Evaluation of atovaquone in the treatment of patients with uncomplicated Plasmodium falciparum malaria

The activity of atovaquone in patients with oligosymptomatic Plasmodium falciparum malaria was assessed in an open, non-comparative clinical study. The patients showed a good clinical response, but there was a high rate of recrudescence. The activity of atovaquone in combination with another antimalarial agent should be investigated.     

Efficacy and safety of atovaquone/proguanil as suppressive prophylaxis for Plasmodium falciparum malaria

Currently recommended prophylactic regimens for Plasmodium falciparum malaria are associated with a high incidence of adverse events and/or suboptimal efficacy. In a double-blind, placebo-controlled, randomized clinical trial in western Kenya, adult volunteers received a treatment course of atovaquone/proguanil hydrochloride (250 mg/100 mg per tablet) to eliminate preexisting infection. Immediately thereafter, subjects were randomized to one of the three prophylactic regimens to receive one atovaquone/proguanil tablet daily (n = 68), two atovaquone/proguanil tablets daily (n = 65), or placebo (n = 65) for 10 weeks. The study endpoint for any subject was the development of parasitemia, evident on blood smear, during prophylaxis. Of the evaluable subjects, all in the low-dose (54 of 54) and high-dose (54 of 54) atovaquone/proguanil groups remained malaria-free during the 10-week prophylaxis period, in contrast to only 48% (26 of 54) in the placebo group (P < .001). Both atovaquone/proguanil prophylactic regimens were as well tolerated as placebo. Thus, atovaquone/proguanil appears to be highly efficacious and safe as prophylaxis for P. falciparum malaria.     

Mannose-binding lectin plasma levels and gene polymorphisms in Plasmodium falciparum malaria

The contribution of mannose-binding lectin (MBL) to protection from malaria was assessed by comparing plasma concentrations of MBL and the frequency of MBL gene polymorphisms in groups of Gabonese children participating in a prospective study of severe and mild malaria due to infection with Plasmodium falciparum. At admission, a higher proportion of patients with severe malaria had a low level of MBL compared with subjects with mild malaria (0.35 vs. 0.19, P = .02). Two mutations in codons 54 and 57 of the MBL gene were detected. They were present at higher frequency in those with severe malaria (0.45 vs. 0.31, P = .04). These results suggest that deficient innate immune responses, in the form of low MBL levels, may be a risk factor for severe malaria in some young children who lack well-developed, clinically protective acquired immune responses.     

Plasma levels of the interleukin-6 cytokine family in persons with severe Plasmodium falciparum malaria

Plasma levels of interleukin (IL)-6, soluble IL-6 receptor, soluble gp130, leukemia inhibitory factor (LIF), and ciliary neutrophic factor (CNTF) were analyzed in 32 patients with severe malaria. Ten had renal failure, 8 had cerebral malaria, and 14 had other causes of severity. Before treatment, the IL-6 and soluble IL-6 receptor plasma levels were significantly higher in persons with cerebral malaria or renal failure than in other groups (P<.01 for both). After initiation of therapy, IL-6 levels dropped within 24 h, but soluble IL-6 receptor levels increased. CNTF levels were significantly reduced in persons with cerebral malaria or renal failure but normalized within 24 h. Plasma concentrations of gp130 and LIF did not differ between the malaria groups or normal controls. Excessive levels of IL-6 could be controlled by a subsequent shedding of the soluble IL-6 receptor, and low-level CNTF expression could contribute to or even result from cerebral malaria or renal failure.     

In vitro activities of novel antifolate drug combinations against Plasmodium falciparum and human granulocyte CFUs

The potency of antimalarial dihydrofolate reductase inhibitors, alone and in synergistic combination with dihydropteroate synthetase inhibitors, against the Kenyan K39 strain of Plasmodium falciparum (pyrimethamine resistant) and against normal replicating human bone marrow cells in in vitro culture has been studied. Therapeutic indices and rank order of synergistic potency were derived. Trimethoprim, pyrimethamine, and the quinazolines WR159412 and WR158122 had the smallest therapeutic indices (1.39, 4.38, 2.56, and 90.0, respectively), while the three triazines clociguanil, WR99210, and chlorcycloguanil had the largest (3,562, 3,000, and 2,000, respectively). In rank order of decreasing activity against P. falciparum, the six most potent drug combinations were WR99210-dapsone, chlorcycloguanil-dapsone, WR158122-dapsone, WR159412-dapsone, WR159412-sulfamethoxazole, and chlorcycloguanil-sulfamethoxazole; pyrimethamine-sulfadoxine was the least potent combination. These experiments form a basis for the selection of rapidly eliminated antifolate combinations for further clinical testing.     

