RASOANAIVO Bioactive compounds to antimalarial plants

Bioresources Towards
Drug Discovery and Development

University of Mauritius, Mauritius
University of Delhi, India

Bioactive compounds
Based on an integrated approach
to antimalarial plants

South-North collaborative work

Philippe Rasoanaivo
Malaria Laboratory
Institut Malgache de Recherches Appliquées

Ethnomedical knowledge of plants:
Source of useful bioactive compounds

Traditional health practitioners
Symptoms

Treatments

Diagnosis

Integrated approach to antimalarial plants
- Synergy
- Preventive
- Cytotoxic
- Antiplasmodial

Chemosensitizers
Prophylactic
Anticancer agents
True antimalarials

Direct effects on parasite viability

ANTIMALARIAL PLANTS
Effects on host-parasite relationships
- Immunostimulants
- Antipyretic
- Unexpected mechanisms

Experimental techniques
Erythrocytic stage

In vitro isotopic microtest
In vivo 4 day suppressive test

Antiplasmodial
Liver stage

Cytotoxicity
Chemosensitization

Immunomodulation

Human hepatocytes
parasitised with sporozoites
of NF54 Pf
P388 cell viability
Isobologram method

Proliferation of splenocytes

Methodology
Plant extracts
In vitro antiplasmodial screening
If negative

If positive
Cytotoxicity

Liver stage

Bioassay-guided fractionation

Immunomodulating tests

Liver stage of Plasmodium malaria
Strychnopsis thouarsii
(Menispermaceae)

OCH3

H3CO

N
H3C

N

OCH3

1

1'

O

CH3

O
OCH3

Limacine & fangchinoline
S. Ratsimamanga, P. Rasoanaivo et al.
(1992) Planta Medica, 58, 485.

Aqueous phase
Active against liver schizonts

New alkaloid
IC50 = 0,5 µg/ml

Cytotoxic versus antiplasmodial
Nuxia genus (Loganiaceae)

Betulinic acid derivatives
IC50 = 1.2 µg/ml

Antiplasmodial versus chemosensitizers
Dalbergia louvelli
EtOAc extract: ~ 2 µg/ml
One fraction : ~ 50 ng/ml
IC50 = 5.8 µg/ml

N. Beldjoudi, L. Mambu, P. Grellier, D. Ramanitrahasimbola, P. Rasoanaivo,
M.-T. Martin & F. Frappier (2003). J. Nat. Prod., 66 (11), 1447-1450.

Chemosensitizers
O
N

CH3

N

O

CH3

O

CH3

Strychnobrasiline

N

CH3

CO2CH3

N
Ac

H
O

Malagashanine
P. Rasoanaivo, S. Ratsimamanga-Urverg, R. Milijaona, H. Rafatro, C. Galeffi,
M. Nicoletti. Planta Medica, 60, 13 (1994).

Oxydation of strychnobrasiline
O

O

N CH3

N CH3

N

Ac

O

N
H

CH3

O

O
O

Peracid/CH2Cl2

N

CH3

N

Me

CH3

N
O

O

N
O

O

O

H3CO

F. Trigalo, M. T. Martin, A. Blond, B. Rasolondratovo, P. Rasoanaivo, F. Frappier.
Tetrahedron, 58, 4555-58 (2002).

Hémisynthesis of malagashanine
O
N CH3
N

Ac

O

N

CH3

N

CO2CH3

N
Ac

6 steps

CH3

O

N

CH3

CO2CH3 +

N
H

H

CH3

CO2CH3

N

H

H

O

H
O

F. Trigalo, R. Joyeau, C.V. Pham, J.J. Youte, P. Rasoanaivo,
F. Frappier. Tetrahedron 2004 (in press).

1,4-diamino bioactive unit

O
N

N

N

CH3

3

Nb
O

21

N
Ac

CH3

Strychnobrasiline

N

CO2CH3

O

CH3

H

H

Ac

O

O
CH3

Spermostrychnine

N

Malagashanine
N

CH3
N

O
O

O
H3CO

N
H

N

CH3

N

CH3

CO2CH3

N
H
O

Hexahydromalagashanine

N
H

N

CH3

1-phenyl,4-amino unit and synthetic chemosensitizers
CN
MeO

Me

Me

N

N Me
Cl

N

MeO
MeO
Verapamil

N Me
H
Desipramine

S

OMe

N

Chloropromazine

N

Cl
HO

F

N

N Me
Me
Amitriptyline

N
Me
Cyproheptadine

CF3

F
Penfluridol

Unifying pharmacophore
in malaria drug reversal
N
N
N

S

(C)n
S = N, Phenyl
n>2

S

chemosensitizing pharmacophore and polyamines
S

NH2

H2N

S

Putrescine
H2N

N
H

NH2

Spermidine
CH3

NC
H3CO

N

Verapamil

H3CO

H2N

OCH3

OCH3

H
N
N
H

NH2

Spermine
P. Rasoanaivo, D. Ramanitrahasimbola, O. Rakotonandrasana, S. Ratsimamanga.
Biodiversity, traditional medicine and resistance modulators.
Ethnopharmacologia 2004 (in press).

Erythroxylum pervillei
and MDR in cancer
CH3
OCH3

OCH3

N

OCH3

H3CO

O
O

H3CO
O

NC
H3CO

H3CO

Pervilleine C

OCH3
O

CH3
N

Verapamil

OCH3

OCH3

B. Cui, G. L. Silva, D. Chavez, M. You, H.-B. Chai, P. Rasoanaivo, S. M. Lynn,
M. O’Neill, J. A. Lewis, J. M. Besterman, A. Monks, N. R. Farnsworth, G. A. Cordell,
J. M. Pezzuto and A. D. Kinghorn. Journal of Natural Products, 64(12) 1514 (2001).

Interestingly,
chloroquine has a 1,4-diamino unit
HN
Cl

N

N

HN

N

Chloroquine
Is the mechanism of chloroquine resistance and its reversal
linked with polyamines ?
Malagashanine

useful biochemical tool.

Mechanism of malagashanine reversal ?
CQ

Malagashanine
CQ

CQ

CQ

D. Ramanitrahasimbola et al. Effects of malagashanine on the chloroquine accumulation
in sensitive and resistant malaria parasites. Biochemical Pharmacology (submitted).

In summary:
Ethnomedical knowledge
Bioactive compounds
Leads

Drug development

Biochemical tool
Synthesis
New activities

Drug
mechanism

CONCLUSION
Broadening plant extracts screening
Anti-malaria screening
Anti-tuberculosis screening

Antiviral Screening

Team at Malaria Laboratory

Dr F. Frappier
Prof J. Brocard

Dr M. T. Martin

Prof M. Nicoletti

Sublime is Science whose aim is to preserve life

Financial support (1994-2004)
Agence Universitaire de la Francophonie
2 PAL+ Projects (French Ministry of Research)
Fonds de Solidarité Prioritaire
CAMPUS/CORUS project
IDRC project
3 IFS projects
Others coming