Efficacies and ADME properties of redox active methylene blue and phenoxazine analogues for use in new antimalarial triple drug combinations with amino-artemisinins.

Efforts to develop new artemisinin triple combination therapies effective against artemisinin-tolerant strains of based on rational combinations comprising artemisone or other amino-artemisinins, a redox active drug and a third drug with a different mode of action have now been extended to evaluation of three potential redox partners. These are the diethyl analogue AD01 of methylene blue (MB), the benzo [α]phenoxazine PhX6, and the thiosemicarbazone DpNEt. IC values against CQ-sensitive and resistant strains ranged from 11.

The Artemiside-Artemisox-Artemisone-M1 Tetrad: Efficacies against Blood Stage Parasites, DMPK Properties, and the Case for Artemiside.

Because of the need to replace the current clinical artemisinins in artemisinin combination therapies, we are evaluating fitness of amino-artemisinins for this purpose. These include the thiomorpholine derivative artemiside obtained in one scalable synthetic step from dihydroartemisinin (DHA) and the derived sulfone artemisone. We have recently shown that artemiside undergoes facile metabolism via the sulfoxide artemisox into artemisone and thence into the unsaturated metabolite M1; DHA is not a metabolite.

Toward New Transmission-Blocking Combination Therapies: Pharmacokinetics of 10-Amino-Artemisinins and 11-Aza-Artemisinin and Comparison with Dihydroartemisinin and Artemether.

As artemisinin combination therapies (ACTs) are compromised by resistance, we are evaluating triple combination therapies (TACTs) comprising an amino-artemisinin, a redox drug, and a third drug with a different mode of action. Thus, here we briefly review efficacy data on artemisone, artemiside, other amino-artemisinins, and 11-aza-artemisinin and conduct absorption, distribution, and metabolism and excretion (ADME) profiling and pharmacokinetic (PK) profiling via intravenous (i.v.

Anti-Melanoma Activities of Artemisone and Prenylated Amino-Artemisinins in Combination With Known Anticancer Drugs.

The most frequently occurring cancers are those of the skin, with melanoma being the leading cause of death due to skin cancer. Breakthroughs in chemotherapy have been achieved in certain cases, though only marginal advances have been made in treatment of metastatic melanoma. Strategies aimed at inducing redox dysregulation by use of reactive oxygen species (ROS) inducers present a promising approach to cancer chemotherapy.

Optimal 10-Aminoartemisinins With Potent Transmission-Blocking Capabilities for New Artemisinin Combination Therapies-Activities Against Blood Stage Including KI3 C580Y Mutants and Liver Stage Parasites.

We have demonstrated previously that amino-artemisinins including artemiside and artemisone in which an amino group replaces the oxygen-bearing substituents attached to C-10 of the current clinical artemisinin derivatives dihydroartemisinin (DHA), artemether and artesunate, display potent activities against the asexual blood stages of (). In particular, the compounds are active against late blood stage gametocytes, and are strongly synergistic in combination with the redox active drug methylene blue. In order to fortify the eventual selection of optimum amino-artemisinins for development into new triple combination therapies also active against artemisinin-resistant mutants, we have prepared new amino-artemisinins based on the easily accessible and inexpensive DHA-piperazine.

The evaluation of the anti-cancer drug elesclomol that forms a redox-active copper chelate as a potential anti-tubercular drug.

The observations that the innate immune system employs copper to eliminate bacterial infection and that resistance to copper enhances virulence of Mycobacterium tuberculosis (Mtb) prompted us to examine the effects the anti-cancer agent elesclomol on Mtb. As a bis-thionohydrazide, elesclomol chelates with copper to form a copper complex in situ that via redox cycling of the metal ion greatly enhances oxidative stress in tumour cells. Here, we demonstrate that elesclomol is relatively potent against Mtb H37Rv with minimum inhibitory concentration of 10 μM (4 mg/L) and against multidrug resistant clinical isolates of Mtb, displays additive interactions with known tuberculosis drugs such as isoniazid and ethambutol, and a synergistic interaction with rifampicin.

Synthesis, antimalarial activities and cytotoxicities of amino-artemisinin-1,2-disubstituted ferrocene hybrids.

Artemisinin-ferrocene conjugates incorporating a 1,2-disubstituted ferrocene analogous to that embedded in ferroquine but attached via a piperazine linker to C10 of the artemisinin were prepared from the piperazine artemisinin derivative, and activities were evaluated against asexual blood stages of chloroquine (CQ) sensitive NF54 and CQ resistant K1 and W2 strains of Plasmodium falciparum (Pf). The most active was the morpholino derivative 5 with IC of 0.86 nM against Pf K1 and 1.

