Speakers

Title: “Characterizing the bioactive species: the relevance of physicochemical properties”

Abstract: A correct definition of the bioactive species is fundamental in the drug discover and design process. On one hand, because it defines a series of molecular determinants that define the ligand binding and recognition. On the other hand, because it establishes the molecular basis for the structure-activity relationships (SAR). However, its prediction is not trivial, since it depends on the interplay between different complicating factors. In the present conference, different projects will be shown that involve the consideration or estimation of this complicating factors, such as the correct definition of the conformational landscape, protonation state, lipophilicity and tautomerism.

Title: “Understanding the selectivity for β-isoforms of direct allosteric modulators of AMPK”

Abstract: AMPK is a key energy sensor regulating the cell metabolism in response to energy supply and demand. The evolutionary adaptation of AMPK to different tissues is accomplished through the expression of distinct isoforms that can form up to 12 complexes, which exhibit notable differences in the sensitivity to allosteric activators.[1] To shed light into the allosteric regulation of this energy sensor, we have assessed the structural and dynamical properties of β1- and β2- containing AMPK complexes formed with small molecules that could act as direct activators, in some cases activating specifically only one of the β-isoforms, while in other cases these ligands could act as a pan-activators.[2,3] So, which are the molecular factors that determine these differences?

We have run molecular simulations of several complexes formed by α2β1 and α2β2 AMPK subunits with A-769662, SC4, PF739 and MT47-100, dissecting the mechanical response leading to active-like enzyme conformations through the analysis of interaction networks between structural domains. The results reveal the mechanical sensitivity of the α2β1 complex, in contrast with a larger resilience of the α2β2 species, especially regarding modulation by A-769662. Furthermore, binding of activators to α2β1 consistently promotes the preorganization of the ATP-binding site, acting as a molecular glue between α- and β- subunits, thus favoring the adoption of activated states of the enzyme. These findings are discussed considering the changes in the residue content of β-subunit isoforms, particularly regarding the β1Asn111--> β2Asp111 substitution as a key factor in modulating the mechanical sensitivity of β1- and β2-containing AMPK complexes. [4,5,6] Our studies pave the way for the design of activators tailored for improving the therapeutic treatment of tissue-specific metabolic disorders.

Title: “Slope-Hunter: A robust Method for Collider Bias Correction in Conditional Genome-wide Association Studies”

Abstract: Studying genetic associations conditioned on a phenotype may be affected by collider bias. An example of this is the study of genetic associations with prognosis (e.g., survival, subsequent events). Selection on disease status can induce associations between causes of incidence with prognosis, potentially leading to collider bias. A current method for adjusting genetic associations for this bias assumes there is no genetic correlation between incidence and prognosis, which may not be a plausible assumption. We propose an alternative, the ‘Slope-Hunter’ approach, which is unbiased even when there is genetic correlation between incidence and prognosis. Our approach uses model-based clustering to identify and utilise the class of variants only affecting incidence to estimate the adjustment factor, assuming this class explains more variation in the incidence than any other variant classes. Simulation studies showed that our approach eliminates the bias and outperforms alternatives in the presence of genetic correlation and performs as well as alternatives under no genetic correlation when its assumptions are satisfied. In a study of fasting blood insulin levels (FI) conditional on body mass index, we eliminate paradoxical associations of the underweight loci: COBLLI; PPARG with increased FI and reveal an association for the locus rs1421085 (FTO). In an analysis of a case-only study for breast cancer mortality, a single region remains associated with more pronounced results.

Title: “Fish mycobacteriosis: diagnostics and prevention in the genomic era”

Abstract: Fish mycobacteriosis is a chronic progressive disease caused by ubiquitous acid-fast bacilli, identified as nontuberculous mycobacteria (NTM). NTM could affect cultured and wild fish worldwide and are associated with high levels of mortality, ranging from 10% to 100% of the infected fish. Rapid and accurate identification is critical for proper diagnosis and management of mycobacteria infections and for outbreak investigation, particularly considering the extraordinary number of species in the genus. While the immune responses against aquatic Mycobacterium sp. have been partly characterized, an effective vaccine against mycobacteriosis has not been developed. In this study, we report the isolation and characterization of two NTM namely M. pseudoshottsii and M. hippocampi, from meagre and European sea bass respectively, originating from aquaculture farms in Western Greece. The genome sequencing of the isolated NTM allowed the design of a PCR-based discriminative assay in one step. The assembled draft genome of M. pseudoshottsii had a length of 5,934,315 bp, with a GC content of 65.6% and for M. hippocampi the length was 6,251,150 bp, with a GC content of 66.7%. Moreover, we applied the integrative in silico approaches of reverse vaccinology on the isolated NTM to identify putative vaccine candidates for further experimentation. Two web-based tools (VaxiJen and Vaxign) and one locally installed program (VacSol) were used to screen and detect prioritized proteins as vaccine candidates. The selection was based on cell topology, secondary structure, presence of epitopes and non-homology to eukaryotic proteins. In silico prediction of vaccine candidates resulted in the selection of 70 proteins from M. pseudoshottsii and 89 from M. hippocampi. Eventually, from this set, we selected 11 genes that can be used for immunization against both Mycobacterium strains. The constructs were assembled by cloning these genes in the pcDNA 3.1 eukaryotic expression vector and their immunogenicity effects were estimated in zebrafish.

Keywords: Mycobacterium hippocampi, Mycobacterium pseudoshottsii, meagre, sea bass, fish disease, immune response

Title: “Drug development and manufacturing process”

Abstract: The presentation is designed to introduce students to the world of biopharmaceutical manufacturing, including innovative and generic products. The aim is to give a clear understanding of the complexity of the drug development and manufacturing process, understanding the requirements for drug approval, distinguishing clinical phase I, II, III, and IV and understanding the balance between medical benefit, medical risk, and economical reward.

The drug life cycle, from discovery to patients, will be covered in this presentation. The different steps of drug discovery, development, pre-clinical and clinical trials, regulatory submission and market authorization will be also introduced.