- Email: [email protected]
Breast cancer detection in axillary sentinel lymph nodes: the impact of the method of pathologic examination
Human Pathology (2014) 45, 2497–2501
www.elsevier.com/locate/humpath
Original contribution
Breast cancer detection in axillary sentinel lymph nodes: the impact of the method of pathologic examination☆ Benjamin C. Calhoun MD, PhD a,1 , Karinn Chambers MD b , Teresa Flippo-Morton MD b , Chad A. Livasy MD a , Edward J. Armstrong III BS b , James T. Symanowski PhD b , Terry Sarantou MD b , Frederick L. Greene MD b , Richard L. White Jr. MD b,⁎ a
Department of Pathology, Levine Cancer Institute, Carolinas Medical Center, Carolinas HealthCare System, Charlotte, NC 28203 Division of Surgical Oncology, Department of Surgery, Levine Cancer Institute, Carolinas Medical Center, Carolinas HealthCare System, Charlotte, NC 28204 b
Received 24 July 2014; revised 10 September 2014; accepted 17 September 2014
Keywords: Breast; Metastasis; Surgery; Pathology; Diagnosis
Summary At Carolinas Medical Center, before 2008, axillary sentinel lymph nodes (SLNs) from breast cancer patients were evaluated with a single hematoxylin and eosin–stained slide. In 2008, the protocol changed to include a limited step sectioning at 500 μm. In this study, we compared the intraoperative and permanent section pathologic findings for SLN biopsies from 2006 to 2007 to those from 2009 to 2010. We hypothesized that evaluating 2 slides would increase the detection of micrometastases and isolated tumor cells (ITCs) on permanent sections and correspondingly decrease the sensitivity of intraoperative touch preparation cytology (IOTPC). From 2006 to 2007, 140 (23.5%) of 597 of SLN permanent sections contained tumor cells: 92 macrometastases (65.7%), 36 micrometastases (25.7%), and 12 ITCs 0.2 mm or less (8.6%). The sensitivity of IOTPC for 2006 to 2007 was 51.4% for any tumor cells and 71.7% for macrometastases. From 2009 to 2010, 160 (21.9%) of 730 SLN permanent sections were positive for any tumor cells: 76 macrometastases (47.5%), 55 micrometastases (34.4%), and 29 ITCs (18.1%). The sensitivity of IOTPC for 2009 to 2010 was 39.4% for any tumor cells and 76.3% for macrometastases. With limited step sectioning, we observed an approximately 10% increase in the detection of both micrometastases and ITCs in SLN. The increased detection of ITCs on permanent sections reached statistical significance (P = .018). However, under current clinical guidelines, patients with limited SLN involvement may not be required to undergo completion axillary lymph node dissection. The ability to detect SLN tumor deposits less than 2 mm must be balanced with the clinical utility of doing so. © 2014 Elsevier Inc. All rights reserved.
☆
Disclosures: There was no specific external funding for this study, and the authors have no relevant financial disclosures. ⁎ Corresponding author. Division of Surgical Oncology, Department of Surgery, Levine Cancer Institute, Carolinas Medical Center, 1021 Morehead Medical Drive #6100, Charlotte, NC 28204. E-mail address: [email protected] (R. L. White). 1 Current address: Department of Pathology, Cleveland Clinic, Cleveland, OH 44195. http://dx.doi.org/10.1016/j.humpath.2014.09.004 0046-8177/© 2014 Elsevier Inc. All rights reserved.
2498
B. C. Calhoun et al.
1. Introduction The development and validation of the sentinel lymph node (SLN) biopsy revolutionized the surgical approach to the axillary staging of breast cancer patients. The technique was rapidly adopted based on the ability to accurately stage breast cancer patients with minimal morbidity. Many patients with negative SLN biopsy results have been able to avoid the potential morbidity associated with a full axillary lymph node dissection (ALND). The role for ALND in patients with low-volume tumor deposits (b2 mm) in SLN has been more controversial. Despite the routine use of SLN biopsy in patients with early stage breast cancer, the technique used in the final pathologic evaluation of removed SLNs shows considerable variation across institutions. This study sought to analyze the effect of different pathologic examination methods on intraoperative and final pathologic diagnoses for axillary SLNs in a large series of breast cancer patients. In 2008, Levine Cancer Institute/Carolinas Medical Center changed the protocol for the pathologic examination of SLN permanent sections from a single hematoxylin and eosin (H&E) slide to 2 H&E slides approximately 500 μm apart. The additional level and the 500-μm interval were based on recommendations published by the American Society of Clinical Oncology [1]. Our hypothesis, based on the results of prior studies of intensive histologic evaluation of SLN [2], was that evaluating an additional deeper level would increase the detection of micrometastases (N0.2 mm and b2.0 mm) and isolated tumor cells (ITCs) 0.2 mm or less on permanent sections and lead to a corresponding decrease in the sensitivity of intraoperative touch preparation cytology (IOTPC). We report the extent of nodal involvement on permanent sections and the sensitivity of IOTPC evaluation for more than 1300 SLN biopsies performed before and after the protocol change.
