efficacy of mefloquine and mefloquine–artesunate for the treatment of uncomplicated plasmodium...

Efficacy of mefloquine and mefloquine–artesunate for the

treatment of uncomplicated Plasmodium falciparum malaria

in the Amazon region of Bolivia

Juan Carlos Avila1, Rodolfo Villaroel1, Wilmer Marquino2, Jorge Zegarra3, Rene Mollinedo1

and Trenton K. Ruebush3

1 Programa Nacional de Vigilancia y Control de la Malaria, Ministerio de Salud Publica y Prevision Social, La Paz, Bolivia2 Instituto Nacional de Salud, Lima, Peru3 Centers for Disease Control and Prevention, US Naval Medical Research Center Detachment, Lima, Peru

Summary We assessed the efficacy of mefloquine monotherapy and mefloquine–artesunate (MQ–AS) combination

therapy for the treatment of Plasmodium falciparum malaria at four sites in the Bolivian Amazon region.

Patients with uncomplicated P. falciparum infections between 5 and 60 years of age were randomly

assigned to be treated with either MQ (15 mg/kg in a single oral dose) or MQ (15 mg/kg) plus AS

(4 mg/kg daily for 3 days). A total of 143 patients were enrolled and followed for 28 days. None of

the 73 patients who received MQ alone or the 70 patients who received MQ–AS combination therapy

had recurrences of parasitaemia during the 28-day follow-up period. Asexual parasite densities fell

significantly more rapidly and the proportion of patients with gametocytes was significantly lower on

days 7–28 in patients treated with MQ–AS than in those treated with MQ alone. All patients tolerated

the medications well. After this study, the Bolivian Ministry of Public Health changed its treatment

policy for uncomplicated P. falciparum malaria in the Amazon region to combination therapy with

MQ–AS to slow or prevent the development of resistance.

keywords Plasmodium falciparum, antimalarial drug resistance, mefloquine, artesunate, Bolivia

Introduction

During the 1990s, Bolivia experienced a major resurgence

of malaria, particularly in the Department of Pando and

the northern third of the Department of Beni, which make

up its Amazon region. The increase in transmission reached

its peak in 1998, with more than 42 000 cases reported

from this area. Twenty-seven percent of cases were caused

by Plasmodium falciparum and these two departments

accounted for more than 99% of all cases of P. falciparum

reported from Bolivia. Although the total number of

malaria cases reported from the Bolivian Amazon region

fell after 2000, the favourable environmental and climatic

conditions, together with an efficient vector, Anopheles

darlingi, and increasing resistance to antimalarial drugs,

have caused concern that malaria could re-emerge as a

major public health problem in this region.

Although little is known about resistance to antimalarial

drugs in the Bolivian Amazon region, two in vivo drug

efficacy studies in the late 1980s and early 1990s suggested

high levels of resistance of P. falciparum to both chloro-

quine and sulphadoxine-pyrimethamine (unpublished

data). Additional evidence of sulphadoxine-pyrimethamine

resistance was provided by a 1994 study that documented

high frequencies of triple and quadruple dihydrofolate

reductase (DHFR) mutations (Plowe et al. 1997). In 1991,

first-line therapy for uncomplicated P. falciparum infec-

tions was changed from sulphadoxine-pyrimethamine to a

7-day course of quinine (30 mg/kg/day in three divided

doses), although sulphadoxine-pyrimethamine continued

to be used in large quantities until the mid-1990s. Because

of poor compliance with the 7-day course of quinine, the

Ministry of Public Health decided to evaluate other drug

regimens more suitable for use in ambulatory patients.

Although mefloquine (MQ) might be considered the

most appropriate alternative in areas with high levels of

resistance to both chloroquine and sulphadoxine-

pyrimethamine, it is now recommended that MQ always

be used in combination with a second drug to delay the

development of resistance and prolong its useful thera-

peutic lifetime (White 1998; World Health Organization

1998). The drugs most commonly recommended for

such combination therapy with MQ are artemisinin

and its derivatives, such as artesunate (AS). Since the

Tropical Medicine and International Health

volume 9 no 2 pp 217–221 february 2004

ª 2004 Blackwell Publishing Ltd 217

National Malaria Control Program of Bolivia had no

previous experience with the use of any of the artemisinin

drugs or with artemisinin drug combination therapy, we

studied the efficacy of combination therapy with MQ plus

AS for uncomplicated P. falciparum infections in the

Bolivian Amazon region. MQ monotherapy was evaluated

at the same time to provide baseline information on the

efficacy that drug.

