Browsing by Subject "drug discovery"

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  • GEMSTONE Working Grp 3 COST Action; Formosa, Melissa M.; Bergen, Dylan J. M.; Gregson, Celia L.; Mäkitie, Outi (2021)
    Genetic disorders of the skeleton encompass a diverse group of bone diseases differing in clinical characteristics, severity, incidence and molecular etiology. Of particular interest are the monogenic rare bone mass disorders, with the underlying genetic defect contributing to either low or high bone mass phenotype. Extensive, deep phenotyping coupled with high-throughput, cost-effective genotyping is crucial in the characterization and diagnosis of affected individuals. Massive parallel sequencing efforts have been instrumental in the discovery of novel causal genes that merit functional validation using in vitro and ex vivo cell-based techniques, and in vivo models, mainly mice and zebrafish. These translational models also serve as an excellent platform for therapeutic discovery, bridging the gap between basic science research and the clinic. Altogether, genetic studies of monogenic rare bone mass disorders have broadened our knowledge on molecular signaling pathways coordinating bone development and metabolism, disease inheritance patterns, development of new and improved bone biomarkers, and identification of novel drug targets. In this comprehensive review we describe approaches to further enhance the innovative processes taking discoveries from clinic to bench, and then back to clinic in rare bone mass disorders. We highlight the importance of cross laboratory collaboration to perform functional validation in multiple model systems after identification of a novel disease gene. We describe the monogenic forms of rare low and high rare bone mass disorders known to date, provide a roadmap to unravel the genetic determinants of monogenic rare bone mass disorders using proper phenotyping and genotyping methods, and describe different genetic validation approaches paving the way for future treatments.
  • Lohtaja, Milka (Helsingfors universitet, 2016)
    Chlamydia pneumoniae is an intracellular bacterium that causes a variety of respiratory infections to humans such as pneumonia and bronchitis. In addition C. pneumoniae -infection has been associated with multiple chronic diseases of which the most important are atherosclerosis and vascular diseases, asthma, chronic obstructive pulmonary disease and different kinds of neurological disorders. C. pneumoniae is a very common pathogen that has the ability to hide in the system in a persistent chronic form out of reach of the immune defences. C. pneumoniae has been shown to infect many other cell types besides bronchial epithelial cells. These cells include monocytes, macrophages and vascular endothelial cells. C. pneumoniae induces the secretion of different kinds of cytokines and cell signalling molecules and the expression of adhesion molecules in all of these cell types. Too strong cytokine and immune response is detrimental to cells and to whole system. Currently available antibiotics aren't effective enough against C. pneumoniae -infection, especially against its chronic form. Furthermore, the lack of effective anti-chlamydial drugs impairs the research of the association between C. pneumoniae and chronic diseases. The aim of this study was to investigate the effect of anti-chlamydial compounds on the release of cytokines and cell signaling molecule, nitric oxide, induced by C. pneumoniae -infection in different cell types. These anti-chlamydial compounds are currently under the investigation in the faculty of pharmacy. In addition the anti-inflammatory properties of the compounds were further investigated with the help of lipopolysaccharide of another gram-negative bacterium E. coli. The groups of compounds investigated in this study were β2,2-amino acid derivatives, Schisandra chinensis -lignans, TE-compounds synthesized in Vienna and benzimidazole compounds synthesized in the faculty of pharmacy. There were four cell types used in this study, HL- and BEAS-2B-epithelial cells, THP-1-monocytes/macrophages and RAW264.7-macrophages. The study focused on the determination of vascular endothelial growth factor VEGF and interleukins IL-8, IL-10 and IL-12. The concentrations of cytokines in the cell medium were measured after infection using ELISA-method. Nitric oxide measurements were also determined from the medium using Griess' reagent. Immunofluorescence labeling was used to confirm the infection and the infection was verified by fluorescence microscope. In addition some of the compounds were tested for the cell viability using resazurin assay. All the groups of compounds showed desired effects on the release of cytokines and nitric oxide. Especially β2,2-amino acid derivatives reduced clearly the release of both cytokines and nitric oxide. β2,2-amino acid derivatives could thus be potential drug candidates for the development of anti-chlamydial and anti-inflammatory drugs. Schisandra chinensis -lignans inhibited especially the release of nitric oxide in both C. pneumoniae -infected and LPS-stimulated cells which may tell about their broad anti-inflammatory properties. There were also found desired results with TE-compounds and benzimidazole compounds. Interleukins were not secreted by any of the studied cells so that part needs more research and further investigation. Based on the results found in this study it can be concluded that the studied compounds could be potential lead compounds in the discovery of anti-chlamydial drugs and drugs that specifically inhibit C. pneumoniae -infection. Further research is needed concerning the effects of these compounds on cytokines and especially on chronic infection.