Facile Preparation of -Glycosylated 10-Piperazinyl Artemisinin Derivatives and Evaluation of Their Antimalarial and Cytotoxic Activities.

According to the precepts that C-10 amino-artemisinins display optimum biological activities for the artemisinin drug class, and that attachment of a sugar enhances specificity of drug delivery, polarity and solubility so as to attenuate toxicity, we assessed the effects of attaching sugars to N-4 of the dihydroartemisinin (DHA)-piperazine derivative prepared in one step from DHA and piperazine. -Glycosylated DHA-piperazine derivatives were obtained according to the Kotchetkov reaction by heating the DHA-piperazine with the sugar in a polar solvent. Structure of the D-glucose derivative is secured by X-ray crystallography.

Artemisone and Artemiside Are Potent Panreactive Antimalarial Agents That Also Synergize Redox Imbalance in Plasmodium falciparum Transmissible Gametocyte Stages.

The emergence of resistance toward artemisinin combination therapies (ACTs) by the malaria parasite has the potential to severely compromise malaria control. Therefore, the development of new artemisinins in combination with new drugs that impart activities toward both intraerythrocytic proliferative asexual and transmissible gametocyte stages, in particular, those of resistant parasites, is urgently required. We define artemisinins as oxidant drugs through their ability to oxidize reduced flavin cofactors of flavin disulfide reductases critical for maintaining redox homeostasis in the malaria parasite.

Synthesis, in vitro antimalarial activities and cytotoxicities of amino-artemisinin-ferrocene derivatives.

Novel derivatives bearing a ferrocene attached via a piperazine linker to C-10 of the artemisinin nucleus were prepared from dihydroartemisinin and screened against chloroquine (CQ) sensitive NF54 and CQ resistant K1 and W2 strains of Plasmodium falciparum (Pf) parasites. The overall aim is to imprint oxidant (from the artemisinin) and redox (from the ferrocene) activities. In a preliminary assessment, these compounds were shown to possess activities in the low nM range with the most active being compound 6 with IC values of 2.

Activities of 11-Azaartemisinin and N-Sulfonyl Derivatives against Asexual and Transmissible Malaria Parasites.

Dihydroartemisinin (DHA), either used in its own right or as the active drug generated in vivo from the other artemisinins in current clinical use-artemether and artesunate-induces quiescence in ring-stage parasites of Plasmodium falciparum (Pf). This induction of quiescence is linked to artemisinin resistance. Thus, we have turned to structurally disparate artemisinins that are incapable of providing DHA on metabolism.

Activities of 11-Azaartemisinin and N-Sulfonyl Derivatives against Neospora caninum and Comparative Cytotoxicities.

Neosporosis caused by the apicomplexan parasite Neospora caninum is an economically important disease that induces abortion in dairy and beef cattle. There are no vaccines or drugs available on the market for control or treatment of the disease in bovines. The peroxide artemisinin and its derivatives used clinically for treatment of malaria are active against N.

Preliminary Evaluation of Artemisinin-Cholesterol Conjugates as Potential Drugs for the Treatment of Intractable Forms of Malaria and Tuberculosis.

To evaluate the feasibility of developing drugs that may be active against both malaria and tuberculosis (TB) by using in part putative cholesterol transporters in the causative pathogens and through enhancement of passive diffusion in granulomatous TB, artemisinin-cholesterol conjugates were synthesized by connecting the component molecules through various linkers. The compounds were screened in vitro against Plasmodium falciparum (Pf) and Mycobacterium tuberculosis (Mtb). Antimalarial activities (IC ) against Pf drug-sensitive NF54, and drug-resistant K1 and W2 strains ranged from 0.

Elimination of Schistosoma mansoni in infected mice by slow release of artemisone.

The current treatment of schistosomiasis is based on the anti-helminthic drug praziquantel (PZQ). PZQ affects only the adult stages of schistosomes. In addition, resistance to PZQ is emerging.

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

We sought to establish if methylene homologues of artemisone are biologically more active and more stable than artemisone. The analogy is drawn with the conversion of natural O- and N-glycosides into more stable C-glycosides that may possess enhanced biological activities and stabilities. Dihydroartemisinin was converted into 10β-cyano-10-deoxyartemisinin that was hydrolyzed to the α-primary amide.

Repurposing of antiparasitic drugs: the hydroxy-naphthoquinone buparvaquone inhibits vertical transmission in the pregnant neosporosis mouse model.

