Prediction of Nipple and Areola Complex Invasion in Breast Cancer Patients. Clinical and Pathological Study of Surgical Specimens

Breast conservation surgery (BCS) and nipple-areola-sparing (NAS) mastectomy have been recognized as two milestones in this period. This study included 60 Egyptian female patients with breast cancer, all of them were subjected to modified radical mastectomy operation. Methods: This study included female patients > 18 years old who have breast cancer with healthy looking non invaded skin of nipple and areola and excluded patients < 18 years old, patients unfit for surger, patients previously subjected to chemo or radiotherapy for breast cancer. We peformed clinical examination of 60 patients with breast cancer. We studied the relevant factors that affect NAC invasion such as patient’s age, menstrual state, family history, tumor size, tumor location (central vs peripheral), tumor to nipple distance, lymphovascular invasion of NAC, lymph node metastasis, histological tumor type, tumor stage, multifocal/multicentric tumors and (ER, PR, HER2) status. Result: In our study, we have shown that NAC invasion is strongly associated with: 1) Nipple retraction as a patient’s complaint; 2) Tumor site; 3) Tumor-nipple-distance ≤ 4 cm; 4) Multifocal/multicentric tumor; 5) Tumor grade (grade III tumors); 6) Positive lymph node invasion; 7) ER and PR re-ceptors negativity; 8) HER2 positivity. This helps in preoperative planning for selecting patients


Introduction
Breast cancer remains the most frequently diagnosed cancer among women. Surgical techniques have evolved from radical mastectomy to less invasive and cosmetically acceptable surgical approach in recent years [1].
Breast conservation surgery (BCS) and nipple-areola-sparing (NAS) mastectomy have been recognized as two milestones in this period [2]. Oncoplastic surgery involves attention to tumor oncology as well as breast aesthetics. It has become popular amongst breast surgeons, combining oncologic principles with plastic surgical techniques [2] [3].
NAS mastectomy entails removal of all breast tissues with preservation of the entire skin of the breast and the nipple-areola complex (NAC) [4].
NAS mastectomy gained great popularity among breast cancer patients, either for treatment or risk reduction. The prophylactic treatment is indicated in women with BRCA1/BRCA2 mutations, personal or family history of breast cancer and severe fibrocystic disease with a strong family history of cancer [5].

Aim of the Work
The aim of this work is to study the clinical and pathological criteria upon which a decision can be made for nipple areola sparing mastectomy in breast cancer patients.

Patients and Methods
This study was carried out in the departments of general surgery and oncology unit, through the period from January 2017 to July 2020.
The study included 60 Egyptian female patients with breast cancer all of them were subjected to modified radical mastectomy operation.  Patients previously subjected to chemo or radiotherapy for breast cancer.

Patient's Selection
 Inflammatory breast cancer cases.
 Patients who had skin changes involving nipple areola complex.

