Antagonism of PI 3-kinase signalling by PTEN and SHIP2

Lead Research Organisation: Heriot-Watt University
Department Name: Sch of Engineering and Physical Science

Abstract

The human body is made up of cells which form the tissues and organs. The different tissues of the body have developed to perform specialised functions which must be coordinated for the organism as a whole to function efficiently and survive. Cell signalling is the process by which cells in our bodies communicate with one another and signal transduction is the means by which a specific signal (perhaps a hormone, such as insulin), arriving at its target tissue, is interpreted to elicit a particular response. Defects in cell signalling are common causes of important human diseases such as cancer and diabetes. We are studying the details of a signal transduction process which malfunctions in more than 50% of human tumours and which accounts for many of the effects of insulin produced after a meal. The central character of this response is a fatty substance or lipid called PIP3 which is made by enzymes called PI 3-kinases. When this substance is produced at the correct time, in the right part of the cell and in small, but sufficient amounts it triggers normal cell responses, but too much, in the wrong place or at an inappropriate time can lead to or promote the development of a tumour. On the other hand, producing too little in response to insulin can be a cause of diabetes. Maintaining this delicate balance of PIP3 involves the PI 3-kinase enzymes which make PIP3 and enzymes called phosphatases which remove it. We are studying the factors which regulate two classes of PIP3 phosphatase called PTEN and SHIP2. PTEN is a tumour suppressor that is mutated or absent in many different kinds of human tumour. We use tissue culture cells to study PTEN and SHIP2 regulation and epithelial cells (the source of most solid tumours) as simple models of disease to examine the consequences of defects in PIP3 phosphatase activity. Lastly, we are developing unique mouse models harbouring specific defects in the PTEN gene to validate the physiological significance of our work using cultured cells. Because of its importance in human disease many pharmaceutical companies are developing drugs which block the PI 3-kinase signalling pathway including inhibitors, currently in clinical trials as anti-cancer or anti-inflammatory agents, of PI 3-kinases themselves. My group has a longstanding, active collaboration with a consortium of 5 international companies to accelerate their drug discovery endeavours in this field.

Technical Summary

Class I phosphoinositide 3-kinases (PI 3-kinases) synthesise phosphatidylinositol 3,4,5-trisphosphate (PtdInsP3), a lipid second messenger which controls cell growth, motility, survival and metabolism in response to a wide variety of hormonal and environmental cues. Defects in this regulatory system contribute to the pathology of important human diseases including many forms of cancer, type II diabetes and inflammatory disorders. The integrity of PI 3-kinase signalling depends upon the correct temporal and spatial distribution of inositol lipid second messengers determined by the regulation and targeting of Class I PI 3-kinases and PtdInsP3 phosphatases. Moreover, the metabolism of PtdInsP3 serves at least two distinct purposes. Firstly, the termination of PtdInsP3 signalling via removal of the 3-phosphate by the tumour suppressor phosphatase, PTEN. Secondly, the co-ordinated synthesis of additional lipid signals, including phosphatidylinositol 3,4-bisphosphate (PtdIns(3,4)P2) and phosphatidylinositol 3-phosphate via removal of the 5-phosphate by one or more members of the phosphoinositide 5-phosphatase family such as SHIP and SHIP2. Whilst the functions and molecular targets of PtdInsP3 are well understood, the relative contributions of PtdInsP3 and/or PtdIns(3,4)P2 to PI 3-kinase signalling and which, if any of these functions are driven primarily by PtdIns(3,4)P2 remain ill-defined. We aim to develop our understanding of the distinct biological functions of PtdInsP3 metabolism under four thematic headings. (i) How do cells exert independent temporal and spatial control of PtdInsP3 versus PtdIns(3,4)P2 when the latter is a metabolic product of the former? (ii) The identification, characterisation and biological functions of PtdIns(3,4)P2 effector proteins. (iii) The roles of PTEN and SHIP2 in the development and maintenance of cell polarity in epithelial cells and other relevant models. How does loss of PTEN or oncogenic mutation of p110alpha affect PtdInsP3 distribution and cell polarity? (iv) What, if any, is the role of PTEN s specificity for both PtdInsP3 and phosphopeptide substrates in mediating its tumour suppressor function? Our approach is multidisciplinary, involving molecular characterisation of expressed and purified components; studies of endogenous and exogenously expressed components in cell lines, 3D epithelial cell cultures and other model systems; and the generation of unique mouse models arising from original discoveries in our laboratory concerning substrate recognition by PTEN. We will continue to collaborate with major pharmaceutical companies via the Division of Signal Transduction Therapy of which CPD is a co-director and which aims to accelerate drug discovery in the area of protein and lipid kinases and phosphatases.

