Dose ratio proton radiography using the proximal side of the Bragg peak

Purpose: In recent years there has been a movement towards single-detector proton radiography, due to its potential ease of implementation within the clinical environment. One such single-detector technique is the dose ratio method, in which the dose maps from two pristine Bragg peaks are recorded beyond the patient. To date, this has only been investigated on the distal side of the lower energy Bragg peak, due to the sharp fall-off. We investigate the limits and applicability of the dose ratio method on the proximal side of the lower energy Bragg peak, which has the potential to allow a much wider range of water-equivalent thicknesses (WET) to be imaged. Comparisons are made with the use of the distal side of the Bragg peak. Methods: Using the analytical approximation for the Bragg peak we generated theoretical dose ratio curves for a range of energy pairs, and then determined how an uncertainty in the dose ratio would translate to a spread in the WET estimate. By defining this spread as the accuracy one could achieve in the WET estimate, we were able to generate look-up graphs of the range on the proximal side of the Bragg peak that one could reliably use. These were dependent on the energy pair, noise level in the dose ratio image and the required accuracy in the WET. Using these look-up graphs we investigated the applicability of the technique for a range of patient treatment sites. We validated the theoretical approach with experimental measurements using a complementary metal oxide semiconductor active pixel sensor (CMOS APS), by imaging a small sapphire sphere in a high energy proton beam. Results: Provided the noise level in the dose ratio image was 1% or less, a larger spread of WETs could be imaged using the proximal side of the Bragg peak (max 5.31 cm) compared to the distal side (max 2.42 cm). In simulation it was found that, for a pediatric brain, it is possible to use the technique to image a region with a square field equivalent size of 7.6 cm2, for a required accuracy in the WET of 3 mm and a 1% noise level in the dose ratio image. The technique showed limited applicability for other patient sites. The CMOS APS demonstrated a good accuracy, with a root-mean-square-error of 1.6 mm WET. The noise in the measured images was found to be σ =1.2% (standard deviation) and theoretical predictions with a 1.96σ noise level showed good agreement with the measured errors. Conclusions: After validating the theoretical approach with measurements, we have shown that the use of the proximal side of the Bragg peak when performing dose ratio imaging is feasible, and allows for a wider dynamic range than when using the distal side. The dynamic range available increases as the demand on the accuracy of the WET decreases. The technique can only be applied to clinical sites with small maximum WETs such as for pediatric brains

The glutathione biosynthetic pathway of Plasmodium is essential for mosquito transmission

1Infection of red blood cells (RBC) subjects the malaria parasite to oxidative stress. Therefore, efficient antioxidant and redox systems are required to prevent damage by reactive oxygen species. Plasmodium spp. have thioredoxin and glutathione (GSH) systems that are thought to play a major role as antioxidants during blood stage infection. In this report, we analyzed a critical component of the GSH biosynthesis pathway using reverse genetics. Plasmodium berghei parasites lacking expression of gamma-glutamylcysteine synthetase (γ-GCS), the rate limiting enzyme in de novo synthesis of GSH, were generated through targeted gene disruption thus demonstrating, quite unexpectedly, that γ-GCS is not essential for blood stage development. Despite a significant reduction in GSH levels, blood stage forms of pbggcs− parasites showed only a defect in growth as compared to wild type. In contrast, a dramatic effect on development of the parasites in the mosquito was observed. Infection of mosquitoes with pbggcs− parasites resulted in reduced numbers of stunted oocysts that did not produce sporozoites. These results have important implications for the design of drugs aiming at interfering with the GSH redox-system in blood stages and demonstrate that de novo synthesis of GSH is pivotal for development of Plasmodium in the mosquito

The Synthetic Plasmodium falciparum Circumsporozoite Peptide PfCS102 as a Malaria Vaccine Candidate: A Randomized Controlled Phase I Trial

