer with naturally or non naturally
occurring nucleobases and/or pentafuranosyl groups and/or
native phosphodiester bonds,
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. Bj is a j-mer organic oligocation moiety, with j = 1 to 50,
where B is selected from the group comprising
• -HPO3-R1-(X-R2n)n1-X-R3-O-, where R1, R2n and
R3, identical or different, are lower alkylene, X is
NH or NC(NH2)2, n varies from 1 to 5 and n1 = 2
to 20,
• -HPO3-R4-CH(R5X1)-R6-O-, where R4 is lower
alkylene, R5 and R6, identical or different, are
lower alkylene and X1 is putrescine, spermidine or
spermine residue,
• -HPO3-R7-(aa)n2-R8-O-, where R7 is lower alkylene
and R8 is lower alkylene, serine, a natural
aminoalcohol, (aa)n2 is a peptide containing natural
aminoacids with cationic side chains, such as
Arginine, Lysine, Ornithine, Histidine,
Diaminopropionic acid and n2= 2 to 20.

(Behr 2:15–29).
3. Behr teaches “A is for example selected from the group
comprising deoxyribo, ribo, locked (LNA) nucleotides as well as their
chemical modifications or substitutions” (Behr 3:1–2).
4. Behr teaches synthesis of oligomers with spermines where
“neutral oligomer N10S3 and the cationic oligomers N10S4, N10S5 and N10S6
(with or without DMT group) were purified” (Behr 13:15–16).
5. Behr teaches a “5’-cationic fluorescein-conjugated
oligonucleotide F-S18N19 (where N19 is TCGAAGTACTCAGCGTAAG)
formulation was prepared in sterile PBS. It was added to the cells to a final
concentration of 2μM . . . cells became fluorescent, with some fluorescence
located in intracellular vacuoles and, most importantly, also spread
throughout the cytoplasm and nucleus” (Behr 19:15–21).
6. Behr teaches “intracellular delivery does not require complex
formation with cationic carrier molecules” (Behr 19:31–32).
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7. Behr teaches “[i]n view of their enhanced hybridization and cell
permeation properties, they are also useful for therapeutic approaches, such
as those mediated by antisense and siRNA degradation of messenger RNA”
(Behr 3:32–34).
8. Behr teaches, regarding duplexes of spermine containing
oligodeoxyribonucleotides, that
[t]he natural duplex melted at Tm= 30°C in 10 mM HEPES pH
7.4 (Figure 5). Conjugation of increasing numbers of
spermines led to remarkable Tm increases. N10S6•C10 melted at
Tm = 75.2°C, some 45°C higher than the natural duplex. The
Tm= f(n) curve showed a sigmoidal shape with an inflection for
the neutral N10S3 oligonucleotide.
(Behr 15:19–23).
9. Neuberg teaches methods for “delivery of oligonucleotides,
particularly small interfering RNA (designated as siRNA thereafter) leading
to RNA interference (RNAi), to eukaryotic cells in culture, ex vivo or in
vivo” (Neuberg 1:14–17).
10. Neuberg teaches “[s]mall interfering RNAs are short double-
stranded RNAs (dsRNAs) having a length preferably ranging from 19 to 29
nucleotides . . . and have RNAi activity in mammalian cell culture systems
. . . . Short dsRNAs, when base-paired, with unpaired 3’ overhanging ends,
act as the guide for sequence-specific mRNA degradation” (Neuberg 1:29–
37).
11. Neuberg teaches “non-viral vectors based on cationic lipid-
mediated transfection . . . are marketed for siRNAs delivery. In contrast to
cationic polymer-based systems, cationic lipids were shown to release the
nucleic acid in the cytoplasm” (Neuberg 1:54–59).
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12. Neuberg teaches a working example where the “luciferase gene
silencing of A549GL3Luc cells was superior to 80% for siRNA
concentration ≥ 1 nM and selective as showed by the absence of luciferase
inhibition when the unrelated GL2Luc siRNA was transfected” (Neuberg
37:12–15).
Principles of Law
“Once an applicant makes a non-frivolous argument that cited prior
art is not enabling . . . the examiner must address that challenge.” In re
Morsa, 713 F.3d 104, 110 (Fed. Cir. 2013).
Analysis
Appellant contends “the Declaration evidence of record demonstrates
that Behr (WO2007/069092) does not describe a method to make
oligoribonucleotide-oligocation molecules or cationic siRNA molecules of
the presently claimed invention” (Appeal Br. 10). Appellant cites the
Erbacher Declaration5 which states the DNA synthesis procedure in Behr
is not compatible with the cationic siRNA chemistry which
used UltraMild conditions of oligonucleotide chemistry. This
oligonucleotide chemistry was adapted to support and
protection on the nucleobases used. This avoids the use of
concentrated ammonia solution at high temperatures of 55°C.
In addition RNA chemistry requires the protection at the 2'OH
position, which is not described in WO2007/069092. All of the
support cleavage, nucleobase deprotection and ribose
deprotection requires a treatment at room temperature and using
successively diluted aqueous ammonia solutions under mild
conditions and TBAF solution to remove the TDMS protection
group on the ribose.

