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Original Article |
1 Department of Surgery, Columbia University Comprehensive Breast Center, Atchley Pavilion, 10th Floor, 161 Fort Washington Avenue, New York, New York 10032
2 Department of Epidemiology, Columbia University Comprehensive Breast Center, Women at Risk, 601 W. 168th Street, New York, New York 10032
3 Department of Pathology, Columbia University, VC10-209, 630 W. 168th Street, New York, New York 10032
4 Department of Biostatistics, Columbia University, 722 W. 168th Street, New York, New York 10032
Correspondence: Address correspondence and reprint requests to: Mahmoud El-Tamer, MD, FACS; E-mail: me180{at}columbia.edu.
| ABSTRACT |
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Methods: Using the Columbia University breast cancer database, we identified all DCIS patients who had eight or more axillary nodes dissected and free of metastasis. Five-micrometer sections from all paraffin blocks containing lymph node tissue were stained with an anticytokeratin antibody cocktail (AE1/AE3 and KL1). The results of the CK-IHC and updated database were anonymized and merged. Survival of CK-IHCpositive and negative patients was compared by using Kaplan-Meier curves and log-rank tests.
Results: CK-IHC was performed on 301 DCIS patients, who had an average of 16.7 axillary nodes dissected. Eighteen (6%) of 301 patients tested positive by CK-IHC. Seventy patients with bilateral breast cancer and 2 patients without any follow-up data were excluded, for a final study population of 229 patients. Among the 216 patients with negative CK-IHC, 18 patients died, compared with 1 of 13 patients with positive CK-IHC. The median follow-up for the study group was 127 months. Kaplan-Meier overall and breast cancerspecific survival estimates were similar for CK-IHCpositive and negative patients (P = .81 and P = .73, respectively).
Conclusions: CK-IHC increases the incidence of positive nodes by 6% in DCIS patients. A positive node by CK-IHC does not seem to affect survival in these patients. These results raise concerns regarding the clinical significance of positive nodes by CK-IHC in DCIS patients.
Key Words: Lymph node metastasis DCIS Cytokeratin immunohistochemistry Prognosis
| INTRODUCTION |
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In the past, axillary dissection was routinely performed in the management of DCIS. Axillary node metastasis was detected in 1% of patients by hematoxylin and eosin stains (H&E).79 With such a low incidence of nodal metastasis, axillary nodal dissection was therefore abandoned when treating patients with DCIS.6,10,11
Sentinel lymph node biopsy (SLNB) is currently an acceptable alternative to axillary nodal dissection, and it has yielded an effective and accurate method of evaluating lymph node metastasis. It has been estimated that the diagnostic yield with SLNB may be increased by 10% to 15% in comparison to routine axillary dissection, particularly with cytokeratin immunohistochemical staining (CK-IHC).12 SLNB is appealing because it can be performed with patients under local anesthesia, through a small incision, and has carried a lower morbidity than axillary nodal dissection.1315 Recently, several investigators have reported the use of SLNB in the management of patients with DCIS. These studies have reported a 6% to 23% rate of axillary metastasis in patients with DCIS by using SLNB and CK-IHC.1618 The aim of our study was to find the incidence of axillary nodal metastasis detected by CK-IHC in DCIS patients who underwent a formal axillary node dissection and to estimate the effect of these positive nodes on survival.
| PATIENTS AND METHODS |
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Histological Examination
The histopathology examination used standard techniques for thorough sampling of all nodal tissue. After thorough dissection, all nodal tissue was embedded according to protocol at Columbia University. If the entire lymph node was too big for a single cassette, it was embedded in multiple cassettes. Between 1980 and 1990, every lymph node was examined at three levels. From 1990 on, each node was bisected, and one level was examined from each section; deeper levels were cut at the pathologists discretion. Lymph node blocks from patients with nodes reported negative on original pathology review were obtained and refaced, and a single 5-µm section from each node block was examined. After microwave antigen retrieval in citrate buffer (pH 6.0), immunostaining was performed by using an anti-cytokeratin (CK) antibody cocktail (AE1/AE3 and KL1). Antibody binding was detected with the Envision Plus system (Dako, Carpinteria, CA), with diaminobenzidine as the chromogen.
Cells were considered positive only when they showed distinct cytoplasm staining and had an epithelial appearance. All staining was performed with multiple positive controls. In addition, most slides showed positive staining of dendritic mesenchymal cells, which were not included as positive. AE1/AE3 was used because it is the standard "pancytokeratin" with reactivity against both basic and acidic keratins. In our experience, AE1/AE3 does not stain CK 8/18 very well. Thus, we added another pancytokeratin antibody, KL1, which has a strong spectrum for CK8/18 immunostaining. This cocktail stained all CKs, especially when used with heat-induced epitope retrieval. Therefore, there was no need to add another CK antibody. The positive CK-IHC cells were counted. A group of four or more positive cells was defined as a cluster irrespective of its size. Removing all patient identifiers anonymized the database, and the CK-IHC results were merged with the follow-up database.
Statistical Methods
To avoid any confounding due to breast cancerrelated deaths within the survival analysis, we excluded all patients with bilateral breast cancer. The patients identified with a positive CK stain in the axillary lymph nodes were grouped together, and their survival was compared with the survival of those without axillary metastasis. The analysis was irrespective of the type of surgery performed (mastectomy or breast preservation). The end points compared were overall survival and disease-specific survival. Overall survival was defined from the date of diagnosis to the date of death or last follow-up. For disease-specific survival, patients who did not die from breast cancer were censored at the time of death. Patients who died of unknown causes but harbored metastatic disease before death were included as deaths from breast cancer. Patients who died from unknown causes without evidence of disease were considered as deaths from other causes.
