PURPOSE:The purpose of this phase 2 clinical study is to investigate the safety and feasibility of a single fraction stereotactic partial breast irradiation (S-PBI) for early-stage breast cancer in the preoperative setting and to evaluate tumor response to a single large radiation dose through pathologic examination and immunohistochemistry analysis of the surgical specimen. METHODS AND MATERIALS:This single arm, phase 2 clinical trial includes patients in postmenopausal status, over the age of 50 years, with early-stage (cT1-T2 cN0) breast cancer, luminal type, any grade, unifocal tumor, and suitable for breast conserving surgery. The gross tumor volume includes the tumor. The clinical target volume corresponds to gross tumor volume. The planning target volume is created by adding 3 mm symmetrical margins from the clinical target volume. Treatment is delivered through GammaPod technology as single fraction radiosurgery, to a total dose of 30 to 36 Gy. Surgery is performed 8 to 28 weeks after S-PBI. Pathologic response is classified as pathologic complete response (pCR), near complete response with <10% of residual disease, pathologic partial response with 10% to 90% of residual disease, or stable disease with >90% of residual disease. We further group pCR and near complete response together as "Major Response." RESULTS:From January 2022 to November 2023, 49 patients were enrolled and underwent S-PBI followed by breast conserving surgery. The rate of Major Response was 37%, including pCR in 18% of cases. The mean Ki-67 index was reduced from 9.5% pre-S-PBI to 2% post-S-PBI. CONCLUSIONS:Preoperative single fraction S-PBI appears to be associated with a promising rate of "Major Response," including cases of complete response.

Background and Objectives: Breast lesions of uncertain malignant potential identified on biopsy, known as “B3 lesions,” constitute a significant portion of diagnoses in numerous published studies. These lesions are associated with a variable risk of coinciding malignant tumors, and current guidelines recommend complete excision, which can occasionally lead to an upgrade in the resection specimen. However, alternative, less invasive treatment strategies, such as clinical follow-up, may be considered. In this study, we retrospectively analyzed diagnostic biopsies from our institution to determine the upgrade rate of each B3 lesion subgroup to breast malignancy following complete excision. Materials and Methods: All breast biopsies conducted at our institution from 1 January 2018 to 30 November 2022 and classified as B3 lesions were included in this study. The lesions were categorized into groups and subgroups based on their growth pattern and histopathological features. To determine the upgrade rate to ductal carcinoma in situ (DCIS) or invasive breast cancer (IBC) for each B3 lesion subgroup, we assessed the histological concordance between the biopsy and the resection specimen. Results: During the study period, 10,531 biopsies were performed, of which 1045 (9.93%) were classified as B3 lesions. Among these, 795 (76.08%) were subsequently resected, either through surgical procedures (98.32%) or using the Vacuum-Assisted Excision technique (1.68%). Histological examination revealed that 89 (11.19%) of the resected B3 lesions were upgraded to breast malignancy, with 59 cases (7.42%) progressing to DCIS, 22 cases (2.76%) to IBC, and 8 cases (1.01%) to borderline or malignant phyllodes tumor. The upgrade rate varied among histopathological subgroups, being lowest in complex sclerosing lesions without atypia (4.95%, 95% CI: 2.5–8.7%) and highest in intraductal papillomas with atypia (58.82%; 95% CI: 32.9–81.6%). Conclusions: Statistically significant differences were observed between B3 lesion subgroups, with a higher risk of upgrade in lesions exhibiting atypia. As our understanding of B3 lesions evolves, there is potential to implement therapeutic strategies tailored to the specific risk associated with each subgroup. This approach could allow for less invasive management options, such as clinical or radiological follow-up, thereby sparing patients from unnecessary invasive procedures when appropriate.

