Breast Cancer Prevention Conference Full Abstracts

1. Breast cancer polygenic risk scores derived in White European (WE) populations are not calibrated for women of Ashkenazi Jewish (AJ) descent. 

Eleanor Roberts1, Elke M van Veen2,3, Helen Byers2,3, Ofra Barnett-Griness4, Naomi Gronich4,5, Flavio Lejbkowicz4, Mila Pinchev4, Miriam J. Smith2,3, Anthony Howell1,6,7 , William G. Newman2,3, Emma R. Woodward2,3, Elaine F Harkness 6,8, Adam R Brentnall9, Jack Cuzick9,Gad Rennert4,5, Sacha J Howell1,6,7*, D. Gareth Evans1,2,3,6,7*

1Division of Cancer Sciences, Faculty of Biology, Medicine and Health, University of Manchester, Manchester Academic Health Science Centre, Manchester, UK.
2Manchester Centre for Genomic Medicine, Manchester University Hospitals NHS Foundation Trust, Manchester, UK.
3Division of Evolution, Infection and Genomics, School of Biological Sciences, Faculty of Biology, Medicine and Health, University of Manchester, Manchester Academic Health Science Centre, Manchester, UK.
4Department of Community Medicine and Epidemiology, Carmel Medical Center, Haifa 3436217, Israel.
5The Ruth and Bruce Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa 3200003, Israel.
6Nightingale/Prevent Breast Cancer Centre, Wythenshawe Hospital, Manchester University NHS Foundation Trust, Manchester, UK.
7Manchester Breast Centre, Manchester Cancer Research Centre, The Christie Hospital, Manchester, UK.
8Division of Informatics, Imaging & Data Sciences, School of Health Sciences, Faculty of Biology, Medicine and Health, University of Manchester, Manchester, UK.
9Queen Mary University of London, Centre for Cancer Prevention, Wolfson Institute of Population Health, Charterhouse Square, London, UK
*Equal contribution senior authors 

Importance: PRS are vital for accurate BC risk prediction but require ethnicity-specific calibration. Those of Ashkenazi-Jewish (AJ) descent are assumed to be of White-European (WE) origin in commercially available PRS despite substantially different effect-allele-frequencies (EAFs) between these ethnicities. 

Design: We conducted a retrospective case-control study of WE and AJ women from the PROCAS study. A separate AJ case-control validation series were recruited as part of BCINIS in Northern-Israel between 2000-2022. All women underwent DNA extraction and SNP analysis using Illumina Oncoarray. The previously reported BC PRS’ SNP142/SNP78 were calculated for cases and controls in each population. EAFs were obtained from gnomAD.  

Setting: PROCAS recruited women aged 46-73 years for comprehensive BC risk assessment at mammographic screening attendance in Manchester, UK. The validation series of Israeli AJs were from the BCINIS population-based case-control study which recruited all newly diagnosed BCs in an area of Northern Israel and randomly-selected matched controls. 221 WE (cases=111;controls=110) and 221 AJ (cases=121;controls=110) women were included from PROCAS. 2045 AJ (cases=1331;controls=714) women were included from BCINIS, after exclusion of women from both cohorts with the common Ashkenazi pathogenic variants in BRCA1/2 

Results: In the Manchester cohort, mean SNP142-PRS demonstrated good calibration and discrimination in the WE population, with mean PRS in cases=1.33(1.18-1.48);controls=1.01(0.89-1.13). In the AJ population, mean PRS in cases=1.54(1.38-1.70);controls=1.20(1.08-1.32) demonstrating maintenance of discrimination but poor calibration and overestimation of BC risk. After adjusting for AJ EAF, calibration was corrected (mean SNP142-PRS cases=1.30(1.16-1.44); controls=1.02(0.92-1.12)). This was recapitulated in the larger Israeli dataset. Similar findings were observed for SNP78. The mean PRS adjustment ratio was calculated for the population, and this appropriately corrected for individuals.  

Conclusions: Women of AJ heritage should not be given BC risk prediction based on PRS from WE populations. PRS need to be recalibrated using AJ-derived EAF and a simple recalibration using the mean PRS adjustment ratio performs well. 