Univariate comparisons between the CK-positive and -negative patients were performed with Pearsons
2 and Students t-tests. The probability of survival and disease-specific survival was estimated by using the Kaplan-Meier method and was compared by using two-sided log-rank tests. The 95% confidence intervals for the survival estimates were calculated by using the Greenwood method. P < .05 was considered significant. All analyses were performed with Stata 8.0 (Stata Corp., College Station, TX).
| RESULTS |
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For the survival analysis, we excluded 70 patients who had bilateral breast cancer and 2 patients who were lost to follow-up. A total of 229 patients were used for the survival comparison. All patients in the survival analysis were women, with a median age of 54 years (range, 3085 years). The vast majority of patients200 (87%) of 229had undergone a modified radical mastectomy, and the rest had lumpectomy and axillary nodal dissection. The average number of nodes dissected was 16.7 (range, 849). The median follow-up was 127 months. In 13 of the 229 patients, the CK stain was positive, for an incidence of 5.7%. The pattern of positive CK stain is described in Table 1
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| DISCUSSION |
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Most metastasis with SLNB has been detected with CK-IHC. Using CK-IHC, we have detected nodal metastasis in 6% of DCIS cases that were previously labeled as negative by H&E stains after axillary dissection. We have not reviewed the slides of the breast primary tumor in these patients, and some microinvasive disease cases may have been included in the study. In a similar retrospective study, Lara et al.25 reviewed 102 pure DCIS patients and identified 13 cases of nodal metastasis detected by CK-IHC, for an incidence of 13%. The higher rate detected by that study may be related to more extensive sectioning of paraffin blocks at seven levels with 100 to 150 µm between levels. In our study, only one section from each nodal block was stained.
We found no differences in prognosis with nodal metastasis detected by CK-IHC in DCIS patients. With a median follow-up of 127 months (10.6 years), the overall and disease-specific survival was similar for the CK-IHCpositive and negative node groups. In the CK-IHCpositive group, only 1 of the 13 patients had died. That patient died at the age of 98 years, 12.4 years after diagnosis, without any evidence of disease. It is interesting to note that she was the only patient with two positive lymph nodes. Lara et al.25 reported 15 deaths (15%) among 102 pure DCIS patients, with a mean follow-up of 19 years; none of the deaths was associated with breast cancer. In Tamhane and colleagues study,24 6 of 26 DCIS patients who had undergone a mastectomy with some axillary node dissection had positive nodes. With a relatively short mean follow-up of 5 years, none of the patients had died, had a recurrence, or developed metastasis. So far, the positive node detected by CK-IHC alone has consistently been an insignificant predictor of outcome in DCIS patients. This finding is inconsistent with what has been established for nodal metastasis detected by H&E. This inconsistency raises many questions. Are CK-IHC metastatic cells real? Are they mechanically disseminated and lack the biologic ability to metastasize?
It is clear from our study and others24,25 that the increase in detection of nodal metastasis by CK-IHC in DCIS patients is not a phenomenon exclusive to SLN procedures. All patients included in this study had had a form of biopsy to diagnose cancer before axillary node dissection. Carter et al.26 and Youngson et al.27 have reported the migration of benign and malignant cells to axillary lymph nodes after simple procedures such as a breast needle biopsies. Moore et al.28 reviewed their experience with 4016 SLN procedures and concluded that the frequency of CK-IHCpositive nodes is not related to the usual predictors of nodal metastasis and that it increases proportionately with the degree of preoperative manipulation. It is quite possible that some nodal metastases detected by CK-IHC in our study were iatrogenic. These single cells or clusters of cells may travel passively to a single lymph node and may die or be destroyed by the host before nesting and later metastasizing if they carry that biologic potential.
Whatever the reasons for positive axillary nodes by CK-IHC in DCIS patients, these metastases do not seem to have a significant effect on survival with a median of 10 years of follow-up. This observation is consistent with those of other studies with long-term follow-up.24,25 Including the 13 patients from this study, 13 patients from Lara and associates study,25 and 6 patients in Tamhane and colleagues study,24 none of the 32 CK-IHCpositive patients in all 3 studies died of breast cancer. CK-IHC in nodes of DCIS patients may not be detecting genuine nodal metastasis related to an active biologic process that will be responsible for a compromised outcome if not treated. One may attribute the lack of effect on survival to the possible therapeutic effect of axillary node dissection. However, the survival of DCIS patients, with or without axillary dissection, has been close to 100%.19,29 The disease-specific survival for this study population (99%) mirrors previously reported DCIS survival rates.
In our study, a formal node dissection was performed, and an average of 16.7 nodes were removed. We can assume that in the vast majority of our cases, the SLN was included in the specimen, and the data can be safely extrapolated to SLNBs. Although SLNB carries a low morbidity, a positive node in DCIS patients may have a major effect on the patients quality of life. Patients may experience psychological trauma and frequently may also undergo a complete axillary node dissection with chemotherapy, hormonal treatment, or both. Perhaps SLNB should be limited to DCIS patients who undergo mastectomy, who have palpable disease,30 or who have proven or questionable microinvasion. Furthermore, this study advises caution in using routine CK-IHC on SLNs from DCIS patients unless it is part of a protocol. The available data do not support the use of a positive SLN detected by CK-IHC alone in the clinical management of DCIS patients.
| ACKNOWLEDGMENTS |
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Received for publication May 4, 2004. Accepted for publication November 5, 2004.
| REFERENCES |
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