Biomedical Engineering Reference
In-Depth Information
tumors in mice, it exhibited dose-dependent tumor-suppressing effects without
severely toxic side effects [38c]. When tumor-implanted mice were treated with
1.16 mg/kg of P61-A6 six times per week for 44 days, tumor weights were reduced
to approximately 32% of control. Moreover, the cytosolic fraction of RhoA and Rap1
was significantly higher in tumor samples from P61-A6-treated animals than in sam-
ples taken from untreated animals, suggesting that the inhibitor was reaching the
tumors. P61-A6 was found to affect neither the vasculature nor levels of apoptosis
in the tumors, and the levels of DNA synthesis measured in tumor cells suggests that
the compound exerts its effects through inhibition of cellular proliferation.
Optimization of the second initial hit, P3-E5, led to the discovery of P61-E7,
a more potent antiproliferative inhibitor than P61-A6 (e.g., 49% inhibition at 1.25
M for Panc-1 cells compared with 10% for P61-A6) [38d]. Treatment with P61-E7
increased the levels of the cyclin-dependent kinase inhibitors p21
CIP1/WAF1
and p27
Kip1
and induced a variety of morphology changes in Panc-1 cells, such as disorganiza-
tion of the actin cytoskeleton, cell rounding, inhibition of focal adhesion assembly,
and impairment of anchorage-independent growth. When a farnesyl-accepting RhoA
mutant was introduced, the effects of the P61-E7 on p21
CIP1/WAF1
and p27
Kip1
levels
and cell proliferation were partially reversed and the cell morphology changes were
minimized.
These new inhibitors of protein prenylation were derived from a DOS library
and developed in a series of studies to expand the collection of nonpeptidomimetic
inhibitors of protein prenylation. The new compounds have utility both as probes for
studying geranylgeranylation of prenylation substrates and as lead compounds for
the development of new anticancer therapies.
18.4.4 Inhibitors of Core Self-Association in Hepatitis C Virus: Probes with
a Novel Mechanism
Hepatitis C virus (HCV) poses a significant public health burden, with a 2.2 to 3.0%
global prevalence of infection [83]. The chronic liver disease caused by the virus can
develop into cirrhosis or liver cancer. The current standard of treatment is suboptimal,
being effective in only about half of patients, and side effects have resulted in the
discontinuation of treatment in 10 to 20% of patients [84]. While the HCV protease
inhibitors boceprevir and telaprevir have recently expanded the options for treatment,
there remains a need to develop new therapeutics that target different stages of the
HCV life cycle.
Penin, Lindenbach, Kota, and co-workers have used a DOS-based strategy to
discover alkaloid-like small molecules that inhibit dimerization of the HCV nucleo-
capsid protein core. HCV core is one of 10 mature proteins coded by the virus, and
dimerization of the protein is a crucial step in the HCV life cycle [85]. To model
core dimerization, a glutathione
S
-transferase (GST)-core106/FLAG-core106 system
was established using core106 (a more soluble, truncated version of the core protein)
and further developed into a time-resolved fluorescence resonance energy transfer
(TR-FRET) high-throughput assay (Figure 18.6) [51b-d,85c]. The TR-FRET assay
was initiated by mixing GST-core106 protein and FLAG-core106 protein. Next, two
