Informe Anual 2017-2018

Background. The Chilean population has experienced in a very short period large changes in the socio-environmental setting, which permitted the effective control of infectious diseases and undernourishment, thereby dramatically enhancing life expectancy. However, the population quality of life is currently threatened by an “epidemic” of risk factors related to chronic diseases (CDs). CDs now account for 84% of all deaths in Chile, the most prominent being cardiovascular diseases (CVD, 27%), followed by cancer (26%) (WHO – Non-communicable Diseases Country Profiles, 2014). These diseases affect mostly people over 45 years of age, even though the pathological process initiates earlier in life. CDs are the result of changes in socio-environmental factors which determined changes in lifestyle at the population level, in particular a significant decrease in physical activity and modification of diet towards one rich in calories, refined carbohydrates and saturated fats. The resulting metabolic alterations and ensuing chronic inflammation represent a common mechanism implicated in the genesis of CVD, diabetes, obesity, and cancer, among others. The measures taken to change people’s behavior, have been largely unsuccessful in Chile. Although CDs share common risk factors worldwide, the interventions should consider particulars of every disease, and the local ethnic, cultural and socio-economic environment. Thus, successful interventions should consider the high risk approach, that is to identify and protect those at highest risk, but long-term solutions will only emerge by identifying and reducing risk factor levels. Our Center is collaborating to generate data at both strategic levels: identifying individual at highest risk and the structural conditions that facilitate individual’s exposure.

Advanced Center for Chronic Diseases (ACCDiS). This is the first Chilean Center of Excellence for the study of CVD and cancer. ACCDiS is a bi-institutional center involving the Universidad de Chile (UCH) and Pontificia Universidad Católica de Chile (PUC), supported by the Funds for Research Centers in Priority Areas (FONDAP) program of CONICYT. ACCDiS goals are to observe and analyze the natural history of CVD and cancer in Chile. The Center includes six research lines with laboratories located on the UCH North Campus (Faculties Medicine/Chemical and Pharmaceutical Sciences/Dentistry) and in the PUC (Faculties Medicine and Biological Sciences). The Center has also an Administrative Office, located on the North Campus (UCH), two Core Facilities (Bioinformatics and Functional and Experimental Animal and Biodata), and a Core Project (MAUCO cohort). The main entities involved in directing and evaluating the center are the Academic Council, composed by the six Principal Investigators (PIs), the International Scientific Committee and the National Advisory Committee. The six Research Lines (RLs) and PIs are: a) Metabolism and cardiovascular signaling (S Lavandero, UCH), b) Biomarkers in heart failure and remodeling (P Castro, PUC), c) Mechanisms of tumor cell migration and metastasis (A Quest, UCH), d) Development of biomarkers for early detection of tumors (A Corvalán, PUC), e) Natural history of gallbladder cancer (C Ferreccio, PUC), f) Nanomedicine and nanotheranostics (M Kogan, UCH).

Integration strategyTo promote the integration of research efforts in cancer and cardiovascular  diseases, we adopted the following strategies: a) Identify research areas of common interest: exosomes, angiotensin-(1-9), nanotechnology, epidemiology of CDs and cancer,  b) secure that PhD, MSc students, and post-doctoral fellows recruited in the Center work and are tutored in at least two research lines, c) Implement a large population based Cohort study (MAUCO) and biobank in which all lines can learn about the natural history of the CDs and their risk factors, d) Core facilities: Bioinformatics and microRNA analysis and Functional and Experimental Animal Core Facility.

Results. After 4.4 years in operation (December 2013 to May 2018), ACCDiS has consolidated itself in the Chilean community as an important center for the study of cancer & CVD. ACCDiS personnel (247 persons) work in the 6 RLs, MAUCO and/or the two core facilities and includes 6 PIs, as well as 24 Associate Investigators (AIs). ACCDiS organization includes an Administrative Office, an Academic Council, an International Scientific Committee and a National Advisory Committee. Both Committees visited the Center annually and made recommendations to improve ACCDiS functioning.

1) In Advanced Training of Human Resources. Currently, after 4.4 years, 42 postdoctoral fellows (20 finished), 114 PhD (50 finished) and 60 MSc (39 finished) students, together with 56 undergraduate (39 finished) students are working on their projects in the center. From this number, 40% of the postdoctoral fellows, 11% of the PhD and 7% of the MSc. students are co-mentored between ACCDiS members from different RLs. Nearly 10% of all students in training and 13% of postdocs at the Center are foreigners. ACCDiS PIs and AIs coordinated and taught in various PhD, MSc and undergraduate courses of both universities. MAUCO incorporated academics and undergraduate students from regional universities in Maule in the field activities.

