manzamine alkaloids: isolation, cytotoxicity, antimalarial activity and sar studies

Q1

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Reviews�POSTSCREEN

Drug Discovery Today � Volume 00, Number 00 � June 2014 REVIEWS

Manzamine alkaloids: isolation,cytotoxicity, antimalarial activity andSAR studies

Penta Ashok1, Swastika Ganguly2 and Sankaranarayanan Murugesan1

1Medicinal Chemistry Research Laboratory, Department of Pharmacy, Birla Institute of Technology & Science, Pilani 333031, India2Department of Pharmaceutical Sciences, Birla Institute of Technology, Mesra 835215, India

The infectious disease Malaria is caused by different species of the genus Plasmodium. Resistance to

quinoline antimalarial drugs and decreased susceptibility to artemisinin-based combination therapy

have increased the need for novel antimalarial agents. Historically, natural products have been used for

the treatment of infectious diseases. Identification of natural products and their semi-synthetic

derivatives with potent antimalarial activity is an important method for developing novel antimalarial

agents. Manzamine alkaloids are a unique group of b-carboline alkaloids isolated from various species of

marine sponge displaying potent antimalarial activity against drug-sensitive and -resistant strains of

Plasmodium. In this review, we demonstrate antimalarial potency, cytotoxicity and antimalarial SAR of

manzamine alkaloids.

3

IntroductionMalaria is one of the major health problems in developing coun-

tries. According to a WHO report, approximately 3.4 billion people

were living in at risk places for malaria in 2012. Nearly 80% of cases

and 90% of deaths are reported from sub-Saharan Africa and

children under age of five and pregnant women are severely

affected [1]. Human malaria is caused by five species of parasites

of the genus Plasmodium (Plasmodium falciparum, Plasmodium

vivax, Plasmodium ovale, Plasmodium malariae and Plasmodium

knowlesi) [2,3]. The majority of the malarial infections in Africa

are caused by the more lethal P. falciparum, whereas P. vivax

malaria is widespread globally and rarely life-threatening [1,4].

Malaria is transmitted by more than 30 species of female anophe-

line mosquitoes. The control methods of malaria include vector

control, prophylatic drugs and vaccination. Malaria vaccine de-

velopment has a long research history. In the 1970s malaria

vaccine was prepared by irradiating the sporozoites of P. falciparum

[5,6]. But further vaccine development has been hampered, owing

to the complex parasite lifecycle and different mechanisms of

induction of host immune responses. Hence, to date no complete-

ly effective vaccine is available for the control and prevention of

malaria.

Please cite this article in press as: Ashok, P. et al., Manzamine alkaloids: isolation, cytotoxicity, aj.drudis.2014.06.010

Corresponding author: Murugesan, S. ([email protected])

1359-6446/06/$ - see front matter � 2014 Published by Elsevier Ltd. http://dx.doi.org/10.1016/j.drudis.2014.

In addition to these control methods, malarial chemotherapy

plays a vital part in malaria control. Quinine is a quinoline alkaloid

isolated from Cinchona bark, and was the first drug used to treat

malaria. Methylene blue was the first synthetic drug used to treat

human malaria [7]. By modifying the chemical structure of meth-

ylene blue, the 8-aminoquinoline derivative pamaquine was de-

veloped in 1925. It was the first drug with the ability to prevent the

relapses seen with P. vivax malaria [8]. Chloroquine (CQ), a 4-

aminoquinoline derivative, was used to treat malaria from 1946

and was the drug of choice for malarial treatment for decades. CQ

was used extensively throughout the 1950s and 1960s; but, be-

cause of its excess prophylactic use, parasites developed resistance

to CQ that restricts the usage of CQ in modern therapy against

malaria [1]. Amodiaquine is structurally related to CQ, but the use

of amodiaquine has been limited because of toxic effects such as

agranulocytosis and hepatotoxicity [9]. Mefloquine was intro-

duced to treat CQ-resistant malaria; however, its use has been

limited because of acquired resistance and neuropsychiatric side

effects [10]. Besides 4-aminoquinolines, the antifolate combina-

tion sulfadoxine–pyrimethamine is the recommended medication

for the treatment of malaria. However, adverse effects such as QStevens–Johnson syndrome and developed resistance have limited

the use of this combination therapy [11]. Antibiotics such

as rifampicin, doxycycline and clindamycin are also used in

ntimalarial activity and SAR studies, Drug Discov Today (2014), http://dx.doi.org/10.1016/

06.010 www.drugdiscoverytoday.com 1

http://dx.doi.org/10.1016/j.drudis.2014.06.010


mailto:[email protected]


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REVIEWS Drug Discovery Today �Volume 00, Number 00 � June 2014

