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Antimicrobial resistance (AMR) in general and antibacterial resistance (ABR) in particular have become one of the biggest health challenges of this era, causing yearly millions of deaths worldwide. The overuse and misuse of antibiotic drugs since their discovery have exacerbated the problem, as bacteria evolved and survived different antibiotics through resistance mechanisms. A worrying increase in the emergence of “superbugs” and the stagnation in antibiotic discovery since the 1980s require drastic measures to curb the life-threatening aspect and the economic burden of such infections. Many strategies and action plans have been launched by different organizations such as the WHO, CDC, and ECDC to combat ABR on different fronts, including public awareness, AMR surveillance, improving sanitation, unnecessary usage reduction, etc. Nonetheless, the availability of novel antibiotics with different modes of action are urgently needed to address this issue.

In our work, we propose multiple methodologies for the discovery and design of peptidic compounds with antibacterial activities, that will subsequently be optimized to improve on their activities and pharmaceutical properties. One strategy involves the modification of known natural antibacterial peptidic secondary metabolites through rational analysis and SAR studies. A second strategy requires the synthesis of fragments from known antimicrobial peptides (AMPs), where in silico prediction tools are employed to dissect the structure of these larger bioactive AMPs. Another strategy uses the rational design of protein-protein interaction (PPI) modulators of important protein complexes in bacterial infections through in silico analysis. One more strategy relies on genome mining for the discovery of unknown peptidic natural products from bacteria, as these so-called silent genes might only be expressed naturally under very specific conditions. 

Regardless of the exact strategy involved, the aim is to chemically synthesize the target peptides as well as an analogue series for each, enabling testing and evaluation of these compounds against a panel of bacteria and fungi, including AMR ones. In each case, a feedback loop will enable the redesign of another round of compounds based on the observed bioactivities. In order to overcome the poor stability and bioavailability of peptidic structures, chemical modifications and the use of peptidomimetics will be used to circumvent these problems. Recently, there has been a renewed interest in peptides and peptidomimetics as potential therapeutics emanating from their wide range of biological activity and their higher affinity and specificity towards molecular targets.

Recently, peptides have been gaining more attention as they display a wide range of biological activities. More particularly, there has been an increased interest in the development of AMPs as antiinfectives, not only because of their innate biological activity, but also their important structural features and immune modulation capabilities. They have also shown a very low prevalence towards resistance development. Despite their promise, AMPs have poor in vivo stability and their clinical use is limited due to toxicity.

One such interesting AMP, lactomodulin, has both antibiotic and anti-inflammatory activity but is 53 amino acids long, making it additionally quite expensive to synthesize. In this project, we are analyzing the composition and structure of the parent peptide using in silico tools in order to generate shorter truncated versions based on AMP potential, helical propensity, and toxicity predictions. Out of nine fragments, one 15-mer in particular and its cyclic version have shown improved activity against both Gram-positive (0.15─0.45 µM) and Gram-negative bacteria (3─5.1 µM). Time killing assays further showed a fast bactericidal activity whereas the peptides were shown to be non-cytotoxic against human cell lines. Further peptide derivatives are currently being synthesized and tested for SAR study. Interestingly, other peptides from the initial library showed 50% retention of anti-inflammatory activity. These results pave the way for the development of unique peptides with Gram-positive, Gram-negative, or anti-inflammatory activities.

Another AMP of interest is laterosporulin 10 (LS10), a 52 amino acid peptides with three disulfide bonds. It has quite potent activity against MRSA and Mycobacterium tuberculosis (Mtb) resistant strain. Using the same strategy, shorter versions within the peptide were predicted to have AMP character and were synthesized and evaluated. Preliminary results show about 5 peptides having a 5 µM activity against MRSA. Currently, we are exploring more derivatives of these peptides for further evaluation.

The prevalence of nosocomial infections in ICU patients is very common, where ventilator-associated pneumonia contributes to longer hospitalization times and a higher mortality rate, not the least because of the high prevalence of multi-drug resistant strains. Acinetobacter baumannii is a leading cause of these infections, especially in the Middle East. Carbapenem-resistant A. baumannii (CRAB) is classified by the world health organization (WHO) as a critical priority 1 pathogen requiring the development of novel antibacterial agents. One key nutrient for the persistence and survival of A. baumannii in the lungs is the amino acid histidine. As histidine is not available in high enough quantities in the lungs of the host to be sequestered, it is biosynthesized by the bacteria through an energetically demanding pathway.1 The first reaction in this pathway is catalyzed by the ATP phosphoribosyltransferase complex (ATPPRTc), a heterooctameric complex of 2 subunits HisG and HisZ. The latter also serves as the allosteric binding site of the final product histidine, a negative feedback inhibitor. When histidine binds to the complex, ATPPRT is deactivated and the flux-controlling step of the histidine biosynthesis is slowed down immensely. Tricking the bacteria into halting its histidine biosynthesis affects its viability in the lungs, leading to bacterial death. In this work, we are developing various peptidic molecules, by independently targeting two modes of inhibition: allosteric binding and protein-protein interaction (PPI) inhibition. The design is aided by computational tools and guided by structure-activity relationship. A first library of 15 dipeptides and tripeptides were synthesized and their inhibitory activity is being assessed and compared to the initial hit dipeptide His-Trp, as allosteric binding site inhibitors.2 On the other hand, five de novo predicted 10-mer peptides are also being tested as PPI inhibitors of the ATPPRT enzymatic complex. The peptidic hit compounds will be further modified chemically to generate peptidomimetic or hybrid derivatives while improving their binding affinity, stability, and physicochemical properties.

