Halichondrin B and homohalichondrin B, marine natural products binding in the vinca domain of tubulin. Discovery of tubulin-based mechanism of action by analysis of differential cytotoxicity data

Data generated in the new National Cancer Institute drug evaluation program, which is based on inhibition of cell growth in 60 human tumor cell lines, were used to compare new compounds with agents of known mechanism of action in terms of their differential cytotoxicity. Two marine natural products, halichondrin B and homohalichondrin B, appeared repeatedly when the data base was probed with known antimitotic agents. We confirmed that both compounds were highly cytotoxic (IC50 values for L1210 murine leukemia cells of 0.3 and 1 nM, respectively), with accumulation of cells arrested in mitosis at toxic concentrations, that both inhibited the polymerization of purified tubulin, and that both inhibited microtubule assembly dependent on microtubule-associated proteins. Limited amounts of homohalichondrin B, the less active agent, were available, so only halichondrin B was studied in detail. Halichondrin B did not interfere with colchicine binding to tubulin, but it was a noncompetitive inhibitor of the binding of vinblastine to tubulin (apparent Ki, 5.0 microM). Halichondrin B was therefore compared with other agents which interfere with the binding of vinca alkaloids to tubulin (vinblastine, maytansine, dolastatin 10, phomopsin A, rhizoxin) in terms of its effects on tubulin polymerization, inhibition of GTP hydrolysis, inhibition of nucleotide exchange, and stabilization of tubulin, as well as the quantitative assessment of its effects on vinca alkaloid binding and inhibition of cell growth. Since halichondrin B was originally isolated from the same organism as the phosphatase inhibitor okadaic acid, and since it is about 50-fold more effective than okadaic acid as an inhibitor of L1210 cell growth, perturbations of cellular microtubules observed following treatment with okadaic acid should be interpreted cautiously.     

Proteomic identification of divalent metal cation binding proteins in plant mitochondria

Kinetic analyses of cpTat-mediated protein transport across the thylakoid membrane were conducted, revealing three important characteristics of this translocation pathway. First, transport via the cpTAT system displays a non-Michaelis–Menten, sigmoidal rate–substrate relationship with an apparent Hill coefficient of 1.80, indicative of positive homotropic cooperativity. Second, the presence of transport-incompetent substrates was found not to competitively inhibit the translocation of transport-competent substrates. However, the presence of low concentrations of transport-incompetent protein enhances the transport of wild type substrate. Together, these findings suggest that interaction between Tat machinery components and both transport-competent and transport-incompetent protein may elicit a cooperative effect on the translocation rate.     

Impact of solar radiation in disinfecting drinking water contaminated with Giardia duodenalis and Entamoeba histolytica/dispar at a point-of-use water treatment

Aims:  To determine the impact of natural sunlight in disinfecting water contaminated with cysts of Giardia duodenalis and Entamoeba histolytica/dispar using plastic containers.
Methods and Results:  Known quantities of Giardia duodenalis and Entamoeba histolytica/dispar cysts in sterile water were exposed to the sun. Containers were made of polyethylene terephthalate, eight painted black on one side, one not painted and another cut open at the top and the last was a high density polypropylene container. Viability testing was performed using vital and fluorescent dyes. The same assays were conducted under cloudy conditions. Thermal control tests were also performed using heat without ultra violet light from the sun. Results show that 99·9% of parasites was inactivated when water temperatures reached 56°C after sunlight exposure.
Conclusion:  Both solar radiation and heat produced by the sun have a synergistic effect in killing cysts of Giardia duodenalis and Entamoeba histolytica/dispar when temperatures rise above 50°C, with complete death at 56°C, using painted 2-l PET containers.
Significance and Impact of the Study:  Solar disinfection system using PET containers painted black on one side can be used to disinfect water against Giardia duodenalis and Entamoeba histolytica/dispar using natural sunlight.     

Group 2 Innate Lymphoid Cell Proportions Are Diminished in Young Helminth Infected Children and Restored by Curative Anti-helminthic Treatment

BackgroundGroup 2 Innate lymphoid cells (ILC2s) are innate cells that produce the TH2 cytokines IL-5 and IL-13. The importance of these cells has recently been demonstrated in experimental models of parasitic diseases but there is a paucity of data on ILC2s in the context of human parasitic infections and in particular of the blood dwelling parasite Schistosoma haematobium.ConclusionThis study demonstrates that ILC2s are diminished in young helminth infected children and restored by removal of the parasites by treatment, indicating a previously undescribed association between a human parasitic infection and ILC2s and suggesting a role of ILC2s before the establishment of protective acquired immunity in human schistosomiasis.     

