|Year : 2021 | Volume
| Issue : 2 | Page : 266-272
p53 Expression in Breast Carcinoma and Its Association with Tumor Aggressiveness
Sushree Swagata Dash, Alaka Sahu, Anupa Toppo
Department of Pathology, VSSIMSAR, Sambalpur, Odisha, India
|Date of Submission||08-Jan-2021|
|Date of Decision||27-Feb-2021|
|Date of Acceptance||18-Apr-2021|
|Date of Web Publication||18-Oct-2021|
Dr. Alaka Sahu
Department of Pathology, VSSIMSAR, Burla, Sambalpur - 768 017, Odisha
Source of Support: None, Conflict of Interest: None
Background: p53 mutations are the most frequent genetic alterations in breast cancer, observed in 30% of breast carcinomas. Their distribution is highly linked to molecular subtypes. Aim: The aim of the study was to establish an association between p53expression with various tumor variables such as age, histological grade, molecular subtypes, ki67, and Nottingham prognostic index (NPI). Materials and Methods: This was a cross-sectional analysis in VSSIMSAR, Burla, Odisha, India, from November 2015 to October 2017. Immunohistochemical study was done using DAKO primary antibody for estrogen receptors, progesterone receptors, human epidermal growth factor receptor (HER2/neu), Ki-67, and p53. Results: A total number of 62 cases were studied. p53 was positive in 10 (55.6%) cases of T2 tumors, 15 (60%) cases of T3 tumors, and 13 (76.5%) cases of T4 tumors. Among 26 basal-like tumors, 16 (61.5%) were p53 positive. In 12 HER2 enriched tumors, 10 (83.3%) were p53 positive, and in 8 luminal B tumors, only 2 (25%) cases came out to be p53 positive. p53 expression has a very significant association with lymph node metastasis (P < 0.01). p53 positivity increases with higher histological grades, and its reactivity is significantly associated with Ki-67 expression (P < 0.05) and NPI (P < 0.01). Conclusion: p53 association is mostly associated with more adverse phenotype and worse overall survival. There is an increased interest in p53 based therapy.
Keywords: Breast cancer, molecular subtype, p53 mutation
|How to cite this article:|
Dash SS, Sahu A, Toppo A. p53 Expression in Breast Carcinoma and Its Association with Tumor Aggressiveness. J Datta Meghe Inst Med Sci Univ 2021;16:266-72
| Introduction|| |
Breast carcinoma is the most common cancer and the leading cause of cancer death in women worldwide, including India., In India, for the year 2018, an estimated 162,468 women were newly detected with breast cancer, and 87,090 women died of breast cancer. Breast carcinoma is a heterogeneous disease both clinically and biologically. It can be classified according to conventional methods such as histopathological/morphological appearance, degree of differentiation (grade), the extent of tumor spread (stage), and also based on gene expression profiling and molecular subtyping. Every approach tries to stratify breast cancer by risk and prognosis. According to the World Health Organization Classification of Tumours of the Breast (4th edition), invasive cancers that fail to exhibit sufficient characteristics to achieve classification as a specific histological type (e.g., lobular, tubular, etc.) are classified as invasive carcinoma of no special type (NST). It is the most common type, comprising 40%–75% of cases.
The three-tiered (grade 1-3) grading scale also called combined histological grade (the Nottingham score system or the Elston-Ellis modification of the Scarff-Bloom-Richardson grading system) is also used to predict the prognosis of a malignant neoplasm. This method is based on the complex evaluation and semiquantitative scoring of “tubule formation” (glandular differentiation) and nuclear morphology combined with the mitotic activity of the tumor.
The most widely used staging system is the TNM system, which classifies cancers according to their local size and some features of advanced disease (T category), lymph node involvement (N category), and distant spread (M category). Both clinical, as well as pathological staging, is used in breast cancer patients. A combination of detailed clinical examination, radiological imaging, and pathological examination are performed before the initiation of neoadjuvant chemotherapy. The determination of the stage before and after treatment provides important prognostic information.
The more recent molecular subtypes of breast cancer have been identified using gene expression profiling. However, the cost and technical complexities limited the use of gene expression profiling in daily clinical practice. Immunohistochemistry (IHC) based surrogate molecular classification has been advocated, which is endorsed by the St Gallen Consensus Conference in planning individual patient treatment. Immunohistochemical analysis of tumors based on estrogen receptors (ER), progesterone receptors (PR), human epidermal growth factor receptor (HER2), and Ki-67 status is used to identify different subtypes: (1) Luminal A-like (ER+, PR ≥20%, and Ki67 <20%), (2) luminal B-like (ER+, PR <20%, and/or HER2+, and Ki-67 ≥20%), (3) HER2-overexpression (ER-, PR-, and HER2+), and (4 basal-like (triple-negative: ER-, PR-, and HER2-). This method is easier, cost-effective and provides similar results for molecular subtypes.
