Impact on the Transcriptome of Proton Beam Irradiation Targeted at Healthy Cardiac Tissue of Mice.

Proton beam therapy is considered a step forward with respect to electromagnetic radiation, thanks to the reduction in the dose delivered. Among unwanted effects to healthy tissue, cardiovascular complications are a known long-term radiotherapy complication. The transcriptional response of cardiac tissue from xenografted BALB/c nude mice obtained at 3 and 10 days after proton irradiation covering both the tumor region and the underlying healthy tissue was analyzed as a function of dose and time. Three doses were used: 2 Gy, 6 Gy, and 9 Gy. The intermediate dose had caused the greatest impact at 3 days after irradiation: at 2 Gy, 219 genes were differently expressed, many of them represented by zinc finger proteins; at 6 Gy, there were 1109, with a predominance of genes involved in energy metabolism and responses to stimuli; and at 9 Gy, there were 105, mainly represented by zinc finger proteins and molecules involved in the regulation of cardiac function. After 10 days, no significant effects were detected, suggesting that cellular repair mechanisms had defused the potential alterations in gene expression. The nonlinear dose-response curve indicates a need to update the models built on photons to improve accuracy in health risk prediction. Our data also suggest a possible role for zinc finger protein genes as markers of proton therapy efficacy.

Biodistribution Assessment of a Novel 68Ga-Labeled Radiopharmaceutical in a Cancer Overexpressing CCK2R Mouse Model: Conventional and Radiomics Methods for Analysis.

The aim of the present study consists of the evaluation of the biodistribution of a novel 68Ga-labeled radiopharmaceutical, [68Ga]Ga-NODAGA-Z360, injected into Balb/c nude mice through histopathological analysis on bioptic samples and radiomics analysis of positron emission tomography/computed tomography (PET/CT) images. The 68Ga-labeled radiopharmaceutical was designed to specifically bind to the cholecystokinin receptor (CCK2R). This receptor, naturally present in healthy tissues such as the stomach, is a biomarker for numerous tumors when overexpressed. In this experiment, Balb/c nude mice were xenografted with a human epidermoid carcinoma A431 cell line (A431 WT) and overexpressing CCK2R (A431 CCK2R+), while controls received a wild-type cell line. PET images were processed, segmented after atlas-based co-registration and, consequently, 112 radiomics features were extracted for each investigated organ / tissue. To confirm the histopathology at the tissue level and correlate it with the degree of PET uptake, the studies were supported by digital pathology. As a result of the analyses, the differences in radiomics features in different body districts confirmed the correct targeting of the radiopharmaceutical. In preclinical imaging, the methodology confirms the importance of a decision-support system based on artificial intelligence algorithms for the assessment of radiopharmaceutical biodistribution.

An Overview of In Vitro Assays of 64Cu-, 68Ga-, 125I-, and 99mTc-Labelled Radiopharmaceuticals Using Radiometric Counters in the Era of Radiotheranostics.

Radionuclides are unstable isotopes that mainly emit alpha (α), beta (ß) or gamma (γ) radiation through radiation decay. Therefore, they are used in the biomedical field to label biomolecules or drugs for diagnostic imaging applications, such as positron emission tomography (PET) and/or single-photon emission computed tomography (SPECT). A growing field of research is the development of new radiopharmaceuticals for use in cancer treatments. Preclinical studies are the gold standard for translational research. Specifically, in vitro radiopharmaceutical studies are based on the use of radiopharmaceuticals directly on cells. To date, radiometric ß- and γ-counters are the only tools able to assess a preclinical in vitro assay with the aim of estimating uptake, retention, and release parameters, including time- and dose-dependent cytotoxicity and kinetic parameters. This review has been designed for researchers, such as biologists and biotechnologists, who would like to approach the radiobiology field and conduct in vitro assays for cellular radioactivity evaluations using radiometric counters. To demonstrate the importance of in vitro radiopharmaceutical assays using radiometric counters with a view to radiogenomics, many studies based on 64Cu-, 68Ga-, 125I-, and 99mTc-labeled radiopharmaceuticals have been revised and summarized in this manuscript.

Proton boron capture therapy (PBCT) induces cell death and mitophagy in a heterotopic glioblastoma model.