Efficacy of artemisinin and mefloquine combinations against Plasmodium falciparum. In vitro simulation of in vivo pharmacokinetics

The activity of artemisinin in combination with mefloquine was tested in vitro against a chloroquine-sensitive (F32) strain of Plasmodium falciparum. A method of repetitive dosing and extending the culture observation period to 28-30 days was used to mimic the in vivo pharmacokinetic situation. Plasmodium falciparum was exposed to artemisinin from 10(-8) to 10(-5) M, mefloquine from 3 x 10(-9) to 10(-5) M and their combinations. The exposure time for artemisinin was 3 hours twice daily and for mefloquine 24 hours. The drug-dosing duration was 3 days. Neither artemisinin nor mefloquine alone provided radical clearance of P. falciparum, even when maximum concentrations (10(-5) M) were applied. The antiparasitic activity of artemisinin and mefloquine were significantly higher when dosed alone. Effective concentrations for different degrees of inhibition (EC 50, 90 and 99) of both artemisinin and mefloquine respectively were significantly lower when used in combination. At concentrations normally reached in vivo, this effect was clearly synergistic (P = 0.016) Our in vitro model of intermittent dosing of artemisinin and mefloquine combinations for 3 days provides significant evidence of positive interaction between the two compounds. Lower combination concentrations around the MIC-values for the individual compounds showed synergistic effect, and high concentrations showed additive effect. This indicates that such drug combinations may provide radical clearance at concentrations lower than those required for single-drug treatment.     

Liver function tests in adults with Plasmodium falciparum infection

This study examines the levels of serum bilirubin, aspartate transaminase and alkaline phosphatase in adults with Plasmodium falciparum malaria. One hundred and six sets of liver function tests were obtained, and 63 (59.4%) patients had one of the above indices elevated outside the local reference range. Serum bilirubin and aspartate transaminase were relatively higher than alkaline phosphatase. Neither duration of illness nor severity of infection showed any significant correlation with any of the indices measured.     

Control of gene expression in Plasmodium falciparum

Myocardial contusion is an infrequent, but sometimes serious complication in patients who experienced deceleration (blunt) trauma. We investigated the assessment of the new cardiac markers troponin I (cTnI) and troponin T (cTnT) in relation to the conventional CKMB-activity, the CKMB-activity/CK-total ratio, CKMB-mass and the CKMB-mass/CK-total ratio for the detection of myocardial contusion in 89 patients with blunt trauma (38 patients with thoracic injuries and 51 patients without thoracic injuries). All parameters were analysed at admission (t1) and 24 h after admission (t2). For the patients with thoracic injuries, at t1 cTnI was elevated in three, and cTnT in four patients; at t2 both cTnI and cTnT were elevated in nine patients. At t1, eighteen to thirty patients had increased levels of the conventional parameters; at t2 this was true for six to thirty-five patients. For the patients without thoracic injuries all cTnI and cTnT levels were within the reference ranges at t1. At t2 one patient, who experienced an acute myocardial infarction, had elevated cTnI and cTnT levels. At t1, five to thirty-five patients had increased levels of the conventional parameters; at t2 this was true for four to forty-two patients. From this study we conclude that the conventional parameters are not useful for the detection of myocardial contusion in patients experiencing blunt trauma. The parameters cTnI and cTnT are equally accurate and more reliable for the selection of patients who require intensive cardiac monitoring. If at admission the cTnI or the cTnT levels are within the reference ranges, a second analysis after admission is necessary to reach a reliable conclusion concerning myocardial contusion as a result of trauma on basis of the troponin levels.     

Risk factors of chloroquine resistance in Plasmodium falciparum malaria

Sources of asbestos in drinking water may be natural deposits or the use of asbestos cement for water distribution. 50 water samples were selected in Austria to detect fibre contamination from either geology or asbestos cement by comparison with control areas and by comparison of raw and treated water. Standardized EPA/BGA methodology with transmission electron microscopy, energy dispersive X-ray analysis and selected area electron diffraction was used to quantify concentrations of different sized amphibole and chrysotile fibres. In 10 areas with asbestos deposits and in 14 areas with use of asbestos cement pipes asbestos concentrations in drinking water were low and not significantly different from 6 control areas (median 32,000 total asbestos fibres per litre). The relative highest concentration was found in an area with natural deposits at the source of the water supply (190,000 per litre). In areas without natural deposits the increase of asbestos concentrations from origin to consumer of water was not significant and unrelated to water aggressiveness, age and length of asbestos cement pipes. This could be mainly due to the fact that in areas with aggressive water asbestos cement pipes have been coated in Austria. A sample from a cistern, however, showed considerable asbestos contamination and raises concern about the use of surface water for room air humidification.     

