An immunohistochemical panel to differentiate


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An immunohistochemical panel to differentiate

  1. 1. Original Articles An Immunohistochemical Panel to DifferentiateMetastatic Breast Carcinoma to Skin From Primary Sweat Gland Carcinomas With a Review of the LiteratureMarian Rollins-Raval, MD, MPH; Mamatha Chivukula, MD; George C. Tseng, ScD; Drazen Jukic, MD, PhD; David J. Dabbs, MDN Context.—Approximatelymetastases. Sweat gland breastcancer develop cutaneous 25% of patients with carci- neoplasm cases, and 2 primary breast cancer cases were retrieved and analyzed with the following IHC panel:nomas (SGCs) account for about 0.05% of all cutaneous mammaglobin, gross cystic disease fluid protein (GCDFP)neoplasms. Cutaneous metastases of breast carcinoma 15, p63, basal cytokeratins (CK5, CK14, and CK17),(CMBCs) (especially the ductal type) can be difficult to androgen receptor, and PAX5.distinguish from SGCs. Treatment and prognoses for these Results.—The p63 was only weakly expressed in 1 of 122 types of tumors differ radically, making accurate CMBC cases (8.3%), whereas it was strongly expressed inhistologic diagnosis crucial. Although a few studies attempt 10 of 11 SGC cases (90.9%) (P , .001). Basal cytokeratinsto differentiate these entities employing immunohisto- demonstrated a similar immunoprofile in the SGC group,chemical (IHC) studies (some of which we review here), to with 10 of 11 cases (90.9%) expressing all 3 markers, and adate, no panel of IHC stains exists, to our knowledge, to variable immunoprofile in the CMBC group with 0%distinguish these entities. (CK14) (P , .001) to 16.7% (2 of 12 cases; CK5 and CK17) Objective.—To devise a panel of IHC stains to distin- (P , .001) expression. Mammaglobin was expressed in 8 ofguish CMBC from SGC. 12 cases (66.7%) of CMBC. Design.—Twelve cases of ductal CMBCs (11 not Conclusions.—Together, these 5 IHC stains were com-otherwise specified type, and 1 basal phenotype), 11 cases bined to make a panel that was 100% sensitive and 91%of SGCs (5 eccrine carcinomas, 3 porocarcinomas, and 3 specific in distinguishing between CMBC and SGC.microcystic adnexal carcinomas), 2 benign sweat gland (Arch Pathol Lab Med. 2011;135:975–983)Approximately accountmetastases. Sweat gland carci- 25% of patients with breast cancer may develop cutaneousnomas (SGCs) 1,2 for 0.05% of all cutaneous reports continue to demonstrate this potential diagnostic pitfall.4,5 We recently received 2 diagnostically challenging cases presenting similar challenges.neoplasms.3 Clinically, the presentation of these 2 entitiesis often distinct. Ductal cutaneous metastases of breastcarcinoma (CMBCs) present as multiple lesions in patientswith a previous diagnosis of primary breast carcinoma(PBC), whereas SGC presents as a single cutaneous lesionin patients with no known history of breast cancer.However, CMBCs can be difficult to distinguish fromSGCs histologically, and complex clinical circumstancescan further complicate a diagnosis. Several recent case Accepted for publication October 12, 2010. From the Department of Pathology, University of Pittsburgh MedicalCenter, Pittsburgh, Pennsylvania (Drs Rollins-Raval, Chivukula, Jukic,and Dabbs); and the Department of Biostatistics, University of Pittsburgh(Dr Tseng). Dr Jukic is now with the Division of Dermatopathology,Department of Pathology and Laboratory Medicine, James A. HaleyVeterans’ Hospital, Tampa, Florida. The authors have no relevant financial interest in the products orcompanies described in this article. Presented in part at the annual meeting of the United States andCanadian Academy of Pathology, Denver, Colorado, March 5, 2008. Figure 1. Eccrine carcinoma ex spiradenocylindroma with inset Reprints: Marian Rollins-Raval, MD, MPH, Department of Pathology, showing the eccrine carcinoma ex spiradenocylindroma at higherUniversity of Pittsburgh Medical Center, A711 Scaife Hall, 3550 Terrace power (hematoxylin-eosin, original magnifications 3100 and 3400St, Pittsburgh, PA 15261 (e-mail: [inset]).Arch Pathol Lab Med—Vol 135, August 2011 IHC Panel of CMBC Versus SGC—Rollins-Raval et al 975
