TER286 [gamma-glutamyl-alpha-amino-beta(2-ethyl-N,N,N',N'-tetrakis(2-chloro
ethyl) phosphorodiamidate)-sulfonyl-propionyl-(R)-(-) phenylglycine] is a n
ovel nitrogen mustard prodrug that is preferentially activated by glutathio
ne S-transferase P1-1 (GSTP1-1). A human promyelocytic leukemia /TER286-res
istant cell line was selected by chronic, long-term exposure to the prodrug
. Although resistance was not readily achieved, eventually a 5-fold resista
nt clone was isolated. Cross-resistance to melphalan occurred, but not to d
oxorubicin (Adriamycin), taxol, and gamma-glutamyl-S-(benzyl)cysteinyl-R(-)
-phenyl glycine diethyl ester, a GSTP1-1 inhibitor. The protein and transcr
ipt levels and enzymatic activity of GSTP1-1 were reduced significantly in
the selected resistant line. GST alpha levels were unchanged, and GST mu wa
s undetectable. Although glutathione levels were elevated in human promyelo
cytic leukemia/TER286 cells, no changes in the expression of thiol-related
genes including gamma-glutamylcysteine synthetase, gamma-glutamyl transpept
idase, or multidrug resistance protein were found. A 7-fold increase in cat
alase expression in the resistant cell line indicated an adaptive response
to oxidative and electrophilic stress, and this was also reflected in the l
ower prevalence of drug-induced DNA single-strand breaks in the resistant c
ells. Mouse embryo fibroblast GSTP1-1(-/-) cells exhibited 2-fold resistanc
e to TER286 compared with GSTP1-1(+/+) cells. NIH3T3 cells transfected with
combinations of gamma-GCS and multidrug resistance protein exhibited enhan
ced resistance to TER286, although the degree of resistance was impaired by
cotransfection of GSTP1-1. These results are consistent with responses in
the TER286-resistant cells indicative of GSTP1-1-mediated mechanism of acti
vation. In consequence, these data support the rationale that tumors expres
sing high levels of GSTP1-1 will be more sensitive to the cytotoxic effects
of the drug.