Despite aggressive therapeutic regimens, glioblastoma (GBM) represents a deadly brain tumor with significant aggressiveness, radioresistance and chemoresistance, leading to dismal prognosis. Hypoxic microenvironment, which characterizes GBM, is associated with reduced therapeutic effectiveness. Moreover, current irradiation approaches are limited by uncertain tumor delineation and severe side effects that comprehensively lead to unsuccessful treatment and to a worsening of the quality of life of GBM patients. Proton beam offers the opportunity of reduced side effects and a depth-dose profile, which, unfortunately, are coupled with low relative biological effectiveness (RBE). The use of radiosensitizing agents, such as boron-containing molecules, enhances proton RBE and increases the effectiveness on proton beam-hit targets. We report a first preclinical evaluation of proton boron capture therapy (PBCT) in a preclinical model of GBM analyzed via µ-positron emission tomography/computed tomography (µPET-CT) assisted live imaging, finding a significant increased therapeutic effectiveness of PBCT versus proton coupled with an increased cell death and mitophagy. Our work supports PBCT and radiosensitizing agents as a scalable strategy to treat GBM exploiting ballistic advances of proton beam and increasing therapeutic effectiveness and quality of life in GBM patients.

Chelation of Theranostic Copper Radioisotopes with S-Rich Macrocycles: From Radiolabelling of Copper-64 to In Vivo Investigation.

Copper radioisotopes are generally employed for cancer imaging and therapy when firmly coordinated via a chelating agent coupled to a tumor-seeking vector. However, the biologically triggered Cu2+-Cu+ redox switching may constrain the in vivo integrity of the resulting complex, leading to demetallation processes. This unsought pathway is expected to be hindered by chelators bearing N, O, and S donors which appropriately complements the borderline-hard and soft nature of Cu2+ and Cu+. In this work, the labelling performances of a series of S-rich polyazamacrocyclic chelators with [64Cu]Cu2+ and the stability of the [64Cu]Cu-complexes thereof were evaluated. Among the chelators considered, the best results were obtained with 1,7-bis [2-(methylsulfanyl)ethyl]-4,10,diacetic acid-1,4,7,10-tetraazacyclododecane (DO2A2S). DO2A2S was labelled at high molar activities in mild reaction conditions, and its [64Cu]Cu2+ complex showed excellent integrity in human serum over 24 h. Biodistribution studies in BALB/c nude mice performed with [64Cu][Cu(DO2A2S)] revealed a behavior similar to other [64Cu]Cu-labelled cyclen derivatives characterized by high liver and kidney uptake, which could either be ascribed to transchelation phenomena or metabolic processing of the intact complex.

A New Preclinical Decision Support System Based on PET Radiomics: A Preliminary Study on the Evaluation of an Innovative 64Cu-Labeled Chelator in Mouse Models.

The 64Cu-labeled chelator was analyzed in vivo by positron emission tomography (PET) imaging to evaluate its biodistribution in a murine model at different acquisition times. For this purpose, nine 6-week-old female Balb/C nude strain mice underwent micro-PET imaging at three different time points after 64Cu-labeled chelator injection. Specifically, the mice were divided into group 1 (acquisition 1 h after [64Cu] chelator administration, n = 3 mice), group 2 (acquisition 4 h after [64Cu]chelator administration, n = 3 mice), and group 3 (acquisition 24 h after [64Cu] chelator administration, n = 3 mice). Successively, all PET studies were segmented by means of registration with a standard template space (3D whole-body Digimouse atlas), and 108 radiomics features were extracted from seven organs (namely, heart, bladder, stomach, liver, spleen, kidney, and lung) to investigate possible changes over time in [64Cu]chelator biodistribution. The one-way analysis of variance and post hoc Tukey Honestly Significant Difference test revealed that, while heart, stomach, spleen, kidney, and lung districts showed a very low percentage of radiomics features with significant variations (p-value < 0.05) among the three groups of mice, a large number of features (greater than 60% and 50%, respectively) that varied significantly between groups were observed in bladder and liver, indicating a different in vivo uptake of the 64Cu-labeled chelator over time. The proposed methodology may improve the method of calculating the [64Cu]chelator biodistribution and open the way towards a decision support system in the field of new radiopharmaceuticals used in preclinical imaging trials.

The Proton-Boron Reaction Increases the Radiobiological Effectiveness of Clinical Low- and High-Energy Proton Beams: Novel Experimental Evidence and Perspectives.

