Ex Parte Bossard et al

Patent Trial and Appeal Board

Dec 12, 2017

12991402 (P.T.A.B. Dec. 12, 2017)

United States Patent and Trademark Office
UNITED STATES DEPARTMENT OF COMMERCE
United States Patent and Trademark Office
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APPLICATION NO. FILING DATE FIRST NAMED INVENTOR ATTORNEY DOCKET NO. CONFIRMATION NO.
12/991,402 01/18/2011 Mary J. Bossard SHE0169.00 8407
21968 7590 12/13/2017
NEKTAR THERAPEUTICS
455 Mission Bay Blvd., South, Suite 100
San Francisco, CA 94158
EXAMINER
COFFA, SERGIO
ART UNIT PAPER NUMBER
1675
MAIL DATE DELIVERY MODE
12/13/2017 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 MARY J. BOSSARD, HAROLD ZAPPE,
SEOJU LEE, and LAL AJITHA RATNAPALA FERNANDO
Appeal 2017-001123
Application 12/991,402
Technology Center 1600
Before JEFFREY N. FREDMAN, JOHN G. NEW, and
RICHARD J. SMITH, Administrative Patent Judges.
FREDMAN, Administrative Patent Judge.
DECISION ON APPEAL
This is an appeal1 under 35U.S.C. § 134 involving claims to a
composition comprising a di-PEGylated dimer conjugate. The Examiner
rejected the claims as obvious. We have jurisdiction under 35 U.S.C. § 6(b).
We affirm.
Statement of the Case
Background
“Acetylcholine is involved with the transmission of signals from
specialized motor nerves to the skeletal muscle as well as much of the
autonomic nervous system” (Spec. 14). “[Ajcetylcholine is degraded — and
1 Appellants identify the Real Party in Interest as Nektar Therapeutics (see
App. Br. 3).
Appeal 2017-001123
Application 12/991,402
therefore its effects controlled — by acetylcholinesterase located in the
synaptic cleft.” (id.). “The balance of the acetylcholine/acetylcholinesterase
system in the central nervous system can be disrupted through exposure [to]
an acetylcholinesterase inhibitor . . . Such a disruption, if allowed to
continue, can cause any number of deleterious conditions and — if severe —
even death” (id. 1 5). “Exposure to cholinesterase inhibitors can be
remedied by the administration of cholinesterase itself. By effectively
‘saturating’ the biological system with cholinesterase, overall normal
cholinesterase functioning would remain substantially unaffected” (id. 17).
The Claims
Claims 1, 10, 11, 13—16, 23, 24, and 28 are on appeal. Independent
claim 1 is representative and read as follows:
1. A composition comprising a di-PEGylated dimer
conjugate comprising two polyethylene glycol )s attached to a
dimer derived from two separate butyrylcholinesterase
monomers, each monomer that forms the dimer having one
polyethylene glycol) having a total weight-average molecular
weight in the range of greater than 5,000 Daltons to about
100,000 Daltons covalently attached through a cysteine residue
corresponding to Cys66 of butyrylcholinesterase, and further
wherein at least 60% of the conjugates in the composition are
di-PEGylated dimer conjugates.
The Issue
The Examiner rejected claims 1, 10, 11, 13—16, 23, 24, and 28 under
35 U.S.C. § 103(a) as obvious over Huang,2 Rosendahl,3 and Bossard4 (Final
Act. 2-7).
2 Huang et al., WO 2008/019036 A2, published Feb. 14, 2008.
3 Rosendahl, M., WO 2006/063055 A2, published June 15, 2006.
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Application 12/991,402
The Examiner finds Huang teaches “a butyrylcholinesterase (BChE)
protein chemically linked to polyethylene glycol (PEG). . . wherein said
PEG is attached to a thiol group of said BChE” (Final Act. 2—3). The
Examiner find Huang’s “product [is] mostly dimers (thus, about 2 PEGs per
dimer)” (id. at 3).
The Examiner acknowledges Huang doesn’t teach linkage “to Cys66
of butyrylcholinesterase” or the specific polymeric reagent (Final Act. 4).
The Examiner finds Rosendahl teaches “a polyethylene glycol is
attached to at least one lysine or cysteine residue in [a] butyrylcholinesterase
protein” and “the cysteine residue can include, but is not limited to, Cys66
and Cys571” (Final Act. 4). The Examiner finds Rosendahl further explains
“BuChE contains 8 native cysteines, six of which form 3 disulfides (C65-
C92, C252-C263, C400-C519) and two (C66, C571) that remain free (i.e.
cysteine residues not involved in disulfide bonds). . . These free cysteines
are reasonable sites for thiol specific PEGylation” (id. at 5).
The Examiner finds Bossard teaches the specific conjugation agent of
claim 13 (see Final Act. 6).
The Examiner finds it obvious “to PEGylate Cys66 in the invention of
Huang et al and arrive at the instantly claimed invention because Rosendahl
teaches the Cys66 is free and is a reasonable site for thiol specific
PEGylation” (Final Act. 6).
