De Novo Prediction of Drug Targets and Candidates by Chemical Similarity-Guided Network-Based Inference.

Identifying drug-target interactions is a crucial step in discovering novel drugs and for drug repositioning. Network-based methods have shown great potential thanks to the straightforward integration of information from different sources and the possibility of extracting novel information from the graph topology. However, despite recent advances, there is still an urgent need for efficient and robust prediction methods. Here, we present SimSpread, a novel method that combines network-based inference with chemical similarity. This method employs a tripartite drug-drug-target network constructed from protein-ligand interaction annotations and drug-drug chemical similarity on which a resource-spreading algorithm predicts potential biological targets for both known or failed drugs and novel compounds. We describe small molecules as vectors of similarity indices to other compounds, thereby providing a flexible means to explore diverse molecular representations. We show that our proposed method achieves high prediction performance through multiple cross-validation and time-split validation procedures over a series of datasets. In addition, we demonstrate that our method performed a balanced exploration of both chemical ligand space (scaffold hopping) and biological target space (target hopping). Our results suggest robust and balanced performance, and our method may be useful for predicting drug targets, virtual screening, and drug repositioning.

Rational Design of Resveratrol O-methyltransferase for the Production of Pinostilbene.

Pinostilbene is a monomethyl ether analog of the well-known nutraceutical resveratrol. Both compounds have health-promoting properties, but the latter undergoes rapid metabolization and has low bioavailability. O-methylation improves the stability and bioavailability of resveratrol. In plants, these reactions are performed by O-methyltransferases (OMTs). Few efficient OMTs that monomethylate resveratrol to yield pinostilbene have been described so far. Here, we report the engineering of a resveratrol OMT from Vitis vinifera (VvROMT), which has the highest catalytic efficiency in di-methylating resveratrol to yield pterostilbene. In the absence of a crystal structure, we constructed a three-dimensional protein model of VvROMT and identified four critical binding site residues by applying different in silico approaches. We performed point mutations in these positions generating W20A, F24A, F311A, and F318A variants, which greatly reduced resveratrol's enzymatic conversion. Then, we rationally designed eight variants through comparison of the binding site residues with other stilbene OMTs. We successfully modified the native substrate selectivity of VvROMT. Variant L117F/F311W showed the highest conversion to pinostilbene, and variant L117F presented an overall increase in enzymatic activity. Our results suggest that VvROMT has potential for the tailor-made production of stilbenes.

Calculation of accurate interatomic contact surface areas for the quantitative analysis of non-bonded molecular interactions.

SUMMARY: Intra- and intermolecular contact surfaces are routinely calculated for a large array of applications in bioinformatics but are typically approximated from differential solvent accessible surface area calculations and not calculated directly. These approximations do not properly take the effects of neighboring atoms into account and tend to deviate considerably from the true contact surface. We implemented an extension of the original Shrake-Rupley algorithm to accurately estimate interatomic contact surface areas of molecular structures and complexes. Our extended algorithm is able to calculate the contact area of an atom to all nearby atoms by directly calculating overlapping surface patches, taking into account the possible shielding effects of neighboring atoms. Here, we present a versatile software tool and web server for the calculation of contact surface areas, as well as buried surface areas and solvent accessible surface areas (SASA) for different types of biomolecules, such as proteins, nucleic acids and small organic molecules. Detailed results are provided in tab-separated values format for analysis and Protein Databank files for visualization. Direct contact surface area calculation resulted in improved accuracy in a benchmark with a non-redundant set of 245 protein-DNA complexes. SASA-based approximations underestimated protein-DNA contact surfaces on average by 40%. This software tool may be useful for surface-based intra- and intermolecular interaction analyses and scoring function development. AVAILABILITY AND IMPLEMENTATION: A web server, stand-alone binaries for Linux, MacOS and Windows and C++ source code are freely available from http://schuellerlab.org/dr_sasa/. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

A novel fungal gene regulation system based on inducible VPR-dCas9 and nucleosome map-guided sgRNA positioning.

