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Total Syntheses and Preliminary Biological Evaluation of Brominated Fascaplysin and Reticulatine Alkaloids and Their Analogues

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A simple approach toward the synthesis of the marine sponge derived pigment fascaplysin was used to obtain the marine alkaloids 3-bromofascaplysin and 3,10-dibromofascaplysin. These compounds were used for first syntheses of the alkaloids 14-bromoreticulatate and 14-bromoreticulatine. Preliminary bioassays showed that 14-bromoreticulatine has a selective antibiotic (to Pseudomonas aeruginosa) activity and reveals cytotoxicity toward human melanoma, colon, and prostate cancer cells. 3,10-Dibromofascaplysin was able to target metabolic activity of the prostate cancer cells, without disrupting cell membrane’s integrity and had a wide therapeutic window amongst the fascaplysin alkaloids.
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Mar. Drugs 2019, 17, 496; doi:10.3390/md17090496 www.mdpi.com/journal/marinedrugs
Communication
Total Syntheses and Preliminary Biological
Evaluation of Brominated Fascaplysin and
Reticulatine Alkaloids and Their Analogues
Maxim E. Zhidkov 1,*, Polina A. Smirnova 1, Oleg A. Tryapkin 1, Alexey V. Kantemirov 1, Yuliya
V. Khudyakova 2, Olesya S. Malyarenko 2, Svetlana P. Ermakova 2, Valeria P. Grigorchuk 3,
Moritz Kaune 4, Gunhild von Amsberg 4,5 and Sergey A. Dyshlovoy 1,2,4,5
1 Department of organic chemistry and Laboratory of biologically active compounds, School of Natural
Sciences, Far Eastern Federal University, 8 Sukhanov Str., Vladivostok 690950, Russia
2 G.B. Elyakov Pacific Institute of Bioorganic Chemistry, 159 Prospekt 100 Let Vladivostoku, Vladivostok
690022, Russia
3 Federal Scientific Center of the East Asia Terrestrial Biodiversity (Institute of Biology and Soil Science), Far
Eastern Branch of the Russian Academy of Sciences, 159 Prospect 100-Let Vladivostoku, Vladivostok
690022, Russia
4 Department of Oncology, Hematology and Bone Marrow Transplantation with Section Pneumology,
Hubertus Wald-Tumorzentrum, University Medical Center Hamburg-Eppendorf, 20246 Hamburg,
Germany
5 Martini-Klinik Prostate Cancer Center, University Hospital Hamburg-Eppendorf, 20246 Hamburg,
Germany
* Correspondence: mzhidkov@rambler.ru; Tel.: +7-423-265-24-24 (423)
Received: 13 July 2019; Accepted: 22 August 2019; Published: 25 August 2019
Abstract: A simple approach toward the synthesis of the marine sponge derived pigment
fascaplysin was used to obtain the marine alkaloids 3-bromofascaplysin and 3,10-
dibromofascaplysin. These compounds were used for first syntheses of the alkaloids 14-
bromoreticulatate and 14-bromoreticulatine. Preliminary bioassays showed that 14-
bromoreticulatine has a selective antibiotic (to Pseudomonas aeruginosa) activity and reveals
cytotoxicity toward human melanoma, colon, and prostate cancer cells. 3,10-Dibromofascaplysin
was able to target metabolic activity of the prostate cancer cells, without disrupting cell membrane’s
integrity and had a wide therapeutic window amongst the fascaplysin alkaloids.
Keywords: total synthesis; 14-bromoreticulatine; 3,10-dibromofascaplysin; bioactivity
1. Introduction
Fascaplysin, homofascaplysins A–C, and their brominated analogues form the group of marine
alkaloids based on the 12H-pyrido [1–2-a:3,4-b’] diindole ring system [1]. The red pigment fascaplysin
(1, Figure 1) is the first isolated compound among these alkaloids and at the present time is the most
investigated one [2]. This compound could be used in the field of medicinal chemistry due to a broad
range of bioactivities including antibacterial, antifungal, antiviral, and antimalarial properties. In
addition, it is able to inhibit the proliferation of numerous cancer cell lines and reveals anti-
angiogenesis properties on human umbilical vein endothelial cells (HUVEC) [3–9]. Fascaplysin
suppresses the growth of S180 cell-implanted tumors in vivo [10]. Remarkably, it effectively decreases
the growth of small cell lung cancer (SCLC) spheroids derived from circulating tumor cells. In fact,
high numbers of circulating tumor cells are linked to the dismal prognosis of SCLC [11]. Its
mechanisms of action include the selective inhibition of cyclin-dependent kinase 4, which regulates
Mar. Drugs 2019, 17, 496 2 of 12
the G0–G1/S checkpoint of the cell cycle, the intercalation of DNA, and the induction of apoptosis,
partially, as a result of the activation of the TRAIL signaling pathway by the upregulation of DR5
expression [12–15]. It was also found that fascaplysin induced autophagy as a cytoprotective response
via ROS and p8 in vascular endothelial cells (VECs) [16]. A cooperative interaction between apoptotic
and autophagic pathways is exhibited by fascaplysin through the inhibition of PI3K/AKT/mTOR
signaling cascade in human leukemia HL-60 cells [17]. It also causes the downregulation of survivin
and HIF-1α and inhibition of VEGFR2 and TRKA, and sensitizes anti-cancer effects of drugs targeting
AKT and AMPK [18,19]. Fascaplysin could be used as a P-gp inducer for the development of anti-
Alzheimer agents [20]. It may also serve as a “balanced agonist” of the µ-opioid receptor with a
signaling profile that resembles endorphins [21].
Cl Cl
N
H
N+
O
N+
R
N
H
6: X1 = H, X2 = H, R = COOMe
7: X1 = H, X2 = Br, R = COOMe
8: X1 = Br, X2 = Br, R = COOMe
9: X1 = X2 = H, R = COO-
10: X1 =H, X2 = Br, R = COO-
11: X1 = X2 = H, R = OH
12: X1 = H, X2 = Br, R = OH
14
7
3
10
1: X1 =X2 = H
2: X1 = H, X2 = Cl
3: X1 = H, X2 = Br
4: X1 = Br, X2= H
5: X1 = X2 = Br
X1 X1
X2 X2
Figure 1. Structures of fascaplysin (1) and its derivatives (25); reticulatine (6), 14-bromoreticulatine
(7), 7,14-dibromoreticulatine (8), reticulatate (9), 14-bromoreticulatate (10), reticulatol (11), 14–
bromoreticulatol (12).
Remarkably, some derivatives of fascaplysin were found to have an increased therapeutic
potential compared to the parental alkaloid. Thus, methylation of fascaplysin at C-9 results in the
more potent Aβ aggregation inhibitor than alkaloid 1 [22]. The synthetic chloro derivative of
fascaplysin (2) inhibited the VEGF-mediated microvessel sprouting with blood vessel formation in
the matrigel plug of C57/BL6J mice and the tumor growth in ET (solid) mouse tumor model [23]. In
addition, natural 3- and 10-bromofascaplysins (3,4) showed anti-cancer activity at submicromolar
concentrations. This was, at least in part, mediated through the induction of caspase-8, -9, and -3-
dependent apoptosis [24]. Antitumor effects of 3-bromofascaplysin and 10-bromofascaplysin were
comprehensively examined in an in vitro glioma C6 cell model. The cytotoxic efficiency of
compounds 3 and 4 was higher than that of unsubstituted fascaplysin; 3-bromofascaplysin exhibited
the best capacity to kill glioma C6 cells [25]. 3,10-Dibromofascaplysin (5)—the last representative of
fascaplysin alkaloids was synthesized in eight steps from 6-bromoindole and 4-amino-2-
bromotoluene, but the therapeutic potential of that perspective compound has not been investigated
yet [26].
Herein, we report the two-step method for the syntheses of 3-bromofascaplysin and 3,10-
dibromofascaplysin, which has been previously used for the synthesis of fascaplysin. The similarity
in structures lets us to use these compounds as starting materials for the first syntheses of several
alkaloids of reticulatine group (compounds 612, Figure 1, [27]). Also, the bioactivities of the obtained
compounds were investigated.
Mar. Drugs 2019, 17, 496 3 of 12
2. Results
2.1. Chemistry
Several groups have synthesized fascaplysin and its naturally occurring analogs and more than
10 syntheses have been reported to date [28–38]. Among them the two-step scheme by Zhu et al. is
the most suitable for the preparation of the target compounds [36]. To apply that synthetic scheme
for the synthesis of 3,10-dibromofascaplysin, the reaction between 3-bromophenylhydrazine (13) and
4-bromobutanal (14) in an autoclave at 150 °C was used to prepare the mixture of 6-bromotryptamine
(15) and 4-bromotryptamine (16) (Scheme 1). Thereafter, the obtained mixture and 2,4-
dibromoacetophenone (17) were subjected to the cascade coupling protocol, previously developed
by Zhu et al., which included the sequential iodination of the corresponding acetophenone, the
Kornblum oxidation of the intermediate in the presence of DMSO to phenylglyoxal, and its Pictet–
Spengler condensation with the derivative of tryptamine followed by the oxidation of the
intermediate. After chromatography purification, two isomeric 1-benzoyl-β-carbolines (18, 19) were
obtained with the yields of 20% and 19%, respectively. These products were subsequently
transformed to 3,10-dibromofascaplysin (5) and its isomer 20 according to the procedure reported by
the group of Radchenko [31].
Br
NH
NH2
Br N
H
N
Br
Br
O
N
H
N
Br
Br
O
Br
Br N
H
N+
Br
O
N
H
N+
Br
O
Br
NH2
N
H
Br
NH2
Br
N
H
20%
19%
91%
93%
ab
16
13 15
18
19
c
5
20
57%
c
+
Cl-
Cl-
Scheme 1. Reagents and conditions. (a) 4-bromobutanal (14, 4.0 equiv.), EtOH, H2O, autoclave, 150
°C, 1 h; (b) 2,4-dibromoacetophenone (17) (1 equiv.), I2 (0.8 equiv.), DMSO, 110 °C, 1 h, then
tryptamines 15, 16 (1.0 equiv.), DMSO, 110 °C, 4 h; (c) 220 °C, 15 min, then HCl (aq).
3-Bromofascaplysin was prepared in a similar manner from tryptamine (21) and 2,4-
dibromoacetophenone (17) with a total yield of 32%. Taking into account the high biological activity
of synthetic chloro derivatives of fascaplysin, we obtained the corresponding derivative at C-2 (25)
from tryptamine and 2,5-dichloroacetophenone (22) by a similar method (Scheme 2) [20].