2) In Research. a) The MAUCO project. By May 2018, we have enrolled 7,500 participants. Baseline results were presented at scientific meetings. We are preparing manuscripts describing the baseline analysis of the health status in the areas cardiovascular, digestive, neurocognitive, respiratory, psychosocial, occupational and environmental exposures. Various MAUCO-based MSc thesis projects have been completed or are underway. We completed the design of the details of the 2-year follow-up of the cohort and started on January 2017. b) Publications & Patents: The ACCDiS center has published from 2014 to May 2018 a total of 262 ISI papers, exceeding the expected 220 papers after 4.4 years; besides 49 (18.7%) were published in Top Ten journals in their area and 158 (60.3%) in Q1 journals. The average ISI impact factor over this period has been 4.65. The Center also produced 28 non-ISI papers. All progress evaluation indicators exceed the expected values. c) Patents. In this period Center applied for nine patents (six in collaboration between RLs) and four have been granted in Chile and in the USA. The AQ/MK groups applied for a patent for a nano-emulsion to improve the antitumor effect of curcumin. The AC/MK groups applied for a patent related for nanoparticles to detect the methylated Reprimo cell-free DNA. The MK group applied for a patent for the controlled oral administration of bio-macromolecules using a polymeric matrix.

3) In Outreach to Society. To facilitate dissemination of results and improve visibility, the ACCDiS Communication and Public Relations Manager worked together with a professional agency (Ines Llambias Communications). Until May 2018, PIs, as well as AIs and collaborators, have participated in almost 374 media events: 202 website news articles, 113 newspapers articles, 21 articles in magazines, 21 television interviews, 14 radio interviews and 3 social network. Concerning organization of or participation in events: ACCDIS has participated (up to May 2018) in 239 events, involving a total of 34,352 participants, including an outreach program for primary and secondary public school teachers and students, and meetings with businessmen/women and politicians.  Regarding the community, the MAUCO Project developed a high level of communication and outreach to scientists, as well as to the Molina community and to the Ministry of Health.

4) In International Networking. We have active collaborations with outstanding international research centers in CVD, cancer, epidemiology and nanomedicine. These were strengthened by 154 visits of Chilean and foreign investigators (73 visits of ACCDiS members to foreign institutions and 81 international researchers visited ACCDiS in Santiago). Moreover, 59% of our ISI papers were published in collaboration with our international collaborators. Also, we organized the first ACCDiS Cancer Spring Symposium (ACSS) in 2016, which included the participation of 11 international and 6 national invited speakers, the IV national meeting of nanotechnology in 2016, and the “Meeting of Population Cohorts for the study of Chronic Diseases – COPLAS”, held on April, 4th-6th, 2018.

5) In Contribution to Public Policies. ACCDiS/MAUCO has been supported by the Ministry of Health (MH), the Maule Health Service (MHS) and the Molina municipality, which recognize the potential of the cohort to guide in developing health prevention and control strategies for CDs, as well as defining that MAUCO is strategic of the MHS because we will provide key data for public health surveillance in the region.  We participated in a “Cancer Law” seminar with the Senate Health Committee to establish efficient public policies to combat cancer and in the development of the Chilean guidelines for the management of heart failure for MH. A seminar for MH was performed on the policy challenges derived from our studies in cancer. The results of this Seminar were included in the MH “Cancer Strategic Planning 2016”. AC participated in the Seminar Cancer: Task of the State at Senate House. CF is part of the advisory board of cervical cancer prevention guidelines at the MoH and of the PAHO-IARC advisory group for adapting the European Code of Cancer.

