Deciphering bone marrow stroma-induced resistance to targeted therapies in T-cell acute lymphoblastic leukemia using functional genetics.

• Applicant: Dr. Adrian Schwarzer, Ph.D.
• Subject Area: Hematology, Oncology
• Term: since 2021
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 471798038

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive leukemia caused by uncontrolled proliferation of immature T-cell precursors. Despite substantial progress in the understanding of the biology of T-ALL, its treatment remains challenging. In contrast to other types of leukemia, no targeted therapies are available for T-ALL. Therefore, multiple efforts have been leveraged to discover druggable drivers of T-ALL. The most promising candidates identified so far are the NOTCH1- and the PI3K-AKT-mTOR signaling pathways. However, although both NOTCH1 and mTOR are central drivers of T-ALL they yielded only mixed results in the clinical setting. In our preliminary work, we identified interactions between T-ALL cells and protective niches as a key factor for resistance against mTOR- and NOTCH1-inhibition in vivo. We developed a T-ALL – bone marrow stroma co-culture model that recapitulates this effect in vitro and used RNA-interference (RNAi) and CRISPR-Cas9 genome editing to identify genes that mediate stroma-induced therapeutic resistance. We conducted a proof-of-principle RNAi-screen and identified the RAC2-GTPase as a critical mediator of stroma-induced resistance to NOTCH1 and mTOR inhibition. In this project, we will first extend these findings by investigating stroma-induced resistance against targeted therapies in subtype-specific models of T-ALL. By using a novel mouse strain developed in our laboratory together with CRISPR-Cas9 we are able to generate new T-ALL models which better reflect the complex biology of the disease. Using these models, we will investigate whether stroma-induced resistance is a common barrier to anti-NOTCH1 therapy in other subtypes and whether it also induces resistance to inhibition of other signaling pathways, such as JAK/STAT signaling. In the second step, we will perform CRISPR-Cas9 screens in different T-ALL models to obtain a high and unbiased resolution of the intracellular networks involved in stroma-mediated resistance in T-ALL. Third, we will study the molecular mechanisms of genes involved in this process using targeted protein degradation followed by gene expression profiling and proteomics to build a comprehensive model that explains mechanisms and key pathways of cell adhesion-mediated therapeutic resistance. Finally, we will substantiate our findings by testing whether inhibition of the identified candidate genes can abrogate the resistance to NOTCH1- and mTOR- inhibition observed in vivo using our T-ALL models and xenograft models of primary T-ALL from patients. Our work supports the notion that leukemic blasts hijack the bone marrow microenvironment and depend on niche signals for disease progression, self-renewal, and therapy evasion. This project combines functional genetics, bioinformatics, and mouse models to systematically interrogate the interaction of T-ALL cells with bone marrow stroma to find and characterize druggable targets that overcome resistance to targeted therapies in T-ALL.

• DFG Programme: Research Grants
• International Connection: Austria
• Cooperation Partner: Professor Dr. Johannes Zuber