DRUDIS 1438 1–10

2

Review

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ombination with other antimalarials as prophylactic agents but

heir use is restricted in pregnant women and children [12].

lthough atovaquone in combination with proguanil has shown

ynergistic antimalarial activity, long-term efficacy is limited as a

esult of developed resistance [13]. Artemisinin is a natural sesqui-

erpene, isolated from sweet warm wood plant Artemisia annua and

eported for potent antimalarial activity. Presently, artemisinin

nd its semi-synthetic derivatives artemether, arteether and arte-

unate are the first line of antimalarial drugs, especially in regions

here resistance to other antimalarial agents has developed. WHO

ecommends the use of artemisinin-based combination therapy

CT) rather than artemisinin alone to prevent emergence of

esistance and to reduce side effects. Presently, ACT is the preferred

reatment regimen for malaria caused by P. falciparum. Unfortu-

ately, susceptibility to ACT is also decreasing and signs of resis-

ance have been identified from the Thai–Cambodian border in

outheast Asia. Signs of resistance to ACT have increased the need

o develop a novel, potent antimalarial agent(s) before resistance

preads further [1].

Nature remains the important source for compounds with

edicinal importance. The historical use of natural products for

he treatment of parasitic infections is well known. Natural pro-

ucts such as quinine and artemisinin have an important role in

alarial chemotherapy. Hence, identification of natural products

nd their semi-synthetic derivatives with potent antimalarial ac-

ivity is one of the effective methods to develop novel antimalarial

gents. Manzamines are a unique group of b-carboline alkaloids

ith an unusual polycyclic system, present in various species of

arine sponges found in the Indian and Pacific Oceans. Manza-

ine A was the first compound of this group isolated from

kinawa sponge in 1986 [14]. Currently more than 100 manza-

ine natural alkaloids have been isolated from more than 16

pecies of marine sponges belonging to five families distributed

om the Red Sea to Indonesia. Many of these isolated alkaloids

isplay potent antimalarial activity against drug-sensitive and -

esistant strains in in vitro and in vivo studies. A few of these

lkaloids exhibited superior antimalarial potency to antimalarial

rugs CQ and artemisinin [15,16]. This extraordinary antimalarial

otency of these alkaloids has made manzamine one of the

portant structural compounds for developing novel antimalar-

l agents. A major limitation of this group of alkaloid is associated

oxicity with repeated doses in animal studies. Hence, it is essential

o understand the importance of each subunit and the effect of

arious substituents on antimalarial potency to develop novel

anzamine derivatives with potent antimalarial activity and a

etter therapeutic index. In the present review, we describe isola-

ion, antimalarial activity, cytotoxicity and antimalarial SAR of

arious natural and semi-synthetic manzamine derivatives.

anzamine alkaloidsanzamines are structurally complex natural alkaloids with a b-

arboline moiety attached to a pentacyclic diamine ring, having

ight- and 13-membered rings on a pyrrolo[2,3-i]isoquinoline

amework. The manzamine group of alkaloids exhibit a wide

ariety of biological activities such as anticancer, antimalarial,

ntileishmanial, anti-Alzheimer, antimicrobial, insecticidal, an-

i-inflammatory and anti-HIV/AIDS-associated opportunistic

fections [17].

Please cite this article in press as: Ashok, P. et al., Manzamine alkaloids: isolation, cytotoxicityj.drudis.2014.06.010

www.drugdiscoverytoday.com

Isolation of manzamine alkaloids and in vitro antimalarialactivityIn 1986, Higa and co-workers reported the isolation of a novel

antitumor alkaloid manzamine A from Okinawa sponge, genus

Haliclona, and the IC50 value was found to be 0.07 mg/ml against

P388 mouse leukemia cells [14]. The exceptional bioactivity and

complex unique structure of manzamine A made total synthesis of

this alkaloid an attractive but challenging task. First, total synthe-

sis of manzamine A was reported by Winkler and Axten in 1998

[18]. After more than a decade of its initial isolation, manzamine A

was reported for its antimalarial activity. Manzamine A (1) exhib-

ited potent in vitro antimalarial activity against D6 and W2 strains

of P. falciparum with IC50 values of 4.5 and 8.0 ng/ml, respectively

[19,20]. In 1987, Higa’s group reported isolation and structure of

two Q5new alkaloids, manzamine B (2) and C (3) from the genus

Halicona [21]. In subsequent studies, manzamine D (4) was isolated

as a minor component from different marine sponges. In 1988,

Ichiba et al. reported isolation, structure and antimalarial activity

of two new manzamine alkaloids, manzamine E (5) and F (6), from

Okinawan Xestospogia sp. [22]. They exhibited moderate antima-

larial activity against D6 (IC50: 3400 and 780 ng/ml, respectively)

and W2 (IC50: 4760 and 1700 ng/ml, respectively) strains of P.

falciparum [20]. Kondo et al. reported isolation of four novel

manzamine alkaloids, manzamine H (7), manzamine J (8), ircinal

A (9) and ircinal B (10), from Okinawan marine sponge Ircinia sp.

