Prospective study of the real impact of fusion centered genomic assays in patient management in a national collaborative group: the GETHI-XX-16 study.

PURPOSE: Precision medicine represents a paradigm shift in oncology. Access to genetic testing and targeted therapies is frequently limited. Assays based on DNA sequencing can miss druggable alterations. We aimed to determine the impact of a free access program to RNA tests in patient management. METHODS: We designed a multicenter prospective observational study within the Spanish National Group for Translational Oncology and Rare and Orphan Tumors (GETTHI). Eligible patients were adults with solid cancers that had progressed on standard therapies. Tumor samples were analyzed using two RNA sequencing assays (Trailblaze PharosTM and Archer FusionPlex Solid TumorTM). A central committee evaluated the actionability of genetic alterations and reported the findings to attending physicians, who made the final clinical management decisions. RESULTS: Between November 2016 and April 2019, 395 patients with 41 different tumors across 30 hospitals were included. Molecular analysis revealed actionable genetic alterations in 57 individuals (14.4%). Targeted therapies were advised for 23 and seven received a matched targeted therapy: two lung cancers (EML4-ALK and CD74-ROS1 fusion), three glioblastomas (EGFR point mutations), one oligodendroglioma (FGFR3-TACC3 fusion) and a prostate cancer (SND1-BRAF fusion). The outcomes included two tumor responses, one disease stabilization, one early withdrawal due to toxicity, one progression, and one unknown. CONCLUSION: Despite the growing knowledge of cancer biology and its translation to drug development, the overall impact of personalized treatments remains low. Access to comprehensive molecular tests covering properly all known actionable alterations and programs for a wide access to targeted therapies seem to be critical steps.

Phase II clinical trial assessing the efficacy of enzalutamide in advanced non-resectable granulosa cell ovarian tumors: The GREKO III study (GETHI2016-01).

BACKGROUND: Granulosa cell ovarian tumors (GCT) are orphan disease with limited treatments. Hormone therapy is a potential treatment, due to the overexpression of hormone receptors in most tumors. This study explores the activity of the antiandrogen, enzalutamide, in metastatic cases. METHODS: We designed a phase II clinical trial under the Spanish Collaborative Group for Transversal Oncology and Rare and Orphan Tumors (GETTHI). Eligible participants were adult women with advanced GCT. Primary endpoint was objective response rate. Secondary endpoints included clinical benefit rate, progression-free survival, overall survival, and safety profile. Patients received enzalutamide 160 mg once daily. RESULTS: From April 2018 to March 2020, eighteen patients were screened, and sixteen were included across nine institutions. Median age was 56.4 years (range 45-71), and most were Caucasian (14 cases), one Arabian and one Latin. ECOG performance status was zero in 13 cases (81 %) and one in three (19 %). Six patients (38 %) had previously received hormone therapy as adjuvant treatment or for advanced disease, and 15 (94 %) chemotherapy. Median time from metastasis to study entry was 96 months (range 4.5-198). No objective response was observed, but the clinical benefit rate reached 68.8 % (95 % CI [46 %-91.5 %]). Median progression-free survival was 3.8 months (95 % CI [1.36-6.14]). Median overall survival was not reached, with a median follow-up of 6 months (range 2.2-19). At the time of database closure, 14 patients had discontinued treatment, 13 due to disease progression and one by personal choice. Two deaths attributed to disease progression were recorded. Five grade 3 adverse events were reported, with only one (asthenia) deemed related to the therapy. CONCLUSIONS: Although enzalutamide demonstrated modest activity in GCT, durable stabilization was observed in some cases. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT03464201.

Design of an SMA-Based Actuator for Replicating Normal Gait Patterns in Pediatric Patients with Cerebral Palsy.

