Distinct Driver Pathway Enrichments and a High Prevalence of TSC2 Mutations in Right Colon Cancer in Chile: A Preliminary Comparative Analysis.

Colorectal cancer (CRC) is the second leading cause of cancer deaths globally. While ethnic differences in driver gene mutations have been documented, the South American population remains understudied at the genomic level, despite facing a rising burden of CRC. We analyzed tumors of 40 Chilean CRC patients (Chp) using next-generation sequencing and compared them to data from mainly Caucasian cohorts (TCGA and MSK-IMPACT). We identified 388 mutations in 96 out of 135 genes, with TP53 (45%), KRAS (30%), PIK3CA (22.5%), ATM (20%), and POLE (20%) being the most frequently mutated. TSC2 mutations were associated with right colon cancer (44.44% in RCRC vs. 6.45% in LCRC, p-value = 0.016), and overall frequency was higher compared to TCGA (p-value = 1.847 × 10-5) and MSK-IMPACT cohorts (p-value = 3.062 × 10-2). Limited sample size restricts definitive conclusions, but our data suggest potential differences in driver mutations for Chilean patients, being that the RTK-RAS oncogenic pathway is less affected and the PI3K pathway is more altered in Chp compared to TCGA (45% vs. 25.56%, respectively). The prevalence of actionable pathways and driver mutations can guide therapeutic choices, but can also impact treatment effectiveness. Thus, these findings warrant further investigation in larger Chilean cohorts to confirm these initial observations. Understanding population-specific driver mutations can guide the development of precision medicine programs for CRC patients.

Development and internal validation of a multifactorial risk prediction model for gallbladder cancer in a high-incidence country.

Since 2006, Chile has been implementing a gallbladder cancer (GBC) prevention program based on prophylactic cholecystectomy for gallstone patients aged 35 to 49 years. The effectiveness of this prevention program has not yet been comprehensively evaluated. We conducted a retrospective study of 473 Chilean GBC patients and 2137 population-based controls to develop and internally validate three GBC risk prediction models. The Baseline Model accounted for gallstones while adjusting for sex and birth year. Enhanced Model I also included the non-genetic risk factors: body mass index, educational level, Mapuche surnames, number of children and family history of GBC. Enhanced Model II further included Mapuche ancestry and the genotype for rs17209837. Multiple Cox regression was applied to assess the predictive performance, quantified by the area under the precision-recall curve (AUC-PRC) and the number of cholecystectomies needed (NCN) to prevent one case of GBC at age 70 years. The AUC-PRC for the Baseline Model (0.44%, 95%CI 0.42-0.46) increased by 0.22 (95%CI 0.15-0.29) when non-genetic factors were included, and by 0.25 (95%CI 0.20-0.30) when incorporating non-genetic and genetic factors. The overall NCN for Chileans with gallstones (115, 95%CI 104-131) decreased to 92 (95%CI 60-128) for Chileans with a higher risk than the median according to Enhanced Model I, and to 80 (95%CI 59-110) according to Enhanced Model II. In conclusion, age, sex and gallstones are strong risk factors for GBC, but consideration of other non-genetic factors and individual genotype data improves risk prediction and may optimize allocation of financial resources and surgical capacity.

Epigenome-Wide Analysis of Methylation Changes in the Sequence of Gallstone Disease, Dysplasia, and Gallbladder Cancer.

BACKGROUND AND AIMS: Gallbladder cancer (GBC) is a highly aggressive malignancy of the biliary tract. Most cases of GBC are diagnosed in low-income and middle-income countries, and research into this disease has long been limited. In this study we therefore investigate the epigenetic changes along the model of GBC carcinogenesis represented by the sequence gallstone disease → dysplasia → GBC in Chile, the country with the highest incidence of GBC worldwide. APPROACH AND RESULTS: To perform epigenome-wide methylation profiling, genomic DNA extracted from sections of formalin-fixed, paraffin-embedded gallbladder tissue was analyzed using Illumina Infinium MethylationEPIC BeadChips. Preprocessed, quality-controlled data from 82 samples (gallstones n = 32, low-grade dysplasia n = 13, high-grade dysplasia n = 9, GBC n = 28) were available to identify differentially methylated markers, regions, and pathways as well as changes in copy number variations (CNVs). The number and magnitude of epigenetic changes increased with disease development and predominantly involved the hypermethylation of cytosine-guanine dinucleotide islands and gene promoter regions. The methylation of genes implicated in Wnt signaling, Hedgehog signaling, and tumor suppression increased with tumor grade. CNVs also increased with GBC development and affected cyclin-dependent kinase inhibitor 2A, MDM2 proto-oncogene, tumor protein P53, and cyclin D1 genes. Gains in the targetable Erb-B2 receptor tyrosine kinase 2 gene were detected in 14% of GBC samples. CONCLUSIONS: Our results indicate that GBC carcinogenesis comprises three main methylation stages: early (gallstone disease and low-grade dysplasia), intermediate (high-grade dysplasia), and late (GBC). The identified gradual changes in methylation and CNVs may help to enhance our understanding of the mechanisms underlying this aggressive disease and eventually lead to improved treatment and early diagnosis of GBC.