1) History taking:
The history included: a) Personal history: As regard name, age, marital status including number of children, occupation and any special habits. b) Complaint: Painless lump, Painful lump, Pain with no lump, nipple discharge, nipple retraction, Skin changes and axillary lump with its duration. c) Present history: as regard  Assessment of breast mass e.g. onset, course, duration, number, site, size, painful or not, associated with nipple discharge or not, …  Assessment of any axillary swelling(s) e.g. onset, course, duration, number, site, size, …  Assessment of skin and nipple changes e.g. nipple discharge (especially bloody one), nipple erosion, skin nodules, … (refer to exclusion criteria)  Assessment of any symptoms suggestive of metastases e.g. bone-aches, jaundice, hemoptysis, neurological deficit, …  Review of sympoms related to other systems e.g. gastro-intestinal and abdominal symptoms, urological symptoms, genital symptoms, cardio vascular symptoms, respiratory symptoms, nervous system, musculoskeletal system and endocrine system. d) Menstrual history: As regard age at menarche, regularity of the menstrual cycle, age at the first childbearing, age at menopause (for post-menopausal women), and use of contraceptive pills. e) Family history: History of breast cancer especially in relatives (sister, mother or daughter). f) Past history: History of previous operations, allergy and sensitivity, diseases and drugs.
2) Physical examination a) General examination: As regard vital signs (pulse, blood pressure, temperature, and respiration), body built, general appearance and systemic examination of all body systems with special concern looking for signs of metastases. b) Local examination: Local examination of the normal breast and axilla was done first before examination of the affected side.  Examination of any breast mass assessing tenderness, hotness, site, size, shape, surface, number, consistency, edge, relation to underlying structures (fixation to pectoral muscles or chest wall), and relation to overlying skin for evidence of malignant invasion (skin ulceration, peau d'orange, fixity, etc.).  Assessment of the depth of the mass in breast tissue either, superficial or deep.  Examination of the axillary lymph nodes assessing number, size, shape, surface, consistency, matting to each other and fixing underlying structures and which group(s) of lymph node is/are affected.
3) Pre-operative investigations: a) Laboratory investigations:  Complete blood count (CBC).  Bleeding profile (PT, PTT and INR).  Liver function tests (total and direct bilirubin, serum albumin, ALT and AST).  Kidney function tests (blood urea and creatinine).  Random blood sugar.  Hepatitis markers. b) Mammography: All patients were subjected to bilateral soft tissue mammography, where cranio-caudal and oblique views of both breasts were obtained. c) Breast ultrasonography: It was done as a complementary imaging technique to mammography to confirm diagnosis, differentiate cystic swellings from solid ones and help take U/S-guided FNAC from suspicious lesions. Now, in many centers it is superior to mammography and the investigation of choice. d) Fine needle aspiration cytology (FNAC): It was done as an outpatient technique and was done U/S-guided. e) Excisional biopsy: It was done when FNAC was not conclusive to diagnose malignancy. f) Investigations to rule out metastasis:  Chest x-ray.  Pelvi-abdominal U/S. g) ECG and medical fitness for surgery: for patients over 40 years. h) Informed consent for mastectomy was signed by all patients after detailed explanation to them.

Operative Work
After doing all investigations and confirming the stage of the disease and fitness of the patients for surgery, patients were prepared to modified radical mastectomy.

Post-Operative
The specimens sent to the pathology department. Breast specimen were examined by standard hematoxylin and eosin stains under light microscopy. Tissue just underlying nipple-areola complex were examined for evidence of malignancy. All specimens were examined by a single expert pathologist to search for malignancy in the subareolar tissues.
Histopathology Post-operative care: 1) Monitoring of the vital signs of the patients every 6 hours and the amount of the drain every 12 hours.
2) Patients were encouraged for early mobilization from bed after 6 hours and started oral intake. 5) The suction drain was removed when the serous discharge decreased to less than 30 ml in 24 hours.
6) The sutures of the wound were removed from days 10 -12.
7) The patients were referred to the oncology department with the final pathology report to complete their post-operative radiotherapy, chemotherapy and/or hormonal therapy. 8) Follow up of patients for 2 years.

