Dengue Virus Capsid Protein Usurps Lipid Droplets for Viral Particle Formation

Dengue virus is responsible for the highest rates of disease and mortality among the members of the Flavivirus genus. Dengue epidemics are still occurring around the world, indicating an urgent need of prophylactic vaccines and antivirals. In recent years, a great deal has been learned about the mechanisms of dengue virus genome amplification. However, little is known about the process by which the capsid protein recruits the viral genome during encapsidation. Here, we found that the mature capsid protein in the cytoplasm of dengue virus infected cells accumulates on the surface of ER-derived organelles named lipid droplets. Mutagenesis analysis using infectious dengue virus clones has identified specific hydrophobic amino acids, located in the center of the capsid protein, as key elements for lipid droplet association. Substitutions of amino acid L50 or L54 in the capsid protein disrupted lipid droplet targeting and impaired viral particle formation. We also report that dengue virus infection increases the number of lipid droplets per cell, suggesting a link between lipid droplet metabolism and viral replication. In this regard, we found that pharmacological manipulation of the amount of lipid droplets in the cell can be a means to control dengue virus replication. In addition, we developed a novel genetic system to dissociate cis-acting RNA replication elements from the capsid coding sequence. Using this system, we found that mislocalization of a mutated capsid protein decreased viral RNA amplification. We propose that lipid droplets play multiple roles during the viral life cycle; they could sequester the viral capsid protein early during infection and provide a scaffold for genome encapsidation

Loss of constitutive ABCB1 expression in breast cancer associated with worse prognosis

João Marcos de Azevedo Delou,1–3 Giselle Maria Vignal,4 Vanessa Índio-do-Brasil,2,5 Maria Theresa de Souza Accioly,4 Taiana Sousa Lopes da Silva,2 Diogo Nascimento Piranda,2,3 Marcelo Sobral-Leite,2,6 Marcelo Alex de Carvalho,2,7 Márcia Alves Marques Capella,1,8 Rosane Vianna-Jorge2,3,5 1Programa de Bioquímica e Biologia Celular, Instituto de Bioquímica Médica, Universidade Federal do Rio de Janeiro, 2Programa de Farmacologia, Coordenação de Pesquisa, Instituto Nacional de Câncer, 3Instituto de Ciências Biomédicas, Universidade Federal do Rio de Janeiro, 4Divisão de Patologia, Instituto Nacional de Câncer, 5Escola Nacional de Saúde Pública – FIOCRUZ, Rio de Janeiro, Brazil; 6Department of Molecular Pathology, Netherlands Cancer Institute, Amsterdam, The Netherlands; 7Instituto Federal do Rio de Janeiro, 8Instituto de Biofísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil Abstract: ABCB1 gene encodes an adenosine 5’-triphosphate–binding cassette transporter, which not only confers multidrug resistance phenotype in malignant cells, but is also present in several nonmalignant tissues. For the last thirty years, ABCB1 expression in breast cancer has been described by many authors, but the extent of expression differs among the studies, and there is no consensus regarding its potential role in carcinogenesis or in the tumor response to antineoplastic drugs. This study aimed to characterize the expression of ABCB1 in breast tumors as a function of genetic, clinical, and histopathological variables. The ABCB1 expression was also evaluated in nonmalignant mammary tissues adjacent to tumors and in benign lesions. The detection of ABCB1 protein was performed by immunohistochemistry in tissue specimens of excised breasts obtained from a prospective cohort of Brazilian women with breast cancer. The association of ABCB1 protein levels with ABCB1 mRNA, gene polymorphisms, and clinical and histopathological variables was also evaluated. The Kaplan–Meier curves and multivariate Cox regression analyses were conducted to identify independent predictors of disease-free survival of patients with breast cancer. ABCB1 was detected in 86.3% (656) of breast tumors, 98.8% (606) of nonmalignant mammary tissue adjacent to tumors, and 100% (28) of benign lesions. Reduced ABCB1 protein levels in breast tumors was associated with triple-negative subtype (adjusted odds ratio [ORadj] =0.24; 95% confidence interval [CI] =0.13–0.45), lymph node status < pN2 (ORadj =0.27; 95% CI =0.10–0.71), tumor size >2 cm (ORadj =0.55; 95% CI =0.32–0.93), and hypertensive status (ORadj =0.42; 95% CI =0.24–0.73), and it was significantly associated with shorter disease-free survival, either for all breast cancer patients (p log-rank =0.012; hazard ratio [HR] =3.46; 95% CI =1.21–9.91) or for those with triple-negative tumors (p log-rank =0.007; HR =11.41; 95% CI =1.29–100.67). The loss of constitutive ABCB1 expression in breast cancer, especially in triple-negative tumors, seems to indicate a subgroup of worse prognosis. Keywords: multidrug resistance, single-nucleotide polymorphisms, immunohistochemistry, disease-free survival, triple-negative breast cancer, hypertensio

Implementation of a pharmacogenomic program in a Brazilian public institution

This narrative review describes implementation, current status and perspectives of a pharmacogenomic (PGx) program at the Brazilian National Cancer Institute (INCA), targeting the cancer chemotherapeutic drugs – fluoropyrimidines, irinotecan and thiopurines. This initiative, designed as a research project, was supported by a grant from the Brazilian Ministry of Health. A dedicated task force developed standard operational procedures from recruitment of patients to creating PGx reports with dosing recommendations, which were successfully applied to test 100 gastrointestinal cancer INCA outpatients and 162 acute lymphoblastic leukemia pediatric patients from INCA and seven other hospitals. The program has been subsequently expanded to include gastrointestinal cancer patients from three additional cancer treatment centers. We anticipate implementation of routine pre-emptive PGx testing at INCA but acknowledge challenges associated with this transition, such as continuous financing support, availability of trained personnel, adoption of the PGx-informed prescription by the clinical staff and, ultimately, evidence of cost–effectiveness