Specialty Spotlight – The Importance of Immunohistochemistry in Oncology
The Importance of Immunohistochemistry in Oncology
Ariana Verrilli, DVM, (practice limited to Oncology)
It is not uncommon for a cytology or biopsy report to land on our desk where the interpretation reads “malignant neoplasia” or “round cell neoplasia” or some equally broad interpretation or diagnosis. What does this mean? What do we tell our clients about their pet’s diagnosis, treatment plan, and prognosis? When a biopsy says “malignant neoplasia” we can tell the client the tumor is not benign, but the definitive diagnosis could be anything from a carcinoma to a lymphoma. If a biopsy diagnoses “round cell neoplasia” the scope of tumors has been narrowed down somewhat, however, you could be dealing with anything from a histiocytic sarcoma to a benign plasmacytoma, or possibly even a poorly differentiated sarcoma or carcinoma! Which still leaves us with a huge variation in recommended treatments and prognoses. When faced with such little information how do we go about telling our clients what the next steps for treating their pet should be?
The truth is, with such limited information, we cannot possibly prognosticate or treat the neoplasm appropriately. Therefore, steps need to be taken to gain more information about the type of tumor we are dealing with. If the diagnosis was made on cytology the next step would be to collect a biopsy sample, as tissue architecture can often provide additional diagnostic clues. If the diagnosis of neoplasia was made on a biopsy sample, it is likely that the pathologist will recommend additional tests, most often immunohistochemistry, to help obtain a more definitive diagnosis.
Immunohistochemistry (IHC) has become a routine supplement to traditional histopathology and is most often used to help diagnose and characterize neoplastic diseases. IHC allows specific tissue antigens to be recognized in a biopsy sample, and when used in conjunction with tumor morphology, can help lead us to a definitive diagnosis. For this test to be performed no additional samples need to be collected, as it is run on the original tissue sample. Depending on what the pathologist sees morphologically, they will likely recommend a panel of IHC markers, to help rule in or out a possible differential diagnosis. The basic premise of IHC is to identify a specific tissue protein or antigen via the use of an antibody that is specific for that particular substance; this antibody is termed a primary antibody. The formation of an antigen-antibody complex (i.e. the presence of the specific antigen in a given tissue) is detected either through direct or indirect methods. In the direct method, the primary antibody itself is coupled to a label which can be visually appreciated (most often an enzyme or fluorophore). In the indirect method, the primary antibody is bound by a second antibody that is labeled. This method is used most frequently as it can generate a stronger signal and does not require the primary antibody to be modified.
Once the IHC staining is complete the results must be interpreted, which is the responsibility of the pathologist. Interpretation is based not only on positive staining, but also on the cellular localization, tissue distribution, labeling intensity, and percentage of positive cells. For accurate interpretation, the pathologist must have a strong understanding of tumor morphology and the expected antigen staining for a given cell or tissue type. The hope is that with appropriate IHC selection and interpretation we will be able to obtain a definitive diagnosis on the submitted biopsy sample. Unfortunately, this is not always possible with IHC alone, however, tumor types can often be ruled out and sometimes a second panel of more focused IHC markers can be performed to further characterize the neoplasm.
IHC in veterinary medicine does have its limitations and cannot replace traditional morphologic evaluation. Although the number of available antibodies in on the rise, we do not yet have antibodies that will definitively identify every cell and tumor type. Additionally, in veterinary medicine we are dealing with a variety of species and many antibodies are not effective across species. For example, separate canine and feline antibodies have been developed for the leukocyte marker CD18, as the canine antibody will not bind to the feline antigen, and vice versa. We hope that our ability to definitively diagnose difficult tumors will continue to improve as new antibodies are developed and validated. Additionally, these markers will likely have a growing role in determining prognosis and developing a therapeutic plan.
To demonstrate the value of immunohistochemistry, let’s consider a case example. A 10-year old, mixed-breed dog presented for surgical excision of a large liver mass. The histopathology report diagnosed a malignant neoplasia, with priority given to a round cell tumor or a neuroendocrine carcinoma. The pathologist recommended IHC to further characterize the tumor, including leukocyte (CD18), lymphocyte (CD3), epithelial (cytokeratin) and neuroendocrine cell (synaptophysin, chromogranin A) markers. The results were negative for cytokeratin, synaptophysin, and chromogranin A, and positive for the CD18 and CD3, leading to a diagnosis of T-cell lymphoma. Based on this result, the dog was treated with a CHOP-based chemotherapy protocol and obtained a prolonged clinical remission. If IHC had not been submitted in this case, a diagnosis of lymphoma would not have been made, appropriate treatment would never have been initiated, and the dog would have experienced rapid progression of his disease. Thus, highlighting the value and importance of IHC in these tricky cases.
-Ariana Verrilli, DVM (practice limited to Oncology)