Chloroquine Resistance in Malaria: A Deficiency of Chloroquine Binding

Abstract

Abstract.-Chloroquine-14C was used to study the processes which concentrate chloroquine in mouse red blood cells infected with chloroquine-sensitive or with chloroquine-resistant Plasmodium berghei. The initial rates of uptake and exchange of chloroquine-14C were both too fast to measure, yet large concentration gradients were maintained by the cells. When red blood cells were exposed to 10-8 M chloroquine at 220C, with pH between 7.2 and 7.4, steady-state gradients of chloroquine-14C were approximately 600: 1 (cells: medium) for cells infected with chloroquine-sensitive parasites, 100:1 for cells comparably infected with chloroquine-resistant parasites, and 14: 1 for uninfected cells. The processes responsible for these gradients were saturable, in agreement with the proposal of chloroquine binding to cellular constituents. No degradation of chloroquine was detected.The major difference between the chloroquine-sensitive and -resistant parasites was a deficiency of high-affinity binding of chloroquine by cells infected with chloroquine-resistant parasites. This deficiency explains the reduced ability of chloroquine-resistant parasites to concentrate chloroquine, and it suggests that chloroquine resistance is due to a decrease in the number, affinity, or accessibility of chloroquine receptor sites on a constituent of the malaria parasite.Previous work has demonstrated that mouse red blood cells infected with chloroquine-sensitive Plasmodium berghei can achieve concentration gradients of chloroquinel of 100: 1 (cells: plasma) in vivo, whereas cells infected with a chloroquine-resistant variant of this same strain of P. berghei concentrate chloroquine less than half as well.2' 3 Since the partial loss of ability to concentrate chloroquine is associated with chloroquine resistance in this model system, the present work was undertaken to characterize and to compare the concentrative processes of chloroquine-sensitive and -resistant P. berghei.Materials and Methods.-Infections were maintained in mice by weekly passage of chloroquine-sensitive P. berghei and biweekly passage of chloroquine-resistant P. berghei.4Both types of parasites were descendants of the same parent strain of P. berghei, the NYU-2 strain; the chloroquine-resistant variant was the one previously used in studies of the processes which concentrate chloroquine in infected red blood cells in vivo.2, 8 To pass the infections, normal mice were each given an intraperitoneal injection of 0.1 ml of a 1: 25 dilution of blood which had 30-90% of its red blood cells parasitized. The blood used for passage of the chloroquine-resistant parasites was always obtained from mice each one of which was treated daily with 1 mg of chloroquine intraperitoneally for 4 days of each week after inoculation with parasitized blood. This amount of chloroquine did not prevent the fatal progression of malaria caused by chloroquine-resistant parasites although it promptly cures mice infected with chloroquine-sensitive parasites.8 Under our conditions, the chloroquine-sensitive para...

Stage independent chloroquine resistance and chloroquine toxicity revealed via spinning disk confocal microscopy

“…When CQR parasites were found to accumulate substantially less CQ vs. CQ sensitive (CQS) [10], and a lysosomal -like organelle called the digestive vacuole (DV) was found to be the principle site of CQ accumulation [11,12], focus began to shift towards investigation of DV processes. Importantly, Orjih and Fitch found that mid-stage trophozoites with fully formed DV accumulate more CQ than any other stage [13].…”

Chloroquine Resistance in Malaria: Accessibility of Drug Receptors to Mefloquine

“…2) differ in several important ways from those reported for chloroquine accumulation (3,9,12 (3,9,12), and the curve describing the process is sigmoid (9). Erythro- cytes infected with P. berghei CS accumulate more chloroquine than those infected with P. berghei CR, and the curve describing the process is a rectangular hyperbola (3, 9, 11).…”

“…berghei is 10i8 M, and the maximal binding capacity is approximately 26 ,imol/kg of erythrocyte pellet (3,9). DISCUSSION Despite large differences between mefloquine and chloroquine in accessibility to drug receptors and in strengths of binding, we now know that mefloquine competitively inhibits chloroquine accumulation and that chloroquine competitively inhibits mefloquine accumulation.…”

“…The parasites, the characteristics of erythrocytes infected with them, and the methods used to obtain and prepare infected erythrocytes for study have been described previously (3,12). The P. berghei CR line is resistant to 40 mg of chloroquine per kg of body weight of the mouse daily (12).…”

“…Under conditions that prevail in vivo, however, resistant parasites receive less exposure because the infected erythrocytes fail to accumulate chloroquine avidly (3,5,9,12,14). Consequently, chloroquine resistance in this model may be attributed to reduced exposure of the parasites rather than to ineffectiveness of the drug after it reaches the parasites: This mechanism of resistance provides a basis for explaining how drugs in the same chemotherapeutic class with chloroquine could be effective in the treatment of chloroquine-resistant malaria.…”