Tissue Microenvironment Group

We are based within the Division of Cancer and Genetics within the School of Medicine and comprise researchers with specialist interests and expertise in immunology, stromal fibroblastic cells, extracellular vesicles, hypoxia and imaging.

We are collectively interested in modelling these complex systems and devising new approaches to perturb cancer-influence and restore normality in the microenvironment.

The group interacts and collaborates with other researchers throughout the world, including in the USA, Netherlands, Australia, Japan and others. We also provide a hub to support local colleagues within the University and the NHS, facilitating studies in this area.

Mission and vision

Our mission is to undertake multidisciplinary research to reveal the complexities of tissue microenvironments, lead technological innovation, scholarship and the exchange of knowledge.

Our vision is to engage in an integrative cancer research programme addressing basic and translational questions.

Our focus areas include:

• innate tissue heterogeneity and the multi-scalar effects of therapeutic perturbation on tissue dynamics
• immune signatures in tissue and immune modulation strategies
• extracellular vesicles as modulators of local and distant tissues.

Amcanion

Mission and Vision

Our Mission is to undertake multidisciplinary research to reveal the complexities of tissue microenvironments, lead technological innovation, scholarship and the exchange of knowledge.

Our Vision is to engage in an integrative cancer research programme addressing basic and translational questions.

Our Focus areas include:

• Innate tissue heterogeneity and the multi-scalar effects of therapeutic perturbation on tissue dynamics.
• Immune signatures in tissue and immune modulation strategies.
• Extracellular vesicles as modulators of local and distant tissues.

Projects

Tumour hypoxia innate and acquired from standard of care treatment

Hypoxia is a common feature in many tumours including pancreas and prostate tumours and has been shown to drive disease progression towards metastasis. It is, therefore, a rational therapeutic strategy to directly target hypoxic tumour cells in an attempt to improve treatment.

We have developed pre-clinical prostate models to measure, map and profile how treatment-induced hypoxia can drive prostate tumour growth. Together with our collaborators, we have provided the first evidence that OCT1002, a novel unidirectional hypoxia-activated pro-drug (uHAP), has a hypoxia-dependent anti-tumour effect in androgen-sensitive prostate. The activated drug is fluorescent and provides exciting opportunities for micro-drug kinetic studies to understand single cell responses in heterogeneous tumour populations (in time and space).

Collaborators: Dr Declan Mckenna (University of Ulster), Oncotherics Ltd and Axis Bioservices Ltd.

Encoding and tracking of mesenchymal stem cells – Bone-in-a-Dish

The coordinated and heterogeneous behaviour of cellular populations is one of the principal components that dictate the function and evolution of many biological tissues. The collaboration group involved have a strong background in multi-dimensional imaging, cell tracking technologies and mesenchymal stem cell biology, tissue engineering.

To date, the collaboration group has focussed on the development of numerical strategies to stochastically simulate proliferation characteristics of large cellular populations, and the focus is to develop our unified experimental-metrology, to understand bone stem cell niches in health and disease.

Collaborators: Professor Alastair Sloan and Dr Rachel Howard-Jones (Cardiff University), Dr Rowan M Brown (Swansea University).

ER stress and mTOR signalling in tuberous sclerosis

The current research project utilises TSC tumour cell line models to test the effectiveness of novel anti-cancer drugs that could easily be repositioned to treat Tuberous Sclerosis (TS), patients. The project focuses on determining whether drug combinations with nelfinavir could be effective at treating TS patients as well as cancer patients that have tumours displaying aberrant mTOR signalling and ER stress. With the Errington lab and led by Andrew Tee (School of Medicine) we are co-investigating ‘Novel Therapies That Selectively Kill Mammalian Target of Rapamycin (mTOR)-addicted Cancer Cells’ which is funded by Cancer Research Wales.

A key mediator of cell growth in many cancers is through aberrant signal transduction through mammalian target of rapamycin (mTOR). The Tee laboratory has a long-standing research history on dissecting novel mechanisms of mTOR signalling, autophagy and endoplasmic reticulum (ER) stress and then relating this to cancer progression and therapy. The Errington laboratory has a track record in single-cell imaging of ER-stress and apoptosis

leading to the development of a mathematical model to track single cell calcium (Ca(2+)) dynamics to predict the consequences of over-expression of endoplasmic reticulum-located chaperones.