2. Materials and methods In an institutional review board–approved retrospective review, we compared the SLN biopsy findings for all patients with invasive breast cancer who underwent SLN biopsy from 2006 to 2007 and 2009 to 2010. Cases were identified using a natural language search in the Anatomic Pathology Laboratory Information System (Sunquest CoPath) using the search term sentinel and excluding cases that contained the term melanoma. A pathologist or surgeon reviewed the final diagnoses and excluded nonmammary cases. The extent of nodal involvement was classified according to the seventh edition of the American Joint Committee on Cancer Staging System [3] (Fig. 1). The SLN biopsy results for each patient were classified based on the size of the largest tumor deposit. For bilateral SLN evaluation, each axilla was counted as an individual SLN biopsy.
Fig. 1 Representative H&E (A) and cytokeratin-stained (B) sections containing ITCs, less than 0.2 mm (original magnification ×400).
At the time of intraoperative evaluation, SLNs were sectioned longitudinally into slices approximately 2 mm thick. Air-dried touch preparations were made from 1 cut surface of each 2-mm section and stained with Diff-Quik (Fig. 2). Before 2008, 1 H&E slide from each SLN tissue block was examined. In 2008, the protocol was changed to include a second H&E slide approximately 500 μm deeper in the block. Six unstained levels between the 2 H&E slides were retained for possible cytokeratin staining or other ancillary studies. In both time frames, immunohistochemistry for cytokeratins was not performed routinely but was used to evaluate indeterminate cells on H&E slides on a case-by-case basis. An SLN biopsy was performed for all invasive cancers that had not been previously proven node positive based on axillary ultrasound or physical examination and subsequent biopsy. To perform the SLN biopsy, both technetium sulfur colloid and lymphazurin blue dye were injected for lymphatic mapping while the patient was under general anesthesia at the beginning of the operative procedure as previously reported [4]. In 2008, the practices of preoperative axillary ultrasound and ultrasound-guided core biopsy of suspicious axillary nodes became widely adopted at our institution. The overall sensitivity of IOTPC as well as its accuracy in the diagnosis of macrometastases, micrometastases, and ITCs during these 2 periods was compared using Fisher exact test. Frequencies and proportions were calculated for each
Limited step sectioning of breast sentinel lymph nodes
Fig. 2 Two representative images of positive IOTPC preparations (original magnification ×400).
period, and 2-sided P values were calculated based on the corresponding contingency tables.
3. Results The pathologic findings of IOTPC and permanent sections for the 2-year periods before (2006-2007) and after (2009-2010) the protocol change requiring the examination of a second H&E slide 500 μm deeper in the block are shown in Table 1. In 2006 to 2007, 140 (23.5%) of 597 SLN biopsies (from 589 patients) were positive for any Table 1
2499 tumor cells and were further classified as 92 macrometastases (65.7%), 36 micrometastases (25.7%), and 12 ITCs 0.2 mm or less (8.6%). Seventy-three IOTPC evaluations were reported as positive, including 1 false positive (1.4%), 66 macrometastases (90.4%), 3 micrometastases (4.1%), and 3 ITCs (4.1%). In 2009 to 2010, 160 (21.9%) of 730 were positive for any tumor cells. Of the involved SLN biopsies, 76 (47.5%) contained macrometastases; 55 (34.4%), micrometastases; and 29 (18.1%), ITC. Sixty-three IOTPC were reported as positive, including 58 macrometastases (92.1%), 3 micrometastases (4.8%), and 2 ITCs (3.2%). The sensitivity of IOTPC and the extent of nodal involvement in permanent sections of SLN biopsies for the 2 periods are shown in Table 2. The decrease in overall sensitivity of IOTPC for detecting any tumor cells in 2009 to 2010 (39.4%) compared to 2006 to 2007 (51.4%) was statistically significant (P = .048, Fisher exact test). The sensitivity of IOTPC in the diagnosis of macrometastases, micrometastases, and ITC specifically showed no statistically significant difference over the 2 periods (P = .598, P = .678, and P = .139, respectively). The proportion of micrometastases in permanent sections of SLN increased from 25.7% to 34.4%, but this did not reach statistical significance (P = .131). In permanent sections of positive SLN, the decrease in the proportion of macrometastases (65.7% versus 47.5%) and the increase in the proportion of ITC (8.6% versus 18.1%) reached statistical significance (P = .002 and P = .018, respectively).
4. Discussion The largest prospective randomized phase III trial comparing SLN biopsy to ALND in the United States, NSABP B-32, confirmed that an SLN biopsy was a technically feasible and accurate means of staging the axilla in women with breast cancer [5]. As SLN biopsy has become widely adopted practice, the intensive pathologic evaluation of these lymph nodes has identified more ITC and micrometastases. Despite our ability to identify very low volumes of disease in SLN, mounting data from multiple single- and multi-institutional retrospective studies suggested little value in completion ALND [6-10] for these
Pathologic classification of breast SLN biopsies
2006-2007 IOTPC Permanent 2006-2007 IOTPC Permanent
Patients
SLN−
SLN+ (any)
False+ (IOTPC)
Macro
Micro
ITC
597 597
524 457
72 140
1 N/A
66 92
3 36
3 12
730 730
667 570
63 160
0 N/A
58 76
3 55
2 29
NOTE. The number of positive SLN identified on IOTPC and permanent sections is shown for the periods with (2006-2007) and without (2009-2010) step sectioning. Cases with multiple positive SLN were classified based on the largest tumor deposit in an SLN. Abbreviations: SLN, sentinel lymph node; IOTPC, intraoperative touch prep cytology; Macro: macrometastases; Micro: micrometastases; ITC, isolated tumor cells.