5 Declaration of Dr. Patrick Erbacher, dated Feb. 5, 2014.
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(Erbacher Decl. ¶ 5). Dr. Erbacher finds it was “unexpected that the
oligocation tail was completely deprotected with diluted aqueous ammonia
and that the oligocation is not degraded using the TBAF solution” (id.).
The Examiner notes that Neuberg uses siRNA (see FF 9–12), and
finds the Declarant unpersuasive because “the declarant’s mere comparison
between DNA synthesis (DNA chemistry) and RNA synthesis (RNA
chemistry) does not demonstrate that the siRNA synthesis was inoperable
and non-enabled” (Ans. 6). The Examiner states the “fact remains that Behr
expressly taught using the oligonucleotide-spermine molecule for ‘siRNA
degradation of messenger RNA’ . . . and siRNA molecules have been
actively synthesized and used by relevant artisans for many years before the
invention date” (id.).
The Examiner cites Pitsch6 and Griffey7 to show that RNA syntheses
were known with deprotection groups in the prior art (see Ans. 9). We agree
with the Examiner that RNA synthesis,8 and consequently, siRNA synthesis
was known in the prior art, and we agree that Behr provides a presumption
that RNA synthesis with spermine is enabled (see FF 3, 4, 7). See In re
Antor Media Corp., 689 F.3d 1282, 1288 (Fed. Cir. 2012) (“[A] prior art

6 Pitsch et al., Reliable Chemical Synthesis of Oligoribonucleotides
(RNA) with 2’-O-[(Triisopropylsilyl)oxy]methyl(2’-O-tom)-
Protected Phosphoramidites, 84 Helvetica Chimica Acta 3773–94
(2001).
7 Griffey et al., US 2006/0073505 A1, published Apr. 6, 2006.
8 We also agree with the Examiner that the formation of siRNA double
helices formed from the two RNA strands including spermines was not
unexpected in view of Behr’s teaching that DNA double strands could be
formed from two DNA strands including spermines (FF 8) because no
specific mechanistic difference in hybridization is disclosed by Appellant.
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printed publication cited by an examiner is presumptively enabling barring
any showing to the contrary by a patent applicant or patentee.”).
On this record, however, Appellant provided evidence that Behr does
not actually synthesize an mRNA with spermines and, therefore, lacks a
working example showing that the synthesis would work. The Examiner
does not address Dr. Erbacher’s contention, regarding RNA-spermine
conjugates, that it was “unexpected that the oligocation tail was completely
deprotected with diluted aqueous ammonia and that the oligocation is not
degraded using the TBAF solution” (Erbacher Decl. ¶ 5). That is, we
understand Dr. Erbacher to mean that the unexpected aspect of the RNA
synthesis was that the spermines linked to the oligoribonucleotide were not
themselves degraded during RNA synthesis.
Dr. Erbacher’s statement is reasonable, and the Examiner provides no
direct evidence rebutting Dr. Erbacher’s position that the ordinary artisan
would have expected the oligocation tail, the spermines, to be degraded
during RNA synthesis by the deprotecting agents, but that contrary to that
expectation, the process used by Appellant did not result in such degradation
as shown by the examples in the Specification.
Because the Erbacher Declaration properly shifted the burden back to
the Examiner to establish that Behr was enabling with regard to RNA
synthesis with spermines, and the Examiner has not satisfied this burden, we
reverse these rejections.
DECISION
In summary:
Claims
Rejected
35 U.S.C.
§
Reference(s)/Basis Affirmed Reversed
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Claims
Rejected
35 U.S.C.
§
Reference(s)/Basis Affirmed Reversed
16, 17, 19–
24, 40
103(a) Behr, Neuberg 16, 17, 19–
24, 40
16, 17, 19–
24, 40
Nonstatutory
Double Patenting
16, 17, 19–
24, 40
Overall
Outcome
16, 17, 19–
24, 40

REVERSED