BRCA1 and BRCA2 mutations sensitize to chemotherapy in patient-derived pancreatic cancer xenografts

Abstract

Background: Germline mutations of the BRCA tumour suppressors have been associated with increased risk of pancreatic cancer. Clinical evidence suggests that these patients may be more sensitive to treatment with cisplatin. As the frequency of germline BRCA mutations is low, definitive experimental data to support the clinical observations are still missing.

Methods: We tested gemcitabine and cisplatin sensitivity of four BRCA1 and BRCA2 mutant and three BRCA1 and BRCA2 wild-type (WT) patient-derived pancreatic cancer xenografts.

Results: We observed treatment sensitivity to gemcitabine and cisplatin in the BRCA WT and mutant models. The BRCA1 and BRCA2 mutant xenografts were significantly more sensitive to cisplatin although these models also showed sensitivity to gemcitabine. The BRCA1 and BRCA2 WT models showed sensitivity to gemcitabine but not cisplatin. Treatment sensitivity in the xenograft models closely resembled treatment response in the corresponding patients.

Discussion: We have characterised a panel of xenografts derived from pancreatic cancer patients carrying germline BRCA mutations, and shown that their genetic features resemble the patient donor. Our results support further clinical testing of treatment regimens combining gemcitabine and platinum drugs in this patient population, as well as preclinical research aiming to identify mechanisms of cisplatin resistance in BRCA mutant pancreatic cancers.

Figure 1

BRCA1 and BRCA2 mutants. (A) The BRCA1 amino terminus contains a RING domain and the nuclear localisation sequence (NLS). The carboxyl terminus contains the coiled coil domain that facilitates BRCA2 binding, a SQ/TQ cluster domain (SCD) that contains 10 possible ATM phosphorylation sites and a BRCT domain, which is associated with BRCA1 recruitment to the DNA damage site. The mutation found in OCIP217 leads to the expression of a truncated BRCA1 protein that has lost the BRCT domain and parts of the ATM SCD. (B) BRCA2 contains eight BRC repeats between amino acids (AA) 1009 and 2083 that binds RAD51. The carboxyl terminus contains helical domain (H), three oligonucleotide binding (OB) domains, which facilitates binding to both single-stranded and double-stranded DNA, and an NLS. All BRCA2 mutants express truncated proteins that have lost the carboxyl terminus. While OCIP28 maintains seven BRC repeats, OCIP232 and OCIPA1 have lost the RAD51-binding domains. Cellular localisation of (C) RAD51(red) in sections of xenograft tumours treated with vehicle or cisplatin. DAPI (blue) was used to visualise the cell nucleus.

Figure 2

Mutations in BRCA1 and BRCA2 increase treatment sensitivity and survival. (A) Mice were treated for 4 weeks with cisplatin, gemcitabine or vehicle, and tumour volume was evaluated three times a week; mt, mutant. (B) Survival in response to treatment was measured until mice either reached the humane tumour end point or their natural life span in the case of no recurrence of the tumour after treatment. Error bars represent s.d. # indicates no tumours detected. Mice were killed due to old age. (C) CT scans for patient from which OCIP217 was derived. Primary tumour in the body of pancreas with involvement of coeliac axis and splenic artery (i, arrow) and solitary liver metastasis (ii, arrow). Major response to gemcitabine and cisplatin with marked shrinkage of primary tumour (iii, arrow), and radiologic complete response of liver metastasis (iv, arrow).

Figure 3

BRCA1 and BRCA2 mutations result in γH2AX foci accumulation in response to cisplatin treatment. Section of mice treated with a single dose of cisplatin or vehicle were stained for (A) γH2AX and (B) CC3. BRCA mutant (mt) models display a significant increase in γH2AX foci that was not observed in the WT models 24 h after treatment. Cleaved caspase-3 staining, however, was not increased in the BRCA mt models. Error bars represent s.d.