 

2. Breast Cancer Know-How: Investigating Screening Willingness and Risk Factor Knowledge Amongst Female Faculty, Staff, and Graduate Students of a Selected Midwest University in the United State of America 

Esther Oshaji1 and Winfred Avogo1

1Department of Sociology and Anthropology, Illinois State University, Illinois, USA.

Background: Breast Cancer (BC) is the second leading cause of cancer deaths in women worldwide. Early detection remains a primary practical approach to combat the disease. Despite having one of the highest BC rates in the world, there is a dearth of literature on the knowledge of BC risk factors and willingness to undergo screening among university students, faculty, and staff in the U.S. Drawing on the constructs of the Health Belief Model, this study is conducted to determine the knowledge and predictors of BC screening behaviors among a distinct population of female graduate students, faculty, and staff at a U.S. Midwestern University, an essential group given that BC risks increase with age. Understanding the perceived susceptibility, benefits, and risks of BC among this demographic is crucial to determining individual determinants of health behavior and developing critical intervention techniques to reduce BC threats to the younger population as they age and decrease mortality among the elderly population via early detection.  

Method: This study uses the quantitative research approach to draw a systematic sample size of 500 respondents. The online survey collects data using a structured questionnaire. The study instrument includes sections on knowledge of risk factors, barriers to and willingness to undertake screening. Data is analyzed using descriptive and multivariate statistics.  

Results: Preliminary results indicate a significant relationship between variables of knowledge, perceived susceptibility, benefits, barriers, and socio-economic factors and women’s screening behaviors.  

Conclusion: Based on the findings of this study, implementing a health belief model-based educational intervention about BC at different phases of life is vital to fight the disease. 

 

3. An ex-vivo human tissue ex-plant model to enable investigations into breast cancer prevention.  

Hannah Harrison1, Sacha Howell1, and Gillian Farnie2 

1Breast Biology, Oglesby Building, Manchester Cancer Research Centre, 555 Wilmslow Road, M20 4GJ.
2Cancer Research Horizons, The Francis Crick Institute, 1 Midland Road, London, NW1 1AT

New prevention agents require extensive pre-clinical testing before they can be utilised in the clinic. Current in vitro and in vivo models of the normal breast do not fully represent the extracellular matrix and its complex cellular micro-environment. In the oncology setting these limitations contribute to the poor rate of translation of targets from lab-based or animal studies to human trials. To overcome these difficulties, we have developed a unique organotypic in vitro model of the normal human breast. Tissue explants, taken following risk-reduction surgery, maintain the inherent complexities of breast structure and micro-environment with key epithelial, stromal and immune cells surviving for 7-days. Explants are cultured within an animal free hydrogel (VitroGel 3D-RGD) and immunohistochemical analysis shows that there is no loss of organisation within the glandular tissue, nor is there a significant change in apoptosis over 7-days. Of note is that there is also no significant change in proliferation in our model which has proven problematic for other in vitro systems. Oestrogen receptor-alpha (ERalpha) and progesterone receptor (PR) continue to be expressed and immune cells, including helper T-cells (CD4), cytotoxic T-cells (CD8) and macrophages (CD68), are maintained both within the ductal structures and the inter-ductal stroma. Importantly, the tissue explants remain responsive to oestrogen; following addition of 17β-estradiol (E2), proliferation (Ki67) and expression of PR increase, both of which are known downstream effects of oestrogen exposure. These changes are blocked by Fulvestrant (FV) showing that the response is ERalpha specific. Additionally, we see from Gene Set Enrichment Analysis of RNAseq data that in tissue treated with E2 we see a positive enrichment score for genes within the known oestrogen response. This is lost following addition FV. This model will offer a platform for pre-clinical testing which faithfully represents patient biology, allowing translation of findings to the clinical setting. 

 

4. Effect of butylphenylmethylpropional (Lilial) on DNA damage and DNA repair in MCF10A immortalised non-transformed human breast epithelial cells. 

Farasani A1,2 and Darbre PD3. 

1Medical Research Center, Jazan University, Jazan, Saudi Arabia.

