Immunohistochemical detection of BRCA1 and BRCA2 proteins in patients with PCa is easy and cost-effective for countries where molecular testing is still a challenge. In this study, we made a hypothesis that association of these gene proteins which are among the key genes mutated in the process of developing of PCa with clinicopathological prognostic factors such as Gleason score, age and PSA among others may help to determine the prognosis of the disease.
The prevalence of expression of BRCA1 protein in this study was 26.1%. This was higher than 15% and 1.7% that were reported in the USA and Israel, respectively [17, 18]. The low prevalence of 1.7% for expression of BRCA1 which was reported by Giusti among the Ashkenazi patients with PCa in Israel could have been attributed to a number of factors including different methods of detection between the two studies, low level of hereditary type of PCa among the Ashkenazi men and also the difference in aggressiveness of the disease for the two races. All these factors might have contributed greatly to the difference in expression of BRCA1 protein. Moreover, the study which was done in the USA used monoclonal BRCA1 antibody compared to polyclonal BRCA1 antibody that was used in the current study. Therefore, the difference in specificity between the two different types of antibodies used for the two studies could be the reason for the difference in the expression of the antibody for the studies. The difference in clonality of the antibodies between the two studies greatly resulted to the substantial difference in expression.
In 2011, Rabiau et al. reported the prevalence of expression of BRCA1 protein detected by means of IHC as 22.4% which is lower than 26.1% was found in this study . The large percentage of patients with high Gleason score in our study compared to the small percentage of patients with high Gleason score in their study contributed for the difference in expression of the antibody between the two studies. This is because studies have shown that poorly differentiated PCa is associated with high expression of BRCA1 protein . Also the subjectivity in interpretation of the positivity of the antibody may have led to the existing difference in the level of expression.
The prevalence of expression of BRCA2 tumour suppressor gene in patients with PCa is lower than BRCA1 [21, 22]. Detection of both BRCA1/2 proteins by means of IHC gives a high prevalence of expression compared to detection by using molecular tests such as real time-quantitative polymerase chain reaction (RT-qPCR) which are more specific as it was reported in the studies in the USA and Israel. These two previous studies used RT-qPCR test to determine the prevalence of mutation of BRCA2 whereby 15% and 1.7% were reported, respectively [17, 18]. At different occasions, Stephen et al. and Tryggvadottir et al. reported 2.3% and 5.7%, respectively [23, 24].
In the study of Tryggvi et al., it was reported that the prevalence of expression of BRCA2 in the controls was 25% slightly higher than 22.9% that was found in our study and 38% in the cases, also higher than the prevalence in our study . The difference in prevalence could have resulted from the two reasons: the first reason is the difference in the methodology. The previous study employed use of tissue microarray (TMA) technique, which by far, provides high positivity rate of the marker based on the reason that selection of the tissue areas to be stained is optimal. The second reason is that the previous study used the antibody for C-and N-terminal of the BRCA2 protein. These give more positivity compared to the approach used in the current study. The heterogeneous nature of the expression of these tumour suppressor genes has also reported to contribute greatly to the difference in prevalence of expression for the different studies done [19, 25].
In our study, we analysed the association of BRCA1 and BRCA2 proteins expression with age of the patients at diagnosis, pre-treatment PSA and Gleason score as the clinicopathological prognostic factors in our study. Besides, we went further and did analysis of the co-expression of the two antibodies with the prognostic factors. We considered a case to have co-expression of BRCA1 and BRCA2 if it could stain for both proteins simultaneously.
Regarding the association of BRCA1/2 protein based on IHC detection method, the results seem to be contradicting. Edwards et al. reported that BRCA2 protein expression was higher in cases aged ≤ 65 years, but the difference was not statistically significant (P = 0.08) . This is contrary to what was found in the present study where cases expressing BRCA2 had higher median age than those not expressing the protein (74 years versus 71 years) and the difference was not statistically significant (P = 0.091) but is in agreement with the observation in the study conducted by Kirchhoff et al. . In their study, it was reported that presence of BRCA2 expression was higher in old patients compared to younger patients.
Among the possible reasons that can explain these contradicting results, it is the difference of the age of onset of the diseases for the different races. In a study done on the African population, it was found that PCa cases in Africa present approximately a decade earlier than it is the case in western countries . In the present study, it was observed that only few cases (< 20%) were aged less than 64 years. Therefore, this could have attributed to the lack of association between BRCA2 expression and age of the patients at diagnosis. This shows that there was no significantly increased risk for early onset prostate cancer in BRCA1 expression. Besides, the small proportion of patients with early onset of PCa in this study could also be due to lack of screening programmes in the country [14, 15]. This causes delayed detection of the diseases reflected by the small percent of cases with early onset of the disease in the present study.
