Ex Parte Subtil et al

Board of Patent Appeals and Interferences

Oct 26, 2009

10014670 (B.P.A.I. Oct. 26, 2009)

UNITED STATES PATENT AND TRADEMARK OFFICE
____________

BEFORE THE BOARD OF PATENT APPEALS
AND INTERFERENCES
____________

Ex parte AGATHE SUBTIL,
CLAUDE PARSOT, and ALICE DAUTRY-VARSAT
____________

Appeal 2009-0029311
Application 10/014,670
Technology Center 1600
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Decided: October 26, 2009
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Before LORA M. GREEN, RICHARD M. LEBOVITZ, and
FRANCISCO C. PRATS, Administrative Patent Judges.

LEBOVITZ, Administrative Patent Judge.

DECISION ON APPEAL
This is a decision on the appeal by patent applicants from the patent
examiner’s rejection of claims 7-10, 34-37, and 44-47. Jurisdiction for this
appeal is under 35 U.S.C. § 6(b). We affirm.

1 Heard Sep. 17, 2009.

Appeal 2009-002931
Application 10/014,670

STATEMENT OF THE CASE
The claims are directed to methods of expressing and secreting
Chlamydia polypeptides in gram-negative bacteria strains containing a type
III secretion pathway. Chlamydiae are gram-negative bacteria that
proliferate only within eukaryotic host cells, causing diseases such as
pneumonia and pelvic inflammatory disease (Spec. 1). The type III
secretion machinery facilitates the translocation of Chlamydia proteins into
the host cell’s cytoplasm where the bacteria reside (id. at 5).
Claims 7-10, 34-37, and 44-47 are appealed and stand rejected by the
Examiner as follows:
Claims 7-10, 34-37, and 44-47 under 35 U.S.C. § 103(a) as obvious in
view of Griffais et al. (US 6,559,294 B1, issued May 6, 2003), Demers et al.
(WO 99/58714, published Nov. 18, 1999), and Kalman et al. (Comparative
genomes of Chlamydia pneumoniae and C. trachomatis, 21 NATURE
GENETICS 385-389 (1999)) (Ans. 3); and
Claims 7-10, 34-37, and 44-47 under 35 U.S.C. § 103(a) as obvious in
view of Stephens et al. (US 6,822,071 B1, issued Nov. 23, 2004) and
Demers (Ans. 6).
Claims 7, 8, and 9 are representative and read as follows:
7. A method for identifying a secreted Chlamydia polypeptide
wherein said method comprises (a) providing a recombinant expression
vector containing at least DNA coding for the polypeptide of interest; (b)
transforming a Gram-negative strain containing a type III secretion pathway
with said recombinant vector; (c) expressing said vector in said Gram-
negative transformed strain; and (d) detecting the secretion of said DNA
expression product; wherein the secretion of said expression product
indicates that it corresponds to a secreted Chlamydia polypeptide.

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8. A method for identifying a secreted Chlamydia polypeptide
wherein said method comprises (a) providing a recombinant expression
vector containing at least DNA coding for the polypeptide of interest fused
to a reporter gene; (b) transforming a Gram-negative strain containing a type
III secretion pathway with said recombinant vector; (c) expressing this
vector in said transformed Gram-negative strain; and (d) detecting the
secretion of said reporter gene expression product; wherein the secretion of
said expression product indicates that the fused DNA contains at least a
polynucleotide corresponding to a secreted Chlamydia polypeptide.

9. A method according to Claims 7 or 8 wherein said Gram-negative
strain containing a type III secretion pathway is a Shigella strain.

THE CLAIMS
1. Claim 7 is to a method “for identifying a secreted Chlamydia
polypeptide” comprising four steps:
2. (a) providing an expression vector containing a DNA coding for a
polypeptide of interest;
3. (b) transforming a “Gram-negative strain containing a type III
secretion pathway” with the vector;
4. (c) expressing the vector in the Gram-negative strain; and
5. (d) detecting secretion of the DNA “wherein the secretion of said
expression product indicates that it corresponds to a secreted Chlamydia
polypeptide.”
6. Claim 8 is to a similar four step method as in claim 7, but the (a) DNA
coding for a polypeptide of interest is “fused to a reporter gene.”
7. Claim 9 is to the method of claim 7 or 8 in which the gram-negative
strain “is a Shigella strain.”

