In Vivo and Ex Vivo Gene Electrotransfer in Ophthalmological Disorders.

The aim of this document is to present an overview of gene electrotransfer in ophthalmological disorders. In order to ensure an adequate variety of the assessed studies, several electronic databases were considered and studies published between January 1998 and December 2021 were analysed. Three investigators carried out data extraction and analysis, focusing on both technical (i.e., electrical protocol, type of electrode, plasmid) and medical (i.e., type of study, threated disease) aspects and highlighting the main differences in terms of results obtained. Moreover, the IGEA experience in the project "Transposon-based, targeted ex vivo gene therapy to treat age-related macular degeneration" (TargetAMD) was reported in the results section. No clinical trial was found on international literature and on ClinicalTrials.gov. Twelve preclinical studies were found including in vivo and ex-vivo applications. The studied showed that electrotransfer could be very efficient for plasmid DNA transfection. Many attempts such as modification of the electric field, buffers and electrodes have been made and the optimization of electric field setting seems to be very important. Using this technique, gene replacement can be designed in cases of retinal inheritance or corneal disease and a wide range of human eye diseases could, in the future, benefitfrom these gene therapy technologies.

Prediction of Breast Cancer Histological Outcome by Radiomics and Artificial Intelligence Analysis in Contrast-Enhanced Mammography.

PURPOSE: To evaluate radiomics features in order to: differentiate malignant versus benign lesions; predict low versus moderate and high grading; identify positive or negative hormone receptors; and discriminate positive versus negative human epidermal growth factor receptor 2 related to breast cancer. METHODS: A total of 182 patients with known breast lesions and that underwent Contrast-Enhanced Mammography were enrolled in this retrospective study. The reference standard was pathology (118 malignant lesions and 64 benign lesions). A total of 837 textural metrics were extracted by manually segmenting the region of interest from both craniocaudally (CC) and mediolateral oblique (MLO) views. Non-parametric Wilcoxon-Mann-Whitney test, receiver operating characteristic, logistic regression and tree-based machine learning algorithms were used. The Adaptive Synthetic Sampling balancing approach was used and a feature selection process was implemented. RESULTS: In univariate analysis, the classification of malignant versus benign lesions achieved the best performance when considering the original_gldm_DependenceNonUniformity feature extracted on CC view (accuracy of 88.98%). An accuracy of 83.65% was reached in the classification of grading, whereas a slightly lower value of accuracy (81.65%) was found in the classification of the presence of the hormone receptor; the features extracted were the original_glrlm_RunEntropy and the original_gldm_DependenceNonUniformity, respectively. The results of multivariate analysis achieved the best performances when using two or more features as predictors for classifying malignant versus benign lesions from CC view images (max test accuracy of 95.83% with a non-regularized logistic regression). Considering the features extracted from MLO view images, the best test accuracy (91.67%) was obtained when predicting the grading using a classification-tree algorithm. Combinations of only two features, extracted from both CC and MLO views, always showed test accuracy values greater than or equal to 90.00%, with the only exception being the prediction of the human epidermal growth factor receptor 2, where the best performance (test accuracy of 89.29%) was obtained with the random forest algorithm. CONCLUSIONS: The results confirm that the identification of malignant breast lesions and the differentiation of histological outcomes and some molecular subtypes of tumors (mainly positive hormone receptor tumors) can be obtained with satisfactory accuracy through both univariate and multivariate analysis of textural features extracted from Contrast-Enhanced Mammography images.

Radiomic and Artificial Intelligence Analysis with Textural Metrics Extracted by Contrast-Enhanced Mammography and Dynamic Contrast Magnetic Resonance Imaging to Detect Breast Malignant Lesions.

Purpose:The purpose of this study was to discriminate between benign and malignant breast lesions through several classifiers using, as predictors, radiomic metrics extracted from CEM and DCE-MRI images. In order to optimize the analysis, balancing and feature selection procedures were performed. Methods: Fifty-four patients with 79 histo-pathologically proven breast lesions (48 malignant lesions and 31 benign lesions) underwent both CEM and DCE-MRI. The lesions were retrospectively analyzed with radiomic and artificial intelligence approaches. Forty-eight textural metrics were extracted, and univariate and multivariate analyses were performed: non-parametric statistical test, receiver operating characteristic (ROC) and machine learning classifiers. Results: Considering the single metrics extracted from CEM, the best predictors were KURTOSIS (area under ROC curve (AUC) = 0.71) and SKEWNESS (AUC = 0.71) calculated on late MLO view. Considering the features calculated from DCE-MRI, the best predictors were RANGE (AUC = 0.72), ENERGY (AUC = 0.72), ENTROPY (AUC = 0.70) and GLN (gray-level nonuniformity) of the gray-level run-length matrix (AUC = 0.72). Considering the analysis with classifiers and an unbalanced dataset, no significant results were obtained. After the balancing and feature selection procedures, higher values of accuracy, specificity and AUC were reached. The best performance was obtained considering 18 robust features among all metrics derived from CEM and DCE-MRI, using a linear discriminant analysis (accuracy of 0.84 and AUC = 0.88). Conclusions: Classifiers, adjusted with adaptive synthetic sampling and feature selection, allowed for increased diagnostic performance of CEM and DCE-MRI in the differentiation between benign and malignant lesions.

