Stiffness-Controlled Hydrogels for 3D Cell Culture Models.

Nanofibrillated cellulose (NFC) hydrogel is a versatile biomaterial suitable, for example, for three-dimensional (3D) cell spheroid culturing, drug delivery, and wound treatment. By freeze-drying NFC hydrogel, highly porous NFC structures can be manufactured. We freeze-dried NFC hydrogel and subsequently reconstituted the samples into a variety of concentrations of NFC fibers, which resulted in different stiffness of the material, i.e., different mechanical cues. After the successful freeze-drying and reconstitution, we showed that freeze-dried NFC hydrogel can be used for one-step 3D cell spheroid culturing of primary mesenchymal stem/stromal cells, prostate cancer cells (PC3), and hepatocellular carcinoma cells (HepG2). No difference was observed in the viability or morphology between the 3D cell spheroids cultured in the freeze-dried and reconstituted NFC hydrogel and fresh NFC hydrogel. Furthermore, the 3D cultured spheroids showed stable metabolic activity and nearly 100% viability. Finally, we applied a convolutional neural network (CNN)-based automatic nuclei segmentation approach to automatically segment individual cells of 3D cultured PC3 and HepG2 spheroids. These results provide an application to culture 3D cell spheroids more readily with the NFC hydrogel and a step towards automatization of 3D cell culturing and analysis.

Marker-controlled watershed with deep edge emphasis and optimized H-minima transform for automatic segmentation of densely cultivated 3D cell nuclei.

BACKGROUND: The segmentation of 3D cell nuclei is essential in many tasks, such as targeted molecular radiotherapies (MRT) for metastatic tumours, toxicity screening, and the observation of proliferating cells. In recent years, one popular method for automatic segmentation of nuclei has been deep learning enhanced marker-controlled watershed transform. In this method, convolutional neural networks (CNNs) have been used to create nuclei masks and markers, and the watershed algorithm for the instance segmentation. We studied whether this method could be improved for the segmentation of densely cultivated 3D nuclei via developing multiple system configurations in which we studied the effect of edge emphasizing CNNs, and optimized H-minima transform for mask and marker generation, respectively. RESULTS: The dataset used for training and evaluation consisted of twelve in vitro cultivated densely packed 3D human carcinoma cell spheroids imaged using a confocal microscope. With this dataset, the evaluation was performed using a cross-validation scheme. In addition, four independent datasets were used for evaluation. The datasets were resampled near isotropic for our experiments. The baseline deep learning enhanced marker-controlled watershed obtained an average of 0.69 Panoptic Quality (PQ) and 0.66 Aggregated Jaccard Index (AJI) over the twelve spheroids. Using a system configuration, which was otherwise the same but used 3D-based edge emphasizing CNNs and optimized H-minima transform, the scores increased to 0.76 and 0.77, respectively. When using the independent datasets for evaluation, the best performing system configuration was shown to outperform or equal the baseline and a set of well-known cell segmentation approaches. CONCLUSIONS: The use of edge emphasizing U-Nets and optimized H-minima transform can improve the marker-controlled watershed transform for segmentation of densely cultivated 3D cell nuclei. A novel dataset of twelve spheroids was introduced to the public.

Automated daily quality control analysis for mammography in a multi-unit imaging center.

BACKGROUND: The high requirements for mammography image quality necessitate a systematic quality assurance process. Digital imaging allows automation of the image quality analysis, which can potentially improve repeatability and objectivity compared to a visual evaluation made by the users. PURPOSE: To develop an automatic image quality analysis software for daily mammography quality control in a multi-unit imaging center. MATERIAL AND METHODS: An automated image quality analysis software using the discrete wavelet transform and multiresolution analysis was developed for the American College of Radiology accreditation phantom. The software was validated by analyzing 60 randomly selected phantom images from six mammography systems and 20 phantom images with different dose levels from one mammography system. The results were compared to a visual analysis made by four reviewers. Additionally, long-term image quality trends of a full-field digital mammography system and a computed radiography mammography system were investigated. RESULTS: The automated software produced feature detection levels comparable to visual analysis. The agreement was good in the case of fibers, while the software detected somewhat more microcalcifications and characteristic masses. Long-term follow-up via a quality assurance web portal demonstrated the feasibility of using the software for monitoring the performance of mammography systems in a multi-unit imaging center. CONCLUSION: Automated image quality analysis enables monitoring the performance of digital mammography systems in an efficient, centralized manner.

