(216g) Targeted Nitric Oxide Donor Enhances Efficacy of Chemotherapeutics Used to Treat Glioblastoma Multiforme

Nitric oxide (NO) plays a
ubiquitous role in human physiology. Research has shown that at sufficient concentrations NO can induce apoptosis as well as increase chemosensitization in tumor cells. However, thus far NO
role in cancer therapy has been limited because of its short half-life and the
variety of effects it can have on numerous biological function.  Previously
we have shown that by derivatizing chlorotoxin (CTX), a protein specific to Glioblastoma multiforme (GBM)
cells, we can synthesize targeted NO donors, which deliver NO only to the
therapeutic target, thereby significantly decreasing their viability. GBM is
the most common malignant central nervous system tumors and patients diagnosed
with the disease have very poor prognoses. The two most popular
chemotherapeutics used in treatment are temozolomide
(TMZ) and carmustine (BCNU), which have both shown only modest improvements in
patient survival. In this study we investigated
the effect of the targeted NO donor, CTX-NO on the chemosensitivity
of GBM cells to the chemotherapeutics BCNU and TMZ.

To assess the
effect of NO on cell viability, T98G and U-87MG human glioblastoma
cells were incubated with varying concentration of CTX-NO, with and without
BCNU for 48 hours, trypsinized, and counted. At a NO
concentration of 2 µM, CTX-NO alone reduced glioma cell viability to 81.4 ±
5.5% of the control cell number. When cells were incubated with only BCNU,
glioma cell viability was not significantly reduced. However when cells were
incubated with a combination of CTX-NO and BCNU, the glioma cell viability was
decreased to 45.3 ± 1.5% (Fig 1A). When these cells were reseeded in fresh
media their proliferation was hindered by the prior exposure to the combination
therapy whereas the proliferation of cells previously exposed to only CTX-NO or
only BCNU was not significantly affected (Fig 1B). In contract BCNU decreased
the viability of U-87MG to 33.8 ± 2.1% and whereas the combination therapy of
BCNU and the NO donor decreased cell viability to 31.2 ± 2.7%.  Thus the NO donor does not affect the
sensitivity of U-87MG cells to BCNU.

We have shown that the targeted NO
donor, CTX-NO, can enhance
the effect of carmustine on T98G cells but not in U-87MG cells. Additionally
the combination therapy affected the proliferation ability of glioma cells even
after the therapeutic agents had been removed. To further elucidate the
mechanism of this NO induced chemosensitivity, the
affect of NO on O6-methylguanine DNA methyltransferase
activity will be examined.

Figure
1:  (A) Cell viability of T98G cells
incubated with varying concentration of CTX-NO, with and without carmustine.
Tumor cell viability is decreased dramatically when cells are exposed to a
combination of CTX-NO and carmustine. (B) Cell proliferation of cells that were
exposed to CTX-NO, with and without carmustine, for 48 hours, trysinized, incubated with fresh media for 48 hours and
then counted. The ability to proliferate was severely hindered by the
combination therapy.

Error bars
indicate standard deviation, *p <0.05