2500
B. C. Calhoun et al.
Table 2 Sensitivity of IOTPC and extent of SLN involvement on permanent sections 2006-2007 IOTPC sensitivity Any tumor cells Macrometastases Micrometastases ITCs Permanent sections Macrometastases Micrometastases ITCs
2009-2010
51.4% (72/140) 39.4% (63/160) 71.7% (66/92) 76.3% (58/76) 8.3% (3/36) 5.5% (3/55) 25.0% (3/12) 6.9% (2/29)
P .048 ⁎ .598 .678 .139
65.7% (92/140) 47.5% (76/160) .002 ⁎ 25.7% (36/140) 34.4% (55/160) .131 8.6% (12/140) 18.1% (29/160) .018 ⁎
NOTE. The sensitivity of IOTPC for any tumor cells, macrometastases, micrometastases, and ITCs is shown for the periods with (2006-2007) and without (2009-2010) step sectioning. Cases with multiple positive SLNs were classified based on the largest tumor deposit in an SLN. Abbreviations: IOTPC, intraoperative touch prep cytology; SLN, sentinel lymph node; ITC, isolated tumor cell. ⁎ Statistically significant P b .05 (Fisher exact test).
patients. More recently, the ACOSOG Z0011 investigators [11,12], IBCSG 23-01 investigators [13], NSABP B-32 investigators [5,14], and others [15,16] have concluded that completion ALND is not required for patients with minimal involvement of SLNs. The utilization of intraoperative evaluation of SLNs in breast cancer has changed dramatically secondary to publication of the results of NSABP B-32 and ACOSOG Z0011 [11,12]. Before the publication of ACOSOG Z0011, IOTPC was routinely performed for all breast cancer patients undergoing SLN biopsy at our institution. Now, the use of IOTPC at our institution is mainly reserved for select patients who would not have met the eligibility criteria for ACOSOG Z0011. The study design for NSABP B-32 included the pathologic assessment of lymph nodes. Additional H&E levels and cytokeratin immunohistochemistry were performed in SLN that were negative on initial examination. These additional slides revealed occult tumor in 15.9% of 3887 patients: 11.1% with ITC clusters, 4.4% with micrometastases, and 0.4% with macrometastases [5]. Our experience with limited wide interval step sectioning is consistent with the finding that ITC accounts for a significant proportion of the nodal involvement detected with more intensive evaluation [2]. A possible rationale for maintaining limited step sectioning would be to ensure the detection of all macrometastases in SLN tissue sections that, due to variations in gross sectioning, are more than 2 mm thick [17]. The 5-year follow-up from NSABP B-32 showed a small, statistically significant difference (1.2%) in the overall survival of patients with occult metastases in SLN that were negative on initial examination [5]. The 10-year followup showed no statistically significant difference in overall survival or local-regional recurrence for patients with occult metastases in SLN that were negative on initial examination [14]. The NSABPB-32 investigators concluded that their “…findings argue against analysis of additional tissue levels
or routine immunohistochemical analysis for sentinellymph-node evaluation” [5] and that ALND for occult nodal involvement offers no benefit [14]. This is consistent with the most recent American Society of Clinical Oncology Clinical Practice Guideline Update for SLN biopsy in patients with early stage breast cancer [18]. Specifically, the Update Committee concluded in Recommendation 2.1 that “clinicians should not recommend ALND for women with early stage breast cancer who have one or two SLN metastases and will receive breast-conserving surgery (BCS) with conventionally fractionated whole-breast radiotherapy” [18]. The unproven benefit of intensive SLN evaluation argues against the practice, particularly in an era of cost reduction in health care. The cost of producing an H&E slide is much lower than other aspects of multidisciplinary breast cancer care (eg, operating room time, chemotherapy, and magnetic resonance imaging studies). However, laboratories are often viewed as cost centers, and the headcount for histotechnologists is frequently capped. Our limited step-sectioning protocol doubled the number of H&E slides produced from SLN tissue blocks. If we assume an average of 3.5 SLN per procedure and an average of 1.5 blocks per SLN with a cost of $5 per H&E slide, our step-sectioning protocol added approximately $19 000 to the cost of evaluating the 730 SLN biopsies in 2009 and 2010. The retrospective single-institution nature of this study and lack of long-term follow-up represent limitations of this study. In addition, the implementation of routine axillary ultrasound around the time of the change in the pathology protocol may have further reduced the likelihood of detecting macrometastases intraoperatively and in permanent sections. Analyzing SLNs with 2 H&E slides approximately 500 μm apart led to an increase in final pathologic diagnosis of microscopic metastatic disease in SLNs and subsequent upstaging in breast cancer patients with tumor deposits greater than 0.2 mm. We observed an approximately 10% increase in both micrometastases and ITCs with limited step sectioning. The higher proportion of micrometastases and ITC in 2009 to 2010 was associated with a decrease in the overall sensitivity of IOTPC. Both of these results were anticipated at the time the decision was made to review a second slide 500 μm deeper in the block. The lower proportion of SLN macrometastases in the 2009 to 2010 period may be related to increased utilization of preoperative axillary ultrasound and ultrasound-guided lymph node biopsy to document macrometastases at the time of diagnosis. In addition, the increase in micrometastases and ITC in 2009 to 2010 very likely diluted the macrometastases that were detected in a slightly larger pool of all metastases. Early stage breast cancer patients with limited SLN involvement are not required to undergo completion ALND [18]. The ability of pathologists to detect microscopic tumor in SLN must be balanced with the clinical significance of the findings and implications for treatment.