The structural correlation of ircinal alkaloids gave further insights

into the biogenetic pathway of manzamine alkaloids [23].

8-Hydroxymanzamine-A Q5(11) was isolated from a sponge, Pachy-

pellzna sp., in 1994 and its structure was confirmed by preparing 8-

methoxymanzmine-A (12). 8-Hydroxymanzamine-A exhibited

potent antimalarial activity against D6 and W2 strains of P.

falciparum (IC50: 6.0 and 8.0 ng/ml, respectively) [24]. Two 8-

hydroxymanzamine-A derivatives, 1,2,3,4-tetrahydro-2-N-meth-

yl-8-hydroxymanzamine-A (13) and 1,2,3,4-tetrahydro-8-hydro-

xymanzamine-A (14) were also isolated from two different

haplosclerid sponges of the genera Petrosia and Cribochalina by

Crews et al. [25]. In the same year, Kobayashi et al. reported

isolation of new manzamine alkaloids, manzamine Y (15) and

3,4-dihydromanzamine-A (16), from Okinawan marine sponge

Amphimedon sp. Among these alkaloids, manzamine Y showed

significant antimalarial activity against D6 and W2 strains of P.

falciparum (IC50: 420 and 850 ng/ml, respectively) [26,27]. In

subsequent studies on the same genus of the sponge, Tsuda

et al. reported keramaphidin C (17) and keramamine C (18) as

plausible biogenetic precursors of manzamine C [26,27]. Kobaya-

shi et al. also reported four novel manzamine alkaloids, xestoman-

zamine A (19), xestomanzamine B (20), manzamine X (21) and

manzamine Y (22), from Okinawan marine sponges of Xestospon-

gia sp. Among these, xestomanzamine B, manzamine X and man-

zamine Y exhibited weak cytotoxicity against KB cell lines [28]. An

unsymmetrical manzamine dimer, kauluamine (23) alkaloid was

isolated from Indonesian marine sponge Prianos sp. by Ohtani

et al., showing moderate immunosuppressive activity [29]. Tsuda

et al. reported isolation of a new manzamine alkaloid called

manzamine L (24). It showed moderate cytotoxicity against KB

and L I210 cell lines [30]. Four new manzamine alkaloids, 6-

deoxymanzamine-X (25), manzamine J N-oxide (26), 3,4-dihydro-

manzamine-A N-oxide (27) and manzamine A N-oxide (28), were

, antimalarial activity and SAR studies, Drug Discov Today (2014), http://dx.doi.org/10.1016/



6


DRUDIS 1438 1–10


isolated from Philippine marine sponge Xestospongia ashmorica by

Edrada et al. in 1996. These alkaloids exhibited moderate cytotox-

icity against L5178y cell lines [31].

In 1998, Watanabe et al. reported isolation of manzamine M

(29), 3,4-dihydromanzamine-J (30) and 3,4-dihydro-6-hydroxy-

manzamine-A (31) from the Okinawan marine sponge Amphime-

don sp. These newly isolated manzamines exhibited moderate

cytotoxicity against L1210 cell lines [32]. In the same year, man-

zamine alkaloid maeganedin A (32), with a tetrahydro-b-carboline

scaffold having a methylene carbon bridge between N-2 and N-27,

was also isolated from an Okinawan marine sponge Amphimedon

sp. Maeganedin A exhibited moderate cytotoxicity against L1210

cell lines [33]. Zhou et al. reported two new manzamine alkaloids,

epimanzamine D (33) and N-methyl epimanzamine D (34), from a

Palaun sponge. Epimanzamine D and N-methyl epimanzamine D

exhibited moderate cytotoxicity against HeLa cell lines [34]. El

Sayed et al. reported the isolation and antiparasitic activity of

manzamine alkaloids ent-8-hydroxymanzamine-A (35), ent-man-

zamine-F (36) and unprecedented manzamine dimer neo-kaulua-

mine (37) from genus of Prianos sp. In vivo antimalarial potency of

these alkaloids will be discussed below [35]. In 2002, Yousaf et al.

reported the isolation of three new manzamine alkaloids, ent-

12,34-oxamanzamine-E (38), ent-12,34-oxamanzamine-F (39)

and 12,34-oxamanzamine-A (40), from three Indo-Pacific sponges.

