T 0108/98 (HSV glycoproteins/DADE BEHRING) 27-03-2003

I. An appeal was lodged by the patentee (appellant) against the interlocutory decision of the opposition division, whereby the European patent No. 0 139 416 with the title "Molecularly cloned diagnostic product and method of use" was maintained on the basis of the fifth auxiliary request filed at the oral proceedings before the opposition division. The patent claimed priority from three US applications filed on 30 August 1983, 31 October 1983 and 9 March 1984, respectively.

II. The patent had been opposed under Article 100(a) EPC for lack of novelty and inventive step. The opposition division decided that the main request (claims as granted) lacked novelty under Article 54(3) EPC. The first auxiliary request was found not to fulfill the requirements of Article 56 EPC. The second auxiliary request was refused under Article 123(3) EPC, whereas the third auxiliary request was refused under Article 84 EPC. The fourth auxiliary request was refused under Articles 123(2), 84 and 56 EPC and was considered not to meet the requirements of Rule 57a EPC.

III. Claim 1 as granted (main request) read as follows:

"1. A process which comprises producing in a recombinant, stable, mammalian, continuous cell line a gC or gD glycoprotein polypeptide of Herpes simplex virus type 1 or type 2 whereby the glycoprotein has exposed antigenic determinants capable of specifically binding complementary antibodies of Herpes simplex virus type 1 and/or type 2.".

Claims 2 and 3 were dependent on claim 1 and defined the association of the glycoprotein with the cell membrane. Claim 4 was directed to a process according to claim 1, wherein there was initially produced a truncated derivative of a membrane-bound gC or gD glycoprotein polypeptide of Herpes simplex virus type 1 or type 2 devoid of membrane-binding domain, whereby the derivative polypeptide was free of said membrane and had said exposed antigenic determinants. Claims 5 to 14 were dependent claims directed to further embodiments of the process of claims 1 to 4. Independent claim 15 was directed to a diagnostic test kit comprising the truncated, membrane-free polypeptide as obtainable by the process according to claim 4, claims 16 to 24 defining further embodiments thereof. Independent claims 25 and 29 were concerned with different methods of detection using said truncated, membrane-free polypeptide, claims 26 to 28 and claims 30 to 31 defining further embodiments thereof, respectively.

IV. The fifth auxiliary request accepted by the opposition division was concerned only and exclusively with subject-matter related to Herpes simplex virus type 2 glycoprotein C (gC-2).

V. With the statement of grounds of appeal on 28 March 1998, the appellant filed a second, fourth and fifth auxiliary requests. A first and a third auxiliary requests were filed on 25 February 2003 and on 21. October 2002, respectively. The fifth auxiliary request was the one accepted by the opposition division (cf Section IV supra), whereas all other auxiliary requests corresponded to the ones treated during the opposition proceedings but with the correction of several omissions ("specifically" and "complementary") which were noted by the opposition division and in the decision under appeal.

VI. Independent claim 1 of the first auxiliary request was concerned with subject-matter relating to Herpes simplex virus type 2 glycoprotein C (gC-2), whereas independent claim 2 read as follows:

"2. A process which comprises producing in a recombinant, stable, mammalian, continuous cell line a gC or gD glycoprotein polypeptide of Herpes simplex virus type 1 or type 2, wherein there is initially produced a polypeptide devoid of membrane-binding domain, whereby the derivative polypeptide is free of membrane and has exposed antigenic determinants, and is capable of specifically binding complementary antibodies of Herpes simplex virus type 1 and/or type 2."

VII. The second auxiliary request comprised the twenty-nine claims of the first auxiliary request, wherein, however, the process claims of the first auxiliary request had been changed to claims directed to the use of the corresponding polypeptides in a diagnostic assay.

VIII. Independent claim 1 of the third auxiliary request read as follows:

"1. A process which comprises producing in a recombinant, stable, mammalian, continuous cell line a gC or gD glycoprotein polypeptide of Herpes simplex virus type 1 or type 2 whereby the polypeptide is a type-specific fraction of the glycoprotein having exposed antigenic determinants capable of specifically binding complementary antibodies of Herpes simplex virus type 1 or type 2.".

Claim 4 was directed to the process of claim 1, wherein there was initially produced a truncated derivative of a membrane-bound gC or gD glycoprotein polypeptide of Herpes simplex virus type 1 or type 2 devoid of membrane-binding domain, whereby the derivative polypeptide was free of said membrane and had said exposed antigenic determinants.

IX. Claims 1 and 2 of the fourth auxiliary request were exclusively concerned with processes relating to Herpes simplex virus type 2 glycoprotein C (gC-2), whereas independent claim 3 read as follows:

"3. A process which comprises producing in a recombinant, stable, mammalian, continuous cell line a type-specific fragment of gC glycoprotein polypeptide of Herpes simplex virus type 1 or type 2 whereby the glycoprotein has exposed antigenic determinants capable of specifically binding complementary antibodies of Herpes simplex virus type 1 or type 2, respectively, and thereby capable of distinguishing between HSV-1 and HSV-2.".

