Rare Driver Mutations in Advanced, Oncogene-Addicted Non-Small Cell Lung Cancer: A North Italian, Real-World, Registry Experience.

The real-world, retrospective, NEROnE registry investigated the impact of next-generation sequencing (NGS) in advanced non-small-cell lung cancer (NSCLC) patients (pts) at three oncology units in the north of Italy between January 2020 and December 2022. We focused on the clinical characterization and outcomes of NSCLC with rare molecular alterations: EGFR exon 20 insertion, non-activating EGFR mutations, BRAF V600E and non-V600, ROS1 and RET rearrangements, MET, ErbB2, and FGFR mutations. Overall, these represented 6.4% (62/970) of the pts analysed with NGS in the daily practice. The most heavily represented rare alterations were ROS1 rearrangement (15 pts-24%) and MET exon 14 skipping mutation (11 pts-18%). No associations were found with the demographic and clinical features. Forty-nine pts received targeted therapies, of which 38.8% were first- and 9.8% were second-line. The remaining pts received chemotherapy and/or immunotherapy. In terms of the clinical outcomes, although not statistically significant, a tendency toward shorter OS was seen when therapies other than specific targeted therapies were used (HR: 1.84, 95% CI: 0.79-4.33, p = 0.158). The pts with co-mutations (19.4%) seemed to receive an advantage from the front-line chemotherapy-based regimen. Finally, an NLR score (a well-known inflammatory index) ≥ 4 seemed to be related to shorter OS among the pts treated with immunotherapy alone or in combination with chemotherapy (HR: 2.83, 95% CI: 1.08-7.40, p = 0.033). Prospective evaluations need to be performed to clarify whether these indexes may help to identify patients with oncogene-addicted NSCLC who could benefit from immunotherapy.

Prostate cancer risk stratification via eNose urine odor analysis: a preliminary report.

Introduction: Prostate cancer (PCa) is known for its highly diverse clinical behavior, ranging from low-risk, slow-growing tumors to aggressive and life-threatening forms. To avoid over-treatment of low-risk PCa patients, it would be very important prior to any therapeutic intervention to appropriately classify subjects based on tumor aggressiveness. Unfortunately, there is currently no reliable test available for this purpose. The aim of the present study was to evaluate the ability of risk stratification of PCa subjects using an electronic nose (eNose) detecting PCa-specific volatile organic compounds (VOCs) in urine samples. Methods: The study involved 120 participants who underwent diagnostic prostate biopsy followed by robot assisted radical prostatectomy (RARP). PCa risk was categorized as low, intermediate, or high based on the D'Amico risk classification and the pathological grade (PG) assessed after RARP. The eNose's ability to categorize subjects for PCa risk stratification was evaluated based on accuracy and recall metrics. Results: The study population comprised 120 participants. When comparing eNose predictions with PG an accuracy of 79.2% (95%CI 70.8 - 86%) was found, while an accuracy of 74.2% (95%CI 65.4 - 81.7%) was found when compared to D'Amico risk classification system. Additionally, if compared low- versus -intermediate-/high-risk PCa, the eNose achieved an accuracy of 87.5% (95%CI 80.2-92.8%) based on PG or 90.8% (95%CI 84.2-95.3%) based on D'Amico risk classification. However, when using low-/-intermediate versus -high-risk PCa for PG, the accuracy was found to be 91.7% (95%CI 85.2-95.9%). Finally, an accuracy of 80.8% (95%CI72.6-87.4%) was found when compared with D'Amico risk classification. Discussion: The findings of this study indicate that eNose may represent a valid alternative not only for early and non-invasive diagnosis of PCa, but also to categorize patients based on tumor aggressiveness. Further studies including a wider sample population will be necessary to confirm the potential clinical impact of this new technology.

Non-Small Cell Lung Cancer Testing on Reference Specimens: An Italian Multicenter Experience.

