Aptamer-Drug conjugates for a targeted and synergistic anticancer Response: Exploiting T30923-5-fluoro-2'-deoxyuridine (INT-FdU) derivatives.

One of the most appealing approaches for cancer treatment is targeted therapy, which is based on the use of drugs able to target cancer cells without affecting normal ones. This strategy lets to overcome the major limitation of conventional chemotherapy, namely the lack of specificity of anticancer drugs, which often leads to severe side effects, decreasing the therapy effectiveness. Delivery of cell-killing substances to tumor cells is one-way targeted drug therapy can work. Generally, monoclonal antibodies are combined with chemotherapeutic drugs, allowing cellular uptake through the binding to their targets on the surface of cancer cells. Aptamer-drug conjugates represent a promising alternative solution to antibodies to minimize off-target effects, considering the remarkable selective binding capabilities of aptamers. In this study, to enhance the therapeutic efficacy of the antineoplastic agent 5-fluoro-2'-deoxyuridine (FdU) in various cancer cells, we focused on the development of a novel conjugate using the antiproliferative aptamer T30923 (INT) as a drug vehicle. Three derivatives composed of T30923 conjugated with a different number of FdU units were synthesized, and their structural and biological properties were thoroughly characterized, highlighting their potential for targeted and synergistic anticancer responses.

TRIB3 silencing promotes the downregulation of Akt pathway and PAX3-FOXO1 in high-risk rhabdomyosarcoma.

Rhabdomyosarcoma (RMS), such as other childhood tumors, has witnessed treatment advancements in recent years. However, high-risk patients continue to face poor survival rates, often attributed to the presence of the PAX3/7-FOXO1 fusion proteins, which has been associated with metastasis and treatment resistance. Despite efforts to directly target these chimeric proteins, clinical success remains elusive. In this study, the main aim was to address this challenge by investigating regulators of FOXO1. Specifically, we focused on TRIB3, a potential regulator of the fusion protein in RMS. Our findings revealed a prominent TRIB3 expression in RMS tumors, highlighting its correlation with the presence of fusion protein. By conducting TRIB3 genetic inhibition experiments, we observed an impairment on cell proliferation. Notably, the knockdown of TRIB3 led to a decrease in PAX3-FOXO1 and its target genes at protein level, accompanied by a reduction in the activity of the Akt signaling pathway. Additionally, inducible silencing of TRIB3 significantly delayed tumor growth and improved overall survival in vivo. Based on our analysis, we propose that TRIB3 holds therapeutic potential for treating the most aggressive subtype of RMS. The findings herein reported contribute to our understanding of the underlying molecular mechanisms driving RMS progression and provide novel insights into the potential use of TRIB3 as a therapeutic intervention for high-risk RMS patients.

Defined covalent attachment of three cancer drugs to DNA origami increases cytotoxicity at nanomolar concentration.

DNA nanostructures have captured great interest as drug delivery vehicles for cancer therapy. Despite rapid progress in the field, some hurdles, such as low cellular uptake, low tissue specificity or ambiguous drug loading, remain unsolved. Herein, well-known antitumor drugs (doxorubicin, auristatin, and floxuridine) were site-specifically incorporated into DNA nanostructures, demonstrating the potential advantages of covalently linking drug molecules via structural staples instead of incorporating the drugs by noncovalent binding interactions. The covalent strategy avoids critical issues such as an unknown number of drug-DNA binding events and premature drug release. Moreover, covalently modified origami offers the possibility of precisely incorporating several synergetic antitumor drugs into the DNA nanostructure at a predefined molar ratio and to control the exact spatial orientation of drugs into DNA origami. Additionally, DNA-based nanoscaffolds have been reported to have a low intracellular uptake. Thus, two cellular uptake enhancing mechanisms were studied: the introduction of folate units covalently linked to DNA origami and the transfection of DNA origami with Lipofectamine. Importantly, both methods increased the internalization of DNA origami into HTB38 and HCC2998 colorectal cancer cells and produced greater cytotoxic activity when the DNA origami incorporated antiproliferative drugs. The results here present a successful and conceptually distinct approach for the development of DNA-based nanostructures as drug delivery vehicles, which can be considered an important step towards the development of highly precise nanomedicines.

