34 3 ART for Antitumor
evident by monitoring the shift of absorbance from heme (A 415 ) to ART-heme
adducts (A 476 ). Accordingly, it was observed that a transient elevation of A 415 is
accompanied with a synchronous burst of NO and a higher rate of survival fol-
lowing incubation of tumor cells with 50 μM ART. In contrast, ART at above
100 μM leads to a lower NO level along with a lower survival rate, strongly
implying that NOS may represent an important target of ART for killing tumor
cells. Although CAT as a hemoprotein also interacts with ART, no remarkable
fluctuation of CAT activity was observed after the incubation of tumor cells with
50, 100, or 200 μM ART for 48 h, suggesting that ART kills tumor cells unlikely
through the H 2 O 2 -mediated apoptosis mechanism. Furthermore, we observed
that a combination of ART with FLU exhibits a synergistic effect in killing
tumor cells, perhaps by abrogating the NO-conferred cytoprotective role. These
results should pave an avenue toward the eventual application of ART in clinic
antitumor treatment.
3.3 Pro-oxidant Agents Synergize ART in Killing Tumor Cells
3.3.1 Purposes and Significance
Although ART exhibits antitumor activity at some extent, it cannot be compara-
tive to any conventional chemotherapeutics because of its relative lower cytotox-
icity. Nevertheless, ART has a potential of killing tumor cells with resistance to
common antitumor drugs. The suggested mechanisms of ART combating tumor
cells include: (1) antiproliferation and antiangiogenesis; (2) apoptosis; (3) oxida-
tive stress; (4) oncogenes and tumor suppressor genes; and (5) multidrug resist-
ance (Efferth 2006 ).
Given ART exerting a tumor-killing role via its endoperoxide, any compounds
that stabilize the unique structure should serve as synergists, whereas any com-
pounds that destabilize the unique structure might function as antagonists.
In other words, a milieu tending to oxidation should enhance ART’s antitumor
activity, whereas a milieu beneficial to antioxidation should suppress ART’s
antitumor activity. In fact, the IC 50 of ART was observed to correlate with the
expression levels of 170 genes involved in the oxidative stress response and
metabolism in 60 cell lines (Efferth and Oesch 2004 ).
To test whether oxidative stress benefits ART to exert more potent antitumor
effects, we combined ART with pro-oxidants to constitute a drug combination,
known as “ART-sensitizing compounds” (ASC), for antitumor evaluations in vitro
in the hepatoma cell line HepG 2 and in vivo in tumor-bearing nude mice. In addi-
tion to ART per se, ASC include three pro-oxidants: the GSH exhauster diethyl
maleate (DM); the GSH-POX inhibitor mercaptosuccinic acid (MA); and the CAT
inhibitor aminotriazole (AT).