A multinational Delphi consensus to end the COVID-19 public health threat.

Despite notable scientific and medical advances, broader political, socioeconomic and behavioural factors continue to undercut the response to the COVID-19 pandemic1,2. Here we convened, as part of this Delphi study, a diverse, multidisciplinary panel of 386 academic, health, non-governmental organization, government and other experts in COVID-19 response from 112 countries and territories to recommend specific actions to end this persistent global threat to public health. The panel developed a set of 41 consensus statements and 57 recommendations to governments, health systems, industry and other key stakeholders across six domains: communication; health systems; vaccination; prevention; treatment and care; and inequities. In the wake of nearly three years of fragmented global and national responses, it is instructive to note that three of the highest-ranked recommendations call for the adoption of whole-of-society and whole-of-government approaches1, while maintaining proven prevention measures using a vaccines-plus approach2 that employs a range of public health and financial support measures to complement vaccination. Other recommendations with at least 99% combined agreement advise governments and other stakeholders to improve communication, rebuild public trust and engage communities3 in the management of pandemic responses. The findings of the study, which have been further endorsed by 184 organizations globally, include points of unanimous agreement, as well as six recommendations with >5% disagreement, that provide health and social policy actions to address inadequacies in the pandemic response and help to bring this public health threat to an end.

Brazil's unified health system: the first 30 years and prospects for the future.

In 1988, the Brazilian Constitution defined health as a universal right and a state responsibility. Progress towards universal health coverage in Brazil has been achieved through a unified health system (Sistema Único de Saúde [SUS]), created in 1990. With successes and setbacks in the implementation of health programmes and the organisation of its health system, Brazil has achieved nearly universal access to health-care services for the population. The trajectory of the development and expansion of the SUS offers valuable lessons on how to scale universal health coverage in a highly unequal country with relatively low resources allocated to health-care services by the government compared with that in middle-income and high-income countries. Analysis of the past 30 years since the inception of the SUS shows that innovations extend beyond the development of new models of care and highlights the importance of establishing political, legal, organisational, and management-related structures, with clearly defined roles for both the federal and local governments in the governance, planning, financing, and provision of health-care services. The expansion of the SUS has allowed Brazil to rapidly address the changing health needs of the population, with dramatic upscaling of health service coverage in just three decades. However, despite its successes, analysis of future scenarios suggests the urgent need to address lingering geographical inequalities, insufficient funding, and suboptimal private sector-public sector collaboration. Fiscal policies implemented in 2016 ushered in austerity measures that, alongside the new environmental, educational, and health policies of the Brazilian government, could reverse the hard-earned achievements of the SUS and threaten its sustainability and ability to fulfil its constitutional mandate of providing health care for all.

Epithelioid Rhabdomyosarcoma: Report of a Cutaneous Case and Literature Review of a Recently Described Variant of Rhabdomyosarcoma.

Rhabdomyosarcoma affects mainly pediatric patients and is currently classified into 4 categories: embryonal, alveolar, pleomorphic, and spindle cell/sclerosing. Epithelioid rhabdomyosarcoma is a recently described variant of rhabdomyosarcoma in which primary cutaneous presentation is infrequent. In this brief report, we describe a rare case of epithelioid rhabdomyosarcoma in an 81-year-old man, presenting as a skin lesion in the neck, which increased in size in 1 month. After imaging evaluation, a solid cervical mass was discovered. A biopsy was performed, and the diagnosis of epithelioid rhabdomyosarcoma was rendered. The patient died due to rapid progression of the tumor. To make an accurate diagnosis and ensure appropriate patient management, it is necessary to be aware of this variant and use proper immunohistochemical stains when facing an epithelioid malignancy, expanding the differential diagnosis of epithelioid neoplasms.

Reproducibility of the NanoString 22-gene molecular subgroup assay for improved prognostic prediction of medulloblastoma.