  • Garcia-Romero, Noemi; Gonzalez-Tejedo, Carmen; Carrion-Navarro, Josefa; Esteban-Rubio, Susana; Rackov, Gorjana; Rodriguez-Fanjul, Vanessa; Oliver-De La Cruz, Jorge; Prat-Acin, Ricardo; Peris-Celda, Maria; Blesa, David; Ramirez-Jimenez, Laura; Sanchez-Gomez, Pilar; Perona, Rosario; Escobedo-Lucea, Carmen; Belda-Iniesta, Cristobal; Ayuso-Sacido, Angel (2016)
    Human gliomas harbour cancer stem cells (CSCs) that evolve along the course of the disease, forming highly heterogeneous subpopulations within the tumour mass. These cells possess self-renewal properties and appear to contribute to tumour initiation, metastasis and resistance to therapy. CSC cultures isolated from surgical samples are considered the best preclinical in vitro model for primary human gliomas. However, it is not yet well characterized to which extent their biological and functional properties change during in vitro passaging in the serum-free culture conditions. Here, we demonstrate that our CSC-enriched cultures harboured from one to several CSC clones from the human glioma sample. When xenotransplanted into mouse brain, these cells generated tumours that reproduced at least three different dissemination patterns found in original tumours. Along the passages in culture, CSCs displayed increased expression of stem cell markers, different ratios of chromosomal instability events, and a varied response to drug treatment. Our findings highlight the need for better characterization of CSC-enriched cultures in the context of their evolution in vitro, in order to uncover their full potential as preclinical models in the studies aimed at identifying molecular biomarkers and developing new therapeutic approaches of human gliomas.
  • Boije af Gennäs, Gustav; Talman, Virpi; Yli-Kauhaluoma, Jari; Tuominen, Raimo K.; Ekokoski, Elina (2011)
    The second messenger diacylglycerol (DAG) plays a central role in the signal transduction of G-protein coupled receptors and receptor tyrosine kinases by binding to C1 domain of effector proteins. C1 domain was first identified in protein kinase C (PKC) which comprises a family of ten isoforms that play roles in diverse cellular processes such as proliferation, apoptosis and differentiation. Aberrant signaling through PKC isoforms and other C1 domain-containing proteins has been implicated in several pathological disorders. Drug discovery concerning C1 domains has exploited both natural products and rationally designed compounds. Currently, molecules from several classes of C1 domain-binding compounds are in clinical trials; however, still more have the potential to enter the drug development pipeline. This review gives a summary of the recent developments in C1 domain-binding compounds.
  • Pohjavaara, Saana (Helsingin yliopisto, 2021)
    Dilated cardiomyopathy is a non-ischemic cardiac disorder predisposing to heart failure, and the characteristics of dilated cardiomyopathy emerge under normal loading conditions. Dilated cardiomyopathy can be consequence of various conditions e.g. genetic mutations, virus infection or toxin exposures. One of the significant causes of familial dilated cardiomyopathy in Finland is mutation S143P in LMNA-gene, coding for A type lamins. Current drug therapy for dilated cardiomyopathy aims to alleviation of symptoms, prevention of complications and progression of the disease, however, efficacy of current therapy is insufficient, and novel therapy strategies are urgently required. Transcription factors are fundamental regulators of gene expression, and GATA4 is a crucial transcription factor both in embryonic and in adult heart and thus an intriguing target for therapeutic manipulation. Compounds targeting GATA4 have shown anti-hypertrophic and cardioprotective effects. Here, effects of two different hypertrophic stimuli, endothelin-1 and mechanical stretch, on human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) were examined with high-content analysis and quantitative reverse transcription PCR (qRT-PCR), respectively. One hiPSC-CM line was used as a healthy control, whereas the other carried the S143P mutation in LMNA-gene (DCM-CMs). Additionally, effects of GATA4-targeting compound C-2021 on cardiomyocytes were investigated. In summary, according to proBNP staining, DCM-CMs are more hypertrophied at baseline. DCM-CMs seemed to be less susceptible to mechanical stretch-induced enhancement in BNP gene expression. In addition, compound C 2021 may have anti-hypertrophic properties suggesting it to be a potential drug candidate in cardiac diseases. Finally, lamin A seemed to mislocalize to nucleoplasm instead of nuclear lamina in DCM-CMs.
