Chlorophyllin (CHL), an antimutagenic and anticarcinogenic water-soluble derivative of
chlorophyll, was recently found to be highly effective as a chemopreventive agent in a high-risk population exposed unavoidably to
aflatoxin B(1) in the diet (P. A. Egner et al., Proc. Natl. Acad. Sci. USA, 98: 14601-14606, 2001). The current study examined the response of HCT116 human
colon cancer cells to CHL treatment. Cells exposed to concentrations in the range 0.0625-0.5 mM CHL underwent growth arrest and apoptosis after 24 h, with the formation of a sub-G(1) peak in the attached cell population and nuclear condensation in the floating cell population. There was a concentration-dependent attenuation of mitochondrial membrane potential (deltapsi(m)) without the release of
cytochrome c or activation of the
caspase-9/
caspase-3/
poly(ADP-ribose) polymerase pathway. However,
apoptosis-inducing factor was released from mitochondria into the cytosol and translocated to the nucleus, leading to concentration-dependent cleavage of nuclear
lamins. The upstream mediators of this CHL-induced apoptosis pathway were identified as
caspase-8/
caspase-6 and truncated Bid, acting in conjunction with other proapoptotic members of the Bcl-2 family, such as Bak. These findings suggest that CHL might trigger apoptosis via interaction with putative "
death receptors" in the plasma membrane of
cancer cells, leading to initial cleavage of
procaspase-8 and activation of subsequent downstream events, resulting in the destruction of nuclear
lamins. Importantly,
E-cadherin and
alkaline phosphatase, which are indicators of cell differentiation, were strongly induced at all concentrations of CHL. Thus, in addition to being an effective blocking agent during the initiation phase, these findings support a role for CHL as a suppressing agent and as a possible novel therapeutic strategy directed toward aberrant cell proliferation in the colon.