Stability of the Antimalarial Drug Dihydroartemisinin under Physiologically Relevant Conditions: Implications for Clinical Treatment and Pharmacokinetic and In Vitro Assays

Abstract

f Artemisinins are peroxidic antimalarial drugs known to be very potent but highly chemically unstable; they degrade in the presence of ferrous iron, Fe(II)-heme, or biological reductants. Less documented is how this translates into chemical stability and antimalarial activity across a range of conditions applying to in vitro testing and clinical situations. Dihydroartemisinin (DHA) is studied here because it is an antimalarial drug on its own and the main metabolite of other artemisinins. The behaviors of DHA in phosphate-buffered saline, plasma, or erythrocyte lysate at different temperatures and pH ranges were examined. The antimalarial activity of the residual drug was evaluated using the chemosensitivity assay on Plasmodium falciparum, and the extent of decomposition of DHA was established through use of high-performance liquid chromatography with electrochemical detection analysis. The role of the Fe(II)-heme was investigated by blocking its reactivity using carbon monoxide (CO). A significant reduction in the antimalarial activity of DHA was seen after incubation in plasma and to a lesser extent in erythrocyte lysate. Activity was reduced by half after 3 h and almost completely abolished after 24 h. Serum-enriched media also affected DHA activity. Effects were temperature and pH dependent and paralleled the increased rate of decomposition of DHA from pH 7 upwards and in plasma. These results suggest that particular care should be taken in conducting and interpreting in vitro studies, prone as their results are to experimental and drug storage conditions. Disorders such as fever, hemolysis, or acidosis associated with malaria severity may contribute to artemisinin instability and reduce their clinical efficacy.A rtemisinin derivatives have been the most potent antimalarials available to date; they are highly active against all Plasmodium species and parasite stages, including young gametocytes, and constitute the backbone of malaria case management for severe and uncomplicated malaria (as a component of artemisinin combination therapy [ACT]) (1). The emergence and spread of artemisinin resistance in Southeast Asia is, therefore, a serious threat to malaria control (2, 3).All artemisinin derivatives share a distinctive 1,2,4-trioxane pharmacophore, which confers their antimalarial activity (the corresponding acyclic analogues lacking the endoperoxide bridge are inactive) (4, 5) but also makes these molecules particularly highly reactive and thus difficult to quantify in plasma or blood. Of the various artemisinin-type compounds in use, dihydroartemisinin (DHA), the reduced lactol derivative of artemisinin, is an antimalarial compound on its own (currently coformulated with piperaquine) and the main bioactive metabolite of artesunate and artemether. DHA itself is chemically fragile and displays a marked propensity to undergo ring opening of the lactol and rearrangement under neutral conditions, leading to a new, biologically active peroxide, which in turn rapidly decays to the inert end product deoxyartemisi...

Genetic screens reveal a central role for heme biosynthesis in artemisinin susceptibility

“…), washing out the compound after different lengths of time, and assessing viability as monolayer lysis 72 h post invasion (Figure 1c). Based on these results, we used 5 h treatments for both intracellular and extracellular parasites 34 .…”

Methylene Homologues of Artemisone: An Unexpected Structure–Activity Relationship and a Possible Implication for the Design of C10‐Substituted Artemisinins

“…ChemMedChem 2016, 11,1469 -1479 www.chemmedchem.org broughta bout by inserting methylene units into glycolipid 12 to generate analogue 13.I nt erms of absorption, the thiomorpholine S,S-dioxide group bestows acceptable physicochemical properties on artemisone, [20,21] including an enhancement in polarity.H owever,t he group may have another less apparent but important role in enhancing antimalarial activity.According to one model put forward for the antimalarial mechanismo f action of artemisinins,n amely the cofactor model, [4,46,49] hydride transfer to O1 of the peroxideb ridge from FADH 2 is coupled with heterolytic cleavage of the peroxide and synchronous unzipping with loss of the (protonated)C 10 amino group in providing ring-opened tricarbonyl compound 22,w hich undergoes closure to the deoxydihydroartemisininp roduct 23 and its epimer 24 (Scheme 5A). [4] Significantly,a nalogous processes occur for artemether (4)a nd artesunate (5), that is, no products are obtained from this heterolytic process that retain the original group attached to C10. [4] In the more specialized cases of C10 aminoartemisinins, hydride transfer must be facilitated by protonation of the nitrogen atom, as illustrated for artemisonei nS cheme 5A.T hus, in terms of the cofactor model, the feebler antimalarial activities of the methylene homologues becomes apparent-there is no leaving group at C10, and therefore, this process terminates with the formation of amino intermediate 25,w hich transforms into deoxy product 26 and the various other products arising from 25 by intramolecular condensation or dehydration pathways.…”

“…[2] DHA undergoes facile phase Im etabolism [3] and rearranges under physiological conditions at pH 7.4 to peroxyhemiacetal 5, [4] which is observed in the plasma of patients treated with artesunate. [5] Artesunate is rapidly hydrolyzed to DHA in vivo, [6] but likely because of the protectivee ffect of the ester group against first-pass metabolism, it is ab etter sourceo fD HA in plasma than DHA itself. [7] The facile metabolism of artemether to DHA is reflected in the detection of the latter compound in subjects administered with artemether.…”

“…[2] The thermal stability of artemisone is similar to that of artesunate. Althougha tp H7.2 artemisone is hydrolytically stable (in contrast to artesunate), at pH 1.2, just like artesunate, [5] it is relativelyr apidly hydrolyzed to DHA. [33] The hydrolysis likely is initiated by protonation of the C10 nitrogen atom followed by cleavaget oo xonium ion 10 that adds water to generateD HA (Scheme1).…”

Intestinal Permeability of Artesunate-Loaded Solid Lipid Nanoparticles Using the Everted Gut Method

“…It is highly unstable in aqueous acidic and neutral conditions [12, 13]. Its active metabolite (DHA) is rapidly decomposed to the inert end product, deoxyartemisinin, and is also easily reduced by biological molecules at physiological pH 7.4 [14]. Artesunate (C 19 H 28 O 8 ) is also a weak acid (Figure 1); hence its solubility, dissolution, and permeability are affected by altered pH conditions.…”