DNA methyltransferase inhibition overcomes diphthamide pathway deficiencies underlying CD123-targeted treatment resistance

J Clin Invest. 2019 Nov 1;129(11):5005-5019. doi: 10.1172/JCI128571.


The interleukin-3 receptor α subunit, CD123, is expressed in many hematologic malignancies including acute myeloid leukemia (AML) and blastic plasmacytoid dendritic cell neoplasm (BPDCN). Tagraxofusp (SL-401) is a CD123-targeted therapy consisting of interleukin-3 fused to a truncated diphtheria toxin payload. Factors influencing response to tagraxofusp other than CD123 expression are largely unknown. We interrogated tagraxofusp resistance in patients and experimental models and found that it was not associated with CD123 loss. Rather, resistant AML and BPDCN cells frequently acquired deficiencies in the diphthamide synthesis pathway, impairing tagraxofusp's ability to ADP-ribosylate cellular targets. Expression of DPH1, encoding a diphthamide pathway enzyme, was reduced by DNA CpG methylation in resistant cells. Treatment with the DNA methyltransferase inhibitor azacitidine restored DPH1 expression and tagraxofusp sensitivity. We also developed a drug-dependent ADP-ribosylation assay in primary cells that correlated with tagraxofusp activity and may represent an additional novel biomarker. As predicted by these results and our observation that resistance also increased mitochondrial apoptotic priming, we found that the combination of tagraxofusp and azacitidine was effective in patient-derived xenografts treated in vivo. These data have important implications for clinical use of tagraxofusp and led to a phase 1 study combining tagraxofusp and azacitidine in myeloid malignancies.

Keywords: Hematology; Leukemias; Molecular biology; Oncology; Toxins/drugs/xenobiotics.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antineoplastic Combined Chemotherapy Protocols / pharmacology*
  • Azacitidine / pharmacology
  • Cell Line, Tumor
  • DNA Methylation
  • Dendritic Cells / metabolism*
  • Dendritic Cells / pathology
  • Drug Delivery Systems*
  • Female
  • Hematologic Neoplasms* / drug therapy
  • Hematologic Neoplasms* / metabolism
  • Hematologic Neoplasms* / pathology
  • Humans
  • Interleukin-3 Receptor alpha Subunit / metabolism*
  • Leukemia, Myeloid, Acute* / drug therapy
  • Leukemia, Myeloid, Acute* / metabolism
  • Leukemia, Myeloid, Acute* / pathology
  • Male
  • Mice
  • Mice, Nude
  • Minor Histocompatibility Antigens / metabolism
  • Neoplasm Proteins / metabolism*
  • Recombinant Fusion Proteins / pharmacology
  • Tumor Suppressor Proteins / metabolism
  • Xenograft Model Antitumor Assays


  • DPH1 protein, human
  • IL3RA protein, human
  • Interleukin-3 Receptor alpha Subunit
  • Minor Histocompatibility Antigens
  • Neoplasm Proteins
  • Recombinant Fusion Proteins
  • Tumor Suppressor Proteins
  • tagraxofusp
  • Azacitidine