Claim 7 related to a process according to any one of claims 1 to 4, wherein there was initially produced a truncated derivative of a membrane-bound gC glycoprotein polypeptide devoid of membrane-binding domain, whereby the derivative polypeptide was free of said membrane and had said exposed antigenic determinants.

X. All the remaining claims of the first, second, third, fourth and fifth auxiliary requests essentially corresponded to the ones of the main request but referring to the specific glycoprotein polypeptides as defined above.

XI. The Board issued a communication pursuant to Article 11(2) of the Rules of Procedure of the Boards of Appeal outlining the points to be discussed. In particular, it was indicated that documents published after the first and second priority dates but earlier than the third priority date were relevant for assessing the inventive step of subject-matter not entitled to said priorities. With reference to decision G 9/92 (OJ EPO 1994, 875), the parties were reminded that neither the Board of Appeal nor the opponent could challenge the maintenance of the patent as amended in accordance with the interlocutory decision, as no appeal had been filed by the opposing party.

XII. Oral proceedings were held on 27 March 2003. As announced in its letter dated 25 February 2003, the appellant did not attend the oral proceedings. For reasons of procedural economy, and to give the absent appellant the benefit of any available doubt, the Board postponed discussions of issues arising under Articles 123 and 84 EPC until after discussion of issues of novelty and inventive step on subject-matter common to all requests.

XIII. The documents referred to in the present decision are the following:

D1a: EP-A-0 100 521 published on 15 February 1984 (filing date 27 July 1983);

D2: R.J. Watson et al., Science, 22 October 1982, Vol. 218, pages 381 to 384;

D2a: J.H. Weis et al., Nature, 3 March 1983, Vol. 302, pages 72 to 74;

D3: R.J. Frink et al., J. Virol., February 1983, Vol. 45(2), pages 634 to 647;

D6: J.K. Rose and J.E. Bergmann, Cell, October 1982, Vol. 30, pages 753 to 762;

D7: E. Amann et al., Gene, 1984, Vol. 32, pages 203 to 215;

D12: B. Norrild and B. Pedersen, J. Virol., August 1982, Vol. 43(2), pages 395 to 402;

D14: P.W. Berman et al., Science, 4 November 1983, Vol. 222, pages 524 to 527;

D16: US 4 399 216;

D17: M. Zoler and T. Wilson, Bio/Technology, April 1983, pages 146 to 147;

D31: N. Sarver et al., Mol. Cell. Biol., June 1981, Vol. 1(6), pages 486 to 496;

D32: J.T. Elder et al., Ann. Rev. Genet., 1981, Vol. 15, pages 295 to 340;

A1: A.D. Levinson, Methods in Enzymology, 1990, Vol. 185, pages 485 to 579.

XIV. The appellant's submissions in writing insofar as they are relevant to the present decision may be summarized as follows:

Article 54 EPC (Novelty)

Document D1a was concerned with the preparation of herpes simplex virus (HSV) antigens suitable for diagnostic and therapeutic uses and it was exemplified by cloning a genome section of HSV-1 corresponding to the gC glycoprotein (gC-1). The document referred, in general, to the expression of gC-1 in prokaryotic and eukaryotic host cells (cf end of Sections 3 and 5 and claim 7) but there was, however, no disclosure of any actual expression. In view of the important technical problems and difficulties found in expressing the gC-1 in prokaryotic host cells (cf document D7 and Exhibit 1, enclosed to the patentee's letter of 23 May 1994), the skilled person would not have contemplated the expression in eukaryotic host cells.

Documents D31 and D32, cited in document D1a, disclosed expression vectors for mammalian cells but there was no reference to the expression of any glycoprotein, let alone of any HSV glycoprotein. Document D31 disclosed minimal levels of expression using the BPV vector and thus, it would not have encouraged to use this BPV system for practical purposes such as the production of vaccines or diagnostic products. Moreover, the disclosed BPV vector did not provide a regulatory element to drive the transcription of the inserted gene but it used the one from the inserted gene. However, the isolation of a suitable HSV regulatory transcription element was not straightforward. In any case, even as late as in 1990 (cf document A1), neither the BPV nor the SV40 eukaryotic expression systems were considered to produce stable transformed cell lines in a reliable manner.

Thus, the claimed subject-matter was novel over document D1a.

Article 56 EPC (Inventive step)

Document D3 was concerned with gC-1, however, it did not disclose the expression of gC-1 in host cells but only the in vitro translation of gC-1 mRNA in a rabbit reticulocyte system. It was neither obvious nor reasonable to expect the skilled person to go from an in vitro expression system to the recombinant, stable, mammalian, continuous cell lines of document D16. In fact, document D16 was a general disclosure of co-transformation for inserting DNA into eukaryotic mammalian cell lines but it was completely silent on HSV glycoproteins, let alone on gC-1 or on its possible use for diagnostic. In view of the additional technical problems and uncertainties (purification, toxicity, etc...), there was no reason to use the expression system of document D16 instead of other well-known and more established expression systems, such as E. coli, yeast or fungi. In particular, the toxic effects found in expressing HSV glycoproteins in E. coli were expected to be greater and more adverse for the amplification systems of document D16. Moreover, there was no indication in document D3 that the resulting gC-1 glycoprotein had the appropriate quality and immunoreactivity for its use in diagnostic assays. The strong immunogenicity and better diagnostic properties resulting from the expression in the system of the contested patent were completely unexpected.

