Establishment and characterization of a novel acute promyelocytic leukemia cell line (UF-1) with retinoic acid-resistant features

Blood ◽  
1996 ◽  
Vol 88 (5) ◽  
pp. 1824-1833 ◽  
Author(s):  
M Kizaki ◽  
H Matsushita ◽  
N Takayama ◽  
A Muto ◽  
H Ueno ◽  
...  
Keyword(s):  
Retinoic Acid ◽  
Cell Line ◽  
Acute Promyelocytic Leukemia ◽  
Leukemia Cell ◽  
Leukemic Cell ◽  
Fusion Transcript ◽  
Promyelocytic Leukemia ◽  
Leukemia Cell Line ◽  
Human Leukemia

All-trans retinoic acid (RA) induces complete remission in a high proportion of patients with acute promyelocytic leukemia (APL). Nevertheless, most of these patients develop RA resistance and relapse. The mechanisms of RA resistance by APL cells are still unclear. To understand the characteristics of human leukemia, human leukemic cell lines are useful tools for study. APL cells have a strikingly low proliferation potential in vitro; thus, only one APL cell line has been established. We developed a novel APL cell line (UF-1) from a patient clinically resistant to all-trans RA. Cell surface markers in the UF-1 cells were positive for CD7, CD13, CD33, and CD38. Cytogenetic analyses revealed additional abnormalities, 46XX, add(1)(q44), add(6)(q12), add(7)(q36), t(15;17) (q21;q21). Molecular analyses showed a PML/RAR alpha fusion transcript. Sequence analysis of the RAR alpha gene in RA- resistant HL-60 cells disclosed a point mutation in codon 411 (C to T substitution), whereas UF-1 cells showed the normal sequence. All-trans RA did not change morphological features of the cell, NBT reduction activity, or their expression of CD11b antigens as determined by FACS analysis except at 10(-6) mol/L. RA also did not alter the growth curve of the cells as determined by the MTT assay. These findings suggest that the UF-1 cell is the first permanent cell line with spontaneous RA- resistant APL cells. This RA-resistant APL cell line may be a useful model for molecular studies on the block of leukemic cell differentiation and as a means to investigate the mechanisms of RA resistance.

scholarly journals Establishment and characterization of a novel acute promyelocytic leukemia cell line (UF-1) with retinoic acid-resistant features

Blood ◽  
1996 ◽  
Vol 88 (5) ◽  
pp. 1824-1833 ◽  
Author(s):  
M Kizaki ◽  
H Matsushita ◽  
N Takayama ◽  
A Muto ◽  
H Ueno ◽  
...  
Keyword(s):  
Retinoic Acid ◽  
Cell Line ◽  
Acute Promyelocytic Leukemia ◽  
Leukemia Cell ◽  
Leukemic Cell ◽  
Fusion Transcript ◽  
Promyelocytic Leukemia ◽  
Leukemia Cell Line ◽  
Human Leukemia

Abstract All-trans retinoic acid (RA) induces complete remission in a high proportion of patients with acute promyelocytic leukemia (APL). Nevertheless, most of these patients develop RA resistance and relapse. The mechanisms of RA resistance by APL cells are still unclear. To understand the characteristics of human leukemia, human leukemic cell lines are useful tools for study. APL cells have a strikingly low proliferation potential in vitro; thus, only one APL cell line has been established. We developed a novel APL cell line (UF-1) from a patient clinically resistant to all-trans RA. Cell surface markers in the UF-1 cells were positive for CD7, CD13, CD33, and CD38. Cytogenetic analyses revealed additional abnormalities, 46XX, add(1)(q44), add(6)(q12), add(7)(q36), t(15;17) (q21;q21). Molecular analyses showed a PML/RAR alpha fusion transcript. Sequence analysis of the RAR alpha gene in RA- resistant HL-60 cells disclosed a point mutation in codon 411 (C to T substitution), whereas UF-1 cells showed the normal sequence. All-trans RA did not change morphological features of the cell, NBT reduction activity, or their expression of CD11b antigens as determined by FACS analysis except at 10(-6) mol/L. RA also did not alter the growth curve of the cells as determined by the MTT assay. These findings suggest that the UF-1 cell is the first permanent cell line with spontaneous RA- resistant APL cells. This RA-resistant APL cell line may be a useful model for molecular studies on the block of leukemic cell differentiation and as a means to investigate the mechanisms of RA resistance.

scholarly journals Retinoic acid is required for and potentiates differentiation of acute promyelocytic leukemia cells by nonretinoid agents

Blood ◽  
1994 ◽  
Vol 84 (7) ◽  
pp. 2122-2129 ◽  
Author(s):  
A Chen ◽  
JD Licht ◽  
Y Wu ◽  
N Hellinger ◽  
W Scher ◽  
...  
Keyword(s):  
Retinoic Acid ◽  
Cell Line ◽  
Acute Promyelocytic Leukemia ◽  
Drug Exposure ◽  
Primary Cultures ◽  
Leukemia Cell ◽  
Promyelocytic Leukemia ◽  
Leukemia Cell Line ◽  
Granulocytic Differentiation ◽  
Nb4 Cells