  2. 2. Figure 2. Infiltrating carcinoma shown with hematoxylin-eosin stain (A), as well as immunohistochemical stains for p63 (B), AE1/AE3 (C), CK5/6(D), epithelial membrane antigen (E), and CK7 (F) (original magnifications 3200). Case 1 7 years after her bilateral mastectomies. Although the morphology of the PBC and the skin lesions was similar, A 75-year-old woman with a history of bilateral immunohistochemical (IHC) studies were performed onmastectomies for infiltrating ductal carcinoma of both all lesions. For the right PBC, the tumor cells demonstrat-breasts, colectomy for carcinoma of the rectum, and ed strong positivity for cytokeratin (CK) 7, estrogenhysterectomy with bilateral salpingo-oophorectomy for receptor (ER), and ERBB2 (formerly HER2/neu) and focalbenign disease presented with skin lesions on her back positivity for progesterone receptor (PR). For the left PBC,976 Arch Pathol Lab Med—Vol 135, August 2011 IHC Panel of CMBC Versus SGC—Rollins-Raval et al
  3. 3. Table 1. Demographics of Cutaneous Metastatic in Figures 2, B through F, respectively), and were negative Breast Carcinoma (CMBC) and Sweat Gland Carcinoma for S100, ER, PR, CK20, GCDFP-15, and mammaglobin. (SGC) Cases Based on this information alone, it was unclear whether Mean Age the lesion was a CMBC or an SGC, making the devel- Case Group No. of Cases % Female (Age Range), y opment of a treatment plan difficult. As the treatment and prognosis for CMBC and SGC PBC 2 100 68 (53–83) CMBC 12 100 57 (40–80)a differ significantly, developing a panel of IHC markers to BAN 2 50 48.5 (34–63) differentiate these 2 entities would be of high clinical SGC 11 58.3 73.6 (49–93) value. Numerous studies have evaluated these entitiesAbbreviations: BAN, benign adnexal neoplasm; PBC, primary breast either individually6–15 or comparatively using variouscarcinoma. (IHC) stains,16–23 but there has been no study, to oura Age at presentation of metastatic lesion, not primary breast lesion. knowledge, to determine the most useful IHC panel for differentiating CMBC from SGC. Several IHC stains have shown promise in their abilitythe tumor cells were strongly positive for CK7, ER, and PR. to differentiate CMBC and SGC. Mammaglobin, a proteinBoth PBCs were negative for CK903, S100, cross cystic fluid frequently overexpressed in breast cancer,11 has not beenprotein–15 (GCDFP-15), and CK20. On hematoxylin-eosin thoroughly examined in either CMBC or SGC. GCDFP-15,stain, the skin lesion, which demonstrated areas of tight a glycoprotein originally isolated in human breast grossintermingling between carcinoma and a spiradenocylin- cystic fluid, although present in most primary anddroma, was more compatible with a malignant neoplasm metastatic breast cancers, has also been reported to bearising in the background of a benign mixed tumor rather expressed in a few cases of SGC.16,24 Androgen receptorthan metastasis to the benign tumor (Figure 1). In the (AR), although often found in high-grade invasive breastbenign component, CK7 was positive only in the internal cancer, has also been identified in some SGCs.7–9,13mature cells and negative in basal cells, ERBB2 demon- The basal cytokeratin stains (CK5, CK14, and CK17),strated weak and less than 1+ positivity, and CK903 was recently shown to be expressed in the basal phenotype ofstrongly positive. CK20, ER, PR, and S100 were all negative breast carcinoma, were not present in most other types ofin the tumor cells. In the malignant component, CK903 was PBC.25,26 These stains have also been shown to be presentstrongly positive, whereas CK7 and S100 were focally in a few SGC cases.14,19,20 However, with the exception ofpositive. CK20, GCDFP-15, ER, PR, and ERBB2 were all CMBC with a basal phenotype, these markers are morenegative. Although these findings supported the diag- likely to be seen in SGC; p63, a homologue of the p53 genenosis of a primary eccrine carcinoma (EC) arising in a and expressed primarily by basal and myoepithelial cellsspiradenocylindroma, the possibility of a CMBC to the of the skin, would be more likely to be seen in cases of SGCbenign neoplasm could not be ruled out. Molecular studies than in cases of CMBC.10,21,22performed on the paraffin-embedded tissues showed In addition, 1 of the authors (M.R.