A single-cell metabolism-based assay for circulating tumour cell enumeration and clinical validation in metastatic breast cancer patients

Breast cancer (BC) is the most common cancer among women worldwide, and it ranks as the second leading cause of female cancer-related death. To improve the clinical outcome and survival of BC patients, early diagnosis, tailored treatment and monitoring of response are critical factors. In the last decades, several studies have reported that circulating tumour cells (CTCs) could meet all these criteria. CTCs in the peripheral blood of metastatic cancer patients are associated with overall survival and treatment outcomes. A hallmark of many cancers is an altered glucose metabolism, which leads to the acidification of the tumour microenvironment. Our aim was to evaluate a method for detecting CTCs that exploits the abnormal metabolic behaviour of cancer cells in patients with metastatic breast cancer (MBC). This assay exploits a droplet microfluidic technology, which allows to compartmentalize single-cell into a droplet and detect metabolically active cells by pH measurements of the extracellular space. Using breast cancer cell lines with different metastatic potential and normal blood cells, we established a functional cut-off, i.e.: the pH value, for discriminating CTCs from white blood cells in clinical samples. We assessed the potential of enumerating metabolically active CTCs in a cohort of MBCs and healthy donor volunteers and we compared our method to the gold standard CellSearch®. The number of detected metabolically active CTCs was significantly higher than metabolically active cells in healthy donors. Interestingly, our method was able to predict both overall and progression free survival similarly to what observed with the CellSearch, although the concordance among the two methods was not high. However, the comparison with the golden standard for CTC enumeration suggested that the two methods recognize partially overlapping populations, suggesting the combined use of both methods to better predict the patient outcome

Radiation recall dermatitis induced by COVID-19 vaccination in breast cancer patients treated with postoperative radiation therapy

Background: and purpose: Radiation recall dermatitis is an adverse event predominantly due to systemic therapy administration after a previous radiation therapy course. Few case reports describe radiation recall dermatitis in breast cancer patients treated with postoperative radiation therapy following COVID-19 vaccination. In this study we investigated the incidence and severity of radiation recall dermatitis after COVID-19 vaccination in irradiated breast cancer patients. Methods: Patients that received at least one COVID-19 vaccination dose during the year after the end of postoperative breast radiation therapy were included in this observational monocentric study. Local symptoms occurring inside the radiation field after vaccination were patient-reported and scored according to the PRO-CTCAE questionnaire. Descriptive data of radiation recall dermatitis incidence and severity, and potential risk factors were evaluated. Results: A cohort of 361 patients with 756 administered COVID-19 vaccinations was analyzed. Breast symptoms were reported by 7.5% of patients, while radiation recall dermatitis was considered for 5.5%. The incidence of radiation recall dermatitis per single dose of vaccine was 2.6%, with a higher risk for the first dose compared to the second/third (4.4% vs 1%, p = 0.003), especially when administered within the first month after the end of irradiation (12.5% vs 2.2%, p = 0.0004). Local symptoms were generally self-limited and a few cases required anti-inflammatory drugs. Conclusions: Radiation recall dermatitis is an uncommon but not rare phenomenon in breast cancer patients that received COVID-19 vaccination within one year after breast irradiation. However, symptoms severity were generally low/mild and reversible. These findings can be useful for patient counseling

Assessment of the mutational status of NSCLC using hypermetabolic circulating tumor cells

Molecular characterization is currently a key step in NSCLC therapy selection. Circulating tumor cells (CTC) are excellent candidates for downstream analysis, but technology is still lagging behind. In this work, we show that the mutational status of NSCLC can be assessed on hypermetabolic CTC, detected by their increased glucose uptake. We validated the method in 30 Stage IV NSCLC patients: peripheral blood samples were incubated with a fluorescent glucose analog (2-NBDG) and analyzed by flow cytometry. Cells with the highest glucose uptake were sorted out. EGFR and KRAS mutations were detected by ddPCR. In sorted cells, mutated DNA was found in 85% of patients, finding an exact match with primary tumor in 70% of cases. Interestingly, in two patients multiple KRAS mutations were detected. Two patients displayed different mutations with respect to the primary tumor, and in two out of the four patients with a wild type primary tumor, new mutations were highlighted: EGFR p.746_750del and KRAS p.G12V. Hypermetabolic CTC can be enriched without the need of dedicated equipment and their mutational status can successfully be assessed by ddPCR. Finally, the finding of new mutations supports the possibility of probing tumor heterogeneity

Polymorphisms in Pepsinogen C and miRNA Genes Associate with High Serum Pepsinogen II in Gastric Cancer Patients

Background: Pepsinogen (PG) II (PGII) is a serological marker used to estimate the risk of gastric cancer but how PGII expression is regulated is largely unknown. It has been suggested that PGII expression, from the PGC (Progastricsin) gene, is regulated by microRNAs (miRNA), but how PGII levels vary with Helicobacter pylori (H. pylori) infection and miRNAs genotype remains unclear. Methods: Serum levels of PGI and PGII were determined in 80 patients with gastric cancer and persons at risk for gastric cancer (74 first-degree relatives of patients, 62 patients with autoimmune chronic atrophic gastritis, and 2 patients with dysplasia), with and without H. pylori infection. As control from the general population, 52 blood donors were added to the analyses. Associations between PGII levels and genetic variants in PGC and miRNA genes in these groups were explored based on H. pylori seropositivity and the risk for gastric cancer. The two-dimensional difference in gel electrophoresis (2D-DIGE) and the NanoString analysis of messenger RNA (mRNAs) from gastric cancer tissue were used to determine the pathways associated with increased PGII levels. Results: PGII levels were significantly higher in patients with gastric cancer, and in those with H. pylori infection, than in other patients or controls. A PGI/PGII ratio ≤ 3 was found better than PGI p = 0.02 and p = 0.01, respectively), but not among other study subjects. Moreover, a strict relation between rs9471643 C-allele with H. pylori infection and gastric cancer was underlined. Fold change in gene expression of mRNA isolated from gastric cancer tissue correlated well with polymorphism, H. pylori infection, increased PGII level, and pathway for bacteria cell entry into the host. Conclusions: Serum PGII levels depend in part on an interaction between H. pylori and host miRNA genotypes, which may interfere with the cut-off of PGI/PGII ratio used to identify persons at risk of gastric cancer. Results reported new findings regarding the relation among H. pylori, PGII-related host polymorphism, and genes involved in this interaction in the gastric cancer setting

Additional file 3: of An improved sequencing-based strategy to estimate locus-specific DNA methylation

Validation of the plasmid DNA standards. (PDF 277 kb

Dataset related to article "Biofeedback as an Adjunctive Treatment for Post-stroke Dysphagia: A Pilot-Randomized Controlled Trial"

Nel file Excel con i raw data sono presenti le caratteristiche demografiche del campione, e per ogni paziente se sia stato assegnato al trattamento sperimentale o di controllo (codificati come 1;0). Si trovano inoltre i punteggi ai vari test che il paziente ha ottenuto nei 3 momenti di valutazione (v1=baseline; v2=valutazione post trattamento; v3=follow up) e per le diverse consistenze provate (solido, semisolido, liquido). Le scale nominate nel dataset sono: PAS-Penetration Aspiration Scale); Pooling Score (suddiviso in P-score e PSCA); FOIS (Functional Oral Intake Scale). Si riporta inoltre la presenza o assenza di cannula tracheale e nutrizione enterale (1;0)