Anti-IFN-α/-ω neutralizing antibodies from COVID-19 patients correlate with downregulation of IFN response and laboratory biomarkers of disease severity.

A significant number of COVID-19 patients were shown to have neutralizing antibodies (NAB) against IFN; however, NAB specificity, fluctuation over time, associations with biochemical and hematological parameters, and IFN gene expression are not well characterized. Binding antibodies (BAB) to IFN-α/-ß were screened in COVID-19 patients' serum. All BAB positive sera, and a subset of respiratory samples, were tested for NAB against IFN-α/-ß/-ω, using an antiviral bioassay. Transcript levels of IFN-α/-ß/-ω and IFN-stimulated genes (ISGs) were quantified. Anti-IFN-I BAB were found in 61 out of 360 (17%) of patients. Among BAB positive sera, 21.3% had a high NAB titer against IFN-α. A total of 69.2% of anti-IFN-α NAB sera displayed cross-reactivity to IFN-ω. Anti-IFN-I NAB persisted in all patients. NAB to IFN-α were also detected in 3 out of 17 (17.6%) of respiratory samples. Anti-IFN-I NAB were higher in males (p = 0.0017), patients admitted to the ICU (p < 0.0001), and patients with a fatal outcome (p < 0.0001). NAB were associated with higher levels of CRP, LDH, d-Dimer, and higher counts of hematological parameters. ISG-mRNAs were reduced in patients with persistently NAB titer. NAB are detected in a significant proportion of severe COVID-19. NAB positive patients presented a defective IFN response and increased levels of laboratory biomarkers of disease severity.

Assessment of sST2 Behaviors to Evaluate Severity/Clinical Impact of Acute Pulmonary Embolism.

Pulmonary embolism (PE) is a potentially life-threatening disorder. Beyond its usefulness in the prognostic stratification of heart failure, sST2 can represent a biomarker with high utility in several acute conditions. Our study was aimed to investigate whether sST2 can be used as a clinical marker of severity and prognostic outcome in acute PE. We enrolled 72 patients with documented PE and 38 healthy subjects; we measured the plasma concentrations of sST2 to evaluate the prognostic and severity performance of different levels of sST2 according to its association with the pulmonary embolism severity index (PESI) score and several parameters of respiratory function. PE patients had significantly higher levels of sST2 compared with healthy subjects (87.74 ± 17.1 vs. 17.1 ± 0.4 ng/mL, p < 0.001); we found higher PESI scores and serum lactate values in the group of patients with sST2 > 35 ng/mL compared with patients with sST2 < 35 ng/mL (138.7 ± 14.9 vs. 103.7 ± 15.1 and 2.43 ± 0.69 vs. 1.025 ± 0.05 mmol/L, respectively; p < 0.05). Patients with sST2 > 35 ng/mL showed higher radiological severity of PE compared with patients with sST2 < 35 ng/mL. Moreover, sST2 was the strongest parameter with a discriminative capacity for the development of acute respiratory failure and a PESI score >106 with respect to C reactive protein (CRP), creatinine, d-dimer, and serum lactate. We clearly demonstrated that sST2 significantly increased in PE and that its elevation was associated with disease severity. Therefore, sST2 may be used as a clinical marker in the evaluation of PE severity. However, further studies with larger patient populations are required to confirm these findings.

Circulating tumor cells in high-risk nonmetastatic colorectal cancer.

The identification of patients at higher risk of recurrence after primary colorectal cancer resection is currently one of the challenges facing medical oncologists. Circulating tumor cell (CTC) may represent a surrogate marker of an early spread of disease in patients without overt metastases. Thirty-seven high-risk stages II-III colorectal cancer patients were evaluated for the presence of CTC. Enumeration of CTCs in 7.5 ml of blood was carried out with the FDA-cleared CellSearch system. CTC count was performed after primary tumor resection and before the start of adjuvant therapy. CTC was detected in 22 % of patients with a significant correlation with regional lymph nodes involvement and stage of disease. No significant correlation was found among the presence of CTC and other clinicopathological parameters. These data suggest that CTCs detection might help in the selection of high-risk stage II colorectal cancer patient candidates for adjuvant chemotherapy.