-R.) noticed a strongmultiple allelic losses in the PBC and no genetic alterations cytoplasmic and/or membranous staining with PAX5, ain the EC. The patient subsequently developed additional B-cell–specific activator protein of the basal layer ofmetastases to lymph nodes and additional skin sites from healthy epithelium, in an unrelated research study andthe EC. Although a definitive diagnosis was rendered in decided to use the antibody in the current study,this case, a simplified IHC panel would have benefited hypothesizing that the staining pattern might be similarboth the pathologist and patient. in SGC. The PAX5 expression has been previously studied in B lymphocytes, in most small cell carcinomas, and in Case 2 Merkel cell carcinomas.27,28 An 84-year-old woman with a history of bilateral PBC The goal of this study was to construct a panel of IHC20 years earlier presented with a skin lesion of the right stains that would be highly sensitive and specific inbreast. The morphology of the previous PBC was not distinguishing these 2 morphologically similar entities.consistent with the new skin lesion, which showed aninfiltrating carcinoma (Figure 2, A). Numerous IHC stains MATERIALS AND METHODSwere performed with the following results: the tumor cells Literature Reviewwere positive for p63, AE1/AE3, CK5/6, and epithelial A MEDLINE (US National Library of Medicine, Bethesda,membrane antigen, were focally positive for CK7 (shown Maryland) search was performed from 1995 to 2009 to identify Table 2. Immunohistochemistry Antibody Information a Antibody Clone Dilution Vendor Location Pretreatment GCDFP 23A3 1:25 Vector Labs Burlingame, California CC1b mild and standard AR AR441 1:100 Dako North America Carpinteria, California CC1 mild and standard Mammaglobin 304-1A5 and 31A5 Predilute Zeta Corporation Sierra Madre, California CC1 mild p63 4A4 1:200 Neomarkers Inc Fremont, California CC1 mild and standard PAX5 24 1:25 BD Biosciences San Jose, California CC1 mild and standard CK5 XM26 1:25 Novocastra Newcastle Upon Tyne, CC1 standard Laboratories Ltd United Kingdom CK14 LL002 Predilute Ventana Medical Tucson, Arizona CC1 standard Systems Inc CK17 E3 1:20 Dako North America Carpinteria, California CC1 mild and protease 3a All immunostains were performed on the BenchMark XT using the iView/DAB detection system (Ventana Medical Systems, Tucson, Arizona).b Ventana Medical Systems, Tucson, Arizona.Arch Pathol Lab Med—Vol 135, August 2011 IHC Panel of CMBC Versus SGC—Rollins-Raval et al 977
  4. 4. Table 3. Comparative Review of Additional Similar Studies and the Immunohistochemical Results Study Wallace et al,16 1995 Wick et al,17 1998 Busam et al,18 1999 Plumb et al,19 2004 a b IHC Marker MAC CMBC EC PBC MAC PC EC CMBC MAC CMBCc ER 2/8 1/15 9/27 31/59 2/8 0/4 2/13 10/30 PR 5/8 15/15 1/4 27/59 2/8 0/4 2/13 8/30 c-erbB-2(HER2/neu) 9/27 12/59 AR GCDFP 1/6 7/15 5/27 41/59 CEA 21/27 5/59 EGF-R 5/8 4/4 11/13 5/30 CK5/6 3/3 2/17 CK5 CK7 CK14 CK17 CK20 p63 (nuclear) S100 12/27 27/59 Podoplanin PAX5 (C/M)g MammaglobinAbbreviations: 1A, 1 apocrine case; 1N, 1 not otherwise specified case; AR, androgen receptor; CEA, carcinoembryonic antigen; CK, cytokeratin; C/M, cytoplasmic/membranous; CMBC, cutaneous metastatic breast carcinoma (ductal); EC, eccrine carcinoma; EGF-R, epidermal growth factorreceptor; ER, estrogen receptor; ES, ex spiradenocylindroma; GCDFP, gross cystic disease fluid protein; IHC, immunohistochemistry; MAC,microcystic adnexal carcinoma; PBC, primary breast carcinoma; PC, porocarcinoma; PR, progesterone receptor.a In this study, EC refers to 23 eccrine carcinomas, as well as 4 apocrine lesions in the same category that were only separated by their staining for GCDFP-15, where 1 of 23 cases of eccrine carcinoma was positive whereas 4 of 4 cases of the apocrine lesions were positive for GCDFP.b These authors looked at primary breast carcinomas as opposed to cutaneous metastases.c There were 6 cases with weak staining, considered negative, for CK5/6.d Of the 3, 1 was mucinous and stained only 1+ for p63 and was thus counted as negative here; the other 2 were strongly positive.e There were 5 cases: 2N, 1A, 2ES.f Two of 3 were graded as 1+.g PAX5 showed a strong C/M staining pattern noted previously in normal epithelium which is reported here.studies similar to our own, comparing primary SGC to CMBC (or paraffin-embedded tissues; and samples were analyzed with thein 1 study, to PBC) using IHC stains. We selected a panel of 8 IHC following IHC stains: mammaglobin, GCDFP-15, p63, CK5, CK14,stains to investigate their staining patterns in CMBC and SGC. CK17, AR, and PAX5.We did not repeat IHC studies that appeared in previous studies All IHC stain analysis was performed on the Ventanato be less useful. BenchMark, XT (Ventana Medical Systems, Inc, Tucson, Arizona) using the iView DAB Detection System (Ventana Medical) with Case Identification commercially available antibodies according to standard proto- A retrospective search for CMBC and SGC cases using a Natural cols (Table 2). All negative and positive controls demonstratedLanguage Search was performed in the University of Pittsburgh appropriate immunolabeling.Medical Center (Pittsburgh, Pennsylvania) CoPath Plus database The IHC stain results were semiquantitated as follows: AR,(Cerner DHT, Inc, Waltham, Massachusetts) for the 9 years from 1998 PAX5, and p63 were nuclear stains, and strong nuclear positivityto 2007. Based on the results of the database search, 27 cases were was considered positive staining. The intensity of immunostain-retrieved from the University of Pittsburgh Medical Center archives ing was graded as negative (0), weak (1), moderate (2), andfor the study: 12 cases of ductal CMBC (44%), which included 11 strong (3), and the proportion of positive staining cells wascases with morphologies not otherwise specified and 1 case of basal recorded as 0% through 5% (1), 6% through 20% (2), 21% throughphenotype; 11 cases of SGC (41%), which included 5 cases of EC, 3 80% (3), and greater than 80% (4).cases of porocarcinoma (PC), and 3 cases of microcystic adnexal Cytoplasmic staining was considered positive for GCDFP-15carcinoma (MAC); and 4 additional, randomly selected cases (15%), and mammaglobin, and the intensity of immunostaining waswhich included 2 primary cutaneous adnexal benign neoplasms (a graded as weak, moderate, or strong.poroma and an apocrine adenoma) and 2 cases of PBC. Basal cytokeratins—CK5, CK14, and CK17—were scored as positive if any cytoplasmic or membranous staining was Demographics observed in the tumor cells. The patients in PBC and metastatic breast cancer groups were all A strong cytoplasmic and/or membranous staining patternwomen, whereas the SGC group consisted of both men and women. was considered positive for PAX5.The mean ages for patients with ductal CMBC and SGC were57 years and 73.6 years, respectively (Table 1). The difference in ages Statistical Analysiswas significantly different. Although all the patients with CMBC (12 A statistical analysis of the positive and negative results wasof 12; 100%) had documented PBCs, only 1 (case 1, described above) performed with R statistical software 2.10.1 (R Project forof the 11 patients with SGC (9%) had a history of PBC. Statistical Computing, Wien, Austria). The SGCs were all compiled into one group for the statistical analysis. Comparisons of the IHC staining profiles in the CMBC and SGC groups were Immunohistochemistry performed using a z test to calculate the P values for each Hematoxylin-eosin slides were reviewed; a representative tumor IHC stain. Subsequently, a simple computer algorithm wasblock was selected; sections were obtained on formalin-fixed, created to examine the effectiveness of a combination of978 Arch Pathol Lab Med—Vol 135, August 2011 IHC Panel of CMBC Versus SGC—Rollins-Raval et al
  5. 5. Table 3. Extended 20 22 Study Qureshi et al, 2004 Ivan et al, 2005 Liang et al,23 2007 This Study d IHC Marker PC CMBC MAC EC CMBC PC CMBC MAC PC ECe CMBC ER PR c-erbB-2(HER2/neu) AR 0/3 2/3 0/5 5/12 GCDFP 0/3 0/3 1/5 (ES) 5/12 CEA EGF-R CK5/6 3/3 0/6 CK5 3/3 3/3 4/5 2/12 CK7 2/3f 6/6 CK14 3/3 3/3 4/5 0/12 CK17 3/3 3/3 4/5 2/12 CK20 0/3 0/6 p63 (nuclear) 3/3 0/6 4/4 2/3 0/12 3/3 3/3 4/5 1/12 S100 Podoplanin 6/6 0/11 PAX5 (C/M)g 3/3 2/3 2/5 (1A + 2N) 0/12 Mammaglobin 0/3 0/3 2/5 (ES) 7/12the near statistically significant and statistically significant with CK5/6, with 6 cases (35%) having weak positivebiomarkers. staining, whereas all 3 MACs (100%) expressed strong positive staining. This difference, although not evaluated RESULTS specifically in the study,19 revealed the diagnostic poten- Literature Review tial of CK5/6 in differentiating these lesions. Wallace et al16 investigated the IHC staining character- Qureshi et al20 also examined the diagnostic potential ofistics of 15 cases (from 12 patients) of CMBC and several IHC stains, including p63, CK5/6, CK7, and CK20,compared them to a series of primary eccrine tumors, to differentiate metastatic carcinomas from primaryincluding 8 MACs. The authors16 concluded that using cutaneous adnexal neoplasms. Several benign adnexalstandard IHC markers, such as ER, PR and GCDFP-15, neoplasms as well as malignant neoplasms, such aswould not reliably distinguish these 2 populations. hidradenocarcinoma, adenoid cystic carcinoma, seba- Wick et al17 examined 59 cases of ductal PBCs and ceous carcinoma, digital papillary adenocarcinoma, syr-compared them with 27 cases of ductal SGC, which were ingomatous