Glioblastoma (GBM) is the most common and lethal primary brain tumour. The prognosis for GBM patients remains poor due to rapid tumour recurrence and resistance to conventional treatments. Small Rho GTPase proteins, which regulate cell shape and motility, are critical for GBM aggressive growth and infiltration into the surrounding brain parenchyma. Hence, small-molecule inhibitors targeting them represent an appealing opportunity to hinder the infiltration behaviour of GBM. Here, a synergistic experimental and computational approach allowed us to identify an inhibitor that reduces migration in patient-derived GBM cell lines. Computational and in vitro functional assays reveal that this compound inhibits Rho GTPases function by targeting multiple allosteric sites thereby enhancing flexibility of key functional regions and hindering their interaction with protein regulators. Our research unveiled a novel hit molecule targeting Rho GTPases with significant potential to improve the treatment of GBM and other highly aggressive tumours.

Risk factors for intrahepatic recurrence after hepatectomy for HCC are incompletely understood. Intrahepatic lymphatic drainage through the Glisson’s sub-serosal system may represent a pathway favoring intrahepatic spread of cancer cells. This study investigates the prognostic role of Glisson’s capsule invasion by hepatocellular carcinoma (HCC) after hepatectomy. Of 192 patients undergoing curative hepatectomy at a single institution between 2010 and 2023, 84 met the inclusion criteria and were retrospectively analyzed. Primary endpoints of intra-hepatic recurrence rate and pattern, and secondary survival endpoints were analyzed between patients with microscopic presence vs. absence of Glisson’s capsule invasion by HCC. Microscopic tumor invasion of the Glisson’s capsule was found in 39 (46.4

ABSTRACT Objective This study aimed to identify risk factors associated with the development of metastases in breast cancer patients, to investigate survival rates, and the relationship between local recurrences and distant metastases. Methods This retrospective case‐cohort study included women with breast cancer who were treated at a certified Breast Unit between 2001 and 2015. Cases who developed distant metastases were compared to controls based on diagnosis year, stage, and age at diagnosis. Comprehensive information on patient characteristics, tumor biology, and treatment options was gathered. Results The study included 412 patients who developed distant metastases and 433 controls who remained metastasis‐free over a median follow‐up of 150 months (interquartile range 87–202). The 20‐year overall survival was 99.23% for the control group and 23.62% for those with metastasis (p < 0.01). Significant risk factors for metastasis included lobular invasive carcinoma (odds ratio (OR) 2.26, p < 0.001), triple‐negative subtype (OR 4.06, p = 0.002), high tumor grade (OR 2.62, p = 0.004), larger tumor size (OR 1.02, p < 0.001), lymph node involvement (p < 0.001), and loco‐regional recurrence (OR 4.32, p < 0.001). Progesterone receptor (PR) expression was protective (OR 0.52, 95% confidence interval 0.34–0.81, p = 0.003). Machine learning models supported these findings, though their clinical significance was limited. Conclusions Lobular invasive carcinoma, specific tumor subtypes, high grade, large tumor size, lymph node involvement, and loco‐regional recurrence are all significant risk factors for distant metastasis, whereas PR expression is protective. The potential of machine learning in predicting metastasis was explored, showing promise for future personalized risk assessment.

Glioblastoma Multiforme (GBM) is the most aggressive primary tumor of the Central Nervous System (CNS) with a low survival rate. The malignancy of GBM is sustained by a bidirectional crosstalk between tumor cells and the Tumor Microenvironment (TME). This mechanism of intercellular communication is mediated, at least in part, by the release of exosomes. Glioma-Derived Exosomes (GDEs) work, indeed, as potent signaling particles promoting the progression of brain tumors by inducing tumor proliferation, invasion, migration, angiogenesis and resistance to chemotherapy or radiation. Given their nanoscale size, exosomes can cross the blood–brain barrier (BBB), thus becoming not only a promising biomarker to predict diagnosis and prognosis but also a therapeutic target to treat GBM. In this review, we describe the structural and functional characteristics of exosomes and their involvement in GBM development, diagnosis, prognosis and treatment. In addition, we discuss how exosomes can be modified to be used as a therapeutic target/drug delivery system for clinical applications.