Cerebral Palsy refers to a group of incurable motor disorders affecting 0.22% of the global population. Symptoms are managed by physiotherapists, often using rehabilitation robotics. Exoskeletons, offering advantages over conventional therapies, are evolving to be more wearable and biomimetic, requiring new flexible actuators that mimic human tissue. The main objective behind this article is the design of a flexible exosuit based on shape-memory-alloy-based artificial muscles for pediatric patients that replicate the walking cycle pattern in the ankle joint. Thus, four shape-memory-alloy-based actuators were sewn to an exosuit at the desired actuation points and controlled by a two-level controller. The loop is closed through six inertial sensors that estimate the real angular position of both ankles. Different frequencies of actuation have been tested, along with the response of the actuators to different walking cycle patterns. These tests have been performed over long periods of time, comparing the reference created by a reference generator based on pediatric walking patterns and the response measured by the inertial sensors. The results provide important measurements concerning errors, working frequencies and cooling times, proving that this technology could be used in this and similar applications and highlighting its limitations.

Proteomic profiles of peritoneal fluid-derived small extracellular vesicles correlate with patient outcome in ovarian cancer.

Cancer-derived small extracellular vesicles (sEVs) are capable of modifying the tumor microenvironment and promoting tumor progression. Ovarian cancer (OvCa) is a lethal malignancy that preferentially spreads through the abdominal cavity. Thus, the secretion of such vesicles into the peritoneal fluid could be a determinant factor in the dissemination and behavior of this disease. We designed a prospective observational study to assess the impact of peritoneal fluid-derived sEVs (PFD-sEVs) in OvCa clinical outcome. For this purpose, 2 patient cohorts were enrolled: patients with OvCa who underwent a diagnostic or cytoreductive surgery and nononcological patients, who underwent abdominal surgery for benign gynecological conditions and acted as the control group. Systematic extraction of PFD-sEVs from surgical samples enabled us to observe significant quantitative and qualitative differences associated with cancer diagnosis, disease stage, and platinum chemosensitivity. Proteomic profiling of PFD-sEVs led to the identification of molecular pathways and proteins of interest and to the biological validation of S100A4 and STX5. In addition, unsupervised analysis of PFD-sEV proteomic profiles in high-grade serous ovarian carcinomas (HGSOCs) revealed 2 clusters with different outcomes in terms of overall survival. In conclusion, comprehensive characterization of PFD-sEV content provided a prognostic value with potential implications in HGSOC clinical management.

Design and Control of a Soft Knee Exoskeleton for Pediatric Patients at Early Stages of the Walking Learning Process.

Pediatric patients can suffer from different motor disorders that limit their neurological and motor development and hinder their independence. If treated at the very early stages of development, those limitations can be palliated or even removed. However, manual interventions are not completely effective due to the restrictions in terms of time, force, or tracking experienced by the physiotherapists. The knee flexo-extension is crucial for walking and often affected by disorders such as spasticity or lack of force in the posterior chain. This article focuses on the development of a knee exosuit to follow angular trajectories mimicking the maximum and minimum peaks present in the knee flexo-extension profiles of healthy individuals during walking. The proposed exosuit is based on shape memory alloy actuators along with four inertial sensors that close the control loop. The whole device is controlled through a two-level controller and has an hybrid rigid-flexible design to overcome the different issues present in the literature. The device was proven to be feasible for this type of application, with replicable and consistent behavior, reducing the price and weight of existing exosuits and enhancing patient comfort.

Pain assessment and management among adult patients in a gynecology ward: a best practice implementation project.

OBJECTIVES: This project aimed to improve compliance with evidence-based practice in pain assessment and management in a gynecology ward. INTRODUCTION: Effective pain control is important to prevent the negative consequences of pain that is poorly managed. However, it remains undervalued and inadequately treated. Applying evidence-based practices to correctly evaluate and manage pain is essential to improve pain relief. METHODS: This project followed the JBI Evidence Implementation Framework. A baseline audit of 41 women admitted to the gynecology ward was conducted and measured against 5 best practice criteria, along with a patient satisfaction questionnaire. Targeted strategies were then implemented and a follow-up audit was conducted using the same criteria, methods, and sample size as the baseline audit. RESULTS: The baseline audit revealed gaps between current and best practice. Barriers to implementation were identified and strategies to resolve the barriers were designed and implemented (nurse education, informative materials, electronic patient records system improvements). Comprehensive pain assessment, including dynamic and static pain assessment, use of a validated tool, and education provided to patients and carers, improved in the follow-up audit. There was no change in patient satisfaction levels; however, the discrepancy between pain measured by nurses and pain measured by patients was reduced. CONCLUSIONS: The JBI methodology was useful in improving compliance with evidence-based practice criteria. It also facilitated adaptation to new barriers, such as the COVID-19 pandemic. Improving nurses' knowledge of pain assessment can lead to more accurate assessment. Inadequate records systems also made it difficult to record the care that was provided. Subsequent audits will assess sustainability and the project will be escalated to other wards.