Mar. Drugs 2019, 17, 496 4 of 12
N
H
NH2
N
H
N+
O
N
H
N
O
CH3
O
ab
3: X1 = H, X2 = Br, 84%
25: X1 = Cl, X2 = H, 70%
Cl-
21
+
X1
X2
X3
17, 23: X1=H, X2=Br, X3=Br
22, 24: X1=Cl, X2=H, X3=Cl
17, 22
X1
X2
X3
23, 38%
24, 44%
X1
X2
Scheme 2. Reagents and conditions. (a) I2 (0.8 equiv.), DMSO, 110 °C, 1 h, then tryptamine (1.0 equiv.),
DMSO, 110 °C, 4 h; (b) 220 °C, 15 min, then HCl (aq).
Previously zwitter-ionic β-carboline 26 was obtained from fascaplysin that was treated with
aqueous solution of NaOH or 30% NH4OH [39]. After optimization of the reaction conditions 14-
bromoreticulatate (10) and its dibromo analog (27, not isolated from marine organisms) were
obtained from compounds 3 and 5 in DMF at r.t. with 86% and 80% yields, respectively (Scheme 3).
Different conditions for methylation of compounds 10 and 27 were investigated, including (i) the
interaction with diazomethane; (ii) with POCl3 and following treatment with methanol; (iii) the
reaction with dimethyl sulfate. In the latter case, best results were achieved. However, 7,14-
dibromoreticulatine (8) was not obtained after methylation of compound 27. Instead, the product of
dimethylation (28) was obtained. Because of the insolubility of compound 28 in most solvents, only
MS and 1H NMR were used to identify its structure. The spectral characteristics of synthetic 3-
bromofascaplysin, 3,10-dibromofascaplysin, 14-bromoreticulatate, and 14-bromoreticulatine were
identical to those of the natural products.
Cl
Cl
Cl
O
N+
N
H
N+
OCH3
O
N
H
Br
N+
O
O
N
H
N
N+
OCH3
O
CH3
Br
Br
14
14
b
7
14
7
3
10
1: X1 = X2 = H,
3: X1 = H, X2 = Br,
5: X1 = X2 = Br
26: X1 = X2 = H, 54% from [39]
10: X1 = H, X2 = Br, 86%
27: X1 = X2 = Br, 80%
a
b
for 10
for 27
X1
X2 X2
X1
7, 52%
for 3, 5
28, 48%
Scheme 3. Reagents and conditions–(a) NaOH (4 equiv.), DMF, r.t., 0.5 h; (b) (CH3)2SO4 (4 equiv.),
CH3CN, 1 h.
2.2. Biology
The bioactivities of obtained compounds were investigated using fascaplysin (1) as a standard.
First, the cytotoxic effects of the compounds against human colorectal carcinoma (HT-29), human
breast cancer (T-47D), and melanoma (SK-MEL-28) cell lines were evaluated by MTS assay (Table 1).
The cells were incubated with different concentrations of the respective compounds (0–5 µM) for 24
h. The concentration that caused inhibition of 50% of cell viability (IC50) was 5 µM for compound 1
against T-47D cells. Other investigated compounds were less cytotoxic against this type of cancer
cells at concentrations up to 5 µM. However, the IC50 of 1, 3, and 7 were detected at concentrations
ranging from 1.1 to 1.9 µM against SK-MEL-28 cells. Among the investigated cancer cells, the most
Mar. Drugs 2019, 17, 496 5 of 12
resistant cell line to the cytotoxic effect of the compounds was found to be breast cancer cells T-47D,
while the most sensitive were melanoma cells SK-MEL-28. It was shown that compounds 1 and 3
possessed comparable IC50 against colorectal carcinoma cells HT-29. Our results indicated that the
investigated compounds reveal selective cytotoxic effects to different cancer cell lines, with highest
efficacy in melanoma cells SK-MEL-28.
Table 1. Cytotoxic activities of fascaplysin and its derivatives. Values are indicated as mean ±
standard deviation.
Compound
Inhibiting Concentration, (IC50), µM IC50
(ViCell)/IC50
(MTT) for
22Rv1 Cells
HT-29 a T-47D
a SK-MEL-28
a PC-3
b 22Rv1
b 22Rv1
c
Fascaplysin (1) 2.7 ± 0.05 5 ± 0.2 1.3 ± 0.08 0.78 ± 0.16 0.24 ± 0.04 0.34 ± 0.11 1.39
3-Bromofascaplysin (3) 3.3 ± 0.12 >5 1.9 ± 0.04 10 ± 1.75 0.42 ± 0.29 0.24 ± 0.14 0.58
Compound 20 >5 >5 >5 1.39 ± 0.43 0.21 ± 0.04 0.26 ± 0.05 1.24
Compound 25 >5 >5 1.8±0.02 0.91 ± 0.06 0.27 ± 0.01 0.5 ± 0.19 1.88
14-Bromoreticulatate
(10) >5 >5 >5 n/d n/d n/d n/d
14-Bromoreticulatine
(7) >5 >5 1.2 ± 0.1 > 50 35.72 ± 10.1 n/d n/d
3,10-
Dibromofascaplysin
(5)
>5 >5 >5 7.28 ± 0.73 0.69 ± 0.05 5.14 ± 1.16 7.45
IC50, the concentration of compounds that caused a 50% reduction in cell viability of tested normal
and cancer cells; a MTS assay was used; b MTT assay was used; c ViCell assay (trypan blue exclusion)
was used, n/d—not determined.
We have also investigated the effect of the synthesized compounds on the viability and the
growth of human prostate cancer drug-resistant PC-3 and 22Rv1 cells. IC50s of the substances have
been determined by both, MTT and trypan blue exclusion assay (ViCell assay) (Table 1, Figure 2). It
is known that MTT assay accesses the metabolic activity of the cells, while the trypan blue exclusion
assay shows the alive cells with either intact (non-stained) or disrupted (stained) membranes.
Compound 20 was identified to be the most active among the tested fascaplysin derivatives.
However, its cytotoxicity determined by MTT assay was within the range of compounds 3 and 25
and fascaplysin (1). Interestingly, compound 5, having a higher IC50 of 0.69 ± 0.14 µM, had a very
smooth cytotoxicity profile, suggesting a wide therapeutic window (Figure 2). Moreover, for
compound 5, the IC50 determined by trypan blue exclusion assay was ~8-fold higher than the IC50
accessed using MTT test. In contrast, for the other compounds the difference of the IC50s generated
by the two different methods was distinctly less pronounced. This may indicate an antimetabolic
effect of compound 5 rather an effect on the cell membrane integrity (necrotic-like cell death).
Compound 5 starts to suppress cancer cell viability/proliferation already at 0.1 µM, while the ranges
of active concentrations for the other two tested compounds were rather narrow. Fascaplysin (1)
started to suppress cancer cell viability/proliferation at 0.125 µM. Remarkably, for this compound no
difference between IC50s generated with the two different methods was observed. The high potential
of compound 5 for therapeutic assays was also confirmed by its low cytotoxity (IC50 50 µM) against
normal MRC-9 lung cells.
Mar. Drugs 2019, 17, 496 6 of 12
Figure 2. Effect of the compounds on viability of 22Rv1 cells. The effect was accessed using MTT
assay. Cells were treated with the compounds for 48 h. The values are presented as mean expression
levels ± SD are shown.
It is known that fascaplysins exhibit potent but nonselective antibiotic activities. To evaluate
activity of reticulatines in comparison to known fascaplysin derivatives, compounds 1, 3, 7, 25 were
studied in vitro for antibiotic activity against several microbes using the disk diffusion soft agar assay
as shown in Table 2. 14-Bromoreticulatine (7) showed potent activity against Pseudomonas aeruginosa
while it exhibited low activity or no activity at all against other tested microbes. As expected, high
and non-selective antibiotic activities were demonstrated for the other tested compounds (1, 3, 25).
Table 2. Antibiotic activity of compounds 1, 3, 25, and 7.
Compound Conc.,
mg/disc
Zone Unit Differentials in the Disk Diffusion Soft Agar Assay a
Bacillus
subtilis
(KMM 430)
Staphylococcus
aureus (ATCC
21027)
Pseudomonas
aeruginosa
(KMM 433)
Escherichia coli
(ATCC 15034)
Candida
albicans
(KMM 455)
1 0.4 25 25 >35 20 n/a
3 0.1 25 20 >35 20 *
25 0.2 20 20 >35 20 n/a
7 0.2 10 n/a >35 10 n/a
a Measured in mm; * fungistatic effect; n/a, not active.
3. Materials and Methods
3.1. Chemistry
All starting materials are commercially available. Commercial reagents were used without any
purification. The products were isolated by MPLC: Buchi B-688 pump, glass column C-690 (15 × 460
mm) with Silica gel (particle size 0.015–0.040 mm), UV-detector Knauer K-2001. The analytical
examples were purified by Shimadzu HPLC system (model: LC-20AP) equipped with a RID detector
(model: RID 10A) using Supelco C18 (5 µm, 4.6 × 250 mm) column using ACN:water (20:80, 50:50,
70:30) mobile phase by isocratic elution at flow rate of 1 mL/min. NMR spectra were recorded with a
NMR instrument operating at 400 MHz (1H) and 100 MHz (13C). Proton spectra were referenced to
TMS as internal standard, in some cases, to the residual signal of used solvents. Carbon chemical
shifts were determined relative to the 13C signal of TMS or used solvents. Chemical shifts are given
on the δ scale (ppm). Coupling constants (J) are given in Hz. Multiplicities are indicated as follows: s
(singlet), d (doublet), t (triplet), q (quartet), m (multiplet), or br (broadened). The original spectra of
the relative compounds could be found in Supplementary Materials. High-resolution mass spectra
(HRMS) were obtained with a time-of-flight (TOF) mass spectrometer equipped with an electrospray
source at atmospheric pressure ionization (ESI).
3.1.1. Preparation of Mixture of Tryptamines 15 and 16
A mixture of 4-bromobutanal (1.33 g, 8.8 mmol), 3–bromophenylhydrazine hydrochloride (0.50
g, 2.2 mmol), EtOH (3 mL), and H2O (1 mL) was placed into an autoclave and heated at 150 °C for 1
h. After cooling, the mixture was poured into H2O (100 mL) and extracted with EtOAc (3 × 50 mL).
Then aqueous solution was treated with NaOH to pH 12 and extracted with CH2Cl2 (3 × 50 mL). The
combined organic layer was washed with brine (2 × 100 mL), dried over Na2SO4, and evaporated.
After flash column chromatography (EtOAc, then EtOH/NH3), compounds 15 and 16 were isolated
as a mixture in ratio of 1:1 (brown oil, 300 mg, 57%).