Collaborators: Dr Andrew Tee (Cardiff University)

Images-to-numbers: bringing rigour through measurement science

An important consideration in developing measurement standards or assurance appropriate with the regulations that govern the production and use of commercial cell therapy products is the development of robust and reliable measurements to monitor the quality of such products. Therefore, the joint long term goal in our case is to design an analysis framework for ensuring the reproducibility of cell-based metrology using imaging approaches (fit for translational imaging) that are necessary for preclinical drug screening, or patient-derived cell evaluation/screening or indeed the production of cell therapeutics.

The purpose of this work is to incorporate and formalise manufacturing standards into a Cardiff-derived cell-based measurement process. A critical component of this is to understand the National Institute of Standards and Technology (NIST) requirements of cause and effect elements which define variability in control processes, and the sensitivity of process outputs. There are many contributing factors which influence both the confidence and robustness of such an undertaking such as the completeness of experimental description (simply termed curation), with associated traceability (including reference materials and different layers of validation) and finally incorporating uncertainty analysis with multi-user/laboratory -and instrument differences.

We focus on the manufacture of a ‘multiwell imaging plate’ that addresses the metrology pipelines from two perspectives:

• the ability to isolate, image and measure a mesenchymal stem cell (from human tissue) and
• to establish a drug screening plate for the detection of cellular phenotypes using ‘tumour cell painting’.

The Cardiff ProtocolNavigator challenge, in particular, is to capture the audit trail through curation, process design traceability and incorporation of imaging standards and to finally output a cause and effect analysis. The clinical application of this measurement science can be best communicated through the quote of LP Freedman “The development of novel therapeutics depends and builds upon the validity and reproducibility of previously published data and findings.

Yet irreproducibility is pervasive in preclinical life science research and can be traced to cumulative errors or flaws in several areas, including reference materials, study design, laboratory protocols, and data collection and analysis. The expanded development and use of consensus-based standards and well-documented best practices are needed to both enhance reproducibility and drive therapeutic innovations.”

Thus there are clear challenges in achieving reproducibility, accuracy and comparability in biological assays all identified as major roadblocks to development, manufacturing and regulatory approval of patient-derived cell diagnostics and cell therapy products (CTPs). Furthermore, data scientists (big data scientists) who analyse these experimental data to gain insight usually have little or no understanding about the choices made in defining the study design.

We hypothesise that the analysis of data without the knowledge of the derivation process often leads to misinterpretations and therefore loss of data value. Therefore, from a translational research perspective, there is an imperative that data, as well as the biological product,  needs to be traceable and with context. Making

ProtocolNavigator fit for a translational and manufacturing purpose and fit for addressing reproducibility issues is the joint undertaking.

Collaborators: Drs John Elliott and Michael Halter (National Institute of Standards and Technology). With Professor Alastair Sloan and Dr Rachel Howard-Jones (Cardiff University) and Dr Imtiaz Khan (Cardiff Metropolitan)

Innovation and translation - novel fluorescent probes and sensors

The research in the Errington laboratory is driven by the intellectual drivers to establish systems engineering principles to study dynamic cellular models – which we term a systems cytometry approach; unifying the biology, the technology platform and the data analysis (biomaterial analytics) as inter-linked, dependent elements of the measurement process; with the long term goal of providing robust cellular and organotypic model systems for high-throughput technologies to progress research from bench to bedside.

Like all complex multi-cellular systems the ability to quantify emergent behaviour is dependent upon robust spatiotemporal measurement over relatively significant periods. Developments in microscopy and flow cytometry permit the measurement of >105 individual cells over extended periods allowing cellular interactions, motion and response to a perturbation to be realized.

Cardiff led research has resulted in a number of biophotonic tools that enable time-series single cell tracking, and analysis of cellular phenotype incorporating the context of the micro-environment. One aspect of the multi-disciplinary programme has led to the development of a family of novel far-red fluorescent dyes. The DRAQ™ probes and more recently sensors which are now used in a wide range of laboratory tests, transforming practice in clinical, commercial and research sectors through the spinout Biostatus Ltd.

Collaborators: Dr Klaus Pors (University of Bradford) and the team at Biostatus Ltd.

