Targeting persistent leukemic cells by combined treatments with NKG2D ligand inducers and engineered NK/T cells

• Applicants: Professorin Dr. Claudia Lengerke; Dr. Christian Seitz
• Subject Area: Hematology, Oncology ; Immunology
• Term: since 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 539826039

Refractoriness to initial treatment and relapse from persistent leukemic stem cells (LSCs) are the main drivers of poor outcome in patients with acute myeloid leukemia (AML). We have previously shown that LSCs not only exhibit enhanced resistance to chemotherapy, but also selectively evade natural killer (NK) cell immune surveillance, e.g., by suppressing the surface expression of ligands for activating NKG2D receptors on NK and T cells. We also found that PARP1 inhibition partly re-induced NKG2DL expression on CD34+ LSCs, sensitizing them to clearance by NK cells in a pre-clinical patient derived xenograft (PDX) model (Paczulla et al., 2019). However, some CD34+ putative LSCs still failed to express NKG2DL upon PARP1 inhibition, and LSCs from CD34 non-expressing AML patients did not respond at all. The NKG2D-NKG2DL axis is of particular interest for cancer immunotherapy, since NKG2DL are selectively expressed on malignant but not on healthy cells. This is particularly valuable in AML, where antigens are difficult to identify for targeted immunotherapy. In this proposal, we will further investigate the role of NKG2DL in targeting LSCs. In Aim 1, we explore new methods that can induce NKG2DL on LSCs with poor or no response to PARP1 inhibition alone. To this end, we use a CRISPR/Cas9-mediated screening approach to identify new pathways that regulate NKG2DL surface expression on leukemic cells. We then seek related drugs and test these, alone or in combination with PARP1 inhibitors, for their ability to induce NKG2DL on CD34 non-expressing AML LSCs, and respectively to further augment PARP1 inhibitor treatment efficacy in CD34 expressing AML. In Aim 2, we generate NKG2D-based chimeric immune receptors (CIR) or AdCAR NK/T cells and test their ability to lyse AML cells. We hypothesize that these engineered immune cells will be able to target persistent LSCs with modest NKG2DL induction. In Aim 3, we are testing whether a combination of NKG2DL inducers and NKG2DL-targeted genetically modified NK/T cells can efficiently kill persistent LSCs in immunosuppressive niches. We are using in vitro co-culture assays of immune and AML cells to identify the most promising co-treatment and then test it in established PDX models. The results of this project will enhance our understanding of the NKG2D-NKG2DL axis in AML, contribute to the development of new cellular therapies, and provide a treatment rationale for an improved targeting of persistent LSCs in CD34-expressing as well as non-expressing AML.

• DFG Programme: Research Grants