Experimental infection and protection against ... - TI Pharma

198 Chapter 10
procedures described in [10]) or an estimated ten year risk of >5% of developing
a cardiac event as estimated by the Systematic Coronary Evaluation (SCORE)
system. All volunteers gave written informed consent prior to inclusion. The trial
was performed in accordance with Good Clinical Practice and approved by the
Central Committee for Research Involving Human Subjects of The Netherlands
(CCMO NL24193.091.09). Clinicaltrials.gov identifier: NCT00757887.
Procedures
Both the six immunised, previously immune individuals, and the five malarianaive
control volunteers were exposed to the bites of five Anopheles stephensi
mosquitoes infected with the NF54 strain of Pf for ten minutes. Infected
mosquitoes were obtained by feeding on gametocytes of NF54 as described
previously [11]. The salivary glands of all blood-engorged mosquitoes were
dissected, confirming the presence of sporozoites in 97%, with a mean of 88,000
sporozoites per mosquito. When necessary, feeding sessions were repeated with
a smaller number of mosquitoes until precisely five infected mosquitoes had
bitten. (Five volunteers required one blood feeding session, five volunteers two
sessions and one volunteer three sessions, raw data provided in Table 1).
Starting from day five post-challenge infection, volunteers were subjected to an
intense follow-up regime with multiple daily visits to our out-patient clinical
research department. All signs and symptoms (solicited and unsolicited) were
recorded and graded by the attending physician as follows: mild (easily
tolerated), moderate (interferes with normal activity), or severe (prevents
normal activity), or in case of fever grade 1 (>37.5°C – 38.0°C), grade 2 (>38.0°C –
39.0°C) or grade 3 (>39.0°C). Haematological and biochemical parameters were
monitored daily. Because of a previously reported serious cardiac adverse event
after a malaria challenge infection in a separate study [12], particular attention
was paid to markers of coagulation or cardiac damage with daily follow-up of
highly-sensitive troponin, platelets, d-dimer and lactate dehydrogenase during
the period when blood stage parasitemia could be expected. Whenever
abnormal, blood samples were checked for the presence of fragmentocytes and
von Willebrand cleaving protease activity, since previous reports have
highlighted the occurrence of von Willebrand factor activation in experimental
malaria [13].
As soon as a blood-slide was found to contain parasites, volunteers were treated
with a curative regimen of atovaquone/proguanil 1000/400mg once daily for
three days. Volunteers who remained free of parasites by blood-slide until day