**12. State-of-the-art of immunology in canine melanoma**

Canine melanoma is an immunogenic tumor, and investigations of different aspects associated with immune cells in tumor development and progression have been reported in the literature [39–41]. Moreover, some studies have evaluated different cancer immunology aspects of canine melanoma, podoplanin [42], and chondroitin sulfate proteoglycan-4 (CSPG4) [40]. Podoplanin is a type I transmembrane protein that is expressed in different cells of the immune system, including lymphatic endothelial cells. Podoplanin overexpression has been investigated in several cancers, including canine melanomas. Since the development and application of a monoclonal antibody against podoplanin, these markers have been recognized as important for canine melanoma immunotherapy [43]. CSPG4 has become very important for canine melanoma owing to the number of vaccines produced against this protein [39, 44, 45].

Its expression has been reported in canine melanoma, and different clinical trials based on vaccines or electrogene therapy have been conducted [39, 44, 45]. A review of the PubMed database for the past ten years is provided in **Table 2** summarizing clinical trials involving dogs with melanoma treated with immunotherapies.

In addition to evaluating immunotargets for canine melanoma, several studies have investigated the association between immune markers and melanoma prognosis,



#### **Table 2.**

*Summary of the articles published in the past ten years with clinical studies evaluating different immunotherapies for canine melanoma.*

including intratumoral infiltration of immune cells. The prognostic significance of CD3+ and CD20+ cell infiltration of canine oral melanoma has been investigated; and high infiltration of CD20+ cells is associated with metastasis, frequency of recurrence, shorter survival time, and high rate of tumor-related deaths, and disease-free interval [51]. Therefore, a high infiltration of CD20+ cells is associated with several negative prognostic factors. Yasumaru et al. [52, 53] evaluated the presence of CD8+ and CD4+ infiltrating T cells in oral melanoma using flow cytometry and concluded that tumor-infiltrating lymphocytes predict the aggressiveness and prognosis of patients with oral melanoma.
