Vascular anomalies are rare soft tissue tumours and malformations formed by abnormal vascular elements of various types, and mainly affect infants, children and young adults. These lesions are painful, many lead to bleeding, infections, organ dysfunction, and they can damage and metastasise to other tissues. Current treatment strategies are invasive, not fully efficient, and there is a high risk of recurrence. To date no specific targeted therapies have been developed. Most of the studies on vascular anomalies are descriptive and focused on their clinical aspects; at the moment, there is a lack of molecular understanding and pre-clinical studies on the field of vascular anomalies. Thus, identifying the causative molecular alterations of vascular anomalies and understanding their biology will lead to more refined diagnoses and will provide better and more directed therapies. Recent published and unpublished observations from our laboratory show that activation of phosphoinositide 3-kinase (PI3K) signalling pathway drive to the developments of a fraction of vascular anomalies; however, the underlying molecular and cellular mechanisms remain enigmatic. In this proposal we aim to (i) unveil alterations in the PI3K signalling pathway components that are involved in the pathogenesis of vascular anomalies, (ii) understand the role of PI3K pathway activation in the development and maintenance of these lesions, and (iii) explore the potential of repurposing PI3K pathway inhibitors for these conditions. For this, we have set up collaboration with clinicians who will provide a large collection of vascular anomalies from patients. Also, in collaboration with Pharma, we aim to use PI3K inhibitors for these diseases. Importantly, these collaborations will allow us to translate our findings into the clinic for future clinical trials. This project proposal seeks to broad our understanding of PI3K activation in vascular biology and to ultimately improve patients’ quality of life.

Precise regulation of gene expression is achieved through the coordinated action of genomic cis-regulatory modules (CRMs). The identification of CRMs has long been a goal of functional genomics as CRM dysregulation can have devastating consequences for health and development such as autism. For example, mutations in the MeCP2 gene, which encodes protein that binds methylated DNA and regulates gene expression in neurons, results an Autism Spectrum Disorder termed Rett Syndrome. While thought to modulate CRM activity as a repressor, MeCP2’s function remains ambiguous and would benefit from a functional genomics characterization. To identify CRMs I recently developed a novel approach called FIREWACh. Here, I propose to utilize FIREWACh first to identify active CRMs within a homogenous population of neurons, identifying genomic loci of CRMs whose function may be compromised by MeCP2 mutations. I will adapt FIREWACh to allow the quantification of CRM output in a new method I propose to call ROQ-WACh (regulatory output quantification within accessible chromatin) by addition of barcodes to reporter mRNAs. This will allow high-throughput readout of CRM activity globally in vivo and will be broadly applicable to many biological fields. Lastly, I aim to combine the above approaches to quantify the changes in CRM output in response to pathological mutations in MeCP2. This Marie Sklodowska-Curie Action will allow me (the Experienced Researcher, Matthew Murtha) mobility to Spain to join the laboratory of Dr. Manel Esteller (Host Supervisor), renowned epigeneticist and expert in DNA-methylation biology to perform the research. Together the approaches and data generated by this MSCA action will provide key insights into the relationship between methylated DNA, precise control of gene expression, and high-order phenotypes such as cognitive behaviors.

Autoimmune encephalitides (AE) are a category of diseases that result from an immune attack against proteins present in patient’s own brain cells (antigens). Clinical symptoms include both neurological and psychiatric manifestations, that differ greatly between patients and during disease progression. Upon prompt diagnosis and immunotherapy symptoms gradually resolve. Current epidemiological studies estimate an annual incidence of encephalitis of any aetiology is 2-3 patients per 100.000 per year. However, given that new autoantibodies are being discovered on a half-yearly basis, the incidence of AE is increasing over time and is nowadays comparable to infectious encephalitis. Methods to diagnose AE rely on immunodetection of autoantigens, using rat brain tissue or transfected cells as a source of neural cell surface proteins. Despite being successful in identifying known autoantibodies, in 50% of all AE cases the affected antigen is unknown and cannot be detected using current technologies. Therefore, there is an urgent clinical need for a diagnostic kit that is able to detect all cell surface neuronal antigens. Thanks to the skills acquired during the ERC-StG, we have developed a platform to identify the presence of autoantibodies against neuronal surface antigens in patient serum or CSF using human induced pluripotent stem cell (hiPSC) derived neural cells. Moreover, first experiments validate that our technology can detect an autoantigen in seronegative samples. In the context of this project, we aim to conduct the necessary steps to bring a successful diagnostic approach developed in the bench towards its application as a diagnostic device at the bedside. To this end, we will develop a minimum viable product (MVP), to assess its capability to detect new AE-associated antigens, to distribute it to reference diagnostic laboratories for benchmarking and to explore the possibility of licensing the invention to a third-party company.

Cancer is a disease of the elderly and chemotherapy remains the mainstay of treatment. The benefits of chemotherapy include increased overall survival, improvement in quality of life, and palliation of symptoms. However, older patients are more susceptible to specific toxicities of chemotherapy, like myelosuppression and life-threatening neutropenia. Among the tissues strongly affected by chemotherapy, the bone marrow sinusoidal endothelial cells constitute the most important supportive niche for aged hematopoietic stem cells function and for myelopoietic recovery in the elderly. Up to now, few data are available about how aged sinusoidal niches regenerate upon chemotherapy damage and whether it is possible to rejuvenate vascular endothelial stem cells and improve the regeneration of the old sinusoidal niche as an effective strategy to improve HSC function and prevent myelosuppression and life-threatening neutropenia in the elderly. Here I hypothesize that the reduced regenerative capacity of aged sinusoidal niches can be improved by rejuvenating vascular endothelial stem cells in vivo via targeting Cdc42 activity and the Notch:Jag2 signaling. The current proposal investigates whether improving the regeneration of the aged sinusoidal niche might represent an important target to enhance the hematopoietic recovery and increase the survival after chemotherapy in the elderly. By combining several ground-breaking approaches ranging from single-cell sequencing, whole-mount bone marrow imaging, deep learning strategies for data analysis and integration, stem cell sorting techniques and specific mouse models, this research project will demonstrate that aging is not irreversible and that targeting the stem cell niche could represent an unprecedented innovative strategy to improve the regenerative capacity not only of hematopoietic stem cells.