Abstract 6050: Transient and selective depletion of MDSCs and Tregs as an effective immunotherapy against triple-negative breast cancer
Authors: Sadiya Parveen, Sumit Siddharth, Laurene Cheung, John R. Murphy, Dipali Sharma, William R. Bishai
Journal: Cancer Research (2020)

Abstract Background: Checkpoint inhibitors (CPI) have revolutionized the field of cancer immunotherapy. However, CPIs have shown only modest efficacy in clinical trials for TNBC which may be attributed to the presence of immunosuppressive cells both myeloid-derived suppressor (MDSCs) and T regulatory (Treg) cells in the tumor microenvironment (TME). We hypothesized that transient depletion of Tregs and MDSCs along with the administration of checkpoint inhibitors would markedly reduce immunosuppressive signals to the effector arm of the immune system and thereby lead to a more robust anti-tumor response causing tumor regression and a better disease outcome. To target Treg/MDSCs cells, we have developed two diphtheria-toxin based fusion protein toxins and are currently evaluating the activity of these molecules both as monotherapies and dual therapies in combination with CPIs. Methods: To target MDSCs and TNBC tumor cells, we genetically constructed, expressed and purified a diphtheria toxin-based fusion protein, s-DAB389mIL-4 that targets the IL-4 receptor (IL-4Rα) on these cell populations. This fusion protein toxin was tested in vitro against murine TNBC 4T1 cells and in vivo in a syngeneic murine adenocarcinoma model both as a monotherapy and in combination with s-DABIL-2(V6A) which targets Tregs cells through high and intermediate affinity IL-2 receptors. Results: We found that s-DAB389mIL-4 is highly potent against 4T1 cells in vitro (IC50 ranging from 10 pM to ~3 nM), and adversely affects 4T1 cell survival, proliferation, invasion, and migratory abilities. Recombinant IL-4 and anti-IL-4 antibody block the cytotoxic action of s-DAB389mIL-4 confirming that the fusion protein's activity is mediated through the IL-4Rα. s-DAB389mIL-4 treatment in a 4T1 cell-based murine adenocarcinoma model resulted in a significant reduction of (a) tumor volume and weight, (b) MDSCs, (c) splenomegaly, (d) 4T1 metastasis to the lung and, (d) increased infiltration of IFNϒ+ effector T-cells. Sequential combination therapy with both s-DABIL-4 and s-DABIL-2(V6A) resulted in (1) reduction in both tumor volume and weight, (2) transient depletion of activated Tregs, and (3) increased infiltration of T-effector cells in the TME. NanoString analysis of RNA isolated from tumor tissues treated with these two fusion protein toxins show enrichment of gene signatures which enhance the infiltration of anti-tumor immune cells in the TME and concomitant inhibition of processes which encourage tumor growth and metastasis. Conclusions: These findings suggest that s-DABmIL-4 both as a monotherapy and in combination with s-DABIL-2(V6A) results in transient reduction of both MDSCs and Tregs. These observations suggest that these novel fusion protein toxins may serve as an effective immunotherapeutic regimen for the treatment of TNBC. Based upon these results, we are currently testing these fusion protein toxins in combination with checkpoint blockade therapy. Citation Format: Sadiya Parveen, Sumit Siddharth, Laurene Cheung, John R. Murphy, Dipali Sharma, William R. Bishai. Transient and selective depletion of MDSCs and Tregs as an effective immunotherapy against triple-negative breast cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6050.