3.1.2. Preparation of Substituted 1-Benzoyl-β-Carbolines 18, 19, 23, 24
Corresponding acetophenone (0.458 mmol) and iodine (92 mg, 0.366 mmol) were added to 2 mL
of DMSO, and the resulting solution was heated at 90 °C for 1 h. After that tryptamine, its derivative
or their mixture (0.458 mmol) was added to the solution and this solution was stirred at the same
Mar. Drugs 2019, 17, 496 7 of 12
temperature for 3–4 h till completion of reaction (monitored by TLC). Then the reaction mixture was
cooled to room temperature followed by the addition of water (50 mL) and extraction with EtOAc (2
× 25 mL). The extract was washed with 10% Na2S2O3, dried over Na2SO4, filtered and evaporated
under reduced pressure. The residue was purified by MPLC using benzene and benzene/hexanes as
eluent to give the desired product.
For 1-(2,4-dibromobenzoyl)-7-bromo-β-carboline (18): yellow solid, 20%. 1H-NMR (400 MHz, CDCl3):
δ 10.45 (br. s, 1H), 8.57 (d, J = 4.9 Hz, 1H), 8.15 (d, J = 4.9 Hz, 1H), 8.05 (d, J = 8.3 Hz, 1H), 7.88 (d, J =
1.7 Hz, 1H), 7.79 (d, J = 1.1 Hz, 1H), 7.62 (dd, J = 8.3, 1.7 Hz, 1H), 7.50 (dd, J = 8.3, 1.5 Hz, 1H), 7.46 (d,
J = 8.2 Hz, 1H). 13C-NMR (100 MHz, CDCl3): δ 196.7, 141.4, 138.9, 138.4, 136.5, 135.2, 134.7, 130.9, 130.6,
129.8, 124.4, 124.2, 122.9, 122.6, 120.5, 119.2, 118.9, 114.8. HRMS-ESI, m/z: [M + H]+ calculated for
C18H1079Br3N2O+: 506.8340, found 506.8345.
For 1-(2,4-dibromobenzoyl)-5-bromo-β-carboline (19): yellow solid, 19%. 1H-NMR (400 MHz, CDCl3):
δ 10.55 (br. s, 1H), 8.81 (d, J = 5.1 Hz, 1H), 8.62 (d, J = 5.1 Hz, 1H), 7.88 (d, J = 1.8 Hz, 1H), 7.63 (dd, J =
3.2, 1.4 Hz, 1H), 7.61 (dd, J = 2.9, 1.4 Hz, 1H), 7.54–7.58 (m, 1H), 7.51 (d, J = 7.80 Hz, 1H), 7.46 (d, J =
8.2 Hz, 1H). 13C-NMR (100 MHz, CDCl3): δ 197.5, 142.4, 142.4, 139.4, 139.1, 137.0, 135.9, 135.1, 131.6,
131.3, 130.4, 130.2, 125.4, 125.1, 121.5, 121.2, 118.5, 111.3. HRMS-ESI, m/z: [M + H]+ calculated for
C18H1079Br3N2O+ 506.8340, found 506.8347.
For 1-(2,4-dibromobenzoyl)-β-carboline (23): yellow solid, 38%. 1H-NMR (400 MHz, DMSO-d6): δ
12.23 (br. s, 1H, NH), 8.48 (d, J = 4.9, 1H, H-3), 8.44 (d, J = 4.9, 1H, H-4), 8.34 (d, J = 7.9, 1H, H-5), 8.02
(d, J = 1.9, 1H, H-3’), 7.85 (d, J = 8.0, 1H, H-8), 7.76 (dd, J = 8.3, 1.9, 1H, H-5’), 7.64 (ddd, J = 7.2, 7.2, 1.0,
1H, H-7), 7.57 (d, J = 8.3, 1H, H-6’), 7.35 (ddd, J = 7.2, 7.2, 1.0, 1H, H-6). 13C-NMR (100 MHz, DMSO-
d6): δ 195.9, 142.0, 140.5, 137.9, 135.3, 134.8, 134.3, 131.4, 131.0, 130.3, 129.2, 128.3, 123.2, 121.9, 120.5,
120.0, 119.8, 113.1. HRMS-ESI, m/z: [M + H]+ calculated for C18H1179Br2N2O+ 428.9235, found 428.9239.
For 1-(2,5-dichlorobenzoyl)-β-carboline (24): yellow solid, 44%. 1H-NMR (400 MHz, CDCl3): δ 10.38
(br. s, 1H), 8.57 (d, J = 4.9 Hz, 1H), 8.19 (m, 2H), 7.65 (m, 2H), 7.57 (t, J = 1.4 Hz, 1H), 7.43 (d, J = 1.1 Hz,
2H), 7.39 (m, 1H). 13C-NMR (100 MHz, CDCl3): δ 196.1, 141.2, 139.6, 139.0, 136.8, 134.9, 132.5, 132.0,
131.1, 131.1, 130.1, 129.7, 129.6, 122.0, 121.2, 120.7, 119.5, 112.2. HRMS-ESI, m/z: [M + H]+ calculated
for C18H1135Cl2N2O+ 341.0247, found 341.0242.
3.1.3. Preparation of Fascaplysin Derivatives
Substituted 1-benzoyl-β-carboline (0.326 mmol) was heated in sealed tube at 220 °C for 15 min.
After cooling, the reaction mixture was washed with EtOAc (3 × 3 mL) and H2O (3 × 10 mL). The
combined aqueous layer was acidified with hydrochloric acid and evaporated under reduced
pressure to give target product as a red powder.
For 3,10-dibromofascaplysin (5): red solid, 91%. 1H NMR (400 MHz, MeOH-d4): δ 9.38 (d, J = 6.4, 1H,
H-6), 8.97 (d, J = 6.4, 1H, H-7), 8.69 (bs, 1H, H-4), 8,41 (d, J = 8.8, 1H, H-8), 8.05 (d, J = 1.4, 1H, H-11),
7.97 (d, J = 0.8 × 2, 2H, H-1, H-2), 7.71 (dd, J = 8.6, 1.7, 1H, H-9). 13C-NMR (100 MHz, MeOH-d4): δ
180.2, 147.7, 147.6, 140.8, 134.0, 131.4, 130.7, 128.7, 126.6, 126.4, 126.0, 124.8, 122.7, 119.5, 119.5, 118.7,
118.4, 115.9. 13C-NMR (100 MHz, DMSO-d6): δ 181.3, 148.0, 147.8, 140.2, 134.4, 131.2, 130.5, 128.2,
127.7, 127.1, 126.6, 126.1, 123.5, 123.3, 120.8, 119.6, 118.6, 116.4. HRMS-ESI, m/z: [M]+ calculated for
C18H979Br2N2O+ 426.9079, found 426.9085.
For compound 20: red solid, 93%. 1H-NMR (400 MHz, MeOH-d4): δ 9.44 (d, J = 4.7 Hz, 1 H), 9.36 (d,
J = 4.5 Hz, 1 H), 8.76 (s, 1 H), 7.98 (s, 2 H), 7.76–7.88 (m, 3 H). 13C-NMR (100 MHz, MeOH-d4): δ 180.1,
148.1, 147.5, 140.1, 134.5, 134.2, 131.4, 130.7, 126.8, 126.0, 122.7, 122.4, 120.3, 119.5, 118.9, 118.9, 118.5,
112.2. HRMS-ESI, m/z: [M]+ calculated for C18H979Br2N2O+ 426.9079, found 426.9083.
For 3-bromofascaplysin (3): red solid, 84%. 1H-NMR (400 MHz, MeOH-d4): δ 9.35 (d, J = 6.2, 1H, H-
6), 8.95 (d, J = 6.2, 1H, H-7), 8.68 (s, 1H, H-4), 8.48 (d, J = 8.1, 1H, H-8), 7.93 (s, 2H, H-1, H-2), 7.88 (t, J
= 7.6, 1H, H-10), 7.79 (d, J = 8.1, 1H, H-11), 7.52 (t, J = 7.6, 1H, H-9). 13C-NMR (100 MHz, MeOH-d4): δ
Mar. Drugs 2019, 17, 496 8 of 12
182.0, 149.4, 148.9, 143.1, 136.0, 135.6, 132.3, 132.2, 127.7, 127.6, 125.1, 124.5, 124.5, 123.8, 121.1, 120.9,
120.3, 114.5. HRMS-ESI, m/z: [M]+ calculated for C18H1079BrN2O+ 348.9974, found 348.9980.
For compound 25: red solid, 70%. 1H-NMR (400 MHz, MeOH-d4): δ 9.36 (d, J = 5.8, 1H), 8.96 (d, J =
6.0, 1H), 8.49 (d, J = 8.0, 1H), 8.35 (d, J = 8.6, 1H), 8.05 (d, J = 1.2, 1H), 7.98 (d, J = 7.5, 1H), 7.90 (d, J =
6.8, 1H), 7.82 (m, 1H), 7.55 (t, J = 7.6, 1H). 13C-NMR (100 MHz, MeOH-d4): δ 180.3, 147.2, 145.3, 136.7,
135.8, 134.3, 127.6, 126.5, 126.2, 126.1, 125.8, 124.9, 123.6, 123.0, 121.9, 119.5, 116.3, 112.9. HRMS-ESI,
m/z: [M]+ calculated for C18H1035ClN2O+ 305.0480, found 305.0486.
3.1.4. Preparation of Compounds 10, 27
A solution of compound 3 or 5 (0.15 mmol) in DMF (10 mL) was treated with solution of NaOH
(24 mg, 0.6 mmol) in 0.1 mL of H2O at room temperature for 0.5 h. The mixture was neutralized with
AcOH and evaporated under reduced pressure. The residue was washed with Et2O and dried.
For 14-bromoreticulatate (10): yellow solid, 86%. 1H-NMR (400 MHz, MeOH-d4): δ 9.37 (s, 1H), 8.75
(d, J = 6.3, 1H), 8.59 (d, J = 6.5, 1H), 8.47 (d, J = 8.1, 1H), 8.17 (d, J = 8.4, 1H), 8.06 (s, 1H), 8.00 (d, J = 8.4,
1H), 7.83 (m, 2H), 7.50 (t, J = 7.4, 1H). 13C-NMR (100 MHz, MeOH-d4): δ 152.1, 144.6, 142.4, 134.2,
133.4, 133.4, 133.1, 132.1, 131.9, 130.0, 129.1, 123.0, 122.7, 121.5, 119.5, 119.0, 116.0, 112.1. HRMS-ESI,
m/z: [M]+ calculated for C18H1279BrN2O2+ 367.0079, found 367.0084.
For compound 27: insoluble in most solvents ivory solid, 80%. It was introduced into next step
without further purification.
