Ex Parte Usui et al

Patent Trial and Appeal Board

Oct 12, 2016

13059703 (P.T.A.B. Oct. 12, 2016)

UNITED STA TES p A TENT AND TRADEMARK OFFICE
APPLICATION NO. FILING DATE
13/059,703 02/18/2011
127226 7590 10/14/2016
Birch, Stewart, Kolasch & Birch, LLP
P.O. Box 747
Falls Church, VA 22040-0747
FIRST NAMED INVENTOR
Kanako Usui
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
ATTORNEY DOCKET NO. CONFIRMATION NO.
l 422-0780PUS 1 2424
EXAMINER
OYEYEMI, OLAYINKAA
ART UNIT PAPER NUMBER
1637
NOTIFICATION DATE DELIVERY MODE
10/14/2016 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
following e-mail address( es):
mailroom@bskb.com
PTOL-90A (Rev. 04/07)
UNITED STATES PATENT AND TRADEMARK OFFICE
BEFORE THE PATENT TRIAL AND APPEAL BOARD
Ex parte KANAKO USUI, TAKASHI UEMORI, HIROYUKI MUKAI, and
IKUNOSHINKAT0. 1
Appeal2015-000056
Application 13/059,703
Technology Center 1600
Before FRANCISCO C. PRATS, JOHN G. NEW and
JOHN E. SCHNEIDER, Administrative Patent Judges.
SCHNEIDER, Administrative Patent Judge.
DECISION ON APPEAL
This is an appeal under 35 U.S.C. § 134(a) involving claims to a
method for detecting RNA which have been rejected as obvious. We have
jurisdiction under 35 U.S.C. § 6(b ).
We affirm.
STATEMENT OF THE CASE
The present invention is directed to a method for detecting RNA
comprising synthesizing cDNA from an RNA template using reverse
transcription ("RT"); combining the cDNA with a thermostable DNA
polymerase, a thermostable RNase H, and an intercalating dye; carrying out
1 Appellants identify the Real Party in Interest as Takara Bio Inc. Br. 1.
Appeal2015-000056
Application 13/059,703
a polymerase chain reaction ("PCR") using the cDNA from the RT step and
amplifying the nucleic acids; and measuring the florescent signal intensity
from the intercalating dye. Spec. ii 10. Use of the thermostable DNA
polymerase and the thermostable RNase H together results in an
amplification reaction that is more specific and efficient that earlier methods.
Spec. ii 8.
Claims 7-9 and 15 are on appeal. Claim 7 is illustrative and reads as
follows:
7. A method for detecting RNA, comprising the steps of:
(A) preparing a composition comprising a reverse
transcriptase, at least one oligonucleotide primer, at least one
deoxyribonucleotide, and RNA that serves as a template of a
reverse transcription reaction;
(B) incubating the composition prepared in the step (A)
to synthesize cDNA;
(C) preparing a composition comprising the cDNA
synthesized in the step (B), a thermostable DNA polymerase, a
thermostable ribonuclease H, and an intercalating dye;
(D) carrying out polymerase chain reaction using the
composition prepared in the step
(C), and amplifying nucleic acids; and
(E) measuring a fluorescent signal intensity from the
intercalating dye, thereby detecting the nucleic acids amplified
in the step (D),
wherein the thermostable ribonuclease H is not contained in the
composition comprising the cDNA synthesized in step (B) prior
to preparing the composition of step (C).
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Application 13/059,703
The claims stand rejected as follows:
Claims 7-9 and 15 stand rejected under 35 U.S.C. § 103(a) as
unpatentable over W ang2 as evidenced by SuperScript3 in view of
Epicenter4, Dahl5 and Schneeberger6.
DISCUSSION
Issue
In rejecting the pending claims as obvious, the Examiner finds that
Wang as evidenced by Superscript teaches a Reverse Transcription -
Polymerase Chain Reaction "R T-PCR") system comprising the steps of
(A) preparing a composition for a reverse transcriptase PCR
(RT-PCR) method (see pg 2, left col, section entitled "RT-
PCR", 1st para), the composition comprising a reverse
transcriptase (i.e. Superscript II reverse transcriptase of a
Superscript first-strand synthesis system), at least one
oligonucleotide primer (see random hexamers), at least one
deoxyribonucleotide (see 10 mM dNTP mix in Superscript
first-strand synthesis system), and RNA (see total RNA
template: or specifically 5 µg of total RNA) that serves as a
template of a reverse transcription reaction.
2 Wang et al., A PCR primer bank for quantitative gene expression analysis,
31 NUCLEIC ACIDS RESEARCH 1-8 (2003) ("Wang").
3 Super Scriptâ„¢ First Strand Synthesis System for RT-PCR, Invitron by Life
technologies (2007) ("Superscript").
4 Hybridaseâ„¢ Thermostable RNase H Product Information (1995)
("Epicenter")
5 Dahl et al., US 5,268,289, issued Dec. 7, 1993 (:Dahl").
6 Schneeberger et al., Quantitative Detection of Reverse Transcriptase-PCR
Products by means of a Novel and Sensitive DNA Stain, 4 PCR METHODS
APPL. 234 (1995) ("Schneeberger").