Appellants contend “Rosendahl’s teachings that Cys66 is a ‘free
cysteine’ and that ‘free cysteines are reasonable sites for thiol-specific
PEGylation’ fails to portray the complete story” (App. Br. 6). Appellants
4 Bossard et al., US 2006/0036080 Al, published Feb. 16, 2006.
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Application 12/991,402
contend “Rosendahl’s failure is evident at least in view of Lockridge et al.
(1987) JBiol. Chem. 262(27): 12945—12952 (referenced in paragraph [0169]
of Appellant’s specification” and Appellants’ cite Lockridge as teaching:
Residue 66 may be a free sulfhydryl. However, we were unable
to specifically alkylate it with [3H Jiodoacetic acid. Cys66 was
successfully alkylated only under conditions which alkylated all
8 cysteines, that is following reduction with dithiothreitol in the
presence of 6 M guanidine chloride, 1 mM EDTA, pH 8.0.
(App. Br. 7, citing Lockridge 12948, col. 1). Appellants contend
“Rosendahl’s discussion of preparing conjugates of butrylcholinesterase at
its ‘free cysteines,’ such as Cys66, using conventional methods is more than
simply not enabled; Rosendahl’s discussion is contrary to the teachings of
those researchers — such as Lockridge et al. — that demonstrated the
inaccessible nature of native butyrylcholinesterase’s Cys66” (App. Br. 7).
The issue with respect to this rejection is: Does the evidence of record
support the Examiner’s conclusion that Huang and Rosendahl render
obvious the linkage of PEG to Cys66 of butyrylcholinesterase as required by
claim 1?
Findings of Fact
1. Huang teaches “PEGylated (meaning attached to PEG -
polyethylene glycol) recombinant butyrylcholinesterase (PEG-BChE)”
(Huang 4:11—13).
2. Huang teaches “PEG is attached to a thiol group of said BChE”
and “the PEG has a molecular weight of 5,000 to 500,000 kilodaltons”
(Huang 18:32 to 19:11).
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Application 12/991,402
3. Huang teaches “a suitable ratio of activated PEG to BChE is
about 80 to 1, which is found to produce a 1:1 ratio of PEG to BChE
monomeric unit, with the product mostly dimers (thus, about 2 PEGs per
dimer)” (Huang 20:8—10).
4. Rosendahl teaches “a polyethylene glycol is attached to at least
one lysine or cysteine residue in said butyrylcholinesterase protein. For
example, the cysteine residue can include, but is not limited to, Cys66 and
Cys571” (Rosendahl 3:17-19).
5. Rosendahl teaches “BuChE contains 8 native cysteines, six of
which form 3 disulfides (C65-C92, C252-C263, C400-C519) and two (C66,
C571) that remain free (i.e. cysteine residues not involved in disulfide
bonds) (all positions given are relative to SEQ ID NO: 1). These free
cysteines are reasonable sites for thiol specific PEGylation” (Rosendahl
13:32 to 14:2).
6. Rosendahl teaches the “cysteine muteins must be partially
reduced with dithiothreitol (DTT), Tris (2-carboxyethyl) phosphine-HCI
(TCEP) or other disulfide disrupting agent, in order to expose the free
cysteine for PEGylation and allow the PEGylation reaction to proceed
efficiently” (Rosendahl 37:17—20).
7. Rosendahl cites, among other references, “Lockridge, O.,
Adkins, S. and La Du, B. (1987) J. Biol. Chem., 262:12945-12952. Location
of disulfide bonds within the sequence of human serum cholinesterase”
(Rosendahl 44:29-30).
8. Lockridge teaches: “Residue 66 may be a free sulfhydryl.
However, we were unable to specifically alkylate it with [3H]iodoacetic acid.
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Cys66 was successfully alkylated only under conditions which alkylated all 8
cysteines, that is following reduction with dithiothreitol in the presence of 6
M guanidine chloride, 1 mM EDTA, pH 8.0” (Lockridge 12948, col. 1).
9. Lockridge teaches:
Attempted Alkylation of Free Sidfhydryl-1) 6.5 mg of
cholinesterase in 1.5 ml of 6 M guanidine HC1,50 mM Tris-Cl, 1
mM EDTA, pH 8.4, was made anaerobic by blowing nitrogen
for 1.5 h. 25 pi of 6.66 mM [3H]iodoacetic acid (150 Ci/mol,
Amersham Corp.) was added, and the reaction was allowed to
proceed for 1 h.
(Lockridge 12946, col. 1). Lockridge finds no radioactivity associated with
“any particular peptide”, indicating no reaction with cys66 (see id.).
10. The Specification teaches
it is possible to carry out a reducing step in a method for
preparing conjugates. A reducing step can be carried out using
techniques known to one of ordinary skill in the art. Lor
example, a reducing step can be carried out by subjecting a
protein to reducing conditions, e.g., addition of a reducing agent
such as 2-mercaptoethanol, dithiothreitol, or tris(2-
carboxyethyl (phosphine.
