BACKGROUND: Proliferation markers and profiles have been recommended for guiding 
the choice of systemic treatments for breast cancer. However, the best molecular 
marker or test to use has not yet been identified. We did this study to identify 
factors that drive proliferation and its associated features in breast cancer 
and assess their association with clinical outcomes and response to 
chemotherapy.
METHODS: We applied an artificial neural network-based integrative data mining 
approach to data from three cohorts of patients with breast cancer (the 
Nottingham discovery cohort (n=171), Uppsala cohort (n=249), and Molecular 
Taxonomy of Breast Cancer International Consortium [METABRIC] cohort; n=1980). 
We then identified the genes with the most effect on other genes in the 
resulting interactome map. Sperm-associated antigen 5 (SPAG5) featured 
prominently in our interactome map of proliferation and we chose to take it 
forward in our analysis on the basis of its fundamental role in the function and 
dynamic regulation of mitotic spindles, mitotic progression, and chromosome 
segregation fidelity. We investigated the clinicopathological relevance of SPAG5 
gene copy number aberrations, mRNA transcript expression, and protein expression 
and analysed the associations of SPAG5 copy number aberrations, transcript 
expression, and protein expression with breast cancer-specific survival, 
disease-free survival, distant relapse-free survival, pathological complete 
response, and residual cancer burden in the Nottingham discovery cohort, Uppsala 
cohort, METABRIC cohort, a pooled untreated lymph node-negative cohort (n=684), 
a multicentre combined cohort (n=5439), the Nottingham historical early stage 
breast cancer cohort (Nottingham-HES; n=1650), Nottingham early stage oestrogen 
receptor-negative breast cancer adjuvant chemotherapy cohort 
(Nottingham-oestrogen receptor-negative-ACT; n=697), the Nottingham 
anthracycline neoadjuvant chemotherapy cohort (Nottingham-NeoACT; n=200), the MD 
Anderson taxane plus anthracycline-based neoadjuvant chemotherapy cohort (MD 
Anderson-NeoACT; n=508), and the multicentre phase 2 neoadjuvant clinical trial 
cohort (phase 2 NeoACT; NCT00455533; n=253).
FINDINGS: In the METABRIC cohort, we detected SPAG5 gene gain or amplification 
at the Ch17q11.2 locus in 206 (10%) of 1980 patients overall, 46 (19%) of 237 
patients with a PAM50-HER2 phenotype, and 87 (18%) of 488 patients with 
PAM50-LumB phenotype. Copy number aberration leading to SPAG5 gain or 
amplification and high SPAG5 transcript and SPAG5 protein concentrations were 
associated with shorter overall breast cancer-specific survival (METABRIC cohort 
[copy number aberration]: hazard ratio [HR] 1·50, 95% CI 1·18-1·92, p=0·00010; 
METABRIC cohort [transcript]: 1·68, 1·40-2·01, p<0·0001; and 
Nottingham-HES-breast cancer cohort [protein]: 1·68, 1·32-2·12, p<0·0001). In 
multivariable analysis, high SPAG5 transcript and SPAG5 protein expression were 
associated with reduced breast cancer-specific survival at 10 years compared 
with lower concentrations (Uppsala: HR 1·62, 95% CI 1·03-2·53, p=0·036; 
METABRIC: 1·27, 1·02-1·58, p=0·034; untreated lymph node-negative cohort: 2·34, 
1·24-4·42, p=0·0090; and Nottingham-HES: 1·73, 1·23-2·46, p=0·0020). In patients 
with oestrogen receptor-negative breast cancer with high SPAG5 protein 
expression, anthracycline-based adjuvant chemotherapy increased breast 
cancer-specific survival overall compared with that for patients who did not 
receive chemotherapy (Nottingham-oestrogen receptor-negative-ACT cohort: HR 
0·37, 95% CI 0·20-0·60, p=0·0010). Multivariable analysis showed high SPAG5 
transcript concentrations to be independently associated with longer distant 
relapse-free survival after receiving taxane plus anthracycline neoadjuvant 
chemotherapy (MD Anderson-NeoACT: HR 0·68, 95% CI 0·48-0·97, p=0·031). In 
multivariable analysis, both high SPAG5 transcript and high SPAG5 protein 
concentrations were independent predictors for a higher proportion of patients 
achieving a pathological complete response after combination cytotoxic 
chemotherapy (MD Anderson-NeoACT: OR 1·71, 95% CI, 1·07-2·74, p=0·024; 
Nottingham-ACT: 8·75, 2·42-31·62, p=0·0010).
INTERPRETATION: SPAG5 is a novel amplified gene on Ch17q11.2 in breast cancer. 
The transcript and protein products of SPAG5 are independent prognostic and 
predictive biomarkers that might have clinical utility as biomarkers for 
combination cytotoxic chemotherapy sensitivity, especially in oestrogen 
receptor-negative breast cancer.
FUNDING: Nottingham Hospitals Charity and the John and Lucille van Geest 
Foundation.