Introduction

Breast cancer is an important cause of cancer mortality in Indian women [1]. Biologic markers like histological grade of the tumor, Estrogen and progesterone hormone receptor expression, HER2 overexpression, and/or amplification and genomic panels are the important predictors of survival in breast carcinoma [2]. But in a remote resource poor setting, the TNM staging classification still continues to be an important predictor of cancer survival [3]. Lymph node metastasis is an important prognostic factor which plays a vital role in staging and management of patients [4]. Fine needle aspiration cytology is a reliable method for the diagnosis of breast carcinoma. There are various cytological grading systems for breast carcinoma of which Robinson’s grading system is one which has 6 cytological parameters [5]. Studies done in the past have shown correlation between cytological grading on FNAC and histological tumor grade [5-9]. These studies have concluded that the cytological tumor grading on FNAC can predict the tumor prognosis, especially for patients who have received neo-adjuvant therapy. Few studies have shown correlation between cytological tumor grade and axillary lymph nodal metastasis [7,9,10]. The present study is designed to explore the prognostic role of FNAC in Invasive Ductal Carcinoma of breast by correlating the cytological grade with axillary lymph node status.

Materials and Methods:

This is a retrospective study conducted in the department of Pathology in a tertiary rural teaching hospital in Mandya, Karnataka. The study group included 68 cases of Invasive Ductal Carcinoma of breast diagnosed on FNAC who subsequently underwent modified radical mastectomy in our hospital. FNAC slides of the breast tumor and the histopathology slides of axillary lymph nodes of all the cases were retrieved. The FNAC slides of all 68 cases of IDC were assessed for cytological grade using Robinson's grading system. (Table 1) The scores for all individual cytological criteria were recorded for all the cases.

The histopathology slides of axillary lymph nodes stained by Hematoxylin-and-eosin were analyzed for the presence of metastasis. Statistical analysis was done by SPSS statistical software package, version 14.0. Finally, the cytological tumor grade and all individual cytological features were correlated with the presence or absence of axillary lymph node metastasis using χ² test and P < 0.05 was considered statistically significant.

Results:

The study group included a total number of 68 cases. Cytological grading of all the 68 cases were done using Robinson’s method (Table 1). 24 cases were assigned grade 1, 17 cases were assigned grade 2 and 27 cases were assigned grade 3 (Table 2) (Figure 1, 2 &amp; 3). Of the 68 cases of IDC breast, 51 cases (75%) showed metastasis to axillary lymph node (Figure 4) and 17 cases (25 %) did not show axillary lymph nodal metastasis (Table 3). Of the 24 cases with cytological grade 1, 14 cases (58.3%) showed lymph node positivity and 10 cases (41.6%) did not show lymph node positivity. Of the 17 cases with cytological grade 2, 13 cases (76.4%) showed lymph node positivity and 4 cases (23.5%) did not show lymph node positivity. Of the 27 cases with cytological grade 3, lymph node positivity was seen in 24 cases (88.8%) and negative in 3 cases (11.1%). (Table 3).

The cytological grade assigned were correlated with the presence and absence of lymph node metastasis using chi square test and a P value of <0.05 was obtained which was statistically significant.

Finally, the score of individual cytological criterias of Robinson’s system which are Cell dissociation, Nuclear size, Cell uniformity, Nucleoli, Nuclear margin and Chromatin pattern were correlated with the presence or absence of axillary lymph nodal metastasis (Table 4).

For Nuclear size cytological scores 1, 2 and 3, lymph node positivity was seen in 6 (11.7%), 16 (31.3%) and 29 (56.8%) cases respectively, whereas lymph nodes were negative in 1 (5.8%), 12 (70.5%) and 4 (23.5%) cases respectively. For Cell uniformity cytological scores 1, 2 and 3, lymph node positivity was seen 6 (11.7%), 20 (39.2%) and 25 (49%) cases respectively, whereas lymph nodes were negative in 7 (41.1%), 6 (35.2%) and 4 (23.5%) cases respectively.

For Nucleoli cytological scores 1, 2 and 3, lymph node positivity was seen in 9 (17.6%), 18 (35.2%) and 24 (47%) cases respectively, whereas lymph nodes were negative in 8 (47%), 7 (41.1%) and 2 (11.7%) cases respectively. For Nuclear margin cytological scores 1, 2 and 3, lymph node positivity was seen in 7 (13.7%),43 (84.3%) and 1 (1.9%) cases respectively, whereas lymph nodes were negative in 7 (23.5%), 6 (11.7%) and 4 (5.8%) cases respectively.