2Department of Medical Laboratory Technology, Faculty of Applied Medical Sciences, Jazan University, Jazan 45142, Saudi Arabia.

3School of Biological Sciences, University of Reading, Reading RG6 6UB, UK.

Presenter e-mail: [email protected] 

Chemical constituents of personal care products may be a contributory factor in breast cancer development. Butylphenylmethylpropional (Lilial) is added for purposes of fragrance to a range of cosmetic products, but as it is classed as a group II allergen, it is limited to 2.5% in products intended for skin contact.  It has been shown previously to possess oestrogenic activity and exposure to oestrogen is a risk factor for breast cancer. However, it has not previously been investigated for effects on genomic stability which is an enabling characteristic underlying cancer development. We report here that exposure of MCF10A immortalised, non-transformed human breast epithelial cells to Lilial causes DNA damage as measured in a Comet assay. Furthermore, long-term (19-22 weeks) exposure to Lilial results in loss of expression of the breast cancer susceptibility genes BRCA1 and BRCA2 which are key genes in repair of DNA in breast cells. Alterations to expression of other DNA repair genes will be discussed. Research is now needed to determine the extent to which this compound can enter human breast tissues. If it is present in the human breast and can both damage DNA and compromise DNA repair systems, then there is the potential for it to impact on breast carcinogenesis. 

 

5. Multifactorial impacts on early breast carcinogenesis- assessing the combined effect of Endocrine disruptors and fatty acids. 

Gideon Enimah1, Elisabete Silva1 

1Department of Life Sciences, College of Health, Medicine and Life Sciences, Brunel University London, Uxbridge, UB8 3PH, United Kingdom.

Introduction: Breast Cancer is the most common cancer in the world, accounting for nearly 2.1 million new cases in 2020. This high global incidence cannot be solely due to hereditary factors, as evidence has shown that lifestyle and environmental factors (e.g., endocrine disrupting chemicals, EDCs) play a role in breast cancer initiation and progression. The role of EDCs in breast carcinogenesis is still unclear. Similarly, the role of diet, especially fatty acids-rich diets on breast cancer remains inconclusive and the mechanisms responsible for this effect are not fully understood. We have investigated the impact of mixtures of EDCs and fatty acids on early breast carcinogenesis. 

Method: Non-tumorigenic human mammary epithelial cells (MCF12A) were cultured in a 3D system and treated with a mixture of 12 EDCs and fatty acids. The EDCs and fatty acids were mixed at the concentrations present in human serum. In the 3D system, MCF12A cells form structured acini that resemble normal mammary tissue architecture. Early breast carcinogenic events are recapitulated by the formation of large and deformed acini. Different parameters of the acini were measured after exposure to a mixture of EDCs, a mixture of fatty acids and as combination of the two mixtures, representative or real-life exposures. EDCs and fatty mixture were also tested on a panel of 45 genes.  

Results: The 3-dimensional model was able to generate acini similar to in vivo mammary acini. The mixture of EDCs disrupted the growth of acini and the transcription of important genes involved in breast carcinogenesis.  

Conclusion: We showed that our model has the potential to be used in breast cancer related studies. We also showed EDCs can act together at their tissue concentration to disrupt normal breast acini, upregulate important cancer genes suggesting potential role in breast carcinogenesis.  

 

6. Ex Vivo treatment of coronary artery endothelial cells with serum post-exercise training offers limited protection against in vitro exposure to FEC-T chemotherapy 

Marie Mclaughlin1,2, Katie L. Hesketh3, Sarah L. Horgan1, Geraint Florida-James1, Matthew Cocks3, Juliette A. Strauss3 and Mark Ross1,4 

1School of Applied Sciences, Edinburgh Napier University, Edinburgh, United Kingdom
2School of Health and Life Sciences, University of the West of Scotland, Lanarkshire, United Kingdom,  
3Liverpool John Moores University, School of Sport and Exercise Sciences, Liverpool, United Kingdom,  
4School of Energy, Geoscience, Infrastructure and Society, Heriot Watt University, Edinburgh, United Kingdom  

