Immunohistochemical localization of carcinoembryonic antigen (CEA) in salivary gland tumors Miriam

Alfaro,

CD,” and Michael

TEMPLE

UNIVERSITY

AND

TEMPLE

Carrozza, UNIVERSITY

CT (ASCP)b

Philadelphia,

pa.

HOSPITAL

Carcinoembryonic antigen (CEA) was first isolated from colonic carcinoma and has been used as a diagnostic marker. CEA has also been observed in a variety of epithelial tumors and normal tissues. In this study, CEA was localized by means of immunohistochemical procedures in benign and malignant salivary gland tumors, as well as in normal parotid gland, indicating that CEA is not a reliable marker for differentiation between benign and malignant salivary gland neoplasms. (ORAL

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C arcinoembryonic antigen glycoprotein with a molecular

(CEA) is an oncofetal weight of 200,000 daltons, which is 40% to 60% carbohydrate.‘, 2 It is widely used as a monitor for detection, staging, checking recurrence, and determining response of therapy in patients with carcinoma of the digestive tract.3-5 McDicken and Scott6 reported that more than 50% of salivary gland tumors, both benign and malignant, were positive for CEA. Sumitomo and colleagues7 found a high incidence of positive reactivity with antibodies to CEA in material secreted into the luminal cavities and in luminal borders of tumor cells of pleomorphic adenomas, cystadenolymphomas, adenoid cystic carcinomas, and acinic cell carcinomas. Saito and colleagues8 found that CEA was present in both malignant and benign tumors from human minor salivary glands; however, the distribution of CEA was relatively more common and with increased staining intensity in malignant tissues. The purpose of this study was to determine whether there was a difference in expression of CEA between benign and malignant salivary gland tumors of human beings.

sOral Pathology Section, Department of Pathology, School Medicine, Temple University. bDepartment of Pathology, Temple University Hospital. l/14/15277

of

MATERIAL

AND METHODS

Twenty-three salivary gland neoplasms accessioned from 1985 through 1988 were retrieved from the files of the Oral Pathology Laboratory, Temple University School of Medicine. Seven tumors were located in the upper lip, six in the buccal mucosa, five in the hard palate, two in the floor of the mouth, two in the lower lip, and one in the parotid gland. All the tumors, except one, were described as raised lesions and three of them as painful. Microscopic slides were reviewed by three pathologists to confirm the diagnoses. In addition, three normal accessory mucous glands and two normal parotid glands were used in the study. All specimens had been fixed in phosphate-buffered neutral formalin and processed for routine histologic sectioning. Sections were cut at 4krn from each block and mounted on glass slides by means of a commercial white glue diluted to 1% as an adhesive. The avidin-biotin technique was used for staining CEA.” Sections were deparaffinized and quenched in 0.3% peroxide/methanol to eliminate endogenous peroxidase activity. The sections were incubated with goat serum for 20 minutes (as a blocking serum) and washed in phosphate-buffered saline (PBS), pH 7.4, for 10 minutes. A 1: 1000 dilution of the primary antibody (rabbit antihuman) was applied for 45 minutes and washed in PBS. Biotinylated secondary antibody (goat antirabbit) was then applied to each specimen and incubated with avidin-biotin complex for 90 minutes. Sections were washed in PBS and incubated in 0.1% diaminobenzidine in PBS for 5 minutes. The 479

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Fig. 1. Pleomorphic adenoma stained with avidin-biotin technique for CEA. Positive staining is seen in the secretory material within the duct lumen and in the luminal membrane of the ductal cells (arrowsj. (Original magnification, X 10.)

Fig. 2. Adenoid cystic carcinoma stained with avidin-biotin technique for CEA. Positive staining is seen in the secretory material within the duct lumen and in the luminal membrane of the ductal cells ~urrowsj. (Original magnification, X40.)

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CEA in salivary gland

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sections were then counterstained with Gill’s No. 2 hematoxylin for 30 seconds, cleared, and mounted. The antibody was applied to each specimen and to known controls simultaneously. A positive control consisted of colonic carcinoma and negative control runs employed normal rabbit serum in lieu of test antibody (Dako Corp., Santa Barbara, California) diluted to match primary antibody concentrations. RESULTS

Twenty-three salivary gland tumors were identified (Table I). The results of CEA reactivity were expressed in grades, namely negative (-) and positive (+,++,+++), according to the absence or presence of stain deposits in the cells and the intensity of the staining. CEA was present in six pleomorphic adenomas (Fig. I), one adenoid cystic carcinoma (Fig. 2), and the two major salivary glands (parotid). The stronger positive reactivity (+i- and +++) was present in two pleomorphic adenomas whereas the remaining cases presented the same reactivity (+). The brown reaction product was present in the luminal membrane of the ductal cells, and positive stained material was seen in the duct lumens. DISCUSSION

Monoclonal and palyclonal antibodies against CEA have been used in the differential diagnosis of various epithelial neoplasms. lo. ’ ’ They have also been used to evaluate normal and inflamed parotid glands and parotid obstructive adenitis.12 It has been established that CEA is also present in nonneoplastic salivary gland conditions and in normal saliva.13 It has been suggested that CEA can be used as a tool to distinguish between epithelial and mesenchyma1 tumors and to establish a prognosis.’ A study of CEA in mammary tumors disclosed that patients with CEA-negative tumors had significantly higher S- and IO-year survival rates,14 supporting the contention that CEA markers can be used as a prognostic indicator in breast cancer.” We were able to detect CEA immunohistochemically in only a few acini of the control series of normal major salivary glands, a finding that is consistent with that of McDicken and Scott,6 who reported CEA present in normal saliva and who suggested that CEA is not primarily related to the histologic type of tumor, but rather is dependent on the degree of differentiation of the neoplastic ceils and the development of well-formed ducts. We conclude from this study that CEA is not a reliable marker to differentiate benign from malignant salivary gland tumors. Dr.

