The region of Podkarpacie is located in the South-East corner of Poland, bordering Ukraine and Slovakia. In our previous study, we identified 10 of 158 (6.3%) of unselected cases of ovarian cancer from this region carried one of 13 founder mutations in the BRCA1 or BRCA2 genes [9]. This is less than in other regions of Poland where the frequency of BRCA1 causative founder variants was observed in about 10-13.5% of ovarian cancer patients [2,3,4,5,6,7,8]. In this study, we performed the NGS study of BRCA1/2 genes in the same group of 158 women affected with ovarian cancer and diagnosed 18 (11.4%) BRCA1/2 mutation carriers. The frequency of individual BRCA1/2 mutations observed in ovarian cancer patients is shown in Table 1. All ten founder mutations were confirmed with our observations performed with a genetic test based on the BRCA1/2 founder mutations characteristic for the Polish population [9]. In addition, we diagnosed 1 carrier of BRCA1 and 7 carriers of the BRCA2 gene. These 8 mutations were detectable by whole sequencing only. Like in other regions of Poland, the most frequent mutation was the BRCA1 c.5266dupC mutation observed in 30% (6/18) of all carriers and the BRCA1 c.181T > G mutation found in 15% (3/18). Other 9 BRCA1/2 mutations were observed in single patients and are rare in the Polish population. In contrary to our former observation the frequency of BRCA1/2 mutation carriers in the group of ovarian cancer patients is only slightly lower than in other regions of Poland. However, we observed a significantly lower frequency of founder mutations, in particular, BRCA1 c.5266dupC and to a lesser extent BRCA1 c.181T > G. This phenomenon can be caused in general by the lower frequency of these mutations in South-East Poland. However, it should be noted that in this region the extensive genetic testing of BRCA1/2 genes has been carried out in patients with ovarian and breast cancer, as well as, in healthy patients since the year 2000. The testing focused mainly on the detection of founder mutations. As a result, several hundred families with BRCA1 founder mutations have been diagnosed so far and several hundred prophylactic adnexectomies have been performed. It should be taken into account that these BRCA1 founder mutations carriers were thus protected against ovarian cancer, and therefore, we observe their lower representation among BRCA1/2 mutation carriers who have now developed ovarian cancer.

The mean age at diagnosis in the 11 cases with BRCA1 mutation was 55.8 years, and of the 7 patients with BRCA2 mutation was 60.1 years. In both groups, the mean age at diagnosis was slightly higher than the observed in BRCA1/2 carriers from other regions of Poland [6,7,8]. Possibly there are lifestyle/environmental factors which may influence the later age of diagnosis in BRCA1/2 carriers. However, for non-carriers, the mean age at diagnosis was similar in the region of Podkarpacie and the rest of Poland (58.77 vs. 56.2–62.3 years) [6,7,8].

We observed strong family history in 10 of 18 (55.5%) mutation carriers, which is slightly more frequent than in other regions [6,7,8]. This applies to families with mutations in the BRCA1 as well as the BRCA2 gene. It can be explained by the relatively larger number of family members in an average family, in this region. However, the frequency of BRCA1/2 mutation carriers with negative family history is so high (44.5%), both in groups with recurrent founder mutations as well as with non-founder mutations, that we cannot recommend limiting performing the BRCA1 and BRCA2 gene testing based on NGS to cases with a burdened family history only. Also, the ovarian cancer age of onset is not a factor facilitating the qualification for this study. Taking into account our observations, it should be stated that performing a test based on the detection of Polish founder mutations in ovarian cancer patients from the Podkarpacie region is associated with relatively low sensitivity (55.5%). In turn, performing the NGS test in all subsequent patients with ovarian cancer is associated with a significant increase in costs. Application of NGS tests only in familial cases is associated also with low sensitivity of 55.5%. One of the compromise solutions would be to perform a standard genetic test based on the detection of founder mutations in all patients, and then if no mutation is detected, perform NGS in cases with a family history. With this algorithm of procedure, the sensitivity of detecting the BRCA1/2 gene mutation in ovarian cancer patients would increase to 77.8%, at a relatively low cost.

However, taking into account the current diagnostic standards of patients with ovarian cancer in the context of determining the optimal treatment and qualifying patients for treatment with PARP inhibitors, the BRCA1/2 gene is routinely tested using the NGS method in DNA extracted from tumour cells. It seems, therefore, that the most justifiable algorithm for detecting a germinal mutation in these patients is to start testing BRCA1/2 genes using the NGS method in DNA extracted from tumour cells. Then, if a mutation is found, the test should be performed in the patient’s peripheral blood to verify whether it is a germline or somatic mutation. If germline mutation is confirmed, the study should be extended to other relatives. However, if such a protocol is used, it should be taken into account that up to nearly 10% of cancer patients with negative NGS results performed in DNA isolated from neoplastic tissue cells, in fact, may carry germline mutation [11]. Lincoln et al. [11] indicated that major reasons for the mutations omissions include: (i) somatic variant interpretation guidelines differ from germline variant interpretation guidelines [12, 13]; (ii) high-quality germline tests can detect a broad spectrum of pathogenic variant types but present technical challenges particularly when analyzing archival specimen types often encountered in oncology e.g., paraffin-embedded formalin-fixed tissues [14,15,16,17], (iii) tumor tests may not include all genes of potential germline relevance in any given patient. Since the material for somatic mutation tests is mainly derived from tissues embedded in paraffin blocks, a significant percentage of the isolated DNA is of poor quality and unsuitable for NGS tests. It should be noted that patients with inconclusive tumor tests may also be carriers of germline mutations [18]. Therefore, the results of NGS tests carried out in somatic tissues should be interpreted with special care.