Abstract Patients with acute promyelocytic leukemia (APL) associated with the t(15;17) translocation and fusion of the promyelocytic leukemia (PML) and retinoic acid receptor-alpha (RAR-alpha) genes achieve complete remission but not cure with all-trans retinoic acid (RA), NB4, a cell line derived from a patient with t(15;17) APL that undergoes granulocytic differentiation when treated with pharmacologic doses of RA, was used as a model for differentiation therapy of APL. We found that NB4 cells are resistant to differentiation by nonretinoid inducers such as hexamethylene bisacetamide (HMBA), butyrates, vitamin D3, or hypoxanthine, all of which can induce differentiation in the commonly used HL60 leukemia cell line. Preexposure of NB4 cells to low concentrations of RA for a period as short as 30 minutes abolished resistance to nonretinoids and potentiated differentiation. Sequential RA and HMBA treatment yielded maximal differentiation by 3 days of drug exposure, whereas the effect of RA alone peaked after 6 days and yielded a smaller percentage of differentiated cells. RA also reversed NB4 cell resistance to butyrates and allowed for synergistic differentiation by these agents. Pretreatment with HMBA before exposure to RA failed to stimulate differentiation. Sequential RA/HMBA treatment also markedly increased the extent of differentiation of primary cultures of bone marrow and peripheral blood mononuclear cells from three APL patients. In one case RA/HMBA treatment overcame resistance to RA in vitro. Together, these results suggest that intermittent low doses of RA followed by either HMBA or butyrates may be a useful combination in the treatment of APL. This clinical strategy may help prevent or overcome RA resistance in APL.

scholarly journals Retinoic acid is required for and potentiates differentiation of acute promyelocytic leukemia cells by nonretinoid agents

Blood ◽  
1994 ◽  
Vol 84 (7) ◽  
pp. 2122-2129 ◽  
Author(s):  
A Chen ◽  
JD Licht ◽  
Y Wu ◽  
N Hellinger ◽  
W Scher ◽  
...  
Keyword(s):  
Retinoic Acid ◽  
Cell Line ◽  
Acute Promyelocytic Leukemia ◽  
Drug Exposure ◽  
Primary Cultures ◽  
Leukemia Cell ◽  
Promyelocytic Leukemia ◽  
Leukemia Cell Line ◽  
Granulocytic Differentiation ◽  
Nb4 Cells

Patients with acute promyelocytic leukemia (APL) associated with the t(15;17) translocation and fusion of the promyelocytic leukemia (PML) and retinoic acid receptor-alpha (RAR-alpha) genes achieve complete remission but not cure with all-trans retinoic acid (RA), NB4, a cell line derived from a patient with t(15;17) APL that undergoes granulocytic differentiation when treated with pharmacologic doses of RA, was used as a model for differentiation therapy of APL. We found that NB4 cells are resistant to differentiation by nonretinoid inducers such as hexamethylene bisacetamide (HMBA), butyrates, vitamin D3, or hypoxanthine, all of which can induce differentiation in the commonly used HL60 leukemia cell line. Preexposure of NB4 cells to low concentrations of RA for a period as short as 30 minutes abolished resistance to nonretinoids and potentiated differentiation. Sequential RA and HMBA treatment yielded maximal differentiation by 3 days of drug exposure, whereas the effect of RA alone peaked after 6 days and yielded a smaller percentage of differentiated cells. RA also reversed NB4 cell resistance to butyrates and allowed for synergistic differentiation by these agents. Pretreatment with HMBA before exposure to RA failed to stimulate differentiation. Sequential RA/HMBA treatment also markedly increased the extent of differentiation of primary cultures of bone marrow and peripheral blood mononuclear cells from three APL patients. In one case RA/HMBA treatment overcame resistance to RA in vitro. Together, these results suggest that intermittent low doses of RA followed by either HMBA or butyrates may be a useful combination in the treatment of APL. This clinical strategy may help prevent or overcome RA resistance in APL.

scholarly journals Occurrence of resistance to retinoic acid in the acute promyelocytic leukemia cell line NB4 is associated with altered expression of the pml/RAR alpha protein

Blood ◽  
1993 ◽  
Vol 82 (5) ◽  
pp. 1573-1577 ◽  
Author(s):  
S Dermime ◽  
F Grignani ◽  
M Clerici ◽  
C Nervi ◽  
G Sozzi ◽  
...  
Keyword(s):  
Retinoic Acid ◽  
Cell Line ◽  
Acute Promyelocytic Leukemia ◽  
Leukemia Cell ◽  
Promyelocytic Leukemia ◽  
Leukemia Cell Line ◽  
Altered Expression ◽  
Trans Retinoic Acid ◽  
All Trans Retinoic Acid ◽  
Promyelocytic Leukemia Cell Line

The mechanism(s) by which acute promyelocytic leukemia (APL) cells acquire resistance to all-trans retinoic acid (ATRA) is poorly understood. We describe here an APL cell line, named NB4.306, that shows resistance to the anti-proliferative action of ATRA. This cell line is also operationally resistant to most ATRA-induced phenotypic modifications (CD11b, CD11c, CD13, and CD33). No significant differences in ATRA intracellular accumulation, efflux, or metabolism were found between NB4.306 and the parent NB4 cell line that could explain the observed resistance of the NB4.306 line. The NB4.306 cell line was found to be positive for the t15;17 translocation and showed the usual pml/RAR alpha fusion bands in both Southern and Northern blot assays, but expressed no detectable amount of the usual pml/RAR alpha protein, as assayed by Western blot analysis using an anti-RAR alpha antibody. These results were confirmed in 14 of 14 clones obtained from the NB4.306 cell line, while 30 of 30 clones obtained from the parental NB4 line expressed the usual 110-Kd fusion polypeptide. It is concluded that the occurrence of resistance to ATRA in the NB4.306 cell line is closely associated to the loss of expression of the intact pml/RAR alpha protein.

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