The Impact of potential 'confounders' on the diagnostic sensitivity of circulating free DNA in management of FIT+ patients: a pilot study.

Cell-free DNA (cfDNA) has long been established as a useful diagnostic and prognostic tool in a variety of clinical settings, ranging from infectious to cardiovascular and neoplastic diseases. However, non-neoplastic diseases can act as confounders impacting on the amount of cfDNA shed in bloodstream and on technical feasibility of tumour derived free circulating nucleic acids selecting patients with cancer. Here, we investigated the potential impact of other pathological processes in the clinical stratification of 637 FIT+ patients. A single and multiple logistic regression yielded similar results. Crude sensitivity was 75.9% versus adjusted sensitivity of 74.1%, relative risk 0.9761 (0.8516 to 1.1188), risk difference 0.0181 (-0.0835 to 0.1199) and OR 0.9079 (0.5264 to 1.5658). Potential confounding effect from other source of cfDNA plays a pivotal role in the clinical stratification of FIT+ patients.

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.

Liquid biopsy: A promising tool for driving strategies and predicting failures in patients with classic Hodgkin lymphoma.

Classic Hodgkin lymphoma (cHL) consists of a heterogeneous group of haematological disorders that covers undifferentiated B cell neoplasms originating from germinal centre B cells. The HL molecular characterization still represents an ongoing challenge due to the low fraction of tumour Hodgkin and Reed-Sternberg cells mixed with a plethora of non-tumour haematological cells. In this scenario, next generation sequencing of liquid biopsy samples is emerging as a useful tool in HL patients' management. In this review, we aimed to overview the clinical and methodological topics regarding the implementation of molecular analysis in cHL, focusing on the role of liquid biopsy in diagnosis, follow-up, and response prediction.

Novel insights into the biomolecular mechanism of action of 4'-geranyloxyferulic acid, a colon cancer chemopreventive agent.

In this manuscript the biomolecular mechanism of action of the natural colon cancer chemopreventive agent 4'-geranyloxyferulic acid in cultured Caco-2 cells has been investigated. It was first demonstrated how the application of this phytochemical led to a time- and dose-dependent decrease of cell viability and in parallel to a massive generation of reactive oxygen species and induction of caspases 3 and 9, finally providing apoptosis. This event is accompanied by deep modifications in key pro-apoptotic targets like CD95, DR4 and 5, cytochrome c, Apaf-1, Bcl-2, and Bax. Such effects can explain the large apoptosis recorded in Caco-2 cells treated with 4'-geranyloxyferulic acid.

Protection of Mitochondrial Potential and Activity by Oxyprenylated Phenylpropanoids.

A series of five naturally occurring oxyprenylated phenylpropanoids, namely, the coumarins auraptene (7-geranyloxycoumarin) 1 and 7-isopentenyloxycoumarin 2, and the coumaric acid and ferulic acid derivatives, 4'-isopentenyloxycoumaric acid 3, boropinic acid 4, and 4'-geranyloxyferulic acid 5 were tested for their effects on mitochondrial functionality using the organophosphate pesticides glyphosate and chlorpyrifos, and resveratrol, as the reference. While not showing an appreciable in vitro antioxidant activity, and virtually no or a little effect on the viability of non-cancer cell lines BEAS-2B and SHSY-5Y, all phytochemicals exhibited a marked protective effect on mitochondrial potential and activity, with values that were comparable to resveratrol. Auraptene 1 and 7-isopentenyloxycoumarin 2 were seen to be the most effective secondary metabolite to this concern, in particular in being able to completely abolish the decrease of mitochondrial potential induced by increasing concentration of both glyphosate and chlorpyrifos. All the compounds tested also exhibited a protective effect on mitochondrial activity. The potency displayed will shed more light on the molecular basis of the beneficial effects of auraptene, 7-isopentenyloxycoumarin, and structurally related oxyprenylated phenylpropanoids reported to date in the literature.

Microsatellite instability evaluation of patients with solid tumour: routine practice insight from a large series of Italian referral centre.

DNA mismatch repair complex is involved in the maintenance of DNA stability. In the recent years, a plethora of technical approaches for microsatellite instability (MSI) analysis emerged. Here, we review the results of our MSI status evaluation by adopting a customised workflow on microfluidic system obtained in 4 years of diagnostic routine practice. Data from MSI status were retrieved from our institutional archive covering the period from January 2017 to December 2021. Microfluidic analysis was carried out on microfluidic platform. Results were inspected with a proprietary software. Overall, microsatellite stability (MSS) and MSI-high (MSI-H) profile was detected in n=423/458 (92.36%) and n=35/458 (7.64%) patients with metastatic CRC (mCRC), respectively. In addition, n=78/86 (90.70%) and n=8/86 (9.30%) patients without CRC showed an MSS and MSI-H profile. This review highlights the suitability of microfluidic approach in patients with cancer for MSI testing.