Gallstones, Body Mass Index, C-Reactive Protein, and Gallbladder Cancer: Mendelian Randomization Analysis of Chilean and European Genotype Data.

BACKGROUND AND AIMS: Gallbladder cancer (GBC) is a neglected disease with substantial geographical variability: Chile shows the highest incidence worldwide, while GBC is relatively rare in Europe. Here, we investigate the causal effects of risk factors considered in current GBC prevention programs as well as C-reactive protein (CRP) level as a marker of chronic inflammation. APPROACH AND RESULTS: We applied two-sample Mendelian randomization (MR) using publicly available data and our own data from a retrospective Chilean and a prospective European study. Causality was assessed by inverse variance weighted (IVW), MR-Egger regression, and weighted median estimates complemented with sensitivity analyses on potential heterogeneity and pleiotropy, two-step MR, and mediation analysis. We found evidence for a causal effect of gallstone disease on GBC risk in Chileans (P = 9 × 10-5 ) and Europeans (P = 9 × 10-5 ). A genetically elevated body mass index (BMI) increased GBC risk in Chileans (P = 0.03), while higher CRP concentrations increased GBC risk in Europeans (P = 4.1 × 10-6 ). European results suggest causal effects of BMI on gallstone disease (P = 0.008); public Chilean data were not, however, available to enable assessment of the mediation effects among causal GBC risk factors. CONCLUSIONS: Two risk factors considered in the current Chilean program for GBC prevention are causally linked to GBC risk: gallstones and BMI. For Europeans, BMI showed a causal effect on gallstone risk, which was itself causally linked to GBC risk.

[The contribution of omic sciences for the management of cancer in Chile]. / Planificación sanitaria a partir de los datos ómicos: ¿es posible esta idea para Chile?

With or without a COVID19 pandemic, cancer is and will continue to be one of the greatest health challenges on the planet. In Chile, during 2016, this disease was the second cause of death in the country and during 2019, it was the first cause in seven Chilean regions, surpassing cardiovascular diseases. With the advent of precision medicine as a powerful tool for cancer control, it is necessary to have genomic, proteomic, and molecular data in general, ideally on a population scale. This is essential for decision-making, for example in public and private oncology, to be as cost-effective as possible. Chile has a mass of high-quality researchers in cancer. However, until today the investment in research and development is far below the peers in the OECD. In this work we put into perspective the role of precision medicine and omic sciences as essential tools for public health. We offer a brief national diagnosis of the knowledge collected to date by the local scientific community regarding onco-genomic data from our own population. We finally discuss the potential behind the strengthening of this scientific knowledge, aiming to optimize the comprehensive management of cancer.

Subtypes of Native American ancestry and leading causes of death: Mapuche ancestry-specific associations with gallbladder cancer risk in Chile.

Latin Americans are highly heterogeneous regarding the type of Native American ancestry. Consideration of specific associations with common diseases may lead to substantial advances in unraveling of disease etiology and disease prevention. Here we investigate possible associations between the type of Native American ancestry and leading causes of death. After an aggregate-data study based on genome-wide genotype data from 1805 admixed Chileans and 639,789 deaths, we validate an identified association with gallbladder cancer relying on individual data from 64 gallbladder cancer patients, with and without a family history, and 170 healthy controls. Native American proportions were markedly underestimated when the two main types of Native American ancestry in Chile, originated from the Mapuche and Aymara indigenous peoples, were combined together. Consideration of the type of Native American ancestry was crucial to identify disease associations. Native American ancestry showed no association with gallbladder cancer mortality (P = 0.26). By contrast, each 1% increase in the Mapuche proportion represented a 3.7% increased mortality risk by gallbladder cancer (95%CI 3.1-4.3%, P = 6×10-27). Individual-data results and extensive sensitivity analyses confirmed the association between Mapuche ancestry and gallbladder cancer. Increasing Mapuche proportions were also associated with an increased mortality due to asthma and, interestingly, with a decreased mortality by diabetes. The mortality due to skin, bladder, larynx, bronchus and lung cancers increased with increasing Aymara proportions. Described methods should be considered in future studies on human population genetics and human health. Complementary individual-based studies are needed to apportion the genetic and non-genetic components of associations identified relying on aggregate-data.

