1. Novel vitamin D3 analog, 21-(3-methyl-3-hydroxy-butyl)-19-nor D3, that modulates cell growth, differentiation, apoptosis, cell cycle, and induction of PTEN in leukemic cells
by Jun-ichi Hisatake, James O'Kelly, Milan R. Uskokovic, Shigeru Tomoyasu, and H. Phillip Koeffler
Blood
Volume 97(8):
April 15, 2001
©2001 by American Society of Hematology

2. Chemical structures of 1,25(OH)2D3 and Gemini compounds.
Jun-ichi Hisatake et al. Blood 2001;97:
©2001 by American Society of Hematology

3. Dose-response effects of vitamin D3compounds on clonal proliferation of cell lines.Results are expressed as a mean (± SD) percentage of control plates containing no vitamin D3 analogs.  = , 1,25(OH)2D3; c, Gemini-5,6-trans; Δ, Gemini-3-epi; O, Gemini-19-nor...
Dose-response effects of vitamin D3compounds on clonal proliferation of cell lines.Results are expressed as a mean (± SD) percentage of control plates containing no vitamin D3 analogs.  = , 1,25(OH)2D3; c, Gemini-5,6-trans; Δ, Gemini-3-epi; O, Gemini-19-nor; X, Gemini-1-F-25-OH. Each point represents a mean of 3 independent experiments with triplicate dishes.
Jun-ichi Hisatake et al. Blood 2001;97:
©2001 by American Society of Hematology

4. Dose-response effects of vitamin D3compounds on clonal proliferation of normal bone marrow-committed myeloid stem cells (CFU-GM).Results are expressed as a mean (± SD) percentage of control plates containing no vitamin D3 analogs.  = , 1,25(OH)2D3; O, Gemin...
Dose-response effects of vitamin D3compounds on clonal proliferation of normal bone marrow-committed myeloid stem cells (CFU-GM).Results are expressed as a mean (± SD) percentage of control plates containing no vitamin D3 analogs.  = , 1,25(OH)2D3; O, Gemini-19-nor. Each point represents a mean of 3 independent experiments with triplicate dishes.
Jun-ichi Hisatake et al. Blood 2001;97:
©2001 by American Society of Hematology

5. Induction of cell surface antigens on HL-60 cells by vitamin D3 compounds.HL-60 cells were treated for 4 days with different concentrations (10−8 or 10−7 M) of either 1,25(OH)2D3 or Gemini-19-nor and then analyzed for expression of either CD11b or CD14 usin...
Induction of cell surface antigens on HL-60 cells by vitamin D3 compounds.HL-60 cells were treated for 4 days with different concentrations (10−8 or 10−7 M) of either 1,25(OH)2D3 or Gemini-19-nor and then analyzed for expression of either CD11b or CD14 using flow cytometry. (A) Dashed line indicates negative control antibody; solid line, CD11b or CD14. (B) Column indicates mean (± SD) of 3 independent experiments. *P < .05; **P < .01 as determined by Student t test difference compared with the control group. Histograms show representative results from one experiment.
Jun-ichi Hisatake et al. Blood 2001;97:
©2001 by American Society of Hematology

6. Effect of vitamin D3 compounds on reduction of NBT by HL-60 cells
Effect of vitamin D3 compounds on reduction of NBT by HL-60 cells.HL-60 cells were treated for 4 days with different concentrations (10−8 or 10−7 M) of either 1,25(OH)2D3 or Gemini-19-nor and then analyzed for reduction of NBT. Column indicates mean (± SD) ...
Effect of vitamin D3 compounds on reduction of NBT by HL-60 cells.HL-60 cells were treated for 4 days with different concentrations (10−8 or 10−7 M) of either 1,25(OH)2D3 or Gemini-19-nor and then analyzed for reduction of NBT. Column indicates mean (± SD) of 3 independent experiments. *P < .05; **P < .01 as determined by Student t test difference compared with the control group.
Jun-ichi Hisatake et al. Blood 2001;97:
©2001 by American Society of Hematology

7. Induction of apoptosis by vitamin D3compounds
Induction of apoptosis by vitamin D3compounds.HL-60 cells were treated for up to 4 days with different concentrations (10−8 or 10−7 M) of either 1,25(OH)2D3 or Gemini-19-nor and then analyzed by flow cytometry for cells that were annexin V-FITC+ and PI−, wh...
Induction of apoptosis by vitamin D3compounds.HL-60 cells were treated for up to 4 days with different concentrations (10−8 or 10−7 M) of either 1,25(OH)2D3 or Gemini-19-nor and then analyzed by flow cytometry for cells that were annexin V-FITC+ and PI−, which represent those in the early stages of apoptosis. (A) The upper right quadrant of each histogram shows necrotic cells that were PI+ and annexin V+. The lower right quadrant shows apoptotic cells that were not necrotic (PI−) and annexin V+. Histograms show representative data from one experiment. (B) Percentage of apoptotic cells after 4 days. Column indicates mean (± SD) of 3 independent experiments. *P < .05 as determined by Studentt test difference compared with the control group.
Jun-ichi Hisatake et al. Blood 2001;97:
©2001 by American Society of Hematology

8. Cell cycle modulation by vitamin D3compounds
Cell cycle modulation by vitamin D3compounds.HL-60 cells were cultured for 4 days with either 1,25(OH)2D3 (10−7 M) or Gemini-19-nor (10−7 M), fixed, and stained with PI, and the cell cycle status was analyzed using flow cytometry.
Cell cycle modulation by vitamin D3compounds.HL-60 cells were cultured for 4 days with either 1,25(OH)2D3 (10−7 M) or Gemini-19-nor (10−7 M), fixed, and stained with PI, and the cell cycle status was analyzed using flow cytometry. Column indicates mean (± SD) of 3 independent experiments. *P < .05 as determined by Student t test difference compared with the control group.
Jun-ichi Hisatake et al. Blood 2001;97:
©2001 by American Society of Hematology

9. Induction of expression of p27kip1 and PTEN by vitamin D3 compounds
Induction of expression of p27kip1 and PTEN by vitamin D3 compounds.(A) Dose-dependent study of p27kip1 and PTEN expression in HL-60 cells analyzed by Western blot.
Induction of expression of p27kip1 and PTEN by vitamin D3 compounds.(A) Dose-dependent study of p27kip1 and PTEN expression in HL-60 cells analyzed by Western blot. Cells were either untreated (control) or cultured with 10−9 to 10−7 M of either 1,25(OH)2D3 or Gemini-19-nor for 4 days. GAPDH was used as a loading control. (B) Time course study of p27kip1 and PTEN expression in HL-60 cells studied by Western blot. Cells were either untreated (Control) or cultured with Gemini-19-nor (10−7 M) for 0.5 to 4 days. GAPDH was used as a loading control. The densities of the bands were measured using densitometery. (C) Induction of PTEN expression by TPA and ATRA in HL-60 cells. Cells were either untreated (control) or cultured with either TPA (10−9 M) or ATRA (10−7 M) for 4 days. GAPDH was analyzed as a loading control.
Jun-ichi Hisatake et al. Blood 2001;97:
©2001 by American Society of Hematology