Ex Parte Huisman et al

Patent Trial and Appeal Board

Nov 5, 2013

10607903 (P.T.A.B. Nov. 5, 2013)

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.
10/607,903 06/27/2003 Gjalt W. Huisman MBX 025 DIV CON 7536
116248 7590 11/06/2013
Pabst Patent Group
1545 Peachtree Street
Atlanta, GA 30309-2492
EXAMINER
HUTSON, RICHARD G
ART UNIT PAPER NUMBER
1652
MAIL DATE DELIVERY MODE
11/06/2013 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 PATENT TRIAL AND APPEAL BOARD
__________

Ex parte GJALT W. HUISMAN, LAURA Z. LUO,
and OLIVER P. PEOPLES
__________

Appeal 2012-005719
Application 10/607,903
Technology Center 1600
__________

Before DEMETRA J. MILLS, ERIC GRIMES, and
JEFFREY N. FREDMAN, Administrative Patent Judges.

MILLS, Administrative Patent Judge.

DECISION ON APPEAL

This is an appeal under 35 U.S.C. § 134. The Examiner has rejected
the claims for obviousness. We have jurisdiction under 35 U.S.C. § 6(b).
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STATEMENT OF CASE
Claims 1 and 7 are representative and read as follows:

1. A bacterial strain producing polyhydroxyalkanoates wherein the
bacterial strain is selected from the group consisting of Ralstonia eutropha,
Pseudomonas putida and Escherichia coli and is genetically modified to
express a heterologous nuclease gene, wherein the nuclease is secreted into
the periplasmic space and released when the bacteria is lysed, wherein the
bacteria expresses a nuclease which is secreted into the periplasmic space in
an amount effective to degrade at least 95% of all of the nucleic acid
released following lysis of the cells in less than 24 hours and reduce the
viscosity of a cell lysate in a bacterial cell culture having a density of at least
50 g/l so that recovery of product is enhanced.

7. A bacterial strain producing polyhydroxyalkanoates, wherein the
bacterial strain is selected from the group consisting of Ralstonia eutropha,
Pseudomonas putida and Escherichia coli and is genetically modified to
express a heterologous nuclease gene integrated into the chromosome of the
bacterial host, wherein the nuclease is secreted into the periplasmic space
and released when the bacteria is lysed, wherein the bacteria expresses a
nuclease which is secreted into the periplasmic space in an amount effective
to degrade at least 95% of all of the nucleic acid released following lysis of
the cells in less than 24 hours.

Cited References

Greer, WO 94/10289, published May 11, 1994.

Miller et al., Secretion and Processing of Staphylococcal Nuclease by
Bacillus subtilis, 169 J. BACTERIOLOGY 3508-3514 (1987).

Bernard Atkinson & Ferda Mavituna, BIOCHEMICAL ENGINEERING AND
BIOTECHNOLOGY HANDBOOK (Stockton Press, 2nd. ed., 1991).1

1 We note that page 7 of the Answer includes a typographical error
indicating that, “Atkinson et al. teach that Alcaligenes eutrophus (Ralstonia
eutrophus) has been studied in detail due to its ability to accumulate large
amounts of P(3HB) (i.e. ability to grow to cell densities of approximately 85
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Liebl et al., Expression, Secretion, and Processing of Staphylococcal
Nuclease by Corynebacterium glutamicum, 174 J. BACTERIOLOGY 1854-
1861 (1992).

Sang Yup Lee & Ho Nam Chang, Production of Poly(hydroxyalkanoic
acid), 52 ADVANCES IN BIOCHEMICAL ENGINEERING/BIOTECHNOLOGY 28-58
(1995).

Boynton et al., Reduction of Cell Lysate Viscosity during Processing of
Poly(3-Hydroxyalkanoates) by Chromosomal Integration of the
Staphylococcal Nuclease Gene in Pseudomonas putida, 65 APPLIED AND
ENVIRONMENTAL MICROBIOLOGY 1524-1529 (1999).

