Co‐occurrence of breast cancer and neuroendocrine tumours: New genetic insights beyond Multiple Endocrine Neoplasia syndromes

Abstract Objective Age‐standardized incidence of female breast cancer is 145.1 per 100000/year and 5.86 per 100000/year for neuroendocrine tumours (NET) in Canada. Evidence is scarce about gene variants that may predispose patients to develop both neoplasms. The objective of this study was to identify germline gene variants associated with this combination of tumours. Design and patients A retrospective chart review (2007‐2018) in a tertiary NET referral centre was completed. A series of 9 female patients with concurrent breast cancer and NET is presented. All patients underwent a 37 gene hereditary cancer next‐generation sequencing panel. Results Mean age was 61.4 years (35‐85) at breast cancer diagnosis and 63.4 years (51‐89) at NET diagnosis. Four patients had a pancreatic, three had a small bowel and two had a lung NET. Two patients were known cases of MEN1, and one patient was found to harbour a pathogenic variant in MEN1 and a variant of unknown significance (VUS) in ATM. A second patient was found to harbour a pathogenic variant in APC. A third patient was found to carry a pathogenic variant in PALB2 as well as a VUS in FANCM, MLH1 and STK11. Another patient was found to harbour a VUS in MSH2. One patient was found to carry a pathogenic variant in NTHL1. Conclusion The first cases of a PALB2, an APC and a NTHL1 pathogenic variants in patients with both breast cancer and NET were presented. NGS testing should be considered in specific patients with this combination of neoplasms, as certain germline variants beyond MEN1, have important implications for cancer surveillance.


| INTRODUC TI ON
Age-standardized incidence of female breast cancer was 145.1 per 100000/year in 2012 1 in Ontario, Canada. Neuroendocrine tumours (NET) are less common, with an incidence of 5.86 per 100000/year in 2009 in the same population. 2 Evidence is scarce about the possible germline (inherited) genetic variants that may predispose patients with NET to develop breast cancer or vice versa. Multiple Endocrine Neoplasia type 1 (MEN1) syndrome, [3][4][5][6] Multiple Endocrine Neoplasia type 4 (MEN4, caused by CDKN1B) 7,8 and Cowden syndrome (PTEN) 9 have been associated with both NET and breast cancer in case reports or case series. A whole-genome sequencing study of 102 pancreatic NET patients by Scarpa et al reported novel pathogenic mutations in DNA repair genes MUTYH, CHEK2 and BRCA2, which are well-known breast cancer susceptibility genes. 10 However, NET are not incorporated into genetic testing guidelines for breast cancer patients. 11 Recently, a SEER database retrospective study 12 showed that the incidence of pancreatic NET is higher in patients with a first primary cancer of the pancreas, bladder, thymus and female breast cancer. However, multiple institutional cohort studies have reported a higher incidence of gastrointestinal and genitourinary second primary malignancies in patients with NETs [13][14][15] and an Italian national database study 16 of patients with a primary diagnosis of bronchopulmonary NET has shown a higher incidence of thyroid neoplasms in women and a higher incidence of kidney and bladder tumours in men. Importantly, these studies did not show a higher incidence of breast cancer in NET patients. We here report a case series of nine female patients with a concurrent diagnosis of breast cancer and NET (lung, GI tract or pancreatic primary) evaluated at a tertiary NET referral centre between 2007 and 2018 as well as the results of a 37 gene next-generation sequencing panel that was performed on all patients. To our knowledge, this is the first study using this approach to report on possible gene variants linking the co-occurrence of breast cancer and NET.

| ME THODS
Three hundred and fifty-three patients with a NET (lung, pancreas or gastrointestinal) were evaluated at our tertiary NET referral centre between 2007 and 2018 (Princess Margaret Cancer Centre, Toronto, Ontario, Canada). Of these, 206 were female of whom 21 were diagnosed with breast cancer either prior to NET diagnosis or during follow-up. Nine patients accepted to take part in the study and are detailed in this report. Four were lost to follow-up prior to the accrual period and eight declined to participate in the study. The breast cancer could have been diagnosed prior to the NET diagnosis, synchronously (here defined as within 6 months before of after the NET diagnosis) or metachronously (more than 6 months after the NET diagnosis). Patients' data including clinical data (age, sex, age at diagnosis of each tumour and follow-up time for each tumour), breast cancer data (affected breast, surgical intervention, hormone receptor status, surgical pathology reports, imaging reports, treatments including chemotherapy, radiation therapy, aromatase inhibitor therapy, trastuzumab and other therapies) were recorded from the electronic medical record and clinic chart. Given that our NET clinic is a provincial tertiary referral centre, some patients had their breast cancer treated in another hospital; therefore, some details on the breast cancer data were unavailable. Patient's NET data including primary site, surgical intervention, WHO grade, surgical pathology reports, imaging reports, treatments including somatostatin analogue therapy, mTor inhibitor therapy (everolimus), peptide receptor radionuclide therapy (PRRT with 177-Lutetium), capecitabine-temozolomide chemotherapy and other therapies, presence and location of metastases and treatment modalities of metastases were recorded from the electronic medical record. All patients un-