Plasmodium falciparum and Plasmodium yoelii: effect of the iron chelation prodrug dexrazoxane on in vitro cultures

To determine if an iron-chelating prodrug that must undergo intracellular hydrolysis to bind iron has antimalarial activity, we examined the action of dexrazoxane on Plasmodium falciparum cultured in human erythrocytes and P. yoelii cultured in mouse hepatocytes. Dexrazoxane was recently approved to protect humans from doxorubucin-induced cardiotoxicity. Using the fluorescent marker calcein, we confirmed that the iron-chelating properties of dexrazoxane are directly related to its ability to undergo hydrolysis. As a single agent, dexrazoxane inhibited synchronized cultures of P. falciparum in human erythrocytes only at suprapharmacologic concentrations (> 200 microM). In combination with desferrioxamine B, dexrazoxane in pharmacologic concentrations (100-200 microM) moderately potentiated inhibition by approximately 20%. In contrast, pharmacologic concentrations of dexrazoxane (50-200 microM) as a single agent inhibited the progression of P. yoelli from sporozoites to schizonts in cultured mouse hepatocytes by 45 to 69% (P < 0.001). These results are consistent with the presence of a dexrazoxane-hydrolyzing enzyme in hepatocytes but not in erythrocytes or malaria parasites. Furthermore, these findings suggest that dexrazoxane must be hydrolyzed to an iron-chelating intermediate before it can inhibit the malaria parasite, and they raise the possibility that the iron chelator prodrug concept might be exploited to synthesize new antimalarial agents.     

Treatment with interleukin 12 in combination with atovaquone or clindamycin significantly increases survival of mice with acute toxoplasmosis

The capacity of interleukin 12 (IL-12) to potentiate drugs in the treatment of murine toxoplasmosis was examined. IL-12 (100 ng/injection), atovaquone (10 mg/kg of body weight/day), or clindamycin (5 mg/kg/day) administered alone caused delayed time to death or minimal survival rates. In contrast, significant survival rates resulted when the same dose of IL-12 was used in combination the same doses of atovaquone (P=0.01) or clindamycin (P=0.001). Infected mice treated with IL-12 plus drug produced significantly higher levels of gamma interferon than controls. Although IL-12 was effective only when administered before infection, these results suggest that this cytokine may be a useful adjunct in the therapy of human toxoplasmosis in situations when cysts reactivate and tachyzoites start multiplying in immunocompromised patients.     

Malaria in humans: Plasmodium falciparum blood infection levels are linked to chromosome 5q31-q33

Blood serum concentration of IGF-I was analyzed to determine its relationship with individual postweaning feed efficiency (gain/feed) of 36 crossbred steer calves fed at three levels of feed intake (n = 12 at each level). Diets consisted of a corn silage-based growing diet for 84 d followed by a 91% concentrate finishing diet for 56 d. Dietary intake levels were at 80, 90, or 100% of ad libitum. Diets were formulated to ensure equal daily intake of protein, vitamins, and minerals across intake treatment levels. Intake was measured daily; ADG, DMI, and feed efficiency were calculated at 28-d intervals, through d 140. Individual weights and serum samples were collected at the beginning of the study and at 28-d intervals thereafter. The IGF-I concentrations were determined with a RIA. Data were analyzed as a multivariate split-plot in time. Imposed dietary intake restrictions did not affect serum IGF-I concentration (P = .90) or individual feed efficiency (P = .36), even though the least squares means for IGF-I concentration tended to decrease and the feed efficiency means tended to increase under the restricted intake levels. Serum IGF-I concentration, ADG, and feed efficiency were affected (P < .001) by collection date. Residual correlations between IGF-I concentrations at adjacent 28-d sampling times averaged .72. Diet intake level x sampling time interactions existed for ADG (P = .02) and feed efficiency (P < .001). Positive residual correlations of .28 (P < .001) and .16 (P = .07) existed between IGF-I and ADG and between IGF-I and feed efficiency, respectively. Regression analysis indicated that a 1 ng/mL increase in serum IGF-I concentration was associated with a .00135 kg/d increase in ADG (P < .001) and a .0001 kg gain/kg feed increase in feed efficiency (P = .04). These results support the hypothesis that serum IGF-I plays a role in growth and in efficiency of feed utilization in beef cattle.     