Limited step sectioning of breast sentinel lymph nodes
References [1] Lyman GH, Giuliano AE, Somerfield MR, et al. American Society of Clinical Oncology guideline recommendations for sentinel lymph node biopsy in early-stage breast cancer. J Clin Oncol 2005;23: 7703-20. [2] Viale G, Mastropasqua MG, Maiorano E, Mazzarol G. Pathologic examination of the axillary sentinel lymph nodes in patients with earlystage breast carcinoma: current and resolving controversies on the basis of the European Institute of Oncology experience. Virchows Arch 2006;448:241-7. [3] Edge SB, Byrd DR, Compton CC, et al, editors. AJCC Cancer Staging Manual. 7th ed. New York: Springer; 2010. [4] Stell VH, Flippo-Morton TS, Norton HJ, White Jr RL. Effect of intraoperative radiocolloid injection on sentinel lymph node biopsy in patients with breast cancer. Ann Surg Oncol 2009;16:2300-4. [5] Weaver DL, Ashikaga T, Krag DN, et al. Effect of occult metastases on survival in node-negative breast cancer. N Engl J Med 2011;364: 412-21. [6] Reed J, Rosman M, Verbanac KM, et al. Prognostic implications of isolated tumor cells and micrometastases in sentinel nodes of patients with invasive breast cancer: 10-year analysis of patients enrolled in the prospective East Carolina University/Anne Arundel Medical Center Sentinel Node Multicenter Study. J Am Coll Surg 2009;208:333-40. [7] Hansen NM, Grube B, Ye X, et al. Impact of micrometastases in the sentinel node of patients with invasive breast cancer. J Clin Oncol 2009;27:4679-84. [8] Kumar S, Bramlage M, Jacks LM, et al. Minimal disease in the sentinel lymph node: how to best measure sentinel node micrometastases to predict risk of additional non-sentinel lymph node disease. Ann Surg Oncol 2010;17:2909-19. [9] Cortesi L, Proietto M, Cirilli C, et al. Prognosis and treatment of micrometastatic breast cancer sentinel lymph node: a population-based study. J Surg Oncol 2012;106:399-405.
2501 [10] Sola M, Alberro JA, Fraile M, et al. Complete axillary lymph node dissection versus clinical follow-up in breast cancer patients with sentinel node micrometastasis: final results from the multicenter clinical trial AATRM 048/13/2000. Ann Surg Oncol 2013;20:120-7. [11] Giuliano AE, Hunt KK, Ballman KV, et al. Axillary dissection vs no axillary dissection in women with invasive breast cancer and sentinel node metastasis: a randomized clinical trial. JAMA 2011;305:569-75. [12] Giuliano AE, McCall L, Beitsch P, et al. Locoregional recurrence after sentinel lymph node dissection with or without axillary dissection in patients with sentinel lymph node metastases: the American College of Surgeons Oncology Group Z0011 randomized trial. Ann Surg 2010; 252:426-32. [13] Galimberti V, Cole BF, Zurrida S, et al. Axillary dissection versus no axillary dissection in patients with sentinel-node micrometastases (IBCSG 23-01): a phase 3 randomised controlled trial. Lancet Oncol 2013;14:297-305. [14] Julian TB, Anderson SJ, Krag DN, Weaver DL. Ten-year follow-up results of occult detected sentinel node disease: NSABP B-32, a randomized phase III clinical trial to compare sentinel node resection (SNR) to conventional axillary dissection (AD) in clinically node-negative breast cancer patients. San Antonio Breast Cancer Symposium; 2013. [15] Fournier K, Schiller A, Perry RR, Laronga C. Micrometastasis in the sentinel lymph node of breast cancer does not mandate completion axillary dissection. Ann Surg 2004;239:859-63. [16] Yegiyants S, Romero LM, Haigh PI, DiFronzo LA. Completion axillary lymph node dissection not required for regional control in patients with breast cancer who have micrometastases in a sentinel node. Arch Surg 2010;145:564-9. [17] Weaver DL, Le UP, Dupuis SL, et al. Metastasis detection in sentinel lymph nodes: comparison of a limited widely spaced (NSABP protocol B-32) and a comprehensive narrowly spaced paraffin block sectioning strategy. Am J Surg Pathol 2009;33:1583-9. [18] Lyman GH, Temin S, Edge SB, et al. Sentinel lymph node biopsy for patients with early-stage breast cancer: American Society of Clinical Oncology clinical practice guideline update. J Clin Oncol 2014;32:1365-83.