The p53 gene, first described in 1979, and initially believed to be an oncogene, was the first tumor suppressor gene to be identified. p53 functions to eliminate and inhibit the proliferation of abnormal cells, thereby preventing neoplastic development. Abrogation of the negative growth regulatory functions of p53 occurs in many, perhaps all, human tumors. The p53 signaling pathway is in “standby” mode under normal cellular conditions. Activation occurs in response to cellular stresses, and several independent pathways of p53 activation have been identified that appear to be dependent on distinct upstream regulatory kinases. Loss of heterozygosity in the p53 gene was shown to be a common event in primary breast carcinomas.
The present study was conducted to evaluate p53 overexpression in known cases of breast carcinoma and to provide prognostic and therapeutic information to clinicians. The IHC for ER, PR, HER2/neu, and Ki-67 were applied to classify breast cancer into molecular subgroups, which further help the clinicians for proper management.
| Materials and Methods|| |
This study was conducted in the department of pathology at VSS Institute of Medical Science and Research, Burla, India, from November 2015 to October 2017. All histologically proven cases of invasive breast carcinoma cases irrespective of age were included in the study. Trucut biopsy specimens and samples with extensive necrosis were excluded from the study. Ultimately 62 cases fulfilling the above criteria were kept under study. Following gross and microscopic examinations, the slides with the most representative tumor tissue were selected for immunohistochemical studies using DAKO primary antibodies such as ER, PR, HER2/neu, Ki67, and p53. The expression of ER and PR were studied using Allred or quick score system. The expression of HER2/neu was studied according to ASCO and CAP guidelines 2013. p53 staining was interpreted as positive when nuclear staining was ≥10% of tumor cells and negative when it was <10%. Correlation between p53 and other clinicopathological prognostic factors was analyzed using the Chi-square test. The results were considered statistically significant if the P value was < 0.05.
| Results|| |
The age of presentation in our study ranged from 34 to 75 years, with a mean age of 51 years. The majority of the patients fall into >40 years of age group (85.5%) and are mostly postmenopausal (62.9%) [Figure 1].
The maximum number of tumors was >5 cm of size. We observed 84% of cases of invasive ductal carcinoma (IDC) breast–NST not otherwise specified (NOS). Other histological types include lobular, papillary, medullary, sebaceous, apocrine carcinoma, and small cell carcinoma [Figure 2].
|Figure 2: Distribution of cases according to tumor size and histologic type|
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According to the Scarff-Bloom-Richardson grading system, the majority were grade II tumors (50%) followed by grade III (37.1%) and grade I tumors (12.9%). 77.4% of cases showed lymph node metastasis [Figure 3].
Figure 3: Distribution of cases according to tumor grade and LN-metastasis
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Based on molecular subtypes, we found the maximum number of cases falling into triple-negative/basal-like [Figure 4].
Immunohistochemistry by p53 stain
IHC was done on a total of 62 cases. Total 61.3% of cases showed p53 positivity, while 38.7% were negative [Figure 5]. Various clinicopathological parameters were studied in association with p53 staining [Table 1].
|Table 1: p53 association with various demographic and histopathological parameters (total=62)|
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p53 status in relation to age and menstrual status
We found a number of p53 positive patients as ≥40 years of age and postmenopausal.
p53 status in relation to tumor stage
Table 1 shows p53 negative in all the T1 tumors. Among 18 T2 tumors, p53 was positive in 55.6% of cases. In 25 T3 tumors, p53 positivity was found in 60% cases. Finally, among 17 T4 tumors, p53 came out to be positive in 76.5% of cases.
p53 status in relation to lymph node metastasis
p53 was positive in 28.6% of 14 N0 tumors. Among 42 N1 tumors, 66.7% were p53 positive. All the 6 N2 tumors came out to be p53 positive. In this case, p53 expression has got highly significant association with lymph node metastasis (P < 0.01).
p53 status in relation to histologic grades
p53 was positive in exactly half of the 8 grade I tumors. In 31 grade II tumors 54.8% were p53 positive. 73.9% of 23 Grade III tumors were p53 positive.
p53 status in relation to Ki 67 expression
Among those tumors which had a high Ki 67 index (exactly half of them), p53 became positive in 74.2% of them. But those tumors which had a low Ki 67 index showed 48.4% of p53 positivity. Using Chi-square test, it was found that p53 reactivity is significantly associated with Ki 67 expression (P < 0.05).
