Targeting Integrin-Linked (pseudo) Kinase in glioblastoma
Lead Research Organisation:
University of Edinburgh
Department Name: Edinburgh Cancer Research Centre
Abstract
ILK (Integrin-Linked Kinase) is a protein found at sites where cells adhere to their environment. For many years ILK was considered to be a classical enzyme whose activity could be inhibited, but it is now known that instead, it works as a scaffold to bind other proteins in complexes that control how cells behave. In examining several important proteins at adhesion sites in mouse brain progenitor cells that harbour cancer-causing mutations, so as to model the most aggressive brain cancer known as glioblastoma (GBM), we have found that loss of ILK protein causes profound effects associated with reduced malignancy. In brief, cells and tumours can no longer grow properly, and the cells lose their ability to invade surrounding material. Also, the cells lose their 'stem-like' state, often associated with problematic cells in tumours that do not respond the treatment. Other researchers have used unbiased genetic screens and also found that ILK is a promising protein to change the cancerous behaviour of human GBM cells.
In this project, we propose to use the most up to date form of gene editing to remove the ILK gene, and so protein, from human GBM cells that were recently derived from patients in Edinburgh. The idea here is to perform proof-of-principle experiments that will tell us whether, and if so how, the ILK protein plays important roles in human GBM cells. In beginning of our experiments in human GBM cells, we made an intriguing finding, namely that one of the proteins that binds to ILK is perturbed. We will investigate whether or not this is an important change, and might predict if the cells have adopted new ways to function.
As well as addressing ILK function in human GBM cells in depth, we will also try to find out how best to combine loss of ILK with drugs that enhance the biological effects, and the duration of effects. For this, we will use state-of-the-art drug discovery methods that "paint" cells and study their shape using high-throughput microscopy. We have used this before to successfully identify combinations between deletion of other adhesion proteins and drugs in a different cancer type. A series of well-understood chemical (drug) libraries that we have built for such purposes in our Institute will be used. Agents that work together with ILK loss to enhance advantageous biological effects will be studied in more detail and the strongest hits will be prioritised for further study to understand how they are working.
Finally, we propose to take the bold approach of generating a novel kind of drug that binds to ILK and cause its protein complexes to disintegrate. The strategy will ensure that biological effects are guiding the synthesis of chemical series (so selecting for agents that get into GBM cells and have good effects ('selecting the winners') early in the drug discovery process. These will be optimised by further chemical modification and then testing in GBM cells. We think this strategy will work because we already have agents that bind to the region of ILK that is needed to maintain the levels of its protein partners, and these can be further chemically modified to improve them. As proof this can work, we have recently used this strategy to develop drug candidates licensed to Pharma.
Our vision is to deepen understanding of GBM cell biology and the role of one important adhesion protein, namely ILK; this is much-needed because no new treatments for this dismal disease have been forthcoming over the past two decades. ILK is not a new target, but we have substantial new information in more physiologically relevant systems to study GBM cells than used previously, implying that ILK may be an excellent target for therapy. New experiments to fully understand its role in GBM, and bold new approaches to: a) determine how best to combine ILK-deficiency with other drugs, and b) make our own small molecule inhibitors for onward translation, will further our aim of ultimately providing benefit to patients.
In this project, we propose to use the most up to date form of gene editing to remove the ILK gene, and so protein, from human GBM cells that were recently derived from patients in Edinburgh. The idea here is to perform proof-of-principle experiments that will tell us whether, and if so how, the ILK protein plays important roles in human GBM cells. In beginning of our experiments in human GBM cells, we made an intriguing finding, namely that one of the proteins that binds to ILK is perturbed. We will investigate whether or not this is an important change, and might predict if the cells have adopted new ways to function.
As well as addressing ILK function in human GBM cells in depth, we will also try to find out how best to combine loss of ILK with drugs that enhance the biological effects, and the duration of effects. For this, we will use state-of-the-art drug discovery methods that "paint" cells and study their shape using high-throughput microscopy. We have used this before to successfully identify combinations between deletion of other adhesion proteins and drugs in a different cancer type. A series of well-understood chemical (drug) libraries that we have built for such purposes in our Institute will be used. Agents that work together with ILK loss to enhance advantageous biological effects will be studied in more detail and the strongest hits will be prioritised for further study to understand how they are working.