3.1.5. Preparation of 14-Bromoreticulatine (7) and Compound 28
A mixture of compound 10 or 27 (0.08 mmol), acetonitrile (1 mL), sodium carbonate (0.57 mmol)
and dimethyl sulfate (0.32 mmol) was stirred at room temperature for 0.5 h. The mixture was
evaporated under reduced pressure and washed with H2O (3 mL). The resulting oil was triturated
with Et2O and dried.
For 14-bromoreticulatine (7): yellow solid, 52%. 1H-NMR (400 MHz, MeOH-d4): δ 9.40 (s, 1H), 8.76
(d, J = 6.4 Hz, 1H), 8.60 (d, J = 6.4 Hz, 1H), 8.48 (d, J = 8.1 Hz, 1H), 8.20 (d, J = 8.5 Hz, 1H), 8.12 (d, J =
1.9 Hz, 1H), 8.05 (dd, J = 8.5, 1.8 Hz, 1H), 7.79–7.87 (m, 2H), 7.50 (ddd, J = 7.5, 7.5, 0.9 Hz, 1H), 3.64 (s,
3H). 13C-NMR (100 MHz, MeOH-d4): δ 162.8, 144.8, 143.2, 134.2, 134.2, 133.7, 133.1, 132.8, 132.4, 130.8,
130.0, 127.1, 124.5, 122.8, 121.7, 119.0, 116.1, 112.3, 51.5. HRMS-ESI, m/z: [M]+ calculated for
C19H1479BrN2O2+ 381.0235, found 381.0242.
For compound 28: yellow solid 48%. 1H-NMR (400 MHz, MeOH-d4): δ 8.90 (br. s, 1H), 8.43 (d, J = 6.1
Hz, 1H), 8.20 (d, J = 8.7 Hz, 1H), 8.05 (d, J = 6.3 Hz, 1H), 7.91 (s, 1H), 7.77–7.88 (m, 3H), 7.26 (d, J = 8.4
Hz, 1H), 3.69 (s, 4 H), 3.35 (s, 3H). HRMS-ESI, m/z: [M]+ calculated for C20H1579Br2N2O2+ 473.0096, found
473.0103.
3.2. Biological Evaluation
McCoy’s 5A Medium (McCoy), Roswell Park Memorial Institute Medium (RPMI 1640),
Dulbecco’s Modified Eagle Medium (DMEM), phosphate buffered saline (PBS), L-glutamine,
penicillin–streptomycin solution, trypsin, fetal bovine serum (FBS), sodium hydrocarbonate
(NaHCO3), and agar were purchased from “Biolot” (Russia).
3.2.1. Cell Lines and Culture Conditions
Human colorectal carcinoma HT-29 (ATCC® no. HTB-38™), human breast cancer T-47D (ATCC®
no. HTB-133™), and melanoma SK-MEL-28 (ATCC® no. HTB-72™) cell lines were gifted by Hormel
Institute University of Minnesota (Austin, MN, USA). Human prostate cancer PC-3 (ATCC® no. CRL-
1435™) and 22Rv1 (ATCC® no. CRL-2505) cells were purchased from ATCC (Manassas, VA, USA).
Human colorectal carcinoma HT-29, human breast cancer T-47D, and melanoma SK-MEL-28 cell lines
were cultured in McCoy, RPMI-1640, and DMEM medium, respectively. Medium were
Mar. Drugs 2019, 17, 496 9 of 12
supplemented with 5% and 10% fetal bovine serum (FBS), 200 mM L-glutamine, and penicillin-
streptomycin solution. The cell cultures were maintained at 37 °C in humidified atmosphere
containing 5% CO2. The human prostate cancer PC-3 and 22Rv1 cells were cultured according to the
manufacturer’s instructions in RPMI media (Invitrogen), supplemented with GlutamaxTM-I
(Invitrogen, Paisley, UK) and contained 10% FBS (Invitrogen) and 1% penicillin/streptomycin
(Invitrogen). Cells were continuously kept in culture for a maximum of 3 months, and were routinely
checked for contamination with mycoplasma and inspected microscopically for stable phenotype.
Several test cultures were used to determine antibiotic activity, including Bacillus subtilis (KMM 430),
Staphylococcus aureus (ATCC 21027), P. aeruginosa (KMM 433), Escherichia coli (ATCC 15034), and
Candida albicans (KMM 455). All cultures are stored in the collection of marine microorganisms of the
PIBOC FEB RAS, the official acronym of CMM [40]. Antibiotic activity was determined with the disk
diffusion soft agar assay as described before [41].
3.2.2. Cytotoxicity Assays
MTS and MTT assays were used as an indicator of cell viability as determined by mitochondrial-
dependent reduction of formazan or its salts. For MTS assay, the cells were seeded in density of 1.0 ×
104 cells/200 µL of complete medium in 96-well plates. After incubation for 24 h attached cells were
treated with various concentrations of the compounds (0.05; 0.1; 0.5; 1; 5 µM), while the control was
treated with the complete McCoy, RPMI-1640, and DMEM medium only. Cells were cultured for
additional 24 h at 37 °C in 5% CO2 incubator. After incubation, MTS-reagent (20 µL) was added to
each well, and then cells were incubated for 3 h at 37 °C in 5% CO2. Absorbance was measured at
490/630 nm by microplate reader (Power Wave XS, American). All tested samples were carried out
in triplicates. MTT assay was performed as previously described with the 48 h drug treatment [42].
The trypan-blue-based viability assay (ViCell assay) was performed using Beckman Coulter Vi-CELL
(Beckman Coulter, Krefeld, Germany) as has been described before [43].
3.2.3. Statistical Analysis
Statistical analyses were performed using GraphPad Prism software v. 5.01 (GraphPad Prism
software Inc., La Jolla, CA, USA). Data are presented as mean ± SD. The unpaired Student’s t-test was
used for the comparison of two groups. Statistical significance was represented as * p < 0.05 and ** p
< 0.01.
4. Conclusions
Thus, the two-step approach toward the synthesis of the marine sponge derived pigment
fascaplysin was used to obtain the marine alkaloids 3-bromofascaplysin and 3,10-
dibromofascaplysin. These compounds were used as the starting materials for first syntheses of the
alkaloids 14-bromoreticulatine and 14-bromoreticulatate. Preliminary bioassays showed that 14-
bromoreticulatine reveals selective antibiotic (to P. aeruginosa) and cytotoxic (to melanoma SK-MEL-
28 cell line) activities. It was also demonstrated that 3,10-dibromofascaplysin was able to suppress
the cell metabolism at concentrations at least 7 times lower than the cytotoxic concentrations, which
induced cell membrane disruption. The examination of biological activity of the synthesized
compounds showed that even minimal modification of fascaplysin structure has a significant effect
on the bioactivity of this lead compound. At the present time, the biological activities of a large series
of novel synthetic derivatives of fascaplysin are being investigated thoroughly. This should open new
opportunities for the detailed studies of the structure–activity relationships among these potent and
promising biologically active substances.
Supplementary Materials: The following are available online at www.mdpi.com/1660-3397/17/9/496/s1,
Comparison of 1H-NMR data of synthetic and natural 3-bromofascaplysin, 3.10-dibromofascaplysin, 14-
bromoreticulatate and 14-bromoreticulatine. Spectra Data.
Author Contributions: P.A.S., O.A.T., A.V.K., and M.E.Z. performed the chemical research. Y.V.K., O.S.M.,
S.P.E., M.K., G.v.A., and S.A.D. performed the biological research. V.P.G. analyzed the data. M.E.Z. was
Mar. Drugs 2019, 17, 496 10 of 12
responsible for the funding of project, the design of the research, and the writing of the manuscript. All authors
read and approved the final manuscript.
Funding: This research was funded by the FEFU Endowment Foundation grant number D-349-17. And the APC
was funded by grant D-349-17.
Conflicts of Interest: The authors declare no conflict of interest.
References
1. Bharate, S.B.; Manda, S.; Mupparapu, N.; Battini, N.; Vishwakarma, R.A. Chemistry and Biology of
Fascaplysin, a Potent Marine-derived CDK 4 Inhibitor. Mini Rev. Med. Chem. 2012, 12, 650–664,
doi:10.2174/138955712800626719.
2. Roll, D.M.; Ireland, C.M.; Lu, H.S.M.; Clardy, J. Fascaplysin, an Unusual Antimicrobial Pigment from the
Marine Sponge Fascaplysinopsis sp. J. Org. Chem. 1988, 53, 3276–3278, doi:10.1021/jo00249a025.
3. Jimenez, C.; Quinoa, E.; Adamczeski, M.; Hunter, L.M.; Crews, P. Novel Sponge-Derived Amino Acids. 12.
Tryptophan-Derived Pigments and Accompanying Sesterterpenes from Fascaplysinopis reticulata. J. Org.
Chem. 1991, 56, 3403–3410, doi:10.1021/jo00010a041.
4. Schmidt, E.W.; Faulkner, D.J. Palauolol, a New Anti-inflammatory Sesterterpene from the Sponge
Fascaplysinopsis sp. from Palau. Tetrahedron Lett. 1996, 37, 3951–3954, doi:10.1016/0040-4039(96)00718-6.
5. Kirsch, G.; Konig, G.M.; Wright, A.D.; Kaminsky, R. A New Bioactive Sesterterpene and Antiplasmodial
Alkaloids from the Marine Sponge Hyrtios cf. erecta. J. Nat. Prod. 2000, 63, 825–829.
6. Charan, R.D.; McKee, T.C.; Gustafson, K.R.; Pannell, L.K.; Boyd, M.R. Cytotoxic Alkaloids from the Marine
Sponge Thorectandra sp. Tetrahedron Lett. 2002, 43, 5201–5204, doi:10.1080/14786410310001622077.
7. Popov, A.M.; Stonik, V.A. Physiological activity of fascaplisine—An unusual pigment from tropical sea
fishes. Antibiot. Chemoter. 1991, 36, 12–14.
8. Hamilton, G. Cytotoxic Effects of Fascaplysin against Small Cell Lung Cancer Cell Lines. Mar. Drugs 2014,
12, 1377–1389, doi:10.3390/md12031377.
9. Zheng, Y.L.; Lu, X.L.; Lin, J.; Chen, H.M.; Yan, X.J.; Wang, F.; Xu, W.F. Direct effects of fascaplysin on
human umbilical vein endothelial cells attributing the anti-angiogenesis activity. Biomed. Pharmacother.
2010, 64, 527–533, doi:10.1016/j.biopha.2009.04.046.