Radiomics Metrics Combined with Clinical Data in the Surgical Management of Early-Stage (cT1-T2 N0) Tongue Squamous Cell Carcinomas: A Preliminary Study.

Objective: To predict the risk of metastatic lymph nodes and the tumor grading related to oral tongue squamous cell carcinoma (OTSCC) through the combination of clinical data with radiomics metrics by computed tomography, and to develop a supportive approach in the management of the lymphatic cervical areas, with particular attention to the early stages (T1−T2). Between March 2016 and February 2020, patients with histologically confirmed OTSCC, treated by partial glossectomy and ipsilateral laterocervical lymphadenectomy and subjected to computed tomography (CT) before surgery, were identified by two centers: 81 patients (49 female and 32 male) with 58 years as the median age (range 19−86 years). Univariate analysis with non-parametric tests and multivariate analysis with machine learning approaches were used. Clinical, hematological parameters and radiological features extracted by CT were considered individually and in combination. All clinical parameters showed statistically significant differences (p < 0.05) for the Kruskal−Wallis test when discriminating both the tumor grading and the metastatic lymph nodes. DOI, PLR, SII, and SIRI showed an accuracy of 0.70 (ROC analysis) when identifying the tumor grading, while an accuracy ≥ 0.78 was shown by DOI, NLR, PLR, SII, and SIRI when discriminating metastatic lymph nodes. In the context of the analysis of radiomics metrics, the original_glszm_HighGrayLevelZoneEmphasis feature was selected for identifying the tumor grading (accuracy of 0.70), while the wavelet_HHH_glrlm_LowGrayLevelRunEmphasis predictor was selected for determining metastatic lymph nodes (accuracy of 0.96). Remarkable findings were also obtained when classifying patients with a machine learning approach. Radiomics features alone can predict tumor grading with an accuracy of 0.76 using a logistic regression model, while an accuracy of 0.82 can be obtained by running a CART algorithm through a combination of three clinical parameters (SIRI, DOI, and PLR) with a radiomics feature (wavelet_LLL_glszm_SizeZoneNonUniformityNormalized). In the context of predicting metastatic lymph nodes, an accuracy of 0.94 was obtained using 15 radiomics features in a logistic regression model, while both CART and CIDT achieved an asymptotic accuracy value of 1.00 using only one radiomics feature. Radiomics features and clinical parameters have an important role in identifying tumor grading and metastatic lymph nodes. Machine learning approaches can be used as an easy-to-use tool to stratify patients with early-stage OTSCC, based on the identification of metastatic and non-metastatic lymph nodes.

Reduction of muscle contraction and pain in electroporation-based treatments: An overview.

BACKGROUND: In the first studies of electrochemotherapy (ECT), small cutaneous metastases were treated and only mild or moderate pain was observed; therefore, pain was not considered a significant issue. As the procedure began to be applied to larger cutaneous metastases, pain was reported more frequently. For that reason, reduction of both muscle contractions and pain have been investigated over the years. AIM: To present an overview of different protocols described in literature that aim to reduce muscle contractions and pain caused by the electroporation (EP) effect in both ECT and irreversible EP treatments. METHODS: Thirty-three studies published between January 1999 and November 2020 were included. Different protocol designs and electrode geometries that reduce patient pain and the number of muscle contractions and their intensity were analysed. RESULTS: The analysis showed that both high frequency and bipolar/biphasic pulses can be used to reduce pain and muscle contractions in patients who undergo EP treatments. Moreover, adequate electrode design can decrease EP-related morbidity. Particularly, needle length, diameter and configuration of the distance between the needles can be optimised so that the muscle volume crossed by the current is reduced as much as possible. Bipolar/biphasic pulses with an inadequate pulse length seem to have a less evident effect on the membrane permeability compared with the standard pulse protocol. For that reason, the number of pulses and the voltage amplitude, as well as the pulse duration and frequency, must be chosen so that the dose of delivered energy guarantees EP efficacy. CONCLUSION: Pain reduction in EP-based treatments can be achieved by appropriately defining the protocol parameters and electrode design. Most results can be achieved with high frequency and/or bipolar/biphasic pulses. However, the efficacy of these alternative protocols remains a crucial point to be assessed further.

A Systematic Review about Imaging and Histopathological Findings for Detecting and Evaluating Electroporation Based Treatments Response.

