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Thyroid cancer (TC) is the most common endocrine malignancy. Recent progress in thyroid cancer biology revealed a certain degree of intratumoral heterogeneity, highlighting the coexistence of cellular subpopulations with distinct proliferative capacities and differentiation abilities. Among those subpopulations, cancer stem-like cells (CSCs) are hypothesized to drive TC heterogeneity, contributing to its metastatic potential and therapy resistance. CSCs principally exist in tumor areas with specific microenvironmental conditions, the so-called stem cell niches. In particular, in thyroid cancer, CSCs’ survival is enhanced in the hypoxic niche, the immune niche, and some areas with specific extracellular matrix composition. In this review, we summarize the current knowledge about thyroid CSCs, the tumoral niches that allow their survival, and the implications for TC therapy.

Introduction: Neuroendocrine transdifferentiation (NED) of prostate cancer (PC) cells is associated with the development of resistance to antiandrogen therapy and poor prognosis in patients with castration-resistant PC (CRPC). Many of the molecular events, involved in NED, appear to be mediated by epigenetic mechanisms. In this study, we evaluated the antitumor activity and epigenetic modulation of 2 epigenetic drugs, such as the demethylating agent 5-aza-2′-deoxycytidine (AZA) and the methyl donor S-adenosylmethionine (SAM), in 2 human CRPC cell lines with NED (DU-145 and PC-3). Methods: The effects of AZA and SAM on cell viability, cell cycle, apoptosis, migration, and genome-wide DNA methylation profiling have been evaluated. Results: Both drugs showed a prominent antitumor activity in DU-145 and PC-3 cells, through perturbation of cell cycle progression, induction of apoptosis, and inhibition of cell migration. AZA and SAM reversed NED in DU-145 and PC-3, respectively. Moreover, AZA treatment modified DNA methylation pattern in DU-145 cells, sustaining a pervasive hypomethylation of the genome, with a relevant effect on several pathways involved in the regulation of cell proliferation, apoptosis, and cell migration, in particular Wnt/β-catenin. Conclusions: A relevant antitumor activity of these epigenetic drugs on CRPC cell lines with NED opens a new scenario in the therapy of this lethal variant of PC.

In spite of the hazard due to the radiation exposure, preparation of 90Y- and 177Lu-labelled radiopharmaceuticals is still mainly performed using manual procedures. In the present study the performance of a commercial automatic synthesizer based on disposable cassettes for the labelling of 177Lu- and 90Y-DOTA-conjugated biomolecules (namely, DOTATOC and PSMA-617) was evaluated and compared to a manual and a semiautomated approach. The dose exposure of the operators was evaluated as well. More than 300 clinical preparations of both 90Y- and 177Lu-labelled radiopharmaceuticals have been performed using the three different methods. The mean radiochemical yields for 90Y-DOTATOC were 96.2±4.9%, 90.3±5.6%, and 82.0±8.4%, while for 177Lu-DOTATOC they were 98.3% ± 0.6, 90.8% ± 8.3, and 83.1±5.7% when manual, semiautomated, and automated approaches were used, respectively. The mean doses on the whole hands for yttrium-90 preparations were 0.15±0.4 mSv/GBq, 0.04±0.1 mSv/GBq, and 0.11±0.3 mSv/GBq for manual, semiautomated, and automated synthesis, respectively, and for lutetium-177 preparations, they were 0.02±0.008 mSv/GBq, 0.01±0.03 mSv/GBq, and 0.01±0.02 mSv/GBq, respectively. In conclusion, the automated approach guaranteed reliable and reproducible preparations of pharmaceutical grade therapeutic radiopharmaceuticals in a decent RCY. The radiation exposure of the operators remained comparable to the manual approach mainly due to the fact that a dedicated shielding was still not available for the system.

