Use of a murine xenograft model for canine transmissible venereal tumor

OBJECTIVE: To develop a murine model for canine transmissible venereal tumor (CTVT). ANIMALS: Thirty-three 6-week-old NOD/LtSz-scid (NOD/SCID) mice and seven 6-week-old C57BL/6J mice. PROCEDURE: Samples of CTVT were excised from a 3-year-old dog and inoculated SC into ten 6-week-old NOD/SCID mice to induce growth of xenograft transmissible venereal tumor (XTVT). To establish mouse-to-mouse transmission, samples of XTVT were removed and inoculated SC into 4 groups of 6-week-old NOD/SCID mice and into a control group. Samples of CTVT were also inoculated into immunocompetent C57BL/6J mice for a mouse antibody production (MAP) test. The canine and xenografted tumors were evaluated cytologically and histologically, and polymerase chain reaction was performed for detection of the rearranged LINE/c-MYC junction. RESULTS: 8 of 10 NOD/SCID mice that were inoculated with CTVT developed tumors 3 to 10 weeks after inoculation. In the second-generation xenograft, all mice developed tumors by postinoculation day 47; 1 X 10(6) of XTVT cells were enough to create a xenograft. Metastases developed in 4 of 20 mice. Xenografted and metastatic tumors retained cytologic, histologic, and molecular characteristics of CTVT. Results of the MAP test were negative for all pathogens. CONCLUSION: We established an NOD/SCID murine model for XTVT and metastasis of CTVT. This model should facilitate study of tumor transplantation, progression, and metastasis and should decrease or eliminate the need for maintaining allogenic transfer in dogs.     

Heat shock proteins in canine transmissible venereal tumor

SDS-PAGE, Western blot analysis and immunohistochemical staining were used to detect heat shock proteins (HSPs) 60, 70 and 90 in canine transmissible venereal tumor (CTVT). Tissues tested for HSPs included: (1) tissues from different growth phases of CTVT tumors artificially induced in dogs; (2) tissues from other canine tumors; (3) normal dog tissues. Our results indicate that HSP 60 was consistently higher in CTVT cells in regressing phase than those in progressing phase. However, no detectable antibody response specific to the tested HSPs was found in the sera from CTVT-laden dogs in different growth phases. Although levels of the HSPs were all detectable in CTVT cells, only 60 and 70 were higher in CTVT cells than in normal tissues. In addition, none of the HSPs were detected in cells from five other canine tumors. These data suggest that canine HSP 60 and 70 are potential markers for CTVT and HSP 60 is appear to be involved in CTVT regression.PCR was used to confirm the existence of CTVT cells using primers designed to cover the sequence between the 5' end of c-myc near the first exon and the 3' end outside the LINE gene. Only CTVT samples were positive for this sequence; samples from other tumors and normal tissues were negative. The sequenced PCR products indicated that CTVT from Taiwan and other countries exhibited over 98% sequence homology. This reconfirms that, worldwide, all CTVT cells are very similar.     

A case of ocular canine transmissible venereal tumor

A 1-year-old, intact female mixed-breed dog was presented to St. George’s University Small Animal Clinic in Grenada for a third eyelid mass. The dog was diagnosed with a rare ocular transmissible venereal tumor (TVT) and concurrent anaplasmosis, ehrlichiosis and dirofilariasis. Treatment with vincristine sulfate resulted in complete resolution of the TVT.     

Immunohistochemical characterization of intraocular metastasis of a canine transmissible venereal tumor

Little has been published on intraocular metastasis of transmissible venereal tumors (TVT) in dogs. This report presents a 4-year-old male Labrador Retriever with a previous history of subcutaneous TVT which underwent total remission after treatment with vincristine. The dog presented with clinical signs of uveitis and increased intraocular pressure (IOP) in both eyes. After enucleation of the left eye, a diagnosis of TVT was made based on morphology, histology and immunohistochemistry (IHC). IHC staining for vimentin, S-100 protein, cytokeratin and HMB45 was performed to differentiate this lesion from TVT, lymphoma, melanoma, carcinomas, neurogenic tumors and fibrosarcoma. The IHC findings supported the diagnosis of TVT for this round cell tumor.     