PSA has been studied extensively as one of the clinical biomarkers that can predict the clinical outcomes of the patients with PCa [28, 29]. However, its ability to predict aggressiveness and progression of the disease is controversial due to the fact that there are many conditions which may cause rising of its level. Neil et al. reported that rising of PSA before diagnosis of PCa is associated with poor clinical outcomes. Additionally, they reported that short doubling time of PSA among patients with PCa is associated with biochemical recurrence and even poor prognosis [30, 31]. In settings where screening of PCa is not routinely done, patients are usually diagnosed with very high PSA level. This is one of the factors explaining the poor outcomes of patients with PCa in settings where screening is not a routine practice where Uganda is included [14, 15].
Fiorentino et al. reported the association of BRCA1 expression with PSA. Patients with BRCA1-positive had median PSA level of 27.0 ng/mL) higher than median PSA level of 10.2 ng/mL) in patients who were BRCA1-negative and the difference was statistically significant (P = 0.0056) . This finding is different from our finding which showed that although there was higher median PSA in BRCA1-positive cases compared to BRCA1-negative cases (449.8 ng/mL versus 389.4 ng/mL), but the difference was not statistically significant (P = 0.446). The difference could have been due to the fact that most our cases had high PSA level unlike the patients in the previous study.
When we analysed the association between BRCA2 protein expression and the PSA level, we found that the median PSA level for BRCA2-negative cases was higher than the BRCA2-positive cases (455.1 ng/mL versus 363.2 ng/mL). This was a converse trend of the level of PSA in terms of progression of the disease, and the difference was not statistically significant (P = 0.399). This finding is similar to the finding in the study of Thorne et al. which also reported that there was no association between PSA and BRCA2 protein expression in patients with PCa (P = 0.006) although BRCA2-positive cases had higher median PSA than BRCA2-negative cases (56.2 ng/mL versus 100 ng/mL) . In another study of Gallager et al., it was found that there was no association between PSA and BRCA2 mutation in patients with BRCA2 wild-type (P = 0.99) and those with BRCA2 mutant gene and both groups had the same median PSA of 7 ng/mL .
There was a significant statistical difference between BRCA-positive with Gleason score ≤ 7 and BRCA-positive cases with Gleason score ≥ 8 (P = 0.013). There were many BRCA1-positive with Gleason score score ≥ 8 than BRCA1-positive cases with Gleason score ≤ 7 (42 cases versus 7 cases). This was similar to the finding reported in the study of Fiorentino et al. who reported that BRCA1-positivity was associated with Gleason score (P = 0.004). They found that cases with Gleason score 8–10 were 21 compared to Gleason score 4–6, 3 + 4 = 7 and 4 + 3 = 7 which comprised 10, 19 and 10 BRCA1-positive cases, respectively .
The high Gleason score for BRCA1 and BRCA2 positive cases found in the current study was also consistently observed in Castro’s study which indicated aggressive form of prostate cancer for both BRCA1 and BRCA2 with poor clinical outcome . The association of BRCA2 protein expression with Gleason score in this study was also in agreement with the findings in the studies of Tryggvadottir et al. (P = 0.001) and Rabiau et al. (P < 0.012) [19, 24]. Moreover, it has been reported that PCa patients containing BRCA1/2 gene are more likely to have poor survival, recurrence and metastasis [4, 9].
This association could be explained by the fact that PCa with BRCA mutations have abnormal epithelial cells with high proliferative rate and extensive genetic instability which eventually could lead to inefficient DNA repair mechanisms. For example, Fiorentino et al. reported that BRCA1 may help to induce cell cycle arrest to allow for DNA repair . However, some studies have reported the contradicting results. Edwards et al. reported that there was no association between BRCA2 status and grade of prostate cancer (P = 0.071) . The findings obtained from this study particularly the association of Gleason score with BRCA1/2 using IHC detection method should pave for the need of having studies that involve large sample size as well as controls in countries where testing for mutations is challenging due to a number of factors including financial constraints and lack of enough and competent trained individuals to run the tests.
Limitations of the study
Because of using retrospective data, we were not able to obtain data on lymph node metastasis and other distant organs which lead to failure to run association of such important prognostic factors and expression of BRCA1/2 genes. Another limitation of our study was inability to perform molecular testing of mutation of the two genes due to financial constraints.