Finally, we propose to take the bold approach of generating a novel kind of drug that binds to ILK and cause its protein complexes to disintegrate. The strategy will ensure that biological effects are guiding the synthesis of chemical series (so selecting for agents that get into GBM cells and have good effects ('selecting the winners') early in the drug discovery process. These will be optimised by further chemical modification and then testing in GBM cells. We think this strategy will work because we already have agents that bind to the region of ILK that is needed to maintain the levels of its protein partners, and these can be further chemically modified to improve them. As proof this can work, we have recently used this strategy to develop drug candidates licensed to Pharma.
Our vision is to deepen understanding of GBM cell biology and the role of one important adhesion protein, namely ILK; this is much-needed because no new treatments for this dismal disease have been forthcoming over the past two decades. ILK is not a new target, but we have substantial new information in more physiologically relevant systems to study GBM cells than used previously, implying that ILK may be an excellent target for therapy. New experiments to fully understand its role in GBM, and bold new approaches to: a) determine how best to combine ILK-deficiency with other drugs, and b) make our own small molecule inhibitors for onward translation, will further our aim of ultimately providing benefit to patients.
Technical Summary
ILK is a pseudokinase and one of the key nodes of the consensus integrin adhesome. We have identified it as a potential target for therapeutic intervention in glioblastoma (GBM). We have found that CRISPR/Cas9-mediated ILK gene deletion from a transformed neural stem cell model of GBM causes substantial phenotypes, including destabilization of the ILK-Pinch-Parvin (IPP) complex through which ILK functions and profound morphological changes, suppression of invasion in vitro that is linked to inhibition of putative invadopodia, reduced expression of invasion- and stem cell- associated genes and inhibition of tumour growth in vivo. Using whole genome CRISPR screen, others have identified ILK as a key vulnerability in patient-derived GBM stem cells, providing support for ILK as a therapeutic target in GBM. We have found that there is a previously unrecognised Pinch isoform substitution in the IPP complex in some mesenchymal human GBM cells, which we wish to understand.
Aims
1) examine how genetic deletion of ILK influences cancer-associated phenotypes across a panel of genomically-annotated human GBM stem cells in vitro and in vivo
2) examine the status of the ILK-Pinch-Parvin (IPP complex - in which we have identified an interesting substitution in mesenchymal GBM cells)
3) perform chemical-genetic screens for possible synergistic combinations with ILK loss using target-annotated chemical libraries
4) develop small molecule inhibitors that degrade the IPP complex in GBM cells via a novel (but proven) chemistry-phenotypic iteration strategy using starting material of agents that dock into the (pseudo)'active' site of ILK
This will establish whether ILK is indeed a high-value target in human GBM and determine how it functions; we will address how best to combine ILK loss with existing drugs and generate small molecule probes or drugs that destabilise the IPP complex and mimic ILK deletion, the latter judged by high-content morphometric fingerprinting.
Aims
1) examine how genetic deletion of ILK influences cancer-associated phenotypes across a panel of genomically-annotated human GBM stem cells in vitro and in vivo
2) examine the status of the ILK-Pinch-Parvin (IPP complex - in which we have identified an interesting substitution in mesenchymal GBM cells)
3) perform chemical-genetic screens for possible synergistic combinations with ILK loss using target-annotated chemical libraries
4) develop small molecule inhibitors that degrade the IPP complex in GBM cells via a novel (but proven) chemistry-phenotypic iteration strategy using starting material of agents that dock into the (pseudo)'active' site of ILK
This will establish whether ILK is indeed a high-value target in human GBM and determine how it functions; we will address how best to combine ILK loss with existing drugs and generate small molecule probes or drugs that destabilise the IPP complex and mimic ILK deletion, the latter judged by high-content morphometric fingerprinting.
