Structure-based drug design of small molecule peptide deformylase inhibitors to treat cancer

Article abstract of DOI:10.3390/molecules21040396

Human peptide deformylase (HsPDF) is an important target for anticancer drug discovery. In view of the limited HsPDF, inhibitors were reported, and high-throughput virtual screening (HTVS) studies based on HsPDF for developing new PDF inhibitors remain to be reported. We reported here on diverse small molecule inhibitors with excellent anticancer activities designed based on HTVS and molecular docking studies using the crystal structure of HsPDF. The compound M7594-0037 exhibited potent anticancer activities against HeLa, A549 and MCF-7 cell lines with IC_50s of 35.26, 29.63 and 24.63 μM, respectively. Molecular docking studies suggested that M7594-0037 and its three derivatives could interact with HsPDF by several conserved hydrogen bonds. Moreover, the pharmacokinetic and toxicity properties of M7594-0037 and its derivatives were predicted using the OSIRIS property explorer. Thus, M7594-0037 and its derivatives might represent a promising scaffold for the further development of novel anticancer drugs.

Full text of DOI:10.3390/molecules21040396

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molecules
Article
Structure-Based Drug Design of Small Molecule
Peptide Deformylase Inhibitors to Treat Cancer
Jian Gao 1,2,*, Tao Wang 1, Shengzhi Qiu 1, Yasheng Zhu 1, Li Liang 1 and Youguang Zheng 1
1
Jiangsu Key Laboratory of New Drug Research and Clinical Pharmacy, Xuzhou Medical College,
Xuzhou 221004, Jiangsu, China; lswangtao@163.com (T.W.); neoevens@hotmail.com (S.Q.);
18151865223@163.com (Y.Z.); 18852143485@163.com (L.L.); zhengyouguang@xzmc.edu.cn (Y.Z.)
Jiangsu Center for the Collaboration and Innovation of Cancer Biotherapy, Xuzhou Medical College,
2
Xuzhou 221004, Jiangsu, China
*
Correspondence: gaojian@xzmc.edu.cn; Tel.: +86-0516-8326-2137
Academic Editor: Derek J. McPhee
Received: 1 February 2016 ; Accepted: 21 March 2016 ; Published: 23 March 2016
Abstract: Human peptide deformylase (HsPDF) is an important target for anticancer drug discovery.
In view of the limited HsPDF, inhibitors were reported, and high-throughput virtual screening (HTVS)
studies based on HsPDF for developing new PDF inhibitors remain to be reported. We reported here
on diverse small molecule inhibitors with excellent anticancer activities designed based on HTVS
and molecular docking studies using the crystal structure of HsPDF. The compound M7594_0037
exhibited potent anticancer activities against HeLa, A549 and MCF-7 cell lines with IC50s of 35.26,
29.63 and 24.63 µM, respectively. Molecular docking studies suggested that M7594_0037 and its
three derivatives could interact with HsPDF by several conserved hydrogen bonds. Moreover, the
pharmacokinetic and toxicity properties of M7594_0037 and its derivatives were predicted using the
OSIRIS property explorer. Thus, M7594_0037 and its derivatives might represent a promising scaffold
for the further development of novel anticancer drugs.
Keywords:
high-throughput virtual screening; human peptide deformylase; vanillin
N-hydroxyacetamide; anticancer
1. Introduction
Peptide deformylase (PDF), a metalloenzyme containing Fe2+, is responsible for the removal of
the N-formyl group from the terminal methionine residue in nascent proteins, which is essential
to the synthesis of proteins [
microorganisms has been considered as a potential target for antibacterial drug discovery, and plenty
of PDF inhibitors have been widely reported [ ]. However, recent studies have demonstrated
that peptide deformylase also can be found in most eukaryotes, including parasites, plants, and
mammals [ 10]. Human genes of peptide deformylase (def) have been cloned and expressed,
and the resulting human peptide deformylase (HsPDF) has been studied extensively [ 11 13].
HsPDF distributes over the mitochondrion in human cells, and also functions similarly to the
PDF in pathogenic microorganisms, which catalyzes the deformylation of polypeptides [14 17].
