Mesenchymal epithelial transition receptor (
) alterations, including
exon 14 skipping mutation, are oncogenic in non-small cell lung cancer (NSCLC) and may confer sensitivity to targeted therapy. ...Given the rarity and the diversity of exon 14 skipping mutations, diagnosis may be challenging on small-biopsy specimens.
Between March 2014 and May 2018, tissue samples from patients with metastatic NSCLC were analysed for
exon 14 skipping mutation as part of routine practice in the Pathology Department of the Hospices Civils de Lyon, France. Over the study period, Sanger sequencing and/or two different DNA-based next generation sequencing (NGS) assays were used.
Genomic alterations of
exon 14 were detected in 2.6% (62/2,369) samples of NSCLC analysed for
exon 14 mutations. Patients were mainly women (38/62, 61%) without smoking history (22/39, 56%) and the median age was 75 years.
exon 14 skipping mutations were diagnosed by NGS in 50 cases and by classical Sanger sequencing in 12 cases. The frequency of
mutations was 15.4% when Sanger sequencing was performed at the request of the clinician and 4.1% when the DNA-based NGS assay coverage included the 3' and 5' parts of the
exon 14 and performed systematically.
The frequency of genomic alterations is highly dependent on patient selection and the technical approach.
Corneal endothelial diseases are the leading cause of corneal transplantation. The global shortage of donor corneas has resulted in the investigation of alternative methods, such as cell therapy and ...tissue-engineered endothelial keratoplasty (TEEK), using primary cultures of human corneal endothelial cells (hCECs). The main challenge is optimizing the hCEC culture process to increase the endothelial cell density (ECD) and overall yield while preventing endothelial-mesenchymal transition (EndMT). Fetal bovine serum (FBS) is necessary for hCEC expansion but contains TGF-βs, which have been shown to be detrimental to hCECs. Therefore, we investigated various TGF-β signaling pathways using inhibitors to improve hCEC culture. Initially, we confirmed that TGF-β1, 2, and 3 induced EndMT on confluent hCECs without FBS. Using this TGF-β-induced EndMT model, we validated NCAM as a reliable biomarker to assess EndMT. We then demonstrated that, in a culture medium containing 8% FBS for hCEC expansion, TGF-β1 and 3, but not 2, significantly reduced the ECD and caused EndMT. TGF-β receptor inhibition had an anti-EndMT effect. Inhibition of the ROCK pathway, notably that of the P38 MAPK pathway, increased the ECD, while inhibition of the ERK pathway decreased the ECD. In conclusion, the presence of TGF-β1 and 3 in 8% FBS leads to a reduction in ECD and induces EndMT. The use of SB431542 or LY2109761 may prevent EndMT, while Y27632 or Ripasudil, and SB203580 or SB202190, can increase the ECD.
Letter to the Editor SMARCE1 (SWI/SNF related, matrix associated, actin dependent regulator of chromatin, subfamily e, member 1) is a constant component of the SWI/SNF (SWItch/Sucrose ...Non-Fermentable) chromatin remodeling complexes. The biallelic hit resulting in the loss of protein expression 5 further argues in favor of a driver role of SMARCE1 in our case. ...the tumor we report enlarges the spectrum of SMARCE1-related brain tumors to a so far undescribed high-grade neoplasm. ...a CCM could be formally ruled out because the tumor was not dura matter based, microscopic findings are not consistent with clear cell meningioma, immunohistochemical features were those of a tumor without SSTR2A expression and methylation profiling was clearly in disfavor of this diagnosis.