  • Johansson, Niklas G; Dreano, Loic; Vidilaseris, Keni; Khattab, Ayman; Liu, Jianing; Lasbleiz, Arthur; de Castro Ribeiro, Orquidea Marilia; Kiriazis, Alexandros; Boije af Gennäs, Gustav; Meri, Seppo; Goldman, Adrian; Yli-Kauhaluoma, Jari; Xhaard, Henri (2021)
    Inhibition of membrane-bound pyrophosphatase (mPPase) with small molecules offer a new approach in the fight against pathogenic protozoan parasites. mPPases are absent in humans, but essential for many protists as they couple pyrophosphate hydrolysis to the active transport of protons or sodium ions across acidocalcisomal membranes. So far, only few nonphosphorus inhibitors have been reported. Here, we explore the chemical space around previous hits using a combination of screening and synthetic medicinal chemistry, identifying compounds with low micromolar inhibitory activities in the Thermotoga maritima mPPase test system. We furthermore provide early structure-activity relationships around a new scaffold having a pyrazolo[1,5-a]pyrimidine core. The most promising pyrazolo[1,5-a]pyrimidine congener was further investigated and found to inhibit Plasmodium falciparum mPPase in membranes as well as the growth of P. falciparum in an ex vivo survival assay.
  • Bakula, Daniela; Ablasser, Andrea; Aguzzi, Adriano; Antebi, Adam; Barzilai, Nir; Bittner, Martin-Immanuel; Jensen, Martin Borch; Calkhoven, Cornelis F.; Chen, Danica; de Grey, Aubrey D. N. J.; Feige, Jerome N.; Georgievskaya, Anastasia; Gladyshev, Vadim N.; Golato, Tyler; Gudkov, Andrei V.; Hoppe, Thorsten; Kaeberlein, Matt; Katajisto, Pekka; Kennedy, Brian K.; Lal, Unmesh; Martin-Villalba, Ana; Moskalev, Alexey A.; Ozerov, Ivan; Petr, Michael A.; Reason, Matthew; Rubinsztein, David C.; Tyshkovskiy, Alexander; Vanhaelen, Quentin; Zhavoronkov, Alex; Scheibye-Knudsen, Morten (2019)
    An increasing aging population poses a significant challenge to societies worldwide. A better understanding of the molecular, cellular, organ, tissue, physiological, psychological, and even sociological changes that occur with aging is needed in order to treat age-associated diseases. The field of aging research is rapidly expanding with multiple advances transpiring in many previously disconnected areas. Several major pharmaceutical, biotechnology, and consumer companies made aging research a priority and are building internal expertise, integrating aging research into traditional business models and exploring new go-to-market strategies. Many of these efforts are spearheaded by the latest advances in artificial intelligence, namely deep learning, including generative and reinforcement learning. To facilitate these trends, the Center for Healthy Aging at the University of Copenhagen and Insilico Medicine are building a community of Key Opinion Leaders (KOLs) in these areas and launched the annual conference series titled "Aging Research and Drug Discovery (ARDD)" held in the capital of the pharmaceutical industry, Basel, Switzerland ( This ARDD collection contains summaries from the 6th annual meeting that explored aging mechanisms and new interventions in age-associated diseases. The 7th annual ARDD exhibition will transpire 2nd-4th of September, 2020, in Basel.
  • Cazaly, Emma; Saad, Joseph; Wang, Wenyu; Heckman, Caroline; Ollikainen, Miina; Tang, Jing (2019)
    Epigenetic research involves examining the mitotically heritable processes that regulate gene expression, independent of changes in the DNA sequence. Recent technical advances such as whole-genome bisulfite sequencing and affordable epigenomic array-based technologies, allow researchers to measure epigenetic profiles of large cohorts at a genome-wide level, generating comprehensive high-dimensional datasets that may contain important information for disease development and treatment opportunities. The epigenomic profile for a certain disease is often a result of the complex interplay between multiple genetic and environmental factors, which poses an enormous challenge to visualize and interpret these data. Furthermore, due to the dynamic nature of the epigenome, it is critical to determine causal relationships from the many correlated associations. In this review we provide an overview of recent data analysis approaches to integrate various omics layers to understand epigenetic mechanisms of complex diseases, such as obesity and cancer. We discuss the following topics: (i) advantages and limitations of major epigenetic profiling techniques, (ii) resources for standardization, annotation and harmonization of epigenetic data, and (iii) statistical methods and machine learning methods for establishing data-driven hypotheses of key regulatory mechanisms. Finally, we discuss the future directions for data integration that shall facilitate the discovery of epigenetic-based biomarkers and therapies.
