Comprehensive dosimetric characterization of novel silicon carbide detectors with UHDR electron beams for FLASH radiotherapy.

BACKGROUND: The extremely fast delivery of doses with ultra high dose rate (UHDR) beams necessitates the investigation of novel approaches for real-time dosimetry and beam monitoring. This aspect is fundamental in the perspective of the clinical application of FLASH radiotherapy (FLASH-RT), as conventional dosimeters tend to saturate at such extreme dose rates. PURPOSE: This study aims to experimentally characterize newly developed silicon carbide (SiC) detectors of various active volumes at UHDRs and systematically assesses their response to establish their suitability for dosimetry in FLASH-RT. METHODS: SiC PiN junction detectors, recently realized and provided by STLab company, with different active areas (ranging from 4.5 to 10 mm2) and thicknesses (10-20 µm), were irradiated using 9 MeV UHDR pulsed electron beams accelerated by the ElectronFLASH linac at the Centro Pisano for FLASH Radiotherapy (CPFR). The linearity of the SiC response as a function of the delivered dose per pulse (DPP), which in turn corresponds to a specific instantaneous dose rate, was studied under various experimental conditions by measuring the produced charge within the SiC active layer with an electrometer. Due to the extremely high peak currents, an external customized electronic RC circuit was built and used in conjunction with the electrometer to avoid saturation. RESULTS: The study revealed a linear response for the different SiC detectors employed up to 21 Gy/pulse for SiC detectors with 4.5 mm2/10 µm active area and thickness. These values correspond to a maximum instantaneous dose rate of 5.5 MGy/s and are indicative of the maximum achievable monitored DPP and instantaneous dose rate of the linac used during the measurements. CONCLUSIONS: The results clearly demonstrate that the developed devices exhibit a dose-rate independent response even under extreme instantaneous dose rates and dose per pulse values. A systematic study of the SiC response was also performed as a function of the applied voltage bias, demonstrating the reliability of these dosimeters with UHDR also without any applied voltage. This demonstrates the great potential of SiC detectors for accurate dosimetry in the context of FLASH-RT.

3D Bioprinting and Microfluidic-Based Devices for Cancer Detection and Drug Treatment: Focus on Prostate Cancer.

The burden of increasing cancer incidence among the population, and, in particular, of prostate cancer in men living in highly developed countries, brings with it, on one hand, the need for new devices that allow a faster and earlier diagnosis, ideally in a non-invasive way and with low consumption of expensive reagents, and on the other the need for the assessment of new in vitro models that allow a more reliable assessment of cancer features, including its microenvironment and sensibility to different drugs. At the crossroads of these features, microfluidic devices are found. These, taking advantage of the chemical-physical properties of cells and human samples, have demonstrated great sensitivity and sensibility at an on-chip scale. Many fields of biomedical sciences have tried to exploit all their potentialities: from the detection of antigens in the early phases of the disease (when they are very low concentrated, but the treatment is more effective) to isolation and characterization of circulating tumor cells. However, the development of in vitro 3D models to better assess and comprehend the fundamental dynamics of tumor microenvironment and metastasis using 3D bioprinting techniques. The aim of the present review is to describe the potential of these two different cutting-edge technologies for the detection and treatment of prostate cancer, in the perspective of a possible future combination of them that allows scientists to fill the gaps present in the field to improve patient care and treatment.

Single-Ion Counting with an Ultra-Thin-Membrane Silicon Carbide Sensor.

In recent times, ion implantation has received increasing interest for novel applications related to deterministic material doping on the nanoscale, primarily for the fabrication of solid-state quantum devices. For such applications, precise information concerning the number of implanted ions and their final position within the implanted sample is crucial. In this work, we present an innovative method for the detection of single ions of MeV energy by using a sub-micrometer ultra-thin silicon carbide sensor operated as an in-beam counter of transmitted ions. The SiC sensor signals, when compared to a Passivated Implanted Planar Silicon detector signal, exhibited a 96.5% ion-detection confidence, demonstrating that the membrane sensors can be utilized for high-fidelity ion counting. Furthermore, we assessed the angular straggling of transmitted ions due to the interaction with the SiC sensor, employing the scanning knife-edge method of a focused ion microbeam. The lateral dimension of the ion beam with and without the membrane sensor was compared to the SRIM calculations. The results were used to discuss the potential of such experimental geometry in deterministic ion-implantation schemes as well as other applications.

Microfluidic/HPLC combination to study carnosine protective activity on challenged human microglia: Focus on oxidative stress and energy metabolism.