10. Yan, X.; Chen, H.; Lu, X.; Wang, F.; Xu, W.; Jin, H.; Zhu, P. Fascaplysin exert anti-tumor effects through
apoptotic and anti-angiogenesis pathways in sarcoma mice model. Eur. J. Pharm. Sci. 2011, 43, 251–259,
doi:10.1016/j.ejps.2011.04.018.
11. Rath, B.; Hochmair, M.; Plangger, A.; Hamilton, G. Anticancer Activity of Fascaplysin against Lung Cancer
Cell and Small Cell Lung Cancer Circulating Tumor Cell Lines. Mar. Drugs 2018, 16, 383,
doi:10.3390/md16100383.
12. Soni, R.; Muller, L.; Furet, P.; Schoepfer, J.; Stephan, C.; Zunstein-Mecker, S.; Fretz, H.; Chaudhuri, B.
Inhibition of Cyclin-Dependent Kinase 4 (Cdk4) by Fascaplysin, a Marine Natural Product. Biochem.
Biophys. Res. Commun. 2000, 275, 877–884, doi:10.1006/bbrc.2000.3349.
13. Hörmann, A.; Chaudhuri, B.; Fretz, H. DNA Binding Properties of the Marine Sponge Pigment Fascaplysin.
Bioorg. Med. Chem. 2001, 9, 917–921, doi:10.1016/S0968-0896(00)00313-8.
14. Lu, X.L.; Zheng, Y.L.; Chen, H.M.; Yan, X.J.; Wang, F.; Xu, W.F. Anti-proliferation of human cervical cancer
HeLa cell line by fascaplysin through apoptosis induction. Acta Pharm. Sin. 2009, 44, 980–986.
15. Wang, F.; Chen, H.; Yan, X.; Zheng, Y. Fascaplysin sensitizes cells to TRAIL-induced apoptosis through
upregulating DR5 expression. Chin. J. Oceanol. Limnol. 2013, 31, 560–569, doi:10.1007/s00343-013-2215-y.
16. Meng, N.; Mu, X.; Lv, X.; Wang, L.; Li, N.; Gong, Y. Autophagy represses fascaplysin-induced apoptosis
and angiogenesis inhibition via ROS and p8 in vascular endothelia cells. Biomed. Pharmacother. 2019, 114,
108866, doi:10.1016/j.biopha.2019.108866.
17. Kumar, S.; Guru, S.K.; Pathania, A.S.; Manda, S.; Kumar, A.; Bharate, S.B.; Vishwakarma, R.A.; Malik, F.;
Bhushan, S. Fascaplysin Induces Caspase Mediated Crosstalk Between Apoptosis and Autophagy Through
the Inhibition of PI3K/AKT/mTOR Signaling Cascade in Human Leukemia HL-60 Cells. J. Cell. Biochem.
2015, 116, 985–997, doi:10.1002/jcb.25053.
18. Oh, T.I.; Lee, Y.M.; Nam, T.J.; Ko, Y.S.; Mah, S.; Kim, J.; Kim, Y.; Reddy, R.H.; Kim, Y.J.; Hong, S.; Fascaplysin
Exerts Anti-Cancer Effects through the Downregulation of Survivin and HIF-1α and Inhibition of VEGFR2
and TRKA. Int. J. Mol. Sci. 2017, 18, 2074–2089, doi:10.3390/ijms18102074.
Mar. Drugs 2019, 17, 496 11 of 12
19. Oh, T.I.; Lee, J.H.; Kim, S.; Nam, T.J.; Kim, Y.S.; Kim, B.M.; Yim, W.J.; Lim, J.H. Fascaplysin Sensitizes Anti-
Cancer Effects of Drugs Targeting AKT and AMPK. Molecules 2018, 23, 42, doi:10.3390/molecules23010042.
20. Manda, S.; Sharma, S.; Wani, A.; Joshi, P.; Kumar, V.; Guru, S.K.; Bharate, S.S.; Bhushan, S.; Vishwakarma,
R.A.; Kumar, A. Discovery of a marine-derived bis-indole alkaloid fascaplysin, as a new class of potent P-
glycoprotein inducer and establishment of its structure-activity relationship. Eur. J. Med. Chem. 2016, 107,
1–11, doi:10.1016/j.ejmech.2015.10.049.
21. Johnson, T.A.; Milan-Lobo, L.; Che, T.; Ferwerda, M.; Lambu, E.; McIntosh, N.L.; Li, F.; He, L.; Lorig-Roach,
N.; Crews, P. Identification of the First Marine-Derived Opioid Receptor “balanced” Agonist with a
Signaling Profile That Resembles the Endorphins. ACS Chem. Neurosci. 2017, 8, 473–485,
doi:10.1021/acschemneuro.6b00167.
22. Sun, Q.; Liu, F.; Sang, J.; Lin, M.; Ma, J.; Xiao, X.; Yan, S.; Naman, C.B.; Wang, N.; He, Sh.; Yan, X. 9-
Methylfascaplysin Is a More Potent Aβ Aggregation Inhibitor than the Marine-Derived Alkaloid,
Fascaplysin, and Produces Nanomolar Neuroprotective Effects in SH-SY5Y Cells. Mar. Drugs 2019, 17, 121,
doi:10.3390/md17020121.
23. Sharma, S.; Guru, S.K.; Manda, S.; Kumar, A.; Mintoo, M.J.; Prasad, V.D.; Sharma, P.R.; Mondhe, D.M.;
Bharate, S.B.; Bhushan, S. A marine sponge alkaloid derivative 4-chloro fascaplysin inhibits tumor growth
and VEGF mediated angiogenesis by disrupting PI3K/Akt/mTOR signaling cascade. Chem. Biol. Interact.
2017, 275, 47–60, doi:10.1016/j.cbi.2017.07.017.
24. Kuzmich, A.S.; Fedorov, S.N.; Shastina, V.V.; Shubina, L.K.; Radchenko, O.S.; Balaneva, N.N.; Zhidkov,
M.E.; Park, J.-I.; Kwak, J.Y.; Stonik, V.A. The anticancer activity of 3- and 10-bromofascaplysins is mediated
by caspase-8, -9, -3-dependent apoptosis. Bioorg. Med. Chem. 2010, 18, 3834–3840,
doi:10.1016/j.bmc.2010.04.043.
25. Lyakhova, I.A.; Bryukhovetsky, I.S.; Kudryavtsev, I.V.; Khotimchenko, Y.S.; Zhidkov, M.E.; Kantemirov,
A.V. Antitumor Activity of Fascaplysin Derivatives on Glioblastoma Model In Vitro. Bull. Exp. Biol. Med.
2018, 164, 666–672.
26. Zhidkov, M.E.; Baranova, O.V.; Balaneva, N.N.; Fedorov, S.N.; Radchenko, O.S.; Dubovitskii, S.V. The first
syntheses of 3-bromofascaplysin, 10-bromofascaplysin and 3,10-dibromofascaplysin—Marine alkaloids
from Fascaplysinopsis reticulata and Didemnum sp. by application of a simple and effective approach to the
pyrido [1,2-a:3,4-b] diindole system. Tetrahedron Lett. 2007, 48, 7998–8000, doi:10.1016/j.tetlet.2007.09.057.
27. Segraves, N.L.; Robinson, S.J.; Garcia, D.; Said, S.A.; Fu, X.; Schmitz, F.J.; Pietraszkiewicz, H.; Valeriote,
F.A.; Crews, P. Comparison of Fascaplysin and Related Alkaloids: A Study of Structures, Cytotoxicities,
and Sources. J. Nat. Prod. 2004, 67, 783–792, doi:10.1021/np049935+.
28. Gribble, G.W.; Pelcman, B. Total Syntheses of the Marine Sponge Pigments Fascaplysin and
Homofascaplysin B and C. J. Org. Chem. 1992, 57, 3636–3642, doi:10.1021/jo00039a024.
29. Rocca, P.; Marsais, F.; Godart, A.; Quéguiner, G. A Short Synthesis of the Antimicrobial Marine Sponge
Pigment Fascaplysin. Tetrahedron Lett. 1993, 34, 7917–7918, doi:10.1016/S0040-4039(00)61510-1.
30. Molina, P.; Fresneda, P.M.; Garcia-Zafra, S.; Almendros, P. Iminophosphorane—Mediated Syntheses of the
Fascaplysin Alkaloid of Marine Origin and Nitramarine. Tetrahedron Lett. 1994, 35, 8851–8854,
doi:10.1016/S0040-4039(00)78515-7.
31. Radchenko, O.S.; Novikov, V.L.; Elyakov, G.B. A Simple and Practical Approach to the Synthesis of the
Marine Sponge Pigment Fascaplysin and Related Compounds. Tetrahedron Lett. 1997, 38, 5339–5342,
doi:10.1016/S0040-4039(97)01167-2.
32. Waldmann, H.; Eberhardt, L.; Wittstein, K.; Kumar, K. Silver catalyzed cascade synthesis of alkaloid ring
systems: Concise total synthesis of fascaplysin, homofascaplysin C and analogues. Chem. Commun. 2010,
46, 4622–4624, doi:10.1039/C001350A.
33. Zhidkov, M.E.; Baranova, O.V.; Kravchenko, N.S.; Dubovitskii, S.V. A new method for the synthesis of the
marine alkaloid fascaplysin. Tetrahedron Lett. 2010, 51, 6498–6499, doi:10.1016/j.tetlet.2010.09.120.
34. Bharate, S.B.; Manda, S.; Joshi, P.; Singh, B.; Vishwakarma, R.A. Total synthesis and anti-cholinesterase
activity of marine-derived bisindole alkaloid fascaplysin Med. Chem. Commun. 2012, 3, 1098–1103,
doi:10.1039/C2MD20076G.
35. Zhidkov, M.E.; Kaminskii, V.A. A new method for the synthesis of the marine alkaloid fascaplysin based
on the microwave-assisted Minisci reaction. Tetrahedron Lett. 2013, 54, 3530–3532,
doi:10.1016/j.tetlet.2013.04.113.
Mar. Drugs 2019, 17, 496 12 of 12
36. Zhu, Y.P.; Liu, M.C.; Cai, Q.; Jia, F.C.; Wu, A.X. A Cascade Coupling Strategy for One-Pot Total Synthesis
of β-Carboline and Isoquinoline-Containing Natural Products and Derivatives. Chem. Eur. J. 2013, 19,
10132–10137, doi:10.1002/chem.201301734.
37. Zhidkov, M.E.; Kantemirov, A.V.; Koisevnikov, A.V.; Andin, A. N.; Kuzmich, A.S. Syntheses of the marine
alkaloids 6-oxofascaplysin, fascaplysin and their derivatives. Tetrahedron Lett. 2018, 59, 708–711.
