Modelling TYK2 germline mutations predisposing for pediatric acute lymphoblastic leukemia

Introduction

Acute lymphoblastic leukemia (ALL) is the most common cancer diagnosed in children. Lately, two germline tyrosine kinase 2 (TYK2) activating mutations were identified in pediatric patients developing multiple ALL forms. The TYK2 mutations are located at a highly conserved ATP binding site within the pseudokinase domain and lead to constitutive active TYK2 and downstream STAT signalling. TYK2 activating somatic mutations and fusion proteins were previously found in several cancers suggesting that aberrant TYK2 acts as an oncogene. Our ultimate goal is to introduce activating TYK2 mutations into the germline of mice in order to study the oncogenic potential and partners of this Janus kinase (JAK) family member in vivo.

Material and methods

Murine and human TYK2 cDNA expression constructs harbouring the described germline mutations were transfected into TYK2-deficient and wildtype cell lines and downstream signalling was analyzed. Transforming capacities were further investigated in IL-3 dependent cell lines 32D and Ba/F3. Bone marrow cells of mice harbouring already a first oncogenic hit mutation were virally transduced with TYK2 constructs and adoptively transferred into immunodeficient mice. Ultimately, CRISPR/Cas9 technology will be applied to generate a gene edited mouse model expressing these two TYK2 mutations.

Results and discussion

Introduction of the two point mutations in either human or murine TYK2 leads to constitutive phosphorylation of STATs 1, 3, 5 and ERK1/2. Human TYK2 mutants transform 32D and Ba/F3 cells into survival factor independent growth. Ongoing in vivo experiments will show disease onset and enable identification of the cell types involved in TYK2 driven ALL. Together with the gene edited mouse these cell and mouse models will be important tools for mimicking the pathology of the genetically predisposed ALL patients. They will give us the possibility to study the molecular and cellular mechanisms during tumor development and test the efficacy of TYK2 inhibitors.

Conclusion

We demonstrate that two TYK2 mutations identified in the germline of cancer diseased children have the capacity to constitutively switch on TYK2 signalling and to confer for IL-3 independent growth on bone marrow and pro-B cell lines. Our studies will give insight into the oncogenic mechanisms of TYK2 in the pathogenesis of ALL and may allow for personalized therapies with small molecule tyrosine kinase inhibitors.

This work is funded by The Austrian Science Fund FWF DK W1212 and SFB F6101 and F6106.