Publications

10 25 50
publication icon
Álvarez-Garcia V (2019) Mechanisms of PTEN loss in cancer: It's all about diversity. in Seminars in cancer biology

publication icon
Wise HM (2017) Prostate cancer, PI3K, PTEN and prognosis. in Clinical science (London, England : 1979)

publication icon
Van Diepen MT (2009) MyosinV controls PTEN function and neuronal cell size. in Nature cell biology

publication icon
Spinelli L (2015) PTEN inhibitors: an evaluation of current compounds. in Advances in biological regulation

publication icon
Spinelli L (2015) Assaying PTEN catalysis in vitro in Methods

publication icon
Spinelli L (2016) Assays to Measure PTEN Lipid Phosphatase Activity In Vitro from Purified Enzyme or Immunoprecipitates. in Methods in molecular biology (Clifton, N.J.)

publication icon
Rodríguez-Escudero I (2015) Yeast-based methods to assess PTEN phosphoinositide phosphatase activity in vivo. in Methods (San Diego, Calif.)

publication icon
Pozuelo-Rubio M (2010) Mechanism of activation of PKB/Akt by the protein phosphatase inhibitor Calyculin A. in Cell biochemistry and biophysics

publication icon
Moult PR (2010) Leptin regulates AMPA receptor trafficking via PTEN inhibition. in The Journal of neuroscience : the official journal of the Society for Neuroscience

publication icon
Leslie NR (2016) Inherited PTEN mutations and the prediction of phenotype. in Seminars in cell & developmental biology

publication icon
Leslie NR (2011) Non-genomic loss of PTEN function in cancer: not in my genes. in Trends in pharmacological sciences

publication icon
Leslie NR (2012) Distinct inactivation of PI3K signalling by PTEN and 5-phosphatases. in Advances in biological regulation

publication icon
Leslie NR (2013) Cell biology. Where is PTEN? in Science (New York, N.Y.)

 
Description Scottish government brain tumour chamber debate and policy review committee
Geographic Reach Local/Municipal/Regional 
Policy Influence Type Participation in a national consultation
 
Description Heriot Watt University 'Theme Studentship'
Amount £105,000 (GBP)
Organisation Heriot-Watt University 
Sector Academic/University
Country United Kingdom
Start 01/2013 
End 08/2017
 
Description James Watt Studentship
Amount £65,000 (GBP)
Organisation Heriot-Watt University 
Sector Academic/University
Country United Kingdom
Start 01/2013 
End 12/2015
 
Description MRS 4yr Studentship
Amount £112,439 (GBP)
Funding ID PhD-1034-2106 
Organisation Medical Research Scotland 
Sector Charity/Non Profit
Country United Kingdom
Start 09/2017 
End 08/2021
 
Description New Ideas Award
Amount £66,924 (GBP)
Funding ID GN-000344 
Organisation The Brain Tumour Charity 
Sector Charity/Non Profit
Country United Kingdom
Start 10/2016 
End 10/2017
 