Programmable transcriptional regulation is a powerful tool to study gene functions. Current methods to selectively regulate target genes are mainly based on promoter exchange or on overexpressing transcriptional activators. To expand the discovery toolbox, we designed a dCas9-based RNA-guided synthetic transcription activation system for Aspergillus nidulans that uses enzymatically disabled "dead" Cas9 fused to three consecutive activation domains (VPR-dCas9). The dCas9-encoding gene is under the control of an estrogen-responsive promoter to allow induction timing and to avoid possible negative effects by strong constitutive expression of the highly active VPR domains. Especially in silent genomic regions, facultative heterochromatin and strictly positioned nucleosomes can constitute a relevant obstacle to the transcriptional machinery. To avoid this negative impact and to facilitate optimal positioning of RNA-guided VPR-dCas9 to targeted promoters, we have created a genome-wide nucleosome map from actively growing cells and stationary cultures to identify the cognate nucleosome-free regions (NFRs). Based on these maps, different single-guide RNAs (sgRNAs) were designed and tested for their targeting and activation potential. Our results demonstrate that the system can be used to regulate several genes in parallel and, depending on the VPR-dCas9 positioning, expression can be pushed to very high levels. We have used the system to turn on individual genes within two different biosynthetic gene clusters (BGCs) which are silent under normal growth conditions. This method also opens opportunities to stepwise activate individual genes in a cluster to decipher the correlated biosynthetic pathway. Graphical abstract KEYPOINTS: • An inducible RNA-guided transcriptional regulator based on VPR-dCas9 was established in Aspergillus nidulans. • Genome-wide nucleosome positioning maps were created that facilitate sgRNA positioning. • The system was successfully applied to activate genes within two silent biosynthetic gene clusters.

Exploring DNA dynamics within oligonucleosomes with coarse-grained simulations: SIRAH force field extension for protein-DNA complexes.

Describing the regulation of chromatin segments by protein recognition events constitute a major goal in biology and biotechnology. Despite astonishing experimental developments, achieving nearly atomistic spatial/temporal resolution on such macromolecular systems remains a big challenge owing to the intrinsic flexibility of large biological assemblies. Although computer simulations have become a reliable complement to experimental techniques, computational cost limits their routine applications to relatively small systems. However, the development of accurate and cost-effective coarse-grained (CG) models helps to bridge the gap between molecular dynamics simulations and biologically relevant scales. Performing an exhaustive search on a set of well-resolved crystallographic protein-DNA complexes, we introduced improvements on the CG SIRAH force field to describe protein-DNA interfaces. Modifications were validated against a set of non redundant structures and applied to the simulation of the longest DNA segment in complex with proteins that has been crystallized to date, i.e. a tetranucleosome. Multimicrosecond simulation of this small chromatin segment evidences a large mobility of the external DNA filaments, which is consistent with results from FRET experiments in solution. Moreover, we found that the sub-microsecond dynamics of DNA is strongly modulated by the quaternary structure, partially overcoming the intrinsic dynamics dictated by the primary structure.

PDIviz: analysis and visualization of protein-DNA binding interfaces.

UNLABELLED: Specific recognition of DNA by proteins is a crucial step of many biological processes. PDIviz is a plugin for the PyMOL molecular visualization system that analyzes protein-DNA binding interfaces by comparing the solvent accessible surface area of the complex against the free protein and free DNA. The plugin provides three distinct three-dimensional visualization modes to highlight interactions with DNA bases and backbone, major and minor groove, and with atoms of different pharmacophoric type (hydrogen bond donors/acceptors, hydrophobic and thymine methyl). Each mode comes in three styles to focus the visual analysis on the protein or DNA side of the interface, or on the nucleotide sequence. PDIviz allows for the generation of publication quality images, all calculated data can be written to disk, and a command line interface is provided for automating tasks. The plugin may be helpful for the detailed identification of regions involved in DNA base and shape readout, and can be particularly useful in rapidly pinpointing the overall mode of interaction. AVAILABILITY AND IMPLEMENTATION: Freely available at http://melolab.org/pdiviz/ as a PyMOL plugin. Tested with incentive, educational, and open source versions of PyMOL on Windows, Mac and Linux systems. CONTACT: aschueller@bio.puc.cl SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Andrographolide activates the canonical Wnt signalling pathway by a mechanism that implicates the non-ATP competitive inhibition of GSK-3ß: autoregulation of GSK-3ß in vivo.