Impact of the Identification of Nonhuman Genetic Signatures in the Diagnosis and Management of Carcinoma of Unknown Primary.

This report presents the case of a 62-year-old woman who was diagnosed in 1999 with stage I cervical carcinoma treated by surgical resection. In 2021, she presented to the emergency department with a complaint of predominantly right-sided lower back pain. A CT scan of the lumbosacral region revealed a bone lesion in the L5 vertebra and retroperitoneal lymphadenopathies suggestive of malignancy. Histology of the L5 vertebra biopsy showed a poorly differentiated carcinoma with an inconclusive immunophenotypic profile. Treatment for carcinoma of unknown primary was started with a combination of carboplatin and paclitaxel every 21 days. A genomic study of the biopsy specimen performed on the FoundationOne CDx platform identified a nonhuman genetic signature compatible with HPV. The presence of HPV 18 DNA in the specimen was confirmed by PCR-reverse dot blot, and the immunophenotypic profile was expanded, revealing strong and diffuse p16 expression, thus corroborating the molecular findings. In view of these findings, the case was reclassified as a recurrence of the cervical adenocarcinoma that had been diagnosed and treated 23 years earlier. Based on the new results, and according to first-line cervical carcinoma protocols, bevacizumab at 15 mg/kg every 21 days was added to her chemotherapy regimen. The identification of HPV DNA sequences by next-generation sequencing facilitated the correct diagnosis and led to a modification of the first-line therapeutic approach.

Clinical Experience with Abemaciclib in Patients Previously Treated with Another CDK 4/6 Inhibitor in a Tertiary Hospital: A Case Series Study.

The three approved cyclin-dependent kinase 4/6 (CDK4/6) inhibitors, including abemaciclib, have shown differences in their preclinical, pharmacological, and clinical data. Abemaciclib stands out for its broader target range and more rapid and intense activity. It has demonstrated efficacy as a monotherapy or in combination with tamoxifen in endocrine-refractory metastatic breast cancer (MBC) patients with prior chemotherapy. However, the clinical data on abemaciclib after exposure to previous CDK4/6 inhibitors are limited. In this single-center retrospective case series, we identified all patients who received abemaciclib until February 2022 after experiencing documented progression on palbociclib or ribociclib. The safety profile and clinical outcomes of abemaciclib treatment in this specific patient cohort were evaluated. Eleven patients were included in this retrospective case series, nine receiving abemaciclib with tamoxifen. Eight patients had visceral involvement, and the median age was 69 (ranging from 42 to 84). The median time from the end of prior CDK4/6 inhibitor treatment to abemaciclib initiation was 17.5 months (ranging from 3 to 41 months). Patients had undergone a median of three prior therapies (ranging from 1 to 7), including chemotherapy in 54.5% of cases. The median follow-up time was six months (ranging from 1 to 22 months). The median progression-free survival (PFS) was 8 months (95% CI 3.9-12). Five patients continued abemaciclib treatment, and one patient with liver metastases achieved a complete hepatic response. The most common adverse events were diarrhea (72.7%, no grade ≥ 3) and asthenia (27.3%, no grade ≥ 3). Our preliminary findings suggest that abemaciclib could be an effective and safe treatment option for MBC patients who have previously received palbociclib or ribociclib.

PBRM1 and KDM5C cooperate to define high-angiogenesis tumors and increased antiangiogenic response in renal cancer.

