Project ID CM-HD2024_03


Co Supervisor 1A Faculty of Life Sciences & Medicine, School of Cancer & Pharmaceutical Sciences, Institute of Pharmaceutical ScienceWebsite

Co Supervisor 1B Faculty of Life Sciences & Medicine, School of Cancer & Pharmaceutical Sciences, Institute of Pharmaceutical ScienceWebsite

Targeted imaging of neutrophils and platelets in inflammation

Neutrophils and platelets are regarded as key mediators of inflammation, participating as first line players to the innate immune response. This project relies on the development of targeted imaging probes for optical visualisation of neutrophils and platelets, enabling a detailed understanding of their activation and functions in inflammatory processes and host defence mechanisms. To this end, a library of probes will be generated as selective binders of two classes of G-protein coupled receptors (GPCRs) – i.e. formyl peptide receptors (FPRs) and purinergic P2Y receptors (both classes overexpressed on the surface of neutrophils/platelets and important for their activation during inflammation).

The project will involve chemical synthesis (via solution chemistry and bio-conjugation reactions with selected fluorescent tags), purification, analytical and biophysical characterisation of the probes (in Cilibrizzi’s and Rahman’s labs, KCL; during year 1). The biological activity, binding affinity and selectivity of the probes will be assessed with GPCR functional tests in vitro (e.g. G-LISA, Western blots) and in cells (i.e. chemotaxis, Ca2+ levels, RhoA, Rac-1), along with flow cytometry measurements (in Cilibrizzi’s, Rahman’s and Pitchford’s labs, KCL; during year 2). Microscopy experiments will be carried out to assess relevant aspects linked to neutrophil/platelet activation, signalling, trafficking and translocation (at the Nikon Centre, KCL; during year 3). Lastly, through in vivo models of inflammation, we plan to profile the movement and functions of fluorescently labelled human neutrophils/platelets and investigate the migration of activated cells across tissues under simulated inflammation stimuli, along with in vivo imaging experiments to observe, spatially and temporally, and quantify the influx of cells (in Pitchford’s lab, KCL; during year 4).

This project will further our understanding of how neutrophils and platelets participate to endogenous inflammatory responses, with a focus on diseases of the airways (e.g. viral/bacterial infections leading to pneumonia, asthma, COPD), to inform new therapeutic opportunities.

Representative Publications

Nielsen, C.D.T.; Dhasmana, D.; Floresta, G.; Wohland, T.; Cilibrizzi. A. Illuminating the path to target GPCR structures and functions. Biochemistry 2020, 59(40): 3783-3795. Vergelli, C.; Schepetkin, I.A.; Ciciani, G.; Cilibrizzi, A.; Crocetti, L.; Giovannoni, M.P; Guerrini, G.; Iacovone, A.; Kirpotina, L.N.; Ye, R.D.; Quinn, MT. Synthesis of Five‐ and Six‐Membered N‐Phenylacetamido Substituted Heterocycles as Formyl Peptide Receptor Agonists. Drug Dev. Res. 2016, 78 (1), 49-62. Schepetkin, I. A.; Khlebnikov, A. I.; Giovannoni, M. P.; Kirpotina, L. N.; Cilibrizzi, A.; Quinn, M.T. Development of small molecule non-peptide formyl peptide receptor (FPR) ligands and molecular modeling of their recognition. Curr. Med. Chem. 2014, 21 (13): 1478-1504. DOI: 10.2174/0929867321666131218095521
Arkless, K. L., Pan, D., Shankar-Hari, M., Amison, R. T., Page, C. P., Rahman, K. M., & Pitchford, S. C. (2023). Stimulation of platelet P2Y1 receptors by different endogenous nucleotides leads to functional selectivity via biased signalling. British Journal of Pharmacology, 1– 16. RT Amison, S Jamshidi, KM Rahman, CP Page, SC Pitchford. (2018). Diverse signalling of the platelet P2Y1 receptor leads to a dichotomy in platelet function. European journal of pharmacology 827, 58-70 Dingxin Pan, Khondaker Miraz Rahman, Simon Pitchford. Exploring bias in platelet P2Y1 signalling: host defence versus haemostasis. Authorea. January 17, 2023. DOI: 10.22541/au.167396408.87937183/v1