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OBVIOUSNESS – GRIFFAIS, DEMERS, & KALMAN
Claims 7-10, 34-37, and 44-47 stand rejected under 35 U.S.C. §
103(a) as obvious in view of Griffais, Demers, and Kalman (Ans. 3).

Facts
Demers
8. “Type III secretion machinery is present in numerous gram-negative
bacteria (including members of the species Shigella, Salmonella, Yersinia,
Escherichia, Pseudomonas, Xanthomonas, Ralstonia, and Erwinia)” (1:11-
13).
9. Demers describes methods of identifying molecules which activate or
inhibit secretion in wild-type strains of gram-negative bacteria (3:1-3).
10. The methods involve expressing a reporter gene fused to a promoter of a
gene regulated by type III secretion machinery in wild-type gram-negative
bacteria and “mutants of these bacteria that constitutively secrete proteins
via the type III secretion machinery or are deficient for secretion” (3:4-13).
11. The presence or activity of the reporter gene “can be used as an
indicator of the secretion activity of the type III secretion machinery” (3:13-
16).
12. Demers states that “[a]ny gram-negative bacterial containing type III
secretion machinery may be used in the methods” (3:23-26), and describes
specific examples using Shigella (6:1-3).
13. Demers describes the detection of 15 proteins secreted by Shigella in its
model system (7:2-25).

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Griffais
14. Griffais describes host cells transformed with a vector comprising
Chlamydia nucleotide sequences, and the expression and secretion of the
polypeptides encoded by the sequences (col. 1, ll. 15-22; col. 48, ll. 25-65;
col. 50, ll. 41-51).
15. The secreted polypeptides include type III secreted polypeptides which
“may be detected by techniques known to persons skilled in the art, such as
for example the techniques using cloning combined with vectors allowing
the expression of the said polypeptides fused to export markers such as the
luc gene for luciferase or the PhoA gene for alkaline phosphatase.” (Col. 40,
ll. 20-27.)
16. The “vectors comprise the elements necessary to allow the expression
and/or the secretion of the said nucleotide sequences in a given host cell”
and may include “particular signals specifying the secretion of the translated
protein” (col. 46, ll. 14-23).
17. “[P]referred” host cells are gram-negative bacteria and “a bacterium
belonging to the Chlamydia family” (col. 48, ll. 59-64).
Kalman
18. Kalman describes various Chlamydia polypeptides, including type III
secreted polypeptides (p. 385, col. 2).

Differences between the claimed invention and the prior art
19. As found by the Examiner, Griffais taught the expression vector,
transformation of a gram-negative host with the vector, expression of the
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vector, and the detecting steps recited in steps (a) through (d) of claims 7 and
8 method (F14-F17; Ans. 4).
20. The secreted polypeptides described in Griffais were Chlamydia
polypeptides (F14) as in the claims.
21. Griffais also disclosed that the Chlamydia polypeptide could be fused to
an “export marker” (F15) which served the same purpose as the “reporter
gene” of claim 8. Demers also taught the use of a reporter gene to indicate
secretion activity of type III secretion machinery (F10-11).
22. The Examiner found that Griffais did not describe utilizing a Shigella
host as recited in claim 9, but found that Demers taught such an expression
host (F12) in a similar expression method, and concluded that it would have
been obvious to have used it as a “known technique” to “yield predictable
results.” (Ans. 4 & 10-11.)

Level of ordinary skill in the art
23. Based on the teachings of Demers (F12-13) and Griffais (F17), persons
of ordinary skill in the art would have believed it reasonably predictable that
Chlamydia polypeptides would be successfully secreted in other gram-
negative bacteria, including Shigella.

Statement of Issue
Appellants contend that the Examiner erred for two reasons (App. Br.
6):
• Persons of ordinary skill in the art would not have used Demers’
secretion system because Demers’ “entire disclosure is directed to looking
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for agents that block secretion or change expression patterns NOT for
determining whether a certain Chlamydia protein is one that can be secreted
through the type III pathway.” (App. Br. 7.)
• The “cited art provides no reason to believe that the expression of
Chlamydia proteins would, in fact, work in other gram negative strains such
as Shigella.” (App. Br. 7.)
Therefore, the issues in this rejection are whether Appellants
established that the Examiner erred in combining Demers with Griffais and,
in doing so, also erred in concluding that there was a reasonable expectation
of success.