Carnosine (ß-alanyl-L-histidine) is a naturally occurring endogenous peptide widely distributed in excitable tissues such as the brain. This dipeptide possesses well-demonstrated antioxidant, anti-inflammatory, and anti-aggregation properties, and it may be useful for treatment of pathologies characterized by oxidative stress and energy unbalance such as depression and Alzheimer's disease (AD). Microglia, the brain-resident macrophages, are involved in different physiological brain activities such synaptic plasticity and neurogenesis, but their dysregulation has been linked to the pathogenesis of numerous diseases. In AD brain, the activation of microglia towards a pro-oxidant and pro-inflammatory phenotype has found in an early phase of cognitive decline, reason why new pharmacological targets related to microglia activation are of great importance to develop innovative therapeutic strategies. In particular, microglia represent a common model of lipopolysaccharides (LPS)-induced activation to identify novel pharmacological targets for depression and AD and numerous studies have linked the impairment of energy metabolism, including ATP dyshomeostasis, to the onset of depressive episodes. In the present study, we first investigated the toxic potential of LPS + ATP in the absence or presence of carnosine. Our studies were carried out on human microglia (HMC3 cell line) in which LPS + ATP combination has shown the ability to promote cell death, oxidative stress, and inflammation. Additionally, to shed more light on the molecular mechanisms underlying the protective effect of carnosine, its ability to modulate reactive oxygen species production and the variation of parameters representative of cellular energy metabolism was evaluated by microchip electrophoresis coupled to laser-induced fluorescence and high performance liquid chromatography, respectively. In our experimental conditions, carnosine prevented LPS + ATP-induced cell death and oxidative stress, also completely restoring basal energy metabolism in human HMC3 microglia. Our results suggest a therapeutic potential of carnosine as a new pharmacological tool in the context of multifactorial disorders characterize by neuroinflammatory phenomena including depression and AD.

Radiation Hardness Study of Silicon Carbide Sensors under High-Temperature Proton Beam Irradiations.

Silicon carbide (SiC), thanks to its material properties similar to diamond and its industrial maturity close to silicon, represents an ideal candidate for several harsh-environment sensing applications, where sensors must withstand high particle irradiation and/or high operational temperatures. In this study, to explore the radiation tolerance of SiC sensors to multiple damaging processes, both at room and high temperature, we used the Ion Microprobe Chamber installed at the Ruder Boskovic Institute (Zagreb, Croatia), which made it possible to expose small areas within the same device to different ion beams, thus evaluating and comparing effects within a single device. The sensors tested, developed jointly by STLab and SenSiC, are PIN diodes with ultrathin free-standing membranes, realized by means of a recently developed doping-selective electrochemical etching. In this work, we report on the changes of the charge transport properties, specifically in terms of the charge collection efficiency (CCE), with respect to multiple localized proton irradiations, performed at both room temperature (RT) and 500 °C.

Impact of buffer composition on biochemical, morphological and mechanical parameters: A tare before dielectrophoretic cell separation and isolation.

Dielectrophoresis (DEP) represents an electrokinetic approach for discriminating and separating suspended cells based on their intrinsic dielectric characteristics without the need for labeling procedure. A good practice, beyond the physical and engineering components, is the selection of a buffer that does not hinder cellular and biochemical parameters as well as cell recovery. In the present work the impact of four buffers on biochemical, morphological, and mechanical parameters was evaluated in two different cancer cell lines (Caco-2 and K562). Specifically, MTT ([3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide]) assay along with flow cytometry analysis were used to evaluate the occurring changes in terms of cell viability, morphology, and granulocyte stress formation, all factors directly influencing DEP sorting capability. Quantitative real-time PCR (qRT-PCR) was instead employed to evaluate the gene expression levels of interleukin-6 (IL-6) and inducible nitric oxide synthase (iNOS), two well-known markers of inflammation and oxidative stress, respectively. An additional marker representing an index of cellular metabolic status, i.e. the expression of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene, was also evaluated. Among the four buffers considered, two resulted satisfactory in terms of cell viability and growth recovery (24 h), with no significant changes in cell morphology for up to 1 h in suspension. Of note, gene expression analysis showed that in both cell lines the apparently non-cytotoxic buffers significantly modulated IL-6, iNOS, and GAPDH markers, underlining the importance to deeply investigate the molecular and biochemical changes occurring during the analysis, even at apparently non-toxic conditions. The selection of a useful buffer for the separation and analysis of cells without labeling procedures, preserving cell status, represents a key factor for DEP analysis, giving the opportunity to further use cells for additional analysis.

