Individuals with a genetic tumour risk syndrome are at high risk to develop cancer. One of these syndromes is PTEN hamartoma tumour syndrome (PHTS). PHTS is associated with pathogenic variants (PVs) in the PTEN gene. Unfortunately, in the majority of patients suspected of PHTS no PV in PTEN is identified. These patients are considered PHTS-like. The low diagnosis rate in this group of patients suggests the possibility of PVs in regions of PTEN that are not covered in routine diagnostics or suggests that PVs in other genes in the PI3K/AKT/mTOR pathway result in PHTS-like disease. In order to identify the missing heritability in PHTS I aim to develop a long-read sequencing (LRS) approach for PHTS-like patients, which will go beyond the currently state-of-the-art short-read sequencing approaches. I will O1) develop a PTEN-targeted LRS assay and O2) perform this assay on a highly selective cohort of PHTS-like families (n=94). I then will O3) perform whole genome LRS on 25 families that were not solved by targeted LRS to identify novel gene-disease associations. At last, I will O4) validate these new associations in a wider cohort. Via this work, I will improve the diagnostic yield for PHTS by identifying novel genetic mechanisms and genes underlying disease in PHTS-like patients. The unique interdisciplinary structure of the Department of Human Genetics at Radboudumc will allow to rapidly implement genetic testing for the newly identified genetic causes for PHTS. Identification of the precise underlying cause of PHTS will enable cascade genetic testing of at-risk family members to determine their status for the causal variant and thus, risk of disease to them and any future off-spring. Furthermore, this approach will allow the implementation of preventive care strategies for the family members at risk. Overall, my proposal will facilitate a personalized care approach for PHTS(-like) patients and their families for improved clinical outcomes and reduced disease morbidity

Although breast cancer screening and improvements in treatment have reduced breast cancer mortality by 30%, this disease still kills over 600,000 women yearly worldwide. To further reduce its morbidity and mortality, new and improved imaging modalities that improve the detection, diagnosis, and treatment of breast cancer are needed. Over the last two decades, changes in x-ray detector technology have revolutionized x-ray-based breast imaging. Among other developments, new digital detectors made feasible the ERC Consolidator project BREAST4D during which a new imaging modality, dynamic contrast enhanced dedicated breast computed tomography (4D DCE-BCT) is being developed to personalize treatment of breast cancer. However, x-ray source technology has remained essentially unchanged for the last 100 years. The continued use of the x-ray tube as the source of x rays restricts the reduction in dose and the quantitative accuracy achievable with advances in the rest of the system. Very recently, however, a new monochromatic x-ray source has been developed that provides the necessary flux, x-ray energy, field of view, and portability, to be used in x-ray breast imaging devices. In MONOBREAST, the capabilities, applicability, and innovativeness of BREAST4D will be extended by introducing these monochromatic x-ray sources to breast CT imaging and combining them with the advanced image reconstruction method developed during BREAST4D. Resulting in a dose reduction of at least two thirds, such a monochromatic breast CT system will be applicable to every stage in the breast cancer chain, from screening with non-contrast BCT at doses equivalent to those of current screening with digital mammography and digital breast tomosynthesis, to static CE-BCT for diagnosis and the originally planned 4D DCE-BCT for treatment personalization. In addition, these new sources will not only result in substantially lower doses, but also in substantially higher accuracy with lower algorithmic complexity.

Serotonin plays a prominent role in a wide range of psychiatric disorders. Importantly, serotonin (5-HT) is not only a neurotransmitter but also a neurotrophic factor during brain development, contributing to neurodevelopmental disorders (NDDs). One largely neglected source of 5-HT affecting development is maternal serotonergic genotype, which can influence offspring through the placenta and by changing maternal care behaviour. As a pediatrician, I am interested in the latter, as it -unlike the biological route- can be modified, providing an opening for early intervention for children with NDDs. Here, I will delineate for the first time how maternal serotonin transporter (5-HTT) genotype affects maternal care behaviour and impacts offspring development from an individual perspective. To understand the influence of maternal 5-HTT genotype on offspring’s development related to sex and genotype, I will use the well-established 5-HTT knockout (5-HTT-/-) rat modeling 5-HTT gene variance in humans and affecting maternal care. First, I will examine maternal care behaviours in wild-type and 5-HTT-/- mothers, and the developmental trajectories of male and female offspring with varying 5-HTT genotypes by tests assessing social and emotional behaviour from infancy to adulthood. Included are highly sophisticated analyzes of ultrasonic vocalizations, which I am an expert in. Second, I will elucidate the offspring’s brain-wide neuronal activity pattern using cutting-edge brain clearing and 3D lightsheet microscopy. To test causality and provide a lead for an early intervention, I will determine if enrichment ameliorates maternal care and improves offspring development. This project is expected to uncover how mother genotype influences offspring development in the context of behaviours relevant to NDDs and whether the developmental trajectory can be improved through a non-pharmacological intervention.