38. Zhidkov, M.E.; Sidorova, M.A.; Lyakhova, I.A. One-step transformation of the marine alkaloid fascaplysin
into homofascaplysins B and B-1. The first syntheses of 3-bromohomofascaplysin B and 3–
bromohomofascaplysin B-1. Tetrahedron Lett. 2018, 59, 1417–1420.
39. Fretz, H.; Ucci-Stoll, K.; Hug, P.; Schoepfer, J.; Lang, M. Investigations on the reactivity of fascaplysin. Part
II. General stability considerations and products formed with nucleophiles. Helv. Chim. Acta 2001, 84, 867–
873, doi:10.1002/1522-2675(20010418)84:4<867::AID-HLCA867>3.0.CO;2-A.
40. Laboratory of Microbiology. Available online: http://www.piboc.dvo.ru/en/structure/biosintez/lab6.php
(accessed on 24 June 2019).
41. Bilay, T.I. Methods of Experimental Mycology; Naukova Dumka: Kiev, Ukraine, 1982; p. 550.
42. Dyshlovoy, S.A.; Tabakmakher, K.M.; Hauschild, J.; Shchekaleva, R.K.; Otte, K.; Guzii, A.G.; Makarieva,
T.N.; Kudryashova, E.K.; Fedorov, S.N.; Shubina, L.K.; et al. Guanidine Alkaloids from the Marine Sponge
Monanchora pulchra Show Cytotoxic Properties and Prevent EGF-Induced Neoplastic Transformation in
Vitro. Mar. Drugs 2016, 14, 133, doi:10.3390/md14070133.
43. Dyshlovoy, S.A.; Madanchi, R.; Hauschild, J.; Otte, K.; Alsdorf, W.H.; Schumacher, U.; Kalinin, V.I.;
Silchenko, A.S.; Avilov, S.A.; Honecker, F.; et al. The marine triterpene glycoside frondoside A induces p53-
independent apoptosis and inhibits autophagy in urothelial carcinoma cells. BMC Cancer 2017, 17, 93,
doi:10.1186/s12885-017-3085-z.
© 2019 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access
article distributed under the terms and conditions of the Creative Commons Attribution
(CC BY) license (http://creativecommons.org/licenses/by/4.0/).
... Recently, the two-step approach toward the synthesis of fascaplysin was utilized by our group to generate 3-bromofascaplysin and 3,10-dibromofascaplysin (8), which were previously isolated from natural sources [27]. Remarkably, 3,10-dibromofascaplysin was able to suppress the cell metabolism at non-cytotoxic concentrations [28]. Further studies to determine the mechanism of the antitumor activity conducted in myeloid leukemia cells revealed that 8 activates the transcription factor E2F1 and decreases the expression of several genes responsible for cancer cell survival [29]. ...
... In addition, in human prostate cancer cells, we identified JNK1/2 to be one of the primary molecular targets of 8. Additionally, 8 could synergize with PARP-inhibitor Olaparib, presumably due to the induction of ROS production and consequent oxidative DNA damage mediated by the drug [30]. Additionally, 8 increased the effects of well-established drugs such as cytarabine, cisplatin, carboplatin, as well as docetaxel and cabazitaxel [28,29]. Despite some promising results published in recent years, the cytotoxicity to normal (non-cancer) cells was assumed to be the main challenge for fascaplysin and related compounds on their way to clinically approved drugs. ...
... To apply this synthetic approach for the synthesis of 3,10-dibromofascaplysin (8), the reaction between 3-bromophenylhydrazine (24) and 4-bromobutanal (25) in an autoclave at 150 °C was used to prepare the mixture of 6-bromotryptamine (26) and 4-bromotryptamine (27). After chromatography purification, two isomeric 1-benzoyl-β-carbolines (28a and 28b) were subsequently transformed to 3,10-dibromofascaplysin (8) and its isomer 9 with the procedure described above for fascaplysin (Scheme 2) [28]. To simplify the preparation of variety of fascaplysin derivatives at cycle A, we initially used the interaction of fascaplysin at C-9 with molecular bromine and molecular chlorine in acetic acid, as described in the literature [45]. ...
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Marine alkaloid fascaplysin and its derivatives are known to exhibit promising anticancer properties in vitro and in vivo. However, toxicity of these molecules to non-cancer cells was identified as a main limitation for their clinical use. Here, for the very first time, we synthesized a library of fascaplysin derivatives covering all possible substituent introduction sites, i.e., cycles A, C and E of the 12H-pyrido[1-2-a:3,4-b’]diindole system. Their selectivity towards human prostate cancer versus non-cancer cells, as well as the effects on cellular metabolism, membrane integrity, cell cycle progression, apoptosis induction and their ability to intercalate into DNA were investigated. A pronounced selectivity for cancer cells was observed for the family of di- and trisubstituted halogen derivatives (modification of cycles A and E), while a modification of cycle C resulted in a stronger activity in therapy-resistant PC-3 cells. Among others, 3,10-dibromofascaplysin exhibited the highest selectivity, presumably due to the cytostatic effects executed via the targeting of cellular metabolism. Moreover, an introduction of radical substituents at C-9, C-10 or C-10 plus C-3 resulted in a notable reduction in DNA intercalating activity and improved selectivity. Taken together, our research contributes to understanding the structure–activity relationships of fascaplysin alkaloids and defines further directions of the structural optimization.
... In addition, the brominated derivatives, i.e., 3-and 10-bromofascaplysins induce caspase-dependent apoptosis in human leukemia cells at nanomolar concentrations [31]. 3,10-Dibromofascaplysin (DBF) is a novel halogenated fascaplysin alkaloid initially isolated from the marine sponge Fascaplysinopsis reticulata, which has been recently synthesized by our group [32]. Treatment with DBF affects cellular metabolism and causes death of prostate cancer cells [32]. ...
... 3,10-Dibromofascaplysin (DBF) is a novel halogenated fascaplysin alkaloid initially isolated from the marine sponge Fascaplysinopsis reticulata, which has been recently synthesized by our group [32]. Treatment with DBF affects cellular metabolism and causes death of prostate cancer cells [32]. In addition, we have reported DBF to inhibit growth of human drug-resistant prostate cancer cells [33] and identified JNK1/2 kinase to be one of the potential targets of this natural compound. ...
... The marine alkaloid 3,10-dibromofascaplysin (DBF) was synthesized and purified as previously reported [32]. 3-Bromofascaplysin and 10-bromofascaplysin were synthesized and purified as previously reported (Zhidkov, M.E. ...
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Myeloid leukemia is a hematologic neoplasia characterized by a clonal proliferation of hematopoietic stem cell progenitors. Patient prognosis varies depending on the subtype of leukemia as well as eligibility for intensive treatment regimens and allogeneic stem cell transplantation. Although significant progress has been made in the therapy of patients including novel targeted treatment approaches, there is still an urgent need to optimize treatment outcome. The most common therapy is based on the use of chemotherapeutics cytarabine and anthrayclines. Here, we studied the effect of the recently synthesized marine alkaloid 3,10-dibromofascaplysin (DBF) in myeloid leukemia cells. Unsubstituted fascaplysin was early found to affect cell cycle via inhibiting CDK4/6, thus we compared the activity of DBF and other brominated derivatives with known CDK4/6 inhibitor palbociclib, which was earlier shown to be a promising candidate to treat leukemia. Unexpectedly, the effect DBF on cell cycle differs from palbociclib. In fact, DBF induced leukemic cells apoptosis and decreased the expression of genes responsible for cancer cell survival. Simultaneously, DBF was found to activate the E2F1 transcription factor. Using bioinformatical approaches we evaluated the possible molecular mechanisms, which may be associated with DBF-induced activation of E2F1. Finally, we found that DBF synergistically increase the cytotoxic effect of cytarabine in different myeloid leukemia cell lines. In conclusion, DBF is a promising drug candidate, which may be used in combinational therapeutics approaches to reduce leukemia cell growth.
... At the same time, the synthetic analog 9-methylfascaplysin reduced the formation of Aβ oligomers by interacting with negatively charged Aβ42 residues, attenuating the toxic effect of Aβ42 on SH-SY5Y cells [376]. At the PIBOC center (Vladivostok), routes for chemical synthesis of fascaplysin and its derivatives 6-oxofascaplysin, homofascaplysins B, 3-bromohomofascaplysin B, 3-bromohomofascaplysin B-1, and 3,10-dibromofascaplysin have been developed for subsequent assessment of their biological activities including those targeting AD [377][378][379]. ...
... Oxysterols Inhibition of BACE1; reduction in ROS production; increase in neuronal survival; neuroprotective effects [312,314] Sea urchin Scaphechinus mirabilis Echinochrome Antioxidant and neuroprotective effects [353,[355][356][357] Starfish Hippasteria kurilensis, Linckia laevigata, Aphelasterias japonica, Leptasterias ochotensis, Linckia hylodes reticulata, Lethasterias fusca, Lethasterias nanimensis chelifera, Aphelasterias japonica Asterosaponines Anti-inflammatory, neuritogenic, and neuroprotective effects [359][360][361][362][363][364][365][366][367][368][369][370] Sponge Lissodendoryx florida Lissodendoric acids A and B Decrease in ROS [374] Sponge Fascaplysinopsis sp. Fascaplysin and its synthetic analogs Inhibition of AChE; inhibition of Aβ42 fibrils [377][378][379] Crabs Paralithodes camtschaticus, Paralithodes platypus, Chionoecetes opilio, Chionoecetes angulatus, Chionoecetes japonicus Docosahexaenoic acid, eicosapentaenoic acid Reduction in tau phosphorylation; reduction in the levels of proinflammatory cytokines; prevention of neuroinflammation; antioxidant effects; activation of remyelination; improvement of motor and cognitive functions; neuroprotective effects [220,239,244] Hepatopancreas of crab Paralithodes camtschaticus, liver of squid Berryteuthis magister, liver of stingray Bathyraja parmifera 1-O-alkyl-glycerols Reduction in expression of proinflammatory cytokines; prevention of activation of M1 microglia [230,233,235] Oncorhynchus gorbuscha (and other pacific salmon) N-docosahexanoylethanolamine Suppression of TNF-α expression, NO production, and neuroinflammation; activation of synaptogenesis; neuroprotective effects [252,253,257] Pacific saury Cololabis saira Phosphatidylcholine, phosphatidylinositol, phosphatidylethanolamine Suppression of Aβ42 release [259] 1 Marine organisms' classification is presented in accordance with the World Register of Marine Species. ...