Multicellular dosimetric chain for molecular radiotherapy exemplified with dose simulations on 3D cell spheroids.

PURPOSE: Absorbed radiation dose-response relationships are not clear in molecular radiotherapy (MRT). Here, we propose a voxel-based dose calculation system for multicellular dosimetry in MRT. We applied confocal microscope images of a spherical cell aggregate i.e. a spheroid, to examine the computation of dose distribution within a tissue from the distribution of radiopharmaceuticals. METHODS: A confocal microscope Z-stack of a human hepatocellular carcinoma HepG2 spheroid was segmented using a support-vector machine algorithm and a watershed function. Heterogeneity in activity uptake was simulated by selecting a varying amount of the cell nuclei to contain 111In, 125I, or 177Lu. Absorbed dose simulations were carried out using vxlPen, a software application based on the Monte Carlo code PENELOPE. RESULTS: We developed a schema for radiopharmaceutical dosimetry. The schema utilizes a partially supervised segmentation method for cell-level image data together with a novel main program for voxel-based radiation dose simulations. We observed that for 177Lu, radiation cross-fire enabled full dose coverage even if the radiopharmaceutical had accumulated to only 60% of the spheroid cells. This effect was not found with 111In and 125I. Using these Auger/internal conversion electron emitters seemed to guarantee that only the cells with a high enough activity uptake will accumulate a lethal amount of dose, while neighboring cells are spared. CONCLUSIONS: We computed absorbed radiation dose distributions in a 3D-cultured cell spheroid with a novel multicellular dosimetric chain. Combined with pharmacological studies in different tissue models, our cell-level dosimetric calculation method can clarify dose-response relationships for radiopharmaceuticals used in MRT.

Analysis of MCNP simulated gamma spectra of CdTe detectors for boron neutron capture therapy.

The next step in the boron neutron capture therapy (BNCT) is the real time imaging of the boron concentration in healthy and tumor tissue. Monte Carlo simulations are employed to predict the detector response required to realize single-photon emission computed tomography in BNCT, but have failed to correctly resemble measured data for cadmium telluride detectors. In this study we have tested the gamma production cross-section data tables of commonly used libraries in the Monte Carlo code MCNP in comparison to measurements. The cross section data table TENDL-2008-ACE is reproducing measured data best, whilst the commonly used ENDL92 and other studied libraries do not include correct tables for the gamma production from the cadmium neutron capture reaction that is occurring inside the detector. Furthermore, we have discussed the size of the annihilation peaks of spectra obtained by cadmium telluride and germanium detectors.

Prompt gamma and neutron detection in BNCT utilizing a CdTe detector.

In this work, a novel sensor technology based on CdTe detectors was tested for prompt gamma and neutron detection using boronated targets in (epi)thermal neutron beam at FiR1 research reactor in Espoo, Finland. Dedicated neutron filter structures were omitted to enable simultaneous measurement of both gamma and neutron radiation at low reactor power (2.5 kW). Spectra were collected and analyzed in four different setups in order to study the feasibility of the detector to measure 478 keV prompt gamma photons released from the neutron capture reaction of boron-10. The detector proved to have the required sensitivity to detect and separate the signals from both boron neutron and cadmium neutron capture reactions, which makes it a promising candidate for monitoring the spatial and temporal development of in vivo boron distribution in boron neutron capture therapy.

Boron neutron capture therapy (BNCT) in Finland: technological and physical prospects after 20 years of experiences.

Boron Neutron Capture Therapy (BNCT) is a binary radiotherapy method developed to treat patients with certain malignant tumours. To date, over 300 treatments have been carried out at the Finnish BNCT facility in various on-going and past clinical trials. In this technical review, we discuss our research work in the field of medical physics to form the groundwork for the Finnish BNCT patient treatments, as well as the possibilities to further develop and optimize the method in the future. Accordingly, the following aspects are described: neutron sources, beam dosimetry, treatment planning, boron imaging and determination, and finally the possibilities to detect the efficacy and effects of BNCT on patients.

