Abstract P4-09-16: A monoclonal antibody with exceptional specificity across major breast cancer subtypes
Authors: L Das Roy, R Zhou, L Dillon, LJ Moore, R Puri, JR Marks, HK Lyerly, P Mukherjee
Journal: Cancer Research (2016)

Abstract Background: Breast cancer (BC) remains the second leading cause of cancer-related deaths for women in the United States and is recognized to be a heterogeneous disease. Advances in technologies such as whole genome sequencing are leading the way to precision medicine and the leading researchers are envisioning personalized therapies in the not too distant future. However, given the diversity of cancer cell populations, that remains a challenging task at best. The tumor form of MUC1 (designated tMUC1), a transmembrane glycoprotein, is aberrantly glycosylated and overexpressed in ∼95% of BC. We have developed an antibody (TAB004) that specifically recognizes tMUC1 across all major subtypes of BC and importantly does not recognize normal breast epithelia. This is a significant development in light of the challenges faced in treating triple negative BC. Methods: A panel of thirty BC cell lines was obtained from ATCC. The following techniques were used to assess the specificity of TAB 004 to the major subtypes based on ER, PR and Her2 expression: 1) Flow cytometry to quantify membrane bound expression of tMUC1 using Cy7-conjugated TAB004; 2) Western blotting to detect molecular weight patterns of tMUC1 in whole cell lysate; 3) A TAB004 based GMP-grade ELISA kit to measure shed tMUC1 in the supernatant and 4) In vivo imaging of tumors in mice using TAB 004 conjugated to Indocyanine Green (ICG). Specificity and sensitivity was further confirmed using primary human serum and tissue samples from all major BC subtypes obtained from bio-repositories at Duke University Cancer Center, Fox Chase Cancer Center and Carolinas Health Care System. Shed tMUC1 in serum samples were tested using the TAB 004 ELISA kit and tissue sections were analyzed using immunohistochemical (IHC) staining with TAB 004 conjugated to HRP. Results: 1) Flow cytometry data shows that TAB 004 recognized tMUC1 on all major BC subtypes: 25 out of 30 BC cell lines tested had higher expression than a normal epithelial breast cell line; 2) Western blotting also detected tMUC1 on all BC subtypes with distinct molecular weight patterns; 3) ELISA showed high levels of shed tMUC1 by most BC cells and correlated with bound/cytoplasmic levels. 4) In vivo imaging shows clear localization of TAB004-ICG to the tumors expressing tMUC1. Primary human breast cancer patient data shows that shed tMUC1 was detected in the serum obtained from all major BC subtypes and showed statistically significant differentiation from normal/benign. IHC results show strong tMUC1 expression in malignant tissue with excellent differentiation from adjacent normal tissue. Conclusion: TAB004 antibody's extraordinary specificity across major BC subtypes has been confirmed with flow cytometry, western blotting, ELISA and Immunohistochemistry. A number of clinical applications are under development: (a) An ELISA test as a supplement to mammography for the early detection of BC in women with dense breasts; (b) serum monitoring during treatment and to detect disease recurrence; and, (c) targeted antibody-drug/antibody-imaging agent based therapies and imaging modalities particularly for triple negative BC. Citation Format: Das Roy L, Zhou R, Dillon L, Moore LJ, Puri R, Marks JR, Lyerly HK, Mukherjee P. A monoclonal antibody with exceptional specificity across major breast cancer subtypes. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P4-09-16.