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Neurodegenerative diseases are growing to become one of humanity’s biggest health problems, given the number of individuals affected by them. They cause enough mortalities and severe economic impact to rival cancers and infections. With the current diversity of pathophysiological mechanisms involved in neurodegenerative diseases, on the one hand, and scarcity of efficient prevention and treatment strategies, on the other, all possible sources for novel drug discovery must be employed. Marine pharmacology represents a relatively uncharted territory to seek promising compounds, despite the enormous chemodiversity it offers. The current work discusses one vast marine region—the Northwestern or Russian Pacific—as the treasure chest for marine-based drug discovery targeting neurodegenerative diseases. We overview the natural products of neurological properties already discovered from its waters and survey the existing molecular and cellular targets for pharmacological modulation of the disease. We further provide a general assessment of the drug discovery potential of the Russian Pacific in case of its systematic development to tackle neurodegenerative diseases.
... Fascaplysin (30), a bis-indole alkaloid with multiple bioactivities, was originally discovered from marine sponge Fascaplysinopsis bergquist [44]. Zhidkov et al. designed and synthesized a series of brominated fascaplysins, and compound 31 exhibited potent and selective inhibitory activity toward P. aeruginosa with clearance zone more than 35 mm at the concentration of 0.2 mg/disc [45]. It also demonstrated cytotoxic activity against melanoma cells SK-MEL-28 with an IC 50 value of 1.2 µM. ...
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Pseudomonas aeruginosa, one of the most intractable Gram-negative bacteria, has become a public health threat due to its outer polysaccharide layer, efflux transporter system, and high level of biofilm formation, all of which contribute to multi-drug resistance. Even though it is a pathogen of the highest concern, the status of the antibiotic development pipeline is unsatisfactory. In this review, we summarize marine natural products (MNPs) isolated from marine plants, animals, and microorganisms which possess unique structures and promising antibiotic activities against P. aeruginosa. In the last decade, nearly 80 such MNPs, ranging from polyketides to alkaloids, peptides, and terpenoids, have been discovered. Representative compounds exhibited impressive in vitro anti-P. aeruginosa activities with MIC values in the single-digit nanomolar range and in vivo efficacy in infectious mouse models. For some of the compounds, the preliminary structure-activity-relationship (SAR) and anti-bacterial mechanisms of selected compounds were introduced. Compounds that can disrupt biofilm formation or membrane integrity displayed potent inhibition of multi-resistant clinical P. aeruginosa isolates and could be considered as lead compounds for future development. Challenges on how to translate hits into useful candidates for clinical development are also proposed and discussed.
... [53]. Then synthesis of fascaplysin and its derivatives was elaborated in a relatively short period, and this process is still ongoing [54,55]. Fascaplysin exerts various biological activities including selective kinase 4 (CDK-4) inhibition, DNA binding, and antiangiogenic effects [56]. ...
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The search for new chemical compounds with antitumor pharmacological activity is a necessary process for creating more effective drugs for each specific malignancy type. This review presents the outcomes of screening studies of natural compounds with high anti-glioma activity. Despite significant advances in cancer therapy, there are still some tumors currently considered completely incurable including brain gliomas. This review covers the main problems of the glioma chemotherapy including drug resistance, side effects of common anti-glioma drugs, and genetic diversity of brain tumors. The main emphasis is made on the characterization of natural compounds isolated from marine organisms because taxonomic diversity of organisms in seawaters significantly exceeds that of terrestrial species. Thus, we should expect greater chemical diversity of marine compounds and greater likelihood of finding effective molecules with antiglioma activity. The review covers at least 15 classes of organic compounds with their chemical formulas provided as well as semi-inhibitory concentrations, mechanisms of action, and pharmacokinetic profiles. In conclusion, the analysis of the taxonomic diversity of marine species containing bioactives with antiglioma activity is performed noting cytotoxicity indicators and to the tumor cells in comparison with similar indicators of antitumor agents approved for clinical use as antiglioblastoma chemotherapeutics.
... [53]. Then synthesis of fascaplysin and its derivatives was elaborated in a relatively short period and this process is still ongoing [54,55]. Fascaplysin exerts various biological activities including selective kinase 4 (CDK-4) inhibition, DNA binding, and antiangiogenic effects [56]. ...
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The search for new chemical compounds with antitumor pharmacological activity is a necessary process for creating more effective drugs for each specific malignancy type. This review presents the outcomes of screening studies of natural compounds with high anti-glioma activity. Despite significant advances in cancer therapy, there are still some tumors currently considered completely incurable including brain gliomas. This review covers the main problems of the glioma chemotherapy including drug resistance, side effects of common anti-glioma drugs, and genetic diversity of brain tumors. The main emphasis is made on the characterization of natural compounds isolated from marine organisms because taxonomic diversity of organisms in seawaters significantly exceeds that of terrestrial species. Thus, we should expect greater chemical diversity of marine compounds and greater likelihood of finding effective molecules with antiglioma activity. The review covers at least 15 classes of organic compounds with their chemical formulas provided as well as semi-inhibitory concentrations, mechanisms of action, and pharmacokinetic profiles. In conclusion, the analysis of the taxonomic diversity of marine species containing bioactives with antiglioma activity is performed noting cytotoxicity indicators and to the tumor cells in comparison with similar indicators of antitumor agents approved for clinical use as antiglioblastoma chemotherapeutics.
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A new methodology for the synthesis of fascaplysin derivatives has been established. A library of diversified fascaplysins can be efficiently and quickly prepared by regioselective Suzuki‐Miyaura coupling and subsequent quaternization for the first time. The minimum inhibitory concentration assessment showed that the unique 5‐ring coplanar aromatic system plays a key role in the resulting bioactivity, and functional groups at different substitution position exert differentiable impact on the bactericidal activity. some synthesized fascaplysin derivatives exhibited higher antibacterial activities than the pristine fascaplysin and the positive controls, especially against Gram‐negative bacteria Escherichia coli (ATCC 25922). These novel fascaplysin derivatives may have promising application potentials as bactericidal drugs. Novel fascaplysin derivatives have been synthesized via Suzuki‐Miyaura coupling with high regioselectivity, efficiency and simpleness. These derivatives exhibit potent antibacterial activity, especially against Gram‐negative bacterial Escherichia coli (ATCC 25922).
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Since early 2020, disease caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has become a global pandemic, causing millions of infections and deaths worldwide. Despite rapid deployment of effective vaccines, it is apparent that the global community lacks multipronged interventions to combat viral infection and disease. A major limitation is the paucity of antiviral drug options representing diverse molecular scaffolds and mechanisms of action. Here we report the antiviral activities of three distinct marine natural products─homofascaplysin A (1), (+)-aureol (2), and bromophycolide A (3)─evidenced by their ability to inhibit SARS-CoV-2 replication at concentrations that are nontoxic toward human airway epithelial cells. These compounds stand as promising candidates for further exploration toward the discovery of novel drug leads against SARS-CoV-2.
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Methicillin-resistant Staphylococcus aureus (MRSA) is considered as one of the most dangerous clinical pathogens. Biofilms forming ability of MRSA is also a major cause of drug resistance. Hence, it is in urgent need to develop novel antibacterial/antibiofilm drugs. Fascaplysin with a unique cationic five-ring coplanar backbone is emerging as a potential antibacterial compound. In this study, aiming at developing novel and more effective agents, a series of fascaplysin derivatives and their corresponding β-carboline precursors have been synthesized. Then their antibacterial/antibiofilm activity and mechanisms against MRSA were investigated for the first time. The results showed that most fascaplysins rather than β-carboline precursors exhibit superior antimicrobial activity against MRSA ATCC43300, demonstrating the important role of cationic five-ring coplanar backbone playing in antibacterial activity. Among them, 14 and 18 are the most potent compounds with MIC value of 0.098 μg/ml (10-fold lower than vancomycin), and 18 featuring the lowest toxicity. Subsequent mechanisms exploration indicates that 18 have relatively stronger ability to destroy bacterial cell wall and membrane, higher binding affinity to bacterial genomic DNA. Molecular docking study revealed that besides the key role of cationic five-ring coplanar backbone, introduction of N-aryl amide at 9-position of fascaplysin promoted the combination of compound 18 and DNA via additional π-π stacking and hydrogen bonding of the naphthyl group. Moreover, fascaplysins could inhibit MRSA biofilm formation in vitro and bacterial infection in vivo. All these results illustrate that fascaplysin derivative 18 is a strong and safe multi-target antibacterial agent, which makes it an attractive candidate for the treatment of MRSA and its biofilm infections.
Chapter
Neuroinflammation is one of the key events in the progression of multiple neurological disorders. The blood–brain barrier and blood–nerve barrier are responsible for the development of neuroinflammation via metabolic alteration, free radical generation, and lipid peroxidation. Furthermore, overexpression of biological proteins such as proinflammatory and proapoptotic mediators, activation of glial, astrocyte, oligodendrocyte, and Schwann cells also plays a key role in the development of neuroinflammation. Moreover, in some cases, alteration of cellular enzymes, ion channels, and prion proteins are also used to enhance neuroinflammation. The natural source of medicines such as plant, animal, marine, and mineral drugs plays a critical role in ameliorating the free radical, lipid peroxidation, and inflammatory cytokine-associated neuroinflammation. Some conventional medicines are involved in the regulation of neurotransmitters and ion channel function in the nervous system. However, the clinical use of conventional medicines is still questionable due to its low safety, efficacy, and higher intolerable adverse effects. The recent drug discovery process has paid greater attention toward natural medicines especially marine drugs for neuroinflammatory disorders. Some of the marine drugs have a promising role in the management of neurovascular disorders via potential antiinflammatory actions. However, the relationship between chemical structure and their biological activity remains to be explored. It is an essential part of bringing potential medicines from nature to health management. Hence, this book chapter is based on exploring the structure–activity relationship of marine drugs for neuroinflammatory disorders.
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Fascaplysin is a natural product isolated from marine sponges that exhibits broad anticancer activity. Previous studies revealed that fascaplysin-induced apoptosis and angiogenesis inhibition in vascular endothelial cells contributed to its anticancer activity. Accumulating evidence indicates that autophagy plays a significant role in mediating the function of vascular endothelial cells (VECs) and the response to cancer therapy. However, the effect of fascaplysin on VEC autophagy and the role of autophagy in fascaplysin-induced vascular endothelial cell apoptosis and angiogenesis inhibition are not clear. The present study found that fascaplysin induced autophagy in vascular endothelial cells. Suppression of autophagy using a pharmacological inhibitor (3-methyladenine) or RNA interference of an essential autophagy gene (ATG5) enhanced the cell death and anti-angiogenesis activity of fascaplysin. We further found that fascaplysin significantly increased p8 protein and reactive oxygen species (ROS) levels and decreased mitochondrial membrane potential but had no effect on the mTOR pathway in VECs. Notably, the ROS scavenger N-acetylcysteine inhibited fascaplysin-induced autophagy and increased p8 protein level. Knockdown of p8 by using RNA interference inhibited the autophagy but increased the level of ROS in VECs. Taken together, these data indicated that fascaplysin activated autophagy as a cytoprotective response via ROS and p8 in VECs. Our findings provided important insight into the response of VECs to fascaplysin and may be useful for improving the anticancer efficacy of fascaplysin.