Materials and methods

The study was conducted at the Guayaramerin and

Riberalta Hospitals in the Department of Beni and in the

Puerto Rico and Porvenir Health Centers in the Depart-

ment of Pando in the Bolivian Amazon region (Figure 1)

during the peak malaria transmission season of 2001. All

four-health facilities draw their patient populations from

the urban and surrounding rural areas of these towns.

Malaria transmission in the Bolivian Amazon region is

unstable with a peak between February and May. All age

groups are affected. Plasmodium vivax is the predominant

species; P. falciparum accounts for 20–25% of infections.

Although the majority of infections are symptomatic,

severe malaria is quite uncommon. The principal vector is

Anopheles darlingi. Chloroquine, sulphadoxine-pyrimeth-

amine, and quinine are sold without prescription in local

pharmacies and shops; MQ and artemisinin drugs were not

available commercially at the time the study was carried out.

The study was approved by the institutional review

board of the US Navy. Guidelines of the Bolivian Ministry

of Public Health governing research involving human

subjects were followed. Our methods followed those

described previously (Marquino et al. 2003). The sample

size was calculated assuming an expected rate of treatment

failure with either MQ monotherapy or MQ–AS combi-

nation therapy of <5% in the study population, a precision

of 5%, and a 5% level of significance. Using a table of

random numbers, patients at each site were treated

with MQ alone or MQ–AS combination therapy. MQ

(Mephaquin�, Mepha Ltd, Aesch-Basel, Switzerland) was

administered in a single oral dose of 15 mg/kg on day 0.

Patients assigned to the MQ–AS group received the same

dose of MQ on day 0, together with AS (Plasmotrin�,

Mepha Ltd.) in a dose of 4 mg/kg daily on days 0, 1 and 2.

All drugs were administered by study staff and subjects

were observed for vomiting for 30 min after ingesting the

drugs. Patients were then followed up on days 1–3, 7, 14,

21 and 28.

Parasitological and therapeutic responses to treatment

were measured as defined by the World Health Organiza-

tion (Bruce-Chwatt et al. 1986; World Health Organiza-

tion 2002). Statistical analysis was carried out using SPSS

(SPSS Inc., Chicago, IL, USA). Dichotomous variables were

compared with chi-square or Fisher’s exact tests. The

Kolmogorov–Smirnov test was used to test for normality

of continuous variables and Student’s t-test or Mann–

Whitney U-test to compare mean values. Relative risk (RR)

was used to evaluate incidence rates.

Results

Of the 7146 patients with suspected malaria who were

screened during the 22-week enrolment period, 149 were

enrolled in the trial, 61 in Puerto Rico, 41 in Riberalta, 38

in Guayaramerin, and nine in Porvenir. Ninety-six (64%)

were males; their mean age was 28.4 ± 15.3 years. Eighty-

four (56.4%) of the subjects had a documented fever

(axillary temperature ‡37.5 �C) on enrolment and 87%

had a history of fever during the previous 48 h. The mean

duration of illness before enrolment was 4.1 ± 3.3 days.

Their geometric mean parasite density was 4,489/ll.

Of the 149 enrolled patients 143 (95.9%) completed

their 28-day follow-up. Six subjects were excluded from

analysis, three in the MQ group and three in the MQ–AS

group. Four of these subjects were lost to follow-up on day

3 (three patients) and day 14, and two had parasite

densities below 250/ll on re-examination of their day 0

blood smears.

PANDO

BRAZIL

BENI

BOLIVIA

PERU

CHILE

ARGENTINA

PARAGUAY

Guayaramerin

Riberalta

Porvenir

Puerto Rico

Figure 1 Study sites in the Departments of Beni and Pando in the

Amazon region of Bolivia.

Tropical Medicine and International Health volume 9 no 2 pp 217–221 february 2004

J. C. Avila et al. Mefloquine–artesunate for falciparum malaria in Bolivia

218 ª 2004 Blackwell Publishing Ltd

The characteristics of the 73 subjects who received MQ

monotherapy and the 70 who were treated with the

combination of MQ–AS are shown in Table 1. No signi-

ficant differences were observed on enrolment between

subjects enrolled at the four sites or between patients in the

two treatment groups, except that only one (12.5%) of the

patients enrolled in Porvenir had a documented fever on

enrolment compared with 58.5% of patients from the

other three sites.

All subjects responded well to therapy and by day 2, only

three (4.2%) of the patients in the MQ group and two

(2.9%) of patients in the MQ–AS group still had fever; by

day 3, all patients were afebrile. Asexual parasitaemia

density fell significantly faster in the subjects who received

combination therapy than in those treated with MQ alone.

