| Summary: | Every year an estimated 1 million people die of malaria, with the majority of these deaths
reported in Africa. The disease caused by Plasmodium falciparum affects an approximate
250 million people annually and with the emergence of monodrug and multidrug resistance it
has seen resurgence in the last decade. The decline in effectiveness of chloroquine in the
treatment of drug resistant malaria has contributed to the doubling of malaria specific
mortality in the last fifteen years. Since the quinoline drug family represents the basis of
malaria chemotherapy for much of the past 50 years. This spread of resistance to existing
antimalarial drugs such as chloroquine, mefloquine, sulfadoxine and pyrimethamine has
driven the search for new drugs that might circumvent parasite resistance mechanisms. The
mechanism of chloroquine resistance is associated with reduced accumulation of the drug
inside the digestive vacuole, which is connected to a Plasmodium falciparum chloroquine
resistance transporter (PfCRT) or ATP-dependant P-glycoprotein efflux pump (Pgh1). The
PfCRT protein demonstrates a structural specificity for the chloroquine side chain, which
allows for changes in the structures of drugs to have different affinities for the transporter.
New drugs with structural modifications that result in reduced affinity for PfCRT may be able
to avoid reduced drug accumulation. Despite resistance, the aminoquinoline pharmacophore
remains an attractive scaffold in the design of new drugs, since it demonstrates a unique
affinity for haematin. This is a desirable feature since the quinoline antimalarial drugs inhibit
conversion of haematin to hemozoin. The 4-aminoquinoline antimalarial drugs are also weak
bases which traverse down the pH gradient to concentrate inside the acidic food vacuole.
The protonation of these drugs inside the vacuole makes them membrane impermeable and
increases their accumulation, which allows for the high concentrations required for hemozoin
inhibition.
The aim of this study was to synthesise a series of bisquinoline and
bispyrrolo[1,2a]quinoxaline compounds containing various polyamines, which may act as
potential protonation sites in the hope of increasing their accumulation via pH-trapping. In
order to achieve this aim twelve bisquinolines 4 - 15 and five bispyrrolo[1,2a]quinoxalines 16
- 20 were synthesised and their structures confirmed by nuclear magnetic resonance
spectroscopy (NMR) and mass spectroscopy (MS). The aqueous solubility (Sw) and
distribution coefficients (logD) were experimentally determined in phosphate buffered saline
(pH 5.5) to mimic the parasitic digestive vacuole environment. The compounds were
screened for antimalarial activity alongside chloroquine (CQ) against chloroquine-sensitive
(CQS) D10 and the moderately chloroquine-resistant (CQR) Dd2 strains of P. falciparum.
The series were also tested for cytotoxicity against Chinese Hamster Ovarian (CHO) cells, using emetine as reference drug. The most active compounds against P. falciparum were
screened for anticancer activity against the TK10 (renal), UACC62 (melanoma) and MCF7
(breast) cancer cells.
The bisquinoline- and bispyrrolo[1,2a]quinoxaline compounds were found to be more
hydrophilic than chloroquine (SW = 0.033 mM) itself with aqueous solubility varying in the
18.94 - 38.86 mM range. Irrespective of the series, the aqueous solubility increases with the
increase in potential protonation sites (N atoms) in the polyamine bridge. However, this
effect is overruled if the carbon-carbon chain separating two nitrogen atoms in the polyamine
also increases.
The in vitro data revealed seven of the twelve bisquinoline compounds to be significantly
more potent against the CQR (Dd2) strain compared to chloroquine. Compounds 8 (7-
chloro-4-[10-(7-chloroquinolin-4-yl)-1,4,7,10-tetraazadecan-1-yl]quinoline) (IC50 = 35.49 nM)
and 9 (7-chloro-4-[12-(7-chloroquinolin-4-yl)-1,5,8,12-tetraazadodecan-1-yl]quinoline) (IC50 =
49.48 nM) featuring the triethylenetetramine or N,N’-bis(3-aminopropyl)ethylenediamine
linkers respectively, were the most active of all synthesised compounds. They were found
significantly more potent than CQ (IC50 = 242.3 nM) against the Dd2 strain. However, they
were as potent as CQ (IC50 = 48.35 nM) against the D10 strain. This potent activity against
the CQR strain could possibly be as result of enhanced pH-trapping inside the digestive
vacuole, since they contain increased protonation sites that also enhance their hydrophilicity.
These compounds also displayed the best drug profile based on toxicity and antimalarial
activity, both demonstrating good selectivity towards parasitic cells with a selectivity index of
greater than 90.
The bis-(7-chloroquinoline)-series displayed the most potent antimalarial activity and were
subsequently screened for potential anticancer activity. The series showed potent growth
inhibitory activity against all 3 cancer cell lines. Presumably the polyamine bridges of
bisquinoline compounds provide increased ionisation of structures that allows for increased
van der Waals interactions with the highly polar phosphate backbone of the parasite DNA.
These interactions possibly interfere with cell replication and cause DNA strand scission,
since bisquinolines are known to bind by external attachment to the AT-rich sequences of
DNA, which is less stable and easier to pull apart. Compound 4 (7-chloro-N-[2-({2-[(7-
chloroquinolin-4-yl)amino]ethyl}amino)ethyl]quinolin-4-amine), 6 (7-chloro-N-[3-({3-[(7-
chloroquinolin-4-yl)amino]propyl}amino)propyl]quinolin-4-amine) and 7 (bis({3-[(7-
chloroquinolin-4-yl)amino]propyl})(methyl)amine) showed significantly more potent growth
inhibition efficacy against breast (MCF7) cancer cells compared to etoposide (TGI > 100 μM) with TGI-values in the range of 0.55 - 0.69 μM. Compounds 4, 6 and 7 were also the most
potent against TK10 (renal) and melanoma (UACC62) cancer cells with TGI-values of 0.6,
2.05 and 1 μM against TK10 cells respectively, compared to etoposide (TGI = 43.33 μM).
Against melanoma cells the TGI values were 0.59 for 4, 0.74 for 6 and 0.64 μM for 7,
compared to 4 μM for etoposide. The results reveal that a two C-C chain, and a three C-C
chain with or without methyl substitution is the optimal linker to separate the identical nonintercalating
pharmacophores for potent anticancer activity. All of the compounds in the
series warrant further investigation in search of more potent anticancer agents. === Thesis (MSc (Pharmaceutical Chemistry))--North-West University, Potchefstroom Campus, 2012
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