Canine Cutaneous Squamous Cell Carcinomas Research Articles

Altered E-cadherin/β-catenin expression in feline cutaneous squamous cell carcinomas

Cutaneous squamous cell carcinoma (CSCC) is the most common malignant skin tumour in cats and the nature of the molecular mechanisms involved is poorly defined. Included among the molecular mechanisms in human and canine CSCCs is altered expression of E-cadherin/β-catenin. This study aimed to explore the immunohistochemical expression pattern of E-cadherin and β-catenin in 43 samples of feline CSCC by using a tissue microarray to elucidate whether expression of these molecules is dysregulated. Membrane expression of E-cadherin and membrane and cytoplasmic expression of β-catenin were significantly reduced in the CSCCs. Cytoplasmic expression of E-cadherin and nuclear expression of β-catenin were also found in some CSCCs. These findings indicate that altered expression of E-cadherin and β-catenin is a frequent event in feline CSCCs, suggesting that these molecules play an important role in acquisition of the malignant phenotype in feline patients with CSCC. The results also suggest the existence of a subpopulation of feline patients with CSCC in which the Wnt pathway may contribute to epidermal carcinogenesis.

Mutations in exon 5 of the p53 gene in canine cutaneous squamous cell carcinomas

Mutations in exon 5 of the p53 gene in canine cutaneous squamous cell carcinomas

VEGF Expression, Cellular Infiltration, and Intratumoral Collagen Levels after Electroporation-Based Treatment of Dogs with Cutaneous Squamous Cell Carcinoma.

Canine cutaneous squamous cell carcinoma (SCC) is the most common type of skin cancer in tropical countries and is generally associated with exposure to solar ultraviolet light. It has a low metastatic rate, and local treatments, such as electrochemotherapy (ECT), promote long-term control or even complete remission. This study aimed to evaluate pre- and post-ECT treatment expression levels of vascular endothelial growth factor (VEGF) and CD31, cellular infiltration, and intratumoral collagen levels in dogs with cutaneous SCC. A prospective nonrandomized clinical study was performed using dogs with spontaneous SCC treated with ECT. Eighteen lesions from 11 dogs were included in the study. The expression levels of VEGF and CD31; cellular infiltration; and intratumoral collagen levels, as determined by Masson’s trichrome staining, were not significantly different from pre-treatment measurements on day 21 (p > 0.05). However, among cellular infiltration, the mixed subtype was correlated with better overall survival time when compared to lymphoplasmacytic and neutrophilic infiltration (p < 0.05). In conclusion, ECT had no effect on VEGF expression, cellular infiltration, or intratumoral collagen levels in dogs with cutaneous SCC at the time of evaluation, suggesting that early and late post-ECT-treatment phases should be considered.

Persistent activation of the mammalian target of rapamycin signalling pathway in cutaneous squamous cell carcinomas in cats.

Cutaneous squamous cell carcinoma (CSCC) represents the most common malignant tumour of the feline skin. Emerging evidence suggests that the phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin (PI3K/Akt/mTOR) signalling pathway may represent a potential target for pharmacological intervention in human and canine CSCC. The present study aimed to explore the expression pattern and status of activation of relevant signalling proteins of the PI3K/Akt/mTOR signalling pathway in feline CSCC. The expression of pEGFRTyr1068 , pAktSer473 , pS6Ser235/236 combined with Ki-67, and the tumour suppressor protein PTEN was evaluated by immunohistochemical analysis in 45 samples of feline CSCC, using a tissue microarray. The immunodetection using phosphospecific antibodies to detect the activated forms of signalling proteins showed that the PI3K/Akt/mTOR signalling pathway is frequently activated in feline CSCCs, and may be independent of the activation of EGFR. The results also showed that PTEN expression is not significantly altered in feline CSCCs. Our study shows that the persistent activation of the PI3K/Akt/mTOR signalling pathway represents a key event in feline CSCC, pointing to this signalling pathway being a potential therapeutic target in feline patients with CSCC.

