MICA: RIP1 kinase inhibition to reduce kidney injury

The human kidney is essential to life and has key roles of clearing the blood of toxins, maintaining salt and water balance and controlling blood pressure. The kidneys are sensitive to infection and other types of injury and this can lead to temporary or permanent loss of function. Kidney diseases and injury are common and place a large financial burden on health systems. The human cost of kidney disease is also important, often requiring long hospital stays, loss of income, and in the worse case scenarios the requirement for dialysis or transplantation.

A protein has recently been discovered called Receptor-interacting serine/threonine-protein kinase 1 or the shortened version RIP1 kinase. RIP1 kinase has emerged as an important protein which can regulate how cells respond when they are injured. When a cell is injured it may survive or if the injury is too severe it may die. A cell can die in a number of different ways and the way in which a cell dies is important because it determines the degree of inflammation that is caused. Cells dying by a process called apoptosis (shrivelling up) cause very little inflammation whereas cells dying by a process of necrosis (bursting apart) cause a lot of inflammation. There is also an intermediate route of cell death called necroptosis which is also associated with less inflammation that necrosis. The degree of inflammation affects the function of the organ and also influences the completeness and speed of repair. RIP1 kinase has a key strategic role in directing injured cells towards apoptosis and necroptosis depending on the precise circumstances and mechanism of cell injury. Early work has shown that blocking the effects of RIP1 kinase can reduce the amount of cell death a kidney experience when it is injured. Our own and others work has suggested that blocking RIP1 kinase may be a good approach to reducing injury and improving the function of kidneys exposed to situations that would normally cause serious injury and loss of function.

We have established a collaboration between the University of Edinburgh and GlaxoSmithKline, a major global pharmaceutical company to investigate the effects of novel RIP1 kinase blocking drugs on kidney function using cell and mouse models of kidney injury. The proposed project is being undertaken in partnership with Kidney Research UK the leading UK charity involved in research into kidney disease. The proposed work will test two new drugs and compare them with a naturally occurring RIP1 kinase inhibitor called necrostatin. The project will explore how important RIP1 kinase is in kidney injury both in the mouse model but also in human patients who have undergone kidney transplantation. It will investigate how the drug works using a genetically modified mouse with an inactive RIP1 kinase gene and will also look at the effect of the new RIP1 kinase blocking drugs on other molecular pathways in the cell. The project will also study the optimum timing and dosage of drug to get the best protective effect in out model. All of this information will be important in deciding the optimum targets for a drug to influence the RIP1 kinase pathway, the dosage and delivery of a drug and allow prediction of possible side effects. This information will be crucial to future development of this class of drugs as they are moved into large animal or human trials.

This project will also provide support and an opportunity for academic career development for a young surgeon with an excellent early academic track record who wishes to train as a specialist kidney surgeon and also to develop the research experience and skills which will allow him to continue research into clinically important kidney diseases.

Importance

Acute kidney injury (AKI) is a major health problem affecting 2000 patients per million per year in the UK. In addition, ischemic injury to kidneys associated with partial nephrectomy, transplantation vascular and cardiac surgery contributes significant mortality and morbidity. No active pharmacological agent is currently in use to reduce AKI.

Solution

Regulation of inflammation and cell death and survival pathways is a crucial determinant of cell fate in response to injury. Necroptosis is a newly recognized, regulated form of necrosis, which contributes to kidney injury in renal injury and transplantation. Receptor-interacting protein-1 kinase (RIPK1) has emerged as a kinase with strategic importance in directing cell fate toward necroptosis. This project is testing two inhibitors of RIPK1 in order to reduce kidney damage and improve kidney function as well as survival following renal IRI.

Objectives

1. Determine efficacy of RIPK1 inhibitors in human/murine cell-based models of renal injury.
2. Determine protective effect from renal IRI in RIPK1-kinase dead (RIPK1-KD) mice compared with wild-type (RIPK1-WT) mice and determine if RIPK1 inhibition abrogates renal injury.
3. Determine evidence for expression and activation of RIPK1 in human acute kidney injury.

Research plan

Cell culture experiments will determine RIP1K phosphorylation after injury and define dose response curves of RIP1K inhibitors. Renal ischemia reperfusion injury in RIPK1-KD mice and RIP1K-WT mice will establish whether inactivation of RIPK1 results in renal protection. RIPK1 inhibitors and necrostatin will be used in both RIPK1-KD and RIPK1-WT mice to demonstrate potential non-RIPK1 effects. The extent RIPK1 inhibitors influence other pathways will be determined by transcriptome analysis. RIPK1 expression and phosphorylation will be explored in human kidney tissue.

Patients and the health care sector

Manipulation of necroptosis has the potential to reduce the injury sustained as a consequence of acute kidney injury (AKI). Developing drugs in this area have significant potential to improve patient care in those who have suffered AKI. Agents may also improve the quality of transplanted organs and convert organs previously deemed unsuitable for use to organs that are usable. Other applications include use during partial nephrectomy surgery for kidney cancer, vascular surgery, and cardiac surgery. Requirement for renal replacement therapy after AKI is associated with significant mortality. Interventions in this area are greatly needed and could reduce mortality and morbidity associated renal ischemia reperfusion injury.

National economic impact

AKI is a major health problem with a significant economic impact that has been underestimated in ths past. With a prevalence of 2000-3000 per million population per year in Europe, AKI is associated with 40000 excess inpatient deaths in England. In addition, a significant proportion of those affected by AKI require renal replacement therapy (RRT) with costs estimated to exceed $10 billion in the US. Any intervention which reduces the prevalence of AKI has the potential to make an significant economic impact.

Academic impact

Necroptosis and RIPK1 and RIPK3 biology are exciting new areas of research. The mechanistic studies described in this project will provide new knowledge in this area with a clear focus on the potential to intervene in this process. This information will be of considerable scientific and clinical value for the future development of drugs targeting necroptosis. There is also the potential that this work could be extended to look at other clinical indications involving ischemia. Functional insights in necroptosis pathways may provide avenues for research by other groups that can be freely emulated after dissemination.

Collaboration with industry

The interaction of academic institutions with industry is important. This Fellowship represents a unique opportunity to broaden the traditional interface by which these two bodies collaborate. By formalising industry collaboration in early years training awards, such as a Clinical Training Fellowship, young talented investigators are exposed to a breadth of training beyond that offered by an academic institution alone. Furthermore, the benefits to the academic institution from this direct link with industry are tangible: examples include access to a broad range of candidate molecules, multiplex approaches to target validation and future opportunities for collaboration beyond that of the immediate project. Our industrial collaborators tell us of the benefits to them; in particular, the closer links with front-line clinicians, patients and treatments provides direction not necessarily easily accessible otherwise. By developing these ideas and broader industry collaboration, relationships between academia and pharmaceutical companies can be exploited, ultimately for the benefit of patients.