Description PrCa UK Project Grant
Amount £300,626 (GBP)
Funding ID PG14-006 
Organisation Prostate Cancer UK 
Sector Charity/Non Profit
Country United Kingdom
Start 08/2015 
End 07/2018
 
Title Antibodies 
Description Purified polyclonal antibodies produced during research programme 
Type Of Material Antibody 
Year Produced 2006 
Provided To Others? Yes  
Impact Research publications and progress 
 
Title Expression construct collection 
Description An extensive range of expression constructs (plasmids and viral vectors) for use in several systems. 
Type Of Material Technology assay or reagent 
Year Produced 2006 
Provided To Others? Yes  
Impact We frequently supply other academic research groups with expression constructs (approx 10-15 times per year). Some of these interactions take the forms of collaborations, others simply acknowledged reagent supply. These reagents have been used in many subsequent published projects. 
 
Title Lipid Kinase selectivity screen 
Description This is a service provided to our pharmaceutical collaborators and the broader academic community supporting drug discovery efforts in the area of PI3K. It is the first of its kind developed within an academic setting. 
Type Of Material Technology assay or reagent 
Year Produced 2009 
Provided To Others? Yes  
Impact Supports intensive research in 5 global pharmaceutical companies developing novel cancer treatments targeting the PI 3-kinase pathway. Will develop a large data matrix relating compound structures to efficacy against a large number of lipid kinase enzymes. 
URL http://www.kinase-screen.mrc.ac.uk/kinase-lipid-kinase-panel
 
Title Method of PIK3CA mutation detection by standard Q-PCR 
Description We describe a method using standard SYBR-Green Q-PCR to detect PIK3CA H1047R and E545K mutations. These are the two most common specific mutations in breast cancer and their detection has previously required more complex, expensive approaches. Described in Alvarez-Garcia et al, Sci Rep 2018 
Type Of Material Technology assay or reagent 
Year Produced 2018 
Provided To Others? Yes  
Impact Not yet clear 
URL http://www.nature.com/articles/s41598-018-22473-9
 
Description Acadamic collaboration - Rick Randall 
Organisation University of St Andrews
Department Centre for Biomolecular Sciences
Country United Kingdom 
Sector Academic/University 
PI Contribution Sharing data and analysing samples
Collaborator Contribution Sharing unpublished data
Impact Publications: PM ID: 18029356 and 20133840
Start Year 2006
 
Description Academic collaboration - Britta Eickholt 
Organisation Charité - University of Medicine Berlin
Country Germany 
Sector Academic/University 
PI Contribution Extensive academic collaboration, involving reagent exchange, sharing ideas and data. One postdoctoral researcher worked in the Eickholt lab in Berlin for 2 months in 2013.
Collaborator Contribution The Eickholt lab have provided reagents and shared unpublished data to allow collaborative projects. They also hosted an NRL lab postdoctoral researcher working in the Eickholt lab in Berlin for 2 months in 2013
Impact 3 research Publications: PubMed IDs: 19767745, 23940795, 23085752
 
Description Academic collaboration - Britta Eickholt 
Organisation King's College London
Department MRC Centre for Developmental Neurobiology
Country United Kingdom 
Sector Academic/University 
PI Contribution Extensive academic collaboration, involving reagent exchange, sharing ideas and data. One postdoctoral researcher worked in the Eickholt lab in Berlin for 2 months in 2013.
Collaborator Contribution The Eickholt lab have provided reagents and shared unpublished data to allow collaborative projects. They also hosted an NRL lab postdoctoral researcher working in the Eickholt lab in Berlin for 2 months in 2013
Impact 3 research Publications: PubMed IDs: 19767745, 23940795, 23085752
 
Description Academic collaboration - Doreen Cantrell 
Organisation University of Dundee
Department College of Life Sciences
Country United Kingdom 
Sector Academic/University 
PI Contribution 3 transgenic mouse lines with mutations at the Pten locus. Expertise in the analysis of Pten function. One of the postdocs working part-time on the project.
Collaborator Contribution Expertise in T-cell biology and oncogenesis2 .Transgenic mouse lines required for T-cell specific manipulation of Pten. One postdoc working part-time on the project.
Impact Unpublished data
Start Year 2012
 