p53 status in relation to Nottingham prognostic index
In breast cancer patients with good Nottingham prognostic index (NPI) (8 cases), all tumors were p53 negative. In patients with moderate NPI (43 cases), 62.8% were p53 positive. Among patients with poor NPI (11 cases), all were p53 positive. In this case, p53 expression was found to be highly significant (P < 0.01).
p53 status in relation to molecular subtypes
Table 1 shows p53 status in different molecular subtypes. Among 26 basal-like tumors, 61.5% were p53 positive. In 12 HER2 enriched tumors, 83.3% were p53 positive. 62.5% of the 16 Luminal-A tumors were p53 positive, and only 25% of 8 Luminal-B tumors came out to be p53 positive.
| Discussion|| |
Breast cancer comprises a heterogeneous group of tumors that vary significantly in their biology, presentation, and response to treatment. There are certain tumor variables frequently used for the assessment of breast cancer in terms of prognosis and therapy. Lymph node status, tumor size, and histological grade are now supplemented with measurements of steroid hormone receptors, HER2/neu receptor status, proliferation index, tumor suppressor genes, growth factors, oncogenes, and oncogene products. Tumor size and axillary lymph node status are the most important classical variables to predict breast cancer prognosis.
It is well documented that the advancement in age increases the risk of breast cancer, although there is evidence that Indian women are more likely to develop breast cancer at an earlier age (in their thirties and forties) than their Western counterparts. In our study, we found a majority of the patients as postmenopausal (62.9%).
Tumor size is one of the most powerful prognostic markers in breast cancer. In node-negative breast cancer cases, the single most important prognostic factor is tumor size and one of the strongest predictors for dissemination and rate of relapse in these cases. However, axillary node status is the single most important prognostic factor for patients with early breast cancer. Many studies had shown that treatment outcome is very poor in cases, which had axillary lymph node metastasis as compared to node-negative breast cancer cases. In the present study, maximum numbers of tumors are of size >5 cm, i.e. T3 stage, which is much higher as compared to the studies of Gupta et al. and Raina et al. This may be due to a late presentation to clinics due to a lack of awareness among patients in this part of the country. The absence of symptoms such as pain or nipple discharge may also contribute to this outcome. We observed 84% of cases to be of IDC-NOS type. Other types include lobular, medullary, papillary, sebaceous, apocrine, and small cell carcinoma, comprising the remaining 16% of cases. This finding is comparable to the findings of Yang et al. (77.3%) and Gupta et al. (86.1%),
Tumor grade is the description of a tumor based on how abnormal the tumor cells and tumor tissue look under a microscope and indicates how quickly the tumor is likely to grow and spread. It's a reflection of the degree of malignancy in morphology. The present study shows that the maximum cases of grade II tumors (50%) are similar to those of Zafrani et al. and Peiró et al. Lymph node metastases were seen in 67.8% of cases in our study, which is comparable to the study of Yang et al.(74.2%).
From a therapeutic point of view, breast cancer is divided into molecular/intrinsic subtypes based on hormone receptors (ER, PR) and HER2/neu status. Accordingly, there are four types: Luminal A, luminal B, HER2/neu enriched, and basal-like (Triple-negative). All these have targeted therapies except triple-negative breast cancer (TNBC), which has only got chemotherapy as its mainstay of treatment and therefore it has the worst prognosis among all. It is documented that Luminal A is the most common and Basal-like (TNBC) is the least common intrinsic subtype (10%–15% of all breast cancers). In our study, we found Basal-like (TNBC) as the most common type (41.9%) and Luminal B being the least common type (12.9%).
Immunohistochemical evaluation of p53 had been done to study its association with different tumor variables. p53 is a product of tumor suppressor gene TP53, which is not detected in normal cells due to its short half-life; but if mutated, it can be detected by IHC or other methods. It is established that p53 expression is associated with a more aggressive breast cancer phenotype in terms of higher tumor grade and stage, lymph node metastases, basal-like subtype, high proliferation index, and poor prognostic index. In the present study, p53 positive cases (expression in >10% of tumor nuclei) accounted for 38 out of 62 cases i.e., 61.3% [Figure 5], which have a higher value as compared to the study of Gammon et al.(44.4%), but Gupta et al. found a much higher percentage of p53 positivity (88.9%).
Table 1 shows the association of p53 with different demographic and histopathological parameters. There was no significant association between p53 status and different age groups in our study. Irrespective of the age of breast cancer patients, p53 was expressive in more than half of the cases. p53 status was not significantly associated with menstrual status as well and was almost equally expressive in both pre- and postmenopausal patients.
p53 was found to be negative in all the T1 tumors. Among T4 tumors, p53 came out to be positive in 76.5% of cases. Percentage of p53 expression increases with an increase in tumor size. Although the maximum number of cases fall within the category T3 with tumor size >5 cm, there was no significant association between p53 and tumor size (P > 0.05) as in the studies of Sekar et al., and Song et al.