For these four cytological criteria, as the scores increased from 1 to 3 the proportion of cases with lymph node positivity also increased and the proportion of cases negative for lymph node metastasis decreased. This correlation between cytological scores and lymph node status showed statistically significant correlation.

The other two cytological criteria, cell dissociation and chromatin pattern did not show statistically significant correlation with the lymph nodal metastasis. For Cell dissociation cytological scores 1, 2 and 3, lymph node positivity was seen in 15 (29.4%), 25 (49%) and 11 (21.5%) cases respectively, whereas lymph nodes were negative in 7 (41.1%), 9 (52.90%) and 1 (5.8%) cases respectively. For Chromatin pattern cytological scores 1, 2 and 3, lymph node positivity was seen in 8(15.6%), 39 (76.4%) and 4 (7.8%) cases respectively, whereas lymph nodes were negative in 2 (11.7%), 14 (82.3%) and 1 (5.8%) cases respectively.

Figure 1: IDC with cytological grade 1 (H&E, X400). The cells are arranged in clusters in a hemorrhagic background.	Figure 2: IDC with cytological grade 2 (H&E, X400). The cells are arranged in clusters and in singles.

Figure 3: IDC with cytological grade 3 (H&E, X400). The cells are arranged in singles.	Figure 4: Axillary lymph node with IDC metastasis (H&E, X400).

Discussion:

Of the 68 cases of IDC breast, 51 cases (75%) showed metastasis to axillary lymph node and 17 cases (25 %) did not show axillary lymph nodal metastasis. Of the 68 cases, cytological grade 1, 2 and 3 were assigned to 24, 17 and 27 cases respectively (Figure 1, 2 &amp; 3). 0f the 24 cases with cytological grade 1, axillary lymph nodal metastasis was present in 14 cases (58.3%) and absent in 10 cases (41.6%). 0f the 17 cases with cytological grade 2, axillary lymph nodal metastasis was present in 13 cases (76.4%) and absent in 4 cases (23.5%). 0f the 27 cases with cytological grade 3, axillary lymph nodal metastasis was present in 24 cases (88.8%) and absent in 3 cases (11.1%). In our study, it is observed that as the cytological grading of the tumor increased from 1 to 3, the number of cases showing metastasis to lymph node increased and this correlation is statistically significant (p < 0.05).

Similar studies done in the past also showed a statistically significant association between cytological grade and presence of axillary metastasis (p < 0.05) [7, 9, 11, 12].

Few studies have shown that the cytological grade correlates well with the histological grade obtained by modified Nottingham- Bloom–Richardson system [ 13, 14].

It is observed in our study that the cytological scores of four cytological criteria of Robinson’s system which are Nuclear size, Cell uniformity, Nucleoli and Nuclear margin correlated well with the axillary lymph nodal metastasis and was found to be statistically significant, whereas the cytological scores of other two criteria, cell dissociation and chromatin pattern did not show significant correlation with lymph node metastasis. (Table 4).

Another study by Sinha A et al, showed that among the six criteria, cell uniformity and nuclear size showed significant positive correlation with the presence of metastasis in axillary lymph nodes [11].

In a study by Robles-Frias et al, three features, that is, cell uniformity, cell dissociation, and nuclear margin showed a positive correlation with metastasis to axillary lymph node [7]. Similar to Robles-Frias et al, our study also showed that cell uniformity and nuclear margin showed a positive correlation with metastasis to axillary lymph node, but cell dissociation was not found to have significant correlation to lymph nodal metastasis in our study.

Similar to a study by Sinha A et al, both cell uniformity and nuclear size showed significant positive correlation with the presence of metastasis in axillary lymph nodes in our study [11]. In addition to these criteria, the present study also found that nucleoli showed positive correlation with the presence of metastasis in axillary lymph nodes.

The present study showed that the cytological scores of Nuclear size, Cell uniformity, Nucleoli and Nuclear margin correlated well with the axillary lymph nodal metastasis. Nuclear pleomorphism is an important component of Bloom Richardson histological grading system and correlates with prognosis [15, 16]. Nuclear size and cell uniformity are the features of pleomorphism. Tumors with increased cytological scores of nuclear size and cell uniformity will have greater pleomorphism, which explains their correlation with axillary nodal metastasis.

Nuclei are required for DNA replication or transcription, and nucleoli constitutes a major part of nuclei. Few studies have demonstrated that nucleoli and nuclear margin are extremely important in differentiating cytological grades of malignant tumors [17, 18].

But the cytological scores of nucleoli and nuclear margins did not show statistically significant correlation with the axillary nodal metastasis in our study. This result is similar to the results of previous studies [7, 11].

