NIHRDataInsights/HRCSData huggingface dataset:
Title: Elucidation and characterisation of the CREBBP-FOXM1 signalling axis as a biomarker of response and resistance to CDK4/6 inhibition in breast cancer
Funder: Breast Cancer Now

Abstract: Aims We aim to address two fundamental aspects and inadequately addressed needs relevant to palbociclib use in breast cancer, i) identification of additional patient populations who would benefit from treatment and ii) investigation of resistance mechanisms using clinically relevant disease models. Background Palbociclib is a selective inhibitor of CDK4/6 and a proven therapeutic strategy for oestrogen receptor positive (ER+) breast cancers (BC). However the molecular determinants of sensitivity and resistance to CDK4/6 inhibition are unclear. We have identified a robust association with loss of CREB-binding protein (CREBBP) and sensitivity to palbociclib in triple negative breast cancer (TNBC), mediated through increased FOXM1 protein expression, a downstream target of CDK4/6 and potent oncogenes. Lower CREBBP expression is associated with particularly aggressive subtypes of TNBC. Our preliminary data also suggest that dysregulation of the CREBBP-FOXM1 axis is involved in resistance to palbociclib in ER+ BC. Techniques and methodology We will use a combination of in vitro, ex vivo and in vivo experimental approaches to assess the efficacy of palbociclib, in patient derived organoid and xenograft models alone and in combination with taxanes and metabolic inhibitors. Using mouse intraductal (MIND) models of ER+ and TNBC patient derived material, we will define the mechanism of resistance at RNA and protein levels, cross-reference these and identify novel treatment strategies. Evaluation of CREBBP and FOXM1 gene activity will be assessed in clinical trial cohorts of both ER+ and TNBC using PALLET and Chemo-NEAR. Impact on breast cancer research Results from this work will (a) have significant impact on the treatment of aggressive subgroups of TNBC for whom currently there is no targeted therapy, (b) identify and validate robust biomarkers of response to CDK4/6 inhibition in patient populations with ER+ BC, and (c) provide a fundamental understanding of how to identify and treat patients with CDK4/6 inhibitors.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS;4.1 DISCOVERY AND PRECLINICAL TESTING OF MARKERS AND TECHNOLOGIES
Health categories: CANCER AND NEOPLASMS
Title: Phase Ib Study of Palbociclib and Avelumab in Metastatic AR+ triple negative breast cancer
Funder: Breast Cancer Now

Abstract: Gene expression studies have demonstrated a distinct luminal androgen receptor (LAR) sub-type of triple negative breast cancer (TNBC), which accounts for approximately 20% of cases characterised by high AR expression and low E type cyclin expression. Although targeting of the androgen receptor has been evaluated in 3 small phase studies of bicalutamide, enzalutamide and abiraterone, response rates have been disappointing. The CDK4/6 inhibitor, palbociclib has significant efficacy combined with endocrine therapy in patients with metastatic ER positive, HER2 negative BC. We have shown that palbociclib has substantial activity in breast cancer cell lines of the LAR sub-type of TNBC (Asghar et al., 2015), with similar levels of activity to ER positive breast cancer cell lines. PDL-1 expression, a mechanism of evading immune system recognition, is relatively common in TNBC, therefore inhibitors of PD-1 and PDL-1 are undergoing clinical evaluation in TNBC. Avelumab is a human IgG1 anti-PDL-1 mAb, with low activity in unselected breast cancer patients (Dirix et al., SABCS 2015). CDK4/6 inhibition induces an interferon response in tumour cells driver by upregulation of STAT1 (unpublished data). In ER positive breast cancer, neoadjuvant CDK4/6 inhibition induces lymphocytic infiltration in residual cancer (Hurvitz S et al SABCS 2016). CDK4/6 inhibition may be pro-immunogenic with the potential to synergise with PDL1 blocking antibodies. The PAVEMENT study aims to translate our prior pre-clinical findings on LAR TNBC, with a dose finding escalation phase, and an expansion in AR positive TNBC. Although there are no overlapping toxicities, the dose finding phase in up to 18 patients will establish the schedule of avelumab in combination with palbociclib. The expansion phase will screen up to 230 patients with AR immunohistochemistry, to treat 27 patients with the avelumab-palbociclib doublet, and objective response primary endpoint. Sufficient activity in the study would allow a subsquent randomised study.


