Cytological and Histopathological Characteristics of Canine Transmissible Venereal Tumour in Male and Female Dogs Before and After Vincristine Treatment

Canine transmissible venereal tumour (CTVT) is a neoplasia naturally transmitted in susceptible dogs through coitus. CTVT has a worldwide distribution, with a high prevalence in tropical and sub-tropical urban environments. The study aimed at evaluating CTVT lesions in local breeds of dogs and to assess morphological changes based on sex before and after administration of vincristine sulphate. Clinical and gross morphology, fine needle aspirates cytology (FNAC) and routine histopathology methods were used. Two FNAC and histopathological microscopic slide sections from each of the seven sampled dogs were stained with Giemsa stain and Hematoxylin and Eosin. All dogs were treated with vincristine once weekly over a six weeks period after which clinical morphological and histopathological changes were assessed. Grossly, before treatment the tumour masses appeared irregular, cauliflower like with tendency to bleed, sizes ranged from ≥5cm to ≤2cm with or without metastasis to regional lymph nodes. Cytologically, the tumours had homogenous, sheet-like cellular mass. Cytoplasm with punctate vacuoles, anisokaryosis with anisonucleoliosis and coarse to reticulate nuclear chromatin were seen. Lymphocytoid cell pattern was dominant cell type. Histopathology showed sheets of round cells with nuclear and cytoplasmic variations. Histopathology of the treated dog revealed hyper-cellularity, absence of nucleoli, prominent mitotic figures, reduced cell size and presence of inflammatory cells. There was no difference on the cellular changes after vincristine treatment between female and male dogs. Cytology and histopathology showed that vincristine sulphate suppresses the development of tumour through alteration of cellular morphology with no difference between male and female dogs.

Treatment of Canine Transmissible Venereal Tumour using Different Surgico-chemotherapeutic Protocols

Background: Canine transmissible venereal tumour (CTVT) also known as infectious sarcoma, venereal granuloma, transmissible lymphosarcoma or sticker tumour is usually transmitted through coitus and mainly affects the external genitalia of young sexually matured dogs. Surgery, chemotherapy, radiotherapy and immunotherapy are considered as effective treatment protocols. Therefore, depending upon the availability present study was designed to investigate the efficacy of different surgico-chemotherapeutic protocols for treatment of canine transmissible venereal tumour.Methods: The study was conducted during January 2018 to July 2018 at the Teaching Veterinary Clinical Complex (TVCC) and Department of Veterinary Surgery and Radiology, College of Veterinary Science and A.H., Anjora, Durg (C.G.) on 18 canines of various breed, irrespective of age, sex and divided into three groups consisting 6 animals in each group. Group A was treated with surgical excision of tumour only where as Group B and Group C were treated with surgical excision of tumour followed by administration of Doxorubicin (30mg/m2) BSA and Vincristine sulphate (0.025 mg/kg) intravenously alongwith DNS at 7th and 14th post-operative days respectively. Different physiological and haemato-biochemical parameters (Hb, PCV, TLC, TPC, DLC, serum glucose, TSP, SUN, SC, ALT, AST and ALP) were recorded preoperatively, postoperatively and after chemotherapy at 10th, 30th and 60th days intervals.Result: The present investigation showed transient changes in physiological and haemato-biochemical parameters before, post surgery and post chemotherapeutic management and was within normal range. Histopathological examination revealed confluent sheet of tumour cells arranged in large round oval or polyhedral shaped distributed in tight clusters or cords. Group A showed mild to moderated reoccurrence while Group B showed minimum reoccurrence. Group C showed no reoccurrence. Thus, surgery combined with vincristine therapy is most effective for treating dogs suffering with transmissible venereal tumour.

Living infectious agents with the same organic wall assembly of Precambrian early-life fossils discovered in Canine Transmissible Venereal Tumour and human cancer: Giant viruses or living protocells? Evaluating the effects of an anti-cancer vaccine in stray dogs, while challenging the mysteries around the RNA world