Obesity-Associated Hepatic Steatosis, Somatotropic Axis Impairment, and Ferritin Levels Are Strong Predictors of COVID-19 Severity.

The full spectrum of SARS-CoV-2-infected patients has not yet been defined. This study aimed to evaluate which parameters derived from CT, inflammatory, and hormonal markers could explain the clinical variability of COVID-19. We performed a retrospective study including SARS-CoV-2-infected patients hospitalized from March 2020 to May 2021 at the Umberto I Polyclinic of Rome. Patients were divided into four groups according to the degree of respiratory failure. Routine laboratory examinations, BMI, liver steatosis indices, liver CT attenuation, ferritin, and IGF-1 serum levels were assessed and correlated with severity. Analysis of variance between groups showed that patients with worse prognoses had higher BMI and ferritin levels, but lower liver density, albumin, GH, and IGF-1. ROC analysis confirmed the prognostic accuracy of IGF-1 in discriminating between patients who experienced death/severe respiratory failure and those who did not (AUC 0.688, CI: 0.587 to 0.789, p < 0.001). A multivariate analysis considering the degrees of severity of the disease as the dependent variable and ferritin, liver density, and the standard deviation score of IGF-1 as regressors showed that all three parameters were significant predictors. Ferritin, IGF-1, and liver steatosis account for the increased risk of poor prognosis in COVID-19 patients with obesity.

Interleukin 18 and IL-18 BP response to Sars-CoV-2 virus infection.

The immune response to the SARS-CoV-2 infection is crucial to the patient outcome. IL-18 is involved in the lymphocyte response to the disease and it is well established its important role in the complex developing of the host response to viral infection. This study aims at the analysis of the concentrations of IL-18, IL-18BP, INF-γ at the onset of the SARS-CoV-2 infection. The serum levels of measured interleukins were obtained through enzyme-linked immunosorbent assay. Furthermore, the free fraction of IL-18 was numerically evaluated. The enrolled patients were divided in two severity groups according to a threshold value of 300 for the ratio of arterial partial pressure of oxygen and fraction of inspired oxygen fraction and according to the parenchymal involvement as evaluated by computerized tomography at the admittance. In the group of patients with a more severe disease, a significant increase of the IL-18, INF-γ and IL-18BP levels have been observed, whereas the free IL-18 component values were almost constant. The results confirm that, at the onset of the disease, the host response keep the inflammatory cytokines in an equilibrium and support the hypothesis to adopt the IL-18BP modulation as a possible and effective therapeutic approach.

Prognostic Role of Circulating Tumor Cell Trajectories in Metastatic Colorectal Cancer.

BACKGROUND: A large amount of evidence from clinical studies has demonstrated that circulating tumor cells are strong predictors of outcomes in many cancers. However, the clinical significance of CTC enumeration in metastatic colorectal cancer is still questioned. The aim of this study was to evaluate the clinical value of CTC dynamics in mCRC patients receiving first-line treatments. MATERIALS AND METHODS: Serial CTC data from 218 patients were used to identify CTC trajectory patterns during the course of treatment. CTCs were evaluated at baseline, at a first-time point check and at the radiological progression of the disease. CTC dynamics were correlated with clinical endpoints. RESULTS: Using a cut-off of ≥1 CTC/7.5 mL, four prognostic trajectories were outlined. The best prognosis was obtained for patients with no evidence of CTCs at any timepoints, with a significant difference compared to all other groups. Lower PFS and OS were recognized in group 4 (CTCs always positive) at 7 and 16 months, respectively. CONCLUSIONS: We confirmed the clinical value of CTC positivity, even with only one cell detected. CTC trajectories are better prognostic indicators than CTC enumeration at baseline. The reported prognostic groups might help to improve risk stratification, providing potential biomarkers to monitor first-line treatments.

Pulmonary embolism post-Covid-19 infection: physiopathological mechanisms and vascular damage biomarkers.