Vascular endothelial growth factor receptor tyrosine kinase inhibitors (VEGFR-TKIs) are key antiangiogenic drugs for renal cancer treatment. While Von Hippel-Lindau dysfunction constitutes the base for VEGFR-TKIs sensitivity, the role for individual and concurrent mutations in the genes encoding for the chromatin remodelers Polybromo-1 (PBRM1) and Lysine Demethylase 5C (KDM5C) is poorly understood. Here, we analyzed the tumor mutational and expression profiles of 155 unselected clear cell RCC (ccRCC) cases treated with first-line VEGFR-TKIs and the ccRCC cases of IMmotion151 trial were used for validation. We found that concurrent PBRM1 and KDM5C (PBRM1&KDM5C) mutations occurred in 4-9% of cases and were enriched in Memorial Sloan Kettering Cancer Center favorable-risk patients. In our cohort, tumors only mutated in PBRM1 or concurrently mutated in PBRM1 and KDM5C had increased angiogenesis (P=0.0068 and 0.039; respectively), and tumors only mutated in KDM5C showed a similar trend. Best response to VEGFR-TKIs corresponded to PBRM1&KDM5C mutated cases, followed by those mutated only in KDM5C or only in PBRM1 (P=0.050, 0.040 and 0.027 versus non-mutated cases, respectively), with a trend for longer progression free survival (PFS) in the group with only PBRM1 mutated (HR=0.64; P=0.059). Validation in the IMmotion151 trial revealed a similar correlation with increased angiogenesis and the PFS of patients in the VEGFR-TKI-arm was the longest in PBRM1&KDM5C mutated cases, intermediate for only PBRM1 or only KDM5C mutated patients and the shortest in non-mutated cases (P=0.009 and 0.025, for PBRM1&KDM5C and PBRM1 versus non-mutated cases). In conclusion, somatic PBRM1 and KDM5C mutations are common in patients with metastatic ccRCC and likely cooperate increasing tumor angiogenesis and VEGFR-TKI-based antiangiogenic therapy benefit.

Randomized phase II clinical trial of ruxolitinib plus simvastatin in COVID19 clinical outcome and cytokine evolution.

Background: Managing the inflammatory response to SARS-Cov-2 could prevent respiratory insufficiency. Cytokine profiles could identify cases at risk of severe disease. Methods: We designed a randomized phase II clinical trial to determine whether the combination of ruxolitinib (5 mg twice a day for 7 days followed by 10 mg BID for 7 days) plus simvastatin (40 mg once a day for 14 days), could reduce the incidence of respiratory insufficiency in COVID-19. 48 cytokines were correlated with clinical outcome. Participants: Patients admitted due to COVID-19 infection with mild disease. Results: Up to 92 were included. Mean age was 64 ± 17, and 28 (30%) were female. 11 (22%) patients in the control arm and 6 (12%) in the experimental arm reached an OSCI grade of 5 or higher (p = 0.29). Unsupervised analysis of cytokines detected two clusters (CL-1 and CL-2). CL-1 presented a higher risk of clinical deterioration vs CL-2 (13 [33%] vs 2 [6%] cases, p = 0.009) and death (5 [11%] vs 0 cases, p = 0.059). Supervised Machine Learning (ML) analysis led to a model that predicted patient deterioration 48h before occurrence with a 85% accuracy. Conclusions: Ruxolitinib plus simvastatin did not impact the outcome of COVID-19. Cytokine profiling identified patients at risk of severe COVID-19 and predicted clinical deterioration. Trial registration: https://clinicaltrials.gov/, identifier NCT04348695.

Open-label phase II clinical trial of ketoconazole as CYP17 inhibitor in metastatic or advanced non-resectable granulosa cell ovarian tumors: the GREKO (GRanulosa Et KetOconazole) trial, GETHI 2011-03.

BACKGROUND: Granulosa cell ovarian tumor (GCT) is characterized by a pathognomonic mutation in the FOXL2 gene (402 C > G) that leads to an overactivation of steroidogenesis. CYP17 is a key enzyme in such process and can be inhibited by ketoconazole. METHODS: We designed a phase II clinical trial to assess the efficacy of ketoconazole in advanced GCT and conducted several in vitro studies to support the clinical findings. RESULTS: From October 1st 2012 to January 31st 2014, six evaluable patients were recruited in ten hospitals of the Spanish Group for Transversal Oncology and Research in Orphan and Infrequent Tumors" (GETTHI). FOXL2 (402C > G) mutation was confirmed in three; two cases were wild type and it could not be assessed in one. No objective response by RECIST was observed, but five cases achieved stable disease longer than 12 months. Median progression-free survival was 14.06 months (CI 95% 5.43-22.69) for the whole study population (3.38 and 13.47 months for wild-type cases and 14.06, 20.67 and 26.51 for those with confirmed FOXL2 mutation). Median overall survival was 22·99 months (CI 95% 8.99-36.99). In vitro assays confirmed the activity of ketoconazole in this tumor and suggested potential synergisms with other hormone therapies. CONCLUSION: Ketoconazole has shown activity in advanced GCT in clinical and in vitro studies. Based on these data, an orphan designation was granted by the European Medicines Agency for ketoconazole in GCT (EU/3/17/1857). GOV IDENTIFIER: NCT01584297.

