Title: Developing a diagnostic tool, using SPAG5, for predicting clinical benefit from standard anthracycline combination (AC) chemotherapy in breast cancer
Funder: Department of Health and Social Care

Abstract: Rationale: Approximately 70% of BC patients can be spared unnecessary AC-chemotherapy by using SPAG5 as a predictive biomarker of response to AC-chemotherapy. Aim: Our final goal is to develop a diagnostic assay/product for SPAG5 protein that is suitable for IHC staining of FFPE tissue and to validate it as a predictive test for sensitivity to AC-chemotherapy which is suitable for adoption by the NHS. This new approach would enhance patient quality of life and save money by avoiding unnecessary treatment and the costs of managing unnecessary side-effects. Challenge: Given that there is no available SPAG5-MAb that is validated for IHC staining, the challenge is to develop a sensitive, specific, reproducible and cost-effective SPAG5 IHC staining assay/product which can predict sensitivity to AC-chemotherapy sensitivity, to validate its clinical utility and health/economic benefits. A: Development of a diagnostic assay/product for the semi-quantitative IHC-testing for SPAG5-protein in FFPE samples. Production of recombinant-SPAG5 protein/immunogen for the immunisation of mice will be undertaken by Origene. Generation, sequencing and molecular engimeering of the MAbs will be undertaken by BioGenes GmbH (Berlin, Germeny) and Absolute Antibodies (Oxford, UK). The hybridomas producing SPAG5-MAbs will be generated and screened for specificity by Western blot, immunoprecipitation and immunofluorescence staining assays using cell lines which have been transiently transfected with a SPAG5-expression-plasmid and a control-vector. B: Clinical validation study B1: IHC analysis of SPAG5-expression IHC staining using the developed SPAG5-MAb will be performed on the original cohorts who have been tested with the SPAG5-Polyclonal antibody (PAb) 3. The ability of IHC staining with the new SPAG5-MAb to predict recurrence risk (RFS) and response (pCR) after receiving adjuvant-AC and AC-NACT will be validated in new series of 2000 and 600 cases; respectively. B2: Comparison with rival genetic tests The ability of SPAG5 MAb and PAb IHC staining to predict recurrence risk (RFS) after receiving adjuvant-AC will be compared to the Oncotype DX test in 200 cases who received the test as part of their standard of care treatment. The ability of SPAG5 MAb and PAb IHC staining to predict sensitivity to AC-NACT (pCR) will be compared to that of PAM-50, mamma-print, Genomic-Chemo-Sensitivity-Predictor, and the 30-gene-DLDA tests in 200 cases of pre-chemotherapy biopsies. A bespoke gene panel featuring genes used in the aforementioned tests, along with SPAG5, will be developed for the nanoString nCounter&trade; FLEX instrument. C: Commercialisation and Health Economics: C1: Health Economics Conducted from the perspective of the NHS and Personal Social Services, the Net Benefit Framework method4 will be used to build economic models for estimating the added costs and savings, as well as the added patient benefits that are expected to be achieved when switching from the current standard of care to SPAG5-informed management. The economic models developed will take the form of an initial decision tree that feeds into a Markov process to represent longer-term outcomes. Model parameterisation will be sought with inputs from the Patient/Public Advisory Group, clinical expert opinion, prospective audit data, and published literature. C2: New Assay/Product We will engage a commercial partner (Source Biosience; Nottingham, UK) who has the necessary expertise in the Regulatory Requirements for delivering clinically validated diagnostic tests into the NHS setting. This collaboration will enable us to effectively develop a prototype SPAG5 protein testing assay/product, including a semi-quantitative scoring system, Schedule The project comprises 9 work packages (see Gantt chart) and four stop/go points: &ldquo;Do we have a SPAG5-MAb suitable for IHC?&rdquo; (Month 12), &ldquo;Does the new MAb work better than the old PAb?&rdquo; (Month 18) and &ldquo;Does the new MAb work better than the rival commercial genetic tests?&rdquo; and dose it produce a beneficial health economic case (Month 33).

Lay abstract: We are developing an inexpensive test to accurately predict how breast cancer patients will respond to the standard chemotherapy Anthracycline (AC). Only 15-20% of patients have no tumour remaining following AC, so a method of treatment selection is urgently needed. Every year nearly 62,000 people are diagnosed with breast cancer in the UK. One in eight women in the UK will develop breast cancer in their lifetime. Breast cancers are currently treated with a combination of chemotherapy, targeted therapy and surgery. However, breast cancers are not identical; each tumour s individual characteristics affect how they respond to treatment. We recently discovered a new tumour characteristic, a protein which is unusually active in approximately 20% of breast cancers. We found that a patient whose tumour showed high activity often respond well to AC, and vice versa. AC is an aggressive treatment which can potentially cause severe side-effects, including a risk of permanent heart damage. It is important, therefore, to spare those patients who will not benefit from AC the physical and emotional side-effects of this drug. Currently, there is no predictive test for selecting which patients will benefit from AC and which will not. We have shown that we can accurately make this prediction by testing the activity of a protein called SPerm associated AntiGen 5 (SPAG5) in tumour tissue. We aim to develop a clinical SPAG5 testing kit that can be used by hospital laboratories to determine the activity of SPAG5 in the tumour. Our immediate goal is to produce a trial-ready kit in the three years of support offered by this grant. We will then seek further funding to evaluate the clinical performance of the kit and build the case for approval for use in the NHS, where it will help guide treatment decisions and improve outcomes for patients .

Research activity codes: 4.1 DISCOVERY AND PRECLINICAL TESTING OF MARKERS AND TECHNOLOGIES
Health categories: CANCER AND NEOPLASMS