ORIGINAL PAPERS PHF3-speci®c antibody responses in over 60% of patients with glioblastoma multiforme A-K Struss 1 , BFM Romeike 2 , A Munnia 1 , W Nastainczyk 3 , W-I Steudel 4 , J KoÈnig 5 , H Ohgaki 6 , W Feiden 2 , U Fischer 1,7 and E Meese* ,1,7 1 Institut fu Èr Humangenetik, Universita Èt(-skliniken) des Saarlandes, 66421 Homburg/Saar, Germany; 2 Abteilung fu Èr Neuropathologie, Universita Èt(-skliniken) des Saarlandes, 66421 Homburg/Saar, Germany; 3 Institut fu Èr Medizinische Biochemie, Universita Èt(-skliniken) des Saarlandes, 66421 Homburg/Saar, Germany; 4 Neurochirurgische Klinik, Universita Èt(-skliniken) des Saarlandes, 66421 Homburg/Saar, Germany; 5 Institut fu Èr Bioinformatik; Universita Èt(-skliniken) des Saarlandes, 66421 Homburg/ Saar, Germany; 6 International Agency for Research on Cancer, 150 Cours Albert Thomas, 69372 Lyon Cedex 8, France Glioblastoma multiforme (GBM), a malignant astrocytic tumour, represents the most frequent tumour of the human brain. Nevertheless, its molecular pathology is not well understood. We utilized the immune system, which contributes to cancer protection, to help identify new GBM-related genes. By screening a human GBM cDNA library with autologous patient serum (SEREX-approach), we isolated a gene termed PHF3 (PHD finger protein 3). The gene product of PHF3 is immunogenic in GBM as tested in an allogenic patient serum screening demonstrat- ing antibodies in 24 of 39 (61.53%) sera, whereas none of the 14 healthy persons had antibodies against PHF3. While previous SEREX studies revealed allogenic anti- body responses up to 40%, our results for PHF3 represent the highest reported rate for a speci®c antibody response. We show that GBM patients with an antibody response against PHF3 show signi®cant better survival than patients without PHF3-speci®c antibodies. Because the amino acid sequence of PHF3 contains a PHD ®nger (also termed LAP motif), a TFIIS homology, a proline rich region and nuclear localization signals, it supposedly functions as a transcription factor. A polyclonal antibody generated against PHF3 shows nuclear expression in most investigated formalin-®xed, paran embedded tissues. In GBM, PHF3 expression is concentrated in cells surround- ing necroses. Oncogene (2001) 20, 4107 ± 4114. Keywords: humoral immune response; SEREX; glio- blastoma multiforme; polyclonal antibody; PHD ®nger; PHF3 Introduction Glioblastoma multiforme (GBM) represents the most malignant tumour of astrocytic origin, corresponding to WHO grade IV. The median survival time varies between 7 and 11 months (Daumas-Duport et al., 1988; Chinot and Martin, 1996). The heterogeneous genetic pattern of glioma cells entails dysfunction of very dierent proteins or cellular regulation cycles (Biernat et al., 1997; Kleihues and Burger, 2000). As known for several decades, the immune system is able to recognize tumour cells (Foley, 1953; Prehn and Main, 1957). In recent years, the number of known tumour antigens has increased enormously. A variety of methods have been developed for identifying tumour antigens such as SEREX, cloning of T-cells and animal transplantation models (Old, 1981; Sahin et al., 1995; Boon and van der Bruggen, 1996; Suresh, 1996). Brain tumours, located in an immunologically privileged site, were believed for a long time not to be involved in such immune reactions. Today, astrocytes and microglia are considered to function as antigen presenting cells (Shrikant and Benveniste, 1996). However, with astro- cytes as the essential part of the blood brain barrier, the blood brain barrier is most likely aected in astrocytic tumours. Antigens can have contact with immune competent cells and induce immune responses (Weller et al., 1992; Zinkernagel et al., 1997). Applying a combined immunological and molecular approach based on the antibody response against tumour-expressed antigens, we have isolated PHF3, a transcription factor-like gene in GBM whose expressed protein was detected by autologous antibodies. Here we report the relevance of PHF3, which is immuno- genic in over 60% of GBM patients but not in 14 healthy control persons. The signi®cant prolonged survival of GBM patients with antibodies against PHF3 compared to the survival of PHF3-antibody negative patients supports the hypothesis that PHF3 plays an important role in GBM pathology. Results Autologous serum screening and identification of PHF3 antigen PHF3 was identi®ed using a SEREX immunoscreening approach on a cDNA library prepared from a human Oncogene (2001) 20, 4107 ± 4114 ã 2001 Nature Publishing Group All rights reserved 0950 ± 9232/01 $15.00 www.nature.com/onc *Correspondence: E Meese, Institut fuÈr Humangenetik, Bldg. 60, Medical School, University of Saarland, 66421 Homburg/Saar, Germany; E-mail: hgemee@med-rz.uni-sb.de 7 Equal contribution as senior authors Received 23 October 2000; revised 28 March 2001; accepted 4 April 2001