Immune checkpoint therapy responders display early clonal expansion of tumor infiltrating lymphocytes.

Immune checkpoint therapy (ICT) causes durable tumour responses in a subgroup of patients, but it is not well known how T cell receptor beta (TCRß) repertoire dynamics contribute to the therapeutic response. Using murine models that exclude variation in host genetics, environmental factors and tumour mutation burden, limiting variation between animals to naturally diverse TCRß repertoires, we applied TCRseq, single cell RNAseq and flow cytometry to study TCRß repertoire dynamics in ICT responders and non-responders. Increased oligoclonal expansion of TCRß clonotypes was observed in responding tumours. Machine learning identified TCRß CDR3 signatures unique to each tumour model, and signatures associated with ICT response at various timepoints before or during ICT. Clonally expanded CD8+ T cells in responding tumours post ICT displayed effector T cell gene signatures and phenotype. An early burst of clonal expansion during ICT is associated with response, and we report unique dynamics in TCRß signatures associated with ICT response.

Gliomas in children and adolescents: investigation of molecular alterations with a potential prognostic and therapeutic impact.

PURPOSE: Gliomas represent the most frequent central nervous system (CNS) tumors in children and adolescents. However, therapeutic strategies for these patients, based on tumor molecular profile, are still limited compared to the wide range of treatment options for the adult population. We investigated molecular alterations, with a potential prognostic marker and therapeutic target in gliomas of childhood and adolescence using the next-generation sequencing (NGS) strategy. METHODS: We selected 95 samples with initial diagnosis of glioma from patients treated at Pediatric Oncology Institute-GRAACC/UNIFESP. All samples were categorized according to the 2021 World Health Organization Classification of Tumors of the CNS, which included 39 low-grade gliomas (LGGs) and 56 high-grade gliomas (HGGs). Four HGG samples were classified as congenital glioblastoma (cGBM). NGS was performed to identify somatic genetic variants in tumor samples using the Oncomine Childhood Cancer Research Assay® (OCCRA®) panel, from Thermo Fisher Scientific®. RESULTS: Genetic variants were identified in 76 of 95 (80%) tumors. In HGGs, the most common molecular alteration detected was H3F3A c.83A > T variant (H3.3 K27M) and co-occurring mutations in ATRX, TP53, PDGFRA, MET, and MYC genes were also frequently observed. One HGG sample was reclassified as supratentorial ependymoma ZFTA-fusion positive after NGS was performed. In LGGs, four KIAA1549-BRAF fusion transcripts were detected and this alteration was the most recurrent genetic event and favorable prognostic factor identified. Additionally, genetic variants in ALK and NTRK genes, which provide potential targets for therapy with Food and Drug Administration-approved drugs, were identified in two different cases of cGBM that were classified as infant-type hemispheric glioma, a newly recognized subgroup of pediatric HGG. CONCLUSION: Molecular profiling by the OCCRA® panel comprehensively addressed the most relevant genetic variants in gliomas of childhood and adolescence, as these tumors have specific patterns of molecular alterations, outcomes, and effectiveness to therapies.

Molecular profiling of pediatric and adolescent ependymomas: identification of genetic variants using a next-generation sequencing panel.

PURPOSE: Ependymoma (EPN) accounts for approximately 10% of all primary central nervous system (CNS) tumors in children and in most cases, chemotherapy is ineffective and treatment remains challenging. We investigated molecular alterations, with a potential prognostic marker and therapeutic target in EPNs of childhood and adolescence, using a next-generation sequencing (NGS) panel specific for pediatric neoplasms. METHODS: We selected 61 samples with initial diagnosis of EPN from patients treated at Pediatric Oncology Institute-GRAACC/UNIFESP. All samples were divided according to the anatomical compartment of the CNS - 42 posterior fossa (PF), 14 supratentorial (ST), and five spinal (SP). NGS was performed to identify somatic genetic variants in tumor samples using the Oncomine Childhood Cancer Research Assay® (OCCRA®) panel, from Thermo Fisher Scientific®. RESULTS: Genetic variants were identified in 24 of 61 (39.3%) tumors and over 90% of all variants were pathogenic or likely pathogenic. The most commonly variants detected were in CIC, ASXL1, and JAK2 genes and have not been reported in EPN yet. MN1-BEND2 fusion, alteration recently described in a new CNS tumor type, was identified in one ST sample that was reclassified as astroblastoma. Additionally, YAP1-MAMLD1 fusion, a rare event associated with good outcome in ST-EPN, was observed in two patients diagnosed under 2 years old. CONCLUSIONS: Molecular profiling by the OCCRA® panel showed novel alterations in pediatric and adolescent EPNs, which highlights the clinical importance in identifying genetic variants for patients' prognosis and therapeutic orientation.