Description Academic collaboration - Grahame Hardie 
Organisation University of Dundee
Department College of Life Sciences
Country United Kingdom 
Sector Academic/University 
PI Contribution Expertise in manipulating cellular PI3K/PTEN signalling. Cell lines and reagents.
Collaborator Contribution Expertise and reagents to study the regulation of AMP-Kinase. Most of the researcher time committed to this project.
Impact Co-authored research paper positively reviewed at the Biochemical journal and currently being revised.
Start Year 2012
 
Description Academic collaboration - Rory Duncan 
Organisation Heriot-Watt University
Department School of Engineering & Physical Sciences
Country United Kingdom 
Sector Academic/University 
PI Contribution Two PhD students are being co-supervised by myself and Prof Duncan. One is principally supervised by me and funded by a Heriot Watt University studentship. The other student is funded by a large collaborative MRC grant (ESRIC) and is executing a collaboration between my lab and those of Prof Duncan (HWU) and Ted Hupp at the Edinburgh University Cancer Centre. My lab provides research expertise in cancer biology and signal transduction.
Collaborator Contribution Prof Duncan provides expertise in super-resolution imaging and the use of fluorescent probes to investigate molecular interactions in living and fixed cells.
Impact Funding for 2 studentships, both started in 2013
Start Year 2013
 
Description Academic collaboration - Seth Schorr 
Organisation University of Dundee
Department College of Medicine, Dentistry & Nursing
Country United Kingdom 
Sector Academic/University 
PI Contribution Project instigation and direction. Some experiments. Writing and over-seeing publication
Collaborator Contribution Extensive experimnetal collaboration, significant scientific dialog, and experiments mostly performed by rsesearchers funded from collaborators lab.
Impact Publication: Ellis et al, PMID: 20600851
Start Year 2007
 
Description Commercial service collaboration - Novartis 
Organisation Novartis
Country Global 
Sector Private 
PI Contribution We tested several compounds for their ability to inhibit PTEN in vitro
Collaborator Contribution It motivated us to test several control compounds as inhibitors of the tumour suppressor phosphatase PTEN
Impact We obtained scientifically valuable data from several control compounds and supplied data to Novartis regarding their test compounds.
Start Year 2011
 
Description DSTT Pharameutical Consortium 
Organisation Dundee Signal Transduction Therapy (DSTT) Consortium
Country United Kingdom 
Sector Academic/University 
PI Contribution The collaboration is funded by a pharmaceutical consortium, supporting several activities including a research Division sited in the University of Dundee College of Life Sciences. From 1998-2012 this was overseen by the co-directors, Peter Downes, the original PI on this grant and Professors Philip Cohen and Dario Alessi. Dr Leslie and Prof Downes and their research groups also provided advice and expertise to the consortium companies. Since the last renewal of the collaboration in 2012, the grant holders have not been formally involved but occasionally act as informal consultants to the consortium companies, the last instance being in October 2013.
Collaborator Contribution We have had occasional access to novel compounds from these pharmaceutical collaborators for use as experimental enzyme inhibitors in our research. Extensive interactions with the pharmaceutical collaborators has also been effectiove in broadening the translational training of the research team.
Impact The collaboration has supported the drug discovery efforts of 6 major global pharmaceutical companies in the field of PI 3-Kinase signalling, an extrememly active area, especially for cancer. The collaboration has supported 2 or 3 research posts in the core CPD/NRL labs since its inception in 1998 contributing to many of the CPD lab publications over this time. It has also funded a level of around ten active laboratory positions since 1998, with these trained staff going into further academic and commercial positions.
 