MEK inhibitor sensitivity in BRAF fusion-driven prostate cancer.

PURPOSE: The identification of subpopulations harboring druggable targets has become a major step forward in the subclassification of solid tumors into small groups suitable for specific therapies. BRAF fusions represent a paradigm of uncommon and targetable oncogenic events and have been widely correlated to the development of specific malignancies. However, they are only present in a limited frequency across most common tumor types. At this regard, we performed a genomic screening aimed to identifying rare variants associated to advanced prostate cancer development. METHODS: Tumoral tissue genomic screening of 41 patients developing advanced prostate cancer was performed at our center as part of the GETHI XX study. The project, sponsored by the Spanish Collaborative Group in Rare Cancers (GETHI), aims to analyze the molecular background of rare tumors and to discover unfrequent molecular variants in common tumors. RESULTS: Here we present the clinical outcome and an in-deep molecular analysis performed in a case harboring a SND1-BRAF fusion gene. The identification of such rearrangement in a patient refractory to standard therapies led to the administration of trametinib (MEK inhibitor). Despite unsensitive to standard therapies, the patient achieved a dramatic response to trametinib. A comprehensive study of the tumor demonstrated this event to be a trunk alteration with higher expression of MEK in areas of tumor invasion. CONCLUSIONS: Our study describes the patient-driven discovery of the first BRAF fusion-driven prostate cancer effectively treated with trametinib. Consequently, MAPK pathway activation could define a new subtype of prostate cancer susceptible to a tailored management.

Case Report: Bocavirus Infection Radiologically Resembling a Congestive Heart Failure in a Patient with Metastatic Castration-Resistant Prostate Cancer.

Background: Human bocavirus (HBoV) is a viral pathogen from the genus Bocaparvovirus (family Parvoviridae, subfamily Parvovirinae) discovered in 2005. Most of available literature is about HBoV in children and adults with hematological malignancies and in otherwise healthy children with respiratory infections. Information regarding infection in the adult population with solid tumors is scarce. Case Report: We report the case of a 51-year-old male with metastatic castration resistant prostate cancer undergoing chemotherapy treatment who presented with fever, dyspnea, dry cough, and pleuritic pain. Imaging techniques showed signs of congestive heart failure. Symptoms, laboratory tests and echocardiography revealed a more probable infectious etiology. Antibiotic therapy was started. A polymerase chain reaction (PCR) test of nasopharyngeal exudate for respiratory viruses was positive for HBoV. The rest of the microbiological tests were negative. Bronchoalveolar lavage (BAL) was performed. Bacterial culture of BAL was negative while respiratory virus PCR confirmed positivity for HBoV. Antibiotic therapy was discontinued. The patient gradually recovered. Conclusions: Emerging infectious diseases are a notorious threat for immunocompromised populations such as solid tumor patients. This case is unique because to our knowledge this is the first case report article of HBoV in a solid tumor patient and because imaging techniques exhibited signs of congestive heart failure that did not correlate with the rest of the tests. It shows that unusual pathogens should be considered when managing serious clinical complications with uncommon presentations in cancer patients. Notable diagnostic efforts should be made to reach a diagnosis in these cases.

Evidence of exhausted lymphocytes after the third anti-SARS-CoV-2 vaccine dose in cancer patients.