Analysis of Cancer Genomic Amplifications Identifies Druggable Collateral Dependencies within the Amplicon.

The identification of novel therapeutic targets for specific cancer molecular subtypes is crucial for the development of precision oncology. In the last few years, CRISPR/Cas9 screens have accelerated the discovery and validation of new targets associated with different tumor types, mutations, and fusions. However, there are still many cancer vulnerabilities associated with specific molecular features that remain to be explored. Here, we used data from CRISPR/Cas9 screens in 954 cancer cell lines to identify gene dependencies associated with 16 common cancer genomic amplifications. We found that high-copy-number genomic amplifications generate multiple collateral dependencies within the amplified region in most cases. Further, to prioritize candidate targets for each chromosomal region amplified, we integrated gene dependency parameters with both druggability data and subcellular location. Finally, analysis of the relationship between gene expression and gene dependency led to the identification of genes, the expression of which may constitute predictive biomarkers of dependency. In conclusion, our study provides a set of druggable targets specific for each amplification, opening the possibility to specifically target amplified tumors on this basis.

Single-cell profiling of alveolar rhabdomyosarcoma reveals RAS pathway inhibitors as cell-fate hijackers with therapeutic relevance.

Rhabdomyosarcoma (RMS) is a group of pediatric cancers with features of developing skeletal muscle. The cellular hierarchy and mechanisms leading to developmental arrest remain elusive. Here, we combined single-cell RNA sequencing, mass cytometry, and high-content imaging to resolve intratumoral heterogeneity of patient-derived primary RMS cultures. We show that the aggressive alveolar RMS (aRMS) subtype contains plastic muscle stem-like cells and cycling progenitors that drive tumor growth, and a subpopulation of differentiated cells that lost its proliferative potential and correlates with better outcomes. While chemotherapy eliminates cycling progenitors, it enriches aRMS for muscle stem-like cells. We screened for drugs hijacking aRMS toward clinically favorable subpopulations and identified a combination of RAF and MEK inhibitors that potently induces myogenic differentiation and inhibits tumor growth. Overall, our work provides insights into the developmental states underlying aRMS aggressiveness, chemoresistance, and progression and identifies the RAS pathway as a promising therapeutic target.

Desarrollo de los cuidados paliativos pediátricos en Uruguay, 2008-2020: informe de avances / Development of pediatric palliative care in Uruguay, 2008-2020: progress report / Desenvolvimento dos cuidados paliativos pediátricos no Uruguai nos anos 2008-2020: relatório de progresso

Desde el año 2007 en Uruguay los cuidados paliativos (CP) son parte de las prestaciones de salud que todos los ciudadanos que los necesitan tienen derecho a recibir y, desde entonces, ha aumentado significativamente la accesibilidad a estos. Objetivo: describir la situación actual del desarrollo organizativo de servicios de cuidados paliativos pediátricos (CPP) en el país y los pacientes por ellos asistidos desde el inicio de sus actividades hasta el 31/12/2020, las principales fortalezas y desafíos percibidos por los profesionales de dichos equipos. Metodología: se realizó una consulta mediante encuesta online auto administrada enviada a los coordinadores de servicios de CPP del Uruguay. Resultados: se confirmaron 19 equipos en 9/19 departamentos, 5/19 están integrados por profesionales de las cuatro disciplinas básicas recomendadas, el resto por distintas combinaciones de disciplinas, con cargas horarias muy variables. Brindan asistencia en: hospitalización 19/19, policlínica 18/19, atención domiciliaria coordinada 13/19 y retén telefónico 10/19. Fueron asistidos 2957 niños, 23% de los mismos fallecieron. 16/19 equipos reportan como principales fortalezas los valores compartidos y el trabajo en equipo interdisciplinario y 15/19 como principal desafío los déficits de recursos humanos. Conclusiones: persisten importantes inequidades en el acceso a los CPP. Se constató gran variabilidad en la integración de los equipos y la carga horaria de los profesionales. Es necesario que las autoridades sanitarias continúen promoviendo y exigiendo el desarrollo de equipos de CPP en las instituciones y departamentos que no los tienen y el cumplimiento de estándares mínimos de calidad en los ya existentes.

