No abstract

Patients with biallelic truncating mutations in PALB2 have a severe form of Fanconi anaemia (FA-N), with a predisposition for developing embryonal-type tumours in infancy. Here we describe two unusual patients from a single family, carrying biallelic PALB2 mutations, one truncating, c.1676_1677delAAinsG;(p.Gln559ArgfsTer2), and the second, c.2586+1G>A; p.Thr839_Lys862del resulting in an in frame skip of exon 6 (24 amino acids). Strikingly, the affected individuals did not exhibit the severe developmental defects typical of FA-N patients and initially presented with B cell non-Hodgkin lymphoma. The expressed p.Thr839_Lys862del mutant PALB2 protein retained the ability to interact with BRCA2, previously unreported in FA-N patients. There was also a large increased chromosomal radiosensitivity following irradiation in G2 and increased sensitivity to mitomycin C. Although patient cells were unable to form Rad51 foci following exposure to either DNA damaging agent, U2OS cells, in which the mutant PALB2 with in frame skip of exon 6 was induced, did show recruitment of Rad51 to foci following damage. We conclude that a very mild form of FA-N exists arising from a hypomorphic PALB2 allele.

PALB2 is DNA damage response protein that acts in Homologous recombination Repair (HRR) pathway and promotes recruitment of principal HRR components such as BRCA1, BRCA2 and Rad51. Biallelic PALB2 inactivation is associated with the severe form of Fanconi anemia (FA-N), characterised by severe developmental abnormalities and occurrence of embryonal tumours in infancy. Given the rarity of reported patients with biallelic PALB2 inactivation, the full extent of phenotypes associated with PALB2 mutations is still unknown. Here we describe a family with two inherited PALB2 mutations where the affected individuals presented with few of the clinical features typically exhibited by FA-N patients. The patients, age 19 and 15 were lacking severe developmental abnormalities and displayed impaired growth and mild learning difficulties. Interestingly, however, both individuals developed B cell Non-Hodgkin lymphoma (NHL) at the early age of 12 and 3.5 years respectively, responded well to intensive chemotherapy and stayed in complete remission. Their cellular phenotype included a high level of spontaneous chromosomal damage suggestive of DNA repair defect, but otherwise deviated from the classic FA phenotype revealing an intermediate sensitivity to mitomycin C (MMC) by colony forming assay but an unusually high chromosomal radiosensitivity in G2. We addressed the possibility that this atypical clinical and cellular phenotype could be related to the functional properties of the mutant PALB2 protein(s) expressed from the affected alleles. Indeed, we demonstrated that the PALB2 protein (T839_K862del) produced from one of the mutant alleles carried by these patients retained both its N- and C-terminal domain required to interact with BRCA1 and BRCA2 respectively. We also showed that this mutant protein preserved some function compared with previously identifiedpatient-associated mutant PALB2 proteins and allowed low level recruitment of Rad51 foci at sites of DNA damage. We propose that the presence of this mutant protein accounts for the milder MMC hyper-sensitivity exhibited by cells from these patients and the lack of FA defining clinical characteristics. Finally, the shift in tumour spectrum from embryonal tumours towards B cell NHL observed in this family implies PALB2 as a potential suppressor of lymphoid tumourigenesis and raises the possibility that somatic mutations in this gene may contribute to the development of sporadic lymphoid tumours. Disclosures Janic: Baxter: Other: Paid Instructor, Research Funding; Pfizer: Other: Paid Instructor, Research Funding; Bayer: Other: Paid Instructor, Research Funding; Octopharma: Research Funding; Novo Nordisk: Other: Paid Instructor, Research Funding.

PALB2 protein is one of the principal components of Homologous Recombination Repair (HRR) that facilitates recruitment of other HRR proteins such as BRCA1, BRCA2 and Rad51. Biallelic PALB2 inactivation is associated with a severe form of Fanconi Anemia (FA-N), however, biallelic PALB2 mutations also occur in patients with a FA-N syndrome variant characterised by a predisposition to B-cell lymphoma development. The association of PALB2 with FA-N and FA-N variant suggests that PALB2 may play a pathogenic role in a wider range of sporadic haematopoietic malignancies. To address this possibility, the mutational status of the PALB2 gene was assessed in 30 myelodysplastic syndrome (MDS) samples, 23 acute lymphoblastic leukaemias (ALL), 171 chronic lymphocytic leukaemias (CLL) and 24 paediatric non-Hodgkin lymphomas (NHL) of T- or B-cell origin. Overall, pathogenic (truncating or splicing errors)orlikely pathogenic sequence changes (missense alterations not previously reported but predicted to alter protein function) were detected in 17 (6.7%) of these patients. PALB2 was most frequently altered in NHL (25%), followed by MDS (6%), CLL (5%) and ALL (4%). The overall incidence of changes observed for B-cell NHL was higher than the frequency detected in T-cell NHL. PALB2 sequence changes were distributed across the whole gene including the DNA binding, BRCA1-interacting and BRCA2-interacting domains. Of note, a single truncating mutation; in the BRCA1-interacting domain, c.886delA; was detected in three cases, two with progressive CLL and one with MDS and interestingly only became detectable in one of these CLL patients following ibrutinib treatment. Furthermore, a polymorphic variant (c.2993G>A), previously associated with an increased breast cancer risk, was present at higher frequency than in the general population in cohorts of both CLL and ALL patients. HRR defective tumour cells can be targeted by pharmacological inhibition of co-operating DNA repair pathways using the principal of synthetic lethality. To explore whether this concept can be utilised in the context of PALB2 deficiency, we evaluated the effectiveness of inhibition of two pathways; PARP and ATR; that co-operate with PALB2. The cytotoxicity induced by either an ATR inhibitor or a PARP inhibitor was assessed in lymphoblastoid cell lines derived from a paediatric B-cell NHL patient carrying the c.1676_1677delAAinsG and c.2586+1G>A mutations. As predicted, exposure of lymphoid cells with PALB2 mutations to either ATR or PARP inhibitor alone exerted a cytotoxic effect which was enhanced when both inhibitors were applied in combination. In conclusion, our results suggest that PALB2 is altered in a subset of patients with different haematopoietic malignancies. This potentially provides another avenue for targeted therapies utilising the concept of synthetic lethality via application of PARP and ATR inhibitors that are currently being tested in clinical trials. Disclosures Janic: Novo Nordisk: Other: Paid Instructor, Research Funding; Baxter: Other: Paid Instructor, Research Funding; Bayer: Other: Paid Instructor, Research Funding; Pfizer: Other: Paid Instructor, Research Funding; Octopharma: Research Funding.