The identification of the glycoprotein transmembrane regions in documents D2 (gD-1) and D3 (gC-1) was only putative. Document D2a disclosed the expression of a fusion (truncated) gD-1 protein in a prokaryotic host cell but not the expression of a truncated gD-1 protein alone. The expression of a truncated membrane-devoid derivative was expected to interfere in the production of the normal HSV glycoprotein structure, such as the one of the gD trimer, altering thus the antigenic properties. In view of the problems and difficulties outlined in document D17 (presence of non-native structure, problems in obtaining polyvalent complexes for immunogenicity, presence of multiple hydrophobic regions, etc...), the production of a membrane-devoid derivative was not straightforward and obvious. Moreover, there was no reason to use the expression system of document D16, concerned with the expression of unaltered genes, instead of other more established and well-known expression systems. Document D6, disclosing the expression of a membrane-devoid derivative of a totally different virus (vesicular stomatitis virus) in mammalian cells, showed the problems encountered using such a system (no secretion or abnormally slow secretion of recombinant product, significant lower expression, etc...) and one of the exemplified host cells (COS cells with SV40-derived vector) was not a stable and continuous cell line in the sense of the contested patent.

Document D12 referred to gC-1 and gC-2 as being type-specific glycoproteins. However, the patent-in-suit provided for the first time a rational basis for designing gC fragments with type-common and/or type-specific epitopes, allowing thus the diagnostic of sub-clinical HSV-2 infections which had been underdiagnosed with the available methods.

Thus, the claimed subject-matter was inventive over the cited prior art.

XV. The respondent's submissions in writing and during the oral proceedings insofar as they are relevant to the present decision may be summarized as follows:

Article 54 EPC (Novelty)

Document D1a, which was prior art under Article 54(2) EPC, referred to the expression of gC-1 and its use for vaccine and diagnostic purposes, the nucleotide and amino acid sequences of the gC-1 glycoprotein being known in the art (cf document D3). Document D1a exemplified the expression of gC-1 in a prokaryotic host cell. However, there was an explicit teaching for other host cells, such as general eukaryotic host cells and animal cell cultures, and reference was made to several expression vectors for mammalian cells known in the art, such as inter alia those of documents D31 and D32. Document D31 disclosed the BPV expression system with recombinant, stable, mammalian, continuous cell lines expressing significant levels of a recombinant product. The provision of suitable regulatory elements for the expression of genes inserted in this BPV system was within the normal abilities and the common general knowledge of the skilled person. Document D32, concerned with the SV40 eukaryotic expression system, represented a similar disclosure. As the technical difficulties and problems found in the expression of HSV glycoproteins in prokaryotic hosts were known to the skilled person (cf document D7 and Exhibit 1), the production of these glycoproteins using their "natural" eukaryotic (mammalian) host cells, as indicated in document D1a, would have been contemplated by the skilled person.

Thus, the claimed subject-matter was not novel over the disclosure of document D1a.

Article 56 EPC (Inventive step)

Document D3 disclosed the nucleotide and amino acid sequences of gC-1 which was identified as a convenient type-specific marker. Mammalian expression systems were well-known in the prior art (cf document A1) and they were the expression system of choice for glycoproteins such as gC-1. In particular, document D16 referred to the advantages of mammalian expression systems with explicit mention of viral antigens and glycoproteins, and to the specific system used in the contested patent, namely the co-selection with the marker dihydrofolate dehydrogenase (DHFR).

Documents D2, D2a (gD-1) and D3 (gC-1) showed that these HSV glycoproteins were membrane-bound. In view of the known advantageous expression of truncated viral glycoproteins (production of truncated viral glycoproteins with viral antigenic determinants, cf documents D17 and D6), it would have been obvious to express these truncated glycoproteins using their normal host cells, ie using a well-known and available mammalian expression system (cf documents D16 and A1). Moreover, their use in HSV diagnostic assays or test kits was also obvious and it did not require any special inventive contribution. In this respect, the claims of the contested patent did not require the HSV glycoproteins to be immunogenically stronger or to have better diagnostic properties. The presence of type-specific gC-1 epitopes was already known from D12 and it was obvious and easy for the skilled person to find type-specific fragments of a known glycoprotein.

Moreover, it was submitted that the subject-matter concerned with the gC glycoprotein and in particular with the gC-1, was obvious in the light of document D1a combined with document D14. Furthermore, documents D17 and D12 with the common general knowledge rendered the subject matter relating to truncated gC-1 and/or to type-specific gC-1 obvious.

Thus, the claimed subject-matter was not inventive in the light of the cited prior art.

XVI. The appellant (patentee) requested that the decision under appeal be set aside and that the patent be maintained as granted (main request) or on the basis of first auxiliary request filed on 25 February 2003, or second, fourth auxiliary requests filed on 20 March 1998 or the third auxiliary request filed on 21 October 2002.

The respondent (opponent) requested that the appeal be dismissed.