FGFR4 as a potential therapeutic target for monoclonal antibody based intervention in rhabdomyosarcoma

Sivasubramanian Baskar, Nityashree Shivaprasad, Zhongyu Zhu, Dimiter Dimitrov, Mhairi Sigrist, Poul Sorensen, Marielle Yohe, John Shern, John Maris, Crystal Mackall, and Javed Khan

Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA See Original Here

Abstract
Background: Rhabdomyosarcoma (RMS) is most common soft tissue sarcoma in childhood and adolescence. There are two major subtypes of RMS - alveolar RMS (ARMS) and embryonal RMS (ERMS). With current treatment modalities the 5 year survival rate of patients with metastatic disease is only about 30%, thus necessitating development of novel targeted therapeutic strategies. Fibroblast Growth Factor Receptor 4 (FGFR4) is highly differentially expressed gene and activating mutations in FGFR4 is associated with increase in RMS metastasis. We hypothesize that FGFR4 can be exploited as a potential therapeutic target in RMS. Monoclonal antibodies (mAbs) against specific cancer cell surface antigens, have gained importance as therapeutic agents in other cancer types. Therefore, mAbs against FGFR4 may be developed either alone or conjugated with a chemotherapeutic agent for treatment of RMS.

Methods: We have developed 15 mAbs that are of mouse, rabbit and human origin. The immunogen for rabbit antibodies is human FGFR4 extracellular domain (ECD), while human FGFR4-Fc chimeric protein or hFGFR4 transfected cells were used for mouse antibodies and were produced using hybridoma technology. Using recombinant DNA technology, human immunoglobulin library was selected using FGFR4 ECD to derive human anti-FGFR4 mAbs. All mAbs were purified using affinity chromatography and reactivity was confirmed by ELISA. These mAbs were tested for their ability to bind cell surface FGFR4 and internalization of mAb bound FGFR4 (upon incubation at 37°C) by flow cytometry using RMS cell lines. To further characterize these antibodies we have performed immunohistochemistry (IHC) on tissue microarrays (TMA) of normal tissue, xenografts of RMS cell lines, and primary tumor from RMS patients. Finally, to select a potential mAb candidate as therapeutic agent, we tested appropriate secondary antibody-drug conjugate (ADC) using cell based cytotoxic assay.

Results: We observed that tested mAbs bind to both ARMS and ERMS cell lines. The level of FGFR4 expression is variable in both subtypes. Internalization assays demonstrated mAbs bound FGFR4 was internalized in RMS cell lines. Analysis by IHC on TMAs suggests that FGFR4 protein is expressed at a considerably higher level in RMS tumor tissue and xenografts compared to normal tissue. Based on these findings, we are currently screening the mAbs for their ability to elicit a cytotoxic response either alone or in conjunction with secondary ADC.

Conclusion: These results suggest anti-FGFR4 mAbs can be used as therapeutic intervention for RMS. High expression of FGFR4 in other cancers such as prostate, melanoma, lung, breast, colorectal, and gastric cancers suggests the potential use of anti-FGFR4 mAbs and their derivatives in these cancers.

Citation Format: Sivasubramanian Baskar, Nityashree Shivaprasad, Zhongyu Zhu, Dimiter Dimitrov, Mhairi Sigrist, Poul Sorensen, Marielle Yohe, John Shern, John Maris, Crystal Mackall, Javed Khan. FGFR4 as a potential therapeutic target for monoclonal antibody based intervention in rhabdomyosarcoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2488. doi:10.1158/1538-7445.AM2015-2488

©2015 American Association for Cancer Research.