Results of Research Lines. See Table Indicators- Indicators for Research of Excellence

In the RL1 (Lavandero [SL] group): Our work was focused on studying whether the cardiovascular metabolism is involved in the genesis and progression of CVD. Active research collaborations are underway with all ACCDiS groups in the four strategic areas: angiotensin-(1-9); exosomes; MAUCO; onco-cardiology, evidenced by joint PhD students, joint postdocs, joint papers and projects. Our main findings showed that insulin and GLP-1 regulate mitochondrial morphology and metabolism in cardiomyocytes and vascular smooth muscle cells. Alteration in cardiomyocyte mitochondrial dynamics was associated with cell death during ischemia/reperfusion, hypertrophy, and insulin resistance. With P Castro’s group, we described the role of mitochondrial dynamics in the control of pulmonary arterial smooth muscle cell metabolism and proliferation associated with pulmonary hypertension. We also showed that Rcan1 induced mitochondrial fusion in cardiomyocytes. In collaboration with JA Hill (Dallas), we showed that the hexosamine pathway acts downstream of the UPR to limit cardiac I/R injury; the anti-cancer histone deacetylase inhibitor SAHA reduced myocardial infarct size; inhibition of class I HDAC blunted cardiac hypertrophy; polycystin (PC)-1 is a mechanosensor in cardiomyocytes; and cardiomyocyte autophagy is controlled by PC-2. With D Klionsky (U Michigan) we described the new guidelines for monitoring autophagy. In order to associate cancer and CVD, a new line of oncocardiology was developed by studying doxorubicin-induced cardiotoxicity. We (with PC’s group) organized the International Workshop “New frontiers in cardiovascular disease management” in the XXV InterAmerican Congress of Cardiology (2015, Santiago) involving collaborators from The Hatter Cardiovascular Institute (UCL). We also organized the ACCDiS-ICGEB International Course and Symposium “Metabolic Signaling and Chronic Diseases” (2015) and the ICGEB Workshop Autophagy (2016). During this period SL was appointed Associate Editor in Circulation and the new young Faculty V Parra joined the group.

In the RL2 (Castro [PC] group): We focused on evaluating the effect of trimetazidine (TMZ) in patients with stable non-ischemic heart failure on cardiac mitochondrial morphology and metabolism. We showed that TMZ protects cardiomyocytes by modulation of the mitochondrial morphology and function. These observations contrast with previous results by other groups showing that the addition of TMZ to existing optimal medical treatments does not result in significant changes in myocardial function. We set up a Cardio-Oncology group, which includes multidisciplinary clinical care of at-risk oncology patients, advanced echocardiography with speckle tracking analyses and the creation of a prospective database and a bio-bank. This is the first of its class in Chile and will provide support for several new lines of research, such as testing for new biomarkers to predict drug-related cardiotoxicity, identification of genomic determinants of myocardial damage, and implementation of early interventions to minimize chemotherapy-related toxicity. This RL continued the electrocardiography (ECG) examinations in MAUCO and the echocardiography sub-study started in MAUCO subjects, publishing a review paper with CF´s group on heart failure in rural communities. With respect to intellectual property, we applied with SL’s group for 3 patents on angiotensin-(1‑9). We were awarded the grant entitled “Angiotensin-(1-9) a new antihypertensive and cardiovascular and renal protective drug: searching for a business partner”. In collaboration with SL group, we studied the effect of TMZ in the hypoxic response of pulmonary VSMC. In an on-site echocardiography sub-study of 100 subjects, we characterized the relationship between ideal CV health and early CV remodelling. Moreover, our group published a paper related to the gender influence on the adaptation of atrial performance to training and another about the role of a previously unreported miRNA, miR-15b, and the role of galectin-3 in heart failure.

In the RL3 (Quest [AQ] group): Specific objectives of the Quest lab were addressed by several PhD students who have graduated. We showed that survivin expression in cancer cells activates the PI3K/Akt pathway to promote β-catenin/Tcf-Lef-dependent transcription of VEGF and angiogenesis. We showed that PGE2 exposure dissociates CAV1/E-cad complexes to promote CAV1-dependent metastasis. Helicobacter pylori (Hp) infection is associated with decreased expression of SLC5A8 in young children. We showed that Hp infection induces HIF-1α expression via activation of a PI3K-dependent pathway and linked HIF-1α induction to cell cycle arrest. The ability of P gingivalis infection to inhibit apoptosis in gingival epithelial cells and cause chronic periodontitis (inflammation) in afflicted human subjects was linked to the O-antigen region of P. gingivalis LPS. We identified a novel CAV1-Rab5-Rac1 signaling axis and showed that CAV1 expression in melanoma, breast, and colon cancer cells promotes migration, invasion and metastasis by activating this pathway. We showed in cancer cells lacking E-cadherin that hypoxia modulates their migratory properties and identified Rab5 as a key mediator of hypoxia-induced migration, invasion, and metastasis. We showed that CAV1 phosphorylation on tyrosine-14 is required to promote migration invasion, transendothelial migration and metastasis in vivo. We also showed that phorbolester-induced CAV1 in endometrial cancer cells and anti-neoplastic drug-induced CAV1 expression in colon and breast cancer cells promote migration, invasion, and metastasis of those cancer cells. CAV1 expression also favors dendritic cell migration to draining lymph nodes. We identified PTPN14 as the phosphatase that specifically co-immunoprecipitates with and dephosphorylates CAV1 in the presence of E-Cad, preventing CAV1-enhanced migration/invasion and metastasis. We organized with AC the International ACCDiS Cancer Summer Symposium. Joint post-docs with SL (P Diaz, A Sandoval), AC (J Bravo), CF (L Orostica), MK (S Guerrero) and with AI L Leyton (J Diaz, R Perez) were awarded fellowships from CONICYT.