On day 2, 39 (53.4%) of the patients treated with MQ had

negative blood smears compared with 55 (78.6%) of those

who had received MQ–AS [RR 1.47; 95% confidence

interval (CI) 1.15–1.88; P ¼ 0.002]. By day 3, 59 (80.8%)

of the patients treated with MQ had negative blood smears

compared with 64 (91.5%) of those treated with combi-

nation therapy, but this difference was not significant (RR

1.13; 95% CI 0.99–1.29; P ¼ 0.09). All patients were

aparasitaemic by day 7.

On enrolment, there was no significant difference in the

proportion of patients in the two treatment groups who

had gametocytes in their blood (33 or 45.2% vs. 29 or

41.4%), but the proportion of subjects with gametocytes

on days 7, 14, 21 and 28 was significantly lower with

combination therapy than with MQ alone (Figure 2).

Among patients who had no gametocytes in their blood

smears on enrolment, significantly fewer of those treated

with combination therapy than MQ alone had gameto-

cytaemia on day 7 (55.5% vs. 100.0%; P ¼ 0.009), on day

14 (27.8% vs. 83.3%; RR 4.33; 95% CI 1.18–15.86;

P ¼ 0.007), on day 21 (0% vs. 75.0%; RR 4.00; 95% CI

1.50–10.66; P ¼ 0.00001) and on day 28 (0% vs. 41.7%;

RR 1.71; 95% CI 1.06–2.77; P ¼ 0.005).

None of the subjects treated with either MQ monotherapy

or MQ–AS combination therapy who completed the trial

had a recurrence of parasitaemia during the 28 days after

treatment. All patients in both treatment groups were

classified as having Sensitive infections and Adequate

Clinical and Parasitological Responses. The four patients

who were lost to follow-up before day 28 had cleared their

parasitaemia before they dropped out of the trial.

No severe adverse drug reactions were observed. One

32-year-old subject vomited the first but not the second

dose of MQ. On day 1, four (5.5%) of the patients treated

with MQ complained of vomiting compared with eight

(11.6%) of those treated with MQ–AS (P ¼ 0.24). No

other subjects complained of new symptoms nor of an

increase in the severity of pre-existing symptoms after the

initiation of therapy.

Discussion

The rationale for using combination therapy for malaria is

similar to that for the treatment of tuberculosis, cancer and

HIV infections (White 1998). When used alone, drugs with

long half-lives and slow elimination from the blood, such

as MQ, are more likely to select resistant parasites. The

use of a rapidly acting and highly effective drug in combi-

nation with MQ, such as artemisinin or one of its derivatives,

Table 1 Characteristics of patients enrolled in mefloquine (MQ)

and MQ-artesunate (MQ–AS) in vivo drug efficacy trial,

Departments of Beni and Pando, Bolivia, 2001

Characteristic MQ (n ¼ 73) MQ–AS (n ¼ 70)

Mean age (years ± SE) 28.3 ± 15.4 28.4 ± 15.3

Gender (male) 64.4% 62.9%

History of fever 84.9% 90.0%Axillary temperature

‡37.5 �C (day 0)

56.2% 56.5%

Duration of fever (days ± SE) 4.3 ± 3.6 3.9 ± 3.1Geometric mean parasite

density (/ll) (day 0)

4595 4445

50

40

30

20

10

00 2 3 7 14 21 28

DayP

atie

nts

with

gam

etoc

ytem

ia (

%)

MQ

MQ-AS

Figure 2 Proportion of patients with gametocytes treated with

mefloquine (MQ) monotherapy or mefloquine plus artesunate

(MQ–AS) combination therapy. Points marked with an asterisk

indicate a P-value £ 0.05.




greatly reduces the probability of selecting parasites that are

resistant to both drugs. In Thailand, the addition of AS to

MQ therapy for P. falciparum infections has been associated

with a halt in the steady increase in MQ resistance that had

been observed when MQ monotherapy was the first-line

treatment (Nosten et al. 2000). Based on the premise that

combination therapy is the best way to prolong the useful

therapeutic lifetimes of antimalarial drugs and the Thai

experience, it is now generally recommended that antima-

larial drugs always be used in combination (White 1998;

World Health Organization 1998).

Mefloquine, either alone or in combination with an

artemisinin drug is increasingly being used for the treat-

ment of P. falciparum infections in the Brazilian Amazon

region (Ministerio da Saude 2001), and at the time this

study was being conducted, MQ monotherapy was the

most commonly prescribed first-line therapy in the area of

Brazil bordering Bolivia. In November 2001, Peru became

the first country in South America to introduce the

combination of MQ–AS as first-line therapy for uncom-

plicated P. falciparum malaria in its Amazon region

(Marquino et al. 2003). Although in vitro resistance to

strains of MQ has been reported from the Amazon Basin

since the early 1980s (de Souza 1983), only recently have

sporadic, but well-documented cases of RI in vivo resist-

ance to a single dose of 15 mg/kg been reported (Cardoso

et al. 1996; Cerutti et al. 1999). This study in the Bolivian

Amazon region showed no evidence of resistance to either

MQ or MQ–AS; however, at the time the study was carried

out, neither MQ nor artemisinin drugs were commercially

available in Bolivia. In addition, as has been reported

previously, we found that combination therapy with MQ–

AS reduced asexual parasite density and the proportion of

patients with gametocytaemia significantly faster than MQ

alone (Price et al. 1995, 1996; Marquino et al. 2003).