Principles of Law
An obviousness “analysis need not seek out precise teachings directed
to the specific subject matter of the challenged claim, for a court can take
account of the inferences and creative steps that a person of ordinary skill in
the art would employ.” KSR Int’l Co. v. Teleflex Inc., 550 U.S. 398, 418
(2007).
In assessing a claim’s obviousness, “a court must ask whether the
improvement is more than the predictable use of prior art elements according
to their established functions.” Id. at 417. “The combination of familiar
elements according to known methods is likely to be obvious when it does
no more than yield predictable results.” Id. at 416.

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Analysis
Appellants contend that the Examiner erred in combining Demers
with Griffais because the purpose of Demers was to identify agents that
blocked secretion, not to determine whether a Chlamydia polypeptide could
be secreted by a type III pathway. This argument does not persuade us that
the Examiner erred.
First, the Examiner relied upon Demers for its teaching that various
gram-negative hosts, including Shigella, could be used for secreting a type
III polypeptide. Thus, even were the methods to differ between Griffais and
Demers, the Examiner did not solely rely on Demers for meeting the
limitations of steps (a) through (d) of claims 7 and 8.
Secondly, Demers describes steps of expressing and detecting type III
polypeptides in gram-negative bacteria (F10-13) as in claims 7 and 8, and
therefore was properly combined with Griffais, which was directed to the
same type of expression methods. As both references are in the same field
of endeavor, persons of ordinary skill in the art would have found them
pertinent to the problem addressed by the claim of expressing polypeptides
in hosts with type III expression machinery.
Finally, the claimed method recites “comprising” which is open-ended
language that does not exclude additional steps, including a step of
identifying molecules that regulate secretion in gram-negative bacteria as in
the Demers publication (F9). Consequently, Appellants’ argument that
Demers’ purpose is different from the claimed invention is not persuasive
because the claims do not exclude Demers’ molecule identification method.
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Appellants also assert that the inventors’ discovery that Chlamydia
proteins could be expressed and secreted in Shigella would not have been
predicted by the prior art (App. Br. 7). They assert that the protein
sequences utilized in the Chlamydia secretion machinery are not conserved
as compared to other gram-negative bacteria and therefore the secretion
signals would differ (id.). They also cited the abstracts of several
publications which they contend demonstrate that other proteins, such as
chaperone proteins, are needed to achieve secretion in gram-negative
bacteria (id. at 8).
Appellants have not provided adequate evidence that persons of skill
in the art would have viewed expression of a Chlamydia polypeptide in other
gram-negative host as unpredictable. Attorney argument is not evidence. In
re Pearson, 494 F.2d 1399, 1405 (CCPA 1974). Nor can it take the place of
evidence lacking in the record. Meitzner v. Mindick, 549 F.2d 775, 782
(CCPA 1977). For example, Appellants did not provide evidence that the
secretion signals in a Chlamydia protein would not be recognized in Shigella
or other gram-negative bacteria.
The Examiner, on the other hand, introduced Demers’ teaching that
any gram-negative bacteria containing type III secretion machinery could be
utilized in its expression methods (F12-13) and the Griffais statement that
gram-negative bacteria were preferred hosts for Chlamydia polypeptide
secretion (F17), evidencing that persons of ordinary skill in the art believed
it predictable that a Chlamydia polypeptide could be secreted in gram-
negative hosts, including Shigella.
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As to the presence of chaperones,2 the Parsot abstract establishes that
chaperones are involved in the type III secretion pathway of gram-negative
bacteria; the Tucker abstract describes chaperones in Salmonella; and the
Fields and Slepenkin abstracts relate to chaperones in Chlamydia (App. Br.
8). Appellants have not explained how the presence of chaperones in gram-
negative bacteria, including Chlamydia, would make expression in gram-
negative hosts unpredictable. To the contrary, the abstracts provide further
evidence of the similarities in type III secretion pathways in gram-negative
bacteria, further buttressing the evidence of predictability provided by
Demers and Griffais (F23).
For the forementioned reasons, we conclude that Appellants did not
establish that the Examiner erred in reaching the conclusion that the claimed
subject matter would have been obvious persons of ordinary skill in the art.