Background: Chemotherapy treatment for breast cancer associates with well-documented cardiovascular detriments. Exercise has shown promise as a potentially protective intervention against cardiac toxicity. However, there is a paucity of evidence for the benefits of exercise on the vasculature. Objectives: This study aimed to determine the effects of chemotherapy on the vascular endothelium; and if there are protective effects of serological alterations elicited by an exercise training intervention. Methods: 15 women participated in a 12-week home-based exercise intervention consisting of three high-intensity interval sessions per week. Human coronary artery endothelial cells (HCAEC) were exposed to physiological concentrations of 5-fluorouracil, epirubicin, cyclophosphamide (FEC) and docetaxel to determine a dose-response. Twenty-four hours prior to FEC and docetaxel exposure, HCAECs were preconditioned with serum collected pre- and post-training. Annexin V binding and cleaved caspase-3 were assessed using flow cytometry and wound repair by scratch assays. Results: Chemotherapy exposure increased HCAEC Annexin V binding, cleaved caspase-3 expression in a dose-dependent manner; and inhibited wound repair. Compared to pre-training serum, conditioning HCAECs with post-training serum, reduced Annexin V binding (42% vs. 30%, p = 0.01) when exposed to FEC. For docetaxel, there were no within-group differences (pre-vs post-exercise) for Annexin V binding or cleaved caspase-3 expression. There was a protective effect of post-training serum on wound repair for 5-flurouracil (p = 0.03) only. Conclusion: FEC-T chemotherapy drugs cause significant damage and dysfunction of endothelial cells. Preconditioning with serum collected after an exercise training intervention, elicited some protection against the usual toxicity of FEC-T, when compared to control serum. 

 

7. Proximity to Breast Cancer Cells Influences Adipocyte Morphology and is Affected by Menopausal Status  

Abigail Dodson1, Justin J. Rochford2, Valerie Speirs1 

1Institute of Medical Sciences, University of Aberdeen, Aberdeen, United Kingdom  

2Rowett Institute, University of Aberdeen, Aberdeen, United Kingdom  

Obesity is a significant risk factor for breast cancer development but in women this can be strongly influenced by menopausal status. In post-menopausal women, obesity increases the risk of breast cancer. In contrast, obesity may protect against breast cancer in pre-menopausal women; the reasons for this are unclear. During obesity, adipose tissue can expand via the enlargement of pre-existing adipocytes and via the generation of new adipocytes, with dynamic changes in adipocyte morphology. They are also believed to take on a tumour-promoting phenotype in association with breast cancer cells. This study aimed to determine whether morphological changes occur to adipocytes specifically when in proximity to breast cancer cells; moreover, whether these changes could be related to tumour development and growth. Adipocyte size, shape, and proximity to tumour cells was evaluated using whole slide digital images of H&E stained sections of cancerous and normal contralateral breast tissue from 26 pre-menopausal patients and 97 post-menopausal patients. From these, 32,000 adipocytes were analysed. Adipocytes were separated into two groups: edge adipocytes (located on the leading edge of the tumour) and distant adipocytes (located >2mm away from the tumour). Our analyses revealed that edge adipocytes were smaller than distant adipocytes in both pre- and post-menopausal cancerous samples (p<0.0001). In addition, the edge adipocytes from cancerous samples were smaller than adipocytes from the corresponding region of normal contralateral breast tissue from pre-menopausal patients (p<0.0001). Intriguingly, the opposite occurred in post-menopausal samples (p<0.0001). Overall, the data suggests that adipocyte morphology may reflect functional interactions with tumour cells. Further analysis of adipocyte parameters and in patients of varying BMI could reveal more about how breast adipocytes influence tumour development and growth. This could also provide insights regarding how obesity differentially impacts breast cancer development in pre- and post-menopausal women. 

 

8. BPA – a major player in breast cancer development? 

Katharina Kusserow1, Paul Fowler1, Valerie Speirs1 

1Institute of Medical Sciences, University of Aberdeen, UK 

The risk of breast cancer development is often increased by endogenous hormones, such as oestrogen, which act on the mammary glands to stimulate tumour formation. However, breast carcinogenesis can also be influenced by exposure to endocrine disrupting chemicals like bisphenol A (BPA), which can act as a weak oestrogen mimic. Concern has been raised that exposure to BPA may influence hormone-sensitive tissues like the breast as it is found ubiquitously present in the environment and especially sources such as food can linings, food containers and water bottles. This project aims to conclusively assess whether BPA exposure contributes to breast cancer development. 