Multicellular three-dimensional (3D) spheroids represent an experimental model that is intermediate in its complexity between monolayer cultures and patients’ tumor. In the present study, we characterize multicellular spheroids from papillary (PTC) and follicular (FTC) thyroid cancers and from the corresponding normal tissues. We show that these 3D structures well recapitulate the features of the original tissues, in either the differentiated and “stem-like” components. As a second step, we were aimed to test the effects of a small multikinase inhibitor, SP600125 (SP), previously shown to efficiently induce cell death in undifferentiated thyroid cancer monolayer cultures. We demonstrate the potent effect of SP on cell growth and survival in our 3D multicellular cultures. SP exerts its main effects through direct and highly significant inhibition of the ROCK pathway, known to be involved in the regulation of cell migration and β-catenin turnover. Consistently, SP treatment resulted in a significant decrease in β-catenin levels with respect to basal conditions in tumor but not in normal spheroids, indicating that the effect is promisingly selective on tumor cells. In conclusion, we provide the morphological and molecular characterization of thyroid normal and tumor spheroids. In this 3D model we tested in vitro the effects of the multikinase inhibitor SP and further characterized its mechanism of action in both normal and tumor spheroids, thus making it an ideal candidate for developing new drugs against thyroid cancer.

Background Multicellular spheroids represent an interesting experimental model with promising applications in the pre-clinical studies on anticancer drugs. We recently demonstrated that thyroid spheroids recapitulate the features of the original tissues, in either the differentiated and "stem-like" components. Here we were aimed to characterize thyroid spheroids and to investigate in vivo the proangiogenic potential of patient-derived xenografts (PDX) of spheroids obtained from papillary thyroid cancer (PTC) and the matched normal tissues. Methods Spheroids cultures were obtained from 11 PTCs and matched normal tissues and characterized by immunohistochemistry. The expression of p53, involved in the regulation of stem cell homeostasis, was evaluated. The proangiogenic effect of thyroid spheroids was assessed by the injection in zebrafish embryos. Results Thyroid spheroids are enriched in stem-like cells, as shown by the positivity for the stem cell marker OCT4, and by the low level of p53 expression. Interestingly, PTCs and normal thyroid tissues have a detectable p53 expression, whereas the derived spheroids are mainly constituted by cells that express p53 at a lower level. Finally, we show that PDXs derived from PTC or normal spheroids stimulate the migration and the growth of sprouting vessels toward the implant into the zebrafish embryos. Conclusions We report the characterization of multicellular spheroids obtained from PTCs and normal thyroid tissues, showing that they are enriched in stem-like cells. Moreover, we established xenografts of spheroids in zebrafish, demonstrating that they stimulate neoangiogenesis. This in vivo model could be considered as a valuable platform to test the effects of anticancer drugs.

Purpose Peptide Receptor Radionuclide Therapy (PPRT) is nowadays the most effective therapy for treating patients who suffered of neuroendocrine tumours [1] . In clinical practice, absorbed dose calculations are computed based on the Medical Internal Radiation Dose (MIRD) scheme (not planned or optimised for patients’ specific characteristics). To increase the quality and the efficacy of the clinical trials, a more accurate 3D dosimetry is required. Monte Carlo (MC) techniques showed to provide the most accurate approach for the MRT dose calculations. In this work, Raydose MC based pipeline [2] was used to compute the patient specific dose calculation for a clinical trial in PRRT. The dose obtained for lesions and organs at risk (OARs) were then compared to the ones based on the MIRD scheme. Methods One hundred patients were enrolled as part of a clinical trial in PRRT with a prescribed activity between 3.7 and 5.5 GBq of 177Lu-DOTA-Tyr3-octreotide. Patients were scanned five times with a SPECT/CT scanner and the sequential scans were co-registered using a non-rigid registration algorithm [3] . Volumes of interest (VOIs) including lesions and OARs were manually outlined on fused images. 3D dose maps calculated with Raydose were compared to the corresponding ones obtained with OLINDA/EXM software. Differences between calculations were evaluated using a paired samples Wilcoxon-Mann-Witney test (two-sided, p Results Mean doses calculated with OLINDA/EXM were higher than the corresponding mean doses calculated with Raydose. In particular, for the structures studied, the following median percentage differences were obtained: kidneys, −11.70% (Internal Quartile Range (IQR) = 7.14); liver, −14.06% (IQR = 5.62); spleen, −12.50% (IQR = 6.38); lesions, −21.01% (IQR = 18.52). Furthermore, significant differences were observed for kidneys and lesions. Conclusions Dose values obtained with the MC method suggested an underestimation of the dose calculated for OARs and lesions in PRRT treatment. A more accurate patient specific 3D dose calculation could be used to increase the injected activity and to rise the efficiency of the treatment but maintaining a safe level of toxicity.