Schistosome infection intensity is inversely related to auto-reactive antibody levels

In animal experimental models, parasitic helminth infections can protect the host from auto-immune diseases. We conducted a population-scale human study investigating the relationship between helminth parasitism and auto-reactive antibodies and the subsequent effect of anti-helminthic treatment on this relationship. Levels of antinuclear antibodies (ANA) and plasma IL-10 were measured by enzyme linked immunosorbent assay in 613 Zimbabweans (aged 2-86 years) naturally exposed to the blood fluke Schistosoma haematobium. ANA levels were related to schistosome infection intensity and systemic IL-10 levels. All participants were offered treatment with the anti-helminthic drug praziquantel and 102 treated schoolchildren (5-16 years) were followed up 6 months post-antihelminthic treatment. ANA levels were inversely associated with current infection intensity but were independent of host age, sex and HIV status. Furthermore, after allowing for the confounding effects of schistosome infection intensity, ANA levels were inversely associated with systemic levels of IL-10. ANA levels increased significantly 6 months after anti-helminthic treatment. Our study shows that ANA levels are attenuated in helminth-infected humans and that anti-helminthic treatment of helminth-infected people can significantly increase ANA levels. The implications of these findings are relevant for understanding both the aetiology of immune disorders mediated by auto-reactive antibodies and in predicting the long-term consequences of large-scale schistosomiasis control programs.     

Algorithm for diagnosis of early Schistosoma haematobium using prodromal signs and symptoms in pre-school age children in an endemic district in Zimbabwe

IntroductionPrompt diagnosis of acute schistosomiasis benefits the individual and provides opportunities for early public health intervention. In endemic areas schistosomiasis is usually contracted during the first 5 years of life, thus it is critical to look at how the infection manifests in this age group. The aim of this study was to describe the prodromal signs and symptoms of early schistosomiasis infection, correlate these with early disease progression and risk score to develop an easy to use clinical algorithm to identify early Schistosoma haematobium infection cases in resource limited settings.MethodologyTwo hundred and four, preschool age children who were lifelong residence of a schistosomiasis endemic district and at high risk of acquiring schistosomiasis were followed up from July 2019 to December 2019, during high transmission season. The children received interval and standard full clinical evaluations and laboratory investigations for schistosomiasis by clinicians blinded from their schistosomiasis infection status. Diagnosis of S. haematobium was by urine filtration collected over three consecutive days. Signs and symptoms of schistosomiasis at first examination visit were compared to follow-up visits. Signs and symptoms common on the last schistosomiasis negative visit (before a subsequent positive) were assigned as early schistosomiasis infection (ESI), after possible alternative causes were ruled out. Logistic regression identified clinical predictors. A model based score was assigned to each predictor to create a risk for every child. An algorithm was created based on the predictor risk scores and validated on a separate cohort of 537 preschool age children.ResultsTwenty-one percent (42) of the participants were negative for S. haematobium infection at baseline but turned positive at follow-up. The ESI participants at the preceding S. haematobium negative visit had the following prodromal signs and symptoms in comparison to non-ESI participants; pruritic rash adjusted odds ratio (AOR) = 21.52 (95% CI 6.38–72.66), fever AOR = 82 (95% CI 10.98–612), abdominal pain AOR = 2.6 (95% CI 1.25–5.43), pallor AOR = 4 (95% CI 1.44–11.12) and a history of facial/body swelling within the previous month AOR = 7.31 (95% CI 3.49–15.33). Furthermore 16% of the ESI group had mild normocytic anaemia, whilst 2% had moderate normocytic anaemia. A risk score model was created using a rounded integer from the relative risks ratios. The diagnostic algorithm created had a sensitivity of 81% and a specificity of 96.9%, Positive predictive value = 87.2% and NPV was 95.2%. The area under the curve for the algorithm was 0.93 (0.90–0.97) in comparison with the urine dipstick AUC = 0.58 (0.48–0.69). There was a similar appearance in the validation cohort as in the derivative cohort.ConclusionThis study demonstrates for the first time prodromal signs and symptoms associated with early S. haematobium infection in pre-school age children. These prodromal signs and symptoms pave way for early intervention and management, thus decreasing the harm of late diagnosis. Our algorithm has the potential to assist in risk-stratifying pre-school age children for early S. haematobium infection. Independent validation of the algorithm on another cohort is needed to assess the utility further.