PS929 KLF4 EXERTS A TUMOR SUPPRESSOR FUNCTION IN PATIENTS’ B‐ALL CELLS GROWING IN MICE AND IS RE‐UPREGULATED BY AZACYTIDIN
Authors: W.‐H. Liu, D. Senft, K. Vösle, M. Carlet, T. Herold, I. Jeremias
Journal: HemaSphere (2019)

Background: The transcription factor KLF4 is known for its important role in stem cells, such as induced pluripotent stem cells, but its function for B‐ALL remains unclear. Aims: Here, we aimed at deciphering the role of KLF4 for survival, growth and treatment response in patients’ B‐ALL cells and asked whether KLF4 might represent a putative therapeutic target to treat B‐ALL. Methods: As clinic‐close in vivo model, patients primary B‐ALL cells were transplanted into immunocompromised mice for amplification; resulting patient‐derived xenograft (PDX) cells were lentivirally transduced to express to physiologic levels, either wildtype KLF4 or a mutant control KLF4 lacking the DNA binding domain. An inducible transgene expression system was established and fluorochromes used in pairwise competitive in vivo assays. Results: To functionally study the role of KLF4 at a molecular level in patients’ B‐ALL cells, we aimed for an inducible expression system in ALL PDX models. For the first time ever, as far as to our knowledge, we were able to establish such an inducible system in PDX B‐ALL cells which allowed to study the re‐expression of KLF4 at different disease stages. Upon spontaneous tumor growth, KLF4 re‐expression significantly reduced B‐ALL PDX cell proliferation in vivo and induced cell cycle arrest and apoptosis. The reduced cell fitness of KLF4 expressing cells was reflected by impaired homing of wtKLF4‐expressing cells into the in vivo environment upon secondary transplantation. When KLF4 was expressed during systemic poly‐chemotherapy, PDX B‐ALL tumor burden was reduced significantly faster in cells expressing KLF4. In a MRD model, mice carrying PDX B‐ALL samples were treated with conventional chemotherapy over several weeks to reduce tumor burden by several orders of magnitude; when KLF4 was re‐expressed during tumor re‐growth after treatment, it rapidly and completely abrogated pre‐treated PDX B‐ALL cells. Our data show that re‐upregulating KLF4 impairs spontaneous growth, especially after treatment, and sensitizes B‐ALL cells towards conventional treatment. KLF4 might thus represent an interesting therapeutic target in B‐ALL. We next searched for drugs which might be able to upregulate KLF4 levels in B‐ALL. We found that Azacitidine (Aza) in low, clinically relevant concentrations upregulated KLF4. Crispr‐Cas9 induced KLF4 knockout in PDX B‐ALL cells disabled Aza‐induced upregulation of KLF4 and significantly reduced AZA‐induced cell death, suggesting that Aza‐induced cell death in B‐ALL was mediated, at least in part, by KLF4. Thus, Aza uses upregulation of KLF4 in order to eliminate B‐ALL cells and KLF4 upregulation explains, at least in part, the mode of action of Aza. Summary/Conclusion: We conclude that KLF4 plays a tumor suppressor function in B‐ALL which might be utilized therapeutically, especially to prevent relapse. Due to its ability to re‐upregulate KLF4, Azacitidine might be tested in ALL in the future.