Artificial intelligence of imaging and clinical neurological data for predictive, preventive and personalized (P3) medicine for Parkinson Disease: The NeuroArtP3 protocol for a multi-center research study.

BACKGROUND: The burden of Parkinson Disease (PD) represents a key public health issue and it is essential to develop innovative and cost-effective approaches to promote sustainable diagnostic and therapeutic interventions. In this perspective the adoption of a P3 (predictive, preventive and personalized) medicine approach seems to be pivotal. The NeuroArtP3 (NET-2018-12366666) is a four-year multi-site project co-funded by the Italian Ministry of Health, bringing together clinical and computational centers operating in the field of neurology, including PD. OBJECTIVE: The core objectives of the project are: i) to harmonize the collection of data across the participating centers, ii) to structure standardized disease-specific datasets and iii) to advance knowledge on disease's trajectories through machine learning analysis. METHODS: The 4-years study combines two consecutive research components: i) a multi-center retrospective observational phase; ii) a multi-center prospective observational phase. The retrospective phase aims at collecting data of the patients admitted at the participating clinical centers. Whereas the prospective phase aims at collecting the same variables of the retrospective study in newly diagnosed patients who will be enrolled at the same centers. RESULTS: The participating clinical centers are the Provincial Health Services (APSS) of Trento (Italy) as the center responsible for the PD study and the IRCCS San Martino Hospital of Genoa (Italy) as the promoter center of the NeuroartP3 project. The computational centers responsible for data analysis are the Bruno Kessler Foundation of Trento (Italy) with TrentinoSalute4.0 -Competence Center for Digital Health of the Province of Trento (Italy) and the LISCOMPlab University of Genoa (Italy). CONCLUSIONS: The work behind this observational study protocol shows how it is possible and viable to systematize data collection procedures in order to feed research and to advance the implementation of a P3 approach into the clinical practice through the use of AI models.

Chemical and Physical Characterisation of Macroaggregated Human Serum Albumin: Strength and Specificity of Bonds with 99mTc and 68Ga.

BACKGROUND: Macroaggregated human serum albumin (MAA) properties are widely used in nuclear medicine, labelled with 99mTc. The aim of this study is to improve the knowledge about the morphology, size, dimension and physical-chemical characteristics of MAA and their bond with 99mTc and 68Ga. METHODS: Commercial kits of MAA (Pulmocis®) were used. Characterisation through experiments based on SEM, DLS and Stokes' Law were carried out. In vitro experiments for Langmuir isotherms and pH studies on radiolabelling were performed and the stability of the radiometal complex was verified through competition reactions. RESULTS: The study settles the MAA dimension within the range 43-51 µm. The Langmuir isotherm reveals for [99mTc]MAA: Bmax (46.32), h (2.36); for [68Ga]MAA: Bmax (44.54), h (0.893). Dual labelling reveals that MAA does not discriminate different radioisotopes. Experiments on pH placed the optimal pH for labelling with 99mTc at 6. CONCLUSION: Radiolabelling of MAA is possible with high efficiency. The nondiscriminatory MAA bonds make this drug suitable for radiolabelling with different radioisotopes or for dual labelling. This finding illustrates the need to continue investigating MAA chemical and physical characteristics to allow for secure labelling with different isotopes.

Chemical and Physical Characterisation of Human Serum Albumin Nanocolloids: Kinetics, Strength and Specificity of Bonds with 99mTc and 68Ga.

Nanoparticles of Human Serum Albumin (NC) labelled with 99mTc are widely used in Nuclear Medicine and represent the gold-standard for the intraoperative detection of the sentinel lymph node in many kinds of cancer, mainly breast cancer and melanoma. A significant amount of radionuclides can be incorporated into the HSA particle, due to the multiple binding sites, and HSA-based nanocolloid catabolism is a fast and easy process that results in innocuous degradation products. NCs labelled with different isotopes represent an interesting radiopharmaceutical for extending diagnostic accuracy and surgical outcome, but the knowledge of the chemical bond between NCs and isotopes has not been fully elucidated, including information on its strength and specificity. The aim of this study is to investigate and compare the physicochemical characteristics of the bond between NCs and 99mTc and 68Ga isotopes. Commercial kits of HSA-based nanocolloid particles (NanoAlbumon®) were used. For this purpose, we have primarily studied the kinetic orders of NC radiolabelling. Langmuir isotherms and pH effect on radiolabelling were tested and the stability of the radiometal complex was verified through competition reactions carried out in presence of different ligands. The future goal of our research is the development of inexpensive and instant kits, easily labelled with a wide spectrum of diagnostic and therapeutic isotopes, thus facilitating the availability of versatile and multipurpose radiopharmaceuticals.

Effects of Photons Irradiation on 18F-FET and 18F-DOPA Uptake by T98G Glioblastoma Cells.