INTRODUCTION: Biomarker testing is mandatory for the clinical management of patients with advanced non-small cell lung cancer (NSCLC). Myriads of technical platforms are now available for biomarker analysis with differences in terms of multiplexing capability, analytical sensitivity, and turnaround time (TAT). We evaluated the technical performance of the diagnostic workflows of 24 representative Italian institutions performing molecular tests on a series of artificial reference specimens built to mimic routine diagnostic samples. METHODS: Sample sets of eight slides from cell blocks of artificial reference specimens harboring exon 19 EGFR (epidermal growth factor receptor) p.E746_AT50del, exon 2 KRAS (Kirsten rat sarcoma viral oncogene homologue) p.G12C, ROS1 (c-ros oncogene 1)-unknown gene fusion, and MET (MET proto-oncogene, receptor tyrosine kinase) Δ exon 14 skipping were distributed to each participating institution. Two independent cell block specimens were validated by the University of Naples Federico II before shipment. Methodological and molecular data from reference specimens were annotated. RESULTS: Overall, a median DNA concentration of 3.3 ng/µL (range 0.1-10.0 ng/µL) and 13.4 ng/µL (range 2.0-45.8 ng/µL) were obtained with automated and manual technical procedures, respectively. RNA concentrations of 5.7 ng/µL (range 0.2-11.9 ng/µL) and 9.3 ng/µL (range 0.5-18.0 ng/µL) were also detected. KRAS exon 2 p.G12C, EGFR exon 19 p.E736_A750del hotspot mutations, and ROS1 aberrant transcripts were identified in all tested cases, whereas 15 out of 16 (93.7%) centers detected MET exon 14 skipping mutation. CONCLUSIONS: Optimized technical workflows are crucial in the decision-making strategy of patients with NSCLC. Artificial reference specimens enable optimization of diagnostic workflows for predictive molecular analysis in routine clinical practice.

Clinicopathological Features of Non-Small Cell Lung Carcinoma with BRAF Mutation.

(1) Background: BRAF mutations affect 4-5% of lung adenocarcinomas. This study aimed to analyze the clinicopathological features of lung carcinomas with BRAF mutations, focusing on V600E vs. non-V600E and the presence of co-mutations. (2) Methods: All BRAF-mutated lung carcinomas were retrieved from a molecular diagnostic unit (the reference unit for four different hospitals). The samples were analyzed using next-generation sequencing. Statistical analyses included log-rank tests for overall survival (OS) and progression-free survival (PFS). (3) Results: In total, 60 BRAF-mutated lung carcinomas were retrieved: 24 (40.0%) with V600E and 36 (60.0%) with non-V600E mutations, and 21 (35.0%) with other co-mutations and 39 (65.0%) with only BRAF mutations. Survival data were available for 54/60 (90.0%) cases. Targeted therapy was documented in 11 cases. Patients with V600E mutations exhibited a better prognosis than patients with non-V600E mutations (p = 0.008 for OS, p = 0.018 for PFS); this was confirmed in PFS (p = 0.036) when considering only patients who received no targeted therapy. Patients with co-mutations displayed no prognostic difference compared to patients carrying only BRAF mutations (p = 0.590 for OS, p = 0.938 for PFS). (4) Conclusions: BRAF-mutated lung carcinomas with V600E (40.0%) had a better prognosis than those without V600E. Concomitant co-mutations (35.0%) did not affect the prognosis.

Implementation of an electronic nose for real -time identification of odour emission peaks at a wastewater treatment plant.

This paper proposes a novel approach for the real-time monitoring of odour emissions from a WasteWater Treatment Plant (WWTP) using an Instrumental Odour Monitoring System (IOMS). The plant is characterized by unpredictable odour peaks at its arrival tank (AT), generating nuisance and complaints in the population living nearby the plant. Odour peaks are most likely due to the conferment of non-identified and malodorous wastewaters coming from various industrial activities. Due to the high variability of sources collecting their wastewaters to the WWTP, a new methodology to train the IOMS, based on the use of a one-class classifier (OCC), has been exploited. The OCC enables to detect deviations from a "Normal Operating Region" (NOR), defined as to include odour concentrations levels unlikely to cause nuisance in the citizenship. Such deviations from the NOR thus should be representative of the odour peaks. The results obtained prove that the IOMS is able to detect real-time alterations of odour emissions from the AT with an accuracy on independent validation data of about 90% (CI95% 55-100%). This ability of detecting anomalous conditions at the AT of the WWTP allowed the targeted withdrawal of liquid and gas samples in correspondence of the odour peaks, then subjected to further analyses that in turn enabled to investigate their origin and take proper counteractions to mitigate the WWTP odour impact.

