50% HGSOC patients harbour defects in homologous recombination repair (HR), where there are many proteins involved eg. BRCA. Mutations in BRCA genes (BRCAm), affect ∼15% of the HGSOC population. RAD51 is another key protein in HR, binding to broken strands, forming nucleoprotein filaments to facilitate repair. This process can be identified via RAD51 foci, subnuclear complexes which can be viewed microscopically. RAD51 is overexpressed in many cancers including HGSOC; higher levels in FFPE tissue of patients with HGSOC are associated with improved DNA repair efficiency and poorer survival (8, 9). Here we sought to identify RAD51 foci in circulating cancer-associated cells correlating them with BRCA status and survival outcomes.
124 HGSOC patients had blood samples taken pre chemo of whom 99 were suitable for analysis. 53 of these had further samples taken during and after treatment, and in follow up (see CONSORT Fig 1). These were processed using an immunofluorescence protocol. CCs were stained with WT1/RAD51 antibodies and viewed under LEICA microscopy, where RAD51 foci were counted. Results were correlated with clinical data and outcomes.
Of 40 patients (5 BRCAm/HRD+) with paired samples, 32 responded according to CA125 (GCIG criteria). These patients had a significant reduction in CCs (p = 0.02) and RAD51 foci (p = 0.0007). By contrast, 8 non-responding patients had no change in RAD51 foci or CC. The numbers of RAD51 foci seen in platinum resistant patients (n=15) were significantly greater compared with those (n= 56) who were platinum sensitive (PS) both at the beginning and end of chemotherapy. There were no RAD51 foci in 7 BRCA1m/HRD+ compared to BRCAwt (n=42) patients (p=0.009). Median progression free survival (mPFS) had not been reached in those with <3 RAD51 foci (n=39) versus 13 months >3 RAD51 foci (n=32) at the start of treatment, p=0.04. mPFS survival was 12 months in those with <3 RAD51 foci (n=18) versus 3 months for those with >3 RAD51 foci at the end of treatment (n=13), p=0.001.
Here we demonstrate RAD51 foci in CCs from HGSOC patients. Numbers of cells and RAD51 foci change with treatment; fewer CC/RAD51 foci are associated with improved PFS.
Legal entity responsible for the study
East and North Herts NHS Trust, Brunel University London.
Cancer Treatment and Research Trust, John Bush Memorial Fund at Mount Vernon Cancer Centre.
All authors have declared no conflicts of interest.