Characterisation of the signalling pathways involved in cell invasion downstream of extra centrosomes
Lead Research Organisation:
Queen Mary University of London
Department Name: Barts Cancer Institute
Abstract
Despite the progress, cancer is still a serious challenge in medicine. Identifying the differences between tumour and normal cells is imperative to understand the disease and to develop novel selective therapeutic strategies. The presence of extra copies of a cellular organelle called centrosome is a unique feature of tumours. Indeed, the presence of extra centrosomes in tumour cells has been recognized since the late 1800s and was then proposed to be cause of malignancy. Yet, to date how centrosome amplification contributes to tumour progression is still unclear. Our recent work demonstrates that the presence of extra centrosomes can induce cell invasion in cells similarly to overexpression of known oncogenes. This finding highlights the importance that these organelles play in tumour progression. In addition, since cell invasion is one of the first steps required for metastatic dissemination of tumour cells, responsible for 90% of human cancer deaths, understanding this process is crucial.
In this research proposal we aim to determine how centrosome amplification induces cell invasion. We will use a combination of approaches to identify the signalling alterations that occur as a result of centrosome amplification. Next, we will investigate the signalling alterations that are important for invasion using sophisticated 3-D cell culture models that closely mimic the real tissues. Our results will be also validated in human breast cancer samples from patients. This is essential to establish the relevance of our findings to cancer. Importantly, this strategy has the potential for the development of novel biomarkers to identify tumours containing extra centrosomes in the clinic. This is important because we previously showed that cells with extra centrosomes have unique survival requirements and can be selectively killed. Based on our previous observations, new drugs that kill cells with extra centrosomes have been recently developed. Thus, our work will not only contribute to understand the role of centrosomes amplification in tumour progression and invasion but could also have a major impact in the clinic.
In this research proposal we aim to determine how centrosome amplification induces cell invasion. We will use a combination of approaches to identify the signalling alterations that occur as a result of centrosome amplification. Next, we will investigate the signalling alterations that are important for invasion using sophisticated 3-D cell culture models that closely mimic the real tissues. Our results will be also validated in human breast cancer samples from patients. This is essential to establish the relevance of our findings to cancer. Importantly, this strategy has the potential for the development of novel biomarkers to identify tumours containing extra centrosomes in the clinic. This is important because we previously showed that cells with extra centrosomes have unique survival requirements and can be selectively killed. Based on our previous observations, new drugs that kill cells with extra centrosomes have been recently developed. Thus, our work will not only contribute to understand the role of centrosomes amplification in tumour progression and invasion but could also have a major impact in the clinic.
Technical Summary
Centrosome abnormalities, and in particular centrosome amplification, are common features of human tumours. However, to date the contribution of extra centrosomes to tumour progression is still unclear. We recently found that centrosome amplification can induce invasion in non-transformed human cells, similarly to the overexpression of known oncogenes, such as ERBB2. This is in part due to RAC1 activation as a result of increased microtubule nucleation downstream of extra centrosomes. However, our preliminary data suggests that RAC1 activation alone is not sufficient to induce a robust invasive phenotype and that other signalling pathways play a role in this process. In this study, we aim to characterise the signalling alterations that are induced by centrosome amplification. To do so we will preform phosphoproteomics to identify altered phospho-sites in cells with extra centrosomes. Next, using 3D cell culture models, we will determine the signalling alterations that are important for the invasive phenotype, which will further clarify the mechanisms behind these striking morphological changes. Finally, we will test whether our findings can be recapitulated in human breast tumours using breast cancer tissue samples. This strategy has the potential for the development of biomarkers to identify tumours with extra centrosomes that would be sensitive to drugs that specifically target these cells (e.g. HSET inhibitors). We anticipate that the experiments proposed here will contribute to our understanding of how centrosome amplification impacts tumour progression. Importantly, these studies will open new areas of investigation in my laboratory regarding not only the role of centrosome amplification in cancer but also how the centrosome regulate intracellular signalling in normal cells.
Planned Impact
Centrosome amplification is a common feature of human tumours. However, the contribution of extra centrosomes to tumour progression is still unclear. Our recent work uncovered a novel role for centrosome amplification in promoting cell invasion and the proposed project aims to identify the signalling pathways that promote invasion downstream of extra centrosomes.
- We predict that this project will generate major interest in the field of centrosomes and cancer. By dissecting the signalling alterations that occurs in cells with extra centrosomes we will contribute to the understanding of how centrosome amplification impacts tumour progression.
