Projeto Portugal 2030

Sumário

There is an unmet need of new drugs to eradicate Helicobacter pylori (Hp), the major etiological agent of gastric cancer & other gastric ailments[12]. Building on our recent preliminary data supporting that Hp recognizes surface grafted Cholesterol (Chol)(Fig.1-4-ANNEX) & Chol analogues(CA) (ergosterol, Fig.5-ANNEX) in solution, HELIPY AIMs to develop an innovative antibiotic-free strategy to specifically overcome Hp gastric infection based on CA grafted on chitosan nanoparticles(CA-ChNP). Our hypothesis is that CA-ChNP will kill Hp directly by internalization of CA-ChNP & indirectly by blocking its capacity to incorporate Chol from host epithelial cells. We also hypothesize that this bactericidal mechanism will aid in reinstating normal local immune response by abating the destruction of lipid rafts on host epithelial gastric cells & impairing Hp-associated chronic inflammation. This is an extra advantage for preventing gastric pathogenesis. Main objectives: 1)Selection of most effective CA that, after surface grafted, are recognized by Hp A 1st screening will be done w/ self-assembled monolayers(SAMs) of alkanethiols. SAMs technology(Team core expertise) is excellent to evaluate interactions of bacteria w/ grafted ligands, as it allows to engineer chemically well-defined surfaces at nanoscale, allied w/ easy preparation/functionalization & compatibility w/ various surface characterization techniques [13-16]. SAMs will allow to decipher the importance of CA concentration, orientation & exposure from the surface in Hp recognition/adhesion. 2)Development of cytocompatible CA-ChNP for Hp eradication ChNP will be produced & functionalized w/ selected CA using an one-pot microfluidic device recently developed by us(Fig.7-ANNEX[11]). CA-ChNP will be screened in vitro for: i)bactericidal effect on clinically relevant Hp strains & Hp biofilm model optimized by us[17]; ii)cytocompatibility against human gastric cell lines & immune cells; iii)interaction w/ gut microbiota by using representative bacterial strains from the gastro-intestinal tract; iv)ability to cross the mucus layer & compete w/ Chol from gastric cells in an in vitro infection model optimized by us that includes physical(epithelial cells & mucus) & chemical(acid medium) barriers[18]. CA-ChNP w/ good in vitro performance will be tested in vivo (mice model established by us at i3S [19]). 3)Study CA-ChNP ex vivo & effect on local immune response Ex vivo studies will be done on a humanized model. Hp(+) gastric biopsies & immune cells isolated from matched patients blood will be used to study the ability of CA-ChNP to kill Hp & induce immune cells activation. In parallel, studies on Hp(-) gastric biopsies & Hp(-)gastric biopsies colonized w/ other highly aggressive Hp clinical strains (not detected on biopsy) will be done. MAJOR ADVANTAGES: i) prospect for massive eradication ii) antibiotic-free strategy, avoiding resistance iii) Hp targeted specificity, minimizing interference w/ gut microbiota iv) reestablishment of local adaptative immune response & decrease of unresolved inflammation. Overall, we will develop a bold & disruptive bioengineered targeted strategy to treat Hp infection by killing & blocking Hp capacity to extract cholesterol from host cells, crucial for its survival & to downregulate the local immune response. HELIPY will represent a major advance in infection, immunology & bioengineering sciences, delivering a targeted strategy against Hp chronic infection.