Among these alkaloids, ent-12,34-oxamanzamine-F and 12,34-

oxamanzamine-A exhibited moderate (IC50: 840 ng/ml) and weak

(IC50: 4760 ng/ml) antimalarial activity against a P. falciparum D6

clone [36]. In 2003, Hamann’s group isolated five new manzamine

alkaloids: 32,33-dihydro-31-hydroxymanzamine-A (41), 32,33-

dihydro-6-hydroxymanzamine-A-35-one (42), des-N-methylxes-

tomanzamine-A (43), 32,33-dihydro-6,31-dihydroxymanzamine-

A (44) and 1,2,3,4-tetrahydronorharman-1-one (45), along with

six known manzamine alkaloids from an Indonesian sponge. But

none of these alkaloids exhibited antimalarial activity and cyto-

toxicity at tested concentrations [37].

Yousaf et al. reported the isolation of three new manzamine

alkaloids, 12,28-oxamanzamine-A (46), 12,28-oxa-8-hydroxyman-

zamine-A (47) and 31-keto-12,34-oxa-32,33-dihydroircinal-A (48),

from an undescribed species of the genus Acanthostrongylophora

[38]. Rao et al. reported isolation of three new manzamine alka-

loids, 12,34-oxamanzamine-E (49), 8-hydroxymanzamine-J (50)

and 6-hydroxymanzamine-E (51), along with 12 known manza-

mine alkaloids from an Indonesian sponge of the genus Acanthos-

trongylophora. Among these alkaloids, only 6-hydroxymanzamine-

E exhibited moderate in vitro antimalarial activity against P. falci-

parum D6 and W2 strains (IC50: 780 and 870 ng/ml, respectively)

[39]. In subsequent studies, Rao et al. reported isolation of four new

manzamine alkaloids, 12,28-oxamanzamine-E (52), 12,34-oxa-6-

hydroxymanzamine-E (53), 8-hydroxymanzamine-B (54) and

12,28-oxaircinal A (55), from Acanthostrongylophora sp. None of

these alkaloids showed antimalarial activity at the tested concen-

tration [20]. Yamada et al. reported isolation of three new manza-

mine alkaloids, zamamidine C (56), 3,4-dihydro-6-hydroxy-10,11-

epoxymanzamine-A (57) and 3,4-dihydromanzamine-J N-oxide

(58), from an Okinawan marine sponge Amphimedon sp. Zamami-

dine C is a new manzamine alkaloid possessing a second b-carboline

ring connected at N-2 of manzamine D through an ethylene linker.

3,4-Dihydro-6-hydroxy-10,11-epoxymanzamine-A is the first


manzamine alkaloid to possess an epoxide ring at C-10 and C-11.

Zamamidine C displayed moderate activity whereas 3,4-dihydro-

manzamine-J N-oxide showed weak activity against a resistant W2

strain of P. falciparum (IC50: 580 and 7020 ng/ml, respectively)

(Table 1) [40].

In vivo antimalarial activity of natural manzaminealkaloidsManzamine A and 8-hydroxymanzamine-A exhibited potent in

vitro and in vivo antimalarial activity against Plasmodium berghei.

More than 90% inhibition of asexual erythrocytic stages of P.

beighei was observed after a single intraperitoneal injection of

manzamine A and 8-hydroxymanzamine-A into infected mice.

Survival time of infected mice was also increased to more than ten

days with single doses of manzamine A (50 mM/kg) and 8-hydro-

xymanzamine-A (100 mM/kg), when compared with standard

drugs artemisinin and CQ (4 and 6 days, respectively, at

100 mM/kg). Among the five mice treated with 100 mM/kg of

manzamine A, two mice were able to clear the parasitemia

completely. Oral administration of manzamine A with a dose of

100 mM/kg also produced significant reductions in parasitemia,

indicating the good pharmacokinetic properties of these alkaloids

[19]. In 2001, El Sayed et al. reported in vivo antimalarial potency of

ent-8-hydroxymanzamine-A and neo-kauluamine against P. ber-

ghei. The average survival time of P. berghei infected mice was

increased to 9–12 days with single dose (100 mM/kg) of ent-8-

hydroxymanzamine-A and neo-kauluamine when compared with

untreated controls (2–3 days). Three 50 mM/kg i.p. doses of ent-8-

hydroxymanzamine-A completely clear the parasitemia without

any toxicity [35].