www.elsevier.com/locate/humpath
Original contribution
Breast cancer detection in axillary sentinel lymph nodes: the impact of the method of pathologic examination☆ Benjamin C. Calhoun MD, PhD a,1 , Karinn Chambers MD b , Teresa Flippo-Morton MD b , Chad A. Livasy MD a , Edward J. Armstrong III BS b , James T. Symanowski PhD b , Terry Sarantou MD b , Frederick L. Greene MD b , Richard L. White Jr. MD b,⁎ a
Department of Pathology, Levine Cancer Institute, Carolinas Medical Center, Carolinas HealthCare System, Charlotte, NC 28203 Division of Surgical Oncology, Department of Surgery, Levine Cancer Institute, Carolinas Medical Center, Carolinas HealthCare System, Charlotte, NC 28204 b
Received 24 July 2014; revised 10 September 2014; accepted 17 September 2014
Keywords: Breast; Metastasis; Surgery; Pathology; Diagnosis
Summary At Carolinas Medical Center, before 2008, axillary sentinel lymph nodes (SLNs) from breast cancer patients were evaluated with a single hematoxylin and eosin–stained slide. In 2008, the protocol changed to include a limited step sectioning at 500 μm. In this study, we compared the intraoperative and permanent section pathologic findings for SLN biopsies from 2006 to 2007 to those from 2009 to 2010. We hypothesized that evaluating 2 slides would increase the detection of micrometastases and isolated tumor cells (ITCs) on permanent sections and correspondingly decrease the sensitivity of intraoperative touch preparation cytology (IOTPC). From 2006 to 2007, 140 (23.5%) of 597 of SLN permanent sections contained tumor cells: 92 macrometastases (65.7%), 36 micrometastases (25.7%), and 12 ITCs 0.2 mm or less (8.6%). The sensitivity of IOTPC for 2006 to 2007 was 51.4% for any tumor cells and 71.7% for macrometastases. From 2009 to 2010, 160 (21.9%) of 730 SLN permanent sections were positive for any tumor cells: 76 macrometastases (47.5%), 55 micrometastases (34.4%), and 29 ITCs (18.1%). The sensitivity of IOTPC for 2009 to 2010 was 39.4% for any tumor cells and 76.3% for macrometastases. With limited step sectioning, we observed an approximately 10% increase in the detection of both micrometastases and ITCs in SLN. The increased detection of ITCs on permanent sections reached statistical significance (P = .018). However, under current clinical guidelines, patients with limited SLN involvement may not be required to undergo completion axillary lymph node dissection. The ability to detect SLN tumor deposits less than 2 mm must be balanced with the clinical utility of doing so. © 2014 Elsevier Inc. All rights reserved.
☆
Disclosures: There was no specific external funding for this study, and the authors have no relevant financial disclosures. ⁎ Corresponding author. Division of Surgical Oncology, Department of Surgery, Levine Cancer Institute, Carolinas Medical Center, 1021 Morehead Medical Drive #6100, Charlotte, NC 28204. E-mail address: [email protected] (R. L. White). 1 Current address: Department of Pathology, Cleveland Clinic, Cleveland, OH 44195. http://dx.doi.org/10.1016/j.humpath.2014.09.004 0046-8177/© 2014 Elsevier Inc. All rights reserved.
2498
B. C. Calhoun et al.
1. Introduction The development and validation of the sentinel lymph node (SLN) biopsy revolutionized the surgical approach to the axillary staging of breast cancer patients. The technique was rapidly adopted based on the ability to accurately stage breast cancer patients with minimal morbidity. Many patients with negative SLN biopsy results have been able to avoid the potential morbidity associated with a full axillary lymph node dissection (ALND). The role for ALND in patients with low-volume tumor deposits (b2 mm) in SLN has been more controversial. Despite the routine use of SLN biopsy in patients with early stage breast cancer, the technique used in the final pathologic evaluation of removed SLNs shows considerable variation across institutions. This study sought to analyze the effect of different pathologic examination methods on intraoperative and final pathologic diagnoses for axillary SLNs in a large series of breast cancer patients. In 2008, Levine Cancer Institute/Carolinas Medical Center changed the protocol for the pathologic examination of SLN permanent sections from a single hematoxylin and eosin (H&E) slide to 2 H&E slides approximately 500 μm apart. The additional level and the 500-μm interval were based on recommendations published by the American Society of Clinical Oncology [1]. Our hypothesis, based on the results of prior studies of intensive histologic evaluation of SLN [2], was that evaluating an additional deeper level would increase the detection of micrometastases (N0.2 mm and b2.0 mm) and isolated tumor cells (ITCs) 0.2 mm or less on permanent sections and lead to a corresponding decrease in the sensitivity of intraoperative touch preparation cytology (IOTPC). We report the extent of nodal involvement on permanent sections and the sensitivity of IOTPC evaluation for more than 1300 SLN biopsies performed before and after the protocol change.
