C. Carlo-stella et al., Effects of the tyrosine kinase inhibitor AG957 and an anti-Fas receptor antibody on CD34(+) chronic myelogenous leukemia progenitor cells, BLOOD, 93(11), 1999, pp. 3973-3982
The hallmark of chronic myelogenous leukemia (CML) is the Philadelphia (Ph)
chromosome that fuses genetic sequences of the BCR gene on chromosome 22 w
ith c-ABL sequences translocated from chromosome 9. BCR/ABL fusion proteins
have a dysregulated protein tyrosine kinase (PTK) activity exerting a key
role in malignant transformation. Targeting the tyrosine kinase activity of
BCR/ABL or using agents capable of triggering apoptosis might represent at
tractive therapeutic approaches for ex vivo purging. AG957, a member of the
tyrphostin compounds, exerts a selective inhibition of p210(BCR/ABL) tyros
ine phosphorylation. We report here that preincubation of CML or normal CD3
4(+) cells with graded concentration of AG957 (1 to 100 mu mol/L) resulted
in a statistically significant, dose-dependent suppression of colony growth
from multipotent, erythroid, and granulocyte-macrophage progenitors as wel
l as the more primitive long-term culture-initiating cells (LTC-IC). Howeve
r, AG957 doses causing 50% inhibition (ID50) Of CML and normal progenitors
were significantly different for multilineage colony-forming units (CFU-Mix
; 12 v 64 mu mol/L; P = .008), burst-forming unit-erythroid (BFU-E; 29 v 89
mu mol/L; P =.004), colony-forming unit-granulocyte-macrophage (CFU-GM; 34
v 85 mu mol/L; P = .004), and LTC-IC (43 v 181 mu mol/L; P = .004). In 5 o
f 10 patients, analysis of BCR/ABL mRNA on single progenitors by reverse tr
anscription-polymerase chain reaction showed that AG957 at 50 mu mol/L sign
ificantly reduced the mean (+/-SD) percentage of BCR/ABL-positive progenito
rs (92% +/- 10% v 33 +/- 5%; P = .001). Because AG957 treatment resulted in
significantly higher percentages of apoptotic cells (30% v 9%) in the BCR/
ABL-transfected 32DLG7 cells as compared with 32D-T2/93 cells (BCR/ABL-nega
tive), we investigated the combined effects of AG957 with the anti-Pas rece
ptor (Fas-R) monoclonal antibody CH11 that triggers apoptosis. As compared
with AG957 alone, the sequential treatment of CML CD34(+) cells with AG957
(1 mu mol/L) and CH11 (1 mu g/mL) increased CFU-Mix, BFU-E, and CFU-GM grow
th inhibition by 1.6-fold, 3-fold, and 4-fold, respectively. In contrast, t
he treatment of normal CD34(+) cells with AG957 and CH11 failed to enhance
AG957-induced colony growth inhibition. We conclude that (1) AG957 inhibits
in a dose-dependent manner CML CD34-derived colony formation by both primi
tive LTC-IC as well as committed CFU-Mix, BFU-E, and CFU-GM; (2) this growt
h inhibition is associated with the selection of a substantial amount of BC
R/ABL-negative progenitors; and (3) the antiproliferative effect of AG957 i
s dramatically increased by combining th is compound with the anti-Fas-R an
tibody CH11. These data may have significant therapeutic applications. (C)
1999 by The American Society of Hematology.