Medulloblastoma is the most frequent malignant brain tumor in children. Four medulloblastoma molecular subgroups, MBSHH , MBWNT , MBGRP3 and MBGRP4 , have been identified by integrated high-throughput platforms. Recently, a 22-gene panel NanoString-based assay was developed for medulloblastoma molecular subgrouping, but the robustness of this assay has not been widely evaluated. Mutations in the gene for human telomerase reverse transcriptase (hTERT) have been found in medulloblastomas and are associated with distinct molecular subtypes. This study aimed to implement the 22-gene panel in a Brazilian context, and to associate the molecular profile with patients' clinical-pathological features. Formalin-fixed, paraffin-embedded (FFPE) medulloblastoma samples (n = 104) from three Brazilian centers were evaluated. Expression profiling of the 22-gene panel was performed by NanoString and a Canadian series (n = 240) was applied for training phase. hTERT mutations were analyzed by PCR followed by direct Sanger sequencing and the molecular profile was associated with patients' clinicopathological features. Overall, 65% of the patients were male, average age at diagnosis was 18 years and 7% of the patients presented metastasis at diagnosis. The molecular classification was attained in 100% of the cases, with the following frequencies: MBSHH (n = 51), MBWNT (n = 19), MBGRP4 (n = 19) and MBGRP3 (n = 15). The MBSHH and MBGRP3 subgroups were associated with older and younger patients, respectively. The MBGRP4 subgroup exhibited the lowest 5-year cancer-specific overall survival (OS), yet in the multivariate analysis, only metastasis at diagnosis and surgical resection were associated with OS. hTERT mutations were detected in 29% of the cases and were associated with older patients, increased hTERT expression and MBSHH subgroup. The 22-gene panel provides a reproducible assay for molecular subgrouping of medulloblastoma FFPE samples in a routine setting and is well-suited for future clinical trials.

[Primary health care in the Region of the Americas 40 years after the Alma-Ata DeclarationAtenção primária à saúde na Região das Américas 40 anos após a Declaração de Alma-Ata]. / La atención primaria de salud en la Región de las Américas a 40 años de la Declaración de Alma-Ata.

This article reviews the evolution of regional proposals and agreements from the Declaration of Alma-Ata (1978) to the Universal Health Strategy, highlighting how the core tenets of the primary health care strategy have come to be reflected in proposals to strengthen the primary level of care and establish integrated health services networks. Contextual aspects of implementing the strategy within the framework of complex national scenarios are also noted, through a review of some of the milestones of the last 40 years. Factors that hinder implementation of primary health care are described, as well as the advances and the emerging challenges that health systems face in several countries. This article reaffirms the need for a strong primary care level--with coordination and response capacity, close to and involved in the community, and accessible--in order to advance towards realizing the right to health for everyone. It also advocates for practical proposals to relaunch the primary health care strategy 40 years after the Declaration of Alma-Ata.

Este artigo apresenta a evolução das propostas e acordos regionais a partir da Declaração de Alma-Ata (1978) até a Estratégia de saúde universal, destacando a vigência das perspectivas básicas da estratégia de atenção primária, atualmente expressas nas propostas de fortalecimento da atenção primária e formação de redes integradas de serviços de saúde. Salienta-se o caráter contextual da implementação da estratégia em cenários nacionais complexos, ilustrando-se com os marcos alcançados nos últimos 40 anos. São descritos os fatores que freiam a implementação da atenção primária à saúde (APS) e os avanços e desafios emergentes atualmente enfrentados pelos sistemas de saúde em vários países. Enfatiza-se que, somente com um nível de atenção primário que seja forte, articulado e resolutivo, que esteja próximo e inserido na comunidade e de fácil acesso às pessoas, é possível progredir no direito à saúde para todos. O artigo defende a elaboração de propostas práticas para relançar a estratégia de APS após 40 anos da Declaração de Alma-Ata.

Study of hTERT and Histone 3 Mutations in Medulloblastoma.