Relationship of Galectin-3 Expression in Canine Cutaneous Squamous Cell Carcinomas with Histopathological Grading and Proliferation Indices

Squamous cell carcinoma (SCC) is a frequent malignant neoplasm of the skin that usually arises from areas of solar dermatosis. It is characterized by local invasiveness and regional lymph node metastasis, mainly in poorly differentiated tumours. Galectin-3 (Gal-3) is a lectin that is expressed in the nucleus or cytoplasm and has been identified as a prognostic tool for human neoplasms. The purpose of this study was to characterize Gal-3 expression in canine cutaneous SCCs and to investigate its relationship with tumour differentiation and cell proliferation indices. Immunohistochemical analysis of 50 SCCs for Gal-3 revealed no correlation between the localization or intensity of immunolabelling, or number of immunopositive cells, with histological grade of tumour or proliferative activity. The results suggest that Gal-3 expression is not a reliable prognostic marker for cutaneous SCC in dogs.

Gene expression of prostaglandin EP4 receptor in three canine carcinomas

BackgroundChronic inflammation mediated by the cyclooxygenase enzymes, specifically their product prostaglandin E2 (PGE2), can result in the development of cancer. PGE2 promotes cell proliferation, apoptosis, and angiogenesis through interaction with its specific receptors (EP1 receptor - EP4 receptor [EP1R-EP4R]). In multiple human cancers, the expression of EP4R is associated with the development of malignancy and a poor prognosis. The expression of EP4R has not yet been evaluated in canine tumors. The aim of this study was to characterize the mRNA gene expression of EP4R (ptger4) in canine squamous cell carcinoma (SCC), apocrine gland anal sac adenocarcinoma (AGASACA), and transitional cell carcinoma (TCC). Archived tumor samples of canine cutaneous SCC (n = 9), AGASACA (n = 9), and TCC (n = 9), and matched archived normal tissue controls were evaluated for mRNA expression of canine EP4R using RNA in situ hybridization (RNAscope®). Quantification of RNAscope® signals in tissue sections was completed with an advanced digital pathology image analysis system (HALO). Data was expressed as copy number, H-index, and percent tumor cell expression of EP4R.ResultsIn all canine SCC, AGASACA, and TCC samples evaluated, strong universal positive expression of EP4R was identified. For SCC and AGASACA, mRNA EP4R expression was statistically higher than that of their respective normal tissues. The TCC tissues displayed significantly less mRNA EP4R expression when compared to normal bladder mucosa.ConclusionsThese results confirm the mRNA expression of canine EP4R in all tumor types evaluated, with SCC and AGASACA displaying the highest expression, and TCC displaying the lowest expression. This study also represents the first reported veterinary evaluation of EP4R expression using the novel in situ hybridization technique, RNAscope®.

Kallikrein-related Peptidase 5 (KLK5) Expression and Distribution in Canine Cutaneous Squamous Cell Carcinoma

Cutaneous squamous cell carcinoma (cSCC) is one of the most common types of malignant skin cancer in dogs, representing 3.9-10.4% of all canine skin tumours. Although the metastatic potential of cSCC is debated, it appears to mimic that observed in man. In man, predictive histopathological features for metastasis include tumour depth, lesions >5-6mm in depth, and invasion of muscle, cartilage or bone. In dogs, some reports have focused on the clinical features and long-term progression of cSCC, but a gold standard treatment has not yet been developed. We explored the protein expression of kallikrein-related peptidase 5 (KLK5), an important modulator of skin homeostasis, in normal canine skin and in examples of cSCC. KLK5 was highly expressed in the upper stratum granulosum, stratum corneum, hair follicles and sweat glands, skin sites where human KLK5 has been shown to be involved in physiological processes including keratinocyte desquamation, antimicrobial defence, lipid permeability and pigmentation. In cSCC, tumour cells at the deep margin, as well as those in the centre of keratin pearls, displayed cytoplasmic expression of KLK5. Some of the KLK5 immunoreactive cells also expressed vimentin, suggesting that they may be undergoing epithelial-mesenchymal transition and therefore have a more invasive behaviour than those expressing only KLK5. KLK5 may be a novel molecular biomarker useful for predicting prognosis of cSSC in dogs.