Since 2007, palliative care (PC) has been a part of the health benefits that all Uruguayan citizens are entitled to receive and, since then, accessibility has increased significantly. Objective: to describe the present development of pediatric palliative care services (PPC) in Uruguay and the patients assisted by health providers since the beginning of their services until 12/31/2020 and the key strengths and challenges perceived by these palliative care teams. Methodology: a consultation was carried out through a self-administered online survey and sent to the PPC service coordinators in Uruguay. Results: 19 teams were confirmed in 9/19 departments, 5/19 are integrated by professionals from the four recommended basic disciplines, the rest by different combinations of disciplines, with highly variable workloads. They provide assistance in: hospitalization 19/19, clinics 18/19, coordinated home care 13/19 and telephone assistance 10/19. 2957 children were assisted, 23% of them died. 16/19 teams report shared values and interdisciplinary teamwork as their main strengths, and 15/19 report human resource shortage as their main challenge. Conclusions: significant inequality persist regarding access to PPCs. We confirmed a high variability in teams' integration and professional workload. It is necessary for the health authorities to continue to promote and demand the development of PPC teams in the institutions and departments that do not yet have them and the compliance with minimum quality standards in those that already operate.

Desde 2007, os cuidados paliativos (CP) fazem parte dos benefícios de saúde que todos os cidadãos têm direito a receber no Uruguai e, desde então, a acessibilidade a eles tem aumentado significativamente. Objetivo: descrever a situação atual do desenvolvimento organizacional dos serviços de cuidados paliativos pediátricos (CPP) no Uruguai e dos pacientes atendidos desde o início de suas atividades até 31/12/2020 e as principais fortalezas e desafios percebidos pelos profissionais das referidas equipes. Metodologia: foi realizada uma consulta por meio de uma pesquisa online autoaplicável enviada aos coordenadores dos serviços do CPP no Uruguai. Resultados: 19 equipes foram confirmadas em 19/09 departamentos, 19/05 compostas por profissionais das quatro disciplinas básicas recomendadas, o restante por diferentes combinações de disciplinas, com cargas horárias altamente variáveis. Elas atendem em: internação 19/19, policlínica 18/19, atendimento domiciliar coordenado 13/19 e posto telefônico 19/10. 2.957 crianças foram atendidas, 23% delas faleceram. 16/19 equipes relatam valores compartilhados e trabalho em equipe interdisciplinar como suas principais fortalezas, e 15/19 relatam déficits de recursos humanos como seu principal desafio. Conclusões: persistem desigualdades significativas no acesso aos CPP. Verificou-se: grande variabilidade na integração das equipes e na carga de trabalho dos profissionais. É necessário que as autoridades de saúde continuem promovendo e exigindo o desenvolvimento de equipes de CPP nas instituições e departamentos que não as possuem e o cumprimento de padrões mínimos de qualidade nas que já existem.

Integrin alpha9 emerges as a key therapeutic target to reduce metastasis in rhabdomyosarcoma and neuroblastoma.

The majority of current cancer therapies are aimed at reducing tumour growth, but there is lack of viable pharmacological options to reduce the formation of metastasis. This is a paradox, since more than 90% of cancer deaths are attributable to metastatic progression. Integrin alpha9 (ITGA9) has been previously described as playing an essential role in metastasis; however, little is known about the mechanism that links this protein to this process, being one of the less studied integrins. We have now deciphered the importance of ITGA9 in metastasis and provide evidence demonstrating its essentiality for metastatic dissemination in rhabdomyosarcoma and neuroblastoma. However, the most translational advance of this study is to reveal, for the first time, the possibility of reducing metastasis by pharmacological inhibition of ITGA9 with a synthetic peptide simulating a key interaction domain of ADAM proteins, in experimental metastasis models, not only in childhood cancers but also in a breast cancer model.

Dickkopf-1 Inhibition Reactivates Wnt/ß-Catenin Signaling in Rhabdomyosarcoma, Induces Myogenic Markers In Vitro and Impairs Tumor Cell Survival In Vivo.