The distribution of C-bands in canine transmissible venereal tumor cells

Colchicine-arrested metaphase preparations, derived from canine transmissible venereal tumor (TVT) cells grown in culture, were characterized based on the occurrence and distribution of constitutive heterochromatin. Analysis of the data with regard to the distribution of C-bands in the pericentric, interstitial and telomeric segments revealed a nonrandom distribution along the arms of many chromosomes with the bulk of the bands occurring in the centromeric region. The frequency of C-banded regions differed from those reported for normal dog cells and both primary and transplanted tumors. These results suggest that similar nondifferentially-stained TVT karyotypes do not necessarily exhibit identical distribution of constitutive heterochromatin.     

Predictive factors for the regression of canine transmissible venereal tumor during vincristine therapy

Canine transmissible venereal tumor (CTVT) is a neoplasm transmitted by transplantation. Monochemotherapy with vincristine is considered to be effective, but treatment time until complete clinical remission may vary. The aim of this study was to determine which clinical data at diagnosis could predict the responsiveness of CTVT to vincristine chemotherapy. One hundred dogs with CTVT entered this prospective study. The animals were treated with vincristine sulfate (0.025 mg/kg) at weekly intervals until the tumor had macroscopically disappeared. The time to complete remission was recorded. A multivariate Cox regression model indicated that larger tumor mass, increased age and therapy during hot and rainy months were independent significant unfavorable predictive factors retarding remission, whereas sex, weight, status as owned dog or breed were of no predictive relevance. Further studies are necessary to investigate whether these results are due to changes in immunological response mechanisms in animals with a diminished immune surveillance, resulting in delays in tumor regression.     

Disseminated transmissible venereal tumor in a dog.

Transmissible venereal tumor (TVT) is a well-documented transplantable tumor in dogs, with no breed or sex predilection and a low metastatic rate. In this report, a 2-year-old intact female Mastiff that had numerous, rapidly growing masses throughout the subcutis mainly at the dorsal body plane, the caudal half of the ventral abdomen, and around the vulva was euthanized due to poor prognosis. Neoplastic nodules similar to those seen in the subcutis were also noted in the lung, anterior mediastinum, liver, spleen, kidney, and superficial and deep lymph nodes in both abdominal and thoracic cavities. The neoplastic nodules from the subcutis as well as metastatic foci revealed similar cytologic and histologic features, which were consistent with canine TVT. By immunohistochemical staining, the neoplastic cells were positive for lysozyme and vimentin but were negative for cytokeratin, desmin, CD3, and CD79a. The diagnosis of the TVT was further supported by the identification and analysis of long interspersed nuclear elements (LINE) from paraffin-embedded tumor tissue. This case is a rare example of TVT with multiorgan metastasis. In this case, the polymerase chain reaction technique was useful in differential diagnosis of canine round cell tumors because this technique can be applied in retrospective as well as future study.     

Ocular involvement of transmissible venereal tumor in a dog

A case of metastatic transmissible venereal tumor (TVT), with lesions on the penis, conjunctiva, buccal mucosa and skin (lips and trunk), is presented in this case report. The clinical picture is described along with the cytological and histopathological features of the tumor leading to definitive diagnosis of TVT. Possible reasons for the unusual metastatic behavior of TVT and full recovery of the dog after chemotherapy are discussed.     

Evaluation of a genetic assay for canine transmissible venereal tumour diagnosis in Brazil

The canine transmissible venereal tumour (CTVT) is a transmissible cancer that is spread between dogs by the allogeneic transfer of living cancer cells. The infectious agents in CTVT are the living cancer cells themselves, which are transmitted between dogs during coitus. CTVT first arose several thousand years ago and the disease has a global distribution and is frequently observed in dogs from Brazil. We evaluated the utility of a LINE‐MYC quantitative polymerase chain reaction for diagnosis of CTVT cases in Brazil. Our analysis indicated that the LINE‐MYC rearrangement was detectable in all CTVT samples but not in their corresponding hosts. This genetic assay proves to be a useful tool for providing a definitive molecular diagnosis of CTVT, which presents with varying degrees of aggressiveness and invasiveness in different host dogs and can therefore be a diagnostic challenge in some specific cases.     

