RUAN, Ke-He

Patent Trials and Appeals Board

Nov 1, 2019

14885561 - (D) (P.T.A.B. Nov. 1, 2019)

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.
14/885,561 10/16/2015 Ke-He RUAN 2483-01101 4745
23505 7590 11/01/2019
CONLEY ROSE, P.C.
575 N. Dairy Ashford Road
Suite 1102
HOUSTON, TX 77079
EXAMINER
RAMIREZ, DELIA M
ART UNIT PAPER NUMBER
1652
NOTIFICATION DATE DELIVERY MODE
11/01/2019 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):
pathou@conleyrose.com
PTOL-90A (Rev. 04/07)

UNITED STATES PATENT AND TRADEMARK OFFICE
____________

BEFORE THE PATENT TRIAL AND APPEAL BOARD
____________

Ex parte KE-HE RUAN
____________

Appeal 2019-004812
Application 14/885,561
Technology Center 1600
____________

Before DONALD E. ADAMS, ERIC B. GRIMES, and
JENNIFER MEYER CHAGNON, Administrative Patent Judges.

ADAMS, Administrative Patent Judge.

DECISION ON APPEAL
Pursuant to 35 U.S.C. § 134(a), Appellant1 appeals from Examiner’s
decision to reject claims 1–3, 5, 16, 19, and 20 (Appeal Br. 5). We have
jurisdiction under 35 U.S.C. § 6(b).
We AFFIRM.

1 We use the word “Appellant” to refer to “applicant” as defined in 37
C.F.R. § 1.42. Appellant identifies the real party in interest as “University
of Houston System” (Appellant’s December 11, 2018 Appeal Brief (Appeal
Br.) 3).
Appeal 2019-004812
Application 14/885,561

2
STATEMENT OF THE CASE
Appellant’s disclosure “relates to the construction and expression of a
single linked protein molecule that possesses both the enzyme functions of
the native . . . [cyclooxygenase (COX)] and that of . . . [prostacyclin
synthase (PGIS)]” (Spec. ¶ 3). Claim 1 is representative and reproduced
below:
1. An isolated hybrid protein molecule comprising an
enzymatically active human cyclooxygenase isoform-1 (COX-
1) enzyme and a human prostacyclin synthase (PGIS) enzyme
wherein said COX-1 enzyme and said PGIS enzyme are
connected with a linker, wherein said linker connects C-
terminus of the COX-1 enzyme with N-terminus of the PGIS
enzyme; and wherein said hybrid protein molecule converts AA
(arachidonic acid) to PGI2 (prostacyclin).
(Appeal Br. 17.)

Appeal 2019-004812
Application 14/885,561

3
Ground of rejection before this Panel for review:2
Claims 1–3, 5, 16, 19, and 20 stand rejected under 35 U.S.C. § 103(a)
as unpatentable over the combination of Ruan ’05,3 Ruan ’04,4 Sakaki,5
Lindbladh,6 and Hargrave.7

ISSUE
Does the preponderance of evidence relied upon by Examiner support
a conclusion of obviousness?
FACTUAL FINDINGS (FF)
FF 1. Ruan ’05 discloses a “cyclooxygenase (COX)/ prostaglandin I2
synthase (PGIS) coupling reaction system . . . used to determine the

2 Appellant’s claim 1 stands objected on this record (see Examiner’s
September 20, 2018 Non-Final Action (Non-Final Act.) 2). For the reasons
set forth by Examiner, we find this objection to be a Petitionable rather than
Appealable issue and, therefore, decline to discuss this objection further
(Examiner’s April 2, 2019 Answer (Ans.) 8).
3 Ke-He Ruan et al., The N-terminal membrane anchor domain of the
membrane-bound prostacyclin synthase involved in the substrate
presentation of the coupling reaction with cyclooxygenase, 435 Archives of
Biochemistry and Biophysics 372–81 (2005).
4 Ke-He Ruan, Advance in Understanding the Biosynthesis of Prostacyclin
and Thromboxane A2 in the Endoplasmic Reticulum Membrane via the
Cyclooxygenase Pathway, 4 Mini-Reviews in Medicinal Chemistry 639-47
(2004).
5 Toshiyuki Sakaki et al., Kinetic Studies on a Genetically Engineered Fused
Enzyme between Rat Cytochrome P4501A1 and Yeast NADPT-P450
Reductase, 33 Biochemistry 4933–39 (1994).
6 C. Lindbladh et al., Preparation and Kinetic Characterization of a Fusion
Protein of Yeast Mitrochondrial Citrate Synthase and Malate
Dehydrogenase, 33 Biochemistry 11692–98 (1994).
7 P.A. Hargrave et al., The Structure of Bovine Rhodopsin, 9 Biophys.
Struct. Mech. 235–44 (1983).
Appeal 2019-004812
Application 14/885,561