Introduction: Evidence is scant regarding the long-term humoral and cellular responses Q7 triggered by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) mRNA vaccines in cancer patients after repeated booster doses. The possibility of T-cell exhaustion following these booster doses in this population has not yet been fully studied and remains uncertain. Methods: In this single-center prospective observational study, we explored the specific humoral and cellular response to S1 antigen in 36 patients with solid malignancies at baseline, and after the second and third doses of the mRNA-1273 vaccine. Results: A dual behavior was observed: 24 (66.7%) patients showed partial specific IFN-γ response after the second dose that was further enhanced after the third dose; and 11 (30.5%) already showed an optimal response after the second dose and experienced a marked fall-off of specific IFN-γ production after the third (4 patients negativization), which might suggest T cell exhaustion due to repetitive priming to the same antigen. One (2.8%) patient had persistently negative responses after all three doses. Seroconversion occurred in all patients after the second dose. We then studied circulating exhausted CD8+ T-cells in 4 patients from each of the two response patterns, those with increase and those with decrease in cellular response after the third booster. The patients with decreased cellular response after the booster had a higher expression of PD1+CD8+ and CD57+PD1+CD8+ exhausted T cells compared with those with an increased cellular response both in vivo and in vitro. The proportion of PD1+CD8+ and CD57+PD1+CD8+ exhausted T cells inversely correlated with IFN-γ production. Discussion: Our preliminary data show that the two-dose SARS-CoV-2 vaccine regimen was beneficial in all cancer patients of our study. An additional booster seems to be beneficial in suboptimal vaccine seroconverters, in contrast to maximal responders that might develop exhaustion. Our data should be interpreted with caution given the small sample size and highlight the urgent need to validate our results in other independent and larger cohorts. Altogether, our data support the relevance of immunological functional studies to personalize preventive and treatment decisions in cancer patients.

Adverse drug reactions to the three doses of the severe acute respiratory syndrome coronavirus 2 (SARS-COV-2) mRNA-1273 vaccine in a cohort of cancer patients under active treatment of a tertiary hospital in Madrid, Spain.

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccines efficacy and safety have been tested in phase 3 studies in which cancer patients were not included or were underrepresented. Methods: The objective of this study is to evaluate the safety profile of the mRNA-1273 vaccine across cancer patients and its relationship to patients' demographics. This retrospective cohort study included patients 18-years or older with solid malignancies receiving active treatment in our hospital who had received the three-dose schedule of the mRNA9 1273 vaccine and whose side effects after each dose were recorded. Patient electronic medical records were reviewed retrospectively to collect data between April 19, 2021, and December 31, 2021. Patients with documented previous infection by SARS-Cov-2 were excluded from the study. Results: A total of 93 patients met the inclusion criteria. Local adverse drug reactions (ADRs) were reported more frequently after the first and second dose than after the third (41.9%, 43% and 31.1% of the patients respectively), while systemic ADRs followed the opposite pattern (16.1%, 34.4% and 52.6% of the patients respectively). We found a statistically significant association between sex and systemic ADRs after the third dose. Cochran-Armitage test showed a statistically significant linear trend, p = 0.012, with a higher Eastern Cooperative Oncology Group (ECOG) score associated with a lower proportion of patients suffering from systemic side effects. A logistic regression showed that women had 5.79 times higher odds to exhibit systemic ADRs after the third dose (p=0.01) compared to males. Increasing age was associated with a decreased likelihood of exhibiting ADRs (p=0.016). Conclusion: The mRNA-1273 vaccine shows a tolerable safety profile. The likelihood of ADRs appears to be associated with gender and age. Its association with ECOG scores is less evident. Further studies are needed to elucidate this data in cancer patients.

Identification of tumors with NRG1 rearrangement, including a novel putative pathogenic UNC5D-NRG1 gene fusion in prostate cancer by data-drilling a de-identified tumor database.

