Potential Neuraminidase Inhibitors Via Virtual Screening, Kinetic Studies, And Adme Predictions
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Date
2015-10
Authors
Nuwarda, Rina Fajri
Journal Title
Journal ISSN
Volume Title
Publisher
Universiti Sains Malaysia
Abstract
Neuraminidase (NA) activity plays an important role in the infection by
influenza viruses by facilitating the release of the newly formed virions from the host
cell receptor and promotes its infection to other cells or organs. With the emergence
of viral resistance towards the existing NA inhibitors, the discovery of new NA
inhibitors is urgently needed. In this work, docking-based virtual screening of large
compounds from NCI database to rapidly select in silico hits to be potential NA
inhibitors was carried out. A new in vitro method has been attempted to be developed
for NA inhibition assay by utilizing AlphaScreenTM technology with Alpha-1-Acid
glycoprotein as a substrate. Unfortunately, this effort was unsuccessful probably due
to the non-specific cleavage of sialic acid by NA. Subsequently, a traditional
MUNANA assay was performed to investigate the inhibitory activities and kinetic
parameters of the inhibitor compounds. Finally, the selected compounds were studied
for their pharmacokinetic properties in silico. From the NCI database, 1541
compounds have been successfully screened and 40 in silico hits compounds were
obtained, and assayed to determine their IC50s. Ten of them demonstrated over 50%
inhibition against NA and four compounds namely NSC 5069, NSC 83318, NSC
156563, and NSC 134137 were found having IC50 values of 216 μM, 320 μM, 571 μM
and 673 μM, respectively. The kinetic studies showed Km value for MUNANA was
36.44 μM and Vmax for the enzymatic reaction was 551.25 RFU/min. From the Dixon
plot, these four compounds appeared to competitively inhibit the neuraminidase with
Ki values for NSC 5069, NSC 83318, NSC 156563, and NSC 134137 were 100.26, 204.13, 202.90, and 197.75 μM, respectively. Drug-likeness properties assessment
predicting the physicochemical properties such as molecular weight, Log P, number
of hydrogen bond donor and acceptor fulfilled Lipinski's Rule of Five. The results of
pKa value showed that the compounds are weakly acidic and tend to be absorbed in
the intestine (NSC 5069 with pKa value 4.1, NSC 83318 with pKa value 6.3 and), and
stonger acidic compound to be absorbed in the gastrik (NSC 156563 and NSC 134137
with pKa value 2.7). In silico calculations of TPSA showed that NSC 83318 and NSC
156563 fulfill Lipinski's criteria of having TPSA value less than 140 Å2, while NSC
5069 and NSC 134137 were predicted to have poor permeation since their TPSA
values are more than the acceptable upper limit of 140 Å2. Based on the fraction’s
absorbed percentage, NSC 5069 and NSC 83318 were considered to have high fraction
for the absorption. While the other two compounds, NSC 156563 and NSC 134137
had low fraction for the absorption. In predicting volume distribution, the results
showed that the volume distribution of NSC 5069, NSC 83318, NSC 156563, NSC
134137 were 0.86 L/kg, 0.7 L/kg, 0.13 L/kg and 0.12 L/kg, respectively. NSC 83318,
NSC 156563, and NSC 134137 have %PPB more than 90% which means that almost
all the compounds administered may be bound to the plasma protein in the body,
whereas NSC 5069 has %PPB 57%, indicating that 57% of the compound is bound to
the plasma proteins. Lastly, the four top compounds showed no probability as
substrates or inhibitors for almost all CYP isoforms, except for NSC 83318 which
showed relatively high probability to act on CYP 1A2 with value 0.84.
Description
Keywords
Neuraminidase