4
coordination of PGIS with COX for the biosynthesis of prostacyclin (PGI2)
using arachidonic acid (AA) as a substrate in a membrane-bound
environment” (Ruan ’05, Abstract; see id. at 373 (“PGI2 serves as one of the
most important cardiovascular protectors in physiopathological conditions”);
Ans. 3).
FF 2. Ruan ’04’s Figure 8 is reproduced below:

Ruan ’04’s Fig. 8 illustrates “[a] model of the coordination of PGIS and
COX in the biosynthesis of PGI2 in the ER membrane” (Ruan ’04 646: Fig.
8, Legend).
FF 3. Ruan ’05 discloses that “the two isoforms of the COX enzymes,
cyclooxygenase-1 (COX-1) and -2 (COX-2) . . . [and] PGIS . . . are mainly
localized in the endoplasmic reticulum (ER) membrane,” wherein the
catalytic domains of the COX “proteins lie on the luminal side of the ER,
and are anchored to the ER membrane by hydrophobic side chains of the
amphipathic helices A–D” and the catalytic domain of PGIS is “on the
cytoplasmic [side of the ER membrane and] . . . is anchored to the ER
Appeal 2019-004812
Application 14/885,561

5
membrane by the first 20 residues and residues in the helix F/G Loop” (Ruan
’05 373 (endnotes omitted); see also Ruan ’04 646: § 8; see Ans. 3–4).
FF 4. Ruan ’05 discloses that PGIS is topologically arranged opposite the
orientation of COX-1 and -2 with respect to the ER membrane (Ruan ’05
373; see also Ans. 4; Ans. 13 (Examiner finds that Ruan ’04 and Ruan ’05
disclose that the amino acid sequences of human COX-1 and human PGIS
were known in the art at the time of Appellant’s claimed invention)).
FF 5. Examiner finds that neither Ruan ’04 nor Ruan ’05 “teach a fusion
protein comprising COX-1 and PGIS” (Ans. 4).
FF 6. Sakaki discloses a recombinant hybrid fusion protein comprising
membrane-bound yeast NADPH-P450 reductase and rat cytochrome
P4501A1 (see Sakaki 4933; Ans. 4).
FF 7. Sakaki discloses that its “fused enzyme is capable of rapidly
transferring electrons from . . . the reductase part” of the fused enzyme to the
cytochrome portion of the fused enzyme (see Sakaki 4938; Ans. 4).
FF 8. Examiner finds that Sakaki does “not teach a fusion protein
comprising human COX-1 and human PGIS” (Ans. 4).
FF 9. Lindbladh discloses that “many enzymes that act within a metabolic
pathway interact with each other to form organized enzyme complexes” and
that “[a] considerable body of evidence . . . demonstrates these interactions
and shows differences in kinetics of enzymes which catalyze sequential
metabolic reactions when they are in close proximity compared to the
kinetics of the same enzymes in random solution” (Lindbladh 11692; see
Ans. 5).
FF 10. Lindbladh discloses that “[a] number of bienzymatic fusion proteins
which catalyze sequential reactions have been constructed,” which “have
Appeal 2019-004812
Application 14/885,561

6
shown kinetic benefits for the overall reaction” (Lindbladh 11692; see Ans.
5).
FF 11. Examiner finds that Lindbladh discloses “the preparation of a fusion
protein comprising citrate dehydrogenase and malate dehydrogenase joined
by a linker peptide” (Ans. 5 (citing Lindbladh, Abstract)).
FF 12. Examiner finds that Lindbladh does “not teach a fusion protein
comprising human COX-1 and human PGIS” (Ans. 5).
FF 13. Examiner finds that Hargrave discloses “the structure of bovine
rhodopsin,” which “has seven transmembrane alpha helices” that “vary from
21-28 amino acids, and that this is a length that is quite adequate to span the
distances of the fatty acid side chains in the lipid bilayer” (Ans. 5).
FF 14. Examiner finds that Hargrave does “not teach a fusion protein
comprising human COX-1 and human PGIS” (Ans. 5).
FF 15. Appellant discloses Km values of a membrane-bound COX-2–10aa
(linker)–PGIS fusion protein compared to those “previously characterized by
several groups” for individual COX2 and PGIS enzymes as follows:

