Bypass of senescence, immortalization, and transformation of human hematopoietic progenitor cells

Stem Cells. 2005 Oct;23(9):1423-33. doi: 10.1634/stemcells.2005-0390. Epub 2005 Sep 6.

Abstract

We attempted to extend the lifespan of CD34+ stem/progenitor cells in human cord blood (CB) by transduction with lentiviral vectors carrying the human telomerase catalytic subunit (hTERT) and/or the human papillomavirus type 16 (HPV16) E6 and E7 oncogenes. We found that hTERT was incapable of prolonging the replicative capacity of CB cells maintained under serum-free conditions in the presence of stem cell factor, Flt3 ligand, thrombopoietin, and interleukin-3 beyond 4 months (n=3). However, transduced CB cells cultured in the same cytokine cocktail constitutively expressing HPV16 E6/E7 alone (n=2) or in concert with hTERT (n=9) continued to proliferate, giving rise to permanent (>2 years) cell lines with a CD45+ CD34- CD133+/- CD44+ CD235a+ CD71+ CD203+ CD33+ CD13+ myeloerythroid/mast cell progenitor phenotype. Notably, CB cell cultures expressing only HPV16 E6/E7 went through a crisis period, and the resulting oligoclonal cell lines were highly aneuploid. By comparison, the CB cell lines obtained by coexpression of HPV16 E6/E7 plus hTERT exhibited near-diploid karyotypes with minimal chromosomal aberrations, concomitant with stabilization of telomere length, yet were clonally derived. The immortalized E6/E7 plus hTERT-expressing CB cells were not tumorigenic when injected intravenously or subcutaneously into sublethally irradiated immunodeficient nonobese diabetic/severe combined immunodeficient mice but could be converted to a malignant state by ectopic expression of a v-H-ras or BCR-ABL oncogene. These findings provide new insights into the mechanisms governing the senescence checkpoint of primitive human hematopoietic precursors and establish a paradigm for studies of the multistep process of human leukemogenesis.

Publication types

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

MeSH terms

  • Antigens, CD34 / biosynthesis
  • Cell Line, Transformed
  • Cell Transformation, Viral / genetics
  • Cell Transformation, Viral / physiology*
  • Cellular Senescence
  • DNA-Binding Proteins / genetics
  • Fetal Blood / cytology
  • Genetic Vectors / genetics
  • HIV-1 / genetics
  • Hematopoietic Stem Cell Transplantation
  • Hematopoietic Stem Cells / cytology*
  • Hematopoietic Stem Cells / enzymology
  • Hematopoietic Stem Cells / physiology
  • Hematopoietic Stem Cells / virology
  • Humans
  • Lentivirus / genetics
  • Oncogene Proteins, Viral / genetics
  • Papillomavirus E7 Proteins
  • Repressor Proteins / genetics
  • Stem Cells / cytology*
  • Stem Cells / enzymology
  • Stem Cells / physiology
  • Stem Cells / virology
  • Telomerase / genetics
  • Telomerase / metabolism
  • Transduction, Genetic

Substances

  • Antigens, CD34
  • DNA-Binding Proteins
  • E6 protein, Human papillomavirus type 16
  • Oncogene Proteins, Viral
  • Papillomavirus E7 Proteins
  • Repressor Proteins
  • oncogene protein E7, Human papillomavirus type 16
  • Telomerase