CLEVELANDTinkering with NFkB has little effect on the
activity of topoisomerase poisons in non-small cell-lung cancer (NSCLC), but
proteasome inhibitors can increase the efficacy of drugs such as irinotecan and
etoposide. Information on proteasome inhibitors was presented at the Vanderbilt
University Symposium by Ram Ganapathi, MD, staff scientist at the Cleveland
Clinic’s Taussig Cancer Center.
"There is a need for more information about what actually
happens after DNA damage, including questions about apoptosis and about other
mechanisms of cell death," Dr. Ganapathi said. "In a variety of tumor
cells, treatment with topoisomerase poisons produces increased DNA binding
activity of NFkB. We became interested in whether the activation of
required for producing apoptosis and cytotoxicity."
A number of possible factors influence the progression from the
DNA cleavable complex to cell death in tumor cells treated with topoisomerase
poisons. Studies were done using two human non-small-cell lung cancer (NSCLC)
cell lines. One (NSCLC-3) is a large-cell undifferentiated carcinoma from the
lower lobe of the left lung. The other (NSCLC-5) is a poorly differentiated
carcinoma with stage IV metastatic brain lesions. Dr. Ganapathi said the
cell-death pathway was explored using dominant-negative mutants for NFkB and
testing the effects of treatment with SN-38 (the active metabolite of
irinotecan), and etoposide, which is a topoisomerase II poison.
"Both etoposide and SN-38 activate NFkB," Dr.
Ganapathi said. "Paclitaxel (Taxol) and cisplatin (Platinol) do not."
Maximum activation was at 2 to 3 hours. Dr. Ganapathi pointed out that
in NSCLC, NFkB may not be required for apoptosis, since it can be induced by
drugs such as paclitaxel that do
not induce NFkB.
Role of Proteasomes
The role of proteasomes was explored using the proteasome
inhibitor MG-132. "Treatment with MG-132 followed by etoposide or SN-38
dramatically decreases upregulation of NFkB," Dr. Ganapathi said. Shutting
down NFkB inhibited apoptotic pathways in the short-term, but apoptosis had
largely resumed by 24 hours after treatment.
Ultimately, Dr. Ganapathi is trying to determine what signals
drive cells to apoptosis.
"Transfection of NSCLC cells with a dominant negative
mutant for NFkB inhibited the transcription and DNA binding activity of
usually induced by SN-38 or by etoposide, but this did not alter drug-induced
apoptosis," Dr. Ganapathi explained. "Regulation of apoptosis by
mitochondrial release of cytochrome C and activation of pro-caspase-9 followed
by cleavage of poly-ADP-ribose polymerase by effector caspase-3 and caspase-7
was similar in control cells and in the mutant cells treated with SN-38 or
In contrast to pretreatment with the proteasome inhibitor
MG-132, exposure to MG-132 following SN-38 or etoposide enhanced apoptosis
compared to drug alone," he continued (Figure 1). "We conclude from
this that apoptosis induced by topoisomerase poisons in human NSCLC is not
mediated by NFkB but can be manipulated by proteasome inhibitors."