Abnormal spindle-like microcephaly-associated (ASPM) gene expression in posterior fossa brain tumors of childhood and adolescence.

PURPOSE: In neurogenesis, ASPM (abnormal spindle-like microcephaly-associated) gene is expressed mainly in the ventricular zone of posterior fossa and is the major determinant in the cerebral cortex. Besides its role in embryonic development, ASPM overexpression promotes tumor growth, including central nervous system (CNS) tumors. This study aims to investigate ASPM expression levels in most frequent posterior fossa brain tumors of childhood and adolescence: medulloblastoma (MB), ependymoma (EPN), and astrocytoma (AS), correlating them with clinicopathological characteristics and tumor solid portion size. METHODS: Quantitative reverse transcription (qRT-PCR) is used to quantify ASPM mRNA levels in 80 pre-treatment tumor samples: 28 MB, 22 EPN, and 30 AS. The tumor solid portion size was determined by IOP-GRAACC Diagnostic Imaging Center. We correlated these findings with clinicopathological characteristics and tumor solid portion size. RESULTS: Our results demonstrated that ASPM gene was overexpressed in MB (p = 0.007) and EPN (p = 0.0260) samples. ASPM high expression was significantly associated to MB samples from patients with worse overall survival (p = 0.0123) and death due to disease progression (p = 0.0039). Interestingly, two patients with AS progressed toward higher grade showed ASPM overexpression (p = 0.0046). No correlation was found between the tumor solid portion size and ASPM expression levels in MB (p = 0.1154 and r = - 0.4825) and EPN (p = 0.1108 and r = - 0.3495) samples. CONCLUSION: Taking in account that ASPM gene has several functions to support cell proliferation, as mitotic defects and premature differentiation, we suggest that its overexpression, presumably, plays a critical role in disease progression of posterior fossa brain tumors of childhood and adolescence.

Inspiratory muscle function in asthmatic and healthy subjects: influence of age, nutrition and physical activity.

OBJECTIVE: To compare inspiratory muscle function (strength and endurance) between asthmatics and healthy controls, and the influence of age, nutritional status and physical activity on them. METHODS: This is a cross-sectional study. Asthmatic and healthy subjects, aged 6 to 18 years old, recruited from two public schools in Southern Brazil were included in the study. Asthmatic subjects were selected using the criteria presented by the International Study on Asthma and Allergies in Children and control subjects based on the absence of respiratory symptoms. Anthropometric data was measured, body mass index calculated and subjects classified as normal weight, overweight or obese. Physical activity levels, maximum inspiratory pressure (MIP) and inspiratory muscle endurance (IME) were also evaluated. RESULTS: A total of 314 participants were included, separated into control group (181) and asthmatics (133), with a total mean age of 11 years. When both groups were compared, there were no significant differences in both MIP and IME. However, when groups were analyzed subdivided in children and adolescents, IME was significantly reduced (p = 0.003) in asthmatic adolescents. Indeed, when groups were also stratified considering the nutritional status, IME showed a reduction in asthmatic adolescents with overweight (p = 0.042) and obesity (p = 0.041) when compared to healthy controls. No effects of physical activity levels between groups were found. CONCLUSIONS: Results demonstrate a reduction in the IME in asthmatic adolescents with overweight and obesity, indicating an association between asthma, nutritional status and respiratory muscle function.

Peri-implantite: etiologia e tratamento / Periimplantitis: etiology and treatment

A peri-implantite tem sido descrita como uma alteração patológica dos tecidos ao redor dos implantes osseointegrados, sendo a microbiota e o trauma oclusal considerados seus principais fatores etiológicos. A profundidade de sondagem, o sangramento, a mucosa queratinizada, o fluxo do fluido sulcular e o infiltrado inflamatório servem como parâmetros para avaliar a saúde peri-implantar. O tratamento da peri-implantite inclui debridamento mecânico, instrução de higiene oral, destoxificação da superfície implantar, uso de antimicrobianos e terapias ressectivas e regenerativas. Uma correta intervenção parece ser necessária para a resolução da lesão peri-implantar.

The peri-implantitis has been described as a pathological alteration of tissues around dental implants, and the microbiota and occlusal trauma considered its main etiological factors. The probing depth, bleeding, keratinized tissue, the flow of sulcular fluid and inflammatory infiltrate serve as parameters to evaluate the peri-implant health. Treatment of peri-implant includes mechanical debridement, oral hygiene instruction, surface detoxification deploy, use of antibiotics and resective and regenerative therapies. A correct intervention seems necessary for the resolution of the peri-implant injury.

A new method for fixing Biomphalaria glabrata for histologic studies, using shell perforation

A new technique for fixation of Biomphalaria glabrata for histologic studies is described. It consists in performing several external holes in the shell, before placing the entire snail into the fixative. It is a very practical and quick procedure that showed excellent results when compared to the usual techniques