Evaluation of HER 2-overexpression Status in Breast Cancer Patients: Results From a Breast Cancer Registry

Background: The expression level of HER2 gene is low in normal breast tissue, but levels of these receptors are higher in half of cases of breast cancer. Different expression levels of HER2 gene in normal and malignant cells have made this gene an ideal biomarker for therapeutic purposes. In this study, the extent of HER2 overexpression and its relationship with the age and the occurrence of metastasis were evaluated in breast cancer patients. Methods: In this cross-sectional study, 62 patients with breast cancer were evaluated at oncology clinics in Ilam province. Clinical examination for metastasis, examination of tissue samples for HER2 gene expression, and information related to variables were recorded in a breast cancer registry. The obtained data were analyzed using SPSS 20 statistical software by appropriate statistical tests. Results: The mean age of the participants was 48.37 ± 10.98, and 98.4% were women. The prevalence of increased HER2 gene expression was 37.1% in patients. There was an inverse relationship between patients' age and HER2 positivity (P value = 0.02). The chance of metastasis was 9 times higher (OR = 9.82) in cancer patients who had the HER2 gene expression Conclusion: In Ilam province, the prevalence of HER2 positivity in breast cancer patients is almost similar to other parts of the country and is associated with the occurrence of metastasis and low age of breast cancer patients.


Background
Breast cancer is one of the most common types of malignancy in the age group of 45-55 years and the second leading cause of cancer death after lung cancer (1). Annually, more than 1,100,000 women are affected by breast cancer in the world (2)(3)(4). Many factors, such as proliferative breast lesions, radiation history, menstrual status, and pregnancy, have been suggested as risk factors for breast cancer. The status of some biomarkers (Ki67 HER2, PR, and ER) is effective in the prognosis and treatment of breast cancer (5).
One of the most important biomarkers is HER2 oncogene, which is a part of the message path (Signal Transduction Pathway). This protein is involved in the cell growth and plays a key role in the shift towards malignancy (6,7). Different expression levels of HER2 gene in normal and malignant cells have made HER2 to act as an ideal biomarker for therapeutic purposes (3,8). There are different methods to evaluate the expression of HER2 gene, such as measuring IHC of HER2 receptor, nding duplication of HER2 gene using FISH or PCR methods, and examining the extracellular amount of HER2 released into the blood by ELISA (9). Among these, the IHC method is superior to the other ones due to the ease of operation and low price.
An increased focus on quality indicators and the use of clinical registries for breast cancer in real world studies have shown higher compliance to recommend the therapy and better survival.
The HER2 gene is important in the prognosis and treatment of patients with breast cancer, which is highly prevalent in Iran and the world. Therefore, because no study has been done on the overexpression of HER2 gene in Ilam province, this study aims to determine the rate of HER2 overexpression in breast cancer patients and the relationship between the expression of this gene and the rate of metastatic patients with breast cancer.

Materials And Methods
This analytical-descriptive study was performed with focus on the use of clinical registries for breast cancer in 2020.
In order to carry out this research, the necessary permission was obtained from the Vice Chancellor for Inclusion criteria were patient satisfaction, con rmed breast cancer in the pathology report, access to tissue samples, and absence of malignancy in other organs of the body. Exclusion criteria were patient dissatisfaction at each stage of the study and destroyed tissue samples. Tumor grade was statistically analyzed based on the age, histopathological diagnosis, tumor size, and lymph node involvement. Data collected from patients and those of examined tissue samples were analyzed by SPSS software version 20 at a signi cance level of P < 0.05. It should be noted that all ethical standards were observed in this study based on the Hlinsky statement. Besides, patient information was maintained in a con dential manner, and the results and nal information were published in the groups.
According to Table 1, most of the studied breast cancer patients had undergraduate and postgraduate education (33.9%), the majority were married (93.5%), and were mostly housewives (82.3%).
In this study, one of the patients did not report smoking, alcohol, or drug use. The prevalence of increased HER2 gene expression was 37.1% in patients. There was a direct relationship between the occurrence of metastasis and HER2 positivity and age (P value = 0.04). There was an inverse relationship between patients and HER2 positivity (P value = 0.01). The prevalence rates of ER and PR receptors in patients were 64.5% and 58.1%, respectively.
Metastases were observed in 83% of patients that presented the expression of her2 gene (Table 2).
Only the HER2 variable was signi cant among the age, PR, ER, and HER2 variables included into the model.
The chance of metastasis in cancer patients who showed the expression of HER2 gene was 9 times higher (OR=9.82) than that in HER2-negative patients (Table 3).