In areas with very low levels of resistance to MQ, it was

initially recommended by the World Health Organization

that a dose of 15 mg/kg of MQ should be used in

combination with an artemisinin drug (World Health

Organization 1998). Most authorities now believe, how-

ever, that even with combination therapy, MQ should

always be used at a dose of 25 mg/kg to reduce the risk of

selecting resistant organisms. This recommendation is

based on experiences from Thailand, where mathematical

modelling of pharmacokinetic and pharmacodynamic data

has demonstrated that the use of a 15 mg/kg dose of MQ

may increase the risk of selecting resistant mutations and

could lead more rapidly to resistance than a dose of

25 mg/kg (Simpson et al. 2000).

Different commercial preparations of MQ vary in terms

of their bioavailability. Two recent studies have showed

that maximum blood levels and the area under the curve

with Mephaquin� were significantly lower that those with

the reference compound, Lariam� (Weidekamm et al.

1998; Na-Bangchang et al. 2000). Thus, the effective dose

of MQ received by our subjects may have been even lower

than 15 mg/kg. If this is the case, strains of P. falciparum

from the Bolivian Amazon region would appear to be

highly sensitive to MQ, and combination therapy with MQ

at a dose of 25 mg/kg should offer excellent protection

against the selection of resistant strains.

With the objective of slowing or preventing the

development of antimalarial drug resistance, the Ministry

of Public Health of Bolivia recommended replacing quinine

with MQ plus AS as the new first-line therapy for

uncomplicated P. falciparum malaria in the Departments

of Beni and Pando in September 2001 (Ministerio de Salud

y Prevision Social 2002). This regimen was implemented in

late 2001 to early 2002 and is being used in all patients

except pregnant women, who will continue to be treated

with a 7-day course of quinine plus clindamycin. With this

change in treatment policy, Bolivia joins Peru as the first

countries in the Americas to implement combination

therapy with an artemisinin drug as first-line therapy for

P. falciparum malaria (Marquino et al. 2003). The efficacy

of the new combination therapy regimen will be monitored

every 2–3 years by in vivo drug efficacy testing.

Acknowledgements

The authors thank Gladys Nakao, Francisco Ramos, Hugo

Duran, Ruben Salazar, and Ademar Vidaurre Velasquez

for the microscopic diagnoses and Drs Ruben Torres Gil,

Armando Achocalla Chambi, Ricardo Arteaga, Angel

Daniel Caceres, and Alfredo Roda and the staff of the

Guayaramerin and Riberalta Hospitals and the Puerto Rico

and Porvenir Health Centers for their assistance with the

enrollment, treatment, and follow-up of patients. Authors

also acknowledge Drs Julio Alfred Cassab, Susan Brems,

Charles Oliver, German Guillen Vargas, Gerardo

Aramayo, and Cesar Reyes Parada, without whose help

and support this study would not have been possible. The

statistical analysis was carried out by Christian Bautista.

This study was supported by the USAID-Ministerio de

Salud y Prevision Social, Proyecto Integral de Salud

(PROSIN), contract no. 511.0644.02.

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Authors

Juan Carlos Avila, Rodolfo Villaroel and Rene Mollinedo, Programa Nacional de Vigilancia y Control de la Malaria,

Ministerio de Salud Publica y Deportes, Capitan Ravelo 3119, La Paz, Bolivia. Tel.: +591-3-855-3413; Fax: +591-3-855-3413;

E-mail: [email protected], [email protected]

Wilmer Marquino and Jorge Zegarra, Instituto Nacional de Salud, Capac Yupanqui 1400, Jesus Maria, Lima 11, Peru.

Tel.: +511-471-9920; Fax: +511-471-2529; E-mail: [email protected], [email protected]

Trenton K. Ruebush, Division of Parasitic Diseases (F-22), Centers for Disease Control and Prevention,

4770 Buford Highway, Atlanta, GA 30341, USA. Tel.: +1-770-488-3604; Fax: +1-770-488-4203; E-mail: [email protected]

(corresponding author).




efficacy of mefloquine and mefloquine–artesunate for the treatment of uncomplicated plasmodium...

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