TRPM4 channel is involved in regulating epithelial to mesenchymal transition, migration, and invasion of prostate cancer cell lines.

Transient Receptor Potential Melastatin 4 (TRPM4) is a Ca2+ -activated and voltage-dependent monovalent cation channel, which depolarizes the plasma cell membrane, thereby modulating Ca2+ influx across Ca2+ -permeable pathways. TRPM4 is involved in different physiological processes such as T cell activation and the migration of endothelial and certain immune cells. Overexpression of this channel has been reported in various types of tumors including prostate cancer. In this study, a significant overexpression of TRPM4 was found only in samples from cancer with a Gleason score higher than 7, which are more likely to spread. To evaluate whether TRPM4 overexpression was related to the spreading capability of tumors, TRPM4 was knockdown by using shRNAs in PC3 prostate cancer cells and the effect on cellular migration and invasion was analyzed. PC3 cells with reduced levels of TRPM4 (shTRPM4) display a decrease of the migration/invasion capability. A reduction in the expression of Snail1, a canonical epithelial to mesenchymal transition (EMT) transcription factor, was also observed. Consistently, these cells showed a significant change in the expression of key EMT markers such as MMP9, E-cadherin/N-cadherin, and vimentin, indicating a partial reversion of the EMT process. Whereas, the overexpression of TRPM4 in LnCaP cells resulted in increased levels of Snail1, reduction in the expression of E-cadherin and increase in their migration potential. This study suggests a new and indirect mechanism of regulation of migration/invasion process by TRPM4 in prostate cancer cells, by inducing the expression of Snail1 gene and consequently, increasing the EMT.

ADAR1-mediated RNA-editing of 3'UTRs in breast cancer.

BACKGROUND: Whole transcriptome RNA variant analyses have shown that adenosine deaminases acting on RNA (ADAR) enzymes modify a large proportion of cellular RNAs, contributing to transcriptome diversity and cancer evolution. Despite the advances in the understanding of ADAR function in breast cancer, ADAR RNA editing functional consequences are not fully addressed. RESULTS: We characterized A to G(I) mRNA editing in 81 breast cell lines, showing increased editing at 3'UTR and exonic regions in breast cancer cells compared to immortalized non-malignant cell lines. In addition, tumors from the BRCA TCGA cohort show a 24% increase in editing over normal breast samples when looking at 571 well-characterized UTRs targeted by ADAR1. Basal-like subtype breast cancer patients with high level of ADAR1 mRNA expression shows a worse clinical outcome and increased editing in their 3'UTRs. Interestingly, editing was particularly increased in the 3'UTRs of ATM, GINS4 and POLH transcripts in tumors, which correlated with their mRNA expression. We confirmed the role of ADAR1 in this regulation using a shRNA in a breast cancer cell line (ZR-75-1). CONCLUSIONS: Altogether, these results revealed a significant association between the mRNA editing in genes related to cancer-relevant pathways and clinical outcomes, suggesting an important role of ADAR1 expression and function in breast cancer.

The Ski Protein is Involved in the Transformation Pathway of Aurora Kinase A.

Oncogenic kinase Aurora A (AURKA) has been found to be overexpresed in several tumors including colorectal, breast, and hematological cancers. Overexpression of AURKA induces centrosome amplification and aneuploidy and it is related with cancer progression and poor prognosis. Here we show that AURKA phosphorylates in vitro the transcripcional co-repressor Ski on aminoacids Ser326 and Ser383. Phosphorylations on these aminoacids decreased Ski protein half-life. Reduced levels of Ski resulted in centrosomes amplification and multipolar spindles formation, same as AURKA overexpressing cells. Importantly, overexpression of Ski wild type, but not S326D and S383D mutants inhibited centrosome amplification and cellular transformation induced by AURKA. Altogether, these results suggest that the Ski protein is a target in the transformation pathway mediated by the AURKA oncogene.

