Figure S1. Expression of tumor-associated antigens in mouse tumor cell lines; Table S1. Number of complete responses in CT26 tumor-bearing mice that were treated with EphA2-Tub or EphA2-PBD; Figure S2. Activity of EphA2-Tub and EphA2-PBD in the Renca tumor model; Fig. S3. Control IgG-ADCs are minimally active in CT26, MCA205, 4T1, and RENCA tumor models, demonstrating target specificity for EphA2-ADC; Figure S4. CD8+ T cells are important for the efficacy of ADCs in the CT26 model; Table S2. Summary of results from combining ADCs with checkpoint inhibitors or TNFR agonists; Figure S5. Blood lymphocyte count of non-tumor bearing BALB/c and C57BL/6 mice following dosing with EphA2-Tub or EphA2-PBD; Figure S6. Combination of EphA2-ADCs with anti-PD1 or PD-L1; Figure S7. Combination of EphA2-ADCs with GITRL FP or OX40L FP; Figure S8. Activity of ADCs in the MCA205 model; Figure S9. Antitumor activity studies in the Renca model; Figure S10. Normalized CD45+ cells in the Renca model; Fig. S11. Detection of EphA2 in syngeneic mouse models via immunohistochemistry; Figure S12. Immunophenotyping of myeloid cells in the Renca model
ARTICLE ABSTRACT
Immunogenic cell death (ICD) is the process by which certain cytotoxic drugs induce apoptosis of tumor cells in a manner that stimulates the immune system. In this study, we investigated whether antibody–drug conjugates (ADCS) conjugated with pyrrolobenzodiazepine dimer (PBD) or tubulysin payloads induce ICD, modulate the immune microenvironment, and could combine with immuno-oncology drugs to enhance antitumor activity. We show that these payloads on their own induced an immune response that prevented the growth of tumors following subsequent tumor cell challenge. ADCs had greater antitumor activity in immunocompetent versus immunodeficient mice, demonstrating a contribution of the immune system to the antitumor activity of these ADCs. ADCs also induced immunologic memory. In the CT26 model, depletion of CD8+ T cells abrogated the activity of ADCs when used alone or in combination with a PD-L1 antibody, confirming a role for T cells in antitumor activity. Combinations of ADCs with immuno-oncology drugs, including PD-1 or PD-L1 antibodies, OX40 ligand, or GITR ligand fusion proteins, produced synergistic antitumor responses. Importantly, synergy was observed in some cases with suboptimal doses of ADCs, potentially providing an approach to achieve potent antitumor responses while minimizing ADC-induced toxicity. Immunophenotyping studies in different tumor models revealed broad immunomodulation of lymphoid and myeloid cells by ADC and ADC/immuno-oncology combinations. These results suggest that it may be possible to develop novel combinatorial therapies with PBD- and tubulysin-based ADC and immuno-oncology drugs that may increase clinical responses. Cancer Res; 77(10); 2686–98. ©2017 AACR.