Discussion
Due to the increasing prevalence of breast cancer and the need for a special attention, this study examined one of the proto-oncogenes associated with this disease.
Overall, 37.1% of the breast cancer patients in this study were positive for the HER2 gene expression, which is consistent with the results of a study conducted in Tehran by Kadivar et al. (10).
However, 23.3% (11) in Isfahan, 32% (12) in Gilan, 13.6% (13) in Arak, and 30.8% (14) were reported in other studies by different researchers. It seems that the reason for the difference in the prevalence obtained in our study and the above studies is due to three factors of genetic differences in different races, different living environments, and, consequently, different levels of exposure to carcinogens and mutants and differences in how. It is clear that any report of the severity of HER2 overexpression in tissue samples depends on the operator and this may explain some of the differences in the prevalence rates in different studies.
The relationship between patients' ages was examined in the present study. Breast cancer was also treated with the HER2 gene overexpression. According to the results, the rate of HER2 overexpression was directly related to the ages of the patients, which is similar to that of a study by Martinez et al. They reported that increased expression of the HER2 gene in patients under 50 years of age was more than that of patients over 50 years of age. In a study by Mirzaei et al.(24) in Tehran, however, no signi cant relationships were found between the HER2 gene positivity and the ages of patients. This difference in the results can be due to the selection of patients in the mentioned studies, where only patients with In ltrating Ductal Carcinoma were present, while our study consisted of patients with all types of breast cancer.
In the present study, the positive association of the HER2 gene with the occurrence of metastasis was evaluated in the patients. Our study showed that the chance of metastasis in cancer patients who showed the HER2 gene expression was 9 times higher (OR = 9.82) than that in HER2-negative patients.
According to ndings of Khudabakhshi et al. (25), the HER2 gene overexpression was directly related to the occurrence of metastasis in patients with breast cancer. In a study by Naja et al., the chance of metastasis or death in HER2-positive patients was 2.64 times higher than that in HER2-negative patients, and more metastasis was found in breast cancer patients who were HER2 positive.
Findings of Mirzaei et al. showed that HER2 positive patients were exposed to higher-grade tumors. Despite the above three studies that con rmed the results of the present study, Naini et al. observed no relationships between the HER2 gene hypermethylation, tumor size, and grade with the disease stage. This can be caused by a low sample size (20 patients) in the latter study and the evaluation of only one type of breast cancer (invasive ductal carcinoma). In the present study, 62 patients were evaluated in terms of all types of breast cancer.
In this study, the rates of PR and ER receptors were examined in patients, 64.5% and 58.1% of which were ER and PR positive, respectively. The reported prevalence of these receptors were respectively 68.8% and 63.8% (Mirzaei et al.), 72.4% and 64.8% (Semnani et al.,(14), 46.6% and 43.8% (Moradi et al.), 70% and 72.6% (Kuzhan et al., (15), and 75% and 59.5% (Khabaz). Accordingly, the rates obtained in most studies are similar to those of the present study, and the differences may be due to genetic and environmental factors.
Due to the high prevalence of increased HER2 gene expression in patients with breast cancer, its association with the occurrence of metastasis, and its special importance, future studies are recommended to investigate the effectiveness of drugs affecting this receptor.
The diagnosis of the HER2 gene overexpression through tissue sample examination is invasive and costly, on the one hand, and the diagnosis of this disorder is of vital importance in breast cancer patients, on the other hand. It is, therefore, recommended to nd alternative methods to study tissue samples, such as examining the serum level of HER2, and comparing the sensitivity and speci city of these methods with the current method (histological examination).

Conclusion
According to the results of this study, the rate of increase in the HER2 gene expression is 37.1% in patients with breast cancer in Ilam province. Besides, the positiveness of this gene is associated with breast cancer at a younger age and the occurrence of metastasis in these patients. It is a weakening factor in the prognosis in these patients.