Solitomab, an EpCAM/CD3 bispecific antibody construct (BiTE), is highly active against primary uterine serous papillary carcinoma cell lines in vitro.
BACKGROUND
Uterine serous carcinoma is an aggressive form of endometrial cancer that carries an extremely poor prognosis. Solitomab is a novel bispecific single-chain antibody construct that targets epithelial cell adhesion molecule on tumor cells and also contains a CD3 binding region. We evaluated the expression levels of epithelial cell adhesion molecule and the in vitro activity of solitomab against primary uterine serous carcinoma cell lines in vitro and ex-vivo in the ascites of patients with uterine serous carcinoma.


OBJECTIVE
The purpose of this study was to determine the frequency of expression of epithelial cell adhesion molecule on uterine serous carcinoma cell lines and the ability of solitomab to modulate immune responses (T-cell proliferation, activation, cytokine production, and tumor killing) to tumor cells when it is combined with lymphocytes and epithelial cell adhesion molecule-positive cell lines or epithelial cell adhesion molecule-positive ascitic fluid in vitro.


METHODS
Epithelial cell adhesion molecule expression was evaluated by flow cytometry in a total of 14 primary uterine serous carcinoma cell lines. Sensitivity to solitomab-dependent cellular-cytotoxicity was tested against a panel of primary uterine serous carcinoma cell lines that express different levels of epithelial cell adhesion molecule in standard 4-hour chromium release assays. The proliferative activity, activation, cytokine secretion (ie, type I vs type II), and cytotoxicity of solitomab in autologous tumor-associated T cells in the ascitic fluid of patients with uterine serous carcinoma was also evaluated by carboxyfluorescein succinimidyl ester and flow-cytometry assays. Differences in epithelial cell adhesion molecule expression, solitomab-dependent cellular-cytotoxicity levels were analyzed with the use of an unpaired t test. T-cell activation marker increase and cytokine release were analyzed by a paired t test.


RESULTS
Surface expression of epithelial cell adhesion molecule was found in 85.7% (12 of 14) of the uterine serous carcinoma cell lines that were tested by flow cytometry. Epithelial cell adhesion molecule-positive cell lines were found resistant to natural killer cells or T-cell-mediated killing after exposure to peripheral blood lymphocytes in 4-hour chromium-release assays (mean killing ± standard of the mean, 2.7% ± 3.1% after incubation of epithelial cell adhesion molecule-positive cell lines with control bispecific antibody construct). In contrast, after incubation with solitomab, epithelial cell adhesion molecule-positive uterine serous carcinoma cells became highly sensitive to T-cell cytotoxicity (mean killing, 25.7% ± 4.5%; P < .0001) by peripheral blood lymphocytes. Ex vivo incubation of autologous tumor-associated lymphocytes with epithelial cell adhesion molecule that expressed malignant cells in ascites with solitomab resulted in a significant increase in T-cell proliferation in both CD4+ and CD8+ T cells, increase in T-cell activation markers (ie, CD25 and HLA-DR), and a reduction in number of viable uterine serous carcinoma cells in ascites (P < .001).


CONCLUSIONS
Solitomab induces robust immunologic responses in vitro that result in increased T-cell activation, proliferation, production of cytokines, and direct killing of tumor cells. These findings suggest that solitomab may represent a novel, potentially effective agent for the treatment of recurrent/metastatic and/or chemo-resistant uterine serous carcinoma-overexpressing epithelial cell adhesion molecule.
Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT.
Stefania
S
Bellone
Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT.
Jonathan
J
Black
Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT.
Diana P
DP
English
Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT.
Carlton L
CL
Schwab
Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT.
Salvatore
S
Lopez
Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT.
Emiliano
E
Cocco
Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT.
Elena
E
Bonazzoli
Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT.
Federica
F
Predolini
Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT.
Francesca
F
Ferrari
Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT.
Elena
E
Ratner
Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT.
Dan-Arin
DA
Silasi
Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT.
Masoud
M
Azodi
Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT.
Peter E
PE
Schwartz
Department of Obstetrics, Gynecology & Reproductive Sciences, Yale University School of Medicine, New Haven, CT. Electronic address: alessandro.santin@yale.edu.
Alessandro D
AD
Santin
D018033:Antibodies, Bispecific / Q000494:pharmacology*; D000951:Antigens, Neoplasm / Q000032:analysis / Q000187:drug effects* / Q000276:immunology*; D000970:Antineoplastic Agents / Q000494:pharmacology*; D001202:Ascitic Fluid / Q000473:pathology; D017252:CD3 Complex / Q000276:immunology*; D015496:CD4-Positive T-Lymphocytes / Q000187:drug effects / Q000276:immunology; D002291:Carcinoma, Papillary / Q000737:chemistry / Q000188:drug therapy* / Q000276:immunology; D015815:Cell Adhesion Molecules / Q000032:analysis / Q000187:drug effects* / Q000276:immunology*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D018920:Coculture Techniques; D016207:Cytokines / Q000187:drug effects / Q000378:metabolism; D003602:Cytotoxicity, Immunologic / Q000187:drug effects; D000071858:Epithelial Cell Adhesion Molecule; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D008213:Lymphocyte Activation / Q000187:drug effects; D018297:Neoplasms, Cystic, Mucinous, and Serous / Q000737:chemistry / Q000188:drug therapy* / Q000276:immunology; D013602:T-Lymphocytes, Cytotoxic / Q000187:drug effects / Q000276:immunology; D014594:Uterine Neoplasms / Q000737:chemistry / Q000188:drug therapy* / Q000276:immunology
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
CD3; EpCAM; T lymphocyte; bispecific antibody; uterine serous cancer