The genetic era of childhood cancer: Identification of high-risk patients and germline sequencing approaches.

Childhood cancer is a leading cause of death by disease in children ages 5-14, for which there are no preventive strategies. Due to early-age of diagnosis and short period of exposure to environmental factors, increasing evidence suggests childhood cancer could have strong association with germline alterations in predisposition cancer genes but, their frequency and distribution are largely unknown. Several efforts have been made to develop tools to identify children with increased risk of cancer who may benefit from genetic testing but their validation and application on a large scale is necessary. Research on genetic bases of childhood cancer is ongoing, in which several approaches for the identification of genetic variants related to cancer predisposition have been used. In this paper, we discuss the updated efforts, strategies, molecular mechanisms and clinical implications for germline predisposition gene alterations and the characterization of risk variants in childhood cancer.

Complementary genomic approaches highlight the PI3K/mTOR pathway as a common vulnerability in osteosarcoma.

Osteosarcoma is the most common primary bone tumor, yet there have been no substantial advances in treatment or survival in three decades. We examined 59 tumor/normal pairs by whole-exome, whole-genome, and RNA-sequencing. Only the TP53 gene was mutated at significant frequency across all samples. The mean nonsilent somatic mutation rate was 1.2 mutations per megabase, and there was a median of 230 somatic rearrangements per tumor. Complex chains of rearrangements and localized hypermutation were detected in almost all cases. Given the intertumor heterogeneity, the extent of genomic instability, and the difficulty in acquiring a large sample size in a rare tumor, we used several methods to identify genomic events contributing to osteosarcoma survival. Pathway analysis, a heuristic analytic algorithm, a comparative oncology approach, and an shRNA screen converged on the phosphatidylinositol 3-kinase/mammalian target of rapamycin (PI3K/mTOR) pathway as a central vulnerability for therapeutic exploitation in osteosarcoma. Osteosarcoma cell lines are responsive to pharmacologic and genetic inhibition of the PI3K/mTOR pathway both in vitro and in vivo.

Cementum proteins: role in cementogenesis, biomineralization, periodontium formation and regeneration.

Destruction of the periodontium is normally associated with periodontal disease, although many other factors, such as trauma, aging, infections, orthodontic tooth movement and systemic and genetic diseases, can contribute to this process. Strategies (such as guided tissue regeneration) have been developed to guide and control regeneration using bioresorbable membranes and bone grafts. Although effective to a certain point, these strategies have the problem that they are not predictable and do not completely restore the architecture of the original periodontium. To achieve complete repair and regeneration it is necessary to recapitulate the developmental process with complete formation of cementum, bone and periodontal ligament fibers. Detailed knowledge of the biology of cementum is key for understanding how the periodontium functions, identifying pathological issues and for developing successful therapies for repair and regeneration of damaged periodontal tissue. It is the purpose of this review to focus on the role of cementum and its specific components in the formation, repair and regeneration of the periodontium. As cementum is a matrix rich in growth factors that could influence the activities of various periodontal cell types, this review will examine the characteristics of cementum, its composition and the role of cementum components, especially the cementum protein-1, during the process of cementogenesis, and their potential usefulness for regeneration of the periodontal structures in a predictable therapeutic manner.

Germline mutations in pediatric cancer cohort with mixed-ancestry Mexicans.

BACKGROUND: Childhood cancer is one of the primary causes of disease-related death in 5- to 14-year-old children and currently no prevention strategies exist to reduce the incidence of this disease. Childhood cancer has a larger hereditary component compared with cancer in adults. Few genetic studies have been conducted on children with cancer. Additionally, Latin American populations are underrepresented in genomic studies compared with other populations. Therefore, the aim of this study is to analyze germline mutations in a group of mixed-ancestry Mexican pediatric patients with solid and hematological cancers. METHODS: We analyzed genetic variants from 40 Mexican childhood cancer patients and their relatives. DNA from saliva or blood samples was used for whole-exome sequencing. All variants were identified following GATK best practices. RESULTS: We found that six patients (15%) were carriers of germline mutations in CDKN2A, CHEK2, DICER1, FANCA, MSH6, MUTYH, NF1, and SBDS cancer predisposition genes, and additional new variants predicted to be deleterious by in silico algorithms. A population genetics analysis detected five components consistent with the demographic models assumed for modern mixed-ancestry Mexicans. CONCLUSIONS: This report identifies potential genetic risk factors and provides a better understanding of the underlying mechanisms of childhood cancer in this population.