High-grade transformation of a polymorphous adenocarcinoma of the salivary gland: a case report and review of the literature.

Background: Polymorphous adenocarcinoma (PAC) represents the second most widespread neoplasm of the minor salivary glands. These tumors rarely develop a histological progression from low-grade to high-grade malignancy, named "high-grade transformation" (HGT). Only nine cases are described in literature. Case description: Here, we describe the case of a 76-year-old male patient with a PAC recurrence of the oral floor displaying HGT, and we explore the tumor cytomorphological features, genomic profiling, and the patient's clinical management. The tumor mass was characterized by poorly atypical cellular elements with vesicular nuclei and comedonecrosis foci. The growth pattern was predominantly solid, tubular, and cribriform. The lesion did not show microsatellite instability or targeted molecular alterations. The case was successfully treated with radical surgery followed by radiotherapy. Conclusion: We report for the first time the recurrence of a PAC with HGT arising in the oral floor after 20 years from the primary lesion. These preliminary data and the literature analysis enhance the knowledge of this extremely rare disease.

Dynamic Monitoring of Circulating Tumor DNA in Patients With Metastatic Colorectal Cancer.

PURPOSE: Plasma circulating tumor DNA (ctDNA) is a valuable resource for tumor characterization and for monitoring of residual disease during treatment; however, it is not yet introduced in metastatic colorectal cancer (mCRC) routine clinical practice. In this retrospective exploratory study, we evaluated the role of ctDNA in patients with mCRC treated with chemotherapy plus bevacizumab. MATERIALS AND METHODS: Fifty-three patients were characterized for RAS and BRAF status on tumor tissue before the start of treatment. Plasma was collected at baseline, at first clinical evaluation, and at disease progression. ctDNA analysis was performed using Oncomine Colon cfDNA Assay on the Ion S5 XL instrument. RESULTS: At baseline, from a plasma sample, RAS, BRAF, or PIK3CA mutations were detected in 44 patients. A high correspondence was observed between ctDNA and tumor tissue mutations (KRAS 100%, NRAS 97.9%, BRAF 97.9%, PIK3CA 90%). Low baseline variant allele frequency (VAF) was found to be associated with longer median progression-free survival (PFS) compared with those with high VAF (15.9 v 12.2 months, P = .02). A higher PFS {12.29 months (95% CI, 9.03 to 17.9) v 8.15 months (95% CI, 2.76 to not available [NA]), P = .04} and overall survival (34.1 months [95% CI, 21.68 to NA] v 11.1 months [95% CI, 3.71 to NA], P = .003) were observed in patients with large decline in VAF at first evaluation. CONCLUSION: ctDNA analysis is useful for molecular characterization and tumor response monitoring in patients with mCRC. Quantitative variations of released ctDNA are associated with clinical outcomes.

Micrometeorological Methods for the Indirect Estimation of Odorous Emissions.

Odors are typically released into the atmosphere as diffuse emissions from area and volume sources, whose detailed quantification in terms of odor emission rate is often hardly achievable by direct source sampling. Indirect methods, involving the use of micrometeorological methods in order to correlate downwind concentrations to the emission rates, are already mentioned in literature, but rarely found in real applications for the quantification of odor emissions. The instrumentation needed for the development of micrometeorological methods has nowadays become accessible in terms of prices and reliability, thus making the implementation of such methods to industrial applications more and more interesting. For this reason, this work aims to provide an overview of micrometeorological methods and investigate their effective applicability to odors, thereby providing a short description of the physics related to such methods and analyzing the relevant scientific literature. The theoretical basis of these methods is presented, and their advantages and disadvantages are discussed. Moreover, their applicability to the estimation of odor emissions is discussed by providing some suggestions about the suitable ways to evaluate the most critical parameters needed for the calculation of the odor emission rate.