Owing to the potent antimalarial activity at nanomolar con-

centrations, manzamines are one of the important structural

targets for developing novel potent antimalarial agents. In the

literature, a few attempts have been made to develop manzamine

derivatives as potent antimalarial agents with improved therapeu-

tic index. Winkler et al. reported some simplified analogs of

manzamine A (59–62; Fig. 1) to find out the role rings QA and D

have on relative stereochemistry and orientation of the b-carbo-

line nucleus on antimalarial potency. Among these synthetic

analogs, compound 59, which has the same relative stereochem-

istry and b-carboline ring orientation (C-10) to that of manzamine

A, exhibited the most potent antimalarial activity (IC50: 300 and

310 ng/ml against D6 and W2 strains of P. falciparum, respective-

ly), but all these new synthetic analogs are significantly less potent

than manzamine A. These results emphasize the importance of the

A and D rings and orientation of the b-carboline ring on antima-

larial activity of manzamine A [41]. In a subsequent study by the

same group, antimalarial activity of new synthetic manzamine

analogs (63–67; Fig. 1) consisting of monocyclic (B), bicyclic (AB

and BC) and tetracyclic (ABCE) rings with a b-carboline skeleton

was discussed. The simplified manzamine analogs displayed dras-

tically reduced antimalarial activity and a relatively narrow range

of differences in their antimalarial activity [42]. With their con-

tinuous interest in developing potent antimalarial agents, Winkler

et al. reported the antimalarial activity of new manzamine A

analogs (68–70; Fig. 1) lacking either or both of the D and E rings.

These new manzamine analogs exhibited significantly reduced

antimalarial activity. These results suggested that simplified


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DRUDIS 1438 1–10

Please cite this article in press as: Ashok, P. et al., Manzamine alkaloids: isolation, cytotoxicity, antimalarial activity and SAR studies, Drug Discov Today (2014), http://dx.doi.org/10.1016/j.drudis.2014.06.010

TABLE 1

In vitro antimalarial activity ofQ12 manzamine alkaloids

Compound

number

Structure IC50 ng/ml (nM) Cytotoxicity

IC50 (mg/ml)

(D6 clone) (W2 clone)

1

NH

N

N

OH

N

4.5 (8.2) 8 (14.6) 0.07

5

NH

N

N

OH

N

O

3400 (6028.4) 4760 (8439.7) >4.7

6

NH

N

N

OH

N

OH

O

780 (1344.8) 1700 (2931.0) >4.7

8

NH

N

N

OH

HN

H

1300 (2363.6) 750 (1363.6) >4.7

9

NOH

N

H

H

H

CHO 2400 (5853.7) 3100 (7561.0) 4.8

11

NH

N

N

OH

N

OH

6 (10.6) 8 (14.1) 1.1

15

NH

N

N

OH

N

HO420 (744.6) 850 (1507.1) 3.9

4 www.drugdiscoverytoday.com

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DRUDIS 1438 1–10


TABLE 1 (Continued )

Compoundnumber

Structure IC50 ng/ml (nM) CytotoxicityIC50 (mg/ml)


21

NH

N

N

OH

N

O

HO 950 (1637.9) 2000 (3448.3) >4.7

25

NH

N

N

OH

N

O

1300 (2305.0) 1400 (2482.3) 4.7

27

NH

N

N

OH

N

O1600 (2826.9) 3700 (6537.1) >4.7

28

NH

N

N

OH

N

O11 (19.5) 13 (23.0) 4.2

37

NH

N

N

HO

N

HN

N

N

HO

N

OH

OO

HO

HH

HH

1700 (1465.5) 2800 (2413.8) >4.7

39

NH

N

NN

O

O

OH

840 (1453.3) 1100 (1903.1) >4.7





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DRUDIS 1438 1–10

TABLE 1 (Continued )

Compound

number

Structure IC50 ng/ml (nM) Cytotoxicity

IC50 (mg/ml)


40

NH

N

NN

O

HH

4760 (8608.1) NA >4.7

51

NH

N

N

OH

N

O

HO780 (1344.8) 870 (1500.0) 4.3

56

NH

N

N OH

N

HHN

HN

H

– 580 (777.4) 13.6

58

NH

N

NOH

HN

HHO

– 7020 (1231.5) 9.0

Abbreviation: NA,Q13 not available.

6

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tructural analogs of manzamine A might not serve as useful leads

r development of new antimalarial agents [43].