Hotspot activating mutations of the telomerase reverse transcriptase (hTERT) promoter region were recently described in several tumor types. These mutations lead to enhanced expression of telomerase, being responsible for telomere maintenance and allowing continuous cell division. Additionally, there are alternative telomere maintenance mechanisms, associated with histone H3 mutations, responsible for disrupting the histone code and affecting the regulation of transcription. Here, we investigated the clinical relevance of these mechanistically related molecules in medulloblastoma. Sixty-nine medulloblastomas, formalin fixed and paraffin embedded, from a cohort of patients aged 1.5-70 years, were used to investigate the hotspot mutations of the hTERT promoter region, i.e. H3F3A and HIST1H3B, using Sanger sequencing. We successfully sequenced hTERT in all 69 medulloblastoma samples and identified a total of 19 mutated cases (27.5%). c.-124:G>A and c.-146:G>A mutations were detected, respectively, in 16 and 3 samples. Similar to previous reports, hTERT mutations were more frequent in older patients (p < 0.0001), being found only in 5 patients <20 years of age. In addition, hTERT-mutated tumors were more frequently recurrent (p = 0.026) and hTERT mutations were significantly enriched in tumors located in the right cerebellar hemisphere (p = 0.039). No mutations were found on the H3F3A or HIST1H3B genes. hTERT promoter mutations are frequent in medulloblastoma and are associated with older patients, prone to recurrence and located in the right cerebellar hemisphere. On the other hand, histone 3 mutations do not seem to be present in medulloblastoma.

The prognostic impact of TERT promoter mutations in glioblastomas is modified by the rs2853669 single nucleotide polymorphism.

Human hotspot TERT promoter (TERTp) mutations have been reported in a wide range of tumours. Several studies have shown that TERTp mutations are associated with clinicopathological features; in some instances, TERTp mutations were considered as biomarkers of poor prognosis. The rs2853669 SNP, located in the TERT promoter region, was reported to modulate the increased TERT expression levels induced by the recurrent somatic mutations. In this study we aimed to determine the frequency and prognostic value of TERTp mutations and TERT rs2853669 SNP in 504 gliomas from Portuguese and Brazilian patients. TERTp mutations were detected in 47.8% of gliomas (216/452). Glioblastomas (GBM) exhibited the highest frequency of TERTp mutations (66.9%); in this glioma subtype, we found a significant association between TERTp mutations and poor prognosis, regardless of the population. Moreover, in a multivariate analysis, TERTp mutations were the only independent prognostic factor. Our data also showed that the poor prognosis conferred by TERTp mutations was restricted to GBM patients carrying the rs2853669 A allele and not in those carrying the G allele. In conclusion, the presence of TERTp mutations was associated with worse prognosis in GBM patients, although such association depended on the status of the rs2853669 SNP. The status of the rs2853669 SNP should be taken in consideration when assessing the prognostic value of TERTp mutations in GBM patients. TERTp mutations and the rs2853669 SNP can be used in the future as biomarkers of glioma prognosis.

Health-system reform and universal health coverage in Latin America.

Starting in the late 1980s, many Latin American countries began social sector reforms to alleviate poverty, reduce socioeconomic inequalities, improve health outcomes, and provide financial risk protection. In particular, starting in the 1990s, reforms aimed at strengthening health systems to reduce inequalities in health access and outcomes focused on expansion of universal health coverage, especially for poor citizens. In Latin America, health-system reforms have produced a distinct approach to universal health coverage, underpinned by the principles of equity, solidarity, and collective action to overcome social inequalities. In most of the countries studied, government financing enabled the introduction of supply-side interventions to expand insurance coverage for uninsured citizens--with defined and enlarged benefits packages--and to scale up delivery of health services. Countries such as Brazil and Cuba introduced tax-financed universal health systems. These changes were combined with demand-side interventions aimed at alleviating poverty (targeting many social determinants of health) and improving access of the most disadvantaged populations. Hence, the distinguishing features of health-system strengthening for universal health coverage and lessons from the Latin American experience are relevant for countries advancing universal health coverage.

Measuring evolution of income-related inequalities in health and health care utilization in selected Latin American and Caribbean countries.