Inhibition of HsPDF results in mitochondrial membrane depolarization and promotes cell death [13 18].
1,2]. In the past few decades, peptide deformylase of pathogenic
3–7
8
–
1
, –
–
,
HsPDF is over-expressed in breast cancer, colon cancer and lung cancer, and the inhibition of
HsPDF would significantly reduce the proliferation of these cancer cells [19]. Actinonin (Figure 1),
the first found naturally occurring bacterial PDF inhibitor, also shows strong HsPDF inhibitory
activity [11,20]. As suggested above, HsPDF is a promising novel target for the development of
anticancer drugs [12,21–25].
Molecules 2016, 21, 396; doi:10.3390/molecules21040396
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(a)
(b)
Figure 1. Structures of PDF inhibitor actinonin (a) and the shortlisted compound M7594_0037 (b).
So far, reports on inhibitors of HsPDF still remain to be conducted. The few inhibitors at present
include a class of peptide inhibitors based on actinonin reported by Mona’s research group [22] and two
series of non-peptide inhibitors reported by Christophe’s research group [23,24]. In addition, in view
of the fact that the crystal structure of HsPDF has been reported, high-throughput virtual screening
(HTVS) based on HsPDF remains to be reported. Hence, this study was intended to discover novel
small molecule inhibitors with novel structural class based on our drug design strategies involving
high-throughput virtual screening and molecular docking techniques. The resulting new structure
was named M7594_0037, and the bioactivity evaluation substantiated its superior anticancer activity.
To optimize the structure of M7594_0037, our research group designed three derivatives of vanillin
N-hydroxyacetamide. One of them, NA-2, was observed to have better anticancer activities against
MCF7 and HeLa cell lines compared to M7594_0037. Molecular docking studies also indicated that the
derivatives of vanillin N-hydroxyacetamide could have a stable interaction with HsPDF, which was
similar to the binding mode of actinonin. In summary, the derivatives of vanillin N-hydroxyacetamide
designed in this study have the potential to become a new class of anticancer drugs.
2. Results and Discussion
2.1. High-Throughput Virtual Screening
As described in the Materials and Methods section, the crystal structure (3G5K) was utilized
to screen the HsPDF inhibitors from the Topscience ChemDiv database (commercially available).
The top-ranked 100 compounds were shortlisted based on the docking score and hydrogen bond.
Finally, six commercially available compounds were selected from the top-ranked list of 100 by
clustering analysis and were tested experimentally. Among them, M7594_0037 (Figure 1) exhibited
dramatic anticancer activities against HeLa, A549 and MCF-7 human cancer cell lines, and was selected
as a lead compound for further study. The structure of M7594_0037 differed from the common peptidic
inhibitors, and it can be classified as a non-peptidic inhibitor. Non-peptidic inhibitors of HsPDF are
superior to peptidic inhibitors in terms of pharmacokinetics [25].
2.2. Design and Synthesis of Derivatives of M7594_0037
M7594_0037 consisted of the amide at its terminal, the methyl vanillin at its middle section and
the indole at its other terminal. The amide group was responsible for binding with Fe2+ as a metal
chelating group, while the known strong metal chelating groups reported in previous papers are
N-formyl-N-hydroxylamine and hydroxamic acid [25]. In other words, the amide functioning as the
metal chelating group at the terminal of M7594_0037 seemed to be a weaker one, so alteration of this
group will be focused on in this article. The terminal indole ring of M7594_0037 interacted with the
hydrophobic S3 region (described in the section on molecular docking studies) where it can provide
additional binding energy, selectivity, and favorable pharmacokinetic and toxicity properties [25].
Moreover, the S3 regions of the binding sites of HsPDF are generally solvent accessible, and especially

