Objectives
The objective of this study is to study the effect of electrochemical treatment on biofilms developed on titanium dental implants, using a six‐species in vitro model simulating subgingival ...oral biofilms.
Materials and Methods
Direct electrical current (DC) of 0.75 V, 1.5 V, and 3 V (anodic polarization, oxidation processes) and of −0.75 V, −1.5 V, and ‐3 V (cathodic polarization, reduction processes) was applied between the working and the reference electrodes for 5 min on titanium dental implants, which have been previously inoculated with a multispecies biofilm. This electrical application consisted of a three‐electrode system where the implant was the working electrode, a platinum mesh was the counter electrode, and an Ag/AgCl electrode was the reference. The effect of the electrical application on the biofilm structure and bacterial composition was evaluated by scanning electron microscopy and quantitative polymerase chain reaction. A generalized linear model was applied to study the bactericidal effect of the proposed treatment.
Results
The electrochemical construct at 3 V and −3 V settings significantly reduced total bacterial counts (p < .05) from 3.15 × 106 to 1.85 × 105 and 2.92 × 104 live bacteria/mL, respectively. Fusobacterium nucleatum was the most affected species in terms of reduction in concentration. The 0.75 V and −0.75 V treatments had no effect on the biofilm.
Conclusion
Electrochemical treatments had a bactericidal effect on this multispecies subgingival in vitro biofilm model, being the reduction more effective than the oxidative treatment.
Objective
The aim of the study was to evaluate the effect of doxycycline‐ and dexamethasone‐doped collagen membranes on the proliferation and differentiation of osteoblasts.
Background
Collagen ...barrier membranes are frequently used to promote bone regeneration and to boost this biological activity their functionalization with antibacterial and immunomodulatory substances has been suggested.
Methods
The design included commercially available collagen membranes doped with doxycycline (Dox‐Col‐M) or dexamethasone (Dex‐Col‐M), as well as undoped membranes (Col‐M) as controls, which were placed in contact with cultured MG63 osteoblast‐like cells (ATCC). Cell proliferation was assessed by 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium (MTT) assay and differentiation by measuring the alkaline phosphatase (ALP) activity using spectrophotometry. Real‐time quantitative polymerase chain reaction was used to study the expression of the genes: Runx‐2, OSX, ALP, OSC, OPG, RANKL, Col‐I, BMP‐2, BMP‐7, TGF‐β1, VEGF, TGF‐βR1, TGF‐βR2, and TGF‐βR3. Scanning electron microscopy was used to study osteoblast morphology. Data were assessed using one‐way analysis of variance or Kruskal–Wallis tests, once their distribution normality was assessed by Kolmogorov–Smirnov tests (p > .05). Bonferroni for multiple comparisons were carried out (p < .05).
Results
Osteoblast proliferation was significantly enhanced in the functionalized membranes as follows: (Col‐M < Dex‐Col‐M < Dox‐Col‐M). ALP activity was significantly higher on cultured osteoblasts on Dox‐Col‐M. Runx‐2, OSX, ALP, OSC, BMP‐2, BMP‐7, TGF‐β1, VEGF, TGF‐βR1, TGF‐βR2, and TGF‐βR3 were overexpressed, and RANKL was down‐regulated in osteoblasts cultured on Dox‐Col‐M. The osteoblasts cultured in contact with the functionalized membranes demonstrated an elongated spindle‐shaped morphology.
Conclusion
The functionalization of collagen membranes with Dox promoted an increase in the proliferation and differentiation of osteoblasts.
•Image-identified extranodal extension (iENE)-positive tumor represents a more aggressive phenotype.•iENE is an independent prognostic factor for laryngeal and hypopharyngeal carcinoma.•iENE provides ...additional value beyond TNM-8 cN that already includes the clinical extranodal extension parameter.
Clinical extranodal extension (cENE) is a cN modifier in TNM-8 for laryngo-hypopharygeal carcinoma (LHC). We hypothesize that image-detected ENE (iENE) can provide additional prognostic value over cENE in LHC.
Baseline CTs/MRIs of cN+ LHC patients treated with definitive (chemo-)radiotherapy between 2010–2019 were re-reviewed by a neuroradiologist using internationally accepted criteria for iENE-positive/negative (iENE+/iENE–). Overall survival (OS) was compared by iENE status. Multivariable analysis (MVA) was performed to confirm the prognostic value of iENE, adjusted for known potential confounders.
A total of 232 LHC patients were identified, including 154 iENE–/cENE–, 60 iENE+/cENE–, and 18 iENE+/cENE+. A higher proportion of iENE+ (vs iENE–) patients had lymph node (LN) size > 3 cm 53 (67 %) vs 4 (3 %), >=5 LNs 51 (65 %) vs 33 (21 %), and retropharyngeal LN 12 (15 %) vs 6 (4 %) (all p < 0.01). Median follow-up was 4.8 years. iENE+/cENE– and iENE+/cENE+patients had similarly low 5-year OS 28 % (18–44) and 29 % (13–63) vs iENE–/cENE– 53 % (45–62) (p < 0.001). On MVA, mortality risk was higher with iENE+vs iENE– hazard ratio (HR) 2.22 (95 % CI 1.47–3.36). The prognostic value of iENE remained with MVA in larynx (n = 124) (HR 2.51 1.35–4.68, p = 0.004 or hypopharynx (n = 108) (HR 1.87 1.02–3.43, p = 0.04) patients, separately.