The mechanism and consequence of vesicle uptake

The interaction of small cancer-derived vesicles with cells in the microenvironment is highly complex and fundamental to our understanding of how vesicles work. However, the details relating to vesicle binding and subsequent uptake of vesicles into the recipient cells remains poorly understood.

The project aims to define the mechanisms and routes of vesicle uptake by stromal-fibroblastic cells. In particular, we are studying the process of endocytosis. We have developed a simple yet efficient method for stable-fluorescent tracking of vesicles allowing us to monitor their interaction with fibroblasts by fluorescence microscopy.

From these studies, we will gain new insights into the molecules and cellular compartments that control how vesicles are dealt-with by recipient cells. From this, we will learn novel approaches to inhibit these interactions. Furthermore, we may learn how to enhance the uptake and delivery of vesicle-cargo as a means of delivering therapeutic agents to stromal or other cell types.

Collaborators: Professor Arwyn T Jones, Dr Pete Watson, Cardiff University

The transit of extracellular vesicles through 3D-matrix

Small vesicles are secreted by a variety of cell types within the cancer microenvironment. Once outside the cell, their interactions with the surrounding extracellular matrix, and with other cellular components are under-investigated, yet these are fundamental aspects to understanding how vesicles function and spread into the circulation, and throughout the body.

This study will develop imaging approaches to track the mobility of vesicles in a three-dimensional space comprising acellular matrices, and thereafter matrices that include cellular components. The requirement for cells for facilitating the mobility, and how this process functions will be addressed. Mathematical approaches will lead to an in silico model defining the major parameters, and the impact of modulating vesicle and cellular phenotypes (eg by drug treatments) will be studied.

The project will provide a completely new and fundamental understanding of vesicles and their behaviours in the 3D environment- and identify mechanisms for their entry to, and subsequent dissemination through the circulation.

Collaborators: Professor Huw Summers, Swansea University.

Control of stromal cells by growth factors, delivered by vesicles

Stromal fibroblasts surrounding cancer cells are often abnormally activated, in which their homeostasis roles are perturbed, and they instead support disease growth and spread. Cancer cell-derived vesicles are important contributors responsible for fibroblast activation, and they do so by delivering Transforming Growth Factor beta-1 (TGFb1). However, stimulation with a single growth factor is not sufficient to generate a tumour-supporting fibroblastic cell, and we hypothesise that a range of other cytokines/growth factors are co-delivered when a vesicle encounters cells.

The investigations will focus on growth-factor binding molecules on the outer surface of vesicles, termed Heparan Sulphate proteoglycans. By manipulating the expression of HSPG-family members, the repertoire of vesicle factors can be modified, and the aggressivity of the responding fibroblasts can be controlled.

These studies will define the complexity of vesicular growth factor delivery, the specific HSPG-members required, and the co-signalling pathways that culminate in tumour-supporting fibroblasts. This study will highlight hitherto untested therapeutic targets for halting stromal involvement in prostate and potentially other solid cancers.

Collaborators: Professor Guido Jenster (Erasmus MC; Rotterdam).

Targeting cancer-stem cells using the immune system

All too often, cancer treatment may provide a temporary effect, and after a period of remission, the disease may sadly return. Whilst we don’t fully understand why this happens, one explanation is that stem cells that give rise to the tumour cells are particularly resistant to our current treatments (such as drugs or radiation treatments), and it is the action of such stem cells which re-populate cancer, allowing the disease to return.

Our approach, therefore, is to manipulate the body’s own immune system, particularly the T lymphocytes. These cells have a remarkable ability to recognise and destroy cells, and this is a normal part of our own defence against virus infections for example. We, however, intend to train T lymphocytes to recognise the abnormal, cancer-generating stem cells. Together with researchers in Japan, we have identified particular fragments of protein molecules (peptides) that are displayed on the surface of cancer stem cells and not present on the non-stem cells.

These fragments, therefore, become a targeting-mechanism and allow T cells to kill and eliminate the cancer stem cells in a highly selective and hopefully safe fashion. The project currently focusses on prostate cancer, but the approach should be amenable to adapt for other cancer types thereafter.

Collaborators: Professor Dr Toshihiko Torigoe and Dr Takayuki Kanaseki (Sapporo Medical University, Japan)