Grounds of Rejection
Claims 1, 3, 4, 6, and 7 are rejected under 35 U.S.C. § 103(a) as being
unpatentable over Greer, Atkinson, and Lee in view of Liebl or Miller.

FINDINGS OF FACT
The Examiner’s findings of fact are set forth in the Answer at pages 4-
8.

Discussion
ISSUE
The Examiner concludes that
One of ordinary skill in the art would have been
motivated to genetically engineer a Alcaligenes eutrophus
(Ralsotnia [sic] eutropha) polyhydroxyalkanoates producing
bacterial strain, as taught by Atkinson et al., to express the
Staphylococcal aureus nuclease as taught by Liebl et al. or

g/l and produce P(3HB) at 61.5 g/l, or 80% wt/wt of dry cell mass, page 30
through 32).” This reference citation, however, is from Lee page 32, section
3.2, not Atkinson pages 30-32.
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Miller et al. or a homologous nuclease gene that has been
modified to enhance nuclease activity, so that this bacterial
strain would express a nuclease which is secreted into the
periplasmic space in an amount effective to degrade at least
95% of all the nucleic acid released following lysis of the cells
in less than 24 hours and reduce the viscosity of a cell lysate in
a bacterial cell culture having a density of at least 50 g/l so that
recovery of product is enhanced. The expression of such a
nuclease could be from a plasmid DNA or from genetic
material integrated into host chromosome. A nuclease excreted
into the medium as a result of such a genetically engineered
bacterial strain would inherently result in the degradation of at
least 95% of all the nucleic acid released following lysis of the
cells in less than 24 hours. The motivation for producing a
nuclease by a genetically engineered bacterial strain used in the
fermentation process is to reduce the amount of nucleic acids in
the medium which result in an increase in the viscosity of the
medium, causing problems in the downstream processing steps,
as taught by Greer et al. Greer et al. give further motivation for
genetically engineering a bacterial strain to express a nuclease,
because they teach that purified preparations of nucleases are
expensive and a bacterial strain that was genetically engineered
to express a nuclease activity would not require an external
nuclease or hydrogen peroxide to be added to the fermentation.
One would have had a reasonable expectation of success
because both Liebl et al. and Miller et al. were able to express
functional Staphylococcal aureus nuclease in different bacterial
species, specifically Corynebacterium glutamicum and Bacillus
subtilis and Liebl et al. teach that the Staphylococcal aureus
nuclease is a heat-stable biochemically well characterized
enzyme. One would have been further motivated to engineer
the bacterial strain to excrete/secrete the nuclease into the
growth medium following cell lysis, in an effective amount to
enhance the recovery of product from the growth medium.

(Ans. 6-7.)

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Appellants argue that
In this case, there could be no predictable outcome since the
evidence in the application clearly shows that success would
have been possible in such a small number of cases, that absent
screening for organisms with (1) secrete nuclease into the
periplasmic space, (2) make very large amount of nuclease,
and (3) which is not released until the organisms are lyzed,
there was no possibility of success. One cannot ignore the
importance of the amount of nuclease expressed.
The Board’s attention is drawn to the examples. Even if
one selected a Gram negative bacteria that made PHA, and
engineered it to express nuclease, one would not know to select
one expressing high amounts sufficient to degrade the nucleic
acid in the culture medium.

(App. Br. 23.)

Appellants argue that, “Holmes steers one away from nucleases.
Greer provides no disclosure that would steer one back to using nucleases,
let alone recombinantly producing the nuclease as claimed.” (Id. at 21.)
Appellants further argue that, “[n]one of Liebl or Miller discloses
engineering nuclease secretion into the periplasmic space. Liebl and Miller
disclose engineering the bacteria in those references to secrete nuclease into
the cell culture medium.” (Id. at 24.) Appellants cite to the Declaration of
Dr. Wolfgang Liebl, submitted October 13, 2009 as evidence that the
nuclease of Liebl is secreted into the culture medium. (Decl. 2.)
In summary, Appellants argue that, “[t]he prior art in combination
does not even vaguely guide one to recombinantly expressing nuclease in a
Gram negative bacteria which is secreted into and stored in a high amount
until the cells is lysed, to solve downstream processing problems.” (App.
Br. 29.)
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The issue is: Does the cited prior art support the Examiner’s
obviousness rejection?