A novel and efficient concentration of pomegranate juice with enhanced antioxidant activity.

In the present study we investigated the performance of a panel of 13 solid sorbents comprising layered double hydroxides, zirconium phosphate-based materials, and phyllosilicates as heterogeneous supports for the concentration of pomegranate (Punica granatum L.) juice. Mg-containing clays exhibited an almost complete bleaching capacityof pomegranate juice and more interestingly provided blends with an increased antioxidant capacity (around 1.5-fold) respect to the parent juice when assayed for the Trolox equivalent antioxidant capacity (TEAC) coupled to ABTS decolorization test. Such an activity remained practically unaltered after 4 days during which the pomegranate concentrated preparations remained supported on clays. The approach investigated herein and used for the concentration of pomegranate juice and the discovery of the preservation for long periods of the antioxidant activities of pomegranate extracts when supported on solid sorbents have been reported herein for the first time in the literature to the best of our knowledge.

A revised version of the Iwaoka's assay: Application of hyphenated techniques.

Assessing the effects of glutathione peroxidase (GPx)-like catalytic activities of natural, semi-synthetic, and synthetic compounds is nowadays well recognized of importance for the preliminary screening of potential anti-inflammatory, neuroprotective, and anti-cancer agents. To this aim the Iwaoka's assay, relying on 1H NMR data recording, has been the most exploited test. In this short communication we propose an efficient and easy to perform methodology to accomplish the same process based on the application of hyphenated techniques like gas-chromatography coupled to mass spectrometry (GC-MS) and high-performance liquid chromatography with diode array detection (HPLC-DAD). The assay consisted in monitoring and quantifying the oxidation at different times of 1,4-dithiothreitol to 4,5-dihydroxy-1,2-dithiane in the presence of H2O2 and 2-phenyl-1,2-benzoselenazol-3(2H)-one (Ebselen) as the catalyst in EtOAc or MeOH as the solvents. The results we recorded by the application of both GC-MS and HPLC-DAD, showed that the GPx-like activity of Ebselen, used as the reference compound, can be effectively monitored in a time-course manner, and compare favorably to the NMR-based test.

A Novel Auraptene-Enriched Citrus Peels-Based Blend with Enhanced Antioxidant Activity.

The aim of this study was to establish a convenient process to get auraptene-enriched blends from Citrus limon L. Osbeck (Rutaceae) peels powder with functional food and nutraceutical potential and antioxidant properties. The process consisted in the selective adsorption of auraptene by bentonite followed by desorption and evaporation of solvents to get the final product. HPLC analyses revealed an adsorption > 99% from lemon fruit peel powder and a 13.7-fold enrichment of the obtained composition respect to the parent food-derived sample. The antioxidant and radical scavenger activities of the Citrus peels based preparation were assayed by the DPPH and ABTS tests and reveal deffects higher than ascorbic acid and Trolox® used as the references. To the best of our knowledge, the findings described herein is the first example reported in the literature of solid phase adsorption experiments carried out on auraptene as a component of edible fruits.

7-Isopentenyloxycoumarin: What Is New across the Last Decade.

7-Isopentenyloxycoumarin is among the most widespread naturally occurring prenyloxy umbelliferone derivatives. This secondary metabolite of mixed biosynthetic origin has been typically isolated from plants belonging to several genera of the Rutaceae and Apiaceae families, comprising widely used medicinal plants and in general plants with beneficial effects on human welfare, as well as edible fruits and vegetables. Although known for quite a long time (more than 50 years), only in the last two decades has this natural compound been revealed to exert powerful and promising pharmacological properties, such as active cancer chemopreventive, antibacterial, antiprotozoal, antifungal, anti-inflammatory, neuroprotective, and antioxidant properties, among the activities best outlined in the recent literature. The aim of this comprehensive miniature review article is to detail the novel natural sources and the effects described during the last decade for 7-isopentenyloxycoumarin and what has been reported on the mechanisms of action underlying the observed biological activities of this oxyprenylated secondary metabolite. In view of the herein described data, suggestions on how to address future research on the abovementioned natural product and structurally related derivatives in the best ways according to the authors will be also provided.