Wnt/ß-catenin signalling is an important pathway that regulates multiple biological processes, including cell adhesion and determination of cell fate during animal development; in the adult nervous system it regulates the structure and function of synapses. Wnt-signalling dysfunction is associated with several neurodegenerative diseases such as schizophrenia and Alzheimer's disease. The use of natural compounds is an interesting strategy in the search for drugs with the therapeutic potential to activate this signalling pathway. In the present study, we report that andrographolide (ANDRO), a component of Andrographis paniculata, is a potent activator of Wnt signalling. Our results indicate that ANDRO activates this pathway, inducing the transcription of Wnt target genes by a mechanism that bypasses Wnt ligand binding to its receptor. In vitro kinase assays demonstrate that ANDRO inhibits glycogen synthase kinase (GSK)-3ß by a non-ATP-competitive, substrate-competitive mode of action. In silico analyses suggest that ANDRO interacts with the substrate-binding site of GSK-3ß. Finally, we demonstrated that the increase seen in the levels of GSK-3ß phosphorylated at Ser9 is the result of an autoregulatory mechanism of the kinase in vivo, although not through activation of protein phosphatase type 1. Our results suggest that ANDRO could be used as a potential therapeutic drug for disorders caused by Wnt-signalling dysfunction such as neurodegenerative diseases.

State-of-the-art silicon carbide diode dosimeters for ultra-high dose-per-pulse radiation at FLASH radiotherapy.

Objective.The successful implementation of FLASH radiotherapy in clinical settings, with typical dose rates >40 Gy s-1, requires accurate real-time dosimetry.Approach.Silicon carbide (SiC) p-n diode dosimeters designed for the stringent requirements of FLASH radiotherapy have been fabricated and characterized in an ultra-high pulse dose rate electron beam. The circular SiC PiN diodes were fabricated at IMB-CNM (CSIC) in 3µm epitaxial 4H-SiC. Their characterization was performed in PTB's ultra-high pulse dose rate reference electron beam. The SiC diode was operated without external bias voltage. The linearity of the diode response was investigated up to doses per pulse (DPP) of 11 Gy and pulse durations ranging from 3 to 0.5µs. Percentage depth dose measurements were performed in ultra-high dose per pulse conditions. The effect of the total accumulated dose of 20 MeV electrons in the SiC diode sensitivity was evaluated. The temperature dependence of the response of the SiC diode was measured in the range 19 °C-38 °C. The temporal response of the diode was compared to the time-resolved beam current during each electron beam pulse. A diamond prototype detector (flashDiamond) and Alanine measurements were used for reference dosimetry.Main results.The SiC diode response was independent both of DPP and of pulse dose rate up to at least 11 Gy per pulse and 4 MGy s-1, respectively, with tolerable deviation for relative dosimetry (<3%). When measuring the percentage depth dose under ultra-high dose rate conditions, the SiC diode performed comparably well to the reference flashDiamond. The sensitivity reduction after 100 kGy accumulated dose was <2%. The SiC diode was able to follow the temporal structure of the 20 MeV electron beam even for irregular pulse estructures. The measured temperature coefficient was (-0.079 ± 0.005)%/°C.Significance.The results of this study demonstrate for the first time the suitability of silicon carbide diodes for relative dosimetry in ultra-high dose rate pulsed electron beams up to a DPP of 11 Gy per pulse.

Two novel members of Onygenales, Keratinophyton kautmanovae and K. keniense spp. nov. from soil.

Two new Keratinophyton species, K. kautmanovae sp. nov. and K. keniense sp. nov., isolated from soil samples originating from two different geographical and environmental locations (Africa and Europe) are described and illustrated. Phylogenetically informative sequences obtained from the internal transcribed spacer (ITS) region and the nuclear large subunit (LSU) rDNA, as well as their unique phenotype, fully support novelty of these two fungi for this genus. Based on ITS and LSU combined phylogeny, both taxa are resolved in a cluster with eight accepted species, including K. alvearium, K. chongqingense, K. hubeiense, K. durum, K. lemmensii, K. siglerae, K. submersum, and K. sichuanense. The new taxon, K. kautmanovae, is characterized by clavate, smooth to coarsely verrucose conidia, absence of arthroconidia, slow growth at 25 °C, and no growth at 30 °C, while K. keniense is morphologically unique with a high diversity of conidial shapes (clavate, filiform, globose, cymbiform and rhomboid). Both species are described based on their asexual, a chrysosporium-like morph. While the majority of hitherto described Keratinophyton taxa came from Europe, India and China, the new species K. keniense represents the first reported taxonomic novelty for this genus from Africa.

