**1. Introduction**

Pheochromocytomas (PCs) are catecholamine-secreting neuroendocrine tumors arising from the chroma ffin cells of the neural crest [1–3]. PCs are more often seen in dogs and cattle [4] and can be unilateral or bilateral and functional or nonfunctional. While canine pheochromocytomas are usually benign, they can invade adjacent tissues and may be malignant, with metastasis to distant tissues [4]. The behavior of pheochromocytomas is di fficult to predict based on histologic findings [5,6].

Immunohistochemistry (IHC) data has found that canine and human PCs are highly similar, as neoplastic cells in both share the expression of numerous antigens, including S100, synaptophysin (SYN), chromogranin A (CGA), and substance P (SP) [7]. Known genetic mutations are involved in the pathogenesis of approximately 60% of human PCs. Frequently, these are associated with mutations in the succinate dehydrogenase (SDH) family of genes, with mutations in succinate dehydrogenase subunit B (SDHB) associated with a high likelihood of metastasis/malignancy [5]. The sequencing of multiple SDH genes in every pheochromocytoma is economically infeasible in veterinary medicine, making the determination of the genetic basis of PC in dogs uncertain.

Studies in human PCs have found that immunohistochemistry (IHC) is highly correlated with the SDH mutation status. For instance, all samples with mutations in SDH family genes lacked SDHB immunoreactivity [8]. Loss of SDHB protein expression has therefore been used for prognostication in human medicine; in one study, the relationship between the SDH genetic background and SDHB immunohistochemistry sensitivity and specificity in human pheochromocytomas was 94.23% [9].

In veterinary medicine, while a link has been established between brachephalic dogs and pheochromocytoma [2], there has not been a definitive familial inheritance pattern like that described in people. Humans and dogs have a similar structure of SDH family genes [10]. Some mutations have been found in SDH family genes, which may indicate that mutations in these genes may initiate oncogenesis in a similar way to people [2,11]. While IHC has previously been used in canine pheochromocytomas to verify the neuroendocrine origin [4,7], only one study to date has examined the SDH family status in canine pheochromocytomas [11]. IHC would be significantly more cost-e ffective and practical than sequencing for mutation detection. In addition, if IHC is associated with patterns in clinical parameters, such as recurrence, invasion, or metastasis, this could be a valuable adjunct to histopathology.

In this study, we examined the expression of SDHA and SDHB in canine pheochromocytomas and compared these with data on patient age, tumor size, and invasion, in order to determine the utility of SDH IHC in canine pheochromocytoma diagnostics.

#### **2. Materials and Methods**