Covid-19 infection is characterized by several acute complications, as well long-term sequelae, mostly sustained by endothelial dysfunction; several studies show that complications as pulmonary embolism (PE) are described both in the acute phase and after negativization. Aim of research was to evaluate anthropometric, bio-humoral, instrumental parameters in a group of patients affected by PE after recent Covid-19 infection compared to PE patients without previous Covid-19 infection. We enrolled 72 consecutive patients (35M, 37F) with acute PE, distinguished in relation to previous acute Covid-19 infection: 54 pts without previous acute Covid-19 infection and 18 pts with previous Covid-19 infection within negativity at least 2 months before PE diagnosis; 44 healthy subjects (21M, 23F) were recruited as control group. Patients who had previously developed Covid-19 needed hospitalization in high percentage (84%); this group showed significantly higher prevalence of diabetes mellitus than Covid-19-free PE patients, reduced serum levels of C-reactive protein, sST2 and PESI score. In post-Covid-19 PE group, we observed higher mean IMPROVE risk score, whereas in Covid-19-free group lower P/F ratio, higher radiological severity, and worse PESI score and severity index. Covid-19 infection affects not just the lung parenchyma but also other organs; endothelial damage plays pivotal role in long-term alterations; in high thrombotic risk group (recent hospitalization due to acute Covid-19 infection), we have described thrombotic complications characterized by persistent prothrombotic state after recovery, highlighted by well-known markers as PCR and D-Dimer as well as novel vascular marker (sST2).

Fibrosis-4 (FIB-4) Index and mortality in COVID-19 patients admitted to the emergency department.

Liver damage worsens the prognosis of coronavirus 19 disease (COVID-19). However, the best strategy to stratify mortality risk according to liver damage has not been established. The aim of this study is to test the predictive value of the validated Fibrosis-4 (FIB-4) Index and compared it to liver transaminases and to the AST-to-Platelet ratio index (APRI). Multicenter cohort study including 992 consecutive COVID-19 patients admitted to the Emergency Department. FIB-4 > 3.25 and APRI > 0.7 were used to define liver damage. Multivariable Cox regression and ROC curve analysis for mortality were performed. Secondary endpoints were (1) need for high-flow oxygen and (2) mechanical ventilation. 240 (24.2%) patients had a FIB-4 > 3.25. FIB-4 > 3.25 associated with an increased mortality (n = 119, log-rank test p < 0.001 and adjusted hazard ratio (HR) 1.72 (95% confidence interval [95%CI] 1.14-2.59, p = 0.010). ROC analysis for mortality showed that FIB-4 (AUC 0.734, 95% CI 0.705-0.761) had a higher predictive value than AST (p = 0.0018) and ALT (p < 0.0001). FIB-4 > 3.25 was also superior to APRI > 0.7 (AUC 0.58, 95% CI 0.553-0.615, p = 0.0008). Using an optimized cut-off > 2.76 (AUC 0.689, 95% CI 0.659-0.718, p < 0.0001), FIB-4 was superior to FIB-4 > 3.25 (p = 0.0302), APRI > 0.7 (p < 0.0001), AST > 51 (p = 0.0119) and ALT > 42 (p < 0.0001). FIB-4 was also associated with high-flow oxygen use (n = 255, HR 1.69, 95% CI 1.25-2.28, p = 0.001) and mechanical ventilation (n = 39, HR 2.07, 95% CI 1.03-4.19, p = 0.043). FIB-4 score predicts mortality better than liver transaminases and APRI score. FIB-4 score may be an easy tool to identify COVID-19 patients at worse prognosis in the emergency department.

The Influence of Ketone Bodies on Circadian Processes Regarding Appetite, Sleep and Hormone Release: A Systematic Review of the Literature.

Chrononutrition is an emerging branch of chronobiology focusing on the profound interactions between biological rhythms and metabolism. This framework suggests that, just like all biological processes, even nutrition follows a circadian pattern. Recent findings elucidated the metabolic roles of circadian clocks in the regulation of both hormone release and the daily feeding-fasting cycle. Apart from serving as energy fuel, ketone bodies play pivotal roles as signaling mediators and drivers of gene transcription, promoting food anticipation and loss of appetite. Herein we provide a comprehensive review of the literature on the effects of the ketogenic diets on biological processes that follow circadian rhythms, among them appetite, sleep, and endocrine function.