Accurate delineation of the gross tumour volume (GTV) in radiation therapy process is mandatory, especially when sophisticated techniques for radiotherapy delivery are used. Functional imaging, particularly PET and its fusion with anatomical modalities, such as PET/CT, promises to improve detection and characterization of disease. Quantitative accuracy in PET imaging is affected by several factors, which influence PET data so as to accurately delineate volume for radiotherapy planning purpose. Several strategies in using PET for target volume definition in radiotherapy treatment planning are being explored in the last years. In the present paper we give an overview of different techniques proposed to generate PET-based tumour volumes and the potential of their use in the radiation oncology process.

Thyroid cancer is the most common endocrine malignancy with increasing incidence worldwide. The majority of thyroid cancer cases are well differentiated with favorable outcome. However, undifferentiated thyroid cancers are one of the most lethal human malignancies because of their invasiveness, metastatization and refractoriness even to the most recently developed therapies. In this study we show for the first time a significant hyperactivation of ROCK/HDAC6 pathway in thyroid cancer tissues, and its negative correlation with p53 DNA binding ability. We demonstrate that a small compound, SP600125 (SP), is able to induce cell death selectively in undifferentiated thyroid cancer cell lines by specifically acting on the pathogenic pathways of cancer development. In detail, SP acts on the ROCK/HDAC6 pathway involved in dedifferentiation and invasiveness of undifferentiated human cancers, by restoring its physiological activity level. As main consequence, cancer cell migration is inhibited and, at the same time, cell death is induced through the mitotic catastrophe. Moreover, SP exerts a preferential action on the mutant p53 by increasing its DNA binding ability. In TP53-mutant cells that survive mitotic catastrophe this process results in p21 induction and eventually lead to premature senescence. In conclusion, SP has been proved to be able to simultaneously block cell replication and migration, the two main processes involved in cancer development and dissemination, making it an ideal candidate for developing new drugs against anaplastic thyroid cancer., SCOPUS: ar.j, info:eu-repo/semantics/published

Molecular radiotherapy (MRT) is an effective technique for the treatment of several oncological diseases, based on the delivery of radiation to malignant tissues via the interaction of an agent with molecular sites and receptors [1] . In the clinical practice, the calculation of the absorbed dose is performed using the Medical Internal Radiation Dose (MIRD) scheme, based on pre-calculated transfer functions determined on standard anthropomorphic phantoms. The MIRD scheme assumes that radiopharmaceutical activity is distributed uniformly inside the organs [2] . For this reason, dose calculation is not planned or optimised for patients’ specific characteristics. To increase the efficacy and quality of clinical trials, a detailed and reliable 3D patient specific dosimetry is required. Monte Carlo techniques are shown to provide the most accurate approach for the radiotherapy dose calculation. In this work, we used Raydose Monte Carlo code [3] to provide a patient specific 3D dose distribution in MRT at the voxel level for patients enrolled as part of a clinical trial on Peptide Receptor Radiotherapy (PRRT) with 177Lu and we compared doses to lesions and OARs with standard dose calculations obtained using the MIRD method. One hundred patients were enrolled in a clinical trial in PRRT with an activity prescription of 177Lu-DOTA-Tyr3-octreotide. All patients were scanned five times with a SPECT/CT scanner and both lesions and OARs were drawn manually on fused images. Sequential scans were co-registered using a non-rigid registration algorithm. Raydose 3D dose maps were compared with dose calculations obtained using the OLINDA/EXM software. Preliminary results for the first 20 patients show that mean doses calculated with OLINDA/EXM are significantly higher than mean doses calculated with Raydose. Initial data based on Monte Carlo dose calculations suggest a possible underestimation of the dose to OARs in PRRT treatments. More accurate dose calculation could be used on a patient by patient basis to increase the prescribed activity, maintaining a safe level of toxicity. A complete analysis that includes doses to lesions is in progress.