A novel model to culture cells from giant cell tumor of bone using three-dimensional (3D) polycaprolactone scaffold.

Two-dimensional (2D) culture of cells from giant cell tumor of bone (GCTB) is affected by loss of the multinucleated giant cells in subsequent passages. Therefore, there is limited time to study GCTB with all its histological components in 2D culture. Here, we explored the possibility of culturing GCTB cells on a polycaprolactone (PCL)-printed scaffold. We also evaluated the viability of the cultured cells and their adherence to the PCL scaffold at day 14 days using immunofluorescence analysis with calcein, vinculin, and phalloidin. Using the histological technique with hematoxylin and eosin staining, we observed all the histological components of GCTB in this 3D model. Immunohistochemical assays with cathepsin K, p63, and receptor activator of nuclear factor (NF)-κB ligand (RANKL) yielded positive results in this construct, which allowed us to confirm that the seeded cells maintained the expression of GCTB markers. Based on these findings, we concluded that the PCL scaffold is an efficient model to culture GCTB cells, and the cell viability and adherence to the scaffold can be preserved for up to 14 days. Moreover, this model can also be used in subsequent studies to assess in vitro cell-cell interactions and antineoplastic efficacy of certain agents to establish a treatment against GCTB.

Genomics of a pediatric ovarian fibrosarcoma. Association with the DICER1 syndrome.

Ovarian fibrosarcomas are extremely rare tumors with little genomic information available to date. In the present report we present the tumoral exome and transcriptome and the germinal exome of an ovarian fibrosarcoma from a 9-years old child. We found a paucity of mutations (0.77/Mb) and CNV alterations. Of these, the most relevant were a point mutation in the metal-binding site of the microRNA-processing DICER1 enzyme and a frame-shift alteration in the tumor suppressor gene NF1. We validated a germinal truncating mutation in DICER1, which was consistent with a DICER1 Syndrome diagnosis, providing the first example of an ovarian fibrosarcoma as the presenting neoplasia in this syndrome. Network and enrichment analyses showed that both a mesenchymal signature and a Hedgehog cascade could be driving the progression of this tumor. We were also able to find a global lincRNA deregulation, as the number of lincRNAs transcripts expressed in the tumor was decreased, with a concomitant upregulation of previously described non-coding transcripts associated with cancer, such as MALAT1, MIR181A1HG, CASC1, XIST and FENDRR. DICER1 Syndrome should be considered as a possible diagnosis in children ovarian fibrosarcoma. The role of lncRNAs in neoplasias associated with DICER1 alterations need to be studied in more detail.

Analysis of Gene Candidate SNP and Ancestral Origin Associated to Obesity and Postoperative Weight Loss in a Cohort of Obese Patients Undergoing RYGB.