Focussing on human studies only, a systematic review of all existing literature is ongoing. Six different databases were searched with terms including ‘BPA’ or ‘bisphenol A’, ‘mammary gland’ or ‘breast’. After removing duplicates, this returned 611 peer-reviewed publications which are currently being ranked. The literature review will inform future studies and identify gaps in research performed up to the present. In-vitro studies are being performed alongside the literature analysis to investigate long-term low dose level exposures to BPA using a non-cancerous human breast epithelial cell line. Additionally, BPA will be measured in patient-derived samples to inform about potential risks of serum BPA levels. Additionally, publicly available databases and Ingenuity Pathway Analysis will be employed to identify impacted genes and pathways. 

Overall, this project will establish whether there is robust proof about the potential risk BPA poses to breast cancer development. 

 

9. Telomere length elongation by epigenetic modifier drugs: A potential mechanism by which cancer develops chemo-resistance. 

Sarah Al-dulaimi1 

1College of Health and Life Sciences, Brunel University London, Uxbridge, United Kingdom 

Background: Telomeres are specialised structures which are present at the ends of chromosomes. Telomeric DNA is made up of the hexanucleotide repeat TTAGGG which extends for several kilobases in normal healthy cells and shortens by up to 30 nucleotides after each cell division. When telomeric DNA become critically short, normal cells enter senescence and cease cellular growth. Re-activation of telomerase in cancer maintains telomere lengths and allows cells to bypass replicative senescence and gain immortality.   

Aim: To investigate the effect of DNMT inhibitor (5-AZA) and (TSA) on telomere length in breast cancer cell lines.  

Methods: telomere length of treated cells measured by qPCR. DNA damage was determined by H2AX assay and DNA damage at the telomeres was detected by telomere dysfunction induced foci (TIF) assay.  

Results: our results have shown that low concentrations of 5-AZA increased telomere length, DNA damage and damage at telomeres in all the treated breast cancer cells.  

Conclusion: These data provide new insights into the DNMT inhibitor 5-AZA activity in breast cancer cells. 5-AZA increases telomere lengths and DNA damage at the telomeres. This will be further investigated to see what affects this has on cancer cell survival and evolution. 

 

10. Unlocking the glycocalyx in breast cancer 

Diego Sarria1, Miriam Dwek1, Emanuela Volpi1 

1Cancer Research Group, School of Life Sciences, College of Liberal Arts and Sciences, University of Westminster, London, United Kingdom. 

Glycosylation is one of the most important post translational modifications of proteins, including cell membrane proteins, important for protein stability and enabling cell-cell adhesion. Abnormal glycosylation impacts biological functions e.g. protein folding. N-linked glycan structures on glycoproteins can be modified by enzyme activity resulting in the removal or addition of specific monosaccharide residues. Changes in the glycocalyx in cancer can promote both invasion and metastasis. Work is ongoing to gain a better understanding of how the glycocalyx affects response to chemotherapeutics (using glycosidase enzyme inhibitors) in breast cancer.  

The aim of this investigation was to study the affect of inhibiting glycosidases on breast cancer cell behaviour in 2D monolayer cultures.  MCF-7 (ER+), SKBR3 (HER2+), MDA-MB-231, and BT-549 (the latter both triple negative cell lines) were exposed to glycosidase inhibitors: deoxynojirimycin (DNJ),  and novel iminosugars (Phytoquest Ltd).  The effect of glycosidase inhibitors was investigated using lectin fluorescence microscopy for cell glycosylation analysis, Alamar blue assay to assess cellular metabolism and responsiveness to the chemotherapeutic drug doxorubicin and DAPI staining to evaluate changes in nuclear number. A synergistic effect was observed between the chemotherapeutic drug doxorubicin and glycosidase inhibitors in 4 cell lines after a 24-hour treatment with inhibitor. The results indicated changes in N-linked glycosylation in the glycocalyx as all the treated cells exhibited a variation in mannose, fucose and GlcNAc residues (reductions in binding using ConA, UEA-1 and WGA lectins). Triple negative breast cancer cells exhibited changes in glycosylation with changes in responsiveness to doxorubicin compared with the MCF-7 luminal A, ER+, subtype.  