carcinoma, and mucinous carcinoma, werefurther described as demonstrating eccrine morphology included in their study.20 Only their examinations of PCs(23 cases; 85%) and apocrine differentiation (4 cases; 15%). and CMBCs were included in Table 3. Three cases of PCUsing pancytokeratins, carcinoembryonic antigen, S100 were all strongly positive for CK5/6 and p63, whereasprotein, GCDFP-15, ER, PR, and c-erbB-2 protein (ERBB2) CK7 demonstrated positivity in less that 10% of cells for 2IHC stains, their findings17 concluded that the infrequency cases, and CK20 was completely negative in all cases. Theof GCDFP-15 in eccrine sweat gland tumors as well as the 6 CMBC cases were diffusely positive for CK7 and werepaucity of carcinoembryonic antigen in breast carcinomas negative for CK5/6, p63, and CK20. This study20 demon-could be useful in predefined differential diagnostic strated the diagnostic potential of CK5/6 and p63 insettings involving these 2 entities with the appropriate differentiating these 2 entities.clinicopathologic information provided. We included this Ivan et al22 assessed the utility of p63 antibody forstudy in our review because of its similarity to the current differentiating primary cutaneous adnexal neoplasms andstudy, even though it focused on distinguishing PBC from adenocarcinoma metastatic to the skin. In addition toSGC, rather than distinguishing CMBC from SGC. several benign adnexal tumors, the authors22 analyzed 4 Busam et al18 studied 30 cases of CMBC compared with MACs and 3 ECs (1 of the latter with mucinous42 primary SGC cases for their expression of ER, PR, and differentiation) as well as a case of hidradenocarcinomaepidermal growth factor receptor (EGFR). Several addi- and 2 cases of trichilemmal carcinomas. Of the MACs andtional histologic types of SGC were examined in this ECs, only the ECs with mucinous differentiation werestudy,18 including apocrine, hidradenocarcinoma, mucin- negative for nuclear positivity of p63, and none of theous, and basaloid carcinomas, which were not included in CMBCs demonstrated nuclear positivity, confirming theour study. Only the 3 types of tumors included in our usefulness of this marker in distinguishing SGC fromstudy—MAC, PC, and EC—are presented in Table 3. CMBC.Their results18 suggested that the expression of EGFR may Finally, Liang et al23 investigated the use of podoplaninbe diagnostically helpful in differentiating these 2 groups to differentiate metastases to skin from various organof tumors, whereas ER and PR continued to show no sites, including the breast, from primary skin adnexalsignificant difference between the 2 groups. carcinomas. In their study,23 the authors examined 11 Plumb et al19 used CK5/6 to differentiate primary cases of metastatic breast cancer to the skin, all of whichcutaneous adnexal neoplasms, including 3 MACs, from (100%) were completely negative for podoplanin. Theycutaneous metastatic lesions, including 17 CMBCs. Only 2 also examined a total of 40 primary skin adnexalof 17 CMBC cases (12%) displayed strong positive staining carcinomas, only 2 of which (5%; a case of adenoid cysticArch Pathol Lab Med—Vol 135, August 2011 IHC Panel of CMBC Versus SGC—Rollins-Raval et al 979
  6. 6. Table 4. Summary of Immunohistochemistry (IHC) Profile Staining Pattern in Cutaneous Metastatic Breast Carcinoma (CMBC) and Sweat Gland Carcinoma (SGC) Mammaglobin, PAX5 (C/M),b IHC Staina % (No.) p63, % (No.) CK5, % (No.) CK14, % (No.) CK17, % (No.) % (No.) CMBC, N 5 12 66.7 (8) 8.3 (1) 16.7 (2) 0 (0) 16.7 (2) 0 (0) SGC, N 5 11 18.2 (2) 90.9 (10) 90.9 (10) 90.9 (10) 90.9 (10) 54.5 (6) P value .06c ,.001 ,.001 ,.001 ,.001 .01Abbreviation: CK, cytokeratin; C/M, cytoplasmic/membranous.a The differences in staining between CMBC and SGC with androgen receptor (P 5 .25) and gross cystic disease fluid protein–15 (P 5 .19) were not statistically significant.b PAX5 did not show nuclear staining in either tumor but did show strong C/M staining, which is reported here.c Marginally significant.and a case of poorly differentiated adnexal carcinoma) Immunohistochemical Analysiswere negative for podoplanin. However, because the Table 4 provides a summary of IHC profile stainingdistinction between SGC and metastatic adenocarcinomas pattern in CMBC and SGC. The p63 was only weaklymay be equivocal in many cases, the only SGCs included expressed in 8.3% (1 of 12) of the CMBC cases, whereas itin their study were the 6 PCs, which were all (100%) was strongly expressed