Mutations which accelerate gl cooperate with cbfβsmmhc to induce acute leukemia in mice

Yandan Yang, Martin Britos-Bray, Linzhao Cheng, Curt I. Civin, Alan D. Friedman

Research output: Contribution to journalArticle

Abstract

In M4eo AML, the oncoprotein CBFβ-SMMHC dominantly inhibits CBF. Our finding that CBFβ-SMMHC slows Gl progression suggests a model in which mutations which stimulate the Gl to S transition cooperate with CBF oncoproteins in leukemogenesis. To test this idea in vivo, we constructed retroviral vectors encoding CBFβ-SMMHC, HPV16-E7, both, or neither. E7 stimulates Gl by inactivating Rb. Three of the vectors also encode an NGFR-GFP fusion protein, enabling use of NGFR antibody to select highlitre CRE producer cells. For the CBFβ-SMMHC/E7 vector, multiple CRE subclones were screened by viral RNA slot blot analysis. Slot blot analysis with an LTR segment probe indicated that the titre of the CBFβ-SMMHC/E7 virus was 2-fold lower than the other vectors. Unfractionated marrow cells isolated from C57BL/6 mice treated 60 hrs earlier with 150 mg/kg 5-FU were co-cultured with irradiated (40 Gy) packaging lines with IL-3, IL-6, SCF, and polybrene for 48 hrs. 2 x 10" cells were then injected IV into syngeneic recipients which had been irradiated to 9.5 Gy. In our first experiment, 3 CBFβSMMHC/E7 mice developed T-lineage ALL, at 4.5, 4.5, and 6 months. No mice with empty vector developed ALL, and one CBFβ-SMMHC mouse developed T-ALL, but not until 13 months. The E7 vector was not available for this first experiment. In a second experiment, ongoing for 5 months to date, 1 of 12 CBFβ-SMMHC/E7 mice has developed a similar ALL, at 3.5 months, whereas 0/18 E7 mice and 0/13 CBFβ-SMMHC mice have developed ALL. We also transduced marrow isolated from p!6(-/-) C57BL/6 mice with the empty vector and the vector encoding CBFβ-SMMHC. 1/7 of the latter group his developed T-lineage ALL, at 6.5 months. Finally, 2 mice injected with normal marrow transduced with either the empty vector or CBFβ-SMMHC were exposed to a mutagen, EMU. One of the CBFβ-SMMHC mice developed T-ALL. A common feature of these 7 T-cell leukemias is the presence of CBFβ-SMMHC. 3 of the leukemias have been examined for their transplantability into non-irradiated, syngeneic recipeints and for clonality, by TCR-Vβ-subtyping. Each was re-transplantable and clonal. DNA from one leukemia has been evaluated, thus far, for the presence of the retroviral vector by PCR. CBFβ-SMMF C DNA was detectable in this leukemia, and was not detected in marrow from a mouse transplanted with control-transduced marrow. T-ALL rather than AML may have develop ;d in these mice due to use of the MSCV LTR or due to species differences. In summary, our results suggest that mutations which accelerate Gl cooperate with CBFβ-SMMHC in leukemogenesis. In the presence of such mutations, CBF oncoproteins may contribute to transformation by inhibiting differentiation or apoptosis.

Original languageEnglish (US)
Pages (from-to)90a
JournalBlood
Volume96
Issue number11 PART I
StatePublished - Dec 1 2000
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Immunology
  • Hematology
  • Cell Biology

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