Cross-resistance to antifolates in multidrug resistant cell lines with P-glycoprotein or multidrug resistance protein expression

Resistance to some (lipophilic) antifolates has been associated with P-glycoprotein (P-gp)-mediated multidrug resistance (MDR). A possible relationship with non-P-gp MDR has not been established. We studied resistance to antifolates in SW-1573 human lung carcinoma cells, a P-gp overexpressing variant SW-1573/2R160 and a multidrug resistance protein (MRP) overexpressing variant SW-1573/2R120. In this study, thymidylate synthase (TS) inhibitors with different properties concerning the efficiency of membrane transport and the efficiency of polyglutamylation were tested for cross-resistance in SW-1573/2R120 and SW-1573/2R160 cells. Growth inhibition patterns in this cell line panel were measured by the Sulforhodamine B (SRB) assay. Resistance factors for TS inhibitors were: 2.4 and 0.4 for 5-fluorouracil (5FU), 18.8 and 8.8 for ZD1694, 17 and 0.7 for AG337, and 40 and 8.3 for BW1843U89 in SW-1573/2R160 and SW-1573/2R120, respectively. This study showed changes in the TS enzyme kinetics during the induction of doxorubicin resistance in both SW-1573 variants, resulting in 2-fold lower Km values for 2'-deoxyuridine-5'-monophosphate (dUMP) in both resistant variants compared to the parental cell line. TS activity, TS protein induction and TS mRNA expression all had 2-fold increased in the SW-1573/2R120 compared to the SW-1573/2R160. 3H-MTX influx was 2-fold lower in SW-1573/2R160 cells compared to SW-1573/2R120 and SW-1573 cells. In the SW-1573/2R160 cell line, an aberrant intracellular trafficking towards the target TS was observed, compared to SW-1573/2R120 and SW-1573 cells as measured by the TS in situ assay. The rate of TS inhibition by the TS inhibitors used in this study was similar in all cell lines. In conclusion, collateral sensitivity to 5FU and the lipophilic AG337 and cross-resistance to other antifolates were observed in non-P-gp MDR SW-1573/2R120 cells, as well as resistance to all antifolates in P-gp SW-1573/2R160 cells. The mechanism of resistance in SW-1573/2R160 cells possibly involves reduced influx and changes in intracellular trafficking routes. For the SW-1573/2R120 cell line, several changes related to the TS enzyme possibly play a role in the observed cross-resistance and collateral sensitivity pattern.     

Compositional constraints in the extremely GC-poor genome of Plasmodium falciparum

We have analyzed the compositional properties of coding (protein encoding) and non-coding sequences of Plasmodium falciparum, a unicellular parasite characterized by an extremely AT-rich genome. GC% levels, base and dinucleotide frequencies were studied. We found that among the various factors that contribute to the properties of the sequences analyzed, the most relevant are the compositional constraints which operate on the whole genome.     

Identification of novel membrane structures in Plasmodium falciparum infected erythrocytes

Phosphorylation sites were introduced into chimeric monoclonal antibody CC49 (MAb-chCC49) by inserting synthetic fragments encoding two and six phosphorylation sites into an expression vector, pdHL7. The phosphorylation sites were created by using the predicted consensus sequences for phosphorylation by the cAMP-dependent protein kinase to the carboxyl terminus of the heavy chain constant region of the MAb-chCC49. The resultant modified antibodies (MAb-chCC49K1 and MAb-chCC49-6P) were expressed in NS0 cells and purified. The MAb-chCC49K1 protein contains two phosphorylation sites per heavy chain whereas the MAb-chCC49-6P protein contains six sites per heavy chain. Both MAb-chCC49K1 and MAb-chCC49-6P proteins can be phosphorylated by the catalytic subunit of cAMP-dependent protein kinase with [gamma-32P]ATP to high specific activity. The 32P-labeled MAb-chCC49K1 and MAb-chCC49-6P proteins bind to cells expressing TAG-72 antigens. The introduction of phosphorylation sites into a monoclonal antibody provides a reagent for the diagnosis and treatment of cancer. The use of multiple phosphorylation sites provides antibodies with very high specific radioactivity and demonstrates that cassettes of phosphorylation sites can be introduced into proteins without altering their functional activity.     