Corneal endothelial cells (CECs) form a monolayer at the innermost face of the cornea and are the engine of corneal transparency. Nevertheless, they are a vulnerable population incapable of ...regeneration in humans, and their diseases are responsible for one third of corneal grafts performed worldwide. Donor corneas are stored in eye banks for security and quality controls, then delivered to surgeons. This period could allow specific interventions to modify the characteristics of CECs in order to increase their proliferative capacity, increase their resistance to apoptosis, or release immunosuppressive molecules. Delivery of molecules specifically into CECs during storage would therefore open up new therapeutic perspectives. For clinical applications, physical methods have a more favorable individual and general benefit/risk ratio than most biological vectors, but are often less efficient. The delivery of molecules into cells by carbon nanoparticles activated by femtosecond laser pulses is a promising recent technique developed on non-adherent cells. The nanoparticles are partly consummated by the reaction releasing CO and H2 gas bubbles responsible for the shockwave at the origin of cell transient permeation. Our aim was to develop an experimental setting to deliver a small molecule (calcein) into the monolayer of adherent CECs. We confirmed that increased laser fluence and time exposure increased uptake efficiency while keeping cell mortality below 5%. We optimized the area covered by the laser beam by using a motorized stage allowing homogeneous scanning of the cell culture surface using a spiral path. Calcein uptake reached median efficiency of 54.5% (range 50.3-57.3) of CECs with low mortality (0.5%, range (0.55-1.0)). After sorting by flow cytometry, CECs having uptaken calcein remained viable and presented normal morphological characteristics. Delivery of molecules into CECs by carbon nanoparticles activated by femtosecond laser could prove useful for future cell or tissue therapy.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Primary central nervous system lymphoma (PCNSL) is a rare subtype of non‐Hodgkin lymphoma (NHL) with extranodal location affecting only the CNS, meninges and eye, without visceral or lymph node ...involvement. Its incidence has increased sharply over the past three decades, especially in immunocompetent subjects. Most PCNSL cases are diffuse large B‐cell lymphomas (DLBCLs). However, it differs from nodal DLBCL in that it has a worse prognosis. DLBCLs are a heterogeneous entity and according to new genomic discoveries, classifications into prognostic subgroups have been embarked upon. Two prognostic algorithms were then prepared using a panel of immunohistochemical markers (CD10, Bcl6, MUM1/IRF‐4, and Bcl2), thus categorizing DLBCL into two subgroups, GCB (germinal centre B‐cell‐like) or non‐GCB, and into Group 1 or Group 2. Our goal is to apply both of these two sub‐classifications to 39 PCNSLs, in order to assess their usefulness and prognostic relevance. 74.3% of our PCNSLs were of a non‐GCB phenotype, corresponding to an activated postgerminal origin. They were evenly distributed across G1 and G2. Two‐ and 5‐year overall survival rates were 34.8% and 19.6%, respectively. Younger age (<65) and a therapeutic combination of chemotherapy and radiotherapy significantly improved our patients' survival rates. The other clinical or biological markers tested had no prognostic impact. The two classifications did not reveal any significant survival difference. The recent discovery of a specific “transcriptional signature” of PCNSL, marking them out of DLBCL could account for the irrelevance of such prognostic classifications to PCNSL.
We report retrospective data on the feasibility and efficacy of prolonging adjuvant temozolomide (TMZ) more than 6 months after chemoradiotherapy completion in patients with glioblastoma (GBM). ...Molecular prognostic factors were assessed. Data from 46 patients were reviewed. Patients received postoperative irradiation, 60 Gy in 30 fractions, combined with concurrent TMZ, 75 mg/m
2
. Four weeks later, adjuvant TMZ was prescribed, 150–200 mg/m
2
for a total of 24 cycles unless there was progression or toxicity. Tumor samples were tested for the following prognostic factors: EGFR overexpression, 1p19q deletion, p53 overexpression and proliferation index. Overall survival (OS) was 84.8% at 6 months, 54.3% at 12 months, 26.1% at 18 months, and 21.7% at 24 months. Progression-free survival (PFS) was 73.9% at 6 months, 34.8% at 12 months, 15.2% at 18 months and 10.4% at 24 months. In the adjuvant phase, no treatment disruption for toxicity was necessary but eight patients required dose adaptation because of side effects. No significant molecular prognostic factor was evidenced for OS. We found that p53 overexpression was the only significant prognostic factor for PFS, with a median PFS of 9.3 months versus 7 months for patients without p53 overexpression (
P
= 0.031). This study suggests that delivering adjuvant TMZ therapy for more than 6 months is feasible in patients with GBM. Efficacy data warrant further prospective assessment with the focus on molecular prognostic factors, such as p53 overexpression, which was found to be the only significant molecular prognostic factor for outcome.