Introduction/Background In 2020, the European Society of Gynecological Oncology/European Society for Radiotherapy and Oncology/European Society of Pathology (ESGO/ESTRO/ESP) integrated molecular classification into the classic risk stratification based on clinicopathological features of patients with endometrial cancer (EC), thus introducing a new decision-making for clinicians in adjuvant therapy. Therefore, each surgical specimen should be tested for mutations in the tumor protein 53 (p53), in mismatch repair (MMR) proteins and Polimerase Epsilon (POLE). However, this may be too expensive and not always necessary. Betella's algorithm (figure 1) therefore pointed to test only for mutations that would actually change the risk group and consequently the adjuvant treatment: first of all, analysis of MMR proteins; then research for p53 mutation in stages I and II of disease according to International Federation of Gynecology and Obstetrics (FIGO) 2009; finally, the sequencing of POLE when class migration occurred. Currently, available data allow class migration 'only' in stage I-II endometrial cancers, in FIGO grade 3 and when lymphovascular spaces are involved. Methodology The study aimed to externally validate Betella's algorithm by retrospective analysis of EC cases treated between March 2021 and March 2023 at the Gynecology Clinic of S. Maria della Misericordia Hospital in Udine, Italy. Results Of 102 patients underwent surgical staging, complete molecular analysis was obtained in 97%. Integrating molecular classification in the calculation of risk groups resulted in a risk group move in 11.1% of patients: 7 moved to a lower risk group due to POLE mutations, while 4 shifted to a higher risk group due to p53 mutations. By applying the Betella algorithm, we were able to spare POLE sequencing in 65 cases (65.7%) and p53 immunohistochemistry in 17 cases (17.2%) (figure 2). Conclusion The application of this proposed new algorithm appears safe for patients and allows rationalization of resources. Disclosures Authors have no disclosures to declare.

Our study investigates the post-mortem findings of the diaphragm's muscular structural changes in mechanically ventilated COVID-19 patients. Diaphragm samples of the right side from 42 COVID-19 critically ill patients were analyzed and correlated with the type and length of mechanical ventilation (MV), ventilatory parameters, prone positioning, and use of sedative drugs. The mean number of fibers was 550±626. The cross-sectional area was 4120±3280 μm2, while the muscular fraction was 0.607±0.126. The overall population was clustered into two distinct populations (clusters 1 and 2). Cluster 1 showed a lower percentage of slow myosin fiber and higher fast fiber content than cluster 2, 68% versus 82%, p<0.00001, and 29.8% versus 18.8%, p=0.00045 respectively. The median duration of MV was 180 (41-346) hours. In cluster 1, a relationship between assisted ventilation and fast myosin fiber percentage (R2=-0.355, p=0.014) was found. In cluster 2, fast fiber content increased with increasing the length of the controlled MV (R2=0.446, p=0.006). A high grade of fibrosis was reported. Cluster 1 was characterized by fibers’ atrophy and cluster 2 by hypertrophy, supposing different effects of ventilation on the diaphragm but without excluding a possible direct viral effect on diaphragmatic fibers.

Primary Sjögren's syndrome (pSS) is a chronic, systemic, inflammatory autoimmune disease characterised by lymphocyte proliferation and progressive damage to exocrine glands. Salivary gland histopathology based on salivary gland biopsy is relevant for the diagnosis of pSS and therefore broadly applied in clinical practice. Tissue can be obtained from labial salivary glands (LSG) biopsy or from major salivary glands (MSG) biopsy, namely the parotid; in this latter scenario, the procedure can be either an open surgical biopsy or a US guided core needle biopsy.In this review we will: i) present the histopathological findings that may be encountered by pathologists on biopsies from pSS patients; ii) discuss the advantages and disadvantages of the surgical and/or imaging guided procedures to obtain tissues from LSG or MSG; iii) describe the histopathological features of lymphoma of MSG in pSS patients.

