The efficacy of topoisomerase (
Topo) I-active drugs may be improved by better understanding the molecular and cellular responses of
tumor compared to normal cells after genotoxic insults. Ionizing radiation (IR) +
Topo I-active drugs (e.g.,
Topotecan) caused synergistic cell killing in various human
cancer cells, even in cells from highly radioresistant
tumors.
Topo I
poisons had to be added either during or immediately after IR. Synergy was caused by DNA lesion modification mechanisms as well as by concomitant stimulation of two pathways of cell death:
necrosis (IR) + apoptosis (
Topo I
poisons). Cumulative data favor a mechanism of synergistic cell killing caused by altered DNA lesion modification and enhanced apoptosis. However, alterations in cell cycle regulation may also play a role in the synergy between these two agents in certain human
cancers. We recently showed that
NF-kappa B, a known anti-apoptotic factor, was activated in various
cancer cells after
poisoning Topo I using clinically active drugs.
NF-kappa B activation was dependent on initial nuclear DNA damage followed by cytoplasmic signaling events. Cytoplasmic signaling leading to
NF-kappa B activation after
Topo I
poisons was diminished in cytoplasts (lacking nuclei) and in CEM/C2 cells that expressed a mutant
Topo I
protein that did not interact with
Topo I-active drugs.
NF-kappa B activation was intensified in S-phase and blocked by
aphidicolin, suggesting that activation was a result of double-strand break formation due to
Topo I
poisoning and DNA replication. Dominant-negative
I kappa B expression augmented
Topo I
poison-mediated apoptosis. Elucidation of molecular signal transduction pathways after
Topo I
drug-IR combinations may lead to improved
radiotherapy by blocking anti-apoptotic
NF-kappa B responses. Recent data also indicate that synergy caused by IR +
Topo I
poisons is different from radiosensitization by
beta-lapachone (beta-lap), a "reported"
Topo I and II-alpha
poison in vitro. In fact, beta-lap does not kill cells by
poisoning either
Topo I or II-alpha in vivo. Instead, the compound is "activated" by an IR (damage)-inducible
enzyme,
NAD(P)H:
quinone oxidoreductase (NQO1), a gene cloned as x-ray-inducible transcript #3, xip3. Unlike the lesion modification pathway induced by IR +
Topo I drugs, beta-lap kills cells via NQO1 futile cycle metabolism. Downstream apoptosis caused by beta-lap appears to be noncaspase-mediated, involving
calpain or a
calpain-like
protease. Thus, although
Topo I
poisons or beta-lap in combination with IR both synergistically kill
cancer cells, the mechanisms are very different.