Effect of tumor infiltrating lymphocytes on the expression of MHC molecules in canine transmissible venereal tumor cells

Canine transmissible venereal tumor (CTVT) can be allo-transplanted across major histocompatibility complex barriers. The expression of MHC molecules is usually low in the progression (P) stage and then greatly increases during tumor regression (R). We investigated the effects of tumor infiltrating lymphocytes (TIL) on the expression of MHC molecules of CTVT cells. Isolated, viable CTVT cells were inoculated at each of 12 sites (1 x 10(8) CTVT cells per site) on the back of six, mixed-breed dogs. Tumor masses were collected every 2-3 weeks and prepared for histopathologic, immunocytochemistry, flow cytometry and immunoblotting studies. The level of MHC expression on tumor cells from different stages of growth was measured. Initially, expression of MHC I and II molecules in P phase CTVT was low. Twelve weeks post-inoculation (PI), expression increased dramatically and it continued to increase during R phase. Tumor growth slowed after 12 weeks PI and tumors entered R phase around 17 weeks PI. We hypothesize that CTVT evades host immunosurveillance and grows progressively for 12 weeks, when it becomes vulnerable and subject to the host's anti-tumor immune responses. We further demonstrated that R phase, but not P phase, TIL were closely associated with the over-expression of MHC I and II molecules by CTVT cells. The number and proportion of TIL were higher in R phase tumors. Supernatants, from R phase co-cultures (CTVT+TIL) and TIL only, promoted MHC I and II expression on P phase CTVT cells. After culturing alone for 1 month, expression of MHC classes I and II molecules in R phase CTVT cells decreased to the level of P phase CTVT cells. However, the above-mentioned supernatants restored their expression of MHC I and II molecules. In contrast, supernatants from P phase TIL or CTVT cells increased expression slightly or had no effect. Therefore, TIL, not CTVT cells, produce the effective substance (s) to promote the expression of MHC molecules by the tumor cells. Heat treated supernatant was unable to promote the expression of MHC I and II molecules by CTVT cells. In conclusion, TIL isolated from R phase CTVT secreted a heat-sensitive, soluble substance(s) that triggered over-expression of MHC I and II after 12 weeks PI. This caused the tumor to enter R phase and helped stop CTVT growth. Our findings will facilitate the understanding and further investigation of the mechanisms that initiate host immune surveillance against tumors.     

Clinicopathological study of canine transmissible venereal tumour in leishmaniotic dogs

Introduction : Canine transmissible venereal tumour is occasionally observed in leishmaniotic dogs, and Leishmania amastigotes can be harboured in canine transmissible venereal tumour cells. Objectives : The aim of this paper was to investigate the clinicopathological significance of the association of both diseases. Methods : Nineteen dogs affected by canine transmissible venereal tumour and canine leishmaniasis were studied retrospectively. Results : In these dogs, the tumour manifested a large size and often aggressive behaviour (42%) and no predictive sign of spontaneous regression was observed. Sporadic Leishmania amastigotes were found within the canine transmissible venereal tumour in three cases, probably transported by infected macrophages often infiltrating the tumour. A high Leishmania parasitisation of canine transmissible venereal tumour was observed in two other cases and verified by immunohistochemistry. Clinical Significance : Canine transmissible venereal tumour is a tumour of the dog able to harbour a large number of Leishmania parasites. Alternatively, the systemic disease (canine leishmaniasis) may lower the immune defence against malignancy (canine transmissible venereal tumour).     

Immunopathogenic behaviors of canine transmissible venereal tumor in dogs following an immunotherapy using dendritic/tumor cell hybrid

Canine transmissible venereal tumor (CTVT) is a naturally occurring tumor that can be transmitted between dogs via live tumor cell inoculation. It is also a spontaneous self-regression tumor and its behavior is closely related to host immune responses. Since CTVT had been widely used for tumor models in canine cancers, whether this self-regression may overtake the immunity elicited from an exogenous tumor vaccine remains unclear and certainly worthwhile to be investigated. In this study, we used DCs/tumor hybrids as a tumor vaccine to evaluate the CTVT model. We prepared mature allogeneic dendritic cells from bone marrow and then assessed their phenotype (CD80, CD83, CD86, CD1a, CD11c, CD40 and MHC II), antigen uptake and presenting abilities. Fused dendritic cell/CTVT hybrids were then used as a vaccine, administered three times at two-week intervals via subcutaneous injection near the bilateral auxiliary and inguinal lymph nodes. In comparison with unvaccinated dogs (spontaneous regressed group), within a period of 2.5 months, the vaccinations substantially inhibited tumor progression (p<0.05) and accelerated the rate of regression by a mechanism involving amplification of the host tumor-specific adaptive immune responses and NK cytotoxicity (p<0.001). Pathologic examination revealed early massive lymphocyte infiltration resulting in final tumor necrosis. In addition, there are not any detectable effects on routine physical, body temperature or blood chemistry examinations. In conclusion, our data furnishes a reference value showing that CTVT is a model of potential use for the study of immunity elicited by vaccines against tumors, and also enable early-phase evaluation of the dendritic cell/tumor vaccine in terms of raising host immunity.     