BACKGROUND | Renal tubulopathies are a large group of genetic disorders that disturb the function of the kidney tubule. A major challenge for diagnostics and treatment of magnesium (Mg2+) wasting tubulopathies, is absence of functional cell models and tools to study the pathophysiology. Diagnostics of hereditary Mg2+ wasting tubulopathies based on genetic screening, rather than biological measurements of kidney processes. Therapy largely depends on oral supplements because therapeutics that target the cause of disease are not available. VISION | In my vision, better insights in the cellular functional processes in the kidney are the next essential step forward towards better diagnostics, pathophysiological understanding and development of therapy. Development of functional Mg2+ transport measurements and innovative use of patient-specific models is required to increase diagnostic yield and to develop therapy. AIMS AND APPROACH | In this project, I aim to develop a urine-based diagnostic approach using scRNA sequencing and extracellular vesicle proteomics to provide biological diagnosis of renal tubulopathies. Using urine-derived adult stem cells, I will perform the first ever functional studies of ion reabsorption in kidney organoids. This approach allows personalized analysis of the pathophysiology as well as patient-specific drug screenings. Lastly, I will be the first to apply intravital multiphoton imaging in mice to measure Mg2+ reabsorption in the kidney tubule. The combination of patient-derived organoids and mice models will be used to test novel therapeutics. IMPACT AND INNOVATION | IN-THE-KIDNEY establishes the novel concept of biological diagnostics for renal tubulopathies, develops for the first-time patient-derived tissue models for functional analysis and introduces intravital ion transport measurements in mice. The methodology and approaches in this project will result in the identification of novel therapeutics for Mg2+-wasting tubulopathies.

The mechanisms underlying monogenic intellectual disability (ID) and autism spectrum disorder (ASD), affecting >2% of the population, remain poorly understood. Moreover, most ID/ASD syndromes are still untreatable, the exception being some disorders of metabolic origin. Therefore, identifying unappreciated metabolic mechanisms underlying the pathophysiology of ID/ASD provides a promising avenue to treatment. Several lines of evidence suggest that the transcription factors FOXP1/2/4 are ID/ASD genes with such an underappreciated role in brain (energy) metabolism: mitochondrial defects and altered metabolic rate in FOXP animal models, unpublished FoxP-/- brain transcriptome data of my host, and GWAS-based data indicating that FOXP1/2 genes are significantly associated with body mass index in the general population. Therefore, my main aim is to characterize ID/ASD phenotype-relevant metabolic dysregulation in FoxP mutants and ameliorate its effects. Powerful genetic tools in Drosophila allow me to probe specific molecular mechanisms with high spatial and temporal resolution and relate them to clinically relevant phenotypes, including locomotion behaviour habituation learning, and sleep. My objectives are to: 1) Identify phenotype-relevant metabolic target pathways of FoxP, 2) Characterize metabolic dysregulation in FoxP neurons with spatial resolution 3) Reverse behavioural FoxP phenotypes with metabolic or other innovative therapeutic interventions. This interdisciplinary approach further develops the data analyses, molecular, and imaging skills that I acquired through my PhD training and allows me to merge it with unique disease-relevant data, clinical knowledge, and collaborating networks. This project has the potential to provide novel insights into the relation of metabolism, cognition, and behaviour, and revolutionize the treatment of FOXP1/2/4 disorders. These insights may also have implications for further molecularly and clinically related ID/ASD disorders.