Research activity codes: 6.1 PHARMACEUTICALS
Health categories: CANCER AND NEOPLASMS
Title: Preclinical evaluation of drug efficacy using patient-relevant breast explant models: a bench-to-bedside approach
Funder: Breast Cancer Now

Abstract: i) Background Predicting clinical response to anticancer drugs is a major challenge in cancer treatment. Patient relevant preclinical models that retain tumour heterogeneity and the microenvironment are urgently needed coupled with the development of predictive and pharmacodynamic biomarkers. We have invested in a personalized explant culture platform that fulfills these criteria and has emerged as a powerful platform for predicting responses in patients. ii) Aims (including use of Pfizer drugs) We propose to undertake an exploratory study that analyses breast cancer explant response to three Pfizer drugs in early phase development. We will identify molecular markers predictive of response and validate pharmacodynamic biomarkers. Positive correlations will be identified, leading to the design of stratified clinical trials. iii) Techniques and Methodology The University Hospitals of Leicester (UHL) is one of the largest breast cancer referral centres in the UK, with ~800 surgical resections annually. We will derive explants from surgical resections representative of the major histological subtypes and analyse the in situ response of both stroma and tumour areas to Pfizer drugs as single agents and in combination. Pharmacodynamic biomarkers will be validated and Next Generation Sequencing (NGS) whole exome profiles generated to allow genotype-phenotype correlations. An NGS “hotspot” breast cancer panel covering the major genetic changes in our cohorts will be validated within circulating free (cf) DNA. Integration of genetic, clinical, pathological, biomarker and explant data will allow rationale design of personalised clinical studies, using cfDNA genomic profiling to guide patient stratification. iv) Impact on breast cancer research We anticipate accurate prediction of subtype-specific response to Pfizer drugs, and novel information on mechanisms of drug action. These data will provide strong rationales for the design of bespoke clinical interventions, which will be delivered through the Leicester Clinical Trials Facility.


Research activity codes: 5.1 PHARMACEUTICALS
Health categories: CANCER AND NEOPLASMS
Title: Developing strategies for treatment of breast cancer patients following progression on CDK4/6 inhibitors
Funder: Breast Cancer Now

Abstract: Background: Dysregulation of cell cycle control involving CDK4/6 is common in ER+ breast cancer (BC) and the CDK4/6 inhibitor palbociclib plus endocrine-therapy (ET), improves clinical outcome in advanced post-menopausal ER+/HER2-ve disease. However, resistance to CDK4/6 inhibitors is an emerging problem. To ascertain mechanisms of response/resistance to palbociclib, we generated cell line models of acquired palbociclib resistance as monotherapy, or in estrogen-deprivation resistance models. Key to palbociclib resistance is altered CDK signalling, chiefly loss of RB1 and/or amplification/over-expression of CDK2, CDK4, CCNE1 and CDK7. siRNA library screening demonstrated dependence of palbociclib-resistant cells on CDK2, CDK4, CDK9 and CDK7. CDK7 is required for activation of cell cycle CDK1/2/4/6. CDK7 also controls transcription initiation by phosphorylating RNA polymerase II and CDK9, with particular sensitivity of super-enhancers that direct expression of cancer drivers, to CDK7 inhibition. Perturbation of CDK7 prevents transcription of key cell cycle drivers. Aims: We will (i) establish mechanisms of palbociclib resistance, focusing on redundancy of cyclin dependent kinases (CDK) and (ii) address the modulating role of CDK7 and its utility as a therapeutic target in overcoming resistance. Techniques/Methodology: Ectopic expression of CDKs upregulated in palbociclib resistance and specific inhibitors of these CDKs will be used to define their relative contribution in models of palbociclib sensitive and resistant BC. Proteomic, transcriptomic and epigenomic methodologies will identify the importance of cell cycle and transcription inhibition in resistance and potential of CDK7 inhibition as a therapeutic strategy. Findings will be validated in palbociclib resistant patient-derived tumour xenografts/organoids. Impact on Breast Cancer Research: Defining the mechanisms of palbociclib resistance and utility of CDK7 inhibition to overcome this will be an important advance. Importantly, rapid progression to the clinic is feasible, as several CDK7 inhibitors showing remarkable lack of toxicity in pre-clinical in vivo models have been developed, with one already advanced to phase I.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS;5.1 PHARMACEUTICALS
Health categories: CANCER AND NEOPLASMS
Title: Duocarmycin prodrugs to treat breast cancer: single-agent and combination therapy efficacy in 2D and 3D cancer models
Funder: Breast Cancer Now

