Antimalarial treatment and malaria transmission: insights from the field. Conférence de la 8ème édition du Cours international « Atelier Paludisme » - DJIMDE Abdoulaye

1. Atelier Paludisme
Institut Pasteur Antananarivo
14 – 20 Mars 2011
Antimalarial treatment and
malaria transmission:
insights from the field
Abdoulaye DJIMDE PharmD, PhD
Malaria Research and Training Center
University of Bamako, Mali
1

2. Molecular Biology
Immunology Genetics
• Drug Resistance
Policy PK
• Spread of drug
resistance
Pharmacogenomics
Host-Parasite-Vector
Transmission 2

3. Outline
1. Sulfadoxine-Pyrimethamine (SP) and
malaria transmission in the field
2. Impact of SP on P. falciparum
gametocytes infectivity in vitro
3. Artemisinin-based combinations and
malaria transmission in sub-Saharan Africa
Conclusion
3

4. P. falciparum life cycle

5. Current malaria control tools
• Artemisinin-based combination therapies (ACTs)
• Sulfadoxine-pyrimethamine recommended for IPT in
pregnant women
– contemplated for IPTi and IPTc
• Quinine for severe malaria
• Insecticide treated nets & indoor residual spray

6. Sulfadoxine-Pyrimethamine treatment
and malaria transmission in a setting of
high Sulfadoxine-Pyrimethamine efficacy
of Mali

7. Background
• Chloroquine efficacy decreasing
• Need alternative second line drug
• Prospective in vivo tests of CQ,
amodiaquine and SP
7

8. Study site
• Kolle: rural village; 2,500
people
• 55 Km South of Bamako
• P. falciparum malaria
endemic and seasonal
• Parasitemia:
– 40-50% dry season
(October-April)
– 70-85% rainy season
Kolle
(May-September).

9. Study design
• Open randomized drug efficacy trial
• Children 6 months – 5 years
• Three arms: Chloroquine, Amodiaquine, SP
• 28 days of follow up
• In vivo, in vitro, molecular and
pharmacokinetic studies
MIM Antimalarial Drug Resistance Network

10. In vivo SP resistance in Mali

11. Evolution of gametocyte carriage after SP
treatment
Beavogui et al., IJP 2010
11

12. Molecular markers of SP resistance
Adapted from P. Wang et al. :Molecular and Biochemical Parasitology 89 (1997)
161–177

13. Mutations in pre-treatment asexual
vs. post-treatment gametocytes
100
90
80
70 *
60 *
* * p<0.001
% 50 * *
40 Asexual
30 *
Gameto
20
10
0
le
59
51
le
8
7
10
43
ip
up
FR
FR
Tr
FR
PS
dr
H
H
FR
H
ua
H
D
D
D
D
Q
Beavogui et al., IJP 2010
13

14. Impact of large scale use of SP on
spread of drug resistance and
malaria transmission?

15. Methods
• Drug efficacy study
• Screening for gametocyte carriers
• Detect molecular markers of drug resistance
• Include gametocyte carriers aged 6 – 18 y.
• Direct feed starved F1 generation An. gambiae
• Maintain mosquitoes in lab for 8 days
• Presence and number of oocysts measured by
dissection
• Compare the infectivity of pre-treatment vs. post-SP
gametocytes to Anopheles gambiae
• Protocol approved by IRBs in Bamako & Maryland
15

16. 16

17. Direct feeding
17

18. 18

19. Infectivity of Post-treatment
Gametocytes
* P < 0.001
Beavogui et al., IJP 2010
19

20. Summary 1
• SP increase rate of gametocyte carriage
• Post-SP gametocytes carry SP- resistance
mutations
• Post-SP gametocytes were less
transmissible to Anopheles gambiae

21. Mechanism of decreased
infectivity of post-sulfadoxine-
pyrimethamine P. falciparum
gametocytes to anopheline
mosquitoes
21

22. Infectivity of Sulfadoxine-Pyrimethamine treated P. Falciparum
Gametocytes to Anopheline Mosquitoes

23. Gametocyte production
• Serum supplemented RPMI complete culture media
• Gas (O2, CO2 and Nitrogen) humidity (70 - 80%)
Candle Jar Shaker Tipper

24. Gametocytes follow up
Day 8 Day 14 Day 14
Stage II Stage V Exflagellation test

25. Standard Membrane feeding Assay
Feeding of 30
mosquitoes
With each sample

26. Sulfadoxine and Pyrimethamine Plasma concentrations in Mali
S
P
26

27. Drug concentrations used
Drugs Mean conc. Day 3 Day 7 Day 14
Sulfa - ug/ml 61 34 14
------ ------------ ----------- ------------ ---------
Pyr ng/ml 158 67 16