Anti-CK7 and anti-CK20 immunohistochemistry is sometimes used to establish a diagnosis of primary lung cancer. We performed a retrospective study on the value of anti-CK7 and anti-CK20 ...immunohistochemistry in 359 biopsies of patients with suspected lung carcinoma in order to assess the usefulness of these antibodies in the evaluation of lung tumors in biopsies. Our results showed TTF-1 positivity in 73.3% of patients. EGFR mutations and ALK rearrangements were significantly different between TTF-1 positive and TTF-1 negative tumors (p < 0.001 and p = 0.023, respectively). Our results show a significant difference (p < 0.001) between TTF-1 positive and TTF-1 negative carcinomas with a median survival of 21.97 months (CI95% = 17.48−30.9 months) and 6.52 months (CI95% = 3.34−10.3 months), respectively. In the group of TTF-1 negative patients, anti-CK7 and CK20 immunohistochemistry was performed in 70 patients and showed CK7+/CK20- staining in 61 patients (87.1%), CK7-/CK20- in 4 patients (5.7%), CK7+/CK20+ in 3 patients (4.3%), and CK7-/CK20- in 2 patients (2.8%). No specific or molecular pattern was found in these groups of CK7/CK20 combinations. In total, this work brings arguments concerning the uselessness of anti-CK7/CK20 immunohistochemistry in the case of suspicion of primary lung cancer in biopsies.
Bronchial dysplasia is the pre-neoplastic lesion recognized for invasive squamous cell carcinoma. The mechanisms leading to invasive squamous cell carcinoma for this lesion are not fully known. ...Programmed Death-Ligand 1 (PD-L1) expression by the bronchial dysplasia neoplastic epithelium might suggest a response to immunotherapy. The objective of this work is to further characterize PD-L1 and CD8 expression in bronchial dysplasia and bronchial metaplasia compared to normal bronchial epithelium. Immunohistochemical analysis of PD-L1 and CD8 staining were characterized in bronchial dysplasia of 24 patients and correlated with clinical data. We also compared PD-L1 expression in dysplasia samples to 30 normal epithelium and 20 samples with squamous bronchial metaplasia. PD-L1 was never expressed in normal epithelium and in metaplastic epithelium whereas 37.5% of patients with bronchial dysplasia were stained by PD-L1 (p < 0.001). PD-L1 expression was not related to the degree of dysplasia or a medical history of invasive squamous cell carcinoma, while CD8 expression and its localization were related to medical history of squamous cell carcinoma (p = 0.044). Our results show that PD-L1 is expressed in roughly one third of patients with bronchial dysplasia and is not expressed in normal and metaplastic epithelium. This suggests that PD-L1 is expressed in preneoplastic lesions of squamous cell carcinoma.
Corneal endothelial cells (ECs) form a monolayer that controls the hydration of the cornea and thus its transparency. Their almost nil proliferative status in humans is responsible, in several ...frequent diseases, for cell pool attrition that leads to irreversible corneal clouding. To screen for candidate genes involved in cell cycle arrest, we studied human ECs subjected to various environments thought to induce different proliferative profiles compared to ECs in vivo. Donor corneas (a few hours after death), organ-cultured (OC) corneas, in vitro confluent and non-confluent primary cultures, and an immortalized EC line were compared to healthy ECs retrieved in the first minutes of corneal grafts. Transcriptional profiles were compared using a cDNA array of 112 key genes of the cell cycle and analysed using Gene Ontology classification; cluster analysis and gene map presentation of the cell cycle regulation pathway were performed by GenMAPP. Results were validated using qRT-PCR on 11 selected genes. We found several transcripts of proteins implicated in cell cycle arrest and not previously reported in human ECs. Early G1-phase arrest effectors and multiple DNA damage-induced cell cycle arrest-associated transcripts were found in vivo and over-represented in OC and in vitro ECs. Though highly proliferative, immortalized ECs also exhibited overexpression of transcripts implicated in cell cycle arrest. These new effectors likely explain the stress-induced premature senescence that characterizes human adult ECs. They are potential targets for triggering and controlling EC proliferation with a view to increasing the cell pool of stored corneas or facilitating mass EC culture for bioengineered endothelial grafts.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
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