title: Solitomab, an EpCAM/CD3 bispecific antibody construct (BiTE®), is highly active against primary uterine and ovarian carcinosarcoma cell lines in vitro.
abstract: BACKGROUND
  Uterine and ovarian carcinosarcomas (CS) are rare but highly aggressive gynecologic tumors which carry an extremely poor prognosis. We evaluated the expression levels of EpCAM and the in vitro activity of solitomab, a bispecific single-chain antibody construct which targets epithelial-cell-adhesion-molecule (EpCAM) on tumor cells and also contains a CD3 binding region, against primary uterine and ovarian CS cell lines.
  
  
  METHODS
  EpCAM expression was evaluated by flow cytometry in a total of 5 primary CS cell lines. Sensitivity to solitomab-dependent-cellular-cytotoxicity (ADCC) was tested against the panel of primary CS cell lines expressing different levels of EpCAM in standard 4 h (51)Cr release-assays. The proliferative activity, activation, cytokine secretion (i.e., Type I vs Type II) and cytotoxicity of solitomab in autologous tumor-associated-T cells (TAL) in the pleural fluid of a CS patient were also evaluated by CFSE and flow-cytometry assays.
  
  
  RESULTS
  Surface expression of EpCAM was found in 80.0 % (4 out of 5) of the CS cell lines tested by flow cytometry. EpCAM positive cell lines were found resistant to NK or T-cell-mediated killing after exposure to peripheral blood lymphocytes (PBL) in 4-h chromium-release assays (mean killing ± SEM = 1.1 ± 1.6 %, range 0-5.3 % after incubation of EpCAM positive cell lines with control BiTE®). In contrast, after incubation with solitomab, EpCAM positive CS cells became highly sensitive to T-cell-cytotoxicity (mean killing ± SEM of 19.7 ± 6.3 %; range 10.0-32.0 %; P < 0.0001). Ex vivo incubation of autologous TAL with EpCAM expressing malignant cells in pleural effusion with solitomab, resulted in a significant increase in T-cell proliferation in both CD4+ and CD8+ T cells, increase in T-cell activation markers (i.e., CD25 and HLA-DR), and a reduction in number of viable CS cells in the exudate (P < 0.001).
  
  
  CONCLUSIONS
  Solitomab may represent an effective treatment for patients with recurrent/metastatic and/or chemo-resistant CS overexpressing EpCAM.
authors.affiliation: Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT, USA. francesca.ferrari@yale.edu.
authors.forename: Francesca
authors.initials: F
authors.lastname: Ferrari
authors.affiliation: Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT, USA. stefania.bellone@yale.edu.
authors.forename: Stefania
authors.initials: S
authors.lastname: Bellone
authors.affiliation: Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT, USA. jonathan.black@yale.edu.
authors.forename: Jonathan
authors.initials: J
authors.lastname: Black
authors.affiliation: Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT, USA. carlton.schwab@yale.edu.
authors.forename: Carlton L
authors.initials: CL
authors.lastname: Schwab
authors.affiliation: Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT, USA. salvatore.lopez@yale.edu.
authors.forename: Salvatore
authors.initials: S
authors.lastname: Lopez
authors.affiliation: Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT, USA. emiliano.cocco@yale.edu.
authors.forename: Emiliano
authors.initials: E
authors.lastname: Cocco
authors.affiliation: Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT, USA. elena.bonazzoli@yale.edu.
authors.forename: Elena
authors.initials: E
authors.lastname: Bonazzoli
authors.affiliation: Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT, USA. ferica.predolini@yale.edu.
authors.forename: Federica
authors.initials: F
authors.lastname: Predolini
authors.affiliation: Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT, USA. gulden.menderes@yale.edu.
authors.forename: Gulden
authors.initials: G
authors.lastname: Menderes
authors.affiliation: Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT, USA. babak.litkouhi@yale.edu.
authors.forename: Babak
authors.initials: B
authors.lastname: Litkouhi
authors.affiliation: Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT, USA. elena.ratner@yale.edu.
authors.forename: Elena
authors.initials: E
authors.lastname: Ratner
authors.affiliation: Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT, USA. dan-arin.silasi@yale.edu.
authors.forename: Dan-Arin
authors.initials: DA
authors.lastname: Silasi
authors.affiliation: Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT, USA. masoud.azodi@yale.edu.
authors.forename: Masoud
authors.initials: M
authors.lastname: Azodi
authors.affiliation: Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT, USA. peter.shwartz@yale.edu.
authors.forename: Peter E
authors.initials: PE
authors.lastname: Schwartz
authors.affiliation: Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT, USA. alessandro.santin@yale.edu.|, 333 Cedar Street, LSOG 305, PO Box 208063, New Haven, CT, 06520-8063, USA. alessandro.santin@yale.edu.
authors.forename: Alessandro D
authors.initials: AD
authors.lastname: Santin
mesh_terms: D000368:Aged; D018033:Antibodies, Bispecific / Q000494:pharmacology*; D000951:Antigens, Neoplasm / Q000276:immunology*; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017252:CD3 Complex / Q000276:immunology*; D002296:Carcinosarcoma / Q000188:drug therapy*; D015815:Cell Adhesion Molecules / Q000037:antagonists & inhibitors / Q000276:immunology*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D018920:Coculture Techniques; D016207:Cytokines / Q000276:immunology; D003602:Cytotoxicity, Immunologic; D019008:Drug Resistance, Neoplasm; D000071858:Epithelial Cell Adhesion Molecule; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D007694:Killer Cells, Natural / Q000276:immunology; D008213:Lymphocyte Activation / Q000276:immunology; D008875:Middle Aged; D010051:Ovarian Neoplasms / Q000188:drug therapy*; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology; D014594:Uterine Neoplasms / Q000188:drug therapy*
chemical_list: D018033:Antibodies, Bispecific; D000951:Antigens, Neoplasm; D000970:Antineoplastic Agents; D017252:CD3 Complex; D015815:Cell Adhesion Molecules; D016207:Cytokines; C499379:EPCAM protein, human; D000071858:Epithelial Cell Adhesion Molecule; C000607229:Solitomab