Hyperparasitaemia and low dosing are an important source of anti-malarial drug resistance

Abstract

Background: Preventing the emergence of anti-malarial drug resistance is critical for the success of current malaria elimination efforts. Prevention strategies have focused predominantly on qualitative factors, such as choice of drugs, use of combinations and deployment of multiple first-line treatments. The importance of anti-malarial treatment dosing has been underappreciated. Treatment recommendations are often for the lowest doses that produce "satisfactory" results.

Methods: The probability of de-novo resistant malaria parasites surviving and transmitting depends on the relationship between their degree of resistance and the blood concentration profiles of the anti-malarial drug to which they are exposed. The conditions required for the in-vivo selection of de-novo emergent resistant malaria parasites were examined and relative probabilities assessed.

Results: Recrudescence is essential for the transmission of de-novo resistance. For rapidly eliminated anti-malarials high-grade resistance can arise from a single drug exposure, but low-grade resistance can arise only from repeated inadequate treatments. Resistance to artemisinins is, therefore, unlikely to emerge with single drug exposures. Hyperparasitaemic patients are an important source of de-novo anti-malarial drug resistance. Their parasite populations are larger, their control of the infection insufficient, and their rates of recrudescence following anti-malarial treatment are high. As use of substandard drugs, poor adherence, unusual pharmacokinetics, and inadequate immune responses are host characteristics, likely to pertain to each recurrence of infection, a small subgroup of patients provides the particular circumstances conducive to de-novo resistance selection and transmission.

Conclusion: Current dosing recommendations provide a resistance selection opportunity in those patients with low drug levels and high parasite burdens (often children or pregnant women). Patients with hyperparasitaemia who receive outpatient treatments provide the greatest risk of selecting de-novo resistant parasites. This emphasizes the importance of ensuring that only quality-assured anti-malarial combinations are used, that treatment doses are optimized on the basis of pharmacodynamic and pharmacokinetic assessments in the target populations, and that patients with heavy parasite burdens are identified and receive sufficient treatment to prevent recrudescence.

Figure 1

Different levels of anti-malarial drug resistance provide different probabilities of individual parasite survival (inset). The total numbers of parasites in an individual patient change with time. Treatment failure is defined as a failure to clear detectable parasites from the blood (R3, R2) or a subsequent recrudescence (R1). The former WHO classification of resistance translates into growth rates per cycle (PMF) as follows; R3; >0.25, R2; 0.1 to 0.25, R1a; 0.1 to 0.01, and R1b; <0.01. The inset shows the corresponding probabilities of an individual parasite surviving one cycle if exposed to therapeutic drug concentrations.

Figure 2

Total numbers of malaria parasites in the body of two adults (logarithmic scale; vertical axis) who develop similar hyperparasitaemias are shown. The days since acquiring the infection are shown on the horizontal axis. The pre-erythrocytic development is approximately 6 days. On the left A; the parasite multiplication factor (PMF) per asexual cycle is 25 representing highly efficient multiplication. On the right B; the multiplication factor is 6 per cycle, so it takes longer to develop hyperparasitaemia.

Figure 3

Total numbers of malaria parasites in the bodies of two adults (logarithmic scale; vertical axis) with similar rapidly expanding infections who develop hyperparasitaemia (>5% parasitaemia) are shown against time since their infecting mosquito bites on the horizontal axis. The horizontal dotted line is the pyrogenic density (the density threshold for fever). The most likely mitotic division to give rise to de-novo resistance is the one immediately preceding peak parasitaemia. The most likely (i.e. modal) number of resistant parasites present therefore is one. There are correspondingly lower probabilities of there being much larger numbers of de-novo resistant parasites. The inset triangles represent the much rarer situation of earlier de-novo appearance of resistant mutants and the corresponding greater numbers of resistant parasites that would occur at presentation with hyperparasitaemia. In the example on the left the fitness and thus multiplication factor of the mutant parasites is unaffected by the resistance mechanism, whereas on the right their fitness and thus multiplication is reduced relative to their drug sensitive siblings.

Figure 4

Approximate per-parasite probabilities of de-novo emergence of viable and transmissible resistant parasites in clinical infections. Mefloquine resistance results from Pfmdr amplification, pyrimethamine resistance from mutations in Pfdhfr, atovaquone resistance from mutations in Pfcyt b, and resistance to chloroquine from mutations in PfCRT.

Figure 5

The anti-malarial concentration window of selection [18]. The drug concentrations (solid lines) following two doses which define the concentration limits of the window of selection are shown for patients with low parasitaemias (10⁸/body). Total parasites within the body are depicted by dotted and dashed lines. The window opens when following the higher dose (blue) sensitive parasites (blue dotted line) are eliminated but de-novo resistant parasites (blue dashed line) can survive, and successful selection of resistance can take place. Following the lower dose the sensitive parasites can still survive (red dotted line) and later grow to outnumber the drug resistant parasites (blue dashed line) by >9:1, so there is no longer an opportunity for preferential survival of the resistant parasites and thus selection. The window closes. Concentrations between these two boundaries (the "window of selection") are selective.

Figure 6

Parasitaemia profiles in a recrudescent infection. Patients with hyperparasitaemia are at greater risk of subsequent recrudescence even with fully sensitive parasites than patients with lower parasite densities. Each recrudescence provides an opportunity for enrichment (i.e. selection) of any resistant parasites formed. The total number of parasites in the body are shown on the lower vertical axis, drug concentrations of a slowly eliminated anti-malarial are shown in the upper panel. MIC: minimum inhibitory concentration.

Figure 7

Single dose exposure to an artemisinin (arrow) results in a reduction in the number of sensitive parasites by a factor (the PRR) of 10,000 fold. But this is not enough to give a de-novo resistant parasites (red line) any chance of generating sufficient gametocytes for transmission as the sensitive sibling parasites (green lines) remain well ahead in the "race" to transmit.