Our study confirms the independent prognostic importance of iENE for LHC following definitive (chemo-)radiotherapy beyond TNM-8 cN status that already contains the cENE parameter. Further research is needed to explore whether iENE could replace cENE for future cN classification.
Sequencing of tumors is now routine and guides personalized cancer therapy. Mutant allele fractions (MAFs, or the ‘mutation dose’) of a driver gene may reveal the genomic structure of tumors and ...influence response to targeted therapies. We performed a comprehensive analysis of MAFs of driver alterations in unpaired primary and metastatic colorectal cancer (CRC) at our institution from 2010 to 2015 and studied their potential clinical relevance. Of 763 CRC samples, 622 had detailed annotation on overall survival in the metastatic setting (OSmet) and 89 received targeted agents matched to KRAS (MEK inhibitors), BRAF (BRAF inhibitors), or PIK3CA mutations (PI3K pathway inhibitors). MAFs of each variant were normalized for tumor purity in the sample (adjMAFs). We found lower adjMAFs for BRAFV600E and PIK3CA than for KRAS, NRAS, and BRAF non‐V600 variants. TP53 and BRAFV600E adjMAFs were higher in metastases as compared to primary tumors, and high KRAS adjMAFs were found in CRC metastases of patients with KRAS wild‐type primary tumors previously exposed to EGFR antibodies. Patients with RAS‐ or BRAFV600E‐mutated tumors, irrespective of adjMAFs, had worse OSmet. There was no significant association between adjMAFs and time to progression on targeted therapies matched to KRAS, BRAF, or PIK3CA mutations, potentially related to the limited antitumor activity of the employed drugs (overall response rate of 4.5%). In conclusion, the lower BRAFV600E and PIK3CA adjMAFs in subsets of primary CRC tumors indicate subclonality of these driver genes. Differences in adjMAFs between metastases and primary tumors suggest that approved therapies may result in selection of BRAFV600E‐ and KRAS‐resistant clones and an increase in genomic heterogeneity with acquired TP53 alterations. Despite significant differences in prognosis according to mutations in driver oncogenes, adjMAFs levels did not impact on survival and did not help predict benefit with matched targeted agents in the metastatic setting.
A comprehensive analysis of mutant allele fractions of driver oncogene mutations in colorectal cancer suggests that (i) some events, including BRAFV600E, may be subclonal; (ii) standard chemotherapies and EGFR antibodies may change the genomic structure of metastatic lesions, with acquired gene alterations and selection of resistant clones; and (iii) the clonality of events does not affect patient outcome or response to matched targeted agents.
Reactivation of latent imported infections has been periodically reported in migrant patients undergoing immunosuppression. We performed a prospective study at Vall d'Hebron University Hospital ...(Barcelona, Spain). Migrant patients over 16 years with the diagnosis of any oncohematologic disease were included. Patients were tested for soil-transmitted helminths, hepatitis virus, and human immunodeficiency virus, Treponema pallidum, human T-cell lymphotropic virus, latent tuberculosis infection, Toxoplasma spp., Plasmodium infection, Schistosoma spp., Trypanosoma cruzi infection, Leishmania spp., and dimorphic fungi. Patients were treated and followed for 1 year to assess reactivation. A total of 42 patients were included in this study. Median age was 39 (31-51) years. Twenty-five (59.5%) patients were women. More than half of the patients were of Latin American origin. Sixteen patients (38.1%) underwent hematopoietic stem cell transplantation. Of the patients, 71.4% had at least one imported infection. Patients with at least one positive result in the screening did not show any statistically significant association with the studied variables. We did not find any reactivation of the treated latent infections. After specific treatment we did not observe any reactivation. Screening of latent imported infections previous to an immunosuppressive treatment is easy to perform and it may be lifesaving.
Eligibility criteria restrict patient enrollment in clinical trials. A Nature paper applied a machine-learning algorithm in a real-world database to show that relaxing some criteria may not ...jeopardize efficacy and safety. This may enable more patients to have earlier access to new therapies and make results more generalizable to clinical practice.
Eligibility criteria restrict patient enrollment in clinical trials. A Nature paper applied a machine-learning algorithm in a real-world database to show that relaxing some criteria may not jeopardize efficacy and safety. This may enable more patients to have earlier access to new therapies and make results more generalizable to clinical practice.