The Wnt/ß-catenin signaling pathway plays a pivotal role during embryogenesis and its deregulation is a key mechanism in the origin and progression of several tumors. Wnt antagonists have been described as key modulators of Wnt/ß-catenin signaling in cancer, with Dickkopf-1 (DKK-1) being the most studied member of the DKK family. Although the therapeutic potential of DKK-1 inhibition has been evaluated in several diseases and malignancies, little is known in pediatric tumors. Only a few works have studied the genetic inhibition and function of DKK-1 in rhabdomyosarcoma. Here, for the first time, we report the analysis of the therapeutic potential of DKK-1 pharmaceutical inhibition in rhabdomyosarcoma, the most common soft tissue sarcoma in children. We performed DKK-1 inhibition via shRNA technology and via the chemical inhibitor WAY-2626211. Its inhibition led to ß-catenin activation and the modulation of focal adhesion kinase (FAK), with positive effects on in vitro expression of myogenic markers and a reduction in proliferation and invasion. In addition, WAY-262611 was able to impair survival of tumor cells in vivo. Therefore, DKK-1 could constitute a molecular target, which could lead to novel therapeutic strategies in RMS, especially in those patients with high DKK-1 expression.

Dickkopf Proteins and Their Role in Cancer: A Family of Wnt Antagonists with a Dual Role.

The Wnt signaling pathway regulates crucial aspects such as cell fate determination, cell polarity and organogenesis during embryonic development. Wnt pathway deregulation is a hallmark of several cancers such as lung, gastric and liver cancer, and has been reported to be altered in others. Despite the general agreement reached by the scientific community on the oncogenic potential of the central components of the pathway, the role of the antagonist proteins remains less clear. Deregulation of the pathway may be caused by overexpression or downregulation of a wide range of antagonist proteins. Although there is growing information related to function and regulation of Dickkopf (DKK) proteins, their pharmacological potential as cancer therapeutics still has not been fully developed. This review provides an update on the role of DKK proteins in cancer and possible potential as therapeutic targets for the treatment of cancer; available compounds in pre-clinical or clinical trials are also reviewed.

Sinais e sintomas de disfunção temporomandibular em adolescentes e sua associação com o nível socioeconômico familiar e outras variáveis

Justificativa e Objetivos: A adolescência é caracterizada por intensas mudanças físicas, emocionais e sociais, o que pode propiciar a presença de fatores etiológicos e perpetuantes importantes da disfunção temporomandibular (DTM). Assim, o objetivo deste estudo foi avaliar a presença de DTM na adolescência e sua associação com aspectos socioeconômicos e outras variáveis. Métodos: Compuseram a amostra 200 adolescentes de 10 a 19 anos provenientes das clínicas do Curso de Odontologia da Universidade de Araraquara ­ UNIARA e de escolas públicas e particulares do município. O Critério de Classificação Econômica Brasil (ABEP) foi utilizado para a identificação da classe social do adolescente. Para diagnóstico da DTM e da intensidade da dor à palpação usou-se o Research Diagnostic Criteria for Temporomandibular Disorders (RDC/TMD). O número de problemas psicoemocionais, as áreas de dor extrafacial e o número de hábitos parafuncionais foram verificados por um questionário estruturado de autorrelato. Resultados: Houve associação positiva entre as classes D/E e a presença de DTM dolorosa (p= 0,007, OR=24,8; 95%IC=2,28-269,79), principalmente com disfunções musculares (p=0,003, OR=32,0; 95%IC=2,83-362,01). Os fatores que se expressaram significativamente maiores na presença de DTM dolorosa foram: intensidade de dor à palpação (p=0,000), quantidade de áreas de dor extrafacial (p=0,025) e número de hábitos parafuncionais (p=0,000). A Classe D/E relatou mais áreas de dor que as demais (p=0,049). Conclusão: O nível socioeconômico se associou significativamente à presença de DTM dolorosa, especialmente de origem muscular. O único fator que se expressou significativamente maior nas classes sociais mais baixas foi o número de áreas dolorosas extrafaciais.

Two novel effectors of trafficking and maturation of the yeast plasma membrane H+ -ATPase.