The   recent  advances  in  bioinformatics,  data mining tools, and sequencing technologies have transformed  the  discovery  of natural products. These  technologies  are  potentially  capable  of identifying all secondary metabolites produced by any organism, eliminating  many of the problems encountered in the traditional drug discovery process such as silent genes. Non-ribosomal peptide synthases (NRPS)  are  large  multi-modular  enzymes  that catalyze the formation of diverse peptides in living organisms. Using these advances, the primary structure of the resulting peptides peptides  can be  predicted to a certain degree of certainty directly from genomic data. Furthermore, their biosynthetic gene clusters will be annotated with a degree of similarity to known ones, allowing more focus on novelty. In this project, we aim to identify, synthesize, and assess the activity of potential unknown antibacterial peptides encoded within the genomes of natural resources with a focus on bacterial strains.

The survival needs of species and their innate chemical defense mechanisms have shaped the structurally diverse plethora of secondary metabolites they produce to fend off microbial predators. Among those, many peptidic compounds exhibit potent antibacterial activities, some of which have already been used in the clinic including the lipodepsipeptide daptomycin. Lipodepsipeptides are a class of natural antibiotics containing both a lipid chain and an ester linkage within their structure. The  cyclic  lipodepsipeptide lysocin has very potent antibacterial and   anti-tubercular   activity  with a   unique mode of action through either menaquinone or lipid-II binding. Despite its potency,  lysocin   has  poor  pharmaceutical  properties hindering its development as a drug. Our  goal  is  to  generate  de  novo  synthetic  hybrid peptides and peptidomimetics derived from the
natural  product  through  analysis  of   structure-activity relationship (SAR)  studies  and  targeted design  for  the  modulation of  their  chemical, physical, and pharmaceutical properties.

The detection and analysis of biomarkers of pathological states in the human body are challenging and often require long turnover times, time that is sometimes crucial for a rapid diagnosis. Hence, the development of novel chemosensors tailored for selective detection of transition metals and other biomarkers are of urgent need. Additionally, fluorescence sensing has emerged as an appealing and optimal detection technique. Typically, fluorescent sensors comprise a receptor and a fluorophore, where the receptor facilitates selective recognition and binding of the analyte such as the metal ions, while the fluorophore primarily converts recognition events into fluorescent signals. An ideal fluorescent chemosensor must fulfil two fundamental criteria. Firstly, the receptor, serving as the core component of a sensor, should exhibit a strong affinity to the metal analyte. Secondly, the fluorescence signal must remain unaffected by the surrounding environment.

The development of such novel molecules capable of selective interactions with transition metals hold promise as innovative theranostic agents (diagnostic and/or therapeutic) for many diseases such as Alzheimer's disease and heart failure. A hallmark of Alzheimer's disease is the formation of extracellular amyloid plaques containing the amyloid β (Aβ) peptide in the brain. High concentrations of Zn and Cu metal ions, ranging from 20 to 50 μM, are found in these plaques formed by metal-mediated amyloid β (Aβ) peptide aggregation. Cu(II) ions were correlated to stabilization of Aβ42 oligomer species as well as to formation of reactive oxygen species. Consequently, research intensely focuses on modulating the interaction between metal ions and soluble peptide species to mitigate their neurotoxic effects. On the other hand, research has found that copper is involved in regulating mitochondrial biological processes while an overload of it affects function. A recent study suggests that high serum copper levels are significantly associated with heart failure, where cell death can be caused by increased levels of copper (cuproptosis) through affecting the hyperacetylation of mitochondrial proteins. Hence, exploring copper chelators can be of huge interest in heart failure and cell death studies.

Currently used/reported chemosensors still suffer from many drawbacks that research attempts to solve such as low selectivity or sensitivity, and susceptibility to interference from metal ions with similar properties and poor water solubility. To address these issues, one of the major goals of this project is the development of novel chemosensors tailored for selective detection of transition metals and modulation of metal/peptide (or protein) interactions with high precision and sensitivity. 

On the other hand, transition metal complexes are key players in a wide range of other applications from photodynamic therapy to catalysis and photocatalysis. Catalysis is currently used to prepare more than 95% (in volume) of all today’s chemical products with a market size close to $16 billion for industrial catalyst manufacture. Photocatalytic conversions are appealing and widely used since they reduce costs of chemical production by using solar energy, reducing wastes and avoiding the use of stoichiometric reagents. Research is currently focusing on shifting these applications from materials based on noble and rare metals (4d and 5d) to metal systems based on eco-friendlier, less toxic and abundant 3d metals such as zinc, copper, and iron. Several high-impact breakthroughs were reported since 2013 despite the challenging electronic states of 3d complexes and their distortions. This ÐÇ¿Õ´«Ã½ the importance and challenges of developing new 3d metal/ligand systems for catalytic and photocatalytic applications. Developing greener and milder catalytic methods with higher selectivity is another major focus of this thesis, where novel metal complexes, adapted via minor structural changes to meet the criteria of catalysis, will be tested for various applications.