Accuracy of a new electronic nose for prostate cancer diagnosis in urine samples.

OBJECTIVE: To evaluate the accuracy of a new electronic nose to recognize prostate cancer in urine samples. METHODS: A blind, prospective study on consecutive patients was designed. Overall, 174 subjects were included in the study: 88 (50.6%) in prostate cancer group, and 86 (49.4%) in control group. Electronic nose performance for prostate cancer was assessed using sensitivity and specificity. The diagnostic accuracy of electronic nose was reported as area under the receiver operating characteristic curve. RESULTS: The electronic nose in the study population reached a sensitivity 85.2% (95% confidence interval 76.1-91.9; 13 false negatives out of 88), a specificity 79.1% (95% confidence interval 69.0-87.1; 18 false positives out of 86). The accuracy of the electronic nose represented as area under the receiver operating characteristic curve 0.821 (95% confidence interval 0.764-0.879). CONCLUSIONS: The diagnostic accuracy of electronic nose for recognizing prostate cancer in urine samples is high, promising and susceptible to supplemental improvement. Additionally, further studies will be necessary to design a clinical trial to validate electronic nose application in diagnostic prostate cancer nomograms.

An Experimental Apparatus for E-Nose Breath Analysis in Respiratory Failure Patients.

Background: Non-invasive, bedside diagnostic tools are extremely important for tailo ring the management of respiratory failure patients. The use of electronic noses (ENs) for exhaled breath analysis has the potential to provide useful information for phenotyping different respiratory disorders and improving diagnosis, but their application in respiratory failure patients remains a challenge. We developed a novel measurement apparatus for analysing exhaled breath in such patients. Methods: The breath sampling apparatus uses hospital medical air and oxygen pipeline systems to control the fraction of inspired oxygen and prevent contamination of exhaled gas from ambient Volatile Organic Compounds (VOCs) It is designed to minimise the dead space and respiratory load imposed on patients. Breath odour fingerprints were assessed using a commercial EN with custom MOX sensors. We carried out a feasibility study on 33 SARS-CoV-2 patients (25 with respiratory failure and 8 asymptomatic) and 22 controls to gather data on tolerability and for a preliminary assessment of sensitivity and specificity. The most significant features for the discrimination between breath-odour fingerprints from respiratory failure patients and controls were identified using the Boruta algorithm and then implemented in the development of a support vector machine (SVM) classification model. Results: The novel sampling system was well-tolerated by all patients. The SVM differentiated between respiratory failure patients and controls with an accuracy of 0.81 (area under the ROC curve) and a sensitivity and specificity of 0.920 and 0.682, respectively. The selected features were significantly different in SARS-CoV-2 patients with respiratory failure versus controls and asymptomatic SARS-CoV-2 patients (p < 0.001 and 0.046, respectively). Conclusions: the developed system is suitable for the collection of exhaled breath samples from respiratory failure patients. Our preliminary results suggest that breath-odour fingerprints may be sensitive markers of lung disease severity and aetiology.

Endometrioid Cancer Associated With Endometriosis: From the Seed and Soil Theory to Clinical Practice.

Endometriosis is a benign condition characterized by the presence of ectopic endometrial tissue. It is still debated whether endometriosis is a disease that can predispose to the pathogenesis of endometrial cancer outside the uterus. Deficiencies in mismatch repair (MMR) genes are a known risk factor for developing endometrioid cancer. Starting from two cases of patients with abnormal MMR endometrioid carcinoma of the uterus and synchronous endometrioid carcinoma in non-ovarian and ovarian endometriosis, we performed a somatic mutation profile and phylogenetic analysis of the lesions in order to identify if they were metastasis or primary de novo tumors. In the first case, we identified de novo activating mutations in PIK3CA and KRAS in endometrioid cancer lesions but not in endometriosis. Although the acquisition of a de novo mutation in ESR1 and a decrease in mutant allele fraction (MAF) from the endometrial tumor to the localizations in the endometriosis lesions, the clonal relationship was confirmed by the limited number of heteroplasmic mutations in D-loop mitochondrial DNA region. In the other case, the clonal behavior was demonstrated by the overlap of MAF at each site. Our data support the hypothesis of a retrograde dissemination of tumor cells, moving from the primary carcinoma in the endometrium to ectopic sites of endometriosis where localizations of tumor arise.