Background: Val804Met RET is one of the most common genetic alterations in Multiple Endocrine Neoplasia 2 and is considered to confer only a moderate risk for familial medullary thyroid carcinoma (MTC). The associated phenotype can however be much more complex in some cases.Methods: A clinical, genetic, and pathological analysis was conducted on a family cluster of thyroid neoplasms associated with Val804Met RET mutation.Results: All the kindreds who are carriers of the mutated RET received total thyroidectomy + /- VI level dissection. The proband presented with a pT1bN0 MTC, her 29-yo brother showed a concomitant papillary thyroid carcinoma (PTC) and MTC, their father had a pT1a PTC plus a follicular adenoma, while the uncle of the proband showed C-cell hyperplasia. None had clinical or biochemical evidence of parathyroid disorders or pheochromocytoma. Conclusions: In the presence of Val804Met RET several types of thyroid premalignant and malignant should be screened for, and without limiting to MTC.

Advancements in cutting-edge molecular profiling techniques, such as next-generation sequencing and bioinformatic analytic tools, have allowed researchers to examine tumour biology in detail and stratify patients based on factors linked with clinical outcome and response to therapy. This manuscript highlights the most relevant prognostic and predictive biomarkers in kidney, bladder, prostate and testicular cancers with recognised impact in clinical practice. In bladder and prostate cancer, new genetic acquisitions concerning the biology of tumours have modified the therapeutic scenario and led to the approval of target directed therapies, increasing the quality of patient care. Thus, it has become of paramount importance to choose adequate molecular tests, i.e., FGFR screening for urothelial cancer and BRCA1-2 alterations for prostate cancer, to guide the treatment plan for patients. While no tissue or blood-based biomarkers are currently used in routine clinical practice for renal cell carcinoma and testicular cancers, the field is quickly expanding. In kidney tumours, gene expression signatures might be the key to identify patients who will respond better to immunotherapy or anti-angiogenic drugs. In testicular germ cell tumours, the use of microRNA has outperformed conventional serum biomarkers in the diagnosis of primary tumours, prediction of chemoresistance, follow-up monitoring, and relapse prediction.

The treatment response criteria in autoimmune hepatitis (AIH) have been recently updated. This study aimed to assess treatment responses in 39 (16 males) patients with AIH confirmed by histology. Prednisone added to azathioprine or mycophenolate was the most frequent first-line treatment. Serum alanine aminotransferase (ALT) levels were periodically checked for a median of 45 months. Eight (20.5%) patients presented 4 weeks non-response (NR). Baseline lower multiples of ALT above the upper normal limit (UNL) (p = 0.005), Ishak liver fibrosis score > 3 (p = 0.029), and less frequent confluent necrosis > 2 (p < 0.001) were independent predictors of NR. 24 (61.5%) patients achieved complete biochemical response (CBR) at six months. Ishak liver fibrosis score ≤ 3 (p < 0.001), lobular eosinophilic infiltrate (p < 0.001), and ≥50% decrease in serum ALT levels at week 4 (p < 0.001) were independent predictors of CBR. In addition, the GLUCRE score, derived from the multiplication of serum creatinine (mg/dL) and glucose (mg/dL) levels, were identified. A baseline GLUCRE value > 100 strongly predicted CBR failure (p = 0.003) at a follow-up greater than 12 months. In conclusion, the absence of cirrhosis and a ≥50% UNL decrease in serum ALT levels were independent predictors for CBR. A baseline GLUCRE score may help identify patients maintaining longer CBR.

