Original ArticlesIncreased in vitro and in vivo sensitivity of BRCA2-associated pancreatic cancer to the poly(ADP-ribose) polymerase-1/2 inhibitor BMN 673
Introduction
Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal malignancies worldwide [1]. Approximately 80% of new cases are diagnosed late, with advanced disease precluding curative resection [2]. Unfortunately, the therapeutic options currently available for these patients are largely ineffective and even patients who present with operable disease have poor outcomes following resection due to early recurrences [3]. The challenges in identifying therapies with meaningful outcomes may reflect the genetic heterogeneity of PDAC. Therefore, research focused on genetic ‘cataloguing’ of PDAC [4] may identify subsets of patients who will benefit from tailored treatment approaches [5]. Although the full spectrum of PDAC subtypes remains to be characterized, investigating PDAC associated with hereditary syndromes provides an opportunity to characterize the therapeutic sensitivities of PDAC arising from common genetic driver mutations.
Approximately 10% of PDAC cases are associated with strong family histories, with a fraction of these accounted for by the tumor spectrums of recognized hereditary syndromes [6]. These syndromes include hereditary breast and ovarian cancer (HBOC) syndrome, most often caused by germline mutations in the BRCA1 or BRCA2 genes [7]. Since BRCA1- or BRCA2-deficient tumors have impaired homology-directed DNA repair (HDR), therapeutic strategies that exploit defects in HDR may represent an avenue for targeted therapy development for these PDAC cases [8].
The hypothesis that BRCA1- and BRCA2-deficient cells are sensitive to agents that target DNA repair mechanisms is supported by a growing body of literature suggesting increased sensitivity of BRCA1- and BRCA2-associated breast and ovarian cancer to DNA crosslinking agents (DCLs) and poly(ADP)-ribose polymerase inhibitors (PARPis) [9], [10]. Since DCLs cause double-strand DNA breaks (DSBs) that must be repaired by HDR, BRCA1- and BRCA2-deficient cells are vulnerable to these agents. PARPis exploit the dependence of BRCA1- and BRCA2-deficient cells on alternative cellular DNA repair pathways by disrupting the base excision DNA repair (BER) pathway, creating a synthetic lethal environment for cells with impaired HDR.
Despite these promising opportunities for personalized therapies, there is a lack of preclinical data comparing the various DCLs and PARPis to rationalize the selection of agents for clinical trial. In the present study, we present a PDAC case with a germline BRCA2 mutation who exhibited a marked response to platinum-based chemotherapy (FOLFIRINOX [11]). We confirm biallelic BRCA2 inactivating mutations in the patient's tumor and investigated the in vitro cytotoxicities of a panel of DCLs and PARPis in BRCA2-deficient PDAC cell lines followed by in vivo validation of the two most efficacious agents, cisplatin and BMN 673 (a PARPi) in a xenograft model derived from our patient. Specifically, we evaluated the efficacy of the newest generation PARPi, BMN 673 [12], in comparison to a panel of commonly used DCLs as well as veliparib, which is currently under clinical trial evaluation for BRCA-associated PDAC [13]. Our findings support a role for personalized therapeutic strategies for BRCA2-associated PDAC and suggest that BMN 673 be considered for clinical trial in PDAC with impaired HDR.
Section snippets
Cell culture
Capan-1 (HTB-79), MIA PaCa-2 (CRL-1420) and PANC-1 (CRL-1469) were obtained from ATCC (Manassas, USA) and cultured in DMEM (Wisent, St-Bruno, Canada) supplemented with 10% FBS, 5% glutamine and 5% penicillin–streptomycin.
Compounds
Gemcitabine (Enzo Life Sciences, Brockville, Canada), cisplatin (Enzo Life Sciences), oxaliplatin (Sigma Aldrich, Oakville, Canada), carboplatin (Sigma Aldrich), veliparib (Enzo Life Sciences) and BMN 673 (Abmole Biosciences, Hong Kong, China) were resuspended in water or DMSO.
Real-time cell analysis (xCELLigence)
Clinical response to platinum-based therapy in a PDAC case with a germline BRCA2 mutation
Fig. 1A shows partial and complete radiological responses of the primary tumor and liver metastasis, respectively. Whole genome sequencing of the patient's primary tumor revealed biallelic BRCA2 inactivating mutations. Sanger sequencing confirmed that the germline mutation but not the “second hit” somatic mutation is present in the patient's lymphocyte DNA, while both the germline and “second hit” BRCA2 mutations are present in the patient's primary tumor (Fig. 1B). We also confirmed, by Sanger
Discussion
The poor outcome of patients with PDAC reflects the desperate need for improved treatment strategies [24]. In this study, we assessed the efficacy of DCLs and PARPis in BRCA2-associated PDAC. As predicted, we observed increased in vitro sensitivity of BRCA2-deficient (Capan-1) PDAC cells to all agents tested. We also showed that shRNA-mediated reduction of BRCA2 expression in PANC-1 induces sensitivity to cisplatin and BMN 673 but not to veliparib, highlighting the increased potential efficacy
Conflict of interest
The authors have no conflicts of interest to declare.
Acknowledgements
This article is dedicated to the late Rosalind Goodman for her philanthropy and tireless efforts to promote cancer research in our institution and community. We would like to thank Dr. Michel Tremblay for his guidance with the xCELLigence assays, Dr. Dongmei Zuo and the Histology Core at the Goodman Cancer Research Centre for technical support with the immunohistochemistry, and Dr. William Foulkes for facilitating rapid clinical genetic testing and confirmation of the germline mutation in our
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