Protontherapy is a rapidly expanding radiotherapy modality where accelerated proton beams are used to precisely deliver the dose to the tumor target but is generally considered ineffective against radioresistant tumors. Proton-Boron Capture Therapy (PBCT) is a novel approach aimed at enhancing proton biological effectiveness. PBCT exploits a nuclear fusion reaction between low-energy protons and 11B atoms, i.e. p+11B→ 3α (p-B), which is supposed to produce highly-DNA damaging α-particles exclusively across the tumor-conformed Spread-Out Bragg Peak (SOBP), without harming healthy tissues in the beam entrance channel. To confirm previous work on PBCT, here we report new in-vitro data obtained at the 62-MeV ocular melanoma-dedicated proton beamline of the INFN-Laboratori Nazionali del Sud (LNS), Catania, Italy. For the first time, we also tested PBCT at the 250-MeV proton beamline used for deep-seated cancers at the Centro Nazionale di Adroterapia Oncologica (CNAO), Pavia, Italy. We used Sodium Mercaptododecaborate (BSH) as 11B carrier, DU145 prostate cancer cells to assess cell killing and non-cancer epithelial breast MCF-10A cells for quantifying chromosome aberrations (CAs) by FISH painting and DNA repair pathway protein expression by western blotting. Cells were exposed at various depths along the two clinical SOBPs. Compared to exposure in the absence of boron, proton irradiation in the presence of BSH significantly reduced DU145 clonogenic survival and increased both frequency and complexity of CAs in MCF-10A cells at the mid- and distal SOBP positions, but not at the beam entrance. BSH-mediated enhancement of DNA damage response was also found at mid-SOBP. These results corroborate PBCT as a strategy to render protontherapy amenable towards radiotherapy-resilient tumor. If coupled with emerging proton FLASH radiotherapy modalities, PBCT could thus widen the protontherapy therapeutic index.

Local Disease-Free Survival Rate (LSR) Application to Personalize Radiation Therapy Treatments in Breast Cancer Models.

Cancer heterogeneity represents the main issue for defining an effective treatment in clinical practice, and the scientific community is progressively moving towards the development of more personalized therapeutic regimens. Radiotherapy (RT) remains a fundamental therapeutic treatment used for many neoplastic diseases, including breast cancer (BC), where high variability at the clinical and molecular level is known. The aim of this work is to apply the generalized linear quadratic (LQ) model to customize the radiant treatment plan for BC, by extracting some characteristic parameters of intrinsic radiosensitivity that are not generic, but may be exclusive for each cell type. We tested the validity of the generalized LQ model and analyzed the local disease-free survival rate (LSR) for breast RT treatment by using four BC cell cultures (both primary and immortalized), irradiated with clinical X-ray beams. BC cells were chosen on the basis of their receptor profiles, in order to simulate a differential response to RT between triple negative breast and luminal adenocarcinomas. The MCF10A breast epithelial cell line was utilized as a healthy control. We show that an RT plan setup based only on α and ß values could be limiting and misleading. Indeed, two other parameters, the doubling time and the clonogens number, are important to finely predict the tumor response to treatment. Our findings could be tested at a preclinical level to confirm their application as a variant of the classical LQ model, to create a more personalized approach for RT planning.

Evaluation of proton beam radiation-induced skin injury in a murine model using a clinical SOBP.

The purpose of this paper is to characterize the skin deterministic damage due to the effect of proton beam irradiation in mice occurred during a long-term observational experiment. This study was initially defined to evaluate the insurgence of myelopathy irradiating spinal cords with the distal part of a Spread-out Bragg peak (SOBP). To the best of our knowledge, no study has been conducted highlighting high grades of skin injury at the dose used in this paper. Nevertheless these effects occurred. In this regard, the experimental evidence of significant insurgence of skin injury induced by protons using a SOBP configuration will be shown. Skin damages were classified into six scores (from 0 to 5) according to the severity of the injuries and correlated to ED50 (i.e. the radiation dose at which 50% of animals show a specific score) at 40 days post-irradiation (d.p.i.). The effects of radiation on the overall animal wellbeing have been also monitored and the severity of radiation-induced skin injuries was observed and quantified up to 40 d.p.i.

Radiosensitizing effect of curcumin-loaded lipid nanoparticles in breast cancer cells.