in 90.9% (10 of 11) of the SGC casespositive for podoplanin. The authors23 suggest that (P , .001). All 3 basal CKs were expressed in 90.9% (10 ofadditional studies may be necessary in the future to 11) of the SGC cases. In comparison, CMBC casesevaluate more SGCs for podoplanin. From that limited demonstrated a staining profile of 0% (0 of 12) for CK14study,23 podoplanin appeared to show promise in distin- and 16.7% (2 of 12) for CK5 and CK17. One case of CMBCguishing SGC from CMBC. (8.3%) expressed both CK and CK17, whereas 2 more cases In our study, the only 2 SGCs that were positive for AR of CMBC (16.7%) were positive for either CK5 or CK17. Thewere 2 of the 3 PCs (67% of the PCs; 18% of all SGCs). difference in basal CK staining among the SGC cases andAlthough most adnexal carcinomas were negative for the CMBC cases was statistically significant for all 3 stains (PGCDFP-15, only 1 of the 2 ECs (50%), ex spiradenocylin- , .001). Mammaglobin expression was seen in 66.7% (8 ofdroma,29 was positive. That same carcinoma was positive 12) of the CMBC cases, compared with 18.2% (2 of 11) of thefor mammaglobin as well, while staining negative for all SGC group (P 5 .06), which was marginally statisticallyother stains. The second EC ex spiradenocylindroma was significant. The difference in expression for both AR orpositive for mammaglobin, while also staining positive for GCDFP-15 in the 2 groups was not statistically significant.p63, CK5, CK14, and CK17. The appearance of a typical CMBC is demonstrated in Although most of the CMBC cases in this study were Figure 3, A, whereas a typical SGC (a PC) is demonstratednegative for the basal CK markers, 3 of the 12 cases (25%) in Figure 3, B. Using these 2 cases as examples, the mostwere positive for either or both CK5 and CK17. One case prominent staining pattern for each group is representedthat was positive for both of these immunostains was in Figure 4. The CMBCs were generally positive formorphologically of a basal phenotype. mammaglobin and negative for p63, CK5, CK14, andFigure 3. Examples of ductal cutaneousmetastatic breast carcinoma (A) and sweatgland carcinoma (porocarcinoma) (B) (hema-toxylin-eosin, original magnifications 3100[A and B] and 3400 [insets]).980 Arch Pathol Lab Med—Vol 135, August 2011 IHC Panel of CMBC Versus SGC—Rollins-Raval et al
  7. 7. Figure 4. Immunohistochemical panel applied to ductal cutaneous metastatic breast carcinoma: mammaglobin (A), p63 (B), CK5 (C), CK14 (D),and CK17 (E); and to sweat gland carcinoma (porocarcinoma): mammaglobin (F), p63 (G), CK5 (H), CK14 (I), and CK17 (J) (original magnifications3100 [A through J] and 3400 [insets]).CK17 (Figure 4, A through E, respectively), whereas SGCs (left inset), as well as the benign basal layer epithelium(represented in the figure by a PC) were generally showing strong cytoplasmic/membranous staining (rightnegative for mammaglobin and positive for p63, CK5, inset).CK14, and CK17 (Figure 4, F though J, respectively). Only 5 out of the 8 stains (63%) examined by accepted None of the CMBC or SGC tumor cells (0%) demon- criteria demonstrate statistically significant, or nearstrated nuclear staining for PAX5 (as shown in the right statistically significant, results. Incorporating those 5inset of Figure 5, B). However, 54.5% (5 of 11) of the cases IHC markers into a sum score system, we constructed aof SGC expressed a distinct cytoplasmic and/or membra- panel to predict the disease represented in each case. Thenous staining pattern (Figure 5, B). Figure 5, A, demon- conditions set were based on the assumption that breaststrates the faint cytoplasmic blush seen in ductal CMBC cancer was usually expected to demonstrate the following Figure 5. PAX5 cytoplasmic/membranous staining. A, Ductal cutaneous metastatic breast carcinoma showing only faint cyto- plasmic blush (left inset) and a benign basal layer of epithelium showing strong cytoplas- mic/membranous staining (right inset). B, Sweat gland carcinoma (porocarcinoma) showing diffuse cytoplasmic/membranous pattern (left inset) with scattered B lympho- cytes showing the classic nuclear pattern (right inset) (original magnifications 3100 [A and B] and 3400 [insets]).Arch Pathol Lab Med—Vol 135, August 2011 IHC Panel of CMBC Versus SGC—Rollins-Raval et al 981