Electrochemotherapy induces tumor regression and decreases the proliferative index in canine cutaneous squamous cell carcinoma

Canine cutaneous squamous cell carcinoma (cSCC) is the most common skin cancer in dogs, and, due to its low metastatic rate, local treatments, such as electrochemotherapy (ECT), promote disease control or even complete remission (CR). This study aimed to evaluate the gene and protein expression of Bcl-2 and Bcl-2 associated X protein (BAX), the proliferative index and clinical parameters in dogs with cSCC subjected to ECT. A prospective nonrandomized clinical study was performed using dogs with naturally occurring cSCC that was treated with ECT. Eighteen lesions from 11 dogs were selected. The tumor size at day 0 (D0) had no impact on survival or prognosis (P > 0.05). Tumor samples had a lower proliferative index after ECT (D21) than before ECT (P = 0.031). The survival of subjects with Ki67 values lower and higher than the Ki67 median value were not significantly different (P > 0.05). Regarding apoptotic markers, there were no significant differences in the gene and protein expression levels of BAX or Bcl-2 at D0 and D21 (P > 0.05) or in the overall survival of subjects with different levels of apoptotic markers. In conclusion, there was no change in BAX or Bcl-2 gene and protein expression in response to ECT at the time points evaluated, but ECT was able to reduce tumor volume and cellular proliferation in cSCC.

Immunohistochemical expression of selected phosphoproteins of the mTOR signalling pathway in canine cutaneous squamous cell carcinoma

Cutaneous squamous cell carcinoma (cSCC) represents one of the most common malignant skin tumors in dogs. Research aimed at clarifying how the deregulated activity of signalling pathways contributes to cSCC progression can help to identify molecular suitable targets for the development of novel therapies. The present study describes the immunohistochemical expression pattern of two proteins (pAktSer473 and pS6Ser235/236, the latter combined with Ki-67) involved in the phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin (PI3K/Akt/mTOR) signalling pathway in canine specimens of normal epidermis, pre-neoplastic epidermis, and cSCC using tissue microarrays.The results suggest that the PI3K/Akt/mTOR signalling pathway has a low expression in the normal canine epidermis, and that selected molecules involved in this signalling pathway are dysregulated during the canine epidermal carcinogenesis process. These findings provide important evidence that the persistent activation of the PI3K/Akt/mTOR signalling pathway represents one of the key events during cSCC progression in canine patients, pointing to the PI3K/Akt/mTOR pathway as a potential therapeutic target.

Relationship between the inflammatory infiltrate and the degree of differentiation of the canine cutaneous squamous cell carcinoma

The inflammatory response may have pro or anti-neoplastic effects in tumors, depending on the histological type and malignancy level of the tumor. This study aimed to evaluate the profile of predominant inflammatory cells by immunohistochemistry in cutaneus squamous cell carcinoma (SCC) of dogs, comparing it with the degree of differentiation of the tumor. Twenty samples of SCC were analyzed. The tumors were histologically classified into two groups, differentiated SCC (SCCd=12) and undifferentiated SCC (SCCu=08). The tumor inflammatory infiltrate was determined by immunohistochemistry, in order to identify macrophages, lymphocytes and plasma cells. The comparison between groups, SCCd and SCCu, was not significant concerning the density of macrophages (P=1.0), T lymphocytes (P=0.335) and plasma cells (P=0.075). However, when comparing the inflammatory infiltrate in each group, the macrophages were the predominant cell type in both groups, a significant difference was found in the SCCd with plasma cells (P<0.0001). In the SCCu the difference occurred with lymphocytes (P<0.05) and plasma cells (P=0.0006). It could be concluded that the presence of inflammation in cases of SCC does not play a role in the differentiation of the neoplasm, since the inflammatory infiltrate was similar in both groups of SCC.