BACKGROUND: Fully efficient vaccines against malaria pre-erythrocytic stage are still lacking. The objective of this dose/adjuvant-finding study was to evaluate the safety, reactogenicity and immunogenicity of a vaccine candidate based on a peptide spanning the C-terminal region of Plasmodium falciparum circumsporozoite protein (PfCS102) in malaria naive adults. METHODOLOGY AND PRINCIPAL FINDINGS: Thirty-six healthy malaria-naive adults were randomly distributed into three dose blocks (10, 30 and 100 microg) and vaccinated with PfCS102 in combination with either Montanide ISA 720 or GSK proprietary Adjuvant System AS02A at days 0, 60, and 180. Primary end-point (safety and reactogenicity) was based on the frequency of adverse events (AE) and of abnormal biological safety tests; secondary-end point (immunogenicity) on P. falciparum specific cell-mediated immunity and antibody response before and after immunization. The two adjuvant formulations were well tolerated and their safety profile was good. Most AEs were local and, when systemic, involved mainly fatigue and headache. Half the volunteers in AS02A groups experienced severe AEs (mainly erythema). After the third injection, 34 of 35 volunteers developed anti-PfCS102 and anti-sporozoite antibodies, and 28 of 35 demonstrated T-cell proliferative responses and IFN-gamma production. Five of 22 HLA-A2 and HLA-A3 volunteers displayed PfCS102 specific IFN-gamma secreting CD8(+) T cell responses. Responses were only marginally boosted after the 3(rd) vaccination and remained stable for 6 months. For both adjuvants, the dose of 10 microg was less immunogenic in comparison to 30 and 100 microg that induced similar responses. AS02A formulations with 30 microg or 100 microg PfCS102 induced about 10-folds higher antibody and IFN-gamma responses than Montanide formulations. CONCLUSIONS/SIGNIFICANCE: PfCS102 peptide was safe and highly immunogenic, allowing the design of more advanced trials to test its potential for protection. Two or three immunizations with a dose of 30 microg formulated with AS02A appeared the most appropriate choice for such studies. TRIAL REGISTRATION: Swissmedic.ch 2002 DR 1227

The use of a P. falciparum specific coiled-coil domain to construct a self-assembling protein nanoparticle vaccine to prevent malaria.

The parasitic disease malaria remains a major global public health concern and no truly effective vaccine exists. One approach to the development of a malaria vaccine is to target the asexual blood stage that results in clinical symptoms. Most attempts have failed. New antigens such as P27A and P27 have emerged as potential new vaccine candidates. Multiple studies have demonstrated that antigens are more immunogenic and are better correlated with protection when presented on particulate delivery systems. One such particulate delivery system is the self-assembling protein nanoparticle (SAPN) that relies on coiled-coil domains of proteins to form stable nanoparticles. In the past we have used de novo designed amino acid domains to drive the formation of the coiled-coil scaffolds which present the antigenic epitopes on the particle surface. Here we use naturally occurring domains found in the tex1 protein to form the coiled-coil scaffolding of the nanoparticle. Thus, by engineering P27A and a new extended form of the coiled-coil domain P27 onto the N and C terminus of the SAPN protein monomer we have developed a particulate delivery system that effectively displays both antigens on a single particle that uses malaria tex1 sequences to form the nanoparticle scaffold. These particles are immunogenic in a murine model and induce immune responses similar to the ones observed in seropositive individuals in malaria endemic regions. We demonstrate that our P27/P27A-SAPNs induce an immune response akin to the one in seropositive individuals in Burkina Faso. Since P27 is highly conserved among different Plasmodium species, these novel SAPNs may even provide cross-protection between Plasmodium falciparum and Plasmodium vivax the two major human malaria pathogens. As the SAPNs are also easy to manufacture and store they can be delivered to the population in need without complication thus providing a low cost malaria vaccine

Conducting High-Value Secondary Dataset Analysis: An Introductory Guide and Resources

