Development and in vitro validation of anti-mesothelin biobodies that prevent CA125/Mesothelin-dependent cell attachment Lindsay Bergan 1 , Jennifer A. Gross 1 , Barry Nevin, Nicole Urban, Nathalie Scholler * Molecular Diagnositics Program, Public Health Sciences, Fred Hutchinson Cancer Research Center, Seattle, WA, USA Received 2 January 2007; received in revised form 26 April 2007; accepted 30 April 2007 Abstract Preventing peritoneal implantation of carcinoma cells could prolong ovarian cancer patient remission and survival. Per- itoneal cells constitutively express mesothelin, a ligand for CA125 that is expressed by tumor cells. Thus blocking CA125/ mesothelin-dependent cell attachment may prevent or delay peritoneal metastatic recurrence. We developed a high- throughput screening system for reagents able to block CA125/mesothelin-dependent cell attachment with a sensitive quantitative readout. Using a novel yeast-expression system to produce secreted, in vivo biotinylated proteins and biobod- ies (Bbs), we generated anti-mesothelin Bbs. Anti-mesothelin Bbs derived from high affinity yeast-display scFv detected both membrane-bound and soluble mesothelins and inhibited CA125/mesothelin-dependent cell attachment. Published by Elsevier Ireland Ltd. Keywords: Mesothelin; CA125; Biobody; Tumor cell attachment; Ovarian cancer; Peritoneal metastasis 1. Introduction Ovarian cancer represents one-fourth of the malignancies of the female genital tract and is the most common cause of death among women with gynecologic cancer. The American Cancer Society estimates that 22,430 women will be diagnosed with and 15,280 women will die of ovarian cancer in 2007 [1]. The 5-year relative survival rates correlate strongly with stage at diagnosis: 93% for early stage when the cancer is confined to the ovary, 69% for regional stage and 30% for advanced stage. However, only 19% of all ovarian cancers are found at an early stage [2]. Cytoreduction and che- motherapy induce remission in 80% of the cases, but recurrence can be expected in nearly all women with advanced disease at the time of diagnosis. A means to prevent or delay metastatic recurrence is needed. The interaction between mesothelin and CA125 proteins could play an important role in the perito- neal implantation of ovarian tumor cells by promot- ing cell attachment between CA125-expressing 0304-3835/$ - see front matter Published by Elsevier Ireland Ltd. doi:10.1016/j.canlet.2007.04.012 * Corresponding author. Tel.: +1 206 667 3170; fax: +1 206 667 2537. E-mail address: nscholle@fhcrc.org (N. Scholler). 1 These authors contributed equally to the work. Cancer Letters 255 (2007) 263–274 www.elsevier.com/locate/canlet