Description Paul Brennan (University of Edinburgh) Collaboration 
Organisation University of Edinburgh
Department Institute of Evolutionary Biology
Country United Kingdom 
Sector Academic/University 
PI Contribution Leading projects to investigate circulating tumour DNA from glioma patients (CSO funded) and to 3D print brain tumour constructs (Brain Tumour Charity and MRS funded)
Collaborator Contribution Provision of clinical material and advice regarding clinical aspects of brain tumour pathology and management
Impact Multi-disciplinary research teams combine Leslie - Cancer Biology Brennan - Neurosurgeon Kersaudy Kerhoas - Microfluidic Bioengineer Shu - 3D biofabrication Bioengineer
Start Year 2015
 
Description Renishaw 3D Bioprinting Collaboration 
Organisation Renishaw PLC
Country United Kingdom 
Sector Private 
PI Contribution We have been testing and helping Renishaw to develop 3D bioprinting hardware
Collaborator Contribution Renishaw have provided us with a prototype 3D bioprinter which performs at least as will as any commercially available instrument we are aware of. Estimated value £100K.
Impact Collaborative MRS 4 yr studentship (£112 439)
Start Year 2016
 
Title Lipid kinase selectivity screen 
Description The development of a lipid kinase selectivity screen is not strictly a discovery, but it requires a high level of expertise and reputation in the lipid signalling field as well as the resources required to generate large quantities of many enzymes grouped together under a single assay platform. We also have 'first mover' advantage in terms of the marketability of the screen and existing collaborators in the form of the DSTT member pharmaceutical companies. 
IP Reference  
Protection Protection not required
Year Protection Granted 2008
Licensed No
Impact A screen which supports and drives PI 3-kinase pathway drug discovery within 5 major pharmaceutical companies.
 
Description AICR Open Day 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact Public Open Day organised by Association for Cancer Research

Raising awareness of research. Donations to medical resaerch
Year(s) Of Engagement Activity 2010
 
Description College Open Day 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact Several group members presenting science at College Open Day

High attendence (around 300)
Year(s) Of Engagement Activity 2010,2011,2012
 
Description Press and media coverage 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Media (as a channel to the public)
Results and Impact Peter Downes and Nick Leslie have been interviewed by press and television to discuss individual research publications.

MRC funded work in the CPD/NRL laboratory is also of great interest to the pharmaceutical industry and underpined the ground breaking collaboration between Dundee scientists and five leading pharmaceutical companies known as the Division of Signal Transduction Therapy (DSTT). Peter Downes was a founding co-director of DSTT which has attracted much external interest 1998-2012. During the period of support we have hosted at least eight visits from members of the international press and other publishing activities, Scottish and international development agencies and industry.

Elevated profile of MRC funded research and its translation to drug discovery through collaboration with global pharmaceutical companies. The PI3K pathway which is the subject of this funding and which was co-discovered by Peter Downes, is one of the most fertile areas of mechanism based drug dicovery, especially relating to human cancer.
Year(s) Of Engagement Activity 2006,2007,2008,2009,2010,2011,2012,2013
 
Description Prostate Cancer UK Support Group Day Glasgow April 2017 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Supporters
Results and Impact Prostate Cancer UK Support Group Day Glasgow April 2017
Year(s) Of Engagement Activity 2017
 
Description School and early undergraduate project placements 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? Yes
Geographic Reach National
Primary Audience Schools
Results and Impact We regularly host laboratory placements for school and early undergraduate students to experience research

To my knowledge, all of these students have entered University scientific degree programmes or in the case of relevant undregraduates continued a research/scientific career
Year(s) Of Engagement Activity 2006,2007,2008,2009,2010,2011,2013,2014
 
Description TV and radio interviews (Brain Tumour Charity) 
Form Of Engagement Activity A broadcast e.g. TV/radio/film/podcast (other than news/press)
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact Interviews broadcast by BBC Radio and ITV TV news discussing 3D bioprinting research
Year(s) Of Engagement Activity 2016