Shilabin et al. reported the antimalarial activity of 8-hydroxy-

anzamine-A derivatives (12,71,72; Fig. 1). Among these analogs,

-acetylmanzamine-A (71; Fig. 1) and 8-methoxymanzamine-A

2; Fig. 1) exhibited significant antimalarial activity (IC50: 9.6,

0.0 and 37.0, 47.0 ng/ml, respectively, against D6 and W2

trains), comparable with 8-hydroxymanzamine-A with increased

ytotoxicity. Whereas, 8,12-diacetylmanzamine-A (72; Fig. 1) dis-

layed least antimalarial potency (IC50: 1300 and 1200 ng/ml

gainst D6 and W2 strains, respectively) with no cytotoxicity. In

ivo antimalarial potency of 8-acetylmanzamine-A at doses of 16

nd 49 mM/kg � 3 (oral) was evaluated and exhibited potent

ntimalarial activity at 49 mM/kg, similar to 8-hydroxymanza-

ine-A, but toxicity was observed at 49 mM/kg � 3 doses after

4 days of oral administration [44]. Ibrahim et al. reported



antimalarial potency of N-methylmanzamine-A derivatives: 2-N-

methylmanzamine-A trifluoromethanesulfonate, 2-N,12-O-

dimethylmanzamine-A trifluoromethanesulfonate and 9-N-

methylmanzamine-A (73–75, respectively; Fig. 1). Of these ana-

logs, 2-N-methylmanzamine-A trifluoromethanesulfonate

showed moderate antimalarial activity against D6 (IC50: 736 ng/

ml) and weak activity against W2 (IC50: 1011 ng/ml) strains with a

comparable selectivity index to manzamine A. 12-O-Methylation

and 9-N-methylation derivatives of manzamine A were devoid of

antimalarial activity [45].

Wabha et al. reported antimalarial activity of manzamine A

derivatives with amidation on position 6 and 8 of the b-carboline

moiety. Twenty new manzamine A derivatives (76a–j to 77a–j;

Fig. 1) were synthesized and evaluated for their antimalarial

activity. Some of these manzamine A derivatives exhibited potent

antimalarial activity against D6 and W2 strains without any





DRUDIS 1438 1–10


N

N

N

N N

N

N

N

N

N N

N

N

N N

N N N

N

OHOH

OH

N

NN

N

N

N

Manzamine A (1)

OH

A

A

A

B

B B

B

C

E

BC

C

D

E

H

NH N

H

NH

NH

N

N

N

N

N

N

N

N

N

N

N

N

N

H NH

NH

NH

H

H

H

NH

NH

NH

NH

NN

N

N

N

N

N

N

NH

N

RO

R

R

70

71: R = OCH3, R 1 = H 73: R = CH3, R1, R2 = H

78: R = NO2, R1 = H

NNH2

CH2CH 379: R = OCH3, R1 = H80: R = H, R1 = COOCH3

74: R, R1 = CH3, R2 = H75: R, R1 = H, R2 = CH3

12: R = CH3, R 1 = H72: R = OCH3, R 1 = OCH3

OH

OHOH

OHOH

HO HO

HO

N

N

76a-ja

77a-ja 81 82

+ N

O

RR

CF3SO 3

OR1OR1

R1

R2

H

NH

NH

NH

H

H

H

H

H H H

H

H

H

H

H

HHNPr

PrN

PrN

6467 68 69

Cisisomer: 65Trans isomer: 66

H

H

H

HH

HN

HH

H

HH

59 60 61 62 63

HN

O

Drug Discovery Today

FIGURE 1

Structures of manzamineQ10 A derivatives. (a) R = CH3, (b) R = CH2CH3, (c) R = CH2CH2CH3, (d) R = CH(CH3)2, (e) R = CH2(CH2)2CH3, (f) R = CH(CH3)3, (g)R = CH2(CH2)3CH3, (h) R = CH2(CH2)4CH3, (i) R = CH2CH(CH3)3, (j) R = cyclohexyl.





c

a

5

w

3

T

a

p

d

r

d

a

d

m

m

h

z

A

a

a

r

t

m

v

MT

t


DRUDIS 1438 1–10

TABLE 2

In vitro antimalarial activity of synthetic manzamine AQ15 analogs

Compound number IC50 ng/ml (nM) Compound number IC50 ng/ml (nM)

D6 clone W2 clone D6 clone W2 clone

59 300 (813) 310 (840) 76f 231 (357) 417 (644)

60 640 (1734) 630 (1707) 76g 680 (1028) 786 (1189)

61 1800 (4878) 2500 (6775) 76h 696 (1031) 754 (1117)

62 1500 (4065) 1300 (3523) 76i 211 (326) 302 (466)

63 3510 (14153) 5550 (22379) 76j 34 (50) 53 (78)

64 920 (2779) 4190 (12658) 77a 182 (300) 231 (381)

65 930 (2695) 1270 (3681) 77b 123 (198) 87 (140)

66 1020 (2956) 2770 (8029) 77c 35 (55) 121 (191)

67 270 (579) 520 (1115) 77d 158 (249) 205 (323)

68 190 (329) – 77e 53 (81) 49 (75)

69 130 (225) – 77f 170 (262) 232 (358)