In breast cancer (BC) care, radiotherapy is considered an efficient treatment, prescribed both for controlling localized tumors or as a therapeutic option in case of inoperable, incompletely resected or recurrent tumors. However, approximately 90% of BC-related deaths are due to the metastatic tumor progression. Then, it is strongly desirable to improve tumor radiosensitivity using molecules with synergistic action. The main aim of this study is to develop curcumin-loaded solid nanoparticles (Cur-SLN) in order to increase curcumin bioavailability and to evaluate their radiosensitizing ability in comparison to free curcumin (free-Cur), by using an in vitro approach on BC cell lines. In addition, transcriptomic and metabolomic profiles, induced by Cur-SLN treatments, highlighted networks involved in this radiosensitization ability. The non tumorigenic MCF10A and the tumorigenic MCF7 and MDA-MB-231 BC cell lines were used. Curcumin-loaded solid nanoparticles were prepared using ethanolic precipitation and the loading capacity was evaluated by UV spectrophotometer analysis. Cell survival after treatments was evaluated by clonogenic assay. Dose-response curves were generated testing three concentrations of free-Cur and Cur-SLN in combination with increasing doses of IR (2-9 Gy). IC50 value and Dose Modifying Factor (DMF) was measured to quantify the sensitivity to curcumin and to combined treatments. A multi-"omic" approach was used to explain the Cur-SLN radiosensitizer effect by microarray and metobolomic analysis. We have shown the efficacy of the Cur-SLN formulation as radiosensitizer on three BC cell lines. The DMFs values, calculated at the isoeffect of SF = 50%, showed that the Luminal A MCF7 resulted sensitive to the combined treatments using increasing concentration of vehicled curcumin Cur-SLN (DMF: 1,78 with 10 µM Cur-SLN.) Instead, triple negative MDA-MB-231 cells were more sensitive to free-Cur, although these cells also receive a radiosensitization effect by combination with Cur-SLN (DMF: 1.38 with 10 µM Cur-SLN). The Cur-SLN radiosensitizing function, evaluated by transcriptomic and metabolomic approach, revealed anti-oxidant and anti-tumor effects. Curcumin loaded- SLN can be suggested in future preclinical and clinical studies to test its concomitant use during radiotherapy treatments with the double implications of being a radiosensitizing molecule against cancer cells, with a protective role against IR side effects.

Sensitization of chondrosarcoma cells with PARP inhibitor and high-LET radiation.

Chondrosarcoma is a malignant tumor that arises from cartilaginous tissue and is radioresistant and chemoresistant to conventional treatments. The preferred treatment consists of surgical resection, which might cause severe disabilities for the patient; in addition, this procedure might be impossible for inoperable locations, such as the skull base. Carbon ion irradiation (hadron therapy) has been proposed as an alternative treatment, primarily due to its greater biological effectiveness and improved ballistic properties compared with conventional radiotherapy with X-rays. The goal of this study was to characterize the genetic mutations of a grade III chondrosarcoma cell line (CH2879) and examine the cellular responses to conventional radiotherapy (X-rays) and hadron therapy (proton and carbon ions) in the presence of the PARP inhibitor Olaparib. To better understand PARP inhibition, we first analyzed the formation of poly-ADP ribose chains by western blot; we observed an increase in its signal after irradiation, which disappeared on addition of the PARP inhibitor. PARPi enhanced ratio of approximately 1.3, 1.8, and 1.5 following irradiation of cells with X-rays, protons, and C-ions, respectively, as detected by clonogenic assay. The decrease in cell survival was confirmed by proliferation assay. The radiosensitivity of CH2879 cells was associated with mutations in homologous recombination repair genes, such as RAD50, SMARCA2 and NBN. This study demonstrates the capacity of the PARP inhibitor Olaparib to radiosensitize mutated chondrosarcoma cells to conventional photon irradiation, proton and carbon ion irradiation.

Gene Expression Profiles Induced by High-dose Ionizing Radiation in MDA-MB-231 Triple-negative Breast Cancer Cell Line.