  8. 8. Table 5. Immunohistochemistry Condition Panela Case ID Diagnosis MGB+ p632 CK52 CK142 CK172 Score Groupb CMBC-1 Ductal, NOS type 1 0 0 0 0 1 1 CMBC-2 Ductal, NOS type 1 0 0 0 0 1 1 CMBC-3 Ductal, NOS type 0 0 0 0 0 0 1 CMBC-4 Ductal, NOS type 0 0 0 0 0 0 1 CMBC-5 Ductal, NOS type 0 0 0 0 0 0 1 CMBC-6 Ductal, basal phenotype 0 0 1 0 1 2 1 CMBC-7 Ductal, NOS type 1 0 0 0 0 1 1 CMBC-8 Ductal, NOS type 0 0 0 0 0 0 1 CMBC-9 Ductal, NOS type 0 0 0 0 0 0 1 CMBC-10 Ductal, NOS type 1 1 0 0 0 2 1 CMBC-11 Ductal, NOS type 0 0 0 0 1 1 1 CBMC-12 Ductal, NOS type 0 0 1 0 0 1 1 SGC-1 Eccrine carcinoma 1 1 1 1 1 5 2 SGC-2 Eccrine carcinoma, ex 0 0 0 0 0 0 1 spiradenocylindroma SGC-3 Porocarcinoma 1 1 1 1 1 5 2 SGC-4 Porocarcinoma 1 1 1 1 1 5 2 SGC-5 Microcystic adnexal carcinoma 1 1 1 1 1 5 2 SGC-6 Eccrine carcinoma 1 1 1 1 1 5 2 SGC-7 Porocarcinoma 1 1 1 1 1 5 2 SGC-8 Microcystic adnexal carcinoma 1 1 1 1 1 5 2 SGC-9 Microcystic adnexal carcinoma 1 1 1 1 1 5 2 SGC-10 Eccrine carcinoma 1 1 1 1 1 5 2 SGC-11 Eccrine carcinoma, ex 0 1 1 1 1 4 2 spiradenocylindromaAbbreviations: CK, cytokeratin; CMBC, cutaneous metastatic breast carcinoma; ID, identification; MGB, mammaglobin; NOS, not otherwisespecified; SGC, sweat gland carcinoma.a Condition is the presumed result for metastatic breast carcinoma given the significant difference between CMBC and SGC for each of the 5 immunohistochemistry stains (see Table 4).b Group 1, ,3 conditions not met 5 CMBC; group 2, $3 conditions not met 5 SGC.IHC staining profile: mammaglobin+, p632, CK52, CK142, patterns of CMBC and SGC. Ultimately, combiningCK172. mammaglobin, p63, CK5, CK14, and CK17, we construct- For each condition that was not met, 1 point was added. ed the IHC panel described above that consistentlyIf the score was less than 3 of 5 (0, 1, or 2; ,60%), the case differentiates CMBC from SGC in our cases.was defined as CMBC; if it was greater than or equal to 3 of Numerous difficulties have hindered researchers in the5 (3, 4, or 5; $60%), it was defined as SGC. Using this sum identification of a clinically useful IHC panel to distin-score system with these conditions, 12 of 12 patients with guish these entities: the paucity of material, varyingCMBC (100% sensitivity) were correctly identified as were morphologic appearances of the entities, and differences10 of 11 patients with SGC (91% specificity) (Table 5). in IHC staining techniques across laboratories are only a few. At the outset of this study, the number of cases COMMENT identified was few and reflected the rarity of these After reviewing several studies that also attempted to neoplasms. In addition, there was considerable heteroge-differentiate breast cancer and SGC, we attempted to neity among the groups of tumors examined. Within theidentify the most specific antibodies to differentiate these classifications of CMBC and SGC, rare subtypes existed2 neoplasms. The ER, PR, CK7, and CK20 stains were not that were challenging to evaluate. For example, only oneeffective in differentiating these entities. The GCDFP-15, case of basal-phenotype CMBC was included in our study.carcinoembryonic antigen, EGFR, CK5/6, podoplanin, The IHC staining pattern of that CMBC subtype includedand p63 stains all showed potential based on previous known positivity for the basal cytokeratins (CK5, CK14,studies. We further investigated 3 of these 6 IHC stains and CK17).26 Although that one case expressed positivity(GCDFP-15, CK5/6, and p63). In addition, we incorporat- for both CK5 and CK17, it still fulfilled the criteria of theed 5 additional IHC stains (CK14, CK17, AR, mammaglo- panel for classification of the neoplasm as a CMBC.bin, and PAX5), which had not been previously employed, Another rare entity with possible confounding IHCto our knowledge, in differentiating these lesions. Our staining would be metaplastic breast carcinoma. Althoughlimited panel did not include carcinoembryonic antigen, its morphologic characteristics can be quite distinct, it hasEGFR, or podoplanin. Carcinoembryonic antigen and been reported to be positive for p63.10 The staining patternEGFR had not shown as much promise in the studies of that entity for basal CK has not been extensivelyreviewed as other IHC stains we wished to include. The evaluated. Both of these subtypes of breast cancer warrantpromising utility of podoplanin (published after the further investigation of their unique IHC staining pat-completion of our study) was unknown during our terns.investigation. For SGC, 2 cases of EC ex spiradenocylindroma were Our study demonstrated a sustained potential of CK5/6 included. These extremely rare neoplasms demonstrated(or CK5 in our study) and p63 in distinguishing CMBC areas of apocrine differentiation. Undoubtedly, furtherfrom SGC. The GCDFP-15 stain did not reveal a study of a larger cohort of SGC cases with apocrinestatistically significant difference between the staining differentiation would be desirable. However, of these 2 EC982 Arch Pathol Lab Med—Vol 135, August 2011 IHC Panel of CMBC Versus SGC—Rollins-Raval et al
