In Vitro Techniques for Antimalarial Development and Evaluation

Desjardins, Robert E.

doi:10.1007/978-3-642-69251-2_6

Cited by 11 publications

(8 citation statements)

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“…In vitro methods for screening antimalarial agents provide an excellent initial assessment of the potency of compounds but do not necessarily predict their efficacy in animals or in patients (4,5). Among other things, such assays are insensitive to how readily parasites can become resistant to a given drug (4). During a typical in vitro assay with a 96-well plate, a 250-l sample contains fewer than 10 6 parasites and proliferation is assessed within 3 days of treatment.…”

Section: Discussionmentioning

confidence: 99%

Frequency of drug resistance in Plasmodium falciparum: a nonsynergistic combination of 5-fluoroorotate and atovaquone suppresses in vitro resistance

Gassis

Rathod

1996

Antimicrob Agents Chemother

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A combination of 5-fluoroorotate and atovaquone eliminated Plasmodium falciparum in long-term cultures more efficiently than either compound alone. The improved potency came not through synergistic activity but through decreased frequency of drug resistance. In support of this finding, it was shown that 5-fluoroorotate and atovaquone do not act in a synergistic fashion, that 5-fluoroorotate-resistant and atovaquone-resistant P. falciparum organisms generated in vitro do not show cross-resistance, and that the frequency of simultaneous resistance to the two compounds approached the product of their individual resistance frequencies. To demonstrate the last finding, and establish proof of principle, an in vitro method was developed for measuring the frequency of drug resistance in P. falciparum. By this method, it was shown that the frequency of resistance to 10(-7) M 5-fluoroorotate was about 10(-6) and the frequency of resistance to 10(-8) M atovaquone was about 10(-5); the frequency of simultaneous resistance to a combination of 10(-7) M 5-fluoroorotate and 10(-8) M atovaquone was less than 5 x 10(-10). On the basis of additional measurements, it was estimated that the frequency of simultaneous resistance to higher, pharmacologically more relevant, concentrations of 10(-6) M 5-fluoroorotate and 10(-7) M atovaquone would be less than 10(-17). Control experiments demonstrated that these drug combinations did not cause increased toxicity to mammalian cells in culture. On this basis, it is predicted that a combination of 5-fluoroorotate and atovaquone will successfully eliminate typical malarial infections in animals and in human patients at doses that are readily tolerated.

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Section: Discussionmentioning

confidence: 99%

Frequency of drug resistance in Plasmodium falciparum: a nonsynergistic combination of 5-fluoroorotate and atovaquone suppresses in vitro resistance

Gassis

Rathod

1996

Antimicrob Agents Chemother

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“…The potential sources of these uncertainties are numerous. In a critical review of in vitro screens, Desjardins pointed out at least three potential limitations (4). First, a compound can be a potent antimalarial agent in vitro but may also have potent toxicity to the host animal.…”

Section: Discussionmentioning

confidence: 99%

Clonal viability measurements on Plasmodium falciparum to assess in vitro schizonticidal activity of leupeptin, chloroquine, and 5-fluoroorotate

Young

Rathod

1993

Antimicrob Agents Chemother

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Until now, the in vitro activity of potential antimalarial agents has been evaluated primarily by monitoring decreases in parasite proliferation. These traditional assays do not distinguish between compounds that arrest proliferation of parasites and compounds that kill them. In this report, a more complex in vitro cytocidal assay for Plasmodium falciparum is described. This assay measures the clonal viability of P. falciparum after the parasites have been treated with an antimalarial agent. The new assay was used to assess cytocidal activities of three antimalarial agents that work through unrelated mechanisms. Leupeptin, a protease inhibitor, arrested the proliferation of W2 clones of P. fakciparum at a MIC of 50 ,uM, but at least 80%o of leupeptin-treated cells were viable as judged by the cytocidal assay. On the other hand, chloroquine at 1 ,uM, its MIC for W2 cells, not only arrested parasite proliferation but also killed more than 99% of the cells. Earlier studies had shown that treatment of P. fakciparum with 100 nM 5-fluoroorotate for 48 h was sufficient to inhibit parasite proliferation and parasite thymidylate synthase but not enough to cause significant incorporation of 5-fluoropyrimidines in parasite nucleic acids. By using the new schizoticidal assay, these conditions were found to be necessary and sufficient to kill all parasites in culture. Results of these studies are consistent with the hypothesis that 5-fluoroorotate-based inactivation of P. falciparum thymidylate synthase triggers a lethal mechanism against malarial parasites.

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“…This measurement is classically known as the Minimum Inhibitory Concentration (MIC), method which is suitable for distinguishing susceptible and resistant isolates. [5] Other in vitro methods…”

Section: Materials and Methods 3 H Hypoxanthine Uptakementioning

confidence: 99%

Screening of antimalarial drugs: An overview

et al. 2006

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Department of Pharmacology, Efforts to discover and develop new antimalarial drugs have increased dramatically in recent Maulana Azad Medical College, years mainly because of resistance to existing antimalarial drugs. Selection of candidate drugs Bahadur Shah Zafar Marg, for clinical trials in man and the design of clinical protocols are based upon consideration of Delhi-110002. data from a battery of preclinical test systems. All compounds are assessed initially in one or *Malaria Research Center, more primary screening models. A compound which is considered 'active' by well estab-No. 22, Sham Nath Marg lished criteria in primary screening test is considered for further evaluation in successively Delhi-110054. more clinical tests. At the end of each stage of testing, a decision is taken to advance the compound to the next stage or discontinue it. Primary screening tests should have optimal

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In Vitro Techniques for Antimalarial Development and Evaluation

Cited by 11 publications

References 70 publications

Frequency of drug resistance in Plasmodium falciparum: a nonsynergistic combination of 5-fluoroorotate and atovaquone suppresses in vitro resistance

Frequency of drug resistance in Plasmodium falciparum: a nonsynergistic combination of 5-fluoroorotate and atovaquone suppresses in vitro resistance

Clonal viability measurements on Plasmodium falciparum to assess in vitro schizonticidal activity of leupeptin, chloroquine, and 5-fluoroorotate

Screening of antimalarial drugs: An overview

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