2. Materials and methods In an institutional review board–approved retrospective review, we compared the SLN biopsy findings for all patients with invasive breast cancer who underwent SLN biopsy from 2006 to 2007 and 2009 to 2010. Cases were identified using a natural language search in the Anatomic Pathology Laboratory Information System (Sunquest CoPath) using the search term sentinel and excluding cases that contained the term melanoma. A pathologist or surgeon reviewed the final diagnoses and excluded nonmammary cases. The extent of nodal involvement was classified according to the seventh edition of the American Joint Committee on Cancer Staging System [3] (Fig. 1). The SLN biopsy results for each patient were classified based on the size of the largest tumor deposit. For bilateral SLN evaluation, each axilla was counted as an individual SLN biopsy.
Fig. 1 Representative H&E (A) and cytokeratin-stained (B) sections containing ITCs, less than 0.2 mm (original magnification ×400).
At the time of intraoperative evaluation, SLNs were sectioned longitudinally into slices approximately 2 mm thick. Air-dried touch preparations were made from 1 cut surface of each 2-mm section and stained with Diff-Quik (Fig. 2). Before 2008, 1 H&E slide from each SLN tissue block was examined. In 2008, the protocol was changed to include a second H&E slide approximately 500 μm deeper in the block. Six unstained levels between the 2 H&E slides were retained for possible cytokeratin staining or other ancillary studies. In both time frames, immunohistochemistry for cytokeratins was not performed routinely but was used to evaluate indeterminate cells on H&E slides on a case-by-case basis. An SLN biopsy was performed for all invasive cancers that had not been previously proven node positive based on axillary ultrasound or physical examination and subsequent biopsy. To perform the SLN biopsy, both technetium sulfur colloid and lymphazurin blue dye were injected for lymphatic mapping while the patient was under general anesthesia at the beginning of the operative procedure as previously reported [4]. In 2008, the practices of preoperative axillary ultrasound and ultrasound-guided core biopsy of suspicious axillary nodes became widely adopted at our institution. The overall sensitivity of IOTPC as well as its accuracy in the diagnosis of macrometastases, micrometastases, and ITCs during these 2 periods was compared using Fisher exact test. Frequencies and proportions were calculated for each
Limited step sectioning of breast sentinel lymph nodes
Fig. 2 Two representative images of positive IOTPC preparations (original magnification ×400).
period, and 2-sided P values were calculated based on the corresponding contingency tables.
3. Results The pathologic findings of IOTPC and permanent sections for the 2-year periods before (2006-2007) and after (2009-2010) the protocol change requiring the examination of a second H&E slide 500 μm deeper in the block are shown in Table 1. In 2006 to 2007, 140 (23.5%) of 597 SLN biopsies (from 589 patients) were positive for any Table 1
2499 tumor cells and were further classified as 92 macrometastases (65.7%), 36 micrometastases (25.7%), and 12 ITCs 0.2 mm or less (8.6%). Seventy-three IOTPC evaluations were reported as positive, including 1 false positive (1.4%), 66 macrometastases (90.4%), 3 micrometastases (4.1%), and 3 ITCs (4.1%). In 2009 to 2010, 160 (21.9%) of 730 were positive for any tumor cells. Of the involved SLN biopsies, 76 (47.5%) contained macrometastases; 55 (34.4%), micrometastases; and 29 (18.1%), ITC. Sixty-three IOTPC were reported as positive, including 58 macrometastases (92.1%), 3 micrometastases (4.8%), and 2 ITCs (3.2%). The sensitivity of IOTPC and the extent of nodal involvement in permanent sections of SLN biopsies for the 2 periods are shown in Table 2. The decrease in overall sensitivity of IOTPC for detecting any tumor cells in 2009 to 2010 (39.4%) compared to 2006 to 2007 (51.4%) was statistically significant (P = .048, Fisher exact test). The sensitivity of IOTPC in the diagnosis of macrometastases, micrometastases, and ITC specifically showed no statistically significant difference over the 2 periods (P = .598, P = .678, and P = .139, respectively). The proportion of micrometastases in permanent sections of SLN increased from 25.7% to 34.4%, but this did not reach statistical significance (P = .131). In permanent sections of positive SLN, the decrease in the proportion of macrometastases (65.7% versus 47.5%) and the increase in the proportion of ITC (8.6% versus 18.1%) reached statistical significance (P = .002 and P = .018, respectively).
4. Discussion The largest prospective randomized phase III trial comparing SLN biopsy to ALND in the United States, NSABP B-32, confirmed that an SLN biopsy was a technically feasible and accurate means of staging the axilla in women with breast cancer [5]. As SLN biopsy has become widely adopted practice, the intensive pathologic evaluation of these lymph nodes has identified more ITC and micrometastases. Despite our ability to identify very low volumes of disease in SLN, mounting data from multiple single- and multi-institutional retrospective studies suggested little value in completion ALND [6-10] for these
Pathologic classification of breast SLN biopsies
2006-2007 IOTPC Permanent 2006-2007 IOTPC Permanent
Patients
SLN−
SLN+ (any)
False+ (IOTPC)
Macro
Micro
ITC
597 597
524 457
72 140
1 N/A
66 92
3 36
3 12
730 730
667 570
63 160
0 N/A
58 76
3 55
2 29
NOTE. The number of positive SLN identified on IOTPC and permanent sections is shown for the periods with (2006-2007) and without (2009-2010) step sectioning. Cases with multiple positive SLN were classified based on the largest tumor deposit in an SLN. Abbreviations: SLN, sentinel lymph node; IOTPC, intraoperative touch prep cytology; Macro: macrometastases; Micro: micrometastases; ITC, isolated tumor cells.