  • de Vera, Caterina R.; Díaz Crespín, Guillermo; Hernández Daranas, Antonio; Montalvão Looga, Sofia; Lillsunde, Katja-Emilia; Tammela, Päivi; Perälä, Merja; Hongisto, Vesa; Virtanen, Johannes; Rischer, Heiko; Muller, Christian D.; Norte, Manuel; Fernández, José J.; Souto, María L. (2018)
    The study of marine natural products for their bioactive potential has gained strength in recent years. Oceans harbor a vast variety of organisms that offer a biological and chemical diversity with metabolic abilities unrivalled in terrestrial systems, which makes them an attractive target for bioprospecting as an almost untapped resource of biotechnological applications. Among them, there is no doubt that microalgae could become genuine cell factories for the biological synthesis of bioactive substances. Thus, in the course of inter-laboratory collaboration sponsored by the European Union (7th FP) into the MAREX Project focused on the discovery of novel bioactive compounds of marine origin for the European industry, a bioprospecting study on 33 microalgae strains was carried out. The strains were cultured at laboratory scale. Two extracts were prepared for each one (biomass and cell free culture medium) and, thus, screened to provide information on the antimicrobial, the anti-proliferative, and the apoptotic potential of the studied extracts. The outcome of this study provides additional scientific data for the selection of Alexandrium tamarensis WE, Gambierdiscus australes, Prorocentrum arenarium, Prorocentrum hoffmannianum, and Prorocentrum reticulatum (Pr-3) for further investigation and offers support for the continued research of new potential drugs for human therapeutics from cultured microalgae.
  • Liu, Xiaonan; Huuskonen, Sini; Laitinen, Tuomo; Redchuk, Taras; Bogacheva, Mariia; Salokas, Kari; Pohner, Ina; Öhman, Tiina; Tonduru, Arun Kumar; Hassinen, Antti; Gawriyski, Lisa; Keskitalo, Salla; Vartiainen, Maria K.; Pietiainen, Vilja; Poso, Antti; Varjosalo, Markku (2021)
    Treatment options for COVID-19, caused by SARS-CoV-2, remain limited. Understanding viral pathogenesis at the molecular level is critical to develop effective therapy. Some recent studies have explored SARS-CoV-2-host interactomes and provided great resources for understanding viral replication. However, host proteins that functionally associate with SARS-CoV-2 are localized in the corresponding subnetwork within the comprehensive human interactome. Therefore, constructing a downstream network including all potential viral receptors, host cell proteases, and cofactors is necessary and should be used as an additional criterion for the validation of critical host machineries used for viral processing. This study applied both affinity purification mass spectrometry (AP-MS) and the complementary proximity-based labeling MS method (BioID-MS) on 29 viral ORFs and 18 host proteins with potential roles in viral replication to map the interactions relevant to viral processing. The analysis yields a list of 693 hub proteins sharing interactions with both viral baits and host baits and revealed their biological significance for SARS-CoV-2. Those hub proteins then served as a rational resource for drug repurposing via a virtual screening approach. The overall process resulted in the suggested repurposing of 59 compounds for 15 protein targets. Furthermore, antiviral effects of some candidate drugs were observed in vitro validation using image-based drug screen with infectious SARS-CoV-2. In addition, our results suggest that the antiviral activity of methotrexate could be associated with its inhibitory effect on specific protein-protein interactions.
