CD15

Characteristics
CD15 (3-fucosyl-N-acetyl-lactosamine) is a cluster of differentiation antigen – an immunologically significant molecule. CD15 is a carbohydrate adhesion molecule (1) that can be expressed on glycoproteins, glycolipids and proteoglycans. It is a complex cluster of cell surface glycoproteins and glycolipids having a common terminal pentasaccaharide known as the Lewisx (Lex) antigen. It is probable that the core proteins and lipids differ on the different cell types since there is a marked variability in the binding of the antibodies within and among them. CD15 exists in a sialylated and an unsialylated forms. The function of CD15 is not well characterized and is most probably multifactorial.
CD15 is a haemopoietic differentiation antigen expressed on most terminally differentiated myeloid cells including granulocytes, eosinophils, mast cells, monocytes/macrophages, and Langerhans’ cells. It is not expressed in red cells, platelets and lymphocytes (2). There are three CD markers of human leukocytes comprising fucosylated carbohydrate epitopes. Fucosylation of distal polylactosamine chain can be achieved through ɑ-1,3 fucosyltransferase (FUT) to form Lewis antigen (3). It was proposed that expression of CD15 in mature granulocytes is determined by FUT9 while that in promyelocytes and monocytes by FUT4 (3).
CD15 is not substantially expressed on haemopoietic progenitor cells. Up to 1% of bone marrow CD34+ cells, and up to 3.8% of CD34+ cells in peripheral blood progenitor cells express CD15. The vast majority of lymphocytes are CD15 negative but activated lymphocytes (particularly T4) may be positive.
CD15 is considered to be involved in neutrophil functions, that is, cell-cell interactions, adhesion to endothelial cells (4) (5), phagocytosis (6), stimulation of degranulation, and respiratory burst, although the function of CD15 is not clear (7) (8) (9) (2). Its role in activation of monocytes has also been proposed (10).
Apart from leukocytes CD15 is also found in various epithelia such as breast (secretory epithelium), kidney (proximal tubules) (11), lung, pancreas (12), and intestinal tract (including Paneth cells) (13) (14).
In the brain CD15 is constantly present in astrocytes and variably in oligodendrocytes and neurons (15) (16) (17) (18) (11) (19) (20).

Neoplasms
Fig1 CD15-neoplasms

The positivity for CD15 is characteristic of Hodgkin’s cells in classical Hodgkin lymphoma (HL) (Fig.1C). It is expressed in great majority of nodular sclerosis (NS), mixed cellularity (MC), lymphocyte depletion (LD) and lymphocyte rich-classical HD cases, but not in malignant cells of lymphocyte predominance (LP) HD (L&H cells, “popcorn cells”). It has been suggested that if classical HD expresses sialylated form of CD15 (sialyl-CD15), it has worse prognosis. (21) (22) (23) (24)
10-15% of peripheral T-cell lymphomas express CD15 (including occasional cases of mycosis fungoides) (25) (26), while B-cell lymphomas stain in an even lower proportion (usually diffuse large B-cell lymphoma). Rare cases of acute lymphoblastic leukaemia, in which myeloid antigens are often CD15 positive.
Myeloid leukaemia cells express CD15 in a heterogeneous manner. CMLs are regularly CD15 positive.
CD15 is expressed in a varying proportion of epithelial tumours such as adenocarcinomas (particularly from breast (27), lung (28)(Fig.1A & B) and colon (14)), renal cell carcinoma (29), apocrine carcinoma of the skin, papillary and follicular carcinoma of the thyroid, and serous carcinoma of the ovary. It is possible that sialyl-CD15 confer on the tumour cells the capacity to metastasize. Malignant mesothelioma is practically always CD15 negative (positivity has been reported in up to 6%, particularly the desmoplastic variant) (Fig.1D).
In gliomas, CD15 positivity inversely correlates with the grade of malignancy (19) (20). Reifenberger G et al also found CD15-positive cases among metastatic carcinomas, craniopharyngeomas, meningiomas, germinomas and malignant melanomas (19). Among germ cell tumours, CD15 is detected only in mature teratoma.

Application
In haematopathology CD15 is important for the diagnosis of classical HD and characterization of acute leukaemia.
In the differentiation of mesothelioma vs. adenocarcinoma, CD15 can be used in the primary panel.
Hepatocellular, gastric and colonic carcinoma and thyroid medullary carcinoma appear to have worse prognosis if positive for CD15. CD15 may be used for histopathological grading of gliomas and differentiating between malignant gliomas and non-neoplastic glial cells (the latter usually strongly stained).
Expression of CD15 in renal cell carcinoma is closely related to better prognosis. Localization of CD15 remains cytosolic in renal cell carcinoma while its expression in normal kidney is membranous. (29)
Detection of CD15 is frequently performed in diagnostic flow cytometry. In MDS, CD15 as well as CD10 (markers for myeloid cell maturation) are more prevalent on myeloid blasts from low-risk MDS (refractory anaemia [RA] and RA with ringed sideroblasts). A shift to a more immature phenotype with loss of CD10 and/or CD15 expression on myeloid blasts was also documented to accompany MDS progression. In acute myeloid leukaemia, the CD117 (+) CD15 (+) phenotype correlates significantly with complete remission achievement and with the lack of unfavourable chromosome associations (30).

Visualization
There are about 90 mAb clones assigned to CD15 and eight alternate names for CD15. Most of anti-CD15 mAbs, such as MMA, LeuM1 and 80H5, are IgM Abs and do not react with the sialylated form (CD15s or sialyl-CD15). The mAbs FH6 and CSLEX1 are directed against sialylated CD15. The binding of each mAb can be markedly increased by exposure of the cell surface to neuraminidase (muramidase), presumably by desialylating CD15, particularly in acute myeloid leukaemia.
The staining product is either membranous, Golgi-type, or cytoplasmic. Membranous and Golgi-type are far more frequent than cytoplasmic pattern. The pattern is commonly combined. Golgi-type is very frequent in Hodgkin’s cells and it can be seen with or without membranous staining. Occasionally, Hodgkin’s cells express only membranous CD15. Cytoplasmic staining without Golgi-type or membranous staining does not appear in HD. However, adenocarcinomas are seen to express predominately cytoplasmic CD15.
Optimal stains can be obtained with the clones C3D1, Carb3 and MMA (as concentrate), while BY87 has performed less satisfactory in the NordiQC lab (unpublished data), and has now been discontinued.

Datasheets for some of the clones: MMA conc MMA RTU Carb3 conc Carb3 RTU BY87 conc Heat-induced antigen retrieval (HIER) in an alkaline buffer is mandatory.
Fixation is important. It is easier to detect CD15 in B5-fixed tissues than in formalin-fixed tissues. Since this is just the opposite for CD30, it is important to try to sample lymph node tissue for fixation in both fixatives for optimal work up of HD.

Control Tissue
Normal kidney is useful as control: The proximal tubules must show a strong membranous and cytoplasmic reaction (photo).
An additional possibility is a case of HL with known weak expression of CD15. This is traditionally used as the only control in many laboratories. However, in principle normal tissues should be preferred to tumour tissue because of more constant and recognizable reaction patterns, as well as easier access to relevant tissues.
The correct titration should not be based on neutrophils, as these cells are high expressors of CD15 and therefore may react even in insufficient protocols.
Controls for B5-fixed tissue should also be fixed in B5.

Assessment
Run G1 2014

Bibliography
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