The fusion genes containing neuregulin-1 (NRG1) are newly described potentially actionable oncogenic drivers. Initial clinical trials have shown a positive response to targeted treatment in some cases of NRG1 rearranged lung adenocarcinoma, cholangiocarcinoma, and pancreatic carcinoma. The cost-effective large scale identification of NRG1 rearranged tumors is an open question. We have tested a data-drilling approach by performing a retrospective assessment of a de-identified molecular profiling database of 3263 tumors submitted for fusion testing. Gene fusion detection was performed by RNA-based targeted next-generation sequencing using the Archer Fusion Plex kits for Illumina (ArcherDX Inc., Boulder, CO). Novel fusion transcripts were confirmed by a custom-designed RT-PCR. Also, the aberrant expression of CK20 was studied immunohistochemically. The frequency of NRG1 rearranged tumors was 0.2% (7/3263). The most common histologic type was lung adenocarcinoma (n = 5). Also, renal carcinoma (n = 1) and prostatic adenocarcinoma (n = 1) were found. Identified fusion partners were of a wide range (CD74, SDC4, TNC, VAMP2, UNC5D), with CD74, SDC4 being found twice. The UNC5D is a novel fusion partner identified in prostate adenocarcinoma. There was no co-occurrence with the other tested fusions nor KRAS, BRAF, and the other gene mutations specified in the applied gene panels. Immunohistochemically, the focal expression of CK20 was present in 2 lung adenocarcinomas. We believe it should be considered as an incidental finding. In conclusion, the overall frequency of tumors with NRG1 fusion was 0.2%. All tumors were carcinomas. We confirm (invasive mucinous) lung adenocarcinoma as being the most frequent tumor presenting NRG1 fusion. Herein novel putative pathogenic gene fusion UNC5D-NRG1 is described. The potential role of immunohistochemistry in tumor identification should be further addressed.

Expression of enamel proteins in human dental pulp stem cells by the effect of extracellular matrix / Expresión de proteínas de esmalte en células madre de pulpa dental humana por el efecto de matriz extracelular

SUMMARY: Mesenchymal stem cells are present in adult tissues such as the human dental pulp. They are pluripotent and can differentiate into various specialized cell types in vitro through appropriate stimuli. Ameloblasts produce human tooth enamel only during embryonic development before tooth eruption, so endogenous regeneration is not possible. Various efforts have been aimed at generating natural or artificial substitutes for dental enamel with properties similar to the specific components of said tissue. The purpose of this study was to induce human dental pulp stem cells to produce enamel proteins using extracellular matrix derived from the rat tail tendon and pigskin. Primary cultures of human dental pulp stem cells were established and characterized by RT-PCR and immunofluorescence, using mesenchymal cell markers such as CD14, CD40, CD44, CD105, and STRO-1. The cells were then incubated with the extracellular matrix for fourteen days and labeled with specific antibodies to detect the expression of dental enamel proteins such as amelogenin, ameloblastin, enamelisin, tuftelin, and parvalbumin, characteristics of the phenotype of ameloblasts. This work demonstrated a positive effect of the extracellular matrix to induce the expression of enamel proteins in the stem cells of the human dental pulp.

RESUMEN: Las células madre mesenquimales están presentes en los tejidos adultos como la pulpa dental humana. Son pluripotentes y pueden diferenciarse en varios tipos de células especializadas in vitro a través de estímulos adecuados. Los ameloblastos producen esmalte dental humano sólo durante el desarrollo embrionario antes de la erupción dental, por lo que no es posible su regeneración endógena. Varios esfuerzos se han orientado a generar sustitutos naturales o artificiales de esmalte dental con propiedades similares a los componentes específicos de este tejido. El propósito de este estudio fue inducir células madre de pulpa dental humana para producir proteínas del esmalte dental a través del estímulo de matriz extracelular derivada del tendón de la cola de rata y piel de cerdo. Se establecieron cultivos primarios de células madre de pulpa dental humana y se caracterizaron por RT-PCR e inmunofluorescencia utilizando marcadores de células mesenquimales como CD14, CD40, CD44, CD105 y STRO-1. Posteriormente, las células se incubaron con matriz extracelular durante un período de catorce días y se marcaron con anticuerpos específicos para detectar la expresión de proteínas de esmalte dental como amelogenina, ameloblastina, enamelisina, tuftelina y parvalbúmina, las cuales son características del fenotipo de ameloblastos. Este trabajo demostró el efecto positivo que tiene el empleo de la matriz extracelular para inducir la expresión de proteínas de esmalte en las células pluripotenciales de la pulpa dental humana.

Imaging of Endoplasmic Reticulum Ca2+ in the Intact Pituitary Gland of Transgenic Mice Expressing a Low Affinity Ca2+ Indicator.