Further analysis is ongoing using 3D cell cultures.  It is expected that through this approach a greater understanding of the role of tumour associated glycans and glycosidase inhibition will provide a key to unlocking the glycocalyx in breast cancer thereby enabling improving responsiveness to chemotherapeutics. 

 

11. Why are there raised rates of breast cancer in aircrew? 

Susan Michaelis1,2 

1Michaelis Aviation Consulting / Lobular Moonshot Project
2Occupational and Environmental Health Research group, University of Stirling, Stirling, Scotland, United Kingdom 

Raised levels of breast cancer in aircrew has been highlighted in published studies ranging at levels up to two times higher than the population. The various associated factors are reported to be occupational factors including ionizing radiation and time factors (circadian rhythm, night shift). To a lesser extent exposure to environmental chemicals including flame retardants has been suggested as a potential relevant factor. A further area that has received minimal attention as risk factor for breast cancer is exposure to aircraft contaminants leaking from the engines into the aircraft air supply, which is not filtered. Low levels of synthetic engine oils and hydraulic fluids are known to leak into the cabin air in normal flight and during fume events. These fluids contain a thermally degraded complex mixture and organophosphates, which are reported to be endocrine disruptors including those mimicking oestrogen. 

The author was medically retired as a pilot in 1997 after chronic low-level exposure to oil fumes, with evidence of exposure to clastogenic chemicals. In 2013 she was diagnosed with invasive lobular breast cancer that failed to show on mammograms or ultrasound and under-recognised on MRI. 7 cm of grade 2 lobular breast cancer was diagnosed after surgery. Lobular breast cancer, known to behave very differently to the main type of breast cancer, has not been afforded required levels of research and does not have its own specific treatments. Recurrence occurred in 2016 and 2021, with the later becoming metastatic lobular breast cancer. 

Further research is ongoing raising awareness about exposure to aircraft air supply fumes, for which there is no form of detection system to advise crew when this occurs. The under-recognition of chronic low-level effects of air supply contamination and the risk of disease requires further research and protections particularly for female aircrew or those who fly regularly. 

 

12. Bioassay-guided purification of annona muricata for the discovery of novel anti-breast cancer compounds 

Derek Osei Berchie1, Svetlana Ignatova1, Terry Roberts2 and Jonathan Huddleston1 

1College of Engineering, Design and Physical Sciences, Brunel University, Uxbridge, United Kingdom.
2College of Health, Medicine and Life Sciences Brunel University, Uxbridge, United Kingdom. 

Natural Products (NPs) provide a largely unexplored pool of clinically relevant metabolites, shown to combat the limitations of conventional cancer and infectious disease treatments. However, concentrations of active components are consistently low in natural medicines thus requiring elaborate extraction and purification methods to achieve viable concentrations of bioactive compounds for effective scientific research and or therapeutic applications. Developing novel cost-effective and selective methods is key for extraction and purification of NPs in respect of drug development from natural medicines, which are ranked amongst the most ethical. Constituent parts of the Annona Muricata plant are documented as promising raw materials for isolating and characterising novel anti-cancer NPs. This research endeavour aims for the extraction, purification and characterisation of novel compounds for advancement in the treatment of Breast Cancer (BC), and the development of new process methods. Here, a simple, novel, UV-visible light spectroscopy guided solid-liquid, low-pressure, gas-agitated extraction method has been developed, primarily for leaching of heat-sensitive/thermolabile compounds, filed for UK patent, and utilised to produce extracts from Annona Muricata leaves and seeds. The extracts, alongside doxorubicin have been evaluated for potency and efficacy against BT474 human BC cell line and MCF10a human breast epithelia reference cell line in MTT assays in vitro. Positive results obtained from the assays and IC50 values, present incentive for further purification of the extracts by Counter-Current and/or High-Performance Liquid Chromatography, alongside further biological assays, and characterisation by Nuclear Magnetic Resonance Spectroscopy and Mass Spectrometry, to determine the extract compounds responsible for the observed bioactivity. After dereplication of compound characterisation data, exists possibility for the discovery of new anti-breast cancer compounds that can be presented for further studies in vitro and in vivo, and to develop new BC treatments. Currently the research has confirmed a new, economical, high-yielding method for extract production from solid raw material.  