- This project also has the potential to uncover novel signalling pathways that are regulated by the centrosome, an aspect poorly studied. thus it will benefit the scientific community
- In addition to the scientific community, our project will also impact the wider community through the public engagement program at Barts Cancer Institute. Our press office to the local and international media will advertise the work generated in this study.
- Our laboratory participates in lab tours for the public that provides another opportunity to engage with the general public.
We previously found that cells with extra centrosomes have unique requirements for survival, including the kinesin HSET/KIFC1. Because centrosome amplification is a common feature of human tumours, these findings open a new venue for the development of selective cancer therapies. Supporting this, two independent inhibitors against HSET were recently developed. However, the use these drugs in the clinic requires an efficient method to identify tumours with extra centrosomes.
- The work described in this proposal has the potential to develop biomarkers to identify tumours with extra centrosomes. We will investigate the signalling pathways that are altered in cells with supernumerary centrosomes in human breast cancer tissue samples to test if these alterations can predict which tumours have higher percentage of centrosome amplification.
- If we succeed, our work can have a major impact in the clinic. We will develop early clinical trials with the help of the Experimental Cancer Medicine Centre at the Barts Cancer Institute. in this case, our work will have the potential to benefit cancer patients by providing an alternative therapeutic strategy to specifically kill cells with supernumerary centrosomes
- We predict that this project will generate major interest in the field of centrosomes and cancer. By dissecting the signalling alterations that occurs in cells with extra centrosomes we will contribute to the understanding of how centrosome amplification impacts tumour progression.
- This project also has the potential to uncover novel signalling pathways that are regulated by the centrosome, an aspect poorly studied. thus it will benefit the scientific community
- In addition to the scientific community, our project will also impact the wider community through the public engagement program at Barts Cancer Institute. Our press office to the local and international media will advertise the work generated in this study.
- Our laboratory participates in lab tours for the public that provides another opportunity to engage with the general public.
We previously found that cells with extra centrosomes have unique requirements for survival, including the kinesin HSET/KIFC1. Because centrosome amplification is a common feature of human tumours, these findings open a new venue for the development of selective cancer therapies. Supporting this, two independent inhibitors against HSET were recently developed. However, the use these drugs in the clinic requires an efficient method to identify tumours with extra centrosomes.
- The work described in this proposal has the potential to develop biomarkers to identify tumours with extra centrosomes. We will investigate the signalling pathways that are altered in cells with supernumerary centrosomes in human breast cancer tissue samples to test if these alterations can predict which tumours have higher percentage of centrosome amplification.
- If we succeed, our work can have a major impact in the clinic. We will develop early clinical trials with the help of the Experimental Cancer Medicine Centre at the Barts Cancer Institute. in this case, our work will have the potential to benefit cancer patients by providing an alternative therapeutic strategy to specifically kill cells with supernumerary centrosomes
People |
ORCID iD |
Susana Godinho (Principal Investigator) |
Publications
Arnandis T
(2018)
Oxidative Stress in Cells with Extra Centrosomes Drives Non-Cell-Autonomous Invasion.
in Developmental cell
Arnandis T
(2015)
Studying centrosome function using three-dimensional cell cultures.
in Methods in cell biology
Godinho S
(2019)
The principles of spindle bipolarity
in Nature Reviews Molecular Cell Biology
Monteiro P
(2023)
Centrosome amplification fine tunes tubulin acetylation to differentially control intracellular organization.
in The EMBO journal
Monteiro P
(2018)
Structural Centrosomal Abnormalities Push Cells toward Invasion.
in Developmental cell
Rhys AD
(2018)
Loss of E-cadherin provides tolerance to centrosome amplification in epithelial cancer cells.
