AFM13 is an NK-cell engaging CD30/CD16A bi-specific tetravalent TandAb antibody currently in Phase 2 clinical development in Hodgkin Lymphoma (HL) and CD30+ malignancies. Immune checkpoints inhibitors have demonstrated clinical efficacy in a variety of cancers, including HL. NK-cells are also regulated by a number of check-points, prompting us to investigate the combination of AFM13 with several immuno-modulatory antibodies. In previous experiments we were able to demonstrate higher efficacy of AFM13 than several immuno-modulatory antibodies in monotherapy and strong synergy between AFM13 and an anti-PD-1 antibody in vitro, as well as in vivo PDX models with human CD30+ HL tumors. In order to investigate the underlying immunological mechanisms we employed the same PDX model by implanting tumor fragments derived from surgical specimens of HL patients in immuno-deficient mice. After establishing tumors, mice were reconstituted with autologous patient-derived PBMC and treated with AFM13 alone and in combination with anti-PD-1 weekly for a total of three weeks. Tumor size, tumor-infiltrating human lymphocytes, myeloid cells and intra-tumoral cytokines were evaluated on days 30, 44, and 58, i.e. 2, 16 and 30 days after treatment start. While monotherapy with AFM13 was reproducibly more potent than anti-PD-1, significant synergy was observed when both agents were combined. Analysis of the tumors on day 58 revealed a strong correlation between tumor growth inhibition and levels of tumor-infiltrating NK-cells, T-cells, myeloid cells and intra-tumoral cytokines such as IFNg. In contrast to anti-PD-1 monotherapy, which only induced T-cell infiltration, AFM13 monotherapy was able to induce infiltration of NK- and T-cells in the tumors, however the combination further enhanced infiltration of both, NK- and T-cells. AFM13 resulted in stronger infiltration of macrophages than anti-PD-1, which was also increased by the combination of both agents, therefore further supporting cross-talk between innate and adaptive immunity. Furthermore, tumor analyses at earlier time-points (days 30 and 44) showed that the initial immune response is characterized by NK-cell infiltration and activation, as well as infiltration of macrophages, whilst the adaptive immune response by T-cells and activated dendritic cells was more pronounced on day 58. Combining AFM13 and anti-PD-1 augments infiltration and activation of all immune subpopulations.

In conclusion, our data support strong synergistic anti-tumor efficacy when AFM13 is combined with anti-PD-1 checkpoint blockade in HL PDX models, mediated by tumor-infiltrating lymphocytes, macrophages and dendritic cells, and provide strong evidence for cross-talk between innate and adaptive immunity induced by AFM13-recruited human NK-cells.

Citation Format: Xing Zhao, Narendiran Rajasekaran, Uwe Reusch, Jens-Peter Marschner, Martin Treder, Holbrook E. Kohrt. Immune checkpoint inhibition by anti-PD-1 or CD137 co-stimulation enhances cytotoxicity towards CD30+ tumors mediated by the bispecific tetravalent CD30/CD16A TandAb AFM13. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2323.