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Application 13/059,703
Ans. 4. The Examiner also finds that Wang teaches the addition of RNase
after RT is complete meeting the limitation that the RNase is not contained
in the product of the reverse transcriptase steps. Id. The Examiner goes on
to find that Wang teaches a real-time PCR composition comprising a
thermostable DNA polymerase, a ribonuclease H, an intercalating dye and at
least one deoxyribonucleotide. Ans. 4-5. The Examiner next finds that
while Wang does not teach the use of a thermostable RN ase H, Epicenter
teaches that Hybridase TM Thermostable RNase H can be used to eliminate
RNA prior to PCR. Id. The Examiner finds that Schneeberger teaches the
desirability of avoiding heteroduplex complexes in PCR. Ans. 7-9. The
Examiner concludes that
[i]t would have been obvious to a person of ordinary skill in the
art at the time of the filing of the invention to particularly
include thermostable ribonuclease H as a replacement for E.
coil [sic] ribonuclease H in the real time PCR reaction mixture
for vvhich any PJ'.LL\ .. present during the cDN.A amplification
may interfere with the attainment of accurate quantitative
results due to formation of heteroduplexes with DNA.
One of ordinary skill in the art would have been
motivated to perform the substitution in the presence of SYBR
because although Schneeberger et al. taught that sensitivity can
be restored with the presence of SYBR, it would have also be
obvious to also use thermostable RNase H since the Epicentre
manual on E. coil ribonuclease Hand thermostable
ribonuclease H taught that both enzymes had inherent RNase H
activity that specifically degrade RNA of an RNA:DNA
hybrid/heteroduplex, and that the thermostable ribonuclease H
"Hybridaseâ„¢" degrades the hybrids at the temperatures ranges
( 45-95 °C of PCR) that give the highest stringency for specific
DNA:RNA heteroduplexes and hydrolyzes these heteroduplex
RNA without affecting the real time amplification and SYBR
green detection of double stranded DNA (i.e. double stranded
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Appeal2015-000056
Application 13/059,703
or un hybridized RNA are not degraded) thereby minimizing
background from nonspecific binding (of undesired duplexes)
and maximizing sensitivity and selectivity of the amplification
and detection methods. One of ordinary skill in the art would
have had a reasonable expectation of success for the
substitution because Dahl et al. taught high conservation of
sequence and structure for the two RN ase H and also the same
inherent function functions. In view of all of the cited prior art
references, the instant claims 7-9 and 15 are prima facie
obvious.
Ans. 8-9 (emphasis in the original).
Appellants contend that the references do not teach the claimed
method in that none of the references teach the use of a thermostable
RNase during the PCR step. Br. 5. Appellants argue that Wang
teaches the use of RNase before the PCR step begins. Br. 7. With
respect to Schneeberger, Appellants argue that Schneeberger does not
address the issue of removing RNA from a complex of cDNA and
RNA; rather Schneeberger is concerned with heteroduplexes between
different DNA sequences. Br. 8. Appellants conclude by arguing that
one skilled in the art would not have a reasonable expectation that
adding a thermostable RNase during PCR would increase PCR
sensitivity. Br. 10.
The issue with respect to this rejection is whether the Examiner has
established by a preponderance of the evidence that claims 7-9 and 15
would have been obvious over Wang as evidenced by SuperScript in view of
Epicenter, Dahl and Schneeberger under by 35 U.S.C. § 103(a).
Findings of Fact
We adopt as our own the Examiner's findings and analysis. The
following findings are included for emphasis and reference convenience.
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Application 13/059,703
FF 1. Wang teaches a method for Reverse Transcription Polymerase
Chain Reaction (RT-PCR). Wang 2.
FF2. Wang teaches the removal of the RNA template after formation
of the cDNA molecule by the addition of RNase H to the product of the
Reverse Transcriptase process. Id.
FF3. Wang uses the Superscript First-Strand Synthesis System for
reverse transcription. Id.
FF4. Superscript teaches that the sensitivity of the PCR step can be
increased by removing the RNA template form the cDNA:RNA hybrid
molecule by digestion of the RNA after first-strand synthesis and that the
SuperScript First Strand Synthesis System introduces the RNase only when
it is beneficial. Superscript 2.
FF5. Wang teaches that the PCR step is performed at temperatures of
from 50° to 95° C. Wang 2.
FF6. E. Coli RNase used in the Superscript system becomes inactive
when exposed to temperatures at or above 65° C. Epicenter.
FF7. Epicenter discloses a thermostable RNase H which is highly
active and stable at high temperatures. Epicenter.
FF8. Both E coli RNase Hand thermostable RNase H eliminate RNA
prior to second-strand synthesis of cDNA. Epicenter.
FF9. Schneeberger teaches that in competitive R T-PCR,
heteroduplexes formed by cross-hybridization of competitor and wild-type
sequences and that the heteroduplexes interfere with the ability to obtain
accurate results. Schneeberger 235.