Secondary analyses of large datasets provide a mechanism for researchers to address high impact questions that would otherwise be prohibitively expensive and time-consuming to study. This paper presents a guide to assist investigators interested in conducting secondary data analysis, including advice on the process of successful secondary data analysis as well as a brief summary of high-value datasets and online resources for researchers, including the SGIM dataset compendium (www.sgim.org/go/datasets). The same basic research principles that apply to primary data analysis apply to secondary data analysis, including the development of a clear and clinically relevant research question, study sample, appropriate measures, and a thoughtful analytic approach. A real-world case description illustrates key steps: (1) define your research topic and question; (2) select a dataset; (3) get to know your dataset; and (4) structure your analysis and presentation of findings in a way that is clinically meaningful. Secondary dataset analysis is a well-established methodology. Secondary analysis is particularly valuable for junior investigators, who have limited time and resources to demonstrate expertise and productivity

Toward a Surrogate Marker of Malaria Exposure: Modeling Longitudinal Antibody Measurements under Outbreak Conditions

Background: Biomarkers of exposure to Plasmodium falciparum would be a useful tool for the assessment of malaria burden and analysis of intervention and epidemiological studies. Antibodies to pre-erythrocytic antigens represent potential surrogates of exposure. Methods and Findings: In an outbreak cohort of U.S. Marines deployed to Liberia, we modeled pre- and post-deployment IgG against P. falciparum sporozoites by immunofluorescence antibody test, and both IgG and IgM against the P. falciparum circumsporozoite protein by enzyme-linked immunosorbant assay. Modeling seroconversion thresholds by a fixed ratio, linear regression or nonlinear regression produced sensitivity for identification of exposed U.S. Marines between 58-70% and specificities between 87-97%, compared with malaria-naïve U.S. volunteers. Exposure was predicted in 30-45% of the cohort. Conclusion: Each of the three models tested has merits in different studies, but further development and validation in endemic populations is required. Overall, these models provide support for an antibody-based surrogate marker of exposure to malaria

The Role of Age and Exposure to Plasmodium falciparum in the Rate of Acquisition of Naturally Acquired Immunity: A Randomized Controlled Trial

Background: The rate of acquisition of naturally acquired immunity (NAI) against malaria predominantly depends on transmission intensity and age, although disentangling the effects of these is difficult. We used chemoprophylaxis to selectively control exposure to P. falciparum during different periods in infancy and explore the effect of age in the build-up of NAI, measured as risk of clinical malaria.\ud \ud Methods and Findings: A three-arm double-blind randomized placebo-controlled trial was conducted in 349 infants born to Mozambican HIV-negative women. The late exposure group (LEG) received monthly Sulfadoxine-Pyrimethamine (SP) plus Artesunate (AS) from 2.5–4.5 months of age and monthly placebo from 5.5–9.5 months; the early exposure group (EEG) received placebo from 2.5–4.5 months and SP+AS from 5.5–9.5 months; and the control group (CG) received placebo from 2.5–9.5 months. Active and passive case detection (PCD) were conducted from birth to 10.5 and 24 months respectively. The primary endpoint was time to first or only episode of malaria in the second year detected by PCD. The incidence of malaria during the second year was of 0.50, 0.51 and 0.35 episodes/PYAR in the LEG, EEG and CG respectively (p = 0.379 for the adjusted comparison of the 3 groups). The hazard ratio of the adjusted comparison between the LEG and the CG was 1.38 (0.83–2.28, p = 0.642) and that between the EEG and the CG was 1.35 (0.81–2.24, p = 0.743).\ud \ud Conclusions: After considerably interfering with exposure during the first year of life, there was a trend towards a higher risk of malaria in the second year in children who had received chemoprophylaxis, but there was no significant rebound. No evidence was found that the age of first exposure to malaria affects the rate of acquisition of NAI. Thus, the timing of administration of antimalarial interventions like malaria vaccines during infancy does not appear to be a critical determinant