Conclusion:

The Robinson’s cytological grading of IDC breast on FNAC correlates with the axillary lymph nodal metastasis. Hence cytological grading on FNAC can be used as an important prognostic tool. In patients of IDC with higher cytological grades on FNAC the possibility of axillary lymph nodal metastasis has to be considered by the treating oncologist which helps in proper patient management.

References:

1. Pandey P, Dixit A, Chandra S, et al. A comparative and evaluative study of two cytological grading systems in breast carcinoma with histological grading: an important prognostic factor. Analytical Cellular Pathology (Amst) 2014;2014:767215.
2. Giuliano AE, Connolly JL, Edge SB, Elizabeth A, Mittendorf, Rugo HS, et al. Breast cancer – Major changes in the American Joint Committee on Cancer eighth edition cancer staging manual. CA Cancer J Clin. 2017;67:290–303.
3. Saadatmand S, Bretveld R, Siesling S, Tilanus-Linthorst MM. Influence of tumour stage at breast cancer detection on survival in modern times: Population based study in 173797 patients. BMJ. 2015;351:h4901.
4. Shek LM, Codolphin W. Model for breast cancer survival: Relative prognostic roles of axillary nodal status, TNM stage, estrogen receptor concentration and tumor necrosis. Cancer Res. 1988;48:5565–9.
5. Robinson IA, McKee G, Nicholson A, D’Arcy J, Jackson PA, Cook MG, et al. Prognostic value of cytological grading of fine needle aspirates from breast carcinomas. Lancet. 1994;343:947–9.
6. Sinha A, Gill SS. Correlative study of cytological features in grading of invasive breast carcinoma. J Cytol. 2018;35:149–52.
7. Robles-Frias A, Campora RG, Martinez-Parra D et al. Robinson cytologic grading of invasive ductal breast carcinoma. Correlation with histologic grading and regional lymph node metastasis. Acta Cytol. 2005;49:149–53.
8. Walke VA, Gunjkar G. Comparative evaluation of six parametric Robinson and three parametric Howell's modification of Scarf-BloomRichardson grading method on breast aspirates with histopathology: A prospective study. Cytojournal. 2017;14:31.
9. Taniguchi E, Yang Q, Tang W, Nakamura Y, Shan L, Nakamura M, et al. Cytologic grading of invasive breast carcinoma: Correlation with clinico-pathologic variables and predictive value of nodal metastasis. Acta Cytol. 2000;44:587–91.
10. Wani FA, Bhardwaj S, Kumar D, Katoch P, Bandy AF. Relationship between cytological grading of breast cancers and lymph node metastasis. Int J Med Sci Public Health. 2014;3:906–11.
11. Sinha A, Gill SS. Cytological Correlates of Axillary Nodal Involvement in Invasive Ductal Carcinoma of Breast. J Cytol. 2019 Jul-Sep;36(3):142-145. doi: 10.4103/JOC.JOC_197_18.
12. Khan N, Afroz N, Rana F, Khan MA. Role of cytologic grading in prognostication of invasive breast carcinoma. J Cytol. 2009;26:65–8.
13. Pal S, Gupta ML. Correlation between cytological and histological grading of breast cancer and its role in prognosis. J Cytol. 2016 Oct-Dec;33(4):182-186. doi: 10.4103/0970-9371.190449.
14. Phukan JP, Sinha A, Deka JP. Cytological grading of breast carcinoma on fine needle aspirates and its relation with histological grading. South Asian J Cancer. 2015 Jan-Mar;4(1):32-4. doi: 10.4103/2278-330X.149948.
15. Meyer JS, Alvarez C, Milikowski C, et al. Breast carcinoma malignancy grading by Bloom-Richardson system vs proliferation index: reproducibility of grade and advantages of proliferation index. Mod Pathol. 2005;18(8):1067–1078. doi: 10.1038/modpathol.3800388.
16. Theissig F, Kunze KD, Haroske G, et al. Histological grading of breast cancer. Interobserver reproducibility and prognostic significance. Pathol Res Pract 1990;186:732–736.
17. Kashyap A, Jain M, Shukla S, Andley M. Role of nuclear morphometry in breast cancer and its correlation with cytomorphological grading of breast cancer: a study of 64 cases. Journal of Cytology. 2018;35(1):41–45.
18. Akashi S, Kuwabara H, Kurisu Y et al. Fine-needle aspiration cytology of triple-negative basal-like breast cancer. Diagnostic Cytopathology. 2013;41(4):283–287.