Infrequent detection of papillomaviral DNA within canine cutaneous squamous cell carcinomas, haemangiosarcomas and healthy skin on the ventrum of dogs

Canine squamous cell carcinomas (SCCs) most frequently develop on the ventral abdomen and are thought to be caused by ultraviolet (UV) light. Papillomaviruses (PVs) have been associated with cutaneous SCCs in multiple species, including dogs. That PVs act as cofactors in canine UV-induced SCCs. The study was performed on skin from the ventrum of 60 dogs. These samples included 20 SCCs, 20 haemangiosarcomas and 20 samples of clinically normal skin. Two canine viral plaques were included as positive controls for PV. PCR was used to amplify PV DNA from all samples. Primers used included two sets of consensus primers and two sets of primers that were designed specifically to amplify PV DNA sequences detected in the viral plaques. The MY09/11 consensus primers amplified PV DNA from both viral plaques. One plaque contained a DNA sequence (CfPV-JM) that had been previously reported from a dog with multiple cutaneous SCCs. The other plaque contained a previously unreported PV DNA sequence. No PV DNA was amplified by either consensus primer from any of the ventrum skin samples. Primers designed specifically to amplify the CfPV-JM sequence amplified DNA from one SCC, but no other sample. No PV DNA was amplified using the other specific PCR primer set. These results do not support a significant role for PVs in SCC development from the ventrum of dogs. However, they contribute another PV sequence to the list of PVs that have been associated with viral plaque development in dogs.

Tumour Epithelial Vimentin Expression in Canine Cutaneous Squamous Cell Carcinoma

Tumour Epithelial Vimentin Expression in Canine Cutaneous Squamous Cell Carcinoma

Immunohistochemical Detection of COX-2 in Feline and Canine Actinic Keratoses and Cutaneous Squamous Cell Carcinoma

Cyclooxygenase-2 (COX-2) overexpression and its causal role in epidermal carcinogenesis have been demonstrated in human actinic keratoses (AK) and cutaneous squamous cell carcinoma (SCC). The aim of this study was to determine immunohistochemically the level of expression of COX-2 in feline and canine AK (n=18), SCC (n=36) and inflammatory dermatoses (n=24). COX-2 immunoreactivity was detected in all feline and canine SCC. In all specimens, labelled basal and suprabasal neoplastic keratinocytes were localized within and below areas of superficial erosion or ulceration and only scattered deeper tumour cells were positively labelled. In most cases, positive immunoreactivity of keratinocytes was associated with the presence of granulocytes. COX-2 expression was detected in 3/9 canine and 4/9 feline cases of AK and in only one case was associated with inflammation. Inflammatory dermatoses were characterized by positively labelled epidermal and follicular basal and suprabasal keratinocytes that were always associated with granulocyte exocytosis. These results indicate that further study of the effect of using COX-2 inhibitors in the management and prevention of feline and canine cutaneous SCC is warranted. The association between inflammatory cells and COX-2 expressing epidermal cells opens a new line of research regarding the role of COX-2 in SCC oncogenesis. Moreover, further studies should investigate the role of COX-2 in the pathogenesis and management of AK in animals.

Papillomavirus-Associated Cutaneous Neoplasia in Mammals

For humans, strong evidence indicates that some mucosal papillomavirus (PV) types cause genital and oral neoplasia, and weaker evidence suggests that some cutaneous PVs may cause cutaneous squamous cell carcinomas (SCC). For nonhuman species, strong evidence supports a causal role of PVs in the development of feline and equine sarcoids. Likewise, PVs are believed to cause cutaneous SCCs in rabbits, western barred bandicoots, and some rodents. Furthermore, some evidence suggests that PVs may influence the development of both feline and canine cutaneous SCCs. This review discusses the evidence that PVs cause human cutaneous SCCs and the proposed mechanisms for this action. It then reviews preneoplastic and neoplastic skin diseases that are associated with PV infection in nonhuman mammals.

Survivin expression in canine epidermis and in canine and human cutaneous squamous cell carcinomas