Boron neutron capture therapy in the treatment of locally recurred head-and-neck cancer: final analysis of a phase I/II trial.

PURPOSE: To investigate the efficacy and safety of boron neutron capture therapy (BNCT) in the treatment of inoperable head-and-neck cancers that recur locally after conventional photon radiation therapy. METHODS AND MATERIALS: In this prospective, single-center Phase I/II study, 30 patients with inoperable, locally recurred head-and-neck cancer (29 carcinomas and 1 sarcoma) were treated with BNCT. Prior treatments consisted of surgery and conventionally fractionated photon irradiation to a cumulative dose of 50 to 98 Gy administered with or without concomitant chemotherapy. Tumor responses were assessed by use of the RECIST (Response Evaluation Criteria in Solid Tumors) and adverse effects by use of the National Cancer Institute common terminology criteria version 3.0. Intravenously administered L-boronophenylalanine-fructose (400 mg/kg) was administered as the boron carrier. Each patient was scheduled to be treated twice with BNCT. RESULTS: Twenty-six patients received BNCT twice; four were treated once. Of the 29 evaluable patients, 22 (76%) responded to BNCT, 6 (21%) had tumor growth stabilization for 5.1 and 20.3 months, and 1 (3%) progressed. The median progression-free survival time was 7.5 months (95% confidence interval, 5.4-9.6 months). Two-year progression-free survival and overall survival were 20% and 30%, respectively, and 27% of the patients survived for 2 years without locoregional recurrence. The most common acute Grade 3 adverse effects were mucositis (54% of patients), oral pain (54%), and fatigue (32%). Three patients were diagnosed with osteoradionecrosis (each Grade 3) and one patient with soft-tissue necrosis (Grade 4). Late Grade 3 xerostomia was present in 3 of the 15 evaluable patients (20%). CONCLUSIONS: Most patients who have inoperable, locally advanced head-and-neck carcinoma that has recurred at a previously irradiated site respond to boronophenylalanine-mediated BNCT, but cancer recurrence after BNCT remains frequent. Toxicity was acceptable. Further research on novel modifications of the method is warranted.

Transition in occupational radiation exposure monitoring methods in diagnostic and interventional radiology.

Radiation exposure monitoring is a traditional keystone of occupational radiation safety measures in medical imaging. The aim of this study was to review the data on occupational exposures in a large central university hospital radiology organisation and propose changes in the radiation worker categories and methods of exposure monitoring. An additional objective was to evaluate the development of electronic personal dosimeters and their potential in the digitised radiology environment. The personal equivalent dose of 267 radiation workers (116 radiologists and 151 radiographers) was monitored using personal dosimeters during the years 2006-2010. Accumulated exposure monitoring results exceeding the registration threshold were observed in the personal dosimeters of 73 workers (59 radiologists' doses ranged from 0.1 to 45.1 mSv; 14 radiographers' doses ranged from 0.1 to 1.3 mSv). The accumulated personal equivalent doses are generally very small, only a few angiography radiologists have doses >10 mSv per 5 y. The typical effective doses are <10 µSv y(-1) and the highest value was 0.3 mSv (single interventional radiologist). A revised categorisation of radiation workers based on the working profile of the radiologist and observed accumulated doses is justified. Occupational monitoring can be implemented mostly with group dosimeters. An active real-time dosimetry system is warranted to support radiation protection strategy where optimisation aspects, including improving working methods, are essential.

MRI quality assurance using the ACR phantom in a multi-unit imaging center.