In the RL4 (Corvalán [AC] group): We propose that the emerging health burden of neoplastic diseases could be reduced by the development of biomarkers for the early detection. This focus on early detection is expected to reduce cancer deaths by more than 75% in the coming decades. This RL has focused on the development and the understanding of the biology behind biomarkers in the context of the multistep process carcinogenesis. In this scenario, and based on our discovery of a potential biomarker for noninvasive diagnosis of gastric cancer, methylated Reprimo (RPRM) cell‑free DNA, we have characterized RPRM at several levels, including evolution, development, and disease. Since methylated RPRM cell-free DNA can identify early-stages of gastric cancer, we proposed that vasculogenic mimicry (VM), the formation of microvascular channels de novo by tumor cells, might explain the presence of RPRM methylated in the blood. To characterize VM, we have used microarray to identify coding and non-coding genes (microRNAs and long noncoding RNAs) based on an experimental VM system developed in this RL. We characterized the biological role of miRNA-335-5p and proposed it use as a novel biomarker for noninvasive diagnosis of gastric cancer. To explore the dissemination of gastric cancer, extracellular vesicles have been studied by developing in vitro and in vivo mouse models using two gastric cancer cell lines. With a regional grant and the collaboration of Universidad de Talca, we have initiated the evaluation of the diagnostic value of methylated RPRM cell-free DNA and miRNA-335-5p in blood samples from MAUCO participants. To expand these validation activities, we established a collaboration with the Chilean Association of Endoscopy (ACHED) and the Metabolic Epidemiology Branch-DCEG, NCI, NIH to speed up the accrual of high-risk patients across the country.

In the RL5 (Ferreccio [CF] group): Chile had the highest gallbladder cancer (GBC) death rate worldwide and GBC represents the first cancer killer among Chilean women. GBC is among the least studied cancers in the world, and is becoming a national priority. The main causal factor of GBC in Chile are gallstones (GS); nevertheless, only 1-3% of GS carriers will develop cancer. The aims of this RL were to describe the occurrence and evolution of biliary diseases and to identify risk factors of GBC and pre-cancer among people with GS. In 2014, we reported that seven serum inflammatory markers correlate significantly with GBC, a higher prevalence of chronic Salmonella typhi carriers among GBC cases and, as the most novel aspect, a high prevalence of aflatoxin-DNA adducts in the blood of GBC cases; the latter was the first report on aflatoxin adducts in the Chilean population. In collaboration with MK’s group and the post-doctoral fellow C Foerster, we are searching for a more practical and economic biomarker of aflatoxin in urine. We are also currently conducting two large longitudinal studies of people with GS and biliary diseases, one cross-sectional study of people undergoing cholecystectomy, and one statistical study to model GBC trend and predict the effect of triage in the waiting list for cholecystectomy. The longitudinal studies are the biliary diseases cohort nested in MAUCO in the Maule region, and the Chile Biliary Longitudinal Study (Chile BiLS) in Temuco, Araucanía Region. In 2016, we implemented the Chile BiLS cohort using the same ultrasound protocol in a population-based cohort of women aged 50-74 years with GS. Chile BiLS is fully funded by NCI/NIH and GS participants are followed for 6 years. We began a risk stratification study based on our findings on inflammatory markers, among GS patients undergoing cholecystectomy. In 2017, we initiated a collaboration, funded by a Bill and Melinda Gates grant, with infectious disease specialists and microbiologists of UCH, the Chilean Institute of Public Health, and the University of Maryland, to expand the risk-stratification study of inflammatory markers with the additional objective to validate a previously reported association between Salmonella typhi and GBC. Nested in the MAUCO biliary disease sub-cohort, we initiated a study of the incidence and evolution of fatty liver and NAFLD among cholecystectomyzed participants. This new study was funded by FONDECYT/CONICYT with the aim to provide evidence about the causal directionality of the association between NAFLD and the extraction of the gallbladder. To test a complementary hypothesis, in 2017 we established a collaboration with NCI/NIH USA to study the microbiome changes associated with the extirpation of the GB among 1,000 participants. During years 2-4, we collaborated with a group from the Institute of Medical Biometry and Informatics, University of Heidelberg, Germany, to study ancestry markers in participants of our GBC case-control study, funded by the Federal Ministry of Education and Research (BMBF). In collaboration with the Chile Genómico group from UCH and funded by FONDEF, we initiated a genetic study; 4,000 subjects will be genotyped to obtain ancestry informative markers and explore the prevalence of various SNPs of digestive and cardiovascular diseases. In collaboration with University of Lund, we began the measurement of urinary metals and pesticides in environmental and human samples from our biliary disease sub-cohort, as well as from other participants in MAUCO. The first 4.4 years of this RL were dedicated to creating the research infrastructure to obtain definitive answers to the main questions regarding GBC.