Circulating tumour cells lacking cytokeratin in breast cancer: the importance of being mesenchymal.

Circulating tumour cells (CTCs) are independent predictor of prognosis in metastatic breast cancer. Nevertheless, in one third of patients, circulating tumour cells are undetected by conventional methods. Aim of the study was to assess the prognostic value of circulating tumour cells expressing mesenchymal markers in metastatic breast cancer patients. We isolated CTC from blood of 55 metastatic breast cancer patients. CTC were characterized for cytokeratins and markers of epithelial mesenchymal transition. The gain of mesenchymal markers in CTC was correlated to prognosis of patients in a follow-up of 24 months. The presence of mesenchymal markers on CTC more accurately predicted worse prognosis than the expression of cytokeratins alone. Because of the frequent loss of epithelial antigens by CTC, assays targeting epithelial antigens may miss the most invasive cell population. Thus, there is an urgent need to improve detection methods to identify CTC which undergone epithelial mesenchymal transition program.

Genetic Vaccination as a Flexible Tool to Overcome the Immunological Complexity of Invasive Fungal Infections.

The COVID-19 pandemic has highlighted genetic vaccination as a powerful and cost-effective tool to counteract infectious diseases. Invasive fungal infections (IFI) remain a major challenge among immune compromised patients, particularly those undergoing allogeneic hematopoietic bone marrow transplantation (HSCT) or solid organ transplant (SOT) both presenting high morbidity and mortality rates. Candidiasis and Aspergillosis are the major fungal infections among these patients and the failure of current antifungal therapies call for new therapeutic aids. Vaccination represents a valid alternative, and proof of concept of the efficacy of this approach has been provided at clinical level. This review will analyze current understanding of antifungal immunology, with a particular focus on genetic vaccination as a suitable strategy to counteract these diseases.

Ketogenic Diet as a Preventive and Supportive Care for COVID-19 Patients.

Severe obesity is associated with an increased risk of admission to intensive care units and need for invasive mechanical ventilation in patients with COVID-19. The association of obesity and COVID-19 prognosis may be related to many different factors, such as chronic systemic inflammation, the predisposition to severe respiratory conditions and viral infections. The ketogenic diet is an approach that can be extremely effective in reducing body weight and visceral fat in the short term, preserving the lean mass and reducing systemic inflammation. Therefore, it is a precious preventive measure for severely obese people and may be considered as an adjuvant therapy for patients with respiratory compromise.

Sequential Isolation and Characterization of Single CTCs and Large CTC Clusters in Metastatic Colorectal Cancer Patients.

Circulating tumor cells (CTCs) detach from a primary tumor or its metastases and circulate in the bloodstream. The vast majority of CTCs are deemed to die into the bloodstream, with only few cells representing viable metastatic precursors. Particularly, single epithelial CTCs do not survive long in the circulation due to the loss of adhesion-dependent survival signals. In metastatic colorectal cancer, the generation of large CTC clusters is a very frequent occurrence, able to increase the aptitude of CTCs to survive in the bloodstream. Although a deepened analysis of large-sized CTC clusters might certainly offer new insights into the complexity of the metastatic cascade, most CTC isolation techniques are unfortunately not compatible with large-sized CTC clusters isolation. The inappropriateness of standard CTC isolation devices for large clusters isolation and the scarce availability of detection methods able to specifically isolate and characterize both single CTCs and CTC clusters finally prevented in-depth studies on the prognostic and predictive value of clusters in clinical practice, unlike that which has been described for single CTCs. In the present study, we validated a new sequential filtration method for the simultaneous isolation of large CTC clusters and single CTCs in patients with metastatic colorectal cancer at failure of first-line treatments. The new method might allow differential downstream analyses for single and clustered CTCs starting from a single blood draw, opening new scenarios for an ever more precise characterization of colorectal cancer metastatic cascade.

Chemosensitivity profile assay of circulating cancer cells: prognostic and predictive value in epithelial tumors.