OBJECTIVE: To describe the methodology used to measure and explain income-related inequalities in health and health care utilization over time in selected Latin American and Caribbean countries. METHODS: Data from nationally representative household surveys in Brazil, Chile, Colombia, Jamaica, Mexico, and Peru were used to analyze income-related inequalities in health status and health care utilization. Health was measured by self-reported health status, physical limitations, and chronic illness when available. Hospitalization, physician, dentist, preventive, curative, and preventive visits were proxies for health care utilization. Household income was a proxy for socioeconomic status except in Peru, which used household expenditures. Concentration indices were calculated before and after standardization for all dependent variables. Standardized concentration indices are also referred to as horizontal inequity index. Decomposition analysis was used to identify the main determinants of inequality in health care utilization. RESULTS: Results of analysis of the evolution of income-related inequality in health and health care utilization in Brazil, Chile, Colombia, Jamaica, Mexico, and Peru are presented in separate articles in this issue. CONCLUSIONS: The methodology used for analysis of equity in all six country research studies attempts not to determine causality but to describe and explain income-related inequalities in health status and health care utilization over time. While this methodology is robust, it is not free of errors. When possible, errors have been identified and corrected.

Equity in health and health care in Peru, 2004-2008.

OBJECTIVE: This study evaluates whether recent positive economic trends and pro-poor health policies have resulted in more health equity and explores key factors that explain such change. METHODS: This study focuses on the evolution of measures of health status (self-reported morbidity) and use of health care services obtained from the 2004 and 2008 rounds of the Peruvian National Household Survey (Encuesta Nacional de Hogares). It concentrates on health inequalities associated with socioeconomic status and uses interquintile differences (gradient), concentration indices with and without needs-based adjustments, and decomposition analysis. RESULTS: Findings show a low level of inequality in measures of health status, with a slightly pro-poor inequality in self-reported health problems and a slightly pro-rich inequality in self-reported chronic illness. Inequity in the use of curative services declined significantly between 2004 and 2008, while inequity in the use of preventive services increased slightly. Use of hospital and dental services remained unchanged during the same period. CONCLUSIONS: Limitations of self-reported morbidity measures probably underestimate the results of health inequalities across socioeconomic groups. Improved equity in the use of curative health services can be explained by a number of positive factors that occurred concurrently during the analysis-namely, increased mean household income, reduced economic inequality, the Juntos conditional cash transfer program, and gradual expansion of public health insurance, Seguro Integral de Salud (SIS). Given that SIS expansion is the main public policy for promoting health equity in Peru, it is crucial that future steps in expansion come with a strategy to isolate its contribution to health equity improvements from that of other positive socioeconomic trends.

Analysis of the evolution and determinants of income-related inequalities in the Brazilian health system, 1998 - 2008.

OBJECTIVE: To analyze the evolution and determinants of income-related inequalities in the Brazilian health system between 1998 and 2008. METHODS: Data from the National Household Sampling Surveys of 1998, 2003, and 2008 were used to analyze inequalities in health and health care. Health was measured by self-reported health status, physical limitations, and chronic illness. Hospitalization and physician and dentist visits were proxies for health care utilization. Income was a proxy for socioeconomic status. Concentration indices were calculated before and after standardization for all dependent variables. Decomposition analysis was used to identify the main determinants of inequality in health care utilization. RESULTS: In all three periods analyzed, the poor reported worse health status, while the wealthy reported more chronic diseases; health care utilization was pro-rich for medical and dental services. Yet, income-related inequality in health care utilization has been declining. Private health insurance, education, and income are the major contributors to the inequalities identified. CONCLUSIONS: Income-related inequality in the use of medical and dental health care is gradually declining in Brazil. The decline is associated with implementation of pro-equity policies and programs, such as the Community Health Agents Program and the Family Health Program.

Ensuring access to essential health products: Lessons from Colombia's leishmaniasis control and elimination initiative.

