There is an unmet need for simplified in vitro models of malignancy and metastasis that facilitate fast, affordable and scalable gene and compound analysis. “Adherent” cancer cell lines frequently release “free-floating” cells into suspension that are viable and can reattach. This, in a simplistic way, mimics the metastatic process. We compared the gene expression profiles of naturally co-existing populations of floating and adherent cells in SW620 (colon), C33a (cervix) and HeLa (cervix) cancer cells. We found that 1227, 1367 and 1333 genes were at least 2-fold differentially expressed in the respective cell lines, of which 122 were shared among the three cell lines. As proof of principle, we focused on the anti-metastatic gene NM23-H1, which was downregulated both at the RNA and protein level in the floating cell populations of all three cell lines. Knockdown of NM23-H1 significantly increased the number of floating (and viable) cells, whereas overexpression of NM23-H1 significantly reduced the proportion of floating cells. Other potential regulators of these cellular states were identified through pathway analysis, including hypoxia, mTOR (mechanistic target of rapamycin), cell adhesion and cell polarity signal transduction pathways. Hypoxia, a condition linked to malignancy and metastasis, reduced NM23-H1 expression and significantly increased the number of free-floating cells. Inhibition of mTOR or Rho-associated protein kinase (ROCK) significantly increased cell death specifically in the floating and not the adherent cell population. In conclusion, our study suggests that dynamic subpopulations of free-floating and adherent cells is a useful model to screen and identify genes, drugs and pathways that regulate the process of cancer metastasis, such as cell detachment and anoikis.

Immunotherapy has improved survival in patients with advanced hepatocellular carcinoma (HCC); yet, objective radiological responses occur in only about 20% of cases, suggesting variable benefits. This study aimed to identify serologic markers predictive of response to immune checkpoint inhibitors (ICIs). A cohort of 38 advanced HCC patients receiving immunotherapy was prospectively analyzed. Levels of cell-free DNA (cfDNA), circulating tumor DNA (ctDNA), and cytokines were measured pre-treatment and three months post-treatment initiation. Genomic profiling of ctDNA was also conducted. Baseline levels of cfDNA and ctDNA effectively discriminated HCC patients based on their radiological response to ICIs. Additionally, individuals with pathologic mutations in the CDKN2A gene exhibited significantly reduced survival. Patients with progressive disease (PD) as their best radiological response had significantly fewer copy number variations (CNVs) than those with a radiological response. Furthermore, levels of IL10, PD1, and TGFβ assessed after three months of treatment showed significant variations correlating with survival status. In conclusion, the analysis of cfDNA, ctDNA, and cytokines may improve treatment selection for HCC patients by predicting their expected response to immunotherapies.

Summary: Background: The WHO has set a goal to decrease viral hepatitis‐related fatalities by 65% by 2030.Aims: To locate and retrieve to care all individuals diagnosed with hepatitis B, D or C, and investigate why they were not linked to appropriate medical management.Methods: We conducted a retrospective‐prospective search for patients with hepatitis B, D or C virus (HBV, HDV and HCV) infection in the central laboratory database of the Barcelona northern health area (catchment population, 450,000). We reviewed records and contacted candidates for retrieval who were offered a specialist medical visit.Results: We reviewed records of 3731 patients with viral hepatitis (January 2019–December 2022): 1763 HBsAg+, 69 anti‐HDV+ and 1899 HCV‐RNA+. Among these, 644 (37%) HBV, 20 (29%) HDV and 1116 (56%) HCV patients were not currently linked to care. The proportion of patients receiving appropriate care was higher in HBV and HDV (p < 0.0001), and a higher percentage of unlinked hepatitis C patients had low life expectancy/comorbidities (39%; p < 0.0001). After implementing the linkage strategy, 254 HBV, 16 HDV and 54 HCV patients were successfully reintegrated into care. Among 1780 patients requiring linkage, 638 (35.8%) had moved to another health area or were missing essential contact data.Conclusions: Among patients with viral hepatitis who required appropriate specialist care and were eligible for contact, 64% with HBV, 100% with HDV and 27% with HCV were successfully reintegrated into care. Overall, 324 (47.2%) eligible patients were linked to care, indicating the success of this strategy.

Sarcopenia is associated with poor outcomes in patients with cirrhosis. While computed tomography (CT)-derived skeletal muscle index (SMI) is considered the gold standard for diagnosis in this setting, its use is limited due to cost, radiation exposure, and availability. Our aim was to evaluate whether simple, non-radiating, easy-to-use methods, including ultrasound (US), bioelectrical impedance analysis (BIA), and anthropometry, could accurately identify sarcopenia in this population. A total of 250 patients with cirrhosis who underwent an abdominal CT scan were enrolled prospectively in this single-center, cross-sectional study. Within 1 month, the patients underwent assessments including anthropometry, handgrip strength, BIA [including body protein index (BPI)], and US of the psoas and thigh. Sarcopenia was defined by CT-based SMI at L3 (<50 cm ² /m ² in men, <39 cm ² /m ² in women). Thigh and psoas muscle thickness (normalized by height ² ) and BPI showed moderate to strong correlations with SMI (ρ=0.42–0.71). Multivariable models combining thigh or psoas US with BMI and calf circumference achieved AUROCs of 0.837 and 0.822, respectively. A model using BPI and BMI showed comparable accuracy (AUROC, 0.844). Interobserver agreement for US measurements was high (Lin’s coefficients 0.66–0.93). Thigh stiffness (2D-SWE) and phase angle did not correlate with SMI. Non-radiating tools, such as US-based thigh and psoas measurements, BIA-derived BPI, and anthropometry, are accurate alternatives to CT for detecting sarcopenia in patients with cirrhosis. These tools are simple, safe, and reproducible, which supports their integration into routine clinical practice for sarcopenia screening and monitoring.

Background: Cell-free DNA (cfDNA) concentrations have been described to be inversely correlated with prognosis in cancer. Mutations in HCC-associated driver genes in cfDNA have been reported, but their relation with patient’s outcome has not been described. Our aim was to elucidate whether mutations found in cfDNA could be representative from those present in HCC tissue, providing the rationale to use the cfDNA to monitor HCC. Methods: Tumoral tissue, paired nontumor adjacent tissue and blood samples were collected from 30 HCC patients undergoing curative therapies. Deep sequencing targeting HCC driver genes was performed. Results: Patients with more than 2 ng/µL of cfDNA at diagnosis had higher mortality (mean OS 24.6 vs. 31.87 months, p = 0.01) (AUC = 0.782). Subjects who died during follow-up, had a significantly higher number of mutated genes (p = 0.015) and number of mutations (p = 0.015) on cfDNA. Number of mutated genes (p = 0.001), detected mutations (p = 0.001) in cfDNA and ratio (number of mutations/cfDNA) (p = 0.003) were significantly associated with recurrence. However, patients with a ratio (number of mutations/cfDNA) above 6 (long-rank p = 0.0003) presented a higher risk of recurrence than those with a ratio under 6. Detection of more than four mutations in cfDNA correlated with higher risk of death (long-rank p = 0.042). Conclusions: In summary, cfDNA and detection of prevalent HCC mutations could have prognostic implications in early-stage HCC patients