Abstract 2727: Bispecific, hexavalent antibodies (HexAbs) targeting IGF-1R and either Trop-2 or CEACAM6 inhibit anchorage-independent growth and invasion of antigen-expressing breast and pancreatic cancer cell lines in vitro
Authors: Wang Yang , Trisal Pretti , Cardillo Thomas M. , Rossi Ed , Goldenberg David M. , Chang Chien-Hsing 
Published: August 2012

Background: Combination therapy using two distinct monoclonal antibodies to achieve improved efficacy without increased toxicity, may be accomplished with a single bispecific antibody. We noted the elevated expression of the type I insulin-like growth factor receptor (IGF-1R), the trophoblast cell-surface marker (Trop-2), and the carcinoembryonic antigen related cell adhesion molecule 6 (CEACAM6) in diverse epithelial cancer cell lines and set forth to explore the potential of 1R-(E1)-(E1) and 1R-(15)-(15), two novel bispecific HexAbs targeting IGF-1R/Trop-2 and IGF-1R/CEACAM6, respectively, for treating breast and pancreatic cancers. Methods: 1R-(E1)-(E1), comprising the full IgG of hR1 (humanized anti-IGF-1R) and four Fabs of hRS7 (humanized anti-Trop-2) was generated with the Dock-and-Lock (DNL) method by reacting the IgG-AD2 module of hR1 with the Fab-DDD2 module of hRS7 under mild redox conditions followed by purification on Protein A. 1R-(15)-(15) was generated in a similar fashion using the Fab-DDD2 module of hMN-15 (humanized anti-CEACAM6). The in vitro effects of 1R-(E1)-(E1) and 1R-(15)-(15) on three triple-negative breast cancer lines of varying invasive activities (MCF-7, low; MDA-MB-468, moderate; MDA-MB-231, high) and two pancreatic cancer lines (Capan-1 and BxPC-3) were determined, which included cell binding by flow cytometry, anchorage-independent growth by soft agar assay, and invasiveness by BD matrigel chambers. Statistical differences (P values) between two populations were determined by Student's t-test. Results: All five cell lines were found to express IGF-1R, Trop-2, and CEACAM6 of sufficient levels, which are higher in MCF-7, MDA-MB-468, and BxPC-3, respectively. When tested at 100 μg/mL, 1R-(E1)-(E1) reduced the invasion of MDA-MB-468 to less than 10% of the untreated control, whereas under the same conditions, MDA-MB-231 appeared to be resistant and the parental antibodies showed no effect. 1R-(15)-(15) at 100 μg/mL potently reduced the invasion of Capan-1, but had little effect on MDA-MB-468. The ability of 1R-(E1)-(E1) to inhibit anchorage-independent growth was demonstrated at 200 nM in MDA-MB-231 with statistically significant difference (P&lt; 0.041) when compared with samples treated with parental antibodies at the same concentrations. Cells treated with 1R-(E1)-(E1) produced few and much smaller colonies, the largest size of which was less than 1/10 of the untreated cells. The parental hR1 alone, but not hRS7, had some effect on inhibiting the growth of MDA-MB-231 in soft agar, presumably resulting from the downregulation of IGF-1R. Conclusions: These promising results, as exemplified by 1R-(E1)-(E1) and 1R-(15)-(15), attest to the potential of bispecific HexAbs for targeted therapy of solid cancers.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2727. doi:1538-7445.AM2012-2727