Large Molecule Therapeutics Antibody-Based Delivery of IL 2 and Cytotoxics Eradicates Tumors in Immunocompetent Mice

Antibody–drug conjugates are increasingly being used for cancer therapy, but little is known about their ability to promote anticancer immunity, which may lead to long-lasting remissions. We investigated the therapeutic effect of antibody-based pharmacodelivery of cemadotin, a cytotoxic drug, and IL2, a strong proinflammatory cytokine. Using the F8 antibody, which selectively localizes to the tumor neovasculature, combination treatment led to tumor eradication, in a process dependent onCD8þT cells and natural killer cells in the C1498 syngeneic mouse model of acute myelogenous leukemia. The clinical combination of antibody– drug conjugates and antibody–cytokine proteins should be facilitated by their orthogonal toxicity profiles.Mol Cancer Ther; 13(7); 1–5. 2014 AACR. Introduction There is a growing interest in pharmaceutical biotechnology to "arm" tumor-targeting antibodieswith bioactive payloads, such as cytotoxic drugs and cytokines, to increase activity at the site of disease and spare normal tissues (1–4). Antibody–drug conjugates (ADC) and antibody–cytokine fusion proteins (immunocytokines) represent two advanced classes of armed antibodies. ADCs are on the market. Adcetris (brentuximab-vedotin) is approved for the treatment of refractory Hodgkin lymphoma and anaplastic large-cell lymphoma, whereas Kadcyla (trastuzumab-emtansine) is applied for the treatment of metastatic Her2þ breast cancer. Over 30 novel ADCs against hematologic and solid tumors are in clinical development. Immunocytokines, carrying IL2 as immunostimulatory payload, have shown selective tumor accumulation and anticancer activity, allowing these agents to progress to phase II clinical trials for the treatment of metastatic solid tumors (5). The most advanced candidates at present are immunocytokines directed against the melanoma and neuroblastoma cellular target GD2 disialoganglioside (h14.18-IL2) or the splice-isoform A1 of tenascin-C (F16IL2) and extradomainBof fibronectin (L19-IL2),which are markers of the tumor neovasculature (5). Recently, we have shown that neovascular-targeting immunocytokines can be efficient for the treatment of hematologic malignancies, including lymphomas and certain acute leukemias (6, 7). ADCs are typically studied in immunocompromised mice, due to a lack of interspecies cross-reactivity for many antibodies. As a consequence, preclinical studies with ADCs focus on cytotoxic activity, without addressing the impact of immune-mediated anticancer mechanisms. However, anticancer immunity (e.g., as a consequence of IL2 treatment) can lead to cures (8, 9) and it is known that certain cytotoxic agents may display immunomodulatory properties that justify their therapeutic application beyond chemotherapy (10, 11). To explore the possibility of improving the therapeutic potential of both ADCs and immunocytokines, we treated immunocompetent mice bearing acute myelogenous leukemia (AML) tumors with a combination of armed antibody products, featuring the cytotoxic drug cemadotin thiol (12) or IL2 as payloads. We chose the F8 antibody as delivery vehicle of proven tumor-homing performance, which recognizes the alternatively spliced extradomain A (EDA) of fibronectin, a marker of angiogenesis, with identical affinity in mouse and man (13). As negative control antibody of irrelevant specificity in the mouse, we used the anti-hen egg lysozyme antibody KSF (14). Materials and Methods Cell lines, animals, and xenograft models The murine AML cell line C1498 was purchased from the ATCC in March 2012 and cultured according to the supplier’s recommendations. Sixto 8-week-old female C57BL/6Jmicewere purchased fromElevage Janvier. For generation of the localized xenograft model, 10 C1498 cells were injected into the flank of 8to 10-week-old C57BL/6J mice. All animal experiments were performed on the basis of project license (42/2012) administered by the "Veterin€aramt des Kantons Z€ urich" and approved by all participating institutions. Authors' Affiliations: Department of Chemistry andAppliedBiosciences, ETH Zürich, Zurich; and Philochem AG, Otelfingen, Switzerland Corresponding Author: Dario Neri, ETH Z€ urich, Wolfgang-Pauli-Strasse 10, Zurich 8093, Switzerland. Phone: 41-44-633-7401; Fax: 41-44-6331358; E-mail: neri@pharma.ethz. ch doi: 10.1158/1535-7163.MCT-14-0105 2014 American Association for Cancer Research. Molecular Cancer Therapeutics www.aacrjournals. org OF1 on June 20, 2017. © 2014 American Association for Cancer Research. mct.aacrjournals. org Downloaded from Published OnlineFirst April 23, 2014; DOI: 10.1158/1535-7163.MCT-14-0105