70 1290 (2135) – 77g 32 (48) 65 (98)

71 9.6 (15) 30 (49) 77h 55 (81) 49 (72)

12 37 (64) 47 (81) 77i 139 (214) 127 (196)

72 1300 (2006) 1200 (1851) 77j 950 (1411) 995 (1478)

73 736 (1307) 1011 (1795) 78 18 (30) 28 (47)

76a 1288 (2128) 1982 (3276) 79 28 (48) 58 (100)

76b 1162 (1874) 1937 (3129) 80 11 (17) 15 (24)

76c 1152 (1816) 1894 (2992) 81 29 (48) 38 (63)

76d 1105 (1745) 1547 (2443) 82 77 (126) 86 (140)

76e 1235 (1908) 1452Q15 (2244)

8

Review

s�P

OSTSCREEN

ytotoxicity. SAR analysis indicated that bulky and cyclic groups

djacent to the amide carbonyl at position 6 (76j; IC50: 34 and

3 ng/ml, respectively) are favored for antimalarial activity,

hereas small and linear alkyl groups at position 8 (77 g; IC50:

2 and 65 ng/ml, respectively) provided the most active amides.

wo potent compounds (76j and 77 g) were screened for in vivo

ntimalarial potency and showed moderate 24% and 62% sup-

ression of parasitemia with oral doses of 44 and 45 mM/kg (once

aily for three days), with no apparent toxicity [46]. Peng et al.

eported antimalarial activity of manzamine A semi-synthetic

erivatives, with substitutions such as hydroxyl, nitro, alkyl and

cetyl groups on the b-carboline ring. Manzamine F is a carbonyl

erivative of 8-hydroxymanzamine-A and exhibits marginal anti-

alarial activity. Hence, manzamine F derivatives were prepared

ainly by modifying the carbonyl functionality into hydroxyl,

ydrazone and alkyl groups. Among the reported synthetic man-

amine derivatives, 6-nitromanzamine-A, 6-methoxymanzamine-

, manzamine A 3-methyl ester, manzamine-F-31-hydrazone and

lkylated manzamine F (78–82; Fig. 1) showed potent antimalarial

ctivity (IC50: 18, 28, 11, 29, 77 and 28, 58, 15, 38, 86 ng/ml,

espectively, against D6 and W2 strains) with no cytotoxicity at

he tested concentration except for 6-methoxymanzamine-A and

anzamine A 3-methyl ester, which displayed cytotoxicity against

ero cells at 0.5 and 0.2 mg/ml, respectively (Table 2) [47].

echanism of actionhe exact mechanism of action of manzamine alkaloids regarding

heir antimalarial activity is not completely defined. But, some



literature states that b-carboline alkaloids inhibit DNA synthesis

through intercalation of DNA base pairs. Therefore, it is hypothe-

sized that compounds containing a b-carboline moiety hamper

the growth of parasites by inhibiting parasitic DNA synthesis [48–

50].

Structure–activity relationshipUndoubtedly, extraordinary antimalarial potency of manzamines

means they are excellent structural targets for developing novel

antimalarial agents, but cytotoxicity is their major drawback.

Hence, a thorough SAR study (Fig. 2) of manzamine alkaloids is

required to understand the importance of each moiety (b-carbo-

line and pentacyclic ring) and the effect of different substituents

on antimalarial activity. Extensive SAR will be useful for the

development of potent antimalarial manzamine derivatives with

better pharmacokinetic properties. After thorough perusal of lit-

erature reports, we have concluded SAR of manzamines for their

antimalarial activity. For easy understanding, we divided SAR into

two objectives, one is the effect of various substitutions on the b-

carboline nucleus and the other is the effect of substitutions on the

pentacyclic ring.

Effect of substitutions on b-carboline nucleus. Iricinal A

is completely devoid of antimalarial activity indicating that the b-

carboline moiety of manzamine alkaloids is responsible for anti-

malarial activity. 9-N Alkylation of the b-carboline ring resulted in

decreased antimalarial activity, indicating that 9-NH is required for

their antimalarial activity. Electron-donating hydroxyl-group sub-

stitution at position 8 of the b-carboline skeleton does not have





DRUDIS 1438 1–10

Electron withdrawing groupsfovors

Electron donating groupsfovors

Ester group favors to retainantimalarial potency

Essential for antimalarial activity

Selective reductions may favorantimalarial activity

Manzamine A

OH

N

N

N

NH

Double bond and electrondonating groups favors

Drug Discovery Today

FIGURE 2

Favorable positions on manzamine A forQ11 antimalarial potency.