2500
B. C. Calhoun et al.
Table 2 Sensitivity of IOTPC and extent of SLN involvement on permanent sections 2006-2007 IOTPC sensitivity Any tumor cells Macrometastases Micrometastases ITCs Permanent sections Macrometastases Micrometastases ITCs
2009-2010
51.4% (72/140) 39.4% (63/160) 71.7% (66/92) 76.3% (58/76) 8.3% (3/36) 5.5% (3/55) 25.0% (3/12) 6.9% (2/29)
P .048 ⁎ .598 .678 .139
65.7% (92/140) 47.5% (76/160) .002 ⁎ 25.7% (36/140) 34.4% (55/160) .131 8.6% (12/140) 18.1% (29/160) .018 ⁎
NOTE. The sensitivity of IOTPC for any tumor cells, macrometastases, micrometastases, and ITCs is shown for the periods with (2006-2007) and without (2009-2010) step sectioning. Cases with multiple positive SLNs were classified based on the largest tumor deposit in an SLN. Abbreviations: IOTPC, intraoperative touch prep cytology; SLN, sentinel lymph node; ITC, isolated tumor cell. ⁎ Statistically significant P b .05 (Fisher exact test).
patients. More recently, the ACOSOG Z0011 investigators [11,12], IBCSG 23-01 investigators [13], NSABP B-32 investigators [5,14], and others [15,16] have concluded that completion ALND is not required for patients with minimal involvement of SLNs. The utilization of intraoperative evaluation of SLNs in breast cancer has changed dramatically secondary to publication of the results of NSABP B-32 and ACOSOG Z0011 [11,12]. Before the publication of ACOSOG Z0011, IOTPC was routinely performed for all breast cancer patients undergoing SLN biopsy at our institution. Now, the use of IOTPC at our institution is mainly reserved for select patients who would not have met the eligibility criteria for ACOSOG Z0011. The study design for NSABP B-32 included the pathologic assessment of lymph nodes. Additional H&E levels and cytokeratin immunohistochemistry were performed in SLN that were negative on initial examination. These additional slides revealed occult tumor in 15.9% of 3887 patients: 11.1% with ITC clusters, 4.4% with micrometastases, and 0.4% with macrometastases [5]. Our experience with limited wide interval step sectioning is consistent with the finding that ITC accounts for a significant proportion of the nodal involvement detected with more intensive evaluation [2]. A possible rationale for maintaining limited step sectioning would be to ensure the detection of all macrometastases in SLN tissue sections that, due to variations in gross sectioning, are more than 2 mm thick [17]. The 5-year follow-up from NSABP B-32 showed a small, statistically significant difference (1.2%) in the overall survival of patients with occult metastases in SLN that were negative on initial examination [5]. The 10-year followup showed no statistically significant difference in overall survival or local-regional recurrence for patients with occult metastases in SLN that were negative on initial examination [14]. The NSABPB-32 investigators concluded that their “…findings argue against analysis of additional tissue levels
or routine immunohistochemical analysis for sentinellymph-node evaluation” [5] and that ALND for occult nodal involvement offers no benefit [14]. This is consistent with the most recent American Society of Clinical Oncology Clinical Practice Guideline Update for SLN biopsy in patients with early stage breast cancer [18]. Specifically, the Update Committee concluded in Recommendation 2.1 that “clinicians should not recommend ALND for women with early stage breast cancer who have one or two SLN metastases and will receive breast-conserving surgery (BCS) with conventionally fractionated whole-breast radiotherapy” [18]. The unproven benefit of intensive SLN evaluation argues against the practice, particularly in an era of cost reduction in health care. The cost of producing an H&E slide is much lower than other aspects of multidisciplinary breast cancer care (eg, operating room time, chemotherapy, and magnetic resonance imaging studies). However, laboratories are often viewed as cost centers, and the headcount for histotechnologists is frequently capped. Our limited step-sectioning protocol doubled the number of H&E slides produced from SLN tissue blocks. If we assume an average of 3.5 SLN per procedure and an average of 1.5 blocks per SLN with a cost of $5 per H&E slide, our step-sectioning protocol added approximately $19 000 to the cost of evaluating the 730 SLN biopsies in 2009 and 2010. The retrospective single-institution nature of this study and lack of long-term follow-up represent limitations of this study. In addition, the implementation of routine axillary ultrasound around the time of the change in the pathology protocol may have further reduced the likelihood of detecting macrometastases intraoperatively and in permanent sections. Analyzing SLNs with 2 H&E slides approximately 500 μm apart led to an increase in final pathologic diagnosis of microscopic metastatic disease in SLNs and subsequent upstaging in breast cancer patients with tumor deposits greater than 0.2 mm. We observed an approximately 10% increase in both micrometastases and ITCs with limited step sectioning. The higher proportion of micrometastases and ITC in 2009 to 2010 was associated with a decrease in the overall sensitivity of IOTPC. Both of these results were anticipated at the time the decision was made to review a second slide 500 μm deeper in the block. The lower proportion of SLN macrometastases in the 2009 to 2010 period may be related to increased utilization of preoperative axillary ultrasound and ultrasound-guided lymph node biopsy to document macrometastases at the time of diagnosis. In addition, the increase in micrometastases and ITC in 2009 to 2010 very likely diluted the macrometastases that were detected in a slightly larger pool of all metastases. Early stage breast cancer patients with limited SLN involvement are not required to undergo completion ALND [18]. The ability of pathologists to detect microscopic tumor in SLN must be balanced with the clinical significance of the findings and implications for treatment.