Appeal 2019-004812
Application 14/885,561

7
(Spec. ¶ 93; see also id. ¶ 68 (Appellant discloses a “COX-2–10aa–PGIS”
fusion protein exhibiting a Km value of “~3.2 µM” (see also Ruan ’06 8
14009: Fig. 6(A) (disclosing a Km of 3.2 µM for a COX-2–10aa–PGIS
fusion protein)))).
ANALYSIS
Examiner finds that the COX-1 and PGIS enzymes are topologically
arranged in an ER membrane to act cooperatively as part of an extremely
important pathway and, thus, those of ordinary skill in this art would have
found it prima facie obvious to characterize the interaction of these enzymes
(see Ans. 6; see also id. at 3–4; FF 1–4). In this regard, Examiner directs
attention to Lindbladh’s disclosure of proximity effects associated with
organized enzyme complexes and, specifically to a fusion protein model
designed to study such proximity effects (see Ans. 6; FF 9–11; see also FF
6–7 (disclosing the interaction of cooperating parts of a dual enzyme fusion
protein)). Looking to the topological orientation of COX-1 and PGIS in the
ER membrane, as disclosed by Ruan ’05 and Ruan ’04, Examiner reasons
that a person of ordinary skill in this art would have found it prima facie
obvious to utilize known methods, such as those disclosed by Sakaki and
Lindbladh, to link the known sequences of the COX-1 and PGIS “proteins
via a peptide linker that is part of a transmembrane domain, such as the
Rhodopsin transmembrane helices,” which are also known in the art, such
that the COX-1 and PGIS “enzymes are in opposite directions at a distance
similar to the distance between PGIS and COX-1 found in nature” (see Ans.

8 Ke-He Ruan et al., Engineering of a Protein with Cyclooxygenase and
Prostacycline Synthase Activities That Converts Arachidonic Acid to
Prostacyclin, 45 Biochemistry 14003–11 (2006).
Appeal 2019-004812
Application 14/885,561

8
6; see also FF 1–4, 6, 7, 9–11, and 13). Thus, Examiner reasons, because
such a fusion protein construct would be
expected to locate to the ER membrane, placing the N-terminus
of human PGIS next to the C-terminus of the linker (Nter-COX-
Cter-linker-Nter-PGIS-Cter) is the only possible configuration that
would allow the N-terminus of human PGIS to be in a position
similar to that found in nature, namely closest to the ER
membrane, by virtue of its proximity to the linker.
(Ans. 7.)
Therefore, based on the combination of Ruan ’05, Ruan ’04, Sakaki,
Lindbladh, and Hargrave, Examiner concludes that, at the time Appellant’s
invention was made, it would have been prima facie obvious to have a
fusion protein comprising human COX-1 and human PGIS, wherein the C-
terminus of the human COX-1 protein is linked to the N-terminus of the
human PGIS by an alpha-helix peptide from the bovine rhodopsin
transmembrane domain (Ans. 6). According to Examiner, “[a] person of
ordinary skill in the art [would have been] . . . motivated to make said
fusion protein, purify it and place it in a buffer solution, for the benefit of
placing together two enzymes of a well-known coupling reaction
system . . . [to] further characterize the interaction of both proteins” (id.).
As discussed above, COX-1, PGIS, and transmembrane linker
sequences were known in the art at the time of Appellant’s claimed
invention (see FF 4 and 13). Therefore, we are not persuaded by Appellant’s
intimation that the components necessary to produce Appellant’s claimed
fusion protein were not known in the art at the time of Appellant’s claimed
invention (see Appeal Br. 13).
As further discussed above, Sakaki and Lindbladh each disclose
methods of producing bienzymatic fusion proteins which catalyze sequential
Appeal 2019-004812
Application 14/885,561