any significant effect on its antimalarial activity (IC50: 6.0 ng/ml)

but cytotoxicity was reduced comparatively. Methoxy substitution

at position 8 resulted in a marginal decrease in antimalarial

potency (IC50: 37 ng/ml), whereas nitro-group substitution at

position 8 significantly decreased the antimalarial potency (IC50

310 ng/ml). Nitro and methoxy groups at position 6 slightly

affected the antimalarial potency with decreased IC50 values (18

and 28 ng/ml). The antimalarial activity was retained upon intro-

duction of a methyl ester group at position 3 of the b-carboline

skeleton (IC50: 11 ng/ml). Conformational planarity and orienta-

tion of b-carboline skeleton also play an important part in anti-

malarial potency of manzamines. Change in conformational

orientation and planarity alteration of the b-carboline skeleton

with reduction of pyridine to piperidine and 2-N-methylation

caused significant reduction in antimalarial potency of manza-

mines. Amide substitution on position 8 and 6 mainly affected

antimalarial potency with significant reduction in cytotoxicity.

A bulky alkyl group (cyclohexyl) adjacent to amide carbonyl

carbon at position 6 (19j) and a linear, short alkyl group adja-

cent to amide carbonyl carbon at position 8 (20 g) are favorable

for antimalarial activity. The 2-N-oxide derivative of manza-

mine A retained its antimalarial potency (IC50: 11 ng/ml),

whereas 2-N-methylation of manzamine A resulted in decreased

antimalarial potency (IC50: 700 and 1000 ng/ml) against D6 and

W2 strains, respectively.

Effect of substitutions on pentacyclic ring. The iricinal

subunit is important for antimalarial activity of manzamines, and

opening or removal of these rings drastically affected the antima-

larial potency. In addition to this, the hydroxyl group at C-12 is

indispensable for antimalarial potency of manzamines. Conver-

sion of the hydroxyl group to ether functionality severely reduced

the antimalarial potency, even some derivatives like 12,28-oxa-

manzamine-A, 12,28-oxa-8-hydroxymanzamine-A and 2-N,12-O-

dimethylmanzamine-A trifluoromethanesulfonate are devoid of


antimalarial activity. Manzamine F is very much structurally

related to potent antimalarial manzamine alkaloid 8-hydroxy-

manzamine-A, with a carbonyl group at C-31 and reduced C-32

double bond, exhibiting significantly reduced antimalarial poten-

cy. Modification of the C-31 carbonyl group to hydrazone and

alkylation greatly enhanced the antimalarial potency (IC50: 29 and

77 ng/ml, respectively) of manzamine F, demonstrating that C-31

carbonyl functionality is not favorable for antimalarial activity.

Simple reduction of the C-31 carbonyl group to alcohol and intro-

duction of a double bond in conjugation with the carbonyl group

did not improve the antimalarial potency. A double bond at C-31 in

the eight-membered ring is required to maintain integrity of the ring

and thereby plays an important part in contributing antimalarial

activity. Reduction of the double bond at C-31 affects integrity of the

ring, resulting in significant reduction in antimalarial potency (IC50:

200 ng/ml) and successive second reduction of the double bond at

C-15 increases the antimalarial potency (IC50: 82 ng/ml). Hence,

selective reduction of the C-15 double bond could enhance antima-

larial potency of manzamine derivatives.

Concluding remarksIn the present review, we presented the antimalarial potency

of manzamine alkaloids. Undoubtedly, manzamines are one of

the important structural targets for the development of novel

antimalarial agents. Among these manzamine alkaloids, manza-

mine A, 8-hydroxymanzamine-A and manzamine-A-N-oxide

exhibited superior antimalarial activity to some of the available

antimalarial drugs. In vivo cytotoxicity with frequent doses of

manzamine is the major limitation of this group of alkaloids.

Hence, necessary modifications are needed to reduce the cytotox-

icity as well as to increase the antimalarial potency. In the present

study, we have drawn up a detailed antimalarial SAR of manza-

mine alkaloids based on the available literature on antimalarial

potency of manzamines. This detailed SAR could be helpful in the





d

t

CT

R

1

1

1

1

1

1

Q91

1

1

1

2

2

2

2

2

2

2

2


DRUDIS 1438 1–10

1

Review

s�P

OSTSCREEN

evelopment of novel synthetic manzamine analogs having po-

ent antimalarial activity with an improved therapeutic index.

onflicts of interesthe authors declare no conflicts of interest.


0 www.drugdiscoverytoday.com

AcknowledgmentsWe acknowledge BITS-Pilani for supporting this work. One of the

authors, P.A. acknowledges the financial support from Q8Council of

Scientific and Industrial Research, New Delhi, in the form of a

Senior Research Fellowship.

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manzamine alkaloids: isolation, cytotoxicity, antimalarial activity and sar studies

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