Objective The aim of the study was to culture vital salivary gland organoids obtained through labial or parotid biopsy of primary Sjogren's syndrome (pSS) patients in order to evaluate their morphological and functional features in basal condition and after stimulation with Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) activator forskolin and phosphodiesterase 4 (PDE4) inhibitor apremilast, their in vitro regenerative capacity and the immune-histological resemblance with original tissue. Methods Salivary gland tissues from five pSS patients were processed to obtain vital organoids; swelling assay and cell proliferation tests were performed after forskolin and apremilast application. Immunochemistry evaluation on original salivary gland tissue and corresponding organoids was performed, and secretomics analysis was conducted to assess their functional status. Results After application of forskolin and apremilast, we observed organoid swelling after 30 minutes, compatible with a positive functional status and enhancement of saliva production. In 3 cases, apremilast induced organoid proliferation. All cases were positive for cytokeratin 14 (CK14) and most for cytokeratin 5 (CK5). All the cases were positive for amylase; its secretion, and thus functional status of organoids, was confirmed by its high concentration in the culture medium. A focal ductal differentiation was found in some cases, highlighted by epithelial membrane antigen (EMA) positivity. The more differentiated EMA positive areas were negative for the staminal marker CK14, showing a sort of "complementary staining". Conclusion Our data highlighted that differentiated cells and vital functional organoids that recapitulate the development of original salivary glands can be obtained from pSS epithelium. For the first time, the direct stimulating effect of PDE4 inhibitor apremilast on pSS human salivary gland organoids is reported, opening new perspectives on targeting oral dryness with drugs that combine secretagogue and immunomodulatory effects.

A 49-year-old woman was referred to the Maxillofacial Surgery Department of our University to evaluate a worsening jaw pain lasting over 3 years. During the day and night, she experienced increasing preauricular pain in the area of the left temporomandibular joint (TMJ) and a progressive limitation of mouth opening over the past few years. The patient denied bruxism or jaw clenching. She had been treated by various health professionals, including physiotherapists, osteopaths, and orthodontists over the years. She also used 4 different splints, with little or no symptomatologic relief. Non-steroidal anti-inflammatory drugs (NSAIDs) and muscle relaxants were taken chronically by the patient with little or no result. The patient did not report any previous trauma, infections, arthritis, or other rheumatological disease. At physical examination, she showed a maximum mouth opening (MMO) of 29 mm with the chin deviating slightly to the left side (Figure 1 A and B). She was painful on palpation of the preauricular and temporal area bilaterally. Furthermore, the masticatory muscles resulted bilaterally painful. Mild swelling in the left TMJ area was observed. An articular crepitus of the left TMJ was perceived. Forcing the MMO, the end feel was rigid and painful. The computed tomography (CT; Figure 2 A and B) and the magnetic resonance imaging (Figure 3 A and B) scans showed an osteoarthritic degeneration of the left TMJ. Fig. 2(A-B) Coronal and sagittal computed tomography scans show a moderate osteoarthritis of the left temporomandibular joint; it is also possible to notice the sclerotic appearance of the left temporomandibular joint bones in contrast with the right side. View Large Image Figure Viewer Download Hi-res image Fig. 3(A-B) Sagittal T2w open mouth scan showing the articular disk damaged and anteriorly displaced with no reduction. Coronal T1w magnetic resonance imaging scans showing on the left temporomandibular joint signal alteration as from arthritic-sclerotic thickening of the mandibular condyle and flattening of the articular surface. View Large Image Figure Viewer Download Hi-res image

Glioma grade 4 (GG4) tumors, including astrocytoma IDH-mutant grade 4 and the astrocytoma IDH wt are the most common and aggressive primary tumors of the central nervous system. Surgery followed by Stupp protocol still remains the first-line treatment in GG4 tumors. Although Stupp combination can prolong survival, prognosis of treated adult patients with GG4 still remains unfavorable. The introduction of innovative multi-parametric prognostic models may allow refinement of prognosis of these patients. Here, Machine Learning (ML) was applied to investigate the contribution in predicting overall survival (OS) of different available data (e.g. clinical data, radiological data, or panel-based sequencing data such as presence of somatic mutations and amplification) in a mono-institutional GG4 cohort. By next-generation sequencing, using a panel of 523 genes, we performed analysis of copy number variations and of types and distribution of nonsynonymous mutations in 102 cases including 39 carmustine wafer (CW) treated cases. We also calculated tumor mutational burden (TMB). ML was applied using eXtreme Gradient Boosting for survival (XGBoost-Surv) to integrate clinical and radiological information with genomic data. By ML modeling (concordance (c)- index = 0.682 for the best model), the role of predicting OS of radiological parameters including extent of resection, preoperative volume and residual volume was confirmed. An association between CW application and longer OS was also showed. Regarding gene mutations, a role in predicting OS was defined for mutations of BRAF and of other genes involved in the PI3K-AKT-mTOR signaling pathway. Moreover, an association between high TMB and shorter OS was suggested. Consistently, when a cutoff of 1.7 mutations/megabase was applied, cases with higher TMB showed significantly shorter OS than cases with lower TMB. The contribution of tumor volumetric data, somatic gene mutations and TBM in predicting OS of GG4 patients was defined by ML modeling.

