Genetic Inactivation of CD33 in Hematopoietic Stem Cells to Enable CAR T Cell Immunotherapy for Acute Myeloid Leukemia

Cell. 2018 May 31;173(6):1439-1453.e19. doi: 10.1016/j.cell.2018.05.013. Epub 2018 May 31.


The absence of cancer-restricted surface markers is a major impediment to antigen-specific immunotherapy using chimeric antigen receptor (CAR) T cells. For example, targeting the canonical myeloid marker CD33 in acute myeloid leukemia (AML) results in toxicity from destruction of normal myeloid cells. We hypothesized that a leukemia-specific antigen could be created by deleting CD33 from normal hematopoietic stem and progenitor cells (HSPCs), thereby generating a hematopoietic system resistant to CD33-targeted therapy and enabling specific targeting of AML with CAR T cells. We generated CD33-deficient human HSPCs and demonstrated normal engraftment and differentiation in immunodeficient mice. Autologous CD33 KO HSPC transplantation in rhesus macaques demonstrated long-term multilineage engraftment of gene-edited cells with normal myeloid function. CD33-deficient cells were impervious to CD33-targeting CAR T cells, allowing for efficient elimination of leukemia without myelotoxicity. These studies illuminate a novel approach to antigen-specific immunotherapy by genetically engineering the host to avoid on-target, off-tumor toxicity.

Keywords: CD33; CRISPR/Cas9 gene editing; acute myeloid leukemia; chimeric antigen receptor T cells; hematopoiesis; immunotherapy; non-human primate hematopoiesis.

Publication types

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

MeSH terms

  • Animals
  • Cell Differentiation
  • Cell Line, Tumor
  • Cell Lineage
  • Electroporation
  • Female
  • Hematopoiesis
  • Hematopoietic Stem Cells / cytology*
  • Humans
  • Immunotherapy / methods*
  • Leukemia, Myeloid, Acute / immunology
  • Leukemia, Myeloid, Acute / therapy*
  • Macaca mulatta
  • Male
  • Mice
  • Mice, Inbred NOD
  • Mice, Knockout
  • Mice, SCID
  • Neoplasm Transplantation
  • RNA, Guide, CRISPR-Cas Systems / genetics*
  • Reactive Oxygen Species
  • Sialic Acid Binding Ig-like Lectin 3 / genetics*
  • T-Lymphocytes / cytology
  • T-Lymphocytes / immunology*


  • CD33 protein, human
  • RNA, Guide, CRISPR-Cas Systems
  • Reactive Oxygen Species
  • Sialic Acid Binding Ig-like Lectin 3