Daam1a mediates asymmetric habenular morphogenesis by regulating dendritic and axonal outgrowth.

Although progress has been made in resolving the genetic pathways that specify neuronal asymmetries in the brain, little is known about genes that mediate the development of structural asymmetries between neurons on left and right. In this study, we identify daam1a as an asymmetric component of the signalling pathways leading to asymmetric morphogenesis of the habenulae in zebrafish. Daam1a is a member of the Formin family of actin-binding proteins and the extent of Daam1a expression in habenular neuron dendrites mirrors the asymmetric growth of habenular neuropil between left and right. Local loss and gain of Daam1a function affects neither cell number nor subtype organisation but leads to a decrease or increase of neuropil, respectively. Daam1a therefore plays a key role in the asymmetric growth of habenular neuropil downstream of the pathways that specify asymmetric cellular domains in the habenulae. In addition, Daam1a mediates the development of habenular efferent connectivity as local loss and gain of Daam1a function impairs or enhances, respectively, the growth of habenular neuron terminals in the interpeduncular nucleus. Abrogation of Daam1a disrupts the growth of both dendritic and axonal processes and results in disorganised filamentous actin and α-tubulin. Our results indicate that Daam1a plays a key role in asymmetric habenular morphogenesis mediating the growth of dendritic and axonal processes in dorsal habenular neurons.

Inhibition of the 3-hydroxy-3-methyl-glutaryl-CoA reductase induces orofacial defects in zebrafish.

BACKGROUND: Orofacial clefts (OFCs) are common birth defects, which include a range of disorders with a complex etiology affecting formation of craniofacial structures. Some forms of syndromic OFCs are produced by defects in the cholesterol pathway. The principal enzyme of the cholesterol pathway is the 3-hydroxy-3-methyl-glutaryl-CoA reductase (HMGCR). Our aim is to study whether defects of HMGCR function would produce orofacial malformation similar to those found in disorders of cholesterol synthesis. METHODS: We used zebrafish hmgcrb mutants and HMGCR inhibition assay using atorvastatin during early and late stages of orofacial morphogenesis in zebrafish. To describe craniofacial phenotypes, we stained cartilage and bone and performed in situ hybridization using known craniofacial markers. Also, we visualized neural crest cell migration in a transgenic fish. RESULTS: Our results showed that mutants displayed loss of cartilage and diminished orofacial outgrowth, and in some cases palatal cleft. Late treatments with statin show a similar phenotype. Affected-siblings displayed a moderate phenotype, whereas early-treated embryos had a minor cleft. We found reduced expression of the downstream component of Sonic Hedgehog-signaling gli1 in ventral brain, oral ectoderm, and pharyngeal endoderm in mutants and in late atorvastatin-treated embryos. CONCLUSION: Our results suggest that HMGCR loss-of-function primarily affects postmigratory cranial neural crest cells through abnormal Sonic Hedgehog signaling, probably induced by reduction in metabolites of the cholesterol pathway. Malformation severity correlates with the grade of HMGCR inhibition, developmental stage of its disruption, and probably with availability of maternal lipids. Together, our results might help to understand the spectrum of orofacial phenotypes found in cholesterol synthesis disorders. Birth Defects Research (Part A) 106:814-830, 2016. © 2016 Wiley Periodicals, Inc.

A Comprehensive Analysis of the Effect of A>I(G) RNA-Editing Sites on Genotoxic Drug Response and Progression in Breast Cancer.

Dysregulated A>I(G) RNA editing, which is mainly catalyzed by ADAR1 and is a type of post-transcriptional modification, has been linked to cancer. A low response to therapy in breast cancer (BC) is a significant contributor to mortality. However, it remains unclear if there is an association between A>I(G) RNA-edited sites and sensitivity to genotoxic drugs. To address this issue, we employed a stringent bioinformatics approach to identify differentially RNA-edited sites (DESs) associated with low or high sensitivity (FDR 0.1, log2 fold change 2.5) according to the IC50 of PARP inhibitors, anthracyclines, and alkylating agents using WGS/RNA-seq data in BC cell lines. We then validated these findings in patients with basal subtype BC. These DESs are mainly located in non-coding regions, but a lesser proportion in coding regions showed predicted deleterious consequences. Notably, some of these DESs are previously reported as oncogenic variants, and in genes related to DNA damage repair, drug metabolism, gene regulation, the cell cycle, and immune response. In patients with BC, we uncovered DESs predominantly in immune response genes, and a subset with a significant association (log-rank test p < 0.05) between RNA editing level in LSR, SMPDL3B, HTRA4, and LL22NC03-80A10.6 genes, and progression-free survival. Our findings provide a landscape of RNA-edited sites that may be involved in drug response mechanisms, highlighting the value of A>I(G) RNA editing in clinical outcomes for BC.

