PS920 PHF6 LOSS DRIVES IL7R ONCOGENE ADDICTION IN TLX1 DRIVEN T‐ALL

Abstract

Background:T‐cell acute lymphoblastic leukemia (T‐ALL) is a genetically heterogeneous disease. The PHF6 gene is frequently targeted by loss‐of‐function mutations or deletions, with the highest prevalence in TLX1 or TLX3 rearranged T‐ALLs.Aims:To gain insights into the putative function of PHF6 as a tumor suppressor in the T‐cell lineage, we investigated the effects of PHF6 knock down during normal and malignant thymocytes both in vitro as in vivo through zebrafish modelling.Methods:We evaluated transcriptomal changes upon PHF6 and TLX1 modulation in vitro. In addition, we set up a T‐ALL acceleration study in vivo by generating a PHF6 knock out and TLX1 over expressing zebrafish line followed by T‐ALL formation monitoring.Results:Notably, we observed broad effects on the investigated transcriptomes in vitro suggesting an important role for PHF6 in gene regulation. Furthermore, IL7R was identified as a common transcriptional target that was significantly upregulated upon PHF6 knockdown in both normal and malignant T cells. IL7R encodes a cytokine receptor critically involved in normal thymic development and which also acts as a bona fide oncogene in subset of primary T‐ALLs. Thus, loss of PHF6 might further boost oncogenic addiction of leukemic T‐cell lymphoblast to IL7‐induced JAK‐STAT signaling.To further explore the role of PHF6 inactivation in TLX1 driven leukemogenesis in vivo, we performed zebrafish modeling. For this, we generated a stable tg(rag2:TLX1, rag2:GFP) overexpressing as well as a phf6 knock out zebrafish line. These lines were crossed and offspring was monitored for T‐ALL formation. Interestingly, 2 out of 70 (2.9%) TLX1 overexpressing only fish developed T‐ALL at an age of 12–15 months. In addition, 3 fish out of a cohort of 64 (4.7%) animals developed leukemia between 10 to 18 months of age. These leukemias originated from the thymus, spreaded throughout the whole body and were transplantable. Thus far, no leukemia was detected in PHF6 mutated only zebrafish. Leukemic cells obtained from tumors that developed in the TLX1 and PHF6/TLX1 animals were subjected to RNA‐, ATAC‐ and H3K27ac ChIP‐sequencing to assess the epigenetic status of the IL7R locus. In addition, exome‐, and CNV‐sequencing was performed to identify somatic lesions that cooperated loss of Phf6 during TLX1 driven T‐cell transformation in zebrafish. Furthermore, additional injections of TLX1 in combination with an activating IL7R mutant into phf6 mutant zebrafish are currently ongoing to monitor additional effects on accelerated tumor formation.Summary/Conclusion:In conclusion, our data suggest that loss of PHF6 drives TLX1 mediated leukemogenesis, at least in part, by increasing surface IL7R expression. Therefore, we believe that increased addiction to oncogenic JAK‐STAT signaling may render PHF6 mutant leukemic cells more sensitive to JAK inhibitors, a notion that we are currently investigating in our TLX1/PHF6 and TLX1/PHF6/IL7R zebrafish models.