in The Journal of cell biology
Description | Dissecting the role of extra centrosomes-induced exosome secretion in PDAC microenvironment |
Amount | £477,195 (GBP) |
Funding ID | MR/T000538/1 |
Organisation | Medical Research Council (MRC) |
Sector | Public |
Country | United Kingdom |
Start | 12/2019 |
End | 11/2022 |
Description | Investigating the impact of amplified centrosomes in cancer |
Amount | £1,489,237 (GBP) |
Funding ID | DCRPGF\100013 |
Organisation | Cancer Research UK |
Sector | Charity/Non Profit |
Country | United Kingdom |
Start | 06/2021 |
End | 05/2027 |
Description | MRC QMUL PhD studentship |
Amount | £84,000 (GBP) |
Funding ID | MIM1030B |
Organisation | Medical Research Council (MRC) |
Sector | Public |
Country | United Kingdom |
Start | 10/2015 |
End | 10/2018 |
Description | Research Prize |
Amount | £200,000 (GBP) |
Funding ID | MIMG1L2R |
Organisation | Lister Institute of Preventive Medicine |
Sector | Charity/Non Profit |
Country | United Kingdom |
Start | 10/2016 |
End | 09/2019 |
Description | Royal Society Research Grant |
Amount | £15,000 (GBP) |
Funding ID | MIMH1A1R |
Organisation | The Royal Society |
Sector | Charity/Non Profit |
Country | United Kingdom |
Start | 03/2016 |
End | 03/2017 |
Description | Understanding exosome-mediated fibrosis to develop novel therapies for PDAC |
Amount | £200,000 (GBP) |
Organisation | Cancer Research UK |
Sector | Charity/Non Profit |
Country | United Kingdom |
Start | 10/2019 |
End | 09/2022 |
Description | WHRI-ACADEMY International Fellowship Programme |
Amount | € 74,500 (EUR) |
Organisation | William Harvey International Translational Research (WHRI) Academy |
Sector | Public |
Country | United Kingdom |
Start | 05/2016 |
End | 05/2018 |
Title | 3D mammary gland organoids |
Description | we established in our lab a 3D culture method to grow organoids obtained from mouse mammary glands. briefly mammary glands are extracted from mouse females and shopped into small pieces which are then cultures in a gel containing matrigel and collagen. this will allow us to measure invasion in a more physiological manner. |
Type Of Material | Technology assay or reagent |
Provided To Others? | No |
Impact | this provides a great system that mimics the mammary gland architecture while reducing the number of mice. |
Title | CDH1 CRISPR HaCAT |
Description | HaCAT cell line knockdown of E-cadherin (CDH1) using CRISPR-Cas9 technology |
Type Of Material | Cell line |
Provided To Others? | No |
Impact | this cell line allowed us to determine that loss of E-cadherin contribute to the survival of cells carrying extra centrosomes |
Title | CDH1 CRISPR MCF10A |
Description | MCF10A cells knockdown for E-cadherin (CDH1) using CRISPR-Cas9 technology |
Type Of Material | Cell line |
Year Produced | 2015 |
Provided To Others? | Yes |
Impact | This cell line allow us to determine that cells carrying extra centrosomes survive better in the absence of E-cadherin |
Title | Generation of a new construct to induce centrosome amplification and label cells with RFP |
Description | this new construct allows the induction of PLK4 overexpression in the presence of doxycycline. at the same time, it contains another cassette that enables the expression of RFP upon PLK4 overexpression to label the cells. this construct was developed to generate a mouse model that enables tracking of cells with extra centrosomes |
Type Of Material | Model of mechanisms or symptoms - mammalian in vivo |
Provided To Others? | No |
Impact | this construct was developed to generate a mouse model that enables tracking of cells with extra centrosomes that can be used to study invasion induced by centrosome amplification in vivo and also to test drugs that can selectively target cancer cells to develop novel therapies. |
Title | Micropatterning technology |
Description | We recently purchase the equipment necessary to implement the micropatterning technique in our laboratory. this will allow us to standardise cell morphology for specific assays, look at cell migration and cell-cell adhesion. |
Type Of Material | Technology assay or reagent |
Year Produced | 2006 |
Provided To Others? | Yes |
Impact | we just implemented the method and we envision that this will not only help our experiments but also of other colleagues that want to take advantage of this technology. |
Title | mouse mammary organoids to study cell invaion |
Description | we established in the laboratory a 3D culture of mouse mammary organoids. these mouse mammary organoids are made from mammary tissue removed from the mouse and function as an in vitro model of primary cells. |
Type Of Material | Model of mechanisms or symptoms - in vitro |
Provided To Others? | No |
Impact | this methodology was established by other and we implemented in our laboratory to study cell invasion. we can manipulate these organoids in 3D and this will enable us to use less animals o study cell invasion in a more physiological relevant model |
Title | zebrafish injections |
Description | we learned how to inject human cells into zebrafish to quantify invasion in vivo. |
Type Of Material | Technology assay or reagent |
Provided To Others? | No |
Impact | this method works very well and is in accordance with the 3Rs, since it can substitute mouse models to assess invaion |
Description | Bioinformatics analyses |
Organisation | Queen Mary University of London |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | We provided a list of annotated human breast cancer cell lines with various degrees of centrosome amplification and a list of genes we identified as being important for invasion |
Collaborator Contribution | Our collaborators at Barts cancer Institute, Claude Chelala and her team, provided bioinformatic analyses of our genes of interest, identified in this study in a panel of human breast cancer cell lines. |
Impact | this collaboration resulted in a publication in Developmental Cell in 2018 |
Start Year | 2017 |
Description | Develop of 3D in vitro systems to study cell invasion |