9
reactions (see FF 6, 7, and 9–11). Lindbladh further discloses that “[a]
number of bienzymatic fusion proteins which catalyze sequential reactions
have been constructed,” which “have shown kinetic benefits for the overall
reaction,” suggesting the routine nature of producing such constructs (FF
10). Thus, at the time of Appellant’s claimed invention, those of ordinary
skill in this art understood how to construct an isolated hybrid protein
molecule such as that required by Appellant’s claimed invention without
undue experimentation. Therefore, we are not persuaded by Appellant’s
contention that Examiner improperly relied upon Official Notice of facts to
establish that methods of producing a fusion protein within the scope of
Appellant’s claimed invention were known in the art at the time of
Appellant’s claimed invention (Appeal Br. 14; see Reply Br. 2 (Appellant
contends that “no prior art can be cited as providing a teaching as to a
method of making the protein”)).
Appellant failed to establish an evidentiary basis to support a
conclusion that the evidence relied upon by Examiner would have been
incapable of producing a functional fusion protein within the scope of
Appellant’s claim 1 (see e.g., FF 4, 6, 7, 9–11, and 13). For the foregoing
reasons, we are not persuaded by Appellant’s unsupported contentions
regarding “a plethora of parameters including,” inter alia, “pH, temperature,
concentration of reagents,” etc., which Appellant contends are required “in
order to achieve the correct folding of each of the protein subunits in which
both enzymatic sites are so positioned that they are complementary to each
other” (Appeal Br. 13; see also Reply Br. 3). For the foregoing reasons, we
are not persuaded by Appellant’s intimation that the combination of
references relied upon by Examiner fails to provide guidance necessary to
Appeal 2019-004812
Application 14/885,561

10
produce the claimed fusion protein (see Appeal Br. 13; Reply Br. 2
(Appellant contends that “one of ordinary skill has no recourse other tha[n]
to undertake the considerable burden of undue experimentation in order to
arrive at the hybrid protein molecule of claim 1”)). See In re Pearson, 494
F.2d 1399, 1405 (CCPA 1974) (“Attorney’s argument in a brief cannot take
the place of evidence.”).
For the foregoing reasons, we are not persuaded by Appellant’s
contention that the combination of prior art relied upon by Examiner fails to
enable the production of Appellant’s claimed fusion protein (see Reply Br.
2).
Appellant contends that “[t]he inventors of the current application
made a number of different fusion proteins before successfully achieving the
claimed isolated fusion protein with the required Km of 3.2µM to 4 µM”
(Appeal Br. 15 (citing Ruan ’06); see generally Reply Br. 3). Appellant
further contends that
in order to obtain a hybrid protein with the same K[m] value as
the hybrid protein molecule of claim 1, the prior art references
must teach an identical hybrid protein sequence and one that is
folded in an identical manner, which would further require the
identical electrostatic environment as dictated by the specific
experimental reagents used and as taught by the instant
[S]pecification.
(Reply Br. 3.) We are not persuaded. Although Appellant discloses Km
data for a COX-2–10aa–PGIS fusion protein, Appellant’s claim 1 is not
limited to this construct. To the contrary, Appellant’s claim 1 relates to a
fusion protein comprising COX-1, not COX-2, and a linker of undefined
length (see Appeal Br. 17). Therefore, we are not persuaded by Appellant’s
Appeal 2019-004812
Application 14/885,561

11
contentions relating to a COX-2–10aa–PGIS fusion protein (see id. at 15–
16).
For the foregoing reasons we are not persuaded by Appellant’s
contention that Examiner relied upon impermissible hindsight (Appeal Br.
13).
CONCLUSION
The preponderance of evidence relied upon by Examiner supports a
conclusion of obviousness. The rejection of claim 1 under 35 U.S.C.
§ 103(a) as unpatentable over the combination of Ruan ’05, Ruan ’04,
Sakaki, Lindbladh, and Hargrave is affirmed. Claims 2, 3, 5, 16, 19, and 20
are not separately argued and fall with claim 1.
DECISION SUMMARY
In summary:
Claims
Rejected
35 U.S.C. § Reference(s)/Basis Affirmed Reversed
1–3, 5, 16,
19, 20
103 Ruan ’05, Ruan ’04,
Sakaki, Lindbladh,
Hargrave
1–3, 5, 16,
19, 20

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