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Identification of Genes Overexpressed in Tumors through Preferential Expression Screening in Trophoblasts¹

Laboratoire d'Immunologie des Tumeurs, CNRS URA 1484, Université René Descartes, Faculté des Sciences Pharmaceutiques et Biologiques, 4, avenue de l'Observatoire, 75006 Paris [D. C., C. D., D. G., J-L. J., J-M. B., D. B., A. K.]; Maternité, Hôpital Bichat, 75018 Paris [J-L. B.]; Service d'Immunologie Moléculaire, Institut Gustave-Roussy, 94805 Villejuif [J-M. B., D. B.]; Maternité, Hôpital Antoine Béclère, 92140 Clamart [H. F., R. F.]; Services d'Anatomie Pathologique B [M. P.], C [G. C.], and A [B. C.], Institut Gustave-Roussy, 94805 Villejuif; Maternité, Hôpital St. Vincent de Paul, 75014 Paris [M. T.]; Service de Chirurgie, Institut Gustave-Roussy, 94805 Villejuif [D. E.]; and Service d'Anatomie Pathologique, Hôpital Kremlin-Bicêtre, 94270 Kremlin-Bicêtre [P. B.], France

Early trophoblastic cells share several features with neoplastic cells. Based on that observation, we attempted to identify genes overexpressed in tumors by analyzing genes preferentially expressed in trophoblasts. A subtracted library enriched in complementary DNA from early cytotrophoblasts was constructed, and the expression level of selected recombinants was analyzed on a large panel of normal and tumor tissues. The library was prepared using a polymerase chain reaction-based complementary DNA subtraction method with 6-week amenorrhea cytotrophoblast endoplasmic reticulum-bound RNA as target, and a mixture of complementary DNA prepared from terminal placenta and activated T-lymphocytes as driver. Two rounds of screening were performed to isolate clones preferentially expressed in early placenta. From a total number of recombinant clones estimated at 32,000 in the subtracted library, 594 inserts were analyzed by Southern blot and 21 sequences were isolated as corresponding to genes highly expressed in early placenta. Eleven encoded known molecules, such as carcinoembryonic antigen, human chorionic gonadotropin, and mitochondrial rRNAs. Ten sequences represented novel genes. Northern blot analysis confirmed that most of these genes were preferentially expressed in early trophoblast in comparison to terminal placenta. Three clones that gave detectable hybridization signals on total RNA were extensively studied and were found to be overexpressed in various tumors. Two of these clones, designated B9 and E4, were later identified as corresponding to genes coding for the putative ribosomal protein S18 and the bifunctional enzyme ADE2H1 involved in purine biosynthesis, respectively. Expression of the third clone, E9, was increased up to 10-fold in breast cancer tissues in comparison with normal counterparts. Present results confirm that many genes expressed in the trophoblast are overexpressed in malignant cells. This approach could provide a general targeted method for the identification of genes overexpressed in various neoplastic cell types.

¹ This study was supported by grants from the Association pour la Recherche sur le Cancer (ARC), Villejuif, France.

² To whom requests for reprints should be addressed, at Laboratoire d'Immunologie des tumeurs, CNRS URA 1484, Université René Descartes, Faculté des Sciences Pharmaceutiques et Biologiques, 4, avenue de l'Observatoire, 75006 Paris, France.

Received 5/20/94. Accepted 8/ 4/94.

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