Metrology for advanced radiotherapy using particle beams with ultra-high dose rates.

Dosimetry of ultra-high dose rate beams is one of the critical components which is required for safe implementation of FLASH radiotherapy (RT) into clinical practice. In the past years several national and international programmes have emerged with the aim to address some of the needs that are required for translation of this modality to clinics. These involve the establishment of dosimetry standards as well as the validation of protocols and dosimetry procedures. This review provides an overview of recent developments in the field of dosimetry for FLASH RT, with particular focus on primary and secondary standard instruments, and provides a brief outlook on the future work which is required to enable clinical implementation of FLASH RT.

Charge collection efficiency of commercially available parallel-plate ionisation chambers in ultra-high dose-per-pulse electron beams.

Objective. This investigation aims to experimentally determine the charge collection efficiency (CCE) of six commercially available parallel-plate ionisation chamber (PPIC) models in ultra-high dose-per-pulse (UHDPP) electron beams.Approach. The CCE of 22 PPICs has been measured in UHDPP electron beams at the National Metrology Institution of Germany (PTB). The CCE was determined for a dose per pulse (DPP) range between 0.1 and 6.4 Gy (pulse duration of 2.5µs). The results obtained with the different PPICs were compared to evaluate the reproducibility, intra- and inter-model variation, and the performance of a CCE empirical model.Main results. The intra-model variation was, on average, 4.0%, which is more than three times the total combined relative standard uncertainty and was found to be greater at higher DPP (up to 20%). The inter-model variation for the PPIC with 2 mm electrode spacing, which was found to be, on average, 10%, was also significant compared to the relative uncertainty and the intra-model variation. The observed CCE variation could not be explained only by the expected deviation of the electrode spacing from the nominal value within the manufacturing tolerance. It should also be noted that a substantial polarity effect, between 0.914(5) and 1.201(3), was observed, and significant intra- and inter-model variation was observed on this effect.Significance. For research and pre-clinical study, the commercially available PPIC with a well-known CCE (directly measured for the specific chamber) and with a small electrode spacing could be used for relative and absolute dosimetry with a lower-limit uncertainty of 1.6% (k= 1) in the best case. However, to use a PPIC as a secondary standard in UHDPP electron beams for clinical purposes would require new model development to reduce the ion recombination, the polarity effect, and the total standard uncertainty on the dose measurement.

Genome analysis of Cephalotrichum gorgonifer and identification of the biosynthetic pathway for rasfonin, an inhibitor of KRAS dependent cancer.

BACKGROUND: Fungi are important sources for bioactive compounds that find their applications in many important sectors like in the pharma-, food- or agricultural industries. In an environmental monitoring project for fungi involved in soil nitrogen cycling we also isolated Cephalotrichum gorgonifer (strain NG_p51). In the course of strain characterisation work we found that this strain is able to naturally produce high amounts of rasfonin, a polyketide inducing autophagy, apoptosis, necroptosis in human cell lines and showing anti-tumor activity in KRAS-dependent cancer cells. RESULTS: In order to elucidate the biosynthetic pathway of rasfonin, the strain was genome sequenced, annotated, submitted to transcriptome analysis and genetic transformation was established. Biosynthetic gene cluster (BGC) prediction revealed the existence of 22 BGCs of which the majority was not expressed under our experimental conditions. In silico prediction revealed two BGCs with a suite of enzymes possibly involved in rasfonin biosynthesis. Experimental verification by gene-knock out of the key enzyme genes showed that one of the predicted BGCs is indeed responsible for rasfonin biosynthesis. CONCLUSIONS: This study identified a biosynthetic gene cluster containing a key-gene responsible for rasfonin production. Additionally, molecular tools were established for the non-model fungus Cephalotrichum gorgonifer which allows strain engineering and heterologous expression of the BGC for high rasfonin producing strains and the biosynthesis of rasfonin derivates for diverse applications.

Computer-based annotation of putative AraC/XylS-family transcription factors of known structure but unknown function.