The role of next-generation sequencing in detecting gene fusions with known and unknown partners: a single-center experience with methodologies' integration.

AIMS: Next-generation sequencing (NGS) is becoming a new gold standard for determining molecular predictive biomarkers. This study aimed to evaluate the reliability of NGS in detecting gene fusions, focusing on comparing gene fusions with known and unknown partners. METHODS: We collected all gene fusions from a consecutive case series using an amplicon-based DNA/RNA NGS platform and subdivided them into two groups: gene fusions with known partners and gene fusions with unknown partners. Gene fusions involving ALK, ROS1 and RET were also examined by immunohistochemistry (IHC) and/or fluorescent in situ hybridization (FISH). RESULTS: Overall, 1174 malignancies underwent NGS analysis. NGS detected gene fusions in 67 cases (5.7%), further subdivided into 43 (64.2%) with known partners and 24 (35.8%) with unknown partners. Gene fusions were predominantly found in non-small cell lung carcinomas (52/67, 77.6%). Gene fusions with known partners frequently involved ALK (20/43, 46.5%) and MET (9/43, 20.9%), while gene fusions with unknown partners mostly involved RET (18/24, 75.0%). FISH/IHC confirmed rearrangement status in most (89.3%) of the gene fusions with known partners, but in only one (4.8%) of the gene fusions with unknown partners, with a significant difference (p < 0.001). In 17 patients undergoing targeted therapy, the log-rank test revealed that the overall survival was higher in the known partner group than in the unknown partner group (p = 0.002). CONCLUSIONS: NGS is a reliable method for detecting gene fusions with known partners, but it is less accurate in identifying gene fusions with unknown partners, for which further analyses (such as FISH) are required.

Drift compensation on electronic nose data for non-invasive diagnosis of prostate cancer by urine analysis.

Diagnostic protocol for prostate cancer (KP) is affected by poor accuracy and high false-positive rate. The most promising innovative approach is based on urine analysis by electronic noses (ENs), highlighting a specific correlation between urine alteration and KP presence. Although EN could be exploited to develop non-invasive KP diagnostic tools, no study has already introduced EN into clinical practice, most probably because of drift issues that hinder EN scaling up from research objects to large-scale diagnostic devices. This study, proposing an EN for non-invasive KP detection, describes the data processing protocol applied to a urine headspace dataset acquired over 9 months, comprising 81 patients with KP and 41 controls, for compensating the drift. It proved effective in mitigating drift on 1-year-old sensors by restoring accuracy from 55% up to 80%, achieved by new sensors not subjected to drift. The model achieved, on double-blind validation, a balanced accuracy of 76.2% (CI95% 51.9-92.3).

Optimization of training and measurement protocol for eNose analysis of urine headspace aimed at prostate cancer diagnosis.

More than one million new cases of prostate cancer (PCa) were reported worldwide in 2020, and a significant increase of PCa incidence up to 2040 is estimated. Despite potential treatability in early stages, PCa diagnosis is challenging because of late symptoms' onset and limits of current screening procedures. It has been now accepted that cell transformation leads to release of volatile organic compounds in biologic fluids, including urine. Thus, several studies proposed the possibility to develop new diagnostic tools based on urine analysis. Among these, electronic noses (eNoses) represent one of the most promising devices, because of their potential to provide a non-invasive diagnosis. Here we describe the approach aimed at defining the experimental protocol for eNose application for PCa diagnosis. Our research investigates effects of sample preparation and analysis on eNose responses and repeatability. The dependence of eNose diagnostic performance on urine portion analysed, techniques involved for extracting urine volatiles and conditioning temperature were analysed. 192 subjects (132 PCa patients and 60 controls) were involved. The developed experimental protocol has resulted in accuracy, sensitivity and specificity of 83% (CI95% 77-89), 82% (CI95% 73-88) and 87% (CI95% 75-94), respectively. Our findings define eNoses as valuable diagnostic tool allowing rapid and non-invasive PCa diagnosis.