BACKGROUND: Magnetic resonance imaging (MRI) instrumentation is vulnerable to technical and image quality problems, and quality assurance is essential. In the studied regional imaging center the long-term quality assurance has been based on MagNET phantom measurements. American College of Radiology (ACR) has an accreditation program including a standardized image quality measurement protocol and phantom. The ACR protocol includes recommended acceptance criteria for clinical sequences and thus provides possibility to assess the clinical relevance of quality assurance. The purpose of this study was to test the ACR MRI phantom in quality assurance of a multi-unit imaging center. MATERIAL AND METHODS: The imaging center operates 11 MRI systems of three major manufacturers with field strengths of 3.0 T, 1.5 T and 1.0 T. Images of the ACR phantom were acquired using a head coil following the ACR scanning instructions. Both ACR T1- and T2-weighted sequences as well as T1- and T2-weighted brain sequences in clinical use at each site were acquired. Measurements were performed twice. The images were analyzed and the results were compared with the ACR acceptance levels. RESULTS: The acquisition procedure with the ACR phantom was faster than with the MagNET phantoms. On the first and second measurement rounds 91% and 73% of the systems passed the ACR test. Measured slice thickness accuracies were not within the acceptance limits in site T2 sequences. Differences in the high contrast spatial resolution between the ACR and the site sequences were observed. In 3.0 T systems the image intensity uniformity was slightly lower than the ACR acceptance limit. CONCLUSION: The ACR method was feasible in quality assurance of a multi-unit imaging center and the ACR protocol could replace the MagNET phantom tests. An automatic analysis of the images will further improve cost-effectiveness and objectiveness of the ACR protocol.

In vitro studies on stability of L-p-boronophenylalanine-fructose complex (BPA-F).

Boron neutron capture therapy (BNCT) is an experimental drug-targeted treatment combining tumour-seeking boronated drug(s) and subsequent ¹°B activation by neutrons. Synthetic amino acid, L-p-boronophenylalanine (BPA), administered as a fructose complex (BPA-F) is used in BNCT trials. We tested the in vitro biological and structural stability of the BPA-F as a function of time (11 days). The BPA-F samples were analyzed using biological bacterial endotoxin and sterility tests. Visual tests were clarity, degree of opalescence and coloration according to European Pharmacopoeia. The structural stability of the BPA-F was monitored via ¹H NMR signal intensity of the aromatic protons of the BPA-F samples. A slight change of BPA-F samples in coloration was observed during storage. BPA-fructose complex remained sterile and bacterial endotoxin tests were negative. In the end of study period, relative intensity of the (1)H NMR signals of the BPA-F sample was ≥ 90% of the initial relative intensity. The biological properties of the BPA-fructose complex and chemical structure of the complex remained the same during the test period. Visually the solutions stayed clear, but there was a slight change in colour. The tests indicate that the prepared batch of BPA-F complex can be used for the patient infusion at least for a week.

L-boronophenylalanine-mediated boron neutron capture therapy for malignant glioma progressing after external beam radiation therapy: a Phase I study.

PURPOSE: To investigate the safety of boronophenylalanine-mediated boron neutron capture therapy (BNCT) in the treatment of malignant gliomas that progress after surgery and conventional external beam radiation therapy. METHODS AND MATERIALS: Adult patients who had histologically confirmed malignant glioma that had progressed after surgery and external beam radiotherapy were eligible for this Phase I study, provided that >6 months had elapsed from the last date of radiation therapy. The first 10 patients received a fixed dose, 290 mg/kg, of L-boronophenylalanine-fructose (L-BPA-F) as a 2-hour infusion before neutron irradiation, and the remaining patients were treated with escalating doses of L-BPA-F, either 350 mg/kg, 400 mg/kg, or 450 mg/kg, using 3 patients on each dose level. Adverse effects were assessed using National Cancer Institute Common Toxicity Criteria version 2.0. RESULTS: Twenty-two patients entered the study. Twenty subjects had glioblastoma, and 2 patients had anaplastic astrocytoma, and the median cumulative dose of prior external beam radiotherapy was 59.4 Gy. The maximally tolerated L-BPA-F dose was reached at the 450 mg/kg level, where 4 of 6 patients treated had a grade 3 adverse event. Patients who were given >290 mg/kg of L-BPA-F received a higher estimated average planning target volume dose than those who received 290 mg/kg (median, 36 vs. 31 Gy [W, i.e., a weighted dose]; p = 0.018). The median survival time following BNCT was 7 months. CONCLUSIONS: BNCT administered with an l-BPA-F dose of up to 400 mg/kg as a 2-hour infusion is feasible in the treatment of malignant gliomas that recur after conventional radiation therapy.