  • Tepsell, Juhani (Helsingfors universitet, 2018)
    During and after myocardial infarction, millions to a billion cells die off. Scar tissue formed by fibroblasts replaces the injured myocardium during recovery. While the newly formed tissue is durable and prevents rupture of the heart, it doesn´t contribute to pump function. Depending on the extent of cardiomyocyte loss, the remaining functional myocardium get strained. Adult mammalian heart has inadequate capacity to regenerate after such injury. In case of sustained substantial increase in workload, the compensatory mechanisms turn into pathological processes including excessive fibrosis and myocyte apoptosis. The progressive decline of hearts contractile function results in heart failure (HF). Current drug treatments for managing HF aim to prevent progression of the disease and relieve symptoms. ACE inhibitors, beta blockers and diuretics are effective along with healthy lifestyle. No practical treatments are available to restore cardiac function yet. Human myocardium normally regenerates, but only 1% or less of myocytes get replaced yearly. Heart’s resident stem/progenitor cells (CPCs) likely play a role in the turnover. The aim of this study was to develop a screening method to identify small molecules that possibly promote differentiation of cardiac progenitor cells to cardiomyocytes. Cell population differentiated from mouse embryonic stem cells (mESCs) was used as a model for CPCs. Directed differentiation protocol of mESCs used here promotes commitment to cells of cardiac mesoderm, part of which will further differentiate to cardiac progenitors. The resulting population at day 6 is heterogenous but many of these cells are progenitors that turn into cardiomyocytes (CMs) by day 8. 10 000 cells per well are plated on 384 well plates at day 5. Test compounds are added at day 6 and removed day 8 for effect in progenitors and day 7-9 for effect in early cardiomyocytes. 0,1% DMSO is used as vehicle and Wnt pathway inhibitor XAV939 as positive control. The effects are quantified with plate reader on day 9. E14 derived mESC reporter line was used. Myl2v-eGFP + SMyHC3-RFP double reporter line allows the specific identification of ventricular CMs with green fluorescence and atrial CMs with red fluorescence. Plate reader measures the total fluorescence of the wells at 485/520nm on day 9, which is used as a readout for ventricular CMs. The fluorescence intensity depends on the amount of GFP+ cells but also on the level of Myl2v expression. Atrial CMs could be quantified similarly but the population doesn´t contain enough RFP+ cells. The assay was shown to reliably point out ‘hits’ that have a strong effect. Any compounds that only produce a moderate effect could be a false negative, however. The effect on cardiac progenitors could likely be increased by simply adding the compounds earlier on day 5. Variability of key reagents causes the main technical troubles through unpredictably affecting cytokine concentrations which decreases the amount of cardiac progenitors. Partially similar screening assays are being used by the big pharma where they cryopreserve progenitors in bulk for later use, thus simplifying and speeding up their method. Same approach could be adopted.
  • Silfvast, Saga (Helsingfors universitet, 2016)
    Heart failure is a major public health problem and a leading cause of mortality worldwide. The most common cause of heart failure is myocardial infarction. Following a myocardial infarction, a large number of cardiomyocytes die and cardiac muscle is replaced by fibrotic scar tissue. Since the adult heart has inadequate endogenous regenerative capacity, loss of muscle tissue often causes a progressive decrease in cardiac function eventually leading to heart failure. At the moment heart transplantation is the only curative treatment for heart failure, but the low number of donor hearts is limiting the use of this treatment option. As current drugs only slow down the progression of the disease, there is a great need for new regenerative treatments. Direct cardiac reprogramming is a new approach for generating cardiomyocytes for cardiac regeneration. Unlike pluripotent stem cell-based strategies, direct reprogramming enables conversion of a terminally differentiated cell type directly into another cell type without first producing a pluripotent intermediate. Due to their abundancy and role in the repair of myocardial injury, fibroblasts represent an attractive starting cell type for direct cardiac reprogramming. Fibroblasts have been directly reprogrammed to induced cardiomyocytes (iCMs) by overexpression of key cardiac transcription factors, microRNAs (miRNA) or by modulating specific signal transduction pathways with small-molecule compounds. Despite successful reports of direct reprogramming both in vitro and in vivo, the efficiency of direct reprogramming remains, however, too low for potential clinical applications. The aim of this M.Sc. thesis work was to establish direct reprogramming of mouse embryonic fibroblasts (MEFs) to iCMs by viral overexpression of cardiac transcription factors Hand2 (H), Nkx2.5 (N) Gata4 (G), Mef2c (M) and Tbx5 (T) and a small-molecule compound screening platform for identifying small-molecule compounds that could enhance the reprogramming efficiency and potentially replace cardiac transcription factors in direct cardiac reprogramming. In accordance with previous publications MEFs were successfully directly reprogrammed to iCMs using both HGMT and HNGMT cardiac transcription factor combinations. The screening platform was tested using the TGF-β inhibitor SB431542, which has recently been reported to increase the cardiac reprogramming efficiency. In line with previous publications, the reprogramming efficiency was significantly increased by treatment with SB431542. Initial tests with other small-molecule compounds did not have a positive effect on the reprogramming efficiency. The results of this M.Sc. thesis work verify previous publications and demonstrate a method for in vitro small-molecule compound screening, which can be used to identify compounds that increase the reprogramming efficiency in direct cardiac reprogramming. However, the results shown here are only preliminary and more replicates are needed in order to confirm the current results. Nonetheless, the results of this thesis work set a foundation for finding small-molecule compounds that in the future might be used to target direct cardiac reprogramming as a regenerative therapy for myocardial infarction and heart failure.