In the RL6 (Kogan [MK] group): Collaboration between different research lines has been fruitful. We developed an ultrasensitive test to measure aflatoxins from urine samples in the MAUCO cohort in collaboration with CF group. We also developed new approaches to detect ultra-sensibly gastric cancer biomarkers as methylated RPRM using nanoparticles and plasmon-enhanced spectroscopy (in collaboration with AC group). In addition, we developed a methodology based on surface plasmon resonance for the detection of galectin-3 in biological samples as a biomarker of cardiac damage in collaboration with G Rivas (Universidad de Córdoba, Argentina) and PC group. In the field of drug delivery we developed a system for the controlled temporal and spatial drug release by using cyclodextrins, different linkers and gold nanoparticles, as well as by using calcium carbonate and polymeric nanoparticles. Also nanoemulsions that improve the activity of curcumin in vivo were developed and shown to prevent melanoma re-incidence and metastasis in a pre-clinical model, with a patent and a license are currently in progress (in collaboration with AQ). In addition we used lipidic nanoparticles for drug delivery of exosomes for targeting of gold nanoparticles to lung tumors (a co-supervised PhD is in progress AQ/MK). To improve the drug delivery of angiotensin-(1-9) for the treatment of cardiovascular disease we explored the use of a nanosystem based on gold and polymeric nanoparticles for the photothermal controlled release of angiotensin-(1-9) and the use of lipid nanoparticles in collaboration with PC and SL groups. In relation with cancer diagnostic we used gold nanoparticles and quantum dots to track tumor cells ex vivo. Three new pieces of equipment have been acquired within this period (SPR in 2015, Nanosight in 2016, EDS for electron microscopy in 2017) with other research grants. We organized the IV Chilean Meeting of Nanotechnology

The epidemiological Maule Cohort (MAUCO) project. The interaction among basic scientists, clinicians, and epidemiologists has improved with joint ongoing projects in CVD and cancer. We developed and initiated the various strategies and methods of this study, and negotiated various agreements with the relevant stake holders at the local, regional and national level. Due to the fact that the MAUCO project takes place in an agricultural area, we have implemented an environmental research line based on the evaluation of pesticide exposure. The MAUCO Project also required a high level of communication and outreach to scientists, as well as to the Molina community and to the Ministry of Health. We published a MAUCO study protocol paper and presented preliminary cross-sectional data in meetings in Chile and abroad. In gallbladder cancer and aflatoxin, we collaborated in the development of an aflatoxin biomarker using urine samples and were recently awarded FONDECYT postdoctoral fellowship for the aflatoxin project. We also collaborate in a clinical trial for the evaluation of the diagnostic value of methylated RPRM cell-free DNA and other biomarkers in MAUCO participants as part of a regional grant in collaboration with Universidad de Talca. To date 7,500 participants have been enrolled, besides the high cardiovascular risk factors described in our previous report, we found a high burden of cancer in this population. Cohort profile: From the cohort, the mean age of the population is 54 years, 51% completed primary school, the average body mass index is 30; diabetes and high blood pressure are present in 14.1%/38.9% of the women and 12.9%/47.6% of the men, respectively. No participant presented all the 7 characteristics of ideal cardiovascular health. During 2017 and 2018, the 2‑year follow-up study of 3,306 patients is being conducted.