Glucagon-like peptide 1 (GLP-1) and PAS kinase (PASK) control glucose and energy homeostasis according to nutritional status. Thus, both glucose availability and GLP-1 lead to hepatic glycogen ...synthesis or degradation. We used a murine model to discover whether PASK mediates the effect of exendin-4 (GLP-1 analogue) in the adaptation of hepatic glycogen metabolism to nutritional status. The results indicate that both exendin-4 and fasting block the Pask expression, and PASK deficiency disrupts the physiological levels of blood GLP1 and the expression of hepatic GLP1 receptors after fasting. Under a non-fasted state, exendin-4 treatment blocks AKT activation, whereby Glucokinase and Sterol Regulatory Element-Binding Protein-1c (Srebp1c) expressions were inhibited. Furthermore, the expression of certain lipogenic genes was impaired, while increasing Glucose Transporter 2 (GLUT2) and Glycogen Synthase (GYS). Moreover, exendin-4 treatment under fasted conditions avoided Glucose 6-Phosphatase (G6pase) expression, while maintaining high GYS and its activation state. These results lead to an abnormal glycogen accumulation in the liver under fasting, both in PASK-deficient mice and in exendin-4 treated wild-type mice. In short, exendin-4 and PASK both regulate glucose transport and glycogen storage, and some of the exendin-4 effects could therefore be due to the blocking of the Pask expression.
Despite major advances in the treatment of metastatic colorectal cancer (mCRC), the survival rate remains very poor. This study aims at exploring the prognostic value of RAS‐mutant allele fraction ...(MAF) in plasma in mCRC. Forty‐seven plasma samples from 37 RAS‐mutated patients with nonresectable metastases were tested for RAS in circulating tumor DNA using BEAMing before first‐ and/or second‐line treatment. RAS MAF was correlated with several clinical parameters (number of metastatic sites, hepatic volume, carcinoembryonic antigen, CA19‐9 levels, primary site location, and treatment line) and clinical outcome progression‐free survival (PFS) and overall survival (OS). An independent cohort of 32 patients from the CAPRI‐GOIM trial was assessed for clinical outcome based on plasma baseline MAF. RAS MAF analysis at baseline revealed a significant correlation with longer OS Hazard ratios (HR) = 3.514; P = 0.00066. Patients with lower MAF also showed a tendency to longer PFS, although not statistically significant. Multivariate analysis showed RAS MAFs as an independent prognostic factor in both OS (HR = 2.73; P = 0.006) and first‐line PFS (HR = 3.74; P = 0.049). Tumor response to treatment in patients with higher MAF was progression disease (P = 0.007). Patients with low MAFs at baseline in the CAPRI‐GOIM group also showed better OS HR = 3.84; 95% confidence intervals (CI) 1.5–9.6; P = 0.004 and better PFS (HR = 2.5; 95% CI: 1.07–5.62; P = 0.033). This minimally invasive test may help in adding an independent factor to better estimate outcomes before initiating treatment. Further prospective studies using MAF as a stratification factor could further validate its utility in clinical practice.
Therapy selection in first‐line treatment of metastatic colorectal cancer (mCRC) depends on tumor RAS mutation status. The use of liquid biopsy, with reported concordance rates between tissue and plasma of around 90%, is an emerging alternative to detect RAS mutations. Beyond mutation status, data support that RAS mutant allele fractions in plasma have independent prognostic value for mCRC survival.
The search for immunotherapy biomarkers in Microsatellite Instability High/Deficient Mismatch Repair system (MSI-H/dMMR) metastatic colorectal cancer (mCRC) is an unmet need. Sixteen patients with ...mCRC and MSI-H/dMMR (determined by either immunohistochemistry or polymerase chain reaction) treated with PD-1/PD-L1 inhibitors at our institution were included. According to whether the progression-free survival with PD-1/PD-L1 inhibitors was longer than 6 months or shorter, patients were clustered into the IT-responder group (
: 9 patients) or IT-resistant group (
: 7 patients), respectively. In order to evaluate determinants of benefit with PD-1/PD-L1 inhibitors, we performed multimodal analysis including genomics (through NGS panel tumour-only with 431 genes) and the immune microenvironment (using CD3, CD8, FOXP3 and PD-L1 antibodies). The following mutations were more frequent in IT-resistant compared with IT-responder groups:
(4/7 versus 2/9),
(2/7 versus 0/9), and biallelic
(3/7 versus 1/9). Biallelic
mutations were found exclusively in the IT-responder group (4/9 patients). Tumour mutational burden did not correlate with immunotherapy benefit, neither the rate of
in homopolymeric regions. Of note, biallelic
mutated tumours had the highest immune infiltration and PD-L1 scores, contrary to tumours with
mutation. Immune microenvironment analysis showed higher densities of different T cell subpopulations and PD-L1 expression in IT-responders. Misdiagnosis of MSI-H/dMMR inferred by discordances between immunohistochemistry and polymerase chain reaction was only found in the IT-resistant population (3/7 patients). Biallelic
mutations and Wnt signalling activation through
mutation were associated with high and low T cell immune infiltrates, respectively, and deserve special attention as determinants of response to PD-1/PD-L1 inhibitors. The non-MSI-H phenotype in dMMR is associated with poor benefit to immunotherapy. Our results suggest that mechanisms of resistance to immunotherapy are multi-factorial.
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