Antibody therapy to human L1CAM in a transgenic mouse model blocks local tumor growth but induces EMT Kai Doberstein 1 , Patrick N. Harter 2 , Uwe Haberkorn 3 , Niko P. Bretz 1 , Bernd Arnold 4 , Rafael Carretero 4 , Gerhard Moldenhauer 1 , Michel Mittelbronn 2 and Peter Altevogt 1 1 Translational Immunology, D015, Tumor Immunology Programme German Cancer Research Center (DKFZ), Heidelberg, Germany 2 Edinger Institute (Neurological Institute), Goethe University Frankfurt, Frankfurt, Germany 3 Department of Nuclear Medicine, University Hospital Heidelberg, Heidelberg, Germany 4 Molecular Immunology, D050, Tumor Immunology Programme German Cancer Research Center (DKFZ), Heidelberg, Germany L1 cell adhesion molecule (L1CAM) is overexpressed in many human cancers, confers bad prognosis and augments cell motil- ity, invasion and metastasis. Results from xenograft mouse models suggested that L1CAM antibodies might be promising tools for cancer therapy. Here, we generated human L1CAM-transgenic mice to study therapeutic efficacy and putative side effects in a model system. We established three transgenic lines (M2, M3 and F4) expressing the human L1CAM transgene in brain, kidney and colon with decreasing intensity (M2, M3 > F4). The expression pattern was similar to that of L1CAM in humans. No interference of the transgene with the expression of endogenous L1CAM was observed. Immunohistochemical analysis revealed correct expression of the transgene in mouse cortex and collective duct of the kidney. Injection of 125 I- labeled L1CAM antibodies resulted in specific enrichment in the kidney but not in the brain. The injection of the therapeutic anti-human L1CAM mAb L1-9.3/2a into transgenic mice even at high doses did not cause behavioral changes or other side effects. Similar results were obtained using a mouse specific L1CAM mAb in normal mice. Tumor therapy experiments were performed using syngeneic mouse tumor cells (RET melanoma and Panc02 pancreatic adenocarcinoma) transduced with human L1CAM. MAb L1-9.3/2a efficiently and specifically attenuated local tumor growth in both model systems without apparent side effects. The therapeutic effect was dependent on immune effector mechanisms. Analysis of Panc02-huL1CAM tumors after therapy showed elevated levels of EGF and evidence of immune-induced epithelial-mesenchymal transition. The results sug- gest that our transgenic mice are valuable tools to study L1CAM-based antibody therapy. The L1 cell adhesion molecule (L1CAM) is a 200–220 kDa transmembrane glycoprotein of the immunoglobulin (Ig) superfamily composed of six Ig-like domains and five fibro- nectin Type III repeats followed by a transmembrane region and a highly conserved cytoplasmic tail. 1 L1CAM can inter- act with itself (homophilic) and with heterophilic ligands such as integrins, CD24, neurocan, neuropilin-1 (NRP-1) and other members of the neural cell adhesion family. Cumulative evidence indicates that L1CAM (CD171) is ectopically expressed at high levels in a variety of cancers, including pancreatic, colorectal, ovarian and endometrial cancers, mela- noma and neuroblastoma. 2–6 Furthermore, high levels of L1CAM were found associated with increased grade of malig- nancy, 5,6 epithelial-mesenchymal transition (EMT), 6 poor patient prognosis 5,7,8 and worse response to chemotherapy. 4,7 These findings prompted extensive research towards develop- ment and preclinical testing of L1CAM specific mAbs for selective targeting of cancers. In line with the role of this molecule in cell motility, invasion and tumorigenesis, 9–11 L1CAM-specific mAbs were shown to suppress tumor cell outgrowth and metastasis in xenograft models. 12–14 The in vivo action of these mAbs was shown to depend on a combi- nation of immunologic and non-immunologic mechanisms, similar to what has been reported for mAbs targeting HER-2 (Herceptin) and CD20 (Rituxan). 13 Taken together, these findings point at the potential of anti-L1CAM mAbs as anti- cancer drugs. The use of mouse xenograft models is limited as only the therapeutic efficacy but not the potentially side effects of can- cer therapy can be evaluated. One way to overcome these problems is the development of antibodies binding with Key words: L1CAM, immunotherapy, transgenic mice, EMT Abbreviations: ADCC: antibody-dependent cellular cytotoxicity; EMT: epithelial-mesenchymal transition; huL1CAM: human L1 cell adhesion molecule; IHC: immunohistochemistry; mAb: mono- clonal antibody; mL1CAM: mouse L1 cell adhesion molecule; qRT-PCR: quantitative real-time PCR; WB: Western blot Additional Supporting Information may be found in the online version of this article. Grant sponsors: DKFZ (to G.M. and P.A.), the Deutsche Krebshilfe (Schwerpunktprogramm Invasion & Metastasis; to P.A.), the Wilhelm Sander Stiftung (Project 2010.094.1; to P.A.) DOI: 10.1002/ijc.29222 History: Received 24 June 2014; Accepted 9 Sep 2014; Online 18 Sep 2014 Correspondence to: Peter Altevogt, D015, Tumor Immunology Programme, German Cancer Research Center, Im Neuenheimer Feld 280, D-69120 Heidelberg, Germany, Tel.: 16221-423714, Fax: 16221-423791, E-mail: P.Altevogt@dkfz.de Tumor Immunology Int. J. Cancer: 00, 00–00 (2014) V C 2014 UICC International Journal of Cancer IJC