Validation of dose planning calculations for boron neutron capture therapy using cylindrical and anthropomorphic phantoms.

In this paper, the accuracy of dose planning calculations for boron neutron capture therapy (BNCT) of brain and head and neck cancer was studied at the FiR 1 epithermal neutron beam. A cylindrical water phantom and an anthropomorphic head phantom were applied with two beam aperture-to-surface distances (ASD). The calculations using the simulation environment for radiation application (SERA) treatment planning system were compared to neutron activation measurements with Au and Mn foils, photon dose measurements with an ionization chamber and the reference simulations with the MCNP5 code. Photon dose calculations using SERA differ from the ionization chamber measurements by 2-13% (disagreement increased along the depth in the phantom), but are in agreement with the MCNP5 calculations within 2%. The (55)Mn(n,gamma) and (197)Au(n,gamma) reaction rates calculated using SERA agree within 10% and 8%, respectively, with the measurements and within 5% with the MCNP5 calculations at depths >0.5 cm from the phantom surface. The (55)Mn(n,gamma) reaction rate represents the nitrogen and boron depth dose within 1%. Discrepancy in the SERA fast neutron dose calculation (of up to 37%) is corrected if the biased fast neutron dose calculation option is not applied. Reduced voxel cell size (6 cm from the phantom surface. Increasing discrepancy along the phantom depth is expected to be caused by the inaccurately determined effective point of the ionization chamber.

Acquisition-weighted MRSI for detection and quantification of BNCT 10B-carrier L-p-boronophenylalanine-fructose complex, a phantom study.

Proton magnetic resonance spectroscopy (1H MRS) is a potential method to detect and quantify a boron neutron capture therapy 10B-carrier compound, L-p-boronophenylalanine (BPA), in the brain. However, optimal positioning of MRS voxel to capture tissue with maximal BPA concentration can be challenging. Three dimensional proton magnetic resonance spectroscopic imaging (3D 1H MRSI) provides spectral data covering a large spatial volume, which is a major advantage in detecting and quantifying BPA. BPA detection limit in phantom conditions was determined at 1.5 T using a 3D 1H MRSI protocol with clinically acceptable nominal spatial resolution and duration. Quantification tests for aqueous phantom were performed using both single voxel MRS and 3D MRSI. In 3D MRSI, BPA detection limit was approximately 1.0 mM and BPA quantification accuracy was better than +/-5%. The results suggest that MRSI would be a feasible method for in vivo BPA evaluation in clinical conditions.

Boron neutron capture therapy in the treatment of locally recurred head and neck cancer.

PURPOSE: Head and neck carcinomas that recur locally after conventional irradiation pose a difficult therapeutic problem. We evaluated safety and efficacy of boron neutron capture therapy (BNCT) in the treatment of such cancers. METHODS AND MATERIALS: Twelve patients with inoperable, recurred, locally advanced (rT3, rT4, or rN2) head and neck cancer were treated with BNCT in a prospective, single-center Phase I-II study. Prior treatments consisted of surgery and conventionally fractionated photon irradiation to a cumulative dose of 56-74 Gy administered with or without concomitant chemotherapy. Tumor responses were assessed using the RECIST (Response Evaluation Criteria in Solid Tumors) criteria and adverse effects using the National Cancer Institute common toxicity grading v3.0. Intravenously administered boronophenylalanine-fructose (BPA-F, 400 mg/kg) was used as the boron carrier. Each patient was scheduled to be treated twice with BNCT. RESULTS: Ten patients received BNCT twice; 2 were treated once. Ten (83%) patients responded to BNCT, and 2 (17%) had tumor growth stabilization for 5.5 and 7.6 months. The median duration of response was 12.1 months; six responses were ongoing at the time of analysis or death (range, 4.9-19.2 months). Four (33%) patients were alive without recurrence with a median follow-up of 14.0 months (range, 12.8-19.2 months). The most common acute adverse effects were mucositis, fatigue, and local pain; 2 patients had a severe (Grade 3) late adverse effect (xerostomia, 1; dysphagia, 1). CONCLUSIONS: Boron neutron capture therapy is effective and safe in the treatment of inoperable, locally advanced head and neck carcinomas that recur at previously irradiated sites.