With higher nodal metastasis, p53 activity increases. A maximum number of p53 positive cases were seen in the N1 stage of our study. Among N1 tumors, 66.7% were p53 positive. All the N2 tumors came out to be p53 positive. We got a highly significant association between p53 and lymph node status (P < 0.01), similar to the study of Gupta et al. In contrast, the study conducted by Song et al. revealed that lymph node metastasis was more in p53 nonexpressors than in p53 overexpressors.
Expression of p53 increases with higher histologic grade. 73.9% of 23 Grade III tumors were p53 positive. There was an equal distribution of Grade II and Grade III tumors among p53 positive cases in this study, but no significant association was noted (P > 0.05) similar to the study of Song et al. However, Gupta et al. and Yamashita et al. found this correlation to be significant.
Among those tumors with a high Ki-67 index, p53 was positive for 74.2% of the tumors. However, those tumors with low Ki-67 indexes showed 48.4% of p53 positivity. A significant association was found between p53 expression and Ki-67 index (P < 0.05). Our study shows that with increased p53 expression, there is a higher proliferative activity of tumor (Ki-67 >14%). Similar findings were noted in the study of Madani et al.
p53 shows a highly significant association with NPI in this study (P < 0.01). Cases with good NPI scores revealed no p53 activity, whereas all the p53 positive cases were distributed among moderate and poor NPI scores. This result was consistent with the study of Kurshumliu et al., the findings of which suggest a significant association between p53 expression and NPI.
Among basal-like tumors, 61.5% were p53 positive. In HER2 enriched tumors 83.4% and among Luminal-A tumors 62.5% were p53 positive. Only 25% of Luminal-B tumors came out to be p53 positive. So, p53 had no significant association with molecular subtypes.
In our study, we had a maximum number of IDC-NOS cases and very few other histopathological types; therefore, we could not find any significant association of p53 with the histological type of tumor. However, p53 expression distribution by histological type has been shown to indicate the lack of any preference for p53 positivity and/or negativity in the case of ductal carcinoma, negativity in lobular carcinoma, and strong positivity in medullary carcinoma and is supported by the study of Sirvent et al. P53 activity in Grade III was higher as compared to grade II IDC-NOC tumors, negative in apocrine tumor, and positive in aggressive tumors such as sebaceous carcinoma [Figure 6], [Figure 7], [Figure 8], [Figure 9], [Figure 10], [Figure 11].
|Figure 6: (a) Malignant ductal epithelial cells in sheets and few tubules: Invasive duct carcinoma – no special type -Grade 2 (H and E, ×100). (b) p53 positive nuclear staining × 400. (c) Human epidermal growth factor receptor positive ×400. (d) Ki 67 (<14%) ×400|
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|Figure 7: (a) Malignant ductal epithelial cells with no tubules, marked nuclear pleomorphism, and increased mitosis: Invasive duct carcinoma – no special type -grade 3 (H and E, ×100). (b) p53 positive nuclear staining ×400. (c) Human epidermal growth factor receptor positive ×400. (d) Ki-67 (>14%) ×400'|
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|Figure 8: (a) Microphotograph showing tumor cells in Indian file pattern: Lobular carcinoma of the breast (H and E, ×400). (b) P53 negative ×400|
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|Figure 9: (a) Microphotograph showing tumor cells in syncytial pattern with lymphoplasmacytic infiltration: Medullary carcinoma of the breast (H and E, ×400). (b) P53 positive nuclear staining ×100|
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|Figure 10: (a) Microphotograph showing apocrine cells (round to polygonal cells with eosinophilic granular cytoplasm and centrally placed round vesicular nuclei) in sheets: Apocrine carcinoma of the breast (H and E, ×400). (b) P53 negative ×100|
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|Figure 11: (a) Microphotograph showing tumor cells with foamy cytoplasm, vesicular nuclei with prominent nucleoli: Sebaceous carcinoma of the breast (H and E, ×400). (b) P53 positive nuclear staining ×400|
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| Conclusions|| |
TP53 mutations are the most frequent genetic mutations in breast carcinoma found in 30% of cases. Their distribution is highly linked to molecular subtypes with the highest expression in basal-like tumors. It is established that p53 expression is associated with more adverse phenotypes. In our study, we also tried to prove the same in cases of breast carcinoma, and it was found that p53 expression is mostly associated with the aggressiveness of cancer. It is studied that p53 accumulation is associated with resistance to endocrine therapy. Our study can therefore assist clinicians in selecting patients for appropriate treatment.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
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