Classification of patients with metastatic colorectal cancer into consensus molecular subtypes into real-world: A pilot study.

BACKGROUND: Colorectal cancer is a complex disease with high mortality rates. Over time, the treatment of metastatic colorectal cancer (mCRC) has gradually improved due to the development of modern chemotherapy and targeted therapy regimens. However, due to the inherent heterogeneity of this condition, identifying reliable predictive biomarkers for targeted therapies remains challenging. A recent promising classification system-the consensus molecular subtype (CMS) system-offers the potential to categorize mCRC patients based on their unique biological and molecular characteristics. Four distinct CMS categories have been defined: immune (CMS1), canonical (CMS2), metabolic (CMS3), and mesenchymal (CMS4). Nevertheless, there is currently no standardized protocol for accurately classifying patients into CMS categories. To address this challenge, reverse transcription polymerase chain reaction (RT-qPCR) and next-generation genomic sequencing (NGS) techniques may hold promise for precisely classifying mCRC patients into their CMSs. AIM: To investigate if mCRC patients can be classified into CMS categories using a standardized molecular biology workflow. METHODS: This observational study was conducted at the University of Chile Clinical Hospital and included patients with unresectable mCRC who were undergoing systemic treatment with chemotherapy and/or targeted therapy. Molecular biology techniques were employed to analyse primary tumour samples from these patients. RT-qPCR was utilized to assess the expression of genes associated with fibrosis (TGF-ß and ß-catenin) and cell growth pathways (c-MYC). NGS using a 25-gene panel (TumorSec) was performed to identify specific genomic mutations. The patients were then classified into one of the four CMS categories according to the clinical consensus of a Tumour Board. Informed consent was obtained from all the patients prior to their participation in this study. All techniques were conducted at University of Chile. RESULTS: Twenty-six patients were studied with the techniques and then evaluated by the Tumour Board to determine the specific CMS. Among them, 23% (n = 6), 19% (n = 5), 31% (n = 8), and 19% (n = 5) were classified as CMS1, CMS2, CMS3, and CMS4, respectively. Additionally, 8% of patients (n = 2) could not be classified into any of the four CMS categories. The median overall survival of the total sample was 28 mo, and for CMS1, CMS2, CMS3 and CMS4 it was 11, 20, 30 and 45 mo respectively, with no statistically significant differences between groups. CONCLUSION: A molecular biology workflow and clinical consensus analysis can be used to accurately classify mCRC patients. This classification process, which divides patients into the four CMS categories, holds significant potential for improving research strategies and targeted therapies tailored to the specific characteristics of mCRC.

Acquired Secondary HER2 Mutations Enhance HER2/MAPK Signaling and Promote Resistance to HER2 Kinase Inhibition in Breast Cancer.

HER2 mutations drive the growth of a subset of breast cancers and are targeted with HER2 tyrosine kinase inhibitors (TKI) such as neratinib. However, acquired resistance is common and limits the durability of clinical responses. Most HER2-mutant breast cancers progressing on neratinib-based therapy acquire secondary mutations in HER2. It is unknown whether these secondary HER2 mutations, other than the HER2T798I gatekeeper mutation, are causal to neratinib resistance. Herein, we show that secondary acquired HER2T862A and HER2L755S mutations promote resistance to HER2 TKIs via enhanced HER2 activation and impaired neratinib binding. While cells expressing each acquired HER2 mutation alone were sensitive to neratinib, expression of acquired double mutations enhanced HER2 signaling and reduced neratinib sensitivity. Computational structural modeling suggested that secondary HER2 mutations stabilize the HER2 active state and reduce neratinib binding affinity. Cells expressing double HER2 mutations exhibited resistance to most HER2 TKIs but retained sensitivity to mobocertinib and poziotinib. Double-mutant cells showed enhanced MEK/ERK signaling, which was blocked by combined inhibition of HER2 and MEK. Together, these findings reveal the driver function of secondary HER2 mutations in resistance to HER2 inhibition and provide a potential treatment strategy to overcome acquired resistance to HER2 TKIs in HER2-mutant breast cancer. SIGNIFICANCE: HER2-mutant breast cancers acquire secondary HER2 mutations that drive resistance to HER2 tyrosine kinase inhibitors, which can be overcome by combined inhibition of HER2 and MEK.