BACKGROUND: This paper identifies opportunities and challenges for leishmaniasis control and elimination in Colombia, emphasizing the role of pooled procurement of essential medicines and supplies. Colombia is among the countries most affected by leishmaniasis globally, and also faces the dual challenge of procuring critically needed medicines in the context of limited national resources. It recently renewed its commitment to the control and elimination of leishmaniasis under its 2022-2031 Public Health Plan (PDSP) through a comprehensive public health approach. METHODOLOGY/PRINCIPAL FINDINGS: The methodology comprises a comprehensive literature review and key informant interviews with leishmaniasis experts from the Colombian national control program and PAHO/WHO, focusing on cutaneous, mucocutaneous, and visceral leishmaniasis. Leishmaniasis is endemic throughout Colombia, with over 11 million people at risk, many of whom live in poverty-stricken, remote and isolated rural areas with limited access to health services. Leishmaniasis care, including medicines, is provided free of charge, but many barriers were nonetheless identified at environmental, population, and health system levels, including the supply of quality-assured medicines. Opportunities to alleviate these barriers were identified, including the support of the PAHO Strategic Fund. Within the context of the sustainable development goals and international leishmaniasis control and elimination targets, Colombian officials have established their own priorities, the highest of which is the reduction of deaths from visceral leishmaniasis. CONCLUSIONS/SIGNIFICANCE: The elimination of leishmaniasis as a public health problem presents significant challenges, given its biological complexity and diversity, physical and clinical manifestations, social and economic impacts, frequently burdensome treatment regimens, and insufficient supply of necessary medicines. However, rigorous prevention and control efforts through strong political commitment and a highly motivated workforce can dramatically reduce its burden. Colombia's new PDSP, which highlights leishmaniasis control, is an opportunity for a revitalized health system response through committed leadership, intersectoral actions, and partnerships with international organizations that share a common vision.

Minimizing COVID-19 disruption: Ensuring the supply of essential health products for health emergencies and routine health services.

Leveraging economies of scale and scope through multi-country pooled procurement enables countries to increase access to quality affordable essential medicines and supplies that meet priority health objectives as well as effectively respond to health emergencies. Strategic partnerships and tools can minimize supply chain disruptions and streamline procurement and deployment in health emergencies, thus mitigating stockouts and ensuring cost efficiencies across various therapeutic areas, including for public health programs at a time when countries may struggle to meet complex needs. As a means to better respond to health emergencies while maintaining priority public health programs, countries should optimize usage of pooled procurement mechanisms facilitated by multilateral technical cooperation and other regional mechanisms, such as the Pan American Health Organization's Strategic Fund. Because few analyses have assessed the role of such regional procurement mechanisms, this Health Policy paper evaluates the key areas of impact of the PAHO Strategic Fund and concludes with lessons learned to help prepare for future health crises while maintaining essential health services.

DNA Methylation Markers from Negative Surgical Margins Can Predict Recurrence of Oral Squamous Cell Carcinoma.

The identification of molecular markers in negative surgical margins of oral squamous cell carcinoma (OSCC) might help in identifying residual molecular aberrations, and potentially improve the prediction of prognosis. We performed an Infinium MethylationEPIC BeadChip array on 32 negative surgical margins stratified based on the status of tumor recurrence in order to identify recurrence-specific aberrant DNA methylation (DNAme) markers. We identified 2512 recurrence-associated Differentially Methylated Positions (DMPs) and 392 Differentially Methylated Regions (DMRs) which were enriched in cell signaling and cancer-related pathways. A set of 14-CpG markers was able to discriminate recurrent and non-recurrent cases with high specificity and sensitivity rates (AUC 0.98, p = 3 × 10-6; CI: 0.95-1). A risk score based on the 14-CpG marker panel was applied, with cases classified within higher risk scores exhibiting poorer survival. The results were replicated using tumor-adjacent normal HNSCC samples from The Cancer Genome Atlas (TCGA). We identified residual DNAme aberrations in the negative surgical margins of OSCC patients, which could be informative for patient management by improving therapeutic intervention. This study proposes a novel DNAme-based 14-CpG marker panel as a promising predictor for tumor recurrence, which might contribute to improved decision-making for the personalized treatment of OSCC cases.

HPV-Induced Oropharyngeal Squamous Cell Carcinomas in Brazil: Prevalence, Trend, Clinical, and Epidemiologic Characterization.