The differential diagnosis between brain tumors recurrence and early neuroinflammation or late radionecrosis is still an unsolved problem. The new emerging magnetic resonance imaging, computed tomography, and positron emission tomography diagnostic modalities still lack sufficient accuracy. In the last years, a great effort has been made to develop radiotracers able to detect specific altered metabolic pathways or tumor receptor markers. Our research project aims to evaluate irradiation effects on radiopharmaceutical uptake and compare the kinetic of the fluorinate tracers. T98G glioblastoma cells were irradiated at doses of 2, 10, and 20 Gy with photons, and 18F-DOPA and 18F-FET tracer uptake was evaluated. Activity and cell viability at different incubation times were measured. 18F-FET and 18F-DOPA are accumulated via the LAT-1 transporter, but 18F-DOPA is further incorporated, whereas 18F-FET is not metabolized. Therefore, time-activity curves (TACs) tend to plateau with 18F-DOPA and to a rapid washout with 18F-FET. After irradiation, 18F-DOPA TAC resembles the 18F-FET pattern. 18F-DOPA activity peak we observed at 20 min might be fictitious, because earlier time points have not been evaluated, and a higher activity peak before 20 min cannot be excluded. In addition, the activity retained in the irradiated cells remains higher in comparison to the sham ones at all time points investigated. This aspect is similar in the 18F-FET TAC but less evident. Therefore, we can hypothesize the presence of a second intracellular compartment in addition to the amino acidic pool one governed by LAT-1, which could explain the progressive accumulation of 18F-DOPA in unirradiated cells.

99mTc-68Ga-ICG-Labelled Macroaggregates and Nanocolloids of Human Serum Albumin: Synthesis Procedures of a Trimodal Imaging Agent Using Commercial Kits.

Recent developments in sentinel lymph node (SLN) and radio occult lesion localization (ROLL) highlight the need for a multimodal contrast agent, providing better presurgical PET imaging and improved intraoperative mapping thanks to fluorescence detection. For this reason, we have studied a trimodal SLN/ROLL targeting agent (99mTc-68Ga-ICG) with commercially available kits of macroaggregated or nanocolloidal albumin (MA/NC-HSA). 68Ga PET imaging does provide better spatial resolution and makes it possible to predict signal intensity during surgery. The presence of 99mTc assesses the efficacy of these compounds in vitro and also during the surgery procedure. The aim of this study was to optimise the labelling and tagging of these two radiopharmaceuticals and assess their yields and stability. Kits of MA/NC-HSA particles (Pulmocis® and NanoAlbumon®) were used for sequential radiolabelling with 99mTc and 68Ga. Fluorescent tagging was performed using indocyanine green, a tricarbocyanine dye. The ITLC radiochemical purity of the trilabelled MA/NC-HSA was >95%. Fluorescent purity was measured by scanning the strips with a PhotoDynamicEye probe. Finally, in vitro stability tests, performed with DTPA and human serum solutions, assessed the efficacy of fluorescent tagging and radiolabelling.

Effects of L-DOPA Pretreatment on the Kinetics, Migration and Carbon Ion Radiation Response of T98G Cells.

BACKGROUND/AIM: Glioblastoma is the most malignant and widespread brain tumor in adults, with a rapid clinical course. Recently, it has been hypothesized that L-DOPA plays a role in the diagnosis and treatment of glioblastoma. The aim of this study was to assess the effects of pretreatment with L-DOPA on the biological behavior of human T98G cells in vitro. MATERIALS AND METHODS: T98G cells were treated with 50 µg/ml or 100 µg/ml of L-DOPA for 4 h and their morphology, growth rate, clonogenic survival and migratory capacity in basal conditions and after carbon ion irradiation were evaluated using standard methods. RESULTS: Treated cells showed a lower growth rate and an increased migratory capacity that correlated with the dose of tested L-DOPA. Treatment with L-DOPA increased the growth rate of carbon ion irradiated T98G cells compared to control non-treated cells exposed to the same radiation dose. CONCLUSION: Our results open further questions about the overall advantage of L-DOPA treatment of glioblastoma.

Evidence of 68Ga-DOTA-NT-20.3 Uptake in Pancreatic Adenocarcinoma AsPC-1 Cell Line - in vitro Study.

BACKGROUND: Neurotensin receptors are overexpressed in several cancer types including pancreatic ductal adenocarcinoma. Three NTR subtypes have been cloned: NTR-1, NTR-2 and NTR-3. The most expressed NTR-1 is not present in normal pancreatic tissue and has a low expression in chronic pancreatitis. OBJECTIVE: Objective of this study was to test in vitro affinity of the new 68Ga labelled neurotensin analogue DOTA-NT-20.3 (fragment 6-13, Ac-Lys(DOTA)-Pro-Arg(N-CH3)-Arg-Pro-Tyr-Tle-Leu) on the human pancreatic ductal adenocarcinoma cell line AsPC-1. METHOD: For the preparation of 68Ga-DOTA-NT-20.3, 68GaCl3 solution (eluted from 68Ge/68Ga generator) and 50 µg of precursor (Iason, Graz, Austria) water dissolved were used in an automatic synthesis module. The labeled compound was added to cell culture flask and incubated at 37°C. At various time points after tracer addition up to 80min, cells were recovered, rinsed and counted for radioactivity. Results were expressed as percent binding normalized to 200000 cells and affinity parameters were calculated. RESULTS: Labeling yield was ≥98 %. The molar ratio between labelled and total peptide was about 1/400. AsPC-1 cell line showed rapid uptake of the tracer including surface and internalized binding, tending to a plateau phase 80 min after tracer addition (11%/200.000 cells). The Kd (7.335 pmol) and Bmax (90.52 kBq) value indicated high tracer affinity for AsPC-1cell line especially if compared with the literature data regarding other malignancies (e.g. colonic cancer cell line). Binding sites were 1.09x106 sites per cell. CONCLUSION: New tracer 68Ga-DOTA-NT-20.3 can be a suitable candidate for the clinical use in patients with pancreatic ductal adenocarcinoma.