Chemokines are small, secreted cytokines that activate membrane receptors belonging to the G protein-coupled receptor (GPCR) superfamily. The chemokine-receptor interactions control crucial physiological processes, but are also implicated in many pathologies, including atherosclerosis, inflammatory diseases, HIV infection or cancer, and hold a great potential for therapeutic intervention. Yet, although GPCRs are the target of about one third of currently marketed drugs, only three of them act on chemokine receptors, essentially due to poor structural understanding of the intricate interactions with their ligands. Indeed, about 50 chemokines and 20 receptors have been identified in humans. Their interactions require precise orchestration, as a chemokine may bind to several receptors, while a single chemokine receptor has multiple ligands. However, the structural determinants dictating ligand specificity and receptor activation remain elusive. Lately, cryo-EM has emerged as a ground-breaking technique for elucidating molecular structures but despite recent advancements, analysis of small proteins like receptor:chemokine complexes (∼50 kDa) is still challenging. In this project, we propose an innovative approach to increase the chemokine size, without altering its functionality, through a rigid insertion into a bulky scaffold protein. Such enlarged chemokines, called Megakines, will facilitate structural analysis of chemokine receptors. To prove this concept, we will first reformat the well-characterized chemokine CCL5 and use it in structural/functional studies with two clinically relevant receptors, CCR5 and ACKR2. The achievement of this project relies on the multidisciplinary combination of the excellent expertise in the chemokine field of the host laboratory and the protein engineering skills of the applicant. The proposed Megakine technology has the potential to revolutionize cryo-EM studies of chemokine receptors and facilitate the therapeutic development.

Mechanisms underlying cancer metastasis are poorly understood and context dependent. An essential feature of disseminating cancer cells is metabolic plasticity which allows cellular adaptation to changing environmental conditions along the metastatic cascade. I discovered in 2016 the phenomenon of formate overflow, an alternative pathway for serine catabolism via folate-mediated one-carbon (1C) cycle. Instead of using formate for nucleotide synthesis (to support proliferation), formate overflow is characterised by formate excretion from cancer cells. Furthermore, I have convincingly demonstrated that mitochondrial 1C metabolism and increased extracellular formate concentrations promote cancer metastasis in a growth-independent manner. However, it remains unknown which intermediates of 1C metabolism contribute to this phenotype, how formate controls metastasis and how it can be targeted therapeutically. To cross the current edge of knowledge, I will (1) take advantage of genetic and newly developed analytical methods to dissect how the formate-dependent effects are propagated in cells; (2) describe intrinsic mechanisms to explain how the formate signal is relayed to promote metastasis and (3) exploit formate overflow for cancer cell killing by directly targeting the formate molecule. 4M8 will explain a novel concept of how a mitochondrial metabolic signal drives cancer cells towards a pro-metastatic phenotype and how this knowledge can be translated into a signature to analyse the metabolic state in human tumour samples. Finally, we will develop a novel tool to target formate directly within cancer cells. In sum, our research will shed light on an unexplored field of cancer metabolism providing the foundation to develop novel treatment approaches against metastatic cancer, the primary cause of cancer death.

Hypertension is an extreme public health problem affecting nearly one third of the World’s adult population. Most cardiovascular diseases are provoked by hypertension resulting in an estimated 9.4 million deaths every year. Surprisingly, the diagnosis of hypertension has changed very little over the past 100 years - just using a stethoscope definitely needs refining. Furthermore, the epidemic proportion of hypertension worldwide pinpoints the necessity for the discovery of fresh antihypertensive therapies. Thus the main objective of the MITO project is to discover novel methods to improve healthcare of hypertension patients. For this we will investigate the circulating pool of microRNAs (small RNA molecules present in the blood, regulating gene expression) - we hypothesize that particularly mitochondrial microRNAs play a critical role in hypertension development. Easily obtainable in the blood, microRNAs constitute an accessible source of biomarkers and therapeutic targets for clinical application. The research plan envisages three scientific work packages dedicated (1) to the discovery of candidate microRNAs associated with hypertension by using small RNA sequencing in 30 hypertensive patients and 30 controls, (2) the validation of candidate microRNAs in a group of 540 participants (paying attention to sex specificities), and (3) to the functional assessment of the role of microRNAs in hypertension development. While the first two parts of the project will be conducted at the host institution in Luxembourg, functional studies will involve three secondments to partner institutions in Denmark, Germany, and France. A successful outcome will have implications ranging from contributing to satisfy the need for novel methods to stop the progression of hypertension, to a better understanding of the role of microRNAs in hypertension development, to the training of a young East European researcher to pioneer development in a research field just emerging in her home country.

Natural Killer (NK) cells have a critical role in anti-cancer immunity and recently the influence of the microbiome in cancer immunotherapy was shown. Comprehending the complex interplay between the expression of NK cells ligands involved in immune evasion mechanisms and the microbiome is crucial for achieving immunotherapy success. Diffuse Large B Cell Lymphoma (DLBCL) and Follicular Lymphoma (FL) are two types of Non-Hodgkin Lymphomas (NHL) characterized by accumulating B lymphocytes in lymph nodes, and extranodal sites. DLBCL and FL usually show aggressive and indolent behavior respectively. DLBCL, includes the known subtypes germinal center B-cells (GCB) and activated B-cells (ABC), and, like the DLBCL-GCB subtype, FL have a germinal origin as well. The mechanism involved in the immune-microbiome interplay are not clearly understood, hampering the improvement of therapeutic possibilities. The objective of this study is to reveal the complex interplay between the NK cells and the microbiome in DLBCL and FL patients and harness its therapeutic potential. To achieve this, I will use single-cell RNA sequencing and flow cytometry to analyze the composition of NK cells dissociated from DLBCL and FL samples. Then, I will use correlative analyses to identify the NK ligands/receptors that are more expressed in patients with low diversity and high variability microbiome. Ultimately, I will address the impact of this correlation in the response to immunotherapy. The results will pave the way for patient risk stratification establishment and new treatment options approaches. Conducting this project at the Tumor Stroma Interactions (TSI) group will provide me a solid scientific basis as well as technical development for further progressing my scientific career.