Liquid Biopsy for EGFR Mutation Analysis in Advanced Non-Small-Cell Lung Cancer Patients: Thoughts Drawn from a Real-Life Experience.

BACKGROUND: Liquid biopsy analysis for EGFR detection in cell-free DNA (cfDNA) from NSCLC patients has become routine. The aim of this study was to explore its applicability in clinical practice. METHODS: We collected data of EGFR-mutated NSCLC patients with liquid biopsy analysis. Data included test timing, concomitant tissue re-biopsy, therapy change, histology, stage, smoking habits, gender and age. All analyses were performed via a real-time PCR method to analyze EGFR mutations at exons 18, 19, 20 and 21. Variant allele frequency was performed for patients with available sequential EGFR mutation analysis in cfDNA. Overall survival was analyzed through the Kaplan-Meier method. We designed flow charts to show the real-life application of liquid biopsy. RESULTS: We found that liquid biopsy is used in treatment-naïve patients as an alternative to EGFR detection in tumor tissue, and in patients with positive or negative EGFR from tumor biopsy. The majority of liquid biopsy analyses were performed in NSCLC patients who were disease progressive during TKI therapy. The presence of EGFR mutation in cfDNA was associated with a worse prognosis. In two patients, VAF of EGFR mutations in cfDNA was concordant with tumor volume changes. CONCLUSION: These findings suggest that liquid biopsy for EGFR detection can continue to be useful.

Simulating Pollutant Dispersion from Accidental Fires with a Focus on Source Characterization.

BACKGROUND: Storage tanks in oil and gas processing facilities contain large volumes of flammable compounds. Once the fuel-air mixture is ignited, it may break out into a large fire or explosion. The growing interest in monitoring air quality and assessing health risks makes the evaluation of the consequences of a fire an important issue. Atmospheric dispersion models, which allow for simulation of the spatial distribution of pollutants, represent an increasingly widespread tool for this type of evaluations. OBJECTIVES: The present study discusses the set up and results of a modeling study relevant to a hypothesized fire in an oil refinery. METHODS: After choosing the most suitable dispersion models, i.e. the Lagrangian model SPRAY and the puff model CALPUFF, estimation of the required input data is discussed, focusing on the source variables, which represent the most uncertain input data. The results of the simulations were compared to regulatory limits to effectively evaluate the environmental consequences. Finally, a sensitivity analysis was employed to identify the most influential variables. RESULTS: The simulation results revealed that ground concentration values were far below the cited long-term limits. However, the most interesting outcome is that depending on the dispersion model and the source type modeled, different results may be obtained. In addition, the sensitivity study indicates that the source area is the most critical variable, since it determines a significantly different behavior depending on the modeled source types, producing, in some cases, variability in the pollutant ground concentrations on selected receptors up to +/- 60%. CONCLUSIONS: Depending on the selected model and the algorithms available to describe the physics of emission, the results showed a different sensitivity to the input variables. Although this can be explained from a mathematical point of view, the problem remains of choosing case by case the option that best approximates the real behavior of the incidental source under investigation. COMPETING INTERESTS: The authors declare no competing financial interests.

Non-carcinogenic occupational exposure risk related to foundry emissions: focus on the workers involved in olfactometric assessments.