Subacute Plasmodium falciparum malaria in 43 patients returning from areas with chloroquine in Africa

PURPOSE: To identify clinical and biological features of subacute falciparum malaria, risk factors, and to evaluate the efficacy of curative treatment. PATIENTS AND METHODS: Diagnostic criteria were the association of apyrexia, anemia, little or no parasitemia and a high titer of anti-Plasmodium antibodies. Forty-three cases were observed in subjects returning from chloroquine-resistant areas in Africa. They were matched with controls for age, country of residence and duration of stay. Controls were missionaries who attended our unit for a routine medical check-up during the study period. RESULTS: The clinical presentation and biological features were similar to "malarial cachexia", a condition mainly described in non-immune children in endemic areas. Splenomegaly was present in 58% of the patients. Biological features included little or no parasitemia, an overall decrease in the blood cell count, an increased erythrocyte sedimentation rate and a high titer of anti-Plasmodium antibodies. This syndrome was not correlated with the frequency of chloroquine resistance, the area of stay (urban or rural) or to the kind of chemoprophylaxis. CONCLUSIONS: This study describes subacute resistant falciparum malaria in patients who had prolonged stay in chloroquine-resistant areas of Africa associating splenomegaly, cytopenia and a low or absent parasitemia. Subacute chloroquine-resistant malaria could be due to host factors which remained to be determined by prospective immunological studies. Curative treatment with mefloquine is effective.     

Treatment of major depression with psychotherapy or psychotherapy-pharmacotherapy combinations

BACKGROUND: Few reliable correlates of treatment response in depression have emerged despite nearly 40 years of research. We examined the correlates of recovery in a "mega-analysis," or meta-analysis of original data, of 595 patients with major depressive disorder enrolled in 6 standardized treatment protocols. METHODS: All patients (mean age, 44 years; 31% male and 69% female) met criteria for nonbipolar, nonpsychotic primary major depressive disorder and were treated for 16 weeks with either cognitive behavior therapy or interpersonal psychotherapy alone (psychotherapy alone; n = 243) or interpersonal psychotherapy plus antidepressant pharmacotherapy (combined therapy; n = 352). The impact of treatment type, severity, study, and other covariates on recovery rates or time to recovery were examined by means of chi 2, log-rank tests, the Cox proportional hazards model, and sensitivity analyses. RESULTS: Whereas combined therapy was not significantly more effective than psychotherapy alone in milder depressions, a highly significant advantage was observed in more severe recurrent depressions. Poorer outcomes were also observed in women and older patients, although these effects were dependent on inclusion of particular studies. CONCLUSIONS: Mega-analysis is a powerful method for comparing the efficacy of treatments and examining correlates of response. Using this method, we found new evidence in support of the widespread clinical impression that combined therapy is superior to psychotherapy alone for treatment of more severe, recurrent depressions.     

Transmission intensity and Plasmodium falciparum diversity on the northwestern border of Thailand

Reproduction depends on the co-ordinated expression of stereotypical behaviors and precisely timed physiological events, yet the neurobiological mechanisms underlying the integration of sensory and hormonal information that is crucial to this process have remained difficult to define. A variety of experimental approaches has provided compelling evidence that the anteroventral periventricular nucleus (AVPV) of the preoptic region plays a particularly important role in the neural control of gonadotropin secretion. It is larger in female rats, contains high densities of neurons that express receptors for ovarian steroid hormones and appears to provide direct projections to gonadotropin releasing hormone neurons in the hypothalamus. Moreover, it receives inputs from a variety of distinct sensory systems known to influence secretion of luteinizing hormone from the anterior pituitary. Thus, the AVPV appears to represent an important nodal point in sexually dimorphic forebrain circuits for the integration of sensory and hormonal information that influence reproduction. Examples of neurohumoral integration at the level of functional neural systems, individual neurons in the AVPV, or at the molecular level have been identified which provide new insight into how the hypothalamus co-ordinates expression of sex specific reproductive behaviors with gonadotropin secretion.     

Considering disparities in resistance of Plasmodium falciparum in Africa in chemoprevention decisions

OBJECTIVES: Assess the efficacy of preventive and curative treatments of imported malaria. METHODS: The in vitro drug susceptibility of mefloquine, chloroquine and cycloguanil was determined against African isolates of Plasmodium falciparum from imported malaria cases by an isotopic in vitro test or a genomic approach. RESULTS: Plasmodium falciparum resistance to mefloquine, chloroquine or to the dihydrofolate reductase inhibitor was present in 5.2%, 46% and 42% of isolates respectively. Plasmodium falciparum drug resistance to chloroquine or antifolinics was more frequent in permanent than in seasonal malarial transmission areas. Simultaneous resistance to chloroquine and antifolinics was observed in 17% of isolates between 1991 and 1994 and in 28% between 1995 and 1997.