Fragiadaki lab Research Interests
My group's long-term goal is to understand the molecular and cellular mechanisms that cause chronic kidney and vascular disease (CKD / CVD). CKD/CVD are leading causes of death worldwide. My lab combines genetic, molecular, high-throughput screening and bioinformatics approaches to address key questions using human and mouse models of disease. Our most recent work is focused on the novel role of RNA binding proteins (RBPs) in the development of Autosomal Dominant Polycystic Kidney Disease (ADPKD), which is the commonest genetic form of renal failure. In addition we are interested in growth promoting signalling pathways, with an emphasis on JAK/STAT signalling and fibrogenesis control. Detailed research interests are listed below.
We grow renal tubular epithelial cells from patients with ADPKD, which are cultured in three-dimensional matrices in which they produce cysts that enlarge over time. With this model we can inhibit gene expression (pharmacologically or with gene silencing strategies) and study the ability of the cells to proliferate and survive. Additionally, gene expression can be enhanced e.g. via cytokine stimulation and/or gene editing. This ex-vivo approach is complementary to our mouse genetic models and studies of human samples.
Three areas of focus:
1) Which signalling pathways are crucial for the development of Polycystic Kidney Disease?
Autosomal Dominant Polycystic Kidney Disease (ADPKD) is a monogenic, multi-organ disease affecting both kidneys and blood vessels, currently lacking a cure. More than 12 million people are affected with this devastating disease and approximately 50% of patients will develop renal failure by the age of 50, requiring lifelong renal replacement therapy or transplantation for survival. ADPKD is due to mutations in one of two genes, known as Pkd1 (85% of cases) and Pkd2 (25% of cases), which encode for Polycystin-1 and polycystin-2, two mechanosensitive ciliary proteins of largely unknown functions. Despite the monogenic nature of this disease, the molecular mechanisms and signalling events that lead to relentless growth of cysts, vascular dysfunction and progressive fibrosis are elusive.
I have recently received funding from the Academy of Medical Sciences (Springboard Fellowship Award - 2018-2020) to combine transcriptomics and functional genomics technologies to identify and characterize key genes involved in cyst growth. My group has provided strong evidence for the involvement of (multiple components of) the JAK/STAT signalling pathway in ADPKD progression. Our next-generation sequencing approaches have revealed roles for additional pathways, signalling pathway cross-talk and development of disease, and these are currently under investigation in the Fragiadaki Lab.
2) How does ANKHD1 and other novel RBPs control renal and vascular dysfunction?
Ankyrin Repeat and Single KH Domain 1 (ANKHD1) is a evolutionarily conserved RNA binding protein (RBP), which we descovered can bind to microRNAs and mRNA and control a number of critical processes. RBPs have fundamental roles in RNA processing (export, decay, splicing, localisation etc), yet their roles in kidney disease are completely unknown.
We performed a genome-wide RNAi screen in Drosophila, which identified Ankyrin Repeat and Single KH Domain 1 (ANKHD1) as a key regulator of JAK/STAT pathway activity. Critically, ANKHD1 controls the transcriptional output while also reducing JAK/STAT cytokine receptor levels (Ref-1, Ref-2). More recently, I discovered a major role for ANKHD1 in the control of renal cell carcinoma cell division and I showed that this was via direct physical interactions of ANKHD1 with tumour-suppressor microRNAs (Fragiadaki et al, 2018 + Figure 2).
I use these findings to better understand the role of ANKHD1 in controlling renal and vascular dysfunction, two pathologies observed in patients with autosomal dominant polycystic kidney disease. This work is funded by Kidney Research UK via an Intermediate Fellowship (2016-2020).
More recently my laboratory has uncovered a role for ANKHD1 in controlling vascular tone and function. Next-generation sequencing and functional genomics approaches will be used by Ms Areej Alahmandi (PhD student) to discover the molecular mechanisms utilised by ANKHD1 to drive vascular dysfunction in the context of atherothrombosis.
Figure 2: ANKHD1 is overexpressed in renal cell
carcinoma patients. A. Arrays of human kidney tissues representing 20 cases of Renal Cell Carcinoma and 3 control non-cancer healthy tissues
were stained with an anti-ANKHD1 antibody and microscopy performed using an upright
Olympus microscope. B. Haematoxylin and Eosin (H&E) staining of the above matching
tissues can be seen. C. qPCR was performed for ANKHD1 normalised to b-actin for noncancer
kidney tissue when compared to renal cell carcinoma. D. Sub-group analysis of the
ANKHD1 expression in the RCC patient population was unable to identify any differences in
the expression in the early (I/II) versus late stages of disease (III/IV), suggesting the early involvement of ANKHD1 in renal cell carcinoma. Error bars show mean
and standard error of the mean.
3) Can Growth-hormone antagonism treat polycystic kidney disease?
I have recently made the novel observation that growth hormone is enhanced by 10-fold in mice with polycystic kidneys (Fragiadaki, et al, 2017). Growth hormone (GH) can activate JAK/STAT signalling via engaging with growth hormone receptors, which are present in the kidney (Figure 3). GH-triggered STAT5 signalling in turn activates proliferation contributing to the relentless growth of cysts. I aim to block GH, in order to stop proliferation in kidney cells which will in turn reduce the growth of cysts. To study proof-of-principle whether GH inhibition can alter polycystic disease, I have received funding in the form of a PhD studentship from the University of Sheffield (2017-2020; held by Ms Fiona MacLeod). Ms MacLeod will generate specific GH antagonists and examine their efficacy in cellular and mouse models of ADPKD.
Figure 3: Serum circulating Growth Hormone (GH) is significantly increased in mice with polycystic kidney disease, when compared with wild-type littermate controls; and critically the receptor for GH is also present in the kidneys of such mice (right hand panel). These data together suggest that the kidney can respond to GH stimulation.
CURRENT GROUP MEMBERS
Dr Hannah Roddie, Post-doctoral Scientist (BHF funded)
Ms Fiona MacLeod, PhD student
Ms Areej Alahmadi (PhD student)
Past members of the Fragiadaki lab
Dr Barbora Ndreca, Research Technician, AMS-funded
Ms Daniela Pirri (PhD student)
Paco Illanes Alvarez, Research Technician (2019)
Foteini Patera, Research Assistant (2018)