Abstract: Background Breast cancer is the most frequently diagnosed cancer in women, with 50,000 cases in women and 400 in men being recorded each year in the UK alone. The projected figure for 2035 is 70,000 which is in accordance with an ageing population and continued improvements in survival. Recurrent breast cancer is largely incurable by currently-used drugs and radiotherapy and hence better treatment options are required for people living with breast cancer. Aims Specifically, this project seeks to determine the efficacy of CYP1A1/1B1-bioactivated duocarmycin prodrugs as single-agent and in combination with radiotherapy or sunitinib (treatment elevates CYP1A1 expression) using a range of 2D and 3D breast cancer models. Techniques and Methodology CYP1A1/1B1 expression will be analysed using a panel of breast cancer cell lines and correlate duocarmycin prodrug efficacy as single-agent or in combination with radiotherapy or sunitinib in 2D and 3D cancer models. CYP1A1/1B1 expression will be analysed using a range of techniques (RT-PCR, immunoblotting, immunohistochemistry and LC/MS multiple reaction monitoring. MTT assays and clonogenicity (crystal violet dissolution assay, manual counting), cell cycle changes (using FACS) and γ-H2AX as a mechanistic endpoint for DNA damage indicative of a cytotoxic event will carried out to correlate duocarmycin prodrug activity. Impact on breast cancer research Successful execution of research objectives in this proposal will help to support the progression of a duocarmycin prodrug towards clinical evaluation. Safely administered and selective duocarmycins can add significant value to the existing treatment options currently available for breast cancer patients, especially those suffering from metastatic disease.


Research activity codes: 5.1 PHARMACEUTICALS
Health categories: CANCER AND NEOPLASMS
Title: Intraoperative spectroscopic evaluation of sentinel lymph nodes in breast cancer
Funder: Breast Cancer Now

Abstract: Background The status of the sentinel lymph node (SLN) in invasive breast cancer (BC) influences subsequent management of the axilla. Current modalities for intraoperative detection of SLN either show low sensitivity, consume whole tissue or are time consuming and laborious. Aims This study aims to develop a novel tool for fast intra-operative assessment of SLN using cutting-edge biophotonics (Raman spectroscopy and auto-fluorescence imaging) combined with advanced statistical modelling and machine learning techniques to develop a medical device that can be used in the surgical theatres to assess SLN status. Techniques and Methodology Develop and validate a diagnostic model based on integrated Raman spectroscopy, guided by auto-fluorescence imaging, and machine learning techniques for diagnosis of SLN (target accuracy >95% sensitivity/specificity). Develop an integrated framework for semi-supervised learning to improve the performance of diagnosis in real-time by including data from new patients. Establish the design of a prototype device which will classify a sample within speed and accuracy levels compatible with clinical use: initial target is 10 minutes at 95% diagnosis sensitivity/specificity. Impact on breast cancer research This novel multimodal spectral imaging technique will provide a cost-effective and accurate intraoperative platform for assessment of SLN.


Research activity codes: 4.1 DISCOVERY AND PRECLINICAL TESTING OF MARKERS AND TECHNOLOGIES
Health categories: CANCER AND NEOPLASMS
Title: Development of a novel, targeted cytotoxic treatment for breast cancer
Funder: Breast Cancer Now

Abstract: Background Despite advances in detection and adjuvant therapies, treatment of metastatic breast cancer (mBC) remains palliative, claiming the lives of 500,000 women worldwide each year, emphasising the magnitude of the unmet clinical need. While targeted therapies exist for individual patient subtypes, cytotoxic chemotherapy remains important for all subgroups. The success of chemotherapy is limited by drug resistance and toxicities. The duocarmycins are ultrapotent cytotoxins, which evade multidrug resistance and DNA repair mechanisms, but proved too toxic for human use. Targeted delivery of duocarmycins has the potential to increase therapeutic index. Aims We will apply the principles of precision medicine to the delivery of a novel, dual-targeted duocarmycin cytotoxic prodrug activated by tumour membrane-type matrix metalloproteinases, and demonstrate pre-clinical proof-of-concept to underpin early clinical trials in women with mBC. Techniques and Methodology We will employ an iterative approach to compound synthesis, informed by detailed LC-MS-based ex vivo metabolism studies developed in our laboratory. These assays will assess prodrug activation in tumour tissues and stability in normal tissues (plasma, liver, kidney, lung) at the bench, and will predict in vivo pharmacokinetics. Two prodrugs will be identified for in vivo evaluation of the maximum-tolerated dose and pharmacokinetics, prior to selecting one candidate for efficacy studies in relevant murine models. Impact on breast cancer research If successful this will be a first-in-class, novel potential therapeutic, and provide a rationale for a clinical trial. This project has significant potential to improve the quantity and quality of life for countless mBC patients, regardless of molecular subtype.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS;5.1 PHARMACEUTICALS
Health categories: CANCER AND NEOPLASMS
Title: Myoepithelial/luminal cross-talk in the progression of DCIS to invasive breast cancer
Funder: Breast Cancer Now