PD-1, PD-L1 and cAMP immunohistochemical expressions are associated with worse oncological outcome in patients with bladder cancer.

PURPOSE: In this study, we aimed to identify prognostic factors of cancer mortality in patients who received radical cystectomy and to identify genomic alterations in a sub-cohort of patients with locally advanced (pT3-4) and/or positive lymph nodes bladder cancer (BC). METHODS: We collected 101 BC samples from 2010 to 2018 who previously received radical cystectomy. Immunohistochemical slides were evaluated for PPAR, cAMP, IMP3, Ki67, CDK4, POU5F1, Cyclin E and MDM2, p65, CD3, CD4, CD8, CD20, CD68, CD163, FOXP3, PD-1 and PD-L1 expression. We calculated a prognostic score (PS) based on the positivity to PD-1, PD-L1 and of cAMP (final score ranging from 0 to 3). DNA of each sample have been used for sequencing by NGS in a sub-cohort of 6 patients with locally advanced (pT3-4) and/or positive lymph nodes BC. RESULTS: PD-1 + (HR [hazard ratio] 2.59; p = 0.04), PD-L1+ (HR = 6.46; p < 0.01) and cAMP+ (HR 3.04; p = 0.02) were independent predictors of cancer-specific mortality (CSM). Increase of PS (score = 0 as reference) was associated with CSM, 0.81 (p = 0.80), 4.72 (p = 0.01) and 10.51 (p < 0.0) for PS 1, 2 and 3, respectively. ERBB2 was the gene most frequently mutated. CONCLUSION: BC exhibited heterogenous protein expression and variable genomic features. Identification of expression of PD-1, PD-L1 and cAMP could help in predicting oncological outcomes.

Label-Free Enrichment of Circulating Tumor Plasma Cells: Future Potential Applications of Dielectrophoresis in Multiple Myeloma.

In multiple myeloma (MM), circulating tumor plasma cells (CTPCs) are an emerging prognostic factor, offering a promising and minimally invasive means for longitudinal patient monitoring. Recent advances highlight the complex biology of plasma cell trafficking, highlighting the phenotypic and genetic signatures of intra- and extra-medullary MM onset, making CTPC enumeration and characterization a new frontier of precision medicine for MM patients, requiring novel technological platforms for their standardized and harmonized detection. Dielectrophoresis (DEP) is an emerging label-free cell manipulation technique to separate cancer cells from healthy cells in peripheral blood samples, based on phenotype and membrane capacitance that could be successfully tested to enumerate and isolate CTPCs. Herein, we summarize preclinical data on DEP development for CTPC detection, as well as their clinical and research potential.

The Role of Dielectrophoresis for Cancer Diagnosis and Prognosis.

Liquid biopsy is emerging as a potential diagnostic tool for prostate cancer (PC) prognosis and diagnosis. Unfortunately, most circulating tumor cells (CTC) technologies, such as AdnaTest or Cellsearch®, critically rely on the epithelial cell adhesion molecule (EpCAM) marker, limiting the possibility of detecting cancer stem-like cells (CSCs) and mesenchymal-like cells (EMT-CTCs) that are present during PC progression. In this context, dielectrophoresis (DEP) is an epCAM independent, label-free enrichment system that separates rare cells simply on the basis of their specific electrical properties. As compared to other technologies, DEP may represent a superior technique in terms of running costs, cell yield and specificity. However, because of its higher complexity, it still requires further technical as well as clinical development. DEP can be improved by the use of microfluid, nanostructured materials and fluoro-imaging to increase its potential applications. In the context of cancer, the usefulness of DEP lies in its capacity to detect CTCs in the bloodstream in their epithelial, mesenchymal, or epithelial-mesenchymal phenotype forms, which should be taken into account when choosing CTC enrichment and analysis methods for PC prognosis and diagnosis.

Investigation of the Young's Modulus and the Residual Stress of 4H-SiC Circular Membranes on 4H-SiC Substrates.

