bearing animals without any deleterious effects on normal tissues. Likewise,
monoclonal antibodies that engage the human Apo2L/TRAIL receptors, DR4 and
DR5, also demonstrate potent antitumor activity without any evidence of toxicity
(Chuntharapai et al., 2001; Ichikawa et al., 2001).
An especially encouraging feature of Apo2L/TRAIL or agonistic antibodies against
death receptors for Apo2L/TRAIL is that they induce apoptosis of tumor cells while
sparing normal tissues. While TRAIL-Rl and TRAIL-R2 are broadly expressed in
most organ systems, normal cells frequently express two additional TRAIL receptors,
TRAIL-R3 (TRID or DcR1) and TRAIL-R4 (TRUNDD or DcR2) that serve as
decoys and confer protection against Apo2L/TRAIL-induced death. The expression
of decoy receptors provides a potential molecular basis for the relative resistance of
normal cells to Apo2L/TRAIL-induced death (Marsters et al., 1999). Other studies
indicate that high levels of FLIP may protect normal cells from death receptor-
induced apoptosis (Kim, K. et al., 2000). Regardless of the specific mechanism(s)
involved, the differential sensitivity of tumor cells and normal tissues to Apo2L/
TRAIL-induced cytotoxicity makes this ligand a promising investigational anticancer
agent. One potential concern was raised by the reported ability of a polyhistidine-
tagged recombinant version of human Apo2L/TRAIL (Apo2L/TRAIL.His) to induce
apoptosis in vitro in isolated human hepatocytes (Jo et al., 2000). This concern was
alleviated by subsequent studies using a Zn-bound homotrimeric version of human
Apo2L/TRAIL that lacks exogenous sequence tags (Apo2L/TRAIL.0), which has
been developed as a candidate for human clinical trials (Lawrence et al., 2001). These
studies demonstrated that Apo2L/TRAIL retains potent antitumor activity but is non-
toxic to human or nonhuman primate hepatocytes in vitro. Moreover, intravenous
administration of Apo2L/TRAIL in cynomolgus monkeys or chimpanzees was well-
tolerated with no evidence of changes in liver enzyme activities, bilirubin, serum
albumin, coagulation parameters, or liver histology (Lawrence et al., 2001).
Although Apo2L/TRAIL can induce apoptosis independently of p53 or Bcl-2,
cancer cell lines exhibit a wide heterogeneity in their sensitivity to Apo2L/TRAIL in
vitro, and certain lines are resistant to this ligand (Ashkenazi et al., 1999). As might
be appreciated from the earlier discussion of the molecular determinants and
regulators of death receptor-induced apoptosis, cancer cells may evade Apo2L/
TRAIL-mediated apoptosis by mutational inactivation of death-signaling genes or
aberrant expression of proteins that block death-signaling pathways. Bax is a frequent
target of mutational inactivation in human cancers that harbor mutations in genes
that govern DNA mismatch repair (MMR) (approximately 15% of human colon,
endometrial, and gastric carcinomas). More than 50% of MMR-deficient colon
adenocarcinomas contain somatic frame-shift mutations in an unstable tract of eight
deoxyguanosines in the third coding exon (spanning codons 38–41)[(G) 8 ] within
Bax (Rampino et al., 1997). In addition, a similar frame-shift mutation results from
loss of a G residue from a repetitive sequence in the second exon (LeBlanc et al.,
2002). MMR-deficient human colon carcinoma cells are rendered completely
resistant to Apo2L/TRAIL by inactivation of Bax (Deng et al., 2002; LeBlanc et al.,
2002; Ravi and Bedi, 2002). While both Bax+/- and Bax-/- sister clones activated apical
30 GENETICS OF APOPTOSIS