The endoplasmic reticulum (ER) is the entry site of proteins into the endomembrane system. Proteins exit the ER via coat protein II (COPII) vesicles in a selective manner, mediated either by direct interaction with the COPII coat or aided by cargo receptors. Despite the fundamental role of such receptors in protein sorting, only a few have been identified. To further define the machinery that packages secretory cargo and targets proteins from the ER to Golgi membranes, we used multiple systematic approaches, which revealed 2 uncharacterized proteins that mediate the trafficking and maturation of Pma1, the essential yeast plasma membrane proton ATPase. Ydl121c (Exp1) is an ER protein that binds Pma1, is packaged into COPII vesicles, and whose deletion causes ER retention of Pma1. Ykl077w (Psg1) physically interacts with Exp1 and can be found in the Golgi and coat protein I (COPI) vesicles but does not directly bind Pma1. Loss of Psg1 causes enhanced degradation of Pma1 in the vacuole. Our findings suggest that Exp1 is a Pma1 cargo receptor and that Psg1 aids Pma1 maturation in the Golgi or affects its retrieval. More generally our work shows the utility of high content screens in the identification of novel trafficking components.

Hedgehog Pathway Inhibition Hampers Sphere and Holoclone Formation in Rhabdomyosarcoma.

Rhabdomyosarcoma (RMS) is the most common type of soft tissue sarcoma in children and can be divided into two main subtypes: embryonal (eRMS) and alveolar (aRMS). Among the cellular heterogeneity of tumors, the existence of a small fraction of cells called cancer stem cells (CSC), thought to be responsible for the onset and propagation of cancer, has been demonstrated in some neoplasia. Although the existence of CSC has been reported for eRMS, their existence in aRMS, the most malignant subtype, has not been demonstrated to date. Given the lack of suitable markers to identify this subpopulation in aRMS, we used cancer stem cell-enriched supracellular structures (spheres and holoclones) to study this subpopulation. This strategy allowed us to demonstrate the capacity of both aRMS and eRMS cells to form these structures and retain self-renewal capacity. Furthermore, cells contained in spheres and holoclones showed significant Hedgehog pathway induction, the inhibition of which (pharmacologic or genetic) impairs the formation of both holoclones and spheres. Our findings point to a crucial role of this pathway in the maintenance of these structures and suggest that Hedgehog pathway targeting in CSC may have great potential in preventing local relapses and metastases.

Frequência de hipotermia não intencional no perioperatório de cirurgias eletivas / Unintentional hypothermia frequency in the perioperative period of elective surgeries / Frequência de hipotermia não intencional sin perioperatório de cirurgias eletivas

Objetivo: Analisar a frequência de hipotermia não intencional em pacientes submetidos a cirurgias eletivas no período perioperatório, pontuando sua relação com o tipo de anestesia. Método: Estudo transversal, descritivo e de abordagem quantitativa, com amostra de 53 pacientes com faixa etária entre 18 e 90 anos, desenvolvido num centro cirúrgico de hospital privado da região Centro-oeste do estado de São Paulo. Resultados: O sexo feminino apareceu como variável preponderante (79,3%) e 96,3% dos pacientes avaliados apresentaram hipotermia, com variação entre leve e moderada. O número de pacientes hipotérmicos submetidos à raquianestesia foi maior quando comparado àqueles submetidos à geral, tanto na sala de operação (93,5%) quanto na sala de recuperação pós-anestésica (96,8%). Conclusão: A hipotermia é um evento comum que acomete uma grande parcela de pacientes submetidos a procedimentos anestésico-cirúrgicos, assim, torna-se imprescindível que o enfermeiro identifique precocemente sua ocorrência, a fim de oferecer uma assistência qualificada e segura a esses pacientes.

Objective: To analyze the frequency of unintended hypothermia in patients undergoing elective surgery in the perioperative period, scoring its relationship with the type of anesthesia. Method: Cross-sectional, descriptive, and quantitative approach study with a sample of 53 patients aged between 18 and 90 years, developed in a surgical center of a private hospital in the Midwest region of the São Paulo State. Results: The female gender appeared as a major variable (79.3%), and 96.3% of the assessed patients presented hypothermia, ranging from mild to moderate. The number of hypothermic patients undergoing spinal anesthesia was higher when compared to those undergoing general anesthesia, both in the operation room (93.5%) and in the postanesthetic room (96.8%). Conclusion: Hypothermia is a common event that affects a large portion of patients undergoing surgical procedures, so it is essential that nurses early identify the occurrence of this phenomenon to provide a qualified and safe care to these patients.