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Principles of Law
"[W]hen a patent claims a structure already known in the prior art that
is altered by the mere substitution of one element for another known in the
field, the combination must do more than yield a predictable result." KSR
Int'! Co. v. Teleflex Inc., 550 U.S. 398, 416 (2007), citing United States v.
Adams, 383 U.S. 39, 50-51 (1966).
"[T]he test of obviousness is not express suggestion of the claimed
invention in any or all of the references but rather what the references taken
collectively would suggest to those of ordinary skill in the art presumed to
be familiar with them." In re Rosselet, 347 F.2d 847, 851, 146 USPQ 183,
186 (CCPA 1965) (emphasis in original).
The obviousness analysis "can take account of the inferences and
creative steps that a person of ordinary skill in the art would employ." KSR
Int'! Co. v. Teleflex Inc., 550 U.S. 398, 418 (2007). "A person of ordinary
skill is also a person of ordinary creativity, not an automaton." Id. at 4 21.
"Obviousness does not require absolute predictability of success ....
For obviousness under§ 103, all that is required is a reasonable expectation
of success." In re O'Farrell, 853 F.2d 894, 903-04 (Fed. Cir. 1988).
Analysis
Claim 7 is representative of the rejected claims and recites a method
for detecting RNA using RT-PCR where, in the disputed step, RNase His
added as a component of the PCR composition.
We agree with the Examiner that the subject matter of claim 7 would
have been obvious to one skilled in the art at the time the invention was
made. Wang discloses a method for Reverse Transcription Polymerase
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Chain Reaction (RT-PCR), which uses RNase H following RT to remove the
RNA template from the RNA:cDNA complex. FFl--4. The RNase used by
Wang is inactivated during the PCR process. FF5-6. Thermostable RNase
H is not inactivated by the temperatures used during PCR. FF5 and 7. Both
E.coli RNase and thermostable RNase perform the same function-
elimination of RNA before second-strand synthesis of cDNA. FF8. We
agree with the Examiner that
[r ]eplacement of mesophilic enzymes with their thermostable
counterparts (e.g. AmpliTaq polymerase in place of a
mesophilic polymerase) is routinely done and obvious to do and
try in amplification assays such as PCR as the optimal assay
conditions including cycling temperature and length of
temperature holding time can vary based on the specifics of the
assay.
In the modified assay of Wang et al., the thermostable
RNase H would remain activated during the hotStart step of the
PCR and would have continued to degrade any RNA present in
DN,,L\ .. :PJ'.LL\ .. hybrids during the PCR cycling method of \Vang et
al. (95°C for l 5s; 60°C for 30s; 68°C for 40s) ensuring that
ssDNA is released from the hybrids and made available for the
quantitative amplification, thereby improving the sensitivity of
the real time assay.
Ans. 13.
Appellants argue that none of the references disclose adding the
thermostable RNase H to the PCR composition. Br. 5. We are not
persuaded. As the Examiner noted, Superscript teaches that the cDNA from
the RT step, along with the RNase Hand any remaining RNA is added to the
PCR composition. Advisory Act. 5; FF4. Thus the references do teach
adding RNase to the PCR composition. In addition, as discussed above,
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Application 13/059,703
substitution of one RNase for another would be a routine matter for one
skilled in the art. KSR Int'! Co. v. Teleflex Inc., 550 U.S. at 416.
Appellants next argue that Schneeberger does not teach the
desirability of using an RNase during the PCR step. Br. 8. Specifically,
Appellants point out that Schneeberger addresses the problem of forming
heteroduplexes between two different DNA strands not RNA/DNA
duplexes. Br. 8-9. Even assuming that Appellants are correct on that point,
our analysis does not rely on the teachings of Schneeberger to support the
combination of references. The motivation to use a thermostable RNase H
in the PCR step stems from a routine desire to optimize conditions in the
R T-PCR process.
Finally, Appellants argue that one skilled in the art would not have
had a reasonable expectation that adding a thermostable RNase to the PCR
composition would result in increased PCR sensitivity. Br. 10-11. We are
unpersuaded. To begin we note that increased sensitivity is not a claim
limitation. In addition, as discussed above, one skilled in the art would have
a reasonable expectation that that the thermostable RNase H would continue
to degrade and RNA present in the PCR composition ensuring that
additional ssDNA is made available for quantitative amplification thereby
improving the sensitivity of the assay.
Conclusion of Law
We conclude that the Examiner has established by a preponderance of
the Evidence that claim 7 would have been obvious over Wang as evidenced
by SuperScript combined with Epicenter, Dahl and Schneeberger under by
35 U.S.C. § 103(a).
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Application 13/059,703
Claims 8, 9, and 15 have not been argued separately and therefore fall
with claim 7. 37 C.F.R. § 41.37(c)(l)(iv).
SUMMARY
We affirm the rejection under 35 U.S.C. § 103(a).
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. § l.136(a).
AFFIRMED
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