Background: COVID-19 patients carry an increased rate of thrombosis. It is controversial to which extent thrombi in the pulmonary arterial tree really contribute to disease severity with hypoxemia secondary to microvascular/ lung parenchymal damage with viral alveolitis considered to play the main role in critical disease.Objectives: The primary objective was to compare post-mortem lung disease from fatal COVID-19 pneumonia in patients with macroscopically evident pulmonary arterial tree thrombosis and patients without, by character-izing the immunohistochemical nature of thrombi, and by comparing clinical and laboratory features of these patients with other COVID-19 patients who died but without evidence of pulmonary arterial thrombosis (controls). Patients and methods: 13 COVID-19 pneumonia cases (mean age +/- standard deviation: 74 +/- 6.5 years) with macroscopically visible pulmonary arterial thrombosis were compared to 14 controls. Hematoxylin and Eosin stained slides were reviewed choosing those with visible pulmonary thrombi which were further characterized by immunohistochemistry, in particular for the inflammatory infiltrates. Ante mortem serum markers relevant to pulmonary embolism were evaluated in both groups.Results: Twenty arterial thrombi (5 cases with multiple thrombi) were selected for study and were composed by white blood cells (WBC) [median, IQR range: 10 % (5-12.25)], mainly neutrophils [58 % (35.2-64.5)]. Cases with thrombosis showed significantly higher levels of platelet count [median, IQR range: 195000/mmc (157750-274,500) vs 143,500 (113000-175,250), p = 0.011], LDH [854 U/L (731-1315) vs 539 (391.5-660), p = 0.003] at admission, and D-dimer at ICU transfer [25,072 FEU (6951-50,531) vs 1024 (620-5501), p = 0.003].Conclusions: Immunothrombotically driven arterial thrombi in COVID-19 patients are associated with D-Dimer and LDH elevations, thus linking inflammation, coagulopathy and organ damage in fatal COVID-19.

Background: Chicken Ovalbumin Upstream Promoter-Transcription Factor I (COUP-TFI) is a member of the steroid/thyroid nuclear receptor superfamily. The aim of this study was to investigate whether absence of this gene affects placental development and fetal growth in a COUP-TFI knockout mouse model. Methods: Placentas of COUP-TFI-knockout (COUP-TFI KO) and wild-type (WT) were collected at 18.5 days post-coitum. The expression level of the following genes known to be involved in different key molecular pathways was evaluated: BCL2Associated X (Bax) and B-cell lymphoma 2 (Bcl-2) (apoptosis), p21, p53 andα subunit of inhibin (INHA) (proliferation and apoptosis), vascular endothelial growth factor A (VEGF-A), placental growth factor (PlGF), hypoxia-inducible factor 1-alpha (HIF1α), Fms related receptor tyrosine kinase 1 (Flt-1), and endoglin (ENG) (angiogenesis). Mouse litter weight at birth was also assessed. Results: RT-qPCR analysis showed increased mRNA expression of VEGF-A and Bax in placental tissue of COUP-TFI KO mice compared to WT mice. We also found a loss in the positive correlation between Bcl-2 and INHA, p21 and ENG, as well as HIF1α and Flt-1 mRNA expression in COUP-TFI mutants. Finally, KO mice were lighter than WT littermates (respectively, the mean weight of COUP-TFI KOmice was 1.3 grams,± 0.13, compared to 1.6 g,± 0.14 ofWTmice, p< 0.05). Conclusions: Our results show that COUP-TFI deletion is associated with a lower birth weight in mice and increased placental transcript expression of pro-apoptotic Bax and pro-angiogenetic VEGF-A genes.