Statistical Analysis
All data were collected, tabulated and statistically analyzed using Statistical package for the social sciences (SPSS). 22.0 for windows (SPSS Inc., Chicago, IL, USA). Table 1: Mean age of breast cancer patients without NAC invasion was non-significantly higher than mean age of breast cancer patients with NAC invasion (Mean: 51.26 vs 49.90 years, p-value = 0.741), also non-significant association between age group and NAC invasion where 26% of patients without NAC invasion was age less than 40 years versus 18.2% in patients with NAC invasion  Table 3 and Table 4). Table 5 and Table 6: 39 out of 50 patients (78%) without NAC invasion had T2 breast cancer versus 10 out of 11 patients (90.9%) with NAC invasion (p = 0.606). There is a significant association between clinical N stage and NAC invasion where 20% of patients without NAC invasion had N3 versus 63.6% of patients with NAC invasion (p-value = 0.014). Mean distance from mass to nipple in patients without NAC invasion was significantly higher than in patients with NAC invasion (Mean: 7.24 vs 3.15 cm, p-value < 0.001), so there's significant association between distance from mass to nipple and NAC invasion. 33 out of 50 masses (66%) without NAC invasion were superficial masses versus 14 out of 20 masses (70%) with NAC invasion (p = 0.748). Table 7 & Figure 1: Optimum cutoff of distance from mass to nipple as a predictor for NAC invasion in breast carcinoma was ≤4 cm. So, 100% of patients with distance > 4 cm, can be expected that they haven't NAC invasion. In other hand no patients with distance > 4 cm, can be expected that they have NAC invasion while 85% of patients with distance ≤ 4 cm, can be expected that they have NAC invasion in other hand 15% of patients with distance ≤ 4 cm, can be expected that they haven't NAC invasion, so we conclude that this cutoff is optimum to exclude NAC invasion (patients with distance > 4 cm) rather than conclude NAC invasion (patients with distance ≤ 4 cm). 32 out of 50 patients (64%) without NAC invasion underwent core biopsy versus 7 out of 11 patients (63.6%) with NAC invasion (p-value = 0.306). 84% of patients without NAC invasion had infiltrating duct carcinoma versus 90.9% of patients with NAC invasion (p-value = 0.624). There is a significant association between histological grade and NAC invasion where 40% of patients without NAC invasion was grade III versus 90.9% of patients with NAC invasion (p-value = 0.002). 60% of patients without NAC invasion were node positive versus 100% of patients with NAC invasion (p-value = 0.011). Number of positive lymph node in patients with NAC invasion was significantly higher than patients without NAC invasion (Mean: 13.72 vs 4.98, p-value = 0.001). There is non-significant difference between patients without and with NAC invasion as regard number of dissected lymph nodes (Mean: 17.98 vs 18.45, p-value = 0.707) ( Table 8). 39 out of 50 patients (78%) without NAC invasion had pT2 breast cancer versus 10 out of 11 Open Journal of Pathology        (Table 9 and Table 10, Figures 2-4).