The prognostic value associated with the detection of circulating tumor cells (CTCs) in metastatic breast cancer by the CellSearch technology raise additional issues regarding the biological value of this information. We postulated that a drug-resistance profile of CTCs may predict response to chemotherapy in cancer patients and therefore could be used for patient selection. One hundred 5 patients with diagnosis of carcinoma were enrolled in a prospective trial. CTCs were isolated from peripheral blood, and positive samples were evaluated for the expression of a panel of genes involved in anticancer drugs resistance. The drug-resistance profile was correlated with disease-free survival (DFS; patients in adjuvant setting) and time to progression (TTP; metastatic patients) in a 24-months follow-up. Objective response correlation was a secondary end point. Fifty-one percent of patients were found positive for CTCs while all blood samples from healthy donors were negative. The drug-resistance profile correlates with DFS and TTP (p < 0.001 in both). Sensitivity of the test: able to predict treatment response in 98% of patients. Specificity of the test: 100%; no sample from healthy subject was positive for the presence of CTCs. Positive and negative predictive values were found to be 96.5 and 100%, respectively. We identified a drug-resistance profile of CTCs, which is predictive of response to chemotherapy, independent of tumor type and stage of disease. This approach may represent a first step toward the individualization of chemotherapy in cancer patients.

Failure of apoptosis and activation on NFkappaB by celecoxib and aspirin in lung cancer cell lines.

Recent studies have demonstrated that antineoplastic activity of Cox-2 inhibitors may depend on targets other than Cox: among those, nuclear factor kappaB (NFkappaB) seems the most promising. Although preclinical studies have suggested that aspirin and Cox-2 inhibitors may influence the progression of lung cancer, the molecular mechanisms of these protective effects in this tumor type has not been fully elucidated. We investigated the effects of celecoxib and aspirin in the induction of apoptosis and in the ability to activate NFkappaB in three non-small cell lung cancer cell lines. Apoptosis was evaluated by FACS, caspase activation assay and expression of apoptosis-related genes by RT-PCR, while NFkappaB activation was assessed by immunofluorescence. No apoptotic response was observed after treatment with both high and low dose of celecoxib. Nevertheless, celecoxib at both concentrations induced a strong NFkappaB activation, with increased expression of NFkappaB-dependent genes, such as bcl-2, bcl-XL and survivin. Similarly, aspirin at both concentrations did not induce any apoptotic response, but activated NFkappaB in a dose-dependent manner. This study supports the hypothesis that NFkappaB activation is an important effect of NSAIDs in lung cancer, leading to apoptosis resistance. This effect of both aspirin and celecoxib may be considered undesirable in lung cancer chemoprevention.

Monitoring PD-L1 positive circulating tumor cells in non-small cell lung cancer patients treated with the PD-1 inhibitor Nivolumab.

Controversial results on the predictive value of programmed death ligand 1 (PD-L1) status in lung tumor tissue for response to immune checkpoint inhibitors do not allow for any conclusive consideration. Liquid biopsy might allow real-time sampling of patients for PD-L1 through the course of the disease. Twenty-four stage IV NSCLC patients included in the Expanded Access Program with Nivolumab were enrolled. Circulating tumor cells (CTCs) were analyzed by CellSearch with anti-human B7-H1/PD-L1 PE-conjugated antibody. PD-L1 expressing CTCs were assessed at baseline, at 3 and 6 months after starting therapy, and correlated with outcome. At baseline and at 3 months of treatment, the presence of CTCs and the expression of PD-L1 on their surface were found associated to poor patients outcome. Nevertheless, the high frequency of PD-L1 expressing CTCs hampered to discriminate the role of PD-L1 in defining prognosis. Conversely although CTCs were found in all patients 6 months after treatment, at this time patients could be dichotomized into two groups based PD-L1 expression on CTCs. Patients with PD-L1 negative CTCs all obtained a clinical benefit, while patients with PD-L1 (+) CTCs all experienced progressive disease. This suggests that the persistence of PD-L1(+) CTCs might mirror a mechanism of therapy escape.

Tenascin C and epidermal growth factor receptor as markers of circulating tumoral cells in bladder and colon cancer.