INTRODUCTION: Obesity is the result of a complex interaction between multiple genetic traits and psychological, behavioral, nutritional and environmental factors. OBJECTIVES: The aims of the study were (a) to comparatively evaluate the presence of 20 candidate gene single nucleotide polymorphisms (SNPs) in morbidly obese patients, (2) their association to comorbid conditions and (3) their impact on weight loss after a Roux-en-Y gastric bypass (RYGB). PATIENTS AND METHODS: Two hundred forty-nine patients were eligible for this study. Clinical, anthropometric, biochemical and demographic variables were analyzed. Body mass index (BMI) and composition were assessed by bioelectrical impedance. Twenty SNPs were included for analysis. RESULTS: There were 168 Mexican mestizos (67.5 %) and 81 (32.5 %) patients with other ancestral origin. One hundred fifty-nine (64.1 %) were females. Mean ± SD age of the general cohort was 41.1 ± 11.3 years (17-71). Preoperative mean ± SD BMI was 42.5 ± 6.5 kg/m2. There were no significant differences between mestizo and non-mestizo for most SNPs except for IFI, LIPC, and ST8SIA2. FTO (OR = 1.71; CI95 % = 1.14-2.57; p = 0.008) and APOB (OR = 0.31; CI95 % = 0.14-0.72; p = 0.004) result is statistically associated to high blood pressure and FTO (OR = 2.0; CI95 % = 1.3-3.1; p = 0.001), GNB3 (OR = 2.69; CI95 % = 1.0-7.2; p = 0.04), IFI30 (OR = 2.0; CI95 % = 1.16-3.6; p = 0.01), and MC4R (OR = 1.81; CI95 % = 1.13-2.9; p = 0.01) to type 2 diabetes (T2D). Based on ANOVA analysis, POMC (rs1042571) was the SNP most significantly associated to a higher weight loss after RYGB. CONCLUSIONS: Obese patients have similar SNP frequencies. Several SNP results are statistically associated to high blood pressure and T2D. POMC was significantly associated to a higher surgically induced weight loss.

Homozygosity for the WRN Helicase-Inactivating Variant, R834C, does not confer a Werner syndrome clinical phenotype.

Loss-of-function mutations in the WRN helicase gene cause Werner syndrome- a progeroid syndrome with an elevated risk of cancer and other age-associated diseases. Large numbers of single nucleotide polymorphisms have been identified in WRN. We report here the organismal, cellular, and molecular phenotypes of variant rs3087425 (c. 2500C > T) that results in an arginine to cysteine substitution at residue 834 (R834C) and up to 90% reduction of WRN helicase activity. This variant is present at a high (5%) frequency in Mexico, where we identified 153 heterozygous and three homozygous individuals among 3,130 genotyped subjects. Family studies of probands identified ten additional TT homozygotes. Biochemical analysis of WRN protein purified from TT lymphoblast cell lines confirmed that the R834C substitution strongly and selectively reduces WRN helicase, but not exonuclease activity. Replication track analyses showed reduced replication fork progression in some homozygous cells following DNA replication stress. Among the thirteen TT homozygotes, we identified a previously unreported and statistically significant gender bias in favor of males (p = 0.0016), but none of the clinical findings associated with Werner syndrome. Our results indicate that WRN helicase activity alone is not rate-limiting for the development of clinical WS.

CEMP1 Induces Transformation in Human Gingival Fibroblasts.

Cementum Protein 1 (CEMP1) is a key regulator of cementogenesis. CEMP1 promotes cell attachment, differentiation, deposition rate, composition, and morphology of hydroxyapatite crystals formed by human cementoblastic cells. Its expression is restricted to cementoblasts and progenitor cell subpopulations present in the periodontal ligament. CEMP1 transfection into non-osteogenic cells such as adult human gingival fibroblasts results in differentiation of these cells into a "mineralizing" cell phenotype. Other studies have shown evidence that CEMP1 could have a therapeutic potential for the treatment of bone defects and regeneration of other mineralized tissues. To better understand CEMP1's biological effects in vitro we investigated the consequences of its expression in human gingival fibroblasts (HGF) growing in non-mineralizing media by comparing gene expression profiles. We identified several mRNAs whose expression is modified by CEMP1 induction in HGF cells. Enrichment analysis showed that several of these newly expressed genes are involved in oncogenesis. Our results suggest that CEMP1 causes the transformation of HGF and NIH3T3 cells. CEMP1 is overexpressed in cancer cell lines. We also determined that the region spanning the CEMP1 locus is commonly amplified in a variety of cancers, and finally we found significant overexpression of CEMP1 in leukemia, cervix, breast, prostate and lung cancer. Our findings suggest that CEMP1 exerts modulation of a number of cellular genes, cellular development, cellular growth, cell death, and cell cycle, and molecules associated with cancer.

Shifts in developmental timing, and not increased levels of experience-dependent neuronal activity, promote barrel expansion in the primary somatosensory cortex of rats enucleated at birth.

