To our knowledge, this study is the first to investigate the relationship between hormone receptor status (ER, PR HER2) and different levels of BIF-1 gene expression in patients with breast cancer. There was no statistically significant relationship between BIF-1 gene expression and estrogen hormone receptor (P> 0.05). Additionally, the relationship between BIF-1 gene expression and progesterone hormone receptor was not statistically significant (P> 0.05). Our findings indicate that the expression in BIF-1 was increased in patients who have at least one hormone receptor. However, in patients with triple-negative hormone receptors, expression was reduced. There was no statistically significant relationship between BIF-1 gene expression and all three hormone receptors (ER, PR, HER2) (P> 0.05).
Alternatively, there was no statistically significant relationship between BIF-1 gene expression in patients with breast cancer (ductal, lobular and ductal & lobular carcinoma) (p> 0.05).
Additionally, there was no statistically significant relationship between BIF-1 gene expression and disease stage (p> 0.05).
Regarding tumor size, 32 patients (64%) had a tumor equal to or more than two centimeters, and 14 patients (28%) had a tumor size less than two centimeters.
According to the results of linear regression, there was no significant relationship between gene expression and tumor size in breast cancer patients (P> 0.05).
Furthermore, BIF-1 can also act as a tumor suppressor because of its role in regulating the BAX gene. BIF-1 accelerates BAX degradation directly by binding to BAX and enhancing apoptosis induction kinetics in response to innate apoptotic signals, thereby increasing the permeability of the mitochondrial outer membrane.
Previous studies correspond with our findings, For example, Cuddeback et al., reported that the BIF-1 protein was silent in 17% (192.33 patients) of all prostate cancer patients. These findings indicate the activity of tumor suppression and pro-apoptotic BIF-1 (22).
Impaired expression of BIF-1 in cancer cells, compared to adjacent healthy tissue, has been observed in various types of cancer, including colorectal cancer (23), prostate cancer (24), pancreatic cancer (25), invasive bladder cancer (26), and gastric cancer (27).
In another study, Coppola et al. found that BIF-1 lacked expression in approximately 45% of patients with malignant pancreatic cancer, but had a high level of expression in patients with benign pancreatic cancer (25).
Similarly, Fan et al., observed that patients with high BIF-1 expression compared to patients with low BIF-1 gene expression, had a shorter survival time, Their study correlates BIF-1 expression to survival time (28).
Takahashi et al. showed that suppression of the BIF-1 gene in mice promoted tumour progression which corresponds with our study’s findings. In this study, it was observed that BIF-1 decreased in expression in patients with tumor size equal to or more than two centimeters and increased in expression in patients with tumor size less than two centimeters. This result suggests the induction function of BIF-1 apoptosis (29).
Unfortunately, few studies have been conducted to quantitatively evaluate the expression of BIF-1. Some studies of BIF-1 gene activity, have been inconsistent. In addition, several studies have compared the expression of BIF-1 in patients' tumor cell lines with that of the control group (30). We contend that this is not a tenable measure for evaluating both tumor tissue of patients and healthy tissue samples. It should be noted that normal tissue (adjacent to the tested tumor) was confirmed by our results.
Current information in molecular markers is also inadequate when evaluating treatment response in breast cancer patients.
Consequently, it is time to develop a reference method to evaluate BIF-1 gene expression. Second, investigation of the role of BIF-1 gene expression in the development of various cancers is necessary. Next, differences in research results may be due to dissimilar methods used to study gene expression. Since various researchers have pointed to this aspect, a reference method for the expression of BIF-1 and its role in the growth of various cancers needs to be developed.
Lastly, some studies examining BIF-1 gene expression in breast cancer have been limited to breast cancer cell lines and normal cell lines. Therefore, in the present study, the expression of BIF-1 gene in tumor and normal human tissue was investigated. It is well known that gene expression in clinical specimens are lower than cell lines.
The lack of a suitable quantitative method to determine the exact amount of gene expression is an important issue that may lead to difficulties with data interpretation. The major methods of gene expression analysis are: Western blotting, immunohistochemistry, flow cytometry, RT PCR, and Real Time RT-PCR. Therefore, given the high sensitivity of the Real Time RT-PCR method, as well as, the ability of the RNA and cDNA of cells to be stored and reused, it also appears to be faster and more efficient than flow cytometry. Real time RT-PCR is an optimal way in evaluating BIF-1 gene at the mRNA level.
Other limitations of our study include the high cost of consumables for molecular analysis, the small number of laboratories equipped with the necessary devices and the unwillingness of some hospitals to cooperate in providing the necessary samples, which led to a limit on the number of patients to perform the necessary tests.