Gallbladder Cancer Risk and Indigenous South American Mapuche Ancestry: Instrumental Variable Analysis Using Ancestry-Informative Markers.

A strong association between the proportion of indigenous South American Mapuche ancestry and the risk of gallbladder cancer (GBC) has been reported in observational studies. Chileans show the highest incidence of GBC worldwide, and the Mapuche are the largest indigenous people in Chile. We set out to assess the confounding-free effect of the individual proportion of Mapuche ancestry on GBC risk and to investigate the mediating effects of gallstone disease and body mass index (BMI) on this association. Genetic markers of Mapuche ancestry were selected based on the informativeness for assignment measure, and then used as instrumental variables in two-sample Mendelian randomization analyses and complementary sensitivity analyses. Results suggested a putatively causal effect of Mapuche ancestry on GBC risk (inverse variance-weighted (IVW) risk increase of 0.8% per 1% increase in Mapuche ancestry proportion, 95% CI 0.4% to 1.2%, p = 6.7 × 10-5) and also on gallstone disease (3.6% IVW risk increase, 95% CI 3.1% to 4.0%), pointing to a mediating effect of gallstones on the association between Mapuche ancestry and GBC. In contrast, the proportion of Mapuche ancestry showed a negative effect on BMI (IVW estimate -0.006 kg/m2, 95% CI -0.009 to -0.003). The results presented here may have significant implications for GBC prevention and are important for future admixture mapping studies. Given that the association between the individual proportion of Mapuche ancestry and GBC risk previously noted in observational studies appears to be free of confounding, primary and secondary prevention strategies that consider genetic ancestry could be particularly efficient.

Chromosomal instability in mouse embryonic fibroblasts null for the transcriptional co-repressor Ski.

Ski is a transcriptional regulator that has been considered an oncoprotein given its ability to induce oncogenic transformation in avian model systems. However, studies in mouse and in some human tumor cells have also indicated a tumor suppressor activity for this protein. We found that Ski-/- mouse embryo fibroblasts exhibit high levels of genome instability, namely aneuploidy, consistent with a tumor suppressor function for Ski. Time-lapse microscopy revealed lagging chromosomes and chromatin/chromosome bridges as the major cause of micronuclei (MN) formation and the subsequent aneuploidy. Although these cells arrested in mitosis after treatment with spindle disrupting drugs and exhibited a delayed metaphase/anaphase transition, spindle assembly checkpoint (SAC) was not sufficient to prevent chromosome missegregation, consistent with a weakened SAC. Our in vivo analysis also showed dynamic metaphase plate rearrangements with switches in polarity in cells arrested in metaphase. Importantly, after ectopic expression of Ski the cells that displayed this metaphase arrest died directly during metaphase or after aberrant cell division, relating SAC activation and mitotic cell death. This increased susceptibility to undergo mitosis-associated cell death reduced the number of MN-containing cells. The presented data support a new role for Ski in the mitotic process and in maintenance of genetic stability, providing insights into the mechanism of tumor suppression mediated by this protein.

Liquid biopsy to detect resistance mutations against anti-epidermal growth factor receptor therapy in metastatic colorectal cancer.

Colorectal cancer (CRC) is a major cause of mortality worldwide, associated with a steadily growing prevalence. Notably, the identification of KRAS, NRAS, and BRAF mutations has markedly improved targeted CRC therapy by affording treatments directed against the epidermal growth factor receptor (EGFR) and other anti-angiogenic therapies. However, the survival benefit conferred by these therapies remains variable and difficult to predict, owing to the high level of molecular heterogeneity among patients with CRC. Although classification into consensus molecular subtypes could optimize response prediction to targeted therapies, the acquisition of resistance mutations to targeted therapy is, in part, responsible for the lack of response in some patients. However, the acquisition of such mutations can induce challenges in clinical practice. The utility of liquid biopsy to detect resistance mutations against anti-EGFR therapy has recently been described. This approach may constitute a new standard in the decision algorithm for targeted CRC therapy.

Increase in ADAR1p110 activates the canonical Wnt signaling pathway associated with aggressive phenotype in triple negative breast cancer cells.