ABSTRACTBackgroundCanine Transmissible Venereal Tumour (CTVT) along with Tasmanian Devil Facial Tumour and transmissible leukaemia in Mya Arenaria soft shell-clams are the only examples of contagious cancers occurring in nature. In particular, CTVT is the oldest contagious cancer present in the wild world. The attempts to detect a transmissible virus as a causative agent in these forms of contagious cancer proved conflicting and the current consensus view is that a transformed somatic cell itself is transmitted and starts the tumor in a new animal, as a parasitic allograft. We modify this perception and report for the first time the isolation of an acutely transforming agent from CTVT.MethodsLarge particles were successfully isolated from CTVT specimens through a sucrose gradient, examined at electron microscopy, fully sequenced, used for transformation tests on NIH-3T3 cells and tumorigenic experiments in dogs. A neutralizing therapeutic vaccine was also administered in dogs with natural, not-induced CTVT.ResultsThe particles, isolated from CTVT, are infectious living entities with large dimension. Electron Microscopy reconstructed an organic wall assemblage pattern typical of early life fossils from the Precambrian age, time at which Earth began to form 4.6 billion years ago. Astonishingly, our agents are not fossils, but unicellular organisms biologically active and acutely transforming. They transformed NIH-3T3 cells in vitro and initiated the typical CTVT lesions in healthy dogs, just one week post-infection. Only the fraction containing these infectious entities were able to induce cancer, while a filtered supernatant did not. This ruled out the presence of conventional filterable viruses. RNA sequencing and bioinformatics analyses disclosed a large genome composed by an almost complete Orphan genes dataset, with retro-elements distinct from the host genome. Five doses of a neutralizing vaccine against these oncogenic organisms, drastically reduced the neoplastic mass in dogs with natural, not-induced CTVT. Analogous infectious agents, acutely transforming were also isolated from several human neoplasms.ConclusionsModifying the current believe that contagious cancers are transmitted by a somatic cells allograft, we identified a living agent that infects and starts the typical CTVT in healthy dogs, while its neutralization with a vaccine induces cancer regression in animals with cancer.Significance StatementThese infectious living single-cell agents establish a new family of oncogenic organisms that resist current classifications and affect humans and animals in the wild. While only a dozen of proteins compose a classic virus, these organisms are small infectious cells, but very distinct from somatic eukaryotic cells. The identification of causative unicellular organisms that start cancer in healthy subjects and the possibility to induce cancer regression with a neutralizing vaccine change some perspectives in cancer. The Precambrian features and the genetic composition suggest that these unicellular entities are infectious living RNA protocells that finally gives form to what was considered only a hypothesis drafted by the Nobel laureate Walter Gilmore: the RNA world, the origin of life and RNA protocells.

Evaluation of a canine transmissible venereal tumour cell line with tumour immunity capacity but without tumorigenic property

Introduction:Canine transmissible venereal tumour (CTVT) is a sexually transmitted tumour affecting dogs worldwide, imposing a financial burden on dog owners. A stable culture cell line in continuous passages for >18 months has only been achieved once. The present study investigated a stable CTVT cell line isolated from a bitch and its potential as a vaccine.Material and Methods:A biopsy from a 2-year-old mongrel bitch with CTVT was obtained for histopathological confirmation and isolation of tumour cells. The isolated cells were cultured to passage 55 and characterised by flow cytometry, with karyotyping by GTG-banding and by PCR detection of myc S-2 and LINE AS1. The isolated CTVT cell line was also used as a preventive vaccine in a canine model.Results:Histopathological analysis of the isolated tumour cells revealed typical CTVT characteristics. Constant proliferation and stable morphological characteristics were observed during culture. Phenotypic analysis determined the expression of HLA-DR+, CD5.1+, CD14+, CD45+, CD83+, CD163+, and Ly-6G-Ly-6C+. GTG-banding revealed a mean of 57 chromosomes in the karyotype with several complex chromosomal rearrangements. LINE-c-myc insertion in the isolated CTVT cell line at 550 bp was not detected. However, a 340-bp band was amplified. Isolated CTVT cell line inoculation at a concentration of 1×108did not induce tumour growth in bitches, nor did a challenge with primary CTVT cells.Conclusion:The present study successfully identified and isolated a stable CTVT cell line that may be useful in CTVT prevention.

Multiple mutations acquired into canine RecQ-like helicases encoded by the aneuploid genome of transmissible sarcoma

ABSTRACTSticker sarcoma – a highly aneuploid, contagious neoplasm circulating in a domestic dog population - is broadly referred as a canine transmissible venereal tumour (CTVT). The karyotype of transmissible Sticker sarcoma appears as a collage of numerical and structural aberrations; the CTVT genome represents the generalized but stable neoplastic aneuploidy of monoclonal origins. Presented is an analysis of genetic events and variants underlying the aneuploid genomic structure of Sticker sarcoma described previously by Murchison et al. (2014) and Decker et al. (2015). Here we explored the above CTVT genomic compendia and mined the existing data - specifically looking for cases of convergence of multiple non-synonymous variants onto a single gene - the mutational patterns indicative for Knudsonian ‘two-hit’ kinetics. A Table I is given, providing theoretical estimates of retaining the intact wild-type copy, expected as a function of a cumulative mutational convergence observed in unphased sequence consensus. We demonstrate that the two canine RecQ-like helicases: Bloom syndrome helicase and RECQL4, encoded by the aneuploid transmissible tumour, have accumulated a multitude of different mutations. Among the sets of most intensely mutated transmissible sarcoma genes, we also identified a canine FANCD2 – yet another previously unnoticed multiple-hit candidate factor. We discuss a possible role of mutated RecQ-like helicases and other cooperating factors, perceivably involved in the maintenance of the neoplastic aneuploidy. We suggest the proposed cooperative actions of CTVT RecQ-like DNA helicases could be relevant interpreting whether variants contributing to RecQ-dependent karyotypic traits, respond to selective pressures that preserve the aneuploid genomic structure of transmissible Sticker sarcoma.