Currently, about 20 crystal structures per day are released and deposited in the Protein Data Bank. A significant fraction of these structures is produced by research groups associated with the structural genomics consortium. The biological function of many of these proteins is generally unknown or not validated by experiment. Therefore, a growing need for functional prediction of protein structures has emerged. Here we present an integrated bioinformatics method that combines sequence-based relationships and three-dimensional (3D) structural similarity of transcriptional regulators with computer prediction of their cognate DNA binding sequences. We applied this method to the AraC/XylS family of transcription factors, which is a large family of transcriptional regulators found in many bacteria controlling the expression of genes involved in diverse biological functions. Three putative new members of this family with known 3D structure but unknown function were identified for which a probable functional classification is provided. Our bioinformatics analyses suggest that they could be involved in plant cell wall degradation (Lin2118 protein from Listeria innocua, PDB code 3oou), symbiotic nitrogen fixation (protein from Chromobacterium violaceum, PDB code 3oio), and either metabolism of plant-derived biomass or nitrogen fixation (protein from Rhodopseudomonas palustris, PDB code 3mn2).

How to Completely Squeeze a Fungus-Advanced Genome Mining Tools for Novel Bioactive Substances.

Fungal species have the capability of producing an overwhelming diversity of bioactive substances that can have beneficial but also detrimental effects on human health. These so-called secondary metabolites naturally serve as antimicrobial "weapon systems", signaling molecules or developmental effectors for fungi and hence are produced only under very specific environmental conditions or stages in their life cycle. However, as these complex conditions are difficult or even impossible to mimic in laboratory settings, only a small fraction of the true chemical diversity of fungi is known so far. This also implies that a large space for potentially new pharmaceuticals remains unexplored. We here present an overview on current developments in advanced methods that can be used to explore this chemical space. We focus on genetic and genomic methods, how to detect genes that harbor the blueprints for the production of these compounds (i.e., biosynthetic gene clusters, BGCs), and ways to activate these silent chromosomal regions. We provide an in-depth view of the chromatin-level regulation of BGCs and of the potential to use the CRISPR/Cas technology as an activation tool.

Characterization of the PTB ultra-high pulse dose rate reference electron beam.

Purpose. This investigation aims to present the characterisation and optimisation of an ultra-high pulse dose rate (UHPDR) electron beam at the PTB facility in Germany. A Monte Carlo beam model has been developed for dosimetry study for future investigation in FLASH radiotherapy and will be presented.Material and methods. The 20 MeV electron beams generated by the research linear accelerator has been characterised both in-beamline with profile monitors and magnet spectrometer, and in-water with a diamond detector prototype. The Monte Carlo model has been used to investigate six different setups to enable different dose per pulse (DPP) ranges and beam sizes in water. The properties of the electron radiation field in water have also been characterised in terms of beam size, quality specifierR50and flatness. The beam stability has also been studied.Results. The difference between the Monte-Carlo simulated and measuredR50was smaller than 0.5 mm. The simulated beam sizes agreed with the measured ones within 2 mm. Two suitable setups have been identified for delivering reference UHPDR electron beams. The first one is characterised by a SSD of 70 cm, while in the second one an SSD of 90 cm is used in combination with a 2 mm aluminium scattering plates. The two set-ups are quick and simple to install and enable an expected overall DPP range from 0.13 Gy up to 6.7 Gy per pulse.Conclusion. The electron beams generated by the PTB research accelerator have shown to be stable throughout the four-months length of this investigation. The Monte Carlo models have shown to be in good agreement for beam size and depth dose and within 1% for the beam flatness. The diamond detector prototype has shown to be a promising tool to be used for relative measurements in UHPDR electron beams.

Absorbed-dose-to-water measurement using alanine in ultra-high-pulse-dose-rate electron beams.

Objective. The aim of the presented study is to evaluate the dose response of the PTB's secondary standard system, which is based on alanine and electron spin resonance (ESR) spectroscopy measurement, in ultra-high-pulse-dose-rate (UHPDR) electron beams.Approach. The alanine dosimeter system was evaluated in the PTB's UHPDR electron beams (20 MeV) in a range of 0.15-6.2 Gy per pulse. The relationship between the obtained absorbed dose to water per pulse and the in-beamline charge measurement of the electron pulses acquired using an integrating current transformer (ICT) was evaluated. Monte Carlo simulations were used to determine the beam quality conversion and correction factors required to perform alanine dosimetry.Main results. The beam quality conversion factor from the reference quality60Co to 20 MeV obtained by Monte Carlo simulation, 1.010(1), was found to be within the standard uncertainty of the consensus value, 1.014(5). The dose-to-water relative standard uncertainty was determined to be 0.68% in PTB's UHPDR electron beams.Significance. In this investigation, the dose-response of the PTB's alanine dosimeter system was evaluated in a range of dose per pulse between 0.15 Gy and 6.2 Gy and no evidence of dose-response dependency of the PTB's secondary standard system based on alanine was observed. The alanine/ESR system was shown to be a precise dosimetry system for evaluating absorbed dose to water in UHPDR electron beams.