BACKGROUND: Imaging methods and the most appropriate criteria to be used for detecting and evaluating response to oncological treatments depend on the pathology and anatomical site to be treated and on the treatment to be performed. This document provides a general overview of the main imaging and histopathological findings of electroporation-based treatments (Electrochemotherapy-ECT and Irreversible electroporation-IRE) compared to thermal approach, such as radiofrequency ablation (RFA), in deep-seated cancers with a particular attention to pancreatic and liver cancer. METHODS: Numerous electronic datasets were examined: PubMed, Scopus, Web of Science and Google Scholar. The research covered the years from January 1990 to April 2021. All titles and abstracts were analyzed. The inclusion criteria were the following: studies that report imaging or histopathological findings after ablative thermal and not thermal loco-regional treatments (ECT, IRE, RFA) in deep-seated cancers including pancreatic and liver cancer and articles published in the English language. Exclusion criteria were unavailability of full text and congress abstracts or posters and different topic respect to inclusion criteria. RESULTS: 558 potentially relevant references through electronic searches were identified. A total of 38 articles met the inclusion criteria: 20 studies report imaging findings after RFA or ECT or IRE in pancreatic and liver cancer; 17 studies report histopathological findings after RFA or ECT or IRE; 1 study reports both imaging and histopathological findings after RFA or ECT or IRE. CONCLUSIONS: Imaging features are related to the type of therapy administrated, to the timing of re-assessment post therapy and to the imaging technique being used to observe the effects. Histological findings after both ECT and IRE show that the treated area becomes necrotic and encapsulated in fibrous tissue, suggesting that the size of the treated lesion cannot be measured as an endpoint to detect response. Moreover, histology frequently reported signs of apoptosis and reduced vital tissue, implying that imaging criteria, which take into account the viability and not the size of the lesion, are more appropriate to evaluate response to treatment.

Design and Characterization of a Minimally Invasive Bipolar Electrode for Electroporation.

OBJECTIVE: To test a new bipolar electrode for electroporation consisting of a single minimally invasive needle. METHODS: A theoretical study was performed by using Comsol Multiphysics® software. The prototypes of electrode have been tested on potatoes and pigs, adopting an irreversible electroporation protocol. Different applied voltages and different geometries of bipolar electrode prototype have been evaluated. RESULTS: Simulations and pre-clinical tests have shown that the volume of ablated area is mainly influenced by applied voltage, while the diameter of the electrode had a lesser impact, making the goal of minimal-invasiveness possible. The conductive pole's length determined an increase of electroporated volume, while the insulated pole length inversely affects the electroporated volume size and shape; when the insulated pole length decreases, a more regular shape of the electric field is obtained. Moreover, the geometry of the electrode determined a different shape of the electroporated volume. A parenchymal damage in the liver of pigs due to irreversible electroporation protocol was observed. CONCLUSION: The minimally invasive bipolar electrode is able to treat an electroporated volume of about 10 mm in diameter by using a single-needle electrode. Moreover, the geometry and the electric characteristics can be selected to produce ellipsoidal ablation volumes.

New Deployable Expandable Electrodes in the Electroporation Treatment in a Pig Model: A Feasibility and Usability Preliminary Study.

The aim of the study is to evaluate the usability aspects of new deployable, expandable, electrode prototypes, in terms of suitability solutions for laparoscopic applications on the liver, endoscopic trans-oral and trans-anal procedures, electroporation segmentation in several steps, mechanical functionality (flexibility, penetrability), visibility of the electrode under instrumental guidance, compatibility of the electrode with laparoscopic/endoscopic accesses, surgical instruments, and procedural room and safety compatibility. The electroporation was performed on an animal model (Sus Scrofa Large White 60 kg) both in laparoscopy and endoscopy, under ultrasound guidance, and in open surgery. Electrodes without divergence, with needles coming out straight, parallel to each other, and electrodes with peripheral needles (four needles), diverging from the electrode shaft axis (electrode with non-zero divergence) have been tested. To cause an evaluable necrosis effect, the number of electrical pulses was increased to induce immediate liver cell death. Histological samples were analyzed by staining with Haematoxylin/Eosin or by immunohistochemical staining to confirm complete necrosis. The prototypes of expandable electrodes, tested in laparoscopy and endoscopy and in open surgery, respectively, are suitable in terms of usability, electroporation segmentation in several steps, mechanical functionality (flexibility, penetrability), visibility under instrumental guidance, compatibility with laparoscopic/endoscopic accesses, surgical instruments and procedural room safety, patient safety (no bleeding and/or perforation), and treatment efficacy (adequate ablated volume). Electroporation treatment using new deployable expandable electrode prototypes is safe and feasible. Moreover, electrode configurations allow for a gradual increase in the ablated area in consecutive steps, as confirmed by histology and immunohistochemistry.