We thank Dr. Arthur Pamela Edmonds

S. Miller, for their

Dr. Sow-Yeh Chen, advice and guidance

and and

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I. CEA reactivity in salivary gland tumors and normal salivary glands

Table

Case No. 1 2 3 4 5 6 7 8 9 10 11 I2 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28

Diagnosis

Age

Sex

Pleomorphic adenoma Pleamorphic adenoma Pleomorphic adenoma Pleomorphic adenoma Pleomorphic adenoma Pleomorphic adenoma Pleomorphic adenoma Pleomorphic adenoma Pleomorphic adenoma Pleomorphic adenoma Pleomorphic adenoma Pleomorphic adenoma Pleomorphic adenoma Adenoid cystic carcmoma Adenoid cystic carcinoma Ductal cystadenoma Ductal cystadenoma Mucoepidermoid carcinoma Mucoepidermoid carcinoma lntraductal papilloma Acinic cell adenocarcinoma Acinic cell adenocarcinoma Clear cell adenoma Normal minor salivary gland Normal minor salivary gland Normal minor salivary gland Normal major salivary gland Normal major salivary gland

36 35 31 25 21 31 40 23 35 28 26 28 37 34 39 37 31 3s IO 37 40 35 22 33 35 15 49 53

F M M M M F M F M F F F M F M F F F M M F M F F F M M M

Mrs. Grace ing technical

Green and assistance.

Mrs.

Eleanora

Satchel1

for

CEA

provid-

FIEFERENCES of CEA. Biochim Biophys Acta 1. Rogers CT. Heterogeneity 1976;458:355-73. PA, Coligen JE, Todd CW. CEA: char2 Terry WD, Henckart acterization and clinical applications. Transplant Rev I 974; IO: I M-29. SO. Demonstration of tumor-specific anti3 Gold P, Freedman gens in human colonic carcinoma by immunological tolerance and absorption technique. J Exp Med 1965;121:439-62. SO. Specific CEA of human digestive sys4 Gold P, Freedman tem 1 Fxp Med 1965:122~467-81. D, Voichl S Carcinoembryonic antigen 5 Tabuchi V. Hiroyuki and carbohydrate antigen IV-9 levels of peripheral and draining venous blood in colorectal cancer patients. Cancer 1988;62:1605-13. 6. McDicken IW, Scott J. The presence and distribution of CEA in tumors of human minor salivary glands. J Oral Path01 198 I; 10.296-303 7 Sumitoma S. Kumasa S, Mitam H, Mori M. Comparison of CEA distribution in lesions and tumors of salivary glands as determined with monoclonal and polyclonal antibodies. Virchows Arch B 1987;53: 133-9. 8 Saito I, Teratani K, Inove M. Saito A, Funatsu K, Moro I. Immunohislochemical characterization of functional markers m human minor salivary gland tumors J Oral Pathot 1984;13:525-34. 9 DeLellis RA. Basic technics of immunohistochemistry. Diag-

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nostic immunohistochemistry. Vol. 1. New York: Masson Publishing USA Inc. 198 1;7- 16 Caselitz J, Jaup T, Seifert G. Immunohistochemical detection of CEA in parotid gland carcinomas. Virchows Arch A 198 1; 394:49-60. Caselitz J, Seifert G, Jaup T. Tumor antigen in neoplasm of the human parottd gland. J Oral Path01 1982;l 1:374-86. Tsukitani K, Nakai M, Tatemoto Y, Hikosaka N, Mori M. Histochemical studies of obstructive adenitis in human submandibular salivary glands. I. Immunohistochemical demonstration of lactoferrin, lysozyme, and carcinoembryonic antigen. J Oral Path01 1985;14:631-8. Martin F, Devan J. CEA in normal human saliva. J Nat1 Cancer Inst 1973;50:1375-9. Theriault RL, Hortobayi GN, Fritsch HA, Frye D, Martinez R, Buzdar AV. The role of serum CEA as a prognostic indi-

cator in adjuvant 15. Shousha bryonic dicator.

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stage II and III breast cancer patients treated with chemotherapy. Cancer 1989;63:828-35. S, Lyssiotis T, Godfrey VM, Schever PJ. Carcinoemantigen in breast-cancer tissue; a useful prognostic inBr Med J 1979;1:777-9.

Reprint requests to: Miriam Alfaro, CD Oral Pathology Section Department of Pathology School of Medicine Temple University 3223 N. Broad St. Philadelphia, PA 19 140