BACKGROUND/AIM: Radiation therapy (RT) represents a therapeutic option in breast cancer (BC). Even if a great number of BC patients receive RT, not all of them report benefits, due to radioresistance that gets activated through several factors, such as the hormone receptor status. Herein, we analyzed the gene expression profiles (GEP) induced by RT in triple-negative BC (TNBC) MDA-MB-231, to study signalling networks involved in radioresistance. MATERIALS AND METHODS: GEP of MDA-MB-231 BC cells treated with a high dose of radiation, went through cDNA microarray analysis. In addition, to examine the cellular effects induced by RT, analyses of morphology and clonogenic evaluation were also conducted. RESULTS: A descriptive report of GEP and pathways induced by IR is reported from our microarray data. Moreover, the MDA-MB-231 Radioresistent Cell Fraction (RCF) selected, included specific molecules able to drive radioresistance. CONCLUSION: In summary, our data highlight, the RT response of TNBC MDA-MB-231 cell line at a transcriptional level, in terms of activating radioresistance in these cells, as a model of late-stage BC.

Betula etnensis Raf. (Betulaceae) Extract Induced HO-1 Expression and Ferroptosis Cell Death in Human Colon Cancer Cells.

Betula etnensis Raf. (Birch Etna) belonging to the Betulaceae family grows on the eastern slope of Etna. Many bioactive compounds present in Betula species are considered promising anticancer agents. In this study, we evaluated the effects of B. etnensis Raf. bark methanolic extract on a human colon cancer cell line (CaCo2). In order to elucidate the mechanisms of action of the extract, cellular redox status, cell cycle, and heme oxygenase-1 (HO-1) expression in ferroptosis induction were evaluated. Cell viability and proliferation were tested by tetrazolium (MTT) assayand cell cycle analysis, while cell death was evaluated by annexin V test and lactic dehydrogenase (LDH) release. Cellular redox status was assessed by measuring thiol groups (RSH) content, reactive oxygen species (ROS) production, lipid hydroperoxide (LOOH) levels and (γ-glutamylcysteine synthetase) γ-GCS and HO-1 expressions. The extract significantly reduced cell viability of CaCo2, inducing necrotic cell death in a concentration-depending manner. In addition, an increase in ROS levels and a decrease of RSH content without modulation in γ-GCS expression were detected, with an augmentation in LOOH levels and drastic increase in HO-1 expression. These results suggest that the B. etnensis Raf. extract promotes an oxidative cellular microenvironment resulting in CaCo2 cell death by ferroptosis mediated by HO-1 hyper-expression.

Gene expression profiling of breast cancer cell lines treated with proton and electron radiations.

OBJECTIVE: Technological advances in radiation therapy are evolving with the use of hadrons, such as protons, indicated for tumors where conventional radiotherapy does not give significant advantages or for tumors located in sensitive regions, which need the maximum of dose-saving of the surrounding healthy tissues. The genomic response to conventional and non-conventional linear energy transfer exposure is a poor investigated topic and became an issue of radiobiological interest. The aim of this work was to analyze and compare molecular responses in term of gene expression profiles, induced by electron and proton irradiation in breast cancer cell lines. METHODS: We studied the gene expression profiling differences by cDNA microarray activated in response to electron and proton irradiation with different linear energy transfer values, among three breast cell lines (the tumorigenic MCF7 and MDA-MB-231 and the non-tumorigenic MCF10A), exposed to the same sublethal dose of 9 Gy. RESULTS: Gene expression profiling pathway analyses showed the activation of different signaling and molecular networks in a cell line and radiation type-dependent manner. MCF10A and MDA-MB-231 cell lines were found to induce factors and pathways involved in the immunological process control. CONCLUSION: Here, we describe in a detailed way the gene expression profiling and pathways activated after electron and proton irradiation in breast cancer cells. Summarizing, although specific pathways are activated in a radiation type-dependent manner, each cell line activates overall similar molecular networks in response to both these two types of ionizing radiation. Advances in knowledge: In the era of personalized medicine and breast cancer target-directed intervention, we trust that this study could drive radiation therapy towards personalized treatments, evaluating possible combined treatments, based on the molecular characterization.

Radiation-Induced Gene Expression Changes in High and Low Grade Breast Cancer Cell Types.