28. Experimental design
A = Gametocytogenesis, B = Gametocyte maturation, 28
Kone A, IJP, 2010
C = Mature gametocyte infectivity

29. Results
29

30. A. Induction of gametocytogenesis
NF 135 Stage II NF135 Stage V
30

31. B. NF 135 Gametocyte development
Controls Pyrimethamine- treated Sulfadoxine- treated
31
Kone A, IJP, 2010

32. C. Effect of S, P and SP on gameto
infectivity
32
Kone A, IJP, 2010

33. Effect of S, P and SP on oocyst density
33
Kone A, IJP, 2010

34. Effect of Day3 levels of S or SP on
mosquito survival
34
Kone A, IJP, 2010

35. Summary 2
• Sulfadoxine and pyrimethamine have complex effects on
the biology of gametocytogenesis.
• Induce differentiation into sexual forms
• Treatment of young gametocytes impaired their further
development into mature stage V gametocytes.
• Drug + mature gametocytes => decreased infectivity
• SP also kills vector A. stephensi
35

36. Artemisinine-based combination
therapies and Malaria transmission
in the field

37. ACTs and malaria transmission
• “A single intramuscular injection of 5 mg/kg artemisinin (to
gametocytemic rhesus monkeys) resulted in complete loss of
mosquito infectivity within 24 h of drug administration” Dutta
GP, et al. 1989
• “artemisinin derivatives reduced gametocyte carriage 18.5 fold”
and “were found to reduce the transmission potential of
falciparum malaria” Price RN et al.,1996
• “Artemesinin-based combination therapies (ACT) for
falciparum malaria reduce gametocyte carriage, and therefore
reduce transmission”.
37

38. ACTs and malaria transmission (2)
• NASBA study in Kenya “data suggest that the potential of
malaria transmission remains high even after treatment
with artemisinin combination therapy” Schneider P. et al,
IJP, 2006
• “An efficacious antimalarial regimen with no specific
gametocytocidal properties but a long prophylactic time
was estimated to be more effective at reducing
transmission than a short-acting ACT in the highest-
transmission setting”. Okell LC, PLoS Med, 2008
38

39. Objective
Measure the impact of AS/AQ, AS/SP
and AR-L on malaria transmission.

40. Day 28 Efficacy
non-Corrected vs. PCR Corrected
* P < 0.05
*

41. Evolution of gametocyte carriage by treatment arm on
follow up days
41

42. How infectious are the post-ACT
gametocytes?
42

43. Methods
• Drug efficacy study
• Screening for gametocyte carriers
• Detect molecular markers of drug resistance
• Include gametocyte carriers aged 6 – 18 y.
• Direct feed starved F1 generation An. gambiae
• Maintain mosquitoes in lab for 8 days
• Presence and number of oocysts measured by dissection
• Compare the infectivity of pre-treatment vs. post-SP
gametocytes to Anopheles gambiae
• Protocol approved by Ethics Committee of FMPOS
43

44. Study site
• Bougoula-Hameau: peri-
urban village; 5000 people
~400 Km South of Bamako
• P. falciparum malaria
hyper endemic
• No Insectaries around!!

45. Bougoula-Hameau, Sikasso, Mali
45

46. 46

47. 47

48. 48

49. Direct feeding
49

50. 50

51. Comparing Ctrl vs. all post-ttt Oocyst positive
45
40 ***
35
30 ***
25
Oocyst +
20 NS
15
10
5
0
l
P
L
Q
tr
SS
R
SA
C
A
A
A
Treatment arms
51

52. Summary 3
• ACTs decreased gametocyte carriage
HOWEVER
• Would all ACTs reduce malaria transmission in
high transmission settings?
52

53. Discussion
• “Overall, infectivity was about three-times higher for
direct feeding than for membrane feeding (p < 0.001)”
Diallo M., et al Malaria Journal 2008
• MFA “Blood cells were separated from plasma by
centrifugation and plasma was replaced by a similar
volume of normal human plasma known to sustain malaria
transmission. This step was performed to avoid the
possible interference (blocking or enhancing) antibodies to
gametocytes in the donor’s plasma”.
53

54. Conclusions
• Malaria eradication/elimination will require
new, safe and truely gametocytocidal drug
• Need more sensitive gametocyte assays that
could indicate gametocyte viability in
addition to prevalence and density.
• Field oriented studies need to be as close as
possible to natural conditions. Too much
cleaning of experimental design may yield
nice results but with little relevance to real
life. 54

55. Acknowledgements
• MRTC • Support
• Study sites & • Government of Mali
participants • MIM/TDR
• Pr. Doumbo O & • NIAID/NIH
MRTC staff • FIC/NIH
• Ministère Français de
• Nijmegen la Recherche (Pal +)
• Adrian Luty • Sanofi-Aventis
• Robert Sauerwein
55