Biology subjects

Scapozza, L.

Publications and source records attributed to Scapozza, L..

4 recordsLinked to original sources

bioRxiv2024

Discovery of the first-efficacious A2AR negative allosteric modulators for high adenosine cancer immunotherapies

Inhibition of adenosine 2A receptor (A2AR) is recognized as a promising immunotherapeutic strategy but is challenged by the ubiquity of A2AR function in the immune system. To develop a safe yet efficacious immunotherapy, the discovery of a novel negative allosteric modulator (NAM) was preferred. Leveraging an in-house, sensitive, high-throughput screening cellular assay, novel A2AR NAM scaffolds were identified followed by an extensive structure-activity relationship (SAR) study, leading to the discovery of potent 2-amino-3,5-dicyanopyridine derivatives. Allosteric mode of action of active compounds was confirmed by shift assay, non-linearity of the Schild plot analysis, biophysical measurements, and retained satisfactory potencies in high-adenosine concentrations. Further correlation of A2AR engagement and downstream signaling was done in a human blood translational assay, clearly showcasing the potential of A2AR allosteric modulation as a novel approach for efficient and safer cancer immunotherapies.

pharmacology and toxicology↗

bioRxiv2022

Targeted protein degradation reveals BET bromodomains as the cellular target of Hedgehog Pathway Inhibitor-1

Target deconvolution of small molecule hits from phenotypic screens presents a major challenge. Illustrative of these are the many screens that have been conducted to find inhibitors for the Hedgehog (Hh) signaling pathway - a major developmental pathway with many implications in health and disease - with many hits but very few identified cellular targets. We here present a strategy for target identification based on Proteolysis-Targeting Chimeras (PROTACs), combined with label-free quantitative proteomics. We developed a PROTAC based on the downstream Hedgehog Pathway Inhibitor-1 (HPI-1), a phenotypic screen hit with unknown cellular target. Using our Hedgehog Pathway PROTAC (HPP) we identified and validated BET bromodomains to be the cellular targets of HPI-1. Furthermore, we found that HPP-9 has a unique mechanism of action as a long-acting Hh pathway inhibitor through prolonged BET bromodomain degradation. Collectively, we provide a powerful PROTAC-based approach for target deconvolution, that has answered the longstanding question of the cellular target of HPI-1 and yielded the first PROTAC that acts on the Hh pathway.

cell biology↗

bioRxiv2022

In silico investigation of cytochrome bc1 molecular inhibition mechanism against Trypanosoma cruzi.

Chagas disease is a neglected tropical disease caused by the kinetoplastid protozoan Trypanosoma cruzi. The only therapies are the nitroheterocyclic chemicals nifurtimox and benznidazole that cause various adverse effects. The need to create safe and effective medications to improve medical care remains critical. The lack of verified T. cruzi therapeutic targets hinders medication research for Chagas disease. In this respect, cytochrome bc1 has been identified as a promising therapeutic target candidate for antibacterial medicines of medical and agricultural interest. Cytochrome bc1 belongs to the mitochondrial electron transport chain and transfers electrons from ubiquinol to cytochrome c1 by the action of two catalytic sites named Qi and Qo. The two binding sites are highly selective, and specific inhibitors exist for each site. Recent studies identified the Qi site of the cytochrome bc1 as a promising drug target against T. cruzi. However, a lack of knowledge of the drug mechanism of action unfortunately hinders the development of new therapies. In this context, knowing the cause of binding site selectivity and the mechanism of action of inhibitors and substrates is crucial for drug discovery and optimization processes. In this paper, we provide a detailed computational investigation of the Qi site of T. cruzi cytochrome b to shed light on the molecular mechanism of action of known inhibitors and substrates. Our study emphasizes the action of inhibitors at the Qi site on a highly unstructured portion of cytochrome b that could be related to the biological function of the electron transport chain complex.

biophysics↗

bioRxiv2018

Identification of novel anti-bacterial compounds from a chemically highly diverse pathways-based library using phenotypic screens in amoebae host models

Legionella pneumophila and tubercular Mycobacteria are the causative agents of potentially fatal diseases due to their pathogenesis but also to the emergence of antibiotic resistance that limits treatment strategies. The aim of our study is to explore the antimicrobial activity of a small ligand-based chemical library of 1,255 structurally diverse compounds. These compounds were screened in a combination of three assays, two monitoring the intracellular growth of the pathogenic bacteria, Mycobacterium marinum and L. pneumophila, and an additional anti-virulence \"plaque\" assay for M. marinum. We set up these assays using two amoeba strains, the genetically tractable Dictyostelium discoideum and the free-living amoeba Acanthamoeba castellanii. In summary, sixty-four compounds showed anti-infective/anti-virulence activity in at least one of the 3 assays. The intracellular assays hit rate varied between 1.7% (n=22) for M. marinum and 2.8% (n=35) for L pneumophila with 7 compounds in common between both pathogens. In parallel, 1.2 % (n= 15) of the tested compounds were able to restore D. discoideum growth in presence of M. marinum spiked in a lawn of Klebsiella pneumoniae. We also validated the generality of the hit compounds identified using the A. castellanii-M. marinum anti-infective screen in the powerful D. discoideum-M. marinum host-pathogen model. The characterization of anti-infective and antibacterial hits in the latter infection model revealed compounds able to reduce intracellular growth more than 50% at 30 M. Our studies underline the relevance of using a combination of low-cost and low-complexity assays with full 3R compliance associated with a rationalized focused library of compounds to help identifying new chemical scaffolds and dissect some of their properties prior to run further compounds development steps.

microbiology↗