The scope of this work is the evaluation of the non-carcinogenic occupational risk related to foundry emissions, focusing on the category of workers involved in olfactometric assessments. Odor pollution from industrial activities such as foundries is a serious environmental concern. Sensorial techniques (e.g. dynamic olfactometry, EN13725:2003) currently represent the preferred method for odor emission characterization. During olfactometric analyses, human assessors are directly exposed to the odor at increasing concentrations, thus requiring the assessment of the associated exposure risk to guarantee workers' safety. This paper presents an investigation aiming to produce an inventory of compounds emitted from foundries together with their odor thresholds and toxicological limits (TLVs), with the final objective to propose a procedure for ensuring workers' safety during olfactometric analyses. Looking at the database resulting from this study, among the >100 compounds emitted by foundries, 8 have a maximum concentration above their TLV. Among those, ammonia, H2S, phenol, toluene and trimethylamine, produce an odor stimulus before they reach a toxic concentration, thus not representing a risk for olfactometric workers. Benzene, formaldehyde and SO2 are identified as the most critical compounds because they may reach toxic concentrations in foundry emissions, and they start being perceived by humans above their TLV. The proposed procedure entails a minimum dilution factor of 27'000 to be applied to odor samples analyzed by olfactometry, which however might result inapplicable in practical cases, thus pointing out the necessity to adopt chemical measurements to investigate specifically the concentration of the most critical compounds identified in this study.

Early Gastric Cancer: identification of molecular markers able to distinguish submucosa-penetrating lesions with different prognosis.

BACKGROUND: Early Gastric Cancer (EGC) reaches 25% of the gastric cancers surgically treated in some areas of Northeastern Italy and is usually characterized by a good prognosis. However, among EGCs classified according to Kodama's criteria, Pen A subgroup is characterized by extensive submucosal invasion, lymph node metastases and worse prognosis, whereas Pen B subgroup by better prognosis. The aim of the study was to characterize the differences between Pen A, Pen B and locally advanced gastric cancer (T3N0) in order to identify biomarkers involved in aggressiveness and clinical outcome. METHODS: We selected 33 Pen A, 34 Pen B and 20 T3N0 tumors and performed immunohistochemistry of mucins, copy number variation analysis of a gene panel, microsatellite instability (MSI), TP53 mutation and loss of heterozygosity (LOH) analyses. RESULTS: Pen A subgroup was characterized by MUC6 overexpression (p = 0.021). Otherwise, the Pen B subgroup was significantly associated with the amplification of GATA6 gene (p = 0.002). The higher percentage of MSI tumors was observed in T3N0 group (p = 0.002), but no significant differences between EGC types were found. Finally, TP53 gene analysis showed that 32.8% of Pen tumors have a mutation in exons 5-8 and 50.0% presented LOH. Co-occurrence of TP53 mutation and LOH mainly characterized Pen A tumors (p = 0.022). CONCLUSIONS: Our analyses revealed that clinico-pathological parameters, microsatellite status and frequency of TP53 mutations do not seem to distinguish Pen subgroups. Conversely, the amplification of GATA6 was associated with Pen B, as well as the overexpression of MUC6 and the TP53mut/LOH significantly characterized Pen A.

Kevetrin induces apoptosis in TP53 wild­type and mutant acute myeloid leukemia cells.

Tumor protein p53 is a key regulator of several cellular pathways, including DNA repair, cell cycle and angiogenesis. Kevetrin exhibits p53­dependent as well as­independent activity in solid tumors, while its effects on leukemic cells remain unknown. The aim of the present study was to analyze the response of acute myeloid leukemia (AML) cell lines (TP53 wild­type: OCI­AML3 and MOLM­13; and TP53­mutant: KASUMI­1 and NOMO­1) to kevetrin at a concentration range of 85­340 µM. The cellular and molecular effects of the treatment were analyzed in terms of cell growth, viability [Annexin V­propidium iodide (PI) staining] and cell cycle alterations (PI staining). Gene expression profiling, western blotting and immunofluorescence were performed to elucidate the pathways underlying kevetrin activity. Pulsed exposure exerted no effect on the wild­type cells, but was effective on mutant cells. After continuous treatment, significant cell growth arrest and apoptosis were observed in all cell lines, with TP53­mutant models displaying a higher sensitivity and p53 induction. Kevetrin also displayed efficacy against TP53 wild­type and mutant primary AML, with a preferential cytotoxic activity against blast cells. Gene expression profiling revealed a common core transcriptional program altered by drug exposure and the downregulation of glycolysis, DNA repair and unfolded protein response signatures. These findings suggest that kevetrin may be a promising therapeutic option for patients with both wild­type and TP53­mutant AML.