BACKGROUND: Tobacco or human papillomavirus (HPV)-related oropharyngeal squamous cell carcinomas (OPSCC) represent different clinical and epidemiologic entities. This study investigated the prevalence of HPV-positive and HPV-negative OPSCC in a reference cancer hospital in Brazil and its association with clinical and demographic data, as well as its impact on overall survival. METHODS: HPV infection was determined by p16-IHC in pre-treatment formalin-fixed paraffin-embedded samples from all patients with OPSCC diagnosed at Barretos Cancer Hospital between 2008 and 2018. The prevalence of HPV-positive cases and its temporal trend was assessed, and the association of clinical and demographic data with HPV infection and the impact on patient overall survival was evaluated. RESULTS: A total of 797 patients with OPSCC were included in the study. The prevalence of HPV-associated tumors in the period was 20.6% [95% confidence interval, 17.5-24.0] with a significant trend for increase of HPV-positive cases over the years (annual percentage change = 12.87). In a multivariate analysis, the variables gender, level of education, smoking, tumor sublocation, region of Brazil, and tumor staging had a significant impact in HPV positivity, and a greater overall survival (OS) was observed in HPV-positive patients (5-year OS: 47.9% vs. 22.0%; P = 0.0001). CONCLUSIONS: This study represents the largest cohort of Brazilian patients with OPSCC characterized according to HPV status. We report significant differences in demographics and clinical presentation according to HPV status, and an increasing trend in prevalence for HPV-induced tumors. IMPACT: These findings can potentially contribute to a better stratification and management of patients as well as assist in prevention strategies.

Human Papillomavirus DNA Detection by Droplet Digital PCR in Formalin-Fixed Paraffin-Embedded Tumor Tissue from Oropharyngeal Squamous Cell Carcinoma Patients.

INTRODUCTION: High-risk human papillomavirus infection impacts staging and prognosis of oropharyngeal squamous cell carcinomas (OPSCCs). Determination of HPV status in tumor tissue by p16-immunohistochemistry (p16-IHC) can be challenging; therefore, complementary methodologies could be useful in a clinical setting. OBJECTIVE: To test for accuracy and clinical relevance of HPV-DNA detection in formalin-fixed and paraffin-embedded (FFPE) tumor samples by droplet digital PCR (ddPCR). MATERIALS AND METHODS: Fifty OPSCCs were tested for p16-IHC status followed by HPV-16/18 DNA detection/quantification in FFPE-recovered DNA using ddPCR. Accuracy for HPV status determination and association with patient information were also evaluated. RESULTS: 32.0% (16/50) of the cases were p16-IHC positive (p16 +), 42.0% (21/50) had detectable levels of HPV-16 DNA, and none were positive for HPV-18 DNA. A higher median viral load of HPV-16 DNA was observed in p16 + cases (p < 0.0001). Concordance between p16-IHC and HPV-16 DNA ranged from 78.0 to 86.0% and accuracy rates were between 78.0 and 86.0%. P16-IHC and HPV-16 DNA detection was associated with gender, smoking status, and tumor subsite, while only HPV-16 DNA was associated with cT stage. The combination of HPV positivity by p16-IHC and ddPCR showed higher overall survival rates in comparison with p16 + /HPV-DNA- and p16 - /HPV-DNA- results. CONCLUSIONS: Type-specific HPV-DNA detection by ddPCR is highly specific but moderately sensitive for the determination of HPV status and showed clinical relevance, mainly when associated with p16-IHC status. Results highlight the importance of performing HPV-DNA testing in combination with p16-IHC for proper identification of HPV-associated OPSCC and to improve clinical management of OPSCC patients.

Focal hyperechoic liver lesions in children: far beyond hemangiomas - pictorial essay.

The aim of this report was to present a selection of focal hyperechoic liver lesions of different etiologies, illustrating the wide spectrum of diagnostic possibilities for such lesions in the pediatric population.

Apresentamos uma seleção de lesões focais hiperecogênicas hepáticas de diversas etiologias, que ilustram o grande espectro de diferentes possibilidades diagnósticas dessas lesões na faixa pediátrica, muito além dos hemangiomas.

Feasibility of methylated ctDNA detection in plasma samples of oropharyngeal squamous cell carcinoma patients.