Survivin, a member of the inhibitor of apoptosis protein (IAP) family, is ubiquitously expressed during tissue development, undetectable in most normal tissues, but re-expressed in most cancers, including skin malignancies. Expression of survivin was evaluated retrospectively in 19 canine cutaneous squamous cell carcinomas (SCCs; one in situ; 16 well differentiated; one invasive, one lymph node metastasis) and 19 well differentiated SCCs from human beings. Seven specimens of normal canine skin were included. Immunohistochemical expression of full-length survivin was determined using a commercially available antibody. In addition, apoptotic rate [Terminal deoxynucleotidyl Transferase Biotin-dUTP Nick End Labelling index (TUNEL) index] and mitotic index (MI), counting mitoses in 10 high power fields (HPF), were determined. Scattered survivin positive nuclei were identified in the epidermal basal cell layer of normal canine skin. Nuclear survivin expression was identified in 18 of 19 human and in all canine SCCs, mainly along the base of the tumour cell population. Cytoplasmic survivin expression was rarely observed in human SCCs and in 84.2% of canine SCCs. The TUNEL index ranged from 0.1 to 2.6 in human beings and from 7.5 to 69.4 in dogs, while MIs ranged from 0 to 4 in human beings and dogs. No correlation was found between survivin expression and apoptotic or mitotic rates. Canine and human tumours showed similar nuclear survivin expression, indicating similar functions of the molecule. We demonstrated survivin expression in normal adult canine epidermis. Increased nuclear survivin expression in pre-neoplastic and neoplastic lesions demonstrates a possible association of survivin with development of SCCs in human beings and dogs.

Nuclear morphometry in relation to tumor grade in canine spontaneous cutaneous squamous cell carcinomas

In a retrospective study on cytological specimens from 17 dogs with histologically confirmed cutaneous squamous cell carcinoma, the morphometric variables were studied and compared to tumors' nuclear grade. The morphometric parameters evaluated in this study were mean nuclear area (MNA, µm2), mean nuclear perimeter (MNP, µm), mean nuclear diameter (D mean, µm), minimum nuclear diameter (D min, µm), and maximum nuclear diameter (D max, µm). Associations between MNA, MNP, D mean, D min, and D max and tumors' grade were assessed using the ANOVA/LSD test (Statistica 6.0, StatSoft, USA) at a level of significance of P < 0.05. The result indicated an increase in the mean values of the nuclear parameters with increasing tumor grade. In the cases examined, significant differences in MNA, MNP, and D mean occurred between tumors in all grades. Besides, no significant differences in D max (between grade I-II and II-III) and D min (grade II-III) were found. In conclusion, nuclear morphometry could be used as an additional tool for differentiating nuclear grade in canine cutaneous squamous cell carcinomas.

Immunohistochemical Expression of Vascular Endothelial Growth Factor and Vascular Endothelial Growth Factor Receptor Associated with Tumor Cell Proliferation in Canine Cutaneous Squamous Cell Carcinomas and Trichoepitheliomas

The expression of 5 markers associated with angiogenesis was studied in canine squamous cell carcinomas (SCCs) (n = 19) and canine trichoepitheliomas (TCPs) (n = 24). SCCs were assigned histologic grades, and tissue sections from both tumor types were immunohistochemially stained for the expression of vascular endothelial growth factor (VEGF), vascular endothelial growth factor receptor-2 (VEGFR-2), as well as intratumoral microvessel density (iMVD), tumor proliferation index (PI), and tumor apoptotic index (AI), using antibodies against VEGF, VEGFR-2, von Willebrand's factor, Ki-67 antigen, and the terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate end-labeling method (TUNEL), respectively. VEGF and VEGFR-2 were detected in 17/19 (89.4%) and 19/19 (100%) SCCs and in 17/24 (70.8%) and 20/24 (83.3%) TCPs, respectively. In SCCs, there was substantial correlation between histologic grade and PI (r = 0.51); and moderate correlation between VEGF and histologic grade (r = 0.43), VEGFR-2 and histologic grade (r = 0.47), VEGF and PI (r = 0.47), and VEGFR-2 and PI (r = 0.47) (Spearman rank correlation coefficient). In TCPs, there was substantial correlation between VEGF and PI (r = 0.51) and a moderate correlation between VEGFR-2 and iMVD (r = 0.36). The median iMVD of SCCs (15.5) was significantly higher than the median iMVD of TCPs (9.05) (P value < .05). It was concluded that VEGF and VEGFR-2 may promote tumor cell proliferation in TCPs and SCCs. An autocrine pathway for VEGF probably operates in canine SCCs and TCPs, as VEGF and VEGFR-2 expression was found in most tumors and was associated with evidence for tumor cell proliferation.