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β-Amyloid (Aβ) is regarded as an important pathogenic target for Alzheimer’s disease (AD), the most prevalent neurodegenerative disease. Aβ can assemble into oligomers and fibrils, and produce neurotoxicity. Therefore, Aβ aggregation inhibitors may have anti-AD therapeutic efficacies. It was found, here, that the marine-derived alkaloid, fascaplysin, inhibits Aβ fibrillization in vitro. Moreover, the new analogue, 9-methylfascaplysin, was designed and synthesized from 5-methyltryptamine. Interestingly, 9-methylfascaplysin is a more potent inhibitor of Aβ fibril formation than fascaplysin. Incubation of 9-methylfascaplysin with Aβ directly reduced Aβ oligomer formation. Molecular dynamics simulations revealed that 9-methylfascaplysin might interact with negatively charged residues of Aβ42 with polar binding energy. Hydrogen bonds and π–π interactions between the key amino acid residues of Aβ42 and 9-methylfascaplysin were also suggested. Most importantly, compared with the typical Aβ oligomer, Aβ modified by nanomolar 9-methylfascaplysin produced less neuronal toxicity in SH-SY5Y cells. 9-Methylfascaplysin appears to be one of the most potent marine-derived compounds that produces anti-Aβ neuroprotective effects. Given previous reports that fascaplysin inhibits acetylcholinesterase and induces P-glycoprotein, the current study results suggest that fascaplysin derivatives can be developed as novel anti-AD drugs that possibly act via inhibition of Aβ aggregation along with other target mechanisms.
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Lung cancer is a leading cause of tumor-associated mortality. Fascaplysin, a bis-indole of a marine sponge, exhibit broad anticancer activity as specific CDK4 inhibitor among several other mechanisms, and is investigated as a drug to overcome chemoresistance after the failure of targeted agents or immunotherapy. The cytotoxic activity of fascaplysin was studied using lung cancer cell lines, primary Non-Small Cell Lung Cancer (NSCLC) and Small Cell Lung Cancer (SCLC) cells, as well as SCLC circulating tumor cell lines (CTCs). This compound exhibited high activity against SCLC cell lines (mean IC50 0.89 µM), as well as SCLC CTCs as single cells and in the form of tumorospheres (mean IC50 0.57 µM). NSCLC lines showed a mean IC50 of 1.15 µM for fascaplysin. Analysis of signal transduction mediators point to an ATM-triggered signaling cascade provoked by drug-induced DNA damage. Fascaplysin reveals at least an additive cytotoxic effect with cisplatin, which is the mainstay of lung cancer chemotherapy. In conclusion, fascaplysin shows high activity against lung cancer cell lines and spheroids of SCLC CTCs which are linked to the dismal prognosis of this tumor type. Derivatives of fascaplysin may constitute valuable new agents for the treatment of lung cancer.
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Fascaplysin, a natural product isolated from marine sponges, is a potential candidate for the development of anti-cancer drugs. However, the mechanism underlying its therapeutic effect of strengthening anti-cancer efficacy of other drugs is poorly understood. Here, we found that fascaplysin increases phosphorylation of protein kinase B (PKB), also known as AKT, and adenosine monophosphate-activated protein kinase (AMPK), which are considered therapeutic targets for cancer treatment due to their anti-apoptotic or pro-survival functions in cancer. A cell viability assay revealed that pharmacological suppression of AKT using LY294002 enhanced the anti-cancer effect of fascaplysin in various cancer cells. Similarly, fascaplysin was observed to have improved anti-cancer effects in combination with compound C, a selective AMPK inhibitor. Another challenge showed that fascaplysin increased the efficacy of methotrexate (MTX)-mediated cancer therapy by suppressing genes related to folate and purine metabolism. Overall, these results suggest that fascaplysin may be useful for improving the anti-cancer efficacy of targeted anti-cancer drugs, such as inhibitors of phosphoinositide 3-kinase AKT signaling, and chemotherapeutic agents, such as MTX.
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Fascaplysin has been reported to exert anti-cancer effects by inhibiting cyclin-dependent kinase 4 (CDK4); however, the precise mode of action by which fascaplysin suppresses tumor growth is not clear. Here, we found that fascaplysin has stronger anti-cancer effects than other CDK4 inhibitors, including PD0332991 and LY2835219, on lung cancer cells that are wild-type or null for retinoblastoma (RB), indicating that unknown target molecules might be involved in the inhibition of tumor growth by fascaplysin. Fascaplysin treatment significantly decreased tumor angiogenesis and increased cleaved-caspase-3 in xenografted tumor tissues. In addition, survivin and HIF-1α were downregulated in vitro and in vivo by suppressing 4EBP1-p70S6K1 axis-mediated de novo protein synthesis. Kinase screening assays and drug-protein docking simulation studies demonstrated that fascaplysin strongly inhibited vascular endothelial growth factor receptor 2 (VEGFR2) and tropomyosin-related kinase A (TRKA) via DFG-out non-competitive inhibition. Overall, these results suggest that fascaplysin inhibits TRKA and VEGFR2 and downregulates survivin and HIF-1α, resulting in suppression of tumor growth. Fascaplysin, therefore, represents a potential therapeutic approach for the treatment of multiple types of solid cancer.
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Background Advanced urothelial carcinomas represent a considerable clinical challenge as they are difficult to treat. Platinum-based combination regimens obtain response rates ranging from 40 to 70% in first-line therapy of advanced urothelial carcinoma. In the majority of cases, however, the duration of these responses is limited, and when progression occurs, the outcome is generally poor. Therefore, novel therapeutic strategies are urgently needed. The purpose of the current research is to investigate the anticancer effects and the mode of action of the marine triterpene glycoside frondoside A in p53-wild type and p53-deficient human urothelial carcinoma cells. Methods Activity of frondoside A was examined in the human urothelial carcinoma cell lines RT112, RT4, HT-1197, TCC-SUP, T-24, and 486p. Effects of frondoside A on cell viability, either alone or in combination with standard cytotoxic agents were investigated, and synergistic effects were analyzed. Pro-apoptotic activity was assessed by Western blotting and FACS, alone and in combination with a caspases-inhibitor. The impact of functional p53 was investigated by siRNA gene silencing and the p53 inhibitor pifithrin-α. Effects on autophagy were studied using LC3B-I/II and SQSTM/p62 as markers. The unpaired Student’s t-test was used for comparison of the data sets. ResultsFrondoside A shows high cytotoxicity in urothelial carcinoma cells with IC50s ranging from 0.55 to 2.33 μM while higher concentrations of cisplatin are required for comparable effects (IC50 = 2.03 ~ 5.88 μM). Induction of apoptosis by frondoside A was associated with the regulation of several pro-apoptotic factors, like caspase-3, -8, and -9, PARP, Bax, p21, DNA fragmentation, and externalization of phosphatidylserine. Remarkably, inhibition of p53 by gene silencing or pifithrin-α pretreatment, as well as caspase inhibition, did not suppress apoptotic activity of frondoside A, while cisplatin activity, in contrast, was significantly decreased. Frondoside A inhibited pro-survival autophagy, a known mechanism of drug resistance in urothelial carcinoma and showed synergistic activity with cisplatin and gemcitabine. ConclusionsA unique combination of properties makes marine compound frondoside A a promising candidate for the treatment of human urothelial carcinomas.
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Antitumor efficiency of fascaplysin synthetic derivatives (7-phenylfascaplysin, 3-chlorofascaplysin, 3-bromofascaplysin, and 10-bromofascaplysin) was compared out in vitro on C6 glioma cells. The cytotoxic efficiency of all tested compounds was higher than that of unsubstituted fascaplysin; 3-bromofascaplysin and 7-phenylfascaplysin exhibited the best capacity to kill glioma C6 cells. Apoptosis was the main mechanism of glioma cell death. The cytotoxic activity of these compounds increased with prolongation of exposure to the substance and increase of its concentration. Fascaplysin derivatives modified all phases of glioma cell vital cycle. The count of viable tumor cell in G0 phase remained minimum by the end of experiment under the effects of 3-bromofascaplysin and 7-phenylfascaplysin.
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A method for one-step conversion of the marine alkaloid fascaplysin into homofascaplysins B and B-1 was elaborated. It was also used for the first syntheses of the marine alkaloids 3-bromohomofascaplysin B and 3-bromohomofascaplysin B-1. The best results were demonstrated under the action of microwave irradiation in presents of dihydroquinone as a reducing agent.
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A simple approach towards the pyrido[1,2-a:3,4-b']diindole system via the reaction of indigo with methylene active compounds was used for the syntheses of the marine alkaloids 6-oxofascaplysin, fascaplysin, and their derivatives. It was also demonstrated that the reaction with ketones led to indigo decomposition and the formation of isatin derivatives. The derivative of fascaplysin with a phenyl substituent at C-7 demonstrated 2-3 times greater inhibitory activity against selected cancer cell lines than fascaplysin.
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Tumor angiogenesis and PI3K/Akt/mTOR pathway are two major molecular objectives for the treatment and management of breast cancer. Here we first time report the molecular mechanism of a marine sponge alkaloid derivative 4-chloro fascapysin (4-CF) for its anticancer and antiangiogenesis potential. It simultaneously targets multiple cancer and angiogenesis dynamics, such as proliferation, chemotaxis cell migration, and invasion, growth factors signaling cascade, autophagy and apoptosis in HUVEC and MDAMB-231 breast cancer cells. It inhibited the VEGF mediated microvessel sprouting and blood vessel formation in the matrigel plug of C57/BL6J mice. It inhibits the tumor growth in ET (solid) mouse tumor model. It significantly inhibited cell survival through PI3K/Akt/mTOR pathway, with attendant effects on key pro-angiogenesis factors like HIF-1α, eNOS and MMP-2/9. The cytotoxicity of 4-CF was reversed by co-treatment with the VEGF and Akt inhibitors sunitinib and perifosine, respectively or by the addition of neutralizing VEGF antibodies. The apoptotic potential of 4-CF was through mitochondrial dependent as illustrated through loss of mitochondrial membrane potential. The safety profile of 4-CF was acceptable as it exhibits five times high cytotoxic IC50 value in normal cells as well as no apparent toxicities in experimental tumor mice at therapeutic doses.