(Spec. 1122).
Principles of Law
“A reference may be said to teach away when a person of ordinary
skill, upon reading the reference, would be discouraged from following the
path set out in the reference, or would be led in a direction divergent from
the path that was taken by the applicant.” In re Gurley, 27 L.3d 551, 553
(Led. Cir. 1994). The degree of teaching away, however, depends on the
particular facts. See id.
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Analysis
We adopt the Examiner’s findings of fact and reasoning regarding the
scope and content of the prior art (Final Act. 2—7; FF 1—10) and agree that
the claims are rendered obvious by Huang, Rosendahl, and Bossard. We
address Appellants’ arguments below.
Appellants contend “even if Rosendahl is correct in stating that ‘free
cysteines are reasonable sites for thiol-specific PEGylation,’ the analysis
does not end there. As confirmed by the teachings of Fockridge et al.,
successful modification of a free cysteine is also contingent upon the free
cysteine being accessible” (App. Br. 7). Appellants contend Fockridge
“demonstrated the inaccessible nature of native butyrylcholinesterase’s
Cys66” (id. ). Appellant conclude “one of ordinary skill in the art would not
have had a reasonable expectation of successfully modifying
butyrylcholinesterase’s Cys66 through application of the general approaches
suggested in each of Huang et al., Rosendahl and Bossard” (id. at 8).
While we agree with Appellants that Fockridge provides some
evidence suggesting difficulties for a process of PEGylation of Cys66 of
butyrylcholinesterase (FF 8—9), Rosendahl was aware of, and actually cites,
Fockridge (FF 7) and still expressly teaches butyrylcholinesterase contains
“two (C66, C571) [cysteine residues] that remain free (i.e. cysteine residues
not involved in disulfide bonds). . . These free cysteines are reasonable sites
for thiol specific PEGylation” (FF 5). Thus, the later filed prior art
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Application 12/991,402
Rosendahl5 reference directly suggests PEGylation at Cys66 even when
aware of Lockridge’s teachings.
Moreover, Rosendahl provides a specific process for efficient
PEGylation that requires the protein “be partially reduced with dithiothreitol
(DTT), Tris(2-c[ar]boxyethyl)phosphine-HCl (TCEP) or other disulfide
disrupting agent” (FF 6). This reduction process is comparable to the
effective reduction process identified by the Specification (FF 10).
Moreover, it is similar to the successful alkylation of Cys66 in Fockridge,
where Fockridge also reduces the protein with dithiothreitol, while also
adding in the denaturant guanidine (FF 8). Fockridge does not provide a
comparison in which only the reducing agent, without the denaturant, was
added, as suggested by both Rosendahl (FF 6) and the instant Specification
(FF 10).
The Rosendahl process is also consistent with Appellants’ argument
that the “specification describes a successful approach for preparing ‘di-
PEGylated dimers of butyrylcholinesterase’ at Cys66. In the approach of
Example 3, a process of ‘reducing’ the individual (i.e., ‘monomer’) form of
native butyrylcholinesterase is described” (App. Br. 7).
5 We note that in the Answer the Examiner cited to Tomlinson et. al., S-
Mercuric-N-Dansyl-Cysteine Labels the Free Sulfhydryl Groups of Human
Serum Cholinesterase, 158 Biochem. Biophys. Res. Comm. 503-7 (1989)
who addressed and disagreed with Fockridge (see Tomlinson 504), instead
finding “Cys-66 appears to be present in the free sulfhydryl form in BChe”
(see Tomlinson 503, abstract). We do not, however, rely upon Tomlinson
because the reference was not addressed in the Examiner’s Final Action,
denying Appellants an opportunity to respond.
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Application 12/991,402
Thus, while Appellants’ arguments and Lockridge provide some
degree of teaching away / reasonable expectation of success concern of
PEGylating Cys66, when these findings are balanced against the specific
reducing process disclosed by Rosendahl for PEGylation (FF 6) and
Rosendahl’s specific identification of Cys66 as a site for PEGylation (FF 5)
while being aware of Fockridge (FF 7), we find that the balance of evidence
supports the Examiner’s obviousness position because “[ojbviousness does
not require absolute predictability of success ... all that is required is a
reasonable expectation of success.'1'’ In re Kubin, 561 F.3d 1351, 1360 (Fed.
Cir. 2009) (citation omitted).
Conclusion of Law
The evidence of record supports the Examiner’s conclusion that
Huang and Rosendahl render obvious the linkage of PEG to Cys66 of
butyrylcholinesterase as required by claim 1.
SUMMARY
In summary, we affirm the rejection of claim 1 under 35 U.S.C.
§ 103(a) as obvious over Huang, Rosendahl, and Bossard. Claims 10, 11,
13—16, 23, 24, and 28 fall with claim 1.
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
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