 

13. RNA Binding Protein ZFP36L1 and Potential Signatures of Enhanced Tumourigenesis in Breast Cancer 

Varsha Teotia, Nahida Bashir, Khalid Akram1, Ekam Kour1, Kevin Roshan Amalanathan1, Usiju Shaldas, Edward Obioha, Nnawuihe Oluigbo, Kanagaraj Radhakrishnan, John J Murphy and Kalpana Surendranath1 

1Genome Engineering Laboratory, Medicines Diagnostics and Disease Modelling research group, School of Life Sciences, University of Westminster, London, United Kingdom 

The heterogeneous nature of breast cancer, a common malignancy in women has prompted the need to explore new genes that can help us understand the complexity associated with this disease. RNA binding proteins such as HuR, SAM68, RBM38 exhibit context dependent roles and have been reported as the key regulatory factors of breast cancer development. Our study focuses on the Adenine Uridine Rich Elements binding protein ZFP36L1 found recurrently mutated in breast cancers.  It is involved in post-transcriptional regulation of genes controlling cell cycle, apoptosis and DNA damage response potentially acting as a tumour suppressor gene across various cancer cell types. We adapted the CRISPR-Cas9-derived ZFP36L1 knock-out approach from our laboratory to create ZFP36L1 deficient breast cancer cell lines and characterized these cell lines through a combination of functional, proteomic and transcriptomics approaches. Loss of ZFP36L1 in breast cancer cell lines has effects on drug sensitivity, proliferation and differential expression of transcriptome in line with our previous observations in the osteosarcoma cell models. Based on our findings we propose that the effect of ZFP36L1 protein in cellular survival and metabolism is diverse and therefore can be involved in different aspects of tumour development suggesting its potential role in prognosis of certain breast cancers. 

 

14. RNA Binding Protein ZFP36L1 Prevents Replication Stress-Induced Genome Instability 

Ahmed Sidali, Nadeen S Solaiman, Varsha Teotia, Nahida Bashir, Khalid Akram1, Kanagaraj Radhakrishnan, John J Murphy, Kalpana Surendranath1 

1Genome Engineering Laboratory, Medicines Diagnostics and Disease Modelling research group, School of Life Sciences, University of Westminster, London, United Kingdom 

The RNA binding protein (RBP) ZFP36L1, which binds to adenylate/uridylate (AU)-rich elements (AREs) in 3’ untranslated region of many messenger RNAs, has been extensively characterised for its role in post transcriptional control of gene expression and is reported as a newly identified breast cancer driver gene. Replication stress (RS) threatens DNA replication fidelity and stability of the genome. Recently, a small number of ARE-RBPs have emerged as key players in the maintenance of genome integrity through mechanisms that govern the replication stress response and DNA repair. Herein, we report that treatment with low doses of aphidicolin results in hallmarks of RS-associated genomic instability in a cellular model depleted of ZFP36L1 using CRISPR/Cas9. We find that loss of ZFP36L1 results in defects in mitosis leading to chromosome missegregation and micronuclei formation. Remarkably, we also find that loss of ZFP36L1 leads to fragile site instability marked by the increase in G1 associated 53BP1 nuclear bodies, ultrafine anaphase bridges and chromosome gaps/breaks. Furthermore, we detected an increase in RPA foci in S/G2 cells indicative of increased ssDNA potentially susceptible to further DNA damage as shown by the increase in γH2AX foci colocalising with 53BP1. Inducible expression of ZFP36L1 demonstrated reversal of these phenotypes associated with loss of ZFP36L1 in the T-REx U2OS cells. Taken together, our work highlights an important, yet previously unidentified role, for ZFP36L1 in preserving genomic stability.

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