Objetivo: Analizar la frecuencia de hipotermia no intencional en pacientes sometidos a cirugías electivas en el período perioperatorio, señalando su relación con el tipo de anestesia. Método: Estudio transversal, descriptivo e de abordaje cuantitativo, con muestra de 53 pacientes en la franja etaria de 18 a 90 anos, desarrollado en un quirófano de hospital privado de la región Centro-oeste del estado de São Paulo. Resultados: El sexo femenino apareció como variable preponderante (79,3%) e 96,3% de los pacientes evaluados presentaron hipotermia, con variación entre leve y moderada. El número de pacientes hipotérmicos sometidos a anestesia raquídea fue mayor al ser comparado a aquellos sometidos a general, tanto en sala de operación (93,5%) como en la sala de recuperación post-anestésica (96,8%). Conclusión: La hipotermia es un evento común que afecta una gran parte de pacientes sometidos a procedimientos anestésico-quirúrgicos, así, se vuelve imprescindible que el enfermero identifique precozmente su ocurrencia, a fin de ofrecer una asistencia calificada y segura a esos pacientes.

Prevalência de maloclusão em escolares de 5 a 12 anos de rede municipal de ensino de Araraquara / Prevalence of malocclusion in children between 5 and 12 years-old in municipal schools in Araraquara

OBJETIVO: avaliar a prevalência de maloclusões em escolares na faixa etária de cinco a 12 anos, matriculadas em escolas municipais de Araraquara. MÉTODO: com base no número total de crianças (7235) realizou-se o cálculo da amostragem, envolvendo sete escolas, num total de 3380 crianças. Um estudo piloto e a calibração dos examinadores precederam o início da pesquisa. Foram excluídas 1934 crianças da amostra por não atenderem aos critérios de inclusão. Na avaliação clínica, foram analisados, as relações inter-arcos nos sentidos transversal, vertical e sagital, a relação intra-arcos, o perfil e padrão de crescimento, a presença de assimetria, além de hábitos deletérios. RESULTADOS: da amostragem total (1446), 80,29% apresentou maloclusão, sendo mais prevalente no gênero feminino (81,34%) e na faixa etária de nove a 12 anos (82,52%). A relação dentária mais prevalente foi a de Classe I (63,27%), o padrão facial mais encontrado foi o Padrão I (92,87%). As alterações oclusais inter-arcos mais encontradas foram a mordida profunda e a mordida aberta, as alterações intra-arcos predominantes foram os diastemas e as giroversões. De acordo com o Teste Qui-quadrado não houve significância entre maloclusão e as variáveis: presença de hábito, assimetria, diastemas e padrão facial. Houve diferença estatisticamente significante na prevalência das variáveis: mordida aberta, mordida profunda, diastemas e dos hábitos de sucção (dedo, chupeta, mamadeira) e onicofagia quando comparada as duas faixas etárias estudadas. CONCLUSÃO: as maloclusões acometem a maior parte das crianças nessa faixa etária, tendo origem predominantemente dentária e com pouco ou nenhum comprometimento facial, evidenciando a necessidade da intervenção precoce.

PURPOSE: to evaluate the prevalence of malocclusion in school children between 5 and 12 years-old enrolled in public schools in Araraquara_S.P. (Brazil). METHOD: based on the total number of children (7235) it was carried out the calculation of sampling, involving seven schools, a total of 3380 children. A pilot study and calibration of examiners preceded the study onset. 1934 children were excluded from the sample because they didn't apply for the inclusion criteria. In clinical evaluation, there were analyzed the inter-arcs in transverse, vertical and sagittal directions, intra-arch relationship, facial profile, growth pattern, presence of asymmetry, and harmful habits. RESULTS: of the total sample (1446), 80.29% had malocclusion, being more prevalent in females (81.34%) from 9 to 12 years-old (82.52%). The most prevalent dental relationship was Class I (63.27%), Standard I was the most found facial pattern (92.87%). The predominant inter-arch malocclusions were deep bite and anterior open bite, and the most prevalent intra-arches occlusal changes were spacing and rotation. According to the Chi-square test, there was no significance between malocclusion and the variables: presence of habit, asymmetry, spacing and facial profile. There was a statistically significant difference in the prevalence of variables: open bite, deep bite, spacing, sucking habits (sucking, pacifier, bottle) and the onychophagia habit, when compared the two age groups studied. CONCLUSION: malocclusions affect the most children in this age group, with predominantly dental origin and with little or no facial involvement, obviating the need for early intervention.