OBVIOUSNESS – STEPHENS AND DEMERS
Claims 7-10, 34-37, and 44-47 stand rejected under 35 U.S.C. §
103(a) as obvious in view of Stephens and Demers (Ans. 6).

Facts
Stephens
24. Stephens describes expression of nucleic acids coding for secreted
Chlamydia polypeptides in host cells using “routine techniques” and

2 Appellants cited abstracts of articles by Parsot, Tuckerdagger et al., Fields
et al. and Slepenkin et al. (App. Br. 8).
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purifying the secreted polypeptide (col. 15, ll. 25-33 & 40-43; col. 16, ll. 4-
15 & 42-44).
25. Stephens states that well-known cells and cell lines, such as E. coli, can
be used as hosts to express the Chlamydia polypeptides (col. 16, ll. 16-20).

Differences between the claimed invention and the prior art
26. Stephens, as found by the Examiner and supported by the evidence,
describes “a method of identifying a secreted Chlamydia polypeptide
comprising using a vector and transforming the cells into host cells (columns
15 and 16),” expression in Escherichia coli which is a gram negative
bacteria (F8), and identifying the secreted polypeptide (Ans. 6; F24-25) as in
claims 7 and 8.
27. The Examiner found that Stephens does not “teach the claim limitation
‘wherein said gram-negative strain containing a type III secretion pathway is
a Shigella strain,” but concluded that it would have been obvious to the
skilled worker because Demers teaches “that gram-negative bacteria contain
type III secretion machinery and can secrete proteins via this machinery
(pages 1 and 2). Demers et al teach that Shigella species can be used to
secrete proteins (pages 1 and 6-9).” (App. Br. 7.) The Examiner’s finding is
supported by the evidence.

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Level of ordinary skill in the art
28. Based on the teachings described in Stephens (F24-25) and Demers
(F12-13), persons of ordinary skill in the art would have believed it
reasonably predictable that Chlamydia polypeptides would be successfully
secreted in other gram-negative bacteria, including Shigella (see App. Br. 7).

Statement of the Issue
Appellants contend that neither “Stephens nor Demers describes a
method for identifying secreted proteins but rather the general methodology
for expressing proteins (which is even acknowledged by Stephens in column
15)” and that therefore the combined references fail to teach or suggest the
claimed invention (App. Br. 9). Appellants also contend that “it is known
that the proteins which make the Chlamydia secretion machinery are not
well conserved compared” to those of other bacteria” (id. at 10).
The issues are therefore whether Appellants have established that the
Examiner erred in finding that Stephens and Demers suggested all the steps
of the claimed method and whether persons of ordinary skill in the art would
have reasonably expected Chlamydia polypeptides to be expressed in gram
negative bacteria, such as Shigella.

Analysis
The Examiner’s findings that Stephens describes steps (a) through (d)
of claim 7 and 8 are supported by factual findings (see F24-26; Ans. 6).
Appellants have not identified any specific error in these findings nor do we
find one. As to Appellants’ argument that it would have been unpredictable
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that a Chlamydia polypeptide would be secreted by other gram-negative
bacteria, such as Shigella, Appellants have not presented any countervailing
evidence or reason to doubt the statements in Stephens (F25) and Demers
(F12-13) that any gram-negative host would be suitable to secrete a
Chlamydia polypeptide. The Examiner’s position that persons of ordinary
skill in the art would have reasonably believed it predictable that Chlamydia
polypeptides would be successfully secreted in other gram-negative bacteria,
including Shigella (F28) is fact-based and supported by the evidence.

CONCLUSIONS OF LAW & SUMMARY
The Examiner did not err in finding all the limitations of claims 7 and
8 to have been taught or suggested by the prior art. The Examiner did not
err in concluding that the ordinary skilled worker would have reasonably
believed that a Chlamydia polypeptide would be successfully secreted in
gram-negative bacteria, including Shigella as recited in claim 9. The
obviousness rejections of claims 7-9 are affirmed.
Claims 10, 34-37, and 44-47 fall with claims 7-9 because separate
reasons for their patentability were not provided. See 37 C.F.R.
§ 41.37(c)(1)(vii).

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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

cdc

OBLON, SPIVAK, MCCLELLAND MAIER & NEUSTADT, L.L.P.
1940 DUKE STREET
ALEXANDRIA VA 22314

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