MAGIC polymer gel for dosimetric verification in boron neutron capture therapy.

Radiation sensitive polymer gels are among the most promising three-dimensional dose verification tools developed to date. Polymer gel dosimeter known by the acronym MAGIC has been tested for evaluation of its use in boron neutron capture (BNCT) dosimetry. We irradiated a large (diameter 10 cm, length 20 cm) cylindrical gel phantom in the epithermal neutron beam of the Finnish BNCT facility at the FiR 1 nuclear reactor. Neutron irradiation was simulated with a Monte Carlo radiation transport code MCNP. Gel samples from the same production batch were also irradiated with 6 MV photons from a medical linear accelerator to compare dose response in the two different types of beams. Irradiated gel phantoms were imaged using MRI to determine their relaxation rate R2 maps. The measured and normalized dose distribution in the epithermal neutron beam was compared to the dose distribution calculated by computer simulation. The results support the feasibility MAGIC gel in BNCT dosimetry.

The impact of PET and SPECT on dosimetry for targeted radionuclide therapy.

Targeted radionuclide therapy (TRT) is an increasingly used treatment modality for a range of cancers. To date, few treatments have involved the use of dosimetry either to plan treatment or to retrospectively ascertain the absorbed dose delivered during treatment. Also the correlation between absorbed dose and biological effect has been difficult to establish. Tomographic methods permit the determination of the activity volume on a macroscopic scale at different time points. Proper attenuation correction in tomographic imaging requires a patient-specific attenuation map. This can be obtained from scintillation-camera transmission scanning, CT or by using segmented scatter-emission images. Attenuation corrections can be performed either on the projection images, on the reconstructed images, or as part of an iterative reconstruction method. The problem of image quantification for therapy radionuclides, particularly for I-131, is exacerbated by the fact that most cameras are optimised for diagnostic imaging with Tc-99m. In addition, problems may arise when high activities are to be measured due to count losses and mis-positioned events, because of insufficient pile-up and dead time correction methods. Sufficient image quantification, however is only possible if all effects that degrade the quantitative content of the image have been corrected for. Monte Carlo simulations are an appealing tool that can help to model interactions occurring in the patient or in the detector system. This is helpful to develop and test correction techniques, or to help to define detectors better suited to quantitative imaging. PET is probably the most accurate imaging method for the determination of activity concentrations in tissue. PET imaging can be considered for pre-therapeutic treatment planning but ideally requires the use of a radioisotope from the same element as that used for treatment (e.g. I-124 for I-131; Y-86 for Y-90). Problems, however are that--some of the positron emitting isotopes have a shorter half-life--non-standard quantification procedures have to be performed--the availability of the radiopharmaceutical is presently limited; Many 3D-tools and -techniques are now available to the physicist and clinician to enable absorbed dose calculations to both target and critical organs-at-risk. The challenge now facing nuclear medicine is to enable this methodology to be routinely available to the clinic, to ensure common standard operating procedures between centres and in particular to correlate response criteria with absorbed dose estimates.

A toolkit for epithermal neutron beam characterisation in BNCT.

Methods for dosimetry of epithermal neutron beams used in boron neutron capture therapy (BNCT) have been developed and utilised within the Finnish BNCT project as well as within a European project for a code of practise for the dosimetry of BNCT. One outcome has been a travelling toolkit for BNCT dosimetry. It consists of activation detectors and ionisation chambers. The free-beam neutron spectrum is measured with a set of activation foils of different isotopes irradiated both in a Cd-capsule and without it. Neutron flux (thermal and epithermal) distribution in phantoms is measured using activation of Mn and Au foils, and Cu wire. Ionisation chamber (IC) measurements are performed both in-free-beam and in-phantom for determination of the neutron and gamma dose components. This toolkit has also been used at other BNCT facilities in Europe, the USA, Argentina and Japan.