BACKGROUND: There is extensive scientific evidence that radiation therapy (RT) is a crucial treatment, either alone or in combination with other treatment modalities, for many types of cancer, including breast cancer (BC). BC is a heterogeneous disease at both clinical and molecular levels, presenting distinct subtypes linked to the hormone receptor (HR) status and associated with different clinical outcomes. The aim of this study was to assess the molecular changes induced by high doses of ionizing radiation (IR) on immortalized and primary BC cell lines grouped according to Human epidermal growth factor receptor (HER2), estrogen, and progesterone receptors, to study how HR status influences the radiation response. Our genomic approach using in vitro and ex-vivo models (e.g., primary cells) is a necessary first step for a translational study to describe the common driven radio-resistance features associated with HR status. This information will eventually allow clinicians to prescribe more personalized total doses or associated targeted therapies for specific tumor subtypes, thus enhancing cancer radio-sensitivity. METHODS: Nontumorigenic (MCF10A) and BC (MCF7 and MDA-MB-231) immortalized cell lines, as well as healthy (HMEC) and BC (BCpc7 and BCpcEMT) primary cultures, were divided into low grade, high grade, and healthy groups according to their HR status. At 24 h post-treatment, the gene expression profiles induced by two doses of IR treatment with 9 and 23 Gy were analyzed by cDNA microarray technology to select and compare the differential gene and pathway expressions among the experimental groups. RESULTS: We present a descriptive report of the substantial alterations in gene expression levels and pathways after IR treatment in both immortalized and primary cell cultures. Overall, the IR-induced gene expression profiles and pathways appear to be cell-line dependent. The data suggest that some specific gene and pathway signatures seem to be linked to HR status. CONCLUSIONS: Genomic biomarkers and gene-signatures of specific tumor subtypes, selected according to their HR status and molecular features, could facilitate personalized biological-driven RT treatment planning alone and in combination with targeted therapies.

Area-based cell colony surviving fraction evaluation: A novel fully automatic approach using general-purpose acquisition hardware.

BACKGROUND: The current methodology for the Surviving Fraction (SF) measurement in clonogenic assay, which is a technique to study the anti-proliferative effect of treatments on cell cultures, involves manual counting of cell colony forming units. This procedure is operator-dependent and error-prone. Moreover, the identification of the exact colony number is often not feasible due to the high growth rate leading to the adjacent colony merging. As a matter of fact, conventional assessment does not deal with the colony size, which is generally correlated with the delivered radiation dose or the administered cytotoxic agent. METHOD: Considering that the Area Covered by Colony (ACC) is proportional to the colony number and size as well as to the growth rate, we propose a novel fully automatic approach exploiting Circle Hough Transform, to automatically detect the wells in the plate, and local adaptive thresholding, which calculates the percentage of ACC for the SF quantification. This measurement relies just on this covering percentage and does not consider the colony number, preventing inconsistencies due to intra- and inter-operator variability. RESULTS: To evaluate the accuracy of the proposed approach, we compared the SFs obtained by our automatic ACC-based method against the conventional counting procedure. The achieved results (r = 0.9791 and r = 0.9682 on MCF7 and MCF10A cells, respectively) showed values highly correlated with the measurements using the traditional approach based on colony number alone. CONCLUSIONS: The proposed computer-assisted methodology could be integrated in laboratory practice as an expert system for the SF evaluation in clonogenic assays.

Cytokine profile of breast cell lines after different radiation doses.

PURPOSE: Ionizing radiation (IR) treatment activates inflammatory processes causing the release of a great amount of molecules able to affect the cell survival. The aim of this study was to analyze the cytokine signature of conditioned medium produced by non-tumorigenic mammary epithelial cell line MCF10A, as well as MCF7 and MDA-MB-231 breast cancer cell lines, after single high doses of IR in order to understand their role in high radiation response. MATERIALS AND METHODS: We performed a cytokine profile of irradiated conditioned media of MCF10A, MCF7 and MDA-MB-231 cell lines treated with 9 or 23 Gy, by Luminex and ELISA analyses. RESULTS: Overall, our results show that both 9 Gy and 23 Gy of IR induce the release within the first 72 h of cytokines and growth factors potentially able to influence the tumor outcome, with a dose-independent and cell-line dependent signature. Moreover, our results show that the cell-senescence phenomenon does not correlate with the amount of 'senescence-associated secretory phenotype' (SASP) molecules released in media. Thus, additional mechanisms are probably involved in this process. CONCLUSIONS: These data open the possibility to evaluate cytokine profile as useful marker in modulating the personalized radiotherapy in breast cancer care.

High-Intensity Focused Ultrasound- and Radiation Therapy-Induced Immuno-Modulation: Comparison and Potential Opportunities.