Discussion
Oncoplastic surgery has become a key aspect of breast cancer treatment, as it considers both oncological and cosmetic outcomes. The idea of sparing the skin of the breast is to facilitate the immediate breast reconstruction [6]. The greatest advantage of nipple areola sparing mastectomy compared to skin sparing mastectomy is the immediate cosmetic result with no need for nipple reconstruction later on. The need to remove the nipple areola complex is based on the concern of occult neoplastic involvement [7].
The concept of the nipple areola sparing mastectomy was popularized by Freeman in the 1960's and 1970's, the procedure was referred to as a subcutaneous mastectomy. The use of NAS mastectomy for the treatment of breast cancer grew in the 1970's and 1980's. The attempts are made to preserve the nipple areola complex since the first paper done by Gerber et al in 2003 [8].
In this study, we evaluated the oncological safety for nipple areola sparing mastectomy procedure by searching for the factors that predict the presence of nipple areola complex invasion. This done by history taking and clinical examination of 60 Egyptian female patients with breast cancer. Then we searched for the presence of the occult malignant cells in the subareolar tissue in the breast specimens of the standard modified radical mastectomy (done by single expert pathologist) for the 60 patients.
In this study, NAC invasion was noted in 9 out of 60 mastectomy specimens. So, the incidence occult NAC invasion was 15%. This rate indicates that even patients who had clinically normal-appearing NAC should be carefully selected for NAS mastectomy. This falls in the same range reported by Gomez et al., (2014) who reported that the incidence of nipple involvement (NI) ranging from 0% to 58% [9].
In our study, the mean age of breast cancer patients without NAC invasion was non-significantly higher than mean age of breast cancer patients with NAC invasion (Mean: 51.26 vs 49.90 years, p-value = 0.741) There was statistically non-significant association between age groups and NAC invasion in which (p-value = 0.121) and this coincides with Zhang et al., (2015) [10].
In this study, 13 out of 50 patients (26%) without NAC invasion had positive family history of breast cancer versus 3 out of 11 patients (27.3%) with NAC invasion. There was statistically non-significant association between family history and NAC invasion in which (p-value = 1.000) and this differs with other studies that reported that up to 5% -10% of breast cancer are due to genetic mutations [12].
In this study, the most common patient's complaints were painless lump (58.3%), painful lump (26.7%) and nipple retraction (8.3%). There was a statistically significant association between nipple retraction and NAC invasion where 2% of patients without NAC invasion complaint from nipple retraction versus 36.7% of patients with NAC invasion (p-value = 0.003).
In this study, there was no statistically significant association between breast size (measured by the bra size) and NAC invasion as mean breast size in patients without NAC invasion was non-significantly lower than mean breast size in patients with NAC invasion (p-value = 0.186) and this coincides with Abou Nagah and El-Sabaa, (2012) [11].
In this study, there was a statistically highly significant association between focality and NAC invasion where none of patients without NAC invasion had multifocal breast cancer versus 81.8% of patients with NAC invasion (p-value < 0.001). This coincides with Zhang et al., (2015) [10]. Wang et al., (2012) [16]. There was a highly significant association between tumor site (centrally located tumors) and NAC invasion, as tumors located in the central areas are more likely to have nipple invasion (NI) than peripheral areas (p-value < 0.001).
In this study, there was no statistically significant association between tumor size neither clinically assisted or US assisted and NAC invasion (p-value = 0.401) (p-value = 0.838). This matches with Lawrence et al., (2018) whose results failed to show any statistically significant association between tumor size and occult nipple involvement, and this differ with Zhang et al., (2015) who reported that the risk of NAC invasion increased significantly in patients with larger tumors [10] [18].
In this study, there was statistically highly significant association between the tumor nipple distance and NAC invasion (p-value < 0.001). This reported by Weidong et al., (2017) [13].
In our study, the optimum cutoff of distance from mass to nipple as a predictor for NAC invasion in breast carcinoma was ≤4 cm. So, all of patients with distance > 4 cm can be expected that they have not NAC invasion. So, we can conclude that a distance > 4 cm is requested for NAS mastectomy. This coincides with Vlajcic et al., (2016) who found that the NAC could be safely preserved with tumor-to-nipple distance > 4 cm and does not match with Zhang et al., (2015) that suggested distance of 2.5 cm from the tumor to the nipple is required to reduce the risk of nipple involvement [10] [19].
In our study, there was no statistically significant association between histological type of the tumor and NAC invasion (p-value = 0.624). This agrees with Zhang et al., (2015) [10].
In this study, there was a statistically significant association between histological grade of the tumor and NAC invasion, where 40% of patients without NAC invasion was grade III versus 90.9% of patients with NAC invasion (p-value = 0.002). This coincides with Eisenberg et al., (2014), but differs with Simmons et al., (2019) who found no significant association in rates of NAC invasion and tumor grades [17] [20].
In this study, there was a statistically significant association between positive lymph node invasion and nipple invasion as 60% of patients without NAC invasion were node positive versus 100% of patients with NAC invasion (p-value = 0.011). Number of positive lymph node in patients with NAC invasion was significantly higher than in patients without NAC invasion (p-value = 0.001). This coincides with study who suggested that NAC invasion was significantly increased in patients with lymph node metastasis and differs with Simmons et al., (2019) their study did not show a higher incidence of NAC invasion in lymph node positive group [17] [21].
In our study, there was a statistically highly significant association between estrogen receptor negativity and NAC invasion where 18% of patients without NAC invasion had negative estrogen receptor versus 90.9% of patients with NAC invasion (p-value < 0.001). Also, there was a statistically significant associ-ation between progesterone receptor negativity and NAC invasion where 24% of patients without NAC invasion had negative progesterone receptor versus 72.7% of patients with NAC invasion (p-value = 0.004). This agrees with Zhang et al., (2015) who reported that the trend toward higher nipple involvement rate was found in patients with negative ER status and negative PR status [10].
In our study, there was a statistically highly significant association between HER2 positivity and NAC invasion where 8% of patients without NAC invasion had positive HER2 versus 90.9% of patients with NAC invasion (p-value < 0.001). This agrees with Zhang et al., (2015) who suggested that patients with positive (HER2) have a higher rate of NAC invasion [10].
In our study, we found that the tumor site and tumor nipple distance affect the prediction of NAC invasion while the age, menstrual state, breast size and tumor type did not directly affect the prediction of presence of subareolar ma-

Conclusions
According to our study, the ideal patients for NAS mastectomy should have these criteria: 1) Clinically normal nipple areola complex.
2) Distance from the tumor to the nipple is >4 cm. So, we recommend taking these criteria in preoperative decision making for NAS mastectomy.