Abstract: Background The mechanisms behind the progression of DCIS to invasive cancer remain poorly understood. Myoepithelial-luminal cell interactions are critical for the maintenance of the breast duct bilayer and become perturbed in breast cancer. Myoepithelial cells in particular dramatically alter their biology in DCIS, where they can adopt a more invasive phenotype, and are unidentifiable in invasive cancer. Aims We hypothesise that changes in cross-talk between myoepithelial and luminal cells underpin breast cancer progression. Identifying these changes will uncover defining events in cancer progression and lead to novel prognostic biomarkers and therapeutic targets. Techniques and Methodology We have developed an in vitro model system of the human breast duct bilayer using patient derived cells. Luminal expression of HER2 creates a DCIS phenotype, while myoepithelial expression of the high-risk DCIS marker ITGB6 promotes cell invasion, revealing a new role for these cells in invasion. We will combine this model with a newly developed proteomic approach to dissect interactive cellular cross-talk in a multi-cellular system. Using lysine biosynthesis enzymes and isotopically labelled precursors we will selectively and continuously label the individual proteomes of myoepithelial, luminal and fibroblast cells. Using this system we will be able to unpick the changes in cellular cross-talk that occurs during destabilisation of the bilayer, and in the initiation of myoepithelial-led invasion. Critical drivers of progression will be validated using genetic/pharmacological approaches and their expression will be investigated in clinical samples of DCIS, to provide strong clinical evidence for their use as markers of progression. Impact on breast cancer research Breast cancer is currently overtreated due to an inability to identify patients whose DCIS will progress to invasive cancer. This project will identify the changes in myoepithelial-luminal cross-talk that occur during cancer progression and will identify markers to be used in the clinic to help determine effective patient therapies.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: CANCER AND NEOPLASMS
Title: TALAZOPARIB AND CHLORAMBUCIL COMBINATION AS A THERAPEUTIC STRATEGY FOR BRCA1/2-DEFICIENT TUMOURS
Funder: Breast Cancer Now

Abstract: BRCA1 and BRCA2 genes are mutated in approximately 25% of the familial cases of breast and ovarian cancers (Futreal et al., 1994; Miki et al., 1994; Wooster et al., 1995). In addition, a significant proportion of sporadic breast and ovarian tumours carry BRCA1 and BRCA2 alterations, as demonstrated by genome sequencing of tumour collections (Cancer Genome Atlas, 2012; Cancer Genome Atlas Research, 2011; Merajver et al., 1995). Cells and tumours with compromised BRCA1 or BRCA2 function accumulate collapsed replication forks and chromosome breaks. However, they retain the ability to proliferate, in spite of the severe genomic instability. This vulnerability is exploited by highly specific therapies that enhance susceptibility of BRCA1/2-deficient tumours to DNA damage, with PARP inhibitors as a prominent example. PARP inhibitors prevent repair of single-stranded DNA breaks and lead to persistent DNA lesions. Alkylating agents represent a class of drugs routinely used in the clinic against tumours, including the BRCA1/2-deficient ones (Vollebergh et al., 2011). They act by introducing intra- and inter-strand DNA crosslinks, which trigger DNA single-stranded and double-stranded breaks (Fu et al., 2012). It is conceivable therefore that combining PARP inhibitors with alkylating agents could be more effective in treating BRCA1/2-mutated cancers than each monotherapy. The aim of the current proposal is to examine whether the combination of the PARP inhibitor talazoparib with the bi-functional alkylating agent chlorambucil will selectively inhibit growth of BRCA1/2-deficient tumoursin vivo. We aim to show that the combination is more effective than either monotherapy, that it is well-tolerated and can induce robust tumour regression. We will also endeavour to identify potential biomarkers for the talazoparib/chlorambucil combination in tumour and non-tumour tissues. If this study meets its objectives, we will propose a phase 1 / 2 clinical trial of the combination in patients with BRCA1/2-mutated tumours.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS;5.1 PHARMACEUTICALS
Health categories: CANCER AND NEOPLASMS
Title: Evaluating the mechanistic role of PARP inhibitor sensitivity in spliceosomal mutant breast cancer
Funder: Breast Cancer Now