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) (citations omitted). In order to determine whether a prima facie
case of obviousness has been established, we consider the factors set forth in
Graham v. John Deere Co., 383 U.S. 1, 17 (1966): (1) the scope and content
of the prior art; (2) the differences between the prior art and the claims at
issue; (3) the level of ordinary skill in the relevant art; and (4) objective
evidence of nonobviousness, if present.
“The combination of familiar elements according to known methods
is likely to be obvious when it does no more than yield predictable results.”
KSR Int’l Co. v. Teleflex Inc., 550 U.S. 398, 416 (2007).
Where . . . the claimed and prior art products are
identical or substantially identical, or are produced by identical
or substantially identical processes, the PTO can require an
applicant to prove that the prior art products do not necessarily
or inherently possess the characteristics of his claimed
product…. Whether the rejection is based on “inherency”
under 35 U.S.C. § 102, on “prima facie obviousness” under 35
U.S.C. § 103, jointly or alternatively, the burden of proof is the
same, and its fairness is evidenced by the PTO’s inability to
manufacture products or to obtain and compare prior art
products.

In re Best, 562 F.2d 1252, 1255 (CCPA 1977) (emphasis added.)
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ANALYSIS
We have decided this case in conjunction with Appeal No. 2012-
004554, Application No. 11/924350. In addition, we have reviewed our
Decisions in related Appeal Nos. 2002-001357, Application No. 09/281,363,
and 2003-001996, Application No. 09/456,940. We further acknowledge
that the claims before us include modifications not present in our earlier
Decisions.
However, in the present case, for the most part, we agree with the
Examiner’s fact finding, statement of the rejection and responses to
Appellants’ arguments as set forth in the Answer. We find that the
Examiner has provided evidence to support a prima facie case of
obviousness, except for the subject matter of claim 7. We provide the
following additional comment to reasoning set forth in the Answer.
Appellants argue that, “Holmes steers one away from nucleases”
citing the expense of nucleases, and that “Greer provides no disclosure that
would steer one back to using nucleases, let alone recombinantly producing
the nuclease as claimed.” (App. Br. 21.)
We agree with the Examiner’s indicated motivation to combine Greer
with Liebl, Miller, Atkinson and Lee, which is substantially the same issue
addressed and decided in our earlier Decision Appeal No. 2003-001996,
pages 11-12. The fact that both Greer and Holmes found that the use of
nucleases is expensive does not mean that the prior art was not aware of the
viscosity problem associated with nucleic acids and fermentation product
recover, or unaware that nucleases could be used in place of peroxide to
address the viscosity problem. The process made obvious by the cited
references involves expressing a nuclease gene in recombinant cells, not
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buying purified nucleases to add to the medium. Appellants have not
explained why the expense of commercially available nucleases would
discourage a skilled artisan from practicing the method suggested by the
prior art. We are further unconvinced by Dr. Oliver P. Peoples Declaration,
submitted February 8, 2011, which argues that the expense and use of
nucleases is prohibitive (Decl. 2) for the reasons discussed above.
Appellants further argue that, “[n]one of Liebl or Miller discloses
engineering nuclease secretion into the periplasmic space. Liebl and Miller
disclose engineering the bacteria in those references to secrete nuclease into
the cell culture medium.” (App. Br. 24.) Appellants cite to the Declaration
of Dr. Liebl as evidence that the nuclease of Liebl is secreted into the culture
medium. (Decl. 2.)
We agree with the Examiner’s response to this argument set forth on
pages 12-14 of the Answer, and find acceptable the Examiner’s use of
Boynton to show that the R. eutropha cells taught by Lee inherently secrete
nuclease into the periplasmic space. The Declaration of Dr. Liebl does not
address or rebut this finding.
Appellants argue that
In this case, there could be no predictable outcome since the
evidence in the application clearly shows that success would
have been possible in such a small number of cases, that absent
screening for organisms with (1) secrete nuclease into the
periplasmic space, (2) make very large amount of nuclease,
and (3) which is not released until the organisms are lyzed,
there was no possibility of success. One cannot ignore the
importance of the amount of nuclease expressed.