Organisation | University College London |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | we have provided the expertise and intellectual input as well as the cell lines to study invasion |
Collaborator Contribution | our collaborator Dr Guillaume Charras has provided us with expertise and microfabricatted devices to study invasion |
Impact | this is a mutidisciplinary collaboration and involves physics and biology. |
Start Year | 2016 |
Description | Generation of novel mouse models to study centrosome amplification |
Organisation | Francis Crick Institute |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | intellectual input, expertise and project develoment |
Collaborator Contribution | Dr Dinis Calado has expertise in genetic engineered mouse models, development of construct to generate a novel mouse for centrosome amplification and mouse generation. |
Impact | we are in the process of developing a mouse model that can be used to assess the in vivo relevance of our findings. we predict that this model will also be useful for other projects, namely testing drugs that can kill cancer cells with extra centrosomes |
Start Year | 2016 |
Description | Proteomics analysis of signalling alterations in cells with extra centrosomes |
Organisation | Queen Mary University of London |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | my team designed the experiment to analyse alterations of the phosphorylated proteins in cells upon induction of centrosome amplification. We also optimise the experiment and performed the collection of extracts to be analysed by mass spectrometry. we further analysed and characterised using bioinformatics the proteins identified by mass spectrometry. |
Collaborator Contribution | our collaborator contributed with discussions to the design of the experiment and performed the proteomics analysis of the samples by mass spectrometry. initial statistical analysis and bioinformatics were performed by our collaborator as well. |
Impact | - identification of the proteins whose phosphorylation is increased or decreased in upon induction of centrosome amplification at different time points (0, 24, 48 and 60 hours) - comprehensive list of altered signalling pathways in cells with extra centrosomes this collaboration resulted in a publication in Developmental Cell in 2018 |
Start Year | 2015 |
Description | Zebrafish model to study invasion |
Organisation | Queen Mary University of London |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | performing the experiments, designing the experiments |
Collaborator Contribution | training staff from my lab on zebrafish injections, advice on experimental design |
Impact | learnt how to inject human cells into zebrafish to assess invasion this collaboration resulted in a publication in Developmental Cell in 2018 |
Start Year | 2015 |
Description | mouse mammary organoids |
Organisation | Francis Crick Institute |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | We used mouse mammary organoids to demonstrate the importance of paracrine signalling mediated by centrosome amplification to induce invasion |
Collaborator Contribution | Our collaborator, Ilaria Malanchi at the Francis Crick Institute, provided us with expertise and biological material to do organoid experiments. Specifically, we obtained CXCR2 knockout mammary glands from her lab which allowed us to test the importance of IL8 in paracrine invasion. In addition, we obtained also primary cells from PyMT tumours purified from Ilaria's lab that we grew in 3D culture. |
Impact | this collaboration resulted in a publication in Developmental Cell in 2018 |
Start Year | 2017 |
Description | CRUK tour |
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 | Supporters |
Results and Impact | At Barts Cancer Institute we have organised tours to allow patients, their families and donors to visit us an learn about cancer and our work. My goal was to provide the scientific talk that covered the basics of what is cancer, what we do at the institute and some aspects of my own research. As usual, the audience was very engaged and wanted to learn more about funding and novel treatments. a a fundamental researcher myself, i always make an effort to explain why we need basic research to understand cancer and develop novel treatments. The feedback i always get from the audience is very positive, with some reporting a better understanding of the disease and also a better understanding of why finding cures is not trivial. They become more aware of timelines and obstacles one needs to overcome o develop new treatments, which i think is important. |
Year(s) Of Engagement Activity | 2018 |
Description | Lab Tour fro CRUK donors |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | Regional |
Primary Audience | Public/other audiences |
Results and Impact | this activity has as main goal to engage CRUK supporter/donors with our institute. as part of the tour, i give a talk about cancer and my work. |
Year(s) Of Engagement Activity | 2016 |
Description | LabTour |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | Local |
Primary Audience | Supporters |
Results and Impact | Labtours are organised by CRUK and aim at explaining the research carried out at our institution to the patients/donors. it is a very rewarding experience as the audience is very engaging |
Year(s) Of Engagement Activity | 2014,2015 |
Description | STEM ambassador |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | Local |
Primary Audience | Schools |
Results and Impact | I am a certified STEM ambassador and as such I participate in tours to local school kids between 6-12 years old. in particular I participate at the Centre of the Cell, part of our school of medicine at QMUL. in these events, i descried my work the students and answer any quetsions they might have. in addition, i stay with them inside the Pod and help them with various games and activities where they learn about cells and research. |
Year(s) Of Engagement Activity | 2018 |