In recent years, high-intensity focused ultrasound (HIFU) has emerged as a new and promising non-invasive and non-ionizing ablative technique for the treatment of localized solid tumors. Extensive pre-clinical and clinical studies have evidenced that, in addition to direct destruction of the primary tumor, HIFU-thermoablation may elicit long-term systemic host anti-tumor immunity. In particular, an important consequence of HIFU treatment includes the release of tumor-associated antigens (TAAs), the secretion of immuno-suppressing factors by cancer cells and the induction of cytotoxic T lymphocyte (CTL) activity. Radiation therapy (RT) is the main treatment modality used for many types of tumors and about 50% of all cancer patients receive RT, often used in combination with surgery and chemotherapy. It is well known that RT can modulate anti-tumor immune responses, modifying micro-environment and stimulating inflammatory factors that can greatly affect cell invasion, bystander effects, radiation tissue complications (such as fibrosis), genomic instability and thus, intrinsic cellular radio-sensitivity. To date, various combined therapeutic strategies (such as immuno-therapy) have been performed in order to enhance RT success in treating locally advanced and recurrent tumors. Recent works suggested the combined use of HIFU and RT treatments to increase the tumor cell radio-sensitivity, in order to synergize the effects reaching the maximum results with minimal doses of ionizing radiation (IR). Here, we highlight the opposite immuno-modulation roles of RT and HIFU, providing scientific reasons to test, by experimental approaches, the use of HIFU immune-stimulatory capacity to improve tumor radio-sensitivity, to reduce the RT induced inflammatory response and to decrease the dose-correlated side effects in normal tissues.

Caveolin-1, breast cancer and ionizing radiation.

Breast cancer (BC) recovery has increased in recent years thanks to efforts of Omics-based research in this field. However, despite the important results obtained, BC remains a complex multifactorial pathology that is difficult to treat appropriately. Caveolin-1 (CAV1), the basic constituent protein of specialized plasma membrane invaginations called caveolae, is emerging as a potential therapeutic biomarker in BC. This factor may modulate BC response to chemotherapy and radiation therapy. In addition, recent reports describe the key role of CAV1 during cell response to oxidative stress. The aim of the present review was to describe the biological roles of CAV1 in BC considering its contrasting dual functions as an oncogene and as a tumor suppressor. In addition, we report on how CAV1 may contribute to tumor cell response to ionizing radiation treatment. Finally, new roles of CAV1 in BC both on epithelium and stroma may be useful as prognostic indicators for patient treatment and help clinicians in the selection of the best personalized therapy.

Gene Expression Profiling of MCF10A Breast Epithelial Cells Exposed to IOERT.

BACKGROUND/AIM: Intraoperative electron radiation therapy (IOERT) is a therapeutic approach that delivers a single high dose of ionizing radiation (IR) directly to the tumor bed during cancer surgery. The main goal of IOERT is to counteract tumor growth by acting on residual cancer cells as well as to preserve healthy surrounding tissue from the side-effects of radiation therapy. The radiobiology of the healthy tissue response to IR is a topic of interest which may contribute to avoiding impairment of normal tissue and organ function and to reducing the risks of secondary cancer. The purpose of the study was to highlight cell and gene expression responses following IOERT treatment in the human non-tumorigenic MCF10A cell line in order to find new potential biomarkers of radiosensitivity/radioresistance. MATERIAL AND METHODS: Gene-expression profiling of MCF10A cells treated with 9 and 23 Gy doses (IOERT boost and exclusive treatment, respectively), was performed by whole-genome cDNA microarrays. Real-time quantitative reverse transcription (qRT-PCR), immunofluorescence and immunoblot experiments were carried out to validate candidate IOERT biomarkers. Clonogenic tests and morphological evaluations to examine cellular effects induced by radiation were also conducted. RESULTS: The study revealed a dose-dependent gene-expression profile and specific key genes that may be proposed as novel markers of radiosensitivity. Our results show consistent differences in non-tumorigenic cell tolerance and in the molecular response of MCF10A cells to different IOERTs. In particular, after 9 Gy of exposure, the selection of a radioresistant cell fraction was observed. CONCLUSION: The possibility of clarifying the molecular strategies adopted by cells in choosing between death or survival after IR-induced damage opens-up new avenues for the selection of a proper personalized therapy schedule.