L-Arginine is the common substrate for two
enzymes,
arginase and
nitric oxide synthase (NOS).
Arginase converts
L-arginine to L-
ornithine, which is the precursor of
polyamines, which are essential components of cell proliferation. NOS converts
L-arginine to produce NO, which inhibits proliferation of many cell lines. Various human
breast cancer cell lines were initially screened for the presence of
arginase and NOS. Two cell lines, BT-474 and MDA-MB-468, were found to have relatively high
arginase activity and very low NOS activity. Another cell line, ZR-75-30, had the highest NOS activity and comparatively low
arginase activity. The basal proliferation rates of MDA-MB-468 and BT-474 were found to be higher than the ZR-75-30 cell line. N-Hydroxy-
L-arginine (NOHA), a stable intermediate product formed during conversion of
L-arginine to NO, inhibited proliferation of the high
arginase-expressing MDA-MB-468 cells and induced apoptosis after 48 h. NOHA arrested these cells in the S phase, increased the expression of p21, and reduced
spermine content. These effects of NOHA were not observed in the ZR-75-30 cell line, which expresses high NOS and relatively low
arginase. The effects of NOHA were antagonized in the presence of L-
ornithine (500 microM), which suggests that in MDA-MB-468 cell line, the
arginase pathway is very important for cell proliferation. Inhibition of the
arginase pathway led to depletion of intracellular
spermine and apoptosis as observed by
terminal deoxynucleotidyl transferase (Tdt)-mediated nick end labeling assay and induction of
caspase 3. In contrast, the ZR-75-30 cell line maintained its viability and its L-
ornithine and
spermine levels in the presence of NOHA. We conclude that NOHA has antiproliferative and apoptotic actions on
arginase-expressing human
breast cancer cells that are independent of NO.