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Molecules 2016, 21, 396
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large in size to accommodate various substituent groups [21]. For the purpose of improving the
anticancer activity, three derivatives of M7594_0037, which were named 3AP-2
designed. The structures of all the derivatives were characterized by H-NMR and mass spectra.
The synthetic routes are shown in Scheme 1.
,
NA-2
,
M-2, were
1
Scheme 1. Synthesis of M7594_0037 derivatives.
2.3. Anticancer Activity of M7594_0037 and Its Derivatives
As we know that HsPDF has been reported to be over-expressed in many cancers, it was proven
that HsPDF inhibitors could specifically be used in cancer treatment. Hence, the anticancer activities
of M7594_0037 and its derivatives were studied on HeLa, A549 and MCF-7 human cancer cell lines
in a dose-dependent manner (Figure 2). The cytotoxic activity was expressed as the mean IC50 of
three independent experiments, and the results were represented in Table 1. It is clearly shown that
M7594_0037 had inhibitory activity against HeLa, A549 and MCF-7, whose IC50 values are 35.26
29.49 2.09 and 24.63 2.19 M, respectively. Its inhibitory activity against HeLa was slightly smaller
than that of actinonin [22] (IC50 = 27.40 M). For further optimization of M7594_0037, the amide at
˘
3.17,
˘
˘
µ
µ
its terminal was replaced with N-formyl-N-hydroxylamine to give three derivatives. Each of them
exhibited obvious anticancer activities. The compound 3AP-2 showed better inhibitory activity against
MCF-7 than M7594_0037 (IC50 = 13.62
inhibition against both HeLa and MCF-7 cell lines (IC50 = 9.60
Thus, M7594_0037 and its derivatives have potential to become lead compounds of a class of novel
˘
3.42
µ
M), while NA-2 was even superior to M7594_0037 in
˘
3.04 and 22.89 3.41 M, respectively).
˘
µ
anticancer drugs.
(a)
Figure 2. Cont.

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(b)
(c)
Figure 2. Inhibition rate of M7594_0037 and analogs against A549 (a); MCF7 (b) and HeLa (c) in
dose-dependence.
Table 1. Effect (IC50) of M7594_0037 and analogs on human cancer cell lines.
Compounds
A549 (µM)
MCF7 (µM)
HeLa (µM)
M7594_0037
3AP-2
29.49 ˘ 2.09
45.37 ˘ 2.08
67.85 ˘ 2.22
119.95 ˘ 3.71
24.63 ˘ 2.19
13.62 ˘ 3.42
22.89 ˘3.41
48.02 ˘ 2.32
35.26 ˘ 3.17
96.57 ˘ 3.73
9.60 ˘ 3.04
36.39 ˘ 3.90
NA-2
M-2
2.4. Binding Modes of M7594_0037 and Its Derivatives
M7594_0037 and its derivatives were docked into the binding site where the ligand actinonin was
placed by the Surflex module in SYBYL 7.1. The binding modes of M7594_0037 and its derivatives with
HsPDF were obtained from the results of Surflex molecular docking. As is vividly shown in Figure 3,
many hydrogen bonds were formed between M7594_0037 and HsPDF, and most of them were allowed
by the amide group at the terminal of M7594_0037. Three hydrogen bonds were networked with
side-chains of Gln-57 and Glu-157, and another three hydrogen bonds were networked with backbones
of Gly-52, Glu-115. The methoxyl group at the vanillin section allowed two hydrogen bonds networked
with Gly-52 and Val-51 as well, and the NH group on the indole ring allowed the hydrogen bond
networked with Glu-76 to be formed. These hydrogen bonds played an important role in the binding
interaction between M7594_0037 and HsPDF. The terminal indole ring was located in the hydrophobic
pocket constituted by Trp-149, Leu-73, Met-87, Arg-85, Cys-77. Analogous to the binding mode of
M7594_0037
and its terminal morpholine was also placed in a similar hydrophobic pocket. In the interaction
between stabilized hydrogen bonds and M7594_0037 NA-2 coincided with the conservative hydrogen
bond interactions which has been reported in the past [21]. For example, actinonin would emerge
, NA-2 also formed stabilized hydrogen bonds with Gln-57, Gly-52, Glu-115, and Glu-157,
,