Numerical modeling of air-vented parallel plate ionization chambers for ultra-high dose rate applications.

PURPOSE: Air-vented ionization chambers have been the secondary standard for radiation dosimetry since the origins of radiation metrology. However, the feasibility of their use in ultra-high dose rate pulsed beams has been a matter of discussion, as large losses are caused by ion recombinations and no suitable theoretical model is available for their correction. The theories developed by Boag and his contemporaries since the 1950s, which have provided the standard ion recombination correction factor in clinical dosimetry, do not provide an accurate description when used under the limit conditions of ultra-high dose rates (UHDRs). Moreover, the high-ion recombination effects of ionization chambers under extreme dose-rate applications are an obstacle to the development of adequate dosimetry standards. METHODS: In this article, the charge carrier transport equations within a parallel plate ionization chamber (PPIC) have been solved numerically with a double aim. First, this numerical model provides a more accurate tool that can be used to evaluate ion recombination correction for established PPICs in pulsed ultra-high dose rate regimes. Second, studying the chamber behavior in detail allow as to explore the limits of new chamber designs in order to improve their performance under UHDRs. The model presented here has been tested by measuring the instantaneous current of one unit of a Roos chamber (i.e., the time-resolved current during and after the irradiation pulse under UHDR conditions) and comparing these results with the absolute value of the simulated current. RESULTS: The experimental data show consistent agreement with the results obtained using the numerical model. The experimental instantaneous current reveals effects such as the variation of the free electron fraction with the dose per pulse that are supported by the numerical model but cannot be explained in the framework of Boag's theory. CONCLUSIONS: Numerical solutions of the charge carrier released and transport in ionization chambers are able to estimate the effects observed when PPICs are irradiated with ultra-high dose rate beams and to provide new insight into processes related to recombination losses at UHDRs. These models can be reliably extended to include regions where current analytical solutions are not valid. An agreement of better than 5 % between the experimental and simulated effective free electron fraction is found. We were able to reproduce the instantaneous current from a Roos chamber. The discrepancies observed between the experimental data and the numerical simulations can be attributed to the uncertainty about the transport parameters involved in the calculation.

Charge collection efficiency, underlying recombination mechanisms, and the role of electrode distance of vented ionization chambers under ultra-high dose-per-pulse conditions.

PURPOSE: Investigating and understanding of the underlying mechanisms affecting the charge collection efficiency (CCE) of vented ionization chambers under ultra-high dose rate pulsed electron radiation. This is an important step towards real-time dosimetry with ionization chambers in FLASH radiotherapy. METHODS: Parallel-plate ionization chambers (PPIC) with three different electrode distances were build and investigated with electron beams with ultra-high dose-per-pulse (DPP) up to 5.4 Gy. The measurements were compared with simulations. The experimental determination of the CCE was done by comparison against the reference dose based on alanine dosimetry. The numerical solution of a system of partial differential equations taking into account charge creations by the radiation, their transport and reaction in an applied electric field was used for the simulations of the CCE and the underlying effects. RESULTS: A good agreement between the experimental results and the simulated CCE could be achieved. The recombination losses found under ultra-high DPP could be attributed to a temporal and spatial charge carrier imbalance and the associated electric field distortion. With ultra-thin electrode distances down to 0.25 mm and a suitable chamber voltage, a CCE greater than 99 % could be achieved under the ultra-high DPP conditions investigated. CONCLUSIONS: Well-guarded ultra-thin PPIC are suited for real-time dosimetry under ultra-high DPP conditions. This allows dosimetry also for FLASH RT according to common codes of practice, traceable to primary standards. The numerical approach used allows the determination of appropriate correction factors beyond the DPP ranges where established theories are applicable to account for remaining recombination losses.