Triple-negative breast cancer (TNBC) represents a challenge in the search for new therapeutic targets. TNBCs are aggressive and generate resistance to chemotherapy. Tumors of TNBC patients with poor prognosis present a high level of adenosine deaminase acting on RNA1 (ADAR1). We explore the connection of ADAR1 with the canonical Wnt signaling pathway and the effect of modulation of its expression in TNBC. Expression data from cell line sequencing (DepMap) and TCGA samples were downloaded and analyzed. We lentivirally generated an MDA-MB-231 breast cancer cell line that overexpress (OE) ADAR1p110 or an ADAR knockdown. Abundance of different proteins related to Wnt/ß-catenin pathway and activity of nuclear ß-catenin were analyzed by Western blot and luciferase TOP/FOP reporter assay, respectively. Cell invasion was analyzed by matrigel assay. In mice, we study the behavior of tumors generated from ADAR1p110 (OE) cells and tumor vascularization immunostaining were analyzed. ADAR1 connects to the canonical Wnt pathway in TNBC. ADAR1p110 overexpression decreased GSK-3ß, while increasing active ß-catenin. It also increased the activity of nuclear ß-catenin and increased its target levels. ADAR1 knockdown has the opposite effect. MDA-MB-231 ADAR1 (OE) cells showed increased capacity of invasion. Subsequently, we observed that tumors derived from ADAR1p110 (OE) cells showed increased invasion towards the epithelium, and increased levels of Survivin and CD-31 expressed in vascular endothelial cells. These results indicate that ADAR1 overexpression alters the expression of some key components of the canonical Wnt pathway, favoring invasion and neovascularization, possibly through activation of the ß-catenin, which suggests an unknown role of ADAR1p110 in aggressiveness of TNBC tumors.

Identification of Circulating lncRNAs Associated with Gallbladder Cancer Risk by Tissue-Based Preselection, Cis-eQTL Validation, and Analysis of Association with Genotype-Based Expression.

Long noncoding RNAs (lncRNAs) play key roles in cell processes and are good candidates for cancer risk prediction. Few studies have investigated the association between individual genotypes and lncRNA expression. Here we integrate three separate datasets with information on lncRNA expression only, both lncRNA expression and genotype, and genotype information only to identify circulating lncRNAs associated with the risk of gallbladder cancer (GBC) using robust linear and logistic regression techniques. In the first dataset, we preselect lncRNAs based on expression changes along the sequence "gallstones → dysplasia → GBC". In the second dataset, we validate associations between genetic variants and serum expression levels of the preselected lncRNAs (cis-lncRNA-eQTLs) and build lncRNA expression prediction models. In the third dataset, we predict serum lncRNA expression based on individual genotypes and assess the association between genotype-based expression and GBC risk. AC084082.3 and LINC00662 showed increasing expression levels (p-value = 0.009), while C22orf34 expression decreased in the sequence from gallstones to GBC (p-value = 0.04). We identified and validated two cis-LINC00662-eQTLs (r2 = 0.26) and three cis-C22orf34-eQTLs (r2 = 0.24). Only LINC00662 showed a genotyped-based serum expression associated with GBC risk (OR = 1.25 per log2 expression unit, 95% CI 1.04-1.52, p-value = 0.02). Our results suggest that preselection of lncRNAs based on tissue samples and exploitation of cis-lncRNA-eQTLs may facilitate the identification of circulating noncoding RNAs linked to cancer risk.

TRPM4 enhances cell proliferation through up-regulation of the ß-catenin signaling pathway.

Altered expression of some members of the TRP ion channel superfamily has been associated with the development of pathologies like cancer. In particular, TRPM4 levels are reportedly elevated in diffuse large B-cell non-Hodgkin lymphoma, prostate, and cervical cancer. However, whether such changes in TRPM4 expression may be relevant to genesis or progression of cancer remains unknown. Here we show that reducing TRPM4 expression decreases proliferation of HeLa cells, a cervical cancer-derived cell line. In this cell line, constitutive TRPM4 silencing promoted GSK-3ß-dependent degradation of ß-catenin and reduced ß-catenin/Tcf/Lef-dependent transcription. Conversely, overexpression of TRPM4 in T-REx 293 cells (a HEK293-derived cell line) increased cell proliferation and ß-catenin levels. Our results identify TRPM4 as an important, unanticipated regulator of the ß-catenin pathway, where aberrant signaling is frequently associated with cancer.