The stress state is a crucial parameter for the design of innovative microelectromechanical systems based on silicon carbide (SiC) material. Hence, mechanical properties of such structures highly depend on the fabrication process. Despite significant progresses in thin-film growth and fabrication process, monitoring the strain of the suspended SiC thin-films is still challenging. However, 3C-SiC membranes on silicon (Si) substrates have been demonstrated, but due to the low quality of the SiC/Si heteroepitaxy, high levels of residual strains were always observed. In order to achieve promising self-standing films with low residual stress, an alternative micromachining technique based on electrochemical etching of high quality homoepitaxy 4H-SiC layers was evaluated. This work is dedicated to the determination of their mechanical properties and more specifically, to the characterization of a 4H-SiC freestanding film with a circular shape. An inverse problem method was implemented, where experimental results obtained from bulge test are fitted with theoretical static load-deflection curves of the stressed membrane. To assess data validity, the dynamic behavior of the membrane was also investigated: Experimentally, by means of laser Doppler vibrometry (LDV) and theoretically, by means of finite element computations. The two methods provided very similar results since one obtained a Young's modulus of 410 GPa and a residual stress value of 41 MPa from bulge test against 400 GPa and 30 MPa for the LDV analysis. The determined Young's modulus is in good agreement with literature values. Moreover, residual stress values demonstrate that the fabrication of low-stressed SiC films is achievable thanks to the micromachining process developed.

Silicon carbide X-ray beam position monitors for synchrotron applications.

In this work, the performance of thin silicon carbide membranes as material for radiation hard X-ray beam position monitors (XBPMs) is investigated. Thermal and electrical behavior of XBPMs made from thin silicon carbide membranes and single-crystal diamond is compared using finite-element simulations. Fabricated silicon carbide devices are also compared with a 12 µm commercial polycrystalline diamond XBPM at the Swiss Light Source at the Paul Scherrer Institute. Results show that silicon carbide devices can reach equivalent transparencies while showing improved linearity, dynamics and signal-to-noise ratio compared with commercial polycrystalline diamond XBPMs. Given the obtained results and availability of electronic-grade epitaxies on up to 6 inch wafers, it is expected that silicon carbide can substitute for diamond in most beam monitoring applications, whereas diamond, owing to its lower absorption, could remain the material of choice in cases of extreme X-ray power densities, such as pink and white beams.

SiCILIA-Silicon Carbide Detectors for Intense Luminosity Investigations and Applications.

Silicon carbide (SiC) is a compound semiconductor, which is considered as a possible alternative to silicon for particles and photons detection. Its characteristics make it very promising for the next generation of nuclear and particle physics experiments at high beam luminosity. Silicon Carbide detectors for Intense Luminosity Investigations and Applications (SiCILIA) is a project starting as a collaboration between the Italian National Institute of Nuclear Physics (INFN) and IMM-CNR, aiming at the realization of innovative detection systems based on SiC. In this paper, we discuss the main features of silicon carbide as a material and its potential application in the field of particles and photons detectors, the project structure and the strategies used for the prototype realization, and the first results concerning prototype production and their performance.

Analysis of the role of the particle-wall interaction on the separation efficiencies of field flow fractionation dielectrophoretic devices.

In this paper we have used both analytical models and finite element simulations to analyze the role of the particle-wall dipole interaction in field-flow fractionation dielectrophoretic (FFF-DEP) devices. We identify the existence of "anomalous" regions where the dielectrophoretic response is altered, independently of the complex dielectric permittivity of the particles and suspending medium. In these regions the interaction between the particle and the conductive (isolating) walls induces cohesive (repulsive) forces, independently of the Clausius-Mossotti term. We quantify the impact of such an effect, which can critically decrease the specificity and sensitivity of both continuous- and batch-mode FFF-DEP. We find a scale invariant relation correlating the particles radius (Rp ) and the electrodes width (Wel ), which permits the design of dielectrophoretic schema capable of avoiding the generation of such regions. Specifically, to avoid the generation of the anomalous DEP regions, Wel should be chosen smaller than ∼5.2 Rp . For this reason, interdigitate schema with electrode widths of 14 µm and gaps of 50 µm could improve the separation efficiency of FFF-DEP devices in the case of rare cells separation in blood samples.

Theoretical and experimental study of the role of cell-cell dipole interaction in dielectrophoretic devices: application to polynomial electrodes.

BACKGROUND: We aimed to investigate the effect of cell-cell dipole interactions in the equilibrium distributions in dielectrophoretic devices. METHODS: We used a three dimensional coupled Monte Carlo-Poisson method to theoretically study the final distribution of a system of uncharged polarizable particles suspended in a static liquid medium under the action of an oscillating non-uniform electric field generated by polynomial electrodes. The simulated distributions have been compared with experimental ones observed in the case of MDA-MB-231 cells in the same operating conditions. RESULTS: The real and simulated distributions are consistent. In both cases the cells distribution near the electrodes is dominated by cell-cell dipole interactions which generate long chains. CONCLUSIONS: The agreement between real and simulated cells' distributions demonstrate the method's reliability. The distribution are dominated by cell-cell dipole interactions even at low density regimes (105 cell/ml). An improved estimate for the density threshold governing the interaction free regime is suggested.