The probe-format graphite calorimeter, Aerrow, for absolute dosimetry in ultrahigh pulse dose rate electron beams.

PURPOSE: The purpose of this investigation is to evaluate the use of a probe-format graphite calorimeter, Aerrow, as an absolute and relative dosimeter of high-energy pulse dose rate (UHPDR) electron beams for in-water reference and depth-dose-type measurements, respectively. METHODS: In this paper, the calorimeter system is used to investigate the potential influence of dose per pulses delivered up to 5.6 Gy, the number of pulses delivered per measurement, and its potential for relative measurement (depth-dose curve measurement). The calorimeter system is directly compared against an Advanced Markus ion chamber. The finite element method was used to calculate heat transfer corrections along the percentage depth dose of a 20-MeV electron beam. Monte Carlo-calculated dose conversion factors necessary to calculate absorbed dose-to-water at a point from the measured dose-to-graphite are also presented. RESULTS: The comparison of Aerrow against a fully calibrated Advanced Markus chamber, corrected for the saturation effect, has shown consistent results in terms of dose-to-water determination. The measured reference depth is within 0.5 mm from the expected value from Monte Carlo simulation. The relative standard uncertainty estimated for Aerrow was 1.06%, which is larger compared to alanine dosimetry (McEwen et al. https://doi.org/10.1088/0026-1394/52/2/272) but has the advantage of being a real-time detector. CONCLUSION: In this investigation, it was demonstrated that the Aerrow probe-type graphite calorimeter can be used for relative and absolute dosimetries in water in an UHPDR electron beam. To the author's knowledge, this is the first reported use of an absorbed dose calorimeter for an in-water percentage depth-dose curve measurement. The use of the Aerrow in quasi-adiabatic mode has greatly simplified the signal readout, compared to isothermal mode, as the resistance was directly measured with a high-stability digital multimeter.

Application of a novel diamond detector for commissioning of FLASH radiotherapy electron beams.

PURPOSE: A diamond detector prototype was recently proposed by Marinelli et al. (Medical Physics 2022, https://doi.org/10.1002/mp.15473) for applications in ultrahigh-dose-per-pulse (UH-DPP) and ultrahigh-dose-rate (UH-DR) beams, as used in FLASH radiotherapy (FLASH-RT). In the present study, such so-called flashDiamond (fD) was investigated from the dosimetric point of view, under pulsed electron beam irradiation. It was then used for the commissioning of an ElectronFlash linac (SIT S.p.A., Italy) both in conventional and UH-DPP modalities. METHODS: Detector calibration was performed in reference conditions, under 60 Co and electron beam irradiation. Its response linearity was investigated in UH-DPP conditions. For this purpose, the DPP was varied in the 1.2-11.9 Gy range, by changing either the beam applicator or the pulse duration from 1 to 4 µs. Dosimetric validation of the fD detector prototype was then performed in conventional modality, by measuring percentage depth dose (PDD) curves, beam profiles, and output factors (OFs). All such measurements were carried out in a motorized water phantom. The obtained results were compared with the ones from commercially available dosimeters, namely, a microDiamond, an Advanced Markus ionization chamber, a silicon diode detector, and EBT-XD GAFchromic films. Finally, the fD detector was used to fully characterize the 7 and 9 MeV UH-DPP electron beams delivered by the ElectronFlash linac. In particular, PDDs, beam profiles, and OFs were measured, for both energies and all the applicators, and compared with the ones from EBT-XD films irradiated in the same experimental conditions. RESULTS: The fD calibration coefficient resulted to be independent from the investigated beam qualities. The detector response was found to be linear in the whole investigated DPP range. A very good agreement was observed among PDDs, beam profiles, and OFs measured by the fD prototype and reference detectors, both in conventional and UH-DPP irradiation modalities. CONCLUSIONS: The fD detector prototype was validated from the dosimetric point of view against several commercial dosimeters in conventional beams. It was proved to be suitable in UH-DPP and UH-DR conditions, for which no other commercial real-time active detector is available to date. It was shown to be a very useful tool to perform fast and reproducible beam characterizations in standard clinical motorized water phantom setups. All of the previously mentioned demonstrate the suitability of the proposed detector for the commissioning of UH-DR linac beams for preclinical FLASH-RT applications.