Despite the state-of-the-art treatment, patients diagnosed with glioblastoma (GBM) have a median overall survival (OS) of 14 months. The insertion of carmustine wafers (CWs) into the resection cavity as adjuvant treatment represents a promising option, although its use has been limited due to contrasting clinical results. Our retrospective evaluation of CW efficacy showed a significant improvement in terms of OS in a subgroup of patients. Given the crucial role of the tumor microenvironment (TME) in GBM progression and response to therapy, we hypothesized that the TME of patients who benefited from CW could have different properties compared to that of patients who did not show any advantage. Using an in vitro model of the glioma microenvironment, represented by glioma-associated-stem cells (GASC), we performed a transcriptomic analysis of GASC isolated from tumors of patients responsive and not responsive to CW to identify differentially expressed genes. We found different transcriptomic profiles, and we identified four genes, specifically down-regulated in GASC isolated from long-term survivors, correlated with clinical data deposited in the TCGA–GBM dataset. Our results highlight that studying the in vitro properties of patient-specific glioma microenvironments can help to identify molecular determinants potentially prognostic for patients treated with CW.

Background Assessment of hepatic steatosis (HS) before transplantation requires the pathologist to read a graft biopsy. A simple method based on the evaluation of images from tissue samples with a smartphone could expedite and facilitate the liver selection. This study aims to assess the degree of HS by analysing photographic images from liver needle biopsy samples. Methods Thirty-three biopsy-images were acquired with a smartphone. Image processing was carried out using ImageJ: background subtraction, conversion to HSB colour space, segmentation of the biopsy area, and evaluation of statistical features of Hue, Saturation, Brightness, Red, Green, and Blue channels on the biopsy area. After feature extraction, correlations were made with gold standard HS percentage assessed at two levels (frozen-section vs glass-slide). Sensitivity, specificity, and accuracy were calculated for each feature. Results Correlations were found for H, S, R. The sensitivity, specificity, and accuracy of the final classifier based on the K* algorithm were 94%, 92%, 94%. Limitations Accuracy assessment was performed considering macrovesicular steatosis on specimens with mostly < 30% HS. Conclusions The steatosis assessment based on needle biopsy images, proved to be an effective and promising method. Deep learning approaches could also be experimented with a larger set of images

Background Colorectal cancer (CRC) represents the fourth leading cause of cancer-related deaths. The heterogeneity of CRC identity limits the usage of cell lines to study this type of tumor because of the limited representation of multiple features of the original malignancy. Patient-derived colon organoids (PDCOs) are a promising 3D-cell model to study tumor identity for personalized medicine, although this approach still lacks detailed characterization regarding molecular stability during culturing conditions. Correlation analysis that considers genomic, transcriptomic, and proteomic data, as well as thawing, timing, and culturing conditions, is missing. Methods Through integrated multi-omics strategies, we characterized PDCOs under different growing and timing conditions, to define their ability to recapitulate the original tumor. Results Whole Exome Sequencing allowed detecting temporal acquisition of somatic variants, in a patient-specific manner, having deleterious effects on driver genes CRC-associated. Moreover, the targeted NGS approach confirmed that organoids faithfully recapitulated patients' tumor tissue. Using RNA-seq experiments, we identified 5125 differentially expressed transcripts in tumor versus normal organoids at different time points, in which the PTEN pathway resulted of particular interest, as also confirmed by further phospho-proteomics analysis. Interestingly, we identified the PTEN c.806_817dup (NM_000314) mutation, which has never been reported previously and is predicted to be deleterious according to the American College of Medical Genetics and Genomics (ACMG) classification. Conclusion The crosstalk of genomic, transcriptomic and phosphoproteomic data allowed to observe that PDCOs recapitulate, at the molecular level, the tumor of origin, accumulating mutations over time that potentially mimic the evolution of the patient's tumor, underlining relevant potentialities of this 3D model.