| Clinical cases: Breast cancer data
Breast cancer data are summarized in Table 1 for all study participants. All patients (n = 9) were female, with a mean age of 61.4 years (35-85) at breast cancer diagnosis. With regards to surgical resection, six patients had a lumpectomy, two had a mastectomy, and one was not a surgical candidate due to her advanced age. In terms of adjuvant therapy, two patients received chemotherapy, four received radiotherapy, six received an aromatase inhibitor and one received trastuzumab. Surgical pathology revealed that five tumours were invasive ductal carcinomas, one was a ductal carcinoma in situ, and two were lobular carcinomas. Mean tumour size was 1.1 cm (0.4-2.0).
Three tumours had a histological grade 2 and one had a grade 3. All tumours were Stage 1a. Hormone receptor status for tumours was the following: seven tumours were ER+, 5 were PR + and only one was HER-2/Neu+. Mean follow-up time for the breast cancer was 7.44 years (2-26) Three patients had locoregional recurrence. Two patients developed breast cancer metastases; one in cervical lymph nodes and bone metastases and one had liver metastases. Four patients had their breast cancer diagnosed before their NET, 2 patients had both malignancies diagnosed synchronously (defined here as within 6 months) and three patients were diagnosed with breast cancer metachronously (ie more than 6 months after NET diagnosis).

| Clinical cases: Neuroendocrine tumour data
Neuroendocrine tumours data are summarized in Table 2

| Clinical Cases: Comprehensive hereditary cancer gene variant panel results
Results of the 37 NGS gene panel are summarized in Table 3

| D ISCUSS I ON
In this study, we report 9 cases of female patients with co-occurrence of breast cancer and a NET (lung, GI tract or pancreas primary) evaluated at a tertiary NET referral centre between 2007 and 2018. During this period, 21 out of 206 female NET patients evaluated at our centre had a concurrent diagnosis of breast cancer; Eight declined to participate in the study and four were lost to follow-up prior to the accrual period. This is in-line with the 1/9 frequency of breast cancer in the general female population. The aim of our study was not to demonstrate that female NET patients have a higher incidence of breast cancer, but rather to illustrate that the were also identified in a whole-genome sequencing study of 102 pancreatic NET patients. 10 Although one recent SEER national database retrospective study showed an increased risk of pancreatic NET in patients with another first primary malignancy including female breast cancer, this was not found in other population studies. [13][14][15][16] Here, we report likely pathogenic and pathogenic variants in APC and PALB2 in cases of co-occurrence of breast cancer and NET. These pathogenic variants are respectively known to be as-

| CON CLUS ION
Patients with one malignancy are more often predisposed to de-

ACK N OWLED G EM ENTS
None.

CO N FLI C T S O F I NTE R E S T
The authors have no conflict of interest to disclose.

AUTH O R S' CO NTR I B UTI O N S
Author Vincent Larouche was the principal investigator of the study. He completed the Ethics Board application, contacted patients, organized patient visits and blood draws, collected data and prepared the manuscript. Author Amit Akirov helped with the literature review, data collection and manuscript preparation. Author Emily Thain is a Genetics Counsellor who helped with literature review, manuscript preparation and proceeded to genetic counselling of all patients who underwent NGS testing. Author Raymond Kim is a Medical Geneticist who contributed to the literature review and manuscript preparation and oversaw the genetic testing of subjects. Author Shereen Ezzat is an Endocrine Oncologist, and he was the main supervisor for the study.

DATA AVA I L A B I L I T Y S TAT E M E N T
The data that support the findings of this study are available on request from the corresponding author. The data are not publicly available due to privacy or ethical restrictions.

E TH I C S S TATEM ENT
All patients signed written consent to be included in this case series, and our institution's research ethics board approved the study.