Tenascin C has been recently suggested as a tumor marker, however, its levels in serum has been evaluated only in patients with head and neck cancer and melanoma. In this study, we investigated Tenascin C expression in blood samples from colorectal and bladder cancer patients, compared to that of epidermal growth factor receptor (EGFR), a known circulating tumor marker in these cancer types. RT-PCR specific for Tenascin C and EGFR was performed on RNAs extracted from blood samples of 60 patients affected by colon or bladder cancer. We then investigated the statistical association between Tenascin C, EGFR expression and disease-free survival using the Kaplan-Meier method. Furthermore, in order to select which variable between EGFR and Tenascin C was the most predictive for recurrence, a Cox model for proportional risk was applied. Among all patients analysed, a significantly higher disease-free time was found in the group negative for both EGFR and Tenascin C expression; EGFR expression was significantly correlated to disease progression in stages III and IV, whereas in all patients with stage I and II disease Tenascin C correlated better with prognosis. Negative expression of both EGFR and Tenascin C identifies a group of patients with poor tendency to disease recurrence and longer relapse-free time. While Tenascin C expression seems to influence prognosis in patients with low-stage disease, EGFR appears a marker of worse prognosis in patients with high-staged tumors.

Persistence of epidermal growth factor receptor and interleukin 10 in blood of colorectal cancer patients after surgery identifies patients with high risk to relapse.

PURPOSE: Despite the great number of studies performed to detect circulating markers of disease progression in colorectal cancer, few have shown a clinical use; among those, epidermal growth factor receptor (EGFR) and, more recently, interleukin (IL)-10. In this article, we sought to investigate how primary surgery could affect expression levels of EGFR, IL-6, and IL-10 in blood from colorectal cancer patients. EXPERIMENTAL DESIGN: We investigated by reverse transcriptase-PCR assay the expression at mRNA level of EGFR, IL-6, and IL-10 in blood samples taken from 56 colorectal cancer patients. Each gene expression was evaluated 1 day before and 20 days after primary surgery. Persistence of each gene in blood after surgery was then correlated to the relapse free time in a follow-up of 3 years. RESULTS: In blood samples taken before surgery, EGFR, IL-6, and IL-10 were found expressed in 62, 100, and 100% of patients, respectively. EGFR expression, but not IL-6 and IL-10, correlates with stage of disease. In the group of 41 patients who underwent follow-up studies, EGFR was found persistently high in 67%; 94% of them had relapse. Persistence of IL-10 after surgery also identifies relapses in 89% of cases. IL-6 persistence was not found to significantly correlate to progression of disease. CONCLUSIONS: Persistence of both EGFR and IL-10 in blood of colorectal cancer patients after surgery identifies patients with high propensity to relapse. These findings may suggest a clinical use of preoperative EGFR/IL-10 reverse transcriptase-PCR assay in the prediction of tumor recurrence.

Detection of CK19, CK20 and EGFR mRNAs in peripheral blood of carcinoma patients: correlation with clinical stage of disease.

Expression of genes such as cytokeratin 19 (CK19), cytokeratin 20 (CK20) and epidermal growth factor receptor (EGFR) has been investigated at mRNA level in peripheral blood of carcinoma patients to detect the presence of circulating tumor cells (CTC). We performed this study because recent literature emphasizes that the importance of CK19, 20 and EGFR mRNAs in CTC as prognostic factors remains unclear especially for breast, head and neck and colon cancer patients. Reverse transcriptase polymerase chain reaction (RT-PCR) followed by Southern blot hybridization was performed in blood samples from 47 subjects (12 colorectal, 15 head and neck and 20 breast carcinoma patients), as well as in 35 healthy donors. The CK19 expression was found in 36/47 patients (9 colorectal, 9 head and neck and 18 breast cancer), two patients (one affected by colorectal and one by head and neck cancer) were positive for CK20 whereas EGFR was found expressed in 9 patients (3 colorectal, 5 head and neck and one breast cancer). Seven of 35 and 4/35 healthy donors displayed positivity for the expression of CK19 and CK20 genes respectively, whereas no EGFR mRNA was found in this group. The correlation of the detection of CTC in peripheral blood with progression of the disease in a follow-up period of 40 months did not show any prognostic value to the presence of mRNAs of these biomarkers in blood. We believe that research should be addressed, at least for breast cancer, to the identification of occult metastases in sentinel lymph nodes, such as recently performed in melanoma patients.