Abstract: i) Aims (including use of Pfizer drug) We aim to address two fundamental aspects and inadequately addressed needs relevant to talazoparib use in breast cancer: i) identification of additional patient populations who would benefit from treatment and ii) investigation of resistance mechanisms in these novel patient populations using clinically relevant disease models. ii) Background Novel mutations in SF3B1, a component of the RNA splicing machinery underpin 3-5% of ER+ breast cancers, promote breast cancer growth and are associated with a poor response to endocrine therapy. Through synthetic-lethal drug screens we have identified a robust association of SF3B1 hotspot mutations and sensitivity to talazoparib and defects in the replication stress response. iii) Techniques and Methodology We will use a combination of in vitro, ex vivo and in vivo experimental approaches to assess the efficacy of talazoparib, in patient derived organoid, xenograft and syngeneic models alone and in combination with transcriptional inhibitors. We will evaluate the mechanism of transcriptional stress induced by talazoparib using ChIP and RNA sequencing approaches to profile RNAPolII transcription rates; define the role of dysregulated ER transcription in SF3B1 driven tumorigenesis under talazoparib treatment; and investigate whether these alterations can be used as patient selection biomarkers. Finally, we will investigate the mechanism of resistance to single and combination approaches at the chromatin, single cell RNA and protein level and cross-reference these to identify novel treatment strategies. iv) Impact on breast cancer research Results from this work will (a) have significant impact on the treatment of an aggressive subgroup of ER+ breast cancer who fail current therapies, (b) identify effective combination strategies, patient selection biomarkers and potential resistance mechanisms; and (c) provide a fundamental understanding of how to identify and treat ER+ BC patients harbouring somatic SF3B1 mutations with PARP inhibitors.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: CANCER AND NEOPLASMS
Title: Validating a new combinatorial approach for Palbociclib to target triple-negative breast cancer
Funder: Breast Cancer Now

Abstract: The Cdk4/6 inhibitor Palbociclib shows promising therapeutic results in women with hormone receptor-positive advanced breast cancer but this approach is less successful in triple-negative breast cancer (TNBC). This necessitates the development of novel combined therapy approaches that could be applied to specific subgroups of TNBC. This proposal aims to investigate a novel regulatory pathway that links Greatwall kinase with Cdk4/6 in the control of cell cycle progression from G1- to S-phase in triple-negative breast cancer (TNBC) cells. Based on our preliminary results we hypothesise that Greatwall kinase supports Cdk4/6 in promoting pRB phosphorylation in early G1 phase, thereby elevating levels of Cdk2:Cyclin E activity and S-phase initiation. We aim to: 1) Analyse the molecular mechanism of this new cell cycle control circuit. 2) Validate the synergistic effects of Greatwall inhibitors and Palbociclib as a potential target for cancer therapy in TNBC in vitro and in vivo. We have put together an interdisciplinary approach to tackle these questions, involving single-cell analysis, phosphoproteomics and in vivo experiments. We specifically aimed to maintain a balance between advancing our basic understanding of tumour biology and developing clinical applications that could lead to patient survival. Upon completion, our work will have two major outcomes: Firstly, we will generate information on a novel cell cycle control pathway that regulates the transition from G1 to S-phase. Secondly, we will provide a novel strategy to apply Cdk4/6 inhibition in a combinatorial approach for TNBC cancer cells and potentially other tumour types that show resistance to Palbociclib. Our mechanistic insight will suggest biomarkers to stratify TNBC that could be amenable to combined Palbociclib treatment and Greatwall inhibition. Overall, this study will validate a promising novel combined therapy approach to apply the Cdk4/6 inhibitor Palbociclib in TNBC patients.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: CANCER AND NEOPLASMS