Ultrastructural characteristics of canine transmissible venereal tumor at various stages of growth and regression.

Ultrastructural study of canine transmissible tumors in developing, mature, and regressing stages from 6 dogs revealed the presence of healthy and degenerating tumor cells in all neoplasms. The total number of neoplastic cells seemed to decrease, and the number of degenerating neoplastic cells seemed to increase in mature tumors. Macrophages, lymphocytes, and plasma cells infiltrated mature and regressing tumors. Alteratons in degenerating tumor cells consisted mainly of cytoplasmic changes in early stages and of both nuclear and cytoplasmic changes in cells in which degeneration was more advanced. Amounts of endoplasmic reticulum and ribosomes were decreased. There were swelling and vacuolation of mitochondria. Nuclear chromatin was clumped along the nuclear envelope, and the perinuclear space was widened. Degenerating cells often contained membrane-bound granules and clusters. Lamellar complexes were observed in tumor cells from 2 dogs. Virus particles were not seen.     

Cutaneous transmissible venereal tumor without genital involvement in a prepubertal female dog

An 11-month-old prepubertal crossbreed female dog was presented with multiple nodular lesions disseminated over the cervical, back, flank, and abdominal regions. The lesions were ulcerated and cauliflowerlike, or nodular and subcutaneous, measuring up to 13 cm in diameter. Cytologic preparations of one of the lesions revealed a uniform population of round to oval cells, with lightly basophilic cytoplasm that contained multiple distinct vacuoles. Frequent mitotic figures and occasional lymphocytes were also observed. The cytologic diagnosis was cutaneous transmissible venereal tumor (TVT) in a progressing growth phase. This was confirmed by histologic and immunohistochemical findings. Vaginal TVT was diagnosed later in the dog's mother. TVT is a contagious neoplasm of sexually mature dogs that usually is transmitted by coitus and affects the genital mucosa. To our knowledge, this is the first report of naturally occurring multicentric TVT in a prepubertal female dog and also is unique in its exclusively cutaneous (no mucosal) involvement. We speculate that transmission of neoplastic cells occurred during cohabitation and social/mothering behavior between the dogs. Despite the atypical clinical presentation, response to chemotherapy with vincristine was excellent, leading to complete regression of the neoplasm without relapse after 6 months.     

Role of exfoliative cytology in the diagnosis of canine transmissible venereal tumour

The objectives of this study were to evaluate the role of exfoliative cytology in the diagnosis of canine transmissible venereal tumour and to improve the success rate of surgical excision of this tumour. The technique was used to screen 360 dogs and, at the time of surgery, 34 clinical cases. Seventy-five per cent of the cases detected by screening were described as early stage disease in comparison with 23 per cent of cases reported by owners. The value of exfoliative cytology at the time of surgery to determine tumour cell removal was demonstrated by the reduction in local tumour recurrence from 22 per cent to 8 per cent. The technique is, therefore, recommended for use in screening dogs for canine transmissible venereal tumour and in determining the extent of surgical removal. However, more work is needed to assess its applicability in practice by determining its sensitivity, specificity and predictive values of a positive and negative test answer.     

Cytology of canine cutaneous round cell tumors. Mast cell tumor, histiocytoma, lymphosarcoma and transmissible venereal tumor.

Sixty-four canine cutaneous round cell tumors were divided into 25 mast cell tumors, 15 histiocytomas, nine cutaneous lymphosarcomas and 15 transmissible venereal tumors. The final diagnosis was made from cytologic, clinical and histologic findings. Cytologic features were significantly distinctive in mast cell tumor, transmissible venereal tumor, and most cases of histiocytoma and lymphosarcoma to allow a diagnostic opinion. This opinion was supported by subsequent histologic examination. In some instances cytology was considered essential in rendering a diagnostic opinion even though histology was available.     

Presence of the canine transmissible venereal tumor in the nasal cavity of dogs in the area of dakar (senegal)

Canine transmissible venereal tumors were observed in the nasal passages of three dogs from Dakar, Senegal. Genital tumors were not present in these dogs. These observations, combined with those of few previous reports, stress the necessity to include this neoplasm in the differential diagnosis of nasal tumors in the dog.