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Homologousits previously reported expression in PBC.11 This finding carcinomas of the breasts, skin, and salivary glands: a histologic and immunohistochemical comparison of ductal mammary carcinoma, ductal sweatmay indicate preservation of this marker from PBC to the gland carcinoma, and salivary duct carcinoma. Am J Clin Pathol. 1998;109(1):metastases. 75–84. Also, the novel interpretation of PAX5 in a cytoplas- 18. Busam KJ, Tan LK, Granter SR, et al. Epidermal growth factor, estrogen, and progesterone receptor expression in primary sweat gland carcinomas andmic and/or membranous staining pattern may provide primary and metastatic mammary carcinomas. Mod Pathol. 1999;12(8):786–793.a specific marker for tumors of adnexal origin. The 19. Plumb SJ, Argenyi ZB, Stone MS, De Young BR. Cytokeratin 5/6known role of PAX5 as a transcription factor was immunostaining in cutaneous adnexal neoplasms and metastatic adenocarcino-functionally consistent with its commonly described ma. Am J Dermatopathol. 2004;26(6):447–451. 20. Qureshi HS, Ormsby AH, Lee MW, Zarbo RJ, Ma CK. The diagnostic utilitynuclear localization in B lymphocytes. However, the of p63, CK5/6, CK 7, and CK 20 in distinguishing primary cutaneous adnexalcytoplasmic and/or membranous staining pattern was neoplasms from metastatic carcinomas. J Cutan Pathol. 2004;31(2):145–152.striking and raises the possibility of alternative func- 21. Ivan D, Nash JW, Prieto VG, et al. Use of p63 expression in distinguishing primary and metastatic cutaneous adnexal neoplasms from metastatic adenocar-tions in cellular pathways. This staining pattern war- cinoma to skin. J Cutan Pathol. 2007;34(6):474–480.rants further investigation. 22. Ivan D, Hafeez Diwan A, Prieto VG. Expression of p63 in primary cutaneous adnexal neoplasms and adenocarcinoma metastatic to the skin. Mod CONCLUSIONS Pathol. 2005;18(1):137–142. 23. Liang H, Wu H, Giorgadze TA, et al. Podoplanin is a highly sensitive and We formed a highly sensitive and specific IHC panel, specific marker to distinguish primary skin adnexal carcinomas from adenocar-composed of mammaglobin, p63, and 3 basal cytokeratins, cinomas metastatic to skin. Am J Surg Pathol. 2007;31(2):304–310. 24. Ormsby AH, Snow JL, Su WP, Goellner JR. Diagnostic immunohisto-with sufficient power to aid in the differentiation between chemistry of cutaneous metastatic breast carcinoma: a statistical analysis of theCMBC and SGC. We recommend the use of this panel to utility of gross cystic disease fluid protein-15 and estrogen receptor protein. J Amdifferentiate most cases of these 2 entities in routine Acad Dermatol. 1995;32(5, pt 1):711–716. 25. Bhargava R, Beriwal S, McManus K, Dabbs DJ. CK5 is more sensitive thanclinical practice. CK5/6 in identifying the ‘‘basal-like’’ phenotype of breast carcinoma. Am J Clin Pathol. 2008;130(5):724–730. We thank Cary Sipos, HT (ASCP) and Kim McManus, HT 26. Dabbs DJ, Chivukula M, Carter G, Bhargava R. Basal phenotype of ductal(ASCP) for their technical assistance. We would also like to carcinoma in situ: recognition and immunohistologic profile. Mod Pathol. 2006;thank Jay S. Raval, MD (Department of Pathology, University of 19(11):1506–1511.Pittsburgh Medical Center) and Darice Y. Wong, PhD (Depart- 27. Feldman AL, Dogan A. Diagnostic uses of Pax5 immunohistochemistry.ment of Bioengineering, University of California, Los Angeles) Adv Anat Pathol. 2007;14(5):323–334.for thoughtful discussions and critical reviews of earlier versions 28. Mhawech-Fauceglia P, Saxena R, Zhang S, et al. Pax-5 immunoexpression in various types of benign and malignant tumours: a high-throughput tissueof this manuscript. Financial support for this article was microarray analysis. J Clin Pathol. 2007;60(6):709–714.provided through the University of Pittsburgh Medical Center’s 29. Carlsten JR, Lewis MD, Saddler K, et al. Spiradenocylindrocarcinoma: aDepartment of Pathology. malignant hybrid tumor. J Cutan Pathol. 2005;32(2):166–171.Arch Pathol Lab Med—Vol 135, August 2011 IHC Panel of CMBC Versus SGC—Rollins-Raval et al 983