The three anti-malarial drugs artemiside, artemisone, and mefloquine, and the naphthoquinone buparvaquone known to be active against theileriosis in cattle and Leishmania infections in rodents, were assessed for activity against Neospora caninum infection. All four compounds inhibited the proliferation of N. caninum tachyzoites in vitro with IC50 in the sub-micromolar range, but artemisone and buparvaquone were most effective (IC50 = 3 and 4.

The Case for Development of 11-Aza-artemisinins for Malaria.

The current treatment regimens for uncomplicated malaria comprise an artemisinin in combination with another drug (ACT). However, the recent emergence of resistance to ACTs in South East Asia dramatically emphasizes the need for new artemisinins. The current artemisinins have been in use since the late 1970s and have relatively poor thermal, chemical and metabolic stabilities - all are metabolized or hydrolyzed in vivo to dihydroartemisinin (DHA) that itself undergoes facile decomposition in vivo.

Interactions between artemisinins and other antimalarial drugs in relation to the cofactor model--a unifying proposal for drug action.

Artemisinins are proposed to act in the malaria parasite cytosol by oxidizing dihydroflavin cofactors of redox-active flavoenzymes, and under aerobic conditions by inducing their autoxidation. Perturbation of redox homeostasis coupled with the generation of reactive oxygen species (ROS) ensues. Ascorbic acid-methylene blue (MB), N-benzyl-1,4-dihydronicotinamide (BNAH)-MB, BNAH-lumiflavine, BNAH-riboflavin (RF), and NADPH-FAD-E.

Synthesis of artemiside and its effects in combination with conventional drugs against severe murine malaria.

This research describes the use of novel antimalarial combinations of the new artemisinin derivative artemiside, a 10-alkylamino artemisinin. It is a stable, highly crystalline compound that is economically prepared from dihydroartemisinin in a one-step process. Artemiside activity was more pronounced than that of any antimalarial drug in use, both in Plasmodium falciparum culture and in vivo in a murine malaria model depicting cerebral malaria (CM).

A partial convergence in action of methylene blue and artemisinins: antagonism with chloroquine, a reversal with verapamil, and an insight into the antimalarial activity of chloroquine.

Artemisinins rapidly oxidize leucomethylene blue (LMB) to methylene blue (MB); they also oxidize dihydroflavins such as the reduced conjugates RFH₂ of riboflavin (RF), and FADH₂ of the cofactor flavin adenine dinucleotide (FAD), to the corresponding flavins. Like the artemisinins, MB oxidizes FADH₂, but unlike artemisinins, it also oxidizes NAD(P)H. Like MB, artemisinins are implicated in the perturbation of redox balance in the malaria parasite by interfering with parasite flavoenzyme disulfide reductases.

Artemisone uptake in Plasmodium falciparum-infected erythrocytes.

Artemisone is one of the most promising artemisinin derivatives in clinical trials. Previous studies with radiolabeled artemisinin and dihydroartemisinin have measured uptake in Plasmodium falciparum-infected erythrocytes. Uptake is much greater in infected than in uninfected erythrocytes, but the relative contributions of transport, binding, and metabolism to this process still await definition.

Facile oxidation of leucomethylene blue and dihydroflavins by artemisinins: relationship with flavoenzyme function and antimalarial mechanism of action.

The antimalarial drug methylene blue (MB) affects the redox behaviour of parasite flavin-dependent disulfide reductases such as glutathione reductase (GR) that control oxidative stress in the malaria parasite. The reduced flavin adenine dinucleotide cofactor FADH(2) initiates reduction to leucomethylene blue (LMB), which is oxidised by oxygen to generate reactive oxygen species (ROS) and MB. MB then acts as a subversive substrate for NADPH normally required to regenerate FADH(2) for enzyme function.

Preparation of N-sulfonyl- and N-carbonyl-11-azaartemisinins with greatly enhanced thermal stabilities: in vitro antimalarial activities.

As the clinically used artemisinins do not withstand the thermal stress testing required to evaluate shelf life for storage in tropical countries where malaria is prevalent, there is a need to develop thermally more robust artemisinin derivatives. Herein we describe the attachment of electron-withdrawing arene- and alkanesulfonyl and -carbonyl groups to the nitrogen atom of the readily accessible Ziffer 11-azaartemisinin to provide the corresponding N-sulfonyl- and -carbonylazaartemisinins. Two acylurea analogues were also prepared by treatment of the 11-azaartemisinin with arylisocyanates.