Limited step sectioning of breast sentinel lymph nodes
References [1] Lyman GH, Giuliano AE, Somerfield MR, et al. American Society of Clinical Oncology guideline recommendations for sentinel lymph node biopsy in early-stage breast cancer. J Clin Oncol 2005;23: 7703-20. [2] Viale G, Mastropasqua MG, Maiorano E, Mazzarol G. Pathologic examination of the axillary sentinel lymph nodes in patients with earlystage breast carcinoma: current and resolving controversies on the basis of the European Institute of Oncology experience. Virchows Arch 2006;448:241-7. [3] Edge SB, Byrd DR, Compton CC, et al, editors. AJCC Cancer Staging Manual. 7th ed. New York: Springer; 2010. [4] Stell VH, Flippo-Morton TS, Norton HJ, White Jr RL. Effect of intraoperative radiocolloid injection on sentinel lymph node biopsy in patients with breast cancer. Ann Surg Oncol 2009;16:2300-4. [5] Weaver DL, Ashikaga T, Krag DN, et al. Effect of occult metastases on survival in node-negative breast cancer. N Engl J Med 2011;364: 412-21. [6] Reed J, Rosman M, Verbanac KM, et al. Prognostic implications of isolated tumor cells and micrometastases in sentinel nodes of patients with invasive breast cancer: 10-year analysis of patients enrolled in the prospective East Carolina University/Anne Arundel Medical Center Sentinel Node Multicenter Study. J Am Coll Surg 2009;208:333-40. [7] Hansen NM, Grube B, Ye X, et al. Impact of micrometastases in the sentinel node of patients with invasive breast cancer. J Clin Oncol 2009;27:4679-84. [8] Kumar S, Bramlage M, Jacks LM, et al. Minimal disease in the sentinel lymph node: how to best measure sentinel node micrometastases to predict risk of additional non-sentinel lymph node disease. Ann Surg Oncol 2010;17:2909-19. [9] Cortesi L, Proietto M, Cirilli C, et al. Prognosis and treatment of micrometastatic breast cancer sentinel lymph node: a population-based study. J Surg Oncol 2012;106:399-405.
2501 [10] Sola M, Alberro JA, Fraile M, et al. Complete axillary lymph node dissection versus clinical follow-up in breast cancer patients with sentinel node micrometastasis: final results from the multicenter clinical trial AATRM 048/13/2000. Ann Surg Oncol 2013;20:120-7. [11] Giuliano AE, Hunt KK, Ballman KV, et al. Axillary dissection vs no axillary dissection in women with invasive breast cancer and sentinel node metastasis: a randomized clinical trial. JAMA 2011;305:569-75. [12] Giuliano AE, McCall L, Beitsch P, et al. Locoregional recurrence after sentinel lymph node dissection with or without axillary dissection in patients with sentinel lymph node metastases: the American College of Surgeons Oncology Group Z0011 randomized trial. Ann Surg 2010; 252:426-32. [13] Galimberti V, Cole BF, Zurrida S, et al. Axillary dissection versus no axillary dissection in patients with sentinel-node micrometastases (IBCSG 23-01): a phase 3 randomised controlled trial. Lancet Oncol 2013;14:297-305. [14] Julian TB, Anderson SJ, Krag DN, Weaver DL. Ten-year follow-up results of occult detected sentinel node disease: NSABP B-32, a randomized phase III clinical trial to compare sentinel node resection (SNR) to conventional axillary dissection (AD) in clinically node-negative breast cancer patients. San Antonio Breast Cancer Symposium; 2013. [15] Fournier K, Schiller A, Perry RR, Laronga C. Micrometastasis in the sentinel lymph node of breast cancer does not mandate completion axillary dissection. Ann Surg 2004;239:859-63. [16] Yegiyants S, Romero LM, Haigh PI, DiFronzo LA. Completion axillary lymph node dissection not required for regional control in patients with breast cancer who have micrometastases in a sentinel node. Arch Surg 2010;145:564-9. [17] Weaver DL, Le UP, Dupuis SL, et al. Metastasis detection in sentinel lymph nodes: comparison of a limited widely spaced (NSABP protocol B-32) and a comprehensive narrowly spaced paraffin block sectioning strategy. Am J Surg Pathol 2009;33:1583-9. [18] Lyman GH, Temin S, Edge SB, et al. Sentinel lymph node biopsy for patients with early-stage breast cancer: American Society of Clinical Oncology clinical practice guideline update. J Clin Oncol 2014;32:1365-83.