Birth-enucleated rodents display enlarged representations of whiskers (i.e., barrels of the posteromedial subfield) in the primary somatosensory cortex. Although the historical view maintains that barrel expansion is due to incremental increases in neuronal activity along the trigeminal pathway during postnatal development, recent evidence obtained in experimental models of intramodal plasticity challenges this view. Here, we re-evaluate the role of experience-dependent neuronal activity on barrel expansion in birth-enucleated rats by combining various anatomical methods and sensory deprivation paradigms. We show that barrels in birth-enucleated rats were already enlarged by the end of the first week of life and had levels of metabolic activity comparable to those in control rats at different ages. Dewhiskering after the postnatal period of barrel formation did not prevent barrel expansion in adult, birth-enucleated rats. Further, dark rearing and enucleation after barrel formation did not lead to expanded barrels in adult brains. Because incremental increases of somatosensory experience did not promote barrel expansion in birth-enucleated rats, we explored whether shifts of the developmental timing could better explain barrel expansion during the first week of life. Accordingly, birth-enucleated rats show earlier formation of barrels, accelerated growth of somatosensory thalamocortical afferents, and an earlier H4 deacetylation. Interestingly, when H4 deacetylation was prevented with a histone deacetylases inhibitor (valproic acid), barrel specification timing returned to normal and barrel expansion did not occur. Thus, we provide evidence supporting that shifts in developmental timing modulated through epigenetic mechanisms, and not increased levels of experience dependent neuronal activity, promote barrel expansion in the primary somatosensory cortex of rats enucleated at birth.

Trastorno por atracón, depresión, ansiedad y gen del neuropéptido relacionado con Agouti en pacientes con cirugía bariátrica / Binge eating disorder, depression, anxiety and Agouti gen related neuropeptide in patients with bariatric surgery

Resumen El objetivo de este estudio retroprospectivo fue analizar la relación de la sintomatología de depresión, de ansiedad y el trastorno por atracón (TPA) con el gen del neuropéptido relacionado con Agouti en pacientes sometidos a cirugía bariátrica. Participó una cohorte de 249 adultos (edad media = 41.1, DE =11.3), 64.1% mujeres y 35.9% hombres. La evaluación de la sintomatología depresiva, de ansiedad y de TPA se llevó a cabo a través de una entrevista semiestructurada. Además, se calculó el índice de masa corporal y se tomaron muestras de sangre para realizar un análisis de discriminación alélica. Del total de pacientes, un 20.2% fueron diagnosticados con TPA, encontrando una asociación de este trastorno con una menor pérdida de peso posterior a la cirugía bariátrica a los 6,12, 18 y 24 meses. Las medidas de depresión y de ansiedad no difirieron entre pacientes con TPA vs. sin TPA. Los pacientes con un alelo mutante en el gen del neuropéptido relacionado con Agouti tuvieron un riesgo 2.6 veces mayor de presentar TPA (IC 95% 1.0-6.8; p = 0.04). Además, el TPA parece ser más frecuente en pacientes con el gen del neuropéptido relacionado con Agouti mutado. Destaca la necesidad de que en el estudio de la obesidad se aborden tanto los aspectos psicológicos como los genéticos.

Abstract The objective of this retrospective study was to analyze the relationship between the symptoms of depression, anxiety and binge eating disorder (BED) with the gene related to the Agouti neuropeptide in patients undergoing bariatric surgery. A cohort of 249 adults (average age = 41.1, SD = 11.3), 64.1% women and 35.9% men, were included. The assessment of depression, anxiety and BED symptoms was carried out through a semi-structured interview. In addition, the body mass index was calculated, and blood samples were taken for an allelic discrimination analysis. Of the total number of patients 20.2% were diagnosed with BED, finding an association of this disorder with a lower weight loss after bariatric surgery at 6, 12, 18 and 24 months. The measures of depression and anxiety did not differ between patients with BED vs. without BED. Patients with a mutant allele in the gene related to the Agouti neuropeptide were 2.6 times more likely to present BED (95% C11.0-6.8, P = 0.04). In addition, BED appears to be more frequent in patients with a gene related to the Agouti neuropeptide mutated. When obesity is studied, it is emphasized the need to address both psychological and genetic factors.