(App. Br. 23.)
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We agree with the Examiner’s response to the expectation of success
and predictability arguments set forth on pages 10-12 of the Answer. In
particular, we agree with the Examiner that Appellants’ disclosure in the
Specification that about 10% of their nuclease expressing clones expressed a
high level of nuclease is not unpredictable in the screening art. With respect
to unpredictability in the screening art, it has been held that some
experimentation, even a considerable amount, is not “undue” if, e.g., it is
merely routine. (compare, In re Wands, 858 F.2d 731, 737 (Fed. Cir.
1988)(addressing the predictability issue with respect to the enablement
requirement)).
With respect to the claimed level of nuclease production, secretion
and degradation, we agree with the Examiner that, in Boynton,
PHA producing R. eutropha cells were generated using the same
nuclease encoding gene taught by Leibl [sic] et al. above (See
Boynton et al. Materials AND Methods, p 1524 and Figure 3, p 1527
and supporting text p 1526). Boynton et al. teach that the
transformant “R. eutropha secreted nuclease into the periplasm but not
into the growth medium” (p1526, Construction of nuclease integrants
of other PHA producers). Thus the bacterial strain that is obvious
over the above references inherently produces the nuclease such that it
is secreted into the periplasmic space because they are gram negative
bacteria. Boynton et al. further characterize the nuclease activity of
the periplasmic nuclease of the transformed R. eutropha cells in the
data provided in Figure 3 and the supporting text, in which they show
the result of chromosomal DNA treatment with periplasmic fractions
of R. eutropha MBX917 (::nuckan) (lane 4) and R. eutropha
NCIMB40124HD untransformed parent strain (lane 5) at 37°C for 1
hour. As is evident from the results of Figure 3, after only one hour of
incubation with the transformed R. eutropha periplasmic fraction, all
of the visible high molecular weight DNA was digested to smaller
molecular weight fragments. Based upon this level of digestion of
high molecular weight chromosomal DNA for only one hour at 37°C,
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clearly the level of nuclease produced in the transformed R. eutropha
strain would digest at least 95% of all nucleic acid released following
lysis of the bacterial cells in a 24 hour period.

(Ans. 13-14.) In other words, we find consistent with our earlier Decision in
Appeal No. 2003-001996, page 13, that, “[u]nder these circumstances, it is
incumbent upon appellants to establish through objective evidence that such
levels of overexpression of nuclease [as in the cited references] do not
necessarily provide the function set forth in claim 1 on appeal.”

Claim 7
Appellants provide separate argument for claim 7 on page 31 of the
Brief. Claim 7 requires a strain “genetically modified to express a
heterologous nuclease gene integrated into the chromosome of the bacterial
host.” Appellants argue that, “[a]s noted above, Miller and Liebl disclose
plasmid expression. No reason is provided for why one would go to the
trouble of integrating the nuclease into the chromosome.” (App. Br. 32.)
In response, the Examiner argues that
It is routine knowledge in the art that nuclear material
integrated into the host chromosome is more stable than that
nuclear material which remains on an extra-chromosomal
plasmid. As all of these methods are routine in the art, it
appears that the integration of the nuclease encoding gene into
the host chromosome would be so as to increase the stability of
the nuclease encoding material in the transformed host cell.

(Ans. 17-18.)
We are not persuaded by the Examiner’s argument. The Examiner
fails to support this rejection with evidence in the prior art to support his
position that it is routine knowledge in the art that nuclear material
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integrated into the host chromosome is more stable than that nuclear material
which remains on an extra-chromosomal plasmid. Therefore, we are
constrained to reverse the rejection of claim 7, as unsupported by evidence
in the record.
CONCLUSION OF LAW
The cited references support the Examiner’s obviousness rejection,
except the rejection of claim 7 is reversed.
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-IN-PART

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