The adenohypophysis contains five secretory cell types (somatotrophs, lactotrophs, thyrotrophs, corticotrophs, and gonadotrophs), each secreting a different hormone, and controlled by different hypothalamic releasing hormones (HRHs). Exocytic secretion is regulated by cytosolic Ca2+ signals ([Ca2+]C), which can be generated either by Ca2+ entry through the plasma membrane and/or by Ca2+ release from the endoplasmic reticulum (ER). In addition, Ca2+ entry signals can eventually be amplified by ER release via calcium-induced calcium release (CICR). We have investigated the contribution of ER Ca2+ release to the action of physiological agonists in pituitary gland. Changes of [Ca2+] in the ER ([Ca2+]ER) were measured with the genetically encoded low-affinity Ca2+ sensor GAP3 targeted to the ER. We used a transgenic mouse strain that expressed erGAP3 driven by a ubiquitous promoter. Virtually all the pituitary cells were positive for the sensor. In order to mimick the physiological environment, intact pituitary glands or acute slices from the transgenic mouse were used to image [Ca2+]ER. [Ca2+]C was measured simultaneously with Rhod-2. Luteinizing hormone-releasing hormone (LHRH) or thyrotropin releasing hormone (TRH), two agonists known to elicit intracellular Ca2+ mobilization, provoked robust decreases of [Ca2+]ER and concomitant rises of [Ca2+]C. A smaller fraction of cells responded to thyrotropin releasing hormone (TRH). By contrast, depolarization with high K+ triggered a rise of [Ca2+]C without a decrease of [Ca2+]ER, indicating that the calcium-induced calcium-release (CICR) via ryanodine receptor amplification mechanism is not present in these cells. Our results show the potential of transgenic ER Ca2+ indicators as novel tools to explore intraorganellar Ca2+ dynamics in pituitary gland in situ.

Apraxia de fala na infância: para além das questões fonéticas e fonológicas / Speech apraxia in childhood: beyond phonetic and phonological issues / Apraxia de habla en la infancia: mas allá de las cuestiones fonéticas y fonológicas

Este artigo propõe a análise e a discussão, com base na Neurolinguística Discursiva, dos aspectos neurofisiológicos, psíquicos, cognitivos, linguísticos e sociais pertinentes ao processo de aquisição de linguagem de crianças ouvintes com "Apraxia de Fala". A partir da revisão bibliográfica da área, recobrindo avaliação e tratamento fonoaudiológico tradicionalmente destinados àquelas crianças, ampliamos, com base em uma abordagem discursiva de língua e linguagem, nossas reflexões sobre a criança com apraxia. Observamos, assim, que, para além das alterações fonoarticulatórias, estão envolvidas na criança questões importantes, pertinentes e referentes ao seu período de balbucio, à conformação neurofuncional da memória dos gestos articulatórios e aos processos constitutivos do diálogo enquanto matriz de significação.

This article proposes the analysis and discussion, based on the Discursive Neurolinguistics (ND), of the neurophysiological, psychic, cognitive, linguistic and social aspects pertinent to the process of language acquisition of hearing children with "Speech Apraxia". Based on the bibliographical review of the area, covering the evaluation and speech therapy traditionally intended for those children, we expanded, based on a discursive approach of language, our reflections on the child with apraxia. Thus, in addition to the phonoarticulatory alterations, the child is involved in important issues related to his or her babbling period, the neurofunctional conformation of the articulatory gestures memory and the constitutive processes of dialogue as a matrix of signification.

En este artículo se propone el análisis y la discusión, bajo la base de la Neurolinguística Discursiva, de los aspectos neurofisiológicos, psíquicos, cognitivos, lingüísticos y sociales pertinentes al proceso de adquisición del lenguaje de niños oyentes con "Apraxia delHabla". A partir de la revisión bibliográfica del áreasobre evaluación y tratamiento fonoaudiológico tradicionalmente destinados a aquellos niños, ampliamos, con base en un abordaje discursivo de la lengua y del lenguaje, nuestras reflexiones sobre el niño con apraxia. Observamos que, mas allá de las alteraciones fonoarticulatorias, están involucradas en el niño cuestiones importantes, pertinentes y referentes a su período de balbuceo, a la conformación neurofuncional de la memoria de los gestos articulatorios ya los procesos constitutivos del diálogo como matriz de significación.