BACKGROUND: Oropharyngeal squamous cell carcinomas (OpSCCs) are commonly associated with high rates of treatment failure. OBJECTIVES: To evaluate methylation-based markers in plasma from OpSCC patients as emerging tools for accurate/noninvasive follow-up. METHODS: Pretreatment formalin-fixed paraffin-embedded (FFPE) biopsies (n = 52) and paired plasma (n = 15) were tested for the methylation of CCNA1, DAPK, CDH8, and TIMP3 by droplet digital PCR (ddPCR). RESULTS: Seventy-one percent (37/52) of the biopsies showed methylation of at least one of the evaluated genes and tumor CCNA1 methylation was associated with recurrence-free survival. Methylated circulating tumor DNA (meth-ctDNA) was detected in 11/15 (73.3%) plasma samples; conversely, plasma samples from healthy controls were all negative for DNA methylation (area under the curve = 0.867; 95% confidence interval = 0.720-1.000). Additionally, preliminary results on the detection of meth-ctDNA in plasma collected during follow-up closely matched patient outcome. CONCLUSIONS: The results suggest the feasibility of detecting meth-ctDNA in plasma using ddPCR and a possible application on routine setting after further validation.

Loss of SPINT2 expression frequently occurs in glioma, leading to increased growth and invasion via MMP2.

PURPOSE: High-grade gliomas (HGG) remain one of the most aggressive tumors, which is primarily due to its diffuse infiltrative nature. Serine proteases and metalloproteases are known to play key roles in cellular migration and invasion mechanisms. SPINT2, also known as HAI-2, is an important serine protease inhibitor that can affect MET signaling. SPINT2 has been found to be frequently downregulated in various tumors, whereby hypermethylation of its promoter appears to serve as a common mechanism. Here, we assessed the clinical relevance of SPINT2 expression and promoter hypermethylation in pediatric and adult HGG and explored its functional role. METHODS: A series of 371 adult and 77 pediatric primary HGG samples was assessed for SPINT2 protein expression (immunohistochemistry) and promoter methylation (methylation-specific PCR) patterns. After SPINT2 knockdown and knock-in in adult and pediatric HGG cell lines, a variety of in vitro assays was carried out to determine the role of SPINT2 in glioma cell viability and invasion, as well as their mechanistic associations with metalloprotease activities. RESULTS: We found that SPINT2 protein expression was frequently absent in adult (85.3%) and pediatric (100%) HGG samples. The SPINT2 gene promoter was found to be hypermethylated in approximately half of both adult and pediatric gliomas. Through functional assays we revealed a suppressor activity of SPINT2 in glioma cell proliferation and viability, as well as in their migration and invasion. These functions appear to be mediated in part by MMP2 expression and activity. CONCLUSIONS: We conclude that dysregulation of SPINT2 is a common event in both pediatric and adult HGG, in which SPINT2 may act as a tumor suppressor.

Expression of GNAS, TP53, and PTEN Improves the Patient Prognostication in Sonic Hedgehog (SHH) Medulloblastoma Subgroup.

Medulloblastoma (MB) is the most common malignant brain tumor in children. It is currently classified in four main molecular subgroups with different clinical outcomes: sonic hedgehog, wingless, group 3, and group 4 (MBSHH, MBWNT, MBGRP3, or MBGRP4). Presently, a 22-gene expression panel has been efficiently applied for molecular subgrouping using nCounter technology. In this study, formalin-fixed, paraffin-embedded samples from 164 Brazilian medulloblastomas were evaluated, applying the 22-gene panel, and subclassified into the low and high expression of nine key medulloblastoma-related genes. In addition, TP53 mutation status was assessed using TruSight Tumor 15 Panel, and its correlation with expression and prognostic impact was evaluated. Samples from 149 of 164 patients (90%) were classified into MBSHH (47.7%), MBWNT (16.1%), MBGRP3 (15.4%), and MBGRP4 (20.8%). GNAS presented the highest expression levels, with higher expression in MBSHH. TP53, MYCN, SOX2, and MET were also up-regulated in MBSHH, whereas PTEN was up-regulated in MBGRP4. GNAS, TP53, and PTEN low expression was associated with the unfavorable patient outcome only for MBSHH (P = 0.04, P = 0.01, and P = 0.02, respectively). TP53 mutations were detected in 28.57% of MBSHH cases and exhibited association with lower expression and worse clinical outcome, although not statistically significant. The 22-gene panel for molecular classification of medulloblastoma associated with the expression of GNAS, TP53, and PTEN improves the patient prognostication in MBSHH subgroup and can be easily incorporated in the 22-gene panel without any additional costs.